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ORGANIC SPECTROSCOPY

Read all about Organic Spectroscopy on ORGANIC SPECTROSCOPY INTERNATIONAL 

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DR ANTHONY MELVIN CRASTO Ph.D

DR ANTHONY MELVIN CRASTO Ph.D

DR ANTHONY MELVIN CRASTO, Born in Mumbai in 1964 and graduated from Mumbai University, Completed his Ph.D from ICT, 1991,Matunga, Mumbai, India, in Organic Chemistry, The thesis topic was Synthesis of Novel Pyrethroid Analogues, Currently he is working with GLENMARK PHARMACEUTICALS LTD, Research Centre as Principal Scientist, Process Research (bulk actives) at Mahape, Navi Mumbai, India. Total Industry exp 30 plus yrs, Prior to joining Glenmark, he has worked with major multinationals like Hoechst Marion Roussel, now Sanofi, Searle India Ltd, now RPG lifesciences, etc. He has worked with notable scientists like Dr K Nagarajan, Dr Ralph Stapel, Prof S Seshadri, Dr T.V. Radhakrishnan and Dr B. K. Kulkarni, etc, He did custom synthesis for major multinationals in his career like BASF, Novartis, Sanofi, etc., He has worked in Discovery, Natural products, Bulk drugs, Generics, Intermediates, Fine chemicals, Neutraceuticals, GMP, Scaleups, etc, he is now helping millions, has 9 million plus hits on Google on all Organic chemistry websites. His friends call him Open superstar worlddrugtracker. His New Drug Approvals, Green Chemistry International, All about drugs, Eurekamoments, Organic spectroscopy international, etc in organic chemistry are some most read blogs He has hands on experience in initiation and developing novel routes for drug molecules and implementation them on commercial scale over a 30 year tenure till date Dec 2017, Around 35 plus products in his career. He has good knowledge of IPM, GMP, Regulatory aspects, he has several International patents published worldwide . He has good proficiency in Technology transfer, Spectroscopy, Stereochemistry, Synthesis, Polymorphism etc., He suffered a paralytic stroke/ Acute Transverse mylitis in Dec 2007 and is 90 %Paralysed, He is bound to a wheelchair, this seems to have injected feul in him to help chemists all around the world, he is more active than before and is pushing boundaries, He has 9 million plus hits on Google, 2.5 lakh plus connections on all networking sites, 50 Lakh plus views on dozen plus blogs, He makes himself available to all, contact him on +91 9323115463, email amcrasto@gmail.com, Twitter, @amcrasto , He lives and will die for his family, 90% paralysis cannot kill his soul., Notably he has 19 lakh plus views on New Drug Approvals Blog in 216 countries......https://newdrugapprovals.wordpress.com/ , He appreciates the help he gets from one and all, Friends, Family, Glenmark, Readers, Wellwishers, Doctors, Drug authorities, His Contacts, Physiotherapist, etc

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AKN 028


img

AKN-028
CAS 1175017-90-9
Chemical Formula: C17H14N6
Molecular Weight: 302.33

N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine

N2-(1H-indol-5-yl)-6-(pyridin-4-yl)pyrazine-2,3-diamine

  • Originator Swedish Orphan Biovitrum
  • Developer Akinion Pharmaceuticals
  • Class Antineoplastics; Small molecules
  • Mechanism of Action Fms-like tyrosine kinase 3 inhibitors; Proto oncogene protein c-kit inhibitors
  • Phase I/II Acute myeloid leukaemia
  • 01 Mar 2016 Akinion Pharmaceuticals terminates phase I/II trial in Acute myeloid leukaemia in Czech Republic, Poland, Sweden and United Kingdom (NCT01573247)
  • 17 Sep 2015 AKN 028 is still in phase I/II trials for Acute myeloid leukaemia in Czech Republic, Poland and Sweden
  • 09 Apr 2014 AKN 028 is still in phase I/II trials for Acute myeloid leukaemia in Czech Republic, Poland and Sweden

AKN-028, a novel tyrosine kinase inhibitor (TKI), is a potent FMS-like receptor tyrosine kinase 3 (FLT3) inhibitor (IC(50)=6 nM), causing dose-dependent inhibition of FLT3 autophosphorylation. Inhibition of KIT autophosphorylation was shown in a human megakaryoblastic leukemia cell line overexpressing KIT. In a panel of 17 cell lines, AKN-028 showed cytotoxic activity in all five AML cell lines included. AKN-028 triggered apoptosis in MV4-11 by activation of caspase 3. In primary AML samples (n=15), AKN-028 induced a clear dose-dependent cytotoxic response (mean IC(50) 1 μM). However, no correlation between antileukemic activity and FLT3 mutation status, or to the quantitative expression of FLT3, was observed. Combination studies showed synergistic activity when cytarabine or daunorubicin was added simultaneously or 24 h before AKN-028. In mice, AKN-028 demonstrated high oral bioavailability and antileukemic effect in primary AML and MV4-11 cells, with no major toxicity observed in the experiment. (source: Blood Cancer J. 2012 Aug 3;2:e81. doi: 10.1038/bcj.2012.28.)

SYN

WO 2013/089636

Clip

Development of a Synthesis of Kinase Inhibitor AKN028

 R&D DepartmentMagle Chemoswed, P.O. Box 839, SE 201 80 Malmö, Sweden
 Recipharm OT ChemistryVirdings Allé 32 B, SE 754 50 Uppsala, Sweden
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.8b00092
*Telephone: +46 704473035. E-mail: johan.docera@gmail.com
Abstract Image

The novel tyrosine kinase inhibitor AKN028 has demonstrated promising results in preclinical trials. An expedient protocol for the synthesis of the compound at kilogram scale is described, including an SNAr reaction with high regioselectivity and a Suzuki coupling. Furthermore, an efficient method for purification and removal of residual palladium is described.

yellow or faint-orange powder. Mp 300 °C (dec.);

IR 3133 broad, 1689, 1597, 1554, 1480 cm–11H NMR (DMSO-d6) δ 11.01 (s, 1H), 8.62–8.50 (m, 2H), 8.22 (s, 1H), 8.15 (s, 1H), 8.06 (s, 1H), 7.89–7.82 (m, 2H), 7.39 (d, J = 2.0 Hz, 2H), 7.32 (t, J = 2.7 Hz, 1H), 6.77 (s, 2H), 6.42 (dd, J1 = 8.7 Hz, J2 = 2.0 Hz, 1H);

13C NMR (DMSO-d6) δ 149.9, 145.2, 145.0, 139.6, 132.8, 132.4, 132.2, 128.4, 127.6, 125.6, 118.7, 116.1, 111.2, 111.0, 101.0.

PATENT

 WO 2009095399

https://patentscope.wipo.int/search/ko/detail.jsf;jsessionid=074E97C06EF8C2088428DECCA2CD2EBA.wapp1nB?docId=WO2009095399&recNum=208&office=&queryString=&prevFilter=%26fq%3DOF%3AWO%26fq%3DICF_M%3A%22C07D%22%26fq%3DDP%3A2009&sortOption=Pub+Date+Desc&maxRec=3425

PATENT

WO 2013089636

https://patents.google.com/patent/WO2013089636A1/ko

Protein kinases are involved in the regulation of cellular metabolism, proliferation, differentiation and survival. The FLT-3 (fms-like tyrosine kinase) receptor is a member of the class III subfamily of receptor tyrosine kinases and has been shown to be involved in various disorders such as haematological disorders, proliferative disorders, autoimmune disorders and skin disorders.

In order to function effectively as an inhibitor, a kinase inhibitor needs to have a certain profile regarding its target specificity and mode of action. Depending on factors such as the disorder to be treated, mode of administration etc. the kinase inhibitor will have to be designed to exhibit suitable properties. For instance, compounds exhibiting a good plasma stability are desirable since this will provide a pharmacological effect of the compounds extending over time. Another example is oral administration of the inhibitor which may require that the inhibitor is transformed into a prodrug in order to improve the bioavailability.

WO 2009/095399 discloses pyrazine compounds acting as inhibitors of protein kinases, especially FTL3, useful in the treatment of haematological disorders, proliferative disorders, autoimmune disorders and skin disorders. This document discloses methods for manufacturing such compounds. However these methods are not suitable for large scale processes and the chemical yields are moderate. Furthermore, the compounds obtained by these methods are in amorphous form.

n one aspect of the invention, there is provided a process for preparing a compound of formula (I)

said process comprises the steps of:

a) reacting a compound of formula (1) with a compound of formula (2) in an inert solvent and in the presence of an (C1-6alkyl)3amine, providing a compound of formula (3):


, b) Suzuki coupling of a compound of formula (3) and a compound of formula (4) in an inert solvent and in the presence of a palladium catalyst and a base, providing a crude product comprising a compound of formula (I) and palladium

and

c) removing the palladium from the crude product in step b).

The compound of formula (I) may be obtained in amorphous or crystalline form using the processes outlined below.

Step 1:

Reaction of 2-amino-3,5-dibromopyrazine (1) and 5-aminoindole (2) in a

nucleophilic substitution reaction in the presence of a C1-6alkylamine and an inert polar solvent yields 3-bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine (3). Examples of inert polar solvents are DMSO, water and NEP. Examples of (C1-6alkyl)3amine are triethylamine, trimethylamine and tributylamine. The reaction may be performed at reflux temperature or at about 100-130°C.

Step 2:

A Suzuki coupling of 3-bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine) (3) and 4- pyridyl-boronic acid (4) in an inert polar solvent in the presence of a palladium catalyst and a base yields N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine (I) in amorphous form. Examples of inert solvents are DMF, water and DMA. Examples of palladium catalysts are Pd(dppf) and Pd(OAc)2-DTB-PPS. Example of a base is

K2CO3 The reaction may be performed under inert and oxygen-free atmosphere such as nitrogen or argon.

Heating may take place during step 1 and/or step 2. Steps 1 and 2 may be performed at reflux or in a temperature range of from 100 to 140°C, such as from 105 to 135°C, such as from 110 to 130°C, such as from 130-135°C, such as from 110-115ºC.

Step 3:

A compound of formula (I), also denominated N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine, in amorphous form may be dissolved in acetic acid (HOAc) after which potassium hydroxide (KOH) is added. The compound of formula (I) in amorphous form may be obtained from the process outlined in steps 1 and 2.

Alternatively, the compound of formula (I) may be obtained according to the process described in WO 2009/095399. The obtained crystalline form is removed from the slurry by, for instance, filtration. Step 3 may be repeated. Step 3 may be performed at a temperature of about 40°C followed by cooling to room temperature.

The process for preparing a compound according to formula (I) may comprise an additional step (step i) between step 2 and step 3 in order to remove palladium from the crude product of the compound of formula (I). The step comprises; forming a slurry comprising an acid and the compound according to formula (I) in a solvent, adding a siloxane compound to said slurry, removing the solvent from the slurry and adding an organic solvent, such as DMF and/or toluene, to the solid formed whereby a mixture is formed and then potassium hydroxide is added to the formed mixture, Alternatively, palladium may be removed from the crude product comprising (I) using a palladium scavenger such as TMT and/or 3-mercaptopropyl ethyl sulfide silica.

The crystalline form of the compound according to formula (I) may also be prepared from an amorphous form of the compound according to formula (I) by dissolving said amorphous form of the compound in a solvent mixture of

dichloromethane/methanol followed by evaporation of the solvent in a rotary evaporator. The amorphous form of the compound of formula (I) may obtained using the process disclosed in WO 2009/095399.

Example 1. Preparation of 5-Bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine (compound 3)

DMSO (10 L, 11 kg), 2-amino-3,5-dibromopyrazine (1) (4.5 kg, 17.8 mol, 1 eq.), 5- amino indole (2) (3.06 kg, 23.15 mol, 1.3 eq.) and triethylamine (7.4 L, 5.4 kg, 53.36 mol, 3 eq.) were charged to a reactor. The reaction mixture was heated to 95°C while agitated. After 12 hours, the heating was discontinued and the conversion was 88% of 2-amino-3,5-dibromopyrazine. The reaction was heated again to 95°C and

agitated for an additional 2.5 hours. There was no improvement in conversion. The reaction mixture was agitated at ambient temperature overnight. Triethylamine (3.5 kg) was removed under vacuum and the remaining reaction mixture was transferred to a stainless steel container from which it was charged into another reactor.

Subsequently, 18.4 kg of 50% acetic acid (aq.) was introduced over a period of 20 minutes under agitation, followed by purified water (61 L) charged over a period time of 60 minutes. The slurry was then filtered and the isolated material was washed with 2 x 20 L of 1% acetic acid (aq.).

The isolated 3-bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine) (3) was transferred to a drying cabinet and dried to invariable weight at 40 ±3°C, (19 hours), to afford 4.36 kg, 14.34 mol, 81 % yield, with a purity of 96% by HPLC.

The reaction temperature in the batch record was set to be 130-135°C. However, at 95°C the reaction mixture was at reflux.

Example 2. Preparation of N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3- diamine (compound I)

To a reactor was charged N,N-dimethylformamide (46.7 L, 45 kg), 4-pyridylboronic acid (4) (2.64 kg, 21.5 mol, 1.5 eq.) and 5-bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3- diamine (3) (4.36 kg, 14.3 mol). The reactor was then flushed with nitrogen prior to the charging of Pd(dppf)Cl2-catalyst (0.47 kg, 0.55 mol, 0.04 eq.). To reactor was then charged, over a period of 20 minutes, 24.9 kg of a 2 M solution of potassium carbonate (aq.). The reactor was flushed with nitrogen and heated under agitation to 110-115°C for 1.5 hours, after which 98.3% conversion of (3) was showed. The reaction mixture was quenched by addition of purified water (180 L) under vigorous agitation. The precipitated material was isolated on a hastalloy filter and washed with purified water (50 L), The isolated material was transferred to a drying cabinet and dried to invariable weight at 40 ±3°C (18 hours), to afford a compound of formula (5), i.e. a compound of formula (!) also denominated N-3-(1H-lndol-5-yl)-5-pyhdin-4-yl-pyrazine-2,3-diamine, (3.64 kg, 12.1 mol, 85 % yield).

During the process precipitated material was observed in the solutions, after the reactions, in both steps not previously seen in lab-scale. These impurities were not removed.

Example 3. Purification and crystallisation

In order to remove residual solvents from the material, two consecutive re-precipitations of the material from acetic acid were performed. This also gave crystallinity of the isolated substance. The purification is performed in order to remove palladium.

Purification

To a 1 L round bottomed flask was added 37.8 g of a compound according to formula (I) followed by 600 mL 2 M HOAc (aq.). The material was stirred at RT until a clear, dark red solution was obtained. To the solution was added 30 g Hyflo Super Celite and the slurry was filtered. The filter cake was washed with 25 mL 2 M HOAc

(aq) and 2×35 mL purified water. The obtained filtrate was transferred to a 2 L round bottomed flask containing 950 mL of Me-THF. The mixture was then stirred and heated to 40°C for 30 minutes. To the solution was then added 290 mL 8 M KOH (aq.) at 40°C and pH in the solution was 14.

The aqueous phase was removed and the organic phase washed with 2×100 mL of purified water. The remaining organic phase was then transferred to a 2 L round bottomed flask, followed by 95 mL of DMF, 20 g scavenger 3-Mercaptopropyl ethyl sulphide silica, Phosphonics LTD and 20 g scavenger 2-Mercaptoethyl ethyl sulfide silica purchased from Phosphonics LTD. The solution was vigorously stirred and heated at 60°C. A sample was withdrawn from the slurry after 12 hours, and showed 6 ppm of palladium remaining in the solution. The mixture was allowed to cool and was then filtered to remove the scavenger. The round bottomed flask and filter were rinsed with a mixture of 90 mL Me-THF and 10 mL DMF. Me-THF was then removed on a rotary evaporator and the remaining slurry was azeotropically dried with two portions of 100 mL toluene. To the remaining slurry was then added 85 mL of DMF to a total of 185 mL DMF (5ml DMF/g substance). To the clear solution was then added, slowly, while agitated, 1500 mL of toluene which produced a heavy precipitate. The slurry was filtered off and washed with 2×50 mL of toluene where after the material was dried overnight at 35°C under vacuum to afford 30.9 g of a compound according to formula (I) in a yield of 82%.

Crystallisation:

Example i

1. First re-precipitation

The N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine material (30.9 g) was added to a 1 L round bottomed flask and 450 mL 2 M HOAc (aq.) was added. The slurry was agitated and heated to 40°C for 1 hour, until the material had dissolved. To the solution was then added 158 mL 8 M KOH (aq.) at 40°C. The pH in the solution was 11.4. The slurry was then allowed to cool to 25°C and filtered. The filter cake was washed with 3x 80 mL of purified water and the material was dried overnight at 95°C under vacuum to afford 28.7g N-3-(1H-indol-5-yl)-5-pyridin-4-yl- pyrazine-2,3-diamine in a yield of 93%.

2. Second re-precipitation

N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine material (28.7 g) was added to a 1L round bottomed flask and 430 mL 2 M HOAc (aq) was added. The slurry was agitated and heated to 40°C for 1 hour, until the material had dissolved. To the solution was then added 15 mL 8M KOH (aq) at 40°C. The pH in the solution was 12.3. The slurry was then allowed to cool to 25°C and filtered. The filter cake was washed with 5×50 mL of purified water, and the solid was then dried overnight at 95°C under vacuum to afford 28.3 g N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3- diamine in a yield of 99%.

Example ii

The N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine material (2.1 kg, 7 mol) was added to a reactor, followed by 2M HOAc (aq.) (59.6 L, 60.2 kg) . The solution in the reactor was then heated to 40°C and stirred for 20 minutes. To the clear solution was then charged, slowly, 30% KOH (aq.) (25 kg) under vigorous agitation. The slurry was agitated for 15 minutes. pH in the solution was 6.2, and a total of 1.5 kg 30% KOH (aq.) was then added to the solution to give pH 12.1. The precipitated material was isolated on a Hastelloy filter and washed with purified water (5×30 L). The solid was then transferred to a drying cabinet and dried to invariable weight at 85 ±3°C under vacuum (16 hours; a sample was withdrawn after 16 hours, showing 1400 ppm HOAc and 75 ppm DMF), to afford N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine (2.0 kg, 7 mol, 95 % yield).

Hence, N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine is obtained in an uniform crystalline form, which was achieved by precipitating the product from aqueous acetic acid by introduction of aqueous potassium hydroxide.

Example 5. Synthesis of 5-Bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine (compound 3)

2-Amino-3,5-dibromopyrazine (45 g, 1.0 eq.), 5-aminoindole (30,6 g, 1.3 eq.), 67.5 mL NEP, i.e. 1-ethyl-2-pyrrolidone, and 74.5 mL triethylamine were added to a 250 mL reactor. The jacket temperature was set to 130°C and the reaction mixture was stirred for 22 h. HPLC after 22 h showed 87% conversion of the 2-amino-3,5-dibromopyrazine. After 24 h HPLC showed 92% conversion and the reaction slurry was cooled to 80°C and quenched by addition of addition of 50% HOAc(aq) and water. The obtained slurry was then allowed to cool to room temperature over night while agitated. The material was isolated on a glass filter funnel and was washed with water. The material was dried at 80 °C under vacuum until dry to afford 71% of the compound 5-bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine as a dark brown powder. The purity was 99.8% as measured by HPLC.

Example 6. Synthesis of N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine (Compound I)

5-Bromo-N-3-(1H-indol-5-yl)-pyrazine-2,3-diamine (15.0 g, 49 mmol, 1.0 eq.), 4-pyridyl boronic acid (6.6 g, 59 mmol, 1.2 eq.), Pd(OAc)2 (166 mg, 0.74 mmol, 0.015 eq.), DTB-PPS, i.e. 3-(di-tert-butylphosphino)propane-1-sulfonic acid, (199 mg, 0.74 mmol, 0.015 eq.), and DMA, i.e. N,N-dimethylacetamide, (75 mL) were added to a three-necked round-bottomed flask equipped with a mechanical stirrer,

thermometer, and a nitrogen atmosphere. Through a septa was added 2M K2CO3 (aq) (27 ml, 54 mmol, 1.1 eq.) with a syringe. The temperature was increased to 100 °C. Samples for HPLC-analysis of the conversion were drawn and when the conversion had reached 100% the temperature was cooled to 25 °C. At that temperature a water solution of 0.5 M L-cysteine (150 ml) was added by a syringe pump over 1 hour with a rate of 2.5 mL/minute. After 3 hours maturing time at room temperature the material was isolated on a glass filter funnel and was washed with water. The material was dried at 40 °C under vacuum over the weekend, and 15 grams of N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine (101%) were obtained as a brown powder.

Example 7. Purification of N-3-(1H-indol-5-yl)-5-pyridin-4-yl-pyrazine-2,3-diamine (Compound I)

The crude (7.0 g, 23 mmol) and 2M HOAc (98 mL) was added to a 250 mL round-bottomed flask. To this was added TMT, i.e. trithiocyanuric acid, (1.4 g) and SPM32, i.e. 3-mercaptopropyl ethyl sulfide silica, (1.4 g). The mixture was stirred in room temperature for 24 hours. After 24 hour a polish filtration through hyflo super cel was performed. To the clear filtrate was added 50 mL 5 M KOH(aq) under 15 minutes to precipitate the product. After 18 hours maturing time at room temperature the material was isolated on a glass filter funnel and was washed with 2×20 mL water. The first was being a slurry wash and the second a displacement wash. The material was dried at 40 °C under vacuum over the weekend, and 3.9 grams (56%) was obtained as a light yellow powder. The Pd content was 3.7 ppm.

PATENT

US 8436171

PATENT

WO 2016008433

PATENT

WO 2016015604

PATENT

WO 2016015597

PATENT

WO 2016015605

PATENT

WO 2016015598

PATENT

WO 2017146794

PATENT

WO 2017146795

https://patents.google.com/patent/WO2017146795A1/en

PATENT

US 20180071302

REFERENCES

1: Eriksson A, Hermanson M, Wickström M, Lindhagen E, Ekholm C, Jenmalm Jensen A, Löthgren A, Lehmann F, Larsson R, Parrow V, Höglund M. The novel tyrosine kinase  inhibitor AKN-028 has significant antileukemic activity in cell lines and primary cultures of acute myeloid leukemia. Blood Cancer J. 2012 Aug 3;2:e81. doi: 10.1038/bcj.2012.28. PubMed PMID: 22864397; PubMed Central PMCID: PMC3432483.

////////////AKN028 , AKN-028 , AKN 028, phase 2, Swedish Orphan Biovitrum,  Akinion Pharmaceuticals,  Acute myeloid leukaemia

NC1=NC=C(C2=CC=NC=C2)N=C1NC3=CC4=C(NC=C4)C=C3

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Taladegib (LY-2940680),


Taladegib.png

Taladegib

LY2940680; 1258861-20-9; Taladegib; LY-2940680; UNII-QY8BWX1LJ5; QY8BWX1LJ5

CAS 1258861-20-9 FREE , CAS HCL 1258861-21-0
4-Fluoro-N-methyl-N-{1-[4-(1-methyl-1H-pyrazol-5-yl)-1-phthalazinyl]-4-piperidinyl}-2-(trifluoromethyl)benzamide
Benzamide, 4-fluoro-N-methyl-N-[1-[4-(1-methyl-1H-pyrazol-5-yl)-1-phthalazinyl]-4-piperidinyl]-2-(trifluoromethyl)-
LY 2940680

4-fluoro-N-methyl-N-[1-[4-(2-methylpyrazol-3-yl)phthalazin-1-yl]piperidin-4-yl]-2-(trifluoromethyl)benzamide

Molecular Formula: C26H24F4N6O
Molecular Weight: 512.513 g/mol

Taladegib is an orally bioavailable small molecule antagonist of the Hedgehog (Hh)-ligand cell surface receptor smoothened (Smo) with potential antineoplastic activity. Taladegib inhibits signaling that is mediated by the Hh pathway protein Smo, which may result in a suppression of the Hh signaling pathway and may lead to the inhibition of the proliferation of tumor cells in which this pathway is abnormally activated. The Hh signaling pathway plays an important role in cellular growth, differentiation and repair; constitutive activation of this pathway is associated with uncontrolled cellular proliferation and has been observed in a variety of cancers.

Taladegib has been used in trials studying the treatment of Solid Tumor, COLON CANCER, BREAST CANCER, Advanced Cancer, and Rhabdomyosarcoma, among others.

Image result for Taladegib

  • Originator Eli Lilly
  • Developer Eli Lilly; Ignyta
  • Class Antineoplastics; Benzamides; Fluorobenzenes; Phthalazines; Piperidines; Pyrazoles; Small molecules
  • Mechanism of Action Hedgehog cell-signalling pathway inhibitors; SMO protein inhibitors

Highest Development Phases

  • Phase I/II Oesophageal cancer; Small cell lung cancer
  • Phase I Ovarian cancer; Solid tumours
  • Preclinical Basal cell cancer
  • No development reported Cancer

Most Recent Events

  • 04 Nov 2017 No recent reports of development identified for phase-I development in Solid-tumours(Late-stage disease, Second-line therapy or greater) in Japan (PO, Tablet)
  • 02 Jun 2017 Adverse events data from a phase I/II trial in Ovarian cancer (Solid tumours) presented at the 53rd Annual Meeting of the American Society of Clinical Oncology (ASCO-2017)
  • 23 Mar 2017 Ignyta amends its license, development and commercialisation agreement with Eli Lilly for taladegib

SYN

PATENT

WO 2010147917

Preparation 1 ter?-Butyl 1 -(4-chlorophthalazin- 1 -yl)piperidin-4-yl(methyl)carbamate

Heat a mixture of potassium carbonate (21.23 g, 153.6 mmol), 1,4-dichlorophthalazine (26 g, 128 mmol) and methyl-piperidin-4-yl carbamic acid ter?-butyl ester (30.01 g, 134.4 mmol) in N-methylpyrrolidine (200 mL) at 80 0C overnight. Pour the reaction mixture into water, extract with dichloromethane, dry over Na2SC”4, and concentrate under reduced pressure. Add diethylether and filter off the resulting solid (4-chlorophethalazin-1-ol from starting material impurity). Concentrate the filtrate. Purify the resulting residue by flash silica gel chromatography (hexane : ethyl acetate = 2 : 1) to X-18698

-9- provide the title compound as a white solid (17.66 g, 37%). ES/MS m/z (37Cl) 377.0 (M+ 1).

Preparation 2 fer?-Butyl 1 -(4-chlorophthalazin- 1 -yl)piperidin-4-ylcarbamate

Prepare the title compound by essentially following the procedure described in Preparation 1 , using piperidin-4-yl-carbamic acid tert-butyl ester. Cool the reaction mixture and pour into water (500 mL). Extract with ethyl acetate, wash with water, dry over Na2SC”4, and remove the solvents under reduced pressure to provide the title compound as a yellow solid (36 g, 97%). ES/MS m/z 363.0 (M+l).

Preparation 3 ter?-Butyl methyl( 1 -(4-( 1 -methyl- lH-pyrazol-5 -yl)phthalazin- 1 -yl)piperidin-4- yl)carbamate

Place sodium carbonate (3.82 g, 36.09 mmol), tert-butyl 1 -(4-chlorophthalazin- 1-yl) piperidin-4-yl(methyl)carbamate (6.8 g, 18.04 mmol) and 1 -methyl- lH-pyrazole-5-boronic acid pinacol ester (5.63 g, 27.1 mmol) in a flask with a mixture of toluene (50 mL), ethanol (17 mL), and water (17 mL). Degas the mixture for 10 min with nitrogen gas. Add tetrakis(triphenylphosphine)palladium (0.4 g, 0.35 mmol) and heat the mixture at 74 0C overnight. Cool the mixture to ambient temperature and dilute with dichloromethane. Wash the organic portion with brine, dry over Na2SC”4, and concentrate under reduced pressure. Purify the resulting residue by flash silica gel chromatography X-18698

-10-

(hexane : ethyl acetate : 2 M NH3 in MeOH = 20 : 5 : 1) to provide the title compound as a yellow foam (5.33 g, 70%). ES/MS m/z 423.2 (M+ 1).

Alternate procedure to prepare tert-butyl methyl(l-(4-(l-methyl-lH-pyrazol-5-yl)phthalazin-l-yl)piperidin-4-yl)carbamate: Preparations 4 – 6

Preparation 4

1 ,4-Dibromophthalazine


Charge a pressure tube with phosphorus pentabromide (24.5 g, 54.1 mmol) and

2,3-dihydro-phthalazine-l,4-dione (5.00 g, 30.8 mmol). Seal the tube and heat at 140 0C for 6-7 h. Allow to cool overnight. Carefully open the tube due to pressure. Chisel out the solid and pour into ice water. Allow to stir in ice water and collect the resulting solid by vacuum filtration. Dry in a vacuum oven to obtain the final product (8.31 g, 93%). ES/MS (79Br, 81Br) m/z 288.8 (M+). Ref: Can. J. Chem. 1965, 43, 2708.

Preparation 5 ter?-Butyl 1 -(4-bromophthalazin- 1 -yl)piperidin-4-yl(methyl)carbamate


Combine 1 ,4-dibromophthalazine (0.70 g, 2.38 mmol), N-methylpyrrolidone (7.0 mL), potassium carbonate (395 mg, 2.86 mmol), and methyl-piperidin-4-yl-carbamic acid ter?-butyl ester (532 mg, 2.38 mmol). Heat at 80 0C overnight. Cool and pour into water. Collect the solid and dry in a vacuum oven at ambient temperature overnight to obtain the final product (0.96 g, 95%). ES/MS m/z (81Br) 421.0 (M+ 1).

X-18698

-11-

Preparation 6 fer?-Butyl methyl (l-(4-(l -methyl- lH-pyrazol-5-yl)phthalazin-l-yl)piperidin-4- yl)carbamate


Charge a reaction tube with fer?-butyl l-(4-bromophthalazin-l-yl)piperidin-4-yl(methyl)carbamate (500 mg, 1.2 mmol), 1 -methyl- lH-pyrazole-5-boronic acid pinacol ester (370 mg, 1.8 mmol), sodium carbonate (252 mg, 2.4 mmol), toluene (3.75 mL), ethanol (1.25 mL), and water (1.25 mL). Degas the reaction mixture with nitrogen for 10 min. Add tetrakis (triphenylphosphine) palladium (137.1 mg, 118.7 μmol). Bubble nitrogen through the reaction mixture for another 10 min. Cap the reaction vial and heat at 90 0C overnight. Cool the reaction and filter through a silica gel pad eluting with 5% MeOH : CΗ2CI2. Concentrate the fractions under reduced pressure. Purify the resulting residue using silica gel chromatography (2% 2 N NH3 in MeOHiCH2Cl2) to obtain the final product (345.6 mg, 69%). ES/MS m/z 423.2 (M+ 1).

Preparation 7 ter?-Butyl 1 -(4-( 1 H-pyrazol-5 -yl)phthalazin- 1 -yl)piperidin-4-yl(methyl)carbamate

Prepare the title compound by essentially following the procedure described in Preparation 3, using tert-buty\ l-(4-chlorophthalazin-l-yl)piperidin-4-yl(methyl)carbamate and lH-pyrazole-3-boronic acid pinacol ester to provide 580 mg,

(67%). ES/MS m/z 409.2 (M+ 1).

Preparation 8 X-18698

-12- tert- Butyl 1 -(4-(I -methyl- lH-pyrazol-5-yl)phthalazin- 1 -yl)piperidin-4-ylcarbamate

Prepare the title compound by essentially following the procedure described in Preparation 3, using tert-bυXy\ 1 -(4-chlorophthalazin- 1 -yl)piperidin-4-ylcarbamate to provide 5.92 g (94%). ES/MS m/z 308.8 (M+).

Preparation 9 iV-methyl- 1 -(4-( 1 -methyl- lH-pyrazol-5-yl)phthalazin- 1 -yl)piperidin-4-amine


Dissolve tert-bvAyl methyl(l-(4-(l-methyl-lH-pyrazol-5-yl)phthalazin-l-yl)piperidin-4-yl)carbamate (7.77 g, 18.39 mmol) in dichloromethane (100 mL). Add an excess of 1 M hydrogen chloride in diethyl ether (20 mL, 80 mmol) to the solution and stir at ambient temperature for 2 h. Concentrate under reduced pressure. Purify the resulting residue by flash silica gel chromatography (dichloromethane : 2 M NΗ3 in MeOH = 10 : 1) to provide the title compound as a yellow foam (5.83 g, 98%). ES/MS m/z 323.2 (M+ 1).

Example 1

4-Fluoro-N-methyl-N-(l-(4-(l-methyl-lH-pyrazol-5-yl)phthalazin-l-yl)piperidin-4-yl)-2- (trifluoromethyl)benzamide

Treat a solution of N-methyl-1 -(4-(I -methyl- lH-pyrazol-5-yl)phthalazin-l-yl)piperidin-4-amine (2.8 g, 8.68 mmol) and triethylamine (3.36 mL, 26.1 mmol) in CH2Cl2(30 mL) with 4-fluoro-2-(trifluoromethyl)benzoyl chloride (2.14 mL, 10.42 mmol). Stir for 3 h at ambient temperature. Concentrate the reaction mixture under reduced pressure. Purify the resulting residue by flash silica gel chromatography (hexane : ethyl acetate : 2 M ΝH3 in MeOH = 20 : 5 : 1) to provide the free base as a yellow foam (3.83 g, 86%). ES/MS m/z 513.0 (M+ 1).

Example Ia

4-Fluoro-N-methyl-N-(l-(4-(l-methyl-lH-pyrazol-5-yl)phthalazin-l-yl)piperidin-4-yl)-2- (trifluoromethyl)benzamide hydrochloride X-18698

-14-

Dissolve 4-fluoro-N-methyl-N-(l -(4-(I -methyl- lH-pyrazol-5-yl)phthalazin-l- yl)piperidin-4-yl)-2-(trifluoromethyl)benzamide (7.13 g, 13.91 mmol) in dichloromethane (100 mL) and add excess 1 N HCl in diethyl ether (30 mL, 30 mmol). Remove the solvents under reduced pressure to provide the title compound (7.05 g, 92%). ES/MS m/z 513.0 (M+ 1). NMR showed a 2:l mixture of amide rotamers. Major rotamer; 1H NMR (400 MHz, DMSOd6): δ 8.34 (m, IH), 8.26 (m, 2H), 7.95 (m, IH), 7.75 (m, IH), 7.64 (m, 2H), 7.55 (m, IH), 6.72 (d, IH, J=2Hz), 5.15 (br, IH), 4.71 (m, IH), 4.22 (m, 2H), 3.84 (s, 3H), 3.48 (m, 2H), 2.65 (s, 3H), 2.19 (m, 2H), 1.89 ( m, 2H). Minor rotamer; 1H NMR (400 MHz, DMSOd6): δ 8.27 (m, IH), 8.24 (m, 2H), 7.94 (m, IH), 7.73 (m, IH), 7.63 (m, 3H), 6.70 (d, IH, J=2Hz), 5.15 (br, IH), 4.71 (m, IH), 4.07 ( m, 2H), 3.81 (s, 3H), 3.16 (m, 2H), 2.92 (s, 3H), 1.90 (m, 2H), 1.62 ( m 2H).

PATENT

CN 106279114

Example 5 Preparation of title compound LY-2940680 [0061] Embodiment

[0062] Compound 10 (0.2g, 0.429mmo 1,1 eq.) Was dissolved in a mixed solution of 18mL of toluene, 6 mL of ethanol, 6 mL of water was added to a solution of 0.091g (0.858mmol, 2eq.) Sodium carbonate which ester (CAS No. 847818-74-0) and 0.098g (0.472mmol, 1 · leq.) in 1-methyl -1H- pyrazole-5-boronic acid, degassed with nitrogen for 20min after addition of 60mg of four (triphenylphosphine) palladium, degassed with nitrogen for lOmin, homogeneous reaction was stirred at reflux for 12h at 74 ° C; after completion the reaction was cooled to room temperature, diluted with methylene chloride, the organic phase washed three times with brine, dried no over anhydrous sodium sulfate, and concentrated under reduced pressure to give a crude product, purified by column chromatography (eluent dichloromethane / methanol, a volume ratio of 30: 1) to give the desired product as a pale yellow foam LY-2940680 (0 · 202g, 92% yield).

[0063] The title compound of detection data LY-2940680:

[0064] 1 ^: 951 ^ 4 ^^ (3001 ^, 0) (: 13) 38.09 ((1 (1,1 = 7.7 ^ 11 (17.74 ^, 210,7.85 (111,210, 7.65 (d, J = 1.80 hz, 1H), 7.47-7.28 (m, 3H), 6.59 (d, J = 1.77Hz, 1H), 4.93 (m, lH), 4.21-4.08 (m, 2H), 4.05 (s, 3H), 3.44 -3.35 (m, 2H), 2.76 (s, 3H), 2.35-2.11 (m, 2H), 2.04-1,88 (m, 2H) ppm; 13C NMR (300Mz, CDC13) S168.0,163.8,159.9,147.4 , 138.2,136.7,132.0,131.9, 131.5,129.4,129.0,128.0,126.3,124.6,121.4,119.5,114.5,109.1,56.9,51.4,38.3, 31.8,29.7,28.4ppm; MS (ESI) m / z: [M + H] + = 513.20181.

PATENT

CN 201610630493

PATENT

CN 106831718

str1

Paper

A novel and efficient route for synthesis of Taladegib

Taladegib (LY-2940680), a small molecule Hedgehog signalling pathway inhibitor, was obtained from N-benzyl-4-piperidone via Borch reductive amination, acylation with 4-fluoro-2-(trifluoromethyl)benzoyl chloride, debenzylation, substitution with 1,4-dichlorophthalazine and Suzuki cross-coupling reaction with 1-methyl-1H-pyrazole-5-boronic acid. The advantages of this synthesis route were the elimination of Boc protection and deprotection and the inexpensive starting materials. Furthermore, the debenzylation reaction was achieved with simplified operational procedure using ammonium formate as hydrogen source that provided high reaction yield. This synthetic procedure was suitable for large-scale production of the compound for biological evaluation and further study.

Synthesis of Taladegib (LY-2940680)

purified by flash silica gel chromatography (dichloromethane/MeOH, 30:1) to provide Taladegib as a yellow foam. Yield 0.20 g, 92%; m.p. 95 °C;

1 H NMR (300 MHz, CDCl3 ) δ 8.09 (dd, J = 7.6, 7.7 Hz, 2H), 7.90–7.80 (m, 2H), 7.65 (d, J = 1.8 Hz, 1H), 7.47–7.28 (m, 3H), 6.59 (d, J = 1.8 Hz, 1H), 4.97–4.89 (m, 1H), 4.21–4.08 (m, 2H), 4.05 (s, 3H), 3.44–3.35 (m, 2H), 2.76 (s, 3H), 2.35–2.11(m, 2H), 2.04–1.88 (m, 2H);

13C NMR (75 MHz, CDCl3 ) δ 168.0, 163.8, 159.9, 147.4, 138.2, 136.7, 132.0, 131.9, 131.5, 129.4, 129.0, 128.0, 126.3, 124.6, 121.4, 119.5, 114.5, 109.1, 56.9, 51.4, 38.3, 31.8, 29.7, 28.4; MS calcd for C26H24F4 N6 O [M + H]+: 513.2026; found: 513.2018.

Patent ID

Patent Title

Submitted Date

Granted Date

US2017209574 COMBINATION THERAPIES
2015-10-02
US8273742 DISUBSTITUTED PHTHALAZINE HEDGEHOG PATHWAY ANTAGONISTS
2010-12-23
US2016375142 TARGETED THERAPEUTICS
2016-04-26
US9000023 DISUBSTITUTED PHTHALAZINE HEDGEHOG PATHWAY ANTAGONISTS
2012-08-21
2012-12-13

////////////PHASE 2, Taladegib, LY-2940680,

CN1C(=CC=N1)C2=NN=C(C3=CC=CC=C32)N4CCC(CC4)N(C)C(=O)C5=C(C=C(C=C5)F)C(F)(F)F

NKTR 214


Image result for NKTR 214

CAS  946414-94-4

  • BMS 936558
  • MDX 1106
  • NKTR 214
  • ONO 4538
  • Opdivio
  • NIVOLUMAB

Pegylated engineered interleukin-2 (IL-2) with altered receptor binding

NKTR-214 is a cytokine (investigational agent) that is designed to target CD122, a protein which is found on certain immune cells (known as CD8+ T Cells and Natural Killer Cells) to expand these cells to promote their anti-tumor effects. Nivolumab is a full human monoclonal antibody that binds to a molecule called PD-1 (programmed cell death protein 1) on immune cells and promotes anti-tumor effects.

Protein Sequence

Sequence Length: 1308, 440, 440, 214, 214multichain; modified (modifications unspecified)

NKTR-214 is a CD122-biased cytokine in phase II clinical trials at the M.D. Anderson Cancer Center for the treatment of advanced sarcoma in combination with nivolumab.

 

M.D. Anderson Cancer Center, PHASE 2, SARCOMA

NKTR-214 in combination with OPDIVO® (nivolumab)

RESEARCH FOCUS: Immuno-oncology

DISCOVERED AND WHOLLY OWNED BY NEKTAR

In clinical collaboration withCollaborator

About NKTR-214, Nektar’s Lead Immuno-oncology Candidate

NKTR-214 is a CD122-biased agonist designed to stimulate the patient’s own immune system to fight cancer. NKTR-214 is designed to grow specific cancer-killing T cells and natural killer (NK) cell populations in the body which fight cancer, which are known as endogenous tumor-infiltrating lymphocytes (TILs). NKTR-214 stimulates these cancer-killing immune cells in the body by targeting CD122 specific receptors found on the surface of these immune cells, known as CD8+ effector T cells and Natural Killer (NK) cells. CD122, which is also known as the Interleukin-2 receptor beta subunit, is a key signaling receptor that is known to increase proliferation of these effector T cells.1 In preclinical studies, treatment with NKTR-214 results in a rapid expansion of these cells and mobilization into the tumor micro-environment. NKTR-214 has an antibody-like dosing regimen similar to the existing checkpoint inhibitor class of approved medicines.

In preclinical studies, NKTR-214 demonstrated a mean ratio of 450:1 within the tumor micro-environment of CD8-positive effector T cells, which promote tumor destruction, compared with CD4-positive regulatory T cells, which are a type of cell that can suppress tumor-killing T cells.2Furthermore, a single dose of NKTR-214 resulted in a 500-fold AUC exposure within the tumor compared with an equivalent dose of the existing IL-2 therapy, enabling, for the first time, an antibody-like dosing regimen for a cytokine.2 In dosing studies in non-human primates, there was no evidence of severe side effects such as low blood pressure or vascular leak syndrome with NKTR-214 at predicted clinical therapeutic doses.2 NKTR-214 has a range of potential uses against multiple tumor types, including melanoma (the most serious type of skin cancer), kidney cancer and non-small cell lung cancer (the most common form of lung cancer).

A Phase 1 study evaluating NKTR-214 as a single agent in patients with locally recurrent or metastatic solid tumors including melanoma, renal cell carcinoma (RCC), bladder, colorectal and other solid tumors is ongoing with patient enrollment complete. Results from this Phase 1 trial were presented at the Society for Immunotherapy of Cancer (SITC) 2016 Annual Meeting and showed encouraging evidence of anti-tumor activity, and a favorable safety and tolerability profile. (Poster #387)

In September 2016, Nektar entered into a clinical collaboration with Bristol-Myers Squibb to evaluate NKTR-214 as a potential combination treatment regimen with Opdivo (nivolumab) in five tumor types and eight potential indications. The Phase 1/2 PIVOT clinical trials, known as PIVOT-02 and PIVOT-04 will enroll up to 260 patients and will evaluate the potential for the combination of Opdivo (nivolumab) and NKTR-214 to show improved and sustained efficacy and tolerability above the current standard of care in melanoma, kidney, triple-negative breast cancer, bladder and non-small cell lung cancer patients.

In May 2017, Nektar entered into a research collaboration with Takeda to explore the combination of NKTR-214 with five oncology compounds from Takeda’s cancer portfolio including a SYK-inhibitor and a proteasome inhibitor. The collaboration will explore the anti-cancer activity of NKTR-214 combined with five different targeted mechanisms in preclinical tumor models of lymphoma, melanoma and colorectal cancer to identify which combination treatment regimens show the most promise for possible advancement into the clinic.

Under the terms of the collaboration, the companies will share costs related to the preclinical studies and each will contribute their respective compounds to the research collaboration. Nektar and Takeda will each maintain global commercial rights to their respective drugs and/or drug candidates.

Additional development plans for NKTR-214 include combination studies with additional checkpoint inhibitors, cell therapies and vaccines.

About the Excel NKTR-214 Phase 1/2 Study

The dose-escalation stage of the Excel Phase 1/2 study is designed to evaluate safety, efficacy, and define the recommended Phase 2 dose of NKTR-214 in approximately 20 patients with solid tumors. In addition to a determination of the recommended Phase 2 dose, the study will assess preliminary anti-tumor activity, including objective response rate (ORR). The immunologic effect of NKTR-214 on tumor-infiltrating lymphocytes (TILs) and other immune infiltrating cells in both blood and tumor tissue will also be assessed. Enrollment in the dose escalation study is completed. More information on the Excel Phase 1/2 study can be found on clinicaltrials.gov.

About the PIVOT Phase 1/2 Program: NKTR-214 in combination with OPDIVO® (nivolumab)

The dose escalation stage of the PIVOT program (PIVOT-02 Phase 1/2 study) is underway and will determine the recommended Phase 2 dose of NKTR-214 administered in combination with nivolumab. The study is first evaluating the clinical benefit, safety, and tolerability of combining NKTR-214 with nivolumab in approximately 30 patients with melanoma, renal cell carcinoma or non-small cell lung cancer. Once the recommended Phase 2 dose is achieved, the study will expand into additional patients for each tumor type. The second phase of the expansion cohorts in the PIVOT program (PIVOT-04 Phase 2 study) will evaluate safety and efficacy of the combination in up to 260 patients, in five tumor types and eight indications, including first and second-line melanoma, second-line renal cell carcinoma in immune-oncology therapy (IO) naïve and IO-relapsed patients, second-line non-small cell lung cancer in IO-naïve and IO-relapsed patients, first-line urothelial carcinoma, and second-line triple negative breast cancer. This study is expected to initiate in the second quarter of 2017.

Information on the PIVOT-02 study can be found on clinicaltrials.gov.

Pivot

About the PROPEL Phase 1/2 Program: NKTR-214 in combination with TECENTRIQ® (atezolizumab) or KEYTRUDA®(pembrolizumab)

The dose escalation stage of the PROPEL program will determine the recommended Phase 2 dose of NKTR-214 administered in combination with anti-PD-L1 agent, atezolizumab or anti-PD-1 agent, pembrolizumab. The study will evaluate the clinical benefit, safety and tolerability of combining NKTR-214 with atezolizumab or pembrolizumab and will enroll patients into two separate arms concurrently. The first arm will evaluate an every three-week dose regimen of NKTR-214 in combination with atezolizumab in up to 30 patients in approved treatment settings of atezolizumab, including patients with non-small cell lung cancer or bladder cancer. The second arm will evaluate an every three-week dose regimen of NKTR-214 in combination with pembrolizumab in up to 30 patients in approved treatment settings of pembrolizumab, including patients with melanoma, non-small cell lung cancer or bladder cancer.

Information on the PROPEL study can be found on clinicaltrials.gov.

References

1Boyman, J., et al., Nature Reviews Immunology, 2012, 12, 180-190.

2Charych, D., et al., Clin Can Res; 22(3) February 1, 2016

http://www.nektar.com/application/files/7714/7887/7212/2016_SITC_NKTR-214-clinical_poster.pdf

https://www.google.co.in/patents/WO2015125159A1?cl=en

Inventors Murali Krishna AddepalliDeborah H. CharychSeema KantakSteven Robert Lee
Applicant Nektar Therapeutics (India) Pvt. Ltd.Nektar Therapeutics

////////////946414-94-4, BMS 936558, MDX 1106, NKTR 214, ONO 4538, Opdivio, NIVOLUMAB, PHASE 2

AMISELIMOD


Image result for AMISELIMOD

AMISELIMOD

UNII-358M5150LY; CAS 942399-20-4; 358M5150LY; MT-1303; Amiselimod, MT-1303

Molecular Formula: C19H30F3NO3
Molecular Weight: 377.448 g/mol

2-amino-2-[2-[4-heptoxy-3-(trifluoromethyl)phenyl]ethyl]propane-1,3-diol

Phase II Crohn’s disease; Multiple sclerosis; Plaque psoriasis

Image result for AMISELIMOD

AMISELIMOD HYDROCHLORIDE

  • Molecular FormulaC19H31ClF3NO3
  • Average mass413.902 Da
1,3-Propanediol, 2-amino-2-[2-[4-(heptyloxy)-3-(trifluoromethyl)phenyl]ethyl]-, hydrochloride (1:1)
2-Amino-2-{2-[4-(heptyloxy)-3-(trifluoromethyl)phenyl]ethyl}-1,3-propanediol hydrochloride (1:1)
942398-84-7 [RN]
MT-1303
UNII-AY898D6RU1
2-amino-2-[2-[4-(heptyloxy)-3-(trifluoromethyl)phenyl]ethyl]-1,3-propanediol, monohydrochloride
  • Originator Mitsubishi Tanabe Pharma Corporation
  • Class Propylene glycols; Small molecules
  • Mechanism of Action Immunosuppressants; Sphingosine-1-phosphate receptor antagonist

Highest Development Phases

  • Phase II Crohn’s disease; Multiple sclerosis; Plaque psoriasis
  • Phase I Autoimmune disorders; Inflammation; Systemic lupus erythematosus
  • No development reported Inflammatory bowel diseases

Most Recent Events

  • 04 Nov 2017 No recent reports of development identified for phase-I development in Autoimmune-disorders in Japan (PO, Capsule)
  • 04 Nov 2017 No recent reports of development identified for phase-I development in Autoimmune-disorders in USA (PO, Capsule)
  • 04 Nov 2017 No recent reports of development identified for phase-I development in Inflammation in Japan (PO, Capsule)
  • Image result

Amiselimod, also known as MT1303, is a potent and selective immunosuppressant and sphingosine 1 phosphate receptor modulator. Amiselimod may be potentially useful for treatment of multiple sclerosis; inflammatory diseases; autoimmune diseases; psoriasis and inflammatory bowel diseases. Amiselimod is currently being developed by Mitsubishi Tanabe Pharma Corporation

Mitsubishi Tanabe is developing amiselimod, an oral sphingosine-1-phosphate (S1P) receptor antagonist, for treating autoimmune diseases, primarily multiple sclerosis, psoriasis and inflammatory bowel diseases, including Crohn’s disease.

WO2007069712

EU states expire 2026, and

Expire in the US in June 2030 with US154 extension.

Inventors Masatoshi KiuchiKaoru MarukawaNobutaka KobayashiKunio Sugahara
Applicant Mitsubishi Tanabe Pharma Corporation

In recent years, calcineurin inhibitors such as cyclosporine FK 506 have been used to suppress rejection of patients receiving organ transplantation. While doing it, certain calcineurin inhibitors like cyclosporin can cause harmful side effects such as nephrotoxicity, hepatotoxicity, neurotoxicity, etc. For this reason, in order to suppress rejection reaction in transplant patients, development of drugs with higher safety and higher effectiveness is advanced.

[0003] Patent Documents 1 to 3 are useful as inhibitors of (acute or chronic) rejection in organ or bone marrow transplantation and also useful as therapeutic agents for various autoimmune diseases such as psoriasis and Behcet’s disease and rheumatic diseases 2 aminopropane 1, 3 dioly intermediates are disclosed.

[0004] One of these compounds, 2-amino-2- [2- (4-octylphenel) propane] 1, 3 diol hydrochloride (hereinafter sometimes referred to as FTY 720) is useful for renal transplantation It is currently under clinical development as an inhibitor of rejection reaction. FTY 720 is phosphorylated by sphingosine kinase in vivo in the form of phosphorylated FTY 720 [hereinafter sometimes referred to as FTY 720-P]. For example, 2 amino-2-phosphoryloxymethyl 4- (4-octafil-el) butanol. FTY720 – P has four types of S1 P receptors (hereinafter referred to as S1 P receptors) among five kinds of sphingosine – 1 – phosphate (hereinafter sometimes referred to as S1P) receptors It acts as an aggroove on the body (other than S1P2) (Non-Patent Document 1).

[0005] It has recently been reported that S1P1 among the S1P receptors is essential for the export of mature lymphocytes with thymus and secondary lymphoid tissue forces. FTY720 – P downregulates S1P1 on lymphocytes by acting as S1P1 ghost. As a result, the transfer of mature lymphocytes from the thymus and secondary lymphatic tissues is inhibited, and the circulating adult lymphocytes in the blood are isolated in the secondary lymphatic tissue to exert an immunosuppressive effect Has been suggested (

Non-Patent Document 2).

[0006] On the other hand, conventional 2-aminopropane 1, 3 dioly compounds are concerned as transient bradycardia expression as a side effect, and in order to solve this problem, 2-aminopropane 1, 3 diiori Many new compounds have been reported by geometrically modifying compounds. Among them, as a compound having a substituent on the benzene ring possessed by FTY 720, Patent Document 4 discloses an aminopropenol derivative as a S1P receptor modulator with a phosphate group, Patent Documents 5 and 6 are both S1P Discloses an amino-propanol derivative as a receptor modulator. However, trihaloalkyl groups such as trifluoromethyl groups are not disclosed as substituents on the benzene ring among them. In any case, it is currently the case that it has not yet reached a satisfactory level of safety as a pharmaceutical.

Patent Document 1: International Publication Pamphlet WO 94 Z 08943

Patent Document 2: International Publication Pamphlet WO 96 Z 06068

Patent Document 3: International Publication Pamphlet W 0 98 z 45 429

Patent Document 4: International Publication Pamphlet WO 02 Z 076995

Patent document 5: International public non-fret WO 2004 Z 096752

Patent Document 6: International Publication Pamphlet WO 2004 Z 110979

Non-patent document 1: Science, 2002, 296, 346-349

Non-patent document 2: Nature, 2004, 427, 355-360

Reference Example 3

5 bromo 2 heptyloxybenzonitrile

(3- 1) 5 Synthesis of bromo-2 heptyloxybenzonitrile (Reference Example Compound 3- 1)

1-Heptanol (1.55 g) was dissolved in N, N dimethylformamide (24 ml) and sodium hydride (0.321 g) was added at room temperature. After stirring for 1 hour, 5 bromo-2 fluoborosyl-tolyl (2.43 g) was added and the mixture was further stirred for 50 minutes. The reaction solution was poured into water, extracted with ethyl acetate, washed with water, saturated brine, dried over anhydrous sodium sulfate, and the solvent was distilled off under reduced pressure. After eliminating the 5 bromo 2 fluconate benzonitrile as a raw material, the reaction was carried out again under the same conditions and purification was carried out by silica gel column chromatography (hexane: ethyl acetate = 50: 1 to 5: 1) to obtain the desired product (3.10 g ) As a colorless oil.

– NMR (CDCl 3) δ (ppm): 0.89 (3H, t, J = 6.4 Hz), 1.24-1.35 (6H, m

J = 8.8 Hz), 1.48 (2H, quint, J = 7.2 Hz), 1.84 7.59 (1 H, dd, J = 8.8, 2.4 Hz), 7.65 (1 H, d, J = 2.4 Hz).

Example 1

2 Amino 2- [2- (4-heptyloxy-3 trifluoromethylph enyl) propane-1, 3-diol hydrochloride

(1 – 1) {2, 2 Dimethyl 5- [2- (4 hydroxy 3 trifluoromethylfuethyl) ethyl] 1,3 dioxane 5 mercaptothenylboronic acid t butyl ester (synthesis compound 1 1)

Reference Example Compound 2-5 (70.3 g) was dissolved in tetrahydrofuran (500 ml), t-butoxycallium (13.Og) was added, and the mixture was stirred for 1 hour. To the mixed solution was dropwise added a solution of the compound of Reference Example 1 (15.Og) in tetrahydrofuran (100 ml) under ice cooling, followed by stirring for 2 hours under ice cooling. Water was added to the reaction solution, the mixture was extracted with ethyl acetate, washed with water, saturated brine, dried with anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by silica gel column chromatography (hexane: ethyl acetate = 3: D to obtain 31. Og of a pale yellow oily matter.) The geometric isomer ratio of the obtained product was (E : Z = 1: 6).

This pale yellow oil was dissolved in ethyl acetate (200 ml), 10% palladium carbon (3.00 g) was added, and the mixture was stirred under a hydrogen atmosphere at room temperature for 7 hours. After purging the inside of the reaction vessel with nitrogen, the solution was filtered and the filtrate was concentrated. The residue was washed with diisopropyl ether to obtain the desired product (2.2 g) as a colorless powder.

1 H-NMR (CDCl 3) δ (ppm): 1. 43 (3H, s), 1.44 (3H, s), 1. 47 (9H, s), 1

(2H, m), 91- 1. 98 (2H, m), 2. 50-2.66 (2H, m), 3. 69 (2H, d, J = Il. 6 Hz), 3. 89 J = 8.2 Hz), 7. 22 (1 H, dd J = 8 Hz), 5. 02 (1 H, brs), 5. 52 . 2, 1. 7 Hz), 7. 29 (1 H, d, J = l. 7 Hz).

(1-2) {2,2 Dimethyl-5- [2- (4heptyloxy-3 trifluoromethyl) ethyl] 1,3 dioxane 5-mercaptobutyric acid t-butyl ester Synthesis (compound 1 2)

Compound 1-1 (510 mg) was dissolved in N, N dimethylformamide (10 ml), potassium carbonate (506 mg) and n-heptyl bromide (0.235 ml) were added and stirred at 80 ° C. for 2 hours. Water was added to the reaction solution, the mixture was extracted with ethyl acetate, washed with water and saturated brine, dried with anhydrous sulfuric acid

The resultant was dried with GENSCHUM and the solvent was distilled off under reduced pressure to obtain the desired product (640 mg) as a colorless oil.

– NMR (CDCl 3) δ (ppm): 0.89 (3H, t, J = 6.8 Hz), l.30-1.37 (6H, m

(2H, m), 1.91-1.98 (2H, m), 1.42-1.50 (2H, m), 1.42 (3H, s), 1.44 (3H, s), 1.47 J = 16.6 Hz), 4.00 (2H, t, J = 6.4 Hz), 4.9 8 (2H, d, J = 11.6 Hz), 3.69 1 H, brs), 6.88 (1 H, d, J = 8.5 Hz), 7.26 – 7.29 (1 H, m), 7.35 (1 H, d, J = 1.5 Hz).

(1-3) Synthesis of 2-amino-2- [2- (4heptyloxy 3 trifluoromethyl) ethyl] propane 1, 3 diol hydrochloride (Compound 1- 3)

Compound 12 (640 mg) was dissolved in ethanol (15 ml), concentrated hydrochloric acid (3 ml) was caught and stirred at 80 ° C. for 2 hours. The reaction solution was concentrated, and the residue was washed with ethyl ether to give the desired product (492 mg) as a white powder.

MS (ESI) m / z: 378 [M + H]

– NMR (DMSO-d) δ (ppm): 0.86 (3H,

6 t, J = 6.8 Hz), 1.24 – 1.39 (6

(4H, m), 3.51 (4H, d, J = 5. lHz), 4.06 (2H, m), 1.39-1.46 (2H, m), 1.68-1.78 (4H, m), 2.55-2.22 , 7.32 (2H, t, J = 5.1 Hz), 7.18 (1 H, d, J = 8.4 Hz), 7.42 – 7.45 (2 H, m), 7.76 (3 H, brs;).

PATENT

WO 2009119858

JP 2011136905

WO 2017188357

PATENT

WO-2018021517

Patent Document 1 discloses 2-amino-2- [2- (4-heptyloxy-3-trifluoromethylphenyl) ethyl] propane- 1,3 which is useful as a medicine excellent in immunosuppressive action, rejection- – diol hydrochloride is disclosed.
The production method includes the step of reducing 4-heptyloxy-3-trifluoromethylbenzoic acid (Ia) to 4-heptyloxy-3-trifluoromethylbenzyl alcohol (IIa). However, until now, there has been a problem such that the conversion is low and the by-product (IIa ‘) in which the trifluoromethyl group is reduced together with the compound (IIa) is generated in this step.
[Chemical formula 1]
 In particular, since a series of analogous substances derived from by-products (IIa ‘) are difficult to be removed in a later process, it is necessary to suppress strict production thereof in the manufacture of drug substances requiring high quality there were.

Patent Document 1: WO2007 / 069712

[Chemical formula 3]

(2-amino-2- [2- (4-heptyloxy-3-trifluoromethylphenyl) ethyl] propane- 1,3-diol hydrochloride) From
the compound (IIa), the following scheme Based on the route, 2-amino-2- [2- (4-heptyloxy-3-trifluoromethylphenyl) ethyl] propane-1,3-diol hydrochloride was prepared.
[Chemical Formula 9]

STR1
Example 2
Synthesis of 4-heptyloxy-3-trifluoromethylbenzyl chloride (Step A) A
few drops of N, N-dimethylformamide was added to a solution of compound (IIa) (26.8 g) in methylene chloride (107 mL), and 0 At 0 ° C., thionyl chloride (8.09 mL) was added dropwise. The mixture was stirred at the same temperature for 2 hours, and water (50 mL) was added to the reaction solution. The organic layer was separated and extracted, washed with water (50 mL), saturated aqueous sodium bicarbonate solution (70 mL), dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure to give 4-heptyloxy-3-trifluoromethylbenzyl Chloride (28.3 g) as white crystals.
1H-NMR (CDCl 3) δ (ppm): 0.89 (3H, t, J = 6.5 Hz), 1.26-1.54 (8H, m), 1.77-1.86 (2H, m , 4.49 (2H, t, J = 6.4 Hz), 4.56 (2H, s), 6.96 (IH, d, J = 8.6 Hz), 7.49 (IH, dd, J = 2.0 Hz, 8.5 Hz), 7.58 (1 H, d, J = 1.9 Hz)
Example 3
Synthesis of dimethyl (4-heptyloxy-3-trifluoromethylbenzyl) phosphonate (Step B) To
a solution of N, N (3-trifluoromethylbenzyl ) phosphonate of 4-heptyloxy-3-trifluoromethylbenzyl chloride (6.00 g, 19.4 mmol) (2.57 g, 23.3 mmol), cesium carbonate (7.60 g, 23.3 mmol) and tetrabutylammonium iodide (7.54 g, 20.4 mmol) were added to a dimethylformamide (36 mL) And the mixture was stirred at 25 ° C. for 1 day. Toluene (36 mL) and water (18 mL) were added for phase separation, and the resulting organic layer was washed twice with a mixture of N, N-dimethylformamide (18 mL) and water (18 mL). After concentration under reduced pressure, column purification using hexane and ethyl acetate gave 4.71 g of dimethyl (4-heptyloxy-3-trifluoromethylbenzyl) phosphonate.
1
H-NMR (CDCl 3) δ (ppm): 0.89 (3 H, t, J = 6.9 Hz), 1.20 – 1.41 (6 H, m) , 1.43-1.49 (2H, m), 1.72-1.83 (2H, m), 3.09 (IH, s), 3.14 (IH, s), 3.68 (3H , 7.41 – 7.44 (2 H, t, J = 6.4 Hz), 6.94 (1 H, d, J = 8.4 Hz), 3.70 (3 H, s), 4.02 (2H, m)
Example 4
tert-Butyl (E) – {2,2-dimethyl-5- [2- (4-heptyloxy-3-trifluoromethylphenyl) vinyl] -1, 3-dioxan-5- yl} carbamate Ester synthesis (Step C) A
solution of dimethyl (1.18 g, 3.09 mmol ) (4-heptyloxy-3-trifluoromethylbenzyl) phosphonate in 1.25 mL of N, N- dimethylformamide and (2, -dimethyl-5-formyl-1,3-dioxan-5-yl) carbamic acid tert-butyl ester (961 mg, 3.71 mmol) in tetrahydrofuran (4 mL) was treated with potassium tert-butoxide (1.28 g, 4 mmol) in tetrahydrofuran (7 mL), and the mixture was stirred at 0 ° C. for 6 hours. Heptane (7 mL) and water (3 mL) were added and the layers were separated, and the obtained organic layer was washed twice with water (3 mL) and concentrated. Heptane was added and the mixture was cooled in an ice bath. The precipitated crystals were collected by filtration and dried under reduced pressure to give (E) – {2,2-dimethyl-5- [2- (4-heptyloxy- Phenyl) vinyl] -1, 3-dioxan-5-yl} carbamic acid tert-butyl ester.
1
H-NMR (CDCl 3) δ (ppm): 0.89 (3 H, t, J = 6.9 Hz), 1.29 – 1.38 (6 H, m) , 1.44 – 1.59 (17 H, m), 1.77 – 1.83 (2 H, m), 3.83 – 3.93 (2 H, m), 3.93 – 4.08 (4 H, J = 16.5 Hz), 6.48 (1 H, d, J = 16.5 Hz), 6.91 (1 H, d, J), 5.21 (1 H, brs), 6.10 J = 8.5 Hz), 7.44 (1 H, dd, J = 8.6, 2.1 Hz), 7.55 (1 H, d, J = 2.0 Hz)
Example 5
Synthesis of 2-amino-2- [2- (4-heptyloxy-3-trifluoromethylphenyl) ethyl] propane-1,3-diol hydrochloride (Step D)
(E) – {2, -dimethyl-5- [2- (4-heptyloxy-3-trifluoromethylphenyl) vinyl] -1,3-dioxan- 5-yl} carbamic acid tert-butyl ester (6.50 g, 12.6 mmol) Methanol (65 mL) solution was heated to 50 ° C., a solution of concentrated hydrochloric acid (2.55 g) in methanol (5.3 mL) was added dropwise, and the mixture was stirred at 60 ° C. for 6 hours. The mixture was cooled to around room temperature, 5% palladium carbon (0.33 g) was added thereto, and the mixture was stirred under a hydrogen gas atmosphere for 3 hours. After filtration and washing the residue with methanol (39 mL), the filtrate was concentrated and stirred at 5 ° C. for 1 hour. Water (32.5 mL) was added and the mixture was stirred at 5 ° C for 1 hour, and the precipitated crystals were collected by filtration. Washed with water (13 mL) and dried under reduced pressure to obtain 4.83 g of 2-amino-2- [2- (4-heptyloxy-3-trifluoromethylphenyl) ethyl] propane-1,3-diol hydrochloride .
MS (ESI) m / z: 378 [M + H]

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PATENTS

Patent ID

Patent Title

Submitted Date

Granted Date

US2017029378 KINASE INHIBITOR
2016-10-12
US2014296183 AMINE COMPOUND AND USE THEREOF FOR MEDICAL PURPOSES
2014-06-17
2014-10-02
Patent ID

Patent Title

Submitted Date

Granted Date

US2017253563 KINASE INHIBITORS
2017-05-24
US9499486 Kinase inhibitor
2015-10-01
2016-11-22
US9751837 KINASE INHIBITORS
2015-10-01
2016-04-14
US8809304 Amine Compound and Use Thereof for Medical Purposes
2009-05-28
US2017209445 KINASE INHIBITORS
2015-10-01

////////////AMISELIMOD, Phase II, Crohn’s disease, Multiple sclerosis, Plaque psoriasis,  MT-1303,  MT1303,  MT 1303, Mitsubishi Tanabe Pharma Corporation, Mitsubishi , JAPAN, PHASE 2

CCCCCCCOC1=C(C=C(C=C1)CCC(CO)(CO)N)C(F)(F)F

GSK2248761A , IDX899, Fosdevirine


Image result for GSK2248761A , IDX899, FosdevirineChemSpider 2D Image | fosdevirine | C20H17ClN3O3P

GSK2248761A , IDX899, Fosdevirine,

Fosdevirine; IDX899; IDX-899; GSK2248761; cas 1018450-26-4; GSK-2248761, IDX 12899

1018450-26-4 CAS
R FORM ROTATION (-)
Molecular Formula: C20H17ClN3O3P
Molecular Weight: 413.798 g/mol
 Phosphinic acid, P-[2-(aminocarbonyl)-5-chloro-1H-indol-3-yl]-P-[3-[(1E)-2-cyanoethenyl]-5-methylphenyl]-, methyl ester, [P(R)]-
Methyl (R)-(2-carbamoyl-5-chloro-1H-indol-3-yl){3-[(E)-2-cyanovinyl]-5-methylphenyl}phosphinate
Phosphinic acid, P-[2-(aminocarbonyl)-5-chloro-1H-indol-3-yl]-P-[3-[(E)-2-cyanoethenyl]-5-methylphenyl]-, methyl ester, (R)-
5DV
Methyl (R)-(2-carbamoyl-5-chloro-1H-indol-3-yl)[3-(2-cyanoethyl)-5-methylphenyl]phosphinate

[R(P)]-(2-Carbamoyl-5-chloro-1H-indol-3-yl)[3-(2-cyanovinyl)-5-methylphenyl]phosphinic acid methyl ester

Phase II clinical trials for the treatment of HIV infection

Idenix (Originator)

Fosdevirine, also known as GSK2248761 and IDX899, a Highly Potent Anti-HIV Non-nucleoside Reverse Transcriptase Inhibitor having an EC50 of 11 nM against the Y181C/K103N double mutant. GSK2248761 is a novel, once-daily (QD), next-generation nonnucleoside reverse transcriptase inhibitor (NNRTI) with activity against efavirenz-resistant strains. GSK2248761 at 100 to 800 mg QD for 7 days was well tolerated, demonstrated potent antiviral activity in treatment-naive HIV-infected subjects, and had favorable PK and resistance profiles. GSK2248761 is no longer in clinical development.

IDX-12899 is a non-nucleoside reverse transcriptase inhibitors (NNRTI) originated by Idenix (acquired by Merck & Co.). It had been in phase II clinical trials for the treatment of HIV infection. However, in 2010, the compound was placed on clinical hold by the FDA. In 2009, the compound was licensed by Idenix to GlaxoSmithKline for the treatment of HIV infection on a worldwide basis.

PATENT

WO2008/042240 A2, 2008, Compound III

compound 66a: racemic form

5-chloro-3-[ methyl 3-((Zζ)-2-cyanovinyl)-5-methylphenyl] phosphinoyl-l//-indole-2- carboxamide.

Figure imgf000091_0003

[00258] Compound 66a was synthesized according to method AL. White solid, 1H NMR (CDCl3, 300 MHz) δ 2.40 (s, 3H), 3.88 (d, J= 11.7 Hz, 3H), 5.89 (d, J= 16.5 Hz, IH), 5.97 (brs, IH), 7.33-7.67 (m, 7H), 10.46 (s, IH), 10.89 (brs, IH), 31P NMR (CDCl3, 121.49 MHz) δ 31.54. MS (ES+) m/z = 414 (MH+).

Example 8

Figure imgf000126_0001

Preparation of Compound HI

Figure imgf000127_0001
Figure imgf000127_0002

305

1 (-)cιnchonιdιne, Acetone

2 1N HCI1 EtOAc

Figure imgf000127_0003

Compound 302

[00348] A suitable reactor was charged Compound 301 (10Og, 0.23mol) and tetrahydrofuran (IL). The resulting solution was chilled between -90° to -100°C under nitrogen using a LN2 / IPA slush bath, then was treated with n-butyl lithium (2.5M in Hexanes, 99ml, 0.25mol) added over 10 minutes. To this was added diethyl chlorophosphite (37.1g, 0.24mol) over 10 minutes. HPLC (Method 001, RT = 18.9 min) showed no starting material and ca. 85% product. The reaction was then diluted with ethyl acetate (IL) and was allowed to warm to -4O0C. The mix was then treated with hydrochloric acid (0.5M, 590ml) and was allowed to warm to ambient temperature and stir for 30 minutes. The resulting layers were separated and the aqueous extracted with ethyl acetate (500ml). The organics were combined and washed with brine (500ml) dried over sodium sulfate, filtered and concentrated to an oil. 88% HPLC AUC (Method 20, RT = 5.8 min) 115g, >100% yield due to impurities and solvent. Used as is in the next step. Compound 303

[00349] A suitable reactor was charged with Compound 302 (111 g, estimated 0.18mol), iodocinnamonitrile (47.1g, 0.175mol), triethylamine (29.3ml, 0.21mol) and toluene (800ml). The resulting mix was degassed by sparging with a stream of nitrogen for 10 minutes at ambient temperature, after which time tetrakis(triphenylphosphine) palladium(O) (10. Ig, 0.0088mol) was added. The mix was sparged for an additional 5 minutes, then was heated to 80°C for 2 hours. HPLC (Method 20, RT = 6.5 min) showed a complete reaction. The mix was cooled to ambient and was filtered through celite and washed with ethyl acetate (400ml). The combined organics were washed with brine (2 x 500ml) then dried over sodium sulfate, filtered and concentrated to a volume of 350ml. The concentrate was cooled to O0C and was stirred for 1 hour, during which time the product crystallized. The solids were filtered and washed with hexane:toluene (2:1, 150ml). Dried to leave 95g, 90% yield, HPLC AUC 98% (Method 20). Used as is in the next reaction. [00350] 303: C29H26ClN2O6PS 597.02gmol‘ m/z (ESI+): 597.0 (MH+, 100%), 599.0 (MH+, 35%) 1H NMR δH (400 MHz, CDCl3): 1.38, 1.48 (2 x 3H, 2 x t, COOCH2CH3, POOCH2CH3), 2.41 (3Η, s, Ar-CH3), 4.09-4.16 (2Η, m, POOCH2CH3), 4.52 (2H, q, COOCH2CH3), 5.93 (IH, d, CH=CHCN), 7.33-7.38 (3Η, m, CH=CHCN, 2 x Ar-H), 7.52 (2Η, t, 2 x Ar-H), 7.64 (1Η, t, Ar-H), 7.74, 7.77 (2 x 1Η, 2 x d, 2 x Ar-H), 7.85 (1Η, d, Ar- H), 7.94 (1Η, dd, Ar-H), 8.08 (2Η, d, 2 x Ar-H) 1H NMR δH (400 MHz, d6-DMSO): 1.26, 1.33 (2 x 3H, 2 x t, COOCH2CH3, POOCH2CH3), 2.34 (3Η, s, Ar-CH3), 3.95-4.10 (2Η, m, POOCH2CH3), 4.40 (2H, q, COOCH2CH3), 6.52 (IH, d, CH=CHCN), 7.52 (1Η, dd, Ar-H), 7.60-7.84 (8Η, m, CH=CHCN, 7 x Ar-H), 8.07 (3 x 1Η, m, 3 x Ar-H)

Compound 304

[003511 A suitable reactor was charged with Compound 303 (537g, 0.90mol) and methylene chloride (2.0L). The resulting solution was cooled to O0C, and was treated with bromotrimethylsilane (45Og, 2.9mol) added over 15 minutes. The reaction was then warmed to 400C for 1.5 hours. ΗPLC (Method 20, RT = 4.4 min) indicated a complete reaction. The excess TMSBr was stripped under vacuum (40 – 45°C) and the resulting sticky solid was resuspended in DCM (2.5L) and chilled to 00C. Oxalyl chloride (156ml, 1.8mol) was added over 15 minutes, followed by N,N-dimethylformamide (13.7ml, 0.18mol) both added at O0C. Gas evolution was observed during the DMF addition. After 1 hour, ΗPLC (Method 20, RT = 6.2 min, sample quenched with anhydrous methanol prior to injection) showed a complete reaction. The solvents were stripped again to remove residual oxalyl chloride and the mix resuspended in chilled methanol (3.0L) at 0° – 5°C, and then was allowed to warm to ambient. After two hours, HPLC indicated a complete reaction (HPLC Method 20, RT = 6.2 min). The solution was concentrated to a volume of 1.5L, and the resulting thin slurry was cooled to 0°C, and was diluted with an aqueous solution of sodium bicarbonate (126g, 3L water). After 2 hours at 50C, the product was filtered and washed with cold water / methanol (2:1, 1.5L) then dried to leave 50Og Compound 304. HPLC (Method 20) purity 92% used as is.

Compound 305

[00352] A suitable reactor was charged with Compound 304 (ca. 28Og, 0.48mol) and tetrahydrofuran (2.8L). The resulting solution was then cooled to 5°C and was treated with lithium hydroxide monohydrate (45g, 1.07mol) added in one portion. The reaction was allowed to warm to ambient, during which time the color lightened and a white precipitate formed. After overnight stirring, HPLC indicated an incomplete reaction (Method 20, product RT = 4.3, partially deprotected RT = 5.1, major impurity RT = 3.8). An additional 10% LiOH-H2O was added, but after 10 hours, the partially deprotected intermediate remained at 5%, and the impurity peak at 3.8 minutes had increased to ca. 25%. The reaction was cooled to 50C and was acidified with hydrochloric acid (5N, 280ml) then was diluted with ethyl acetate (2L). The layers were separated and the aqueous extracted with ethyl acetate (500ml). The combined organics were washed with brine (IL) and dried with sodium sulfate, then concentrated to leave a crude oily solid, Compound 305. Ca. 300g, HPLC AUC 57%.

[00353] The crude product was taken up in acetonitrile (1.2L) at 4O0C, and the product triturated w/ water (1.2L). The resulting slurry was cooled to 50C and was allowed to granulate for 30 minutes, after which time the product was filtered and washed with ACN:H2O (1 :1, 100 ml). Ca. 103g, 88% by HPLC. The product was then recrystallized from 360ml ACN at 400C and 360ml water as before. Filtered, washed and dried to leave 75g Compound 305. HPLC AUC 97%. Used as is in the next step.

Compound 306 (chiral resolution)

[00354] A suitable reactor was charged with Compound 305 (28Og, 0.66mol) and acetone (4.2L). The resulting thin slurry was then treated with (-)-cinchonidine (199g, 0.66mol) added in one portion. After one hour, a solution had formed, and after an additional hour, a white solid precipitated, and the mix was left to stir for an additional two hours (four hours total) after which time the solids were filtered, washed with acetone (200ml) and dried to leave 199g Crude Compound 306 cinchonidine salt. HPLC showed an isomer ratio of 96:4.

[00355] The crude salt was then slurried in ethyl acetate (3L) and hydrochloric acid (IN, 3L). The two phase solution was vigorously stirred for 2 hours at ambient temperature. The layers were separated, and the aqueous extracted with ethyl acetate (3L). The organics were combined, dried with sodium sulfate, and concentrated to leave the free base Compound 306, 107g, 95:5 by chiral HPLC.

[00356] The crude Compound 306 was then suspended in acetone (1.07L) and treated with (-)-cinchonidine (76g, 0.26 mol.) After 4 hours total stir time (as above) the solids were filtered, washed with acetone (200ml) and dried to leave 199g of the salt. HPLC 98.6:1.4.

[00357] The salt was broken by dissolving in ethyl acetate (3L) and hydrochloric acid (IN, 3L). The two phase solution was stirred for 2 hours at ambient temperature. The layers were separated, and the aqueous extracted with ethyl acetate (2L). The organics were combined, dried with sodium sulfate, and concentrated to leave the free base Compound 306, 98g, 98.6:1.4 by chiral HPLC. 70% recovery of the desired isomer, 35% yield from the racemic Compound 306. #6: C20H16ClN2O4P 414.78gmol‘ m/z (ESI+): 415.1 (MH+, 100%), 417.0 (MH+, 35%) [α]D 25 : -47.51 (c, 10.66mgml‘ in EtOAc) [Opposite enantiomer [α]D 25 : +47.26 (c, 9.60mgml‘ in EtOAc)] 1H NMR δH (400 MHz, d6-DMSO): 2.33 (3 H, s, Ar-CH3), 3.71 (3H, d, CH3OP), 6.50 (1Η, d, CH=CHCN), 7.36 (1Η, dd, H-6), 7.57 (1Η, d, H-I), 7.66-7.71 (2Η, m, H-4, Ar-Hortho), 7.67 (1Η, d, CH=CHCN), 7.84 (IH, d, Ar-Hortho), 7.98 (1Η, s, Ar-Hpara), 12.97 (1Η, s, N-H), 14.38 (1Η, br-s, COOH) Multiple δc values indicate splitting of carbon signal due to P. 13C NMR δc (100 MHz, d6-DMSO): 20.68 (Ar-CH3), 51.70 (CH3OP), 98.15 (CH=CHCN), 102.33, 103.85, 1 14.98, 120.91 (3 x Q, 118.47 (CN), 125.39 (C), 126.78 (Q, 127.74, 127.86 (C- Hortho), 129.78, 129.88 (Q, 131.25 (Q, 132.06 (Q, 133.44, 133.55 (Q, 133.89, 134.05 (Q, 134.62, 134.75 (Q, 135.47, 135.66 (Q, 138.78, 138.91 (Q, 149.62 (CH=CHCN), 160.40 (C=O) 31P NMR δP (162 MHz, d6-DMSO): 33.50 (IP, s)

Compound HI

[00358] A suitable reactor was charged with Compound 306 (0.63g, O.OOHmol) and 1 ,2-dimethoxyethane (10ml.) The mix was treated with 1,1-carbonyldiimidazole (0.47g, 0.0028mol) added in one portion, and the mix was allowed to stir at ambient temperature until gas evolution ceased (ca. 1.5 hours.) The solution was then cooled to 50C, and was sparged with ammonia gas for 5 minutes. HPLC (Method 20, product RT=5.0 min) showed a complete reaction after one hour at ambient. The reaction was quenched by the addition of 1Og crushed ice, and was concentrated under reduced pressure to remove the DME. The resulting slurry was stirred for one hour at 50C to granulate the product. The solids were filtered and dried to leave pure Compound III ((2-Carbamoyl-5-chloro-4-fluoro-lH-indol-3- yl)-[3-((E)-2-cyano-vinyl)-5-methyl-phenyl]-(S)-phosphinic acid methyl ester) as a white solid 0.56g, 89% yield. HPLC (Method 20) chemical purity 98.5%. Chiral purity 97%. [00359] A suitable reactor was charged with Compound 306 (1Og, 0.024mol) and 1,2- dimethoxyethane (150ml). The mix was treated with 1,1-carbonyldiimidazole (7.8g, 0.048mol) added in one portion, and the mix was allowed to stir at ambient temperature until gas evolution ceased. The solution was then cooled to 5°C, and was sparged with ammonia gas for 5 minutes. HPLC (Method 20, product RT=5.0 min) showed a complete reaction after one hour. The reaction was quenched by the addition of lOOg crushed ice, and was concentrated under reduced pressure to remove the DME. The resulting oily solid (in water) was diluted with methanol (20ml) and stirred for one hour at 50C to granulate the product. The solids were filtered and dried to leave pure Compound III ((2-Carbamoyl-5- chloro-4-fluoro-lH-indol-3-yl)-[3-((E)-2-cyano-vinyl)-5-methyl-phenyI]-(S)-phosphinic acid methyl ester). 9.8g, 98% yield. HPLC (Method 20) chemical purity 99.5%. Chiral purity 94.3%.

Compound III: C20Hi7ClN3O3P 413.79gmol‘ m/z (ESI+): 414.1 (MH+, 100%), 416.1 (MH+, 35%)

vmax (KBr disc) (cm“1) 1620.0 (amide I), 1670.6 (amide II), 2218.7 (CN), 3125.5, 3291.9 (N-H)

[α]D 20 : -75.08 (c, 9.04mgmr’ in CHCl3)

m.p.: 144- 1480C transition to opaque semi-solid, 209-2100C melts

Elemental analysis: C20H17ClN3O3P calculated C 58.05%, H 4.14%, N 10.15%, Cl 8.57%, P 7.49%. Found C 58.13%, H 4.08%, N 10.16%, Cl 8.69%, P 7.44% 

1H NMR δH (400 MHz, d6-DMSO): 2.32 (3H, s, Ar-CH3), 3.74 (3Η, d, CH3OP), 6.52 (1Η, d, CH=CHCN), 7.30 (1Η, dd, H-6), 7.53-7.58 (3Η, m, H-4, H-7, H-6′), 7.68 (1Η, d, CH=CHCN), 7.73 (IH, s, H-4′), 7.75 (1Η, d, H-2′), 8.02, 10.15 (2 x 1Η, 2 x s, NH2), 12.80 (1Η, s, N-H) Multiple δc values indicate splitting of carbon signal due to P. 

13C NMR δc(100 MHz, d6-DMSO): 20.77 (Ar-CH3), 51.75, 51.81 (CH3OP), 98.39, 98.91 (C-3), 98.44 (CH=CHCN), 1 15.05 (C-7), 1 18.53 (CN), 119.96 (C-4), 124.73 (C-6), 126.68 (C-5), 127.15, 127.26 (C-2′), 129.25, 129.35 (C-9), 131.37 (C-4′), 132.45, 134.04 (C-I ‘), 132.69, 132.80 (C-6′), 133.92 (C-8), 134.30, 134.44 (C-3′), 139.33, 139.46 (C-5’), 139.96, 140.17 (C-2), 149.55 (CH=CHCN), 160.65 (C=O)

 31P NMR δP (162 MHz, d6-DMSO): 33.72 (IP, s)

PATENT

http://www.google.ch/patents/WO2009120914A1?cl=en&hl=de

Figure imgf000003_0001

(2-carbamoyl-5-chloro-lH-indol-3-yl)-[3-((E)-2-cyano-vinyl)-5-methyl-phenyl]- (7?)-phosphinic acid methyl ester (I):

WO2008042240A2 * 28. Sept. 2007 10. Apr. 2008 Idenix Pharmaceuticals, Inc. Enantiomerically pure phosphoindoles as hiv inhibitors
US20060074054 * 16. Sept. 2005 6. Apr. 2006 Richard Storer Phospho-indoles as HIV inhibitors

Figure 7 provides an infrared spectrum of Form I.

Paper

Development of an Efficient Manufacturing Process to GSK2248761A API

 GlaxoSmithKline, Medicines Research Center, Gunnels Wood Road, Stevenage, Hertfordshire, SG1 2NY, U.K.
 Merck & Co. Inc., 126 East Lincoln Avenue, Rahway, New Jersey 07065, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.7b00357
Abstract Image

Amidation of indole 2-carboxylate 1 with ammonia gas via the imidazolide 2 gave GSK2248761A API 3, which was in development for the treatment of HIV. Three significant impurities, namely the phosphinic acid 4, the N-acyl urea 8, and the indoloyl carboxamide 6, were formed during the reaction, and the original process was unable to produce API within clinical specification when run at scale. Investigation into the origin, fate, and control of these impurities led to a new process which was able to produce API within clinical specification.

1H NMR (500 MHz, CDCl3) δ ppm 2.37 (s, 3H), 3.86 (d, J = 15.0 Hz, 3H), 5.86 (d, J = 15.0 Hz, 1H), 5.94 (s, 1H), 7.33 (dd, J = 9.0 Hz, J = 2.0 Hz, 1H), 7.34 (d, J = 15.5 Hz, 1H), 7.39 (s, 1H), 7.49 (dd, J = 9.0 Hz, J = 1.5 Hz, 1H) 7.60 (d, J = 13.5 Hz, 1H), 7.64 (d, J = 13.5 Hz, 1H), 7.65 (d, J = 1.5 Hz, 1H), 10.40 (s, 1H), 10.88 (s, 1H); 
13C NMR (126 MHz, CDCl3) δ 21.3, 52.1, 98.1, 100.5 (d, J = 152.5 Hz), 113.9, 117.6, 120.9, 126.2, 126.5 (d, J = 11.3 Hz) 128.7, 129.9 (d, J = 10.1 Hz), 131.7, 133.0 (d, J = 151.2 Hz), 133.2 (d, J = 8.8 Hz), 133.4 (d, J = 10.1 Hz), 134.1 (d, J= 15.1 Hz), 138.7, 139.9, 149.2 and 161.2;
 31P NMR (202 MHz, CDCl3) δ 31.4.
IR ν (cm–1) 3280, 3065, 1679, 1619, 1402, 1195 and 1010.
HRMS calcd for C20H18ClN3O3P: 414.0769; HRMS found [M + H]+: 414.0760.
PAPER

Development and Scale-Up of a Manufacturing Route for the Non-nucleoside Reverse Transcriptase Inhibitor GSK2248761A (IDX-899): Synthesis of an Advanced Key Chiral Intermediate

 GlaxoSmithKline, Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire SG1 2NY, U.K.
 Merck & Co., Inc.,126 East Lincoln Avenue, Rahway, New Jersey 07065, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.7b00356

Abstract

Abstract Image

A new and improved synthetic route to an intermediate in the synthesis of the phosphinate ester GSK2248761A is described. In the key step, we describe the first process-scale example of a palladium-catalyzed phosphorus–carbon coupling to give the entire backbone of GSK2248761A in one telescoped stage in 65% average yield on a 68 kg scale. This unusual chemistry enabled the route to be reduced from six chemistry stages to four and eliminated a number of environmentally unfriendly reagents and solvents.

REFERENCES

1: Dousson C, Alexandre FR, Amador A, Bonaric S, Bot S, Caillet C, Convard T, da Costa D, Lioure MP, Roland A, Rosinovsky E, Maldonado S, Parsy C, Trochet C, Storer R, Stewart A, Wang J, Mayes BA, Musiu C, Poddesu B, Vargiu L, Liuzzi M, Moussa A, Jakubik J, Hubbard L, Seifer M, Standring D. Discovery of the Aryl-phospho-indole IDX899, a Highly Potent Anti-HIV Non-nucleoside Reverse Transcriptase Inhibitor. J Med Chem. 2016 Feb 3. [Epub ahead of print] PubMed PMID: 26804933.

2: Margolis DA, Eron JJ, DeJesus E, White S, Wannamaker P, Stancil B, Johnson M. Unexpected finding of delayed-onset seizures in HIV-positive, treatment-experienced subjects in the Phase IIb evaluation of fosdevirine (GSK2248761). Antivir Ther. 2014;19(1):69-78. doi: 10.3851/IMP2689. Epub 2013 Oct 24. PubMed PMID: 24158593.

3: Ölgen S. Recent development of new substituted indole and azaindole derivatives as anti-HIV agents. Mini Rev Med Chem. 2013 Oct;13(12):1700-8. Review. PubMed PMID: 23895189.

4: Castellino S, Groseclose MR, Sigafoos J, Wagner D, de Serres M, Polli JW, Romach E, Myer J, Hamilton B. Central nervous system disposition and metabolism of Fosdevirine (GSK2248761), a non-nucleoside reverse transcriptase inhibitor: an LC-MS and Matrix-assisted laser desorption/ionization imaging MS investigation into central nervous system toxicity. Chem Res Toxicol. 2013 Feb 18;26(2):241-51. doi: 10.1021/tx3004196. Epub 2012 Dec 20. PubMed PMID: 23227887.

5: Zala C, St Clair M, Dudas K, Kim J, Lou Y, White S, Piscitelli S, Dumont E, Pietropaolo K, Zhou XJ, Mayers D. Safety and efficacy of GSK2248761, a next-generation nonnucleoside reverse transcriptase inhibitor, in treatment-naive HIV-1-infected subjects. Antimicrob Agents Chemother. 2012 May;56(5):2570-5. doi: 10.1128/AAC.05597-11. Epub 2012 Feb 6. PubMed PMID: 22314532; PubMed Central PMCID: PMC3346662.

6: Piscitelli S, Kim J, Gould E, Lou Y, White S, de Serres M, Johnson M, Zhou XJ, Pietropaolo K, Mayers D. Drug interaction profile for GSK2248761, a next generation non-nucleoside reverse transcriptase inhibitor. Br J Clin Pharmacol. 2012 Aug;74(2):336-45. doi: 10.1111/j.1365-2125.2012.04194.x. PubMed PMID: 22288567; PubMed Central PMCID: PMC3630753.

7: La Regina G, Coluccia A, Silvestri R. Looking for an active conformation of the future HIV type-1 non-nucleoside reverse transcriptase inhibitors. Antivir Chem Chemother. 2010 Aug 11;20(6):213-37. doi: 10.3851/IMP1607. Review. PubMed PMID: 20710063.

8: Klibanov OM, Kaczor RL. IDX-899, an aryl phosphinate-indole non-nucleoside reverse transcriptase inhibitor for the potential treatment of HIV infection. Curr Opin Investig Drugs. 2010 Feb;11(2):237-45. Review. PubMed PMID: 20112173.

9: Zhou XJ, Garner RC, Nicholson S, Kissling CJ, Mayers D. Microdose pharmacokinetics of IDX899 and IDX989, candidate HIV-1 non-nucleoside reverse transcriptase inhibitors, following oral and intravenous administration in healthy male subjects. J Clin Pharmacol. 2009 Dec;49(12):1408-16. doi: 10.1177/0091270009343698. Epub 2009 Sep 23. PubMed PMID: 19776293.

10: Zhou XJ, Pietropaolo K, Damphousse D, Belanger B, Chen J, Sullivan-Bólyai J, Mayers D. Single-dose escalation and multiple-dose safety, tolerability, and pharmacokinetics of IDX899, a candidate human immunodeficiency virus type 1 nonnucleoside reverse transcriptase inhibitor, in healthy subjects. Antimicrob Agents Chemother. 2009 May;53(5):1739-46. doi: 10.1128/AAC.01479-08. Epub 2009 Feb 17. PubMed PMID: 19223643; PubMed Central PMCID: PMC2681571.

11: Mascolini M, Larder BA, Boucher CA, Richman DD, Mellors JW. Broad advances in understanding HIV resistance to antiretrovirals: report on the XVII International HIV Drug Resistance Workshop. Antivir Ther. 2008;13(8):1097-113. PubMed PMID: 19195337.

12: Dalton P. Two new NNRTIs enter the pipeline. Proj Inf Perspect. 2008 Sep;(46):13. PubMed PMID: 19048672.

13: Sweeney ZK, Klumpp K. Improving non-nucleoside reverse transcriptase inhibitors for first-line treatment of HIV infection: the development pipeline and recent clinical data. Curr Opin Drug Discov Devel. 2008 Jul;11(4):458-70. Review. PubMed PMID: 18600563.

/////////////GSK2248761A , IDX899, Fosdevirine, PHASE 2

CC1=CC(=CC(=C1)C=CC#N)P(=O)(C2=C(NC3=C2C=C(C=C3)Cl)C(=O)N)OC

OLINCIGUAT


img2D chemical structure of 1628732-62-6

OLINCIGUAT

cas 1628732-62-6
Chemical Formula: C21H16F5N7O3
UNII-PD5F4ZXD21
Molecular Weight: 509.4

Olinciguat is a guanylate cyclase activator drug candidate.

(2R)-3,3,3-trifluoro-2-{[(5-fluoro-2-{1-[(2-fluorophenyl)methyl]- 5-(1,2-oxazol-3-yl)-1H-pyrazol- 3-yl}pyrimidin-4-yl)amino]methyl}-2-hydroxypropanamide

  • Originator Ironwood Pharmaceuticals
  • Class Antifibrotics; Cardiovascular therapies
  • Mechanism of Action Soluble guanylyl cyclase agonists
  • Orphan Drug StatusNo
  • New Molecular EntityYes

Highest Development Phases

  • Phase II Gastrointestinal disorders; Sickle cell anaemia
  • Phase I Cardiovascular disorders; Fibrosis

Most Recent Events

  • 03 Jan 2018 Pharmacodynamics data from a preclinical trial in Cardiovascular disorders presented at the 59th Annual Meeting and Exposition of the American Society of Hematology (ASH-2017)
  • 21 Dec 2017 Phase-II clinical trials in Sickle cell anaemia in USA (PO)
  • 09 Dec 2017 Adverse events, pharmacokinetic and pharmacodynamics data from a phase Ib trial in healthy volunteers presented at the 59th Annual Meeting and Exposition of the American Society of Hematology

IW-1701

Currently in Phase II Clinical Development

Area of focus:

Achalasia and Sickle Cell Disease
Dysregulation of the nitric oxide-soluble guanylate cyclase-cyclical guanosine monophosphate (NO-sGC-cGMP) signaling pathway is believed to be linked to multiple vascular and fibrotic diseases, such as achalasia and sickle cell disease.

Our candidate:

IW-1701 is an investigational soluble guanylate cyclase (sGC) stimulator from Ironwood’s diverse library of sGC stimulators, which are being investigated for their potential effects on vascular and fibrotic diseases. The compound has been shown in nonclinical studies to modulate the NO-sGC-cGMP signaling pathway and is currently being evaluated in a Phase II study in achalasia. IW-1701 is wholly-owned by Ironwood Pharmaceuticals.

sGC is the primary receptor for NO in vivo. sGC can be activated via both NO-dependent and NO-independent mechanisms. In response to this activation, sGC converts Guanosine-5′-triphosphate (GTP) into the secondary messenger cGMP. The increased level of cGMP, in turn, modulates the activity of downstream effectors including protein kinases, phosphodiesterases (PDEs) and ion channels.

In the body, NO is synthesized from arginine and oxygen by various nitric oxide synthase (NOS) enzymes and by sequential reduction of inorganic nitrate. Three distinct isoforms of NOS have been identified: inducible NOS (iNOS or NOS II) found in activated macrophage cells; constitutive neuronal NOS (nNOS or NOS I), involved in neurotransmission and long term potentiation; and constitutive endothelial NOS (eNOS or NOS III) which regulates smooth muscle relaxation and blood pressure. Experimental and clinical evidence indicates that reduced concentrations orbioavailability of NO and/or diminished responsiveness to endogenously produced NO contributes to the development of disease.

NO-independent, heme -dependent sGC stimulators, have shown several important differentiating characteristics, when compared to sGC activators, including crucial dependency on the presence of the reduced prosthetic heme moiety for their activity, strong synergistic enzyme activation when combined with NO and stimulation of the synthesis of cGMP by direct stimulation of sGC, independent of NO. The benzylindazole compound YC-1 was the first sGC stimulator to be identified. Additional sGC stimulators with improved potency and specificity for sGC have since been developed.

Compounds that stimulate sGC in an NO-independent manner offer considerable advantages over other current alternative therapies that target the aberrant NO pathway. There is a need to develop novel, well-characterized stimulators of sGC. Compound I is an sGC stimulator that has demonstrated efficacy for the treatment of a number of NO related disorders in preclinical models. Compound I was previously described in WO2014144100, Example 1, as a light orange solid. Compound I may be present in various crystalline forms and may also form several pharmaceutically acceptable salts.

Compounds which enhance eNOS transcription: for example those described in WO

02/064146, WO 02/064545, WO 02/064546 and WO 02/064565, and corresponding patent documents such as US2003/0008915, US2003/0022935, US2003/0022939 and US2003/0055093. Other eNOS transcriptional enhancers including those described in US20050101599 (e.g. 2,2-difluorobenzo[l,3]dioxol-5-carboxylic acid indan-2-ylamide, and 4-fluoro-N-(indan-2-yl)-benzamide), and Sanofi-Aventis compounds AVE3085 and AVE9488 (CA Registry NO. 916514-70-0; Schafer et al., Journal of Thrombosis and Homeostasis 2005; Volume 3, Supplement 1 : abstract number P 1487);

NO independent heme-independent sGC activators, including, but not limited to: -2667 (see patent publication DE19943635)

HMR-1766 (ataciguat sodi

S 3448 (2-(4-chloro-phenylsulfonylamino)-4,5-dimethoxy-N-(4-(thiomoφholine-4-sulfonyl)-phenyl)-benzamide (see patent publi

HMR-1069 (Sanofi-Aventis).

(7) Heme-dependent sGC stimulators including, but not limited to:

YC-1 (see patent publications EP667345 and DE19744026)

Riociguat (BAY 63-2521, Adempas, commercial product, described in DE19834044)

Neliciguat (BAY 60-4552, described in WO 2003095451)

Vericiguat (BAY 1021189, clinical backup to Riociguat),

BAY 41-2272 (described in DE19834047 and DE19942809)

BAY 41-8543 (described in DE I 9834044)

Etriciguat (described in WO 2003086407)

CFM-1571 (see patent publicatio

A-344905, its acrylamide analo analogue A-778935.

A-344905;

Compounds disclosed in one of publications: US20090209556, US8455638, US20110118282 (WO2009032249), US20100292192, US20110201621, US7947664, US8053455 (WO2009094242), US20100216764, US8507512, (WO2010099054) US20110218202 (WO2010065275),

US20130012511 (WO2011119518), US20130072492 (WO2011149921), US20130210798

(WO2012058132) and other compounds disclosed in Tetrahedron Letters (2003), 44(48): 8661-8663.

Pictorial synthesis

FROM PATENTS

CONSTRUCT YOUR OWN

SIDE CHAIN SHOWN ABOVE

                     FINAL STEP SHOWN ABOVE  OLINCIGUAT

PATENT

WO2014144100, Example 1

Inventors Takashi NakaiJoel MooreNicholas Robert PerlRajesh R. IyengarAra MermerianG-Yoon Jamie ImThomas Wai-Ho LeeColleen HudsonGlen Robert RENNIEJames JiaPaul Allen RENHOWETimothy Claude BardenXiang Y YuJames Edward SHEPPECKKarthik IyerJoon JungLess «
Applicant Takashi NakaiJoel MooreNicholas Robert PerlIyengar Rajesh RAra MermerianG-Yoon Jamie ImThomas Wai-Ho LeeColleen HudsonRennie Glen RobertJames JiaRenhowe Paul AllenTimothy Claude BardenXiang Y YuSheppeck James EdwardKarthik IyerJoon JungLess «

PATENT

WO 2016044447

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2016044447

Inventors Timothy Claude BardenJames Edward SHEPPECKGlen Robert RENNIEPaul Allan RenhoweNicholas PerlTakashi NakaiAra MermerianThomas Wai-Ho LeeJoon JungJames JiaKarthik IyerRajesh R. IyengarG-Yoon Jamie Im
Applicant Ironwood Pharmaceuticals, Inc.

Compound 195

lntermediate-36 Compound 195

[00463] lntermediate-36 (35 mg, 0.09 mmol),

(R)-2-(aminomethyl)-3,3,3-trifluoro-2-hydroxypropanamide (60 mg, 0.35 mmol) and

N-ethyl-N-isopropylpropan-2-amine (0.10 mL, 0.56 mmol) were mixed in dimethylsulfoxide (1.5 mL) and heated at 95°C for 8 hr. The solution was cooled to room temperature, diluted with water (2 mL) and the pH taken to 2-3 with 1 N (aq) HC1. The solution was mixed with ethyl acetate (50 mL) and the organic phase was washed with water (2 x 5 mL), brine, then dried over Na2S04, filtered and concentrated by rotary evaporation. The residue was subjected to preparative reverse phase HPLC

. . . t . + + . using a giauiciu ui water acetonitri e . tni uoroacetic aci as e uant to give me iouu i s a wnite solid (11 mg, 23% yield). ¾-NMR (400 MHz, CD3OD) δ 8.83 (br s, 1H), 8.27 (br s, 1H), 7.49 (br s,

1H), 6.9-7.0 (m, 2H), 6.5-6.6 (m, 2H), 5.86 (s, 2H), 4.35 (d, 1H), 4.16 (d, 1H) ppm. Note: exchangable protons all appeared under the residual HOD peak at 4.91 ppm.

PATENT

WO-2018009609

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2018009609&recNum=1&maxRec=&office=&prevFilter=&sortOption=&queryString=&tab=PCTDescription

Novel crystalline solid forms of olinciguat (presumed to be IW-1701), an SGC stimulator and their salts, such as hydrochloride acid (designated as Forms A, B, D, E, F, H and G), processes for their preparation and compositions comprising them are claimed. Also claimed are processes for preparing the crystalline forms. Further claimed are their use for treating cancer, sickle cell disease, osteoporosis, dyspepsia, Duchenne muscular dystrophy, amyotrophic lateral sclerosis and spinal muscle atrophy

In one aspect, the invention relates to crystalline solid forms of Compound I, depicted below:

Compound I

[0009] For purposes of this disclosure, “Compound I,” unless otherwise specifically indicated, refers to the free base or to the hydrochloric acid salt of the structure denoted above. Compound I, as its crystalline free base, is highly polymorphic and known to have seven crystalline forms (Forms A, B, D, E, F, G and H) as well as multiple solvates. Compound I was previously described in

WO2014144100, Example 1, as a light orange solid.

[0010] In one embodiment, the crystalline solid forms of Compound I here disclosed are polymorphs of the free base. In another embodiment, a crystalline solid form of Compound I is the hydrochloric acid salt. In one embodiment, the polymorphs of Compound I are crystalline free base forms. In another embodiment, they are solvates.

[001 1] In another aspect, also provided herein are methods for the preparation of the above described crystalline free forms and salts of Compound I.

[0012J In another aspect, the invention relates to pharmaceutical compositions comprising one or more of the polymorphs of Compound I herein disclosed, or the hydrochloric acid salt of Compound I, and at least one pharmaceutically acceptable excipient or carrier. In another embodiment, the invention relates to pharmaceutical dosage forms comprising said pharmaceutical compositions.

[0013] In another embodiment, the invention relates to a method of treating a disease, health condition or disorder in a subject in need thereof, comprising administering, alone or in combination therapy, a therapeutically effective amount of a polymorph of Compound I herein disclosed, or a mixture of polymorphs thereof, or its hydrochloric acid salt , to the subject; wherein the disease or disorder is one that may benefit from sGC stimulation or from an increase in the concentration of NO and/or cGMP.

EXAMPLES

Example 1: Preparation of crude Compound I

i): Coupling of Compound (1′) and 7V,0-Dimethylhydroxylamine to provide N-methoxy-N-methylisoxazole-3-carboxamide (2′)

[00238] Isooxazole-3-carboxylic acid ((l’)> 241.6 g, 2137 mmoles, 1.0 equiv.), toluene (1450 mL) and DMF (7.8 g, 107 mmoles, 0.05 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The resulting slurry was heated to 45-50 °C. Oxalyl chloride (325 g, 2559 mmoles, 1.2 equiv.) was then charged via an addition funnel over the course of 2 h while maintaining the reaction temperature between 45 to 50 °C and a vigorous gas evolution was observed. A brown mixture was obtained after addition. The brown mixture was heated to 87 to 92 °C over 1 h and stirred at 87 to 92 °C for 1 h. The reaction was completed as shown by HPLC. During heating, the brown mixture turned into a dark solution. The reaction was monitored by quenching a portion of the reaction mixture into piperidine and monitoring the piperidine amide by HPLC. The dark mixture was cooled to 20-25 °C and then filtered through a sintered glass funnel to remove any insolubles. The dark filtrate was concentrated under reduced pressure to a volume of 400 mL dark oil.

[00239] Potassium carbonate (413 g, 2988 mmoles, 1.4 equiv.) and water (1000 mL) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The reaction solution was cooled to -10 to -5 °C. N,0-dimethylhydroxyamine hydrochloride (229 g, 2348 mmoles, 1.1 equiv.) was charged to a suitable reaction vessel and dissolved in water (1000 mL). The N,0-dimethylhydroxyamine solution and dichloromethane (2500 mL) were then charged to the potassium carbonate solution.

[00240] The above dark oil (400 mL) was then charged slowly via an addition funnel while maintaining the reaction temperature -10 to 0 °C. The addition was slightly exothermic and a brown mixture was obtained after addition. The mixture was stirred at 0 to 5 °C over 20 min. and then warmed to 20 to 25 °C. The bottom organic layer was collected and the top aq. layer was extracted with dichloromethane (400 mL). The combined organic layers were washed with 15% sodium chloride solution (1200 mL). The organic layer was dried over magnesium sulfate and then filtered. The filtrate was concentrated under reduced pressure to give intermediate (2′) as a dark oil (261.9 g, 97 wt%, 76% yield, 3 wt% toluene by Ή-ΝΜΡν, 0.04 wt % water content by KF). Ή-ΝΜΡν (500 MHz, CDC13) δ ppm 8.48 (s, 1 H); 6.71(s, 1 H); 3.78 (s, 3 H); 3.38 (s, 3 H).

ii): alkylation of Compound (2′) and ethyl propiolate to provide (E)-ethyl 4-(isoxazol-3-yl)-2-(methox methyl)amino)-4-oxobut-2-enoate (3′)

(2′) (3′)

[00241] Intermediate (2′) (72.2 g, 96 wt%, 444 mmoles, 1.0 equiv.), ethyl propiolate (65.7 g, 670 mmoles, 1.5 equiv.) and anhydrous THF (650 mL) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The solution was cooled to -65 to -55 °C. Sodium bis(trimethylsilyl)amide in THF (1 M, 650 mL, 650 mmoles, 1.46 equiv.) was then charged slowly via an addition funnel while maintaining the reaction temperature -65 to -55 °C. The mixture was stirred below -55 °C over 10 min. after addition was complete. Then 1 N HC1 (650 mL, 650 mmoles, 1.46 equiv.) was charged to quench the reaction while maintaining the reaction temperature below -20 °C followed immediately with the addition of ethyl acetate (1500 mL) and water (650 mL). The top ethyl acetate layer was collected and the bottom aqueous layer was extracted with ethyl acetate (800 mL). The combined organic layers were washed with 10% citric acid (1000 mL) and saturated sodium chloride solution (650 mL). The organic layer was concentrated under reduced pressure to give a dark oil.

[00242] The dark oil was dissolved in a solution of dichloromethane/ethyl acetate/heptane

(150mL/100mL/100mL). The solution was loaded on a silica pad (410 g) and the silica pad was eluted with ethyl acetate/heptane (1/1 v/v). The filtrate (~ 3000 mL) was collected and then concentrated under reduced pressure to a volume of 150 mL to give a slurry upon standing. Heptane (200 mL) was then added to the slurry and the slurry was concentrated under reduced pressure to a volume of 150 mL. The resulting slurry was filtered, and the filter cake was washed with heptane (150 mL). The filter cake was then air dried overnight to furnish intermediate (3′) as a brown solid (63.4 g, 56% yield, >99% pure by HPLC). i-NMR (500 MHz, CDC13) δ ppm 8.42 (d, J=1.53 Hz, 1 H); 6.76 (d, J=1.53 Hz, 1 H); 6.18 (s, 1 H); 4.47 (q, J=7.07 Hz, 2H); 3.75 (s, 3 H); 3.21 (s, 3 H); 1.41 (t, J=7.17 Hz, 3 H). iii): cyclization of Compound 3′ and 2-fluorobenzylhydrazine to provide ethyl l-(2-fluorobenz l)-5-(isoxazol-3-yl)-lH-pyrazole-3-carboxylate (4′)

[00243] Intermediate (3′) (72.9 g, 287 mmoles, 1.0 equiv.) and absolute ethanol (730 mL) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The mixture was cooled to 0 to 5 °C. 2-Fluorobenzylhydrazine (48.2 g, 344 mmoles, 1.2 equiv.) was then charged to the mixture. The mixture was stirred at 0 to 10 °C over 1 h and then warmed to 20 to 25 °C and stirred at 20 to 25 °C over 16 h. The reaction was completed by HPLC. Concentrated HCl (33.9 g, 37 wt%, 344 mmoles, 1.2 equiv.) was charged to the reaction mixture over 1 min and the batch temperature exothermed from 20 °C to 38 °C. A slurry was obtained. The mixture was cooled to 0 to 10 °C over 1 h and stirred at 0-10 °C for 1 h. The resulting slurry was filtered, and the filter cake was washed with ethanol (200 mL). The filter cake was dried under vacuum at 30 to 40 °C over 16 h to furnish intermediate (4′) as an off-white solid (81.3 g, 90% yield, >99% pure by HPLC). ¾-NMR (500 MHz, CDC13) δ ppm 8.47 (d, J=1.68 Hz, 1 H); 7.15 – 7.26 (m, 2 H); 6.94 – 7.08 (m, 2H); 6.77 – 6.87 (m, 1 H); 6.55 (d, J=1.68 Hz, 1 H); 5.95 (s, 2 H); 4.43 (q, J=7.02 Hz, 2 H); 1.41 (t, J=7.17 Hz, 3 H).

iv): amination of Compound (4′) to provide l-(2-fluorobenzyl)-5-(isoxazol-3-yl)-lH-pyrazole-3-carboximidamide hydrochloride (5’B)

[00244] Anhydrous ammonium chloride (267 g, 4991 mmoles, 5.0 equiv.) and toluene (5400 mL) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. Trimethylaluminum in toluene (2 M, 2400 mL, 4800 mmoles, 4.8 equiv.) was charged

slowly via an addition funnel while maintaining the reaction temperature at 20 to 40 °C (Note:

Methane gas evolution was observed during addition). Then the mixture was heated to 75 to 80 °C over 30 min. and a clear white solution was obtained. Intermediate (4′) (315 g, 999 mmoles, 1.0 equiv.) was charged to reaction mixture in four equal portions over 1 h at 75 to 90 °C. The reaction was stirred at 80 to 90 °C over 30 min. and then heated to 100 to 110 °C and stirred at 100 to 110 °C over 3 h. The reaction was completed by HPLC. The reaction mixture was cooled to 10 to 20 °C and methanol (461 g, 14.4 moles, 14.4 equiv.) was charged slowly via an addition funnel while

maintaining the reaction temperature 10-40 °C. Note the quenching was very exothermic and a lot gas evolution was observed. A thick slurry was obtained. A 3N HQ (6400 mL, 3 N, 19.2 moles, 19.2 equiv.) was then charged slowly via an addition funnel while maintaining the reaction temperature at 20 to 45 °C. The mixture was heated to 80 to 85 °C and stirred at 80 to 85 °C over 10 min. to obtain a clear biphasic mixture. The mixture was cooled to 0 to 5 °C over 3 h and stirred at 0 to 5 °C over 1 h. The resulting slurry was filtered, and the filter cake was washed with water (3000 mL). The filter cake was dried under vacuum at 40 to 50 °C over 24 h to furnish intermediate (5’B) as an off-white solid (292 g, 91% yield, >99% pure by HPLC). ¾-ΝΜΡν (500 MHz, DMSO- 6) δ ppm 9.52 (s, 2 H); 9.33 (s, 2 H); 9.18 (d, J=1.53 Hz, 1 H); 7.88 (s, 1 H); 7.29 – 7.38 (m, 1 H); 7.19 – 7.25 (m, 1 H); 7.10 – 7.16 (m, 1 H); 7.03 (d, J=1.53 Hz, 1 H); 6.92 – 6.98 (m, 1 H); 5.91 (s, 2 H). M.P. 180-185 °C.

v): cyclization of Compound (5’B) and diethyl fluoromalonate to provide 5-fluoro-2-(l-(2-fluorobenz l)-5-(isoxazol-3-yl)-lH-pyrazol-3-yl)pyrimidine-4,6-diol (6′)

(5’B) (6·)

[00245] Intermediate (5’B) (224.6 g, 698 mmoles, 1.0 equiv.), methanol (2250 mL) and diethyl fluoromalonate (187 g, 1050 mmoles, 1.5 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. Then sodium methoxide in methanol solution (567 g, 30 wt %, 3149 mmoles, 4.5 equiv.) was charged via an addition funnel while maintaining the reaction temperature 20 to 35 °C. The mixture was stirred at 20 to 35 °C over 30 min. and a light suspension was obtained. The reaction was completed by HPLC. A solution of 1.5 N HQ (2300 mL, 3450 mmoles, 4.9 equiv.) was charged via an addition funnel over 1 h while maintaining the reaction temperature 20 to 30 °C. A white suspension was obtained. The pH of the reaction mixture was to be ~1 by pH paper. The slurry was stirred at 20 to 30 °C over 30 min. The resulting slurry was filtered, and the filter cake was washed with a pre-mixed solution of methanol and water (500 mL/500 mL), and then with water (1000 mL). The filter cake was dried under vacuum at 50 to 60 °C over 16 h to furnish intermediate (6′) as an off-white solid (264 g, 97% yield, >99% pure by HPLC). ¾-NMR (500 MHz,

DMSO- s) δ ppm 12.82 (br. s., 1 H); 12.31 (br. s., 1 H); 9.14 (d, J=1.53 Hz, 1 H); 7.55 (s, 1 H); 7.31 -7.37 (m, 1 H); 7.18 – 7.25 (m, 1 H); 7.10 – 7.15 (m, 2 H); 6.97 – 7.02 (t, J=7.55 Hz, 1 H); 5.88 (s, 2 H).

vi): chlorination of Compound (6′) to provide 3-(3-(4,6-dichloro-5-fluoropyrimidin-2-yl)-l-(2-fluorobenz l)-lH-pyrazol-5-yl)isoxazole (7′)

(6«) (7«)

[00246] Intermediate (6′) (264 g, 71 1 mmoles, 1.0 equiv.), acetonitrile (4000 mL) and N,N-dimethylaniline (138 g, 1 137 mmoles, 1.6 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The slurry mixture was heated to 70-80 °C. Then phosphorous oxychloride (655 g, 4270 mmoles, 6.0 equiv.) was charged via an addition funnel over 1 h while maintaining the reaction temperature 70 to 80 °C. The mixture was stirred at 75 to 80 °C over 22 h and a brown solution was obtained. The reaction was completed by HPLC. Then the mixture was cooled to between 0 and 5 °C and cotton like solids precipitated out at 25 °C. Water (3000 mL) was charged slowly via an addition funnel while maintaining the reaction temperature at 0 to 10 °C. The slurry was stirred at 0 to 10 °C over 30 min. The resulting slurry was filtered, and the filter cake was washed with a pre-mixed solution of acetonitrile and water (500 mL/500 mL). The filter cake was dried under vacuum at 35 to 45 °C over 16 h to furnish intermediate (7′) as an off-white solid (283 g, 98% yield, >99% pure by HPLC). ‘H-NMR (500 MHz, CDC13) δ ppm 8.48 (d, J=1.68 Hz, 1 H); 7.44 (s, 1 H); 7.19 – 7.25 (m, 1 H); 6.96 – 7.08 (m, 2 H); 6.81 – 6.88 (m, 1 H); 6.60 (d, J=1.68 Hz, 1 H); 6.03 (s, 2 H).

vii): substitution of Compound (7′) with meth oxide to provide 3-(3-(4-chloro-5-fluoro-6-m

(7′) (8′)

[00247] Methanol (3400 mL) and sodium methoxide in methanol (154 mL, 5.4 M, 832 mmoles,

1.2 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The reaction mixture was heated to 23 to 27 °C. Intermediate (7′) (283 g, 693 mmoles, 1.0 equiv.) was charged to the mixture in small portions (5-10 g each portion) over 40 min while maintaining the reaction temperature 23 to 27 °C. The slurry was stirred at 23 to 27 °C over 30 min. The reaction was completed by HPLC. The resulting slurry was filtered, and the filter cake was washed with methanol (850 mL) and then water (850 mL). The filter cake was dried under vacuum at 35 to 45 °C over 16 h to furnish intermediate (8′) as an off-white solid (277 g, 99% yield, 97% pure by HPLC). i-NMR (500 MHz, CDCl3) 5 ppm 8.47 (d, J=1.83 Hz, 1 H); 7.38 (s, 1 H); 7.18 – 7.25 (m, 1 H); 7.01 – 7.08 (m, 1 H); 6.94 – 7.00 (m, 1 H); 6.81 – 6.88 (m, 1 H); 6.60 (d, J=1.68 Hz, 1 H); 6.00 (s, 2 H); 4.21 (s, 3 H).

viii): hydrogenation of Compound (8′) to provide 3-(3-(5-fluoro-4-methoxypyrimidin-2-yl)-l-(2-fluorobenz l)-lH-pyrazol-5-yl)isoxazole (9′)

[00248] Intermediate (8′) (226 g, 560 mmoles, 1.0 equiv.), palladium (10% on activated carbon, nominally 50% water wet, 22.6 g, 0.01 moles, 0.018 equiv), tetrahydrofuran (3400 mL) and triethylamine (91 g, 897 mmoles, 1.6 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. Nitrogen was bubbled into the reaction mixture via teflon tubing over 10 min. at 20 to 30 °C. Then the mixture was heated to 40 to 50 °C and hydrogen gas was bubbled into the reaction mixture via teflon tubing over 6 h while maintaining the reaction temperature 40 to 50 °C. The reaction was completed by HPLC. Nitrogen was then bubbled into the reaction mixture via teflon tubing over 10 min. at 40 to 50 °C The reaction mixture was hot filtered through Hypo Supercel™ and the filter cake was washed with tetrahydrofuran (2000 mL). The filtrate was concentrated under reduced pressure to a volume of -1300 mL to give a slurry. Tetrahydrofuran was then solvent exchanged to methanol under reduced pressure via continuously feeding methanol (3000 mL). The final volume after solvent exchange was 1300 mL. The resulting slurry was filtered, and the filter cake was washed with methanol (500 mL). The filter cake was dried under vacuum at 20 to 25 °C over 16 h to furnish intermediate (9′) as a white solid (192 g, 93% yield, 98% pure by HPLC). ¾-NMR (500 MHz, CDC13) δ ppm 8.47 (d, J=1.68 Hz, 1 H); 8.41 (d, J=2.59 Hz, 1 H); 7.36 (s, 1 H); 7.17 – 7.24 (m, 1 H); 6.95 – 7.07 (m, 2 H); 6.83 – 6.90 (m, 1 H); 6.60 (d, J=1.68 Hz, 1 H); 5.99 (s, 2 H); 4.19 (s, 3 H).

ix: demethylation of Compound (9′) to provide 5-fluoro-2-(l-(2-fluorobenzyl)-5-(isoxazol-3-yl)-lH-pyrazol-3-yl)pyrimidin-4-ol (10′)

[00249] Intermediate (9′) (230 g, 623 mmoles, 1.0 equiv.), Me OH (3450 mL) and cone. HC1

(307 g, 37 wt%, 3117 mmoles, 5.0 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The mixture was heated to 60 to 65 °C and a solution was obtained. The mixture was then stirred at 60 to 65 °C over 17 h and a slurry was obtained. The reaction was completed by HPLC. The slurry was cooled to 20 to 25 °C over 2 h and stirred at 20 to 25 °C over 30 min. The resulting slurry was filtered, and the filter cake was washed with methanol (1000 mL). The filter cake was dried under vacuum at 35 to 45 °C over 16 h to furnish intermediate (10′) as a white solid (214 g, 97% yield, >99% pure by HPLC). ¾-NMR (500 MHz, DMSO-t/6) δ ppm 12.90 – 13.61 (br. s., 1 H); 9.11 (d, J=1.68 Hz, 1 H); 8.16 (s, 1 H); 7.64 (s, 1 H); 7.29 – 7.42 (m, 1 H); 7.17 – 7.28 (m, 2 H); 7.08 – 7.15 (m, 1 H); 6.97 (s, 1 H); 5.91 (s, 3 H).

x): chlorination of Compound (10′) to provide 3-(3-(4-chloro-5-fluoropyrimidin-2-yl)-l-(2-fluorobenzyl)-lH-pyrazol-5-yi)isoxazole (Formula IV)


Formula IV

[00250] Intermediate (10′) (214 g, 602 mmoles, 1.0 equiv.), acetonitrile (3000 mL) and NN-dimethylaniline (109 g, 899 mmoles, 1.5 equiv.) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The slurry mixture was heated to 70 to 80 °C. Then phosphorous oxychloride (276 g, 1802 mmoles, 3.0 equiv.) was charged via an addition funnel over 30 min. while maintaining the reaction temperature 70-80 °C. The mixture was stirred at 75 to 80 °C over 2 h and a green solution was obtained. The reaction was completed by HPLC. Then the mixture was cooled to 0 to 5 °C. Water (1500 mL) was charged slowly via an addition funnel while maintaining the reaction temperature at 0 to 10 °C. The slurry was stirred at 0 to 10 °C over 30 min. The resulting slurry was filtered, and the filter cake was washed with a pre-mixed solution of

acetonitrile and water (500 mL/500 mL) and water (500 mL). The filter cake was dried under vacuum at 30 to 40 °C over 16 h to furnish intermediate of Formula IV as an off-white to pink solid (214 g, 95% yield, >99% pure by HPLC). 1H NMR (500 MHz, CDC13) 5 ppm 8.65 (s, 1 H); 8.48 (d, J=1.68 Hz, 1 H); 7.44 (s, 1 H); 7.21 – 7.25 (m, 1 H); 6.97 – 7.06 (m, 2 H); 6.83 – 6.87 (m, 1 H); 6.61 (d, J=1.68 Hz, 1 H); 6.03 (s, 2 H).

a): Cyanation of intermediate (15) to provide 2-(bromomethyl)-3,3,3-trifluoro-2-((trimethylsilyl)oxy)propanenitrile (16)

(15) (16)

[00251 ] Trimethylsilanecarbonitrile ( 153 g, 1.54 moles, 0.97 equiv) and triethylamine (4.44 mL,

3.22 g, 0.032 mole, 0.02 equiv) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The mixture was cooled to 5 °C. 3-Bromo-l, l, l-trifluoropropan-2-one ((15), 304 g, 1.59 moles, 1.0 equiv) was charged via an addition funnel over 35 min, while maintaining the reaction temperature between 10 to 20 °C. The mixture was stirred at 20 to 30 °C over 3 h after the addition to furnish intermediate (16) as a dense oil which was used directly in the next step. 1H-NMR (500 MHz, CDC13) δ ppm 3.68 (d, J=1 1.14 Hz, 1 H); 3.57 (d, J=11.14 Hz, 1 H), 0.34 – 0.37 (m, 9 H).

b): Conversion of nitrile Compound (16) to amide to provide 2-(bromomethyl)-3,3,3-trifluoro-2-hydroxypropanamide (17)

2

(16) (17)

[00252] Concentrated sulfuric acid (339 mL, 6.37 moles, 4.0 equiv) was stirred in a suitable reaction vessel equipped with a mechanical stirrer, digital thermometer and an addition funnel. The sulfuric acid was heated to 45 °C. The above intermediate (16) was added via an addition funnel over 50 min, while keeping the temperature below 75 °C. The reaction mixture was stirred at 75 °C for 2 h and then allowed to cool to room temperature. ¾-NMR indicated reaction complete. The reaction mixture was cooled to -15 °C and diluted with ethyl acetate (1824 mL) via an addition funnel over 45 min (very exothermic), while keeping the temperature between -15 to 5 °C. Water ( 1520 mL) was added slowly via an addition funnel for 1 h 20 min. (very exothermic) between -10 to 0 °C. The layers were separated and the organic layer was washed with 15% aqueous sodium chloride solution ( 1520

mL), 25% aqueous sodium carbonate solution (911 mL) followed by 15% aqueous sodium chloride solution (911 mL). The organic layer was filtered and concentrated under reduced pressure to get 348 g of intermediate (17) as light yellow oil. This oil was dissolved in methanol (1200 mL) and concentrated to furnish 380 g of intermediate (17). (296 g adjusted weight, 79% yield). i-NMR (500 MHz, CDC13) 5 6.61 – 6.94 (m, 1 H); 5.92 – 6.26 (m, 1 H); 3.93 – 4.00 (m, 1 H); 3.68 (d, J=l 1.14 Hz, 1 H).

c): N-Alkylation of compound (17) to provide of 2-(aminomethyl)-3,3,3-trifluoro-2-hydroxypropanamide (14)

(17) (14)

[00253] A 7 N solution of ammonia in methanol (600 mL, 4.28 moles, 10 equiv) was charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The solution was cooled to 0 to 5 °C. Then the intermediate (17) (102 g, 0.432 moles, 1 equiv) was added via an addition funnel over 30 min at 0 to 5 °C. The reaction mixture was warmed to 20 to 25 °C over 1 h and held for 72 h. The reaction was completed by HPLC. The reaction mixture was cooled to 0 to 5 °C and sodium methoxide (78 mL, 5.4 M, 0.421 moles, 0.97 equiv) was added over 2 min. The reaction mixture was then concentrated under reduced pressure to a volume of 300 mL. 2 L of ethyl acetate was added and concentration was continued under reduced pressure to a volume to 700 mL to get a slurry. 700 mL of ethyl acetate was added to the slurry to make the final volume to 1400 mL. 102 mL of water was added and stirred for 2 min to get a biphasic solution. The layers were separated. The ethyl acetate layer was concentrated under reduced pressure to a volume of 600 mL. Then the ethyl acetate layer was heated to > 60 °C and heptane (600 mL) was added slowly between 55 to 60 °C. The mixture was cooled to 15 to 20 °C to give a slurry. The slurry was stirred at 15 to 20 °C for 2 h and filtered. The solids were dried under vacuum at 25 °C for 16 h to furnish amine (14) as white solid (48 g, 64% yield). ‘H-NMR (500 MHz, MeOH-d4) δ ppm 2.94 (d, J= 13.73 Hz, 1H); 3.24 (d, J= 13.58 Hz, 1H).

d): chiral resolution of amine (14) as the 1:1 salt of (R)-2,2-dimethyl-5- (trifluoromethyl)oxazolidine-5-carboxamide (R)-2-hydroxysuccinate (18A) and (D)-malic acid.

(14) (ISA)

[00254] Amine (14) (105 g, 0.608 moles, 1.0 equiv.), (D)-Malic acid (82 g, 0.608 moles, 1.0 equiv.) and acetone (1571 mL) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The reaction mixture was stirred at 20 to 25 °C for 16 h. The resulting slurry was filtered, and the wet cake was washed with acetone (300 mL). The wet cake was charged back to the reaction vessel, and acetone (625 mL) was charged. The slurry was heated to 53 °C and held for 6 h. The slurry was cooled to 20 to 25 °C and held at this temperature for 16 h. The slurry was filtered, and the wet cake was washed with acetone (200 mL). The wet cake was dried under vacuum at 40 °C for 4 h to furnish 82.4 g of the 1 : 1 salt of (18A) and (D)-malic acid as a white solid (82.4 g, 39% yield, 97% ee). i-NMR (500 MHz, D20) δ ppm 4.33 (br, s, 1H); 3.61 (br, d, J= 13.58 Hz, 1H); 3.40 – 3.47 (m, 1H); 2.76 (br, d, J= 15.87 Hz, 1H); 2.53 – 2.63 (m, 1H); 2.16 (br, s, 4H).

e): Coupling of the 1:1 (D)-malic acid salt of intermediate (18A) and Formula IV to provide (R)-3,3,3-trifluoro-2-(((5-fluoro-2-(l-(2-fluorobenzyl)-5-(isoxazol-3-yl)-lH-pyrazol-3-yl)pyrimidin-4-yl)amino)methyl)-2-hydroxypropanamide (Compound I)

Formula IV Compound I

[00255] The 1: 1 salt of intermediate (18A) and (D)-malic acid (74.1 g, 0.214 moles, 2.5 equiv) and water (44.8 mL) were charged to a suitable reaction vessel equipped with a mechanical stirrer and a digital thermometer. The reaction mixture was heated to 70 °C and stirred for 20 min. Acetone generated during the reaction was removed by blowing with nitrogen. The reaction mixture was cooled to 30 to 40 °C and Formula IV (32 g, 0.086 moles, 1.0 equiv), DMSO (448 mL) and Hunig’s base (44.7 mL, 0.257 moles, 3.0 equiv) were charged. The reaction mixture was heated to 90 °C and stirred at 90 °C over 17 h. The reaction was complete by HPLC. Then the mixture was cooled to 60 °C. Another portion of Hunig’s base (104 mL, 0.599 moles, 7.0 equiv) was charged followed by water (224 mL) at 55 to 62 °C. The reaction mixture was stirred for 15 min at 55 to 60 °C to form the seed bed. Water (320 mL) was added via addition funnel at 55 to 62 °C over the course of 30 min, and the resultant slurry was stirred for 1 h at 55 to 60 °C. The resulting slurry was filtered, and the filter cake was washed with a pre-mixed solution of methanol and water (320 mL/320 mL) followed by water (640 mL). The filter cake was then dried under vacuum at 40 °C over 16 h to furnish Compound I as an off-white solid (40 g, 92% yield, 99% pure by HPLC, 98% ee). ¾-NMR (500 MHz, DMSO-t/6) δ ppm 9.10 (s, 1 H); 8.33 (d, J=2.90 Hz, 1 H); 7.93 (s, br, 1 H); 7.90 (s, 1 H); 7.78 (s, br, 1 H); 7.69 (s, br, 1 H); 7.52 (s, 1 H); 7.33 (q, J=7.02 Hz, 1 H); 7.17 – 7.25 (m, 1 H); 7.17 – 7.25

(m, 1 H); 7.10 (t, J=7.48 Hz, l H); 6.98 (t, J=7.55 Hz, 1 H); 5.90 (s, 2 H); 3.92-4.05 (m, 2 H).

////////////OLINCIGUAT, IW-1701, phase 2, ironwood

NC(=O)[C@](O)(CNc1nc(ncc1F)c2cc(c3ccon3)n(Cc4ccccc4F)n2)C(F)(F)F

TRILACICLIB, G1T28


ChemSpider 2D Image | Trilaciclib | C24H30N8OTrilaciclib.png

Trilaciclib

  • Molecular FormulaC24H30N8O
  • Average mass446.548 Da
  • G1T 28
CAS 1374743-00-6
2′-{[5-(4-Methyl-1-piperazinyl)-2-pyridinyl]amino}-7′,8′-dihydro-6’H-spiro[cyclohexane-1,9′-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one
G1T28, SHR 6390
Spiro[cyclohexane-1,9′(6’H)-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one, 7′,8′-dihydro-2′-[[5-(4-methyl-1-piperazinyl)-2-pyridinyl]amino]-
  • 7′,8′-Dihydro-2′-[[5-(4-methyl-1-piperazinyl)-2-pyridinyl]amino]spiro[cyclohexane-1,9′(6’H)-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one
  • 2′-[[5-(4-Methylpiperazin-1-yl)pyridin-2-yl]amino}-7′,8′-dihydro-6’H-spiro[cyclohexane-1,9′-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one
UNII:U6072DO9XG

Reduction of Chemotherapy-Induced Myelosuppression

Trilaciclib dihydrochloride
1977495-97-8

2D chemical structure of 1977495-97-8

In phase II clinical development as a chemoprotectant at G1 Therapeutics for first- or second-line treatment in patients with metastatic triple negative breast cancer, in combination with gemcitabine and carboplatin

logo

PATENT, WO 2014144326Compound 89 (also referred to as Compound T)

WO2014144847A3
Inventors Norman E. SharplessJay Copeland StrumJohn Emerson BisiPatrick Joseph RobertsFrancis Xavier Tavares
Applicant G1 Therapeutics, Inc.
Norman Sharpless
Norman Sharpless official photo.jpg
Born Norman Edward Sharpless
September 20, 1966 (age 51)
Greensboro, North Carolina
Nationality American
Other names Ned Sharpless
Occupation Director, Lineberger Comprehensive Cancer Center Founder, G1 Therapeutics ($GTHX)
Notable work Wellcome Distinguished Professor, American Society of Clinical Investigation Member, Association of American Cancer Institute board of directors,

NCI Director Dr. Norman E. SharplessPinterest

NCI Director Dr. Norman E. Sharpless, Credit: National Institutes of Health

Norman E. “Ned” Sharpless, M.D., was officially sworn in as the 15th director of the National Cancer Institute (NCI) on October 17, 2017. Prior to his appointment, Dr. Sharpless served as the director of the University of North Carolina (UNC) Lineberger Comprehensive Cancer Center, a position he held since January 2014.

Dr. Sharpless was a Morehead Scholar at UNC–Chapel Hill and received his undergraduate degree in mathematics. He went on to pursue his medical degree from the UNC School of Medicine, graduating with honors and distinction in 1993. He then completed his internal medicine residency at the Massachusetts General Hospital and a hematology/oncology fellowship at Dana-Farber/Partners Cancer Care, both of Harvard Medical School in Boston.

After 2 years on the faculty at Harvard Medical School, he joined the faculty of the UNC School of Medicine in the Departments of Medicine and Genetics in 2002. He became the Wellcome Professor of Cancer Research at UNC in 2012.

Dr. Sharpless is a member of the Association of American Physicians as well as the American Society for Clinical Investigation (ASCI), the nation’s oldest honor society for physician–scientists, and served on the ASCI council from 2011 to 2014. Dr. Sharpless was an associate editor of Aging Cell and deputy editor of the Journal of Clinical Investigation. He has authored more than 150 original scientific papers, reviews, and book chapters, and is an inventor on 10 patents. He cofounded two clinical-stage biotechnology companies: G1 Therapeutics and HealthSpan Diagnostics.

In addition to serving as director of NCI, Dr. Sharpless continues his research in understanding the biology of the aging process that promotes the conversion of normal self-renewing cells into dysfunctional cancer cells. Dr. Sharpless has made seminal contributions to the understanding of the relationship between aging and cancer, and in the preclinical development of novel therapeutics for melanoma, lung cancer, and breast cancer.

Record ID Title Status Phase
NCT03041311 CarboplatinEtoposide, and Atezolizumab With or Without Trilaciclib (G1T28), a CDK 4/6 Inhibitor, in Extensive Stage Small Cell Lung Cancer (SCLC) Recruiting 2
NCT02978716 Trilaciclib (G1T28), a CDK 4/6 Inhibitor, in Combination With Gemcitabineand Carboplatin in Metastatic Triple Negative Breast Cancer (mTNBC) Recruiting 2
NCT02514447 Trilaciclib (G1T28), a CDK 4/6 Inhibitor, in Patients With Previously Treated Extensive Stage SCLC Receiving Topotecan Chemotherapy Recruiting 2
NCT02499770 Trilaciclib (G1T28), a CDK 4/6 Inhibitor, in Combination With Etoposide and Carboplatin in Extensive Stage Small Cell Lung Cancer (SCLC) Active, not recruiting 2

Synthesis

WO  2016040858

Trilaciclib (G1T28)

Trilaciclib is a potential first-in-class short-acting CDK4/6 inhibitor in development to preserve hematopoietic stem cells and enhance immune system function during chemotherapy. Trilaciclib is administered intravenously prior to chemotherapy and has the potential to significantly improve treatment outcomes.

G1 is currently evaluating trilaciclib in four Phase 2 clinical trials: three studies in patients with small-cell lung cancer (SCLC), and one study in patients with triple-negative breast cancer (TNBC). Preliminary data from the SCLC trials were presented at the American Society of Clinical Oncology 2017 Annual Meeting and at the 2016 World Conference on Lung Cancer.

Data from a Phase 1 trial in healthy volunteers were presented at the American Society of Clinical Oncology 2015 Annual Meeting and published in Science Translational Medicine. Trilacicilib has been extensively studied in animals; these preclinical data have been presented at several scientific meetings and published in Molecular Cancer Therapeutics, Science Translational Medicine, and Cancer Discovery.

Trilaciclib is a small molecule, competitive inhibitor of cyclin dependent kinases 4 and 6 (CDK4/6), with potential antineoplastic and chemoprotective activities. Upon intravenous administration, trilaciclib binds to and inhibits the activity of CDK4/6, thereby blocking the phosphorylation of the retinoblastoma protein (Rb) in early G1. This prevents G1/S phase transition, causes cell cycle arrest in the G1 phase, induces apoptosis, and inhibits the proliferation of CDK4/6-overexpressing tumor cells. In patients with CDK4/6-independent tumor cells, G1T28 may protect against multi-lineage chemotherapy-induced myelosuppression (CIM) by transiently and reversibly inducing G1 cell cycle arrest in hematopoietic stem and progenitor cells (HSPCs) and preventing transition to the S phase. This protects all hematopoietic lineages, including red blood cells, platelets, neutrophils and lymphocytes, from the DNA-damaging effects of certain chemotherapeutics and preserves the function of the bone marrow and the immune system. CDKs are serine/threonine kinases involved in the regulation of the cell cycle and may be overexpressed in certain cancer cell types. HSPCs are dependent upon CDK4/6 for proliferation.

Trilaciclib (G1T28) is a CDK4/6 inhibitor in phase II clinical development as a chemoprotectant at G1 Therapeutics for first- or second-line treatment in patients with metastatic triple negative breast cancer, in combination with gemcitabine and carboplatin. Also, phase II trials are ongoing in newly diagnosed, treatment-naive small-cell lung cancer patients, in combination with carboplatin, etoposide, and atezolizumab and phase I trials in previously treated small-cell lung cancer patients, in combination with topotecan.

U.S. Patent Nos. 8,822,683; 8,598,197; 8,598,186, 8,691,830, 8,829,102, 8,822,683, 9, 102,682, 9,499,564, 9,481,591, and 9,260,442, filed by Tavares and Strum and assigned to Gl Therapeutics describe a class of N-(heteroaryl)-pyrrolo[3,2-d]pyrimidin-2-amine cyclin dependent kinase inhibitors including those of the formula with variables as defined therein):

U.S. Patent Nos. 9,464,092, 9,487,530, and 9,527,857 which are also assigned to Gl Therapeutics describe the use of the above pyrimidine-based agents in the treatment of cancer.

These patents provide a general synthesis of the compounds that is based on a coupling reaction of a fused chloropyrimidine with a heteroaryl amine to form the central disubstituted amine. Such coupling reactions are sometimes referred to as Buchwald coupling (see WO Ί56 paragraph 127; reference WO 2010/020675). The lactam of the fused chloropyrimidine, for example, a 2-chloro-spirocyclo-pyrrolo[2,3-d]pyrimidine-one such as Intermediate K as shown below can be prepared by dehydration of the corresponding carboxylic acid. The reported process to prepare intermediate IK requires seven steps.


(Intermediate IK; page 60, paragraph 215 of WO Ί56)

WO 2013/148748 (U.S. S.N. 61/617,657) entitled “Lactam Kinase Inhibitors” filed by Tavares, and also assigned to Gl Therapeutics likewise describes the synthesis of N-(heteroaryl)-pyrrolo[3,2-d]pyrimidin-2-amines via the coupling reaction of a fused chloropyrimidine with a heteroaryl amine to form the central disubstituted amine.

WO 2013/163239 (U.S. S.N. 61/638,491) “Synthesis of Lactams” describes a method for the synthesis of this class of compounds with the variation that in the lactam preparation step, a carboxylic acid can be cyclized with a protected amine in the presence of a strong acid and a dehydrating agent, which can be together in one moiety as a strong acid anhydride. The purported improvement is that cyclization can occur without losing the protecting group on the amine before cyclization. The typical leaving group is “tBOC” (t-butoxycarbonyl). The application teaches (page 2 of WO 2013/163239) that the strong acid is, for example, trifluoroacetic acid anhydride, tribromoacetic acid anhydride, trichloroacetic acid anhydride or mixed anhydrides. An additional step may be necessary to take off the N-protecting group. The dehydrating agent can be a carbodiimide-based compound such as DCC (Ν,Ν-dicyclohexylcarbodiimide), EDC (l-ethyl-3-(3-dimethylaminopropyl)carbodiimide, or DIC (Ν,Ν-diisopropylcarbodiimide). DCC and DIC are in the same class of reagents-carbodiimides. DIC is sometimes considered better because it is a liquid at room temperature, which facilitates reactions.

WO 2015/061407 filed by Tavares and licensed to Gl Therapeutics also describes the synthesis of these compounds via the coupling of a fused chloropyrimidine with a heteroaryl amine to form the central disubstituted amine. WO ‘407 focuses on the lactam production step and in particular describes that the fused lactams of these compounds can be prepared by treating the carboxylic acid with an acid and a dehydrating agent in a manner that a leaving group on the amine is not removed during the amide-forming ring closing step.

Other publications that describe compounds of this general class include the following. WO 2014/144326 filed by Strum et al. and assigned to Gl Therapeutics describes compounds and methods for protection of normal cells during chemotherapy using pyrimidine based CDK4/6 inhibitors. WO 2014/144596 filed by Strum et al. and assigned to Gl Therapeutics describes compounds and methods for protection of hematopoietic stem and progenitor cells against ionizing radiation using pyrimidine based CDK4/6 inhibitors. WO 2014/144847 filed by Strum et al. and assigned to Gl Therapeutics describes HSPC-sparing treatments of abnormal cellular proliferation using pyrimidine based CDK4/6 inhibitors. WO2014/144740 filed by Strum et al. and assigned to Gl Therapeutics describes highly active anti -neoplastic and anti-proliferative pyrimidine based CDK 4/6 inhibitors. WO 2015/161285 filed by Strum et al. and assigned to Gl Therapeutics describes tricyclic pyrimidine based CDK inhibitors for use in radioprotection. WO 2015/161287 filed by Strum et al. and assigned to Gl Therapeutics describes analogous tricyclic pyrimidine based CDK inhibitors for the protection of cells during chemotherapy. WO 2015/161283 filed by Strum et al. and assigned to Gl Therapeutics describes analogous tricyclic pyrimidine based CDK inhibitors for use in HSPC-sparing treatments of RB-positive abnormal cellular proliferation. WO 2015/161288 filed by Strum et al. and assigned to Gl Therapeutics describes analogous tricyclic pyrimidine based CDK inhibitors for use as anti -neoplastic and anti-proliferative agents. WO 2016/040858 filed by Strum et al. and assigned to Gl Therapeutics describes the use of combinations of pyrimidine based CDK4/6 inhibitors with other anti-neoplastic agents. WO 2016/040848 filed by Strum et al. and assigned to Gl Therapeutics describes compounds and methods for treating certain Rb-negative cancers with CDK4/6 inhibitors and topoisomerase inhibitors.

Other biologically active fused spirolactams and their syntheses are described, for example, in the following publications. Griffith, D. A., et al. (2013). “Spirolactam-Based Acetyl-CoA Carboxylase Inhibitors: Toward Improved Metabolic Stability of a Chromanone Lead Structure.” Journal of Medicinal Chemistry 56(17): 7110-7119, describes metabolically stable spirolactams wherein the lactam resides on the fused ring for the inhibition of acetyl-CoA carboxylase. WO 2013/169574 filed by Bell et al. describes aliphatic spirolactams as CGRP receptor antagonists wherein the lactam resides on the spiro ring. WO 2007/061677 filed by Bell et al. describes aryl spirolactams as CGRP receptor antagonists wherein the lactam resides on the spiro ring. WO 2008/073251 filed by Bell et al. describes constrained spirolactam compounds wherein the lactam resides on the spiro ring as CGRP receptor antagonists. WO 2006/031606 filed by Bell et al. describes carboxamide spirolactam compounds wherein the spirolactam resides on the spiro ring as CGRP receptor antagonists. WO 2006/031610, WO 2006/031491, and WO 2006/029153 filed by Bell et al. describe anilide spirolactam compounds wherein the spirolactam resides on the spiro ring; WO 2008/109464 filed by Bhunai et al. describes spirolactam compounds wherein the lactam resides on the spiro ring which is optionally further fused.

Given the therapeutic activity of selected N-(heteroaryl)-pyrrolo[3,2-d]pyrimidin-2-amines, it would be useful to have additional methods for their preparation. It would also be useful to have new intermediates that can be used to prepare this class of compounds.

PATENT

WO 2014144596

PATENT

WO 2014144326

Compound 89 (also referred to as Compound T)

WO2014144847A3
Inventors Norman E. SharplessJay Copeland StrumJohn Emerson BisiPatrick Joseph RobertsFrancis Xavier Tavares
Applicant G1 Therapeutics, Inc.

EXAMPLES

Intermediates B, E, K, L, 1A, IF and 1CA were synthesized according to US 8,598,186 entitled CDK Inhibitors to Tavares, F.X. and Strum, J.C..

The patents WO 2013/148748 entitled Lactam Kinase Inhibitors to Tavares, F.X., WO 2013/163239 entitled Synthesis of Lactams to Tavares, F.X., and US 8,598,186 entitled CDK Inhibitors to Tavares, F.X. and Strum, J.C. are incorporated by reference herein in their entirety. Example 1

Synthesis of tert-butyl N- [2- [(5-bromo-2-chloro-pyrimidin-4yl)amino] ethyl] carbamate, Compound 1

Figure imgf000106_0001

To a solution of 5-bromo-2,4-dichloropyrimidine (3.2 g, 0.0135 mol) in ethanol (80 mL) was added Hunig’s base (3.0 mL) followed by the addition of a solution of N-(tert- butoxycarbonyl)-l,2-diaminoethane (2.5 g, 0.0156 mole) in ethanol (20 mL). The contents were stirred overnight for 20 hrs. The solvent was evaporated under vacuum. Ethyl acetate (200 mL) and water (100 mL) were added and the layers separated. The organic layer was dried with magnesium sulfate and then concentrated under vacuum. Column chromatography on silica gel using hexane/ethyl acetate (0- 60%) afforded tert-butyl N-[2-[(5-bromo-2-chloro-pyrimidin-4- yl)amino]ethyl]carbamate. 1HNMR (d6-DMSO) δ ppm 8.21 (s, 1H), 7.62 (brs, 1H), 7.27 (brs, 1H), 3.39 (m, 2H), 3.12 (m, 2H), 1.34 (s, 9H). LCMS (ESI) 351 (M + H).

Example 2

Synthesis of tert-butyl N-[2-[[2-chloro-5-(3,3-diethoxyprop-l-ynyl)pyrimidin-4- yl] amino] ethyl] carbamate, Compound 2

Figure imgf000106_0002

To tert-butyl N-[2-[(5-bromo-2-chloro-pyrimidin-4-yl)amino]ethyl]carbamate (1.265 g, 6 mmol) in THF (10 mL) was added the acetal (0.778 mL, 5.43 mmol), Pd(dppf)CH2Cl2 (148 g), and triethylamine (0.757 mL, 5.43 mmol). The contents were degassed and then purged with nitrogen. To this was then added Cul (29 mg). The reaction mixture was heated at reflux for 48 hrs. After cooling, the contents were filtered over CELITE™ and concentrated. Column chromatography of the resulting residue using hexane/ethyl acetate (0- 30%) afforded tert-butyl N- [2- [ [2-chloro-5 -(3 ,3 -diethoxyprop- 1 -ynyl)pyrimidin-4-yl]amino] ethyl] carbamate. 1HNMR (d6-DMSO) δ ppm 8.18 (s, 1H), 7.63 (brs, 1H), 7.40 (brs, 1H), 5.55 (s, 1H), 3.70 (m, 2H), 3.60 (m, 2H), 3.42 (m, 2H), 3.15 (m, 2H), 1.19 – 1.16 (m, 15H). LCMS (ESI) 399 (M + H).

Example 3

Synthesis of tert-butyl N-[2-[2-chloro-6-(diethoxymethyl)pyrrolo[2,3-d]pyrimidin-7- yl] ethyl] carbamate, Compound 3

Figure imgf000107_0001

To a solution of the coupled product (2.1 g, 0.00526 mole) in THF (30 mL) was added TBAF solid (7.0 g). The contents were heated to and maintained at 65 degrees for 2 hrs. Concentration followed by column chromatography using ethyl acetate/hexane (0-50%) afforded tert-butyl N-[2-[2-chloro-6-(diethoxymethyl)pyrrolo[2,3-d]pyrimidin-7-yl]ethyl]carbamate as a pale brown liquid (1.1 g). 1FiNMR (d6-DMSO) δ ppm 8.88 (s, 1H), 6.95 (brs, 1H), 6.69 (s, 1H), 5.79 (s, 1H), 4.29 (m, 2H), 3.59 (m, 4H), 3.34 (m, 1H), 3.18 (m, 1H), 1.19 (m, 9H), 1.17 (m, 6H). LCMS (ESI) 399 (M + H).

Example 4

Synthesis of tert-buty\ N-[2-(2-chloro-6-formyl-pyrrolo [2,3-d] pyrimidin-7- yl)ethyl] carbamate, Compound 4

Figure imgf000108_0001

To the acetal (900 mg) from the preceeding step was added AcOH (8.0 mL) and water

(1.0 mL). The reaction was stirred at room temperature for 16 hrs. Cone, and column chromatography over silica gel using ethyl acetate/hexanes (0- 60%) afforded tert-butyl N-[2-(2- chloro-6-formyl-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]carbamate as a foam (0.510 g). 1HNMR (d6-DMSO) δ ppm 9.98 (s, 1H), 9.18 (s, 1H), 7.66 (s, 1H), 6.80 (brs, 1H), 4.52 (m, 2H), 4.36 (m, 2H), 1.14 (s, 9H). LCMS (ESI) 325 (M + H).

Example 5

Synthesis of 7- [2-(teri-butoxycarbonylamino)ethyl] -2-chloro-pyrrolo [2,3-d] pyrimidine-6- carboxylic acid, Compound 5

Figure imgf000108_0002

To the aldehyde (0.940 g) from the preceeding step in DMF (4 mL) was added oxone (1.95 g, 1.1 eq). The contents were stirred at room temp for 7 hrs. Silica gel column chromatography using hexane/ethyl acetate (0- 100%) afforded l-\2-(tert- butoxycarbonylamino)ethyl]-2-chloro-pyrrolo[2,3-d]pyrimidine-6-carboxylic acid (0.545 g). 1HNMR (d6-DMSO) δ ppm 9.11 (s, 1H), 7.39 (s, 1H), 4.38 (m, 2H), 4.15 (m, 2H), 1.48 (m, 9H). LCMS (ESI) 341(M + H).

Example 6

Synthesis of methyl 7-[2-(teri-butoxycarbonylamino)ethyl]-2-chloro-pyrrolo[2,3- d]pyrimidine-6-carboxylate, Compound 6

Figure imgf000109_0001

To a solution of 2-chloro-7-propyl-pyrrolo[2,3-d]pyrimidine-6-carboxylic acid (0.545 g, 0.00156 mole) from the preceeding step in toluene (3.5 mL) and MeOH (1 mL) was added TMS- diazomethane (1.2 mL). After stirring overnight at room temperature, the excess of TMS- diazomethane was quenched with acetic acid (3 mL) and the reaction was concentrated under vacuum. The residue was purified by silica gel column chromatography with hexane/ethyl acetate (0- 70%) to afford methyl 7-[2-(tert-butoxycarbonylamino)ethyl]-2-chloro-pyrrolo[2,3- d]pyrimidine-6-carboxylate as an off white solid (0.52 g). 1HNMR (d6-DMSO) δ ppm 9.10 (s, 1H), 7.45 (s, 1H), 6.81 (brs, 1H) 4.60 (m, 2H), 3.91 (s, 3H), 3.29 (m, 2H), 1.18 (m, 9H) LCMS (ESI) 355 (M + H).

Example 7

Synthesis of Chloro tricyclic amide, Compound 7

Figure imgf000109_0002

To methyl 7- [2-(tert-butoxycarbonylamino)ethyl] -2-chloro-pyrrolo [2,3 -d]pyrimidine-6- carboxylate (0.50 g, 0.0014 mole) from the preceeding step in dichloromethane (2.0 mL) was added TFA (0.830 mL). The contents were stirred at room temperature for 1 hr. Concentration under vacuum afforded the crude amino ester which was suspended in toluene (5 mL) and Hunig’s base (0.5 mL). The contents were heated at reflux for 2 hrs. Concentration followed by silica gel column chromatography using hexane/ethyl acetate (0- 50%) afforded the desired chloro tricyclic amide (0.260 g). 1HNMR (d6-DMSO) δ ppm 9.08 (s, 1H), 8.48 (brs, 1H), 7.21 (s, 1H) 4.33 (m, 2H), 3.64 (m, 2H). LCMS (ESI) 223 (M + H).

Example 8

Synthesis of chloro-N-methyltricyclic amide, Compound 8

Figure imgf000110_0001

To a solution of the chloro tricycliclactam, Compound 7, (185 mg, 0.00083 mole) in DMF (2.0 mL) was added sodium hydride (55% dispersion in oil, 52 mg). After stirring for 15 mins, methyl iodide (62 μί, 1.2 eq). The contents were stirred at room temperature for 30 mins. After the addition of methanol (5 mL), sat NaHCOs was added followed by the addition of ethyl acetate. Separation of the organic layer followed by drying with magnesium sulfate and concentration under vacuum afforded the N-methylated amide in quantitative yield. 1FiNMR (d6-DMSO) δ ppm 9.05 (s, 1H), 7.17 (s, 1H) 4.38 (m, 2H), 3.80 (m, 2H), 3.05 (s, 3H). LCMS (ESI) 237 (M + H). Example 9

Synthesis of l-methyl-4-(6-nitro-3-pyridyl)piperazine, Compound 9

Figure imgf000110_0002

To 5-bromo-2-nitropyridine (4.93 g, 24.3 mmole) in DMF (20 mL) was added N- methylpiperazine (2.96 g, 1.1 eq) followed by the addition of DIPEA (4.65 mL, 26.7 mmole). The contents were heated at 90 degrees for 24 hrs. After addition of ethyl acetate (200 mL), water (100 mL) was added and the layers separated. Drying followed by concentration afforded the crude product which was purified by silica gel column chromatography using (0-10%) DCM/Methanol. 1HNMR (d6-DMSO) δ ppm 8.26 (s, 1H), 8.15 (1H, d, J = 9.3 Hz), 7.49 (1H, d, J = 9.4 Hz), 3.50 (m, 4H), 2.49 (m, 4H), 2.22 (s, 3H).

Example 10

Synthesis of 5-(4-methylpiperazin-l-yl)pyridin-2-amine, Compound 10

Figure imgf000111_0001

To l-methyl-4-(6-nitro-3-pyridyl)piperazine (3.4 g) in ethyl acetate (100 mL) and ethanol (100 mL) was added 10%> Pd/C (400 mg) and then the reaction was stirred under hydrogen (10 psi) overnight. After filtration through CELITE™, the solvents were evaporated and the crude product was purified by silica gel column chromatography using DCM/ 7N ammonia in MeOH (0- 5%) to afford 5-(4-methylpiperazin-l-yl)pyridin-2-amine (2.2 g). 1HNMR (d6-DMSO) δ ppm 7.56 (1H, d, J = 3 Hz), 7.13 (1H, m), 6.36 (1H, d, J = 8.8 Hz), 5.33 (brs, 2H), 2.88 (m, 4H), 2.47 (m, 4H), 2.16 (s, 3H).

Example 11

Synthesis of tert-butyl 4-(6-amino-3-pyridyl)piperazine-l-carboxylate, Compound 11

Figure imgf000111_0002

This compound was prepared as described in WO 2010/020675 Al .

Synthesis of Compound 89 (also referred to as Compound T)

Figure imgf000169_0002

Compound 89 was synthesized in a similar manner to that described for compound 78 and was converted to an HCl salt. 1HNMR (600 MHz, DMSO-d6) δ ppm 1.47 (br. s., 6 H) 1.72 (br. s., 2 H) 1.92 (br. s., 2 H) 2.77 (br. s., 3 H) 3.18 (br. s., 2 H) 3.46 (br. s., 2 H) 3.63 (br. s., 2 H) 3.66 (d, J=6.15 Hz, 2 H) 3.80 (br. s., 2 H) 7.25 (s, 1 H) 7.63 (br. s., 2 H) 7.94 (br. s., 1 H) 8.10 (br. s., 1 H) 8.39 (br. s., 1 H) 9.08 (br. s., 1 H) 11.59 (br. s., 1 H). LCMS (ESI) 447 (M + H)

PATENT

WO 2014144740

PATENT

WO 2016040858

Preparation of Active Compounds

Syntheses

The disclosed compounds can be made by the following general schemes:

Scheme 1

In Scheme 1, Ref-1 is WO 2010/020675 Al; Ref-2 is White, J. D.; et al. J. Org. Chem. 1995, 60, 3600; and Ref-3 Presser, A. and Hufher, A. Monatshefte fir Chemie 2004, 135, 1015.

Scheme 2

In Scheme 2, Ref-1 is WO 2010/020675 Al; Ref-4 is WO 2005/040166 Al; and Ref-5 is Schoenauer, K and Zbiral, E. Tetrahedron Letters 1983, 24, 573.

92

93 

3) Pd/C/H2 

Scheme 6

Scheme 7

NHfOH

Scheme 8

In Scheme 8, Ref-1 is WO 2010/020675 Al; Ref-2 is WO 2005/040166 Al; and Ref-3 is Schoenauer, K and Zbiral, E. Tetrahedron Letters 1983, 24, 573.

Alternatively, the lactam can be generated by reacting the carboxylic acid with a protected amine in the presence of a strong acid and a dehydrating agent, which can be together in one moiety as a strong acid anhydride. Examples of strong acid anhydrides include, but are not limited to, trifluoroacetic acid anhydride, tribromoacetic acid anhydride, trichloroacetic acid anhydride, or mixed anhydrides. The dehydrating agent can be a carbodiimide based compound such as but not limited to DCC (Ν,Ν-dicyclohexylcarbodiimide), EDC (l-ethyl-3-(3-

dimethylaminopropyl)carbodiimide or DIC (Ν,Ν-diisopropylcarbodiimide). An additional step may be necessary to take off the N-protecting group and the methodologies are known to those skilled in the art.

Alternatively, the halogen moiety bonded to the pyrimidine ring can be substituted with any leaving group that can be displaced by a primary amine, for example to create an intermediate for a final product such as Br, I, F, SMe, SO2Me, SOalkyl, SO2alkyl. See, for Exmaple PCT /US2013/037878 to Tavares.

Other amine intermediates and final amine compounds can be synthesized by those skilled in the art. It will be appreciated that the chemistry can employ reagents that comprise reactive functionalities that can be protected and de-protected and will be known to those skilled in the art at the time of the invention. See for example, Greene, T.W. and Wuts, P.G.M., Greene’s Protective Groups in Organic Synthesis, 4th edition, John Wiley and Sons.

Scheme 9

CDK4/6 Inhibitors of the present invention can be synthesized according to the generalized Scheme 9. Specific synthesis and characterization of the Substituted 2-aminopyrmidines can be found in, for instance, WO2012/061156.

Compounds T, Q, GG, and U were prepared as above and were characterized by mass spectrometry and NMR as shown below:

Compound T

1H NMR (600 MHz, DMSO- d6) ppm 1.47 (br. s., 6 H) 1.72 (br. s., 2 H) 1.92 (br. s., 2 H) 2.77 (br. s., 3 H) 3.18 (br. s., 2 H) 3.46 (br. s., 2 H) 3.63 (br. s., 2 H) 3.66 (d, J=6.15 Hz, 2 H) 3.80 (br. s., 2 H) 7.25 (s, 1 H) 7.63 (br. s., 2 H) 7.94 (br. s., 1 H) 8.10 (br. s., 1 H) 8.39 (br. s., 1 H) 9.08 (br. s., 1 H) 11.59 (br. s., 1 H). LCMS ESI (M + H) 447.

PATENT

WO-2018005865

Synthesis of N-(heteroaryl)-pyrrolo[3,2-d]pyrimidin-2-amines. The application appears to be particularly focused on methods for the preparation of trilaciclib and an analog of it. Trilaciclib is the company’s lead CDK4/6 inhibitor presently in phase II trials against small-cell lung cancer and triple negative breast cancer. Interestingly, the company is working on a second CDK4/6 inhibitor, G1T38 , which is in a phase II trial against breast cancer.

GENERAL METHODS

The structure of starting materials, intermediates, and final products was confirmed by standard analytical techniques, including NMR spectroscopy and mass spectrometry. Unless otherwise noted, reagents and solvents were used as received from commercial suppliers. Proton nuclear magnetic resonance spectra were obtained on a Bruker AVANCE 500 at 500 MHz in DMSO-dis. HPLC analyses were performed on a Waters HPLC using the below HPLC method.

HPLC Method

Column: Atlantis T3 (150 χ 4.6, 3 μιη)

Column Temperature: 40°C

Flow Rate: 1 mL/min

Detection: UV @ 275 nm

Analysis Time: 36 min

Mobile Phase A: Water (with 0.1% Trifluoroacetic Acid)

Mobile Phase B : Acetonitrile (with 0.1% Trifluoroacetic Acid)

Sample preparation: dissolve PC sample, wet or dry solid (~1 mg of active compound) in acetonitrile/water (1/1) to achieve complete dissolution.

HPLC Method Gradient

Example 1. General Routes of Synthesis

Scheme 1-1 : Starting from an appropriately substituted halo pyrimidine, compounds of the present invention can be prepared. In Step 1 the appropriately substituted halo pyrimidine is subjected to l,4-diazaspiro[5.5]undecan-3-one in the presence of base and heat to afford a substituted spirolactam. In Step 2 the appropriately substituted spirolactam is protected with a group selected from R2. In Step 3 the protected spirolactam is cyclized in the presence of base to afford a fused spirolactam. The fused spirolactam can be optionally oxidized to a sulfoxide or sulfone after Step 3, Step 4, Step 5, or Step 6. Oxidation prior to Step 3 results in undesired byproducts. In Step 4 the hydroxyl group of the fused spirolactam is converted to a leaving group.

In Step 5 the leaving group is dehydrated to afford a compound of Formula IV. In Step 6 the compound of Formula IV is optionally deprotected.

Scheme 1-2: Starting from an appropriately substituted halo pyrimidine compounds of the present invention can be prepared. In Step 1 the appropriately substituted halo pyrimidine is subjected to l,4-diazaspiro[5.5]undecan-3-one in the presence of base and heat to afford a substituted spirolactam. In Step 2 the appropriately substituted spirolactam is protected with a group selected from R2. In Step 3 the protected spirolactam is cyclized in the presence of base to afford a fused spirolactam of Formula IV. The fused spirolactam can be optionally oxidized to a sulfoxide or sulfone after Step 3 or Step 4. Oxidation prior to Step 3 results in undesired byproducts. In Step 4 the compound of Formula IV is optionally deprotected.

Scheme 1-3 : Starting from an appropriately substituted alkyl glycinate, compounds of the present invention can be prepared. In Step 1 the appropriately substituted alkyl glycinate is subjected to cyclohexanone and TMSCN in the presence of base to afford a cyano species. In Step 2 the appropriately substituted cyanospecies is reduced and subsequently cyclized to afford a compound of Formula I.

Scheme 1-4

Scheme 1-4: Starting from an appropriately substituted l-(aminomethyl)cyclohexan-l-amine, compounds of the present invention can be prepared. In Step 1 the appropriately substituted l-(aminomethyl)cyclohexan-l -amine is reductively aminated with an aldehyde. In Step 2 the appropriately substituted cyclohexane amine is optionally deprotected (i.e.: the group selected from R2 if not H is optionally replaced by H). In Step 3 the cyclohexane amine is cyclized to afford a compound of Formula I. In Step 4 the compound of Formula I is optionally protected.

1-5

Conversion

Scheme 1-5: Starting from an appropriately substituted halo pyrimidine, compounds of the present invention can be prepared. In Step 1 the appropriately substituted halo pyrimidine is subjected to l,4-diazaspiro[5.5]undecan-3-one in the presence of base and heat to afford a

substituted spirolactam. In Step 2 the protected spirolactam is cyclized in the presence of base to afford a fused spirolactam. The fused spirolactam can be optionally oxidized to a sulfoxide or sulfone after Step 2, Step 3, Step 4, or Step 5. Oxidation prior to Step 2 results in undesired byproducts. In Step 3 the hydroxyl group of the fused spirolactam is converted to a leaving group. In Step 4 the leaving group is dehydrated to afford a compound of Formula IV. In Step 5 the compound of Formula IV is optionally deprotected.

S

Scheme 1-6: Starting from an appropriately substituted halo pyrimidine compounds of the present invention can be prepared. In Step 1 the appropriately substituted halo pyrimidine is subjected to l,4-diazaspiro[5.5]undecan-3-one in the presence of base and heat to afford a substituted spirolactam. In Step 2 the protected spirolactam is cyclized in the presence of base to afford a fused spirolactam of Formula IV. The fused spirolactam can be optionally oxidized to a sulfoxide or sulfone after Step 2 or Step 3. Oxidation prior to Step 2 results in undesired byproducts. In Step 3 the compound of Formula IV is optionally deprotected.

Scheme 1-7: Starting from compound of Formula IV a CDK4/6 inhibitor can be prepared. In Step 1 a heteroaryl amine is subjected to a base and a compound of Formula IV is added slowly under chilled conditions to afford a nucleophilic substitution reaction. The compound of Formula IV can previously be prepared as described in the schemes herein.

Example 2. Representative Routes of Synthesis

Scheme 2-1

quant, yield 2 steps

isolated

70% yield 2 steps 75% yield 95% yield

isolated isolated isolated

Scheme 2-1 : An ester route is one embodiment, of the present invention. Ideally, the best synthesis scheme would afford crystalline intermediates to provide material of consistent purity without column chromatography, and high yielding steps while using safe and cost effective reagents when possible.

The first step in the ester route is a SNAr nucleophilic substitution of CI group in commercially available ester 3 using spirolactam 4. Due to low reactivity of 4, a reaction temperature of 85-95 °C was required. Because of the temperature requirements, DIPEA and dimethylacetamide were selected as the base and solvent, respectively. The reaction follows second-order kinetics and usually stalls after -85% conversion. Therefore, the reaction was typically stopped after 60 hours by first cooling it to room temperature at which point solid formation was observed. The mixture was then partitioned between MTBE and water and product was filtered with excellent purity with -53% yield of the desired product 5. The obtained

compound 5 was protected with a Boc group using Boc anhydride and DMAP as the catalyst and dichloromethane as the solvent. The intermediate 6 was obtained in a quantitative yield. Due to the semi-solid nature of compound 6, the material was taken to the next step without further purification. The Dieckmann condensation was initially performed with strong bases such as LiHMDS and tBuOK. A similar result to the aldehyde route (Scheme 2-2) was obtained: a partial deprotection of Boc group was observed that required column chromatography. However, the best results were obtained when DBU was used as base and THF as solvent. The reaction outcome was complete, clean conversion of 6 to 7. Moreover, the product crystallized from the reaction mixture upon seeding, and a quantitative yield was obtained for the two steps.

The hydroxyl group of 7 was removed via a two-step procedure. First, compound 7 was converted completely into triflate 8 using triflic anhydride and triethylamine in dichloromethane. The reaction was found to proceed well at 0°C. Due to the potential instability of the triflate intermediate, it was not isolated. It was immediately taken to the next step and reduced with triethylsilane and palladium tetrakis to afford the product 9 after ethyl acetate crystallization in -70% yield. The Boc group of 9 was removed using trifluoroacetic acid in dichloromethane to afford 10. Intermediate 10 was converted into the final sulfone 11 using Oxone™ in acetonitrile/water solvent system.

The obtained sulfone 11 was use-tested in the coupling step and was found to perform well. In conclusion, the route to sulfone 11 was developed which eliminated the use of column chromatography with good to excellent yields on all steps.

Scheme 2-2


Molecular Weight: 421 

Scheme 2-2: The first step of Scheme 2-2 consistently afforded product 13 contaminated with one major impurity found in substantial amount. Thorough evaluation of the reaction impurity profile by LC-MS and 2D MR was performed, which showed the impurity was structurally the condensation of two aldehyde 12 molecules and one molecule of lactam 4. Therefore, column chromatography was required to purify compound 13, which consistently resulted in a modest 30% yield. A solvent screen revealed that sec-butanol, amyl alcohol, dioxane, and tert-butanol can all be used in the reaction but a similar conversion was observed in each case. However, tert-butanol provided the cleanest reaction profile, so it was selected as a solvent for the reaction. Assessing the impact of varying the stoichiometric ratio of 4 and 12 on the reaction outcome was also investigated. The reaction was performed with 4 equivalents of amine 4 in an attempt to disrupt the 2: 1 aldehyde/amine composition of the impurity. The result was only a marginal increase in product 13 formation. The temperature impact on the reaction outcome was evaluated next. The coupling of aldehyde 12 and 4 was investigated at two different temperatures: 50 °C and 40 °C with 1 : 1 ratio of aldehyde/amine. Reactions were checked at 2 and 4 hours and then every 12 hours. The reaction progress was slow at 50°C and was accompanied by growth of other impurities. The reaction at 40°C was much cleaner; however the conversion was lower in the same time period. The mode of addition of the reagents was investigated as well at 80°C with a slow addition (over 6 hours) of either aldehyde 12 or amine 4 to the reaction mixture. The product distribution did not change and an about 1 to 1 ratio was observed between product and impurity when amine 4 was added slowly to the reaction mixture containing aldehyde 12 and

DIPEA at reflux. The product distribution did change when aldehyde 12 was added slowly to the mixture of amine 4 and DIPEA. However, the major product of the reaction was the undesired impurity. Other organic bases were tried as well as different ratios of DIPEA. No product was observed when potassium carbonate was used as a base. The results of the experiments are presented in Table 1 below.

Table 1

Compound 13 was successfully formed in three cases: triethylamine, 2,6-lutidine and DIPEA, with the DIPEA result being the best. The use of Boc protected spirolactam 4 had no effect on the impurity formation as well. Its utilization was speculated to be beneficial in performing the coupling step together with the following step, preparation of compound 14.

The major impurity formed during Step 1 of Scheme 2-2 is:

Chemical Formula:€2)Η(¾ 62ί>2

Molecular Weight: 527.4903

The second step (Boc protection of the free lactam) proceeded well using DMAP as a catalyst in dichloromethane at room temperature. The product 14 is a thick oil, and, therefore, cannot be purified by crystallization. The Boc protected intermediate 14 was cyclized successfully into the desired pentacyclic structure 10 upon treatment with a strong base such as LiHMDS or tBuOK. Surprisingly, the Boc group was partially removed during the reaction. The level of deprotection was independent from the internal reaction temperature and was positively correlated with excess of base used. Therefore the mixture of the desired product 10 and 10-Boc compound was treated with acid to completely deprotect Boc group. The conversion of methyl sulfide into the final sulfone 11 was carried out with Oxone™. Initially a mixture of methanol and water was used for the reaction. As the result, a partial displacement of sulfone by methoxy group was detected. The methanol was replaced with acetonitrile and the sulfone displacement was eliminated.

In summary, the ester route (Scheme 2-1) is preferred because:

1. Formation of the impurity during the first step of Scheme 2-2 was unavoidable and resulted in yields of < 35%.

2. Column purification was required to isolate intermediate 14.

3. The aldehyde starting material was not commercially available and required two synthetic steps from the corresponding ester.

Scheme 2-3 : Starting with cyclohexanone, compounds of the present invention can be prepared. In Step 1 the methyl glycinate is subjected to cyclohexanone and TMSCN in the presence of tri ethyl amine in DCM to afford 15. In Step 2 15 hydrogenated with hydrogen gas in the presence of catalytic platinum oxide and subsequently undergoes an intramolecular cyclization to afford compound 16 which is used in the schemes above.

Scheme 2-4: Starting with compound 17, compounds of the present invention can be prepared. In Step 1 compound 17 is subjected to ethyl 2-oxoacetate in the presence platinum on carbon and hydrogen gas to afford compound 18. In Step 2 compound 18 is Boc-deprotected with hydrochloric acid. In Step 3 compound 18 is cyclized to afford compound 16 which is used in the schemes above.

Scheme 2-5

11 19

Scheme 2-5: Starting from compound 11 the CDK 4/6 inhibitor 19 can be prepared. In Step 1 5-(4-methylpiperazin-l-yl)pyridin-2-amine is subjected to LiHMDS and compound 11 is added slowly under chilled conditions to afford a nucleophilic substitution reaction and compound 19. Compound 11 can be prepared as described in the schemes herein.

Scheme 2-6: Starting from compound 11 the CDK 4/6 inhibitor 20 can be prepared. In Step 1 5-(4-isopropylpiperazin-l-yl)pyridin-2-amine is subjected to LiHMDS and compound 11 is added slowly under chilled conditions to afford a nucleophilic substitution reaction and compound 20. Compound 11 can be prepared as described in the schemes herein.

Preparation of Compound 5:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet, and reflux condenser was charged with ethyl 4-chloro-2-(methylthio)pyrimidine-5-carboxylate 3 (49.2 g, 0.21 mol, 1.00 equiv.), spirolactam 4 (39.2 g, 0.23 mol, 1.10 equiv.), DIPEA (54.7 g, 0.42 mol, 2.00 equiv.), and DMAc (147.6 mL, 3 vol). The batch was heated to 90-95 °C, and after 60 h, IPC confirmed -14% (AUC) of ethyl 4-chloro-2-(methylthio)pyrimidine-5-carboxylate remained. The batch was cooled to RT, and precipitate formation was observed. The suspension was diluted with MTBE (100 mL, 2 vol) and water (442 mL, 9 vol) and stirred for 2 h at RT. The product was isolated by vacuum filtration and washed with MTBE (49 mL, 1 vol). The solid cake was conditioned for 1 h and dried under vacuum at 40 °C for 16 h to afford compound 5 [41.0 g, 53% yield] as an off-white solid with a purity of >99% AUC. ¾ MR (CDCh): δ 8.76 (d, J = 2.0 Hz, 1H), 6.51-6.29 (br, 1H), 4.33 (q, J = 7.0 Hz, 2H), 3.78 (s, 2H), 3.58 (s, 2H), 2.92 (s, 2H), 2.53 (s, 3H), 1.63-1.37 (m, 12H). LCMS (ESI, m/z = 365.3 [M+H]).

Preparation of Compound 6:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged with 5 [41.0 g, 0.11 mol, 1.00 equiv.], Boc-anhydride (36.8 g, 0.17 mol, 1.50 equiv.), DMAP (1.37 g, 0.01 mol, 0.10 equiv.), and dichloromethane (287 mL, 7 vol). The batch was stirred for 3 h at RT. IPC confirmed no starting material remained (AUC). The batch was concentrated into a residue under reduced pressure and taken to the next step (a quantitative yield is assumed for this step). An aliquot (200 mg) was purified by column chromatography (heptanes/ethyl acetate 0 to 100%) to afford compound 6. 1H MR (CDCh): δ 8.64 (s, 1H), 4.31 (q, J = 7.0 Hz, 2H), 4.07 (s, 2H), 3.83 (S, 2H), 3.15 (m, 2H), 2.56 (s, 3H), 172 (m, 3H), 1.59 (m, 15H), 1.42 (t, J= 7.0 Hz, 3H). LCMS (ESI, m/z = 465.2 [M+H]).

Preparation of Compound 7:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged with compound 6 [residue from a previous step, quantitative yield assumed, 52.2 g, 0.11 mol, 1.00 equiv.], and THF (261 mL, 5 vol). The batch was cooled to 0°C and 1,8-diazabicyclo[5.4.0]un-dec-7-ene (17.1 g, 0.11 mmol, 1.00 equiv.) was added keeping the internal temperature in 0-10°C range. After the addition was complete, the cooling bath was removed and the reaction mixture was allowed to warm up to RT and after 2 h, IPC confirmed no starting material remained. The batch was seeded with the product (1.0 g) and was cooled to 0°C. The slurry was stirred at 0°C for 2 h. The product was isolated by vacuum filtration and washed with cold (0°C) THF (50 mL, 1 vol). The solid cake was conditioned for 1 h and dried under vacuum at 40°C for 16 h to afford 7 [47 g, quantitative yield] as a light orange solid with a purity of >99% AUC. The color of the product changed into yellow once the batch was exposed to air for an extended period of time (~ 1 day). Material was isolated with substantial amount DBU, according to proton NMR. However, it did not interfere with the next step. 1H MR (CDCh): δ 8.71 (s, 1H), 4.03 (s, 2H), 2.57 (s, 3H), 1.85 (m, 10H), 1.51 (s, 9H). LCMS (ESI, m/z = 419.2 [M+H]).

Preparation of Compound 8:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged with 7 [40.8 g, 0.10 mol, 1.00 equiv.], triethylamine (31.5 g, 0.31 mol, 3.20 equiv.), and dichloromethane (408 mL, 10 vol). The batch was purged with N2 for 15 min and was cooled to 0°C. Triflic anhydride (44.0 g, 0.16 mol, 1.60 equiv.) was added keeping the

internal temperature in 0-10°C range. The batch was stirred at 0°C and after 3 h, IPC confirmed -7.0% (AUC) of 7 remained. [It was speculated that the product was hydrolyzing back into starting material during the analysis.] Once the reaction was deemed complete, the batch was transferred to a 1 L, separatory funnel and was washed with 50% saturated sodium bicarbonate (200 mL, 5 vol). [It was prepared by mixing saturated sodium bicarbonate (100 mL) with water (100 mL)).] The aqueous layer was separated and was extracted with DCM (2×40 mL, 1 vol). The organic layers were combined and concentrated into a residue under reduced pressure and taken to the next step. LCMS (ESI, m/z = 551.6 [M+H]).

Preparation of Compound 9:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged with compound 8 [residue from a previous step, quantitative yield assumed, 53.7 g, 0.10 mol, 1.00 equiv.], and THF (110 mL, 2 vol). The solvent was removed under vacuum distillation and the procedure was repeated two times. The flask was charged with triethylsilane (22.7 g, 0.20 mol, 2.00 equiv.), and DMF (268 mL, 5 vol). The batch was degassed by five cycles of evacuation, followed by backfilling with nitrogen. The flask was charged with tetrakis(triphenylphosphine)palladium(0) (11.3 g, 0.01 mol, 0.1 equiv.). The batch was heated to 45-50°C, and after 14 h, IPC confirmed no starting material remained. The batch was transferred to a 500 mL, separatory funnel while still warm. The reaction was partitioned between water (5 vol) and ethyl acetate (5 vol). The aqueous layer was extracted with ethyl acetate (3 x3 vol). The organic layers were combined and concentrated down to 2 volumes. The precipitate was filtered and washed with ethyl acetate (2x 1 vol). The solid cake was conditioned for 1 h and dried under vacuum at 40°C for 16 h to afford 9 [27.5 g, 70% yield] as a yellow solid with a purity of -98% AUC. Proton NMR showed some triphenylphosphine oxide present. ¾ NMR (DMSO-i¾):5 9.01 (s, 1H), 7.40 (s, 1H), 4.30 (s, 2H), 2.58 (m, 2H), 2.58 (s, 3H), 1.81 (m, 5H), 1.51 (s, 9H). LCMS (ESI, m/z = 403.4 [M+H]).

Preparation of Compound 10 from the Scheme 2-1 route:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged 9 (12.8 g, 31.8 mmol, 1.00 equiv.) and dichloromethane (64 mL, 5 vol). Trifluoroacetic acid (18.2 g, 159 mmol, 5.00 equiv.) was added over 20 min and the solution was stirred for 2 h at RT. IPC confirmed reaction was complete. The batch was transferred to a 500 mL, separatory funnel and washed with saturated sodium bicarbonate (200 mL). The aqueous layer was extracted with dichlorom ethane (3 x3 vol). The organic layers were combined and concentrated down to 1 volume. The precipitate was filtered and conditioned for 1 h and dried under vacuum at 40 °C for 16 h to afford 9 [6.72 g, 70% yield] as an off-white solid with a purity of 99.1% AUC. ¾ NMR (DMSO-dis): δ 8.95 (s, 1H), 8.32 (s, 1H), 7.15 (s, 1H), 3.68 (d, J = 1.0 Hz, 2H), 2.86 (m, 2H), 2.57 (s, 3H), 1.92 (m, 2H), 1.73 (m, 3H), 1.39 (m, 3H). LCMS, ESI, m/z = 303.2 [M+H]).

Preparation of Compound 10 from Scheme 2-2 route:

A 50 mL, three-neck flask equipped with a magnetic stirring bar, thermocouple, N2 inlet was charged 14 (680 mg, 1.62 mmol, 1.00 equiv.) and THF (6.8 mL, 10 vol). A I M solution of potassium tert-butoxide (3.2 mL, 3.24 mmol, 2.00 equiv.) in THF was added over 10 min and the solution was stirred for 2 h at RT. IPC confirmed reaction was complete. The batch was acidified with 4 N hydrogen chloride solution in dioxane (2.4 mL, 9.72 mmol, 6.00 equiv.) and stirred for additional 1 h. The batch was transferred to a 500 mL, separatory funnel and washed with saturated sodium bicarbonate (100 mL). The aqueous layer was extracted with ethyl acetate (3 x20 vol). The organic layers were combined and concentrated down to 3volumes and product precipitated. The precipitate was filtered and conditioned for 1 h and dried under vacuum at 40 °C for 16 h to afford 9 [489 mg, quantitative yield] as an off-white solid.

Preparation of Compound 11 :

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged with 10 (9.00 g, 29.8 mmol, 1.00 equiv.), and acetonitrile (180 mL, 20 vol). A solution of Oxone™ (45.9 g, 0.15 mol, 5.00 equiv.) in water (180 mL, 20 vol) was added to the batch over 20 min. The batch was stirred for 2 h and IPC confirmed the reaction was complete. The batch was concentrated down to ½ of the original volume and was extracted with dichloromethane DCM (4x 10 vol). The organic layers were combined; polish filtered and concentrated down to -10 vol of DCM. The product was slowly crystallized out by addition of heptanes (-30 vol). The mixture was cooled to 0°C and the product was filtered and dried under vacuum at 40 °C for 16 h to afford 11 [9.45 g, 95% yield] as an off-white solid with a purity of >99% AUC. ¾ NMR (CDCb): 5 9.24 (s, 1H), 7.78 (s, 1H), 7.46 (s, 1H), 3.89 (d, J= 2.0 Hz, 2H), 3.43 (s, 3H), 2.98 (m, 2H), 2.10 (m, 2H), 1.86 (m, 3H), 1.50 (m, 3H). LCMS (ESI, m/z = 335.2 [M+H]).

Preparation of Compound 13:

A 250 mL, single-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet, and reflux condenser was charged with 4-chloro-2-(methylthio)pyrimidine-5-carbaldehyde (2.00 g, 10.6 mmol, 1.00 equiv.), spirolactam 4 (1.96 g, 11.7 mmol, 1.10 equiv.), DIPEA (2.74 g, 21.2 mmol, 2.00 equiv.), and fert-butanol (20 mL, 10 vol). The batch was heated to 80-85 °C, and after 24 h, IPC confirmed no aldehyde 12 remained. The batch was cool to RT and concentrated into a residue, which was loaded on silica gel column. The product was eluted with ethyl acetate/heptanes (0% to 100%). The product containing fractions were pulled out and concentrated to afford 13 [0.98 g, 29% yield] as an off-white solid.

Preparation of Compound 14:

A 500 mL, three-neck flask equipped with a mechanical overhead stirrer, thermocouple, N2 inlet was charged with 13 [0.98 g, 3.00 mmol, 1.00 equiv.], Boc-anhydride (4.90 g, 21.5 mmol, 7.00 equiv.), DMAP (36 mg, 0.30 mmol, 0.10 equiv.), and dichloromethane (7 mL, 7 vol). The batch was stirred for 3 h at RT. IPC confirmed no starting material remained. The batch was cool to RT and concentrated into a residue, which was loaded on silica gel column. The product was eluted with ethyl acetate/heptanes (0% to 100%). The product containing fractions were pulled out and concentrated to afford 14 [0.98 g, 29% yield] as an off-white solid.

Preparation of Compound 15:

To a suspension of methyl glycinate (500 g, 3.98 mol, 1 eq) in DCM (10 L) was added

TEA dropwise at rt under nitrogen atmosphere, followed by the addition of cyclohexanone (781 g, 7.96 mol, 2 eq) dropwise over 15 min. To the resulting mixture was added TMSCN (591 g, 5.97 mol, 1.5 eq) dropwise over 1 hour while maintaining the internal reaction temperature below 35

°C. After stirred at rt for 2 hrs, the suspension became a clear solution. The progress of the reaction was monitored by H- MR.

When the methyl glycinate was consumed completely as indicated by H-NMR analysis, the reaction was quenched by water (5 L). The layers were separated. The aqueous layer was extracted with DCM (1 L). The combined organic phase was washed with water (5 L X 2) and

dried over Na2S04 (1.5 Kg). After filtration and concentration, 1.24 Kg of crude 15 was obtained as oil.

The crude 15 was dissolved in IPA (4 L). The solution was treated with HC1/IPA solution (4.4 mol/L, 1.1L) at RT. A large amount of solid was precipitated during the addition. The resulting suspension was stirred for 2 hrs. The solid product was collected by vacuum filtration and rinsed with MTBE (800 mL). 819 g of pure 15 was obtained as a white solid. The yield was 88.4%. ¾- MR (300 MHz, CD3OD) 4.20 (s, 2H), 3.88 (s, 3H), 2.30-2.40 (d, J = 12 Hz, 2H), 1.95-2.02 (d, J = 12 Hz, 2H), 1.55-1.85 (m, 5H), 1.20-1.40 (m, 1H).

Preparation of Compound 16:

To a solution of 15 (10 g, 43 mmol) in MeOH (100 mL) was added methanolic hydrochloride solution (2 .44 mol/L, 35.3 mL, 2 eq) and Pt02 (0.5 g, 5 wt %). The reaction suspension was stirred with hydrogen bubble at 40 °C for 6 hours. H- MR analysis showed consumption of 15. To the reaction mixture was added K2CO3 (15 g, 108 mmol, 2.5 eq) and the mixture was stirred for 3 hrs. The suspension was filtered and the filtrate was concentrated to dryness. The residual oil was diluted with DCM (100 mL) and resulting suspension was stirred for 3 hrs. After filtration, the filtrate was concentrated to provide crude 16 (6.6 g) as an oil. The crude 16 was diluted with EtOAc/hexane (1 : 1, 18 mL) at rt for 2 hrs. After filtration, 16 (4 g) was isolated. The obtained 16 was dissolved in DCM (16.7 mL) and hexane (100 mL) was added dropwise to precipitate the product. After further stirred for 1 h, 2.8 g of the pure 16 was isolated as a white solid. The yield was 39%. HPLC purity was 98.3%; MS (ESI): 169.2 (MH+); 1 H-NMR (300 MHz, D2O) 3.23 (s, 3H), 3.07 (s, 3H), 1.37-1.49 (m, 10H).

Preparation of compound 19:

5-(4-methylpiperazin-l-yl)pyridin-2-amine (803.1 g; 3.0 equivalents based on sulfone 11) was charged to a 22 L flask. The flask was blanketed with N2 and anhydrous THF added (12.4 kg). The resulting black-purple solution was cooled in an ice bath to < 5°C. 1M LiHMDS (4.7 L; 1.2 equivalents based on sulfone 11) was added via an addition funnel in three equal additions to keep the temperature below 10°C. Upon the completion of the addition, the reaction mixture was warmed to 16°C. The sulfone 11 (455.1 g; 1.00 equivalents) was added in five additions. Reaction proceeded until HPLC analysis of an IPC sample indicated less than 3% of sulfone 11 remained.

To quench the reaction, the contents of the 22L flask were transferred to a 100 L flask containing water. After stirring for 30 minutes at <30°C, the crude product was collected by filtration, washed with water and dried to afford 19 (387 g, 99.1% purity, 63.7% yield).

Preparation of compound 20:

5-(4-isopropylpiperazin-l-yl)pyridin-2-amine (1976.2 g; 3.0 equivalents based on sulfone 11) was charged to a 50 L flask. The flask was blanketed with N2 and anhydrous THF added (10.7 kg). The resulting black-purple solution was cooled in an ice bath to < 5°C. 1M LiHMDS (9.6 kg; 3.6 equivalents based on sulfone) was added via an addition funnel at a rate to keep the temperature below 10°C. Upon the completion of the addition, the reaction mixture was warmed to 16°C over 120 minutes by removing the ice bath. The sulfone (1000 g; 1.00 mol) was added in five additions. The reaction proceeded until HPLC analysis of an IPC sample indicated less than 1% of sulfone 11 remained. After completion of the reaction, ammonium chloride was added to the reaction mixture. The mixture stirred at < 32°C for at least 30 minutes and the solids collected by filtration to afford 20 (900 g, 99.1% purity, 64.2% yield).

Alternate Route to Spirolactam via cyclohexanone:

Scheme 2-7

26

In one embodiment the spirolactam is made via the synthetic scheme above. By reducing the nitrile group before addition of the glycinate group the reaction sequence proceeds in higher yield. The chemistry used in Step 1 is described in the literature (J. Org. Chem. 2005, 70,8027-8034), and was performed on a kilogram scale. The chemistry to convert Compound 24 into the

spirolactam was also demonstrated on kilogram scale. The Boc protection of Compound 23, is carried out at -70°C in order to limit formation of the di-Boc protected product. Experimental details of a 200 g pilot run are described below.

Step 1

200 g of cyclohexanone 21 was converted to 22 using Ti(Oi-Pr)4 /TMSCN/NH3. After work-up, 213 g of 22 was obtained. The H- MR was clean. The yield was 84%. The titanium salts were removed by vacuum filtration. In one embodiment, the titanium salts are removed by centrifugation or Celite filtration.

Step 2

190 g of 22 was mixed with LAH (2 eq) in MTBE for 30 minutes at 45°C. After work-up, 148 g of crude 23 was obtained.

Step 3

136 g of the crude 23 from step 2 was converted to 24 with 0.9 eq of B0C2O at -70°C. The reaction was completed and worked up. After concentration, 188 g of 24 was obtained. The yield was 86%. The H-NMR and C-NMR spectra confirmed that the compound was pure.

Step 4

188 g of 24 was subjected to methyl 2-bromoacetate and K2CO3 in acetonitrile to afford 25. 247 g of crude 25 was obtained.

Step 5

247 g of 25 was subjected to TFA in DCE heated to reflux to afford 26. After work-up, 112 g of 6 as TFA salt was obtained. H- MR was clean.

Step 6

26 27

Compound 26 was stirred in EtOH in the presence at room temperature overnight to afford 27. In one embodiment DCM is used as the solvent instead of EtOH.

Example 3. Purge of residual palladium from Step 5 Scheme 2-1:

Since palladium was used in Step 5 of Scheme 2-1, the levels of residual Pd present in the subsequent synthetic steps was determined. Table 2 below and Figure 3 show the palladium levels in the isolated solids.

Table 2

The material after Step 5 was isolated containing 1.47% (14700 ppm) of residual palladium. This data represents the highest level of palladium in the worst case scenario. The workup conditions of the latter steps purged nearly all of the palladium and the final product, 19 bis HC1 salt, contained 14 ppm of Pd, which is below the standard 20 ppm guidline. The Pd levels will likely be even lower once the catal st loading is optimized in Step 5.

19

The process developed in this route was a significant improvement over the one used for the first generation synthesis. Overall, the scheme consists of seven steps with five isolations, all by crystallization. No silica column chromatography is employed in the synthesis, which makes the process highly scalable. The process workup conditions can successfully purge the 1.47% of residual palladium after step 5 of Scheme 2-1.

Patent ID

Patent Title

Submitted Date

Granted Date

US8829012 CDK inhibitors
2014-01-23
2014-09-09
US8598197 CDK inhibitors
2013-04-24
2013-12-03
US8598186 CDK inhibitors
2013-04-24
2013-12-03
US8691830 CDK inhibitors
2013-04-24
2014-04-08
US2014274896 Transient Protection of Hematopoietic Stem and Progenitor Cells Against Ionizing Radiation
2014-03-14
2014-09-18
Patent ID

Patent Title

Submitted Date

Granted Date

US2015297607 Tricyclic Lactams for Use in the Protection of Normal Cells During Chemotherapy
2015-04-17
2015-10-22
US2015297608 Tricyclic Lactams for Use as Anti-Neoplastic and Anti-Proliferative Agents
2015-04-17
2015-10-22
US9487530 Transient Protection of Normal Cells During Chemotherapy
2014-03-14
2014-09-18
US2017057971 CDK Inhibitors
2016-11-10
US2017037051 TRANSIENT PROTECTION OF NORMAL CELLS DURING CHEMOTHERAPY
2016-10-07
Patent ID

Patent Title

Submitted Date

Granted Date

US2017100405 HSPC-Sparing Treatments for RB-Positive Abnormal Cellular Proliferation
2016-12-21
US2017065597 Transient Protection of Normal Cells During Chemotherapy
2016-11-03
US2016310499 Highly Active Anti-Neoplastic and Anti-Proliferative Agents
2016-07-01
US2016220569 CDK4/6 Inhibitor Dosage Formulations For The Protection Of Hematopoietic Stem And Progenitor Cells During Chemotherapy
2016-02-03
2016-08-04
US2015297606 Tricyclic Lactams for Use in the Protection of Hematopoietic Stem and Progenitor Cells Against Ionizing Radiation
2015-04-17
2015-10-22
Patent ID

Patent Title

Submitted Date

Granted Date

US9717735 Tricyclic Lactams for Use in HSPC-Sparing Treatments for RB-Positive Abnormal Cellular Proliferation
2015-04-17
2015-10-22
US9527857 HSPC-Sparing Treatments for RB-Positive Abnormal Cellular Proliferation
2014-03-14
2014-09-18
US2014271460 Highly Active Anti-Neoplastic and Anti-Proliferative Agents
2014-03-14
2014-09-18
US2017182043 Anti-Neoplastic Combinations and Dosing Regimens using CDK4/6 Inhibitor Compounds to Treat RB-Positive Tumors
2017-03-13
US2017246171 Treatment Of RB-Negative Tumors Using Topoisomerase Inhibitors In Combination With Cyclin Dependent Kinase 4/6 Inhibitors
2017-03-13

///////////////TRILACICLIB, G1T28, G1T 28, SHR 6390, PHASE 2, G1 Therapeutics, Inc.

CN1CCN(CC1)C2=CN=C(C=C2)NC3=NC=C4C=C5C(=O)NCC6(N5C4=N3)CCCCC6

BMS-986020


imgImage result for BMS-986020

BMS-986020

AM-152; BMS-986020; BMS-986202

cas 1257213-50-5
Chemical Formula: C29H26N2O5
Molecular Weight: 482.536

(R)-1-(4′-(3-methyl-4-(((1-phenylethoxy)carbonyl)amino)isoxazol-5-yl)-[1,1′-biphenyl]-4-yl)cyclopropane-1-carboxylic acid

Cyclopropanecarboxylic acid, 1-(4′-(3-methyl-4-((((1R)-1-phenylethoxy)carbonyl)amino)-5-isoxazolyl)(1,1′-biphenyl)-4-yl)-

1-(4′-(3-Methyl-4-(((((R)-1-phenylethyl)oxy)carbonyl)amino)isoxazol-5-yl)biphenyl-4-yl)cyclopropanecarboxylic acid

UNII: 38CTP01B4L

For treatment for pulmonary fibrosis, phase 2, The lysophosphatidic acid receptor, LPA1, has been implicated as a therapeutic target for fibrotic disorders

Lysophospholipids (LPs), including lysophosphatidic acid (LPA), sphingosine 1-phospate (S1P), lysophosphatidylinositol (LPI), and lysophosphatidylserine (LysoPS), are bioactive lipids that transduce signals through their specific cell-surface G protein-coupled receptors, LPA1-6, S1P1-5, LPI1, and LysoPS1-3, respectively. These LPs and their receptors have been implicated in both physiological and pathophysiological processes such as autoimmune diseases, neurodegenerative diseases, fibrosis, pain, cancer, inflammation, metabolic syndrome, bone formation, fertility, organismal development, and other effects on most organ systems.

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  • Originator Amira Pharmaceuticals
  • DeveloperB ristol-Myers Squibb; Duke University
  • Class Antifibrotics; Azabicyclo compounds; Carboxylic acids; Small molecules; Tetrazoles
  • Mechanism of Action Lysophosphatidic acid receptor antagonists
  • Orphan Drug Status Yes – Fibrosis
  • Phase II Idiopathic pulmonary fibrosis
  • Phase IPsoriasis

Most Recent Events

  • 05 May 2016 Bristol-Myers Squibb plans a phase I trial for Psoriasis in Australia (PO, Capsule, Liquid) (NCT02763969)
  • 01 May 2016 Preclinical trials in Psoriasis in USA (PO) before May 2016
  • 14 Mar 2016 Bristol-Myers Squibb withdraws a phase II trial for Systemic scleroderma in USA, Canada, Poland and United Kingdom (PO) (NCT02588625)

BMS-986020, also known as AM152 and AP-3152 free acid, is a potent and selective LPA1 antagonist. BMS-986020 is in Phase 2 clinical development for treating idiopathic pulmonary fibrosis. BMS-986020 selectively inhibits the LPA receptor, which is involved in binding of the signaling molecule lysophosphatidic acid, which in turn is involved in a host of diverse biological functions like cell proliferation, platelet aggregation, smooth muscle contraction, chemotaxis, and tumor cell invasion, among others

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PRODUCT PATENT

GB 2470833, US 20100311799, WO 2010141761

Hutchinson, John Howard; Seiders, Thomas Jon; Wang, Bowei; Arruda, Jeannie M.; Roppe, Jeffrey Roger; Parr, Timothy

Assignee: Amira Pharmaceuticals Inc, USA

Image result for Hutchinson, John Howard AMIRA

John Hutchinson

PATENTS

WO 2011159632

WO 2011159635

PATENT

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2013025733&recNum=1&maxRec=&office=&prevFilter=&sortOption=&queryString=&tab=PCTDescription

WO 2013025733

Synthesis of Compound 74

Synthetic Route (Scheme XLV)

Compound 74 Compound 74a

[0562] Compound XLV-1 was prepared by the same method as described in the synthesis of compound 1-4 (Scheme 1-A).

[0563] To a solution of compound XLV-1 (8 g, 28.08 mmol) in dry toluene (150 mL) was added compound XLV-2 (1.58 g, 10.1 mmol), triethylamine (8.0 mL) and DPPA (9.2 g, 33.6 mmol). The reaction mixture was heated to 80 °C for 3 hours. The mixture was diluted with EtOAc (50 mL), washed with brine, dried over Na2S04, filtered and concentrated. The residue was purified by column chromatography (PE/EA = 10 IX) to give compound XLV-3 (9.4 g, yield: 83 %). MS (ESI) m/z (M+H)+402.0.

[0564] Compound 74 was prepared analogously to the procedure described in the synthesis of Compound 28 and was carried through without further characterization.

[0565] Compound 74a was prepared analogously to the procedure described in the synthesis of Compound 44a. Compound 74a: 1HNMR (DMSO-d6 400MHz) δ 7.81 (d, J = 8.4 Hz, 2H), 7.41 (d, J = 8.4 Hz, 2H), 7.52 (d, J = 8.4 Hz, 2H), 7.29-7.32 (m, 7 H), 5.78 (q, 1 H), 2.15 (s, 3 H), 1.52 (d, J = 6.0 Hz, 3H), 1.28 (br, 2 H), 0.74 (br, 2 H). MS (ESI) m/z (M+H)+ 483.1.

Paper

Development of a Concise Multikilogram Synthesis of LPA-1 Antagonist BMS-986020 via a Tandem Borylation–Suzuki Procedure

Chemical and Synthetic Development, Bristol-Myers Squibb Company, One Squibb Drive, New Brunswick, New Jersey 08903, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.7b00301

http://pubs.acs.org/doi/10.1021/acs.oprd.7b00301

Abstract Image

The process development for the synthesis of BMS-986020 (1) via a palladium catalyzed tandem borylation/Suzuki reaction is described. Evaluation of conditions culminated in an efficient borylation procedure using tetrahydroxydiboron followed by a tandem Suzuki reaction employing the same commercially available palladium catalyst for both steps. This methodology addressed shortcomings of early synthetic routes and was ultimately used for the multikilogram scale synthesis of the active pharmaceutical ingredient 1. Further evaluation of the borylation reaction showed useful reactivity with a range of substituted aryl bromides and iodides as coupling partners. These findings represent a practical, efficient, mild, and scalable method for borylation.

1H NMR (500 MHz, DMSO-d6) δ 1.19 (dd, J = 6.8, 3.8 Hz, 2H), 1.50 (dd, J = 6.8, 3.8 Hz, 2H), 1.56 (br s, 3H), 2.14 (br s, 3H), 5.78 (br s, 1H), 6.9–7.45 (br, 5H), 7.45 (br d, J = 8.3 Hz, 2H), 7.65 (d, J = 8.3 Hz, 2H), 7.79 (br d, 2H), 7.82 (br d, 2H), 8.87 (br s, 0.8H), 9.29 (s, 0.2H), 12.39 (br s, 1H). 13C NMR (126 MHz, DMSO-d6) δ 9.2, 15.8, 22.4, 28.3, 72.8, 113.8, 125.4, 125.6, 126.2, 126.3, 127.1, 127.7, 128.4, 130.9, 137.4, 140.0, 141.5, 142.2, 154.4, 159.6, 160.8, 175.2. HRMS (ESI+) Calculated M + H 483.19145, found 483.19095.

REFERENCES

1: Kihara Y, Mizuno H, Chun J. Lysophospholipid receptors in drug discovery. Exp
Cell Res. 2015 May 1;333(2):171-7. doi: 10.1016/j.yexcr.2014.11.020. Epub 2014
Dec 8. Review. PubMed PMID: 25499971; PubMed Central PMCID: PMC4408218.

//////////////BMS-986020,  AM 152, BMS 986020, BMS 986202, Orphan Drug, BMS, Amira Pharmaceuticals, Bristol-Myers Squibb, Duke University, Antifibrotics, PHASE 2, pulmonary fibrosis

O=C(C1(C2=CC=C(C3=CC=C(C4=C(NC(O[C@H](C)C5=CC=CC=C5)=O)C(C)=NO4)C=C3)C=C2)CC1)O

Funapide, TV 45070, XEN-402, фунапид فونابيد 呋纳匹特


Image result for TV 450702D chemical structure of 1259933-16-8

ChemSpider 2D Image | Funapide | C22H14F3NO5Funapide.png

Funapide TV 45070,  XEN-402,  Funapide, (+)-

фунапид
فونابيد
呋纳匹特
  • Molecular FormulaC22H14F3NO5
  • Average mass429.345 Da

(S)-1′-[(5-Methyl-2-furyl)methyl]spiro[6H-furo[3,2-f][1,3]benzodioxole-7,3′-indoline]-2′-one

Spiro(furo(2,3-F)-1,3-benzodioxole-7(6H),3′-(3H)indol)-2′(1’H)-one, 1′-((5-(trifluoromethyl)-2-furanyl)methyl)-, (3’S)-

(3’S)-1′-((5-(Trifluoromethyl)furan-2-yl)methyl)-2H,6H-spiro(furo(2,3-F)(1,3)benzodioxole-7,3′-indol)-2′(1’H)-one

Spiro[furo[2,3-f]-1,3-benzodioxole-7(6H),3′-[3H]indol]-2′(1’H)-one, 1′-[[5-(trifluoromethyl)-2-furanyl]methyl]-, (7S)-
TV-45070
UNII-A5595LHJ2L
XEN-401-S
XEN402
(3’S)-1′-{[5-(trifluoromethyl)furan-2-yl]methyl}-2H-6H-spiro[furo[2,3-f]-1,3-benzodioxole-7,3′-indol]-2′(1’H)-one
(7S)-1′-{[5-(Trifluoromethyl)-2-furyl]methyl}spiro[furo[2,3-f][1,3]benzodioxole-7,3′-indol]-2′(1’H)-one
1259933-16-8 CAS
UNII-A5595LHJ2L

Phase II clinical trials for Postherpetic neuralgia (PHN)

Treatment of Neuropathic Pain

  • Originator Xenon Pharmaceuticals
  • Developer Teva Pharmaceutical Industries; Xenon Pharmaceuticals
  • Class Benzodioxoles; Fluorobenzenes; Furans; Indoles; Non-opioid analgesics; Small molecules; Spiro compounds
  • Mechanism of Action Nav1.7-voltage-gated-sodium-channel-inhibitors; Nav1.8 voltage-gated sodium channel inhibitors
  • Orphan Drug Status Yes – Erythromelalgia

Highest Development Phases

  • Phase II Erythromelalgia; Postherpetic neuralgia
  • No development reported Dental pain; Pain
  • Discontinued Musculoskeletal pain

Most Recent Events

  • 09 May 2017 Teva Pharmaceutical Industries completes a phase IIb trial for Postherpetic neuralgia in USA (Topical) (NCT02365636)
  • 26 Sep 2016 Adverse events data from a phase II trial in Musculoskeletal pain presented at the 16th World Congress on Pain (PAN – 2016)
  • 19 Aug 2015 No recent reports of development identified – Phase-I for Pain (In volunteers) in Canada (PO)

MP 100 – 102 DEG CENT EP2538919

S ROT  ALPHA 0.99 g/100ml, dimethyl sulfoxide, 14.04, US 20110087027

Funapide (INN) (former developmental code names TV-45070 and XEN402) is a novel analgesic under development by Xenon Pharmaceuticals in partnership with Teva Pharmaceutical Industries for the treatment of a variety of chronic pain conditions, including osteoarthritisneuropathic painpostherpetic neuralgia, and erythromelalgia, as well as dental pain.[1][2][3][4] It acts as a small-moleculeNav1.7 and Nav1.8 voltage-gated sodium channel blocker.[1][2][4] Funapide is being evaluated in humans in both oral and topicalformulations, and as of July 2014, has reached phase IIb clinical trials.[1][3]

Image result for TV 45070

Sodium channels play a diverse set of roles in maintaining normal and pathological states, including the long recognized role that voltage gated sodium channels play in the generation of abnormal neuronal activity and neuropathic or pathological pain. Damage to peripheral nerves following trauma or disease can result in changes to sodium channel activity and the development of abnormal afferent activity including ectopic discharges from axotomised afferents and spontaneous activity of sensitized intact nociceptors. These changes can produce long-lasting abnormal hypersensitivity to normally innocuous stimuli, or allodynia. Examples of neuropathic pain include, but are not limited to, post-herpetic neuralgia, trigeminal neuralgia, diabetic neuropathy, chronic lower back pain, phantom limb pain, and pain resulting from cancer and chemotherapy, chronic pelvic pain, complex regional pain syndrome and related neuralgias.

There have been some advances in treating neuropathic pain symptoms by using medications, such as gabapentin, and more recently pregabalin, as short-term, first-line treatments. However, pharmacotherapy for neuropathic pain has generally had limited success with little response to commonly used pain reducing drugs, such as NSAIDS and opiates. Consequently, there is still a considerable need to explore novel treatment modalities.

There remain a limited number of potent effective sodium channel blockers with a minimum of adverse events in the clinic. There is also an unmet medical need to treat neuropathic pain and other sodium channel associated pathological states effectively and without adverse side effects. PCT Published Patent Application No. WO 2006/110917, PCT Published Patent Application No. WO 2010/045251 , PCT Published Patent Application No. WO 2010/045197, PCT Published Patent Application No. WO 2011/047174 and PCT Published Patent Application No. WO 2011/002708 discloses certain spiro-oxindole compounds. These compounds are disclosed therein as being useful for the treatment of sodium channel-mediated diseases, preferably diseases related to pain, central nervous conditions such as epilepsy, anxiety, depression and bipolar disease;

cardiovascular conditions such as arrhythmias, atrial fibrillation and ventricular fibrillation; neuromuscular conditions such as restless leg syndrome; neuroprotection against stroke, neural trauma and multiple sclerosis; and channelopathies such as erythromelalgia and familial rectal pain syndrome.

Methods of preparing these compounds and pharmaceutical compositions containing them are also disclosed in PCT Published Patent Application No. WO 2006/110917, PCT Published Patent Application No. WO 2010/045251 , PCT

Published Patent Application No. WO 2010/045197, PCT Published Patent Application No. WO 2011/047174 and PCT Published Patent Application No. WO 2011/002708.

Postherpetic neuralgia (PHN) is a rare disorder that is defined as significant pain or abnormal sensation 120 days or more after the presence of the initial rash caused by shingles. This pain persists after the healing of the associated rash. Generally, this affliction occurs in older individuals and individuals suffering from immunosuppression. There are about one million cases of shingles in the US per year, of which 10–20% will result in PHN.
Topical analgesics such as lidocaine and capsaicin are traditionally used to treat this disorder. Both lidocaine and TV-45070 have a mechanism of action that involves the inhibition of voltage-gated sodium ion channels.
TV-45070 (formerly XEN-402) was in-licensed by Teva from Xenon Pharmaceuticals and is reported to be an antagonist of the Nav1.7 sodium ion channel protein.
It is currently in Phase II clinical trials for PHN. Interestingly, the loss of function of the Nav1.7 sodium ion channel was reported to result in the inability to experience pain as a hereditary trait in certain individuals.
Primary erythromelalgia is another rare disease where alterations in Nav1.7 or mutations in the corresponding encoding gene SCN9A have been reported to result in chronic burning pain that can last for hours or even days. Thus, compounds which regulate this protein have potential therapeutic value as analgesics for chronic pain.
Image result for XENON PHARMA
PATENT
US 20100331386
WO 2011106729
US 20110087027
US 20110086899
US 20130143941
US 20130210884
WO 2013154712
 US 20150216794
WO 2016127068
WO 2016109795
CN 106518886
US 20170239183
SYNTHESIS
WO 2013154712
 CONTD…….
Synthesis
CN 106518886
PATENT
US 20100331386
Preparation of the (S)-Enantiomer of the Invention
The (S)-enantiomer of the invention and the corresponding (R)-enantiomer are prepared by the resolution of the compound of formula (I), as set forth above in the Summary of the Invention, using either chiral high pressure liquid chromatography methods or by simulated moving bed chromatography methods, as described below in the following Reaction Scheme wherein “chiral HPLC” refers to chiral high pressure liquid chromatography and “SMB” refers to simulated moving bed chromatography:
Figure US20100331386A1-20101230-C00006
The compound of formula (I) can be prepared by the methods disclosed in PCT Published Patent Application No. WO 2006/110917, by methods disclosed herein, or by methods known to one skilled in the art.
One of ordinary skill in the art would recognize variations in the above Reaction Scheme which are appropriate for the resolution of the individual enantiomers.
Alternatively, the (S)-enantiomer of formula (I-S) and the (R)-enantiomer of formula (I-R), can be synthesized from starting materials which are known or readily prepared using process analogous to those which are known.
Preferably, the (S)-enantiomer of the invention obtained by the resolution methods disclosed herein is substantially free of the (R)-enantiomer or contains only traces of the (R)-enantiomer.
The following Synthetic Examples serve to illustrate the resolution methods disclosed by the above Reaction Schemes and are not intended to limit the scope of the invention.
Synthetic Example 1Synthesis of 1-{[5-(trifluoromethyl)furan-2-yl]methyl}spiro[furo[2,3-f][1,3]benzodioxole-7,3′-indol]-2′(1′H)-one (Compound of formula (I))
Figure US20100331386A1-20101230-C00007
To a suspension of spiro[furo[2,3-f][1,3]benzodioxole-7,3′-indol]-2′(1′H)-one (1.0 g, 3.6 mmol), which can be prepared according to the methods disclosed in PCT Published Patent Application No. WO 2006/110917, and cesium carbonate (3.52 g, 11 mmol) in acetone (50 mL) was added 2-bromomethyl-5-trifluoromethylfuran (1.13 g, 3.9 mmol) in one portion and the reaction mixture was stirred at 55-60° C. for 16 hours. Upon cooling to ambient temperature, the reaction mixture was filtered and the filtrate was evaporated under reduced pressure. The residue was subjected to column chromatography, eluting with ethyl acetate/hexane (1/9-1/1) to afford 1′-{[5-(trifluoromethyl)furan-2-yl]methyl}spiro[furo[2,3-f][1,3]benzodioxole-7,3′-indol]-2′(1 ′H)-one, i.e., the compound of formula (I), (1.17 g, 76%) as a white solid: mp 139-141° C.;
1H NMR (300 MHz, CDCl3) δ 7.32-6.97 (m, 5H), 6.72 (d, J=3.3 Hz, 1H), 6.66 (s, 1H), 6.07 (s, 1H), 5.90-5.88 (m, 2H), 5.05, 4.86 (ABq, JAB=16.1 Hz, 2H), 4.91 (d, J=9.0 Hz, 1H), 4.66 (d, J=9.0 Hz, 1H); 13C NMR (75 MHz, CDCl3) δ 176.9, 155.7, 153.5, 148.8, 142.2, 141.9, 140.8, 140.2, 139.7, 139.1, 132.1, 129.2, 124.7, 124.1, 123.7, 121.1, 120.1, 117.6, 114.5, 114.4, 110.3, 109.7, 103.0, 101.9, 93.8, 80.0, 57.8, 36.9;
MS (ES+) m/z 430.2 (M+1), 452.2 (M+23); Cal’d for C22H14F3NO5: C, 61.54%; H, 3.29%; N, 3.26%; Found: C, 61.51%; H, 3.29%; N, 3.26%.
Synthetic Example 2Resolution of Compound of Formula (I) by Chiral HPLC
The compound of formula (I) was resolved into the (S)-enantiomer of the invention and the corresponding (R)-enantiomer by chiral HPLC under the following conditions:

Column: Chiralcel® OJ-RH; 20 mm I.D.×250 mm, 5 mic; Lot: OJRH CJ-EH001 (Daicel Chemical Industries, Ltd)

Eluent: Acetonitrile/Water (60/40, v/v, isocratic)

Flow rate: 10 mL/min

Run time: 60 min

Loading: 100 mg of compound of formula (I) in 1 mL of acetonitrileTemperature: Ambient

Under the above chiral HPLC conditions, the (R)-enantiomer of the compound of formula (I), i.e., (R)-1′-{[5-(trifluoromethyl)furan-2-yl]methyl}spiro[furo[2,3-f][1,3]-benzodioxole-7,3′-indol]-2′(1′H)-one, was isolated as the first fraction as a white solid; ee (enantiomeric excess)>99% (analytical OJ-RH, 55% acetonitrile in water); mp 103-105° C.; 1H NMR (300 MHz, DMSO-d6) δ 7.32-6.99 (m, 5H), 6.71 (d, J=3.4 Hz, 1H), 6.67 (s, 1H), 6.05 (s, 1H), 5.89 (d, J=6.2 Hz, 2H), 5.13, 5.02 (ABq, JAB=16.4 Hz, 2H), 4.82, 4.72 (ABq, JAB=9.4 Hz, 2H); 13C NMR (75 MHz, CDCl3) δ 177.2, 155.9, 152.0, 149.0, 142.4, 142.0, 141.3, 132.0, 129.1, 123.9, 120.6, 119.2, 117.0, 112.6, 109.3, 108.9, 103.0, 101.6, 93.5, 80.3, 58.2, 36.9; MS (ES+) m/z 430.2 (M+1), [α]D−17.46° (c 0.99, DMSO).

The (S)-enantiomer of the compound of formula (I), i.e., (S)-1′-{[5-(trifluoromethypfuran-2-yl]methyl}spiro-[furo[2,3-f][1,3]benzodioxole-7,3′-indol]-2′(1′H)-one was isolated as the second fraction as a white solid; ee >99% (analytical OJ-RH, 55% acetonitrile in water); mp 100-102° C.; 1H NMR (300 MHz, DMSO-d6) δ 7.32-6.99 (m, 5H), 6.71 (d, J=3.4 Hz, 1H), 6.67 (s, 1H), 6.05 (s, 1H), 5.89 (d, J=6.3 Hz, 2H), 5.12, 5.02 (ABq, JAB=16.4 Hz, 2H), 4.82, 4.72 (ABq, JAB=9.4 Hz, 2H); 13C NMR (75MHz, CDCl3) δ 177.2, 155.9, 152.0, 149.0, 142.4, 142.0, 141.3, 132.0, 129.1, 123.9, 120.6, 119.2, 117.0, 112.6, 109.3, 108.9, 103.0, 101.6, 93.5, 80.3, 58.2, 36.9; MS (ES+) m/z 430.2 (M+1), [α]D+14.04° (c 0.99, DMSO)

Synthetic Example 3Resolution of Compound of Formula (I) by SMB Chromatography

The compound of formula (I) was resolved into the (S)-enantiomer of the invention and the corresponding (R)-enantiomer by SMB chromatography under the following conditions:

Extract: 147.05 mL/min, Raffinate: 76.13 mL/min Eluent: 183.18 mL/min Feed: 40 mL/min Recycling: 407.88 mL/min Run Time: 0.57 min Temperature: 25° C. Pressure: 46 bar

The feed solution (25 g of compound of formula (I) in 1.0 L of mobile phase (25:75:0.1 (v:v:v) mixture of acetonitrile/methanol/trifluoroacetic acid)) was injected continuously into the SMB system (Novasep Licosep Lab Unit), which was equipped with eight identical columns in 2-2-2-2 configuration containing 110 g (per column, 9.6 cm, 4.8 cm I.D.) of ChiralPAK-AD as stationary phase. The first eluting enantiomer (the (R)-enantiomer of the compound of formula (I)) was contained in the raffinate stream and the second eluting enantiomer (the (S)-enantiomer of the compound of formula (I)) was contained in the extract stream. The characterization data of the (S)-enantiomer and the (R)-enantiomer obtained from the SMB resolution were identical to those obtained above utilizing chiral HPLC.

The compound of formula (I) was resolved into its constituent enantiomers on a Waters preparative LCMS autopurification system. The first-eluting enantiomer from the chiral column was brominated (at a site well-removed from the stereogenic centre) to give the corresponding 5′-bromo derivative, which was subsequently crystallized to generate a single crystal suitable for X-ray crystallography. The crystal structure of this brominated derivative of the first-eluting enantiomer was obtained and its absolute configuration was found to be the same as the (R)-enantiomer of the invention. Hence, the second-eluting enantiomer from the chiral column is the (S)-enantiomer of the invention. Moreover, the material obtained from the extract stream of the SMB resolution had a specific optical rotation of the same sign (positive, i.e. dextrorotatory) as that of the material obtained from the aforementioned LC resolution.

Patent

WO 2013154712

EXAMPLE 8

Synthesis of (7S)-1 ‘-{[5-(trifluoromethyl)furan-2- yllmethylJspirotfurop.S-flll .Sl enzoclioxole-y.S’-indoll-Zil ‘Wi-one

Compound of formula (ia1 )

Figure imgf000095_0001

To a cooled (0 °C) solution of (3S)-3-(6-hydroxy-1 ,3-benzodioxol-5-yl)-3- (hydroxymethyl)-1-{[5-(trifluoromethyl)furan-2-yl]methyl}-1 ,3-dihydro-2H-indol-2-one prepared according to the procedure described in Example 7 (16.4 mmol) and 2- (diphenylphosphino)pyridine (5.2 g, 20 mmol) in anhydrous tetrahydrofuran (170 mL) was added di-ferf-butylazodicarboxylate (4.5 g, 20 mmol). The mixture was stirred for 2 h at 0 °C, then the reaction was diluted with ethyl acetate (170 mL), washed with 3 N hydrochloric acid (7 x 50 mL) and brine (2 x 100 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was dissolved in ethanol (80 mL), decolorizing charcoal (15 g) was added and the mixture was heated at reflux for 1 h. The mixture was filtered while hot through a pad of diatomaceous earth. The filtrate was concentrated in vacuo and the residue triturated in a mixture of diethyl ether/hexanes to afford (7S)-1 ‘-{[5-(trifluoromethyl)furan-2-yl]methyl}spiro- [furo[2,3-/][1 ,3]benzodioxole-7,3’-indol]-2′(1 ‘H)-one (1.30 g) as a colorless solid in 18% yield. The mother liquor from the trituration was concentrated in vacuo, trifluoroacetic acid (20 mL) was added and the mixture stirred for 3 h at ambient temperature. The mixture was diluted with ethyl acetate (100 mL), washed with saturated aqueous ammonium chloride (100 mL), 3 N hydrochloric acid (4 x 60 mL) and brine (2 x 100 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by column chromatography, eluting with a gradient of ethyl acetate in hexanes to afford further (7S)-1 ‘-{[5-(trifluoromethyl)furan-2-yl]methyl}spiro- [furo[2,3- ][1 ,3]benzodioxole-7,3’-indol]-2′(1 ‘H)-one (2.6 g) as a colorless solid (37% yield, overall yield 55% over 2 steps): H NMR (300 MHz, CDCI3) £7.29-6.96 (m, 4H), 6.73 (s, 1 H), 6.50 (s, 1 H), 6.38 (s, 1 H), 6.09 (s, 1 H), 5.85 (br s, 2H), 5.06 (d, J = 16.0 Hz, 1 H), 4.93-4.84 (m, 2H), 4.68-4.65 (m, 1 H); MS (ES+) m/z 429.8 (M + 1 ); ee (enantiomeric excess) >99.5% (HPLC, Chiralpak IA, 2.5% acetonitrile in methyl tert- butyl ether).

EXAMPLE 9

Synthesis of 1-(diphenylmethyl)-1 H-indole-2,3-dione

Compound of formula (15a)

Figure imgf000096_0001

A. To a suspension of hexanes-washed sodium hydride (34.0 g, 849 mmol) in anhydrous Λ/,/V-dimethylformamide (400 mL) at 0 °C was added a solution of isatin (99.8 g, 678 mmol) in anhydrous Λ/,/V-dimethylformamide (400 mL) dropwise over 30 minutes. The reaction mixture was stirred for 1 h at 0 °C and a solution of benzhydryl bromide (185 g, 745 mmol) in anhydrous N-dimethylformamide (100 mL) was added dropwise over 5 minutes. The reaction mixture was allowed to warm to ambient temperature, stirred for 16 h and heated at 60 °C for 2 h. The mixture was cooled to 0 °C and water (500 mL) was added. The mixture was poured into water (2 L), causing a precipitate to be deposited. The solid was collected by suction filtration and washed with water (2000 mL) to afford 1-(diphenylmethyl)-1H-indole-2,3- dione (164 g) as an orange solid in 77% yield.

B. Alternatively, to a mixture of isatin (40.0 g, 272 mmol), cesium carbonate (177 g, 543 mmol) and A/./V-dimethylformamide (270 mL) at 80 °C was added dropwise a solution of benzhydryl bromide (149 g, 544 mmol) in N,N- dimethyiformamide (200 mL) over 30 minutes. The reaction mixture was heated at 80 °C for 3 h, allowed to cool to ambient temperature and filtered through a pad of diatomaceous earth. The pad was rinsed with ethyl acetate (1000 mL). The filtrate was washed with saturated aqueous ammonium chloride (4 x 200 mL), 1 N

hydrochloric acid (200 mL) and brine (4 x 200 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was triturated with diethyl ether to afford 1 -(diphenylmethyl)-1 H-indole-2,3-dione (59.1 g) as an orange solid in 69% yield. The mother liquor from the trituration was concentrated in vacuo and the residue triturated in diethyl ether to afford a further portion of 1-(diphenylmethyl)-1 H- indole-2,3-dione (8.2 g) in 10% yield: 1H NMR (300 MHz, CDCI3) £7.60 (d, J = 7.4 Hz, 1 H), 7.34-7.24 (m, 1 1 H), 7.05-6.97 (m, 2H), 6.48 (d, J = 8.0 Hz, 1 H); MS (ES+) m/z 313.9 (M + 1 ).

C. Alternatively, a mixture of isatin (500 g, 3.4 mol) and anhydrous N,N- dimethylformamide (3.5 L) was stirred at 15-35 °C for 0.5 h. Cesium carbonate (2.2 kg, 6.8 mol) was added and the mixture stirred at 55-60 °C for 1 h. A solution of benzhydryl bromide (1.26 kg, 5.1 mol) in anhydrous N, A/-dimethylformamide (1.5 L) was added and the resultant mixture stirred at 80-85 °C for 1 h, allowed to cool to ambient temperature and filtered. The filter cake was washed with ethyl acetate (12.5 L). To the combined filtrate and washes was added 1 N hydrochloric acid (5 L). The phases were separated and the aqueous phase was extracted with ethyl acetate (2.5 L). The combined organic extracts were washed with 1 N hydrochloric acid (2 * 2.5 L) and brine (3 χ 2.5 L) and concentrated in vacuo to a volume of approximately 750 mL. Methyl ferf-butyl ether (2 L) was added and the mixture was cooled to 5-15 °C, causing a solid to be deposited. The solid was collected by filtration, washed with methyl ferf- butyl ether (250 mL) and dried in vacuo at 50-55 °C for 16 h to afford 1- (diphenylmethyl)-1 H-indole-2,3-dione (715 g) as an orange solid in 67% yield: 1H NMR (300 MHz, CDCI3) 7.60 (d, J = 7.4 Hz, H), 7.34-7.24 (m, 1 H), 7.05-6.97 (m, 2H), 6.48 (d, J = 8.0 Hz, 1 H); MS (ES+) m/z 313.9 (M + 1 ).

EXAMPLE 10

Synthesis of 1-(diphenylmethyl)-3-hydroxy-3-(6-hydroxy-1 ,3-benzodioxol-5-yl)-1 ,3- dihydro-2H-indol-2-one

Compound of formula (16a1 )

Figure imgf000097_0001

A. To a solution of sesamol (33.1 g, 239 mmol) in anhydrous

tetrahydrofuran (500 mL) at 0 °C was added dropwise a 2 M solution of

isopropylmagnesium chloride in tetrahydrofuran (104 mL, 208 mmol), followed by 1 – (diphenylmethyl)-1H-indole-2,3-dione (50.0 g, 160 mmol) and tetrahydrofuran (100 mL). The reaction mixture was stirred at ambient temperature for 5 h, diluted with ethyl acetate (1500 mL), washed with saturated aqueous ammonium chloride (400 mL) and brine (2 x 400 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was triturated with a mixture of diethyl ether and hexanes to afford 1- (diphenylmethyl)-3-hydroxy-3-(6-hydroxy-1 ,3-benzodioxol-5-yl)-1 ,3-dihydro-2H-in

2- one (70.7 g) as a colorless solid in 98% yield: 1H NMR (300 MHz, CDCI3) <59.12 (br s, 1 H), 7.45-7.43 (m, 1 H), 7.30-7.22 (m, 10H), 7.09-7.07 (m, 2H), 6.89 (s, 1 H), 6.56- 6.55 (m, 1 H), 6.47-6.46 (m, 1 H), 6.29-6.28 (m, 1 H), 5.86 (s, 2H), 4.52 (br s, 1 H); MS (ES+) m/z 433.7 (M – 17).

B. Alternatviely, a mixture of sesamol (0.99 kg, 7.2 mol) and anhydrous tetrahydrofuran (18 L) was stirred at 15-35 °C for 0.5 h and cooled to -5-0 °C.

Isopropyl magnesium chloride (2.0 M solution in tetrahydrofuran, 3.1 L, 6.2 mol) was added, followed by 1-(diphenylmethyl)-1 H-indole-2,3-dione (1.50 kg, 4.8 mol) and further anhydrous tetrahydrofuran (3 L). The mixture was stirred at 15-25 °C for 5 h. Ethyl acetate (45 L) and saturated aqueous ammonium chloride (15 L) were added. The mixture was stirred at 15-25 °C for 0.5 h and was allowed to settle for 0.5 h. The phases were separated and the organic phase was washed with brine (2.3 L) and concentrated in vacuo to a volume of approximately 4 L. Methyl ferf-butyl ether (9 L) was added and the mixture concentrated in vacuo to a volume of approximately 4 L. Heptane (6 L) was added and the mixture was stirred at 15-25 °C for 2 h, causing a solid to be deposited. The solid was collected by filtration, washed with methyl tert- butyl ether (0.3 L) and dried in vacuo at 50-55 °C for 7 h to afford 1-(diphenylmethyl)-3- hydroxy-3-(6-hydroxy-1 ,3-benzodioxol-5-yl)-1 ,3-dihydro-2H-indol-2-one (2.12 kg) as an off-white solid in 98% yield: 1H NMR (300 MHz, CDCI3) 9.12 (br s, 1 H), 7.45-7.43 (m, 1 H), 7.30-7.22 (m, 10H), 7.09-7.07 (m, 2H), 6.89 (s, 1 H), 6.56-6.55 (m, 1 H), 6.47-6.46 (m, 1 H), 6.29-6.28 (m, 1 H), 5.86 (s, 2H), 4.52 (br s, 1 H); MS (ES+) m/z 433.7 (M – 17).

EXAMPLE 1 1

Synthesis of 3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-1-(diphenylmethyl)-3-hydroxy-1 ,3- dihydro-2H-indol-2-one

Compound of formula (17a1)

Figure imgf000098_0001

A. A mixture of 1-(diphenylmethyl)-3-hydroxy-3-(6-hydroxy-1 ,3- benzodioxol-5-yl)-1 ,3-dihydro-2H-indol-2-one (30.0 g, 66.5 mmol), benzyl bromide (8.3 mL, 70 mmol), and potassium carbonate (18.4 g, 133 mmol) in anhydrous N,N- dimeihylformamide (100 mL) was stirred at ambient temperature for 16 h. The reaction mixture was filtered and the solid was washed with /V,A/-dimethylformamide (100 mL). The filtrate was poured into water (1000 mL) and the resulting precipitate was collected by suction filtration and washed with water to afford 3-[6-(benzyloxy)-1 ,3-benzodioxol- 5-yl]-1-(diphenylmethyl)-3-hydroxy-1 ,3-dihydro-2H-indol-2-one (32.0 g) as a beige solid in 83% yield: 1H NMR (300 MHz, CDCI3) 7.42-7.28 (m, 9H), 7.22-7.14 (m, 6H), 7.10- 6.93 (m, 3H), 6.89-6.87 (m, 2H), 6.53 (d, J = 7.6 Hz, 1 H), 6.29 (br s, 1 H), 5.88 (s, 1 H), 5.85 (s, 1 H), 4.66 (d, J = 14.2 Hz, 1 H), 4.51 (d, J = 14.1 Hz, 1 H), 3.95 (s, 1 H); MS (ES+) m/z 542.0 (M + 1), 523.9 (M – 17).

B. Alternatively, to a solution of 1-(diphenylmethyl)-3-hydroxy-3-(6- hydroxy-1 ,3-benzodioxol-5-yl)-1 ,3-dihydro-2H-indol-2-one (2.1 kg, 4.6 mol) in anhydrous A/,A/-dimethylformamide (8.4 L) at 20-30 °C was added potassium carbonate (1.3 kg, 9.2 mol), followed by benzyl bromide (0.58 L, 4.8 mol). The mixture was stirred at 20-30 °C for 80 h and filtered. The filter cake was washed with

A/,/V-dimethylformamide (0.4 L) and the filtrate was poured into water (75 L), causing a solid to be deposited. The mixture was stirred at 15-25 °C for 7 h. The solid was collected by filtration, washed with water (2 L) and dried in vacuo at 50-60 °C for 48 h to afford 3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-1-(diphenylmethyl)-3-hydroxy-1 ,3- dihydro-2H-indol-2-one (2.1 1 kg) as an off-white solid in 84% yield; 1H NMR (300

MHz, CDCI3) £7.42-7.28 (m, 9H), 7.22-7.14 (m, 6H), 7.10-6.93 (m, 3H), 6.89-6.87 (m, 2H), 6.53 (d, J = 7.6 Hz, 1 H), 6.29 (br s, 1 H), 5.88 (s, 1 H), 5.85 (s, 1 H), 4.66 (d, J = 14.2 Hz, 1 H), 4.51 (d, J = 14.1 Hz, 1 H), 3.95 (s, 1 H); MS (ES+) m/z 542.0 (M + 1 ).

EXAMPLE 12

Synthesis of 3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-1 -(diphenylmethyl)-l ,3-dihydro-2H- indol-2-one

Compound of formula (18a1 )

Figure imgf000099_0001

A. To a solution of 3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-1- (diphenylmethyl)-3-hydroxy-1 ,3-dihydro-2H-indol-2-one (32.0 g, 57.7 mmol) in dichloromethane (100 mL) was added trifluoroacetic acid (50 mL) followed by triethylsilane (50 mL). The reaction mixture was stirred at ambient temperature for 2 h and concentrated in vacuo. The residue was dissolved in ethyi acetate (250 mL), washed with saturated aqueous ammonium chloride (3 x 100 mL) and brine (3 x 100 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was triturated with diethyl ether to afford 3-[6-(benzyloxy)-1 ,3-benzodioxol-5- yl]-1-(diphenylmethyl)-1 ,3-dihydro-2H-indol-2-one (19.0 g) as a colorless solid in 61 % yield: 1H NMR (300 MHz, CDCI3) 7.31 -7.23 (m, 15H), 7.10-6.88 (m, 4H), 6.50-6.45 (m, 3H), 5.86 (s, 2H), 4.97-4.86 (m, 3H); MS (ES+) m/z 525.9 (M + 1).

B. Alternatively, to a solution of 3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-1- (diphenylmethyl)-3-hydroxy-1 ,3-dihydro-2H-indol-2-one (2.0 kg, 3.7 mol) in

dichloromethane (7 L) at 20-30 °C was added trifluoracetic acid (2.5 L), followed by triethylsilane (3.1 L). The mixture was stirred at 15-35 °C for 4 h and concentrated in vacuo to dryness. To the residue was added ethyl acetate (16 L) and the mixture was stirred at 15-35 °C for 0.5 h, washed with saturated aqueous ammonium chloride (3 x 7 L) and brine (3 χ 7 L) and concentrated in vacuo to a volume of approximately 7 L. Methyl ferf-butyl ether (9 L) was added and the mixture concentrated in vacuo to a volume of approximately 9 L and stirred at 10-20 °C for 2.5 h, during which time a solid was deposited. The solid was collected by filtration, washed with methyl te/t-butyl ether (0.4 L) and dried in vacuo at 50-55 °C for 7 h to afford 3-[6-(benzyloxy)-1 ,3- benzodioxol-5-yl]-1-(diphenylmethyl)-1 ,3-dihydro-2H-indol-2-one (1 .26 kg) as an off-white solid in 65% yield: 1H NMR (300 MHz, CDCI3) £7.31 -7.23 (m, 15H), 7.10- 6.88 (m, 4H), 6.50-6.45 (m, 3H), 5.86 (s, 2H), 4.97-4.86 (m, 3H); MS (ES+) m/z 525.9 (M + 1).

EXAMPLE 13

Synthesis of (3S)-3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-3-[(benzyloxy)methyl]-1 –

(diphenylmethyl)-1 ,3-dihydro-2H-indol-2-one

Compound of formula (19a1 )

Figure imgf000100_0001

A. To a nitrogen-degassed mixture of 50% w/w aqueous potassium hydroxide (69.6 mL, 619 mmol), toluene (100 mL), and (9S)-1 -(anthracen-9- ylmethyl)cinchonan-1 -ium-9-ol chloride (0.50 g, 0.95 mmol) cooled in an ice/salt bath to an internal temperature of -18 °C was added a nitrogen-degassed solution of 3-[6- (benzyloxy)-l ,3-benzodioxol-5-yl]-1 -(diphenylmethyl)-l ,3-dihydro-2H-indol-2-one (10.0 g, 19.0 mmol) and benzyl chloromethyl ether (2.9 mL, 21 mmol) in

toluene/tetrahydrofuran (1 :1 v/v, 80 mL) dropwise over 1 h. The reaction mixture was stirred for 3.5 h and diluted with ethyl acetate (80 mL). The organic phase was washed with 1 N hydrochloric acid (3 x 150 mL) and brine (2 x 100 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo to afford (3S)-3-[6-(benzyloxy)-1 ,3- benzodioxol-5-yl]-3-[(benzyloxy)methyl]-1-(diphenylmethyl)-1 ,3-dihydro-2/-/-indol-2-one (12.6 g) as a colorless solid in quantitative yield: 1H NMR (300 MHz, CDCI3) 7.42 (d, 2H), 7.24-6.91 (m, 21 H), 6.69-6.67 (m, 2H), 6.46 (d, J = 7.7 Hz, 1 H), 6.15 (s, 1 H), 5.83- 5.81 (m, 2H), 4.53-4.31 (m, 3H), 4.17-4.09 (m, 3H); MS (ES+) m/z 646.0 (M + 1); ee (enantiomeric excess) 90% (HPLC, Chiralpak IA, 2.5% acetonitrile in methyl tert-butyl ether).

B. Alternatively, a mixture of 50% w/v aqueous potassium hydroxide (4.2 kg), toluene (12 L) and (9S)-1 -(anthracen-9-ylmethyl)cinchonan-1 -ium-9-ol chloride (0.06 kg, 0.1 mol) was degassed with dry nitrogen and cooled to -18 to -22 °C. To this mixture was added a cold (-18 to -22 °C), nitrogen-degassed solution of 3-[6-

(benzyloxy)-l ,3-benzodioxol-5-yl]-1 ~(diphenylmethyl)-1 ,3-dihydro-2H-indol-2-one (1.2 kg, 2.3 mol) and benzyl chloromethyl ether (0.43 kg, 2.8 mol) in toluene (10 L) and tetrahydrofuran (10 L) at -18 to 22 °C over 3 h. The mixture was stirred at -18 to -22 °C for 5 h, allowed to warm to ambient temperature and diluted with ethyl acetate (10 L). The phases were separated and the organic layer was washed with 1 N

hydrochloric acid (3 χ 8 L) and brine (2 χ 12 L) and concentrated in vacuo to dryness to afford (3S)-3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-3-[(benzyloxy)methyl]-1- (diphenylmethyl)-1 ,3-dihydro-2H-indol-2-one (1.5 kg) as a colorless solid in quantitative yield: 1H NMR (300 MHz, CDCI3) £7.42 (d, 2H), 7.24-6.91 (m, 21 H), 6.69-6.67 (m, 2H), 6.46 (d, J = 7.7 Hz, 1 H), 6.15 (s, 1 H), 5.83-5.81 (m, 2H), 4.53-4.31 (m, 3H), 4.17- 4.09 (m, 3H); MS (ES+) m/z 646.0 (M + 1); ee (enantiomeric excess) 90% (HPLC, ChiralPak IA). EXAMPLE 14

Synthesis of (3S)-1-(diphenylmethyl)-3-(6-hydroxy-1 ,3-benzodioxol-5-yl)-3- (hydroxymethyl)-1 ,3-dihydro-2/-/-indol-2-one

Compound of formula (20a1)

Figure imgf000102_0001

A. A mixture of (3S)-3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-3- [(benzyloxy)methyl]-1 -(diphenylmethyl)-1 ,3-dihydro-2/-/-indol-2-one (8.8 g, 14 mmol), 10% w/w palladium on carbon (50% wetted powder, 3.5 g, 1.6 mmol), and acetic acid (3.9 ml_, 68 mmol) in a nitrogen-degassed mixture of ethanol/tetrahydrofuran (1 : 1 v/v, 140 mL) was stirred under hydrogen gas (1 atm) at ambient temperature for 4 h. The reaction mixture was filtered through a pad of diatomaceous earth and the pad was rinsed with ethyl acetate (100 mL). The filtrate was concentrated in vacuo to afford (3S)-1-(diphenylmethyl)-3-(6-hydroxy-1 ,3-benzodioxol-5-yl)-3-(hydroxymethyl)-1 ,3- dihydro-2H-indol-2-one as a colorless solid that was carried forward without further purification: H NMR (300 MHz, CDCI3) 9.81 (br s, 1 H), 7.35-7.24 (m, 1 1 H), 7.15- 7.01 (m, 3H), 6.62 (s, 1 H), 6.54-6.47 (m, 2H), 5.86-5.84 (m, 2H), 4.76 (d, J = 1 1.0 Hz, 1 H), 4.13-4.04 (m, 1 H), 2.02 (s, 1 H); MS (ES+) m/z 465.9 (M + 1); ee (enantiomeric excess) 93% (HPLC, Chiralpak IA, 2.5% acetonitrile in methyl ie t-butyl ether).

B. Alternatively, a glass-lined hydrogenation reactor was charged with (3S)-3-[6-(benzyloxy)-1 ,3-benzodioxol-5-yl]-3-[(benzyloxy)methyl]-1 -(diphenylmethyl)- 1 ,3-dihydro-2H-indol-2-one (0.1 kg, 0.15 mol), tetrahydrofuran (0.8 L), ethanol (0.4 L), acetic acid (0.02 L) and 20% w/w palladium (li) hydroxide on carbon (0.04 kg). The reactor was purged three times with nitrogen. The reactor was then purged three times with hydrogen and was then pressurized to 50-55 lb/in2 with hydrogen. The mixture was stirred at 20-30 °C for 5 h under a 50-55 lb/in2 atmosphere of hydrogen. The reactor was purged and the mixture was filtered. The filtrate was concentrated in vacuo to a volume of approximately 0.2 L and methyl te/t-butyl ether (0.4 L) was added. The mixture was concentrated in vacuo to a volume of approximately 0.2 L and methyl ie/t-butyl ether (0.2 L) was added, followed by heptane (0.25 L). The mixture was stirred at ambient temperature for 2 h, during which time a solid was deposited. The solid was collected by filtration, washed with heptane (0.05 L) and dried in vacuo at a temperature below 50 °C for 8 h to afford (3S)-1 -(diphenylmethyl)-3-(6-hydroxy- 1 ,3-benzodioxol-5-yl)-3-(hydroxymethyl)-1 ,3-dihydro-2H-indol-2-one (0.09 kg) as a colorless solid in 95% yield: 1H NMR (300 MHz, CDCI3) 9.81 (br s, 1 H), 7.35-7.24 (m, 1 1 H), 7.15-7.01 (m, 3H), 6.62 (s, 1 H), 6.54-6.47 (m, 2H), 5.86-5.84 (m, 2H), 4.76 (d, J = 1 1.0 Hz, 1 H), 4.13-4.04 (m, 1 H), 2.02 (s, 1 H); MS (ES+) m/z 465.9 (M + 1); ee (enantiomeric excess) 91% (HPLC, ChiralPak IA).

EXAMPLE 15

Synthesis of (7S)-1′-(diphenylmethyl)spiro[furo[2,3-/][1 ,3]benzodioxole-7,3′-indol]-

2′(1 ‘tf)-one

Compound of formula (21 a1 )

Figure imgf000103_0001

A. To a cooled (0 °C) solution of (3S)-1 -(diphenylmethyl)-3-(6-hydroxy-1 ,3- benzodioxol-5-yl)-3-(hydroxymethyl)-1 ,3-dihydro-2H-indol-2-one prepared according to the procedure described in Example 14 (13.6 mmol) and 2-

(diphenylphosphino)pyridine (4.3 g, 16 mmol) in anhydrous tetrahydrofuran (140 mL) was added di-tert-butylazodicarboxylate (3.8 g, 17 mmol). The reaction mixture was stirred at 0 °C for 3 h, diluted with ethyl acetate (140 mL), washed with 3 N

hydrochloric acid (6 * 50 mL) and brine (2 χ 100 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was triturated with a mixture of diethyl ether and hexanes to afford (7S)-1 ‘-(diphenylmethyl)spiro[furo[2,3- ][1 ,3]benzodioxole-7,3’-indol]-2′(1 ‘H)-one (4.55 g) as a colorless solid in a 75% yield over 2 steps: 1H NMR (300 MHz, CDCI3) 7.34-7.24 (m, 10H), 7.15-7.13 (m, 1 H), 7.04 (s, 1 H), 6.99-6.95 (m, 2H), 6.50-6.48 (m, 2H), 6.06 (s, 1 H), 5.85-5.83 (m, 2H), 4.96 (d, J = 8.9 Hz, 1 H), 4.69 (d, J = 8.9 Hz, 1 H); MS (ES+) m/z 447.9 (M + 1); ee

(enantiomeric excess) 93% (HPLC, Chiraipak IA, 2.5% acetonitrile in methyl te/f-butyl ether).

B. Alternativel, to a cooled (0-5 °C) solution of (3S)-1-(diphenylmethyl)-3- (6-hydroxy-1 ,3-benzodioxol-5-yl)-3-(hydroxymethyl)-1 ,3-dihydro-2 -/-indol-2-one (1 .0 kg, 2.1 mol) and 2-(diphenylphosphino)pyridine (0.66 kg, 2.5 mol) in anhydrous tetrahydrofuran (20 L) was added over 2 h a solution of di-terf-butylazodicarboxylate (0.62 kg, 2.7 mmol) in anhydrous tetrahydrofuran (5 L). The mixture was stirred for 4 h at 0-5 °C and was allowed to warm to ambient temperature. The mixture was diluted with ethyl acetate (20 L), washed with 3 N hydrochloric acid (6 * 8 L) and brine (2 x 12 L) and concentrated in vacuo to a volume of approximately 1.5 L. Methyl rert-butyl ether (4 L) was added and the mixture concentrated in vacuo to a volume of

approximately 1.5 L. Methyl terf-butyl ether (2 L) and heptane (2 L) were added and the mixture was stirred at ambient temperature for 2 h, during which time a solid was deposited. The solid was collected by filtration, washed with heptane (0.5 L) and dried in vacuo below 50 °C for 8 h to afford (7S)-1′-(diphenylmethyl)spiro[furo[2,3- f][1 ,3]benzodioxole-7,3′-indol]-2′(1’H)-one (0.76 kg) as a colorless solid in 79% yield: 1H NMR (300 MHz, CDCI3) 7.34-7.24 (m, 10H), 7.15-7.13 (m, 1 H), 7.04 (s, 1 H), 6.99- 6.95 (m, 2H), 6.50-6.48 (m, 2H), 6.06 (s, 1 H), 5.85-5.83 (m, 2H), 4.96 (d, J = 8.9 Hz, 1 H), 4.69 (d, J = 8.9 Hz, 1 H); MS (ES+) m/z 447.9 (M + 1 ); ee (enantiomeric excess) 92% (HPLC, ChiralPak IA).

EXAMPLE 16

Synthesis of (7S)-spiro[furo[2,3-f][1 ,3]benzodioxole-7,3′-indol]-2′(1 ‘H)-one

Compound of formula (22a1)

Figure imgf000104_0001

A. To a solution of (7S)-1′-(diphenylmethyl)spiro[furo[2,3- f][1 ,3]benzodioxole-7,3′-indol]-2′(1’H)-one (4.55 g, 10.2 mmol) in trifluoroacetic acid (80 ml_) was added triethylsilane (7 ml_). The reaction mixture was heated at reflux for 2.5 h, allowed to cool to ambient temperature and concentrated in vacuo. The residue was triturated with a mixture of diethyl ether and hexanes to afford

(7S)-spiro[furo[2,3-/][1 ,3]benzodioxole-7,3,-indol]-2′(1’W)-one (2.30 g) as a colorless solid in 80% yield: 1H NMR (300 MHz, CDCI3) £8.27 (br s, 1 H), 7.31-7.26 (m, 1 H), 7.17-7.15 (m, 1 H), 7.07-7.02 (m, 1 H), 6.96-6.94 (m, 1 H), 6.53-6.52 (m, 1 H), 6.24-6.23 (m, 1 H), 5.88-5.87 (m, 2H), 4.95 (d, J = 8.6 Hz, 1 H), 4.68 (d, J = 8.9 Hz, 1 H); MS (ES+) m/z 281.9 (M + 1 ); ee (enantiomeric excess) 99% (HPLC, Chiralpak IA, 2.5% acetonitrile in methyl fert-butyl ether). B. Alternatively, a mixture of (7S)-1 ‘-(diphenylmethyl)spiro[furo[2,3- /Kl^benzodioxole^-indol^ r^-one (0.70 kg, 1.6 mol), trifluoroacetic acid (12 L) and triethylsilane (1.1 L) was heated at reflux under nitrogen atmosphere for 3 h, allowed to cool to ambient temperature and concentrated in vacuo to dryness. To the residue was added ethyl acetate (0.3 L), methyl fert-butyl ether (1 L) and heptane (3.5 L), causing a solid to be deposited. The solid was collected by filtration, taken up in dichloromethane (3 L), stirred at ambient temperature for 1 h and filtered. The filtrate was concentrated in vacuo to dryness. The residue was taken up in ethyl acetate (0.3 L), methyl ferf-butyl ether (1 L) and heptane (3.5 L), causing a solid to be deposited. The solid was collected by filtration and dried in vacuo below 50 °C for 8 h to afford (7S)-spiro[furo[2,3- ][1 ,3]benzodioxole-7,3’-indol]-2′(1 ‘ -/)-one (0.40 kg) as a colorless solid in 91 % yield: 1H NMR (300 MHz, CDCI3) 8.27 (br s, 1 H), 7.31-7.26 (m, 1 H), 7.17-7.15 (m, 1 H), 7.07-7.02 (m, 1 H), 6.96-6.94 (m, 1 H), 6.53-6.52 (m, 1 H), 6.24-6.23 (m, 1 H), 5.88-5.87 (m, 2H), 4.95 (d, J = 8.6 Hz, 1 H), 4.68 (d, J = 8.9 Hz, 1 H); MS (ES+) m/z 281.9 (M + 1); ee (enantiomeric excess) 98.6% (HPLC, ChiralPak IA).

EXAMPLE 17

Synthesis of of (7S)-1 ‘-{[5-(trifluoromethyl)furan-2- yl]methyl}spiro[furo[2,3- ][1 ,3]benzodioxole-7,3’-indol]-2′(rH)-one

Compound of formula (Ia1)

Figure imgf000105_0001

A. To a mixture of (7S)-6H-spiro[[1 ,3]dioxolo[4,5-f]benzofuran-7,3′-indolin]- 2′-one (1.80 g, 6.41 mmol) and 2-(bromomethyl)-5-(trifluoromethyl)furan (1.47 g, 6.41 mmol) in acetone (200 mL) was added cesium carbonate (3.13 g, 9.61 mmol). The reaction mixture was heated at reflux for 2 h and filtered while hot through a pad of diatomaceous earth. The filtrate was concentrated in vacuo to afford (7S)-1′-{[5- (trifluoromethyOfuran^-yllmethy^spiroIfurop.S- ltl .Slbenzodioxole^.S’-indol^ rH)- one (2.71 g) as a colorless solid in quantitative yield (97% purity by HPLC). The product was crystallized from a mixture of methanol and hexanes to afford (7S)-1 ‘-{[5- (trifluoromethy furan^-yllmethylJspirotfuro^.S- lfl .Slbenzodioxole^.S’-indoll^ rH)- one (1.46 g) as colorless needles in 53% yield. The mother liquor was concentrated in vacuo and subjected to a second crystallization in methanol and hexanes to afford further (7S)-1 ‘-{[5-(trifluoromethyl)furan-2-yl]methyl}spiro[furo[2,3-/][1 ,3]benzodioxole- 7,3’-indol]-2′(1 ‘H)-one (0.469 g) as a colorless solid in 17% yield (total yield 70%): 1H NMR (300 MHz, CDCI3) δ 7.29-6.96 (m, 4H), 6.73 (s, 1 H), 6.50 (s, 1 H), 6.38 (s, 1 H), 6.09 (s, 1 H), 5.85 (br s, 2H), 5.06 (d, J = 16.0 Hz, 1 H), 4.93-4.84 (m, 2H), 4.68-4.65 (m, 1 H); MS (ES+) m/z 429.8 (M + 1); ee (enantiomeric excess) >99.5% (HPLC, Chiralpak IA, 2.5% acetonitrile in methyl tert-butyl ether).

B. Alternatively, to a solution of (7S)-spiro[furoI2,3-f][1 ,3]benzodioxole-7,3′- indol]-2′(1’H)-one (0.40 kg, 1.4 mol) in anhydrous N, W-dimethylformamide (5 L) was added cesium carbonate (1.2 kg, 3.4 mol), followed by 2-(bromomethyl)-5- (trifluromethyl)furan (0.24 L, 1.7 mol). The mixture was heated at 80-85 °C for 3 h, allowed to cool to ambient temperature and filtered through a pad of diatomaceous earth. The pad was washed with ethyl acetate (8 L). The combined filtrate and washes were washed with water (4 L), saturated aqueous ammonium chloride (2 * 4 L) and brine (2 * 4 L) and concentrated in vacuo to dryness. The residue was purified by recrystallization from te/t-butyl methyl ether (0.4 L) and heptane (0.8 L), followed by drying of the resultant solid in vacuo at 40-50 °C for 8 h to afford (7S)-1 ‘-{[5- (trifluoromethyl)furan-2-yl]methyl}spiro[furo[2,3-f][1 ,3]benzodioxole-7,3’-indol]-2′(1 ‘H)- one (0.37 kg) as a colorless solid in 61% yield: 1H NMR (300 MHz, CDCI3) δ 7.29-6.96 (m, 4H), 6.73 (s, 1 H), 6.50 (s, 1 H), 6.38 (s, 1 H), 6.09 (s, 1 H), 5.85 (br s, 2H), 5.06 (d, J = 16.0 Hz,1 H), 4.93-4.84 (m, 2H), 4.68-4.65 (m, 1 H); MS (ES+) m/z 429.8 (M + 1 ); ee (enantiomeric excess) > 99% (HPLC, Chiralpak IA).

PATENT
CadieuxJ.-J.ChafeevM.ChowdhuryS.FuJ.JiaQ.AbelS.El-SayedE.HuthmannE.IsarnoT. Synthetic Methods For Spiro-Oxindole Compounds. U.S. Patent 8,445,696, May 21, 2013.
PATENT
SunS.FuJ.ChowdhuryS.HemeonI. W.GrimwoodM. E.MansourT. S. Asymmetric Syntheses of Spiro-Oxindole Compounds Useful As Therapeutic Agents. U.S. Patent 9,487,535, Nov 08, 2016.
PAPER
Abstract Image

TV-45070 is a small-molecule lactam containing a chiral spiro-ether that has been reported as a potential topical therapy for pain associated with the Nav1.7 sodium ion channel encoded by the gene SCN9A. A pilot-scale synthesis is presented that is highlighted by an asymmetric aldol coupling at ambient temperature, used to create a quaternary chiral center. Although only a moderate ee is obtained, the removal of the undesired isomer is achieved through preferential precipitation of a near racemic mixture from the reaction, leaving the enantiopure isomer in solution. Cyclization to form the final API uses an uncommon diphenylphosphine-based leaving group which proved successful on the neopentyl system when other traditional leaving groups failed.

The First Asymmetric Pilot-Scale Synthesis of TV-45070

Chemical Process Research and Development, Analytical Research and Development, Teva Branded Pharmaceutical Products R&D Inc., 383 Phoenixville Pike, Malvern, Pennsylvania 19355, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.7b00237
Publication Date (Web): September 8, 2017
Copyright © 2017 American Chemical Society

*E-mail: jasclafan@yahoo.com.

(S)-1′-[(5-Methyl-2-furyl)methyl]spiro[6H-furo[3,2-f][1,3]benzodioxole-7,3′-indoline]-2′-one (1)

1H NMR (DMSO, 400 MHz) δ 7.32 (t, J = 7.7 Hz, 1H), 7.20 (m, 3H), 7.07 (t, J = 7.3 Hz, 1H), 6.77 (d, J= 3.3 Hz, 1H), 6.72 (s, 1H), 6.10 (s, 1H), 5.94 (d, J = 9.1 Hz, 1H), 5.94 (d, J = 9.1 Hz, 1H), 5.13 (d, J = 16.5 Hz, 1H), 5.02 (d, J = 16.5 Hz, 1H), 4.82 (d, J = 9.5 Hz, 1H), 4.73 (d, J = 9.5 Hz, 1H).
13C NMR (100 MHz, DMSO-d6): 176.48, 155.28, 153.02, 148.40, 141.80, 141.51, 139.54 (q, JCF = 41.9 Hz), 131.63, 128.79, 123.64, 123.29, 119.69, 118.92 (q, JCF = 266.4 Hz), 114.01 (q, JCF = 2.9 Hz) 109.86, 109.21, 102.55, 101.44, 93.31, 79.52, 57.41, 36.44.

References

  1. Jump up to:a b c Bagal, Sharan K.; Chapman, Mark L.; Marron, Brian E.; Prime, Rebecca; Ian Storer, R.; Swain, Nigel A. (2014). “Recent progress in sodium channel modulators for pain”. Bioorganic & Medicinal Chemistry Letters24 (16): 3690–9. ISSN 0960-894XPMID 25060923doi:10.1016/j.bmcl.2014.06.038.
  2. Jump up to:a b Stephen McMahon; Martin Koltzenburg; Irene Tracey; Dennis C. Turk (1 March 2013). Wall & Melzack’s Textbook of Pain: Expert Consult – Online. Elsevier Health Sciences. p. 508. ISBN 0-7020-5374-0.
  3. Jump up to:a b Xenon Pharma. “TV-45070: A Small Molecule for the Treatment of the Orphan Disease EM and Other Pain Disorders”.
  4. Jump up to:a b Xenon Pharma (2012). “Teva and Xenon Announce Teva’s World Wide License of Xenon’s Pain Drug XEN402”.

External links

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US2016326184 SYNTHETIC METHODS FOR SPIRO-OXINDOLE COMPOUNDS 2016-01-06
US2017095449 PHARMACEUTICAL COMPOSITIONS OF SPIRO-OXINDOLE COMPOUND FOR TOPICAL ADMINISTRATION AND THEIR USE AS THERAPEUTIC AGENTS 2016-10-11
Patent ID

Patent Title

Submitted Date

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US2015216794 METHODS OF TREATING PAIN ASSOCIATED WITH OSTEOARTHRITIS OF A JOINT WITH A TOPICAL FORMULATION OF A SPIRO-OXINDOLE COMPOUND 2015-02-05 2015-08-06
US9682033 METHODS OF TREATING POSTHERPETIC NEURALGIA WITH A TOPICAL FORMULATION OF A SPIRO-OXINDOLE COMPOUND 2016-02-05 2016-08-11
US2016166541 Methods For Identifying Analgesic Agents 2016-01-27 2016-06-16
US2017066777 ASYMMETRIC SYNTHESES FOR SPIRO-OXINDOLE COMPOUNDS USEFUL AS THERAPEUTIC AGENTS 2016-09-14
US2017073351 ENANTIOMERS OF SPIRO-OXINDOLE COMPOUNDS AND THEIR USES AS THERAPEUTIC AGENTS 2016-09-28
Patent ID

Patent Title

Submitted Date

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US8742109 Synthetic methods for spiro-oxindole compounds 2012-09-14 2014-06-03
US8883840 Enantiomers of spiro-oxindole compounds and their uses as therapeutic agents 2012-09-14 2014-11-11
US9260446 SYNTHETIC METHODS FOR SPIRO-OXINDOLE COMPOUNDS 2014-05-07 2014-11-13
US9278088 Methods for Identifying Analgesic Agents 2013-04-11 2013-08-15
US9480677 ENANTIOMERS OF SPIRO-OXINDOLE COMPOUNDS AND THEIR USES AS THERAPEUTIC AGENTS 2014-10-09 2015-01-22
Patent ID Patent Title Submitted Date Granted Date
US8450358 ENANTIOMERS OF SPIRO-OXINDOLE COMPOUNDS AND THEIR USES AS THERAPEUTIC AGENTS 2010-12-30
US2011086899 PHARMACEUTICAL COMPOSITIONS FOR ORAL ADMINISTRATION 2011-04-14
US8445696 SYNTHETIC METHODS FOR SPIRO-OXINDOLE COMPOUNDS 2011-04-14
US9487535 ASYMMETRIC SYNTHESES FOR SPIRO-OXINDOLE COMPOUNDS USEFUL AS THERAPEUTIC AGENTS 2013-03-11 2013-10-17
US9504671 PHARMACEUTICAL COMPOSITIONS OF SPIRO-OXINDOLE COMPOUND FOR TOPICAL ADMINISTRATION AND THEIR USE AS THERAPEUTIC AGENTS 2011-02-25 2013-06-06
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Reference
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2 E.J. COREY; M.C. NOE, ORG. SYNTH., vol. 80, 2003, pages 38 – 45
3 GARST, J. F.; UNGVARY, F.: “Grignard Reagents”, 2000, JOHN WILEY & SONS, article “Mechanism of Grignard reagent formation“, pages: 185 – 275
4 GREENE, T.W.; P.G.M. WUTS: “Greene’s Protective Groups in Organic Synthesis, 4th Ed.,“, 2006, WILEY
5 GREENE, T.W.; WUTS, P.G.M.: “Greene’s Protective Groups in Organic Synthesis, 4th Ed.“, 2006, WILEY
6 HUGHES, D.L., ORG. PREP., vol. 28, 1996, pages 127 – 164
7 KUMARA SWAMY, K.C. ET AL.: “Mitsunobu and Related Reactions: Advances and Applications“, CHEM. REV., vol. 109, 2009, pages 2551 – 2651, XP055023394, DOI: doi:10.1021/cr800278z
8 MERSMANN, A.: “Crystallization Technology Handbook; 2nd ed.“, 2001, CRC
9 SMITH, M.; BAND J. MARCH: “Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 5th edition“, December 2000, WILEY
10 SMITH, M.B.; J. MARCH: “Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 5th edition“, December 2000, WILEY
11 * TAKASHI OOI ET AL: “Recent Advances in Asymmetric Phase-Transfer Catalysis“, ANGEWANDTE CHEMIE INTERNATIONAL EDITION, vol. 46, no. 23, 4 June 2007 (2007-06-04), pages 4222 – 4266, XP055060024, ISSN: 1433-7851, DOI: 10.1002/anie.200601737
Citing Patent Filing date Publication date Applicant Title
WO2016109795A1 31 Dec 2015 7 Jul 2016 Concert Pharmaceuticals, Inc. Deuterated funapide and difluorofunapide
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US9682033 5 Feb 2016 20 Jun 2017 Teva Pharmaceuticals International Gmbh Methods of treating postherpetic neuralgia with a topical formulation of a spiro-oxindole compound
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Funapide
Funapide.svg
Clinical data
Routes of
administration
By mouthtopical
ATC code
  • None
Identifiers
CAS Number
PubChem CID
ChemSpider
UNII
Chemical and physical data
Formula C22H14F3NO5
Molar mass 429.34547 g/mol
3D model (JSmol)
//////////TV 45070,  XEN 402, TEVA, XENON, Postherpetic neuralgia, PHN, PHASE 2, Funapide, фунапид , فونابيد , 呋纳匹特 , Orphan Drug Status
C1C2(C3=CC=CC=C3N(C2=O)CC4=CC=C(O4)C(F)(F)F)C5=CC6=C(C=C5O1)OCO6

Prexasertib , прексасертиб , بريكساسيرتيب , 普瑞色替 ,


Prexasertib.svg

Prexasertib

Captisol® enabled prexasertib; CHK1 Inhibitor II; LY 2606368; LY2606368 MsOH H2O

5-(5-(2-(3-aminopropoxy)-6-methoxyphenyl)-1H-pyrazol-3-ylamino)pyrazine-2-carbonitrile

2-Pyrazinecarbonitrile, 5-[[5-[2-(3-aminopropoxy)-6-methoxyphenyl]-1H-pyrazol-3-yl]amino]-

Name Prexasertib
Lab Codes LY-2606368
Chemical Name 5-({5-[2-(3-aminopropoxy)-6-methoxyphenyl]-1H-pyrazol-3-yl}amino)pyrazine-2-carbonitrile
Chemical Structure ChemSpider 2D Image | prexasertib | C18H19N7O2
Molecular Formula C18H19N7O2
UNII UNII:820NH671E6
Cas Registry Number 1234015-52-1
OTHER NAMES
прексасертиб [Russian] [INN]
بريكساسيرتيب [Arabic] [INN]
普瑞色替 [Chinese] [INN]
Originator Array BioPharma
Developer Eli Lilly, National Cancer Institute
Mechanism Of Action Checkpoint kinase inhibitors, Chk-1 inhibitors
Who Atc Codes L01X-E (Protein kinase inhibitors)
Ephmra Codes L1H (Protein Kinase Inhibitor Antineoplastics)
Indication Breast cancer, Ovarian cancer, Solid tumor, Head and neck cancer, Leukemia, Neoplasm Metastasis, Colorectal Neoplasms, Squamous Cell Carcinoma

Image result for Array BioPharma

Image result for ELI LILLY

Image result for Prexasertib2100300-72-7 CAS

Image result for Prexasertib

Prexasertib mesylate hydrate
CAS#: 1234015-57-6 (mesylate hydrate)
Chemical Formula: C19H25N7O6S
Molecular Weight: 479.512, CODE LY-2940930
LY-2606368 (free base)

Image result for Prexasertib

Prexasertib mesylate ANHYDROUS
CAS#: 1234015-55-4 (mesylate)
Chemical Formula: C19H23N7O5S
Molecular Weight: 461.497

2D chemical structure of 1234015-54-3

Prexasertib dihydrochloride
1234015-54-3. MW: 438.3169


LY2606368 is a small-molecule Chk-1 inhibitors invented by Array and being developed by Eli Lilly and Company. Lilly is responsible for all clinical development and commercialization activities. Chk-1 is a protein kinase that regulates the tumor cell’s response to DNA damage often caused by treatment with chemotherapy. In response to DNA damage, Chk-1 blocks cell cycle progression in order to allow for repair of damaged DNA, thereby limiting the efficacy of chemotherapeutic agents. Inhibiting Chk-1 in combination with chemotherapy can enhance tumor cell death by preventing these cells from recovering from DNA damage.

Originator Array BioPharma; Eli Lilly

Developer Eli Lilly; National Cancer Institute (USA)

Class Antineoplastics; Nitriles; Pyrazines; Pyrazoles; Small molecules

Mechanism of Action Checkpoint kinase 1 inhibitors; Checkpoint kinase 2 inhibitors

Highest Development Phases

  • Phase II Breast cancer; Ovarian cancer; Small cell lung cancer; Solid tumours
  • Phase I Acute myeloid leukaemia; Colorectal cancer; Head and neck cancer; Myelodysplastic syndromes; Non-small cell lung cancer

Most Recent Events

  • 10 Apr 2017 Eli Lilly completes a phase I trial for Solid tumours (Late-stage disease, Second-line therapy or greater) in Japan (NCT02514603)
  • 10 Mar 2017 Phase-I clinical trials in Solid tumours (Combination therapy, Metastatic disease, Inoperable/Unresectable) in USA (IV) (NCT03057145)
  • 22 Feb 2017 Khanh Do and AstraZeneca plan a phase H trial for Solid tumour (Combination therapy, Metastatic disease, Inoperable/Unresectable) in USA (NCT03057145)

Prexasertib (LY2606368) is a small molecule checkpoint kinase inhibitor, mainly active against CHEK1, with minor activity against CHEK2. This causes induction of DNA double-strand breaks resulting in apoptosis. It is in development by Eli Lilly[1]

A phase II clinical trial for the treatment of small cell lung cancer is expected to be complete in December 2017.[2]

an aminopyrazole compound, or a pharmaceutically acceptable salt thereof or a solvate of the salt, that inhibits Chkl and is useful for treating cancers characterized by defects in deoxyribonucleic acid (DNA) replication, chromosome segregation, or cell division.

Chkl is a protein kinase that lies downstream from Atm and/or Atr in the DNA damage checkpoint signal transduction pathway. In mammalian cells, Chkl is phosphorylated in response to agents that cause DNA damage including ionizing radiation (IR), ultraviolet (UV) light, and hydroxyurea. This phosphorylation which activates Chkl in mammalian cells is dependent on Atr. Chkl plays a role in the Atr dependent DNA damage checkpoint leading to arrest in S phase and at G2M. Chkl phosphorylates and inactivates Cdc25A, the dual-specificity phosphatase that normally dephosphorylates cyclin E/Cdk2, halting progression through S-phase. Chkl also phosphorylates and inactivates Cdc25C, the dual specificity phosphatase that dephosphorylates cyclin B/Cdc2 (also known as Cdkl) arresting cell cycle progression at the boundary of G2 and mitosis (Fernery et al, Science, 277: 1495-1, 1997). In both cases, regulation of Cdk activity induces a cell cycle arrest to prevent cells from entering mitosis in the presence of DNA damage or unreplicated DNA. Various inhibitors of Chkl have been reported. See for example, WO 05/066163,

WO 04/063198, WO 03/093297 and WO 02/070494. In addition, a series of aminopyrazole Chkl inhibitors is disclosed in WO 05/009435.

However, there is still a need for Chkl inhibitors that are potent inhibitors of the cell cycle checkpoints that can act effectively as potentiators of DNA damaging agents. The present invention provides a novel aminopyrazole compound, or a pharmaceutically acceptable salt thereof or solvate of the salt, that is a potent inhibitor of Chkl . The compound, or a pharmaceutically acceptable salt thereof or a solvate of the salt, potently abrogates a Chkl mediated cell cycle arrest induced by treatment with DNA damaging agents in tissue culture and in vivo. Furthermore, the compound, or a pharmaceutically acceptable salt thereof or a solvate of the salt, of the present invention also provides inhibition of Chk2, which may be beneficial for the treatment of cancer. Additionally, the lack of inhibition of certain other protein kinases, such as CDKl, may provide a -2- therapeutic benefit by minimizing undesired effects. Furthermore, the compound, or a pharmaceutically acceptable salt thereof or a solvate of the salt, of the present invention inhibits cell proliferation of cancer cells by a mechanism dependent on Chkl inhibition.

Inventors Francine S. FarouzRyan Coatsworth HolcombRamesh KasarSteven Scott Myers
Applicant Eli Lilly And Company

WO 2010077758

Preparation 8

tert-Butyl 3-(2-(3-(5-cyanopyrazin-2-ylamino)-lH-pyrazol-5-yl)-3- methoxyphenoxy)propylcarbamate

Figure imgf000025_0002

A solution of tert-butyl 3-(2-(3-(5-bromopyrazin-2-ylamino)-lH-pyrazol-5-yl)-3- methoxyphenoxy)propylcarbamate (0.378 g, 0.730 mmol) and zinc cyanide (0.10 g, 0.870 mmol) in DMF (10 mL) is degassed with a stream of nitrogen for one hour and then -25- heated to 80 0C. To the reaction is added Pd(Ph3P)4 (0.080 g, 0.070 mmol), and the mixture is heated overnight. The reaction is cooled to room temperature and concentrated under reduced pressure. The residue is purified by silica gel chromatography (CH2Cl2/Me0H) to give 0.251 g (73%) of the title compound.

Preparation 12 tert-Butyl 3-(2-(3-(5-cyanopyrazin-2-ylamino)-lH-pyrazol-5-yl)-3- methoxyphenoxy)propylcarbamate

Figure imgf000028_0001

A 5 L flange-neck round-bottom flask equipped with an air stirrer rod and paddle, thermometer, pressure-equalizing dropping funnel, and nitrogen bubbler is charged with 5-(5-(2-hydroxy-6-methoxy-phenyl)-lH-pyrazol-3-ylamino)-pyrazine-2-carbonitrile (47.0 g, 152 mmol) and anhydrous THF (1.2 L). The stirred suspension, under nitrogen, is cooled to 0 0C. A separate 2 L 3 -necked round-bottom flask equipped with a large -28- magnetic stirring bar, thermometer, and nitrogen bubbler is charged with triphenylphosphine (44.0 g; 168 mmol) and anhydrous THF (600 mL). The stirred solution, under nitrogen, is cooled to 0 0C and diisopropylazodicarboxylate (34.2 g; 169 mmol) is added and a milky solution is formed. After 3-4 min, a solution of7-butyl-N-(3- hydroxypropyl)-carbamate (30.3 g, 173 mmol) in anhydrous THF (100 mL) is added and the mixture is stirred for 3-4 min. This mixture is then added over 5 min to the stirred suspension of starting material at 0 0C. The reaction mixture quickly becomes a dark solution and is allowed to slowly warm up to room temperature. After 6.5 h, more reagents are prepared as above using PPh3 (8 g), DIAD (6.2 g) and carbamate (5.4 g) in anhydrous THF (150 mL). The mixture is added to the reaction mixture, cooled to -5 0C and left to warm up to room temperature overnight. The solvent is removed in vacuo. The resulting viscous solution is loaded onto a pad of silica and product is eluted with ethyl acetate. The concentrated fractions are separately triturated with methanol and resulting solids are collected by filtration. The combined solids are triturated again with methanol (400 mL) and then isolated by filtration and dried in vacuo at 50 0C overnight to give 31.3 g of desired product. LC-ES/MS m/z 466.2 [M+ 1]+.

Example 2

5 -(5 -(2-(3 -Aminopropoxy)-6-methoxyphenyl)- 1 H-pyrazol-3 -ylamino)pyrazine-2- carbonitrile dihydrogen chloride salt

Figure imgf000029_0001

A 5 L flange-neck, round-bottom flask equipped with an air stirrer rod and paddle, thermometer, and air condenser with bubbler attached, is charged with tert-bvXyl 3-(2-(3- (5-cyanopyrazin-2-ylamino)-lH-pyrazol-5-yl)-3-methoxyphenoxy)propylcarbamate (30.9 g, 66.3 mmol) and ethyl acetate (3 L). The mechanically stirred yellow suspension is cooled to just below 10 0C. Then hydrogen chloride from a lecture bottle is bubbled in -29- vigorously through a gas inlet tube for 15 min with the ice-bath still in place. After 5 h the mixture is noticeably thickened in appearance. The solid is collected by filtration, washed with ethyl acetate, and then dried in vacuo at 60 0C overnight to give 30.0 g. 1H NMR (400 MHz, DMSO-d6) δ 2.05 (m, 2H), 2.96 (m, 2H), 3.81 (s, 3H), 4.12 (t, J = 5.8 Hz, 2H), 6.08 (br s, 3H), 6.777 (d, J = 8.2 Hz, IH), 6.782 (d, J = 8.2 Hz, IH), 6.88 (br s, IH), 7.34 (t, J = 8.2 Hz, IH), 8.09 (br s, IH), 8.55 (br s, IH), 8.71 (s, IH), 10.83 (s, IH), 12.43 (br s, IH). LC-ES/MS m/z 366.2 [M+lf.

Example 3 5 -(5 -(2-(3 -Aminopropoxy)-6-methoxyphenyl)- 1 H-pyrazol-3 -ylamino)pyrazine-2- carbonitrile

Figure imgf000030_0001

5-(5-(2-(3-Aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-ylamino)pyrazine-2- carbonitrile dihydrogen chloride salt (3.0 g, 6.84 mmol) is suspended in 200 mL of CH2Cl2. 1 N NaOH is added (200 mL, 200 mmol). The mixture is magnetically stirred under nitrogen at room temperature for 5 h. The solid is collected by filtration and washed thoroughly with water. The filter cake is dried in vacuo at 50 0C overnight to give 2.26 g (90%) of the free base as a yellow solid. 1H NMR (400 MHz, DMSO-d6) δ 1.81 (m, 2H), 2.73 (t, J = 6.2 Hz, 2H), 3.82 (s, 3H), 4.09 (t, J = 6.2 Hz, 2H), 6.76 (t, J = 8.2 Hz, 2H), 6.93 (br s, IH), 7.31 (t, J = 8.2 Hz, IH), 8.52 (br s, IH), 8.67 (s, IH). LC- MS /ES m/z 366.2 [M+ 1]+.

Example 4

5 -(5 -(2-(3 -Aminopropoxy)-6-methoxyphenyl)- 1 H-pyrazol-3 -ylamino)pyrazine-2- carbonitrile methanesulfonic acid salt -30-

Figure imgf000031_0001

5-(5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-ylamino)pyrazine-2- carbonitrile (1.0 g, 2.74 mmol) is suspended in MeOH (100 mL). A I M solution of methanesulfonic acid in MeOH (2.74 mL, 2.74 mmol) is added to the mixture dropwise with stirring. The solid nearly completely dissolves and is sonicated and stirred for 15 min, filtered, and concentrated to 50 mL. The solution is cooled overnight at -15 0C and the solid that forms is collected by filtration. The solid is dried in a vacuum oven overnight to give 0.938 g (74%) of a yellow solid. 1H NMR (400 MHz, DMSO-d6) δ 1.97 (m, 2H), 2.28 (s, 3H), 2.95 (m, 2H), 3.79 (s, 3H), 4.09 (t, J = 5.9 Hz, 2H), 6.753 (d, J = 8.4 Hz, IH), 6.766 (d, J = 8.4 Hz, IH), 6.85 (br s, IH), 7.33 (t, J = 8.4 Hz, IH), 7.67 (br s, 3H), 8.49 (br s, IH), 8.64 (s, IH), 10.70 (s, IH), 12.31 (s, IH). LC-ES/MS m/z 366.2 [M+l]+.

Preparation 18 tert-Butyl 3-(2-(3-(5-cyanopyrazin-2-ylamino)-lH-pyrazol-5-yl)-3- methoxyphenoxy)propylcarbamate

Figure imgf000035_0001

5-(5-(2-Hydroxy-6-methoxyphenyl)-lH-pyrazol-3-ylamino)pyrazine-2- carbonitrile (618 g, 1.62 mol) is slurried in tetrahydrofuran (6.18 L, 10 volumes) and chilled to -5 to 0 0C with an acetone/ice bath. Triethylamine (330 g, 3.25 mol) is added through an addition funnel over 30 – 40 min at -5 to 5 0C. The resulting slurry is stirred at -5 to 5 0C for 60 – 90 min. The insoluble triethylamine hydrochloride is filtered and the solution of the phenol ((5-(2-hydroxy-6-methoxyphenyl)-lH-pyrazol-3- ylamino)pyrazine-2-carbonitrile) collected in an appropriate reaction vessel. The cake is rinsed with THF (1.24 L). The THF solution of the phenol is held at 15 to 20 0C until needed.

Triphenylphosphine (1074 g, 4.05 mol) is dissolved at room temperature in THF (4.33 L). The clear colorless solution is cooled with an acetone/ice bath to -5 to 5 0C. Diisopropylazodicarboxylate (795 g, 3.89 mol) is added dropwise through an addition funnel over 40 – 60 min, keeping the temperature below 10 0C. The resulting thick white slurry is cooled back to -5 to 0 0C. tert-Butyl 3-hydroxypropylcarbamate (717g, 4.05 moles) is dissolved in a minimum of THF (800 mL). The tert-butyl 3- hydroxypropylcarbamate/THF solution is added, through an addition funnel, over 20 – 30 -35- min at -5 to 5 0C to the reagent slurry. The prepared reagent is stirred in the ice bath at -5 to 0 0C until ready for use.

The prepared reagent slurry (20%) is added to the substrate solution at 15 to 20 0C. The remaining reagent is returned to the ice bath. The substrate solution is stirred at ambient for 30 min, then sampled for HPLC. A second approximately 20% portion of the reagent is added to the substrate, stirred at ambient and sampled as before. Addition of the reagent is continued with monitoring for reaction completion by HPLC. The completed reaction is concentrated and triturated with warm methanol (4.33 L, 50 – 60 0C) followed by cooling in an ice bath. The resulting yellow precipitate is filtered, rinsed with cold MeOH (2 L), and dried to constant weight to provide 544 g (72%) of the title compound, mp 214 – 216 0C; ES/MS m/z 466.2 [M+l]+.

Example 5

2-Pyrazinecarbonitrile, 5-[[5-[-[2-(3-aminopropyl)-6-methoxyphenyl]-lH-pyrazol-3- yl]amino] monomesylate monohydrate (Chemical Abstracts nomenclature)

Figure imgf000036_0001

tert-Butyl 3-(2-(3-(5-cyanopyrazin-2-ylamino)-lH-pyrazol-5-yl)-3- methoxyphenoxy)propylcarbamate (1430 g, 3.07 mol) is slurried with acetone (21.5 L) in a 30 L reactor. Methanesulfonic acid (1484 g, 15.36 mol) is added through an addition funnel in a moderate stream. The slurry is warmed to reflux at about 52 0C for 1 to 3 h and monitored for reaction completion by HPLC analysis. The completed reaction is cooled from reflux to 15 to 20 0C over 4.5 h. The yellow slurry of 2-pyrazinecarbonitrile, 5-[[5-[-[2-(3-aminopropyl)-6-methoxyphenyl]-lH-pyrazol-3-yl]amino] dimesylate salt is filtered, rinsed with acetone (7 L) and dried in a vacuum oven. The dimesylate salt, (1608 g, 2.88 mol) is slurried in water (16 L). Sodium hydroxide (aqueous 50%, 228 g, 2.85 mol) is slowly poured into the slurry. The slurry is -36- heated to 60 0C and stirred for one hour. It is then cooled to 16 0C over 4 h and filtered. The wet filter cake is rinsed with acetone (4 L) and dried to constant weight in a vacuum oven at 40 0C to provide 833 g (94%) of 2-pyrazinecarbonitrile, 5-[[5-[-[2-(3- aminopropyl)-6-methoxyphenyl]-lH-pyrazol-3-yl]amino] monomesylate monohydrate. mp 222.6 0C; ES/MS m/z 366.2 [M+l]+.

Example 5a

2-Pyrazinecarbonitrile, 5-[[5-[-[2-(3-aminopropyl)-6-methoxyphenyl]-lH-pyrazol-3- yl] amino] monomesylate monohydrate (Chemical Abstracts nomenclature)

Crude 2-pyrazinecarbonitrile, 5 -[ [5 – [- [2-(3 -aminopropyl)-6-methoxyphenyl]- IH- pyrazol-3-yl] amino] monomesylate monohydrate is purified using the following procedure. The technical grade 2-pyrazinecarbonitrile, 5-[[5-[-[2-(3-aminopropyl)-6- methoxyphenyl]-lH-pyrazol-3-yl] amino] mono mesylate mono hydrate (1221 g, 2.55 mol) is slurried in a solvent mixture of 1: 1 acetone/water (14.7 L). The solid is dissolved by warming the mixture to 50 – 55 0C. The solution is polish filtrated while at 50 – 55 0C through a 0.22μ cartridge filter. The solution is slowly cooled to the seeding temperature of about 42 – 45 0C and seeded. Slow cooling is continued over the next 30 – 60 min to confirm nucleation. The thin slurry is cooled from 38 to 15 0C over 3 h. A vacuum distillation is set up and the acetone removed at 110 – 90 mm and 20 – 30 0C. The mixture is cooled from 30 to 15 0C over 14 h, held at 15 0C for 2 h, and then filtered. The recrystallized material is rinsed with 19: 1 water/acetone (2 L) and then water (6 L) and dried to constant weight in a vacuum oven at 40 0C to provide 1024 g (83.9%) of the title compound, mp 222.6 0C; ES/MS m/z 366.2 [M+l]+. X-ray powder diffraction (XRPD) patterns may be obtained on a Bruker D8

Advance powder diffractometer, equipped with a CuKa source (λ=l.54056 angstrom) operating at 40 kV and 40 mA with a position-sensitive detector. Each sample is scanned between 4° and 35° in °2Θ ± 0.02 using a step size of 0.026° in 2Θ ± 0.02 and a step time of 0.3 seconds, with a 0.6 mm divergence slit and a 10.39 mm detector slit. Primary and secondary Soller slits are each at 2°; antiscattering slit is 6.17 mm; the air scatter sink is in place. -37-

Characteristic peak positions and relative intensities:

Figure imgf000038_0001

Differential scanning calorimetry (DSC) analyses may be carried out on a Mettler- Toledo DSC unit (Model DSC822e). Samples are heated in closed aluminum pans with pinhole from 25 to 350 0C at 10 °C/min with a nitrogen purge of 50 mL/min. Thermogravimetric analysis (TGA) may be carried out on a Mettler Toledo TGA unit (Model TGA/SDTA 85Ie). Samples are heated in sealed aluminum pans with a pinhole from 25 to 350 0C at 10 0C /min with a nitrogen purge of 50 mL/min.

The thermal profile from DSC shows a weak, broad endotherm form 80 – 1400C followed by a sharp melting endotherm at 222 0C, onset (225 0C, peak). A mass loss of 4% is seen by the TGA from 25 – 140 0C.

PATENT

US 20110144126

WO 2017015124

WO 2017100071

WO 2017105982

WO 2016051409

PATENT

WO 2017100071

Preparation 1

tert-Butyl (E)-(3-(2-(3-(dimethylamino)ac^’loyl)-3-me1hoxyphenox50propyl)carbamate

L _l H

Combine l-(2-hydroxy-6-methox>’phenyl)e1han-l-one (79.6 kg, 479 mol) and 1,1-<iimethoxy-N,N-dimemylmethanamino (71.7 kg, 603.54 mol) with DMF (126 kg). Heat to 85-90 °C for 12 hours. Cool the reaction mixture containing intermediate (E)-3-(dimethylamino)-l-(2-hydroxy-6-methoxyphenyl)prop-2-en-l-one (mp 84.74 °C) to ambient temperature and add anhydrous potassium phosphate (136 kg, 637.07 mol) and tert-butyl (3-bromopropyl)carbamate (145 kg, 608.33 mol). Stir the reaction for 15 hours at ambient temperature. Filter the mixture and wash the filter cake with ΜΓΒΕ (3 χ , 433 kg, 300 kg, and 350 kg). Add water (136 kg) and aqueous sodium chloride (25% solution, 552 kg) to the combined MTBE organic solutions. Separate the organic and aqueous phases. Back-extract the resulting aqueous phase with MTBE (309 kg) and add the MTBE layer to the organic solution. Add an aqueous sodium chloride solution (25% solution, 660 kg) to the combined organic extracts and separate the layers. Concentrate the organic extracts to 1,040 kg – 1,200 kg and add water (400 kg) at 30-35 °C to the residue. Cool to ambient temperature and collect material by filtration as a wet cake to give the title product (228.35 kg, 90%). ES/MS (m/z): 379.22275 (M+l).

Preparation 2

tert-Butyl (3-(2-(2-cyanoacetyl)-3-methoxyphenoxy)propyl)carbamate

“9 o


 

Combine ethanol (1044 kg), hydroxyl amino hydrochloride (30 kg, 431.7 mol), and terr-butyl (E)-(3-(2-(3-(^me%lamino)acryloyl)-3-

methoxyphenoxy)propyl)carbamate (228.35 kg, 72% as a wet water solid, 434.9 mol) to form a solution. Heat the solution to 35 – 40 °C for 4-6 hours. Cool the reaction to ambient temperature and concentrate to a residue. Add MTBE (300 kg) to the residue and concentrate the solution to 160 kg – 240 kg. Add MTBE (270 kg) and concentrate the solution. Add MTBE (630 kg), water (358 kg), and sodium chloride solution (80 kg, 25% aqueous) and stir for 20 minutes at ambient temperature. Let the mixture stand for 30 minutes. Separate the aqueous layer. Add water (360 kg) and sodium chloride solution (82 kg, 25% sodium chloride) to the organic phase. Stir for 20 minutes at ambient temperature. Let the mixture stand for 30 minutes. Separate the aqueous portion. Add sodium chloride solution (400 kg, 25 % aqueous) to the organic portion. Stir for 20 minutes at ambient temperature. Let the mixture stand for 30 minutes at ambient temperature. Separate the aqueous portion. Concentrate the organic portion to 160 kg – 240 kg at 40 °C. Add ethanol (296 kg) to the organic portion. Concentrate the solution to 160 kg to 240 kg at 40 °C to provide an intermediate of tert-butyl (3-(2-(isoxazol-5-yl)-3-methox>’phenoxy)propyl)carbamate. Add ethanol (143 kg) and water (160 kg) to the concentrated solution. Add potassium hydroxide (31.8 kg) at 40 °C. Add ethanol (80 kg) and adjust the temperature to 45-50 °C. Stir for 4-6 hours at 45-50 °C and concentrate to 160 kg – 240 kg at 40 °C. Add water to the concentrate (160 kg) and acetic acid (9.0 kg) drop-wise to adjust the pH to 10-12 while mamtaining the temperature of the solution at 25 to 35 °C. Add ethyl acetate (771 kg) and acetic acid drop-wise to adjust the pH to 5-7 while maintaining the temperature of the solution at 25-35 °C. Add sodium chloride solution (118 kg, 25% aqueous solution). Stir the mixture for 20 minutes at ambient temperature. Let the solution stand for 30 minutes at ambient temperature. Separate Ihe aqueous portion. Heat the organic portion to 30-35 °C. Add water (358 kg) drop-wise. Stir the solution for 20 minutes while maintaining the temperature at 30 to 35 °C. Let the mixture stand for 30 minutes and separate the aqueous portion. Wash the organic portion with sodium chloride solution (588 kg, 25% aqueous) and concentrate the organic portion to 400 kg – 480 kg at 40-50 °C. Heat the concentrated solution to 50 °C to form a solution. Maintain the solution at 50 °C and add M-heptane (469 kg) drop-wise. Stir the solution for 3 hours at 50 °C before slowly cooling to ambient temperature to crystallize the product. Stir at ambient temperature for 15 hours and filter the crystals. Wash the crystals with ethanol/«-heptane (1 :2, 250 kg) and dry at 45 °C for 24 hours to provide the title compound (133.4 kg, 79.9%), rap. 104.22 °C,

Example 1

5-(5-(2-(3-Ammopropoxy)-6-memoxyphenyl)-lH-pyrazol-3-ylammo)pyrazine-2- carbonitrile (S)-lactate monohydrate

Combine a THJF solution (22%) of fcrt-butyl (3-(2-(2-cyanoacetyl)-3-memoxyphenoxy)propyl)carbamate (1.0 eqv, this is define as one volume) with hydrazine (35%, 1.5 eqv), acetic acid (glacial, 1.0 eqv), water (1 volume based on the THF solution) and methanol (2 volumes based on the THF solution). This is a continuous operation. Heat the resulting mixture to 130 °C and 1379 kPa with a rate of V/Q = 70 minutes, tau = 60. Extract the solution with toluene (4 volumes), water (1 volume), and sodium carbonate (10% aqueous, 1 eqv). Isolate Ihe toluene layer and add to DMSO (0.5 volumes). Collect a solution of the intermediate compound tert-butyl (3-(2-(3-amino-lH-pyrazol-5-yl)-3-methoxyphenoxy) propyl)carbamate (26.59 kg, 91%) in 10 days, mp = 247.17 °C as a DMSO solution (3 volumes of product). N-Eftylmorpholine (1.2 eqv) and 5-chloropyrazine-2-carbonitrile (1.15 eqv) in 2 volumes of DMSO is combined in a tube reactor at 80 °C, V/Q = 3 and tau = 170 minutes at ambient pressure. Add the product stream to methanol (20 vol). As a continuous process, filter the mixture and wash with methanol followed by MTBE. Air dry the material on the filter to give tert-butyl (3-(2-(3-((5-cyanopyrazm-2-yl)arnino)-lH-pyrazol-5-yl)-3-methox>’phenoxy) propyl)carbamate in a continuous fashion (22.2 kg, 88.7%, 8 days). Dissolve a solution of fcrt-butyl (3-(2-(3-((5-cyanopyrazin-2-yl)amino)-lH-pyrazol-5-yl)-3-methoxyphenoxy) propyl)carbamate in formic acid (99%, 142 kg) at ambient temperature and agitate for 4 hours to provide an intermediate of 5-((5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-yl)amino)pyrazine-2-carbonitrile formate. Dilute the solution with water (55 kg), (S)-lactic acid (30%, 176 kg) and distill the resulting mixture until < 22 kg formic acid remains. Crystallize the resulting residue from THF and wash with a THF -water (0.5% in THF) solution. Dry the wet cake at 30 °C at >10% relative humidity to give the title product as a white to yellow solid (24.04 kg, 85-90%), mp. 157 °C.

Alternate Preparation Example 1

5-(5-(2-(3-Ammopropoxy)-6-memoxyphenyl)-lH-pyrazol-3-ylammo)pyrazine-2- carbonitrile (S)-lactate monohydrate

Add 5-({3-[2-(3-aminopropoxy)-6-methoxyphenyl]-lH-pyrazol-5-yl}ammo)pyrazine-2-carbonitrile (4.984 g, 13.33 mmol, 97.7 wt%) to n-PrOH (15.41 g, 19.21 mL) to form a slurry. Heat the slurry to 60 °C. Add (S)-lactic acid (1.329 g, 14.75 mmol) to water (19.744 mL) and add this solution to the slurry at 58 °C. Heat the solution to 60 °C and add n-PrOH (21.07 g, 26.27 mL). Seed the solution with 5-((5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-yl)ammo)pyrazme-2-carbom^ (S)-lactate monohydrate (48.8 mg, 0.1 mmol) and cool the solution to 40 °C over 35 minutes. Add H-PrOH (60.5 mL) to the slurry at 40 °C via a syringe pump over 2 hours and maintain the temperature at 40 °C. Once complete, air cool the slurry to ambient temperature for 2 hours, the cool the mixture in ice-water for 2 hours. Filter the product, wash the wet cake with 6:1 (v/v) rc-PrOH : H20 (15 mL), followed by n-PrOH (15 mL) and dry the wet cake for 20 minutes. Dry the solid overnight at 40 °C in vacuo to give the title compound as a white to yellow solid (5.621 g, 89.1%), m.p. 157 °C.

Crystalline Example 1

Crystalline 5-(5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3- ylamino)pyrazine-2-carbonitrile (S)-lactate monohydrate Prepare a slurry having 5-(5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3 -y lamino)py razine-2-carbonitrile (368 mg, 1.0 mmol) in a 10:1 THF-water (5 mL) solution and stir at 55 °C. Add (S)-lactic acid (110 mg, 1.22 mmol) dissolved in THF (1 mL). A clear solution forms. Stir for one hour. Reduce Ihe temperature to 44 °C and stir until an off-white precipitate forms. Filter the material under vacuum, rinse with THF, and air dry to give the title compound (296 mg, 80%).

X-Ray Powder Diffraction, Crystalline Example 1 Obtain the XRPD patterns of the crystalline solids on a Bruker D4 Endeavor X-ray powder diffractometer, equipped with a CuKa source (λ = 1.54060 A) and a Vantec detector, operating at 35 kV and 50 mA. Scan the sample between 4 and 40° in 2Θ, with a step size of 0.0087° in 2Θ and a scan rate of 0.5 seconds/step, and with 0.6 mm divergence, 5.28mm fixed anti-scatter, and 9.5 mm detector slits. Pack the dry powder on a quartz sample holder and obtain a smooth surface using a glass slide. It is well known in the crystallography art that, for any given crystal form, the relative intensities of the diffraction peaks may vary due to preferred orientation resulting from factors such as crystal morphology and habit. Where the effects of preferred orientation are present, peak intensities are altered, but the characteristic peak positions of the polymorph are unchanged. See, e.g. The U. S. Pharmacopeia 35 – National Formulary 30 Chapter <941> Characterization of crystalline and partially crystalline solids by XRPD Official December 1, 2012-May 1, 2013. Furthermore, it is also well known in the

crystallography art that for any given crystal form the angular peak positions may vary slightly. For example, peak positions can shift due to a variation in the temperature or humidity at which a sample is analyzed, sample displacement, or the presence or absence of an internal standard. In the present case, a peak position variability of ± 0.2 in 2Θ will take into account these potential variations without hindering the unequivocal identification of the indicated crystal form Confirmation of a crystal form may be made based on any unique combination of distinguishing peaks (in units of ° 2Θ), typically the more prominent peaks. The crystal form diffraction patterns, collected at ambient temperature and relative humidity, were adjusted based on NIST 675 standard peaks at 8.85 and 26.77 degrees 2-theta,

Characterize a prepared sample of crystalline 5-(5-(2-(3-aminopropoxy)-6-methoxyphenyl)- lH-pyrazol-3-ylamino)pyrazine-2-carbonitrile (S)-lactate monohydrate by an XPRD pattern using CuKa radiation as having diffraction peaks (2-theta values) as described in Table 1 below. Specifically the pattern contains a peak at 12.6 in

combination with one or more of the peaks selected from the group consisting of 24.8, 25.5, 8.1, 6.6, 12.3, and 16.3 with a tolerance for the diffraction angles of 0.2 degrees.

PATENT

WO 2017105982

Example 1

5-(5-(2-(3-Aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-ylamino)pyrazine-2- carbonitrile S)-lactate monohydrate

Combine a THF solution (22%) of i<?ri-butyl (3-(2-(2-cyanoacetyl)-3-methoxyphenoxy)propyl)carbamate (1.0 eqv, this is define as one volume) with hydrazine (35%, 1.5 eqv), acetic acid (glacial, 1.0 eqv), water (1 volume based on the THF solution) and methanol (2 volumes based on the THF solution). As this is a continuous operation, grams or kg is irrelevant in this processing methodology. Heat the resulting mixture to 130 °C and 1379 kPa with a rate of V/Q = 70 minutes (where V refers to the volume of the reactor and Q refers to flow rate), tau = 60. Extract the solution with toluene (4 volumes), water (1 volume), and sodium carbonate (10% aqueous, 1 eqv). Isolate the toluene layer and add to DMSO (0.5 volumes). Collect a solution of the intermediate compound i<?ri-butyl (3-(2-(3-amino- lH-pyrazol-5-yl)-3-methoxyphenoxy)

propyl)carbamate (26.59 kg, 91%) in 10 days, mp = 247.17 °C as a DMSO solution (3 volumes of product). N-ethylmorpholine (1.2 eqv) and 5-chloropyrazine-2-carbonitrile (1.15 eqv) in 2 volumes of DMSO is combined in a tube reactor at 80 °C, V/Q = 3 and tau = 170 minutes at ambient pressure. Add the product stream to methanol (20 vol). As a continuous process, filter the mixture and wash with methanol followed by MTBE. Air dry the material on the filter to give i<?ri-butyl (3-(2-(3-((5-cyanopyrazin-2-yl)amino)-lH-pyrazol-5-yl)-3-methoxyphenoxy) propyl)carbamate in a continuous fashion (22.2 kg, 88.7%, 8 days). Dissolve a solution of i<?ri-butyl (3-(2-(3-((5-cyanopyrazin-2-yl)amino)-lH-pyrazol-5-yl)-3-methoxyphenoxy) propyl)carbamate in formic acid (99%, 142 kg) at ambient temperature and agitate for 4 hours to provide an intermediate of 5-((5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-yl)amino)pyrazine-2-carbonitrile formate. Dilute the solution with water (55 kg), (S)-lactic acid (30%, 176 kg) and distill the resulting mixture until < 22 kg formic acid remains. Crystallize the resulting residue from THF and wash with a THF -water (0.5% in THF) solution. Dry the wet cake at 30 °C at >10% relative humidity to give the title product as a white to yellow solid (24.04 kg, 85-90%), m.p. 157 °C.

Alternate Preparation Example 1

5-(5-(2-(3-Aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-ylamino)pyrazine-2- carbonitrile (S)-lactate monohydrate

Add 5-({3-[2-(3-aminopropoxy)-6-methoxyphenyl]-lH-pyrazol-5-yl}amino)pyrazine-2-carbonitrile (4.984 g, 13.33 mmol, 97.7 wt%) to n-PrOH (15.41 g, 19.21 mL) to form a slurry. Heat the slurry to 60 °C. Add (S)-lactic acid (1.329 g, 14.75 mmol) to water (19.744 mL) and add this solution to the slurry at 58 °C. Heat the solution to 60 °C and add n-PrOH (21.07 g, 26.27 mL). Seed the solution with 5-((5-(2-(3-aminopropoxy)-6-methoxyphenyl)-lH-pyrazol-3-yl)amino)pyrazine-2-carbonitrile (S)-lactate monohydrate (48.8 mg, 0.1 mmol) and cool the solution to 40 °C over 35 minutes. Add ft-PrOH (60.5 mL) to the slurry at 40 °C via a syringe pump over 2 hours and maintain the temperature at 40 °C. Once complete, air cool the slurry to ambient temperature for 2 hours, then cool the mixture in ice-water for 2 hours. Filter the product, wash the wet cake with 6:1 (v/v) n-PrOH : H20 (15 mL), followed by n-PrOH (15 mL)

and dry the wet cake for 20 minutes. Dry the solid overnight at 40 °C in vacuo to give the title compound as a white to yellow solid (5.621 g, 89.1%), m.p. 157 °C.

Clip

Kilogram-scale prexasertib monolactate monohydrate synthesis under continuous-flow CGMP conditions

Science  16 Jun 2017:
Vol. 356, Issue 6343, pp. 1144-1150
DOI: 10.1126/science.aan0745

science 20173561144

Kilogram-Scale Prexasertib Monolactate Monohydrate Synthesis under Continuous-Flow CGMP Conditions


A multidisciplinary team from Eli Lilly reports the development and implementation of eight continuous unit operations for the synthesis of ca. 3 kg API per day under CGMP conditions (K. P. Cole et al., Science 20173561144). The recent drive toward more potent APIs that have a low annual demand (<100 kg) has made continuous synthesis a viable alternative to traditional batch processes with advantages which include reducing equipment footprint and worker exposure. In this report the authors describe the enablement of three continuous synthetic steps followed by a salt formation, using surge tanks between steps to allow each step to be taken offline if online PAT detects a loss in reaction performance. A combination of MSMPRs (mixed-suspension, mixed-product removal) vessels, plug-flow reactors, and dissolve-off filters were used to perform the chemistry, with an automated 20 L rotary evaporator used to concentrate process streams and perform solvents swaps. This paper gives an excellent account of the potential solutions to continuous API synthesis and is well worth a read for anyone contemplating such methodology.
str1 str2 str3

Integrated flow synthesis and purification process for prexasertib meets high industry standards

Photograph of continuous crystallizers during processing

Source: © Eli Lilly and Company

Continuous crystallisation, shown here, and subsequent filtration have been the most difficult-to-develop part of the prexasertib production process

Eli Lilly has taken an important step away from traditional batch process drug manufacturing by using an industry-first continuous process to make a compound for phase I and II clinical trials. Workers at Lilly’s Kinsale site in Ireland, did three steps involved in producing cancer drug candidate prexasertib continuously, under current good manufacturing practice (CGMP) standards that ensure safety for human consumption.

Continuous processing relies on chemical and physical changes happening as substances flow through pipes. Isolated steps of this type are already well-established in the pharmaceutical industry. However, Lilly ‎principal research scientist Kevin Cole stresses that a series including reaction and purification steps like this has not been demonstrated before. And the company wants to go much further.

‘We envision entire synthetic routes consisting of many reaction and separation unit operations being executed simultaneously in flow, with heavy reliance on design space understanding, process analytical technologies and process modelling to ensure quality,’ Cole says. ‘We think this will drastically change the environment for pharmaceutical manufacturing.’

A scheme showing a continuous manufacturing production route for prexasertib monolactate monohydrate

Source: © Science / AAAS

The complex synthesis of prexasertib even requires the use of toxic hydrazine – used as a rocket fuel. As a result, and because of prexasertib’s toxicity, the drug was a good candidate to test out a comprehensive flow chemistry setup

In batch processes different chemical reaction and purification steps are typically done in large, costly vessels. However, this can be uneconomical when small amounts of drug molecules are needed for early stage clinical trials and, because drugs are getting more potent, increasingly in mainstream production.

By contrast, small volume continuous flow processing runs in more compact equipment in fume hoods. Flow systems can adapt to different processes, with cheap parts that can either be dedicated to specific drugs or readily replaced. The US Food and Drug Administration (FDA) has also been promoting continuous manufacturing because it integrates well with advanced process analytical technology. This helps pharmaceutical companies make high quality drugs with less FDA oversight.

Lilly chose prexasertib as its test case for such a process because it’s challenging to make. It is a chain of three aromatic rings, and one challenge comes because its central ring is formed using hydrazine. Hydrazine is used as a component in rocket fuel, and is also highly toxic. A second challenge comes from prexasertib itself, which, as a potent kinase inhibitor, is toxic to healthy cells, as well as cancerous ones, even at low doses. Lilly therefore wants to minimise its workers’ exposure.

Feeding the plant

Cole and his colleagues at Lilly’s labs in Indianapolis, US, have developed flow processes for three of the seven steps involved in prexasertib production. They start with the hydrazine step, which they could safely speed up by super-heating in the continuous process. After aqueous workup purification the solution of the two-ring intermediate solution runs into a ‘surge tank’. From there the solution flows intermittently into a rotary evaporator that removes solvents to concentrate it.

The second continuous flow step adds the third of prexasertib’s rings. In this case, the Lilly team purified the intermediate by crystallising it and filtering it out, washing away impurities. They could then redissolve the pure intermediate in formic acid, which also removes a protecting group, giving the desired prexasertib molecule. Automating this was probably the hardest part, Cole says. ‘Development of a predictive filtration model, equipment design and identification of formic acid as the solvent were keys to success,’ he explains. The final flow step then starts converting prexasertib to its final lactate salt form.

Photograph of deprotection gas/liquid reactor during processing

Source: © Eli Lilly and Company

This coil of tubes forms a low-cost deprotection gas/liquid reactor Eli Lilly uses during continuous processing of prexasertib

After developing the processes and systems in Indianapolis, Lilly shipped them to be equipped in an existing facility at its Kinsale manufacturing site at the cost of €1 million (£870,000). Once the prexasertib system was installed, the company was able to make 3kg of raw material per day for clinical trials. Cole describes the level of manual intervention needed as ‘moderate’.

Klavs Jensen from the Massachusetts Institute of Technology calls the paper describing the work ‘terrific’. ‘This work marks an important milestone in the continuous manufacturing of pharmaceuticals by demonstrating the feasibility of producing a modern kinase inhibitor under CGMP conditions,’ he says.

Likewise, Brahim Benyahia from Loughborough University, UK, calls this achievement ‘very interesting’. ‘The paper is another example that demonstrates the benefits and feasibility of the integrated continuous approach in pharma,’ he says.

Cole adds that Lilly has several other similar projects in advanced stages of development intended for the €35 million small-volume continuous plant it recently built in Kinsale. ‘We are committed to continuous manufacturing as well as full utilisation of our new facility,’ he says.

Correction: This article was updated on 16 June 2017 to clarify the chronology of the completion of the Kinsale, Ireland plant

References

REFERENCES

1: Lowery CD, VanWye AB, Dowless M, Blosser W, Falcon BL, Stewart J, Stephens J, Beckmann RP, Bence Lin A, Stancato LF. The Checkpoint Kinase 1 Inhibitor Prexasertib Induces Regression of Preclinical Models of Human Neuroblastoma. Clin Cancer Res. 2017 Mar 7. pii: clincanres.2876.2016. doi: 10.1158/1078-0432.CCR-16-2876. [Epub ahead of print] PubMed PMID: 28270495.

2: Zeng L, Beggs RR, Cooper TS, Weaver AN, Yang ES. Combining Chk1/2 inhibition with cetuximab and radiation enhances in vitro and in vivo cytotoxicity in head and neck squamous cell carcinoma. Mol Cancer Ther. 2017 Jan 30. pii: molcanther.0352.2016. doi: 10.1158/1535-7163.MCT-16-0352. [Epub ahead of print] PubMed PMID: 28138028.

3: Ghelli Luserna Di Rorà A, Iacobucci I, Imbrogno E, Papayannidis C, Derenzini E, Ferrari A, Guadagnuolo V, Robustelli V, Parisi S, Sartor C, Abbenante MC, Paolini S, Martinelli G. Prexasertib, a Chk1/Chk2 inhibitor, increases the effectiveness of conventional therapy in B-/T- cell progenitor acute lymphoblastic leukemia. Oncotarget. 2016 Aug 16;7(33):53377-53391. doi: 10.18632/oncotarget.10535. PubMed PMID: 27438145; PubMed Central PMCID: PMC5288194.

REFERENCES

1: Zeng L, Beggs RR, Cooper TS, Weaver AN, Yang ES. Combining Chk1/2 inhibition with cetuximab and radiation enhances in vitro and in vivo cytotoxicity in head and neck squamous cell carcinoma. Mol Cancer Ther. 2017 Jan 30. pii: molcanther.0352.2016. doi: 10.1158/1535-7163.MCT-16-0352. [Epub ahead of print] PubMed PMID: 28138028.

2: Ghelli Luserna Di Rorà A, Iacobucci I, Imbrogno E, Papayannidis C, Derenzini E, Ferrari A, Guadagnuolo V, Robustelli V, Parisi S, Sartor C, Abbenante MC, Paolini S, Martinelli G. Prexasertib, a Chk1/Chk2 inhibitor, increases the effectiveness of conventional therapy in B-/T- cell progenitor acute lymphoblastic leukemia. Oncotarget. 2016 Aug 16;7(33):53377-53391. doi: 10.18632/oncotarget.10535. PubMed PMID: 27438145; PubMed Central PMCID: PMC5288194.

3: King C, Diaz HB, McNeely S, Barnard D, Dempsey J, Blosser W, Beckmann R, Barda D, Marshall MS. LY2606368 Causes Replication Catastrophe and Antitumor Effects through CHK1-Dependent Mechanisms. Mol Cancer Ther. 2015 Sep;14(9):2004-13. doi: 10.1158/1535-7163.MCT-14-1037. PubMed PMID: 26141948.
4: Hong D, Infante J, Janku F, Jones S, Nguyen LM, Burris H, Naing A, Bauer TM, Piha-Paul S, Johnson FM, Kurzrock R, Golden L, Hynes S, Lin J, Lin AB, Bendell J. Phase I Study of LY2606368, a Checkpoint Kinase 1 Inhibitor, in Patients With Advanced Cancer. J Clin Oncol. 2016 May 20;34(15):1764-71. doi: 10.1200/JCO.2015.64.5788. PubMed PMID: 27044938.

Prexasertib
Prexasertib.svg
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CAS Number
ChemSpider
UNII
Chemical and physical data
Formula C18H19N7O2
Molar mass 365.40 g·mol−1
3D model (JSmol)

////////////Prexasertib, прексасертиб , بريكساسيرتيب , 普瑞色替 , PHASE 2, LY-2606368, LY 2606368

N#CC1=NC=C(NC2=NNC(C3=C(OC)C=CC=C3OCCCN)=C2)N=C1
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