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DRUG APPROVALS BY DR ANTHONY MELVIN CRASTO .....FOR BLOG HOME CLICK HERE

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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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FDA approves treatment Inrebic (fedratinib) for patients with rare bone marrow disorder


FDA approves treatment Inrebic (fedratinib) for patients with rare bone marrow disorder

Today, the U.S. Food and Drug Administration approved Inrebic (fedratinib) capsules to treat adult patients with certain types of myelofibrosis.

“Prior to today, there was one FDA-approved drug to treat patients with myelofibrosis, a rare bone marrow disorder. Our approval today provides another option for patients,” said Richard Pazdur, M.D., director of the FDA’s Oncology Center of Excellence and acting director of the Office of Hematology and Oncology Products in the FDA’s Center for Drug Evaluation and Research. “The FDA is committed to encouraging the development of treatments for patients with rare diseases and providing alternative options, as not all patients respond in the same way.”

Myelofibrosis is a chronic disorder where scar tissue forms in the bone marrow and the production of the blood cells moves from the bone marrow to the spleen and liver, causing organ enlargement. It can cause extreme fatigue, shortness of breath, pain below the ribs, fever, night sweats, itching and bone pain. When myelofibrosis occurs on its own, it is called primary myelofibrosis. Secondary myelofibrosis occurs when there is excessive red blood cell production (polycythemia vera) or excessive platelet production (essential thrombocythemia) that evolves into myelofibrosis.

Jakafi (ruxolitinib) was approved by the FDA in 2011. The approval of Inrebic for intermediate-2 or high-risk primary or secondary (post-polycythemia vera or post-essential thrombocythemia) myelofibrosis was based on the results of a clinical trial where 289 patients with myelofibrosis were randomized to receive two different doses (400 mg or 500 mg daily by mouth) of fedratinib or placebo. The clinical trial showed that 35 of 96 patients treated with the fedratinib 400 mg daily dose (the dose recommended in the approved label) experienced a significant therapeutic effect (measured by greater than or equal to a 35% reduction from baseline in spleen volume at the end of cycle 6 (week 24) as measured by an MRI or CT scan with a follow-up scan four weeks later). As a result of treatment with Inrebic, 36 patients experienced greater than or equal to a 50% reduction in myelofibrosis-related symptoms, such as night sweats, itching, abdominal discomfort, feeling full sooner than normal, pain under ribs on left side, and bone or muscle pain.

The prescribing information for Inrebic includes a Boxed Warning to advise health care professionals and patients about the risk of serious and fatal encephalopathy (brain damage or malfunction), including Wernicke’s, which is a neurologic emergency related to a deficiency in thiamine. Health care professionals are advised to assess thiamine levels in all patients prior to starting Inrebic, during treatment and as clinically indicated. If encephalopathy is suspected, Inrebic should be immediately discontinued.

Common side effects for patients taking Inrebic are diarrhea, nausea, vomiting, fatigue and muscle spasms. Health care professionals are cautioned that patients may experience severe anemia (low iron levels) and thrombocytopenia (low level of platelets in the blood). Patients should be monitored for gastrointestinal toxicity and for hepatic toxicity (liver damage). The dose should be reduced or stopped if a patient develops severe diarrhea, nausea or vomiting. Treatment with anti-diarrhea medications may be recommended. Patients may develop high levels of amylase and lipase in their blood and should be managed by dose reduction or stopping the mediation. Inrebic must be dispensed with a patient Medication Guide that describes important information about the drug’s uses and risks.

The FDA granted this application Priority Review designation. Inrebic also received Orphan Drug designation, which provides incentives to assist and encourage the development of drugs for rare diseases. The FDA granted the approval of Inrebic to Impact Biomedicines, Inc., a wholly-owned subsidiary of Celgene Corporation.

LINK

http://s2027422842.t.en25.com/e/es?s=2027422842&e=245172&elqTrackId=376c7bc788024cd5a73d955f2e3dcbdc&elq=2a5deafa24e642ce8b78e60dd7bc7120&elqaid=9163&elqat=1

///////Inrebic , fedratinib, FDA 2019, Priority Review , Orphan Drug, Biomedicines, Celgene , bone marrow disorder

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Tanzisertib


Tanzisertib.png

ChemSpider 2D Image | Tanzisertib | C21H23F3N6O2

Tanzisertib

CAS 899805-25-5

trans-4-((9-((3S)-Tetrahydrofuran-3-yl)-8-((2,4,6-trifluorophenyl)amino)-9H-purin-2-yl)amino)cyclohexanol

4-[[9-[(3S)-oxolan-3-yl]-8-(2,4,6-trifluoroanilino)purin-2-yl]amino]cyclohexan-1-ol

C21-H23-F3-N6-O2, 448.4467

9557
Cyclohexanol, 4-[[9-[(3S)-tetrahydro-3-furanyl]-8-[(2,4,6-trifluorophenyl)amino]-9H-purin-2-yl]amino]-, trans-
  • CC 930
  • CC-930
  • Tanzisertib
  • UNII-M5O06306UO
  • A c-Jun amino-terminal kinase inhibitor.UNII, M5O06306UO

Treatment of Idiopathic Pulmonary Fibrosis (IPF)

  • Originator Celgene Corporation
  • Class Antifibrotics; Small molecules
  • Mechanism of ActionJ NK mitogen-activated protein kinase inhibitors
  • Orphan Drug Status Yes – Idiopathic pulmonary fibrosis
  • Discontinued Discoid lupus erythematosus; Idiopathic pulmonary fibrosis
  • 16 Jul 2012 Celgene Corporation terminates a phase II trial in Discoid lupus erythematosus in USA (NCT01466725)
  • 23 Feb 2012 Celgene initiates enrolment in a phase II trial for Discoid lupus erythematosus in the USA (NCT01466725)
  • 08 Nov 2011The Committee for Orphan Medicinal Products (COMP) recommends orphan drug designation for tanzisertib in European Union for Idiopathic pulmonary fibrosis

Tanzisertib has been granted orphan drug status by the FDA for the treatment of idiopathic pulmonary fibrosis. A positive opinion has been received from the EU Committee for Orphan Medicinal Products (COMP

Tanzisertib has been used in trials studying the treatment of Fibrosis, Discoid Lupus, Pulmonary Fibrosis, Interstitial Lung Disease, and Lung Diseases, Interstitial, among others.

PATENT

https://patents.google.com/patent/US20090048275A1/de

Image result for US 20090048275

Image result for US 20090048275

PATENT

WO 2006076595

US 20070060598

WO 2008057252

US 20080021048

US 20140094456

WO 2014055548

PATENT

WO 2015153683

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2015153683

/////////Tanzisertib, CC 930,  Idiopathic Pulmonary Fibrosis, Orphan Drug, phase II, CELGENE

c1c(c(c(cc1F)F)Nc2n(c3nc(ncc3n2)N[C@H]4CC[C@@H](CC4)O)[C@@H]5COCC5)F

FDA approves first treatment Soliris (eculizumab) for neuromyelitis optica spectrum disorder, a rare autoimmune disease of the central nervous system


The U.S. Food and Drug Administration today approved Soliris (eculizumab) injection for intravenous use for the treatment of neuromyelitis optica spectrum disorder (NMOSD) in adult patients who are anti-aquaporin-4 (AQP4) antibody positive. NMOSD is an autoimmune disease of the central nervous system that mainly affects the optic nerves and spinal cord.

“Soliris provides the first FDA-approved treatment for neuromyelitis optica spectrum disorder, a debilitating disease that profoundly impacts patients’ lives,” said Billy Dunn, M.D., director of the Division of Neurology Products in the FDA’s Center for Drug Evaluation and Research. “This approval changes the landscape of therapy for patients with NMOSD. Having an approved therapy for this condition is the culmination of extensive work we have engaged in with drug companies to …

June 27, 2019

The U.S. Food and Drug Administration today approved Soliris (eculizumab) injection for intravenous use for the treatment of neuromyelitis optica spectrum disorder (NMOSD) in adult patients who are anti-aquaporin-4 (AQP4) antibody positive. NMOSD is an autoimmune disease of the central nervous system that mainly affects the optic nerves and spinal cord.

“Soliris provides the first FDA-approved treatment for neuromyelitis optica spectrum disorder, a debilitating disease that profoundly impacts patients’ lives,” said Billy Dunn, M.D., director of the Division of Neurology Products in the FDA’s Center for Drug Evaluation and Research. “This approval changes the landscape of therapy for patients with NMOSD. Having an approved therapy for this condition is the culmination of extensive work we have engaged in with drug companies to expedite the development and approval of safe and effective treatments for patients with NMOSD, and we remain committed to these efforts for other rare diseases.”

In patients with NMOSD, the body’s immune system mistakenly attacks healthy cells and proteins in the body, most often in the optic nerves and spinal cord. Individuals with NMOSD typically have attacks of optic neuritis, which causes eye pain and vision loss. Individuals also can have attacks resulting in transverse myelitis, which often causes numbness, weakness, or paralysis of the arms and legs, along with loss of bladder and bowel control. Most attacks occur in clusters, days to months to years apart, followed by partial recovery during periods of remission. Approximately 50% of patients with NMOSD have permanent visual impairment and paralysis caused by NMOSD attacks. According to the National Institutes of Health, women are more often affected by NMOSD than men and African Americans are at greater risk of the disease than Caucasians. Estimates vary, but NMOSD is thought to impact approximately 4,000 to 8,000 patients in the United States.

NMOSD can be associated with antibodies that bind to a protein called aquaporin-4 (AQP4). Binding of the anti-AQP4 antibody appears to activate other components of the immune system, causing inflammation and damage to the central nervous system.

The effectiveness of Soliris for the treatment of NMOSD was demonstrated in a clinical study of 143 patients with NMOSD who had antibodies against AQP4 (anti-AQP4 positive) who were randomized to receive either Soliris treatment or placebo. Compared to treatment with placebo, the study showed that treatment with Soliris reduced the number of NMOSD relapses by 94 percent over the 48-week course of the trial. Soliris also reduced the need for hospitalizations and the need for treatment of acute attacks with corticosteroids and plasma exchange.

Soliris has a boxed warning to alert health care professionals and patients that life-threatening and fatal meningococcal infections have occurred in patients treated with Soliris, and that such infections may become rapidly life-threatening or fatal if not recognized and treated early. Patients should be monitored for early signs of meningococcal infections and evaluated immediately if infection is suspected. Use should be discontinued in patients who are being treated for serious meningococcal infections. Health care professionals should use caution when administering Soliris to patients with any other infection. In the NMOSD clinical trial, no cases of meningococcal infection were observed.

Soliris is available only through a restricted program under a Risk Evaluation and Mitigation Strategy (REMS). Prescribers must enroll in the REMS program. Prescribers must counsel patients about the risk of meningococcal infection, provide the patients with the REMS educational materials and ensure patients are vaccinated with meningococcal vaccine(s). The drug must be dispensed with the FDA-approved patient Medication Guide that provides important information about the drug’s uses and risks.

The most frequently reported adverse reactions reported by patients in the NMOSD clinical trial were: upper respiratory infection, common cold (nasopharyngitis), diarrhea, back pain, dizziness, influenza, joint pain (arthralgia), sore throat (pharyngitis) and contusion.

The FDA granted the approval of Soliris to Alexion Pharmaceuticals.

Soliris was first approved by the FDA in 2007. The drug is approved to reduce destruction of red blood cells in adults with a rare blood disease called paroxysmal nocturnal hemoglobinuria, for the treatment of adults and children with a rare disease that causes abnormal blood clots to form in small blood vessels in the kidneys (atypical hemolytic uremic syndrome to inhibit complement-mediated thrombotic microangiopathy), and for the treatment of adults with Myasthenia Gravis who are anti-acetylcholine receptor antibody positive.

The FDA granted this application Priority Review. The use for NMOSD received Orphan Drug designation, which provides incentives to assist and encourage the development of drugs for rare diseases.

https://www.fda.gov/news-events/press-announcements/fda-approves-first-treatment-neuromyelitis-optica-spectrum-disorder-rare-autoimmune-disease-central?utm_campaign=062719_PR_FDA%20approves%20first%20treatment%20for%20NMOSD&utm_medium=email&utm_source=Eloqua

///////////////fda 2019, Soliris, eculizumab, neuromyelitis optica spectrum disorder, Orphan DrugPriority Review

Solriamfetol hydrochloride, ソルリアムフェトル塩酸塩 , солриамфетол , سولريامفيتول , 索安非托 ,


2D chemical structure of 178429-65-7

Solriamfetol hydrochloride

FDA APPROVED 2019/3/20, Sunosi

ソルリアムフェトル塩酸塩; R228060, R 228060

Formula
C10H14N2O2. HCl
CAS
178429-65-7 HCL
Mol weight
230.6913
(2R)-2-Amino-3-phenylpropyl carbamate
(2R)-2-Amino-3-phenylpropylcarbamat
10117
178429-62-4 [RN] FREE FORM
Benzenepropanol, β-amino-, carbamate (ester), (βR)- [
солриамфетол [Russian] [INN]
سولريامفيتول [Arabic] [INN]
索安非托 [Chinese] [INN]
JZP-110
Originator SK Holdings
  • Developer Jazz Pharmaceuticals plc; SK biopharmaceuticals
  • Class Carbamates; Sleep disorder therapies; Small molecules
  • Mechanism of Action Adrenergic uptake inhibitors; Dopamine uptake inhibitors
  • Orphan Drug Status Yes – Narcolepsy
  • Registered Hypersomnia
  • Discontinued Depressive disorders
  • 26 Mar 2019 Discontinued – Phase-I for Depressive disorders (Adjunctive treatment) in USA (PO) (Jazz Pharmaceuticals pipeline, March 2019)
  • 20 Mar 2019 Registered for Hypersomnia (excessive daytime sleepiness) in patients with obstructive sleep apnoea and narcolepsy in USA (PO) – First global approval
  • 20 Mar 2019 US FDA approves solriamfetol to improve wakefulness in adult patients with excessive daytime sleepiness associated with narcolepsy or obstructive sleep apnoea(OSA)
  • New Drug Application (NDA): 211230
    Company: JAZZ PHARMA IRELAND LTD

Solriamfetol, sold under the brand name Sunosi, is a medication used for the treatment of excessive sleepiness associated with narcolepsy and sleep apnea.[1]

Common side effects include headache, nausea, anxiety, and trouble sleeping.[1] It is a norepinephrine–dopamine reuptake inhibitor(NDRI). It is derived from phenylalanine and its chemical name is (R)-2-amino-3-phenylpropylcarbamate hydrochloride.[2]

The drug was discovered by a subsidiary of SK Group, which licensed rights outside of 11 countries in Asia to Aerial Pharma in 2011.[3]

History

The drug was discovered by a subsidiary of SK Group, which licensed rights outside of 11 countries in Asia to Aerial Pharma in 2011.[3]Aerial ran two Phase II trials of the drug in narcolepsy[4] before selling the license to solriamfetol to Jazz in 2014; Jazz Pharmaceuticalspaid Aerial $125 million up front and will pay Aerial and SK up to $272 million in milestone payments, and will pay double digit royalties to SK.[3][5]

In March 2019 the FDA accepted SK’s and Jazz’ NDA for use of solriamfetol to treat excessive sleepiness in people with narcolepsy or obstructuve sleep apnea; the drug has an orphan designation for narcolepsy.[3][6]

Names

During development it has been called SKL-N05, ADX-N05, ARL-N05, and JZP-110.[6]

Research

Solriamfetol had also been tested in animal models of depression, but as of 2017 that work had not been advanced to clinical trials.[7]

PATENT

WO 9607637

https://patents.google.com/patent/WO1996007637A1/e

Organic alkyl carbamates have been effectively used for controlling various central nervous system (CNS) disorders. For example, U.S. Pat. Nos . 2,884,444, 2,937,119 and 3,313,697 disclose function of carbamate in CNS disorders, especially as antiepileptic and centrally acting muscle relaxant.
Phenylethylamine derivatives, one important class of therapeutical medicines useful for managing CNS diseases, have been used mainly to treat obesity, narcolepsy, minimal brain dysfunction and mild depression.
Recent design of pharmacologically useful compounds has been based on amino acids or the derivatives thereof, which is mainly attributable to the fact that many of the compounds found in biological systems come from amino acids or the derivatives thereof. In addition, in most cases, the function of a pharmaceutically useful compound is effected after it binds to an enzyme or receptor, which may trigger the regulatory mechanisms of the enzyme or receptor.

REACTION SCHEME I

REACTION SCHEME II

REACTION SCHEME III

EXAMPLE I
Preparation of N-Benzyloxycarbonyl-D-phenylalaninol

In a 500 mL RB flask equipped with a mechanical stirrer and a dropping funnel, D-phenylalaninol (45.4 g, 300 mmol) was dissolved in 220 mL of distilled water, and cooled in an ice-bath. The pH of the solution was adjusted with 50 % sodium hydroxide to 14. Benzyl chloroformate (49.3 mL, 345 mmol) was charged into the dropping funnel and added slowly to the well stirred solution over 0.5 hr. After the completion of the addition, the reaction mixture was stirred for 1 hr. at 0 *C. The product precipitated from the reaction mixture as a white solid. It was collected by filtration and washed completely with distilled water. After being dried in vacuo, the solid thus obtained weighed 104 grams without any further purification: 99.8% Yield.
Melting point = 90 – 92 *C
[α]D20 = + 43.4 (c = 1.0, EtOH)
Analysis calc: C, 71.56; H, 6.71; N,4.91
Found: C, 71.35; H, 6.71; N,4.91

EXAMPLE II
Preparation of N-Benzyloxycarbonyl-D-phenylalaninol
carbamate

In a 500 mL RB flask, N-benzyloxycarbonyl-D- phenylalaninol (13.56 g, 50 mmol) was charged with antipyrine (11.29 g, 60 mmol) in 250 mL of dry THF under a nitrogen atmosphere. The reaction mixture was cooled in an ice-bath and phosgene (30.3 mL of 1.93 M solution in toluene, 58.5 mmol) was added quickly while vigorously stirring. After stirring for 1 hr. , the formation of a corresponding chloroformate from the starting material was monitored by TLC. The chloroformate solution thus prepared, was slowly added to a well stirred and ice-chilled aqueous ammonium hydroxide solution (75 mL, 28-30 %, 1,190 mmol) via cannula over 0.5 hr. The resulting reaction mixture was stirred for an extra 0.5 hr. The organic phase separated was collected. The aqueous phase was extracted twice with methylene chloride (100 mL). The combined organic phase was washed with brine (50 mL), dried over sodium sulfate, and concentrated to yield 17.8 g (113%) of foamy solid. It was purified a flash column chromatography to give 14.8 g of the title compound, white solid: 94% Yield.
Melting point = 121 – 125 *C
[α]D20 = + 28.6 (c = 2.0, EtOH)
Analysis calc. : C, 65.84; H, 6.14; N, 8.53
Found: C, 66.68; H, 6.21; N, 7.80

EXAMPLE III
Preparation of D-Phenylalaninol carbamate hydrochloric
acid salt In a 160 mL Parr reactor, N-benzyloxycarbonyl-D-phenylalaninol carbamate (9.43 g) was added with 75 mL of anhydrous methanol and 10 % palladium on charcoal (0.32 g). Then, the reactor was closed and purged with hydrogen for 1 in. The reaction was completed in 2 hrs . under 40 psi pressure of hydrogen at 45 #C. The catalyst was filtered off. Thereafter, the organic layer was concentrated into 5.97 g (102 %) of pale yellow thick liquid. The liquid was poured in 50 mL of anhydrous THF and cooled to 0 “C. Anhydrous hydrogen chloride gas was then purged through the solution with slowly stirring for

0.5 hr. 50 mL of anhydrous ether was added, to give a precipitate. Filtration with THF-ether (1:1) mixture provided 6.1 g of the title compound as a white solid: 88 % Yield.
Melting point = 172 – 174 “C
[α]D20 = – 12.9 (c = 2.0, H20)
Analysis calc. : C, 52.60; H, 6.55; N, 12.14; Cl, 15.37
Found: C, 51.90; H, 6.60; N, 12.15; Cl ,

15.52

EXAMPLE IV
Preparation of N-benzyloxγcarbonyl-L-Phenγlalaninol

The title compound was prepared in the same manner as that of Example I, except that (L)-phenylalaninol was used as the starting material.
Melting point = 90 – 92 *C
[α]D20 = – 42.0 (c = 1.0, EtOH)
Analysis calc. : C, 71.56; H, 6.71; N,4.91
Found: C, 70.98; H, 6.67; N,4.95

EXAMPLE V
Preparation of -N-benzyloxycarbonyl-L-Phenylalaninol
carbamate

The title compound was prepared in the same manner as that of Example II, except that N-benzyloxycarbonyl-L-phenylalaninol was used as the starting material.
Melting point = 121 – 128 ‘C
[α]D20 = – 28.9 (c = 2.0, EtOH)
Analysis calc: C, 65.84; H, 6.14; N, 8.53
Found: C, 65.45; H, 6.15; N, 8.32

EXAMPLE VI
Preparation of L-Phenylalaninol carbamate hydrochloric
acid salt

The title compound was prepared in the same manner as that of Example III, except that N-benzyloxycarbonyl-L-phenylalaninol carbamate was used as the starting material.
Melting point = 175 – 177 *C [α]D20 = + 13.1 (c = 1.0, H20)
Analysis calc : C, 52.60; H, 6.55; N, 12.14; Cl, 15.37
Found: C, 51.95; H, 6.58; N, 12.09; Cl , 15.37

EXAMPLE VII
Preparation of N-benzyloxycarbonyl-D,L-Phenylalaninol

The title compound was prepared in the same manner as that of Example I, except that (D,L)-phenylalaninol was used as the starting material.
Melting point = 72 – 75 #C
Analysis calc: C, 71.56; H, 6.71; N,4.91
Found: C, 71.37; H, 6.74; N,4.84

EXAMPLE VIII
Preparation of N-benzyloxycarbonyl-D,L-Phenylalaninol
carbamate

The title compound was prepared in the same manner as that of Example II, except that N-benzyloxycarbonyl-D,L-phenylalaninol was used as the starting material.
Melting point = 130 – 133 *C
Analysis calc: C, 65.84; H, 6.14; N, 8.53
Found: C, 65.85; H, 6.14; N, 8.49 EXAMPLE IX
Preparation of D,L-Phenylalaninol carbamate hydrochloric
acid salt

The title compound was prepared in the same manner as that of Example III, except that N-benzyloxycarbonyl-D,L-phenylalaninol carbamate was used as the starting material.
Melting point = 163 – 165 *C
Analysis calc: C, 52.60; H, 6.55; N, 12.14; Cl, 15.37
Found: C, 51.92; H, 6.56; N, 11.95; Cl , 15.82

PATENT

US 20050080268

PATENT

WO 2018133703

https://patents.google.com/patent/WO2018133703A1/en

Excessive daytime sleepiness (Excessive Daytime Sleepiness, EDS) or pathological somnolence refers to excessive daytime sleep and wakefulness associated with various sleep disorders. These disorders can be the basis for a sleep disorder or sleep have side effects caused by some other medical conditions. Excessive daytime sleep, also known as narcolepsy, sleep clinics is seen mainly in patients with disease that affects 12% of the general population. EDS patients may be manifested as mental distress, poor work or school performance, increasing the risk of accidents, the impact of EDS can debilitating, even life-threatening.

R228060, also known JZP-110, is a selective dopamine and norepinephrine reuptake inhibitor, originally developed by R & D, SK biopharmaceutical, 2014 Sir ownership of the pharmaceutical compound. R228060 has the potential to treat narcolepsy and sleep apnea syndrome, in three multi-center study in two global reached the primary endpoint, and achieved positive results, significantly improved adult obstructive sleep apnea patients excessive sleepiness in patients with narcolepsy and excessive sleep problems.

R228060 chemical name is O- carbamoyl – (D) – phenylalaninol, as shown in the structural formula of formula (I):

Figure PCTCN2018071889-appb-000001

Solid Form different chemicals, can cause varying their solubility and stability, and thus affects the absorption and bioavailability of the drug, and can lead to differences in clinical efficacy. Improve the candidate compound has a solubility by salt way become an important means of drug development. Compared to the free form of the drug, suitable pharmaceutically acceptable salts can improve the solubility of the drug type, increased physical and chemical stability, and also to improve the drug-salt having a melting point, hygroscopicity, crystal type and other physical properties, further development of the pharmaceutical dosage form It plays an important role. Patent Document WO1996007637A1 discloses R228060 hydrochloride and its preparation method, and other characteristics of the obtained having a melting point of 172-174 deg.] C as a white solid, the solid was not given in the text data. Further, the present inventors found no other relevant R228060 hydrochloride polymorph or patent literature. Accordingly, the present need in the art to develop a comprehensive system R228060 hydrochloride polymorph, found to be suitable to the development of crystalline form. The present inventors after many experiments, found that polymorph CS1 R228060 hydrochloride CS2 and a melting point polymorph, Form CS1 and CS2 is Form 183 ℃, much higher than the melting point disclosed in prior art solid. It provides a better alternative preparation of pharmaceutical preparations containing R228060 is, has very important implications for drug development.

PATENT

WO 2019027941

https://patentscope2.wipo.int/search/en/detail.jsf;jsessionid=15B8F200BCC820C3761C600EA64A2018?docId=WO2019027941&recNum=4220&office=&queryString=&prevFilter=%26fq%3DOF%3AWO&sortOption=Pub+Date+Desc&maxRec=3471866

(i?)-2-amino-3-phenylpropyl carbamate (APC) is a phenylalanine analog that has been demonstrated to be useful in the treatment of a variety of disorders, including excessive daytime sleepiness, cataplexy, narcolepsy, fatigue, depression, bipolar disorder, fibromyalgia, and others. See, for example, US Patent Nos. 8,232,315; 8,440,715; 8,552,060; 8,623,913; 8,729,120; 8,741,950; 8,895,609; 8,927,602; 9,226,910; and 9,359,290; and U.S. Publication Nos. 2012/0004300 and 2015/0018414. Methods for producing APC (which also has other names) and related compounds can be found in US Patent Nos. 5,955,499; 5,705,640; 6,140,532 and 5,756,817. All of the above patents and applications are hereby incorporated by reference in their entireties for all purposes.

EXAMPLE 1

Synthesis of Compounds

Compound 8 (110CR002)

1 B 110CR002

[0083] tert- utyl (if)-(l-(Carbamothioyloxy)-3-phenylpropan-2-yl)carbamate (IB): A

60% dispersion of sodium hydride (0.36 g, 4.78 mmol, 1.2 equiv) in mineral oil was added in portions to compound 1A (1.0 g. 3.98 mmol, 1 equiv) in THF (20 mL) at 0 °C. After stirring for 1 hour, carbon disulfide (0.191 g, 4.78 mmol, 1.2 equiv) was added at 0 °C. After an additional hour of stirring, methyl iodide (0.3 mL, 4.78 mmol, 1.2 equiv) was added and the reaction was warmed to room temperature. After stirring two additional hours, concentrated ammonium hydroxide (1.6 mL, 7.98 mmol, 2 equiv) was added and the reaction was stirred overnight at room temperature. The reaction was diluted with water (50 mL) and extracted with dichloromethane (3 x 50 mL). The combined organic layers were dried over sodium sulfate and concentrated under reduced pressure to give crude compound IB. The solid was triturated in diethyl ether (20 mL) to give compound IB (0.17 g, 14% yield) as a light yellow solid.

[0084] (R)-0-(2-Amino-3-phenylpropyl) carbamothioate dihydrochloride (110CR002):

4M HCI in dioxane (0.68 mL, 2.74 mmol, 5 equiv) was added to neat compound IB (0.17 g, 0.548 mmol, 1 equiv) and the reaction was stirred overnight. The solution was diluted with diethyl ether (20 mL) and the resulting suspension was filtered. The solid was triturated in diethyl ether (20 mL) and the filtered solid was dried under vacuum at room temperature for two hours to give compound 110CR003 (140 mg, 93% yield, 96.9% purity) as a white solid.

Compound 9 (110CR003)

Scheme 2

2A 2B 110CR003

[0085] (R)-2-((ter^Butoxycarbonyl)amino)-3-phenylpropyl sulfamate (2B): A solution of sulfamoyl chloride (1.15 g, 9.95 mmol, 2.5 equiv) in acetonitrile (2 mL) was added dropwise to a solution of compound 2 A (1.0 g, 3.98 mmol, 1 equiv) and triethylamine (2.1 mL, 14.95 mmol, 3.75 equiv) in N,N-dimethylacetamide (20 mL) at 0 °C. After stirring at room temperature for 4 hours, additional triethylamine (2.1 mL, 14.95 mmol, 3.75 equiv) and sulfamoyl chloride (1.15 g, 9.95 mmol, 2.5 equiv) in acetonitrile (2 mL) was added at 0 °C. The reaction was stirred at room temperature overnight, at which point LCMS indicated a 3 :2 mixture of product to starting material. Additional triethylamine (2.1 mL, 14.95 mmol, 3.75 equiv) and sulfamoyl chloride (1.15 g, 9.95 mmol, 2.5 equiv) in acetonitrile (2 mL) was added at 0 °C and the reaction was stirred at room temperature for an additional 6 hours. LCMS indicated a 4: 1 mixture of product to starting material. The reaction was quenched with saturated sodium bicarbonate (5 mL) and stirred for an additional hour at room temperature. The reaction was diluted with saturated sodium bicarbonate (25 mL) and extracted with ethyl acetate (3 x 50 mL). The combined organic layers were dried over sodium sulfate and concentrated under reduced pressure. The product still contained unreacted starting material which could not be easily separated. Sulfamoyl chloride (1.15 g, 9.95 mmol, 2.5 equiv) in acetonitrile (2 mL) was added dropwise to a solution of crude compound 2B (0.9 g) and triethylamine (2.1 mL, 14.95 mmol, 3.75 equiv) in N,N-dimethylacetamide (20 mL) at 0 °C. After stirring at room temperature for two hours, the reaction was quenched with saturated sodium bicarbonate (5 mL) and the reaction was stirred for an additional hour at room temperature. The reaction was diluted with saturated sodium bicarbonate (25 mL) and extracted with ethyl acetate (3 x 50 mL). The combined organic layers were dried over sodium sulfate and concentrated under reduced pressure. The residue was purified on an AnaLogix automated system (Redisep 24 g silica gel column), eluting with a gradient of 25 to 50% ethyl acetate in heptanes, to give compound 2B (0.37 g, 28% yield) as a white solid.

[0086] (R)-2-Amino-3-phenylpropyl sulfamate hydrochloride (110CR003): 4M HC1 in dioxane (1.4 mL, 5.6 mmol, 5 equiv) was added to neat compound 2B (0.37 g, 1.12 mmol, 1 equiv) and the reaction was stirred overnight. The solution was diluted with diethyl ether (20 mL) and the resulting suspension was filtered. The solid was triturated in diethyl ether (20 mL) and the filtered solid was dried under a vacuum at room temperature for two hours to give compound 110CR003 (250 mg, 84% yield, 97.8% purity) as a white solid.

Com ound 3 (110CR007)

[0087] (Benzyl (R)-(l-phenyl-3-ureidopropan-2-yl)carbamate) (3B): Concentrated hydrochloric acid (0.06 mL, 0.68 mmol, 0.12 equiv) was added to a solution of benzyl (ft)-(l -amino-3-phenylpropan-2-yl)carbamate ( 1.5 g, 5.28 mmol, 1 equiv) and urea (1.26 g, 21.21 mmol, 4 equiv) in toluene (150 mL) under nitrogen. After refluxing overnight, LCMS indicated the reaction was complete. The reaction was concentrated under reduced pressure, diluted with water (150 mL) and stirred for 30 minutes. The resulting solid was filtered and washed with water (25 mL) to give crude compound 3B (1.4 g, 4.27 mmol, 80% yield) as a white solid, which was used sequentially.

[0088] ((R)-l-(2-mino-3-phenylpropyl)urea) (3C): Compound 3B (0.5 g, 1.5 mmol, 1 equiv) and 10% palladium on carbon (0.09 g) in methanol (60 mL) was hydrogenated at 30 psi for 1 hour at which time LC-MS determined that the reaction was incomplete. The solution was filtered and fresh catalyst (0.09 g) was added. The solution was hydrogenated at 30 psi for an additional 45 minutes resulting in complete conversion. Two identical scale reactions were run for 105 minutes each, both resulting in complete conversion. The three runs were combined and filtered through celite, which was washed with methanol (50 mL). The filtrate was concentrated under reduced pressure to give crude compound 3C (0.9 g), which was used sequentially.

[0089] (R)-l-(2-Amino-3-phenylpropyI)urea hydrochloride (110CR007): Compound 3C (0.88 g, 4.58 mmol, 1 equiv) was dissolved diethyl ether (10 mL) and 4 N HCl in dioxane (2.31 mL, 9.27 mmol, 2 equiv) was added. The reaction was stirred overnight and then concentrated under reduced pressure to give crude 110CR007 as a white solid. The material was twice recrystallized from 10% methanol in ethanol (30 mL) to give 110CR007 (0.163 g, 16 % yield, 93.7 % purity) as a white solid.

Compound 4 (110CR009)

Scheme 4

[0090] Ethyl (R^)-4-((tert-butoxycarbonyI)amino)-5-phenylpent-2-enoate (4B): A solution of compound 4A (4.0 g, 16.1 mmol, 1 equiv) and ethyl (triphenylphos-phoranylidene)acetate (5.6 g, 16.1 mmol, 1 equiv) in dichloromethane (40 mL) was stirred at room temperature overnight. The reaction was concentrated under reduce pressure to remove the organic solvent and the resulting residue was purified on an AnaLogix automated system (40 g Sorbtech silica gel column), eluting with gradient of 50 to 100% ethyl acetate in heptanes, to give compound 4B (4.8 g, 94% yield) as a white solid.

[0091] (R^E)-4-((te *i-ButoxycarbonyI)amino)-5-phenylpent-2-enoic acid (4C): Lithium hydroxide (1.4 g, 60 mmol, 4 equiv) in water (15 mL) was added to compound 4B (4.8 g, 15 mmol, 1 equiv) in THF (60 mL) at room temperature and the reaction was stirred overnight. After 16 hours, the reaction was adjusted to pH 4 with IN hydrochloric acid. The organic layer was removed and the aqueous layer was extracted with ethyl acetate (2 x 50 mL). The combined organic layers was washed with saturated brine (50 mL), dried over sodium sulfate and concentrated under reduced pressure to give compound 4C (4.2 g, 97% yield) as a light cream solid, which was used subsequently.

[0092] Methyl (R E)-4-((½ -i-butoxycarbonyl)amino)-5-phenylpent-2-enoate (4D1):

Isobutyl chloro formate (1.3 mL, 10 mmol, 1 equiv) in THF (4 mL) was added dropwise to a solution of compound 4C (3.0 g, 10 mmol, 1 equiv) and N-methyl-morpholine (1.1 mL, 10 mmol, 1 equiv) in THF (12 mL) at -15 °C. After 30 minutes of stirring, LCMS indicated complete conversion to the anhydride intermediate. 2M Ammonia in methanol (5 mL, 10 mmol, 1 equiv) was added dropwise over 20 minutes, keeping the internal temperature between -25 to -15 °C. After 30 minutes of stirring, the reaction was warmed to room

temperature and stirred overnight. The reaction mixture was concentrated at reduced pressure to remove the organic solvent. The resulting residue was dissolved in ethyl acetate (50 mL) and washed with water (100 mL). The aqueous layer was extracted with ethyl acetate (2 x 50 mL). The combined organic layers were washed with saturated brine (50 mL), dried over sodium sulfate and concentrated under reduced pressure. The residue was purified on an AnaLogix automated system (80 g Sorbtech silica gel column), eluting with a gradient of 25 to 50% ethyl acetate in heptanes, to give compound 4D1 (1.1 g, 35 % yield) as a white solid.

[0093] Methyl (S)-4-((te^-butoxycarbonyl)amino)-5-phenylpentanoate (4D2): A mixture of compound 4D1 (1.1 g, 3.6 mmol, 1 equiv) and 10% palladium on carbon (0.33 g, 50% wet) in methanol (40 mL) was hydrogenated at 40 psi at room temperature for 4 hours. The mixture was filtered through celite, which was washed with methanol (100 mL). The filtrate was concentrated under reduced pressure to give compound 4D2 (1.1 g, 99% yield) as a white solid.

[0094] (S)-4-((ii? i-Butoxycarbonyl)amino)-5-phenylpentanoic acid (4D3): Lithium hydroxide (73 mg, 3 mmol, 1.5 equiv) in water (1 mL) was added to compound 4B (0.6 g, 2 mmol, 1 equiv) in THF (9 mL) at room temperature. After stirring overnight, the reaction was adjusted to pH 4 with IN hydrochloric acid. The organic layer was removed and the aqueous layer was extracted with ethyl acetate (3 x 25 mL). The combined organic layers was washed with saturated brine (25 mL), dried over sodium sulfate and concentrated under reduced pressure to give compound 4D3 (0.56 g, 98% yield) as a white solid, which was used subsequently.

[0095] tert-Butyl (S)-(5-amino-5-oxo-l-phenylpentan-2-yl)carbamate (4E): Isobutyl chloroformate (0.23 mL, 1.8 mmol, 1 equiv) in THF (0.5 mL) was added drop-wise to a solution of compound 4C (0.54 g, 1.8 mmol, 1 equiv) and N-methylmorpholine (0.2 mL, 1.8 mmol, 1 equiv) in THF (1 mL) at -15 °C. After 20 minutes of stirring, LCMS indicated complete conversion to the anhydride intermediate. 0.4M Ammonia in THF (9 mL, 3.6 mmol, 2 equiv) was added drop-wise over 20 minutes, keeping the internal temperature between -25 to -15 °C. After 30 minutes of stirring the reaction was warmed to room temperature and stirred overnight. The reaction mixture was concentrated under reduced pressure to remove the organic solvent. The resulting residue was dissolved in ethyl acetate (25 mL) and washed with water (25 mL). The organic layer was separated and the aqueous layer was extracted with ethyl acetate (2 x 25 mL). The combined organic layers were washed with saturated brine (50 mL), dried over sodium sulfate and concentrated under

reduced pressure to give compound 4E (0.5 g, 93% yield) as a white solid, which was used subsequently.

[0096] (S)-4-Amino-5-phenylpentanamide hydrochloride (110CR009): 4M HC1 in dioxane (6 mL, 25 mmol, 10 equiv) was added to compound 4E (0.73 g, 1.12 mmol, 1 equiv) After stirring overnight at room temperature, the reaction was diluted with diethyl ether (20 mL) and stirred for 6 hours. The resulting suspension was filtered and the solid was washed with diethyl ether (20 mL). The filtered solid was dried under vacuum at room temperature for two hours to give compound 110CR009 (340 mg, 60% yield, 97.9 % purity) as a white solid.

Compound 10 (110CR012)

[0097] tert-Butyl (R)-(l-(carbamoylthio)-3-phenyIpropan-2-yI)carbamate (5B):

Compound 5 A (0.15 g, 0.56 mmol, 1 equiv) was dissolved in THF (8 mL) and sparged with nitrogen for 15 minutes. Trichloroacetyl isocyanate (0.1 mL, 0.84 mmol, 1.5 equiv) was added and the solution stirred for 3 hours, at which point TLC (30% ethyl acetate in heptane) indicated absence of starting material. The reaction was cooled to 0°C and concentrated ammonium hydroxide (0.15 mL) was added. After stirring overnight at room temperature, TLC indicated that the reaction was complete. The reaction was washed with a 10% ammonium hydroxide (10 mL). The organic layer was concentrated under reduced pressure. The residue was purified on an AnaLogix automated system (12 g silica gel column), eluting with a gradient of 0 to 30% ethyl acetate in heptane, to give compound 5B. This reaction was repeated an additional two times 0.15 g and 0.18 g). The products were to give compound 5B (0.35 g, 1.12 mmol, 62.2% yield) as a white solid.

[0098] (R)-S-(2-Amino-3-phenylpropyl) carbamothioate hydrochloride (110CR012):

Compound 5B (0.35 g, 1.12 mmol, 1 equiv) was dissolved in 4N HCI in dioxane (2 mL). The reaction was stirred for two hours and then concentrated under reduced pressure to give crude 110CR012 as a white solid. The material was triturated in diethyl ether (15 mL) to give 110CR012 (0.215 g, 78 % yield, 98.0 % purity) as a white solid.

References

  1. Jump up to:a b “SUNOSI™ (solriamfetol) Tablets, for Oral Use. Full Prescribing Information” (PDF). Jazz Pharmaceuticals. 2019. Retrieved 21 March2019.
  2. ^ Abad, VC; Guilleminault, C (2017). “New developments in the management of narcolepsy”Nature and Science of Sleep9: 39–57. doi:10.2147/NSS.S103467PMC 5344488PMID 28424564.
  3. Jump up to:a b c d Ji-young, Sohn (5 March 2018). “SK Biopharmaceuticals’ narcolepsy drug on track to hitting US market”The Korea Herald.
  4. ^ Sullivan, SS; Guilleminault, C (2015). “Emerging drugs for common conditions of sleepiness: obstructive sleep apnea and narcolepsy”. Expert Opinion on Emerging Drugs20 (4): 571–82. doi:10.1517/14728214.2015.1115480PMID 26558298.
  5. ^ Garde, Damian (January 14, 2014). “Jazz bets up to $397M on Aerial’s narcolepsy drug”FierceBiotech.
  6. Jump up to:a b “Solriamfetol – Jazz Pharmaceuticals/SK Biopharmaceuticals”. AdisInsight. Retrieved 15 April 2018.
  7. ^ de Biase, S; Nilo, A; Gigli, GL; Valente, M (August 2017). “Investigational therapies for the treatment of narcolepsy”. Expert Opinion on Investigational Drugs26 (8): 953–963. doi:10.1080/13543784.2017.1356819PMID 28726523.
Solriamfetol
Solriamfetol.svg
Clinical data
Trade names Sunosi
Synonyms SKL-N05, ADX-N05, ARL-N05, and JZP-110; (R)-2-amino-3-phenylpropylcarbamate hydrochloride
Routes of
administration
By mouth
ATC code
Pharmacokinetic data
Bioavailability ~95%
Protein binding 13.3–19.4%
Metabolism negligible
Elimination half-life ~7.1 h
Excretion urine (95% unchanged)
Identifiers
CAS Number
PubChem CID
ChemSpider
UNII
KEGG
Chemical and physical data
Formula C10H14N2O2
Molar mass 194.234 g/mol g·mol−1
3D model (JSmol)

///////////Solriamfetol hydrochloride, Solriamfetol, ソルリアムフェトル塩酸塩; солриамфетол , سولريامفيتول 索安非托 JZP-110, Orphan Drug, fda 2019, R228060, R 228060

RISDIPLAM , リスジプラム


Risdiplam.svg

Image result for RISDIPLAM

RISDIPLAM

RG-7916, RO-7034067, リスジプラム

Formula
C22H23N7O
Cas
1825352-65-5
Mol weight
401.4643
US9969754

7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[1,2-b]pyridazin-6-yl)pyrido[1,2-a]pyrimidin-4-one

WHO 10614

RG-7916

HY-109101

RO7034067

CS-0039501

EX-A2074

RG7916

The compound was originally claimed in WO2015173181 , for treating spinal muscular atrophy (SMA). Roche , under license from PTC Therapeutics , and Chugai , are developing risdiplam (RO-7034067; RG-7916), a small-molecule survival motor neuron (SMN)2 gene splicing modulator and a lead from an SMN2 gene modulator program initiated by PTC Therapeutics in collaboration with the SMA Foundation , for the oral treatment of spinal muscular atrophy

The product was granted orphan drug designation in the U.S., E.U. and in Japan for the treatment of spinal muscular atrophy. In 2018, it also received PRIME designation in the E.U. for the same indication.

Risdiplam (RG7916RO7034067) is a highly potent, selective and orally active small molecule experimental drug being developed by F. Hoffmann-La RochePTC Therapeutics and SMA Foundation to treat spinal muscular atrophy (SMA). It is a pyridazine derivative that works by increasing the amount of functional survival of motor neuron protein produced by the SMN2 gene through modifying its splicing pattern.[1][2]

As of September 2018, risdiplam is undergoing late-stage clinical trials across the spectrum of spinal muscular atrophy[3][4][5] where it has shown promising preliminary results.[6][7]

PATENT

WO2015173181

https://patentscope.wipo.int/search/en/detail.jsf;jsessionid=B8D897794EC02E2BBFD5D2280B3E1883.wapp1nC?docId=WO2015173181&recNum=9&office=&queryString=&prevFilter=%26fq%3DOF%3AKR%26fq%3DICF_M%3A%22C07D%22%26fq%3DPAF_M%3A%22F.+HOFFMANN-LA+ROCHE+AG%22&sortOption=Pub+Date+Desc&maxRec=912

Example 20

7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6- yl)pyrido[l,2-a]pyrimidin-4-one

In a sealed tube, 2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-7-fluoro-pyrido[l,2-a]pyrimidin-4-one (Intermediate 2; 50 mg, 0.162 mmol), DIPEA (0.22 mL, 1.29 mmol, 4 eq.) and 4,7-diazaspiro[2.5]octane dihydrochloride (32 mg, 0.320 mmol, 3.0 eq.) were stirred in

DMSO (2 mL) at 130°C for 48 hours. The solvent was removed under high vacuum. The residue was taken up in CH2CI2 and washed with an aqueous saturated solution of NaHC03. The organic layer was separated and dried over Na2S04 and concentrated in vacuo. The crude was purified by column chromatography (Si02, CH2Cl2/MeOH=98/2 to 95/5) to afford the title product (12 mg, 18%) as a light yellow solid. MS m/z 402.3 [M+H+].

PATENT

WO-2019057740

Process for the preparation of risdiplam and its derivatives.

Scheme 1:

Scheme 3:

Scheme 4:

xample 1: tert-Butyl 7-(6-chloro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate

5-Bromo-2-chloropyridine (85.0 g, 442 mmol), tert-butyl 4,7-diazaspiro[2.5]octane-4-carboxylate (102 g, 442 mmol) and Me-THF (722 g) were charged into a reaction vessel. After 10 minutes stirring, most of the solids were dissolved and [Pd(Xantphos)Cl2] (3.34 g) was added followed after 5 minutes by a solution of sodium tert-butanolate (56.3 g, 574 mmol) in Me-THF (173 g). The reaction mixture was stirred at 70 °C for 1.25 hours, cooled to room temperature and water (595 g) and 1-propylacetate (378 g) were added. After vigorous stirring, the phases were separated, the organic phase was washed with a second portion of water (425 g) and with a mixture of water (425 g) and brine (25 mL). The organic phase was treated with active charcoal (6.8 g), filtered and concentrated under reduced pressure to afford a brown oil, which was dissolved in tert-amyl-methyl-ether (347 g) at reflux. The solution was cooled slowly to room temperature. After stirring 18 hours at room temperature, n-heptane (205 g) was added and the suspension was further cooled to -10 °C. The precipitate was filtered off and dried under high vacuum to afford tert-butyl 7-(6-chloro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (110.9 g, 77.5%) as a beige solid.

Ή-ΝΜΡν (CDC13, 600 MHz): 7.95 (d, 1H); 7.18 – 7.14 (m, 1H); 7.13 – 7.09 (m, 1H); 3.79 – 3.63 (m, 2H); 3.24 – 3.12 (m, 2H); 2.96 (s, 2H); 1.47 (s, 9H); 1.11 – 1.04 (m, 2H); 0.90 -0.79 (m, 2H); LCMS: 324.15, 326.15 (M+H+)

Example 2: tert-butyl 7-(6-amino-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate

An autoclave equipped with an ascending pipe was filled with ammonia (78.7 g, 15 eq; 10 eq are sufficient) at -70 °C. Another autoclave was charged with tert-butyl 7-(6-chloro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (100 g, 309 mmol), sodium tert-butanolate (32.6 g, 340 mmol) and dioxane (800 mL). After 10 minutes stirring at room temperature under Ar, a solution of Pd2(dba)3 (1.41 g, 1.54 mmol) and tBuBrettPhos (1.50 g, 3.09 mmol) in dioxane (180 mL) was added. Thereafter, the connected ammonia vessel was warmed with a warm water bath and the connecting valve was opened. The autoclave was warmed to 30 °C and the reaction mixture stirred 5 hours at this temperature. The ammonia vessel was closed and disconnected. The excess ammonia was washed out of the autoclave with Argon. The reaction solution was poured into a separating funnel, the autoclave washed with ethyl acetate (300 mL) and water (100 mL) and these two solvent portions were added to the separating funnel. The biphasic mixture was further diluted with ethyl acetate (900 mL) and water (1000 mL). After vigorous stirring, the phases were separated. The organic phase was washed with a mixture of water (500 mL) and brine (10 mL). The combined aqueous phases were extracted twice with ethyl acetate (500 mL). The combined organic phases were treated with active charcoal (3.70 g, 309 mmol), filtered and the filtrate was concentrated under reduced pressure to afford a thick brown oil. This oil was dissolved in 1 -propyl acetate (160 mL) at 45-50°C and n-heptane (940 mL) was added drop wise within 1.5 hours. The suspension was cooled slowly to -5°C, stirred 4 hours at -5 °C and filtered. The precipitate was washed with cold n-heptane and dried under high vacuum at 50°C to afford tert-butyl 7-(6-amino-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (81.4 g, 86.5%) as a beige solid.

Ή-ΝΜΡν (CDCb, 600 MHz): 7.71 (d, 1H); 7.12 (dd, 1H); 6.47 (d, 1H); 4.18 (br s, 2H); 3.74 – 3.58 (m, 2H); 3.09 – 2.94 (m, 2H); 2.81 (s, 2H); 1.52 – 1.39 (m, 9H); 1.17 – 0.98 (m, 2H); 0.92 – 0.75 (m, 2H); LCMS: 305.20 (M+H+)

Example 3: tert-butyl 7-(6-amino-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate

An autoclave was charged with tert-butyl 7-(6-chloro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (339 mg, 1 mmol), sodium tert-butanolate (109 mg, 1.1 mmol) and dioxane (5 mL). After 5 minutes stirring at room temperature under Argon [Pd(allyl)(tBuBrettPhos)]OTf (4 mg, 5 μιηοΐ) was added. Thereafter, the autoclave was closed and connected to an ammonia tank, the valve was open and ammonia (230 mg, 13.5 mmol) was introduced into the autoclave. The valve was closed and the autoclave disconnected. The autoclave was warmed to 30 °C and the reaction mixture stirred 4 hours at this temperature. Then the autoclave was opened and the excess ammonia was washed out of the autoclave with Argon. The reaction solution was poured into a flask and taken to dryness under reduced pressure. The residue was purified by chromatography over silica gel (eluent: dichloromethane/ethyl acetate to dichloromethane/methanol). After evaporation of the solvents tert-butyl 7-(6-amino-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (283 mg, 93%) was isolated as a brown oil containing 4% dichloromethane and 3% ethyl acetate.

Example 4: tert-butyl 7-(6-nitro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate

tert-Butyl 4,7-diazaspiro[2.5]octane-4-carboxylate oxalate salt (2.46 kg, 8.13 mol), 5-bromo-2-nitro-pyridine (1.50 kg, 7.39 mol) and dimethyl sulfoxide (7.80 L) were char; into a reaction vessel pre-heated to 35 °C. With stirring, and keeping the temperature below 40°C, lithium chloride (1.25 kg, 25.6 mol) was added portion- wise followed by tetramethylguanidine (2.98 kg, 25.9 mol). Dimethyl sulfoxide (450 mL) was used to rinse the feed line. The reaction mixture was stirred at 79 °C for 8 hours, cooled to 70°C and water (2.48 L) was added within 2 hours. After stirring at 70 °C for an additional 1 hour, the precipitate was filtered off and washed with water (4.5 L) three times. The precipitate was dissolved in ethyl acetate (15 L) and water (7.5 L) at reflux temperature. The phases were separated at 60°C and n-heptane (7.5 L) was added to the organic layer at 60°C within 30 minutes. The solution was cooled to 0°C in 2 hours and further stirred at 0°C for 1 hour. The precipitate was filtered off, washed with a mixture of ethyl acetate (750 mL)/n-heptane (375 mL) twice and dried under reduced pressure to afford 1.89 kg (76.4%) of tert-butyl 7-(6-nitro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate as a yellow to light brown solid.

!H-NMR (CDCls, 600 MHz): 8.16 (d, 1H); 8.07 (d, 1H); 7.15 (dd, 1H); 3.80 – 3.72 (m, 2H); 3.49 – 3.41 (m, 2H); 3.23 (s, 2H); 1.48 (s, 9H); 1.16 – 1.08 (m, 2H); 0.92 – 0.85 (m, 2H); LCMS: 335.17 (M+H+)

Example 5: tert-butyl 7-(2-hydroxy-4-oxo-pyrido[l,2-a]pyrimidin-7-yl)-4,7-diazaspiro[2.5]octane-4-carboxylate

tert-Butyl 7-(6-amino-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (80.0 g, 263 mmol) was dissolved in anisole (800 mL) and di-tert-butyl malonate (71.1 g, 315 mmol) was added. The solution was stirred 3.5 hours at 145 °C then cooled to room temperature. The precipitate was filtered off, washed with toluene (in portions, 320 mL in total) and dried under high vacuum at 50°C to afford tert-butyl 7-(2-hydroxy-4-oxo-pyrido[l,2-a]pyrimidin-7-yl)-4,7-diazaspiro[2.5]octane-4-carboxylate (65.6 g, 67%) as a light pink powder.

Ή-ΝΜΡν (CDCI3, 600 MHz): 8.46 (d, 1H); 7.74 (dd, 1H); 7.52 (d, 1H); 5.37 (s, 2H); 3.83 – 3.69 (m, 2H); 3.23 (t, 2H); 3.01 (s, 2H); 1.48 (s, 9H); 1.17 – 1.03 (m, 2H); 0.95 – 0.75 (m, 2H); LCMS: 373.19 (M+H+)

Example 6: tert-butyl 7-(2-hydroxy-4-oxo-pyrido[l,2-a]pyrimidin-7-yl)-4,7-diazaspiro[2.5]octane-4-carboxylate

tert-Butyl 7-(6-nitro-3-pyridyl)-4,7-diazaspiro[2.5]octane-4-carboxylate (950 g, 2.84 mol), Pt 1%, V 2% on active charcoal (95.1 g, 2 mmol) and ethyl acetate (9.5 L) were charged into an autoclave that was pressurized with hydrogen gas to 3 bar. The reaction mixture was stirred at room temperature for 6 hours. The excess hydrogen was vented. The reaction mixture was filtered, the catalyst was washed with ethyl acetate (0.95 L) three times. The filtrate was concentrated under reduced pressure and the solvent exchanged to anisole (add two portions of 2.85 L and 5.18 L) by distillation. Di tert-butyl malonate (921.7 g, 4.26 mol) was added and the charging line was rinsed with anisole (618 mL) and the reaction mixture was stirred at 125-135 °C for 8 hours. It may be necessary to distill off the by-product tert-butanol to reach this temperature. The progress of the reaction was followed eg.by HPLC. If the reaction stalls, the temperature is increased to 135-145°C and checked for progress after 1 hour. When the reaction was complete, the batch was cooled to room temperature and stirred at room temperature for 4 hours. The precipitate was filtered off, washed with toluene (3.55 L) and dried under vacuum at 60°C to afford tert-butyl 7-(2-hydroxy-4-oxo-pyrido[l,2-a]pyrimidin-7-yl)-4,7-diazaspiro[2.5]octane-4-carboxylate (861.0 g, 81.4%) as a yellow to light brown solid.

Example 7: tert-butyl 7-[4-oxo-2-(p-tolylsulfonyloxy)pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate

A reactor was charged with tert-butyl 7-(2-hydroxy-4-oxo-pyrido[l,2-a]pyrimidin-7-yl)-4,7-diazaspiro[2.5]octane-4-carboxylate (920 g, 2.47 mol) and then triethylamine (325 g, 3.21 mol), followed by tosyl chloride (527.1 g, 2.77 mol) and dichloromethane (4.6 L). The reaction mixture was stirred at 20-25 °C for at least three hours. Upon complete reaction, the organic solution was washed with a prepared solution of HC1 (32%, 247.8 mL) and water (4.6 L), followed by a prepared solution of sodium hydroxide (432.3 mL of a 30% stock solution) and water (3.9 L) in that order. The organic phase was finally washed with water (4.8 L) and then dichloromethane was nearly completely distilled off under reduced pressure at 50-55°C. Ethyl acetate (920 mL) was added and distilled twice at this temperature under reduced pressure, and then ethyl acetate (4.8 L) was added and the suspension cooled to 20-25 °C over two hours. n-Heptane (944.4 mL) was added and the mixture was cooled to 0-5 °C and then stirred for an additional 3 hours. The precipitate was filtered off, washed with a prepared solution of ethyl acetate (772.8 mL) and n-heptane (147.2 mL), and then twice with n-heptane (2.6 L). The solid was dried under vacuum at 45-50°C to afford 1122.6 g (86.3%) tert-butyl 7-[4-oxo-2-(p-tolylsulfonyloxy)pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate as yellow crystals.

!H-NMR (CDCls, 600 MHz): 8.32 (d, 1H); 8.00 – 7.89 (m, 2H); 7.66 (dd, 1H); 7.50 (d, 1H); 7.36 (d, 2H); 6.04 (s, 1H); 3.80 – 3.68 (m, 2H); 3.23 (t, 2H); 3.01 (s, 2H); 1.48 (s, 9H); 1.15 – 1.04 (m, 2H); 0.92 – 0.82 (m, 2H); LCMS: 527.20 (M+H+)

Example 8: 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine

6-Chloro-2,8-dimethylimidazo[l,2-b]pyridazine (40.0 g, 220 mmol), bis pinacol diborane (69.9 g, 275 mmol) and potassium acetate (43.2 g, 440 mmol) were suspended in acetonitrile (440 mL). The suspension was heated to reflux and stirred 30 minutes at reflux, then a suspension of PdCl2(dppf) (4.03 g, 5.51 mmol) and dppf (610 mg, 1.1 mmol) in acetonitrile (40 mL) was added. The vessel was rinsed with acetonitrile (20 mL), which were also poured into the reaction mixture. The orange suspension was further stirred at reflux, whereby acetonitrile (50 mL) were distilled off. After 4 hours, the reaction mixture was filtered off, the filter was washed with several portions of acetonitrile (in total 150 mL). The filtrate was diluted to obtain a volume of 700 mL. The 314 mmolar solution of 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine in acetonitrile was used as such in the next step.

Example 9: 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine

6-chloro-2,8-dimethylimidazo[l,2-b]pyridazine (29.0 g, 22.8 mmol), bis pinacol diborane (44.6, 25.1 mmol) and potassium acetate (31.3 g, 45.6 mmol) were suspended in 1-propyl acetate (365 mL). The suspension was heated to 80°C and a solution of

tricyclohexylphosphine (448 mg, 0.23 mmol) and Pd(OAc)2 (179 mg, 0.11 mmol) in 1-propyl acetate (37 mL) was added within 20 minutes. After 2.5 hours further stirring at 80°C, the suspension was cooled to 40°C and filtered at this temperature. The precipitate was washed with 1-propyl acetate (200 mL). The filtrate corresponds to 516.4 g of a 8.5% solution of 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine in 1 -propyl acetate.

Example 10: Isolation of 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[ 1 ,2-b]pyridazine

In another experiment, the above solution obtained was cooled to 0-5 °C within 3 hours. The precipitate was filtered off, washed with cold 1 -propyl acetate and dried under high vacuum at 60°C to afford 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine (24. Og, 55%) as a colourless solid.

lH NMR (CDCls, 600 MHz, ) δ ppm 7.86 (d, J=0.7 Hz, 1 H), 7.20 (d, J=1.0 Hz, 1 H), 2.63 (d, J=1.0 Hz, 3 H), 2.51 (d, J=0.7 Hz, 3 H), 1.33 – 1.49 (m, 12 H)

Example 11: (step 6) tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate

tert-Butyl 7-[4-oxo-2-(p-tolylsulfonyloxy)pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5] octane-4-carboxylate (25 g, 47.5 mmol), 2,8-dimethyl-6-(4,4,5,5-tetramethyl- 1,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine (314 mM in acetonitrile, 191 mL, 59.8 mmol), PdCi2(dppf) (868 mg, 1.19 mmol) and aqueous potassium carbonate 4.07 M (17.1 mL, 69.8 mmol) were charged into a reaction vessel. The reaction mixture was stirred at reflux for 3 hours, cooled overnight to room temperature and filtered. The precipitate was washed with several portions of acetonitrile (146 mL in total), then suspended in methyl-THF (750 mL) and methanol (75 mL). Aqueous sodium hydrogen carbonate 5% (250 mL) was added, the mixture was vigorously stirred at 35°C. The phases were separated, the organic phase was washed again with aqueous sodium hydrogen carbonate 5% (250 mL). The organic phase was treated with active charcoal for 1 hour at room temperature, filtered and the filtrate was concentrated under reduced pressure at 60 °C to a volume of 225 mL, heated to reflux then cooled to room temperature, stirred at room temperature for 16 hours, then cooled to 0°C and stirred at 0°C for 3 hours. The precipitate was filtered off, washed with n-heptane (60 mL) and dried under high vacuum at 55°C to afford tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (20.13 g, 84.5%) as a yellow solid.

This solid could be recrystallized in the following manner: 15 g of the above solid was dissolved at reflux in toluene (135 mL) and ethanol (15 mL). The solution was slowly cooled to room temperature, stirred 16 hours at room temperature, then cooled to 0°C and stirred at 0°C for 4 hours. The precipitate was filtered off, washed with cold toluene and dried under high vacuum at 55°C to afford tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (11.92 g, 79.5%) as a yellow-green solid.

!H-NMR (CDCls, 600 MHz): 8.44 (d, 1H); 7.93 (d, 1H); 7.96 – 7.89 (m, 1H); 7.80 (d, 1H); 7.76 – 7.72 (m, 1H); 7.70 – 7.63 (m, 1H); 7.38 (s, 1H); 3.85 – 3.69 (m, 2H); 3.28 (t, 2H); 3.07 (s, 2H); 2.74 (d, 3H); 2.55 (s, 3H); 1.49 (s, 9H); 1.16 – 1.09 (m, 2H); 0.93 – 0.86 (m, 2H); LCMS: 502.26 (M+H+)

Example 12: tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate

6-chloro-2,8-dimethylimidazo[l,2-b]pyridazine (4.14 g, 22.8 mmol), bis pinacol diborane (6.37g, 25.1 mmol) and potassium acetate (4.47 g, 45.6 mmol) were suspended in 1-propyl acetate (59 mL). The suspension was heated to 80°C and a solution of

tricyclohexylphosphine (63.9 mg, 0.23 mmol) and Pd(OAc)2 (25.6 mg, 0.11 mmol) in 1-propyl acetate (6 mL) was added within 20 minutes. After 2.5 hours further stirring at 80°C, the suspension was cooled to 40°C and filtered at this temperature. The precipitate was washed with 1-propyl acetate (32 mL). The filtrate corresponds to 74.6 g of a 8.5% solution of 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine in 1-propyl acetate.

A reaction vessel was charged with tert-butyl 7-[4-oxo-2-(p-tolylsulfonyloxy)pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (10.0 g, 19.0 mmol), tricyclohexylphosphine (58.6 mg, 0.21 mmol) and Pd(OAc)2 (21.3 mg, 0.10 mmol) and 1-propyl acetate (42 mL) and a solution of potassium carbonate (5.25 g, 38.0 mmol) in water (19.0 mL) was added. The suspension was heated to 70°C and the solution of 2,8-dimethyl-6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)imidazo[l,2-b]pyridazine in 1-propyl acetate was added within 30 minutes. The mixture was stirred for 2 hours at 70-75°C. The suspension was cooled to 40°C, water (10 mL) was added. The suspension was aged for 30 minutes. The crude product was filtered off and rinsed with 1-propyl acetate (41 mL). The crude product was taken up in toluene (100 mL), 5% aqueous NaHC03-solution (30 mL) and 1-propanol (20.0 mL). The mixture was heated to 60-65 °C, the phases were separated and the organic phase was washed with 2 more portions of water (30.0 mL). The organic phase was filtered on active charcoal, the filter washed with toluene (60.0 mL). The filtrate was concentrated under reduced pressure to a volume of ca. 120 mL, heated to reflux and 1-propanol (0.8 mL) was added to obtain a solution. The solution was cooled to 0-5°C within 4-6 hours, stirred at 0-5°C for 1 hour. The precipitate was filtered off, washed with toluene (30 mL) and dried under reduced pressure at 70-80°C to afford tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (7.7 g, 80.8%) as a yellowish solid.

Example 13: 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)pyrido[l,2-a]pyrimidin-4-one di-hydrochloride salt

To prepare a solution of HC1 in in 1-propyl acetate/ 1-propanol, acetyl chloride (15.8 g, 199 mmol) was slowly added to a mixture of 1-propyl acetate (60 mL) and 1-propanol (30 mL) at 0°C, and stirring was pursued for an additional 2 hours at room temperature.

tert-Butyl 7-[2-(2,8-dimethylimidazo[ 1 ,2-b]pyridazin-6-yl)-4-oxo-pyrido[ 1 ,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (20 g, 39.9 mmol) was suspended in 1-propyl acetate (60 mL) and 1-propanol (30 mL) at room temperature and the HC1 solution in 1-propyl acetate and 1-propanol was added. The reaction mixture was heated within 3 hours to 70°C and stirred 16 hours at this temperature, then cooled to 20°C. The precipitate was filtered off, washed with 1-propyl acetate (50 mL) in several portions and dried under vacuum at 55 °C to afford 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)pyrido[l,2-a]pyrimidin-4-one hydrochloride salt (18.8 g, 99%) as yellow crystals.

^-NMR (CDCls, 600 MHz): 8.34 (s, 1H); 8.22(s, 1H); 8.05 (s, 1H); 8.01 (dd, 1H); 7.80 (d, 1H); 7.16 (s, 1H); 3.71 – 3.67 (m, 2H); 3.64 – 3.59 (m, 2H); 3.52 (s, 2H); 2.69 (s, 3H); 2.54 (s, 3H); 1.23- 1.20 (m, 2H); 1.14 – 1.08 (m, 2H); LCMS: 402.20 (M+H+)

Example 14: 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)pyrido[ 1 ,2-a]pyrimidin-4-one

To a suspension of tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (25 g, 50 mmol) in 1-propyl acetate (375 mL) was added a solution of HC1 in 1-propanol (prepared by adding slowly at 5°C acetyl chloride (18.0 mL) to 1-propanol (37.6 mL) and stirring 1 hour at room temperature). The stirred suspension was heated to 75°C within 10 hours and stirred a further 5 hours at 75 °C. Water (160.0 mL) was added and the phases were separated at 75°C. Aqueous sodium hydroxide 32% (27.8 mL) was added to the aqueous phase. The suspension obtained was cooled to room temperature within 5 hours and stirred one hour at room temperature. The precipitate was filtered off, washed with water (100.0 mL) and dried under reduced pressure at 50°C for 18 hours to afford 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)pyrido[l,2-a]pyrimidin-4-one (19.7 g, 98.3%) as yellow crystals.

!H-NMR (CDCb, 600 MHz): 8. 45 (d, 1H); 7.92 (d, 1H); 7.80 (s, 1H); 7.75 – 7.71 (m, 1H); 7.71 – 7.67 (m, 1H); 7.37 (s, 1H); 3.31 – 3.24 (m, 2H); 3.22 – 3.16 (m, 2H); 3.09 (s, 2H); 2.73 (s, 3H); 2.55 (s, 3H); 0.82- 0.76 (m, 2H); 0.71 – 0.63 (m, 2H); LCMS: 402.20

(M+H+)

Example 15: 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)pyrido[ 1 ,2-a]pyrimidin-4-one

A suspension of tert-butyl 7-[2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)-4-oxo-pyrido[l,2-a]pyrimidin-7-yl]-4,7-diazaspiro[2.5]octane-4-carboxylate (13.5 g, 26.9

in toluene (237.0 g) was stirred at 75°C and a 21.9% solution of HCl in 1-propanol (21.4 g, 134.5 mmol) was added within 2.5 hours. The reaction mixture was stirred further at 75 °C until complete conversion. The reaction mixture was cooled to 20-25°C. Water (70 g) was added. The biphasic mixture was stirred another 10 minutes at 20-25 °C and the phases were separated. The organic phase was extracted with water (17 g) twice and the combined aqueous phases were added into mixture of aqueous sodium hydroxide 28% (15.0 g) and water (45.0 g). The suspension obtained was cooled to 20°C. The precipitate was filtered off , washed with water (25 g) three times and dried under reduced pressure at 60°C to afford 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[l,2-b]pyridazin-6-yl)pyrido[l,2-a]pyrimidin-4-one (9.5 g, 95.1%) as yellow crystals.

Example 16: 4-bromo-6-chloro-pyridazin-3-amine

3-amino-6-chloropyridazine (20 g, 154 mmol), sodium bicarbonate (25.9 g, 309 mmol) and methanol (158 g) were charged in a reaction vessel and cooled to 0-10°C. Bromine (34.5 g, 216 mmol) was added drop wise and the reaction mixture was stirred 3 days at room temperature. 10% Aqueous sodium sulfate was added. The suspension was filtered off. The filtrate was washed with ethyl acetate (300 mL) twice. The combined organic layers were dried and evaporated. A suspension of the residue in methanol (50 mL) was heated to reflux, water (120 mL) was added and the suspension was stirred 16 hours at room temperature. The precipitate was filtered off and dried. The residue was suspended in n-heptane (50 mL), stirred 2 hours at room temperature, filtered off and dried to afford 4-bromo-6-chloro-pyridazin-3-amine (14.5 g, 46.2%) as a light brown solid.

!H-NMR (CDCls, 600 MHz): 7.55 (s, 1H); 5.83-4.89 (m, 2H); LCMS: 209.93 (M+H+)

Example 17: 4-bromo-6-chloro-pyridazin-3-amine

3-amino-6-chloropyridazine (50 g, 360 mmol), acetic acid (5.8 g, 96.5 mmol), sodium acetate (28.7 g, 289.5 mmol) and methanol (395 g) were charged in a reaction vessel and heated to 25-35°C. Dibromodimethylhydatoin (66.0 g, 231.6 mmol) was added in several portions and the reaction mixture was stirred 3 hours at 30°C. Completion is checked by IPC and if the conversion is incomplete, dibromodimethylhydantoin is added (5.5g). At reaction completion, 38% aqueous sodium sulfate (77.2 mmol NaHS03) was added slowly. The suspension was concentrated under reduced pressure and water (500 g) was added slowly at 45°C, then 30% aqueous sodium hydroxide (31.5 g, 231.6 mmol NaOH) was added at 20°C to adjust pH to 7-8. The precipitate was filtered off, washed with water and dried under reduced pressure to afford 4-bromo-6-chloro-pyridazin-3-amine (50.2 g, 62.5%) as a grey solid.

Example 18: 6-chloro-4-methyl-pyridazin-3-amine

4-bromo-6-chloro-pyridazin-3-amine (3.0 g, 14.4 mmol) and

tetrakis(triphenylphosphine)palladium (1666 mg, 144 μιηοΐ) were suspended in THF (13.2 g) and a solution of zinc chloride in Me-THF (2.0 M, 9 mL, 18 mmol) was added. The reaction mixture was cooled to -5°C and methyllithium in diethoxymethane (3.1 M, 11.6 mL, 36 mmol) was added. The reaction mixture was stirred at 45°C for 4 hours. Sodium sulfate decahydrate (11.7 g, 36 mmol) was added at room temperature, the mixture was stirred 1.5 hours at 60°C, diluted with water (100 mL) and after 30 minutes the precipitate was filtered off. The precipitate was dissolved in aqueous HC1 2M (100 mL) and ethyl acetate (140 mL). The biphasic system was filtered, the phases were separated and the pH of the water layer adjusted to 7 with aqueous NaOH 32% (18 mL). The precipitate was filtered and dried. The solid obtained was digested twice in methanol (20 mL) at room temperature. The two filtrates were combined, evaporated and dried under high vacuum to afford 6-chloro-4-methyl-pyridazin-3-amine (1.2 g, 58.1%) as a red solid.

Ή-ΝΜΡν (CDCb, 600 MHz): 7.09 (d, 1H); 4.90 (br s, 2H), 2.17 (d, 3H)

Example 19: 6-chloro-4-methyl-pyridazin-3-amine

4-bromo-6-chloro-pyridazin-3-amine (30.02 g, 143 mmol) and THF (180 mL) were charged into a reaction vessel. Methylmagnesium chloride (22% in THF, 50.0 mL, 1.03 eq.) was added at 20°C over 60 minutes, followed by zinc chloride in Me-THF (25%, 37 mL, 0.50 eq.) and palladium tetrakis(triphenyphosphine) (1.66 g, lmol%). The reaction mixture was heated to 50°C and methylmagnesium chloride (22% in THF, 81 mL, 1.7 eq.) was added slowly. The reaction mixture was stirred at 50°C until complete conversion, then at 10°C for 14.5 hours and poured into a mixture of water (90 g), aqueous HCl 33% (52.5 g) and toluene (150 mL) maintained at 20-30°C. The aqueous phase was separated and the organic phase was extracted with a solution of aqueous HCl 33% (2.0 g) and water (45 g). The aqueous layers were combined and washed with toluene (30 mL) twice and the pH was adjusted by addition of 25% aqueous ammonia solution. When a pH of 2.4 was reached, seeding crystals were added, the mixture was stirred further for 15 minutes and thereafter the pH was brought to 4.0. The suspension was stirred at 20°C for 2 hours, the precipitate was filtered off, washed with water (20 mL) three times to afford crude 6-chloro-4-methyl-pyridazin-3-amine (29 g) as a brown solid.

29 g crude product was transferred to a reaction vessel and methanol (20 mL) was added. The mixture was refluxed for 30 minutes and 12 g water was added. The solution was cooled to 0°C and stirred for 2 hours at this temperature. The precipitate was filtered off, washed with water three times and dried under reduced pressure at 40°C to afford purified 6-chloro-4-methyl-pyridazin-3-amine (13.8 g, 66%) as a light brown solid.

Alternative purification:

50 g crude 6-chloro-4-methyl-pyridazin-3-amine were dissolved in methanol (250 mL) and active charcoal (4.0 g) and diatomaceous earth (2.5 g) were added. The suspension was stirred at 45°C for 1 hour, cooled to 30°C and potassium hydrogenophosphate (2.1 g) was added. The suspension was stirred at 30°C for another 90 minutes, filtered and the precipitate washed with methanol (100 mL). The filtrate was concentrated to a residual volume of 175 mL and water (120 mL) was added. The resulting suspension was heated

to reflux affording a solution which was cooled to 20°C resulting in a suspension. The precipitate was filtered off, washed with water (90 mL) and dried under reduced pressure to afford pure 6-chloro-4-methyl-pyridazin-3-amine (38 g, 76%) as a light yellow solid.

Example 20: 6-chloro-2,8-dimethyl-imidazo[l,2-b]pyridazine

6-chloro-4-methyl-pyridazin-3-amine (70.95 kg, 494.2 mol), sodium bromide (35 kg, 345.9 mol), isopropyl acetate (611 kg), isopropanol (28 kg and water (35 kg) were charged into a reaction vessel. The reaction mixture was stirred at 80-85 °C for 8 hours. Isopropyl acetate (310 kg) and water (420 kg) were added. 30% Aqueous NaOH was added at 45-55 °C and the system was stirred for 2 hours. The phases were separated at 25-35 °C. The organic layer was washed with water (370 kg), filtered on diatomite (7 kg) and the filter washed with isopropyl acetate (35 kg). The organic phase was extracted with two portions of 5.4% aqueous sulfuric acid (910 kg followed by 579 kg). The combined aqueous phases were basified with 30% aqueous NaOH (158 kg). The suspension was stirred 2 hours at 15-25 °C. The precipitate was isolated by centrifugation in three portions, each washed with water (31 kg). The wet solid was dissolved in isopropyl acetate (980 kg) at 25-35 °C, the solution washed with water (210 kg), three times. The organic phase was treated with active charcoal for 12 hours at 45-50 °C, concentrated to ca. 300 kg and heated to 70-80 °C to obtain a clear solution. This solution was cooled to 50-60 °C, stirred at this temperature for 1 hour, n-heptane (378 kg) was added and stirring was pursued for 1 hour. The mixture was cooled to -10- -5°C and stirred for another 3 hours. The precipitate was isolated by centrifuging, washed with n-heptane (33 kg) and dried under reduced pressure at 30-50 °C for 15 hours to afford 67.4 kg (76%) 6-chloro-2,8-dimethyl-imidazo[l,2-b]pyridazine as an off-white solid.

XH-NMR (CDCls, 600 MHz): 7.67 (s, 1H); 6.86 (s, 1H); 2.65 (s, 3H), 2.50 (s, 3H)

Paper

https://pubs.acs.org/doi/pdf/10.1021/acs.jmedchem.8b00741

Abstract Image

SMA is an inherited disease that leads to loss of motor function and ambulation and a reduced life expectancy. We have been working to develop orally administrated, systemically distributed small molecules to increase levels of functional SMN protein. Compound 2 was the first SMN2 splicing modifier tested in clinical trials in healthy volunteers and SMA patients. It was safe and well tolerated and increased SMN protein levels up to 2-fold in patients. Nevertheless, its development was stopped as a precautionary measure because retinal toxicity was observed in cynomolgus monkeys after chronic daily oral dosing (39 weeks) at exposures in excess of those investigated in patients. Herein, we describe the discovery of 1 (risdiplam, RG7916, RO7034067) that focused on thorough pharmacology, DMPK and safety characterization and optimization. This compound is undergoing pivotal clinical trials and is a promising medicine for the treatment of patients in all ages and stages with SMA.

 7-(4,7-diazaspiro[2.5]octan-7-yl)-2-(2,8-dimethylimidazo[1,2-b]pyridazin-6-yl)pyrido[1,2-a]pyrimidin-4-one 1 (12 mg, 18%) as a pale yellow solid. 1H NMR (600 MHz,CDCl3) δ ppm 8.45 (d, J = 2.4 Hz, 1H), 7.92 (d, J = 1.0 Hz, 1H), 7.73 (d, J = 9.6 Hz, 1H) 7.80 (s, 1H), 7.70 (dd, J = 9.7, 2.5 Hz, 1H), 7.38 (s, 1H), 3.31–3.22 (m, 2H), 3.20–3.16 (m, 2H), 3.08 (s, 2H), 2.74 (d, J = 0.9 Hz, 3H) 2.55 (s, 3H), 1.68 (br s, 1H), 0.77–0.75 (m, 2H), 0.67–0.64 (m, 2 H);

13C NMR (151 MHz,CDCl3) δ ppm 158.2, 156.3, 148.5, 147.2, 144.1, 142.2, 140.0, 135.6, 131.2, 126.7, 114.9, 114.7, 110.1, 99.3, 56.7, 49.9, 44.5, 36.5, 16.9, 15.0, 13.0. LC–HRMS: m/z = 402.2051 [(M + H)+ calcd for C22H24N7O, 402.2042; Diff 0.9 mDa].

References

  1. ^ Maria Joao Almeida (2016-09-08). “RG7916”. BioNews Services. Retrieved 2017-10-08.
  2. ^ Zhao, Xin; Feng, Zhihua; Ling, Karen K. Y; Mollin, Anna; Sheedy, Josephine; Yeh, Shirley; Petruska, Janet; Narasimhan, Jana; Dakka, Amal; Welch, Ellen M; Karp, Gary; Chen, Karen S; Metzger, Friedrich; Ratni, Hasane; Lotti, Francesco; Tisdale, Sarah; Naryshkin, Nikolai A; Pellizzoni, Livio; Paushkin, Sergey; Ko, Chien-Ping; Weetall, Marla (2016). “Pharmacokinetics, pharmacodynamics, and efficacy of a small-molecule SMN2 splicing modifier in mouse models of spinal muscular atrophy”Human Molecular Genetics25 (10): 1885. doi:10.1093/hmg/ddw062PMC 5062580PMID 26931466.
  3. ^ “Genentech/Roche Releases Clinical Trial Update for RG7916”. CureSMA. 2017-09-15. Retrieved 2017-10-08.
  4. ^ “A Study to Investigate the Safety, Tolerability, Pharmacokinetics, Pharmacodynamics and Efficacy of RO7034067 in Infants With Type1 Spinal Muscular Atrophy (Firefish)”.
  5. ^ “A Study to Investigate the Safety, Tolerability, Pharmacokinetics, Pharmacodynamics and Efficacy of RO7034067 in Type 2 and 3 Spinal Muscular Atrophy Participants (Sunfish)”.
  6. ^ “Updated Preliminary Data from SMA FIREFISH Program in Type 1 Babies Presented at the CureSMA Conference”http://www.prnewswire.com. Retrieved 2018-09-11.
Risdiplam
Risdiplam.svg
Clinical data
Synonyms RG7916; RO7034067
Identifiers
CAS Number
PubChem CID
UNII
KEGG
Chemical and physical data
Formula C22H23N7O
Molar mass 401.474 g/mol g·mol−1
3D model (JSmol)

///////////RISDIPLAM, RG-7916, RO-7034067, リスジプラム , PHASE 3, PRIME designation, ORPHAN DRUG

76RS4S2ET1 (UNII code)

CC1=CC(=NN2C1=NC(=C2)C)C3=CC(=O)N4C=C(C=CC4=N3)N5CCNC6(C5)CC6

Voretigene neparvovec , ボレチジーンネパルボベック;


Voretigene neparvovec
Voretigene neparvovec-rzyl;
Luxturna (TN)

ボレチジーンネパルボベック;

DNA (synthetic adeno-associated virus 2 vector AAV2-hRPE65v2)

CAS: 1646819-03-5
2017/12/19, FDA  Luxturna, SPARK THERAPEUTICS

Vision loss treatment, Retinal dystrophy

AAV2-hRPE65v2
AAV2.RPE65
LTW-888
SPK-RPE65
rAAV.hRPE65v2
rAAV2-CBSB-hRPE65
2SPI046IKD (UNII code)

melting point (°C) 72-90ºC Rayaprolu V. et al. J. Virol. vol. 87. no. 24. (2013)

FDA

https://www.fda.gov/downloads/BiologicsBloodVaccines/CellularGeneTherapyProducts/ApprovedProducts/UCM592766.pdf

LUXTURNA

STN: 125610
Proper Name: voretigene neparvovec-rzyl
Trade Name: LUXTURNA
Manufacturer: Spark Therapeutics, Inc.
Indication:

  • Is an adeno-associated virus vector-based gene therapy indicated for the treatment of patients with confirmed biallelic RPE65 mutation-associated retinal dystrophy. Patients must have viable retinal cells as determined by the treating physician(s).

Product Information

Related Information

Voretigene neparvovec (Luxturna) is a novel gene therapy for the treatment of Leber’s congenital amaurosis.[1] It was developed by Spark Therapeutics and Children’s Hospital of Philadelphia.[2][3] It is the first in vivo gene therapy approved by the FDA.[4]

Leber’s congenital amaurosis, or biallelic RPE65-mediated inherited retinal disease, is an inherited disorder causing progressive blindness. Voretigene is the first treatment available for this condition.[5] The gene therapy is not a cure for the condition, but substantially improves vision in those treated.[6] It is given as an subretinal injection.

It was developed by collaboration between the University of Pennsylvania, Yale University, the University of Florida and Cornell University. In 2018, the product was launched in the U.S. by Spark Therapeutics for the treatment of children and adult patients with confirmed biallelic RPE65 mutation-associated retinal dystrophy. The same year, Spark Therapeutics received approval for the product in the E.U. for the same indication.

Chemistry and production

Voretigene neparvovec is an AAV2 vector containing human RPE65 cDNA with a modified Kozak sequence. The virus is grown in HEK 293 cells and purified for administration.[7]

History

Married researchers Jean Bennett and Albert Maguire, among others, worked for decades on studies of congenital blindness, culminating in approval of a novel therapy, Luxturna.[8]

It was granted orphan drug status for Leber congenital amaurosis and retinitis pigmentosa.[9][10] A biologics license application was submitted to the FDA in July 2017 with Priority Review.[5] Phase III clinical trial results were published in August 2017.[11] On 12 October 2017, a key advisory panel to the Food and Drug Administration (FDA), composed of 16 experts, unanimously recommended approval of the treatment.[12] The US FDA approved the drug on December 19, 2017. With the approval, Spark Therapeutics received a pediatric disease priority review voucher.[13]

The first commercial sale of voretigene neparvovec — the first for any gene therapy product in the US — occurred in March 2018.[14][14][4] The price of the treatment has been announced at $425,000 per eye.[15]

INDICATION

LUXTURNA (voretigene neparvovec-rzyl) is an adeno-associated virus vector-based gene therapy indicated for the treatment of patients with confirmed biallelic RPE65 mutation-associated retinal dystrophy.

Patients must have viable retinal cells as determined by the treating physicians.

IMPORTANT SAFETY INFORMATION FOR LUXTURNA

Warnings and Precautions

  • Endophthalmitis may occur following any intraocular surgical procedure or injection. Use proper aseptic injection technique when administering LUXTURNA, and monitor for and advise patients to report any signs or symptoms of infection or inflammation to permit early treatment of any infection.

  • Permanent decline in visual acuity may occur following subretinal injection of LUXTURNA. Monitor patients for visual disturbances.

  • Retinal abnormalities may occur during or following the subretinal injection of LUXTURNA, including macular holes, foveal thinning, loss of foveal function, foveal dehiscence, and retinal hemorrhage. Monitor and manage these retinal abnormalities appropriately. Do not administer LUXTURNA in the immediate vicinity of the fovea. Retinal abnormalities may occur during or following vitrectomy, including retinal tears, epiretinal membrane, or retinal detachment. Monitor patients during and following the injection to permit early treatment of these retinal abnormalities. Advise patients to report any signs or symptoms of retinal tears and/or detachment without delay.

  • Increased intraocular pressure may occur after subretinal injection of LUXTURNA. Monitor and manage intraocular pressure appropriately.

  • Expansion of intraocular air bubbles Instruct patients to avoid air travel, travel to high elevations or scuba diving until the air bubble formed following administration of LUXTURNA has completely dissipated from the eye. It may take one week or more following injection for the air bubble to dissipate. A change in altitude while the air bubble is still present can result in irreversible vision loss. Verify the dissipation of the air bubble through ophthalmic examination.

  • Cataract Subretinal injection of LUXTURNA, especially vitrectomy surgery, is associated with an increased incidence of cataract development and/or progression.

Adverse Reactions

  • In clinical studies, ocular adverse reactions occurred in 66% of study participants (57% of injected eyes), and may have been related to LUXTURNA, the subretinal injection procedure, the concomitant use of corticosteroids, or a combination of these procedures and products.

  • The most common adverse reactions (incidence ≥5% of study participants) were conjunctival hyperemia (22%), cataract (20%), increased intraocular pressure (15%), retinal tear (10%), dellen (thinning of the corneal stroma) (7%), macular hole (7%), subretinal deposits (7%), eye inflammation (5%), eye irritation (5%), eye pain (5%), and maculopathy (wrinkling on the surface of the macula) (5%).

Immunogenicity

Immune reactions and extra-ocular exposure to LUXTURNA in clinical studies were mild. No clinically significant cytotoxic T-cell response to either AAV2 or RPE65 has been observed.

In clinical studies, the interval between the subretinal injections into the two eyes ranged from 7 to 14 days and 1.7 to 4.6 years. Study participants received systemic corticosteroids before and after subretinal injection of LUXTURNA to each eye, which may have decreased the potential immune reaction to either AAV2 or RPE65.

Pediatric Use

Treatment with LUXTURNA is not recommended for patients younger than 12 months of age, because the retinal cells are still undergoing cell proliferation, and LUXTURNA would potentially be diluted or lost during the cell proliferation. The safety and efficacy of LUXTURNA have been established in pediatric patients. There were no significant differences in safety between the different age subgroups.

Please see US Full Prescribing Information for LUXTURNA.

References:

1. LUXTURNA [package insert]. Philadelphia, PA: Spark Therapeutics, Inc; 2017. 2. Gupta PR, Huckfeldt RM. Gene therapy for inherited retinal degenerations: initial successes and future challenges. J Neural Eng. 2017;14(5):051002. 3. Kay C. Gene therapy: the new frontier for inherited retinal disease. Retina Specialist. March 2017. http://www.retina-specialist.com/CMSDocuments/2017/03/RS/rs0317I.pdf. Accessed November 14, 2017 4. Polinski NK, Gombash SE, Manfredsson FP, et al. Recombinant adeno-associated virus 2/5-mediated gene transfer is reduced in the aged rat midbrain. Neurobiol Aging. 2015;36(2):1110-1120. 5. Moore T. Restoring retinal function in a mouse model of hereditary blindness. PLoS Med. 2005;2(11):e399. 6. McBee JK, Van Hooser JP, Jang GF, Palczewski K. Isomerization of 11-cis-retinoids to all-trans-retinoids in vitro and in vivo. J Biol Chem. 2001;276(51):48483-48493. 7. Thomas CE, Ehrhardt A, Kay MA. Progress and problems with the use of viral vectors for gene therapy. Nat Rev Genet. 2003;4(5):346-358. 8. Trapani I, Puppo A, Auricchio A. Vector platforms for gene therapy of inherited retinopathies. Prog Retin Eye Res. 2014;43:108-128. 9. Russell S, Bennett J, Wellman JA, et al. Efficacy and safety of voretigene neparvovec (AAV2-hRPE65v2) in patients with RPE65-mediated inherited retinal dystrophy: a randomised, controlled, open-label, phase 3 trial. Lancet. 2017;390(10097):849-860.

Illustration of the RPE65 gene delivery method

Illustration of the RPE65 protein production cycle

PAPERS

Progress in Retinal and Eye Research (2018), 63, 107-131

Lancet (2017), 390(10097), 849-860.

References

  1. ^ “Luxturna (voretigene neparvovec-rzyl) label” (PDF). FDA. December 2017. Retrieved 31 December 2017. (for label updates, see FDA index page)
  2. ^ “Spark’s gene therapy for blindness is racing to a historic date with the FDA”Statnews.com. 9 October 2017. Retrieved 9 October 2017.
  3. ^ Clarke,Reuters, Toni. “Gene Therapy for Blindness Appears Initially Effective, Says U.S. FDA”Scientific American. Retrieved 2017-10-12.
  4. Jump up to:a b “First Gene Therapy For Inherited Disease Gets FDA Approval”NPR.org. 19 Dec 2017.
  5. Jump up to:a b “Press Release – Investors & Media – Spark Therapeutics”Ir.sparktx.com. Retrieved 9 October 2017.
  6. ^ McGinley, Laurie (19 December 2017). “FDA approves first gene therapy for an inherited disease”Washington Post.
  7. ^ Russell, Stephen; Bennett, Jean; Wellman, Jennifer A.; Chung, Daniel C.; Yu, Zi-Fan; Tillman, Amy; Wittes, Janet; Pappas, Julie; Elci, Okan; McCague, Sarah; Cross, Dominique; Marshall, Kathleen A.; Walshire, Jean; Kehoe, Taylor L.; Reichert, Hannah; Davis, Maria; Raffini, Leslie; George, Lindsey A.; Hudson, F Parker; Dingfield, Laura; Zhu, Xiaosong; Haller, Julia A.; Sohn, Elliott H.; Mahajan, Vinit B.; Pfeifer, Wanda; Weckmann, Michelle; Johnson, Chris; Gewaily, Dina; Drack, Arlene; et al. (2017). “Efficacy and safety of voretigene neparvovec (AAV2-hRPE65v2) in patients with RPE65 -mediated inherited retinal dystrophy: A randomised, controlled, open-label, phase 3 trial”The Lancet390 (10097): 849–860. doi:10.1016/S0140-6736(17)31868-8PMC 5726391PMID 28712537.
  8. ^ “FDA approves Spark’s gene therapy for rare blindness pioneered at CHOP – Philly”Philly.com. Retrieved 2018-03-24.
  9. ^ “Voretigene neparvovec – Spark Therapeutics – AdisInsight”adisinsight.springer.com.
  10. ^ Ricki Lewis, PhD (October 13, 2017). “FDA Panel Backs Gene Therapy for Inherited Blindness”Medscape.
  11. ^ Lee, Helena; Lotery, Andrew (2017). “Gene therapy for RPE65 -mediated inherited retinal dystrophy completes phase 3”. The Lancet390 (10097): 823–824. doi:10.1016/S0140-6736(17)31622-7PMID 28712536.
  12. ^ “Landmark Therapy to Treat Blindness Gets One Step Closer to FDA Approval”Bloomberg.com. 2017-10-12. Retrieved 2017-10-12.
  13. ^ “Spark grabs FDA nod for Luxturna, a breakthrough gene therapy likely bearing a pioneering price”FiercePharma.
  14. Jump up to:a b “The anxious launch of Luxturna, a gene therapy with a record sticker price”STAT. 2018-03-21. Retrieved 2018-03-24.
  15. ^ Tirrell, Meg (3 January 2018). “A US drugmaker offers to cure rare blindness for $850,000”. CNBC. Retrieved 3 January 2018.

Further reading

Voretigene neparvovec
Gene therapy
Vector Adeno-associated virusserotype 2
Nucleic acid type DNA
Editing method RPE65
Clinical data
Trade names Luxturna
Pregnancy
category
  • US: N (Not classified yet)
Routes of
administration
subretinal injection
ATC code
Legal status
Legal status
Identifiers
KEGG

//////////FDA 2017, Voretigene neparvovec , Voretigene neparvovec-rzyl, Luxturna, ボレチジーンネパルボベック, 1646819-03-5 , FDA  Luxturna, SPARK THERAPEUTICS, Vision loss treatment, Retinal dystrophy., AAV2-hRPE65v2, LTW-888, SPK-RPE65, Orphan drug,

Elapegademase, エラペグアデマーゼ (遺伝子組換え)


AQTPAFNKPK VELHVHLDGA IKPETILYYG RKRGIALPAD TPEELQNIIG MDKPLSLPEF
LAKFDYYMPA IAGSREAVKR IAYEFVEMKA KDGVVYVEVR YSPHLLANSK VEPIPWNQAE
GDLTPDEVVS LVNQGLQEGE RDFGVKVRSI LCCMRHQPSW SSEVVELCKK YREQTVVAID
LAGDETIEGS SLFPGHVKAY AEAVKSGVHR TVHAGEVGSA NVVKEAVDTL KTERLGHGYH
TLEDTTLYNR LRQENMHFEV CPWSSYLTGA WKPDTEHPVV RFKNDQVNYS LNTDDPLIFK
STLDTDYQMT KNEMGFTEEE FKRLNINAAK SSFLPEDEKK ELLDLLYKAY GMPSPA

str1

>>Elapegademase<<<
AQTPAFNKPKVELHVHLDGAIKPETILYYGRKRGIALPADTPEELQNIIGMDKPLSLPEF
LAKFDYYMPAIAGSREAVKRIAYEFVEMKAKDGVVYVEVRYSPHLLANSKVEPIPWNQAE
GDLTPDEVVSLVNQGLQEGERDFGVKVRSILCCMRHQPSWSSEVVELCKKYREQTVVAID
LAGDETIEGSSLFPGHVKAYAEAVKSGVHRTVHAGEVGSANVVKEAVDTLKTERLGHGYH
TLEDTTLYNRLRQENMHFEVCPWSSYLTGAWKPDTEHPVVRFKNDQVNYSLNTDDPLIFK
STLDTDYQMTKNEMGFTEEEFKRLNINAAKSSFLPEDEKKELLDLLYKAYGMPSPA

ChemSpider 2D Image | ELAPEGADEMASE | C10H20N2O5

Elapegademase, エラペグアデマーゼ (遺伝子組換え)

EZN-2279

Protein chemical formula C1797H2795N477O544S12

Protein average weight 115000.0 Da

Peptide

APPROVED, FDA, Revcovi, 2018/10/5

CAS: 1709806-75-6

Elapegademase-lvlr, Poly(oxy-1,2-ethanediyl), alpha-carboxy-omega-methoxy-, amide with adenosine deaminase (synthetic)

L-Lysine, N6-[(2-methoxyethoxy)carbonyl]-
N6-[(2-Methoxyethoxy)carbonyl]-L-lysine

EZN-2279; PEG-rADA; Pegademase recombinant – Leadiant Biosciences; Pegylated recombinant adenosine deaminase; Polyethylene glycol recombinant adenosine deaminase; STM-279, UNII: 9R3D3Y0UHS

  • Originator Sigma-Tau Pharmaceuticals
  • Developer Leadiant Biosciences; Teijin Pharma
  • Class Antivirals; Polyethylene glycols
  • Mechanism of Action Adenosine deaminase stimulants
  • Orphan Drug Status Yes – Immunodeficiency disorders; Adenosine deaminase deficiency
  • Registered Adenosine deaminase deficiency; Immunodeficiency disorders
  • 05 Oct 2018 Registered for Adenosine deaminase deficiency (In adults, In children) in USA (IM)
  • 05 Oct 2018 Registered for Immunodeficiency disorders (In adults, In children) in USA (IM)
  • 04 Oct 2018 Elapegademase receives priority review status for Immunodeficiency disorders and Adenosine deaminase deficiency in USA

検索キーワード:Elapegademase (Genetical Recombination)
検索件数:1


エラペグアデマーゼ(遺伝子組換え)
Elapegademase (Genetical Recombination)

[1709806-75-6]

Elapegademase is a PEGylated recombinant adenosine deaminase. It can be defined molecularly as a genetically modified bovine adenosine deaminase with a modification in cysteine 74 for serine and with about 13 methoxy polyethylene glycol chains bound via carbonyl group in alanine and lysine residues.[4] Elapegademase is generated in E. coli, developed by Leadiant Biosciences and FDA approved on October 5, 2018.[15]

Indication

Elapegademase is approved for the treatment of adenosine deaminase severe combined immune deficiency (ADA-SCID) in pediatric and adult patients.[1] This condition was previously treated by the use of pegamedase bovine as part of an enzyme replacement therapy.[2]

ADA-SCID is a genetically inherited disorder that is very rare and characterized by a deficiency in the adenosine deaminase enzyme. The patients suffering from this disease often present a compromised immune system. This condition is characterized by very low levels of white blood cells and immunoglobulin levels which results in severe and recurring infections.[3]

Pharmacodynamics

In clinical trials, elapegademase was shown to increase adenosine deaminase activity while reducing the concentrations of toxic metabolites which are the hallmark of ADA-SCID. As well, it was shown to improve the total lymphocyte count.[6]

Mechanism of action

The ADA-SCID is caused by the presence of mutations in the ADA gene which is responsible for the synthesis of adenosine deaminase. This enzyme is found throughout the body but it is mainly active in lymphocytes. The normal function of adenosine deaminase is to eliminate deoxyadenosine, created when DNA is degraded, by converting it into deoxyinosine. This degradation process is very important as deoxyadenosine is cytotoxic, especially for lymphocytes. Immature lymphocytes are particularly vulnerable as deoxyadenosine kills them before maturation making them unable to produce their immune function.[3]

Therefore, based on the causes of ADA-SCID, elapegademase works by supplementing the levels of adenosine deaminase. Being a recombinant and an E. coli-produced molecule, the use of this drug eliminates the need to source the enzyme from animals, as it was used previously.[1]

Absorption

Elapegademase is administered intramuscularly and the reported Tmax, Cmax and AUC are approximately 60 hours, 240 mmol.h/L and 33000 hr.mmol/L as reported during a week.[Label]

Volume of distribution

This pharmacokinetic property has not been fully studied.

Protein binding

This pharmacokinetic property is not significant as the main effect is in the blood cells.

Metabolism

Metabolism studies have not been performed but it is thought to be degraded by proteases to small peptides and individual amino acids.

Route of elimination

This pharmacokinetic property has not been fully studied.

Half life

This pharmacokinetic property has not been fully studied.

Clearance

This pharmacokinetic property has not been fully studied.

Toxicity

As elapegademase is a therapeutic protein, there is a potential risk of immunogenicity.

There are no studies related to overdose but the highest weekly prescribed dose in clinical trials was 0.4 mg/kg. In nonclinical studies, a dosage of 1.8 fold of the clinical dose produced a slight increase in the activated partial thromboplastin time.[Label]

FDA label. Download (145 KB)

General References

  1. Rare DR [Link]
  2. Globe News Wire [Link]
  3. NIH [Link]
  4. NIHS reports [File]
  5. WHO Drug Information 2017 [File]
  6. Revcovi information [File]

/////////////Elapegademase, Peptide, エラペグアデマーゼ (遺伝子組換え) , EZN-2279, Elapegademase-lvlr, Orphan Drug, STM 279, FDA 2018

COCCOC(=O)NCCCC[C@H](N)C(=O)O

“ALL FOR DRUGS” CATERS TO EDUCATION GLOBALLY, No commercial exploits are done or advertisements added by me. This is a compilation for educational purposes only. P.S. : The views expressed are my personal and in no-way suggest the views of the professional body or the company that I represent

 

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Calaspargase pegol, カラスパルガーゼペゴル


LPNITILATG GTIAGGGDSA TKSNYTAGKV GVENLVNAVP QLKDIANVKG EQVVNIGSQD
MNDDVWLTLA KKINTDCDKT DGFVITHGTD TMEETAYFLD LTVKCDKPVV MVGAMRPSTS
MSADGPFNLY NAVVTAADKA SANRGVLVVM NDTVLDGRDV TKTNTTDVAT FKSVNYGPLG
YIHNGKIDYQ RTPARKHTSD TPFDVSKLNE LPKVGIVYNY ANASDLPAKA LVDAGYDGIV
SAGVGNGNLY KTVFDTLATA AKNGTAVVRS SRVPTGATTQ DAEVDDAKYG FVASGTLNPQ
KARVLLQLAL TQTKDPQQIQ QIFNQY
(tetramer; disulfide bridge 77-105, 77′-105′, 77”-105”, 77”’-105”’)

Image result for Calaspargase pegol

str3

Calaspargase pegol

Molecular Formula, C1516-H2423-N415-O492-S8 (peptide monomer), Molecular Weight, 10261.2163

APPROVED, Asparlas, FDA 2018/12/20

CAS 941577-06-6

UNII T9FVH03HMZ

カラスパルガーゼペゴル;

(27-Alanine,64-aspartic acid,252-threonine,263-asparagine)-L-asparaginase 2 (EC 3.5.1.1, L-asparagineamidohydrolase II) Escherichia coli (strain K12) tetramer alpha4, carbamates with alpha-carboxy-omega-methoxypoly(oxyethylene)

Asparaginase (Escherichia coli isoenzyme II), conjugate with alpha-(((2,5-dioxo-1-pyrrolidinyl)oxy)carbonyl)-omega-methoxypoly(oxy-1,2-ethanediyl)

List Acronyms
Peptide
  • Calaspargase pegol
  • calaspargase pegol-mknl
  • EZN-2285
  • Used to treat acute lymphoblastic leukemia., Antineoplastic
  • BAX-2303
    SC-PEG E. Coli L-asparaginase
    SHP-663

Calaspargase pegol-mknl (trade name Asparlas) is a drug for the treatment of acute lymphoblastic leukemia (ALL). It is approved by the Food and Drug Administration for use in the United States as a component of a multi-agent chemotherapeutic regimen for ALL in pediatric and young adult patients aged 1 month to 21 years.[1]

Calaspargase pegol was first approved in 2018 in the U.S. as part of a multi-agent chemotherapeutic regimen for the treatment of patients with acute lymphoblastic leukemia.

In 2008, orphan drug designation was assigned in the E.U.

Calaspargase pegol is an engineered protein consisting of the E. coli-derived enzyme L-asparaginase II conjugated with succinimidyl carbonate monomethoxypolyethylene glycol (pegol).[2] The L-asparaginase portion hydrolyzes L-asparagine to L-aspartic acid depriving the tumor cell of the L-asparagine it needs for survival.[2] The conjugation with the pegol group increases the half-life of the drug making it longer acting.

Asparaginase is an important agent used to treat acute lymphoblastic leukemia (ALL) [1]. Asparagine is incorporated into most proteins, and the synthesis of proteins is stopped when asparagine is absent, which inhibits RNA and DNA synthesis, resulting in a halt in cellular proliferation. This forms the basis of asparaginase treatment in ALL [1][2][6].

Calaspargase pegol, also known as asparlas, is an asparagine specific enzyme which is indicated as a part of a multi-agent chemotherapy regimen for the treatment of ALL [3]. The asparagine specific enzyme is derived from Escherichia coli, as a conjugate of L-asparaginase (L-asparagine amidohydrolase) and monomethoxypolyethylene glycol (mPEG) with a succinimidyl carbonate (SC) linker to create a stable molecule which increases the half-life and decreases the dosing frequency [Label][1].

Calaspargase pegol, by Shire pharmaceuticals, was approved by the FDA on December 20, 2018 for acute lymphoblastic anemia (ALL) [3].

Indication

This drug is is an asparagine specific enzyme indicated as a component of a multi-agent chemotherapeutic regimen for the treatment of acute lymphoblastic leukemia in pediatric and young adult patients age 1 month to 21 years [Label].

The pharmacokinetics of calaspargase pegol were examined when given in combination with multiagent chemotherapy in 124 patients with B-cell lineage ALL [3]. The FDA approval of this drug was based on the achievement and maintenance of nadir serum asparaginase activity above the level of 0.1 U/mL when administering calaspargase, 2500 U/m2 intravenously, at 3-week intervals.

Associated Conditions

Pharmacodynamics

The effect of this drug is believed to occur by selective killing of leukemic cells due to depletion of plasma L-asparagine. Leukemic cells with low expression of asparagine synthetase are less capable of producing L-asparagine, and therefore rely on exogenous L-asparagine for survival [Label]. When asparagine is depleted, tumor cells cannot proliferate [6].

During remission induction, one dose of SC-PEG (2500 IU/m2) results in a sustained therapeutic serum asparaginase activity (SAA) without excessive toxicity or marked differences in the proportion of patients with low end-induction minimum residual disease (MRD) [5].

Pharmacodynamic (PD) response was studied through measurement of plasma and cerebrospinal fluid (CSF) asparagine concentrations with an LC-MS/MS assay (liquid chromatography–mass spectrometry). Asparagine concentration in plasma was sustained below the assay limit of quantification for more than 18 days after one dose of calaspargase pegol, 2,500 U/m2, during the induction phase of treatment. Average cerebrospinal asparagine concentrations decreased from a pretreatment concentration of 0.8 μg/mL (N=10) to 0.2 μg/mL on Day 4 (N=37) and stayed decreased at 0.2 μg/mL (N=35) 25 days after the administration of one of 2,500 U/m2 in the induction phase [Label].

Mechanism of action

L-asparaginase (the main component of this drug) is an enzyme that catalyzes the conversion of the amino acid L-asparagine into both aspartic acid and ammonia [Label][2]. This process depletes malignant cells of their required asparagine. The depletion of asparagine then blocks protein synthesis and tumor cell proliferation, especially in the G1 phase of the cell cycle. As a result, tumor cell death occurs. Asparagine is important in protein synthesis in acute lymphoblastic leukemia (ALL) cells which, unlike normal cells, cannot produce this amino acid due to lack of the enzyme asparagine synthase [2][Label].

Pegylation decreases enzyme antigenicity and increases its half-life. Succinimidyl carbamate (SC) is used as a PEG linker to facilitate attachment to asparaginase and enhances the stability of the formulation [4][1]. SC-PEG urethane linkages formed with lysine groups are more hydrolytically stable [2].

Toxicity

Pancreatitis, hepatotoxicity, hemorrhage, and thrombosis have been observed with calaspargase pegol use [Label].

Pancreatitis: Discontinue this drug in patients with pancreatitis, and monitor blood glucose.

Hepatotoxicity: Hepatic function should be tested regularly, and trough levels of this drug should be measured during the recovery phase of the drug cycle [Label].

Hemorrhage or Thrombosis: Discontinue this drug in serious or life-threatening hemorrhage or thrombosis. In cases of hemorrhage, identify the cause of hemorrhage and treat appropriately. Administer anticoagulant therapy as indicated in thrombotic events [Label].

A note on hypersensitivity:

Observe the patient for 1 hour after administration of calaspargase pegol for possible hypersensitivity [Label]. In cases of previous hypersensitivity to this drug, discontinue this drug immediately.

Lactation: Advise women not to breastfeed while taking this drug [Label].

Pregnancy: There are no available data on the use of calaspargase pegol in pregnant women to confirm a risk of drug-associated major birth defects and miscarriage. Published literature studies in pregnant animals suggest asparagine depletion can cause harm to the animal offspring. It is therefore advisable to inform women of childbearing age of this risk. The background risk of major birth defects and miscarriage for humans is unknown at this time [Label].

Pregnancy testing should occur before initiating treatment. Advise females of reproductive potential to avoid becoming pregnant while taking this drug. Females should use effective contraceptive methods, including a barrier methods, during treatment and for at least 3 months after the last dose. There is a risk for an interaction between calaspargase pegol and oral contraceptives. The concurrent use of this drug with oral contraceptives should be avoided. Other non-oral contraceptive methods should be used in women of childbearing potential [Label].

References
  1. Angiolillo AL, Schore RJ, Devidas M, Borowitz MJ, Carroll AJ, Gastier-Foster JM, Heerema NA, Keilani T, Lane AR, Loh ML, Reaman GH, Adamson PC, Wood B, Wood C, Zheng HW, Raetz EA, Winick NJ, Carroll WL, Hunger SP: Pharmacokinetic and pharmacodynamic properties of calaspargase pegol Escherichia coli L-asparaginase in the treatment of patients with acute lymphoblastic leukemia: results from Children’s Oncology Group Study AALL07P4. J Clin Oncol. 2014 Dec 1;32(34):3874-82. doi: 10.1200/JCO.2014.55.5763. Epub 2014 Oct 27. [PubMed:25348002]
  2. Appel IM, Kazemier KM, Boos J, Lanvers C, Huijmans J, Veerman AJ, van Wering E, den Boer ML, Pieters R: Pharmacokinetic, pharmacodynamic and intracellular effects of PEG-asparaginase in newly diagnosed childhood acute lymphoblastic leukemia: results from a single agent window study. Leukemia. 2008 Sep;22(9):1665-79. doi: 10.1038/leu.2008.165. Epub 2008 Jun 26. [PubMed:18580955]
  3. Blood Journal: Randomized Study of Pegaspargase (SS-PEG) and Calaspargase Pegol (SPC-PEG) in Pediatric Patients with Newly Diagnosed Acute Lymphoblastic Leukemia or Lymphoblastic Lymphoma: Results of DFCI ALL Consortium Protocol 11-001 [Link]

References

  1. ^ “FDA approves longer-acting calaspargase pegol-mknl for ALL” (Press release). Food and Drug Administration. December 20, 2018.
  2. Jump up to:a b “Calaspargase pegol-mknl”NCI Drug Dictionary. National Cancer Institute.

FDA label, Download(300 KB)

General References

  1. Angiolillo AL, Schore RJ, Devidas M, Borowitz MJ, Carroll AJ, Gastier-Foster JM, Heerema NA, Keilani T, Lane AR, Loh ML, Reaman GH, Adamson PC, Wood B, Wood C, Zheng HW, Raetz EA, Winick NJ, Carroll WL, Hunger SP: Pharmacokinetic and pharmacodynamic properties of calaspargase pegol Escherichia coli L-asparaginase in the treatment of patients with acute lymphoblastic leukemia: results from Children’s Oncology Group Study AALL07P4. J Clin Oncol. 2014 Dec 1;32(34):3874-82. doi: 10.1200/JCO.2014.55.5763. Epub 2014 Oct 27. [PubMed:25348002]
  2. Appel IM, Kazemier KM, Boos J, Lanvers C, Huijmans J, Veerman AJ, van Wering E, den Boer ML, Pieters R: Pharmacokinetic, pharmacodynamic and intracellular effects of PEG-asparaginase in newly diagnosed childhood acute lymphoblastic leukemia: results from a single agent window study. Leukemia. 2008 Sep;22(9):1665-79. doi: 10.1038/leu.2008.165. Epub 2008 Jun 26. [PubMed:18580955]
  3. Asparlas Approval History [Link]
  4. NCI: Calaspargase Pegol [Link]
  5. Blood Journal: Randomized Study of Pegaspargase (SS-PEG) and Calaspargase Pegol (SPC-PEG) in Pediatric Patients with Newly Diagnosed Acute Lymphoblastic Leukemia or Lymphoblastic Lymphoma: Results of DFCI ALL Consortium Protocol 11-001 [Link]
  6. Medsafe NZ: Erwinaze inj [File]
Calaspargase pegol-mknl
Clinical data
Trade names Asparlas
Synonyms EZN-2285
Legal status
Legal status
Identifiers
CAS Number
DrugBank
UNII
KEGG
ChEMBL

/////////////Calaspargase pegol, Peptide, FDA 2018, EZN-2285, カラスパルガーゼペゴル  , BAX-2303, SC-PEG E. Coli L-asparaginase , SHP-663, orphan drug

CC(C)C[C@@H](C(=O)O)NC(=O)OCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOC.COCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOC(=O)NCCCC[C@@H](C(=O)O)N

IMETELSTAT


Image result for IMETELSTAT

Image result for IMETELSTAT

2D chemical structure of 868169-64-6

IMETELSTAT

CAS 868169-64-6, N163L

Molecular Formula, C148-H211-N68-O53-P13-S13, Molecular Weight, 4610.2379,

Nucleic Acid Sequence

Sequence Length: 135 a 1 c 4 g 3 tmodified

DNA d(3′-amino-3′-deoxy-P-thio)(T-A-G-G-G-T-T-A-G-A-C-A-A) 5′-[O-[2-hydroxy-3-[(1-oxohexadecyl)amino]propyl] hydrogen phosphorothioate]

PHASE 3, GERON, Myelodysplasia

Image result for IMETELSTAT

ChemSpider 2D Image | Imetelstat sodium | C148H197N68Na13O53P13S13

IMETELSTAT SODIUM

CAS 1007380-31-5, GRN163L, GRN 163L Sodium Salt

Molecular Formula: C148H198N68Na13O53P13S13
Molecular Weight: 4895.941 g/mol

5′-(O-(2-hydroxy-3-((1-oxohexadecyl)amino)propyl)phosphorothioate)-d(3′-amino-3′-deoxy-p-thio)(t-a-g-g-g-t-t-a-g-a-c-a-a), sodium salt (13)

DNA, d(3′-amino-3′-deoxy-p-thio)(T-A-G-G-G-T-T-A-G-A-C-A-A), 5′-(o-(2-hydroxy-3-((1-oxohexadecyl)amino)propyl) hydrogen phosphorothioate), sodium salt (1:13)

UNII-2AW48LAZ4I, Antineoplastic

In 2014, Geron entered into an exclusive worldwide license and collaboration agreement with Janssen Biotech for the treatment of hematologic cancers. However, in 2018, the agreement was terminated and Geron regained global rights to the product.

In 2015, imetelstat was granted orphan drug status in the U.S. for the treatment of myelodysplastic syndrome, as well as in both the U.S. and the E.U. for the treatment of myelofibrosis. In 2017, fast track designation was received in the U.S. for the treatment of adult patients with transfusion-dependent anemia due to low or intermediate-1 risk myelodysplastic syndromes (MDS) who are non-del(5q) and who are refractory or resistant to treatment with an erythropoiesis stimulating agent (ESA).

Imetelstat Sodium is the sodium salt of imetelstat, a synthetic lipid-conjugated, 13-mer oligonucleotide N3′ P5′-thio-phosphoramidate with potential antineoplastic activity. Complementary to the template region of telomerase RNA (hTR), imetelstat acts as a competitive enzyme inhibitor that binds and blocks the active site of the enzyme (a telomerase template antagonist), a mechanism of action which differs from that for the antisense oligonucleotide-mediated inhibition of telomerase activity through telomerase mRNA binding. Inhibition of telomerase activity in tumor cells by imetelstat results in telomere shortening, which leads to cell cycle arrest or apoptosis.

Imetelstat sodium, a lipid-based conjugate of Geron’s first-generation anticancer drug, GRN-163, is in phase III clinical trials at Geron for the treatment of myelodysplastic syndrome, as well as in phase II for the treatment of myelofibrosis. 

Geron is developing imetelstat, a lipid-conjugated 13-mer thiophosphoramidate oligonucleotide and the lead in a series of telomerase inhibitors, for treating hematological malignancies, primarily myelofibrosis.

Imetelstat, a first-in-class telomerase inhibitor and our sole product candidate, is being developed for the potential treatment of hematologic myeloid malignancies. Imetelstat is currently in two clinical trials being conducted by Janssen under the terms of an exclusive  worldwide collaboration and license agreement.

Originally known as GRN163L, imetelstat sodium (imetelstat) is a 13-mer N3’—P5’ thio-phosphoramidate (NPS) oligonucleotide that has a covalently bound 5’ palmitoyl (C16) lipid group. The proprietary nucleic acid backbone provides resistance to the effect of cellular nucleases, thus conferring improved stability in plasma and tissues, as well as significantly improved binding affinity to its target. The lipid group enhances cell permeability to increase potency and improve pharmacokinetic and pharmacodynamic properties. The compound has a long residence time in bone marrow, spleen and liver. Imetelstat binds with high affinity to the template region of the RNA component of telomerase, resulting in direct, competitive inhibition of telomerase enzymatic activity, rather than elicit its effect through an antisense inhibition of protein translation. Imetelstat is administered by intravenous infusion.

Preclinical Studies with Imetelstat

A series of preclinical efficacy studies of imetelstat have been conducted by Geron scientists and academic collaborators. These data showed that imetelstat:

  • Inhibits telomerase activity, and can shorten telomeres.
  • Inhibits the proliferation of a wide variety of tumor types, including solid and hematologic, in cell culture systems and rodent xenograft models of human cancers, impacting the growth of primary tumors and reducing metastases.
  • Inhibits the proliferation of malignant progenitor cells from hematologic cancers, such as multiple myeloma, myeloproliferative neoplasms and acute myelogenous leukemia.
  • Has additive or synergistic anti-tumor effect in a variety of cell culture systems and xenograft models when administered in combination with approved anti-cancer therapies, including radiation, conventional chemotherapies and targeted agents.

Clinical Experience with Imetelstat

Over 500 patients have been enrolled and treated in imetelstat clinical trials.

PHASE 1

Six clinical trials evaluated the safety, tolerability, pharmacokinetics and pharmacodynamics both as a single agent and in combination with standard therapies in patients with solid tumors and hematologic malignancies:

  • Single agent studies of imetelstat were in patients with advanced solid tumors, multiple myeloma and chronic lymphoproliferative diseases. Combination studies with imetelstat were with bortezomib in patients with relapsed or refractory multiple myeloma, with paclitaxel and bevacizumab in patients with metastatic breast cancer, and with carboplatin and paclitaxel in patients with advanced non-small cell lung cancer (NSCLC).
  • Doses ranging from 0.5 mg/kg to 11.7 mg/kg were tested in a variety of dosing schedules ranging from weekly to once every 28 days.
  • The human pharmacokinetic profile was characterized in clinical trials of patients with solid tumors and chronic lymphoproliferative diseases. Single-dose kinetics showed dose-dependent increases in exposure with a plasma half-life (t1/2) ranging from 4-5 hours. Residence time in bone marrow is long (0.19-0.51 µM observed at 41-45 hours post 7.5 mg/kg dose).
  • Telomerase inhibition was observed in various tissues where the enzymes’s activity was measurable.

PHASE 2

Imetelstat was studied in two randomized clinical trials, two single arm proof-of-concept studies and an investigator sponsored pilot study:

  • Randomized trials were in combination with paclitaxel in patients with metastatic breast cancer and as maintenance treatment following a platinum-containing chemotherapy regimen in patients with NSCLC.
  • Single arm studies were as a single agent or in combination with lenalidomide in patients with multiple myeloma and as a single agent in essential thrombocythemia (ET) or polycythemia vera (PV).
  • An investigator sponsored pilot study was as a single agent in patients with myelofibrosis (MF) or myelodysplastic syndromes (MDS).

SAFETY AND TOLERABILITY

The safety profile of imetelstat across the Phase 1 and 2 trials has been generally consistent. Reported adverse events (AEs) and laboratory investigations associated with imetelstat administration included cytopenias, transient prolonged activated partial thromboplastin time (aPTT; assessed only in Phase 1 trials), gastrointestinal symptoms, constitutional symptoms, hepatic biochemistry abnormalities, and infusion reactions. Dose limiting toxicities include thrombocytopenia and neutropenia.

A Focus on Hematologic Myeloid Malignancies

Early clinical data from the Phase 2 clinical trial in ET and the investigator sponsored pilot study in MF suggest imetelstat may have disease-modifying activity by suppressing the proliferation of malignant progenitor cell clones for the underlying diseases, and potentially allowing recovery of normal hematopoiesis in patients with hematologic myeloid malignancies.

Results from these trials were published in the New England Journal of Medicine:

Current Clinical Trials

Imetelstat is currently being tested in two clinical trials: IMbark, a Phase 2 trial in myelofibrosis (MF), and IMerge, a Phase 2/3 trial in myelodysplastic syndromes (MDS).

IMbark

IMbark is the ongoing Phase 2 clinical trial to evaluate two doses of imetelstat in intermediate-2 or high-risk MF patients who are refractory to or have relapsed after treatment with a JAK inhibitor.

Internal data reviews were completed in September 2016, April 2017 and March 2018. The safety profile was consistent with prior clinical trials of imetelstat in hematologic malignancies, and no new safety signals were identified. The data supported 9.4 mg/kg as an appropriate starting dose in the trial, but an insufficient number of patients met the protocol defined interim efficacy criteria and new patient enrollment was suspended in October 2016. As of January 2018, median follow up was approximately 19 months, and median overall survival had not been reached in either dosing arm. In March 2018, the trial was closed to new patient enrollment. Patients who remain in the treatment phase of the trial may continue to receive imetelstat, and until the protocol-specified primary analysis, all safety and efficacy assessments are being conducted as planned in the protocol, including following patients, to the extent possible, until death, to enable an assessment of overall survival.

IMerge

IMerge is the ongoing two-part Phase 2/3 clinical trial of imetelstat in red blood cell (RBC) transfusion-dependent patients with lower risk MDS who are refractory or resistant to treatment with an erythropoiesis stimulating agent (ESA). Part 1 is a Phase 2, open-label, single-arm trial of imetelstat administered as a single agent by intravenous infusion, and is ongoing. Part 2 is designed to be a Phase 3, randomized, controlled trial, and has not been initiated.

Preliminary data as of October 2017 from the first 32 patients enrolled in the Part 1 (Phase 2) of IMerge were presented as a poster at the American Society of Hematology Annual Meeting in December 2017.

The data showed that among the subset of 13 patients who had not received prior treatment with either lenalidomide or a hypomethylating agent (HMA) and did not have a deletion 5q chromosomal abnormality (non-del(5q)), 54% achieved RBC transfusion-independence (TI) lasting at least 8 weeks, including 31% who achieved a 24-week RBC-TI. In the overall trial population, the rates of 8- and 24-week RBC-TI were 38% and 16%, respectively. Cytopenias, particularly neutropenia and thrombocytopenia, were the most frequently reported adverse events, which were predictable, manageable and reversible.

Based on the preliminary data from the 13-patient subset, Janssen expanded Part 1 of IMerge to enroll approximately 20 additional patients who were naïve to lenalidomide and HMA treatment and non-del(5q) to increase the experience and confirm the benefit-risk profile of imetelstat in this refined target patient population

PATENT

WO 2005023994

WO 2006113426
WO 2006113470

 WO 2006124904

WO 2008054711

WO 2008112129

US 2014155465

WO 2014088785

PATENT

WO 2016172346

http://appft1.uspto.gov/netacgi/nph-Parser?Sect1=PTO1&Sect2=HITOFF&d=PG01&p=1&u=/netahtml/PTO/srchnum.html&r=1&f=G&l=50&s1=20160312227.PGNR.

PATENT

WO2018026646

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2018026646

Patients of acute myeloid leukemia (AML) have limited treatment options at diagnosis; treatment typically takes the form of chemotherapy to quickly reduce the leukemic cell burden. Invasive leukapheresis procedures to remove large numbers of leukocytes (normal and diseased) may be applied in parallel to chemotherapy to temporarily lower tumor cell burden. Induction phase chemotherapy can be successful but, most healthy cells residing in patient bone marrow are also killed, causing illness and requiring additional palliative therapy to ward off infection and raise leukocyte counts. Additional rounds of chemotherapy can be used in an attempt to keep patients in remission; but relapse is common.

[0005] Telomerase is present in over 90% of tumors across all cancer types; and is lacking in normal, healthy tissues. Imetelstat sodium is a novel, first-in-class telomerase inhibitor that is a covalently-lipidated 13-mer oligonucleotide (shown below) complimentary to the human telomerase RNA (hTR) template region. Imetelstat sodium does not function through an anti-sense mechanism and therefore lacks the side effects commonly observed with such therapies. Imetelstat sodium is the sodium salt of imetelstat (shown below):

Imetelstat sodium

Unless otherwise indicated or clear from the context, references below to imetelstat also include salts thereof. As mentioned above, imetelstat sodium in particular is the sodium salt of imetelstat.

[0006] ABT-199/venetoclax (trade name Venclexta) is an FDA approved Bcl-2 inhibitor for use in chronic lymphocytic leukemia (CLL) patients with dell7p who are relapsed/refractory. ABT-199 is also known as ABT 199, GDC0199, GDC-0199 or RG7601. The chemical name for ABT-199 is 4-[4-[[2-(4-chlorophenyl)-4,4-dimethylcyclohexen-l-yl]methyl]piperazin-l-yl]-N-[3-nitro-4-(oxan-4-ylmethylamino)phenyl]sulfonyl-2-(lH-pyrrolo[2,3-b]pyridin-5-yloxy)benzamide (Cas No. 1257044-40-8). Unless otherwise indicated or clear from the context, references below to ABT-199 also include pharmaceutically acceptable salts thereof. Specifically in the Examples however, ABT-199 was used in the free base form.

[0007] ABT-199, shown below in the free base form, is highly specific to Bcl-2, unlike other first generation inhibitors which show affinity for related Bel family members and induce greater side effects. Inhibition of Bcl-2 blocks the pro-apoptotic signals caused by damage to or abnormalities within cellular DNA and ultimately leads to programmed cell death in treated cells via the caspase cascade and apoptosis through the intrinsic pathway.

ABT-199 (shown in the free base form)

PATENT

WO-2019011829

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2019011829&tab=PCTDESCRIPTION&maxRec=1000

Improved process for preparing imetelstat .  claiming use of a combination comprising a telomerase inhibitor, specifically imetelstat sodium and a Bcl-2 inhibitor, specifically ABT-199 for treating hematological cancer such as acute myeloid leukemia, essential thrombocythemia and polycythemia vera, specifically acute myeloid leukemia.

Imetelstat (SEQ ID NO: 1 ) is a N3′- P5′ thiophosphoramidate oligonucleotide covalently linked to a palmitoyl lipid moiety and has been described in WO-2005/023994 as compound (1 F). The sodium salt of imetelstat acts as a potent and specific telomerase inhibitor and can be used to treat telomerase-mediated disorders, e.g. cancer, including disorders such as myelofibrosis (MF), myelodysplastic syndromes (MDS) and acute myelogenous leukemia (AML).

The structure of imetelstat sodium is shown below :

The structure of imetelstat can also be represented as shown below

imetelstat

The LPT group represents the palmitoyi lipid that is covalently linked to the N3′- P5′ thiophosphor-amidate oligonucleotide. The base sequence of the thirteen nucleotides is as follows :

TAGGGTTAGACAA and is represented by the bases B1 to B13. The -NH-P(=S)(OH)-and -0-P(=S)(OH)- groups of the structure can occur in a salt form. It is understood that salt forms of a subject compound are encompassed by the structures depicted herein, even if not specifically indicated.

Imetelstat sodium can also be represented as follows

o H

LPT = CH3-(CH2)i4-C-N-CH2-(CHOH)-CH2-

The -NH-P(=S)(OH)- group and the thymine, adenine, guanine and cytosine bases can occur in other tautomeric arrangements then used in the figures of the description. It is understood that all tautomeric forms of a subject compound are encompassed by a structure where one possible tautomeric form of the compound is described, even if not specifically indicated.

Prior art

The synthetic scheme used in WO-2005/023994 to prepare imetelstat as compound (1 F) is described in Scheme 1 and Scheme 2. The synthesis of this oligonucleotide is achieved using the solid-phase phosphoramidite methodology with all reactions taking place on solid-phase support. The synthesis of imetelstat is carried out on controlled pore glass (LCAA-CPG) loaded with

3-palmitoylamido-1-0-(4, 4′-dimethoxytrityl)-2-0-succinyl propanediol. The oligonucleotide is assembled from the 5′ to the 3′ terminus by the addition of protected nucleoside 5′-phosphor-amidites with the assistance of an activator. Each elongation cycle consists of 4 distinct, highly controlled steps : deprotection, amidite coupling, sulfurization and a capping step.

Scheme 1 : imetelstat synthetic scheme cycle 1

3. Sulfurization

In Scheme 1 the solid-phase supported synthesis starts with removal of the acid-labile 4,4-dimethoxy-trityl (DMT) protecting group from the palmitoylamidopropanediol linked to the solid-phase support. The first phosphoramidite nucleotide is coupled to the support followed by sulfurization of the phosphor using a 0.1 M solution of phenylacetyl disulfide (PADS) in a mixture of acetonitrile and 2,6-lutidine (1 : 1 ratio). Then a capping step is applied to prevent any unreacted solid-phase support starting material from coupling with a phosphoramidite nucleotide in the following reaction cycles. Capping is done using an 18:1 :1 mixture of THF / isobutyric anhydride / 2,6-lutidine.

After the first cycle on the solid-phase support, chain elongation is achieved by reaction of the 3′-amino group of the support-bound oligonucleotide with an excess of a solution of the protected nucleotide phosphoramidite monomer corresponding to the next required nucleotide in the sequence as depicted in Scheme 2.

Scheme 2 : imetelstat synthetic scheme cycle 2-13

In Scheme 2 the first cycle is depicted of the chain elongation process which is achieved by deprotection of the 3′-amino group of the support-bound oligonucleotide (a), followed by a coupling reaction of the 3′-amino group of the support-bound oligonucleotide (b) with an excess of a solution of a 5′-phosphoramidite monomer corresponding to the next required nucleotide in the sequence of imetelstat. The coupling reaction is followed by sulfurization of the phosphor of the support-bound oligonucleotide (c) and a capping step (see Scheme 3) to prevent any unreacted solid-phase support starting material (b) from coupling with a 5′-phosphoramidite nucleotide in the following reaction cycles. The reaction cycle of Scheme 2 is repeated 12 times before the solid-phase support-bound oligonucleotide is treated with a 1 :1 mixture of ethanol and concentrated ammonia, followed by HPLC purification to obtain imetelstat.

Scheme 3

The capping step using an 18:1 : 1 mixture of THF / isobutyric anhydride / 2,6-lutidine is done to convert after the coupling step any remaining solid-phase support bound oligonucleotide (b) with a primary 3′-amino group into oligonucleotide (e) with a protected (or ‘capped’) 3′-amino group in order to prevent the primary 3′-amino group from coupling with a phosphoramidite nucleotide in the next reaction cycles.

WO-01/18015 discloses in Example 3 with SEQ ID No. 2 a N3’^P5′ thiophosphoramidate oligonucleotide and a process for preparing this oligonucleotide encompassing a capping step.

Herbert B-S et al. discusses the lipid modification of GRN163 (Oncogene (2005) 24, 5262-5268).

Makiko Horie et al. discusses the synthesis and properties of 2′-0,4′-C-ethylene-bridged nucleic acid oligonucleotides targeted to human telomerase RNA subunit (Nucleic Acids Symposium Series (2005) 49, 171-172).

Description of the invention

The coupling reaction in the solid-phase support bound process disclosed in WO-01/18015 and WO-2005/023994 include a capping step to prevent any unreacted primary 3′ amino groups on the support-bound oligonucleotide from reacting during subsequent cycles.

It has now surprisingly been found that the use of a capping step as described in the prior art is superfluous and that imetelstat can be prepared using a 3-step cycle without an additional capping step with nearly identical yield and purity compared to the prior art 4-step cycle that uses a specific capping step. Eliminating the capping step from each cycle benefits the overall process by reducing the number of cycle steps by 22% (from 54 to 42 steps) and consequent reduction of process time. Also, the solvent consumption is reduced due to the reduction of cycle steps which makes for a greener process.

Wherever the term “capping step” is used throughout this text, it is intended to define an additional chemical process step wherein the primary free 3′-amino group on the solid-phase support bound oligonucleotide is converted into a substituted secondary or tertiary 3′-amino group that is not capable of participating in the coupling reaction with a protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropylamino-phosphoramidite monomer in the ensuing coupling step.

In one embodiment, the present invention relates to a method of synthesizing an oligonucleotide N3′ – P5′ thiophosphoramidate of formula

imetelstat

the method comprises of

a) providing a first 3′-amino protected nucleotide attached to a solid-phase support of formula (A) wherein PG is an acid-labile protecting group;

b) deprotecting the protected 3′-amino group to form a free 3′-amino group;

c) reacting the free 3′-amino group with a protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N- diisopropylaminophosphoramidite monomer of formula (B n) wherein n = 2 to form an internucleoside N3′- P5′-phosphoramidite linkage;

mer (B’n)

d) sulfurization of the internucleoside phosphoramidite group using an acyl disulfide to form a N3′- P5′ thiophosphoramidate;

e) repeating 1 1 times in successive order the deprotection step b), the coupling step c) with a protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropylamino-phosphoramidite monomer of formula (B n) wherein the protected nucleoside base B’ in monomer (B n) is successively the protected nucleobase B3 to B13 in the respective 1 1 coupling steps, and the sulfurization step d);

f) removing the acid-labile protecting group PG; and

g) cleaving and deprotecting imetelstat from the solid-phase support;

characterized in that no additional capping step is performed in any of the reaction steps a) to e).

In one embodiment, the present invention relates to a method of synthesizing the N3′ – P5′

thiophosphoramidate oligonucleotide imetelstat of formula

imetelstat

the method comprises of

a) providing a first 3′-amino protected nucleotide attached to a solid-phase support of formula (A) wherein PG is an acid-labile protecting group;

b) deprotecting the protected 3′-amino group to form a free 3′-amino group;

c) reacting the free 3′-amino group with a protected 3′-aminonucleoside-5′-0-cyanoethyl- Ν,Ν-diisopropylaminophosphoramidite monomer of formula (B n), wherein B n with n = 2 is protected A, to form an internucleoside N3′- P5′-phosphoramidite linkage;

mer

d) sulfurization of the internucleoside phosphoramidite group using an acyl disulfide to form a N3′- P5′ thiophosphoramidate;

e) repeating 1 1 times in successive order the deprotection step b), the coupling step c) with a protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropylamino-phosphoramidite monomer of formula (B n) wherein the nucleoside base B’ of monomer (B n) is protected B except when B is thymine, and wherein Bn is successively nucleobase B3 to B13 in the respective 1 1 coupling steps, and the sulfurization step d);

f) removing the acid-labile protecting group PG; and

g) deprotecting and cleaving imetelstat from the solid-phase support;

characterized in that no additional capping step is performed in any of the reaction steps a) to e).

In one embodiment, the present invention relates to a method of synthesizing the N3′ – P5′

thiophosphoramidate oligonucleotide imetelstat of formula

imetelstat

thymine

adenine

guanine


cytosine

9 H

LPT =CH3-(CH2)i4-C-N-CH2-(CHOH)-CH2-

the method comprises of

a) providing a first protected 3′-amino nucleotide attached to a solid-phase support of formula (A) wherein PG is an acid-labile protecting group;

b) deprotecting the PG-protected 3′-amino nucleotide to form a free 3′-amino nucleotide of formula (A’);

c) coupling the free 3′-amino nucleotide with a protected 3′-aminonucleoside-5′-0- cyanoethyl-N,N-diisopropylaminophosphoramidite monomer (B n), wherein B nwith n = 2 is protected A, to form an internucleoside N3′- P5′-phosphoramidite linkage;

monomer (B’n)

d) sulfurizing the N3′- P5′-phosphoramidite linkage using an acyl disulfide to form an internucleoside N3′- P5′ thiophosphoramidate linkage;

e) repeating 1 1 times in successive order:

the deprotecting step b);

the coupling step c) with a protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N- diisopropylamino-phosphoramidite monomer (B n) wherein the nucleoside base B’ of monomer (B n) is protected B except when B is thymine, and wherein Bn is successively nucleobase B3 to B13 in the respective 1 1 coupling steps; and

the sulfurizing step d);

to produce a protected N3′ – P5′ thiophosphoramidate oligonucleotide imetelstat attached to the solid-phase support;

f) removing the 3′-terminal acid-labile protecting group PG from the protected N3′ – P5′ thiophosphoramidate oligonucleotide imetelstat; and

g) deprotecting and cleaving the protected N3′ – P5′ thiophosphoramidate oligonucleotide imetelstat from the solid-phase support to produce imetelstat;

characterized in that no additional capping step is performed in any of the reaction steps a) to e).

A wide variety of solid-phase supports may be used with the invention, including but not limited to, such as microparticles made of controlled pore glass (CPG), highly cross-linked polystyrene, hybrid controlled pore glass loaded with cross-linked polystyrene supports, acrylic copolymers, cellulose, nylon, dextran, latex, polyacrolein, and the like.

The 3′-amino protected nucleotide attached to a solid-phase support of formula (A)

can be prepared as disclosed in WO-2005/023994 wherein a controlled pore glass support loaded with 3-palmitoylamido-1-0-(4, 4′-dimethoxytrityl)-2-0-succinyl propanediol has been coupled with a protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropylaminophosphoramidite monomer of formula (B^ )

monomer (B’-| ) wherein B’-| = T

wherein PG is an acid-labile protecting group. Suitable acid-labile 3′-amino protecting groups PG are, but not limited to, e.g. triphenylmethyl (i.e. trityl or Tr), p-anisyldiphenylmethyl (i.e. mono-methoxytrityl or MMT), and di-p-anisylphenylmethyl (i.e. dimethoxytrityl or DMT).

The protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropylaminophosphoramidite monomers of formula (B n) have a 3′-amino protecting group PG which is an acid-labile group, such as triphenylmethyl (i.e. trityl or Tr), p-anisyldiphenylmethyl (i.e. monomethoxytrityl or MMT), or di-p-anisylphenylmethyl (i.e. dimethoxytrityl or DMT). Furthermore the nucleoside base B’ is protected with a base-labile protecting group (except for thymine).

ed A ed C ed A ed A

B’s = protected A G = guanine

B’g = protected G C = cytosine

The nucleotide monomers and B’2 to B’13 are used successively in the 13 coupling steps starting from the provision of a solid-phase support loaded with 3-palmitoylamido-1-0-(4, 4′-dimethoxytrityl)-2-0-succinyl propanediol and coupled to nucleotide monomer and the following cycle of 12 deprotection, coupling, and sulfurization reactions wherein the nucleotide monomers B’2 to B -I 3 are used.

The 3′-amino protecting group PG can be removed by treatment with an acidic solution such as e.g. dichloroacetic acid in dichloromethane or toluene.

The nucleoside base B’ in the protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropyl-aminophosphoramidite monomers of formula (B n) is protected with a base-labile protecting group which is removed in step g). Suitable base-labile protecting groups for the nucleoside base adenine, cytosine or guanine are e.g. acyl groups such as acetyl, benzoyl, isobutyryl, dimethyl-formamidinyl, or dibenzylformamidinyl. Under the reaction conditions used in oligonucleotide synthesis the thymine nucleoside base does not require protection. Such protected 3′- amino-nucleoside-5′-0-cyanoethyl-N,N-diisopropylaminophosphoramidite monomers of formula (B N) having a 3′-amino protected with an acid-labile group protecting group PG and a nucleoside base B’ protected with a base-labile protecting group are commercially available or can be prepared as described in WO-2006/014387.

The coupling step c) is performed by adding a solution of protected 3′-aminonucleoside-5′-0-cyanoethyl-N,N-diisopropylaminophosphoramidite monomer of formula (BN) and a solution of an activator (or a solution containing the phosphoramidite monomer (BN) and the activator) to the reaction vessel containing the free amino group of an (oligo)nucleotide covalently attached to a solid support. The mixture is then mixed by such methods as mechanically vortexing, sparging with an inert gas, etc. Alternately, the solution(s) of monomer and activator can be made to flow through a reaction vessel (or column) containing the solid-phase supported (oligo)nucleotide with a free 3′-amino group. The monomer and the activator either can be premixed, mixed in the valve-block of a suitable synthesizer, mixed in a pre-activation vessel and preequilibrated if desired, or they can be added separately to the reaction vessel.

Examples of activators for use in the invention are, but not limited to, tetrazole, 5-(ethylthio)-1 H-tetrazole, 5-(4-nitro-phenyl)tetrazole, 5-(2-thienyl)-1 H-tetrazole, triazole, pyridinium chloride, and the like. Suitable solvents are acetonitrile, tetrahydrofuran, dichloromethane, and the like. In practice acetonitrile is a commonly used solvent for oligonucleotide synthesis.

The sulfurization agent for use in step d) is an acyl disulfide dissolved in a solvent. Art know acyl disulfides are e.g. dibenzoyl disulphide, bis(phenylacetyl) disulfide (PADS), bis(4-methoxybenzoyl) disulphide, bis(4-methylbenzoyl) disulphide, bis(4-nitrobenzoyl) disulphide and bis(4-chlorobenzoyl) disulfide.

Phenylacetyl disulfide (PADS) is a commonly used agent for sulfurization reactions that it is best ‘aged’ in a basic solution to obtain optimal sulfurization activity (Scotson J.L. et al., Org. Biomol. Chem., vol. 14, 10840 – 10847, 2016). A suitable solvent for PADS is e.g. a mixture of a basic solvent such as e.g. 3-picoline or 2,6-lutidine with a co-solvent such as acetonitrile, toluene, 1-methyl-pyrrolidinone or tetrahydrofuran. The amount of the basic solvent to the amount of the co-solvent can be any ratio including a 1 :1 ratio. Depending upon the phosphite ester to be converted into its corresponding thiophospate, both ‘fresh’ and ‘aged’ PADS can be used however ‘aged’ PADS has been shown to improve the rate and efficiency of sulfurization. ‘Aged’ PADS solutions are freshly prepared PADS solutions that were maintained some time before usage in the sulfurization reaction. Aging times can vary from a few hours to 48 hours and the skilled person can determine the optimal aging time by analysing the sulfurization reaction for yield and purity.

For the preparation of imetelstat in accordance with the present invention, a PADS solution in a mixture of acetonitrile and 2,6-lutidine, preferably in a 1 :1 ratio, with an aging time of 4 to 14 hours is used. It has been found that when 2,6-lutidine is used, limiting the amount of 2,3,5-collidine (which is often found as an impurity in 2,6-lutidine) below 0.1 % improves the efficiency of sulfurization and less undesirable phosphor oxidation is observed.

In step g) imetelstat is deprotected and cleaved from the solid-phase support. Deprotection includes the removal of the β-cyanoethyl groups and the base-labile protecting groups on the nucleotide bases. This can be done by treatment with a basic solution such as a diethylamine (DEA) solution in acetonitrile, followed by treatment with aqueous ammonia dissolved in an alcohol such as ethanol.

The reaction steps a) to f) of the present invention are carried out in the temperature range of 10°C to 40°C. More preferably, these reactions are carried out at a controlled temperature ranging from 15°C to 30°C. In particular reaction step b) of the present invention is carried out in the temperature range of 15°C to 30°C; more in particular 17°C to 27°C. In particular reaction step d) of the present invention is carried out in the temperature range of 17°C to 25°C; more in particular 18°C to 22°C; even more in particular 19°C. The step g) wherein imetelstat is deprotected and cleaved from the solid-phase support is carried out at a temperature ranging from 30°C to 60°C. Depending upon the equipment and the specific reaction conditions used, the optimal reaction temperature for each step a) to g) within the above stated ranges can be determined by the skilled person.

After each step in the elongation cycle, the solid-phase support is rinsed with a solvent, for instance acetonitrile, in preparation for the next reaction.

After step g), crude imetelstat is obtained in its ammonium salt form which is then purified by a preparative reversed phase high performance liquid chromatography (RP-HPLC) by using either polymeric or silica based resins to get purified imetelstat in triethyl amine form. An excess of a sodium salt is added, and then the solution is desalted by diafiltration thereby yielding imetelstat sodium which is then lyophilized to remove water.

Experimental part

‘Room temperature’ or ‘ambient temperature’ typically is between 21-25 °C.

Experiment 1 (no capping step)

All the reagents and starting material solutions were prepared including 3% dichloroacetic acid (DCA) in toluene, 0.5 M 5-(ethylthio)-1 H-tetrazole in acetonitrile, 0.15 M of all 4 nucleotide monomers of formula (B n) in acetonitrile, 0.2 M phenyl acetyl disulfide (PADS) in a 1 :1 mixture of acetonitrile and 2,6-lutidine and 20% DEA (diethylamine) in acetonitrile.

The oligonucleotide synthesis was performed in the direction of 5′ to 3′ utilizing a repetitive synthesis cycle consisting of detritylation followed by coupling, and sulfurization performed at ambient temperature.

A column (diameter : 3.5 cm) was packed with a solid-support loaded with 3-palmitoylamido-1-0- (4, 4′-dimethoxytrityl)-2-0-succinyl propanediol (3.5 mmol based on a capacity of 400 μιηοΙ/g) that was coupled with the nucleotide monomer B Detritylation was achieved using 3% dichloroacetic acid (DCA) in toluene (amount is between 6.5 and 13.4 column volumes in each detritylation step) and the solid-support bound nucleotide was washed with acetonitrile (amount: 5 column volumes). Coupling with the next nucleotide monomer of formula (B n) was achieved by pumping a solution of 0.5 M 5-(ethylthio)-1 H-tetrazole in acetonitrile and 0.15 M of the next nucleotide monomer of formula (B n) in the sequence, dissolved in acetonitrile, through the column. The column was washed with acetonitrile (amount : 2 column volumes). Then sulfurization was performed by

pumping a solution of 0.2 M phenyl acetyl disulfide (PADS) in a 1 :1 mixture of acetonitrile and 2,6-lutidine mixture through the column followed by washing the column with acetonitrile (amount : 5 column volumes).

The synthesis cycle of detritylation, coupling with the next nucleotide monomer of formula (B n) and sulfurization was repeated 12 times, followed by detritylation using 3% dichloroacetic acid (DCA) in toluene (amount is between 6.5 and 13.4 column volumes).

Upon completion of the synthesis cycle, the crude oligonucleotide on the solid-support support was treated with a diethylamine (DEA) solution followed by treatment with ammonium hydroxide solution: ethanol (3: 1 volume ratio) at a temperature of 55°C. The reaction mixture was aged for

4 to 24 hours at 55°C, cooled to room temperature, and slurry was filtered to remove the polymeric support. The solution comprising imetelstat in its ammonium form was subjected to the HPLC analysis procedure of Experiment 3.

Experiment 2 (with capping step)

All the reagents and starting material solutions were prepared including 3% dichloroacetic acid (DCA) in toluene, 0.5 M 5-(ethylthio)-1 H-tetrazole in acetonitrile, 0.15 M of all 4 nucleotide monomers of formula (B n) in acetonitrile, 0.2 M phenyl acetyl disulfide (PADS) in a 1 :1 mixture of acetonitrile and 2,6-lutidine mixture, 20% N-methylimidazole (NMI) in acetonitrile as capping agent A, isobutryic anhydride in a 1 :1 mixture of acetonitrile and 2,6-lutidine mixture as capping agent B and 20% DEA in acetonitrile.

The oligonucleotide synthesis was performed in the direction of 5′ to 3′ utilizing a repetitive synthesis cycle consisting of detritylation followed by coupling, and sulfurization performed at ambient temperature.

A column (diameter : 3.5 cm) was packed with a solid-support loaded with 3-palmitoylamido-1-0-(4, 4′-dimethoxytrityl)-2-0-succinyl propanediol (3.5 mmol based on a capacity of 400 μιηοΙ/g) that was coupled with the nucleotide monomer B Detritylation was achieved using 3% dichloroacetic acid (DCA) in toluene (amount is between 6.5 and 13.4 column volumes in each detritylation step) and the solid-support bound nucleotide was washed with acetonitrile (amount : 5 column volumes). Coupling with the next nucleotide monomer of formula (B n) was achieved by pumping a solution of 0.5 M 5-(ethylthio)-1 H-tetrazole in acetonitrile and 0.15 M of the next nucleotide monomer of formula (B n) in the sequence, dissolved in acetonitrile, through the column. The column was washed with acetonitrile (amount : 2 column volumes). Then sulfurization was performed by pumping a solution of 0.2 M phenyl acetyl disulfide (PADS) in a 1 :1 mixture of acetonitrile and 2,6-lutidine mixture through the column followed by washing the column with acetonitrile (amount :

5 column volumes).

The sulfurization was followed by a capping step. Each capping in a given cycle used 37-47 equivalents (eq.) of the capping agent NMI, and 9-1 1 equivalents of the capping agent B isobutryic anhydride (IBA), and 1 .4-1.8 equivalents of 2,6 lutidine. Capping agents A and B were pumped through the column with separate pumps at different ratios such as 50:50, 35:65, 65:35.

The synthesis cycle of detritylation, coupling with the next nucleotide monomer of formula (B n) and sulfurization, and capping step was repeated 12 times, followed by detritylation using 3% dichloroacetic acid (DCA) in toluene (amount is between 6.5 and 13.4 column volumes).

Upon completion of the synthesis cycle, the crude oligonucleotide on the solid-support support was treated with a diethylamine (DEA) solution followed by treatment with ammonium hydroxide solution: ethanol (3: 1 volume ratio) at a temperature of 55°C. The reaction mixture was aged for 4 to 24 hours at 55°C, cooled to room temperature, and slurry was filtered to remove the polymeric support. The solution comprising imetelstat in its ammonium form was subjected to the HPLC analysis procedure of Experiment 3.

Experiment 3 : comparision of no-capping vs. capping

Imetelstat obtained in Experiment 1 and Experiment 2 was analysed by HPLC. The amount of the desired full length oligonucleotide having 13 nucleotides was determined and listed in the Table below for Experiment 1 and Experiment 2. Also, the total amount of shortmer, specifically the 12mer, was determined and listed in the Table below for Experiment 1 and Experiment 2.

HPLC analysis method :

column type: Kromasil C18, 3.5 μιτι particle size, 4.6 X 150 mm

eluent:

A: 14.4 mM TEA/386 mM HFIP (hexafluoroisopropanol) /100 ppm(w/v) Na2EDTA in water B: 50% MeOH, 50% EtOH containing 5% IPA

Gradient :

Step Run time (minutes) %B

1 0 10

2 5 10

3 12 26 (linear)

4 35 45 (linear)

5 40 50 (linear)

6 42 50

7 44 10 (linear)

8 50 10

Table : capping vs. no-capping experiments (Experiment 1 was run twice and results are listed as Experiment 1a and 1 b).

The HPLC analysis of Experiment 1 and Experiment 2 demonstrates that yield and purity are comparable for the no-capping experiment vs. the capping experiment.

Main peak % includes Full length oligonucleotide + PO impurities + depurinated impurities.

PO impurities are impurities including one or more oxophosphoramidate internucleoside linkages instead of thiophosphoramidate internucleoside linkages.

Solvent use and reaction time

0.45 L of acetonitrile/mmol is used to prepare capping agent A and capping agent B reagents which corresponds to approximately 25 % of the overall acetonitrile use during the preparation of the reagents. Since each chemical reaction step is followed by a solvent wash, after each capping step too, a solvent wash takes place which is equivalent to about 40 column volumes of the solvent. Considering that about 212 column volumes of the solvent wash is done for a given synthesis run, about 19 % of the wash solvent is used for the capping steps. Each capping step takes between 3 – 6 minutes. This corresponds to about 8 % of the overall synthesis time including the 13 cycles and DEA treatment.

Experiment 4 (detritylation temperature)

The detritylation temperature has an impact in terms of controlling n-1 and depurinated impurities. The temperature of the deblocking solution at the entrance of the synthesizer was chosen between 17.5 and 27 °C (at 3.5 mmol scale) and the selected temperature was kept the same for all detritylation steps. The acetonitrile washing was also kept at the same temperature of the deblocking solution. The % depurinated impurities increased linearly with temperature while n-1 was higher at lower temperatures.

Temperature n-1 % Depurinated Impurity %

17.5 10.7 5.3

19 7.6 6.4

22 5.4 8.7

25 6.1 10.8

27 5.3 12.3

Experiment 5 (sulfurization step temperature)

In the experiments below, the temperature (RT means room temperature) of the PADS solution used in the sulfurization reactions was tested for the % of less favourable PO impurities (these are impurities where phosphor oxidation occurred instead of sulfurization). Lower temperature results in lower PO %.

SEQ ID NO:1 – imetelstat and imetelstat sodium

5′-R-TAGGGTTAGACAA-NH2-3′

wherein R represents palmitoyl [(CH2)1 CH3] amide is conjugated through an aminoglycerol linker to the 5′-thiophosphate group of an N3′ – P5′ thiophosphoramidate (NPS) -linked oligonucleotide.

///////////IMETELSTAT,  GRN163L, PHASE 3, orphan drug, FAST TRACK

CCCCCCCCCCCCCCCC(=O)NCC(COP(=S)([O-])OCC1C(CC(O1)N2C=C(C(=O)NC2=O)C)NP(=S)([O-])OCC3C(CC(O3)N4C=NC5=C4N=CN=C5N)NP(=S)([O-])OCC6C(CC(O6)N7C=NC8=C7N=C(NC8=O)N)NP(=S)([O-])OCC9C(CC(O9)N1C=NC2=C1N=C(NC2=O)N)NP(=S)([O-])OCC1C(CC(O1)N1C=NC2=C1N=C(NC2=O)N)NP(=S)([O-])OCC1C(CC(O1)N1C=C(C(=O)NC1=O)C)NP(=S)([O-])OCC1C(CC(O1)N1C=C(C(=O)NC1=O)C)NP(=S)([O-])OCC1C(CC(O1)N1C=NC2=C1N=CN=C2N)NP(=S)([O-])OCC1C(CC(O1)N1C=NC2=C1N=C(NC2=O)N)NP(=S)([O-])OCC1C(CC(O1)N1C=NC2=C1N=CN=C2N)NP(=S)([O-])OCC1C(CC(O1)N1C=CC(=NC1=O)N)NP(=S)([O-])OCC1C(CC(O1)N1C=NC2=C1N=CN=C2N)NP(=O)(OCC1C(CC(O1)N1C=NC2=C1N=CN=C2N)N)[S-])O.[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+].[Na+]

Selexipag, セレキシパグ ,селексипаг , سيليكسيباق ,


Selexipag.svg

ChemSpider 2D Image | Selexipag | C26H32N4O4S

Selexipag

  • Molecular FormulaC26H32N4O4S
  • Average mass496.622 Da

SelexipagUptravi

475086-01-2 CAS

(C26H32N4O4S, Mr = 496.6 g/mol)

A prostacyclin receptor (PGI2) agonist used to treat pulmonary arterial hypertension (PAH).

NIPPON SHINYAKU….INNOVATOR

セレキシパグ

UNII-5EXC0E384L
селексипаг [Russian] [INN]
سيليكسيباق [Amharic] [INN]
2-{4-[(5,6-diphenylpyrazin-2-yl)(propan-2-yl)amino]butoxy}-N-(methylsulfonyl)acetamide
475086-01-2 [RN]
5EXC0E384L
9231
Acetamide, 2-[4-[(5,6-diphenyl-2-pyrazinyl)(1-methylethyl)amino]butoxy]-N-(methylsulfonyl)-

Selexipag (brand name Uptravi) is a drug developed by Actelion for the treatment of pulmonary arterial hypertension (PAH). Selexipag and its active metaboliteACT-333679 (MRE-269) (the free carboxylic acid), are agonists of the prostacyclin receptor, which leads to vasodilation in the pulmonary circulation.[1]

FDA approves new orphan drug to treat pulmonary arterial hypertension

12/22/2015
On December 21, the U.S. Food and Drug Administration approved Uptravi (selexipag) tablets to treat adults with pulmonary arterial hypertension (PAH), a chronic, progressive, and debilitating rare lung disease that can lead to death or the need for transplantation.

December 22, 2015

On December 21, the U.S. Food and Drug Administration approved Uptravi (selexipag) tablets to treat adults with pulmonary arterial hypertension (PAH), a chronic, progressive, and debilitating rare lung disease that can lead to death or the need for transplantation.

“Uptravi offers an additional treatment option for patients with pulmonary arterial hypertension,” said Ellis Unger, M.D., director of the Office of Drug Evaluation I in the FDA’s Center for Drug Evaluation and Research. “The FDA supports continued efforts to provide new treatment options for rare diseases.”

PAH is high blood pressure that occurs in the arteries that connect the heart to the lungs. It causes the right side of the heart to work harder than normal, which can lead to limitations on exercise ability and shortness of breath, among other more serious complications.

Uptravi belongs to a class of drugs called oral IP prostacyclin receptor agonists. The drug acts by relaxing muscles in the walls of blood vessels to dilate (open) blood vessels and decrease the elevated pressure in the vessels supplying blood to the lungs.

Uptravi’s safety and efficacy were established in a long-term clinical trial of 1,156 participants with PAH. Uptravi was shown to be effective in reducing hospitalization for PAH and reducing the risks of disease progression compared to placebo. Participants were exposed to Uptravi in this trial for a median duration of 1.4 years.

Common side effects observed in those treated with Uptravi in the trial include headache, diarrhea, jaw pain, nausea, muscle pain (myalgia), vomiting, pain in an extremity, and flushing.

Uptravi was granted orphan drug designation. Orphan drug designation provides incentives such as tax credits, user fee waivers, and eligibility for exclusivity to assist and encourage the development of drugs for rare diseases.

Uptravi is marketed by San Francisco-based Actelion Pharmaceuticals US, Inc.

The US FDA granted it Orphan Drug status[2] (for PAH). It was approved by the U.S. FDA on 22 December 2015.[2]

In 2016, the EMA granted marketing authorization in the E.U. for this indication and launch took place shortly after in Germany and the United Kingdom. In Japan, Nippon Shinyaku received approval for the treatment of PAH in 2016.

Selexipag was approved by the U.S. Food and Drug Administration (FDA) on Dec 21, 2015, approved by European Medicine Agency (EMA) on May 12, 2016. It was originally developed by Nippon Shinyaku and then it was licensed to Actelion for co-development. It is marketed as Uptravi® by Actelion in US and EU.

Selexipag is a prostacyclin receptor (PGI2) agonist, which leads to vasodilation in the pulmonary circulation. It is indicated for the treatment of pulmonary arterial hypertension (PAH).

Uptravi® is available as tablets for oral use, containing 200, 400, 600, 800, 1000, 1200, 1400, or 1600 mcg of selexipag. The initial dose is 200 mcg twice daily, and increase the dose by 200 mcg twice daily at weekly intervals to the highest tolerated dose up to 1600 mcg twice daily.

ACT-333679 or MRE-269, the active metabolite of selexipag

SYNTHESIS DEPICT

PATENT

US2012/101276

http://www.google.st/patents/US20120101276?hl=pt-PT&cl=en

The present invention relates to a crystal of 2-{4-[N-(5,6-diphenylpyrazin-2-yl)-N-isopropylamino]butyloxy}-N-(methylsulfonyl)acetamide (hereinafter referred to as “compound A”).

Figure US20120101276A1-20120426-C00001

BACKGROUND OF THE INVENTION

Compound A has an excellent PGI2 agonistic effect and shows a platelet aggregation inhibitory effect, a vasodilative effect, a bronchodilative effect, a lipid deposition inhibitory effect, a leukocyte activation inhibitory effect, etc. (see, for example, in WO 2002/088084 (“WO ‘084”)).

Specifically, compound A is useful as preventive or therapeutic agents for transient ischemic attack (TIA), diabetic neuropathy, diabetic gangrene, peripheral circulatory disturbance (e.g., chronic arterial occlusion, intermittent claudication, peripheral embolism, vibration syndrome, Raynaud’s disease), connective tissue disease (e.g., systemic lupus erythematosus, scleroderma, mixed connective tissue disease, vasculitic syndrome), reocclusion/restenosis after percutaneous transluminal coronary angioplasty (PTCA), arteriosclerosis, thrombosis (e.g., acute-phase cerebral thrombosis, pulmonary embolism), hypertension, pulmonary hypertension, ischemic disorder (e.g., cerebral infarction, myocardial infarction), angina (e.g., stable angina, unstable angina), glomerulonephritis, diabetic nephropathy, chronic renal failure, allergy, bronchial asthma, ulcer, pressure ulcer (bedsore), restenosis after coronary intervention such as atherectomy and stent implantation, thrombocytopenia by dialysis, the diseases in which fibrosis of organs or tissues is involved [e.g., Renal diseases (e.g., tuburointerstitial nephritis), respiratory diseases (e.g., interstitial pneumonia (pulmonary fibrosis), chronic obstructive pulmonary disease), digestive diseases (e.g., hepatocirrhosis, viral hepatitis, chronic pancreatitis and scirrhous stomachic cancer), cardiovascular diseases (e.g, myocardial fibrosis), bone and articular diseases (e.g, bone marrow fibrosis and rheumatoid arthritis), skin diseases (e.g, cicatrix after operation, scalded cicatrix, keloid, and hypertrophic cicatrix), obstetric diseases (e.g., hysteromyoma), urinary diseases (e.g., prostatic hypertrophy), other diseases (e.g., Alzheimer’s disease, sclerosing peritonitis; type I diabetes and organ adhesion after operation)], erectile dysfunction (e.g., diabetic erectile dysfunction, psychogenic erectile dysfunction, psychotic erectile dysfunction, erectile dysfunction associated with chronic renal failure, erectile dysfunction after intrapelvic operation for removing prostata, and vascular erectile dysfunction associated with aging and arteriosclerosis), inflammatory bowel disease (e.g., ulcerative colitis, Crohn’s disease, intestinal tuberculosis, ischemic colitis and intestinal ulcer associated with Behcet disease), gastritis, gastric ulcer, ischemic ophthalmopathy (e.g., retinal artery occlusion, retinal vein occlusion, ischemic optic neuropathy), sudden hearing loss, avascular necrosis of bone, intestinal damage caused by administration of a non-steroidal anti-inflammatory agent (e.g., diclofenac, meloxicam, oxaprozin, nabumetone, indomethacin, ibuprofen, ketoprofen, naproxen, celecoxib) (there is no particular limitation for the intestinal damage so far as it is damage appearing in duodenum, small intestine and large intestine and examples thereof include mucosal damage such as erosion and ulcer generated in duodenum, small intestine and large intestine), and symptoms associated with lumbar spinal canal stenosis (e.g., paralysis, dullness in sensory perception, pain, numbness, lowering in walking ability, etc. associated with cervical spinal canal stenosis, thoracic spinal canal stenosis, lumbar spinal canal stenosis, diffuse spinal canal stenosis or sacral stenosis) etc. (see, for example, in WO ‘084, WO 2009/157396, WO 2009/107736, WO 2009/154246, WO 2009/157397, and WO 2009/157398).

In addition, compound A is useful as an accelerating agent for angiogenic therapy such as gene therapy or autologous bone marrow transplantation, an accelerating agent for angiogenesis in restoration of peripheral artery or angiogenic therapy, etc. (see, for example, in WO ‘084).

Production of Compound A

Compound A can be produced, for example, according to the method described in WO ‘084, and, it can also be produced according to the production method mentioned below.

Figure US20120101276A1-20120426-C00002

Step 1:

6-Iodo-2,3-diphenylpyrazine can be produced from 6-chloro-2,3-diphenylpyrazine by reacting it with sodium iodide. The reaction is carried out in the presence of an acid in an organic solvent (e.g., ethyl acetate, acetonitrile, acetone, methyl ethyl ketone, or their mixed solvent). The acid to be used is, for example, acetic acid, sulfuric acid, or their mixed acid. The amount of sodium iodide to be used is generally within a range of from 1 to 10 molar ratio relative to 6-chloro-2,3-diphenylpyrazine, preferably within a range of from 2 to 3 molar ratio. The reaction temperature varies depending on the kinds of the solvent and the acid to be used, but may be generally within a range of from 60° C. to 90° C. The reaction time varies depending on the kinds of the solvent and the acid to be used and on the reaction temperature, but may be generally within a range of from 9 hours to 15 hours.

Step 2:

5,6-Diphenyl-2-[(4-hydroxybutyl(isopropyl)amino]pyrazine can be produced from 6-iodo-2,3-diphenylpyrazine by reacting it with 4-hydroxybutyl(isopropyl)amine. The reaction is carried out in the presence of a base in an organic solvent (e.g., sulfolane, N-methylpyrrolidone, N,N-dimethylimidazolidinone, dimethyl sulfoxide or their mixed solvent). The base to be used is, for example, sodium hydrogencarbonate, potassium hydrogencarbonate, potassium carbonate, sodium carbonate or their mixed base. The amount of 4-hydroxybutyl(isopropyl)amine to be used may be generally within a range of from 1.5 to 5.0 molar ratio relative to 6-iodo-2,3-diphenylpyrazine, preferably within a range of from 2 to 3 molar ratio. The reaction temperature varies depending on the kinds of the solvent and the base to be used, but may be generally within a range of from 170° C. to 200° C. The reaction time varies depending on the kinds of the solvent and the base to be used and on the reaction temperature, but may be generally within a range of from 5 hours to 9 hours.

Step 3:

Compound A can be produced from 5,6-diphenyl-2-[4-hydroxybutyl(isopropyl)amino]pyrazine by reacting it with N-(2-chloroacetyl)methanesulfonamide. The reaction is carried out in the presence of a base in a solvent (N-methylpyrrolidone, 2-methyl-2-propanol or their mixed solvent). The base to be used is, for example, potassium t-butoxide, sodium t-butoxide or their mixed base. The amount of N-(2-chloroacetyl)methanesulfonamide to be used may be generally within a range of from 2 to 4 molar ratio relative to 5,6-diphenyl-2-[4-hydroxybutyl(isopropyl)amino]pyrazine, preferably within a range of from 2 to 3 molar ratio. The reaction temperature varies depending on the kinds of the solvent and the base to be used, but may be generally within a range of from −20° C. to 20° C. The reaction time varies depending on the kinds of the solvent and the base to be used and on the reaction temperature, but may be generally within a range of from 0.5 hours to 2 hours.

The compounds to be used as the starting materials in the above-mentioned production method for compound A are known compounds, or can be produced by known methods.

PATENT

WO 2002088084

and

http://www.google.fm/patents/WO2009157398A1?cl=en

PAPER

Bioorganic and Medicinal Chemistry, 2007 ,  vol. 15,   21  p. 6692 – 6704

compd 31

PAPER

Bioorganic and Medicinal Chemistry, 2007 ,  vol. 15,   24  p. 7720 – 7725

Full-size image (5 K)2a is the drug

N-Acylsulfonamide and N-acylsulfonylurea derivatives of the carboxylic acid prostacyclin receptor agonist 1 were synthesized and their potential as prodrug forms of the carboxylic acid was evaluated in vitro and in vivo. These compounds were converted to the active compound 1 by hepatic microsomes from rats, dogs, monkeys, and humans, and some of the compounds were shown to yield sustained plasma concentrations of 1 when they were orally administered to monkeys. These types of analogues, including NS-304 (2a), are potentially useful prodrugs of 1.

http://www.sciencedirect.com/science/article/pii/S0968089607007614

PATENT

WO 2011024874

A. Preparation of
Compound A Compound A can be produced , for example, by the method described in Patent Document 1, but can also be produced by the production method described below.
[
Step 2]
6-iodo-2,3-diphenylpyrazine can be produced by reacting 6-chloro-2,3-diphenylpyrazine with sodium iodide. This reaction is carried out in an organic solvent (for example, ethyl acetate, acetonitrile, acetone, methyl ethyl ketone, or a mixed solvent thereof) in the presence of an acid. As the acid to be used, for example, acetic acid, sulfuric acid, or a mixed acid thereof can be mentioned. The amount of sodium iodide used is, for example, suitably in the range of 1 mole to 10 moles, preferably in the range of 2 time moles to 3 times the amount of 1 mole of 6-chloro-2,3-diphenylpyrazine . The reaction temperature varies depending on the raw materials used and the type of acid, but is usually carried out within the range of 60 ° C. to 90 ° C. The reaction time varies depending on the starting materials used, the type of acid and the reaction temperature, but it is usually within the range of 9 hours to 15 hours.Step 2
5,6-diphenyl-2- [4-hydroxybutyl (isopropyl) amino] pyrazine can be prepared by reacting 6-iodo-2,3-diphenylpyrazine with 4-hydroxybutyl (isopropyl) amine. This reaction is carried out in an organic solvent (for example, sulfolane, N-methylpyrrolidone, N, N-dimethylimidazolidinone, dimethylsulfoxide or a mixed solvent thereof) in the presence of a base. Examples of the base used include sodium hydrogencarbonate, potassium hydrogen carbonate, potassium carbonate, sodium carbonate, and mixed bases thereof. The amount of 4-hydroxybutyl (isopropyl) amine to be used is, for example, suitably in the range of 1.5 mol to 5.0 mol per 1 mol of 6-iodo-2,3-diphenylpyrazine, It is within the range of 2 mol to 3 mol. The reaction temperature varies depending on the type of raw material and base used, but is usually carried out within the range of 170 ° C. to 200 ° C. The reaction time varies depending on the type of raw materials and base used and the reaction temperature, but it is usually within the range of 5 hours to 9 hours.Step 3
Compound A can be prepared by reacting 5,6-diphenyl-2- [4-hydroxybutyl (isopropyl) amino] pyrazine with N- (2-chloroacetyl) -methanesulfonamide. This reaction is carried out in an organic solvent (N-methylpyrrolidone, 2-methyl-2-propanol or a mixed solvent thereof) in the presence of a base. Examples of the base to be used include potassium t-butoxide, sodium t-butoxide or mixed bases thereof. The amount of N- (2-chloroacetyl) -methanesulfonamide used is, for example, 2 to 4 mol per 1 mol of 5,6-diphenyl-2- [4-hydroxybutyl (isopropyl) amino] It is suitable within the range, and preferably within the range of 2 mol to 3 mol. The reaction temperature varies depending on the type of raw material and base used, but is usually carried out within the range of -20 ° C. to 20 ° C. The reaction time varies depending on the kinds of raw materials and bases used and the reaction temperature, but it is usually within the range of 0.5 hour to 2 hours.Each compound used as a raw material in the above-mentioned production method of compound A is a known compound or can be produced according to a known method.

[0016]
B. Preparation of salt of the present invention The salt of the
present invention can be obtained, for example, by the following method.
The salt of the present invention can be prepared by dissolving the compound A in an appropriate solvent (for example, an ether solvent (for example, dimethoxyethane, tetrahydrofuran), an ester solvent (for example, isopropyl acetate), an aromatic hydrocarbon (for example, toluene), acetonitrile After dissolving and adding a desired base, if necessary, the mixed solution is left to stand at room temperature or under cooling in the state of concentrating or stirring or leaving it stationary. The precipitate formed is collected by filtration , Followed by washing with an appropriate solvent to obtain the desired salt of the present invention. When cooling, not only cooling but also gradual cooling or rapid cooling may be effective in obtaining good crystals. It is also effective to obtain good crystals by adding an ether solvent (for example, t-butyl methyl ether), an ester solvent (for example, ethyl acetate), and an aromatic hydrocarbon (for example, toluene) There are cases.The amount of the solvent used for dissolving the compound A is suitably in the range of 10 ml to 300 ml with respect to the compound A 1 g, for example.
The amount of the base to be used for preparing the salt of the present invention is suitably in the range of 0.5 mol to 1.2 mol with respect to the mol of the compound A 1.
Further, the salt of the present invention, which is a crystal, can be obtained by, for example, the method described in Examples described later.

Example 1 t- butylamine Form I crystal of the salt
Compound A (40 mg) with 0.5mL dimethoxyethane (hereinafter, referred to as. “DME”) was dissolved in, and t- butylamine (1.1 eq) were added, 25 1 ° C. at 8 it was stirred for hours. Thereafter, the reaction solution was added t- butyl methyl ether (1mL), at -20 ° C. 3 and held hours. It was collected by filtration the precipitated crystals produced, under reduced pressure, and dried, I-form crystals of t- butylamine salt ( 3 to afford 9.9mg). B Powder X-ray diffraction spectrum of type I crystal obtained t- butylamine salt using the apparatus shown in Figure 1.
Melting point: 152.5 ℃
elemental analysis (C 3 0 H 4 3 N 5 O 4 S + 0.0 3 H 2 as O)
calculated value (%) C: 6 3 .1 8 H: 7 . 6 1 N: 12 .2 8 measured value (%) C: 6 2. 8 5 H: 7 . 6 4 N: 12.52 1 H-NMR (DMSO-D 6 ): delta 8 .15 (s, 1H), 7 .55 – 7 . 8 0 (M, 2H), 7 .10- 7 . .45 (M, 10H), 4 7 . 0-4 8 5 (M, 1H), 3 . 6 6 (s, 2H), 3 .4 7 (t, 2H), 3 .45 (t, 2H), 2. 7 3 (s, 3 H), 1.50-1. 7 5 (M, 4H), 1.2 3 (s, 9H), 1.22 (D, 6 H)
Example 2 I-form crystal of the potassium salt
Compound A tetrahydrofuran with (40mg) 12mL (hereinafter, referred to as. “THF”) was dissolved in, 0.1M aqueous potassium hydroxide solution (1.1 eq) was added, 40 ℃ It was heated and stirred in for 15 minutes. After that, it was evaporated under reduced pressure, the solvent. The residue it was added ethyl acetate (200μL). While shaking the mixture heated to 50 ° C. 8 was allowed to cool to 25 ℃ over hours. After repeated two more times this step, at -20 ° C. 3 and held hours. The resulting precipitated crystals were collected by filtration under reduced pressure, and dried to obtain Form I crystal of the potassium salt. B Powder X-ray diffraction spectrum of type I crystal of the obtained potassium salt using the apparatus shown in Fig. 1 H-NMR (DMSO-D 6 ): delta 8 .14 (s, 1H), 7 .1 8 – 7 . 3 8 . (M, 10H), 4 7 . 2-4 8 4 (M, 1H) , 3 . 6 5 (s, 2H), 3 .4 7 (t, 2H), 3 .45 (t, 2H), 2. 7 2 (s, 3 H), 1.55-1. 7 0 ( M, 4H), 1.2 3 (D, 6 H)
Example 3  II-form crystals of the potassium salt
Compound A with (40mg) was dissolved in THF and 12mL, 0.1M aqueous potassium hydroxide solution (1.1 eq) was added and heated with stirring for 15 min at 40 ℃. After that, it was evaporated under reduced pressure, the solvent. The residue it was added ethyl acetate (200μL). While shaking the mixture heated to 50 ° C. 8 was allowed to cool to 25 ℃ over hours. This operation was repeated two more times, at -20 ° C. 3 and held hours. It was collected by filtration the precipitated crystals produced, under reduced pressure, after drying, 40 ℃, relative humidity 7 while 5% of thermo-hygrostat 7 left for days to give crystalline Form II of the potassium salt. B Powder X-ray diffraction spectrum of crystalline Form II of the resulting potassium salt using the apparatus Fig 3 is shown in.

Example 4 III type crystal of the potassium salt
Compound A , in addition to (100mg) acetonitrile (1mL), and stirred with heating, Compound A was dissolved, followed by cooling to 20 ℃. To a solution 3 .5M potassium hydroxide / ethanol solution (1.1 eq) was added and stirred for 200 minutes at 20 ℃. While stirring the mixture 7 after a heated stirring for 1 hour to 0 ° C., and then cooled to 10 ℃ over 10 hours. Further heated while the mixture 6 is heated to 0 ℃, t- butyl methyl ether (0. 3 after adding mL), cooled to 20 ℃ over 10 hours. It was collected by filtration the precipitated crystals produced, under reduced pressure, and dried, III type crystal of the potassium salt ( 7 to afford 5mg). The powder X-ray diffraction spectrum of the type III crystal of the obtained potassium salt using R unit is shown in FIG. Furthermore, in differential scanning calorimetry, of about 7 endothermic peak was observed at around 4 ° C..
Elemental analysis (C 2 6 H 3 1 N 4 O 4 . SK + 0 7 8 H 2 as O)
calculated value (%) C: 5 6 .91 H: 5.9 8 N: 10.21
measured value (%) C: 5 6 . 6 1 H: 5.55 N:. 10 3 6

EXAMPLE 5 IV-type crystal of the potassium salt
Compound A , in addition to (50mg) and ethyl acetate (1mL), and stirred with heating, Compound A was dissolved, followed by cooling to 20 ℃. To a solution 3 .5M potassium hydroxide / ethanol solution (2.2 eq) was added and 2 at 20 ° C. 3 and stirred for hours. It was collected by filtration the precipitated crystals produced, under reduced pressure, and dried to obtain Form IV crystal of the potassium salt (41mg). The powder X-ray diffraction spectrum of crystalline Form IV of the resulting potassium salt using R unit is shown in FIG. Furthermore, in differential scanning calorimetry, an endothermic peak was observed at around approximately 91 ℃.

Paper

J Med Chem 2015, 58(18): 7128

PATENT

WO 2018008042

https://patents.google.com/patent/WO2018008042A1/en

The present invention relates to an improved and novel processes for the preparation of 2- {4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy} -N-(methylsulfonyl) acetamide compound of formula- 1 , which is represented by the following structural formula- l .

Figure imgf000003_0001

Formula-

The present invention also relates to novel crystalline forms of the compound of formula- 1 and process for the preparation thereof.

Background of the Invention:

2- {4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy}-N-(methylsulfonyl) acetamide is known as Selexipag. It is developed by Nippon Shinyaku under the brand name of Uptravi®, for the treatment of pulmonary arterial hypertension.

2- {4-[(5,6-diphenylpyTazin-2-yl)(isopropyl)amino]butoxy}-N-(methylsulfonyl) acetamide was firstly described in US7205302B2 herein after referred as US ‘302. The said patent also describes its process for the preparation. According to this process the final product was obtained with low yield and purity.

US8791 122 (herein after referred as US’ 122) patent describes crystalline form-I, II and III of 2- {4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy} -N-(methylsulfonyl) acetamide. Because of drug compounds having, for example, improved stability, solubility, shelf life and in vivo pharmacology, are consistently sought, there is an ongoing need for new or pure salts, hydrates, solvates and polymorphic forms of existing drug molecules. The novel crystalline forms of 2- {4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy} -N- (methylsulfonyl) acetamide described herein help meet this requirement.

US ‘ 122 patent describes amorphous form of the compound of formula- 1 . This patent does not disclose any detailed process for amorphous form and PXRD pattern of amorphous compound of formula- 1 .

Figure imgf000019_0001

Examples:

Example-1 Preparation of 4-((5)6-diphenylpyrazin-2-yl)(isopropyl)amino)butan-l-ol compound of formula-8

A mixture of 5-chloro-2,3-diphenylpyrazine (25 gm) compound of formula-7a and 4- (isopropyl amino)butan- 1 -ol (108 gm) was heated to 190-195°C and stirred the reaction i mixture for 10- 12 hours at same temperature. Cooled the reaction mixture to 25-35°C. To this reaction mixture n-heptane followed by water were added slowly at 25-30°C and stirred the reaction mixture for 2 hours at the same temperature. Filter the precipitated solid, washed with water and dried to get the title compound.

Yield: 30 gm.

Example-2: Preparation of tert-butyl 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino) butoxy)acetate

Potassium hydroxide solution (96.6 gm of potassium hydroxide dissolved in 175 ml of water) was added to the mixture of 4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butan- l -ol (25 gm) and toluene ( 175 ml) at 25-30°C and stirred the reaction mixture for 30 minutes at the same temperature. Cooled the reaction mixture to 0-5°C. Tert-butyl bromoacetate (94 gm) was slowly added to the reaction mixture at 0-5°C and stirred the reaction for 60 minutes at same temperature. Raised the temperature of the reaction mixture to 25-30°C and maintained for 60 minutes. Both the aqueous and organic layers were separated. The aqueous layer was extracted with toluene and combined the organic layers. Organic layer was washed with hydrochloric acid solution followed by with aqueous sodium bicarbonate solution. Organic layer was dried with sodium sulphate and distilled off the solvent completely from the organic layer under reduced pressure to get the title compound.

Yield: 29 gm.

Example-3: Preparation of 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butoxy) acetic acid compound of formula-6

Aqueous sodium hydroxide solution (7.5 gm of sodium hydroxide was dissolved in 80 ml of water) was added to the solution of tert-butyl 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyl) amino)butoxy)acetate (30 gm) in methanol (290 ml) at 30-35°C. Heated the reaction mixture to reflux temperature and stirred for 3 hours at the same temperature. Distilled off solvent completely from the reaction mixture under reduced pressure and cooled the reaction mixture to 25-30°C. Water was added to the obtained compound and acidified the reaction mixture using diluted hydrochloric acid at the same temperature. Extracted the reaction mixture with ethyl acetate. The organic layer was washed with aqueous sodium chloride solution and dried with sodium sulphate. Distilled off the solvent from the organic layer under reduced pressure. Diisopropyl ether (60 ml) was added to the obtained compound at 25-30°C and stirred for 60 minutes at the same temperature. Filtered the precipitated solid, washed with diisopropyl ether and dried to get the title compound.

Yield: 19 gm.

Example-4: Preparation of 2-{4-[(5,6-diphenylpyrazin-2-yl)(isopropy.)amino]butoxy}- N-(methylsulfonyl) acetamide compound of formula-1

Triethylamine (9.6 gm) was added to the mixture of 2-(4-((5,6-diphenylpyrazin-2- yl)(isopropyl)amino)butoxy)acetic acid (10 gm), dichloro methane (100 ml), N,N- dicyclohexylcarbodiimide (4.9 gm), hydroxybenzotriazole (3.5 gm) and methane sulfonamide (3.39 gm) at 25-30°C and stirred the reaction mixture for 12 hours at the same temperature. Filtered the unwanted compounds from the reaction mixture and washed with dichloromethane. The organic layer was washed with water, followed by with aqueous citric acid solution and then washed with aqueous sodium chloride solution. Distilled off the solvent from the organic layer under reduced pressure. To this residue ethyl acetate (20 ml) and carbon (1 gm) were added at 25-30°C and stirred the reaction mixture for 30 minutes at the same temperature. Filtered the reaction mixture through hyflow bed and washed with ethyl acetate. The obtained filtrate was slowly added to the mixture of n-heptane and water at 25-30°C and stirred for 10 hours. Filtered the precipitated solid, washed with n-heptane and dried to get the title compound.

Yield: 4.5 gm.

Example-5: Preparation of 2-{4-f(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy}- N-(methylsulfonyl) acetamide compound of formula-1

Sodium t-butoxide (96.6 gm) was added to the mixture of n-methy pyrrolidinone (125 ml) and 4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butan-l-ol (25 gm) compound of formula-8 at 0-5°C and stirred the reaction for 20 minutes at the same temperature. 2-chloro- N-(methylsulfonyl)acetamide (23.7 gm) was slowly added to the reaction mixture at 0-5°C and raise the temperature of the reaction mixture to 25-30°C. Stirred the reaction mixture for 10-12 hours at 25-30°C and water was added to it at the same temperature. The reaction mixture was extracted with ethyl acetate. The organic layer was washed with aqueous sodium chloride solution and distilled off the solvent from the organic layer under reduced pressure. To this residue ethyl acetate (50 ml) and carbon (2.5 gm) were added at 25-30°C and stirred the reaction mixture for 30 minutes at the same temperature. Filtered the reaction mixture through hyflow bed and washed with ethyl acetate. The obtained filtrate was slowly added to the mixture of n-heptane and water at 25-30°C and stirred for 10 hours. Filtered the precipitated solid, washed with n-heptane and dried to get the title compound.

Yield: 14 gm.

Example-6: Preparation of 2-chIot*o- -(methylsulfonyl)acetamide

A mixture of methane sulfonamide (100 gm) and chloroacetyl chloride (356.4 gm) was heated to reflux temperature and stirred it for 10 hours at the same temperature. Cooled the reaction mixture to – 10 to -5°C and stirred it for 2 hours at the same temperature. Filtered the precipitated, solid, washed with toluene followed by n-heptane and dried to get the title compound.

Yield: 175 gm.

ExampIe-7: Purification of the compound of formula-1

Methanol (20 ml) was added to the compound of formula-1 (2 gm) at 25-30°C and heated to reflux temperature. Dichloromethane (3 ml) was added to the reaction mixture at reflux temperature and stirred for 15 minutes at the same temperature. Filtered the reaction mixture, distilled off the solvent from the filtrate under reduced pressure to get the title compound. Yield: 2 gm

Example-8: Preparation of N-isopropyI-5,6-diphenylpyrazin-2-amine (Formula-4) Isopropyl bromide (5.5 gm) was added to the mixture of 2-amino -5,6-diphenylpyrazine ( 10 gm), potassium tert-butoxide (9 gm) and dimethylformamide (50 ml) at 25-30°C, slowly heated to 80-85°C and stirred the reaction mixture for 6 hours at same temperature. The reaction mixture was cooled to 10- 15°C, diluted the reaction mixture with water and stirred it for 2 hours at the same temperature. Filtered the obtained solid and dried to get the title compound.

Yield: 9.5 gm

ExampIe-9: Preparation of N-isopropyl-5,6-diphenylpyrazin-2-amine (Formula-4)

A mixture of 5-chloro-2,3-diphenylpyrazine ( 10 gm), isopropyl amine (7.5 gm) and potassium carbonate (10.5 gm) and dioxane (50 ml) were heated to 40-45°C and stirred the reaction mixture for 12 hrs at the same temperature. The reaction mixture was cooled to 10- 15°C, diluted with water and extracted with dichloromethane. Combined the organic layers was washed with aqueous sodium hydrochloride solution and dried over anhydrous sodium sulphate. Distilled off the solvent completely from the organic layer under reduced pressure to provide the title compound.

Yield: 9 gm

Example-10: Preparation of 2-(4-chlorobutoxy)aceticacid (Formula-5a)

2-bromoaceticacid (10 gm) was slowly added to a mixture of l-chlorobutan-4-ol (7.2 gm), potassium carbonate (26.5 gm) and acetonitrile (50 ml) at 25-30°C. The reaction mixture was heated to 75-80°C and stirred the reaction mixture for 6 hours at same temperature. The reaction mixture was cooled to 25-30°C and diluted with , water. Acidified the reaction mixture using diluted hydrochloric acid at 25-30°C. The reaction mixture extracted with dichloromethane. Combined the organic layers was dried over anhydrous sodium sulphate and distilled off the solvent under reduced pressure to provide the title compound.

Yield: 10.5 gm.

Example-11: Preparation of 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino) butoxy)acetic acid (formula-6)

A mixture of N-isopropyl-5,6-diphenylpyrazin-2-amine (8 gm), potassium carbonate (7.5 gm) and acetonitrile (40 ml) was stirred for 1 hr at 25-30°C. A solution of 2-(4-chlorobutoxy) aceticacid (5.4 gm) in acetonitrile (15 ml) was slowly added to the reaction mixture at 25- 30°C. Heated the reaction mixture to reflux and stirred for 12 hours at the same temperature. The reaction mixture was cooled to 10-15°C and diluted with wateT. Acidified the reaction mixture using diluted hydrochloric acid and extracted the reaction mixture using ethyl acetate. Combined the organic layers and dried over sodium sulphate. Distilled off the solvent completely from the organic layer to get the title compound.

Yield: 8.5 gm

Example-12: Preparation of 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyt)amino)butoxy)- N-(methylsulfonyl)acetamide (formula-1)

A mixture of 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butoxy)acetic acid (5 gm), HATU (5.4 gm), triethylamine (1.5 gm) and dimethylformamide (20 ml) was stirred for 1 hr at 5-10°C under nitrogen atmosphere. Methane sulfonamide (5.2 gm) was slowly added to the reaction mixture at 5-10°C and stirred for 12 hrs at the same temperature. The reaction mixture was diluted with water and stirred for 2 hrs. The precipitated solid was filtered and dried to get the title compound.

Yield: 4.5 gm

Example-13: Preparation of 2-(4-((5,6-diphenylpyrazin-2-yl) (isopropyl) amino) butoxy) acetonitrile (Formula-12)

To the mixture of 4-((5,6-diphenylpyrazin-2-yI)(isopropyl)amino)butan-l-ol ( 10 gm), tetrabutyl ammoniumbromide (0.2 gm), potassium carbonate (7.6 gm) and acetone (50 mL), chloroacetonitrile (3.2 gm) was added at 25-30°C. Heated the reaction mixture to reflux temperature and stirred the reaction mixture for 6 hrs at the same temperature. The reaction mixture was cooled to 10- 15°C and filtered the reaction mixture. Distilled off the solvent completely from the filtrate to get the tile compound.

Yield: 9 gm

Example-14: Preparation of 2-(4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butoxy) acetic acid (formula-6)

Sodium hydroxide (3.5 gm) was added to a solution of 2-(4-((5,6-diphenylpyrazin-2-yl) (isopropyl) amino) butoxy) acetonitrile (8 gm) in methanol (60 ml) and water (30 ml). The reaction mixture was heated to 65-70°C and maintained for 6 hrs. The reaction mixture was cooled to 10°C, acidified with diluted hydrochloric acid and stirred at same temperature for 2 hr. The obtained solid was filtered and dried to provide the title compound.

Yield: 7.5 gm

Example-15: Preparation of 2-chloro-N-(methylsulfonyl)acetamide (Formula-16)

The mixture of methane sulfonamide (50 gm) and chloroacetyl chloride (92 gm) was heated to 1 10-1 15°C and stirred the reaction mixture for 7 hours at the same temperature. The reaction mixture was cooled to 25-30°C and dichloromethane was added to the reaction mixture at the same temperature. Cooled the reaction mixture to 15-20°C and stirred for 1 hour at the same temperature. Filtered the precipitated solid and washed with dichloromethane. The obtained solid was recrystallized using dichloromethane to get pure title compound. Yield: 80 gm. M.R.: U0- 1 15°C. Purity by HPLC: 98.85%.

Example-16: Preparation of 4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butan-l-ol (Formula-8)

The mixture of 5-chloro-2,3-diphenylpyrazine ( 100 gm) and 4-(isopropylamino)butan-l -ol (245.5 gm) was heated to 190-195°G and stirred the reaction mixture for 12 hours at the same temperature. The reaction was cooled to 25-30°C and n-heptane was added to the reaction mixture. The reaction mixture was further cooled to 10-15°C, water was slowly added to the reaction mixture and stirred for 2 hours at the same temperature. Filtered the precipitated solid and washed with water. Dichloromethane (300 ml) was added to the obtained solid and stirred for 5 minutes. Both the organic and aqueous layers were separated. The organic layer was dried with sodium sulphate, distilled off the solvent from the organic layer completely under reduced pressure and co-distilled with n-heptane. 400 ml of n-heptane was added to the obtained compound at 25-30°C, heated the reaction mixture to 45-50°C and stirred for 30 minutes at the same temperature. The reaction mixture was cooled to 15-20°C and stirred for 2 hours at the same temperature. Filtered the solid, washed with n-heptane and dried to get the title compound.

Yield: 82 gm. M.R.: 100-105°C. Purity by HPLC: 95.4%.

Example-17: Purification of 4-((5,6-diphenylpyrazin-2-yl)(isopropyl)amino)butan-l-ol (Formula-8)

n-Heptane (750 ml) was slowly added to pre-cooled solution of 4-((5,6-diphenylpyrazin-2- yl)(isopropyl)amino)butan- l -ol (100 gm) in acetone (250 ml) was cooled to 0-5°C. Stirred the reaction mixture for 4 hours at the same tempereature. Filtered the precipitated solid, washed with n-heptane and dried to get the pure title compound.

Yield: 54 gm. Purity by HPLC: 99.92%.

Example-18: Preparation of crystalline form-L of compound of formula-1

Melting the compound of formula-1 (10 gm) at 140-145°C under reduced pressure for 15 minutes. The above obtained oily residue was added to 100 ml of pre-cooled n-heptane at 0- 5°C. Stirred the reaction mixture for 6 hr at 0-5°C. Filtered the precipitated solid, washed with n-heptane and dried to get the title compound. Yield: 9 gm; PXRD of the obtained compound is depicted in figure- 10 and DSC thermogram is depicted in figure- 1 1. Example-19: Preparation of crystalline form-P of compound of formula-1

Melting the compound of formula-1 (10 gm) at 140-145°C under reduced pressure for 15 minutes. The above obtained oily residue was added to 100 ml of pre-cooled n-heptane at 0- 5°C. Stirred the reaction mixture for 36 hours at 0-5°C. Filtered the precipitated solid, washed with n-heptane and dried to get the title compound.

Yield: 9 gm; PXRD of the obtained compound is depicted in figure-7, its IR is depicted in figure-8 and its DSC is depicted in figure-9.

Example-20: Preparation of crystalline form-P of compound of formula-1

Melting the compound of formula-1 (10 gm) at 140-145°C under reduced pressure for 15 minutes. The above obtained oily residue was added to 100 ml of n-heptane at 30-40°C.

Stirred the reaction mixture for 36 hours at 30-40°C. Filtered the precipitated solid, washed with n-heptane and dried to get the title compound.

Yield: 9 gm; PXRD of the obtained compound is similar to the figure-7.

Example-21 : Preparation of amorphous form of compound of formula-1

Melting the compound of formula-1 ( 10 gm) at 140- 145°C under reduced pressure for 15 minutes and the above obtained oily residue was cooled to 0-5°C. Unload the obtained compound and dried to get the title compound. Yield: 9 gm; Purity by HPLC: 99.74%. PXRD of the obtained compound is depicted in figure-5 and IR is depicted in figure-6.

Exaniple-22: Preparation of crystalline form-I of compound of formula-1

Melting the compound of formula-1 (5 gm) at 140-145°C under reduced pressure for 15 minutes. 50 ml of n-heptane was added to the above obtained oily residue at 115-120°C.

Stirred the reaction mixture for 20 minutes at 1 15- 120°C. Cooled the reaction mixture to 25-

30°C and stirred for 60 minutes at the same temperature. Further cooled the reaction mixture to 0-5°C and stirred the reaction mixture for 60 minutes at the same temperature. Filtered the precipitated solid, washed with n-heptane and dried to get the title compound.

Yield: 4 gm; Purity by HPLC: 99.68%.

PATENT

CN 108675964

PATENT

CN 106316967

PATENT

WO 2017029594

PATENT

US8791122

Form-I  II  III

https://patents.google.com/patent/US8791122B2/en

Figure US08791122-20140729-C00002

PATENT

https://patents.google.com/patent/WO2018022704A1/en

Selexipag has the chemical name 2-{4-[(5,6-diphenylpyrazin-2- yl)(isopropyl)amino]butoxy}-N-(methylsulfonyl)acetamide. Selexipag has the following chemical structure:

Figure imgf000002_0001

[0004] Selexipag is being developed by Actelion and Nippon Shinyaku for the treatment of arteriosclerosis obliterans, pulmonary hypertension and Raynaud’s disease secondary to systemic sclerosis.

[0005] Selexipag is disclosed in US 7,205,302. US 8,791,122, US 9,284,280 and US 2014- 0155414 disclose polymorphs of Selexipag, denominated forms I, II and III. WO

2017/040872 discloses form IV and V of Selexipag.

xample 1: Preparation of Selexipag

[00126] A. Route 1

[00127] Crude Selexipag can be obtained by any method known in the art, for example by the method described in US 7,205,302 or according to the following.

[00128] B. Route 2

[00129] Step a: Preparation of 4-((5,6-diphenyl-pyrazin-2-yl)(isopropyl)amino)butan-l-ol

[00130] To 50 g (0.161 mol) of 5-bromo-2,3-diphenylpyrazine, 116 g (0.884 mol, 5.5 eq/mol) of 4-(isopropylamino)-butan-l-ol and 13.33 g of KI (0.080 mol, 0.5 Eq/mol) were added. The reaction mixture was stirred, warmed and then heated up to 140°C for about 18- 20 hrs. The reaction was monitored by TLC up to completion (starting material about 1% by TLC). The reaction mixture was cooled down to room temperature. After the reaction was completed, the following work up step was performed:

[00131] Option 1 : Ethyl acetate was added (500 mL, 10 vol) and the organic phase was washed with water (150 mL, 3 vol). The organic phase was separated and aqueous phase was extracted with ethyl acetate (150 mL, 3 vol). The organic phases were joined and washed with water (200 mL, 2 vol) three times.

[00132] The solvent was distilled off under vacuum at not more than (“NMT”) 40°C until 1 vol (oil appearance).

[00133] Option 2: The material (mixture) was dissolved in acetone (250 mL, 5 vol), the solution obtained was cooled down to 0°C to 5°C and anti-solvent / water was added (1000 mL, 20 vol) for 40 minutes, then the suspension was stirred for about 30 minutes at about 0°C-5°C. The solid material was filtered and washed with water (200 mL, 4 vol). Crude wet product was obtained as yellow solid yielding 101.8 % WY (87 % MY), HPLC purity 90.8% on area at this stage.

[00134] The crude material, obtained in either of the above described options, was purified through crystallization from acetone :«-heptane as follows: to a solution of 4-((5,6-diphenyl- pyrazin-2-yl)(isopropyl)amino)butan-l-ol crude in acetone (175 mL, 3.5 vol) at 0°C – 5°C, hexane (600 mL, 12 vol) dropwise in about 120 min was added, then the precipitated mixture was cooled down to about -10°C and stirred for about 60 min. The product was filtered off and washed with hexane (250 mL, 5 vol) and dried under vacuum at 25°C. Pure product was obtained as yellowish solid yielding overall 77.2%, (66.5% MY), HPLC purity 98.2% on area.

[00135] Step b: Preparation (2-bromo-N-(methylsulfonyl)-acetamide)

[00136] To a suspension of 50 g (0.526 mol) of methanesulfonamide in toluene (625 mL, 12.5 vol) and isopropyl acetate (625 mL, 12.5 vol), 159.1 g (0.789 mol) of bromo-acetyl- bromide (“BAB”) was added under nitrogen atmosphere. The reaction mixture was heated up to about 90°C for about 8 hours under a nitrogen stream. The reaction was monitored by TLC up to completion (starting material about 1% by TLC). The reaction mixture was cooled down to about 40°C and concentrated under vacuum until 10 volumes. Subsequently, toluene was added (250 mL, 5 vol) and distilling off solvents is carried out at NMT 30°C until 10 volumes. Then was added dichloromethane (100 mL, 2 vol) and the mixture was cooled down at 0°C and is stirred for 90 min. The solid was filtered and washed with

dichloromethane (100 mL, 2 vol). Crude product was obtained as beige solid material yielding 187% WY (83% MY), HPLC purity 99.2% at this stage. [00137] The crude material (83 g) was purified through re-slurring with dichloromethane (166 mL, 2 vol; preferably 332 mL, 4 vol) by stirring at about 32°C for about 60 min. The crystallization mixture was cooled down to about 0°C-5°C and stirring for 30 min, filtered off and washed with dichloromethane (100 mL, 2 vol). Subsequently, the material was dried at 35°C for 24 hours. Pure and dried material was obtained as white off solid yielding overall 173%, (77% MY), HPLC purity 99.6 % on area.

[00138] Step c: Preparation of (2-[4-[(5,6-diphenyl-2-pyrazinyl)(l- methylethyl)amino]butoxy]-N-(methylsulfonyl)-acetamide) – Selexipag

[00139] To 10 g (0.028 mol) of 4-((5,6-diphenyl-pyrazin-2-yl)(isopropyl)amino) butan-l-ol was added a strong base (6.0 eq/mol), previously suspended in an appropriate solvent, within a range of from -10°C to 40°C under a nitrogen atmosphere and stirred for 60 min. Then, a solution of 17.9 g (3.0 eq/mol) of 2-bromo-N-(methylsulfonyl)-acetamide, previously dissolved in the same solvent, is added dropwise within a range of from 120 tol 80 min, controlling the exothermic temperature. The reaction was monitored by TLC up to completion. Subsequently, the mixture reaction was cooled down around 5°C and water is added by controlling the exotherm (NMT 15°C). Finally, an acetic acid solution was added and the suspension was stirred for about 60 min at 0°C -5°C. The product (crude) was filtered off and washed with water. An amorphous solid was obtained. The crude product was purified by crystallization from ethanol:THF.

[00140] Step d: Purification of Selexipag

[00141] Crude Selexipag can be purified by crystallization in an organic solvent for example alcohols such as ethanol, iso-amyl alcohol, iso-propyl alcohol, butanol; ethers such as tetrahydrofuran, hydrocarbons such as heptane and mixed solvents thereof.

[00142] C. Route 3

[00143] 33.3 g (0.297 mol, 6.0 eq/mol) of potassium tert-butoxide were dissolved in DMF (2.8 vol) in a flask (500 mL) under nitrogen atmosphere and stirred for 15 min. Then, a solution of 17.9 g (0.049 mol, 1.0 eq/mol) of 4-((5,6-diphenyl-pyrazin-2-yl)(isopropyl) amino) butan-l-ol (SLX-4) dissolved in DMF (1.2 vol) was added in one portion. The reaction mixture was stirred for 60 min within a temperature range from 20°C to 25°C at 150 rpm Then, a solution of 32.1 g (0.15 mol, 3.0 Eq/mol) of 2-bromo-N-(methylsulfonyl)- acetamide (SLX-9), previously dissolved in DMF (1.3 vol), was added dropwise for 120 minutes by controlling the temperature (exothermic process).

[00144] The reaction mixture was quenched with cool water (0.33 vol), transferred into a flask of more capacity (1000 mL) and placed in an ice bath. Cool water (38.32 vol) was added to the reaction mixture and the pH was adjusted to 5.0 with AcOH (0.33 vol). The mixture was stirred at 300 rpm for 40 min. Then, the flask with the reaction mixture was stored in the refrigerator at 8°C. After 8h, the solid was filtered and washed with cool water (5 vol, 2 times). The crude product (yellow solid) was drained (i.e. dried) for 30 min and was stored at 8°C.

Example 2: Preparation of crystalline Selexipag Form IV

[00145] A. Route 1

[00146] 3.0 g of Selexipag was dissolved in dimethylformamide (“DMF”) (12 mL, 4 vol). The obtained solution was added dropwise to a pre-cooled acetic acid solution (0.06 M, 120 mL, from 2°C to 8°C) to obtain a suspension. The suspension was stirred within a range of from 2°C to 8°C for 30 min; then the material was filtered, washed with water (10 mL, 3.3 vol) and drained (i.e. dried) for 10 minutes. The solid material (amorphous) was suspended in heptane (25 mL, 7.5 vol) and the obtained suspension was stirred for 30 minutes at room temperature. The material was filtered, washed with heptane (20 mL, 6.6 vol) and drained (i.e. dried) under vacuum for at least 30 minutes at room temperature to obtain the Form IV Crystal.

[00147] B. Route 2

[00148] Crude Selexipag (1.0 g, amorphous solid, obtained from the synthesis) was dissolved in ethyl acetate (5 vol, 5 mL), then water was added (10 vol, 10 mL) into the solution, the mixture was stirred for about 10 minutes and the pH was adjusted to a range of from 8.0 to 9.0 by titration with K2CO3 solution. The phases were separated; the pH of the aqueous phase was adjusted to a range of from 3.5 to 5.0 by titration with acetic acid. Then, ethyl acetate (10 vol, 10 mL) was added into the aqueous phase, the obtained mixture was stirred and the phases were separated. The organic phase was distilled off under reduced pressure (from 2 to 3 volumes), and a solution was obtained. The obtained concentrated solution was quickly added to a mixture (suspension) of Form IV in ^-heptane (17 mL, 17 vol), over a period of less than 5 minutes, (the suspension temperature was of from 15°C to 25°C), and a suspension was obtained. The obtained suspension was stirred (155rpm) for 90 minutes at a temperature of from 0°C to 5°C. The suspension was filtered, washed with heptane, squeezed for 15 minutes and dried at 25°C, under vacuum, for about 14 hours. The product was analyzed by PXRD – Form IV was obtained.

[00149] The above procedure can be performed by dissolving the crude amorphous starting material in any suitable organic solvent, for example ester solvent. Example 3: Preparation of (2-[4-[(5,6-diphenyl-2-pyrazinyl)(l- methylethyl)amino] butoxy] -N-(methylsulfonyl)-acetamide) – Selexipag

Figure imgf000024_0001

SLX-4 SLX-9 SLX-6

[00150] 9.2 grams (0.082 mol, 5.9 eq/mol) of potassium tert-butoxide were combined with DMF (2.7 vol, 13.5 mL) in a flask (50 mL) under nitrogen atmosphere and a suspension was formed and was stirred for 20 min. Then, 5.0 g (0.014 mol, 1.0 eq/mol) of 4-((5,6-diphenyl- pyrazin-2-yl)(isopropyl)amino)butan-l-ol (SLX-4) as solid powder was added under nitrogen atmosphere. The reaction mixture was stirred for 60 min within a temperature range from 20°C to 25°C and at 170 rpm. Then, a solution of 8.9 g (0.041 mol, 3.0 eq/mol) of 2-bromo- N-(methylsulfonyl)-acetamide (SLX-9), previously dissolved in DMF (1.3 vol, 6.5 mL), was added dropwise for 120 minutes by controlling the temperature (exothermic process). After the end of addition, the reaction was completed, and the reaction mixture was quenched with cold water (0.5 vol, 2.5 mL), subsequently transferred into a flask of more capacity (500 mL) and placed into an ice bath. Cold water (40 vol, 200 mL) was added into the suspension and the pH was adjusted within the range from 4.0 to 5.0 with acetic acid. The obtained mixture was stirred for 120 min. The crude amorphous product was collected by filtration and washed twice with cold water (5 vol, 25 mL). The product was drained (i.e. dried) for 30 min and isolated as a yellow-brown solid which was stored within the range from 2°C to 8°C for approximately 17 hours. Then, the crude amorphous material was dissolved in ethyl acetate (15 vol, 75 mL) and water was added into the solution (30 vol, 150 mL). The pH was adjusted from 8.0 to 9.0 by addition of potassium carbonate solution, the phases were separated and the aqueous phase was washed twice with ethyl acetate (7.5 vol, 37.5 mL). The pH of the final aqueous phase was adjusted to a range from 4.0 to 5.0 with acetic acid. Then, ethyl acetate was added (30 vol, 150 mL) and the phases were separated. The organic phase was washed twice with water (7.5 vol, 37.5 mL). The organic phase was distilled off under reduced pressure (from 6 to 7 volumes, or from 6 to 15 volumes) and a solution was obtained.

[00151] In a different flask (capacity of 250 mL with a PTFE stirrer blade), a suspension of 0.05 g of Selexipag Form IV in ^-heptane (30 volumes, 150 mL) was stirred for 60 minutes within the range 0°C to 5°C and this suspension was added into the above ethyl acetate concentrated solution at room temperature over a period of less than 5 minutes. The final suspension was cooled down to 0°C to 5°C and stirred (220 rpm) for 120 minutes. The solid product was filtered off and washed twice with cold heptane (5 vol, 25 mL). The product was drained (i.e. dried) overnight. The product was analyzed by PXRD – Form VI was obtained, PXRD pattern is depicted in Figure 1.

PATENT

CN105949135

https://patents.google.com/patent/CN105949135A/en

Figure CN105949135AD00052

Example 1

[0027] A) Preparation of [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate:

[0028] 4 – [(tert-butoxycarbonyl) (isopropyl) amino] butanol -1_ (20 (^, 0.09111 〇1) and tert-butyl bromoacetate (21 · lg, 0 llmol) solution. The reaction was stirred for 2 hours to burn dichloromethane (90mL), was added tetrabutylammonium chloride (0.72g, 2.6mmol), potassium hydroxide (7.3g, 0.13mol) and water (12.0g), the reaction mixture was 25 ° C The reaction solution was concentrated under reduced pressure and rotary evaporated to dryness and extracted with ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, a mixed solvent of ethanol and recrystallized from isopropanol to give [4- (tert-butoxycarbonyl) (isopropyl yl) aminobutoxy] acetate, as a pale yellow oil (26.6 g of), a yield of 89.0%, the reaction formula of this step is as follows:

[0029]

Figure CN105949135AD00071

[0030] B) Preparation of [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid:

[0031] [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate (26. (^, 0.075! 11〇1) was dissolved in methanol (50 mL), was added sodium hydroxide solution (NaOH = 3 · 3g, 0 · 08mol; water 9 · 0g), was heated to 80 ° C for 6 hours, cooled to room temperature, after treatment and purification, to give [4- (tert-butoxycarbonyl) (isopropyl ) aminobutoxy] acetic acid (20.7 g of), a yield of 95.0%, the reaction formula of this step is as follows:

Figure CN105949135AD00081

[0033] C) Preparation of 2- [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide:

[0034] [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid (20 (^, 0.07 11〇1) and a hoot “-.! P sitting carbonyldiimidazole (14.0g, 0.09mo 1 ) was dissolved in tetrahydro-thiopyran Misaki (70 mL), with stirring, was added methyl sulfonamide (7.9g, 0.08mol), the reaction mixture was 90 ° C the reaction stirred for 18 hours, the reaction solution was concentrated by rotary evaporation to dryness, extracted with ethyl acetate, over magnesium sulfate, and concentrated by rotary evaporation to dryness, recrystallized from methanol to give 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide as an off-white solid (21.2 g), yield 83.7%, the reaction formula of this step is as follows:

Figure CN105949135AD00082

[0036] D) Preparation of SIPA Seiler: 2- [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide (20 (^, 0.055111〇1. ) and dissolved in methanol (1101 ^), trifluoroacetic acid (6.88,0.06111〇1), 65 ° (: the reaction stirred for 6 hours to complete the reaction, the reaction was added dropwise to a stirred solution of water (200 mL), cooled to 0 ° C crystallization for 3 hours and filtered to give the intermediate compound (2- [4- (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide), and then dissolved in methanol (40 mL), was added 5 – chloro-2,3-diphenyl-pyrazine (16 · 0g, 0 · 06mol), N, N- diisopropylethylamine (15 · 5g, 0 · 12mol), the reaction mixture was stirred reactor 8 100 ° C hours, the reaction was cooled to room temperature, water (40 mL), cooled to -10 ° C crystallization for 3 hours and filtered to give SIPA game music, as a white solid (25.0 g of), a yield of 92.3%, the reaction step formula as follows:

[0037]

Figure CN105949135AD00083

[0038] Example 2

[0039] A) Preparation of [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate:

[0040] 4 – [(tert-butoxycarbonyl) (isopropyl) amino] -1-butanol (23 (^, 0.10111 〇1) and tert-butyl bromoacetate (25 · 2g, 0 · 13mol) was dissolved. burning in 1,2-dichloroethane (110mL), was added tetrabutylammonium bromide (1 · lg, 3 · 5mmol), sodium hydroxide (6.4g, 0.16mol) and water (14.0g), the reaction mixture was 30 ° C The reaction was stirred for 3 hours, the reaction solution was concentrated by rotary evaporation to dryness under reduced pressure and extracted with ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, a mixed solvent of ethanol and recrystallized from isopropanol to give [4- (tert-butoxy butoxycarbonyl) (isopropyl) aminobutoxy] acetate, as a pale yellow oil (30.3 g of), a yield of 88.2%, the reaction of the present step is the same formula as in Example 1;

[0041] B) Preparation of [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid:

[0042] [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate (30. (^, 0.09! 11〇1) was dissolved in ethanol (85 mL), was added potassium hydroxide solution ( 1 (! = 5.78,0.10111〇1 01; 128 water), heated to 75 ° (: 7 hours, cooled to room temperature, after treatment and purification, to give [4- (tert-butoxycarbonyl) (isopropyl ) aminobutoxy] acetic acid, an off-white solid (23.5 g of), a yield of 93.7%, the reaction of the present step is the same formula as in Example 1;

[0043] C) Preparation of 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide:

[0044] [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid (23 (^, 0.08 11〇1) and Chi ^ -! Dicyclohexyl carbodiimide (22. lg, 0. llmol) was dissolved in chloroform (120 mL), with stirring, was added methyl sulfonamide (9.8g, 0. lOmol), the reaction mixture was 80 ° C the reaction stirred for 19 hours, the reaction solution was concentrated by rotary evaporation to dryness, ethyl acetate was added and extracted dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, recrystallized from methanol to give 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide, off-white the solid (24.8 g of), a yield of 85.0%, the reaction of the present step is the same formula as in Example 1;

[0045] D) Preparation of SIPA Seiler: 2- [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide (24 (^, 0.065111〇1. ) and dissolved in ethanol (1601 ^), trifluoroacetic acid (9 (^, 0.08111〇1.), 70 ° (: the reaction was stirred for 7 hours to complete the reaction, the reaction was added dropwise to a stirred solution of water (260 mL of), cooled crystallization to 0 ° C for 3 hours and filtered to give the intermediate compound (2- [4- (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide), and then dissolved in ethanol (90 mL) , 5-chloro-2,3-diphenyl-pyrazine (! 11〇1 21.8 8,0.08), triethylamine (14.98,0.15111〇1), the reaction mixture was 100 ° (: The reaction was stirred for 18 hours, the reaction solution cooled to room temperature, water (40 mL), cooled to -10 ° C crystallization for 3 hours and filtered to give SIPA game music, as a white solid (29.6 g of), a yield of 91.0%, the reaction of the present step is the same formula as in Example 1 .

[0046] Example 3

[0047] A) Preparation of [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate:

[0048] 4 – [(tert-butoxycarbonyl) (isopropyl) amino] -1-butanol (12g, 0.05mol) and t-butyl bromoacetate (12.1g, 0.06mol) was dissolved in chloroform (70mL), was added tetrabutylammonium iodide (0 · 5g, 1 · 3mmol), lithium hydroxide (1 · 7g, 0 · 07mol) and water (6.5 g of), the reaction mixture was stirred 20 ° C for 4 hours, the reaction solution under reduced pressure concentrated by rotary evaporation to dryness, extracted with ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, a mixed solvent of ethanol and recrystallized from isopropanol to give [4- (tert-butoxycarbonyl) (isopropyl) aminobutyrate oxygen yl] acetate, as a pale yellow oil (15.6 g of), a yield of 86.8%, the reaction of the present step is the same formula as in Example 1;

[0049] B) Preparation of [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid:

[0050] [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate (15.0g, 0.04mol) was dissolved in isopropanol (40mL), was added a solution of lithium hydroxide (LiOH = 1 · 3g, 0 · 05mol; water 6 · 0g), was heated to 70 ° C for 8 hours, cooled to room temperature, after treatment and purification, to give [4- (tert-butoxycarbonyl) (isopropyl) aminobutyrate oxy] acetic acid as an off-white solid (11.7 g), 93.0% yield, this step is the same reaction scheme of Example 1;

[0051] C) Preparation of 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide:

[0052] [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid (11 (^, 0.04! 11〇1) and 1- (3-dimethylaminopropyl) -3- ethylcarbodiimide (8.38,0.05111〇1) was dissolved in acetonitrile (4〇1111 ^), with stirring, was added methyl sulfonamide (5.] ^, 0.05mol), the reaction mixture was 95 ° C the reaction stirred for 22 hours, The reaction solution was concentrated by rotary evaporation to dryness, extracted with ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, recrystallized from methanol to give 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide as an off-white solid (11.7 g), yield 84.2%, the reaction of the present step is the same formula as in Example 1;

[0053] D) Preparation of SIPA Seiler: 2- [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide (11 (^, 0.03111〇1. ) and dissolved in dichloromethane (601 ^), trifluoroacetic acid (4.48,0.04111〇1), 50 ° (: the reaction was stirred for 10 hours to water (120 mL completion of the reaction, the reaction liquid was added to a stirred), cooled to crystallization 0 ° C for 3 hours and filtered to give the intermediate compound (2- [4- (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide), and then dissolved in tert-butanol (40 mL ), 5-chloro-2,3-diphenyl-pyrazine (9.68,0.036 11〇1), 4-dimethylaminopyridine (8.18,0.07111〇1), the reaction mixture was 110 ° (:! reaction was stirred for 14 hours the reaction was cooled to room temperature, water (15 mL), cooled to -10 ° C crystallization for 3 hours and filtered to give SIPA game music, as a white solid (13.5 g of), a yield of 90.5%, the reaction in this step is the same formula Example 1.

[0054] Example 4

[0055] A) Preparation of [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate:

[0056] 4 – [(tert-butoxycarbonyl) (isopropyl) amino] -1-butanol (15 (^, 0.065111〇1) and t-butyl bromoacetate (17.78,0.09111〇1) was dissolved in toluene (701] 11 ^), was added tetrabutylammonium hydrogen sulfate (0.888,2.61] 11] 1〇1), potassium carbonate (15.2 area, 0.1 lmol) and water (9.5 g of), the reaction mixture was stirred 40 ° C for 1.5 hours, the reaction solution was concentrated under reduced pressure and rotary evaporated to dryness and extracted with ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, a mixed solvent of ethanol and recrystallized from isopropanol to give [4- (tert-butoxycarbonyl) (isopropyl propyl) aminobutoxy] acetate, as a pale yellow oil (19.6 g of), in the same reaction formula in this step a yield of 87.5% in Example 1;

[0057] B) Preparation of [4_ (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid:

[0058] [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetate (19 (^, 0.055! 11〇1) was dissolved in tert-butanol (60 mL), hydroxide solution of cesium (CsOH = 11. lg, 0.07mol; water, 8.0 g), the reaction was heated to 75 ° C for 6.5 hours cooled to room temperature, after treatment and purification, to give [4- (tert-butoxycarbonyl) (isopropyl ) aminobutoxy] acetic acid, an off-white solid (15.0 g of), a yield of 94.2%, the reaction of the present step is the same formula as in Example 1;

[0059] C) Preparation of 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide:

[0060] [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] acetic acid (15.0g, 0.05mol) and diazabicyclo 1,8_

[5.4.0] – | -7- dilute (9.5 region, 0.06111〇1) was dissolved in toluene (8〇1111 ^), with stirring, was added methyl sulfonamide (5.7 region, 0.06mo 1), the reaction mixture was 105 ° C for 16 hours, the reaction solution was concentrated by rotary evaporation to dryness, extracted with ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, recrystallized from methanol to give 2- [4- (tert-butoxycarbonyl) (isopropyl ) aminobutoxy] -N- (methylsulfonyl) acetamide as an off-white solid (16.6 g of), 87.3% yield, this step is the same reaction scheme of Example 1;

[0061] D) Preparation of SIPA Seiler: 2- [4- (tert-butoxycarbonyl) (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide (16 (^, 0.04111〇. 1) and dissolved in ethyl acetate (^ 1,301,111), trifluoroacetic acid (5.78,0.051] 1〇1), 80 <€ the reaction was stirred for 5 hours to complete the reaction, the reaction was added dropwise to a stirred solution of water (150 mL), was cooled to 0 ° C crystallization for 3 hours and filtered to give the intermediate compound (2- [4- (isopropyl) aminobutoxy] -N- (methylsulfonyl) acetamide), then dissolved in isopropanol (50 mL), was added 5-chloro-2,3-diphenyl-pyrazine (13.58,0.05 11〇1!), a hoot dimethylaniline (12.28,0.10111〇1), the reaction mixture was 95 ° (: The reaction was stirred 12 hours, the reaction was cooled to room temperature, water (40 mL), cooled to -10 ° C crystallization for 3 hours and filtered to give SIPA game music, as a white solid (19.7 g of), a yield of 91.0%, the reaction step formula in Example 1.

//////////////

Selexipag (C26H32N4O4S, Mr = 496.6 g/mol) ist ein Diphenylpyrazin-Derivat. Es wird in der Leber zum aktiven Metaboliten ACT-333679 (MRE-269) biotransformiert. Selexipag unterscheidet sich strukturell von Prostazyklin und anderen Prostazylin-Rezeptor-Agonisten.

References

 

  1. Kuwano et al. NS-304, an orally available and long-acting prostacyclin receptor agonist prodrug. J Pharmacol Exp Ther 2007;322:1181-1188.
  2. Kuwano et al. A long-acting and highly selective prostacyclin receptor agonist prodrug, NS-304, ameliorates rat pulmonary hypertension with unique relaxant responses of its active form MRE-269 on rat pulmonary artery. J Pharmacol Exp Ther 2008;326:691-699.
  3. Simonneau G, Lang I, Torbicki A, Hoeper MM, Delcroix M, Karlocai K, Galie N. Selexipag, an oral, selective IP receptor agonist for the treatment of pulmonary arterial hypertension Eur Respir J 2012; 40: 874-880
  4. Mubarak KK. A review of prostaglandin analogs in the management of patients with pulmonary arterial hypertension. Respir Med 2010;104:9-21.
  5. Sitbon, O.; Morrell, N. (2012). “Pathways in pulmonary arterial hypertension: The future is here”. European Respiratory Review 21 (126): 321–327. doi:10.1183/09059180.00004812PMID 23204120.
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    WO2018015974A12016-07-202018-01-25Mylan Laboratories LimitedPolymorphic forms and amorphous solid dispersion of selexipag
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COMPOUNDS CAPABLE OF MODULATING/PRESERVING ENDOTHELIAL INTEGRITY FOR USE IN PREVENTION OR TREATMENT OF ACUTE TRAUMATIC COAGULOPATHY AND RESUSCITATED CARDIAC ARREST [US2015057325] 2013-03-26 2015-02-26
INHIBITION OF NEOVASCULARIZATION BY SIMULTANEOUS INHIBITION OF PROSTANOID IP AND EP4 RECEPTORS [US2014275200] 2014-03-05 2014-09-18
INHIBITION OF NEOVASCULARIZATION BY INHIBITION OF PROSTANOID IP RECEPTORS [US2014275238] 2014-03-05 2014-09-18
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Selexipag
Selexipag.svg
Names
IUPAC name

2-{4-[(5,6-diphenylpyrazin-2-yl)(propan-2-yl)amino]butoxy}-N-(methanesulfonyl)acetamide
Other names

ACT-293987, NS-304
Identifiers
475086-01-2 Yes
ChEMBL ChEMBL238804 
ChemSpider 8089417 Yes
7552
Jmol interactive 3D Image
KEGG D09994 Yes
PubChem 9913767
UNII P7T269PR6S Yes
Properties
C26H32N4O4S
Molar mass 496.6 g·mol−1

//////////ACT-333679,  MRE-269, Selexipag, セレキシパグ , UNII-5EXC0E384L, селексипаг سيليكسيباق Orphan Drug, fda 2015, NS 304,  ACT 293987,  Uptravi, EU 2016, 

CC(C)N(CCCCOCC(=O)NS(=O)(=O)C)C1=CN=C(C(=N1)C2=CC=CC=C2)C3=CC=CC=C3

Selexipag (Uptravi)

Selexipag and its active metabolite, the corresponding carboxylic acid, are nonprostanoid prostaglandin I2 (PGI-2) receptor agonists (Scheme 8).(24) The N-methylsulfonamide within selexipag is hydrolyzed to the corresponding carboxylic acid in vivo by hepatic microsomes at a rate which provides a slow-release pharmacological effect.(24) The compound was originally discovered by Nippon Shinyaki and later licensed to Actelion for development. The drug was approved in 2015 and first launched for the oral treatment of pulmonary arterial hypertension (PAH) in the U.S. in 2016 to delay disease progression and reduce the risk of hospitalization.(25)
Figure
The synthesis of selexipag began with condensation of commercially available benzil (51) and glycinamide hydrochloride in the presence of concentrated sodium hydroxide in refluxing MeOH to yield hydroxypyrazine 52. This compound was subsequently converted to 5-chloro-2,3-diphenylpyrazine (53) upon treatment with refluxing POCl3 in the presence of a catalytic amount of H2SO4.(26) Chloride 53 was then subjected to neat 4-(isopropylamino)-1-butanol (54, prepared by the reductive alkylation of 4-amino-1-butanol and acetone with hydrogen over PtO2 in EtOH) at 190 °C to give aminopyrazinyl alcohol 55 in 56% yield as colorless crystals. Alcohol 55 was alkylated with tert-butyl bromoacetate using Bu4NHSO4 as a phase-transfer catalyst and 40% aqueous KOH in benzene to give ester 56. Although it is particularly unusual to employ benzene on a production scale, these are the only reported conditions for this transformation. The crude ester 56 was then saponified using methanolic sodium hydroxide to yield the corresponding carboxylic acid 57 in 62% as pale-yellow crystals in two steps from compound 55. Finally, the carboxylic acid 57 was coupled with methanesulfonamide in the presence of CDI and DBU in THF to give selexipag (VI) in 77% yield.(27
  1. 24.AsakiT.KuwanoK.MorrisonK.GatfieldJ.HamamotoT.ClozelM. Selexipag: An Oral and Selective IP Prostacyclin Receptor Agonist for the Treatment of Pulmonary Arterial Hypertension J. Med. Chem. 2015,587128– 7137 DOI: 10.1021/acs.jmedchem.5b00698

  2. 25.Skoro-SajerN.LangI. M. Selexipag for the Treatment of Pulmonary Arterial Hypertension Expert Opin. Pharmacother. 201415429– 436 DOI: 10.1517/14656566.2014.876007

  3. 26.KarmasG.SpoerriP. E. The Preparation of Hydroxypyrazines and Derived Chloropyrazines J. Am. Chem. Soc. 1952741580– 1584 DOI: 10.1021/ja01126a070

  4. 27.AsakiT.HamamotoT.SugiyamaY.KuwanoK.KuwabaraK. Structure-activity Studies on Diphenylpyrazine Derivatives: a Novel Class of Prostacyclin Receptor Agonists Bioorg. Med. Chem. 2007,156692– 6704 DOI: 10.1016/j.bmc.2007.08.010

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