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Lisocabtagene maraleucel

March 15, 2021 10:10 am / 1 Comment on Lisocabtagene maraleucel

U.S. FDA Accepts Priority Review for Lisocabtagene Maraleucel R/R Large B-Cell Lymphoma - Onco'Zine Lisocabtagene maraleucel (liso-cel; JCAR017; Anti-CD19 CAR T-Cells) is an investigational chimeric antigen receptor (CAR) T-cell therapy designed to target CD19, ^[1][2] which is a surface glycoprotein expressed during normal B-cell development and maintained following malignant transformation of B cells. ^[3][4][5] Liso-cel CAR T-cells aim to target and CD-19 expressing cells through a CAR construct that includes an anti-CD19 single-chain variable fragment (scFv) targeting domain for antigen specificity, a transmembrane domain, a 4-1BB costimulatory domain hypothesized to increase T-cell proliferation and persistence, and a CD3-zeta T-cell activation domain. ^{[1][2][6][7][8][9]}The defined composition of liso-cel may limit product variability; however, the clinical significance of defined composition is unknown. ^[1][10] Image Courtesy: 2019/2020 Celgene/Juno Therapeutics / Bristol Meyers Squibb.

REF https://www.oncozine.com/u-s-fda-accepts-priority-review-for-lisocabtagene-maraleucel-r-r-large-b-cell-lymphoma/

Lisocabtagene maraleucel

リソカブタゲンマラルユーセル;

JCAR 017

STN# BLA 125714

Adoptive immunotherapy agent JCAR 017
Autologous anti-CD19 scFv/4-1BB/CD3ζ/CD28 chimeric antigen receptor-expressing CD4+/CD8+ central memory T cell JCAR 017
CAR T-cell JCAR 017

FDA 2021, 2021/2/24, BREYANZI

Juno Therapeutics

Antineoplastic, Anti-CD19 CAR-T cell

An immunotherapeutic autologous T cell preparation expressing a chimeric antigen receptor (CAR) specific to the CD19 antigen (Juno Therapeutics, Inc., Seattle, Washington, USA – FDA Clinical Trial Data)

For the treatment of adult patients with relapsed or refractory large B-cell lymphoma after two or more lines of systemic therapy, including diffuse large B-cell lymphoma (DLBCL) not otherwise specified (including DLBCL arising from indolent lymphoma), high-grade B-cell lymphoma, primary mediastinal large B-cell lymphoma, and follicular lymphoma grade 3B.

Lisocabtagene maraleucel, sold under the brand name Breyanzi, is a cell-based gene therapy used to treat large B-cell lymphoma.^[1]^[3]

Side effects of lisocabtagene maraleucel include hypersensitivity reactions, serious infections, low blood cell counts and a weakened immune system.^[3]

Lisocabtagene maraleucel, a chimeric antigen receptor (CAR) T cell therapy, is the third gene therapy approved by the U.S. Food and Drug Administration (FDA) for certain types of non-Hodgkin lymphoma, including diffuse large B-cell lymphoma (DLBCL).^[3] Lisocabtagene maraleucel was approved for medical use in the United States in February 2021.^[1]^[3]

U.S. Food and Drug Administration Approves Bristol Myers Squibb's Breyanzi (lisocabtagene maraleucel), a New CAR T Cell Therapy for Adults with Relapsed or Refractory Large B-cell Lymphoma | Business Wire

Medical uses

Lisocabtagene maraleucel is indicated for the treatment of adults with relapsed or refractory large B-cell lymphoma after two or more lines of systemic therapy, including diffuse large B-cell lymphoma (DLBCL) not otherwise specified (including DLBCL arising from indolent lymphoma), high-grade B-cell lymphoma, primary mediastinal large B-cell lymphoma, and follicular lymphoma grade 3B.^[1]^[3]

Lisocabtagene maraleucel is not indicated for the treatment of people with primary central nervous system lymphoma.^[3]

Adverse effects

The labeling carries a boxed warning for cytokine release syndrome (CRS), which is a systemic response to the activation and proliferation of CAR T cells, causing high fever and flu-like symptoms and neurologic toxicities.^[3]

History

The safety and efficacy of lisocabtagene maraleucel were established in a multicenter clinical trial of more than 250 adults with refractory or relapsed large B-cell lymphoma.^[3] The complete remission rate after treatment with lisocabtagene maraleucel was 54%.^[3]

The FDA granted lisocabtagene maraleucel orphan drug, regenerative medicine advanced therapy (RMAT) and breakthrough therapy designations.^[3] Lisocabtagene maraleucel is the first regenerative medicine therapy with RMAT designation to be licensed by the FDA.^[3] The FDA granted approval of Breyanzi to Juno Therapeutics Inc., a Bristol-Myers Squibb Company.^[3]

SYN

WO 2018156680

WO 2018183366

Saishin Igaku (2018), 73(11), 1504-1512.

WO 2019148089

WO 2019220369

Leukemia & Lymphoma (2020), 61(11), 2561-2567.

WO 2020097350

WO 2020086943

Journal of Immunotherapy (2020), 43(4), 107-120.

https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/breyanzi-lisocabtagene-maraleucel

CLIP

https://www.fda.gov/drugs/drug-approvals-and-databases/fda-approves-lisocabtagene-maraleucel-relapsed-or-refractory-large-b-cell-lymphoma

On February 5, 2021, the Food and Drug Administration approved lisocabtagene maraleucel (Breyanzi, Juno Therapeutics, Inc.) for the treatment of adult patients with relapsed or refractory (R/R) large B-cell lymphoma after two or more lines of systemic therapy, including diffuse large B-cell lymphoma (DLBCL) not otherwise specified (including DLBCL arising from indolent lymphoma), high-grade B-cell lymphoma, primary mediastinal large B-cell lymphoma, and follicular lymphoma grade 3B.

Lisocabtagene maraleucel is a CD19-directed chimeric antigen receptor (CAR) T cell immunotherapy. It consists of autologous T cells that are genetically modified to produce a CAR protein, allowing the T cells to identify and eliminate CD19-expressing normal and malignant cells.

Efficacy was evaluated in TRANSCEND (NCT02631044), a single-arm, open label, multicenter trial that evaluated lisocabtagene maraleucel, preceded by lymphodepleting chemotherapy, in adults with R/R large B-cell lymphoma after at least two lines of therapy.

Of the 192 patients evaluable for response, the overall response rate (ORR) per independent review committee assessment was 73% (95% CI: 67, 80) with a complete response (CR) rate of 54% (95% CI: 47, 61). The median time to first response was one month. Of the 104 patients who achieved CR, 65% had remission lasting at least 6 months and 62% had remission lasting at least 9 months. The estimated median duration of response (DOR) was not reached (95% CI: 16.7 months, NR) in patients who achieved a CR. The estimated median DOR among patients with partial response was 1.4 months (95% CI: 1.1, 2.2).

Cytokine release syndrome (CRS) occurred in 46% of patients (Grade 3 or higher, 4%) and neurologic toxicity occurred in 35% (Grade 3 or higher, 12%). Three patients had fatal neurologic toxicity. Other Grade 3 or higher adverse reactions included infections (19%) and prolonged cytopenias (31%). FDA approved lisocabtagene maraleucel with a Risk Evaluation and Mitigation Strategy because of the risk of fatal or life-threatening CRS and neurologic toxicities.

The recommended regimen is a single dose containing 50 to 110 x 106 CAR-positive viable T cells with a 1:1 ratio of CD4 and CD8 components, administered by IV infusion and preceded by fludarabine and cyclophosphamide for lymphodepletion. Lisocabtagene maraleucel is not indicated for the treatment of patients with primary central nervous system lymphoma.

References

^ Jump up to:^a ^b ^c ^d “Lisocabtagene maraleucel”. U.S. Food and Drug Administration (FDA). 5 February 2021. Retrieved 5 February 2021. This article incorporates text from this source, which is in the public domain.
^ https://www.fda.gov/media/145711/download
^ Jump up to:^a ^b ^c ^d ^e ^f ^g ^h ⁱ ^j ^k ^l “FDA Approves New Treatment For Adults With Relapsed Or Refractory Large-B-Cell Lymphoma”. U.S. Food and Drug Administration (FDA) (Press release). 5 February 2021. Retrieved 5 February 2021. This article incorporates text from this source, which is in the public domain.

External links

“Lisocabtagene maraleucel”. NCI Drug Dictionary. National Cancer Institute.
Clinical trial number NCT02631044 for “Study Evaluating the Safety and Pharmacokinetics of JCAR017 in B-cell Non-Hodgkin Lymphoma (TRANSCEND-NHL-001)” at ClinicalTrials.gov

Clinical data
Trade names	Breyanzi
Other names	JCAR017
License data	US DailyMed: Lisocabtagene_maraleucel
Routes of administration	Intravenous
ATC code	None
Legal status
Legal status	US: ℞-only ^[1]^[2]
Identifiers
UNII	7K2YOJ14X0
KEGG	D11990
ChEMBL	ChEMBL4297236

///////////Lisocabtagene maraleucel, BREYANZI, FDA 2021, APPROVALS 2021, リソカブタゲンマラルユーセル , Juno Therapeutics, JCAR 017, STN# BLA 125714

#Lisocabtagene maraleucel, #BREYANZI, #FDA 2021, #APPROVALS 2021, #リソカブタゲンマラルユーセル , #Juno Therapeutics, #JCAR 017, #STN# BLA 125714

Casimersen

March 14, 2021 6:37 am / Leave a comment

Casimersen

カシメルセン;

RNA, [P-deoxy-P-(dimethylamino)](2′,3′-dideoxy-2′,3′-imino-2′,3′-seco)(2’a→5′)(C-A-A-m⁵U-G-C-C-A-m⁵U-C-C-m⁵U-G-G-A-G-m⁵U-m⁵U-C-C-m⁵U-G), 5′-[P-[4-[[2-[2-(2-hydroxyethoxy)ethoxy]ethoxy]carbonyl]-1-piperazinyl]-N,N-dimethylphosphonamidate]

Formula	C268H424N124O95P22
CAS	1422958-19-7
Mol weight	7584.4307

FDA 2021/2/25 , Amondys 45, Antisense oligonucleotide
Treatment of Duchenne muscular dystrophy

Nucleic Acid Sequence

Sequence Length: 224 a 7 c 5 g 6 umodified

Exon-45: NG-12-0064
SRP-4045
WHO 10354

Casimersen, sold under the brand name Amondys 45, is an antisense oligonucleotide medication used for the treatment of Duchenne muscular dystrophy (DMD) in people who have a confirmed mutation of the dystrophin gene that is amenable to exon 45 skipping.^[1]^[2]^[3]^[4] It is an antisense oligonucleotide of phosphorodiamidate morpholino oligomer (PMO).^[1]

The most common side effects include upper respiratory tract infections, cough, fever, headache, joint pain and throat pain.^[2]

Casimersen was approved for medical use in the United States in February 2021,^[1]^[2] and it is the first FDA-approved targeted treatment for people who have a confirmed mutation of the DMD gene that is amenable to skipping exon 45.^[2]

Duchenne muscular dystrophy (DMD) is an X-linked recessive allelic disorder characterized by a lack of functional dystrophin protein, which leads to progressive impairment of ambulatory, pulmonary, and cardiac function and is invariably fatal. A related, albeit a less severe, form of muscular dystrophy known as Becker muscular dystrophy (BMD) is characterized by shortened and partially functional dystrophin protein production. Although corticosteroids effectively slow disease progression in both DMD and BMD patients, they do not address the underlying molecular pathogenesis.^1,2,3

The application of antisense oligonucleotides in DMD patients with specific mutations allows for exon skipping to produce truncated BMD-like dystrophin proteins, which restore partial muscle function and slow disease progression.^1,2,4,5,7 Casimersen is a phosphorodiamidate morpholino oligonucleotide (PMO); PMOs are oligonucleotides in which the five-membered ribofuranosyl ring is replaced with a six-membered morpholino ring, and the phosphodiester links between nucleotides are replaced with a phosphorodiamidate linkage.^6,7 In this manner, PMOs are much less susceptible to endo- and exonucleases and exhibit drastically reduced metabolic degradation compared to traditional synthetic oligonucleotides.⁶ Casimersen is the most recent in a line of approved PMOs for treating DMD, including eteplirsen and viltolarsen. However, the specific mutations, and hence the precise exon skipping, targeted by each is different.

Casimersen was granted accelerated FDA approval on February 25, 2021, based on data showing an increase in dystrophin levels in skeletal muscle of patients treated with casimersen; this approval is contingent on further verification in confirmatory trials. Casimersen is currently marketed under the tradename AMONDYS 45™ by Sarepta Therapeutics, Inc.⁷

Casimersen is indicated for the treatment of Duchenne muscular dystrophy (DMD) in patients confirmed to have a DMD gene mutation amenable to exon 45 skipping. This indication represents an accelerated approval based on observed efficacy; continued approval for this indication may be contingent on the verification of safety and efficacy in a confirmatory trial.⁷

Medical uses

Casimersen is indicated for the treatment of Duchenne muscular dystrophy (DMD) in people who have a confirmed mutation of the DMD gene that is amenable to exon 45 skipping.^[1]^[2]

History

Casimersen was evaluated in a double-blind, placebo-controlled study in which 43 participants were randomized 2:1 to receive either intravenous casimersen or placebo.^[2] All participants were male, between 7 and 20 years of age, and had a genetically confirmed mutation of the DMD gene that is amenable to exon 45 skipping.^[2]

The U.S. Food and Drug Administration (FDA) granted the application for casimersen fast track, priority review, and orphan drug designations.^[2]^[5] The FDA granted the approval of Amondys 45 to Sarepta Therapeutics, Inc.^[2]

Pharmacodynamics

Casimersen is an antisense phosphorodiamidate morpholino oligonucleotide designed to bind to exon 45 of the DMD pre-mRNA, preventing its inclusion in mature mRNA and allowing the production of an internally truncated dystrophin protein in patients who would normally produce no functional dystrophin. Due to the need for continuous alteration of mRNA splicing and its relatively short half-life, casimersen is administered weekly.⁷ Although casimersen is associated with mostly mild adverse effects, animal studies suggest a potential for nephrotoxicity, which has also been observed after administration of some oligonucleotides.^4,7 Measurement of glomerular filtration rate before starting casimersen is advised. Serum cystatin C, urine dipstick, and urine protein-to-creatinine ratio should be measured before starting therapy. They should be measured monthly (urine dipstick) or every three months (serum cystatin C and urine protein-to-creatinine ratio) during treatment. Creatinine levels are not reliable in muscular dystrophy patients and should not be used. Any persistent alteration in kidney function should be further investigated.⁷

Mechanism of action

Duchenne muscular dystrophy (DMD) is an X-linked recessive allelic disorder that results in the absence of functional dystrophin, a large protein comprising an N-terminal actin-binding domain, C-terminal β-dystroglycan-binding domain, and 24 internal spectrin-like repeats.^1,2,3 Dystrophin is vital for normal muscle function; the absence of dystrophin leads to muscle membrane damage, extracellular leakage of creatinine kinase, calcium influx, and gradual replacement of normal muscle tissue with fibrous and adipose tissue over time.^1,2 DMD shows a characteristic disease progression with early functional complaints related to abnormal gait, locomotion, and falls that remain relatively stable until around seven years of age. The disease then progresses rapidly to loss of independent ambulatory function, ventilatory insufficiency, and cardiomyopathy, with death typically occurring in the second or third decade of life.^1,2,3

The human DMD gene contains 79 exons spread over approximately 2.4 million nucleotides on the X chromosome.¹ DMD is associated with a variety of underlying mutations, including exon duplications or deletions, as well as point mutations leading to nonsense translation through direct production of an in-frame stop codon, frameshift production of an in-frame stop codon, or aberrant inclusion of an intronic pseudo-exon with the concomitant production of an in-frame stop codon.^1,2 In all cases, no functional dystrophin protein is produced. Becker muscular dystrophy (BMD) is a related condition with in-frame mutations that result in the production of a truncated but partially functional dystrophin protein. BMD patients, therefore, have milder symptoms, delayed disease progression, and longer life expectancy compared to DMD patients.^1,2,3

Casimersen is an antisense phosphorodiamidate morpholino oligonucleotide designed to bind to exon 45 of the DMD pre-mRNA and prevent its inclusion within the mature mRNA before translation.^4,7 It is estimated that around 8% of DMD patients may benefit from exon 45 skipping, in which the exclusion of this exon results in the production of an internally truncated and at least partly functional dystrophin protein.^4,7,5 Although fibrotic or fatty muscle tissue developed previously cannot be improved, this therapy aims to slow further disease progression through the production of partially functional dystrophin and alleviation of the pathogenic mechanism of muscle tissue necrosis.^1,2

TARGET	ACTIONS	ORGANISM
ADMD gene (exon 45 casimersen target site)	binder	Humans

Absorption

DMD patients receiving IV doses of 4-30 mg/kg/week revealed exposure in proportion to dose with no accumulation of casimersen in plasma with once-weekly dosing. Following a single IV dose, casimersen C_max was reached by the end of infusion. Inter-subject variability, as measured by the coefficient of variation, ranged from 12-34% for C_max and 16-34% for AUC.⁷

Pre-clinical studies in nonhuman primates (cynomolgus monkeys) investigated the pharmacokinetics of once-weekly casimersen administered at doses of 5, 40, and 320 mg/kg. On days 1 and 78, the 5 mg/kg dose resulted in a C_max of 19.5 ± 3.43 and 21.6 ± 5.60 μg/mL and an AUC_0-t of 24.9 ± 5.17 and 26.9 ± 7.94 μg*hr/mL. The 40 mg/kg dose resulted in a C_max of 208 ± 35.2 and 242 ± 71.1 μg/mL and an AUC_0-t of 283 ± 68.5 and 320 ± 111 μg*hr/mL. Lastly, the 320 mg/kg dose resulted in a a C_max of 1470 ± 88.1 and 1490 ± 221 μg/mL and an AUC_0-t of 1960 ± 243 and 1930 ± 382 μg*hr/mL.⁴

Volume of distribution

Casimersen administered at 30 mg/kg had a mean steady-state volume of distribution (%CV) of 367 mL/kg (28.9%).⁷

Protein binding

Casimersen binding to human plasma proteins is not concentration-dependent, ranging from 8.4-31.6%.⁷

Metabolism

Casimersen incubated with human hepatic microsomal preparations is metabolically stables and no metabolites are detected in plasma or urine.⁷

Route of elimination

Casimersen is predominantly (more than 90%) excreted in the urine unchanged with negligible fecal excretion.⁷

Half-life

Casimersen has an elimination half-life of 3.5 ± 0.4 hours.⁷

Clearance

Casimersen administered at 30 mg/kg has a plasma clearance of 180 mL/hr/kg.⁷

NAME	DOSAGE	STRENGTH	ROUTE	LABELLER	MARKETING START	MARKETING END
Amondys 45	Injection	50 mg/1mL	Intravenous	Sarepta Therapeutics, Inc.	2021-02-25	Not applicable

Synthesis Reference

Diane Elizabeth Frank and Richard K. Bestwick, “Exon skipping oligomers for muscular dystrophy.” U.S. Patent US20190262375A1, issued August 29, 2019.

PATENT

https://patents.google.com/patent/WO2017205879A2/en

also

WO 2021025899

References

^ Jump up to:^a ^b ^c ^d ^e “Amondys 45- casimersen injection”. DailyMed. Retrieved 1 March 2021.
^ Jump up to:^a ^b ^c ^d ^e ^f ^g ^h ⁱ ^j “FDA Approves Targeted Treatment for Rare Duchenne Muscular Dystrophy Mutation”. U.S. Food and Drug Administration (FDA) (Press release). 25 February 2021. Retrieved 25 February 2021. This article incorporates text from this source, which is in the public domain.
^ “Sarepta Therapeutics Announces FDA Approval of Amondys 45 (casimersen) Injection for the Treatment of Duchenne Muscular Dystrophy (DMD) in Patients Amenable to Skipping Exon 45” (Press release). Sarepta Therapeutics. 25 February 2021. Retrieved 25 February 2021 – via GlobeNewswire.
^ Rodrigues M, Yokota T (2018). “An Overview of Recent Advances and Clinical Applications of Exon Skipping and Splice Modulation for Muscular Dystrophy and Various Genetic Diseases”. Exon Skipping and Inclusion Therapies. Methods in Molecular Biology. 1828. Clifton, N.J. pp. 31–55. doi:10.1007/978-1-4939-8651-4_2. ISBN 978-1-4939-8650-7. PMID 30171533.
^ “Casimersen Orphan Drug Designations and Approvals”. U.S. Food and Drug Administration (FDA). 4 June 2019. Retrieved 25 February 2021.

General References

Wein N, Alfano L, Flanigan KM: Genetics and emerging treatments for Duchenne and Becker muscular dystrophy. Pediatr Clin North Am. 2015 Jun;62(3):723-42. doi: 10.1016/j.pcl.2015.03.008. Epub 2015 Apr 20. [PubMed:26022172]
Verhaart IEC, Aartsma-Rus A: Therapeutic developments for Duchenne muscular dystrophy. Nat Rev Neurol. 2019 Jul;15(7):373-386. doi: 10.1038/s41582-019-0203-3. [PubMed:31147635]
Mercuri E, Bonnemann CG, Muntoni F: Muscular dystrophies. Lancet. 2019 Nov 30;394(10213):2025-2038. doi: 10.1016/S0140-6736(19)32910-1. [PubMed:31789220]
Carver MP, Charleston JS, Shanks C, Zhang J, Mense M, Sharma AK, Kaur H, Sazani P: Toxicological Characterization of Exon Skipping Phosphorodiamidate Morpholino Oligomers (PMOs) in Non-human Primates. J Neuromuscul Dis. 2016 Aug 30;3(3):381-393. doi: 10.3233/JND-160157. [PubMed:27854228]
Rodrigues M, Yokota T: An Overview of Recent Advances and Clinical Applications of Exon Skipping and Splice Modulation for Muscular Dystrophy and Various Genetic Diseases. Methods Mol Biol. 2018;1828:31-55. doi: 10.1007/978-1-4939-8651-4_2. [PubMed:30171533]
Smith CIE, Zain R: Therapeutic Oligonucleotides: State of the Art. Annu Rev Pharmacol Toxicol. 2019 Jan 6;59:605-630. doi: 10.1146/annurev-pharmtox-010818-021050. Epub 2018 Oct 9. [PubMed:30285540]
FDA Approved Drug Products: AMONDYS 45 (casimersen) injection [Link]

External links

“Casimersen”. Drug Information Portal. U.S. National Library of Medicine.
Clinical trial number NCT02500381 for “Study of SRP-4045 and SRP-4053 in DMD Patients (ESSENCE)” at ClinicalTrials.gov

Clinical data
Trade names	Amondys 45
Other names	SRP-4045
License data	US DailyMed: Casimersen
Routes of administration	Intravenous
Drug class	Antisense oligonucleotide
ATC code	None
Legal status
Legal status	US: ℞-only ^[1]^[2]
Identifiers
CAS Number	1422958-19-7
DrugBank	DB14984
UNII	X8UHF7SX0R
KEGG	D11988
Chemical and physical data
Formula	C₂₆₈H₄₂₄N₁₂₄O₉₅P₂₂
Molar mass	7584.536 g·mol⁻¹

////////////Casimersen, FDA 2021, APPROVALS 2021, カシメルセン , Exon-45: NG-12-0064, SRP-4045, WHO 10354, Amondys 45, Antisense oligonucleotide, Duchenne muscular dystrophy

#Casimersen, #FDA 2021, #APPROVALS 2021, #カシメルセン , #Exon-45: NG-12-0064, #SRP-4045, #WHO 10354, #Amondys 45, #Antisense oligonucleotide, #Duchenne muscular dystrophy

Sequence:

1caaugccauc cuggaguucc ug

Sequence Modifications

Type	Location	Description
modified base	c-1	5′-ester
modified base	c-1	modified cytidine
modified base	a-2	modified adenosine
modified base	a-3	modified adenosine
modified base	u-4	m5u
modified base	u-4	modified uridine
modified base	g-5	modified guanosine
modified base	c-6	modified cytidine
modified base	c-7	modified cytidine
modified base	a-8	modified adenosine
modified base	u-9	modified uridine
modified base	u-9	m5u
modified base	c-10	modified cytidine
modified base	c-11	modified cytidine
modified base	u-12	m5u
modified base	u-12	modified uridine
modified base	g-13	modified guanosine
modified base	g-14	modified guanosine
modified base	a-15	modified adenosine
modified base	g-16	modified guanosine
modified base	u-17	modified uridine
modified base	u-17	m5u
modified base	u-18	modified uridine
modified base	u-18	m5u
modified base	c-19	modified cytidine
modified base	c-20	modified cytidine
modified base	u-21	m5u
modified base	u-21	modified uridine
modified base	g-22	modified guanosine
uncommon link	c-1 – a-2	unavailable
uncommon link	a-2 – a-3	unavailable
uncommon link	a-3 – u-4	unavailable
uncommon link	u-4 – g-5	unavailable
uncommon link	g-5 – c-6	unavailable
uncommon link	c-6 – c-7	unavailable
uncommon link	c-7 – a-8	unavailable
uncommon link	a-8 – u-9	unavailable
uncommon link	u-9 – c-10	unavailable
uncommon link	c-10 – c-11	unavailable
uncommon link	c-11 – u-12	unavailable
uncommon link	u-12 – g-13	unavailable
uncommon link	g-13 – g-14	unavailable
uncommon link	g-14 – a-15	unavailable
uncommon link	a-15 – g-16	unavailable
uncommon link	g-16 – u-17	unavailable
uncommon link	u-17 – u-18	unavailable
uncommon link	u-18 – c-19	unavailable
uncommon link	c-19 – c-20	unavailable
uncommon link	c-20 – u-21	unavailable
uncommon link	u-21 – g-22	unavailable

Fosdenopterin hydrobromide

March 12, 2021 6:31 am / 1 Comment on Fosdenopterin hydrobromide

Fosdenopterin hydrobromide

FDA APPR 2021/2/26, NULIBRY

BBP-870/ORGN001

a cyclic pyranopterin monophosphate (cPMP) substrate replacement therapy, for the treatment of patients with molybdenum cofactor deficiency (MoCD) Type A.

ホスデノプテリン臭化水素酸塩水和物;

Formula	C10H14N5O8P. 2H2O. HBr
CAS	2301083-34-9DIHYDRATE
Mol weight	480.1631

2301083-34-9

(1R,10R,12S,17R)-5-amino-11,11,14-trihydroxy-14-oxo-13,15,18-trioxa-2,4,6,9-tetraza-14λ⁵-phosphatetracyclo[8.8.0.0^3,8.0^12,17]octadeca-3(8),4-dien-7-one;dihydrate;hydrobromide

1,3,2-DIOXAPHOSPHORINO(4′,5′:5,6)PYRANO(3,2-G)PTERIDIN-10(4H)-ONE, 8-AMINO-4A,5A,6,9,11,11A,12,12A-OCTAHYDRO-2,12,12-TRIHYDROXY-, 2-OXIDE, HYDROBROMIDE, HYDRATE (1:1:2), (4AR,5AR,11AR,12AS)-

CYCLIC PYRANOPTERIN MONOPHOSPHATE MONOHYDROBROMIDE DIHYDRATE

(4aR,5aR,11aR,12aS)-8-Amino-2,12,12-trihydroxy-4a,5a,6,7,11,11a,12,12aoctahydro-2H-2lambda5-(1,3,2)dioxaphosphinino(4′,5′:5,6)pyrano(3,2-g)pteridine-2,10(4H)-dione, hydrobromide (1:1:2)

1,3,2-Dioxaphosphorino(4′,5′:5,6)pyrano(3,2-g)pteridin-10(4H)-one, 8-amino-4a,5a,6,9,11,11a,12,12a-octahydro-2,12,12-trihydroxy-, 2-oxide, hydrobromide, hydrate (1:1:2), (4aR,5aR,11aR,12aS)-

1,3,2-Dioxaphosphorino(4′,5′:5,6)pyrano(3,2-g)pteridin-10(4H)-one, 8-amino-4a,5a,6,9,11,11a,12,12a-octahydro-2,12,12-trihydroxy-, 2-oxide,hydrobromide, hydrate (1:1:2), (4aR,5aR,11aR,12aS)-

ALXN1101 HBr, UNII-X41B5W735T, X41B5W735T, D11780

Nulibry Approved for Molybdenum Cofactor Deficiency Type A - MPR

ChemSpider 2D Image | Cyclic pyranopterin monophosphate | C10H14N5O8P

C₁₀H₁₄N₅O₈P, Average: 363.223

150829-29-1

ALXN-1101
WHO 11150
Synthesis ReferenceClinch K, Watt DK, Dixon RA, Baars SM, Gainsford GJ, Tiwari A, Schwarz G, Saotome Y, Storek M, Belaidi AA, Santamaria-Araujo JA: Synthesis of cyclic pyranopterin monophosphate, a biosynthetic intermediate in the molybdenum cofactor pathway. J Med Chem. 2013 Feb 28;56(4):1730-8. doi: 10.1021/jm301855r. Epub 2013 Feb 19.

Fosdenopterin (or cyclic pyranopterin monophosphate, cPMP), sold under the brand name Nulibry, is a medication used to reduce the risk of death due to a rare genetic disease known as molybdenum cofactor deficiency type A (MoCD-A).^[1]

Adverse effects

The most common side effects include complications related to the intravenous line, fever, respiratory infections, vomiting, gastroenteritis, and diarrhea.^[1]

Mechanism of action

People with MoCD-A cannot produce cyclic pyranopterin monophosphate (cPMP) in their body.^[1] Fosdenopterin is an intravenous medication that replaces the missing cPMP.^[1]^[2] cPMP is a precursor to molybdopterin, which is required for the enzyme activity of sulfite oxidase, xanthine dehydrogenase/oxidase and aldehyde oxidase.^[3]

History

Fosdenopterin was developed by José Santamaría-Araujo and Guenter Schwarz at the German universities TU Braunschweig and the University of Cologne.^[4]^[5]

The effectiveness of fosdenopterin for the treatment of MoCD-A was demonstrated in thirteen treated participants compared to eighteen matched, untreated participants.^[1]^[6] The participants treated with fosdenopterin had a survival rate of 84% at three years, compared to 55% for the untreated participants.^[1]

The U.S. Food and Drug Administration (FDA) granted the application for fosdenopterin priority review, breakthrough therapy, and orphan drug designations along with a rare pediatric disease priority review voucher.^[1] The FDA granted the approval of Nulibry to Origin Biosciences, Inc., in February 2021.^[1] It is the first medication approved for the treatment of MoCD-A.^[1]

References

^ Jump up to:^a ^b ^c ^d ^e ^f ^g ^h ⁱ ^j “FDA Approves First Treatment for Molybdenum Cofactor Deficiency Type A”. U.S. Food and Drug Administration (FDA) (Press release). 26 February 2021. Retrieved 26 February 2021. This article incorporates text from this source, which is in the public domain.
^ DrugBank DB16628 . Accessed 2021-03-05.
^ Santamaria-Araujo JA, Fischer B, Otte T, Nimtz M, Mendel RR, Wray V, Schwarz G (April 2004). “The tetrahydropyranopterin structure of the sulfur-free and metal-free molybdenum cofactor precursor”. The Journal of Biological Chemistry. 279 (16): 15994–9. doi:10.1074/jbc.M311815200. PMID 14761975.
^ Schwarz G, Santamaria-Araujo JA, Wolf S, Lee HJ, Adham IM, Gröne HJ, et al. (June 2004). “Rescue of lethal molybdenum cofactor deficiency by a biosynthetic precursor from Escherichia coli”. Human Molecular Genetics. 13 (12): 1249–55. doi:10.1093/hmg/ddh136. PMID 15115759.
^ Tedmanson S (5 November 2009). “Doctors risk untried drug to stop baby’s brain dissolving”. TimesOnline.
^ Schwahn BC, Van Spronsen FJ, Belaidi AA, Bowhay S, Christodoulou J, Derks TG, et al. (November 2015). “Efficacy and safety of cyclic pyranopterin monophosphate substitution in severe molybdenum cofactor deficiency type A: a prospective cohort study”. Lancet. 386 (10007): 1955–63. doi:10.1016/S0140-6736(15)00124-5. PMID 26343839. S2CID 21954888.

External links

“Fosdenopterin”. Drug Information Portal. U.S. National Library of Medicine.

Molybdenum cofactor deficiency (MoCD) is an exceptionally rare autosomal recessive disorder resulting in a deficiency of three molybdenum-dependent enzymes: sulfite oxidase (SOX), xanthine dehydrogenase, and aldehyde oxidase.¹ Signs and symptoms begin shortly after birth and are caused by a build-up of toxic sulfites resulting from a lack of SOX activity.^1,5 Patients with MoCD may present with metabolic acidosis, intracranial hemorrhage, feeding difficulties, and significant neurological symptoms such as muscle hyper- and hypotonia, intractable seizures, spastic paraplegia, myoclonus, and opisthotonus. In addition, patients with MoCD are often born with morphologic evidence of the disorder such as microcephaly, cerebral atrophy/hypodensity, dilated ventricles, and ocular abnormalities.¹ MoCD is incurable and median survival in untreated patients is approximately 36 months¹ – treatment, then, is focused on improving survival and maintaining neurological function.

The most common subtype of MoCD, type A, involves mutations in MOCS1 wherein the first step of molybdenum cofactor synthesis – the conversion of guanosine triphosphate into cyclic pyranopterin monophosphate (cPMP) – is interrupted.^1,3 In the past, management strategies for this disorder involved symptomatic and supportive treatment,⁵ though efforts were made to develop a suitable exogenous replacement for the missing cPMP. In 2009 a recombinant, E. coli-produced cPMP was granted orphan drug designation by the FDA, becoming the first therapeutic option for patients with MoCD type A.¹

Fosdenopterin was approved by the FDA on Februrary 26, 2021, for the reduction of mortality in patients with MoCD type A,⁵ becoming the first and only therapy approved for the treatment of MoCD. By improving the three-year survival rate from 55% to 84%,⁷ and considering the lack of alternative therapies available, fosdenopterin appears poised to become a standard of therapy in the management of this debilitating disorder.

Fosdenopterin replaces an intermediate substrate in the synthesis of molybdenum cofactor, a compound necessary for the activation of several molybdenum-dependent enzymes including sulfite oxidase (SOX).¹ Given that SOX is responsible for detoxifying sulfur-containing acids and sulfites such as S-sulfocysteine (SSC), urinary levels of SSC can be used as a surrogate marker of efficacy for fosdenopterin.⁷ Long-term therapy with fosdenopterin has been shown to result in a sustained reduction in urinary SSC normalized to creatinine.⁷

Animal studies have identified a potential risk of phototoxicity in patients receiving fosdenopterin – these patients should avoid or minimize exposure to sunlight and/or artificial UV light.⁷ If sun exposure is necessary, use protective clothing, hats, and sunglasses,⁷ in addition to seeking shade whenever practical. Consider the use of a broad-spectrum sunscreen in patients 6 months of age or older.⁸

Molybdenum cofactor deficiency (MoCD) is a rare autosomal-recessive disorder in which patients are deficient in three molybdenum-dependent enzymes: sulfite oxidase (SOX), xanthine dehydrogenase, and aldehyde dehydrogenase.¹ The loss of SOX activity appears to be the main driver of MoCD morbidity and mortality, as the build-up of neurotoxic sulfites typically processed by SOX results in rapid and progressive neurological damage. In MoCD type A, the disorder results from a mutation in the MOCS1 gene leading to deficient production of MOCS1A/B,⁷ a protein that is responsible for the first step in the synthesis of molybdenum cofactor: the conversion of guanosine triphosphate into cyclic pyranopterin monophosphate (cPMP).^1,4

Fosdenopterin is an exogenous form of cPMP, replacing endogenous production and allowing for the synthesis of molybdenum cofactor to proceed.⁷

Mechler K, Mountford WK, Hoffmann GF, Ries M: Ultra-orphan diseases: a quantitative analysis of the natural history of molybdenum cofactor deficiency. Genet Med. 2015 Dec;17(12):965-70. doi: 10.1038/gim.2015.12. Epub 2015 Mar 12. [PubMed:25764214]
Schwahn BC, Van Spronsen FJ, Belaidi AA, Bowhay S, Christodoulou J, Derks TG, Hennermann JB, Jameson E, Konig K, McGregor TL, Font-Montgomery E, Santamaria-Araujo JA, Santra S, Vaidya M, Vierzig A, Wassmer E, Weis I, Wong FY, Veldman A, Schwarz G: Efficacy and safety of cyclic pyranopterin monophosphate substitution in severe molybdenum cofactor deficiency type A: a prospective cohort study. Lancet. 2015 Nov 14;386(10007):1955-63. doi: 10.1016/S0140-6736(15)00124-5. Epub 2015 Sep 3. [PubMed:26343839]
Iobbi-Nivol C, Leimkuhler S: Molybdenum enzymes, their maturation and molybdenum cofactor biosynthesis in Escherichia coli. Biochim Biophys Acta. 2013 Aug-Sep;1827(8-9):1086-101. doi: 10.1016/j.bbabio.2012.11.007. Epub 2012 Nov 29. [PubMed:23201473]
Mendel RR: The molybdenum cofactor. J Biol Chem. 2013 May 10;288(19):13165-72. doi: 10.1074/jbc.R113.455311. Epub 2013 Mar 28. [PubMed:23539623]
FDA News Release: FDA Approves First Treatment for Molybdenum Cofactor Deficiency Type A [Link]
OMIM: MOLYBDENUM COFACTOR DEFICIENCY, COMPLEMENTATION GROUP A (# 252150) [Link]
FDA Approved Drug Products: Nulibry (fosdenopterin) for intravenous injection [Link]
Health Canada: Sun safety tips for parents [Link]

SYN

Journal of Biological Chemistry (1995), 270(3), 1082-7.

https://linkinghub.elsevier.com/retrieve/pii/S0021925818829696

PATENT

WO 2005073387

PATENT

WO 2012112922

PAPER

Journal of Medicinal Chemistry (2013), 56(4), 1730-1738

https://pubs.acs.org/doi/10.1021/jm301855r

Cyclic pyranopterin monophosphate (1), isolated from bacterial culture, has previously been shown to be effective in restoring normal function of molybdenum enzymes in molybdenum cofactor (MoCo)-deficient mice and human patients. Described here is a synthesis of 1 hydrobromide (1·HBr) employing in the key step a Viscontini reaction between 2,5,6-triamino-3,4-dihydropyrimidin-4-one dihydrochloride and d-galactose phenylhydrazone to give the pyranopterin (5aS,6R,7R,8R,9aR)-2-amino-6,7-dihydroxy-8-(hydroxymethyl)-3H,4H,5H,5aH,6H,7H,8H,9aH,10H-pyrano[3,2-g]pteridin-4-one (10) and establishing all four stereocenters found in 1. Compound 10, characterized spectroscopically and by X-ray crystallography, was transformed through a selectively protected tri-tert-butoxycarbonylamino intermediate into a highly crystalline tetracyclic phosphate ester (15). The latter underwent a Swern oxidation and then deprotection to give 1·HBr. Synthesized 1·HBr had in vitro efficacy comparable to that of 1 of bacterial origin as demonstrated by its enzymatic conversion into mature MoCo and subsequent reconstitution of MoCo-free human sulfite oxidase–molybdenum domain yielding a fully active enzyme. The described synthesis has the potential for scale up.

PAPER

European Journal of Organic Chemistry (2014), 2014(11), 2231-2241.

https://chemistry-europe.onlinelibrary.wiley.com/doi/abs/10.1002/ejoc.201301784

Click to access downloadSupplement

Abstract

The first synthesis of an oxygen‐stable analogue of the natural product cyclic pyranopterin monophosphate (cPMP) is reported. In this approach, the hydropyranone ring is annelated to pyrazine by a sequence comprising ortho‐lithiation/acylation of a 2‐halopyrazine, followed by nucleophilic aromatic substitution. The tetrose substructure is introduced from the chiral pool, from D‐galactose or D‐arabitol.

Abstract

Molybdenum cofactor (Moco) deficiency is a lethal hereditary metabolic disease. A recently developed therapy requires continuous intravenous supplementation of the biosynthetic Moco precursor cyclic pyranopterin monophosphate (cPMP). The limited stability of the latter natural product, mostly due to oxidative degradation, is problematic for oral administration. Therefore, the synthesis of more stable cPMP analogues is of great interest. In this context and for the first time, the synthesis of a cPMP analogue, in which the oxidation‐labile reduced pterin unit is replaced by a pyrazine moiety, was achieved starting from the chiral pool materials D‐galactose or D‐arabitol. Our synthesis, 13 steps in total, includes the following key transformations: i) pyrazine lithiation, followed by acylation; ii) closure of the pyrane ring by nucleophilic aromatic substitution; and iii) introduction of phosphate.

Patent

https://patents.google.com/patent/US9260462B2/en

Molybdenum cofactor (Moco) deficiency is a pleiotropic genetic disorder. Moco consists of molybdenum covalently bound to one or two dithiolates attached to a unique tricyclic pterin moiety commonly referred to as molybdopterin (MPT). Moco is synthesized by a biosynthetic pathway that can be divided into four steps, according to the biosynthetic intermediates precursor Z (cyclic pyranopterin monophosphate; cPMP), MPT, and adenylated MPT. Mutations in the Moco biosynthetase genes result in the loss of production of the molybdenum dependent enzymes sulfite-oxidase, xanthine oxidoreductase, and aldehyde oxidase. Whereas the activities of all three of these cofactor-containing enzymes are impaired by cofactor deficiency, the devastating consequences of the disease can be traced to the loss of sulfite oxidase activity. Human Moco deficiency is a rare but severe disorder accompanied by serious neurological symptoms including attenuated growth of the brain, untreatable seizures, dislocated ocular lenses, and mental retardation. Until recently, no effective therapy was available and afflicted patients suffering from Moco deficiency died in early infancy.

It has been found that administration of the molybdopterin derivative precursor Z, a relatively stable intermediate in the Moco biosynthetic pathway, is an effective means of therapy for human Moco deficiency and associated diseases related to altered Moco synthesis (see U.S. Pat. No. 7,504,095). As with most replacement therapies for illnesses, however, the treatment is limited by the availability of the therapeutic active agent.

Scheme 3.

Scheme 4.

(I).

Scheme 6.

(I).

Scheme 8.

(I).

Scheme 10.

EXAMPLESExample 1Preparation of Precursor Z (cPMP)

Experimental

Air sensitive reactions were performed under argon. Organic solutions were dried over anhydrous MgSO₄and the solvents were evaporated under reduced pressure. Anhydrous and chromatography solvents were obtained commercially (anhydrous grade solvent from Sigma-Aldrich Fine Chemicals) and used without any further purification. Thin layer chromatography (t.l.c.) was performed on glass or aluminum sheets coated with 60 F₂₅₄silica gel. Organic compounds were visualized under UV light or with use of a dip of ammonium molybdate (5 wt %) and cerium(IV) sulfate 4H₂O (0.2 wt %) in aq. H₂SO₄(2M), one of I₂(0.2%) and KI (7%) in H₂SO₄(1M), or 0.1% ninhydrin in EtOH. Chromatography (flash column) was performed on silica gel (40-63 μm) or on an automated system with continuous gradient facility. Optical rotations were recorded at a path length of 1 dm and are in units of 10⁻¹deg cm²g⁻¹; concentrations are in g/100 mL. ¹H NMR spectra were measured in CDCl₃, CD₃OD (internal Me₄Si, δ 0 ppm) or D₂O(HOD, δ 4.79 ppm), and ¹³C NMR spectra in CDCl₃(center line, δ 77.0 ppm), CD₃OD (center line, δ 49.0 ppm) or DMSO d₆(center line δ 39.7 ppm), D₂O (no internal reference or internal CH₃CN, δ 1.47 ppm where stated). Assignments of ¹H and ¹³C resonances were based on 2D (¹H—¹H DQF-COSY, ¹H—¹³C HSQC, HMBC) and DEPT experiments. ³¹P NMR were run at 202.3 MHz and are reported without reference. High resolution electrospray mass spectra (ESI-HRMS) were recorded on a Q-TOF Tandem Mass

Spectrometer. Microanalyses were performed by the Campbell Microanalytical Department, University of Otago, Dunedin, New Zealand.

A. Preparation of (5aS,6R,7R,8R,9aR)-2-amino-6,7-dihydroxy-8-(hydroxymethyl)-3H,4H,5H,5aH,6H,7H,8H,9aH,10H-pyrano[3,2-g]pteridin-4-one mono hydrate (1)

2,5,6-Triamino-3,4-dihydropyrimidin-4-one dihydrochloride (Pfleiderer, W.; Chem. Ber. 1957, 90, 2272; Org. Synth. 1952, 32, 45; Org. Synth. 1963, Coll. Vol. 4, 245, 10.0 g, 46.7 mmol), D-galactose phenylhydrazone (Goswami, S.; Adak, A. K. Tetrahedron Lett. 2005, 46, 221-224, 15.78 g, 58.4 mmol) and 2-mercaptoethanol (1 mL) were stirred and heated to reflux (bath temp 110° C.) in a 1:1 mixture of MeOH—H₂O (400 mL) for 2 h. After cooling to ambient temperature, diethyl ether (500 mL) was added, the flask was shaken and the diethyl ether layer decanted off and discarded. The process was repeated with two further portions of diethyl ether (500 mL) and then the remaining volatiles were evaporated. Methanol (40 mL), H₂O (40 mL) and triethylamine (39.4 mL, 280 mmol) were successively added and the mixture seeded with a few milligrams of 1. After 5 min a yellow solid was filtered off, washed with a little MeOH and dried to give 1 as a monohydrate (5.05 g, 36%) of suitable purity for further use. An analytical portion was recrystallized from DMSO-EtOH or boiling H₂O. MPt 226 dec. [α]_D ²⁰+135.6 (c1.13, DMSO). ¹H NMR (DMSO d₆): δ 10.19 (bs, exchanged D₂O, 1H), 7.29 (d, J=5.0 Hz, slowly exchanged D₂O, 1H), 5.90 (s, exchanged D₂O, 2H), 5.33 (d, J=5.4 Hz, exchanged D₂O, 1H), 4.66 (ddd, J˜5.0, ˜1.3, ˜1.3 Hz, 1H), 4.59 (t, J=5.6 Hz, exchanged D₂O, 1H), 4.39 (d, J=10.3 Hz, exchanged D₂O, 1H), 3.80 (bt, J˜1.8 Hz, exchanged D₂O, 1H), 3.70 (m, 1H), 3.58 (dd, J=10.3, 3.0 Hz, 1H), 3.53 (dt, J=10.7, 6.4 Hz, 1H), 3.43 (ddd, J=11.2, 5.9, 5.9 Hz, 1H), 3.35 (t, J=6.4 Hz, 1H), 3.04 (br m, 1H). ¹³C NMR (DMSO d₆center line 6 39.7): δ 156.3 (C), 150.4 (C), 148.4 (C), 99.0 (C), 79.4 (CH), 76.5 (CH), 68.9 (CH), 68.6 (CH), 60.6 (CH₂), 53.9 (CH). Anal. calcd. for C₁₀H₁₅N₅O₅H₂O 39.60; C, 5.65; H, 23.09; N. found 39.64; C, 5.71; H, 22.83; N.

B. Preparation of Compounds 2 (a or b) and 3 (a, b or c)

Di-tert-butyl dicarbonate (10.33 g, 47.3 mmol) and DMAP (0.321 g, 2.63 mmol) were added to a stirred suspension of 1 (1.5 g, 5.26 mmol) in anhydrous THF (90 mL) at 50° C. under Ar. After 20 h a clear solution resulted. The solvent was evaporated and the residue chromatographed on silica gel (gradient of 0 to 40% EtOAc in hexanes) to give two product fractions. The first product to elute was a yellow foam (1.46 g). The product was observed to be a mixture of two compounds by ¹H NMR containing mainly a product with seven Boc groups (2a or 2b). A sample was crystallized from EtOAc-hexanes to give 2a or 2b as a fine crystalline solid. MPt 189-191° C. [α]_D ²⁰−43.6 (c 0.99, MeOH). ¹H NMR (500 MHz, CDCl₃): δ 5.71 (t, J=1.7 Hz, 1H), 5.15 (dt, J=3.5, ˜1.0, 1H), 4.97 (t, J=3.8, 1H), 4.35 (br t, J=˜1.7, 1H), 4.09-3.97 (m, 3H), 3.91 (m, 1H), 1.55, 1.52, 1.51, 1.50, 1.45 (5s, 45H), 1.40 (s, 18H). ¹³C NMR (125.7 MHz, CDCl₃): δ 152.84 (C), 152.78 (C), 151.5 (C), 150.9 (C), 150.7 (2×C), 150.3 (C), 149.1 (C), 144.8 (C), 144.7 (C), 118.0 (C), 84.6 (C), 83.6 (C), 83.5 (C), 82.7 (3×C), 82.6 (C), 76.3 (CH), 73.0 (CH), 71.4 (CH), 67.2 (CH), 64.0 (CH₂), 51.4 (CH), 28.1 (CH₃), 27.8 (2×CH₃), 27.7 (CH₃), 27.6 (3×CH₃). MS-ESI+ for C₄₅H₇₂N₅O₁₉ ⁺, (M+H)⁺, Calcd. 986.4817. found 986.4818. Anal. calcd. for C₄₅H₇₁N₅O₁₉H₂O 54.39; C, 7.39; H, 6.34; N. found 54.66; C, 7.17; H, 7.05; N. A second fraction was obtained as a yellow foam (2.68 g) which by ¹H NMR was a product with six Boc groups present (3a, 3b or 3c). A small amount was crystallized from EtOAc-hexanes to give colorless crystals. [α]_D ²O −47.6 (c, 1.17, CHCl₃). ¹H NMR (500 MHz, CDCl₃): δ 11.10 (br s, exchanged D₂O, 1H), 5.58 (t, J=1.8 Hz, 1H), 5.17 (d, J=3.4 Hz, 1H), 4.97 (t, J=3.9 Hz, 1H), 4.62 (s, exchanged D₂O, 1H), 4.16 (dd, J=11.3, 5.9 Hz, 1H), 4.12 (dd, J=11.3, 6.4 Hz, 1H), 3.95 (dt, J=6.1, 1.1 Hz, 1H), 3.76 (m, 1H), 1.51, 1.50, 1.49, 1.48, 1.46 (5s, 54H). ¹³C NMR (125.7 MHz, CDCl₃): δ 156.6 (C), 153.0 (C), 152.9 (C), 151.9 (C), 150.6 (C), 149.4 (2×C), 136.2 (C), 131.8 (C), 116.9 (C), 85.0 (2×C), 83.3 (C), 82.8 (C), 82.49 (C), 82.46 (C), 73.3 (CH), 71.5 (CH), 67.2 (CH), 64.5 (CH₂), 51.3 (CH), 28.0, 27.72, 27.68, 27.6 (4×CH₃). MS-ESI+ for C₄₀H₆₄N₅O₁₇ ⁺, (M+H)⁺calcd. 886.4287. found 886.4289.

C. Preparation of Compound 4a, 4b or 4c

Step 1—The first fraction from B above containing mainly compounds 2a or 2b (1.46 g, 1.481 mmol) was dissolved in MeOH (29 mL) and sodium methoxide in MeOH (1M, 8.14 mL, 8.14 mmol) added. After leaving at ambient temperature for 20 h the solution was neutralized with Dowex 50WX8 (H⁺) resin then the solids filtered off and the solvent evaporated.

Step 2—The second fraction from B above containing mainly 3a, 3b or 3c (2.68 g, 3.02 mmol) was dissolved in MeOH (54 mL) and sodium methoxide in MeOH (1M, 12.10 mL, 12.10 mmol) added. After leaving at ambient temperature for 20 h the solution was neutralized with Dowex 50WX8 (H⁺) resin then the solids filtered off and the solvent evaporated.

The products from step 1 and step 2 above were combined and chromatographed on silica gel (gradient of 0 to 15% MeOH in CHCl₃) to give 4a, 4b or 4c as a cream colored solid (1.97 g). ¹H NMR (500 MHz, DMSO d₆): δ 12.67 (br s, exchanged D₂O, 1H), 5.48 (d, J=5.2 Hz, exchanged D₂O, 1H), 5.43 (t, J=˜1.9 Hz, after D₂O exchange became a d, J=1.9 Hz, 1H), 5.00 (br s, exchanged D₂O, 1H), 4.62 (d, J=5.7 Hz, exchanged D₂O, 1H), 4.27 (d, J=6.0 Hz, exchanged D₂O, 1H), 3.89 (dt, J=5.2, 3.8 Hz, after D₂O became a t, J=3.9 Hz, 1H), 3.62 (dd, J=6.0, 3.7 Hz, after D₂O exchange became a d, J=3.7 Hz, 1H), 3.52-3.39 (m, 4H), 1.42 (s, 9H), 1.41 (s, 18H). ¹³C NMR (125.7 MHz, DMSO d₆): δ 157.9 (C), 151.1, (C), 149.8 (2×C), 134.6 (C), 131.4 (C), 118.8 (C), 83.5 (2×C), 81.3 (C), 78.2 (CH), 76.5 (CH), 68.1 (CH), 66.8 (CH), 60.6 (CH₂), 54.4 (CH), 27.9 (CH₃), 27.6 (2×CH₃). MS-ESI+ for C₂₅H₄₀N₅O₁₁ ⁺, (M+H)⁺ calcd. 586.2719. found 586.2717.

D. Preparation of Compound 5a, 5b or 5c

Compound 4a, 4b or 4c (992 mg, 1.69 mmol) was dissolved in anhydrous pyridine and concentrated. The residue was dissolved in anhydrous CH₂Cl₂(10 mL) and pyridine (5 mL) under a nitrogen atmosphere and the solution was cooled to −42° C. in an acetonitrile/dry ice bath. Methyl dichlorophosphate (187 μL, 1.86 mmol) was added dropwise and the mixture was stirred for 2 h 20 min. Water (10 mL) was added to the cold solution which was then removed from the cold bath and diluted with ethyl acetate (50 mL) and saturated NaCl solution (30 mL). The organic portion was separated and washed with saturated NaCl solution. The combined aqueous portions were extracted twice further with ethyl acetate and the combined organic portions were dried over MgSO₄and concentrated. Purification by silica gel flash column chromatography (eluting with 2-20% methanol in ethyl acetate) gave the cyclic methyl phosphate 5a, 5b or 5c (731 mg, 65%). ¹H NMR (500 MHz, CDCl₃,): δ 11.72 (bs, exchanged D₂O, 1H), 5.63 (t, J=1.8 Hz, 1H), 5.41 (s, exchanged D₂O, 1H), 4.95 (d, J=3.2 Hz, 1H), 4.70 (dt, J=12.4, 1.8 Hz, 1H), 4.42 (dd, J=22.1, 12.1 Hz, 1H). 4.15 (q, J=3.7 Hz, 1H), 3.82 (s, 1H), 3.75 (s, 1H), 3.58 (d, J=11.7 Hz, 3H), 2.10 (bs, exchanged D₂₀, 1H+H₂O), 1.50 (s, 9H), 1.46 (s, 18H). ¹³C NMR (125.7 MHz, CDCl₃, centre line δ 77.0): δ 157.5 (C), 151.2 (C), 149.6 (2×C), 134.5 (C), 132.3 (C), 117.6 (C), 84.7 (2×C), 82.8 (C), 77.3 (CH), 74.8 (d, J=4.1 Hz, CH), 69.7 (CH₂), 68.8 (d, J=4.1 Hz, CH), 68.6 (d, J=5.9 Hz, CH), 56.0 (d, J=7.4 Hz, CH₃), 51.8 (CH), 28.1 (CH₃), 27.8 (CH₃). MS-ESI+ for C₂₆H₄₀N₅NaO₁₃P⁺ (M+Na)⁺, calcd. 684.2252. found 684.2251.

E. Preparation of Compound 6a, 6b or 6c

Compound 5a, 5b or 5c (223 mg, 0.34 mmol) was dissolved in anhydrous CH₂Cl₂(7 mL) under a nitrogen atmosphere. Anhydrous DMSO (104 μL, 1.46 mmol) was added and the solution was cooled to −78° C. Trifluoroacetic anhydride (104 μL, 0.74 mmol) was added dropwise and the mixture was stirred for 40 min. N,N-diisopropylethylamine (513 μL, 2.94 mmol) was added and the stirring was continued for 50 min at −78° C. Saturated NaCl solution (20 mL) was added and the mixture removed from the cold bath and diluted with CH₂Cl₂(30 mL). Glacial acetic acid (170 μL, 8.75 mmol) was added and the mixture was stirred for 10 min. The layers were separated and the aqueous phase was washed with CH₂Cl₂(10 mL). The combined organic phases were washed with 5% aqueous HCl, 3:1 saturated NaCl solution:10% NaHCO₃solution and saturated NaCl solution successively, dried over MgSO₄, and concentrated to give compound 6a, 6b or 6c (228 mg, quant.) of suitable purity for further use. ¹H NMR (500 MHz, CDCl₃): δ 5.86 (m, 1 H), 5.07 (m, 1 H), 4.70-4.64 (m, 2 H), 4.49-4.40 (m, 1 H), 4.27 (m, 1 H), 3.56, m, 4 H), 1.49 (s, 9 H), 1.46 (s, 18 H) ppm. ¹³C NMR (500 MHz, CDCl₃): δ 157.5 (C), 151.1 (C), 150.6 (2 C), 134.6 (C), 132.7 (C), 116.6 (C), 92.0 (C), 84.6 (2 C), 83.6 (C), 78.0 (CH), 76.0 (CH), 70.4 (CH₂), 67.9 (CH), 56.2 (CH₃) δ6.0 (CH), 28.2 (3CH₃), 26.8 (6 CH₃) ppm. ³¹P NMR (500 MHz, CDCl₃): δ−6.3 ppm.

F. Preparation of compound 7: (4aR,5aR,11aR,12aS)-1,3,2-Dioxaphosphorino[4′,5′:5,6]pyrano[3,2-g]pteridin-10(4H)-one,8-amino-4-a,5a,6,9,11,11a,12,12a-octahydro-2,12,12-trihydroxy-2-oxide

Compound 6a, 6b or 6c (10 mg, 14.8 μmol was dissolved in dry acetonitrile (0.2 mL) and cooled to 0° C. Bromotrimethylsilane (19.2 μL, 148 μmol) was added dropwise and the mixture was allowed to warm to ambient temperature and stirred for 5 h during which time a precipitate formed. HCl_(aq)(10 μl, 37%) was added and the mixture was stirred for a further 15 min. The mixture was centrifuged for 15 min (3000 g) and the resulting precipitate collected. Acetonitrile (0.5 mL) was added and the mixture was centrifuged for a further 15 min. The acetonitrile wash and centrifugation was repeated a further two times and the resulting solid was dried under high vacuum to give compound 7 (4 mg, 75%). ¹H NMR (500 MHz, D₂O): δ 5.22 (d, J=1.6 Hz, 1H), 4.34 (dt, J=13, 1.6 Hz, 1H), 4.29-4.27 (m, 1H), 4.24-4.18 (m, 1H), 3.94 (br m, 1H), 3.44 (t, J=1.4 Hz, 1H). ³¹P NMR (500 MHz, D₂O): δ −4.8 MS-ESI+ for C₁₀H₁₅N₅O₈P⁺, (M+H)⁺calcd. 364.0653. found 364.0652.

Example 2Comparison of Precursor Z (cPMP) Prepared Synthetically to that Prepared from E. Coli in the In vitro Synthesis of Moco

In vitro synthesis of Moco was compared using samples of synthetic precursor Z (cPMP) and cPMP purified from E. coli. Moco synthesis also involved the use of the purified components E. coli MPT synthase, gephyrin, molybdate, ATP, and apo-sulfite oxidase. See U.S. Pat. No. 7,504,095 and “Biosynthesis and molecular biology of the molybdenum cofactor (Moco)” in Metal Ions in Biological Systems, Mendel, Ralf R. and Schwarz, Gunter, Informa Plc, 2002, Vol. 39, pages 317-68. The assay is based on the conversion of cPMP into MPT, the subsequent molybdate insertion using recombinant gephyrin and ATP, and finally the reconstitution of human apo-sulfite oxidase.

As shown in FIG. 1, Moco synthesis from synthetic cPMP was confirmed, and no differences in Moco conversion were found in comparison to E. coli purified cPMP.

Example 3Comparison of Precursor Z (cPMP) Prepared Synthetically to that Prepared from E. coli in the In vitro Synthesis of MPT

In vitro synthesis of MPT was compared using samples of synthetic precursor Z (cPMP) and cPMP purified from E. coli. MPT synthesis also involved the use of in vitro assembled MPT synthase from E. coli. See U.S. Pat. No. 7,504,095 and “Biosynthesis and molecular biology of the molybdenum cofactor (Moco)” in Metal Ions in Biological Systems, Mendel, Ralf R. and Schwarz, Gunter, Informa Plc, 2002, Vol. 39, pages 317-68. Three repetitions of each experiment were performed and are shown in FIGS. 2 and 3.

As shown in FIGS. 2 and 3, MPT synthesis from synthetic cPMP confirmed, and no apparent differences in MPT conversion were found when compared to E. coli purified cPMP. A linear conversion of cPMP into MPT is seen in all samples confirming the identity of synthetic cPMP (see FIG. 2). Slight differences between the repetitions are believed to be due to an inaccurate concentration determination of synthetic cPMP given the presence of interfering chromophores.

Example 4Preparation of Precursor Z (cPMP)

A. Preparation of Starting Materials

B. Introduction of the protected Phosphate

The formation of the cyclic phosphate using intermediate [10] (630 mg) gave the desired product [11] as a 1:1 mixture of diastereoisomers (494 mg, 69%).

C. Oxidation and Overall Deprotection of the Molecule

Oxidation of the secondary alcohol to the gem-diol did prove successful on intermediate [12], but the oxidized product [13] did show significant instability and could not be purified. For this reason, deprotection of the phosphate was attempted before the oxidation. However, the reaction of intermediate [11] with TMSBr led to complete deprotection of the molecule giving intermediate [14]. An attempt to oxidize the alcohol to the gem-diol using Dess-Martin periodinane gave the aromatized pteridine [15].

Oxidation of intermediate [11] with Dess-Martin periodinane gave a mixture of starting material, oxidized product and several by-products. Finally, intermediate [11] was oxidized using the method described Example 1. Upon treatment, only partial oxidation was observed, leaving a 2:1 mixture of [11]/[16]. The crude mixture was submitted to the final deprotection. An off white solid was obtained and analyzed by ¹H-NMR and HPLC-MS. These analyses suggest that cPMP has been produced along with the deprotected precursor [11].

Because the analytical HPLC conditions gave a good separation of cPMP from the major impurities, this method will be repeated on a prep-HPLC in order to isolate the final material.

CLIP

BridgeBio Pharma And Affiliate Origin Biosciences Announces FDA Acceptance Of Its New Drug Application For Fosdenopterin For The Treatment Of MoCD Type A

Application accepted under Priority Review designation with Breakthrough Therapy Designation and Rare Pediatric Disease Designation previously grantedThere are currently no approved therapies for the treatment of MoCD Type A, which results in severe and irreversible neurological injury for infants and children.This is BridgeBio’s first NDA acceptanceSAN FRANCISCO, September 29, 2020 – BridgeBio Pharma, Inc. (Nasdaq: BBIO) and affiliate Origin Biosciences today announced the US Food and Drug Administration (FDA) has accepted its New Drug Application (NDA) for fosdenopterin (previously BBP-870/ORGN001), a cyclic pyranopterin monophosphate (cPMP) substrate replacement therapy, for the treatment of patients with molybdenum cofactor deficiency (MoCD) Type A.The NDA has been granted Priority Review designation. Fosdenopterin has previously been granted Breakthrough Therapy Designation and Rare Pediatric Disease Designation in the US and may be eligible for a priority review voucher if approved. It received Orphan Drug Designation in the US and Europe. This is BridgeBio’s first NDA acceptance.“We want to thank the patients, families, scientists, physicians and all others involved who helped us reach this critical milestone,” said BridgeBio CEO and founder Neil Kumar, Ph.D. “MoCD Type A is a devastating disease with a median survival of less than four years and we are eager for our investigational therapy to be available to patients, who currently have no approved treatment options. BridgeBio exists to help as many patients as possible afflicted with genetic diseases, no matter how rare. We are grateful that the FDA has accepted our first NDA for priority review and we look forward to submitting our second NDA later this year for infigratinib for second line treatment of cholangiocarcinoma.”About Fosdenopterin
Fosdenopterin is being developed for the treatment of patients with MoCD Type A. Currently, there are no approved therapies for the treatment of MoCD Type A, which results in severe and irreversible neurological injury with a median survival between 3 to 4 years. Fosdenopterin is a first-in-class cPMP hydrobromide dihydrate and is designed to treat MoCD Type A by replacing cPMP and permitting the two remaining MoCo synthesis steps to proceed, with activation of MoCo-dependent enzymes and elimination of sulfites.About Molybdenum Cofactor Deficiency (MoCD) Type A
MoCD Type A is an ultra-rare, autosomal recessive, inborn error of metabolism caused by disruption in molybdenum cofactor (MoCo) synthesis which is vital to prevent buildup of s-sulfocysteine, a neurotoxic metabolite of sulfite. Patients are often infants with severe encephalopathy and intractable seizures. Disease progression is rapid with a high infant mortality rate.Those who survive beyond the first few month’s experience profuse developmental delays and suffer the effects of irreversible neurological damage, including brain atrophy with white matter necrosis, dysmorphic facial features, and spastic paraplegia. Clinical presentation that can be similar to hypoxic-ischemic encephalopathy (HIE) or other neonatal seizure disorders may lead to misdiagnosis and underdiagnosis. Immediate testing for elevated sulfite levels and S-sulfocysteine in the urine and very low serum uric acid may help with suspicion of MoCD.About Origin Biosciences
Origin Biosciences, an affiliate of BridgeBio Pharma, is a biotechnology company focused on developing and commercializing a treatment for Molybdenum Cofactor Deficiency (MoCD) Type A. Origin is led by a team of veteran biotechnology executives. Together with patients and physicians, the company aims to bring a safe, effective treatment for MoCD Type A to market as quickly as possible. For more information on Origin Biosciences, please visit the company’s website at www.origintx.com.

About BridgeBio Pharma
BridgeBio is a team of experienced drug discoverers, developers and innovators working to create life-altering medicines that target well-characterized genetic diseases at their source. BridgeBio was founded in 2015 to identify and advance transformative medicines to treat patients who suffer from Mendelian diseases, which are diseases that arise from defects in a single gene, and cancers with clear genetic drivers. BridgeBio’s pipeline of over 20 development programs includes product candidates ranging from early discovery to late-stage development. For more information visit bridgebio.com.

Clinical data

Trade names	Nulibry
Other names	Precursor Z, ALXN1101
License data	US DailyMed: Fosdenopterin
ATC code	None
Legal status
Legal status	US: ℞-only ^[1]
Identifiers
show IUPAC name
CAS Number	150829-29-1
PubChem CID	135894389
DrugBank	DB16628
ChemSpider	17221217
UNII	4X7K2681Y7
KEGG	D11779
ChEMBL	ChEMBL2338675
CompTox Dashboard (EPA)	DTXSID90934067
Chemical and physical data
Formula	C₁₀H₁₄N₅O₈P
Molar mass	363.223 g·mol⁻¹
3D model (JSmol)	Interactive image
hide SMILESNC1=NC(=O)C2=C(N[C@@H]3O[C@@H]4COP(=O)(O)O[C@@H]4C(O)(O)[C@@H]3N2)N1
hide InChIInChI=1S/C10H14N5O8P/c11-9-14-6-3(7(16)15-9)12-4-8(13-6)22-2-1-21-24(19,20)23-5(2)10(4,17)18/h2,4-5,8,12,17-18H,1H2,(H,19,20)(H4,11,13,14,15,16)/t2-,4-,5+,8-/m1/s1Key:CZAKJJUNKNPTTO-AJFJRRQVSA-N

//////////Fosdenopterin hydrobromide, ホスデノプテリン臭化水素酸塩水和物 , ALXN1101 HBr, UNII-X41B5W735T, X41B5W735T, D11780, BBP-870/ORGN001, Priority Review designation, Breakthrough Therapy Designation, Rare Pediatric Disease Designation, Orphan Drug Designation, molybdenum cofactor deficiency, ALXN-1101, WHO 11150, FDA 2021, APPROVALS 2021

#Fosdenopterin hydrobromide, #ホスデノプテリン臭化水素酸塩水和物 , #ALXN1101 HBr, #UNII-X41B5W735T, X41B5W735T, #D11780, #BBP-870/ORGN001, #Priority Review designation, #Breakthrough Therapy Designation, #Rare Pediatric Disease Designation, #Orphan Drug Designation, #molybdenum cofactor deficiency, #ALXN-1101, #WHO 11150, #FDA 2021, #APPROVALS 2021

C1C2C(C(C3C(O2)NC4=C(N3)C(=O)NC(=N4)N)(O)O)OP(=O)(O1)O.O.O.Br

Melphalan flufenamide hydrochloride

March 11, 2021 2:38 am / Leave a comment

.HCl

Melphalan flufenamide hydrochloride

メルファランフルフェナミド塩酸塩;

L-Phenylalanine, 4-[bis(2-chloroethyl)amino]-L-phenylalanyl-4-fluoro-, ethyl ester, hydrochloride

Formula	C24H30Cl2FN3O3. HCl
CAS	380449-54-7
Mol weight	534.8786

FDA APPROVED PEPAXTO, 2021/2/26

Efficacy	Antineoplastic, Alkylating agent
Disease	Multiple myeloma

Ethyl (2S)-2-[(2S)-2-amino-3-{4-[bis(2-chloroethyl)amino]phenyl}propanamido]-3-(4-fluorophenyl)propanoate
J 1
J 1 (prodrug)
L-Melphalanyl-L-p-fluorophenylalanine ethyl ester
Melflufen

Melphalan flufenamide
Pepaxto
Prodrug J 1

Melflufen

Molecular FormulaC₂₄H₃₀Cl₂FN₃O₃
Average mass498.418 Da
SP ROT +33.0 ° Conc: 1.3 g/100mL; chloroform ; 589.3 nm, Oncology Research 2003, V14(3), P113-132

мелфалана флуфенамид [Russian] [INN]ميلفالان فلوفيناميد [Arabic] [INN]氟美法仑 [Chinese] [INN]380449-51-4[RN]
9493Ethyl 4-[bis(2-chloroethyl)amino]-L-phenylalanyl-4-fluoro-L-phenylalaninate
F70C5K4786L-Phenylalanine, 4-[bis(2-chloroethyl)amino]-L-phenylalanyl-4-fluoro-, ethyl ester

Melphalan flufenamide, sold under the brand name Pepaxto, is an anticancer medication used to treat multiple myeloma.^[3]^[4]

The most common adverse reactions include fatigue, nausea, diarrhea, pyrexia and respiratory tract infection.^[3]

Melphalan flufenamide is a peptidase enhanced cytotoxic (PEnC) that exerts a targeted delivery of melphalan in cells with high expression of aminopeptidases, such as aminopeptidase N, which has been described as over-expressed in human malignancies.Aminopeptidase N plays a functional role in malignant angiogenesis.

Melphalan flufenamide was approved for medical use in the United States in February 2021.^[4]^[5]

Medical uses

Melphalan flufenamide is indicated in combination with dexamethasone for the treatment of adults with relapsed or refractory multiple myeloma, with relapsed or refractory multiple myeloma who have received at least four prior lines of therapy and whose disease is refractory to at least one proteasome inhibitor, one immunomodulatory agent, and one CD-38 directed monoclonal antibody.^[3]^[4]

Metabolism

Melphalan flufenamide is metabolized by aminopeptidase hydrolysis and by spontaneous hydrolysis on N-mustard.^[6] Its biological half-life is 10 minutes in vitro.

Origin and development

Melphalan flufenamide is a peptidase enhanced cytotoxic (PEnC) with a targeted delivery within tumor cells of melphalan, a widely used classical chemotherapeutic belonging to a group of alkylating agents developed more than 50 years ago. Substantial clinical experience has been accumulated about melphalan since then. Numerous derivatives of melphalan, designed to increase the activity or selectivity, have been developed and investigated in vitro or in animal models.^[7] Melphalan flufenamide was synthesized, partly due to previous experience of an alkylating peptide cocktail named Peptichemio^[8] and its anti-tumor activity is being investigated.

Pharmacology

Compared to melphalan, melphalan flufenamide exhibits significantly higher in vitro and in vivo activity in several models of human cancer.^[9]^[10]^[11]^[12]^[13]^[14]^[15]^[16] A preclinical study, performed at Dana–Farber Cancer Institute, demonstrated that melphalan flufenamide induced apoptosis in multiple myeloma cell lines, even those resistant to conventional treatment (including melphalan).^[17] In vivo effects in xenografted animals were also observed, and the results confirmed by M Chesi and co-workers – in a unique genetically engineered mouse model of multiple myeloma – are believed to be predictive of clinical efficacy.^[18]

Structure

Chemically, the drug is best described as the ethyl ester of a dipeptide consisting of melphalan and the amino acid derivative para-fluoro-L-phenylalanine.

Pharmacokinetics

Pharmacokinetic analysis of plasma samples showed a rapid formation of melphalan; concentrations generally exceeded those of melphalan flufenamide during ongoing infusion. Melphalan flufenamide rapidly disappeared from plasma after infusion, while melphalan typically peaked a few minutes after the end of infusion. This suggests that melphalan flufenamide is rapidly and widely distributed to extravasal tissues, in which melphalan is formed and thereafter redistributed to plasma.^[19]

This rapid disappearance from plasma is likely due to hydrolytic enzymes.^[20] The Zn(2+) dependent ectopeptidase (also known as alanine aminopeptidase), degrades proteins and peptides with a N-terminal neutral amino acid. Aminopeptidase N is frequently overexpressed in tumors and has been associated with the growth of different human cancers suggesting it as a suitable target for anti-cancerous therapy.^[21]

Adverse effects

In a human Phase 1 trial, no dose-limiting toxicities (DLTs) were observed at lower doses. At doses above 50 mg, reversible neutropenias and thrombocytopenias were observed, and particularly evident in heavily pretreated patients.^[22] These side-effects are shared by most chemotherapies, including alkylating agents in general.

Drug interactions

No drug interaction studies have been reported. Several in vitro studies indicate that melphalan flufenamide may be successfully combined with standard chemotherapy or targeted agents.^[23]^[24]

Therapeutic efficacy

In a Phase 1/2 trial, in solid tumor patients refractory to standard therapy, response evaluation showed disease stabilization in a majority of patients.^[25]^[26] In relapsed and refractory multiple-myeloma (RRMM) patients, promising activity was seen in heavily pre-treated RRMM patients where conventional therapies had failed; the median Progression-Free Survival was 9.4 months and the Duration of Response was 9.6 months.^[27] An overall response rate of 41% and a clinical benefit rate of 56% were also shown, with similar results seen across patient populations regardless of their refractory status. Hematologic toxicity was common, but manageable with cycle prolongations, dose modifications and supportive therapy, and non-hematologic treatment-related adverse events were infrequent.

History

Efficacy was evaluated in HORIZON (NCT02963493), a multicenter, single-arm trial.^[3] Eligible patients were required to have relapsed refractory multiple myeloma.^[3] Patients received melphalan flufenamide 40 mg intravenously on day 1 and dexamethasone 40 mg orally (20 mg for patients ≥75 years of age) on day 1, 8, 15 and 22 of each 28-day cycle until disease progression or unacceptable toxicity.^[3] Efficacy was evaluated in a subpopulation of 97 patients who received four or more prior lines of therapy and were refractory to at least one proteasome inhibitor, one immunomodulatory agent, and a CD38-directed antibody.^[3]

The application for melphalan flufenamide was granted priority review and orphan drug designations.^[3]

Society and culture

Names

Melphalan flufenamide is the International nonproprietary name (INN).^[28]

PAPER

Organic Process Research & Development (2019), 23(6), 1191-1196.

https://pubs.acs.org/doi/pdf/10.1021/bk-2020-1369.ch005

Ethyl (2S)-2-[(2S)-2-amino-3-[bis-(2-chloroethyl)amino]phenyl]propaneamido]-3-(4-fluorophenyl)propanoate hydrochloride, (melphalan flufenamide or Melflufen), is an alkylating agent intended for the treatment of multiple myeloma. Initially only milligram quantities were synthesized, following a route starting from pharmaceutical-grade melphalan. Along with the pharmaceutical development, adjustments were made to the original medicinal chemistry route. This resulted in material for early clinical trials, but it became obvious that further development was necessary. Development resulted in a route in which two phenyl alanine derivatives were coupled to give a dipeptide. This intermediate was further manipulated to give an aniline which could be converted into the desired compound melflufen. The aniline derivative was converted to the corresponding N,N–bis-chloroethylaniline using chloroacetic acid and borane. Deprotection and conversion to the hydrochloride gave melflufen in good yield and excellent purity. Production was performed without chromatography at multi-kilogram scale to supply the API for Phase III studies and commercial validation batches.

PAPER

Antineoplastics

R.S. Vardanyan, V.J. Hruby, in Synthesis of Essential Drugs, 2006

Melphalan

Melphalan, l-3-[p-[bis-(2-chloroethyl)amino]phenyl]alanine (30.2.1.13), is a structural analog of chlorambucil in which the butyric acid fragment is replaced with an aminoacid fragment, alanine. This drug is synthesized from l-phenylalanine, the nitration of which with nitric acid gives 4-nitro-l-phenylalanine (30.2.1.8). Reacting this with an ethanol in the presence of hydrogen chloride gives the hydrochloride of 4-nitro-l-phenylalanine ethyl ester (30.2.1.9), the amino group of which is protected by changing it to phthalamide by a reaction with succinic anhydride to give 30.2.1.10. The nitro group in this molecule is reduced to an amino group using palladium on calcium carbonate as a catalyst. The resulting aromatic amine (30.2.1.11) is then reacted with ethylene oxide, which forms a bis-(2-hydroxyethyl)-amino derivative (30.2.1.12). The hydroxy groups in this molecule are replaced with chlorine atoms upon reaction with thionyl chloride, after which treatment with hydrochloric acid removes the phthalamide protection, giving melphalan (30.2.13) [47–50].

Melaphalan is used intravenously and orally to treat multiple myeloma and cancers of the breast, neck, and ovaries. A synonym of this drug is alkeran.

The racemic form of this drug, d,l-3-[p-[bis-(2-chloroethyl)amino]phenyl]alanine, is also widely used under the name sarcolysine or racemelfalan.
PATENT WO 2001096367PAPEROncology Research (2003), 14(3), 113-132PATENTWO 2016180740https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2016180740

Alkylating agents, such as drugs derived from nitrogen mustard, that is bis(2-chloroethyl)amine derivatives, are used as chemotherapeutic drugs in the treatment of a wide variety of cancers. Melphalan, or p-bis-(2-chloroethyl)-amino-L-phenylalanine (compound (Id), CAS No. 148-82-3), is an alkylating agent which is a conjugate of nitrogen mustard and the amino acid phenylalanine (US 3,032,584). Melphalan is used clinically in the treatment of metastatic melanomas, but has limited efficacy, dose-limiting toxicities and resistance can develop.

Melphalan flufenamide ethyl ester (L-melphalanyl-L-p-fluorophenylalanine ethyl ester, melflufen, compound (Ib)) is a derivative of melphalan conjugated to the amino acid phenylalanine, creating a dipeptide (WO 01/96367):

The monohydrochloride salt of melflufen (L-melphalanyl-L-p-fluorophenylalanine ethyl ester monohydrochloride; hydrochloride salt of (Ib); CAS No. 380449-54-7) is referred to as melflufen hydrochloride.

When studied in cultures of human tumor cells representing approximately 20 different diagnoses of human cancers, including myeloma, melflufen showed 50- to 100-fold higher potency compared with that of melphalan (http://www.oncopeptides.se/products/melflufen/ accessed 26 March 2015). Data disclosed in Arghya, et al, abstract 2086 “A Novel Alkylating Agent Melphalan Flufenamide Ethyl Ester Induces an Irreversible DNA Damage in Multiple Myeloma Cells” (2014) 5^th ASH Annual Meeting and Exposition, suggest that melflufen triggers a rapid, robust and irreversible DNA damage, which may account for its ability to overcome melphalan-resistance in multiple myeloma cells. Melflufen is currently undergoing phase I/IIa clinical trials in multiple myeloma.

A process for preparing melflufen in hydrochloride salt form is described in WO 01/96367, and is illustrated in Scheme 1, below. In that process N-tert-butoxycarbonyl-L-melphalan is reacted with p-fluorophenylalanine ethyl ester to give N-tert-butoxycarbonyl-L-melphalanyl-L-p-fluorophenylalanine ethyl ester. After purification by gradient column chromatography the yield of that step is 43%.

Scheme 1. Current route to melflufen (in hydrochloride salt form)

As shown in Scheme 1, the known process for preparing melflufen (in hydrochloride salt form) uses the cytotoxic agent melphalan as a starting material, and melflufen is synthesised in a multistep sequence. Melphalan is highly toxic, thus the staring materials and all of the intermediates, and also the waste stream generated, are extremely toxic. That is a major disadvantage in terms of safety, environmental impact and cost when using the process on a large scale. Therefore, an improved and safer method is highly desired, especially for production of melflufen on a large scale. Further, the purity of commercially available melphalan is poor due to its poor stability, the yield in each step of the process is poor, and purity of the final product made by the known process is not high.

A process for preparing melphalan is described in WO 2014/141294. In WO 2014/141294 the step to introduce the bis(2-chloroethyl) group into the molecule comprises conversion of a primary phenyl amine to a tertiary phenyl amine diol, by reaction with ethylene oxide gas. This gives a 52.6% yield. The amine diol is then converted to a bis(2-chloroethyl) phenylamine by reaction with phosphoryl chloride. Using ethylene oxide, or chloroethanol, to convert an aromatic amine to the corresponding bis-(2-hydroxy ethyl) amine, followed by

chlorination of that intermediate, is a common technique for producing aromatic bis-(2-chloroethyl) amines. It is also known to start from a chloroarene and let it undergo a SNAr-reaction with diethanolamine. The present inventors have applied those methods to produce melflufen (in its salt form), shown in Scheme 2 below.

Scheme 2. Alternative pathways to melflufen

The inventors have found that using ethylene oxide in THF (route (a) of Scheme 2), no alkylation occurs at 55 °C; increasing the temperature to 60 °C lead to the dialkylated intermediate being formed, but the reaction was very slow. To increase yield and reaction rate the reaction would require high temperatures, but this would cause increased pressure so that the reaction would need be performed in a pressure reactor. Such conditions are likely lead to formation of side products. Similar reaction conditions but using a 50:50 mixture of ethylene oxide and acetic acid (route (b) of Scheme 2) lead to faster reaction times but formation of side products. Using potassium carbonate and chloroethanol (route (c) of Scheme 2) also lead to formation of side product, possibly due to the chloroethanol undergoing partial trans-esterification with the ethyl ester.

The inventors also attempted chlorination of the di-alkylated compound. Chlorination of the bis-(2-hydroxyethyl) compound (4) of Scheme 2 using thionyl chloride in dichloromethane led to significant de-protected side product formation. Chlorination of the bis-(2-hydroxyethyl) compound (4) of Scheme 2 using POCl₃ required high temperature and long

reaction times. In addition, both thionyl chloride and POCl₃ are challenging to handle at large scale due to safety concerns. The inventors also converted the bis-(2-hydroxyethyl) compound (4) of Scheme 2 to the corresponding dimesylate by treatment with methanesulfonyl chloride and triethylamine. The dimesylate was treated then with sodium chloride in DMF at 120 °C. However, the crude product of this reaction contained significant side products making this route unsuitable to be used economically at scale.

In summary, none of these routes were found to be suitable for large scale production of high purity melflufen. They do not work well for the synthesis of melflufen, resulting in poor yields and are inefficient. Further, the routes shown in Scheme 2 require multiple steps to form the N, N-bis-chloroethyl amine and use toxic reagents.

Example 1 – Synthesis of compound (VIc)

To a reactor with overhead stirring, equipped with nitrogen inlet and reflux condenser, was charged Boc-nitrophenylalanine (compound (IVc)) (35.0 g, 112.8 mmol, 1 eq.), followed by acetone (420 mL), N-methylmorpholine (43.4 mL, 394.8 mmol, 3.5 eq.), fluoro-L-phenylalanine ethyl ester hydrochloride (compound (V)) (28.5 g, 115 mmol, 1.02 eq.), EDC (23.8 g, 124.1 mmol, 1.1 eq.) and HOBt·H₂O (1.7 g, 11.3 mmol, 0.1 eq.). The slurry was stirred at room temperature for 18.5 h which led to full consumption of compound (IVc) according to HPLC. Water (180 mL) and 2-MeTHF (965 mL) were charged. Approximately 640 g solvent was then removed by evaporation (T_J: 35 °C) from the clear two phase orange mixture. 360 mL 2-MeTHF was then added and evaporated off twice. The water phase was acidified to pH 3 via addition of 58 mL 2 M sulfuric acid. The organic layer was heated to 35-40 °C and was then sequentially washed with water (90 mL), twice with saturated aqueous NaHCO₃ solution (90 mL) and then brine (90 mL) and finally water (90 mL). To the 2-MeTHF dissolved product was added heptane (270 mL) drop wise at 35-40 °C before the mixture was allowed to reach room temperature overnight with stirring. Another 135 mL heptane was added drop wise before the beige slurry was cooled to 10 °C. The product was isolated and was rinsed with 100 mL cold 2-MeTHF/heptane 6/4. Product compound (VIc) was stored moist (82.5 g). A small sample of the product was analyzed by limit of detection (LOD) which revealed the solid to contain 43.8% solvent residues. Based on this, the purified product was obtained in a yield of 82 %. The purity was determined by HPLC to be: 99.4 area%.

¹ H-NMR (300 MHz, DMSO-D₆) δ 8.48 (broad d, 1H, J=7.5 Hz), 8.16 (2H, d, J=8.7 Hz), 7.55 (2H, d, J=9 Hz), 7.28 (2H, dd, J=8,7, 8.1 Hz), 7.12-7.02 (3H, m), 4.49 (1H, dd, J=14.4, 7.2 Hz), 4.32-4.24 (1 H, m), 4.04 (2H, dd, J=14.4, 7.2 Hz), 3.08-2.95 (3H,m), 2.84 (1H, dd, J=13.2, 10.8 Hz), 1.27 (s, 9H), 1.11 (3H, t, J=7.2Hz)

¹³C-NMR (75 MHz, DMSO-D₆) δ 171.4 (C=O), 171.2 (C=O), 161.2 (C-F, d, J=242.3 Hz), 155.2 (C=O), 146.6 (C), 146.2 (C), 133.1 (C), 131.1 (2 carbon, CH, d, J=8.3 Hz), 130.6 (2 carbon, CH), 123.1 (C), 114.9 (2 carbon, CH, J=20.4 Hz), 78.1 (C), 60.6 (CH₂), 55.1 (CH), 53.6 (CH), 37.3 (CH₂), 35.9 (CH₂), 28.0 (3 carbons, CH₃), 14.0 (CH₃)

Example 2 – Synthesis of compound (IIc)

To a hydrogenation autoclave was added wet solid product compound (VIc) (approximately 4.9 g dry weight, 9.7 mmol, 1 eq.), 2-MeTHF (75 mL) and 3 w/w% of a 5% Pd/C-catalyst (147 mg, 50% moist). The reaction mixture was degased with nitrogen and then 1 barg hydrogen gas was charged. Stirring was set to 600 rpm and T_J to 36 °C. The reaction was completed in four hours, The hydrogenation autoclave was rinsed with 10 mL 2-MeTHF and the rinsing portion was added to the reaction solution in the E-flask. Charcoal (250 mg, 5 wt%) was then added and the resulting mixture was stirred for 15 minutes at room temperature before it was filtered. The filter was rinsed with 10 mL 2-MeTHF and the rinsing portion was added to the filter. The light yellow/pink filtrate contained white precipitated product. The slurry was heated to approximately 40 °C to dissolve the solid before heptane (42 mL) was added drop wise during one hour. The heating was turned off and the mixture was allowed to reach room temperature with overnight stirring. Additional 21 mL heptane was the added before the mixture was cooled to approximately 7 °C (ice/water bath). The solid was isolated and was washed through with 10 mL cold 2-MeTHF/heptane 6/4. The moist solid (5.7 g) was vacuum dried at 35 °C overnight which gave a dry weight of

compound (IIc) of 4.2 g which corresponds to a yield of 91 %. The purity was determined by HPLC to be 99.1 area%.

¹H-NMR (300 MHz, DMSO-D₆) δ 8.26 (1H, d, J=7.5Hz), 7.26 (dd, 2H, J=8.1, 5.7 Hz), 7.09 (2H, t, J=8.7 Hz), 6.86 (2H, d, J=8.1 Hz), 6.71 (1H, d, J=8.7 Hz), 6.45 (1H, d, J=8.1 Hz), 4.87 (2H, s), 4.45 (1H, dd, J=14.4, 7.5 Hz), 4.07-4.00 (3H, m), 3.06-2.91 (2H, m), 2.71 (1H, dd, J=13.8, 3.9 Hz), 2.54-2.46 (1H, m), 1.31 (s, 9H), 1.11 (3H, t, J=6.9 Hz).

¹³C-NMR (75 MHz, DMSO-D₆) δ 171.4 (C=O), 171.2 (C=O), 161.2 (C-F, d, J=242.3 Hz), 155.1 (C=O), 146.9 (C), 133.2 (C, d, J=3.0 Hz), 131.1 (2 carbon, CH, d, J=8.3 Hz), 129.5 (2 carbon, CH), 124.8 (C), 114.8 (2 carbon, CH, J=21.1 Hz), 113.6 (2 carbon, CH), 77.9 (C), 60.5 (CH₂), 56.0 (CH), 53.5 (CH), 36.7 (CH₂), 35.9 (CH₂), 28.1 (3 carbons, CH₃), 13.9 (CH₃)

The present inventors have repeated Example 2 several times using crude compound (VIc) or recrystallised compound (VIc) (purity: 99.1 area%) as starting material and varying various reaction conditions, e.g. pressure of H₂, w/w% of Pd/C, solvent and temperature. The crude purity (97.2 area%) was a slightly higher when recrystallized compound (VIc) was used as starting material than when using crude compound (VIc), in which case the crude purity is generally 95-96 area%. Final yield and purity is also slightly higher than when starting from crude compound (VIc) (98-98.5 area%).

The present inventors have also repeated Example 2 several times varying the Pd/C w/w%, temperature, pressure of H₂ and concentration using 2-MeTHF as the solvent. A high conversion of Compound (VIc) (>99.5 area%) was achieved for Pd/C w/w% from 3 to 6 bar; temperature ranges from 30 to 40 °C, H₂ pressure from 1 to 6 barg, and for varying reaction concentrations. The resulting crude purity was similar in all attempts (95.3-96.2 area%), as was the purity of the isolated product after crystallization from 2-MeTHF/heptane (98.0-98.5 area%).

Example 3 – Preparation of compound (IIIc)

(i) carried out using BH₃SMe₂ in the presence of chloroacetic acid salt

In a 0.5 L dried reactor with overhead stirrer, compound (IIc) (6.99 g, 14.76 mmol) was added, followed by anhydrous tetrahydrofuran (46 mL), chloroacetic acid (36.3 g, 383.8 mmol), chloroacetic acid sodium salt (17.2 g, 147.6 mmol) at T_I=5-13°C. A solution of

BH₃SMe₂ (14.6 g, 191.9 mmol, 18.2 mL) was then added over 45 minutes. After the addition, the reaction temperature was adjusted to T_I=25-30°C and kept for 2 hr after reaching this temperature. The reaction was slowly quenched with ethanol (17.7 g, 383.8 mmol, 22.4 mL) and was stirred overnight at T_J=5°C and then slowly diluted with distilled water (138 mL) to precipitate the product, compound (IIIc). The temperature was adjusted to T_I=15°C and the stirring rate was increased before addition of a solution of aqueous K₂CO₃ (8.0 M, 27 mL) to pH = 7.0-7.5. The reaction slurry was collected on a filter and reaction vessel and filter-cake were washed with water (2×40 mL). The filter-cake was re-slurred in water (200 mL) for 1 hr at T_J=20°C and then filtered again. Washing with water (50 mL), followed by drying at T_J=35°C under high vacuum, produced the crude white product, compound (IIIc), in 7.85 g (88.8%) uncorrected yield. HPLC purity 97.5 area %.

Crude compound (IIIc) (7.5 gram) prepared according to the described procedure was charged to a reactor and washed down with 2-MeTHF (80 mL). Heating at T_J=50°C dissolved the substance. Heptane (80 mL) was added with stirring at T_I=45-50°C and then stirred before adjusting the temperature to T_J=10°C. The precipitated solid was collected by filtration and dried at T_J=35°C under high vacuum which produced white product, compound (IIIc), in 6.86 g (91.5%). HPLC purity 99.1 area %.

¹H-NMR (300 MHz, DMSO-D₆) δ 8.30 (1H, d, J=7.8 Hz), 7.26 (2H, dd, J=8.1, 6 Hz), 7.09-7.05 (3H, m), 6.79 (1H, d, J=8.9 Hz), 6.63 (2H, d, J=8.4 Hz), 4.49-4.42 (1H, dd, J=14.7, 7.5 Hz), 4.07-3.99 (3H, m), 3.68 (8H, s), 3.06-2.91 (2H, m), 2.76 (1H, dd, J=13.8, 4.2 Hz), 2.56 (1H, m), 1.29 (9H, s), 1.1 (3H, t, J=6.6 Hz)

¹³C-NMR (75 MHz, DMSO-D₆) δ 172.1 (C=O), 171.3 (C=O), 161.2 (C-F, d, J=242.3 Hz), 155.2 (C=O), 144.7 (C), 133.2 (C, d, J=3.0 Hz), 131.1 (2 carbon, CH, d, J=7.5 Hz), 130.2 (2 carbon, CH), 126.1 (C), 114.9 (2 carbon, CH, J=21.1 Hz), 111.6 (2 carbon, CH), 78.0 (C), 60.6 (CH₂), 55.9 (CH), 53.5 (CH), 52.2 (CH₂), 41.2 (CH₂), 36.4 (CH₂), 35.9 (CH₂), 28.1 (3 carbons, CH₃), 14.0 (CH₃)

(ii) Carried out using BH₃SMe₂ in the presence of chloroacetic acid salt

In a 0.5 L dried reactor with overhead stirrer, compound (IIe) (7.5 g, 15.84 mmol) was added, followed by 2-MeTHF (150 mL). The mixture was heated to 45 °C to form a clear solution. The solution was cooled to 4 °C and chloroacetic acid (38.9 g, 411.8 mmol), followed by chloroacetic acid sodium salt (18.4 g, 158.4 mmol) was added at T_I=5-13°C. A solution of BH₃SMe₂ (15.6 g, 205.9 mmol, 19.5 mL) was then added over 90 minutes. After the addition, the reaction temperature was adjusted to T_I=20-25°C and kept for 5 hr after reaching this temperature. The reaction was slowly quenched with water at T_I=15-25 °C (150 g, 8333 mmol, 150 mL), pH=3.5 in water phase, and left overnight without stirring at T_I=6 °C.

Product, compound (IIIc), had precipitated out in the organic phase and the temperature was adjusted to T_I=35 °C while stirring, and two clear phases formed. The phases were allowed to separate and the water phase was removed. The organic phase was washed three times with 20% NaCl(aq). pH in the three water phases were: 1.7, 1.1, and 1.1. After the removal of the third water phase, the organic phase was transferred to a round bottom flask and concentrated to half its volume on an evaporator. Product, compound (IIIc), started to precipitate out and the product slurry was allowed to mature at 6 °C for 19 hr. The slurry was collected on a filter and round bottom flask and filter-cake were washed with 2-MeTHF:n-heptane (2×40 mL), followed by drying at T_J=35 °C under high vacuum, to produce the crude white product, compound (IIIc), in 8.3 g (87.6%) uncorrected yield. HPLC purity 99.4 area % .

(iii) Carried out using borane-tetrahydrofuran in the presence of chloroacetic acid salt

In a 100 mL dried round bottom flask with magnet stirrer bar, compound (IIc) (0.75 g, 1.58 mmol) was added under a slow nitrogen flow followed by anhydrous tetrahydrofuran (6 mL), chloroacetic acid (3.89 g, 41.2 mmol), and chloroacetic acid sodium salt (1.84 g, 15.8 mmol). At T_I=5-13°C °C a 1 M solution of BH₃THF (20.6 mmol, 20.6 mL) was added over 30

_minutes. After the addition the reaction temperature was adjusted between T_I=23-28 °C and kept for 2 hr after reaching this temperature. In process control sample (HPLC) indicated in-complete reaction and the jacket temperature was set to T_J=40°C and when the internal temperature reached T_I=40°C the reaction was kept at this temperature for 2 hr when in-process sample (HPLC) showed 6.7 area% starting material, 7.1% acylation adduct

(impurity) and 84.1% compound (IIIc). The reaction was progressed at T_I=23°C and left for 4 days before slowly quenched with ethanol (2.4 g, 3 mL). Water (100 mL) was added and the pH adjusted with 1 M aqueous K₂CO₃ to pH 7. The reaction slurry was collected on a filter and reaction vessel and filter-cake were washed with water (2×20 mL) followed by drying at T_J=35°C under high vacuum produced the crude colorless product in 0.85 g (89.6%) uncorrected yield. HPLC purity was 94.3 area %, with one major impurity attributed to a chloroacylation adduct of the starting material in 3.8 area %.

(iv) Carried out using BH₃SMe₂ without addition of chloroacetic acid salt

In a 100 mL dried round bottom flask with magnet stirrer bar, compound (IIc) (0.75 gram, 1.58 mmol) was added under a slow nitrogen flow followed by anhydrous tetrahydrofuran (6 mL) and chloroacetic acid (3.89 g, 41.2 mmol). At T_I=5-16°C a solution of BH₃SMe₂ (1.56 g, 20.6 mmol, 2.0 mL) was added over 30. After the addition the reaction temperature was adjusted between T_I=25°C and kept for 2.5 h after reaching this temperature. A process control sample (HPLC) indicated melflufen (Compound (Ib)), the Boc-deprotected form of Compound (IIIc), in 66 area %. The reaction was slowly quenched with ethanol (2.9 g, 3.7 mL). The pH of the reaction was adjusted with 1 M aqueous K₂CO₃ solution to pH=8, followed by addition of EtOAc (40 mL). Layers were separated and the aqueous layer re-extracted with EtOAc (50 mL). The organic layers were combined and reduced at <30 mbar / 35°C to an oil. The oil was re-distilled from EtOAc (30 mL) twice and the residue was dried at T_J=23°C / 5 mbar to leave 1.6 g brownish oil. HPLC purity of Compound (Ib) was 66.1 area %.

Example 4 – Preparation of compound (Ib) as hydrochloride salt

Boc-melflufen (compound (IIIc)) (5.0 g, 8.3 mmol) was charged to a round bottomed flask, equipped with magnet stirrer bar, and nitrogen inlet. 1.3 M HCl (anhydrous) in ethanol (64 mL, 83.5 mmol, 10 eq.) was added. After 19 h the conversion was 99.4%. The solvents were partially distilled at T_J=33°C on a rotary evaporator, followed by the addition of ethanol (18 mL). This was repeated twice. Seed crystals were added and after 30 minutes product had precipitated. The slurry was stirred for 21 h and was then concentrated. Methyl tert-butyl ether (MTBE) (108 mL) was added at room temperature with an even rate over 30 minutes. After 100 minutes of stirring at room temperature the precipitate was collected by vacuum filtration and washed with 2×25 mL ethanol: MTBE (1:6). Drying was performed overnight at T_J=35°C / 5 mbar in vacuum oven. Yield of compound (Ib) in the form of its hydrochloride salt, 4.0 g (90%). HPLC-purity 98.7 area%.

¹H-NMR (300 MHz, MeOH-D₄) δ 7.26 (2H, dd, J=8.4, 8.1 Hz), 7.17 (2H, d, J=8.4 Hz), 7.02 (2H, dd, J=9, 8.4 Hz), 6.74 (2H, d, J=8.4 Hz), 4.69 (1H, dd, J=7.8, 6.3 Hz), 4.15 (2H, dd, J=14.1, 7.2 Hz), 4.04 (1H, dd, J=8.4, 5.4 Hz), 3.76 (4H, dd, J=6.3, 6 Hz), 3.67 (4H, dd, 6.6, 5.7 Hz), 3.17 (2H, dd, J=14.4, 6 Hz), 3.06-2.88 (2H, m), 1.22 (3H, t, J=7.2 Hz)

¹³C-NMR (75 MHz, MeOH-D₄) δ 172.2 (C=O), 169.8 (C=O), 163.4 (C-F, d, J=244.5 Hz), 147.4 (C), 133.9 (C, d, J=3 Hz), 132.1 (2 carbon, CH, d, J=7.5 Hz), 131.8 (2 carbon, CH), 123.4 (C), 116.2 (2 carbon, CH, d, J=21.9 Hz), 113.7 (2 carbon, CH), 62.6 (CH₂), 55.6 (CH), 55.5 (CH), 54.3 (CH₂), 41.6 (CH₂), 37.6 (CH₂), 37.6 (CH₂), 14.5 (CH₃)

Example 4 was repeated successfully in the presence ethyl acetate and with varying concentrations of HCl from 1.3 M to 2.5 M and at varying temperatures from 6 °C to room temperature.PAPERhttps://pubs.acs.org/doi/10.1021/acs.oprd.9b00116 Organic Process Research & Development (2019), 23(6), 1191-1196.Melflufen is a novel cytostatic currently in phase III clinical trials for treatment of multiple myeloma. Development of a process suitable for production is described. The two key features of the novel method are late introduction of the alkylating pharmacophore and an improved method for formation of the bis-chloroethyl group.

1H NMR spectrum of L-Phenylalanine, 4-[bis(2-chloroethyl)amino]-L-phenylalanyl-4-fluoro-, ethyl ester, hydrochloride (1) (in D4–MeOH).

13C NMR spectrum of L-Phenylalanine, 4-[bis(2-chloroethyl)amino]-L-phenylalanyl-4-fluoro-, ethyl ester, hydrochloride (1) (in D4–MeOH).

References

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^ Berglund, Åke; Ullén, A; Lisyanskaya, A; Orlov, S; Hagberg, H; Tholander, B; Lewensohn, R; Nygren, P; Spira, J; Harmenberg, J; Jerling, M; Alvfors, C; Ringbom, M; Nordström, E; Söderlind, K; Gullbo, J (2015). “First-in-human, phase I/IIa clinical study of the peptidase potentiated alkylator melflufen administered every three weeks to patients with advanced solid tumor malignancies”. Investigational New Drugs. 33 (6): 1232–41. doi:10.1007/s10637-015-0299-2. PMID 26553306. S2CID 8207569.
^ Wickström, M; Haglund, C; Lindman, H; Nygren, P; Larsson, R; Gullbo, J (2008). “The novel alkylating prodrug J1: Diagnosis directed activity profile ex vivo and combination analyses in vitro”. Investigational New Drugs. 26 (3): 195–204. doi:10.1007/s10637-007-9092-1. PMID 17922077. S2CID 19915448.
^ Chauhan, D; Ray, A; Viktorsson, K; Spira, J; Paba-Prada, C; Munshi, N; Richardson, P; Lewensohn, R; Anderson, K. C. (2013). “In vitro and in vivo antitumor activity of a novel alkylating agent, melphalan-flufenamide, against multiple myeloma cells”. Clinical Cancer Research. 19 (11): 3019–31. doi:10.1158/1078-0432.CCR-12-3752. PMC 4098702. PMID 23584492.
^ Berglund, Åke; Ullén, A; Lisyanskaya, A; Orlov, S; Hagberg, H; Tholander, B; Lewensohn, R; Nygren, P; Spira, J; Harmenberg, J; Jerling, M; Alvfors, C; Ringbom, M; Nordström, E; Söderlind, K; Gullbo, J (2015). “First-in-human, phase I/IIa clinical study of the peptidase potentiated alkylator melflufen administered every three weeks to patients with advanced solid tumor malignancies”. Investigational New Drugs. 33 (6): 1232–41. doi:10.1007/s10637-015-0299-2. PMID 26553306. S2CID 8207569.
^ Viktorsson, K; Shah, C. H.; Juntti, T; Hååg, P; Zielinska-Chomej, K; Sierakowiak, A; Holmsten, K; Tu, J; Spira, J; Kanter, L; Lewensohn, R; Ullén, A (2016). “Melphalan-flufenamide is cytotoxic and potentiates treatment with chemotherapy and the Src inhibitor dasatinib in urothelial carcinoma”. Molecular Oncology. 10 (5): 719–34. doi:10.1016/j.molonc.2015.12.013. PMC 5423156. PMID 26827254.
^ https://ash.confex.com/ash/2015/webprogram/Paper85666.html
^ World Health Organization (2012). “International nonproprietary names for pharmaceutical substances (INN): recommended INN: list 67”. WHO Drug Information. 26 (1): 72. hdl:10665/109416.

External links

“Melphalan flufenamide”. Drug Information Portal. U.S. National Library of Medicine.
Clinical trial number NCT02963493 for “A Study of Melphalan Flufenamide (Melflufen) in Combination With Dexamethasone in Relapsed Refractory Multiple Myeloma Patients (HORIZON)” at ClinicalTrials.gov

Clinical data

Trade names	Pepaxto
Other names	Melflufen, 4-[Bis-(2-chloroethyl)amino]-L-phenylalanine-4-fluoro-L-phenylalanine ethyl ester, J1^[1]^[2]
License data	US DailyMed: Melphalan_flufenamide
Legal status
Legal status	US: ℞-only ^[3]
Pharmacokinetic data
Metabolism	Aminopeptidase hydrolysis, Spontaneous hydrolyisis on N-mustard
Elimination half-life	10 min in vitro^{[medical citation needed]}
Identifiers
show IUPAC name
CAS Number	380449-51-4
PubChem CID	9935639
DrugBank	DB16627
ChemSpider	8111267
UNII	F70C5K4786
ChEMBL	ChEMBL4303060
Chemical and physical data
Formula	C₂₄H₃₀Cl₂FN₃O₃
Molar mass	498.42 g·mol⁻¹
3D model (JSmol)	Interactive image
hide SMILESCCOC(=O)[C@H](CC1=CC=C(C=C1)F)NC(=O)[C@H](CC2=CC=C(C=C2)N(CCCl)CCCl)N
hide InChIInChI=1S/C24H30Cl2FN3O3/c1-2-33-24(32)22(16-18-3-7-19(27)8-4-18)29-23(31)21(28)15-17-5-9-20(10-6-17)30(13-11-25)14-12-26/h3-10,21-22H,2,11-16,28H2,1H3,(H,29,31)/t21-,22-/m0/s1Key:YQZNKYXGZSVEHI-VXKWHMMOSA-N

//////////Melphalan flufenamide hydrochloride, Melphalan flufenamide, FDA 2021, APPROVALS 2021, PEPAXTO, メルファランフルフェナミド塩酸塩 , J 1

#Melphalan flufenamide hydrochloride, #Melphalan flufenamide, #FDA 2021, #APPROVALS 2021, #PEPAXTO, メルファランフルフェナミド塩酸塩 , #J 1

Evinacumab

February 26, 2021 9:34 am / 1 Comment on Evinacumab

(Heavy chain)
EVQLVESGGG VIQPGGSLRL SCAASGFTFD DYAMNWVRQG PGKGLEWVSA ISGDGGSTYY
ADSVKGRFTI SRDNSKNSLY LQMNSLRAED TAFFYCAKDL RNTIFGVVIP DAFDIWGQGT
MVTVSSASTK GPSVFPLAPC SRSTSESTAA LGCLVKDYFP EPVTVSWNSG ALTSGVHTFP
AVLQSSGLYS LSSVVTVPSS SLGTKTYTCN VDHKPSNTKV DKRVESKYGP PCPPCPAPEF
LGGPSVFLFP PKPKDTLMIS RTPEVTCVVV DVSQEDPEVQ FNWYVDGVEV HNAKTKPREE
QFNSTYRVVS VLTVLHQDWL NGKEYKCKVS NKGLPSSIEK TISKAKGQPR EPQVYTLPPS
QEEMTKNQVS LTCLVKGFYP SDIAVEWESN GQPENNYKTT PPVLDSDGSF FLYSRLTVDK
SRWQEGNVFS CSVMHEALHN HYTQKSLSLS LGK
(Light chain)
DIQMTQSPST LSASVGDRVT ITCRASQSIR SWLAWYQQKP GKAPKLLIYK ASSLESGVPS
RFSGSGSGTE FTLTISSLQP DDFATYYCQQ YNSYSYTFGQ GTKLEIKRTV AAPSVFIFPP
SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ ESVTEQDSKD STYSLSSTLT
LSKADYEKHK VYACEVTHQG LSSPVTKSFN RGEC
(Disulfide bridge: H22-H96, H140-L214, H153-H209, H232-H’232, H235-H’235, H267-H327, H373-H431, H’22-H’96, H’140-L’214, H’153-H’209, H’267-H’327, H’373-H’431, L23-L88, L134-L194, L’23-L’88, L’134-L’194)

Evinacumab

エビナクマブ (遺伝子組換え)

Immunoglobulin G4, anti-(human protein ANGPTL3 (angiopoietin-like 3)) (human monoclonal REGN1500 heavy chain), disulfide with human monoclonal REGN1500 light chain, dimer

Formula	C6480H9992N1716O2042S46
CAS	1446419-85-7
Mol weight	146081.9345

Protein Sequence

Sequence Length: 1334, 453, 453, 214, 214multichain; modified (modifications unspecified)

FDA APPROVED, 2021/2/11, EVKEEZA

Antihyperlipidemic, Anti-angiopietin like 3

Monoclonal antibody
Treatment of dyslipidemia

REGN 1500
REGN-1500
REGN1500

Sequence:

1EVQLVESGGG VIQPGGSLRL SCAASGFTFD DYAMNWVRQG PGKGLEWVSA51ISGDGGSTYY ADSVKGRFTI SRDNSKNSLY LQMNSLRAED TAFFYCAKDL101RNTIFGVVIP DAFDIWGQGT MVTVSSASTK GPSVFPLAPC SRSTSESTAA151LGCLVKDYFP EPVTVSWNSG ALTSGVHTFP AVLQSSGLYS LSSVVTVPSS201SLGTKTYTCN VDHKPSNTKV DKRVESKYGP PCPPCPAPEF LGGPSVFLFP251PKPKDTLMIS RTPEVTCVVV DVSQEDPEVQ FNWYVDGVEV HNAKTKPREE301QFNSTYRVVS VLTVLHQDWL NGKEYKCKVS NKGLPSSIEK TISKAKGQPR351EPQVYTLPPS QEEMTKNQVS LTCLVKGFYP SDIAVEWESN GQPENNYKTT401PPVLDSDGSF FLYSRLTVDK SRWQEGNVFS CSVMHEALHN HYTQKSLSLS451LGK

Sequence:

1DIQMTQSPST LSASVGDRVT ITCRASQSIR SWLAWYQQKP GKAPKLLIYK51ASSLESGVPS RFSGSGSGTE FTLTISSLQP DDFATYYCQQ YNSYSYTFGQ101GTKLEIKRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV151DNALQSGNSQ ESVTEQDSKD STYSLSSTLT LSKADYEKHK VYACEVTHQG201LSSPVTKSFN RGEC

Sequence:

Sequence Modifications

Type	Location	Description
bridge	Cys-22 – Cys-96	disulfide bridge
bridge	Cys-140 – Cys-214”	disulfide bridge
bridge	Cys-153 – Cys-209	disulfide bridge
bridge	Cys-232 – Cys-232′	disulfide bridge
bridge	Cys-235 – Cys-235′	disulfide bridge
bridge	Cys-267 – Cys-327	disulfide bridge
bridge	Cys-373 – Cys-431	disulfide bridge
bridge	Cys-22′ – Cys-96′	disulfide bridge
bridge	Cys-140′ – Cys-214”’	disulfide bridge
bridge	Cys-153′ – Cys-209′	disulfide bridge
bridge	Cys-267′ – Cys-327′	disulfide bridge
bridge	Cys-373′ – Cys-431′	disulfide bridge
bridge	Cys-23” – Cys-88”	disulfide bridge
bridge	Cys-134” – Cys-194”	disulfide bridge
bridge	Cys-23”’ – Cys-88”’	disulfide bridge
bridge	Cys-134”’ – Cys-194”’	disulfide bridge

PATENTS

WO 2017024062

US 20170305999

Evinacumab, sold under the brand name Evkeeza, is a monoclonal antibody medication for the treatment of homozygous familial hypercholesterolemia (HoFH).^[1]^[2]

Evinacumab is a recombinant human IgG4 monoclonal antibody targeted against angiopoietin-like protein 3 (ANGPTL3) and the first drug of its kind. The ANGPTL family of proteins serve a number of physiologic functions – including involvement in the regulation of lipid metabolism – which have made them desirable therapeutic targets in recent years.² Loss-of-function mutations in ANGPTL3 have been noted to result in hypolipidemia and subsequent reductions in cardiovascular risk, whereas increases in function appear to be associated with cardiovascular risk, and it was these observations that provided a rationale for the development of a therapy targeted against ANGPTL3.³

In February 2021, evinacumab became the first-and-only inhibitor of ANGPTL3 to receive FDA approval after it was granted approval for the adjunctive treatment of homozygous familial hypercholesterolemia (HoFH) under the brand name “Evkeeza”.⁸ Evinacumab is novel in its mechanism of action compared with other lipid-lowering therapies and therefore provides a unique and synergistic therapeutic option in the treatment of HoFH.

Common side effects include nasopharyngitis (cold), influenza-like illness, dizziness, rhinorrhea (runny nose), and nausea. Serious hypersensitivity (allergic) reactions have occurred in the Evkeeza clinical trials.^[2]

Evinacumab binds to the angiopoietin-like protein 3 (ANGPTL3).^[2] ANGPTL3 slows the function of certain enzymes that break down fats in the body.^[2] Evinacumab blocks ANGPTL3, allowing faster break down of fats that lead to high cholesterol.^[2] Evinacumab was approved for medical use in the United States in February 2021.^[2]^[3]

NAME	DOSAGE	STRENGTH	ROUTE	LABELLER	MARKETING START	MARKETING END
Evkeeza	Injection, solution, concentrate	150 mg/1mL	Intravenous	Regeneron Pharmaceuticals, Inc.	2021-02-11	Not applicable
Evkeeza	Injection, solution, concentrate	150 mg/1mL	Intravenous	Regeneron Pharmaceuticals, Inc.	2021-02-11	Not applicable

History

The effectiveness and safety of evinacumab were evaluated in a double-blind, randomized, placebo-controlled, 24-week trial enrolling 65 participants with homozygous familial hypercholesterolemia (HoFH).^[2] In the trial, 43 participants received 15 mg/kg of evinacumab every four weeks and 22 participants received the placebo.^[2] Participants were taking other lipid-lowering therapies as well.^[2]

The primary measure of effectiveness was the percent change in low-density lipoprotein (LDL-C) from the beginning of treatment to week 24.^[2] At week 24, participants receiving evinacumab had an average 47% decrease in LDL-C while participants on the placebo had an average 2% increase.^[2]

The U.S. Food and Drug Administration (FDA) granted the application for evinacumab orphan drug, breakthrough therapy, and priority review designations.^[2] The FDA granted approval of Evkeeza to Regeneron Pharmaceuticals, Inc.^[2]

References

^ Jump up to:^a ^b https://www.accessdata.fda.gov/drugsatfda_docs/label/2021/761181s000lbl.pdf
^ Jump up to:^a ^b ^c ^d ^e ^f ^g ^h ⁱ ^j ^k ^l ^m ⁿ “FDA approves add-on therapy for patients with genetic form of severely”. U.S. Food and Drug Administration (FDA). 11 February 2021. Retrieved 12 February 2021. This article incorporates text from this source, which is in the public domain.
^ “FDA Approves First-in-class Evkeeza (evinacumab-dgnb) for Patients with Ultra-rare Inherited Form of High Cholesterol” (Press release). Regeneron Pharmaceuticals. 11 February 2021. Retrieved 12 February 2021 – via PR Newswire.

External links

“Evinacumab”. Drug Information Portal. U.S. National Library of Medicine.

Monoclonal antibody
Type	Whole antibody
Source	Human
Target	Angiopoietin-like 3 (ANGPTL3)
Clinical data
Trade names	Evkeeza
Other names	REGN1500, evinacumab-dgnb
License data	US DailyMed: Evinacumab
Routes of administration	Intravenous
ATC code	None
Legal status
Legal status	US: ℞-only ^[1]^[2]
Identifiers
CAS Number	1446419-85-7
DrugBank	DB15354
ChemSpider	none
UNII	T8B2ORP1DW
KEGG	D11753
Chemical and physical data
Formula	C₆₄₈₀H₉₉₉₂N₁₇₁₆O₂₀₄₂S₄₆
Molar mass	146083.95 g·mol⁻¹

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

#Evinacumab, #Peptide, #APPROVALS 2021, #FDA 2021, #Monoclonal antibody, #dyslipidemia, #エビナクマブ (遺伝子組換え) , #REGN 1500, #REGN-1500, #REGN1500, #Anthony melvin crasto, #world drug tracker. # new drug approvals, #pharma

Tozinameran, Pfizer–BioNTech COVID‑19 vaccine

February 24, 2021 9:24 am / 2 Comments on Tozinameran, Pfizer–BioNTech COVID‑19 vaccine

SEQUENCE1

gagaauaaac uaguauucuu cuggucccca cagacucaga gagaacccgc51caccauguuc guguuccugg ugcugcugcc ucuggugucc agccagugug101ugaaccugac caccagaaca cagcugccuc cagccuacac caacagcuuu151accagaggcg uguacuaccc cgacaaggug uucagaucca gcgugcugca201cucuacccag gaccuguucc ugccuuucuu cagcaacgug accugguucc251acgccaucca cguguccggc accaauggca ccaagagauu cgacaacccc301gugcugcccu ucaacgacgg gguguacuuu gccagcaccg agaaguccaa351caucaucaga ggcuggaucu ucggcaccac acuggacagc aagacccaga401gccugcugau cgugaacaac gccaccaacg uggucaucaa agugugcgag451uuccaguucu gcaacgaccc cuuccugggc gucuacuacc acaagaacaa501caagagcugg auggaaagcg aguuccgggu guacagcagc gccaacaacu551gcaccuucga guacgugucc cagccuuucc ugauggaccu ggaaggcaag601cagggcaacu ucaagaaccu gcgcgaguuc guguuuaaga acaucgacgg651cuacuucaag aucuacagca agcacacccc uaucaaccuc gugcgggauc701ugccucaggg cuucucugcu cuggaacccc ugguggaucu gcccaucggc751aucaacauca cccgguuuca gacacugcug gcccugcaca gaagcuaccu801gacaccuggc gauagcagca gcggauggac agcuggugcc gccgcuuacu851augugggcua ccugcagccu agaaccuucc ugcugaagua caacgagaac901ggcaccauca ccgacgccgu ggauugugcu cuggauccuc ugagcgagac951aaagugcacc cugaaguccu ucaccgugga aaagggcauc uaccagacca1001gcaacuuccg ggugcagccc accgaaucca ucgugcgguu ccccaauauc1051accaaucugu gccccuucgg cgagguguuc aaugccacca gauucgccuc1101uguguacgcc uggaaccgga agcggaucag caauugcgug gccgacuacu1151ccgugcugua caacuccgcc agcuucagca ccuucaagug cuacggcgug1201uccccuacca agcugaacga ccugugcuuc acaaacgugu acgccgacag1251cuucgugauc cggggagaug aagugcggca gauugccccu ggacagacag1301gcaagaucgc cgacuacaac uacaagcugc ccgacgacuu caccggcugu1351gugauugccu ggaacagcaa caaccuggac uccaaagucg gcggcaacua1401caauuaccug uaccggcugu uccggaaguc caaucugaag cccuucgagc1451gggacaucuc caccgagauc uaucaggccg gcagcacccc uuguaacggc1501guggaaggcu ucaacugcua cuucccacug caguccuacg gcuuucagcc1551cacaaauggc gugggcuauc agcccuacag agugguggug cugagcuucg1601aacugcugca ugccccugcc acagugugcg gcccuaagaa aagcaccaau1651cucgugaaga acaaaugcgu gaacuucaac uucaacggcc ugaccggcac1701cggcgugcug acagagagca acaagaaguu ccugccauuc cagcaguuug1751gccgggauau cgccgauacc acagacgccg uuagagaucc ccagacacug1801gaaauccugg acaucacccc uugcagcuuc ggcggagugu cugugaucac1851cccuggcacc aacaccagca aucagguggc agugcuguac caggacguga1901acuguaccga agugcccgug gccauucacg ccgaucagcu gacaccuaca1951uggcgggugu acuccaccgg cagcaaugug uuucagacca gagccggcug2001ucugaucgga gccgagcacg ugaacaauag cuacgagugc gacaucccca2051ucggcgcugg aaucugcgcc agcuaccaga cacagacaaa cagcccucgg2101agagccagaa gcguggccag ccagagcauc auugccuaca caaugucucu2151gggcgccgag aacagcgugg ccuacuccaa caacucuauc gcuaucccca2201ccaacuucac caucagcgug accacagaga uccugccugu guccaugacc2251aagaccagcg uggacugcac cauguacauc ugcggcgauu ccaccgagug2301cuccaaccug cugcugcagu acggcagcuu cugcacccag cugaauagag2351cccugacagg gaucgccgug gaacaggaca agaacaccca agagguguuc2401gcccaaguga agcagaucua caagaccccu ccuaucaagg acuucggcgg2451cuucaauuuc agccagauuc ugcccgaucc uagcaagccc agcaagcgga2501gcuucaucga ggaccugcug uucaacaaag ugacacuggc cgacgccggc2551uucaucaagc aguauggcga uugucugggc gacauugccg ccagggaucu2601gauuugcgcc cagaaguuua acggacugac agugcugccu ccucugcuga2651ccgaugagau gaucgcccag uacacaucug cccugcuggc cggcacaauc2701acaagcggcu ggacauuugg agcaggcgcc gcucugcaga uccccuuugc2751uaugcagaug gccuaccggu ucaacggcau cggagugacc cagaaugugc2801uguacgagaa ccagaagcug aucgccaacc aguucaacag cgccaucggc2851aagauccagg acagccugag cagcacagca agcgcccugg gaaagcugca2901ggacgugguc aaccagaaug cccaggcacu gaacacccug gucaagcagc2951uguccuccaa cuucggcgcc aucagcucug ugcugaacga uauccugagc3001agacuggacc cuccugaggc cgaggugcag aucgacagac ugaucacagg3051cagacugcag agccuccaga cauacgugac ccagcagcug aucagagccg3101ccgagauuag agccucugcc aaucuggccg ccaccaagau gucugagugu3151gugcugggcc agagcaagag aguggacuuu ugcggcaagg gcuaccaccu3201gaugagcuuc ccucagucug ccccucacgg cgugguguuu cugcacguga3251cauaugugcc cgcucaagag aagaauuuca ccaccgcucc agccaucugc3301cacgacggca aagcccacuu uccuagagaa ggcguguucg uguccaacgg3351cacccauugg uucgugacac agcggaacuu cuacgagccc cagaucauca3401ccaccgacaa caccuucgug ucuggcaacu gcgacgucgu gaucggcauu3451gugaacaaua ccguguacga cccucugcag cccgagcugg acagcuucaa3501agaggaacug gacaaguacu uuaagaacca cacaagcccc gacguggacc3551ugggcgauau cagcggaauc aaugccagcg ucgugaacau ccagaaagag3601aucgaccggc ugaacgaggu ggccaagaau cugaacgaga gccugaucga3651ccugcaagaa cuggggaagu acgagcagua caucaagugg cccugguaca3701ucuggcuggg cuuuaucgcc ggacugauug ccaucgugau ggucacaauc3751augcuguguu gcaugaccag cugcuguagc ugccugaagg gcuguuguag3801cuguggcagc ugcugcaagu ucgacgagga cgauucugag cccgugcuga3851agggcgugaa acugcacuac acaugaugac ucgagcuggu acugcaugca3901cgcaaugcua gcugccccuu ucccguccug gguaccccga gucucccccg3951accucggguc ccagguaugc ucccaccucc accugcccca cucaccaccu4001cugcuaguuc cagacaccuc ccaagcacgc agcaaugcag cucaaaacgc4051uuagccuagc cacaccccca cgggaaacag cagugauuaa ccuuuagcaa4101uaaacgaaag uuuaacuaag cuauacuaac cccaggguug gucaauuucg4151ugccagccac acccuggagc uagcaaaaaa aaaaaaaaaa aaaaaaaaaa4201aaaagcauau gacuaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa4251aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaa

Sequence Modifications

Type	Location	Description
modified base	g-1	m7g
modified base	g-1	3′-me
modified base	a-2	am
uncommon link	g-1 – a-2	5′->5′ triphosphate

Tozinameran

Pfizer–BioNTech COVID-19 vaccine

トジナメラン (JAN);
コロナウイルス修飾ウリジンRNAワクチン;

RNA (recombinant 5′-[1,2-[(3′-O-methyl)m⁷G-(5’→5′)-ppp-Am]]-capped all uridine→N¹-methylpseudouridine-substituted severe acute respiratory syndrome coronavirus 2 secretory signal peptide contg. spike glycoprotein S1S2-specifying plus 5′- and 3′-untranslated flanking region-contg. poly(A)-tailed messenger BNT162b2), inner salt

Nucleic Acid Sequence

Sequence Length: 42841106 a 1315 c 1062 g 801 umodified

APPROVED JAPAN Comirnaty, 2021/2/14

CAS 2417899-77-3

Active immunization (SARS-CoV-2)

Tozinameran is mRNA encoding full length of spike protein analog of SARS-CoV-2

Target Severe acute respiratory syndrome coronavirus 2 spike glycoprotein

Coronavirus disease – COVID-19

FORM	ROUTE	STRENGTH
Injection, suspension	Intramuscular	0.23 mg/1.8mL
Suspension	Intramuscular	30 mcg

NAME	INGREDIENTS	DOSAGE	ROUTE	LABELLER	MARKETING START	MARKETING END
Pfizer-BioNTech Covid-19 Vaccine	Pfizer-BioNTech Covid-19 Vaccine (0.23 mg/1.8mL)	Injection, suspension	Intramuscular	Pfizer Manufacturing Belgium NV	2020-12-12	Not applicable

NAME	DOSAGE	STRENGTH	ROUTE	LABELLER	MARKETING START	MARKETING END
Comirnaty		30 mcg	Intramuscular	Bio N Tech Manufacturing Gmb H	2021-01-06	Not applicable
Pfizer-BioNTech Covid-19 Vaccine	Suspension	30 mcg	Intramuscular	Biontech Manufacturing Gmbh	2020-12-14	Not applicable
Pfizer-BioNTech Covid-19 Vaccine	Injection, suspension	0.23 mg/1.8mL	Intramuscular	Pfizer Manufacturing Belgium NV	2020-12-12	Not applicable

The Pfizer–BioNTech COVID‑19 vaccine (pINN: tozinameran), sold under the brand name Comirnaty,^[13] is a COVID-19 vaccine developed by the German company BioNTech in cooperation with Pfizer. It is both the first COVID-19 vaccine to be authorized by a stringent regulatory authority for emergency use^[14]^[15] and the first cleared for regular use.^[16]

It is given by intramuscular injection. It is an RNA vaccine composed of nucleoside-modified mRNA (modRNA) encoding a mutated form of the spike protein of SARS-CoV-2, which is encapsulated in lipid nanoparticles.^[17] The vaccination requires two doses given three weeks apart.^[18]^[19]^[20] Its ability to prevent severe infection in children, pregnant women, or immunocompromised people is unknown, as is the duration of the immune effect it confers.^[20]^[21]^[22] As of February 2021, it is one of two RNA vaccines being deployed against COVID‑19, the other being the Moderna COVID‑19 vaccine. A third mRNA-based COVID-19 vaccine, CVnCoV, is in late-stage testing.^[23]

Trials began in April 2020; by November, the vaccine had been tested on more than 40,000 people.^[24] An interim analysis of study data showed a potential efficacy of over 90% in preventing infection within seven days of a second dose.^[19]^[20] The most common side effects include mild to moderate pain at the injection site, fatigue, and headache.^[25]^[26] As of December 2020, reports of serious side effects, such as allergic reactions, have been very rare,^[a] and no long-term complications have been reported.^[28] Phase III clinical trials are ongoing: monitoring of the primary outcomes will continue until August 2021, while monitoring of the secondary outcomes will continue until January 2023.^[18]

In December 2020, the United Kingdom was the first country to authorize the vaccine on an emergency basis,^[28] soon followed by the United States, the European Union and several other countries globally.^[29]^[30]^[6]^[31]^[32]

BioNTech is the initial developer of the vaccine, and partnered with Pfizer for development, clinical research, overseeing the clinical trials, logistics, finances and for manufacturing worldwide with the exception of China.^[33] The license to distribute and manufacture in China was purchased by Fosun, alongside its investment in BioNTech.^[34]^[35] Distribution in Germany and Turkey is by BioNTech itself.^[36] Pfizer indicated in November 2020, that 50 million doses could be available globally by the end of 2020, with about 1.3 billion doses in 2021.^[20]

Pfizer has advanced purchase agreements of about US$3 billion for providing a licensed vaccine in the United States, the European Union, the United Kingdom, Japan, Canada, Peru, Singapore, and Mexico.^[37]^[38] Distribution and storage of the vaccine is a logistics challenge because it needs to be stored at temperatures between −80 and −60 °C (−112 and −76 °F),^[39] until five days before vaccination^[38]^[39] when it can be stored at 2 to 8 °C (36 to 46 °F), and up to two hours at temperatures up to 25 °C (77 °F)^[40]^[11] or 30 °C (86 °F).^[41]^[42] In February 2021, Pfizer and BioNTech asked the U.S. Food and Drug Administration (FDA) to update the emergency use authorization (EUA) to permit the vaccine to be stored at between −25 and −15 °C (−13 and 5 °F) for up to two weeks before use.^[43]

Development and funding

Before COVID-19 vaccines, a vaccine for an infectious disease had never before been produced in less than several years, and no vaccine existed for preventing a coronavirus infection in humans.^[44] After the COVID-19 virus was detected in December 2019,^[45] the development of BNT162b2 was initiated on 10 January 2020, when the SARS-CoV-2 genetic sequences were released by the Chinese Center for Disease Control and Prevention via GISAID,^[46]^[47]^[48] triggering an urgent international response to prepare for an outbreak and hasten development of preventive vaccines.^[49]^[50]

In January 2020, German biotech-company BioNTech started its program ‘Project Lightspeed’ to develop a vaccine against the new COVID‑19 virus based on its already established mRNA-technology.^[24] Several variants of the vaccine were created in their laboratories in Mainz, and 20 of those were presented to experts of the Paul-Ehrlich-Institute in Langen.^[51] Phase I / II Trials were started in Germany on 23 April 2020, and in the U.S. on 4 May 2020, with four vaccine candidates entering clinical testing. The Initial Pivotal Phase II / III Trial with the lead vaccine candidate ‘BNT162b2’ began in July. The Phase III results indicating a 95% effectiveness of the developed vaccine were published on 18 November 2020.^[24]

BioNTech received a US$135 million investment from Fosun in March 2020, in exchange for 1.58 million shares in BioNTech and the future development and marketing rights of BNT162b2 in China,^[35] Hong Kong, Macau and Taiwan.^[52]

In June 2020, BioNTech received €100 million (US$119 million) in financing from the European Commission and European Investment Bank.^[53] In September 2020, the German government granted BioNTech €375 million (US$445 million) for its COVID‑19 vaccine development program.^[54]

Pfizer CEO Albert Bourla stated that he decided against taking funding from the US government’s Operation Warp Speed for the development of the vaccine “because I wanted to liberate our scientists [from] any bureaucracy that comes with having to give reports and agree how we are going to spend the money in parallel or together, etc.” Pfizer did enter into an agreement with the US for the eventual distribution of the vaccine, as with other countries.^[55]

Clinical trials

Preliminary results from Phase I–II clinical trials on BNT162b2, published in October 2020, indicated potential for its efficacy and safety.^[17]^[56] During the same month, the European Medicines Agency (EMA) began a periodic review of BNT162b2.^[57]

The study of BNT162b2 is a continuous-phase trial in Phase III as of November 2020.^[18] It is a “randomized, placebo-controlled, observer-blind, dose-finding, vaccine candidate-selection, and efficacy study in healthy individuals”.^[18] The early-stage research determined the safety and dose level for two vaccine candidates, with the trial expanding during mid-2020 to assess efficacy and safety of BNT162b2 in greater numbers of participants, reaching tens of thousands of people receiving test vaccinations in multiple countries in collaboration with Pfizer and Fosun.^[20]^[35]

The Phase III trial assesses the safety, efficacy, tolerability, and immunogenicity of BNT162b2 at a mid-dose level (two injections separated by 21 days) in three age groups: 12–15 years, 16–55 years or above 55 years.^[18] For approval in the EU, an overall vaccine efficacy of 95% was confirmed by the EMA.^[58] The EMA clarified that the second dose should be administered three weeks after the first dose.^[59]

Efficacy endpoint	Vaccine efficacy (95% confidence interval) [%]
After dose 1 to before dose 2	52.4 (29.5, 68.4)
≥10 days after dose 1 to before dose 2	86.7 (68.6, 95.4)
Dose 2 to 7 days after dose 2	90.5 (61.0, 98.9)
≥7 days after dose 2 (subjects without evidence of infection prior to 7 days after dose 2)
Overall	95.0 (90.0, 97.9)
16–55 years	95.6 (89.4, 98.6)
≥55 years	93.7 (80.6, 98.8)
≥65 years	94.7 (66.7, 99.9)

The ongoing Phase III trial, which is scheduled to run from 2020 to 2022, is designed to assess the ability of BNT162b2 to prevent severe infection, as well as the duration of immune effect.^[20]^[21]^[22]

Pfizer and BioNTech started a Phase II/III randomized control trial in healthy pregnant women 18 years of age and older (NCT04754594).^[60] The study will evaluate 30 µg of BNT162b2 or placebo administered via intramuscular injection in 2 doses, 21 days apart. The Phase II portion of the study will include approximately 350 pregnant women randomized 1:1 to receive BNT162b2 or placebo at 27 to 34 weeks’ gestation. The Phase III portion of this study will assess the safety, tolerability, and immunogenicity of BNT162b2 or placebo among pregnant women enrolled at 24 to 34 weeks’ gestation. Pfizer and BioNTech announced on 18 February 2021 that the first participants received their first dose in this trial.^[61]

Vaccine technology

The BioNTech technology for the BNT162b2 vaccine is based on use of nucleoside-modified mRNA (modRNA) which encodes part of the spike protein found on the surface of the SARS-CoV-2 coronavirus (COVID‑19), triggering an immune response against infection by the virus protein.^[62]

The vaccine candidate BNT162b2 was chosen as the most promising among three others with similar technology developed by BioNTech.^[18]^[62]^[56] Prior to choosing BNT162b2, BioNTech and Pfizer had conducted Phase I trials on BNT162b1 in Germany and the United States, while Fosun performed a Phase I trial in China.^[17]^[63] In these Phase I studies, BNT162b2 was shown to have a better safety profile than the other three BioNTech candidates.^[63]

Sequence

The modRNA sequence of the vaccine is 4,284 nucleotides long.^[64] It consists of a five-prime cap; a five prime untranslated region derived from the sequence of human alpha globin; a signal peptide (bases 55–102) and two proline substitutions (K986P and V987P, designated “2P”) that cause the spike to adopt a prefusion-stabilized conformation reducing the membrane fusion ability, increasing expression and stimulating neutralizing antibodies;^[17]^[65] a codon-optimized gene of the full-length spike protein of SARS-CoV-2 (bases 103–3879); followed by a three prime untranslated region (bases 3880–4174) combined from AES and mtRNR1 selected for increased protein expression and mRNA stability^[66] and a poly(A) tail comprising 30 adenosine residues, a 10-nucleotide linker sequence, and 70 other adenosine residues (bases 4175–4284).^[64] The sequence contains no uridine residues; they are replaced by 1-methyl-3′-pseudouridylyl.^[64]

Composition

In addition to the mRNA molecule, the vaccine contains the following inactive ingredients (excipients):^[67]^[68]^[8]

ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate)
ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide
1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)
cholesterol
dibasic sodium phosphate dihydrate
monobasic potassium phosphate
potassium chloride
sodium chloride
sucrose
water for injection

The first four of these are lipids. The lipids and modRNA together form nanoparticles. ALC-0159 is a polyethylene glycol conjugate (that is, a PEGylated lipid).^[69]

The vaccine is supplied in a multidose vial as “a white to off-white, sterile, preservative-free, frozen suspension for intramuscular injection“.^[11]^[12] It must be thawed to room temperature and diluted with normal saline before administration.^[12]

Authorizations

Expedited

The United Kingdom’s Medicines and Healthcare products Regulatory Agency (MHRA) gave the vaccine “rapid temporary regulatory approval to address significant public health issues such as a pandemic” on 2 December 2020, which it is permitted to do under the Medicines Act 1968.^[70] It was the first COVID‑19 vaccine to be approved for national use after undergoing large scale trials,^[71] and the first mRNA vaccine to be authorized for use in humans.^[14]^[72] The United Kingdom thus became the first Western country to approve a COVID‑19 vaccine for national use,^[73] although the decision to fast-track the vaccine was criticised by some experts.^[74]

On 8 December 2020, Margaret “Maggie” Keenan, 90, from Fermanagh, became the first person to receive the vaccine.^[75] In a notable example of museums documenting the pandemic, the vial and syringe used for that first dose were saved acquired by The Science Museum in London for its permanent collection.^[76] By 20 December, 521,594 UK residents had received the vaccine as part of the national vaccination programme. 70% had been to people aged 80 or over.^[77]

After the United Kingdom, the following countries expedited processes to approve the Pfizer–BioNTech COVID‑19 vaccine for use: Argentina,^[78] Australia,^[79] Bahrain,^[80] Canada,^[7]^[81] Chile,^[82] Costa Rica,^[83] Ecuador,^[82] Hong Kong,^[84] Iraq,^[85] Israel,^[86] Jordan,^[87] Kuwait,^[88] Mexico,^[89] Oman,^[90] Panama,^[91] the Philippines,^[92] Qatar,^[93] Saudi Arabia,^[32]^[94] Singapore,^[95]^[96] the United Arab Emirates,^[97] and the United States.^[10]

The World Health Organization (WHO) authorized it for emergency use.^[98]

In the United States, an emergency use authorization (EUA) is “a mechanism to facilitate the availability and use of medical countermeasures, including vaccines, during public health emergencies, such as the current COVID‑19 pandemic”, according to the FDA.^[99] Following an EUA issuance, BioNTech and Pfizer are expected to continue the Phase III clinical trial to finalize safety and efficacy data, leading to application for licensure (approval) of the vaccine in the United States.^[99]^[100]^[101] The United States Centers for Disease Control and Prevention (CDC) Advisory Committee on Immunization Practices (ACIP) approved recommendations for vaccination of those aged 16 years or older.^[102]^[103]

Standard

On 19 December 2020, the Swiss Agency for Therapeutic Products (Swissmedic) approved the Pfizer–BioNTech COVID‑19 vaccine for regular use, two months after receiving the application, stating that the vaccine fully complied with the requirements of safety, efficacy and quality. This is the first authorization under a standard procedure.^[1]^[104] On 23 December, a Lucerne resident, a 90-year-old woman, became the first person to receive the vaccine in Switzerland.^[105] This marked the beginning of mass vaccination in continental Europe.^[106]

On 21 December 2020, the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA) recommended granting conditional marketing authorization for the Pfizer–BioNTech COVID‑19 vaccine under the brand name Comirnaty.^[2]^[107]^[108] The recommendation was accepted by the European Commission the same day.^[107]^[109]

On February 23, 2021, the Brazilian Health Regulatory Agency approved the Pfizer–BioNTech COVID-19 vaccine under its standard marketing authorization procedure. It became the first COVID-19 vaccine to receive definitive registration rather than emergency use authorization in the country.^[110]

Adverse effects

The adverse effect profile of the Pfizer–BioNTech COVID‑19 vaccine is similar to that of other adult vaccines.^[20] During clinical trials, the side effects deemed very common^[a] are (in order of frequency): pain and swelling at the injection site, tiredness, headache, muscle aches, chills, joint pain, and fever.^[68] Fever is more common after the second dose.^[68] These effects are predictable and to be expected, and it is particularly important that people be aware of this to prevent vaccine hesitancy.^[111]

Severe allergic reaction has been observed in approximately 11 cases per million doses of vaccine administered.^[112]^[113] According to a report by the US Centers for Disease Control and Prevention 71% of those allergic reactions happened within 15 minutes of vaccination and mostly (81%) among people with a documented history of allergies or allergic reactions.^[112] The UK’s Medicines and Healthcare products Regulatory Agency (MHRA) advised on 9 December 2020, that people who have a history of “significant” allergic reaction should not receive the Pfizer–BioNTech COVID‑19 vaccine.^[114]^[115]^[116] On 12 December, the Canadian regulator followed suit, noting that: “Both individuals in the U.K. had a history of severe allergic reactions and carried adrenaline auto injectors. They both were treated and have recovered.”^[67]

On 28 January 2021, the European Union published a COVID-19 vaccine safety update which found that “the benefits of Comirnaty in preventing COVID‑19 continue to outweigh any risks, and there are no recommended changes regarding the use the vaccine.”^[113]^[117] No new side effects were identified.^[113]

Manufacturing

A doctor holding the Pfizer vaccine

Pfizer and BioNTech are manufacturing the vaccine in their own facilities in the United States and in Europe in a three-stage process. The first stage involves the molecular cloning of DNA plasmids that code for the spike protein by infusing them into Escherichia coli bacteria. In the United States, this stage is conducted at a small pilot plant in Chesterfield, Missouri^[118] (near St. Louis). After four days of growth, the bacteria are killed and broken open, and the contents of their cells are purified over a week and a half to recover the desired DNA product. The DNA is stored in tiny bottles and frozen for shipment. Safely and quickly transporting the DNA at this stage is so important that Pfizer has used its company jet and helicopter to assist.^[119]

The second stage is being conducted at plants in Andover, Massachusetts^[120] in the United States, and in Germany. The DNA is used as a template to build the desired mRNA strands. Once the mRNA has been created and purified, it is frozen in plastic bags about the size of a large shopping bag, of which each can hold up to 5 to 10 million doses. The bags are placed on special racks on trucks which take them to the next plant.^[119]

The third stage is being conducted at plants in Portage, Michigan^[121] (near Kalamazoo) in the United States, and Puurs in Belgium. This stage involves combining the mRNA with lipid nanoparticles, then filling vials, boxing vials, and freezing them.^[119] Croda International subsidiary Avanti Polar Lipids is providing the requisite lipids.^[122] As of November 2020, the major bottleneck in the manufacturing process was combining mRNA with lipid nanoparticles.^[119]

In February 2021, Pfizer revealed this entire sequence initially took about 110 days on average from start to finish, and that the company was making progress on reducing that number to 60 days.^[123] Vaccine manufacturers normally take several years to optimize the process of making a particular vaccine for speed and cost-effectiveness before attempting large-scale production.^[123] Due to the urgency presented by the COVID-19 pandemic, Pfizer began production immediately with the process by which the vaccine had been originally formulated in the laboratory, then started to identify ways to safely speed up and scale up that process.^[123]

BioNTech announced in September 2020 that it had signed an agreement to acquire from Novartis a manufacturing facility in Marburg, Germany, to expand their vaccine production capacity.^[124] Once fully operational, the facility would produce up to 750 million doses per year, or over 60 million doses per month. The site will be the third BioNTech facility in Europe which currently produces the vaccine, while Pfizer operates at least four production sites in the United States and Europe.

Advance orders and logistics

Pfizer indicated in its 9 November press release that 50 million doses could be available by the end of 2020, with about 1.3 billion doses provided globally by 2021.^[20] In February 2021, BioNTech announced it would increase production by more than 50% to manufacture two billion doses in 2021.^[125]

In July 2020, the vaccine development program Operation Warp Speed placed an advance order of US$1.95 billion with Pfizer to manufacture 100 million doses of a COVID‑19 vaccine for use in the United States if the vaccine was shown to be safe and effective.^[34]^[126]^[127]^[128] By mid-December 2020, Pfizer had agreements to supply 300 million doses to the European Union,^[129] 120 million doses to Japan,^[130] 40 million doses (10 million before 2021) to the United Kingdom,^[22] 20 million doses to Canada,^[131] an unspecified number of doses to Singapore,^[132] and 34.4 million doses to Mexico.^[133] Fosun also has agreements to supply 10 million doses to Hong Kong and Macau.^[134] The Hong Kong government said it would receive its first batch of one million doses by the first quarter of 2021.^[135]

BioNTech and Fosun agreed to supply Mainland China with a batch of 100 million doses in 2021, subject to regulatory approval. The initial supply will be delivered from BioNTech’s production facilities in Germany.^[136]

The vaccine is being delivered in vials that, once diluted, contain 2.25 ml of vaccine (0.45 ml frozen plus 1.8ml diluent).^[101] According to the vial labels, each vial contains five 0.3 ml doses, however excess vaccine may be used for one, or possibly two, additional doses.^[101]^[137] The use of low dead space syringes to obtain the additional doses is preferable, and partial doses within a vial should be discarded.^[101]^[138] The Italian Medicines Agency officially authorized the use of excess doses remaining within single vials.^[139] As of 8 January 2021, each vial contains six doses.^[68]^[140]^[141]^[138] In the United States, vials will be counted as five doses when accompanied by regular syringes and as six doses when accompanied by low dead space syringes.^[142]

Temperature the Pfizer vaccine must be kept at to ensure effectiveness, roughly between −80 and −60 °C (−112 and −76 °F)

Logistics in developing countries which have preorder agreements with Pfizer—such as Ecuador and Peru—remain unclear.^[38] Even high-income countries have limited cold chain capacity for ultracold transport and storage of a vaccine that degrades within five days when thawed, and requires two shots three weeks apart.^[38] The vaccine needs to be stored and transported at ultracold temperatures between −80 and −60 °C (−112 and −76 °F),^[39]^[22]^[38]^[143]^[144] much lower than for the similar Moderna vaccine. The head of Indonesia‘s Bio Farma Honesti Basyir stated that purchasing the vaccine is out of the question for the world’s fourth-most populous country, given that it did not have the necessary cold chain capability. Similarly, India’s existing cold chain network can only handle temperatures between 2 and 8 °C (36 and 46 °F), far above the requirements of the vaccine.^[145]^[146]

In January 2021, Pfizer and BioNTech offered to supply 50 million doses of COVID‑19 vaccine for health workers across Africa between March and the end of 2021, at a discounted price of US$10 per dose.^[147]

Name

BNT162b2 was the code name during development and testing,^[17]^[148] tozinameran is the proposed international nonproprietary name (pINN),^[149] and Comirnaty is the brand name.^[1]^[2] According to BioNTech, the name Comirnaty “represents a combination of the terms COVID‑19, mRNA, community, and immunity.”^[150]^[151]

The vaccine also has the common name “COVID‑19 mRNA vaccine (nucleoside-modified)”^[2] and may be distributed in packaging with the name Pfizer–BioNTech COVID‑19 Vaccine.”^[152]

How the Pfizer-BioNTech Vaccine Works

By Jonathan Corum and Carl ZimmerUpdated Jan. 21, 2021Leer en español

The German company BioNTech partnered with Pfizer to develop and test a coronavirus vaccine known as BNT162b2, the generic name tozinameran or the brand name Comirnaty. A clinical trial demonstrated that the vaccine has an efficacy rate of 95 percent in preventing Covid-19.

A Piece of the Coronavirus

The SARS-CoV-2 virus is studded with proteins that it uses to enter human cells. These so-called spike proteins make a tempting target for potential vaccines and treatments.

Spikes

Spike

protein

gene

CORONAVIRUS

Like the Moderna vaccine, the Pfizer-BioNTech vaccine is based on the virus’s genetic instructions for building the spike protein.

mRNA Inside an Oily Shell

The vaccine uses messenger RNA, genetic material that our cells read to make proteins. The molecule — called mRNA for short — is fragile and would be chopped to pieces by our natural enzymes if it were injected directly into the body. To protect their vaccine, Pfizer and BioNTech wrap the mRNA in oily bubbles made of lipid nanoparticles.

Lipid nanoparticles

surrounding mRNA

Because of their fragility, the mRNA molecules will quickly fall apart at room temperature. Pfizer is building special containers with dry ice, thermal sensors and GPS trackers to ensure the vaccines can be transported at –94°F (–70°C) to stay viable.

Entering a Cell

After injection, the vaccine particles bump into cells and fuse to them, releasing mRNA. The cell’s molecules read its sequence and build spike proteins. The mRNA from the vaccine is eventually destroyed by the cell, leaving no permanent trace.

VACCINE

PARTICLES

VACCINATED

CELL

Spike

protein

mRNA

Translating mRNA

Three spike

proteins combine

Spike

Cell

nucleus

Spikes

and protein

fragments

Displaying

spike protein

fragments

Protruding

spikes

Some of the spike proteins form spikes that migrate to the surface of the cell and stick out their tips. The vaccinated cells also break up some of the proteins into fragments, which they present on their surface. These protruding spikes and spike protein fragments can then be recognized by the immune system.

Spotting the Intruder

When a vaccinated cell dies, the debris will contain many spike proteins and protein fragments, which can then be taken up by a type of immune cell called an antigen-presenting cell.

Debris from

a dead cell

Engulfing

a spike

ANTIGEN-

PRESENTING

CELL

Digesting

the proteins

Presenting a

spike protein

fragment

HELPER

T CELL

The cell presents fragments of the spike protein on its surface. When other cells called helper T cells detect these fragments, the helper T cells can raise the alarm and help marshal other immune cells to fight the infection.

Making Antibodies

Other immune cells, called B cells, may bump into the coronavirus spikes on the surface of vaccinated cells, or free-floating spike protein fragments. A few of the B cells may be able to lock onto the spike proteins. If these B cells are then activated by helper T cells, they will start to proliferate and pour out antibodies that target the spike protein.

HELPER

T CELL

Activating

the B cell

Matching

surface proteins

VACCINATED

CELL

B CELL

SECRETED

ANTIBODIES

Stopping the Virus

The antibodies can latch onto coronavirus spikes, mark the virus for destruction and prevent infection by blocking the spikes from attaching to other cells.

ANTIBODIES

VIRUS

Killing Infected Cells

The antigen-presenting cells can also activate another type of immune cell called a killer T cell to seek out and destroy any coronavirus-infected cells that display the spike protein fragments on their surfaces.

ANTIGEN-PRESENTING CELL Presenting a spike protein fragment ACTIVATED KILLER T CELL INFECTED CELL Beginning to kill the infected cell

Remembering the Virus

The Pfizer-BioNTech vaccine requires two injections, given 21 days apart, to prime the immune system well enough to fight off the coronavirus. But because the vaccine is so new, researchers don’t know how long its protection might last.

First dose, 0.3ml

Second dose, 21 days later

A preliminary study found that the vaccine seems to offer strong protection about 10 days after the first dose, compared with people taking a placebo:

Cumulative incidence of Covid-19 among clinical trial participants 2.5% 2.0 People taking a placebo

1.5 1.0 Second dose First dose People taking the

Pfizer-BioNTech vaccine

0.5

Weeks after the first dose

It’s possible that in the months after vaccination, the number of antibodies and killer T cells will drop. But the immune system also contains special cells called memory B cells and memory T cells that might retain information about the coronavirus for years or even decades.

For more about the vaccine, see Pfizer’s Covid Vaccine: 11 Things You Need to Know.

Preparation and Injection

Each vial of the vaccine contains 5 doses of 0.3 milliliters. The vaccine must be thawed before injection and diluted with saline. After dilution the vial must be used within six hours.

A diluted vial of the vaccine at Royal Free Hospital in London.Jack Hill/Agence France-Presse

References

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External links

“Tozinameran”. Drug Information Portal. U.S. National Library of Medicine.

Global Information About Pfizer–BioNTech COVID‑19 Vaccine (also known as BNT162b2) Pfizer
Comirnaty assessment report European Medicines Agency Committee for Medicinal Products for Human Use
A Phase 1/2/3 Study to Evaluate the Safety, Tolerability, Immunogenicity, and Efficacy of RNA Vaccine Candidates Against COVID‑19 in Healthy Individuals Pfizer clinical protocol
Pfizer Vaccince News, updates and tracking of Israel’s vaccinaion campaign
“How the Pfizer-BioNTech Covid-19 Vaccine Works”. The New York Times.

Vaccine description
A vial of the Pfizer–BioNTech COVID‑19 vaccine
Target disease	COVID‑19
Type	mRNA
Clinical data
Trade names	Comirnaty^[1]^[2]
Other names	BNT162b2, COVID-19 mRNA vaccine (nucleoside-modified)
License data	EU EMA: by INNUS DailyMed: Pfizer-BioNTech_COVID-19_Vaccine
Pregnancy category	AU: B1^[3]
Routes of administration	Intramuscular
ATC code	None
Legal status
Legal status	AU: S4 (Prescription only) ^[4]^[5]CA: Authorized by interim order ^[6]^[7]UK: Conditional and temporary authorization to supply ^[8]^[9]US: Unapproved (Emergency Use Authorization)^[10]^[11]^[12]EU: Conditional marketing authorization granted ^[2]CH: Rx-only^{[further explanation needed]}^[1]
Identifiers
CAS Number	2417899-77-3
PubChem SID	434370509
DrugBank	DB15696
UNII	5085ZFP6SJ
KEGG	D11971

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#Tozinameran, #APPROVALS 2021, #JAPAN 2021, Comirnaty, #Coronavirus disease, #COVID-19, #BNT162b2 , #BNT162b2, #SARS-CoV-2 Vaccine, #RNA ingredient BNT-162B2, #corona

The Pfizer-BioNTech COVID-19 vaccine (Tozinameran, INN), also known as BNT162b2, is one of four advanced mRNA-based vaccines developed through “Project Lightspeed,” a joint program between Pfizer and BioNTech.^2,3 Tozinameran is a nucleoside modified mRNA (modRNA) vaccine encoding an optimized full-length version of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein. It is designed to induce immunity against SARS-CoV-2, the virus responsible for causing COVID-19.² The modRNA is formulated in lipid nanoparticles for administration via intramuscular injection in two doses, three weeks apart.^1,3

Tozinameran is undergoing evaluation in clinical trials in both the USA (NCT04368728) and Germany (NCT04380701).^4,5 Tozinameran received fast track designation by the U.S. FDA on July 13, 2020.⁶ On December 11, 2020, the FDA issued an Emergency Use Authorization (EUA) based on 95% efficacy in clinical trials and a similar safety profile to other viral vaccines over a span of approximately 2 months.¹ Tozinameran was granted an EUA in the UK on December 2, 2020,⁸ and in Canada on December 9, 2020⁷ for active immunization against SARS-CoV-2.¹²

Currently, sufficient data are not available to determine the longevity of protection against COVID-19, nor direct evidence that the vaccine prevents the transmission of the SARS-CoV-2 virus from one individual to another.⁹ Fact sheets for caregivers, recipients, and healthcare providers are now available.^10,11

Tozinameran has not yet been fully approved by any country. In both the UK and Canada, Tozinameran is indicated under an interim authorization for active immunization to prevent COVID-19 caused by SARS-CoV-2 in individuals aged 16 years and older.^7,8

On December 11, 2020, the U.S. Food and Drug Administration granted emergency use authorization (EUA) for Tozinameran to prevent COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in patients aged 16 years and above.⁹ Safety and immune response information for adolescents 12-15 years of age will follow, and studies to further explore the administration of Tozinameran in pregnant women, children under 12 years of age, and those in special risk groups will be evaluated in the future.¹

This vaccine should only be administered where appropriate medical treatment for immediate allergic reactions are immediately available in the case of an acute anaphylactic reaction after vaccine administration.¹² Tozinameran administration should be postponed in any individual suffering from an acute febrile illness. Its use should be carefully considered in immunocompromised individuals and individuals with a bleeding disorder or on anticoagulant therapy. Appropriate medical treatment should be readily available in case of an anaphylactic reaction following vaccine administration.^7,8

Tozinameran contains nucleoside modified mRNA (modRNA) encapsulated in lipid nanoparticles that deliver the modRNA into host cells. The lipid nanoparticle formulation facilitates the delivery of the RNA into human cells.¹² Once inside these cells, the modRNA is translated by host machinery to produce the SARS-CoV-2 spike (S) protein antigen, which is subsequently recognized by the host immune system. Tozinameran has been shown to elicit both neutralizing antibody and cellular immune responses to the S protein, which helps protect against subsequent SARS-CoV-2 infection.^7,8

Tozinameran is a nucleoside modified mRNA (modRNA) vaccine encoding an optimized full-length version of the SARS-CoV-2 spike (S) protein, translated and expressed in cells in vaccinated individuals to produce the S protein antigen against which an immune response is mounted. As with all vaccines, protection cannot be guaranteed in all recipients, and full protection may not occur until at least seven days following the second dose.^7,8

In U.S. clinical trials, the vaccine was 95% effective in preventing COVID-19; eight COVID-19 cases occurred in the vaccine group and 162 cases occurred in the placebo group. Of the total 170 COVID-19 cases, one case in the vaccine group and three cases in the placebo group were considered to be severe infections.^1,9

Polack FP, Thomas SJ, Kitchin N, Absalon J, Gurtman A, Lockhart S, Perez JL, Perez Marc G, Moreira ED, Zerbini C, Bailey R, Swanson KA, Roychoudhury S, Koury K, Li P, Kalina WV, Cooper D, Frenck RW Jr, Hammitt LL, Tureci O, Nell H, Schaefer A, Unal S, Tresnan DB, Mather S, Dormitzer PR, Sahin U, Jansen KU, Gruber WC: Safety and Efficacy of the BNT162b2 mRNA Covid-19 Vaccine. N Engl J Med. 2020 Dec 10. doi: 10.1056/NEJMoa2034577. [PubMed:33301246]
Gen Eng News: BNT162 vaccine candidates [Link]
BioNTech BNT162 Update [Link]
Clinical Trial NCT04368728 [Link]
Clinical Trial NCT04380701 [Link]
FDA fast track designation: BNT162b1 and BNT162b2 [Link]
Health Canada Interim Product Monograph: BNT162b2 SARS-CoV-2 Vaccine [Link]
MHRA Interim Product Monograph: BNT162b2 SARS-CoV-2 Vaccine [Link]
FDA News Release: FDA Takes Key Action in Fight Against COVID-19 By Issuing Emergency Use Authorization for First COVID-19 Vaccine [Link]
Pfizer: Fact Sheet for Healthcare Providers Administering Vaccine, Pfizer-BioNtech COVID-19 vaccine [Link]
Pfizer: Fact Sheet for Recipients and Caregivers, Pfizer BioNTech COVID-19 vaccine [Link]
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Umbralisib

February 18, 2021 9:15 am / Leave a comment

Umbralisib tosylate

Formula	C31H24F3N5O3. C7H8O3S
Cas	1532533-72-4 FREE 1532533-67-7
Mol weight	743.7508

FDA APPR 2021/2/5

ウムブラリシブトシル酸塩;

Treatment of cancer and B-cell related disorders

Antineoplastic

RP-5152; RP-5237; PI3K delta inhibitors (cancer), Rhizen/Incozen; PI3K delta inhibitors (B-cell lymphoma/hematological cancers), Incozen/Rhizen; TGR-1202; TG-1202; RV-1001; umbralisib tosylate; umbralisib; RP-5264; RP-5307; dual PI3Kdelta/CK1 inhibitor (cancer), TG Therapeutics; Ukoniq

Umbralisib (TGR-1202) is an orally available PI3K delta inhibitor, targeting the delta isoform with nanomolar potency and several fold selectivity over the alpha, beta, and gamma isoforms of PI3K. The delta isoform of PI3K is strongly expressed in cells of hematopoietic origin and is believed to be important in the proliferation and survival of B-cell lymphocytes. Inhibition of PI3K delta signaling with umbralisib has demonstrated robust activity in numerous pre-clinical models and primary cells from patients with hematologic malignancies. Umbralisib is currently in Phase 3 clinical development in combination with Ublituximab for patients with hematologic malignancies.

Umbralisib, sold under the brand name Ukoniq, is a medication for the treatment of marginal zone lymphoma (MZL) and follicular lymphoma (FL).^[2] It is taken by mouth.^[2]

The most common side effects include increased creatinine, diarrhea-colitis, fatigue, nausea, neutropenia, transaminase elevation, musculoskeletal pain, anemia, thrombocytopenia, upper respiratory tract infection, vomiting, abdominal pain, decreased appetite, and rash.^[2]

Umbralisib is a kinase inhibitor including PI3K-delta and casein kinase CK1-epsilon.^[2]^[3]^[4] Umbralisib was approved for medical use in the United States in February 2021.^[2]^[5]

In April 2019, the FDA granted umbralisib Orphan drug designations for the treatment of nodal MZL, extranodal MZL, and splenic MZL. In January 2019, the FDA granted Breakthrough Therapy Designation for the treatment of MZL in patients who had received at least one prior anti-CD20 regimen, based on the interim data from the MZL umbralisib monotherapy cohort in the UNITY-NHL study. In March 2020, the drug was granted Orphan status for treatment of FL By June 2019, the confirmation of registration path to submit umbralisib for accelerated approval was obtained from the MZL cohort of the UNITY-NHL Phase IIb trial .

In August 2020, the FDA accepted the NDA for review; the MZL indication (patients with previously treated MZL who have received at least one prior anti-CD20 based regimen) was accepted for Priority Review with a PDUFA date of February 15, 2021, while the FL indication (patients with previously treated FL who have received at least two prior systemic therapies) was accepted for standard review with a PDUFA date of June 15, 2021.

In February 2021, the drug was granted accelerated approval by the FDA for second-line MZL and for fourth-line FL, based on results of UNITY-NHL. At that time, commercial launch was expected in the coming days

Medical uses

Umbralisib is indicated for adults with relapsed or refractory marginal zone lymphoma (MZL) who have received at least one prior anti-CD20-based regimen; and adults with relapsed or refractory follicular lymphoma (FL) who have received at least three prior lines of systemic therapy.^[2]^[1]

Umbralisib is a kinase inhibitor. The active pharmaceutical ingredient is umbralisib tosylate with the molecular formula C₃₈H₃₂F₃N₅O₆S and a molecular weight of 743.75 g/mol. The chemical name for umbralisib tosylate is (S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo [3, 4-d] pyrimidin-1-yl)-ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one 4- methylbenzenesulfonate and has the following structure:

UKONIQ™ (umbralisib) Structrual Formula Illustration

Umbralisib tosylate is white to light brown powder that is freely soluble in dimethyl sulfoxide, soluble in methanol, and practically insoluble in water. The ionization constant (pKa) of umbralisib tosylate is 2.71.

UKONIQ tablets are for oral administration. Each tablet contains 200 mg of umbralisib free base equivalent to 260.2 mg of umbralisib tosylate. The tablets also contain inactive ingredients: croscarmellose sodium, hydroxypropyl betadex, hydroxypropyl cellulose, magnesium stearate and microcrystalline cellulose.

The tablet coating film consists of FD&C Blue No. 1, FD&C Yellow No. 5, ferric oxide yellow, hypromellose 2910, polydextrose, polyethylene glycol 8000, titanium dioxide and triacetin.
Indications & Dosage

INDICATIONS

Marginal Zone Lymphoma

UKONIQ is indicated for the treatment of adult patients with relapsed or refractory marginal zone lymphoma (MZL) who have received at least one prior anti-CD20-based regimen.

This indication is approved under accelerated approval based on overall response rate [see Clinical Studies]. Continued approval for this indication may be contingent upon verification and description of clinical benefit in a confirmatory trial(s).

Follicular Lymphoma

UKONIQ is indicated for the treatment of adult patients with relapsed or refractory follicular lymphoma (FL) who have received at least three prior lines of systemic therapy.

Adverse effects

The prescribing information provides warnings and precautions for adverse reactions including infections, neutropenia, diarrhea and non-infectious colitis, hepatotoxicity, and severe cutaneous reactions.^[2]

History

It has undergone clinical studies for chronic lymphocytic leukemia (CLL).^[6]^[7] Three year data (including follicular lymphoma and DLBCL) was announced June 2016.^[8] It is in combination trials for various leukemias and lymphomas, such as mantle cell lymphoma (MCL)^[9]^[10] and other lymphomas.^[11]

Umbralisib was granted breakthrough therapy desgination by the U.S. Food and Drug Administration (FDA) for use in people with marginal zone lymphoma (MZL), a type of cancer with no specifically approved therapies.^[12]

FDA approval was based on two single-arm cohorts of an open-label, multi-center, multi-cohort trial, UTX-TGR-205 (NCT02793583), in 69 participants with marginal zone lymphoma (MZL) who received at least one prior therapy, including an anti-CD20 containing regimen, and in 117 participants with follicular lymphoma (FL) after at least two prior systemic therapies.^[2] The application for umbralisib was granted priority review for the marginal zone lymphoma (MZL) indication and orphan drug designation for the treatment of MZL and follicular lymphoma (FL).^{[2][13][14][15][16]}

SYN

WO 2014071125

clip

First new chemical entity discovered by Indian scientists gets USFDA approval

https://www.businesstoday.in/sectors/pharma/first-new-chemical-entity-discovered-by-indian-scientists-gets-us-fda-approval/story/430693.html

Rhizen has retained commercialisation rights for India while also being the manufacturing and supply partner for Umbralisib. Alembic owns 50 per cent stake in Rhizen

Umbralisib, a novel cancer drug discovered and out-licensed by India’s Alembic Pharmaceuticals and its associate drug discovery company Rhizen Pharmaceuticals, has received the drug regulatory approval for sales in the US market. The drug is touted to be the first new chemical entity (NCE) discovered by Indian scientists to secure a US Food and Drug Administration (FDA) approval.

Switzerland based Rhizen had discovered the molecule in 2012 and two years later was licensed to US based TG Therapeutics, which has worldwide sales rights. Rhizen has retained commercialisation rights for India while also being the manufacturing and supply partner for Umbralisib. Alembic owns 50 per cent stake in Rhizen.

Umbralisib is a novel, next generation, oral, once daily drug for adult patients with relapsed or refractory lymphoma and relapsed or refractory marginal zone lymphoma (MZL) that resists treatments and drugs. Such cancers affect over 3-4 lakh patients in the US every year. The drug is estimated to have a global market worth US$ 1-1.5 billion.

“We are extremely proud of this historic milestone for Rhizen, and of the fact that Umbralisib is the first NCE discovered by Indian scientists to secure a US FDA approval,” said Pranav Amin, Chairman, Rhizen Pharmaceuticals & Managing Director of Alembic Pharmaceuticals.

“We are keen to bring Umbralisib to Indian patients and we plan to initiate activities towards registration and approval there soon,” said Swaroop Vakkalanka, President & CEO of Rhizen Pharmaceuticals.

Ahmedabad-based Zydus Cadila had a few months ago got ‘Fast Track Designation’ by the US Food and Drug Administration (USFDA) for Saroglitazar in the treatment of patients with Primary Biliary Cholangitis (PBC), a liver disorder due to progressive destruction of the bile ducts.

PATENT

WO 2021009509

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2021009509&tab=PCTDESCRIPTION&_cid=P10-KLABTI-88387-1

Umbralisib, having the chemical designation (S)-2-(l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one, is an orally available PI3K delta inhibitor. Umbralisib has the following structure:

Inhibition of PI3K delta signaling with umbralisib has demonstrated activity in several pre-clinical models and primary cells from patients with hematologic malignancies. In a Phase 2 trial, umbralisib provided effective PI3K-delta inhibition and appeared well-tolerated among patients with relapsed/refractory marginal zone lymphoma. Umbralisib is currently in Phase 3 clinical development in combination with ublituximab for patients with hematologic malignancies. Hematologic malignancies are forms of cancer that begin in the cells of blood-forming tissue, such as the bone marrow, or in the cells of the immune system. Examples of hematologic cancer are acute and chronic leukemias, lymphomas, multiple myeloma and myelodysplastic syndromes. Lymphomas can include follicular lymphoma (FL), small lymphocytic lymphoma (SLL), non-Hodgkin lymphoma (NHL), and diffuse large B-cell lymphoma (DLBCL), among others. Leukemia can include chronic lymphocytic leukemia (CLL), among others. The U.S. Food and Drug Administration (FDA) has granted orphan drug designation to umbralisib for the treatment of patients with follicular lymphoma and for the treatment of patients with nodal, extranodal, and splenic marginal zone lymphoma.

U.S. Patent No. 9,150,579 discloses umbralisib and pharmaceutically acceptable salts thereof, such as 4-methylbenzenesulfonate (also known as tosylate), sulphate, hydrochloride, benzenesulfonate, maleate, and camphor sulfonate salts. U.S. Patent Nos. 9,969,740 and 10,414,773 and U.S. Patent Application Publication No. 2019/0382411 disclose solid state forms of a p-toluenesulfonic acid salt (PTSA) of umbralisib. None of these references disclose an amorphous form of umbralisib monotosylate.

An amorphous form of a compound is considered to be a solid state form that lacks long-range order relative to crystalline solid state forms of the compound. The amorphous form is chemically identical to other crystalline solid state forms but can exhibit different physical properties such as intrinsic solubility, rate of dissolution, density, mechanical property, chemical and physical stability, hygroscopicity, and morphology. The differences in intrinsic solubility also may lead to a difference in the rate of absorption, thus impacting bioavailability. Generally, amorphous compounds have a higher solubility than crystalline compounds.

EXAMPLES

Examples 1-3, which follow herein, provide embodiments of the preparation of amorphous umbralisib monotosylate.

Example 1

Preparation of Amorphous Umbralisib Monotosylate by Dry Grinding of Crystalline Umbralisib Tosylate Salt

Form I of umbralisib tosylate salt is dried under vacuum at about 40 °C in an oven for at least about 3 days to remove any residual ethyl acetate. About 30 mg of the dried umbralisib tosylate salt is ground manually using a mortar (about 6 cm in diameter) and pestle for about 3 minutes. The ground umbralisib tosylate salt is identified as being amorphous by XRPD. FIG. 1 is a representative XPRD pattern for amorphous umbralisib monotosylate prepared according to Example 1.

The amorphous umbralisib monotosylate prepared according to Example 1 is characterized by a T_g of about 51 °C, as depicted in the mDSC thermogram contained in FIG. 2.

A DVS of amorphous umbralisib monotosylate prepared according to Example 1 indicates the sample is hygroscopic, with about a 4% weight change between about 0-90% relative humidity, as depicted in FIG. 3, and less than about a 1% weight change in the sample over three cycles, as depicted in FIG. 4.

An XRPD pattern of the sample after DVS indicates that the sample is still amorphous, as depicted in FIG. 5.

Example 2

Preparation of Amorphous Umbralisib Monotosylate by Dissolution of

Crystalline Umbralisib Tosylate Salt in Methanol and Its Evaporation Therefrom

About 470 mg of Form I of umbralisib tosylate salt is dissolved in about 20 mL of methanol at about 50 °C. A solid umbralisib tosylate salt is obtained by evaporation of the solution under vacuum at about 40 °C in an oven overnight. The isolated product is identified as being amorphous umbralisib monotosylate by XRPD. FIG. 6 is a representative XPRD pattern for amorphous umbralisib monotosylate prepared according to Example 2.

The amorphous umbralisib monotosylate prepared according to Example 2 is characterized by a T_g of about 75 °C, as depicted in the mDSC thermogram contained in FIG. 7.

A TGA of amorphous umbralisib monotosylate prepared according to Example 2 shows about a 0.9% weight loss up to about 120 °C, as depicted in FIG. 8.

A DVS of amorphous umbralisib monotosylate prepared according to Example 2 indicates that the sample is hygroscopic, with about a 4% weight change between about 0-90% relative humidity, as depicted in FIG. 9, with about a 0.5% weight change in the sample over three cycles, as depicted in FIG. 10.

An XRPD pattern of the sample after DVS indicates that the sample is still amorphous, as depicted in FIG. 11.

‘ H NMR is carried out on a sample of amorphous umbralisib monotosylate prepared according to Example 2 in DMSO-d₆ which indicates an umbralisib tosylate salt with a 1 :0.9 ratio of free base to acid, as depicted in FIG. 12. The peak at 8.25 ppm is representative of a single proton in the free base and the peaks at 2.30 ppm are the three protons from p-toluenesulfonic acid. A trace amount (about 0.07%) of methanol is observed at 3.16 ppm.

FTIR spectra is collected on amorphous umbralisib monotosylate prepared according to Example 2, as depicted in FIG. 13(a) and on starting crystalline umbralisib tosylate salt, as depicted in FIG. 13(b).

XRPD of amorphous umbralisib monotosylate prepared according to Example 2 after storage at about 40 °C under vacuum conditions for about two weeks indicates that the sample is still amorphous, as depicted in FIG. 14. Further, mDSC of amorphous umbralisib monotosylate after storage at about 40 °C under vacuum conditions for about two weeks indicates that the T_g is increased to about 83 °C, as depicted in FIG. 15.

Example 3

Solution Preparation of Amorphous Umbralisib Monotosylate from Umbralisib

Free Base and p-Toluenesulfonic Acid

Umbralisib free base and p-toluenesulfonic acid are each separately dissolved in MeOH. Specifically, about 72 mg of umbralisib free base is dissolved in about 3mL of MeOH at about 50 °C and about 24 mg of p-toluenesulfonic acid is dissolved in about 0.25 mL of MeOH at about 50 °C. The two solutions are mixed and stirred at room temperature for about 1 hr and then at about 4 °C overnight. The solution is transferred to a vacuum oven at about 40 °C overnight to evaporate the MeOH. Amorphous umbralisib monotosylate, identified by XRPD, is obtained. FIG. 16 is a representative XPRD pattern for amorphous umbralisib monotosylate prepared according to Example 3.

PATENT

WO 2015181728

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2015181728&_cid=P10-KLABVD-88799-1

TGR-1202, chemically known as (S)-2-(l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-3-(3-fluorophenyl)-4H-chromen-4-one, has the following chemical structure:

[04] The preparation of TGR-1202 and its salts is described in International Publication No. WO 2014/006572 and U.S. Patent Publication No. 2014/0011819, each of which is incorporated herein by reference in its entirety for all purposes. TGR-1202 is an investigational drug currently undergoing multiple clinical trials in the area of haematological malignancies.

[05] WO 2014/006572 and US 2014/0011819 describe the synthesis of TGR-1202 (Example B l) and also disclose the therapeutic activity of this molecule to inhibit, regulate and/or modulate the signal transduction of PI3K.

Example 1: Preparation of the PTSA Salt of TGR-1202 (Form A)

[103] 7100 g of TGR-1202 was charged in a reactor containing 56.8 litres of acetone and stirred at ambient temperature. 4680 g of p-toluene sulphonic acid was added and the reaction mixture was heated at a temperature of 60-65° C for about 6 hours. The solvent was removed by distillation under reduced pressure to obtain a wet residue. The wet residue was degassed and allowed to cool to < 20° C. Approximately 142 litres of diethyl ether was then added and the resulting mixture was stirred overnight, then filtered to obtain a solid mass which was washed with diethyl ether and dried in vacuo to yield a solid mass. The solid mass was re-suspended in diethyl ether, stirred for 6 hours, and then filtered to yield a solid mass which was subsequently dissolved in 56.8 litres of acetone, filtered through a HiFlow bed, and concentrated under reduced pressure. The resulting residue mass was stirred with water overnight, then filtered and vacuum dried to yield 6600 g of the PTSA salt of TGR-1202. HPLC: 99.21% and chiral purity of 99.64:0.36 (S:R).

Example 2: Preparation of the PTSA Salt of TGR-1202 (Form B)

1000 g of TGR-1202 was charged in a reactor containing 8 litres of acetone and stirred at ambient temperature. 666 g of p-toluene sulphonic acid was then added and the reaction mixture was heated at a temperature of 60-65 °C for about 6 hours. The solvent was removed by distillation under reduced pressure to obtain a wet residue. The wet residue was degassed and allowed to cool to < 20° C. Approximately 20 litres of diethyl ether was added and the resulting mixture was stirred overnight, then filtered to obtain a solid mass which was washed with diethyl ether and dried in vacuo to yield a solid mass which was then vacuum dried to yield 1150 g of the PTSA salt of TGR-1202. HPLC: 99.33% and chiral purity: 99.61:0.39 (S:R).

PATENT

WO 2014006572

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2014006572&_cid=P10-KLABZ1-89676-1

Intermediate 1

[104] Intermediate 1: 6-fluoro-3-(3-fluorophenyl)-2-(l-hydroxyethyl)-4H-chromen-4-one: To a solution of 2-(l-bromoethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (15.0 g,

40.84 mmol) in DMSO (150 ml), n-butanol (7.5 ml) was added and heated to 120°C for 3h. The reaction mixture was cooled to RT, quenched with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as an off-white solid (7.90 g, 64%). H-NMR (δ ppm, CDC1₃, 400 MHz): 7.85 (dd, J = 8.1, 3 Hz, 1H), 7.54 (dd, J = 9.2, 4.2 Hz, 1H), 7.47-7.37 (m, 2H), 7.15-6.98 (m, 3H), 4.74 (quintet, J = 6.8 Hz, 1H), 2.23 (d, J = 7.4 Hz, 1H), 1.54 (d, J = 6.6 Hz, 3H).

Intermediate 2

[105] Intermediate 2: 2-acetyl-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one: DMSO (5.60 ml, 79.14 mmol) was added to dichloromethane (40 ml) cooled to -78°C, followed by oxalyl chloride (3.40 ml, 39.57 mmol). After 10 min. intermediate 1 (6.00 g, 19.78 mmol) in dichloromethane (54 ml) was added dropwise and stirred for 20 min. Triethylamine (12 ml) was added and stirred for lh. The reaction mixture was quenched with water and extracted with dichloromethane. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow solid (4.2 g, 71%) which was used as such in the next step.

Intermediate 3

[106] Intermediate 3: (S)-6-fluoro-3-(3-fluorophenyl)-2-(l-hydroxyethyl)-4H-chromen-4-one: To intermediate 2 (2.00 g, 6.66 mmol), R-Alpine borane (0.5M in THF, 20 ml) was added and heated to 60°C for 20h. The reaction mixture quenched with aq. 2N HC1, and

extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as an off-white solid (1.51 g, 75%). Enantiomeric excess: 94.2%, enriched in the fast eluting isomer (retention time: 8.78 min.) as determined by HPLC on a chiralpak AD-H column.

Intermediate 4

[107] Intermediate 4: (R)-l-(6-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl 4-chlorobenzoate: To a solution of intermediate 3 (1.45 g, 4.78 mmol) in THF (15 ml), 4-chlorobenzoic acid (0.748 g, 4.78 mmol) and triphenylphosphine (1.88 g, 7.17 mmol) were added and heated to 45 C followed by diisopropylazodicarboxylate (1.4ml, 7.17 mmol). After lh, the reaction mixture was concentrated and the residue was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as an off-white solid (1.81 g, 86%) which was used without purification in the next step.

Intermediate 5

Method A

[108] Intermediate 5: (R)-6-fluoro-3-(3-fluorophenyl)-2-(l-hydroxyethyl)-4H-chromen-4-one: To intermediate 4 (1.75 g, 3.96 mmol) in methanol (17 ml) cooled to 10°C, potassium carbonate (0.273 g, 1.98 mmol) was added and stirred for 30 min. The reaction mixture was concentrated, acidified with 2N HC1 solution, extracted with ethyl acetate, dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow solid (1.05 g, 87%). Enantiomeric excess: 93.6%, enriched in the late eluting isomer (retention time: 11.12 min.) as determined by HPLC on a chiralpak AD-H column.

Method B:

[109] Step-1 : (R)-2-(l-(benzyloxy)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one : To l-(5-fluoro-2-hydroxyphenyl)-2-(3-fluorophenyl)ethanone (11.00 g, 44.31 mmol ) in dichloromethane, HATU (33.7 g, 88.63 mmol) and R-(+)2-benzyloxypropionic acid (9.58 g, 53.17 mmol) were added and stirred for 10 min. Triethylamine (66.7 ml, 0.47 mol) was added dropwise and stirred at RT for 24h. The reaction mixture was quenched with water, extracted with dichloromethane, dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow solid (10.5 g, 60%). ‘H-NMR (δ ppm, CDC1₃, 400 MHz): 7.85 (dd, J = 8.1,3 Hz, 1H), 7.58 (dd, J = 9.1, 4.1 Hz, 1H), 7.47-7.39 (m, 1H), 7.39-7.34 (m, 1H), 7.28-7.20 (m, 3H), 7.20-7.14 (m, 2H), 7.16-7.07 (m, 1H), 6.99-6.89 (m, 2H), 4.50-4.31 (m, 3H), 1.56 (d, J = 6.4 Hz, 3H).

[110] Step-2 : (R)-2-(l-(benzyloxy)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (10.5 g, 26.69 mmol) in dichloromethane (110 ml) cooled to 0°C, aluminium chloride (5.35 g, 40.03 mmol) was added portionwise and stirred at RT for 6h. The reaction mixture was quenched with 2N HC1 solution, extracted with dichloromethane, dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the desired intermediate as a yellow solid (6.1 g, 76%). Enantiomeric excess: 97.7%, enriched in the late eluting isomer (retention time: 11.12 min.) as determined by HPLC on a chiralpak AD-H column.

Intermediate 13

[121] Intermediate 13: 3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-4-amine: To a solution of 3-iodo-lH-pyrazolo[3,4-d]pyrimidin-4-amine (11.0 g, 42.14 mmol) in DMF 110 ml), ethanol (55 ml) and water (55 ml), intermediate 12 (23.4 g, 84.28 mmol) and sodium carbonate (13.3 g, 126.42 mmol) were added and degassed for 30 min. Tetrakis(triphenylphosphine)palladium(0) (2.4 g, 2.10 mmol) was added under nitrogen atmosphere and heated to 80°C. After 12h, the reaction mixture was filtered though celite, concentrated and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was triturated with diethyl ether, filtered and dried under vacuum to afford the title compound as light brown solid (3.2 g, 26% yield) which is used as such for the next step.

Example Bl

(S)-2-(l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l- yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one

[127] To a solution of intermediate 13 (0.134 g, 0.494 mmol) in THF (2.0 ml), intermediate 5 (0.150 g, 0.494 mmol) and triphenylphosphine (0.194 g, 0.741 mml) were added and stirred at RT for 5 min. Diisopropylazodicarboxylate ( 0.15 ml, 0.749 mmol) was added heated to 45°C. After 2h, the reaction mixture was quenched with with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate : petroleum ether to afford the title compound as an off-white solid (0.049 g, 20 %). MP: 139-142°C. Mass : 571.7 (M H-NMR (δ ppm, CDC1_3, 400 MHz): 8.24 (s, 1H), 7.85 (dd, J = 8.2,3.1 Hz, 1H), 7.50-7.29 (m, 5H), 7.14 (t, J = 8.4 Hz, 1H), 7.02 (m, 2H), 6.92 (d, J = 8.4 Hz, 1H), 6.11 (q, J = 7.1 Hz, 1H), 5.40 (s, 2H), 4.66 (quintet, J = 6.1 Hz, 1H), 2.00 (d, J = 7.1Hz, 3H), 1.42 (d, J = 6.1 Hz, 6H). Enantiomeric excess: 89.8% as determined by HPLC

on a chiralpak AD-H column, enriched in the fast eluting isomer (retention time = 10.64min.).

PATENT

US 2014/0011819 describe the synthesis of TGR-1202 (Example B l)

PATENT

US 20150290317

US 20150174263

WO 2014071125

WO 2014006572

WO 2013188763*

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^ “Therapy Focus –- TG Could Benefit From Zydelig Setback”. Seeking Alpha. 29 March 2016.
^ “TG Therapeutics, Inc. Announces First Patient Enrolled in the Registration-Directed UNITY-DLBCL Phase 2b Trial”. TG Therapeutics Inc. June 2016.
^ Clinical trial number NCT02268851 for “A Phase I/Ib Safety and Efficacy Study of the PI3K-delta Inhibitor TGR-1202 and Ibrutinib in Patients With CLL or MCL” at ClinicalTrials.gov
^ “Follow-Up Data for Combination of TGR-1202 (umbralisib) plus Ibrutinib in Patients with Relapsed or Refractory CLL and MCL”(Press release). TG Therapeutics. 14 June 2017 – via Globenewswire.
^ Clinical trial number NCT02793583 for “Study to Assess the Efficacy and Safety of Ublituximab + TGR-1202 With or Without Bendamustine and TGR-1202 Alone in Patients With Previously Treated Non-Hodgkin’s Lymphoma (UNITY-NHL)” at ClinicalTrials.gov
^ Columbus G (22 January 2019). “FDA Grants Umbralisib Breakthrough Designation for Marginal Zone Lymphoma”. OncLive. Archived from the original on 23 January 2019.
^ “Orphan Treatment of extranodal marginal zone lymphoma”. U.S. Food and Drug Administration (FDA). 11 April 2019. Retrieved 5 February 2021.
^ “Orphan Treatment of splenic marginal zone lymphoma”. U.S. Food and Drug Administration (FDA). 11 April 2019. Retrieved 5 February 2021.
^ “Orphan Treatment of Follicular Lymphoma”. U.S. Food and Drug Administration (FDA). 11 April 2019. Retrieved 5 February2021.
^ “Orphan Treatment of nodal marginal zone lymphoma”. U.S. Food and Drug Administration (FDA). 11 April 2019. Retrieved 5 February 2021.

External links

“Umbralisib”. Drug Information Portal. U.S. National Library of Medicine.
“Umbralisib”. NCI Drug Dictionary. National Cancer Institute.

Clinical data

Trade names	Ukoniq
Other names	RP5264; TGR-1202
License data	US DailyMed: Umbralisib
Pregnancy category	Not recommended^[1]
Routes of administration	By mouth
ATC code	None
Legal status
Legal status	US: ℞-only ^[1]^[2]
Pharmacokinetic data
Metabolism	CYP2C9, CYP3A4, and CYP1A2^[1]
Elimination half-life	91 h^[1]
Excretion	Feces, urine^[1]
Identifiers
IUPAC name [show]
CAS Number	1532533-67-7
PubChem CID	72950888
DrugBank	DB14989
ChemSpider	34979945
UNII	38073MQB2A
ChEMBL	ChEMBL3948730
Chemical and physical data
Formula	C₃₁H₂₄F₃N₅O₃
Molar mass	571.560 g·mol⁻¹
3D model (JSmol)	Interactive image
SMILES [hide]CC(C)OC1=C(C=C(C=C1)C2=NN(C3=NC=NC(=C23)N)C(C)C4=C(C(=O)C5=C(O4)C=CC(=C5)F)C6=CC(=CC=C6)F)F

Feb. 9, 2021 04:45 UTC Rhizen Pharmaceuticals AG Announces That Its Partnered Asset, Umbralisib (UKONIQ™), Has Received US FDA Accelerated Approval for Adult Patients With Relapsed or Refractory MZL & FL

Umbralisib (UKONIQ™) granted accelerated approval by US FDA for the treatment of adult patients with relapsed or refractory marginal zone lymphoma (MZL), follicular lymphoma (FL).

Umbralisib, a novel next generation inhibitor of PI3K delta & CK1 epsilon, was discovered by Rhizen Pharmaceuticals and subsequently licensed to TG Therapeutics, who led the asset’s clinical development.

Rhizen and its affiliate Alembic Pharma to support TG Therapeutics towards UKONIQ’s commercialization as its manufacturing & supply partner; Rhizen plans to register and commercialize Umbralisib in India.

BASEL, Switzerland–(BUSINESS WIRE)–Rhizen Pharmaceuticals, a clinical-stage oncology-focused biopharmaceutical company, today announced that its novel next generation PI3K-delta inhibitor, Umbralisib, which was licensed to TG Therapeutics (NASDAQ:TGTX), has secured US FDA accelerated approval for the treatment of:

adult patients with relapsed or refractory marginal zone lymphoma (MZL) who have received at least one prior anti-CD20 based regimen, and

adult patients with relapsed or refractory follicular lymphoma (FL) who have received at least three prior lines of systemic therapy.

Accelerated approval was granted for these indications, under a priority review (MZL), based on the results of the Phase 2 UNITY-NHL Trial (NCT02793583); in MZL, an ORR of 49% with 16% complete responses and in FL an ORR of 43% with 3% complete responses were achieved, respectively. Umbralisib was earlier granted Breakthrough Therapy Designation (BTD) for the treatment of MZL and orphan drug designation (ODD) for the treatment of MZL and FL.

Umbralisib is a novel, next generation, oral, once daily, inhibitor of phosphoinositide 3 kinase (PI3K) delta and casein kinase 1 (CK1) epsilon and was discovered by Rhizen Pharma and subsequently licensed to TG Therapeutics (NASDAQ:TGTX) at an IND stage (TGR 1202) in 2012. In 2014, both parties entered into a licensing agreement as a part of which TGTX obtained worldwide rights and Rhizen has retained commercialization rights for India while also being the manufacturing and supply partner for Umbralisib.

Swaroop Vakkalanka, President & CEO of Rhizen Pharmaceuticals said: “Umbralisib’s approval offers MZL & FL patients a new treatment option and is a huge validation of Rhizen’s drug discovery & development capabilities. This is a momentous occasion in Rhizen’s journey as a successful biotech that speaks of the true ability of our team to discover & develop safe and effective therapies that can last the rigors of drug development. Further, we are keen to bring Umbralisib to Indian patients and we plan to initiate activities towards registration and approval there soon.”

Pranav Amin, Chairman, Rhizen Pharmaceuticals & Managing Director of Alembic Pharmaceuticals Ltd said: “We are extremely proud of this historic milestone for Rhizen, and of the fact that Umbralisib is the first NCE discovered by Indian scientists to secure a US FDA approval. We are committed to working together with TG Therapeutics and Rhizen Pharma to ensure uninterrupted supply of UKONIQ™. Umbralisib is the first discovery asset to come out of Rhizen’s R&D efforts and this approval heralds the promise of the rest of Rhizen’s deep pipeline and continuing efforts.”

About Umbralisib:

Umbralisib is the first and only oral inhibitor of phosphoinositide 3 kinase (PI3K) delta and casein kinase 1 (CK1) epsilon. PI3K-delta is known to play an important role in supporting cell proliferation and survival, cell differentiation, intercellular trafficking and immunity and is expressed in both normal and malignant B-cells. CK1-epsilon is a regulator of oncoprotein translation and has been implicated in the pathogenesis of cancer cells, including lymphoid malignancies. Umbralisib is indicated for the treatment of adult patients with relapsed or refractory marginal zone lymphoma (MZL) who have received at least one prior anti-CD20-based regimen and for the treatment of adult patients with relapsed or refractory follicular lymphoma (FL) who have received at least three prior lines of systemic therapy. These indications are approved under accelerated approval based on overall response rate. Continued approval for this indication may be contingent upon verification and description of clinical benefit in a confirmatory trial. More information on Umbralisib or UKONIQ™ can be found at https://www.tgtherapeutics.com/prescribing-information/uspi-ukon.pdf.

About Alembic Pharmaceuticals Ltd:

Alembic Pharmaceuticals Limited, a vertically integrated research and development pharmaceutical company, has been at the forefront of healthcare since 1907. Headquartered in India, Alembic is a publicly listed company that manufactures and markets generic pharmaceutical products all over the world. Alembic’s state of the art research and manufacturing facilities are approved by regulatory authorities of many developed countries including the USFDA. Alembic is one of the leaders in branded generics in India. Alembic’s products that are marketed through a marketing team of over 5000 are well recognized by doctors and patients.

Information about Alembic can be found at http://www.alembicpharmaceuticals.com/.

(Reuters: ALEM.NS) (Bloomberg: ALPM) (NSE: APLL TD) (BSE: 533573)

About Rhizen Pharmaceuticals A.G.:

Rhizen Pharmaceuticals is an innovative, clinical-stage biopharmaceutical company focused on the discovery and development of novel onco-therapeutics. Since its establishment in 2008, Rhizen has created a diverse pipeline of proprietary drug candidates targeting several cancers and immune associated cellular pathways. Rhizen is headquartered in Basel, Switzerland. For additional information, please visit http://www.rhizen.com.

View source version on businesswire.com: https://www.businesswire.com/news/home/20210208005742/en/ Contacts

////////////ウムブラリシブトシル酸塩 , Umbralisib, fda 2021, 2021 approvals, TGR 1202, TGR-1202-101, RP 5264, Umbralisib tosylate, RP-5307 , TGR-1202, TGR-1202 PTSA, FU8XW5V3FS , RP-5264, AK173784,

old post pasted

TGR 1202, TGR-1202-101, RP 5264, UmbralisibAK173784;(S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one(S)-2-(l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-3-(3-fluorophenyl)-4H-chromen-4-one,2-[(1S)-1-[4-amino-3-(3-fluoro-4-propan-2-yloxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]ethyl]-6-fluoro-3-(3-fluorophenyl)chromen-4-one CAS TOSYLATE 1532533-72-4 Umbralisib tosylateCAS 1532533-67-7, 1514919-95-9

Molecular Formula:	C₃₁H₂₄F₃N₅O₃
Molecular Weight:	571.54917 g/mol

RP-5307
TGR-1202
TGR-1202 PTSA
FU8XW5V3FS (UNII code)
RP-5264 (free base)

A PI3K inhibitor potentially for treatment of chronic lymphocytic leukemia, leukemia，lymphoma，B-cell

TGR‐1202, a next generation PI3K-δ delta inhibitor. TGR-1202 (RP-5264) is a highly specific, orally available, PI3K delta inhibitor, targeting the delta isoform with nanomolar potency and several fold selectivity over the alpha, beta, and gamma isoforms of PI3K.

TG Therapeutics, under license from Rhizen Pharmaceuticals, is developing TGR-1202 (structure shown; formerly RP-5264), a lead from a program of PI3K delta inhibitors, for the potential oral treatment of hematological cancers including Hodgkin lymphoma, non-Hodgkin lymphoma (NHL), chronic lymphocytic leukemia (CLL), B-cell lymphoma and mantle cell lymphoma (MCL)

Incozen Therapeutics Pvt Ltd

TG Therapeutics

TGR-1202 potential to perform as the best PI3K inhibitor in its class and the possible superiority of TG-1101 over Rituxan®.

	Rhizen Pharmaceuticals S.A.
Description	Phosphoinositide 3-kinase (PI3K) delta inhibitor

Leukemia, chronic lymphocytic PHASE 3, TG Therapeutics

Orphan Drug

Umbralisib is a novel phosphatidylinositol 3-kinase delta (PI3Kdelta) inhibitor under development at TG Therapeutics in phase III clinical trials, in combination with ublituximab, for the treatment of chronic lymphocytic leukemia (CLL) and for the treatment of diffuse large B-cell lymphoma (DLBCL). The company refers to the combination regimen of ublituximab and TGR-1202 as TG-1303. The drug is also in phase II clinical development for the oral treatment of hematologic malignancies, as a single agent or in combination therapy. Phase I clinical trials are ongoing in patients with select relapsed or refractory solid tumors, such as adenocarcinoma of the pancreas, adenocarcinoma of the colon, rectum, gastric and GE junction cancer, and GI Stromal Tumor (GIST).

In 2016, orphan drug designation was assigned to the compound in the U.S. for the treatment of CLL. In 2017, additional orphan drug designation was granted in the U.S. for the treatment of CLL and DLBCL, in combination with ublituximab.

Originated by Rhizen Pharmaceuticals, the product was jointly developed by Rhizen Pharmaceuticals and TG Therapeutics since 2012. In 2014, exclusive global development and commercialization rights (excluding India) were licensed to TG Therapeutics.

CLINICAL TRIALS……….https://clinicaltrials.gov/search/intervention=TGR-1202

B-cell lymphoma; Chronic lymphocytic leukemia; Hematological neoplasm; Hodgkins disease; Mantle cell lymphoma; Non-Hodgkin lymphoma

Phosphoinositide-3 kinase delta inhibitor

SYNTHESIS

Rhizen Pharmaceuticals Announces Out-licensing Agreement for TGR-1202, a Novel Next Generation PI3K-delta Inhibitor

Rhizen to receive upfront payment of $8.0 million — Rhizen to retain global manufacturing and supply rights — Rhizen to retain development and commercialization for India

Rhizen to retain development and commercialization for India

http://globenewswire.com/news-release/2014/09/23/667853/10099642/en/Rhizen-Pharmaceuticals-Announces-Out-licensing-Agreement-for-TGR-1202-a-Novel-Next-Generation-PI3K-delta-Inhibitor.html?parent=794070#

September 23, 2014 09:00 ET | Source: Rhizen Pharmaceuticals SA

La Chaux-de-Fonds, Switzerland, Sept. 23, 2014 (GLOBE NEWSWIRE) — Rhizen Pharmaceuticals S.A. today announced an out-licensing agreement for TGR-1202, a novel next generation PI3K-delta inhibitor. TG Therapeutics exercised its option for early conversion to a licensing agreement from a 50:50 joint venture partnership.

In exchange for this licensing agreement, TG Therapeutics will pay Rhizen an upfront payment of $8.0 million ($4.0 million in cash and $4.0 million in TG Therapeutics common stock). In addition to the upfront payment, Rhizen will be eligible to receive regulatory filing, approval and sales based milestones in the aggregate of approximately $240 million, and tiered royalties based on net sales.

Swaroop Vakkalanka, Ph.D. and President of Rhizen stated, “We are extremely happy and take pride in discovering a novel, next generation, once-daily PI3K-delta inhibitor under active development led by TG Therapeutics. We are encouraged by the progress of TRG-1202 to date, and the speed at which TG Therapeutics is developing the asset in various hematological malignancies. We look forward to the day this novel drug reaches cancer patients in need of new and safe therapies.”

About Rhizen Pharmaceuticals S.A.:

Rhizen Pharmaceuticals is an innovative, clinical-stage biopharmaceutical company focused on the discovery and development of novel therapeutics for the treatment of cancer, immune and metabolic disorders. Since its establishment in 2008, Rhizen has created a diverse pipeline of proprietary drug candidates targeting several cancers and immune associated cellular pathways. Rhizen is headquartered in La-Chaux-de-Fonds, Switzerland. For additional information, please visit Rhizen’s website, www.rhizen.com.

TGR-1202.with Idelalisib and IPI-145 (left to right) for comparison.

IPI 145

PATENTS

WO 2011055215

http://www.google.com/patents/WO2011055215A2?cl=en

PATENT

WO 2015181728

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2015181728

TGR-1202, chemically known as (S)-2-(l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-3-(3-fluorophenyl)-4H-chromen-4-one, has the following chemical structure:

Example 1: Preparation of the PTSA Salt of TGR-1202 (Form A)

7100 g of TGR-1202 was charged in a reactor containing 56.8 litres of acetone and stirred at ambient temperature. 4680 g of p-toluene sulphonic acid was added and the reaction mixture was heated at a temperature of 60-65° C for about 6 hours. The solvent was removed by distillation under reduced pressure to obtain a wet residue. The wet residue was degassed and allowed to cool to < 20° C. Approximately 142 litres of diethyl ether was then added and the resulting mixture was stirred overnight, then filtered to obtain a solid mass which was washed with diethyl ether and dried in vacuo to yield a solid mass. The solid mass was re-suspended in diethyl ether, stirred for 6 hours, and then filtered to yield a solid mass which was subsequently dissolved in 56.8 litres of acetone, filtered through a HiFlow bed, and concentrated under reduced pressure. The resulting residue mass was stirred with water overnight, then filtered and vacuum dried to yield 6600 g of the PTSA salt of TGR-1202. HPLC: 99.21% and chiral purity of 99.64:0.36 (S:R).

Example 2: Preparation of the PTSA Salt of TGR-1202 (Form B)

Table 1 lists the XRPD pattern peaks and relative peak intensities for the products of Examples 1 and 2.

TABLE 1

The tablet composition comprising a PTSA salt of TGR-1202 prepared according to Example 2 exhibited a Cmax about 2.5 fold and an area under the curve (AUC) about 1.9 fold greater than that of the tablet composition comprising a PTSA salt of TGR-1202 prepared according to Example 1. The results are provided in Table 8 below.

TABLE 8

PATENT

WO 2014071125

http://www.google.com/patents/WO2014071125A1?cl=en

formula (A) that is a ΡΒΚδ selective inhibitor,

(A)

Synthesis of Compound of Formula A

Unless otherwise stated, purification implies column chromatography using silica gel as the stationary phase and a mixture of petroleum ether (boiling at 60-80°C) and ethyl acetate or dichloromethane and methanol of suitable polarity as the mobile phases. The term “RT” refers to ambient temperature (25-28°C).

Intermediate 1 : 2-( l-bromoethyl)-6-fluoro-3-f3-fluorophenyl)-4H-chromen-4-one

Step-1 [l-(5-Fluoro-2-hydroxyphenyl)-2-(3-fluorophenyl)ethanone]: 3- Fluorophenylacetic acid (7.33 g, 47.56 mmoles) was dissolved in 25 ml dichloromethane. To this mixture, oxalylchloride (7.54 g, 59.46 mmoles) and DMF (3 drops) were added at 0°C and stirred for 30 min. The solvent was evaporated and dissolved in 25 ml dichloromethane. To this mixture, 4-fluoroanisole (5.00 g, 39.64 mmoles) was added and cooled to 0°C. At 0°C A1C1₃ (7.95 g, 59.46 mmoles) was added and the reaction mixture was warmed to RT and stirred for 12 hours. The reaction mixture was quenched by the addition of 2N HC1, extracted with ethyl acetate, dried over sodium sulphate and concentrated. The crude product was purified by column chromatography with ethyl acetate :petroleum ether to afford the title compound as colorless solid (4.5 g, 45% yield). 1H-NMR (δ ppm, DMSO-D₆, 400 MHz): δ 11.34 (s, 1H), 7.75 (dd, J=9.4, 3.1 Hz, 1H), 7.42 (m, 2H), 7.12 (m, 3H), 7.05 (dd, J=9.0, 4.5 Hz, 1H), 4.47 (s, 2H).

Step-2 [2-Ethyl-6-fiuoro-3-(3-fluorophenyl)-4H-chromen-4-one]: l-(5-Fluoro-2- hydroxyphenyl)-2-(3-fluorophenyl)ethanone obtained from Step-1 (3.00 g, 12.08 mmoles) was placed in a round bottom flask and to this triethylamine (25 ml) and propionic anhydride (4.92 g, 37.82 mmoles) were added, and the mixture was refluxed for 24 hours. After cooling to RT, the reaction mixture was acidified by the addition of IN HC1 solution, extracted with ethyl acetate, washed with sodium bicarbonate solution, dried with sodium sulphate and concentrated. The crude product was purified by column chromatography with ethyl acetate :petroleum ether to afford the title compound as off-yellow solid (1.80 g, 52% yield). 1H-NMR (δ ppm, DMSO-D₆, 400 MHz): δ 7.80 (m, 1H), 7.76 (m, 2H), 7.51 (dd, J=8.0, 6.4 Hz), 7.22 (m, 1H), 7.18 (m, 2H), 2.56 (q, J=7.6 Hz, 2H), 1.20 (t, J=7.6 Hz, 3H).

Step-3: To a solution of 2-Ethyl-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one obtained from Step-2 (1.80 g, 6.28 mmoles) in carbon tetrachloride (20 ml), N- bromosuccinimide (1.11 g, 6.28 mmoles) was added and heated to 80°C. Azobisisobutyronitrile (10 mg) was added to the reaction mixture at 80°C. After 12 hours, the reaction mixture was cooled to RT, diluted with dichloromethane and washed with water. The organic layer was dried over sodium sulphate and concentrated under reduced pressure to afford the crude title compound as yellow solid (1.25 g, 55% yield). 1H-NMR (δ ppm, DMSO-D₆, 400 MHz): δ 7.91 (dd, J=9.2, 4.3 Hz, 1H), 7.81 (dt, j=8.2, 2.8 Hz, 1H), 7.74 (dd, J=8.3, 3.1 Hz, 1H), 7.57 (m, 1H), 7.32 (dt, J=8.5, 2.4 Hz, 1H), 7.19 (m, 2H), 5.00 (q, J=6.8 Hz, 1H), 1.97 (d, J=6.8 Hz, 3H).

Intermediate 2: 6-fluoro-3-f3-fluorophenyl)-2-fl-hvdroxyethyl)-4H-chromen-4-one

To a solution of Intermediate 1 (15.0 g, 40.84 mmol) in DMSO (150 ml), n-butanol (7.5 ml) was added and heated to 120°C for 3 hours. The reaction mixture was cooled to RT, quenched with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as an off-white solid (7.90 g, 64%). 1H-NMR (δ ppm, CDC1₃, 400 MHz): 7.85 (dd, J = 8.1, 3 Hz, 1H), 7.54 (dd, J = 9.2, 4.2 Hz, 1H), 7.47-7.37 (m, 2H), 7.15-6.98 (m, 3H), 4.74 (quintet, J= 6.8 Hz, 1H), 2.23 (d, J = 7.4 Hz, 1H), 1.54 (d, J = 6.6 Hz, 3H).

Intermediate 3 : 2-acetyl-6-fluoro-3-( 3-fluorophenyl)-4H-chromen-4-one

DMSO (5.60 ml, 79.14 mmol) was added to dichloromethane (40 ml), and cooled to – 78°C, followed by oxalyl chloride (3.40 ml, 39.57 mmol). After 10 min., intermediate 2 (6.00 g, 19.78 mmol) in dichloromethane (54 ml) was added dropwise and stirred for 20 min.

Triethylamine (12 ml) was added and stirred for 1 hour. The reaction mixture was quenched with water and extracted with dichloromethane. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow solid (4.2 g, 71%) which was used as such in the next step.

Intermediate 4: fS)-6-fluoro-3-f3-fluorophenyl)-2-fl-hvdroxyethyl)-4H-chromen-4-one

To intermediate 3 (2.00 g, 6.66 mmol), R-Alpine borane (0.5 M in THF, 20 ml) was added and heated to 60°C for 20 hours. The reaction mixture quenched with 2N HC1, and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as an off-white solid (1.51 g, 75%).

Enantiomeric excess: 94.2%, enriched in the fast eluting isomer (retention time: 8.78 min.) as determined by HPLC on a chiralpak AD-H column.

Intermediate 5: fR)-l-f6-fluoro-3-f3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl 4- chlorobenzoate

To a solution of intermediate 4 (1.45 g, 4.78 mmol) in THF (15 ml), 4-chlorobenzoic acid (0.748 g, 4.78 mmol) and triphenylphosphine (1.88 g, 7.17 mmol) were added and heated to 45°C followed by diisopropylazodicarboxylate (1.4 ml, 7.17 mmol). After 1 hour, the reaction mixture was concentrated and the residue was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as an off-white solid (1.81 g, 86%) which was used without purification in the next step. Intermediate 6: fR)-6-fluoro-3-f3-fluorophenyl)-2-fl-hvdroxyethyl)-4H-chromen-4-one

Method A

Intermediate 5 (1.75 g, 3.96 mmol) in methanol (17 ml) was cooled to 10°C, potassium carbonate (0.273 g, 1.98 mmol) was added and stirred for 30 min. The reaction mixture was concentrated, acidified with 2N HCl solution, extracted with ethyl acetate, dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow solid (1.05 g, 87% yield). Enantiomeric excess: 93.6%>, enriched in the late eluting isomer (retention time: 11.12 min.) as determined by HPLC on a chiralpak AD-H column.

Method B

Step-1 [(R)-2-(l-(benzyloxy)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one]: To l-(5-fluoro-2-hydroxyphenyl)-2-(3-fluorophenyl)ethanone (11.00 g, 44.31 mmol) in dichloromethane, HATU (33.7 g, 88.63 mmol) and R-(+)2-benzyloxypropionic acid (9.58 g, 53.17 mmol) were added and stirred for 10 min. Triethylamine (66.7 ml, 0.47 mol) was added dropwise and stirred at RT for 24 hours. The reaction mixture was quenched with water, extracted with dichloromethane, dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate:

petroleum ether to afford the title compound as a yellow solid (10.5 g, 60%> yield). 1H-NMR (δ ppm, CDCls, 400 MHz): 7.85 (dd, J = 8.1,3 Hz, 1H), 7.58 (dd, J = 9.1, 4.1 Hz, 1H), 7.47-7.39 (m, 1H), 7.39-7.34 (m, 1H), 7.28-7.20 (m, 3H), 7.20-7.14 (m, 2H), 7.16-7.07 (m, 1H), 6.99-6.89 (m, 2H), 4.50-4.31 (m, 3H), 1.56 (d, J = 6.4 Hz, 3H).

Step-2: (R)-2-(l-(benzyloxy)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one obtained in Step-1 (10.5 g, 26.69 mmol) in dichloromethane (110 ml) was cooled to 0°C, aluminium chloride (5.35 g, 40.03 mmol) was added portionwise and stirred at RT for 6 hours. The reaction mixture was quenched with 2N HCl solution, extracted with dichloromethane, dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford intermediate 6 a yellow solid (6.1 g, 76% yield). Enantiomeric excess: 97.7%, enriched in the late eluting isomer (retention time: 11.12 min.) as determined by HPLC on a chiralpak AD-H column.

Intermediate 7: 4-bromo-2-fluoro-l-isopropoxybenzene

To a solution of 4-bromo-3-fluorophenol (10 g, 52.35 mmol) in THF (100ml), isopropyl alcohol (4.8 ml, 62.62 mmol) and triphenylphosphine (20.6 g, 78.52 mmol) were added and heated to 45°C followed by diisopropylazodicarboxylate (15.4 ml, 78.52 mmol). The mixture was refluxed for 1 hour, concentrated and the residue was purified by column

chromatography with ethyl acetate: petroleum ether to afford the title compound as a colorless liquid (13.1 g, 99% yield), which was used without purification in the next step.

Intermediate 8: 2-f3-fluoro-4-isopropoxyphenyl)-4,4,5.,5-tetramethyl-l,3i2-dioxaborolane

Potassium acetate (10.52 g, 107.2 mmol) and bis(pinacolato)diboron (15 g, 58.96 mmol) were added to a solution of intermediate 7 (10.52 g, 107.2 mmol) in dioxane (125 ml), and the solution was degassed for 30 min. [l, -Bis(diphenylphosphino)ferrocene]dichloro palladium(II) CH₂CI₂ (4.4 g, 5.36 mmol) was added under nitrogen atmosphere and heated to 80°C. After 12 hours, the reaction mixture was filtered through celite and concentrated. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow oil (13.9g, 99%) which was used without purification in the next step.

Intermediate 9: 3-f3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3.,4-dlpyrimidin-4-amine

To a solution of 3-iodo-lH-pyrazolo[3,4-d]pyrimidin-4-amine (11.0 g, 42.14 mmol) in DMF (110 ml), ethanol (55 ml) and water (55 ml), intermediate 8 (23.4 g, 84.28 mmol) and sodium carbonate (13.3 g, 126.42 mmol) were added and degassed for 30 min.

Tetrakis(triphenylphosphine)palladium(0) (2.4 g, 2.10 mmol) was added under nitrogen atmosphere and heated to 80°C. After 12 hours, the reaction mixture was filtered through celite, concentrated and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was triturated with diethyl ether, filtered and dried under vacuum to afford the title compound as light brown solid (3.2 g, 26% yield) which is used as such for the next step.

(RS)- 2-fl-f4-amino-3-f3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3.,4-(ilpyrimi(iin-l- yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one

To a solution of intermediate 9 (0.080 g, 0.293 mmol) in DMF (2 ml), potassium carbonate (0.081 g, 0.587 mmol) was added and stirred at RT for 10 min. To this mixture intermediate 1 (0.215 g, 0.587 mmol) was added and stirred for 12 hours. The reaction mixture was diluted with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with methanol: dichloromethane to afford the title compound as a pale yellow solid (0.045 g). MP: 175-177°C. 1H-NMR (δ ppm, DMSO-D₆, 400 MHz): δ 8.20 (s, 1H), 7.85 (dd, J = 81, 3.0 Hz, 1H), 7.48-7.33 (m, 5H), 7.14 (t, J= 8.3 Hz, 1H), 7.02 (m, 2H), 6.90 (m, 1H), 6.10 (q, J = 7.1 Hz, 1H), 5.42 (s, 2H), 4.64 (quintet, J = 6.0 Hz, 1H), 1.99 (d, J = 7.1 Hz, 3H), 1.42 (d, J= 6.1 Hz, 6H).

fS)-2-fl-f4-amino-3-f3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3.,4-(ilpyrimi(iin-l- yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (“S-isomer”)

To a solution of intermediate 9 (0.134 g, 0.494 mmol) in THF (2.0 ml), intermediate 6 (0.150 g, 0.494 mmol) and triphenylphosphine (0.194 g, 0.741 mml) were added and stirred at RT for 5 min. Diisopropylazodicarboxylate (0.15 ml, 0.749 mmol) was added heated to 45°C. After 2 hours, the reaction mixture was quenched with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate : petroleum ether to afford the title compound as an off-white solid (0.049 g, 20 % yield). MP: 139-142°C. Mass: 571.7 (M⁺). Enantiomeric excess: 89.8% as determined by HPLC on a chiralpak AD-H column, enriched in the fast eluting isomer (retention time = 10.64 min.). fR)-2-fl-f4-amino-3-f3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3.,4-(ilpyrimi(iin-l- yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-ehromen-4-one

To a solution of intermediate 8 (0.284 g, 0.989 mmol) in THF (5.0 ml), intermediate 4 (0.250 g, 0.824 mmol) and tris(4-methoxy)phenylphosphine (0.435 g, 1.23 mml) were added and stirred at RT for 5 min. Diisopropylazodicarboxylate (0.25 ml, 1.23 mmol) was added stirred at RT. After 12 hours, the reaction mixture was quenched with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate :

petroleum ether to afford the title compound as an off-white solid (0.105 g, 22 % yield). MP: 145-148°C. Mass: 571.7 (M⁺). Enantiomeric excess: 95.4% as determined by HPLC on a chiralpak AD-H column, enriched in the late eluting isomer (retention time = 14.83 min.).

PATENT

WO 2014006572

http://www.google.com/patents/WO2014006572A1?cl=en

Figure imgf000005_0001 B1 IS DESIRED

(S)-2- (l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-6- fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (compound-B l)

Intermediate 11

[119] Intermediate 11: 4-bromo-2-fluoro-l-isopropoxybenzene:To a solution of 4-bromo-2- fluorophenol (lOg, 52.35 mmol) in THF (100ml), isopropyl alcohol (4.8ml, 62.62 mmol) and triphenylphosphine (20.6g, 78.52 mmol) were added and heated to 45 C followed by diisopropylazodicarboxylate (15.4ml, 78 52 mmol). The mixture was refluxed for lh, concentrated and the residue was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a colourless liquid (13. lg, 99%) which was used without purification in the next step. Intermediate 12

[120] Intermediate 12: 2-(3-fluoro-4-isopropoxyphenyl)-4,4,5,5-tetramethyl- 1,3,2- dioxaborolane: Potassium acetate (10.52 g, 107.2 mmol) and bis(pinacolato)diboron (15g, 58.96 mmol) were added to a solution of intermediate 11 (10.52 g, 107.2 mmol) in dioxane (125 ml), and the solution was degassed for 30 min. [1,1 ‘- Bis(diphenylphosphino)ferrocene]dichloro palladium(II).CH₂Cl₂ (4.4g, 5.36 mmol) was added under nitrogen atmosphere and heated to 80°C. After 12h the reaction mixture was filtered through celite and concentrated. The crude product was purified by column chromatography with ethyl acetate: petroleum ether to afford the title compound as a yellow oil (13.9g, 99%) which was used without purification in the next step.

Intermediate 13

[121] Intermediate 13: 3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-4- amine: To a solution of 3-iodo-lH-pyrazolo[3,4-d]pyrimidin-4-amine (11.0 g, 42.14 mmol) in DMF 110 ml), ethanol (55 ml) and water (55 ml), intermediate 12 (23.4 g, 84.28 mmol) and sodium carbonate (13.3 g, 126.42 mmol) were added and degassed for 30 min. Tetrakis(triphenylphosphine)palladium(0) (2.4 g, 2.10 mmol) was added under nitrogen atmosphere and heated to 80°C. After 12h, the reaction mixture was filtered though celite, concentrated and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was triturated with diethyl ether, filtered and dried under vacuum to afford the title compound as light brown solid (3.2 g, 26% yield) which is used as such for the next step.

Example Bl

(S)-2-(l-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l- yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one

[127] To a solution of intermediate 13 (0.134 g, 0.494 mmol) in THF (2.0 ml), intermediate 5 (0.150 g, 0.494 mmol) and triphenylphosphine (0.194 g, 0.741 mml) were added and stirred at RT for 5 min. Diisopropylazodicarboxylate ( 0.15 ml, 0.749 mmol) was added heated to 45°C. After 2h, the reaction mixture was quenched with with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate : petroleum ether to afford the title compound as an off-white solid (0.049 g, 20 %). MP: 139- 142°C. Mass : 571.7 (M H-NMR (δ ppm, CDC1_3, 400 MHz): 8.24 (s, 1H), 7.85 (dd, J = 8.2,3.1 Hz, 1H), 7.50-7.29 (m, 5H), 7.14 (t, J = 8.4 Hz, 1H), 7.02 (m, 2H), 6.92 (d, J = 8.4 Hz, 1H), 6.11 (q, J = 7.1 Hz, 1H), 5.40 (s, 2H), 4.66 (quintet, J = 6.1 Hz, 1H), 2.00 (d, J = 7.1Hz, 3H), 1.42 (d, J = 6.1 Hz, 6H). Enantiomeric excess: 89.8% as determined by HPLC on a chiralpak AD-H column, enriched in the fast eluting isomer (retention time = 10.64min.).

NEW DRUG APPROVALS

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PATENT

US 2014/0011819 describe the synthesis of TGR-1202 (Example B l)

http://www.google.co.in/patents/US20140011819

Example B1 (S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one

To a solution of intermediate 13 (0.134 g, 0.494 mmol) in THF (2.0 ml), intermediate 5 (0.150 g, 0.494 mmol) and triphenylphosphine (0.194 g, 0.741 mml) were added and stirred at RT for 5 min. Diisopropylazodicarboxylate (0.15 ml, 0.749 mmol) was added heated to 45° C. After 2 h, the reaction mixture was quenched with with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with ethyl acetate:petroleum ether to afford the title compound as an off-white solid (0.049 g, 20%). MP: 139-142° C. Mass: 571.7 (M⁺).¹H-NMR (δ ppm, CDCl₃, 400 MHz): 8.24 (s, 1H), 7.85 (dd, J=8.2, 3.1 Hz, 1H), 7.50-7.29 (m, 5H), 7.14 (t, J=8.4 Hz, 1H), 7.02 (m, 2H), 6.92 (d, J=8.4 Hz, 1H), 6.11 (q, J=7.1 Hz, 1H), 5.40 (s, 2H), 4.66 (quintet, J=6.1 Hz, 1H), 2.00 (d, J=7.1 Hz, 3H), 1.42 (d, J=6.1 Hz, 6H). Enantiomeric excess: 89.8% as determined by HPLC on a chiralpak AD-H column, enriched in the fast eluting isomer (retention time=10.64 min)

4-Methylbenzenesulfonate Salt of Compound B1 (S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one 4-methylbenzenesulfonate

(S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one 4-methylbenzenesulfonate: To (S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (22.7 g, 39.69 mmol) in isopropanol (600 ml), p-toluenesulphonic acid (8.30 g, 43.66 mmol) was added and refluxed for 1 h. The reaction mixture was concentrated, co-distilled with petroleum ether and dried. To the residue water (300 ml) was added and stirred for 30 min. The solid was filtered, washed with petroleum ether and dried under vacuum to afford the title compound as off-white solid (28.2 g, 95%). MP: 138-141° C. ¹H-NMR (δ ppm, CDCl₃, 400 MHz): 8.11 (s, 1H), 7.85 (dd, J=8.0, 3.0 Hz, 1H), 7.80 (d, J=8.2 Hz, 2H), 7.51 (dd, J=9.3, 4.3 Hz, 1H), 7.45 (dd, J=7.5, 3.1 Hz, 1H), 7.42-7.31 (m, 3H), 7.29 (m, 2H), 7.22 (d, J=8.0 Hz, 2H), 7.16 (t, J=8.3 Hz, 1H), 7.08 (dt, J=8.5, 2.5 Hz, 1H), 6.97 (br s, 1H), 6.88 (br s, 1H), 6.11 (q, J=7.2 Hz, 1H), 4.67 (quintet, J=6.0 Hz, 1H), 2.36 (s, 3H), 2.03 (d, J=7.1 Hz, 3H), 1.43 (d, J=6.0 Hz, 6H). Mass: 572.4 (M⁺+1-PTSA). Enantiomeric excess: 93.4% as determined by HPLC on a chiralpak AD-H column, enriched in the fast eluting isomer (retention time=12.35 min.)

Sulphate Salt of Compound B1 (S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one sulfate

(S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one sulphate: To (S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (15.0 g, 26.24 mmol) in isopropanol (600 ml) was cooled to 0° C. To this Sulphuric acid (2.83 g, 28.86 mmol) was added and stirred at room temperature for 24 h. The reaction mass was filtered and washed with petroleum ether and dried under vacuum. To the solid, water (150 ml) was added and stirred for 30 min. The solid was filtered, washed with petroleum ether and dried under vacuum to afford the title compound as off-white solid (13.5 g, 76%). MP: 125-127° C. ¹H-NMR (δ ppm, CDCl₃, 400 MHz): 8.11 (s, 1H), 7.85 (dd, J=8.0, 3.0 Hz, 1H), 7.51 (dd, J=9.2, 4.2 Hz, 1H), 7.45-7.31 (m, 3H), 7.29 (m, 1H), 7.15 (t, J=8.3 Hz, 1H), 7.08 (dt, J=8.5, 2.4 Hz, 1H), 6.96 (br s, 1H), 6.88 (br s, 1H), 6.09 (q, J=7.1 Hz, 1H), 4.676 (quintet, J=6.1 Hz, 1H), 2.01 (d, J=7.1 Hz, 3H), 1.42 (d, J=6.1 Hz, 6H). Mass: 572.2 (M⁺+1-H₂SO₄). Enantiomeric excess: 89.6% as determined by HPLC on a chiralpak AD-H column, enriched in the fast eluting isomer (retention time=12.08 min.)
Various other acid addition salts of compound B1 were prepared as provided in Table 1.
TABLE 1 Melting PointAcidMethod of preparation(° C.) Hydro-Compound B1 (1 eq.) dissolved in THF,130-132chloricexcess HCl/Et₂O was added, the clearacidsolution obtained was evaporated completely. The residue obtained was washed with water.p-Compound B1 (1 eq.) dissolved in138-141° C.Toluene-isopropyl alcohol (IPA), refluxed forsulfonic30 min., acid (1.1 eq.) in IPA was added,acidthe clear solution obtained was evaporated completely. The residue obtained was washed with water.Benzene-Compound B1 (1 eq.) dissolved in IPA,170-172sulphonicrefluxed for 30 min., acid(1.1 eq.) in IPAacidwas added, the clear solution not obtained, the residue was evaporated completely and was washed with water.MaleicCompound B1 (1 eq.) dissolved in IPA,107-109acidrefluxed for 30 min., acid (1.1 eq.) in IPA was added, the clear solution not obtained, the residue was evaporated completely and was washed with water.CamphorCompound B1 (1 eq.) dissolved in IPA,120-121sulfonicrefluxed for 30 min., acid (1.1 eq.) in IPAacidwas added, the clear solution not obtained, the residue was evaporated completely and was washed with water.SulphuricCompound B1 (1 eq.) dissolved in IPA,125-127acidrefluxed for 30 min., acid(1.1 eq.) in IPA was added, the clear solution obtained was evaporated completely. The residue obtained was washed with water.

REFERENCES

WO 2014/006572 and U.S. Patent Publication No. 2014/0011819,

http://www.tgtherapeutics.com/O’ConnorTGR202Single%20AgentEHA&Lugano2015.pdf

TGR-1202: Phase I/II started 09/28/2015Week in Review, Clinical Status Rhizen Pharmaceuticals S.A., La Chaux-de-Fonds, Switzerland TG Therapeutics Inc. (NASDAQ:TGTX), New York, N.Y. Product: TGR-1202 (formerly RP5264) Business: Cancer Molecular target: Phosphoinositide 3-kinase (PI3K) …
Ublituximab: Phase I/II started 09/28/2015Week in Review, Clinical Status LFB S.A., Les Ulis, France TG Therapeutics Inc. (NASDAQ:TGTX), New York, N.Y. Product: Ublituximab (TGTX-1101, TG-1101, LFB-R603) Business: Cancer Molecular target: CD20 Description: Glycoengineered mAb against CD20 …
COMPANY NEWS: TG rises on SPA for combination CLL therapy 09/17/2015The Daily Extra, Company News TG Therapeutics Inc. (NASDAQ:TGTX) rose $2.65 (23%) to $14.37 after the company said it received an SPA from FDA for the Phase III UNITY-CLL trial of ublituximab (TG-1101) in combination with TGR-1202 to treat chronic …
Targets & Mechanisms: The battle for IRAK 04/23/2015
Nimbus, Aurigene and TG Therapeutics are chasing IRAK4 inhibitors for cancerBC Innovations, Targets & Mechanisms Now that Nimbus has put IRAK4 on the map for B cell lymphoma, several companies are closing in with their own inhibitors, and they’re all on track for IND-enabling studies this year.
TGR-1202: Additional Phase I/II data 01/26/2015Week in Review, Clinical Results Rhizen Pharmaceuticals S.A., La Chaux-de-Fonds, Switzerland TG Therapeutics Inc. (NASDAQ:TGTX), New York, N.Y. Product: TGR-1202 (formerly RP5264) Business: Cancer Molecular target: Phosphoinositide 3-kinase (PI3K) …
Ublituximab: Additional Phase I/II data 01/26/2015Week in Review, Clinical Results LFB S.A., Les Ulis, France TG Therapeutics Inc. (NASDAQ:TGTX), New York, N.Y. Ildong Pharmaceutical Co. Ltd. (KSE:000230), Seoul, South Korea Product: Ublituximab (TGTX-1101, TG-1101, LFB-R603) Business: Cancer …
TGR-1202: Phase I started 12/15/2014Week in Review, Clinical Status Rhizen Pharmaceuticals S.A., La Chaux-de-Fonds, Switzerland TG Therapeutics Inc. (NASDAQ:TGTX), New York, N.Y. Product: TGR-1202 (formerly RP5264) Business: Cancer Molecular target: Phosphoinositide 3-kinase (PI3K) …
Rhizen, TG Therapeutics deal 12/08/2014Week in Review, Deals Rhizen Pharmaceuticals S.A., La Chaux-de-Fonds, Switzerland TG Therapeutics Inc. (NASDAQ:TGTX), New York, N.Y. Business: Cancer TG Therapeutics exercised an option under a 2012 deal to license exclusive, worldwide …

Patent	Submitted	Granted
NOVEL SELECTIVE PI3K DELTA INHIBITORS [US2014011819]	2013-07-02	2014-01-09
Treatment Of Cancers Using PI3 Kinase Isoform Modulators [US2014377258]	2014-05-30	2014-12-25

////////Umbralisib

CC(C)OC1=C(C=C(C=C1)C2=NN(C3=C2C(=NC=N3)N)C(C)C4=C(C(=O)C5=C(O4)C=CC(=C5)F)C6=CC(=CC=C6)F)F

PATENT

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2021164789&_cid=P12-KSZQ3G-94695-1Phosphatidylinositol 3-kinase (phosphatidylinositol-3-kinase, PI3K) is composed of the regulatory subunit p85 or p101, and the catalytic subunit p110 (subdivided into four subtypes: p110a, p110b, p110g, and p110d) Lipid kinase catalyzes the phosphorylation of the inositol ring 3′-OH of phosphatidylinositol 4,5-bisphosphate (phosphatidylinositol 4,5-bisphosphate, PIP2) to phosphatidylinositol 3,4,5-triphosphate (phosphatidylinositol 4,5-bisphosphate, PIP2). 3,4,5-trisphosphate, PIP3) and activate downstream Akt, which plays a key role in cell proliferation, survival and metabolism. In tumor cells, PI3K is overexpressed, which leads to rapid proliferation and growth of tumor cells.
The tumor suppressor gene PTEN (phosphatase, tension homolog deleted on chromosome ten) dephosphorylates PIP3 to generate PIP2, which leads to negative feedback regulation of the PI3K signaling pathway, inhibits cell proliferation and promotes cell apoptosis. PI3K gene mutation and amplification frequently occur in cancer, and PTEN gene deletion in cancer, etc., suggest that PI3K overexpression is closely related to tumorigenesis.
TGR-1202 is a second-generation PI3Kδ inhibitor developed by TG Therapeutic. Compared with the first-generation PI3Kδ inhibitor, it can significantly reduce the toxicity of liver and gastrointestinal tract in clinical trials, and patients with large B-cell lymphoma are also exposed to TGR. -1202 There is a partial response. Patent WO2014006572 discloses the structure of TGR-1202. ACP-196 is a second-generation BTK inhibitor that has been approved for marketing by the FDA. It has been reported in the literature (PLoS ONE 12(2):e0171221.). The combination of a PI3Kδ inhibitor and a BTK inhibitor can inhibit BCR signaling in two ways. Access, thereby playing a synergistic effect.Example 1: Preparation of the compound of formula (I)

Step 1: Synthesis of compound BB-1-3
To a solution of BB-1-1 (23g, 205.17mmol, 1eq) in polyphosphoric acid (23g, 17.84mmol) was added BB-1-2 (43.90g, 266.71mmol, 1.3eq). The reaction solution was stirred at 125°C for 5 hours under the protection of nitrogen. After the completion of the reaction, water (300 mL) was added to the reaction solution to quench the reaction, and a solid precipitated out, which was directly filtered to obtain a filter cake. The filter cake was washed with water (100 mL) once, and then purified by column chromatography (PE:EA=1:1) to obtain the target compound BB-1-3. ¹ H NMR (400MHz, CDCl ₃ ) δ 8.94 (br s, 1H), 7.68 (br d, J=5.3 Hz, 2H), 6.65 (s, 1H), 4.51 (s, 2H).
Step 2: Synthesis of compound BB-1-4
To a solution of BB-1-3 (21.02g, 98.87mmol, 1eq) in glacial acetic acid (210mL) was added NBS (19.36g, 108.75mmol, 1.1eq). The reaction solution was stirred at 25°C for 1 hour under the protection of nitrogen. After the completion of the reaction, water (200 mL) was added to the reaction solution to quench the reaction, and a solid was formed, which was filtered to obtain a filter cake. After washing three times with water (30mL*3), the filter cake was dissolved in dichloromethane (100mL), dried over anhydrous sodium sulfate, concentrated, and then beaten with methyl tert-butyl ether (50mL) once. The filter cake was collected by filtration to obtain the target Compound BB-1-4 batch one. The aqueous phase was extracted with dichloromethane (100mL*3) and combined with the mother liquor obtained by washing with methyl tert-butyl ether, and then subjected to column chromatography (petroleum ether: ethyl acetate = 1:1, target product Rf = 0.43) ) Purification to obtain the target compound BB-1-4 batch two. The two batches were dissolved and combined with dichloromethane and spin-dried to obtain the target compound BB-1-4. ¹ H NMR (400 MHz, CDCl ₃ ) δ 8.93 (dd, J = 1.3, 3.1 Hz, 1H), 7.80-7.69 (m, 2H), 4.74 (s, 2H).
Step 3: Synthesis of compound BB-1-5
To a solution of BB-1-4 (3g, 10.29mmol, 1eq) in N,N-dimethylformamide (30mL), potassium acetate (1.52g, 15.44mmol, 1.5eq) was added. The reaction solution was stirred at 40°C for 3.5 hours under the protection of nitrogen. After the reaction was completed, water (60 mL) was added to the reaction solution to quench the reaction, and a large amount of solid was formed, which was filtered to obtain a filter cake. The filter cake was dissolved in dichloromethane (100 mL), dried over anhydrous sodium sulfate, and concentrated to obtain the target compound BB-1-5 batch one. The aqueous phase was extracted with methyl tert-butyl ether (100 mL*3) to obtain the organic phase, dried over anhydrous sodium sulfate, and concentrated to obtain BB-1-5 batch two, which was obtained by combining the two batches. Used directly in the next reaction. ¹ H NMR (400MHz, CDCl ₃ ) δ 9.07-8.88 (m, 1H), 7.71 (dd, J=1.7, 5.8 Hz, 2H), 5.31-5.26 (m, 2H), 2.22 (s, 3H).
Step 4: Synthesis of compound BB-1-6
To the dioxane (37 mL) solution of BB-1-5 (3.77 g, 11.96 mmol, 1 eq), hydrochloric acid (12M, 3.49 mL, 3.5 eq) was added. The reaction solution was stirred at 40°C for 3.5 hours under the protection of nitrogen. After the completion of the reaction, the reaction solution was concentrated, water (2 mL) was added, the pH was adjusted to 9 with ammonia water, and the filter cake was collected by filtration. After dissolving with dichloromethane (100 mL), drying with anhydrous sodium sulfate, and concentration, the target compound BB-1-6 was obtained. Used directly in the next reaction. ¹ H NMR(400MHz,DMSO-d ₆ )δ8.95(dd,J=2.9,4.6Hz,1H), 8.15(dd,J=2.6,7.0,9.6Hz,1H), 7.86(dd,J=5.3 , 9.6 Hz, 1H), 5.35 (t, J = 5.9 Hz, 1H), 4.58 (d, J = 6.1 Hz, 2H).
Step 5: Synthesis of compound BB-1-7
To BB-1-6 (2.6g, 9.52mmol, 1eq) and 3-fluorophenylboronic acid (2.66g, 19.04mmol, 2eq) in acetonitrile/water (12.5mL, volume ratio: 3/1), add carbonic acid Sodium ( 5.05g, 47.61mmol, 5eq ) and Pd(PPh ₃ ) ₄ (550.15mg, 476.09μmol, 0.05eq). The reaction solution was stirred at 85°C for 4 hours under the protection of nitrogen. After the reaction was completed, dichloromethane (50 mL) was added to the reaction solution, and then water (5 mL) was slowly added to quench the reaction, and then extracted with dichloromethane (50 mL*3). The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (petroleum ether: ethyl acetate=0:1) to obtain BB-1-7. ¹ H NMR(400MHz,DMSO-d ₆ )δ8.94(dd,J=3.1,4.8Hz,1H), 8.12(ddd,J=2.6,7.1,10.0Hz,1H), 7.84(dd,J=5.3 , 10.1 Hz, 1H), 7.62 (s, 1H), 7.27-7.15 (m, 3H), 5.25 (t, J = 5.9 Hz, 1H), 4.28 (d, J = 5.7 Hz, 2H).
Step 6: Synthesis of compound BB-1-8
In a three-neck flask, at -78°C, to a solution of oxalyl chloride (1.85g, 14.57mmol, 1.28mL, 3eq) in dichloromethane (20mL) was added DMSO (2.28g, 29.14mmol, 2.28mL, 6eq). The reaction solution was stirred at -78°C for 1 hour under the protection of nitrogen. A solution of BB-1-7 (1.4g, 4.86mmol, 1eq) in dichloromethane (20mL) was added, and the mixture was stirred at -78°C for 1 hour. Triethylamine (4.91g, 48.57mmol, 6.76mL, 10eq) was added, and after stirring at -78°C for 1 hour, the reaction solution was raised to 25°C and stirred for 1 hour. After the reaction was completed, dichloromethane (20 mL) was added to the reaction solution at 0° C., water (10 mL) was added slowly to quench the reaction, and it was extracted with dichloromethane (30 mL*3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to obtain the target compound BB-1-8. LCMS, m/z=287.0 [M+1].
Step 7: Synthesis of compound BB-1
In a three-neck flask, at 0°C, to a tetrahydrofuran (50 mL) solution of BB-1-8 (1.9 g, 6.64 mmol, 1 eq) was added methyl magnesium bromide (3M, 5.53 mL, 2.5 eq). The reaction solution was stirred at 25°C for 5 hours under the protection of nitrogen. After the reaction was completed, at 0°C, water (10 mL) was slowly added to the reaction solution to quench the reaction, and then extracted with dichloromethane (10 mL*3). The organic phases were combined, dried with anhydrous sodium sulfate, concentrated, and subjected to preparative high performance liquid chromatography (column: Phenomenex Luna C18 200*40mm*10μm; mobile phase: [water (0.1%TFA)-acetonitrile]; B%: 15%- 35%, 10min) Purification (B is acetonitrile) to obtain BB-1.
Step 8: Synthesis of compound WX001-2
To the N,N-dimethylformamide (20mL) solution of WX001-1 (2g, 10.47mmol, 1eq) and potassium carbonate (4.34g, 31.41mmol, 3eq), add 2-iodopropane (3.56g, 20.94 mmol, 2.09mL, 2eq). The reaction solution was stirred at 90°C for 12 hours under the protection of nitrogen. After the reaction was completed, water (20 mL) was added to the reaction solution to quench the reaction, and then extracted with methyl tert-butyl ether (10 mL*3). The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (petroleum ether: ethyl acetate = 5:1) to obtain the target compound WX001-2. ¹ H NMR(400MHz, CDCl ₃ )δ7.23(dd,J=2.4,10.6Hz,1H), 7.16(td,J=1.9,8.8Hz,1H), 6.85(t,J=8.7Hz,1H) , 4.49 (spt, J = 6.1 Hz, 1H), 1.35 (d, J = 6.1 Hz, 6H).
Step 9: Synthesis of compound WX001-3
To the dioxane (20mL) solution of WX001-2 (2g, 8.58mmol, 1eq), add double pinacol borate (2.40g, 9.44mmol, 1.1eq), potassium acetate (1.68g, 17.16 mmol, 2eq) and Pd(dppf)Cl ₂ (627.87mg, 858.09μmol, 0.1eq). The reaction solution was stirred at 90°C for 3 hours under the protection of nitrogen. After the reaction was completed, water (20 mL) was added to the reaction solution to quench the reaction, and then extracted with dichloromethane (30 mL*3). The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and the crude product was purified by column chromatography (petroleum ether: ethyl acetate = 5:1) to obtain the target compound WX001-3. ¹ H NMR (400MHz, CDCl ₃ ) δ7.54-7.44 (m, 2H), 6.95 (t, J = 8.1 Hz, 1H), 4.60 (spt, J = 6.1 Hz, 1H), 1.37 (s, 3H) ,1.36(s,3H),1.33(s,12H).
Step 10: Synthesis of compound WX001-5
To WX001-3 (2.65g, 9.46mmol, 1eq) and WX001-4 (2.47g, 9.46mmol, 1eq) N,N-dimethylformamide/ethanol/water (265mL, volume ratio: 2/1/ 1) In the solution, add Pd(PPh ₃ ) ₄ (546.55 mg, 472.97 μmol, 0.05 eq) and sodium carbonate (3.01 g, 28.38 mmol, 3 eq). The reaction solution was stirred at 80°C for 12 hours under the protection of nitrogen. After the completion of the reaction, the reaction solution was filtered while hot (80℃) to obtain the mother liquor. After the mother liquor was spin-dried, dichloromethane (30mL) and water (30mL) were added. A large amount of insoluble matter was formed. After filtration, it was subjected to preparative high performance liquid chromatography. Purified to obtain the target compound WX001-5. LCMS, m/z=288.1 [M+1].
Step 11: Synthesis of compound WX001-6
To BB-1 (230mg, 760.90μmol, 1eq), WX001-5 (218.60mg, 760.90μmol, 1eq) and PPh ₃ (299.36mg, 1.14mmol, 1.5eq) in tetrahydrofuran (50mL) solution at 25℃ , Add diisopropyl azodicarboxylate (230.79 mg, 1.14 mmol, 221.91 μL, 1.5 eq). The reaction solution was stirred at 45°C for 5 hours under the protection of nitrogen. After the completion of the reaction, the reaction solution was directly concentrated, and purified by a thin-layer chromatography plate (dichloromethane:methanol=15:1) to obtain an isomer mixture WX001-6.
Step 12: Synthesis of the compound of formula (I)
WX001-6 was purified by supercritical fluid chromatography (column: DAICEL CHIRALPAK AD-H (250mm*30mm, 5μm); mobile phase: [0.1% ammonia in ethanol]; B%: 22%-22%, 8min) (B It is 0.1% ammonia in ethanol) to obtain the compound of formula (I) (retention time is 2.29 min), and the structure of the compound of formula (I) is confirmed by a single crystal to be correct. ¹ H NMR (400MHz, CD ₃ OD) δ 8.95 (br s, 1H), 8.03 (s, 1H), 8.00-7.92 (m, 1H), 7.91-7.82 (m, 1H), 7.42-7.29 (m ,2H),7.27-7.09(m,1H),7.22(br t,J=8.6Hz,1H),6.96-6.71(m,2H),6.20(q,J=6.6Hz,1H),4.68(td , J=6.1, 11.9 Hz, 1H), 1.91 (d, J=7.0 Hz, 3H), 1.36 (d, J=5.7 Hz, 6H); LCMS, m/z=572.2 [M+1].

////////////TGR 1202

Inclisiran

January 21, 2021 11:53 am / Leave a comment

Inclisiran

CAS 1639324-58-5

ALN-60212
ALN-PCSsc

US FDA APPROVED

12/22/2021

To treat heterozygous familial hypercholesterolemia or clinical atherosclerotic cardiovascular disease as an add-on therapy, Leqvio

Inclisiran was first developed by Alnylam Pharmaceuticals, Inc. (Cambridge, Massachusetts, US). Development has now been assumed by The Medicines Company (Parsippany, New Jersey, US). One phase I and two phase II trials have been completed. Topline results of two phase III trials were also recently presented while other phase III trials are still ongoing as part of the ORION clinical development program. …..https://www.ncbi.nlm.nih.gov/books/NBK555477/

Inclisiran is a long-acting, synthetic small interfering RNA (siRNA) directed against proprotein convertase subtilisin-kexin type 9 (PCSK9), which is a serine protease that regulates plasma low-density lipoprotein cholesterol (LDL-C) levels. By binding to PCSK9 messenger RNA, inclisiran prevents protein translation of PCSK9, leading to decreased concentrations of PCSK9 and plasma concentrations of LDL cholesterol.^1,2 Lowering circulating plasma LDL-C levels offers an additional benefit of reducing the risk of cardiovascular disease (CVD) and improving cardiovascular outcomes, as hypercholesterolemia is a major known risk factor for CVD.^1,2

On December 11, 2020, the European Commission (EC) granted authorization for marketing inclisiran as the first and only approved siRNA for the treatment of adults with primary hypercholesterolemia (heterozygous familial and non-familial) or mixed dyslipidemia, alone or in combination with other lipid-lowering therapies. It is marketed under the trade name Leqvio ⁸ and is also currently under review by the FDA.

Inclisiran, sold under the brand name Leqvio, is a medication for the treatment of people with atherosclerotic cardiovascular disease (ASCVD), ASCVD risk equivalents and heterozygous familial hypercholesterolemia (HeFH). It is a small interfering RNA that inhibits translation of the protein PCSK9.^[2]^[3]^[4] It is being developed by The Medicines Company which licensed the rights to inclisiran from Alnylam Pharmaceuticals.^[5]

On 15 October 2020, the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA) adopted a positive opinion, recommending the granting of a marketing authorization for the medicinal product Leqvio, intended for the treatment for primary hypercholesterolaemia or mixed dyslipidaemia.^[6] Inclisiran was approved for use in the European Union in December 2020.^[1]

History

In 2019 The Medicines Company announced positive results from pivotal phase III study (all primary and secondary endpoints were met with efficacy consistent with Phase I and II studies). The company anticipates regulatory submissions in the U.S. in the fourth quarter of 2019, and in Europe in the first quarter of 2020.^[7] The Medicines Company is being acquired by Novartis.^[8]

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References

^ Jump up to:^a ^b “Leqvio EPAR”. European Medicines Agency. 13 October 2020. Retrieved 6 January 2021.
^ Fitzgerald K, White S, Borodovsky A, Bettencourt BR, Strahs A, Clausen V, et al. (January 2017). “A Highly Durable RNAi Therapeutic Inhibitor of PCSK9”. The New England Journal of Medicine. 376 (1): 41–51. doi:10.1056/NEJMoa1609243. PMC 5778873. PMID 27959715.
^ Spreitzer H (11 September 2017). “Neue Wirkstoffe: Inclisiran”. Österreichische Apotheker-Zeitung (in German) (19/2017).
^ “Proposed INN: List 114” (PDF). WHO Drug Information. WHO. 29 (4): 531f. 2015.
^ Taylor NP (26 August 2019). “Medicines Company’s PCSK9 drug hits phase 3 efficacy goals”. FierceBiotech.
^ “Leqvio: Pending EC decision”. European Medicines Agency (EMA). 16 October 2020. Retrieved 16 October 2020. Text was copied from this source which is © European Medicines Agency. Reproduction is authorized provided the source is acknowledged.
^ “The Medicines Company Announces Positive Topline Results from First Pivotal Phase 3 Trial of Inclisiran”. The Medicines Company. Retrieved 29 August 2019.
^ “Novartis acquires medicines company”. Novartis. Retrieved 15 January 2020.

External links

“Inclisiran”. Drug Information Portal. U.S. National Library of Medicine.
Clinical trial number NCT03399370 for “Inclisiran for Participants With Atherosclerotic Cardiovascular Disease and Elevated Low-density Lipoprotein Cholesterol (ORION-10)” at ClinicalTrials.gov
Clinical trial number NCT03400800 for “Inclisiran for Subjects With ACSVD or ACSVD-Risk Equivalents and Elevated Low-density Lipoprotein Cholesterol (ORION-11)” at ClinicalTrials.gov

Clinical data
Trade names	Leqvio
Other names	ALN-PCSsc, ALN-60212
Routes of administration	Subcutaneous injection
ATC code	C10AX16 (WHO)
Legal status
Legal status	EU: Rx-only ^[1]
Identifiers
CAS Number	1639324-58-5
DrugBank	DB14901
UNII	UOW2C71PG5
KEGG	D11931
Chemical and physical data
Formula	C₅₂₀H₆₇₉F₂₁N₁₇₅O₃₀₉P₄₃S₆
Molar mass	16248.27 g·mol⁻¹

General References

Kosmas CE, Munoz Estrella A, Sourlas A, Silverio D, Hilario E, Montan PD, Guzman E: Inclisiran: A New Promising Agent in the Management of Hypercholesterolemia. Diseases. 2018 Jul 13;6(3). pii: diseases6030063. doi: 10.3390/diseases6030063. [PubMed:30011788]
German CA, Shapiro MD: Small Interfering RNA Therapeutic Inclisiran: A New Approach to Targeting PCSK9. BioDrugs. 2020 Feb;34(1):1-9. doi: 10.1007/s40259-019-00399-6. [PubMed:31782112]
Doggrell SA: Inclisiran, the billion-dollar drug, to lower LDL cholesterol – is it worth it? Expert Opin Pharmacother. 2020 Nov;21(16):1971-1974. doi: 10.1080/14656566.2020.1799978. Epub 2020 Aug 4. [PubMed:32749892]
Goldstein JL, Brown MS: Regulation of low-density lipoprotein receptors: implications for pathogenesis and therapy of hypercholesterolemia and atherosclerosis. Circulation. 1987 Sep;76(3):504-7. doi: 10.1161/01.cir.76.3.504. [PubMed:3621516]
Pratt AJ, MacRae IJ: The RNA-induced silencing complex: a versatile gene-silencing machine. J Biol Chem. 2009 Jul 3;284(27):17897-901. doi: 10.1074/jbc.R900012200. Epub 2009 Apr 1. [PubMed:19342379]
Leiter LA, Teoh H, Kallend D, Wright RS, Landmesser U, Wijngaard PLJ, Kastelein JJP, Ray KK: Inclisiran Lowers LDL-C and PCSK9 Irrespective of Diabetes Status: The ORION-1 Randomized Clinical Trial. Diabetes Care. 2019 Jan;42(1):173-176. doi: 10.2337/dc18-1491. Epub 2018 Nov 28. [PubMed:30487231]
Cupido AJ, Kastelein JJP: Inclisiran for the treatment of hypercholesterolaemia: implications and unanswered questions from the ORION trials. Cardiovasc Res. 2020 Sep 1;116(11):e136-e139. doi: 10.1093/cvr/cvaa212. [PubMed:32766688]
Novartis: Novartis receives EU approval for Leqvio (inclisiran), a first-in-class siRNA to lower cholesterol with two doses a year [Link]
Summary of Product Characteristics: Leqvio (inclisiran), solution for subcutaneous injection [Link]

Summary

Atherosclerotic cardiovascular disease (ASCVD) remains one of the leading causes of death in Canada. Cholesterol, specifically low-density lipoprotein cholesterol (LDL-C), is a major risk factor for cardiovascular disease (CVD) and is thereby targeted to reduce the likelihood of a cardiovascular event, such as a myocardial infarction (MI) and stroke.
Inclisiran, first developed by Alnylam Pharmaceuticals, Inc. (Cambridge, Massachusetts, US) then by The Medicines Company (Parsippany, New Jersey, US), is a small interfering ribonucleic acid (siRNA) molecule being investigated for the treatment of hypercholesterolemia.
ORION-1 was a phase II, double-blind, placebo-controlled, multi-centre, randomized controlled trial of 501 patients. Patients were included in the trial if they had a history of ASCVD or were at high risk of ASCVD. The treatment arms were administered 200 mg, 300 mg, or 500 mg of inclisiran on day 1, or 100 mg, 200 mg, or 300 mg of inclisiran on days 1 and 90. The comparator was either placebo on day 1 or placebo on days 1 and 90. The primary end point was percentage change in LDL-C at day 180 from baseline.
The ORION-1 study demonstrated that inclisiran, administered at various doses and intervals, compared with placebo, resulted in a statistically significant reduction in LDL-C levels (P < 0.001 for all comparisons versus placebo). The greatest reduction in LDL-C levels was obtained with the 300 mg dose of inclisiran given at days 1 and 90 with a 52.6% (95% confidence interval [CI]: −57.1 to −48.1) reduction at day 180 compared with baseline, and a mean absolute reduction in LDL-C levels of 1.66 (standard deviation 0.54) mmol/L. Results from the ORION-1 trial provided the necessary data to make a decision regarding the dosing regimen to be used in subsequent phase III trials, in particular the ORION-11 phase III trial.
The ORION-11 study was a phase III international, multi-centre, and double-blind trial which randomized 1,617 participants (87% with established ASCVD) to inclisiran 300 mg (n = 810) or placebo (n = 807). An initial inclisiran dose of 300 mg given subcutaneously was administered at day 1, day 90, and then every six months for two doses, that is at days 270 and 450. The mean baseline LDL-C level was 2.8 mmol/L (inclisiran) and 2.7 mmol/L (placebo); 96% of participants were on high-dose statin therapy. There was a 50% time-averaged reduction in LDL-C levels from day 90 to day 540 (P < 0.00001). Pre-specified exploratory cardiovascular composite end point (cardiac death, cardiac arrest, MI, or stroke) occurred in 7.8% of inclisiran treated patients versus 10.3% of patients on placebo; this lower rate was mainly driven by a reduction in MI and stroke. With respect to adverse effects, 4.69% of patients on inclisiran reported an injection site reaction, compared with 0.5% of patients on placebo. All reactions were transient. There was no evidence of liver, kidney, muscle, or platelet toxicity.
Inclisiran may be an option in the future as a cholesterol-lowering medication, where it would likely be used in patients who are unable to achieve their LDL-C targets despite maximally tolerated statin therapy or who are intolerant to statin therapy. However, results from the inclisiran cardiovascular outcome trial (ORION-4), are needed to confirm its efficacy in reducing CVD and its long-term safety.
Inclisiran is not yet approved by any regulatory authority, but its ORION clinical development program identifies the year 2021 as the goal to reach worldwide markets.

///////////Inclisiran, LEQVIO, ALN 60212, ALN PCSsc , NOVARTIS, Leqvio, APPROVALS 2021, FDA 2021

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Setmelanotide

December 14, 2020 3:34 am / 1 Comment on Setmelanotide

ChemSpider 2D Image | Setmelanotide | C49H68N18O9S2

Setmelanotide

Ac-Arg-Cys(1)-D-Ala-His-D-Phe-Arg-Trp-Cys(1)-NH2

Molecular FormulaC₄₉H₆₈N₁₈O₉S₂
Average mass1117.309 Da
N-acetyl-L-arginyl-L-cysteinyl-D-alanyl-L-histidyl-D-phenylalanyl-L-arginyl-L-tryptophyl-L-cysteinamide (2->8)-disulfide

1,2-Dithia-5,8,11,14,17,20-hexaazacyclotricosane-4-carboxamide, 22-[[(2S)-2-(acetylamino)-5-[(diaminomethylene)amino]-1-oxopentyl]amino]-10-[3-[(diaminomethylene)amino]propyl]-16-(1H-imidazol-5-ylmeth yl)-7-(1H-indol-3-ylmethyl)-19-methyl-6,9,12,15,18,21-hexaoxo-13-(phenylmethyl)-, (4R,7S,10S,13R,16S,19R,22R)- [ACD/Index Name]10011920014-72-8[RN]Imcivree [Trade name]N2-acetyl-L-arginyl-L-cysteinyl-D-alanyl-L-histidyl-D-phenylalanyl-L-arginyl-Ltryptophyl- L-cysteinamide, cyclic (2-8)-disulfideN7T15V1FUYRM-493, BIM-22493UNII-N7T15V1FUYсетмеланотид [Russian] [INN]سيتميلانوتيد [Arabic] [INN]司美诺肽 [Chinese] [INN](4R,7S,10S,13R,16S,19R,22R)-22-[[(2S)-2-acetamido-5-(diaminomethylideneamino)pentanoyl]amino]-13-benzyl-10-[3-(diaminomethylideneamino)propyl]-16-(1H-imidazol-5-ylmethyl)-7-(1H-indol-3-ylmethyl)-19-methyl-6,9,12,15,18,21-hexaoxo-1,2-dithia-5,8,11,14,17,20-hexazacyclotricosane-4-carboxamide

FDA 11/25/2020, Imcivree, To treat obesity and the control of hunger associated with pro-opiomelanocortin deficiency, a rare disorder that causes severe obesity that begins at an early age
Drug Trials Snapshot, 10MG/ML, SOLUTION;SUBCUTANEOUS, Orphan

Rhythm Pharmaceuticals Announces FDA Approval of IMCIVREE™ (setmelanotide) as First-ever Therapy for Chronic Weight Management in Patients with Obesity Due to POMC, PCSK1 or LEPR Deficiency Nasdaq:RYTM

update Imcivree EMA APPROVED 2021/7/16

DESCRIPTION

IMCIVREE contains setmelanotide acetate, a melanocortin 4 (MC4) receptor agonist. Setmelanotide is an 8 amino acid cyclic peptide analog of endogenous melanocortin peptide α-MSH (alpha-melanocyte stimulating hormone).

The chemical name for setmelanotide acetate is acetyl-L-arginyl-L-cysteinyl-D-alanyl-Lhistidinyl-D-phenylalanyl-L-arginyl-L-tryptophanyl-L-cysteinamide cyclic (2→8)-disulfide acetate. Its molecular formula is C₄₉H₆₈N₁₈O₉S₂ (anhydrous, free-base), and molecular mass is 1117.3 Daltons (anhydrous, free-base).

The chemical structure of setmelanotide is:

IMCIVREE (setmelanotide) Structrual Formula Illustration

IMCIVREE injection is a sterile clear to slightly opalescent, colorless to slightly yellow solution. Each 1 mL of IMCIVREE contains 10 mg of setmelanotide provided as setmelanotide acetate, which is a salt with 2 to 4 molar equivalents of acetate, and the following inactive ingredients: 100 mg N-(carbonyl-methoxypolyethylene glycol 2000)-1,2-distearoyl-glycero-3phosphoethanolamine sodium salt, 8 mg carboxymethylcellulose sodium (average MWt 90,500), 11 mg mannitol, 5 mg phenol, 10 mg benzyl alcohol, 1 mg edetate disodium dihydrate, and Water for Injection. The pH of IMCIVREE is 5 to 6.

Setmelanotide is a peptide drug and investigational anti-obesity medication which acts as a selective agonist of the MC4 receptor. Setmelanotide binds to and activates MC4 receptors in the paraventricular nucleus (PVN) of the hypothalamus and in the lateral hypothalamic area (LHA), areas involved in the regulation of appetite, and this action is thought to underlie its appetite suppressant effects. Setmelanotide increases resting energy expenditure in both obese animals and humans. Setmelanotide has been reported to possess the following activity profile (cAMP, EC50): MC4 (0.27 nM) > MC3 (5.3 nM) ≈ MC1 (5.8 nM) > MC5 (1600 nM) ≟ MC2 (>1000 nM).

Setmelanotide, sold under the brand name Imcivree, is a medication for the treatment of obesity.^[1]

The most common side effects include injection site reactions, skin hyperpigmentation (skin patches that are darker than surrounding skin), headache and gastrointestinal side effects (such as nausea, diarrhea, and abdominal pain), among others.^[1] Spontaneous penile erections in males and adverse sexual reactions in females have occurred with treatment.^[1] Depression and suicidal ideation have also occurred with setmelanotide.^[1]

SYN

WO 2011060355

Medical uses

Setmelanotide is indicated for chronic weight management (weight loss and weight maintenance for at least one year) in people six years and older with obesity due to three rare genetic conditions: pro-opiomelanocortin (POMC) deficiency, proprotein subtilisin/kexin type 1 (PCSK1) deficiency, and leptin receptor (LEPR) deficiency confirmed by genetic testing demonstrating variants in POMC, PCSK1, or LEPR genes considered pathogenic (causing disease), likely pathogenic, or of uncertain significance.^[1] Setmelanotide is the first FDA-approved treatment for these genetic conditions.^[1]

Setmelanotide is not approved for obesity due to suspected POMC, PCSK1, or LEPR deficiency with variants classified as benign (not causing disease) or likely benign or other types of obesity, including obesity associated with other genetic syndromes and general (polygenic) obesity.^[1]

Setmelanotide binds to and activates MC₄ receptors in the paraventricular nucleus (PVN) of the hypothalamus and in the lateral hypothalamic area (LHA), areas involved in the regulation of appetite, and this action is thought to underlie its appetite suppressant effects.^[2] In addition to reducing appetite, setmelanotide increases resting energy expenditure in both obese animals and humans.^[3] Importantly, unlike certain other MC₄ receptor agonists, such as LY-2112688, setmelanotide has not been found to produce increases in heart rate or blood pressure.^[4]

Setmelanotide has been reported to possess the following activity profile (cAMP, EC₅₀): MC₄ (0.27 nM) > MC₃ (5.3 nM) ≈ MC₁ (5.8 nM) > MC₅ (1600 nM) ≟ MC₂ (>1000 nM).^[5] (19.6-fold selectivity for MC₄ over MC₃, the second target of highest activity.)

History

Setmelanotide was evaluated in two one-year studies.^[1] The first study enrolled participants with obesity and confirmed or suspected POMC or PCSK1 deficiency while the second study enrolled participants with obesity and confirmed or suspected LEPR deficiency; all participants were six years or older.^[1] The effectiveness of setmelanotide was determined by the number of participants who lost more than ten percent of their body weight after a year of treatment.^[1]

The effectiveness of setmelanotide was assessed in 21 participants, ten in the first study and eleven in the second.^[1] In the first study, 80 percent of participants with POMC or PCSK1 deficiency lost ten percent or more of their body weight.^[1] In the second study, 46 percent of participants with LEPR deficiency lost ten percent or more of their body weight.^[1]

The study also assessed the maximal (greatest) hunger in sixteen participants over the previous 24 hours using an eleven-point scale in participants twelve years and older.^[1] In both studies, some, but not all, of participants’ weekly average maximal hunger scores decreased substantially from their scores at the beginning of the study.^[1] The degree of change was highly variable among participants.^[1]

The U.S. Food and Drug Administration (FDA) granted the application for setmelanotide orphan disease designation, breakthrough therapy designation, and priority review.^[1] The FDA granted the approval of Imcivree to Rhythm Pharmaceutical, Inc.^[1]

Research

Setmelanotide is a peptide drug and investigational anti-obesity medication which acts as a selective agonist of the MC₄ receptor.^[6]^[4] Its peptide sequence is Ac-Arg-Cys(1)-D-Ala-His-D-Phe-Arg-Trp-Cys(1)-NH2. It was first discovered at Ipsen and is being developed by Rhythm Pharmaceuticals for the treatment of obesity and diabetes.^[6] In addition, Rhythm Pharmaceuticals is conducting trials of setmelanotide for the treatment of Prader–Willi syndrome (PWS), a genetic disorder which includes MC₄ receptor deficiency and associated symptoms such as excessive appetite and obesity.^[7] As of December 2014, the drug is in phase II clinical trials for obesity and PWS.^[6]^[8]^[9]^{[needs update]} So far, preliminary data has shown no benefit of Setmelanotide in Prader-Willi syndrome.^[10]

PATENT

WO 2007008704

WO 2011060355

WO 2011060352

US 20120225816

PAPER

Journal of Medicinal Chemistry, 61(8), 3674-3684; 2018

PATENT

https://patents.google.com/patent/US9314509

Synthesis of Example 1i.e., Ac-Arg-cyclo(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-NH₂

The title peptide having the above structure was assembled using Fmoc chemistry on an Apex peptide synthesizer (Aapptec; Louisville, Ky., USA). 220 mg of 0.91 mmol/g (0.20 mmoles) Rink Amide MBHA resin (Polymer Laboratories; Amherst, Mass., USA) was placed in a reaction well and pre-swollen in 3.0 mL of DMF prior to synthesis. For cycle 1, the resin was treated with two 3-mL portions of 25% piperidine in DMF for 5 and 10 minutes respectively, followed by 4 washes of 3-mL DMF—each wash consisting of adding 3 mL of solvent, mixing for 1 minute, and emptying for 1 minute. Amino acids stocks were prepared in NMP as 0.45M solutions containing 0.45M HOBT. HBTU was prepared as a 0.45M solution in NMP and DIPEA was prepared as a 2.73M solution in NMP. To the resin, 2 mL of the first amino acid (0 9 mmoles, Fmoc-Cys(Trt)-OH) (Novabiochem; San Diego, Calif., USA) was added along with 2 mL (0.9 mmoles) of HBTU and 1.5 mL (4.1 mmoles) of DIPEA. After one hour of constant mixing, the coupling reagents were drained from the resin and the coupling step was repeated. Following amino acid acylation, the resin was washed with two 3-mL aliquots of DMF for 1 minute. The process of assembling the peptide (deblock/wash/acylate/wash) was repeated for cycles 2-9 identical to that as described for cycle 1. The following amino acids were used: cycle 2) Fmoc-Trp(Boc)-OH (Genzyme; Cambridge, Mass., USA); cycle 3) Fmoc-Arg(Pbf)-OH (Novabiochem); cycle 4) Fmoc-DPhe-OH (Genzyme); cycle 5) Fmoc-His(Trt)-OH (Novabiochem); cycle 6) Fmoc-D-Ala-OH (Genzyme); cycle 7) Fmoc-Cys(Trt)-OH, (Novabiochem); and cycle 8) Fmoc-Arg(Pbf)-OH (Genzyme). The N-terminal Fmoc was removed with 25% piperidine in DMF as described above, followed by four 3-mL DMF washes for 1 minute. Acetylation of the N-terminus was performed by adding 0.5 mL of 3M DIPEA in NMP to the resin along with 1.45 mL of 0.45M acetic anhydride in NMP. The resin was mixed for 30 minutes and acetylation was repeated. The resin was washed with 3 mL of DMF for a total of 5 times followed with 5 washes with 5 mL of DCM each.

To cleave and deprotect the peptide, 5mL of the following reagent was added to the resin: 2% TIS/5% water/5% (w/v) DTT/88% TFA. The solution was allowed to mix for 3.5 hours. The filtrate was collected into 40 mL of cold anhydrous ethyl ether. The precipitate was pelleted for 10 minutes at 3500 rpm in a refrigerated centrifuge. The ether was decanted and the peptide was re-suspended in fresh ether. The ether workup was performed three times. Following the last ether wash, the peptide was allowed to air dry to remove residual ether.

The peptide was dissolved in 10% acetonitrile and analyzed by mass spectrometry and reverse-phase HPLC employing a 30×4.6 cm C18 column (Vydac; Hesperia, Calif., USA) with a gradient of 2-60% acetonitrile (0.1% TFA) over 30 minutes. This analysis identified a product with ˜53% purity. Mass analysis employing electrospray ionization identified a main product containing a mass of 1118.4 corresponding to the desired linear product. The crude product (˜100 mg) was diluted to a concentration of 2 mg/mL in 5% acetic acid. To this solution, 0.5M iodine/methanol was added dropwise with vigorous stirring until a pale yellow color was achieved. The solution was vigorously stirred for another 10 minutes. Excess iodine was then quenched by adding 1.0M sodium thiosulfate under continuous mixing until the mixture was rendered colorless. The peptide was re-examined by mass spectrometry analysis and HPLC. Mass spectrometry analysis identified a main species with a mass of 1116.4 which indicated successful oxidation to form the cyclic peptide. The peptide solution was purified on a preparative HPLC equipped with a C18 column using a similar elution gradient. The purified product was re-analyzed by HPLC for purity (>95%) and mass spectrometry (1116.9 which is in agreement with the expected mass of 1117.3) and subsequently lyophilized. Following lyophilization, 28 mg of purified product was obtained representing a 24% yield.

The other exemplified peptides were synthesized substantially according to the procedure described for the above-described synthetic process. Physical data for select exemplified peptides are given in Table 1.

TABLE 1 Example Mol. Wt. Mol. Wt. Purity Number (calculated) (ES-MS) (HPLC) 1 1117.3 1116.9 95.1% 2 1117.3 1116.8 99.2% 3 1280.5 1280.6 98.0% 5 1216.37 1216.20 99.9%

Preparation of Pamoate Salt of Example 1

The acetate salt of Example 1 (200 mg, 0.18 mmole) was dissolved in 10 mL of water. Sodium pamoate (155 mg, 0.36 mmole) was dissolved in 10 mL of water. The two solutions were combined and mixed well. The precipitates were collected by centrifugation at 3000 rpm for 20 minutes, washed for three times with water, and dried by lyophilization.

References

^ Jump up to:^a ^b ^c ^d ^e ^f ^g ^h ⁱ ^j ^k ^l ^m ⁿ ^o ^p ^q ^r “FDA approves first treatment for weight management for people with certain rare genetic conditions”. U.S. Food and Drug Administration (FDA) (Press release). 27 November 2020. Retrieved 27 November 2020. This article incorporates text from this source, which is in the public domain.
^ Kim GW, Lin JE, Blomain ES, Waldman SA (January 2014). “Antiobesity pharmacotherapy: new drugs and emerging targets”. Clinical Pharmacology and Therapeutics. 95 (1): 53–66. doi:10.1038/clpt.2013.204. PMC 4054704. PMID 24105257.
^ Chen KY, Muniyappa R, Abel BS, Mullins KP, Staker P, Brychta RJ, et al. (April 2015). “RM-493, a melanocortin-4 receptor (MC4R) agonist, increases resting energy expenditure in obese individuals”. The Journal of Clinical Endocrinology and Metabolism. 100 (4): 1639–45. doi:10.1210/jc.2014-4024. PMC 4399297. PMID 25675384.
^ Jump up to:^a ^b Kievit P, Halem H, Marks DL, Dong JZ, Glavas MM, Sinnayah P, et al. (February 2013). “Chronic treatment with a melanocortin-4 receptor agonist causes weight loss, reduces insulin resistance, and improves cardiovascular function in diet-induced obese rhesus macaques”. Diabetes. 62 (2): 490–7. doi:10.2337/db12-0598. PMC 3554387. PMID 23048186.
^ Muniyappa R, Chen K, Brychta R, Abel B, Mullins K, Staker P, et al. (June 2014). “A Randomized, Double-Blind, Placebo-Controlled, Crossover Study to Evaluate the Effect of a Melanocortin Receptor 4 (MC4R) Agonist, RM-493, on Resting Energy Expenditure (REE) in Obese Subjects” (PDF). Endocrine Reviews. Rhythm Pharmaceuticals. 35 (3). Retrieved 2015-05-21.
^ Jump up to:^a ^b ^c Lee EC, Carpino PA (2015). “Melanocortin-4 receptor modulators for the treatment of obesity: a patent analysis (2008-2014)”. Pharmaceutical Patent Analyst. 4 (2): 95–107. doi:10.4155/ppa.15.1. PMID 25853469.
^ “Obesity and Diabetes Caused by Genetic Deficiencies in the MC4 Pathway”. Rhythm Pharmaceuticals. Retrieved 2015-05-21.
^ Jackson VM, Price DA, Carpino PA (August 2014). “Investigational drugs in Phase II clinical trials for the treatment of obesity: implications for future development of novel therapies”. Expert Opinion on Investigational Drugs. 23 (8): 1055–66. doi:10.1517/13543784.2014.918952. PMID 25000213. S2CID 23198484.
^ “RM-493: A First-in-Class, Phase 2-Ready MC4 Agonist: A New Drug Class for the Treatment of Obesity and Diabetes”. Rhythm Pharmaceuticals. Archived from the original on 2015-06-14. Retrieved 2015-05-21.
^ Duis J, van Wattum PJ, Scheimann A, Salehi P, Brokamp E, Fairbrother L, et al. (March 2019). “A multidisciplinary approach to the clinical management of Prader-Willi syndrome”. Molecular Genetics & Genomic Medicine. 7 (3): e514. doi:10.1002/mgg3.514. PMC 6418440. PMID 30697974.

ADDITIONAL INFORMATION

The peptide sequence is Ac-Arg-Cys(1)-D-Ala-His-D-Phe-Arg-Trp-Cys(1)-NH2. It is being researched by Rhythm Pharmaceuticals for the treatment of obesity and diabetes. In addition, Rhythm Pharmaceuticals is conducting trials of setmelanotide for the treatment of Prader–Willi syndrome (PWS), a genetic disorder which includes MC4 receptor deficiency and associated symptoms such as excessive appetite and obesity. As of December 2014, the drug is in phase II clinical trials for obesity and PWS.

L-Cysteinamide, N2-acetyl-L-arginyl-L-cysteinyl-D-alanyl-L-histidyl-D-phenylalanyl-L-arginyl-L-tryptophyl-, cyclic (2->8)-disulfide
Ac-Arg-Cys(1)-D-Ala-His-D-Phe-Arg-Trp-Cys(1)-NH2

REFERENCES

1: Lee EC, Carpino PA. Melanocortin-4 receptor modulators for the treatment of obesity: a patent analysis (2008-2014). Pharm Pat Anal. 2015;4(2):95-107. doi: 10.4155/ppa.15.1. PubMed PMID: 25853469.

2: Chen KY, Muniyappa R, Abel BS, Mullins KP, Staker P, Brychta RJ, Zhao X, Ring M, Psota TL, Cone RD, Panaro BL, Gottesdiener KM, Van der Ploeg LH, Reitman ML, Skarulis MC. RM-493, a melanocortin-4 receptor (MC4R) agonist, increases resting energy expenditure in obese individuals. J Clin Endocrinol Metab. 2015 Apr;100(4):1639-45. doi: 10.1210/jc.2014-4024. Epub 2015 Feb 12. PubMed PMID: 25675384; PubMed Central PMCID: PMC4399297.

3: Clemmensen C, Finan B, Fischer K, Tom RZ, Legutko B, Sehrer L, Heine D, Grassl N, Meyer CW, Henderson B, Hofmann SM, Tschöp MH, Van der Ploeg LH, Müller TD. Dual melanocortin-4 receptor and GLP-1 receptor agonism amplifies metabolic benefits in diet-induced obese mice. EMBO Mol Med. 2015 Feb 4;7(3):288-98. doi: 10.15252/emmm.201404508. PubMed PMID: 25652173; PubMed Central PMCID: PMC4364946.

4: Jackson VM, Price DA, Carpino PA. Investigational drugs in Phase II clinical trials for the treatment of obesity: implications for future development of novel therapies. Expert Opin Investig Drugs. 2014 Aug;23(8):1055-66. doi: 10.1517/13543784.2014.918952. Epub 2014 Jul 7. Review. PubMed PMID: 25000213.

5: Kievit P, Halem H, Marks DL, Dong JZ, Glavas MM, Sinnayah P, Pranger L, Cowley MA, Grove KL, Culler MD. Chronic treatment with a melanocortin-4 receptor agonist causes weight loss, reduces insulin resistance, and improves cardiovascular function in diet-induced obese rhesus macaques. Diabetes. 2013 Feb;62(2):490-7. doi: 10.2337/db12-0598. Epub 2012 Oct 9. PubMed PMID: 23048186; PubMed Central PMCID: PMC3554387.

6: Kumar KG, Sutton GM, Dong JZ, Roubert P, Plas P, Halem HA, Culler MD, Yang H, Dixit VD, Butler AA. Analysis of the therapeutic functions of novel melanocortin receptor agonists in MC3R- and MC4R-deficient C57BL/6J mice. Peptides. 2009 Oct;30(10):1892-900. doi: 10.1016/j.peptides.2009.07.012. Epub 2009 Jul 29. PubMed PMID: 19646498; PubMed Central PMCID: PMC2755620.

External links

“Setmelanotide”. Drug Information Portal. U.S. National Library of Medicine.

Clinical data

Trade names	Imcivree
Other names	RM-493; BIM-22493; IRC-022493; N2-Acetyl-L-arginyl-L-cysteinyl-D-alanyl-L-histidyl-D-phenylalanyl-L-arginyl-L-tryptophyl-L-cysteinamide, cyclic (2-8)-disulfide
ATC code	None
Legal status
Legal status	US: ℞-only
Identifiers
IUPAC name [show]
CAS Number	920014-72-8
PubChem CID	11993702
ChemSpider	10166169
UNII	N7T15V1FUY
KEGG	D11927
Chemical and physical data
Formula	C₄₉H₆₈N₁₈O₉S₂
Molar mass	1117.32 g·mol⁻¹
3D model (JSmol)	Interactive image
SMILES [hide]C[C@@H]1C(=O)N[C@H](C(=O)N[C@@H](C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@@H](CSSC[C@@H](C(=O)N1)NC(=O)[C@H](CCCN=C(N)N)NC(=O)C)C(=O)N)Cc2c[nH]c3c2cccc3)CCCN=C(N)N)Cc4ccccc4)Cc5cnc[nH]5
InChI [hide]InChI=1S/C49H68N18O9S2/c1-26-41(70)63-37(20-30-22-55-25-59-30)46(75)64-35(18-28-10-4-3-5-11-28)44(73)62-34(15-9-17-57-49(53)54)43(72)65-36(19-29-21-58-32-13-7-6-12-31(29)32)45(74)66-38(40(50)69)23-77-78-24-39(47(76)60-26)67-42(71)33(61-27(2)68)14-8-16-56-48(51)52/h3-7,10-13,21-22,25-26,33-39,58H,8-9,14-20,23-24H2,1-2H3,(H2,50,69)(H,55,59)(H,60,76)(H,61,68)(H,62,73)(H,63,70)(H,64,75)(H,65,72)(H,66,74)(H,67,71)(H4,51,52,56)(H4,53,54,57)/t26-,33+,34+,35-,36+,37+,38+,39+/m1/s1Key:HDHDTKMUACZDAA-PHNIDTBTSA-N

///////////Setmelanotide, FDA 2020, 2020 APPROVALS, Imcivree, Orphan, PEPTIDE, ANTIOBESITY, UNII-N7T15V1FUY, сетмеланотид , سيتميلانوتيد , 司美诺肽 , BIM 22493, RM 493

CC1C(=O)NC(C(=O)NC(C(=O)NC(C(=O)NC(C(=O)NC(CSSCC(C(=O)N1)NC(=O)C(CCCN=C(N)N)NC(=O)C)C(=O)N)CC2=CNC3=CC=CC=C32)CCCN=C(N)N)CC4=CC=CC=C4)CC5=CN=CN5

Odevixibat

September 14, 2020 2:51 am / Leave a comment

Odevixibat

A-4250, AR-H 064974

CAS 501692-44-0

BUTANOIC ACID, 2-(((2R)-2-((2-((3,3-DIBUTYL-2,3,4,5-TETRAHYDRO-7-(METHYLTHIO)-1,1-DIOXIDO-5-PHENYL-1,2,5-BENZOTHIADIAZEPIN-8-YL)OXY)ACETYL)AMINO)-2-(4-HYDROXYPHENYL)ACETYL)AMINO)-, (2S)-

(2S)-2-[[(2R)-2-[[2-[(3,3-dibutyl-7-methylsulfanyl-1,1-dioxo-5-phenyl-2,4-dihydro-1λ⁶,2,5-benzothiadiazepin-8-yl)oxy]acetyl]amino]-2-(4-hydroxyphenyl)acetyl]amino]butanoic acid

Molecular Formula	C37H48N4O8S2
Molecular Weight	740.929

- Bylvay
- New Drug Application (NDA): 215498
  Company: ALBIREO PHARMA INC

AZD8294 WHO 10706 AR-H064974 HY-109120 CS-0078340 D11716 US9694018, 5Originator Albireo AB
Developer Albireo AB; Albireo Pharma
ClassAcetamides; Butyric acids; Hepatoprotectants; Small molecules; Sulfones; Thiazepines
Mechanism of Action Sodium-bile acid cotransporter inhibitors

Orphan Drug Status Yes – Primary biliary cirrhosis; Biliary atresia; Intrahepatic cholestasis; Alagille syndrome
New Molecular Entity Yes

Phase III Biliary atresia; Intrahepatic cholestasis
Phase II Alagille syndrome; Cholestasis; Primary biliary cirrhosis
No development reported Non-alcoholic steatohepatitis

22 Jul 2020 Albireo initiates an expanded-access programme for Intrahepatic cholestasis in USA, Canada, Australia and Europe
14 Jul 2020 Phase-III clinical trials in Biliary atresia (In infants, In neonates) in Belgium (PO) after July 2020 (EudraCT2019-003807-37)
14 Jul 2020 Phase-III clinical trials in Biliary atresia (In infants, In neonates) in Germany, France, United Kingdom, Hungary (PO) (EudraCT2019-003807-37)

UPDATE Bylvay, FDA APPROVED2021/7/20 AND EMA 2021/7/16

Odevixibat, sold under the trade name Bylvay, is a medication for the treatment of progressive familial intrahepatic cholestasis (PFIC).^[1]

The most common side effects include diarrhea, abdominal pain, hemorrhagic diarrhea, soft feces, and hepatomegaly (enlarged liver).^[1]

Odevixibat is a reversible, potent, selective inhibitor of the ileal bile acid transporter (IBAT).^[1]^[2]

In May 2021, the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA) recommended granting a marketing authorization in the European Union for odevixibat for the treatment of PFIC in people aged six months or older.^[1]^[3]

A-4250 (odevixibat) is a selective inhibitor of the ileal bile acid transporter (IBAT) that acts locally in the gut. Ileum absorbs glyco-and taurine-conjugated forms of the bile salts. IBAT is the first step in absorption at the brush-border membrane. A-4250 works by decreasing the re-absorption of bile acids from the small intestine to the liver, whichreduces the toxic levels of bile acids during the progression of the disease. It exhibits therapeutic intervention by checking the transport of bile acids. Studies show that A-4250 has the potential to decrease the damage in the liver cells and the development of fibrosis/cirrhosis of the liver known to occur in progressive familial intrahepatic cholestasis. A-4250 is a designated orphan drug in the USA for October 2012. A-4250 is a designated orphan drug in the EU for October 2016. A-4250 was awarded PRIME status for PFIC by EMA in October 2016. A-4250 is in phase II clinical trials by Albireo for the treatment of primary biliary cirrhosis (PBC) and cholestatic pruritus. In an open label Phase 2 study in children with cholestatic liver disease and pruritus, odevixibat showed reductions in serum bile acids and pruritus in most patients and exhibited a favorable overall tolerability profile.

Odevixibat is a highly potent, non-systemic ileal bile acid transport inhibitor (IBATi) that has has minimal systemic exposure and acts locally in the small intestine. Albireo is developing odevixibat to treat rare pediatric cholestatic liver diseases, including progressive familial intrahepatic cholestasis, biliary atresia and Alagille syndrome.

With normal function, approximately 95 percent of bile acids released from the liver into the bile ducts to aid in liver function are recirculated to the liver via the IBAT in a process called enterohepatic circulation. In people with cholestatic liver diseases, the bile flow is interrupted, resulting in elevated levels of toxic bile acids accumulating in the liver and serum. Accordingly, a product capable of inhibiting the IBAT could lead to a reduction in bile acids returning to the liver and may represent a promising approach for treating cholestatic liver diseases.

The randomized, double-blind, placebo-controlled, global multicenter PEDFIC 1 Phase 3 clinical trial of odevixibat in 62 patients, ages 6 months to 15.9 years, with PFIC type 1 or type 2 met its two primary endpoints demonstrating that odevixibat reduced serum bile acids (sBAs) (p=0.003) and improved pruritus (p=0.004), and was well tolerated with a low single digit diarrhea rate. These topline data substantiate the potential for odevixibat to be first drug for PFIC patients. The Company intends to complete regulatory filings in the EU and U.S. no later than early 2021, in anticipation of regulatory approval, issuance of a rare pediatric disease priority review voucher and launch in the second half of 2021.

Odevixibat is being evaluated in the ongoing PEDFIC 2 open-label trial (NCT03659916) designed to assess long-term safety and durability of response in a cohort of patients rolled over from PEDFIC 1 and a second cohort of PFIC patients who are not eligible for PEDFIC 1.

Odevixibat is also currently being evaluated in a second Phase 3 clinical trial, BOLD (NCT04336722), in patients with biliary atresia. BOLD, the largest prospective intervention trial ever conducted in biliary atresia, is a double-blind, randomized, placebo-controlled trial which will enroll approximately 200 patients at up to 75 sites globally to evaluate the efficacy and safety of odevixibat in children with biliary atresia who have undergone a Kasai procedure before age three months. The company also anticipates initiating a pivotal trial of odevixibat for Alagille syndrome by the end of 2020.

For more information about the PEDFIC 2 or BOLD studies, please visit ClinicalTrials.gov or contact medinfo@albireopharma.com.

The odevixibat PFIC program, or elements of it, have received fast track, rare pediatric disease and orphan drug designations in the United States. In addition, the FDA has granted orphan drug designation to odevixibat for the treatment of Alagille syndrome, biliary atresia and primary biliary cholangitis. The EMA has granted odevixibat orphan designation, as well as access to the PRIority MEdicines (PRIME) scheme for the treatment of PFIC. Its Paediatric Committee has agreed to Albireo’s odevixibat Pediatric Investigation Plan for PFIC. EMA has also granted orphan designation to odevixibat for the treatment of biliary atresia, Alagille syndrome and primary biliary cholangitis.

PATENT

https://patents.google.com/patent/US9694018B1/en

Example 5

1,1-Dioxo-3,3-dibutyl-5-phenyl-7-methylthio-8-(N—{(R)-α-[N—((S)-1-carboxypropyl) carbamoyl]-4-hydroxybenzyl}carbamoylmethoxy)-2,3,4,5-tetrahydro-1,2,5-benzothiadiazepine, Mw. 740.94.

This compound is prepared as described in Example 29 of WO3022286.

PATENT

https://patents.google.com/patent/WO2003022286A1/sv

Example 29

1,1-Dioxo-3,3-dibutyl-5-phenyl-7-methylthio-8-(N-((R)-α-[N-((S)- 1-carboxypropyl) carbamoyl]-4-hydroxybenzyl}carbamoylmethoxy)-2,3,4,5-tetrahydro-1,2,5-benzothiadiazepine

A solution of 1,1-dioxo-3,3-dibutyl-5-phenyl-7-methylthio-8-[N-((R)-α-carboxy-4-hydroxybenzyl)carbamoylmethoxy]-2,3,4,5-tetrahydro-1,2,5-benzothiadiazepine (Example 18; 0.075 g, 0.114 mmol), butanoic acid, 2-amino-, 1,1-dimethylethyl ester, hydrochloride, (2S)-(0.031 g, 0.160 mmol) and Ν-methylmorpholine (0.050 ml, 0.457 mmol) in DMF (4 ml) was stirred at RT for 10 min, after which TBTU (0.048 g, 0.149 mmol) was added. After 1h, the conversion to the ester was complete. M/z: 797.4. The solution was diluted with toluene and then concentrated. The residue was dissolved in a mixture of DCM (5 ml) and TFA (2 ml) and the mixture was stirred for 7h. The solvent was removed under reduced pressure. The residue was purified by preparative HPLC using a gradient of 20-60% MeCΝ in 0.1M ammonium acetate buffer as eluent. The title compound was obtained in 0.056 g (66 %) as a white solid. ΝMR (400 MHz, DMSO-d₆): 0.70 (3H, t), 0.70-0.80 (6H, m), 0.85-1.75 (14H, m), 2.10 (3H, s), 3.80 (2H, brs), 4.00-4.15 (1H, m), 4.65 (1H, d(AB)), 4.70 (1H, d(AB)), 5.50 (1H, d), 6.60 (1H, s), 6.65-7.40 (11H, m), 8.35 (1H, d), 8.50 (1H, d) 9.40 (1H, brs).

PATENT

https://patents.google.com/patent/US20140323412A1/en

PATENT

https://patents.google.com/patent/WO2013063526A1/e

PATENT

https://patents.google.com/patent/WO2019245448A1/en

The compound l,l-dioxo-3,3-dibutyl-5-phenyl-7-methylthio-8-(A/-{(R)-a-[A/-((S)-l-carboxypropyl) carbamoyl]-4-hydroxybenzyl}carbamoylmethoxy)-2,3,4,5-tetrahydro-l,2,5-benzothiadiazepine (odevixibat; also known as A4250) is disclosed in WO 03/022286. The structure of odevixibat is shown below.

As an inhibitor of the ileal bile acid transporter (IBAT) mechanism, odevixibat inhibits the natural reabsorption of bile acids from the ileum into the hepatic portal circulation. Bile acids that are not reabsorbed from the ileum are instead excreted into the faeces. The overall removal of bile acids from the enterohepatic circulation leads to a decrease in the level of bile acids in serum and the liver. Odevixibat, or a pharmaceutically acceptable salt thereof, is therefore useful in the treatment or prevention of diseases such as dyslipidemia, constipation, diabetes and liver diseases, and especially liver diseases that are associated with elevated bile acid levels.

According to the experimental section of WO 03/022286, the last step in the preparation of odevixibat involves the hydrolysis of a tert-butyl ester under acidic conditions. The crude compound was obtained by evaporation of the solvent under reduced pressure followed by purification of the residue by preparative HPLC (Example 29). No crystalline material was identified.

Amorphous materials may contain high levels of residual solvents, which is highly undesirable for materials that should be used as pharmaceuticals. Also, because of their lower chemical and physical stability, as compared with crystalline material, amorphous materials may display faster

decomposition and may spontaneously form crystals with a variable degree of crystallinity. This may result in unreproducible solubility rates and difficulties in storing and handling the material. In pharmaceutical preparations, the active pharmaceutical ingredient (API) is for that reason preferably used in a highly crystalline state. Thus, there is a need for crystal modifications of odevixibat having improved properties with respect to stability, bulk handling and solubility. In particular, it is an object of the present invention to provide a stable crystal modification of odevixibat that does not contain high levels of residual solvents, that has improved chemical stability and can be obtained in high levels of crystallinity.

Example 1

Preparation of crystal modification 1

Absolute alcohol (100.42 kg) and crude odevixibat (18.16 kg) were charged to a 250-L GLR with stirring under nitrogen atmosphere. Purified water (12.71 kg) was added and the reaction mass was stirred under nitrogen atmosphere at 25 ± 5 °C for 15 minutes. Stirring was continued at 25 ± 5 °C for 3 to 60 minutes, until a clear solution had formed. The solution was filtered through a 5.0 m SS cartridge filter, followed by a 0.2 m PP cartridge filter and then transferred to a clean reactor.

Purified water (63.56 kg) was added slowly over a period of 2 to 3 hours at 25 ± 5 °C, and the solution was seeded with crystal modification 1 of odevixibat. The solution was stirred at 25 ± 5 °C for 12 hours. During this time, the solution turned turbid. The precipitated solids were filtered through centrifuge and the material was spin dried for 30 minutes. The material was thereafter vacuum dried in a Nutsche filter for 12 hours. The material was then dried in a vacuum tray drier at 25 ± 5 °C under vacuum (550 mm Hg) for 10 hours and then at 30 ± 5 °C under vacuum (550 mm Hg) for 16 hours. The material was isolated as an off-white crystalline solid. The isolated crystalline material was milled and stored in LDPE bags.

An overhydrated sample was analyzed with XRPD and the diffractogram is shown in Figure 2.

Another sample was dried at 50 °C in vacuum and thereafter analysed with XRPD. The diffractogram of the dried sample is shown in Figure 1.

The diffractograms for the drying of the sample are shown in Figures 3 and 4 for 2Q ranges 5 – 13 ° and 18 – 25 °, respectively (overhydrated sample at the bottom and dry sample at the top).

References

^ Jump up to:^a ^b ^c ^d “First treatment for rare liver disease”. European Medicines Agency (EMA) (Press release). 21 May 2021. Retrieved 21 May 2021. Text was copied from this source which is © European Medicines Agency. Reproduction is authorized provided the source is acknowledged.
^ “Odevixibat”. Albireo Pharma. Retrieved 21 May 2021.
^ “Bylvay: Pending EC decision”. European Medicines Agency (EMA). 19 May 2021. Retrieved 21 May 2021.

External links

“Odevixibat”. Drug Information Portal. U.S. National Library of Medicine.

ClinicalTrials.gov

CTID	Title	Phase	Status	Date
NCT04336722	Efficacy and Safety of Odevixibat in Children With Biliary Atresia Who Have Undergone a Kasai HPE (BOLD)	Phase 3	Recruiting	2020-09-02
NCT04483531	Odevixibat for the Treatment of Progressive Familial Intrahepatic Cholestasis		Available	2020-08-25
NCT03566238	This Study Will Investigate the Efficacy and Safety of A4250 in Children With PFIC 1 or 2	Phase 3	Active, not recruiting	2020-03-05
NCT03659916	Long Term Safety & Efficacy Study Evaluating The Effect of A4250 in Children With PFIC	Phase 3	Recruiting	2020-01-21
NCT03608319	Study of A4250 in Healthy Volunteers Under Fasting, Fed and Sprinkled Conditions	Phase 1	Completed	2018-09-19

CTID	Title	Phase	Status	Date
NCT02630875	A4250, an IBAT Inhibitor in Pediatric Cholestasis	Phase 2	Completed	2018-03-29
NCT02360852	IBAT Inhibitor A4250 for Cholestatic Pruritus	Phase 2	Terminated	2017-02-23
NCT02963077	A Safety and Pharmakokinetic Study of A4250 Alone or in Combination With A3384	Phase 1	Completed	2016-11-16

EU Clinical Trials Register

EudraCT	Title	Phase	Status	Date
2019-003807-37	A Double-Blind, Randomized, Placebo-Controlled Study to Evaluate the Efficacy and Safety of Odevixibat (A4250) in Children with Biliary Atresia Who Have Undergone a Kasai Hepatoportoenterostomy (BOLD)	Phase 3	Ongoing	2020-07-29
2015-001157-32	An Exploratory Phase II Study to demonstrate the Safety and Efficacy of A4250	Phase 2	Completed	2015-05-13
2014-004070-42	An Exploratory, Phase IIa Cross-Over Study to Demonstrate the Efficacy	Phase 2	Ongoing	2014-12-09
2017-002325-38	An Open-label Extension Study to Evaluate Long-term Efficacy and Safety of A4250 in Children with Progressive Familial Intrahepatic Cholestasis Types 1 and 2 (PEDFIC 2)	Phase 3	Ongoing
2017-002338-21	A Double-Blind, Randomized, Placebo-Controlled, Phase 3 Study to Demonstrate Efficacy and Safety of A4250 in Children with Progressive Familial Intrahepatic Cholestasis Types 1 and 2 (PEDFIC 1)	Phase 3	Ongoing, Completed

Odevixibat
Clinical data

Trade names	Bylvay
Routes of administration	By mouth
ATC code	None
Identifiers
show IUPAC name
CAS Number	501692-44-0
PubChem CID	10153627
IUPHAR/BPS	11194
DrugBank	DB16261
ChemSpider	8329135
UNII	2W150K0UUC
KEGG	D11716
ChEMBL	ChEMBL4297588
Chemical and physical data
Formula	C₃₇H₄₈N₄O₈S₂
Molar mass	740.93 g·mol⁻¹
3D model (JSmol)	Interactive image
show SMILES
show InChI

////////////odevixibat, Orphan Drug Status, phase 3, Albireo, A-4250, A 4250, AR-H 064974

CCCCC1(CN(C2=CC(=C(C=C2S(=O)(=O)N1)OCC(=O)NC(C3=CC=C(C=C3)O)C(=O)NC(CC)C(=O)O)SC)C4=CC=CC=C4)CCCC

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Nucleic Acid Sequence

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1,3,2-Dioxaphosphorino(4′,5′:5,6)pyrano(3,2-g)pteridin-10(4H)-one, 8-amino-4a,5a,6,9,11,11a,12,12a-octahydro-2,12,12-trihydroxy-, 2-oxide, hydrobromide, hydrate (1:1:2), (4aR,5aR,11aR,12aS)-

1,3,2-Dioxaphosphorino(4′,5′:5,6)pyrano(3,2-g)pteridin-10(4H)-one, 8-amino-4a,5a,6,9,11,11a,12,12a-octahydro-2,12,12-trihydroxy-, 2-oxide,hydrobromide, hydrate (1:1:2), (4aR,5aR,11aR,12aS)-

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BridgeBio Pharma And Affiliate Origin Biosciences Announces FDA Acceptance Of Its New Drug Application For Fosdenopterin For The Treatment Of MoCD Type A

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Melflufen

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