Cidofovirסידופוביר سيدوفوفير

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Mar 072014

CIDOFOVIR

(S)-1-(3-Hydroxy-2-phosphonylmethoxypropyl)cytosine
[(S)-2-(4-Amino-2-oxo-1,2-dihydropyrimidin-2-yl)-1-(hydroxymethyl)ethoxymethyl]phosphonic acid

113852-37-2 CAS

120362-37-0 (Na salt)
149394-66-1 (dihydrate)

launched 1996 Gilead

EMA:Link,

US FDA:link

SYNTHESIS.. CHEMDRUG

Rega Instituut (Originator)

For the treatment of CMV retinitis in patients with acquired immunodeficiency syndrome (AIDS)

US5142051 PATENT

Canada	1340856	1999-12-21	EXPIRY 2016-12-21
United States	5142051	1993-06-26	2010-06-26

Cidofovir is a DNA polymerase inhibitor that was launched in 1996 by Gilead for the intravenous treatment of cytomegaloviral (CMV) retinitis in AIDS patients. Early clinical trials are underway at the National Institute for Allergy & Infectious Disease (NIAID) for the treatment of BK virus nephropathy (BKVN) in patients who have undergone kidney transplants.

Cidofovir suppresses CMV replication by selective inhibition of viral DNA synthesis. Biochemical data support selective inhibition of CMV DNA polymerase by cidofovir diphosphate, the active intracellular metabolite of cidofovir. Cidofovir diphosphate inhibits herpesvirus polymerases at concentrations that are 8- to 600-fold lower than those needed to inhibit human cellular DNA polymerases alpha, beta, and gamma1, 2, 3. Incorporation of cidofovir into the growing viral DNA chain results in reductions in the rate of viral DNA synthesis.

Cidofovir was originally developed under a collaboration between the Academy of Sciences of the Czech Republic and the Rega Institute for Medical Research. In 1991 and 1992, Gilead entered into license agreements with the Rega Institute that covered a large number of nucleotide analogue compounds and structures, including cidofovir. The drug became the subject of a marketing collaboration between Gilead and Pfizer (formerly Pharmacia & Upjohn) in August 1996 that covers all countries outside the U.S.

Cidofovir (brand name Vistide) is an injectable antiviral medication primarily used as a the treatment for cytomegalovirus (CMV) retinitis (an infection of the retina of the eye) in patients with AIDS.^[1]^[2]

Its only indication that has received regulatory approval worldwide is cytomegalovirus retinitis.^[1]^[2] Cidofovir has also shown efficacy in the treatment ofaciclovir-resistant HSV infections.^[3] Cidofovir has also been investigated as a treatment for progressive multifocal leukoencephalopathy with successful case reports of its use.^[4] Despite this meta-analyses have failed to demonstrate any efficacy in AIDS patients,^[5] and the limited data in non-AIDS patients fail to demonstrate any efficacy either.^[6] Cidofovir might have anti-smallpox efficacy and might be used on a limited basis in the event of a bioterror incident involving smallpox cases.^[7] A cidofovir derivative with much higher activity against smallpox that can be taken orally has been developed.^[8] It has inhibitory effects on varicella-zoster virus replication in vitro although no clinical trials have been done to date, likely due to the abundance of safer alternatives such as aciclovir.^[9] Cidofovir shows anti-BK virus activity in a subgroup of transplant patients.^[10] Cidofovir is being investigated as a complementary intralesional therapy against papillomatosis caused by HPV.^[11]^[12]
It first received FDA approval on the 26th of June 1996,^[13] TGA approval on the 30th of April 1998^[2] and EMA approval on the 23rd of April 1997.^[14]

Other

It has been suggested as an antitumour agent, due to its suppression of FGF2.^[15]^[16]

Cidofovir was discovered at the Institute of Organic Chemistry and Biochemistry, Prague, by Antonín Holý, and developed by Gilead Sciences^[20] and is marketed with the brand name Vistide by Gilead in the USA, and by Pfizerelsewhere.

The chemical name of cidofovir is 1-[(S)-3-hydroxy-2-(phosphonomethoxy)propyl]cytosine dihydrate (HPMPC), with the molecular formula of C8H14N3O6P•2H2O and a molecular weight of 315.22 (279.19 for anhydrous). The chemical structure is:

Cidofovir is a white crystalline powder with an aqueous solubility of ≥ 170 mg/mL at pH 6 to 8 and a log P (octanol/aqueous buffer, pH 7.1) value of -3.3.
Cidofovir Injection is a sterile, hypertonic aqueous solution for intravenous infusion only. The solution is clear and colorless. It is supplied in clear glass vials, each containing 375 mg of anhydrous cidofovir in 5 mL aqueous solution at a concentration of 75 mg/mL.
The formulation is pH-adjusted to 7.4 (range 7.1 to 7.7) with sodium hydroxide and/or hydrochloric acid and contains no preservatives. The appropriate volume of Cidofovir Injection must be removed from the single-use vial and diluted prior to administration

INTRODUCTION

Cidofovir’s chemical formula is C₈H₁₄N₃O₆P and its IUPAC name is ({[(S)-1-(4-amino-2-oxo-1,2-dihydropyrimidin-1-yl)-3-hydroxypropan-2-yl]oxy}methyl)phosphonic acid. Cidofovir has also been described as (S)-(1-(4-amino-2-oxopyrimidin-1(2H)-yl)-3-hydroxypropan-2-yloxy)methylphosphonic acid as well as possibly by other chemical names. Its chemical structure is:

Cidofovir was discovered at the Institute of Organic Chemistry and Biochemistry, Prague, and developed by Gilead Sciences. Today, cidofovir is an injectable antiviral medication for the treatment of cytomegalovirus (CMV) retinitis in patients with AIDS. It suppresses CMV replication by selective inhibition of viral DNA polymerase and therefore prevention of viral replication and transcription. It is an acyclic nucleoside phosphonate, and is therefore independent of phosphorylation by viral enzyme, in contrast to, for instance, acyclovir.

Cidofovir is marketed with the brand name Vistide® by Gilead in the United States and by Pfizer in other parts of the world. Vistide® is a sterile, hypertonic aqueous solution for intravenous infusion only. The solution is clear and colorless. It is supplied in clear glass vials, each containing 375 mg of anhydrous cidofovir in 5 mL aqueous solution at a concentration of 75 mg/mL. The formulation is pH-adjusted to 7.4 with sodium hydroxide and/or hydrochloric acid and contains no preservatives. Renal impairment is the major toxicity of Vistide®.

Presently, there are no Orange Book patents listed as having claims which cover Vistide®, although previously U.S. Pat. No. 5,142,051 was listed in the Orange Book for Vistide®. The ‘051 patent is not directed specifically to cidofovir or its crystalline forms. Instead, it broadly discloses N-phosphonylmethoxyalkyl derivatives of pyrimidine and purine bases.

Cytomegalovirus (Cytomegaoviyns, CMV) is one of the biggest dangers of the herpes virus, the body’s infection rates as high as 50% to 80% of the current adult prevalence rate of more than 95%, generally showed a latent infection, most infections had no clinical symptoms, but under certain conditions, the invasion of organs and systems to produce more severe disease. The virus can invade the lung, liver, kidney, salivary gland, mammary gland and other polymorphonuclear leukocytes and lymphocytes, and, since the long-term or intermittent saliva, milk sweat, blood, urine, semen, exclude uterine secretions of the virus. Spread through a variety of ways in the mouth, genital tract, placenta, blood transfusion or organ transplantation.

When the body’s immune dysfunction, such as infected with HIV, cancer patients undergoing radiotherapy, chemotherapy, organ or bone marrow transplantation immunosuppressive anti-rejection etc will stimulate active infection, can cause acute retinitis, interstitial pneumonia, gastroenteritis and encephalitis, blindness or death without treatment rate of over 70%. With the rise in HIV infection rates and organ transplants extensively for anti-CMV drugs is also increasing demand.

cidofovir (cidofovir, HPMPC) are novel ether derivatives of cytidine phosphono chemical name

[5]-NL [(3 – hydroxy-2 – methoxy-phosphonic acid) glycerol]-N4-cytosine, Molecular structure of the formula (I):

Gilead developed by the United States, in May 1996 the FDA approved injectable celecoxib Duofu Wei listed, France and Canada also continued with the approval of the use of the trade name Vistide. Its CAS number is 113852-37-2, formula C8H14N3O6P, the structure of formula (I). Cidofovir for CMV is highly inhibitory activity of certain ganciclovir or foscarnet resistant strains of the virus are also active. And herpes simplex virus (HSV), herpes zoster virus (VZV), human papillomavirus (HPV), also has a strong activity.

Its mechanism of action: cidofovir having a phosphoric acid group, a ring-opening mechanism of the antiviral nucleoside phosphonate compound (ANP) and the consistent cyclic nucleoside analogues are nucleosides or virus in vivo kinase activation into triphosphate metabolite, thereby inhibiting viral replication by DNA polymerase and reverse transcriptase. Unlike the three-step cyclic nucleoside analogues must phosphorylation reaction, ring opening nucleoside phosphonate group containing phosphorus compound itself, eliminating the first step of the phosphorylation reaction speed, and thus a higher activity. Cidofovir is absorbed when the cells in the cell pyrimidine nucleoside phosphorylase kinase (P bandit kinase and NDP kinase) to effect conversion of the active metabolite monophosphate (HPMPCp), diphosphate (HPMPCpp) and a bile acid base adducts. Cidofovir diphosphate inhibits viral DNA polymerase or reverse transcriptase activity, and its corresponding natural dNTP incorporated into the viral DNA chain competition, since no 3 – hydroxy end, continue to extend the DNA chain termination. Can slow the synthesis of DNA, viral DNA and to the loss of stability, thereby inhibiting viral replication, transcription of the ability to reduce viral DNA to exert antiviral activity. Compared with other anti-CMV drugs, cidofovir characteristics: significant and lasting effect, started the first two weeks administered once a week, then only administered once every two weeks, easy to use, and to reduce its toxicity side effects.

Several major techniques are based on the synthesis of cidofovir cytosine as starting material, mainly carried out to improve the synthesis of the side chain.

(I) J. Med Chem, 1989,32,1457 ~ 1463 discloses a synthetic process:

The route to cytosine as the raw material, with a chiral side chain by condensation, deprotection and reduction can be obtained in three steps cidofovir.However, chiral side chain subject to a six-step reaction system. The total yield is low, adverse side. And using Me3SiBr, so that the costs and the risk of surge, is not conducive to industrial production.

(2) US 5591852,1995-1-7; US 2005/023833 & WO 2006/014429 and US 2009/0270618, Tetrahedron Lett 1994,35,3243-3246 and “Chinese Journal of New Drugs”, 2007,16. , 1272-1274 for the synthesis of a lot of improvements:

Benzoyl cytosine with a chiral starting material and trityloxymethyl ethylene oxide condensation, deprotection and hydrolysis was then prepared by deprotection cidofovir group. The synthetic steps to make some shorter, but still use expensive Me3SiBr, adverse ones, the low yield of the security at the cost of industrial production is still unfavorable. (Several different patent protection only in the order of the amino cytosine different!)

(3) Patent Publication No. CN1690065A, CN1690066A, CN1690067A (2005 年 11 月 2 Publication Date) and the “Chinese Journal of Medicinal Chemistry” 2007,17,41-46, reported a new synthetic route:

The route of process steps is too long, the total yield is low, side effects side. But not conducive to industrial production.

(4) Patent No. CN 101205215A (25 June 2008 publicly) announced a halogen epoxy propane as a starting material for the synthesis route:

Use of the route (R) – epihalohydrin reaction with cytosine, cytosine ring because alkaline easily cause epoxy ring-opening reaction of the ring, but side reactions, the purified product is not, nor is suitable for industrial production.

Subsequently, the patent number CN 101525352A (2009 年 9 月 9 Publication Date) discloses (4) based on the modified route through epoxypropionate alkane ether in the form of a direct reaction with cytosine, after a series of similar steps obtain the final product cidofovir.

In view of the clinical application of cidofovir more favorable therapeutic effect in, looking for a high yield and because of economic and practical, easy to control, the risk of small synthetic methods and technology is now more urgent needs.

Synthesis

Brodfuehrer, P; Howell, Henry G.; Sapino, Chester; Vemishetti, Purushotham (1994). “A practical synthesis of (S)-HPMPC”. Tetrahedron Letters 35 (20): 3243. doi:10.1016/S0040-4039(00)76875-4.

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CN 102268040

Figure CN102268040AD00112

, Example 1:

1 Synthesis of 4,4 ‘- dimethoxytrityl methyl – (R) – glycidol (Compound III): The 5 04 g (15 mmoDDMT-Cl grain port 0 20 g (1 52 mmol… ) 4_ dimethylaminopyridine (DMAP) was dissolved in 100 mL CH2C12 cooled to 0 ° C, was added dropwise 10 mL TEA was slowly added 2. 00 g (27mmol) hydroxymethyl chiral oxirane (Compound II ) addition was completed, the reaction warmed to room temperature naturally. fly 4 h, until TLC until the disappearance of the detection DMT-Cl, the reaction was stopped by filtration, the filtrate was washed with saturated NaHC03 solution (50mLX2), saturated NaCl solution (50 mLX2), anhydrous Na2S04 dried, filtered, and concentrated to a viscous colorless directly, i.e., 5 08 g of 4,4 ‘-dimethoxy-triphenylmethyl _ -.. (R) – glycidol (Compound III), yield 90 %, HPLC purity 99%.

2, Synthesis (S)-N1_ [(2 – hydroxy-3 – (dimethoxytrityl) propyl] cytosine (Compound IV):. Under nitrogen to 3 56 g (32 mmol) of cytosine was added 150 mL of anhydrous N, N-dimethylformamide (DMF), and at room temperature, was added portionwise 1. 24 g (31 mmol, molar concentration of 60%) NaH, 0. 5 h after adding 11 92 g (31 mmol) 4,4 ‘-. dimethoxytrityl methyl – (R) – glycidol (Compound III), plus finished warming up to 10 (Tll (TC reaction . 6-8 h and then filtered, and the filtrate evaporated under reduced pressure DMF, the remaining solid phase was added 500 mL of ethyl acetate and 50 mL of water, separated and the organic layer was washed with saturated NaHC03 solution (50 mL X 2), saturated NaCl solution (50 mL X 2), dried over anhydrous Na2S04 filtered and dried, and concentrated to give 13 90 g of a white solid, S Jie (S)-Nl-[(2 -.. hydroxy-3 – (methoxy-dimethoxytrityl ) propyl] cytosine (Compound IV), yield 92%, HPLC purity 98%.

3 Synthesis ⑶-Nl-{[2_ (phosphonic acid methoxy diethoxy) -3 – (methoxy-dimethoxytrityl)] propyl} cytosine (Compound V):

75 ~ 80 ° C under the conditions, 48 76 g (0 100 mol.) (S) _N1_ [(2 – hydroxy-3 – (dimethoxytrityl) propyl]. Cytosine (Compound IV) was added to 150 mL anhydrous DMF, and then inputs 8. 5g (0. 050 mol) tert-butoxide, magnesium reaction 0.5-1 h, tosyloxy added diethyl 32 methylsulfinyl . 2 g (0. 100 mol), the reaction epileptic 8 h, p-toluenesulfonic acid was added to neutralize the excess alkali to neutral distilled DMF, ethyl acetate (300 mLX 3) washing the combined ethyl acetate phase was concentrated to give a solid, i.e., synthetic 58 18 g (S)-Nl-. {[2 – (diethoxy-phosphono-methoxy) -3 – (methoxy-dimethoxytrityl)] propyl} cytosine (Compound V), yield 89%, HPLC purity greater than 95%.

4 Synthesis of (S)-Nl-{[2_ (phosphonic acid methoxy diethoxy) -3 – hydroxy] propyl} cytosine (Compound VI): The 10 g (S)-Nl- {[2 – (phosphono-methoxy ethoxy) -3 – (methoxy-dimethoxytrityl)] propyl}-cell

Pyrimidine (compound V) was dissolved in a concentration of 70 mL of 80% acetic acid solution, 90 ° C reaction. After 5 h, cooled to room temperature, 50 mL of water and 30 mL of dichloromethane, and the organic phase washed with water (30 mL X2) and the combined aqueous phase was concentrated to give crude 9. 5 g, can be performed directly in the next reaction.

can also be separated by flash column chromatography (CH2C12 = MeOH = 10: 1), 4.6 g obtained as a pale yellow oil, i.e. (S)-Nl-{[2 – (methoxy diethoxy phosphono ) -3 – hydroxy] propyl} cytosine (Compound VI), yield 90%.

5 was synthesized ⑶-Nl-{[2_ (diphosphonic acid methoxy) -3 – hydroxy] propyl} cytosine (Compound I):

The 9.5g (S)-Nl-{[2 – (methoxy diethoxy phosphonomethyl) -3 – hydroxy] propyl} cytosine (Compound VI) into a crude product containing 5 76 g (0.. 045 mol) solution of hydrogen iodide, hydroiodic acid, and after reflux for 4-5 h. (50 mLX 2) wash solution was separated with ethyl acetate. The aqueous phase was added sodium hydroxide to adjust pH between 3 Γ3 6, filtered, recrystallized from methanol to give 3.81 g of white crystalline solid, S Jie (S)-Ni-{[2 -.. (Diphosphonic acid methoxy yl) -3 – hydroxy] propyl} cytosine (Compound I), yield 88% (containing two crystal water), HPLC purity greater than 99%.

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POLYMORPHS

US20120277191

Example 7 Amorphous Cidofovir

Intermediate 5 (FIG. 7; 0.5 g, 0.054 mol) was heated with a solution of sodium methoxide in methanol (0.5 M, 15 mL, 7.5 mmol) at 72° C. for 14.5 h then at 90° C. for 5.5 h. The reaction mixture was quenched with water (10 mL) and filtered through a bed of ion exchange resin Dowex® 50WX8 100-200 (H). The filtrate was cycled through the ion exchange bed (2 times) then washed successively with 1:1 methanol:water (40 mL), methanol (40 mL) and 4% triethylamine:methanol (50 mL). This ion-exchange bed was further washed with 48:48:4 methanol:water:triethylamine (100 mL) until no UV absorbance was detected in the filtrate. This reaction produced intermediate 7 (FIG. 7) together with cyclic cidofovir impurity. This mixture was then dissolved in 6 N HCl and heated to 65° C. After cooling the reaction mixture to room temperature, ethyl acetate was charged and stirred and the aqueous layer separated. The aqueous was stirred with ethanol (50 mL). The precipitated material was filtered and the solid was washed with ethanol. The ethanol filtrate was concentrated. The concentrated material was taken up in acetonitrile and stirred with trimethylsilyl bromide (19 mL) at room temperature for 18 h. The reaction mixture was filtered and the filtrate concentrated. The residue was taken up in toluene (30 mL) and ammonium hydroxide (28%, 50 mL) was charged and stirred at room temperature. The organic phase was separated and the aqueous phase was concentrated to dryness. Water (20 mL) and ethanol (15 mL) were added to the residue. The mixture pH was 6 and was adjusted to pH 3 with concentrated HCl (2 mL) then adjusted to pH 4 to 4.5 with 28% NH₄OH. After stirring for 0.5 h, the mixture was cooled, filtered and the solids washed with 2:1 EtOH:H₂O and dried under vacuum for 18 h. The isolated solid was taken up in water (10 mL) and 28% NH₄OH added to give a solution. Concentrated HCl was added to the solution until pH 4 was reached. Ethanol (13 mL) was charged and the mixture stirred at −17° C. for 18 h, filtered and the solids washed with 2:1 EtOH:water (2×8 mL), dried under vacuum at 35° C. The cidofovir isolated in this manner was determined to be in the amorphous form by XRPD.

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Journal of the American Chemical Society, 2011 , vol. 133, 7 p. 2264 – 2274

http://pubs.acs.org/doi/abs/10.1021/ja109823e

Abstract Image

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11	*	PETR ALEXANDER AND ANTONÍN HOL: “General Method of Preparation of N-[(S)-(3-Hydroxy-2-phosphonomethoxypropyl )] Derivatives of Heterocyclic Bases“, COLLECTION OF CZECHOSLOVAK CHEMICAL COMMUNICATIONS, INSTITUTE OF ORGANIC CHEMISTRY & BIOCHEMISTRY, PRAGUE; CZ, vol. 58, no. 5, 1 May 1993 (1993-05-01), pages 1151-1163, XP008152404, ISSN: 0010-0765, DOI: 10.1135/CCCC19931151
12	*	SNOECK, ROBERT ET AL: “(S)-1-(3-Hydroxy-2-phosphonylmethoxypropy l)cytosine, a potent and selective inhibitor of human cytomegalovirus replication“, ANTIMICROBIAL AGENTS AND CHEMOTHERAPY , 32(12), 1839-44 CODEN: AMACCQ; ISSN: 0066-4804, 1988, XP002677023,
13	*	SYMERSK AND A HOL J: “Structure of 1-(S)-(3-hydroxy-2-phosphonylmethoxypropyl )cytosine; an antiviral agent“, ACTA CRYSTALLOGRAPHICA SECTION C. CRYSTAL STRUCTURE COMMUNICATIONS, MUNKSGAARD, COPENHAGEN, DK, vol. 47, no. 10, 15 October 1991 (1991-10-15), pages 2104-2107, XP008152403, ISSN: 0108-2701, DOI: 10.1107/S0108270190013257
14	*	ULRIKA ERIKSSON: “Synthesis and biological evaluation of novel cidofovir prodrugs targeting HPepT1“, SYNTHESIS AND BIOLOGICAL EVALUATION OF NOVEL CIDOFOVIR PRODRUGS TARGETING HPEPT1- A DISSERTATION PRESENTED TO THE FACULTY OF THE GRADUATE SCHOOL UNIVERSITY OF SOUTHERN CALIFORNIA IN PARTIAL FULFILLMEN , 1 January 2005 (2005-01-01), pages 1-287, XP008152406, Retrieved from the Internet: URL:http://search.proquest.com/pqdtscieng/docview/305421788/fulltextPDF/1371C222DB27F96BE2D/2?accountid=29404
15	*	WEBB, ROBERT R., II ET AL: “Synthesis of (S)-N1-(3-hydroxy-2-phosphonylmethoxy)prop ylcytosine, (S)-HPMPC“, TETRAHEDRON LETTERS , 29(43), 5475-8 CODEN: TELEAY; ISSN: 0040-4039, 1988, XP002677025

CN1559429A *	Feb 18, 2004	Jan 5, 2005	肖广常	注射用西多福韦冻干粉针剂
CN1690066B *	Apr 19, 2004	Apr 28, 2010	横店集团成都分子实验室有限公	抗病毒剂西多福韦新衍生物
CN101525352A *	Feb 16, 2009	Sep 9, 2009	苏州凯达生物医药技术有限公司	西多福韦及其中间体的制备方法
CN102268040A *	Sep 5, 2011	Dec 7, 2011	扬州三友合成化工有限公司	抗病毒药物西多福韦的一种合成方法
US5142051	Jul 17, 1987	Aug 25, 1992	Ceskoslovenska Akademie Ved	N-phosphonylmethoxyalkyl derivatives of pyrimidine and purine bases and a therapeutical composition therefrom with antiviral activity

Uncialamycin

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Mar 052014

Uncialamycin

(1aS,11S,11aR,18R)-3,18-Dihydroxy-11a-[1(R*)-hydroxyethyl]-9,10,11,11a-tetrahydro-4H-11,1a-[3]heptene[1,5]diynonaphtho[2,3-h]oxireno[c]quinoline-4,9-dione

439.4163

C26 H17 N O6

870471-83-3 cas

WO2007038868A2, WO2013122823A1,

University of British Columbia (Originator)

uncialamycin, an enediyne natural product isolated from the Streptomyces uncialis, bacteria present on the surface of the lichen Cladonia uncialis.

Laboratory cultures of an undescribed streptomycete obtained from the surface of a British Columbia lichen produce uncialamycin (1), a new enediyne antibiotic.Uncialamycin exhibits potent in vitro antibacterial activity against Gram-positive and Gram-negative human pathogens, including Burkholderia cepacia, a major cause of morbidity and mortality in patients with cystic fibrosis.

Uncialamycin is an enediyne antibiotic with some unprecedented activity. The isolationists have filed a patent application almost right away. The total synthesis by Nicolaou [ACIE2007, 46, 4704] goes along nearly the same lines that have been predicted, and similar to Myers’ synthesis of dynemicin A [JACS 1997, 119, 6072], only it is not paper chemistry but the real one.

They have easily constructed the quinoline system with required functionality and subjected it to AllocCl-assisted acetylide addition (if I interpreted correctly “92% yield based on 80% conversion”). 5-alkoxyquinoline system was later advanced to iminoquinone and the two remaining rings were again attached by Hauser annulation with 3-cyanophthalide. The final product turned out be different from the one reported, more precisely, it was a C26-epimer. It is funny that I have accidentally drawn the correct structure with R-configuration at C-26 in the previous post.

The synthetic scheme allowed to easily invert this stereocenter via oxidation/reduction sequence on the last compound shown on the scheme below. The spectral properties of the final product thus obtained matched the reported data, and the structure of uncialamycin was confirmed by X-ray, despite it was isolated as an oil. The structure on the right is the revised one. The remaining details, including the chemistry behind DNA-cleaving Bergmann cycloaromatization,

Total Synthesis and Stereochemistry of Uncialamycin

K. C. Nicolaou, Hongjun Zhang, Jason S. Chen, James Crawford, Laxman Dasunoori

¹Department of Chemistry and, The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA
²Department of Chemistry and Biochemistry, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA

A new tot synth of Uncialamycin by Nicolaou. This is a natural occurring enediyne. Because the stereochemistry of C26 was unknown, both diastereomers as shown were synthesized. The retrosyn led back to simpler fragments 2, 3, and 4.

The following scheme illustrates the route to fragment 2. The key transformation was the two-step Friedlander quinoline synthesis (7 to 9).

Then fragment 2 was used in the following sequence. The key steps in the sequence involved installation of enediyne fragment 3 to give 11, the closure of the macrocycle to give 15, and the Hauser annulation in the last step to give 1a from 16.

In this case, it was found that the final product’s spectrum (1a) did not match the reported value. And therefore, the other isomer was synthesized. This was easily done using fragment 12 through oxidation-reduction sequence to give 18 with the opposite stereochemistry at C26. Sequence in Scheme 3 was then repeated on this fragment.

And 1b was found to match spectrum of the natural isomer. This natural compound was found to be stable as a solid and as solutions in a variety of solvents. But in presence of dray HCl in CH2Cl2 solution at rt, it rapidly converts to hexacyclic 19 through a cascade of Bergman cycloaromatization reaction. This cascade of reactions is believed to be responsible mode of action in damaging DNA and killing cells.

The enediynes are a family of antibiotics that possess a distinctive strained nine- or ten-member ring system comprising a Z-carbon-carbon double bond and two carbon- carbon triple bonds, usually arranged with the latter two flanking the former. The enediynes are potent damagers of DNA, causing single and double strand cuts. Their potency is attributed to their ability to bind to DNA and undergo a Bergmann rearrangement in which the strained ring system is converted into a highly reactive 1 ,4-benzenoid diradical, which damages the DNA by abstracting hydrogens from it.

Uncialamycin is an enediyne isolated from a Streptomyces strain found on the lichen Cladonia uncialis (Davies et al. 2005; 2007). (Full citations for references cited in this specification by first named author or inventor and year are provided in the section entitled “REFERENCES” later herein.)

Uncialamycin

The structure of uncialamycin has been confirmed by total synthesis (Nicolaou et al. 2007a; 2007b). In the course of the synthesis, it was noted that the unnatural 26(S) epimer was almost as active as the natural 26(R) epimer – that is, the stereochemistry of the C27 methyl had a minor effect on biological activity. Both epimers were active against several ovarian tumor cell lines. The IC₅₀ values rang ed from 9 x 10^“12 to 1 x 10^“10, depending on the epimer and cell line or sub-line (Nicolaou et al, 2008).

Conjugates are an important method for the delivery of anti-cancer drugs, which are often highly cytotoxic and might otherwise be problematic to administer due to the risk of systemic toxicity. In a conjugate, the drug is conjugated (covalently linked) to a targeting moiety that specifically or preferentially binds to a chemical entity characteristic of the cancer cell, thus delivering the drug there with high specificity. Further, the drug is held in an inactive form until released from the conjugate, usually by cleavage of the covalent linker.

Typically, the targeting moiety is an antibody or an antigen-binding portion thereof, whose antigen is overexpressed or uniquely expressed by a cancer cell (“tumor associated antigen”). In such instances, the resulting conjugate is sometimes refered to as an “immunoconjugate” or an “antibody-drug conjugate” (ADC). Preferably the tumor associated antigen is located on the surface of the cancer cell, but also can be one that is secreted into the vicinal extracellular space. Upon binding, the antigen-conjugate complex is internalized and eventually finds its way inside a vesicular body such as a lysosome, where the covalent linker is cleaved, liberating active drug to exert its chemotherapeutic effect.

Advantageously, the covalent linker is designed such that cleavage is caused by a factor prevalent inside a cancer cell but not in plasma. One such factor is the low lysosomal pH, so that the covalent linker can be an acid-sensitive group such as a hydrazone. Another such factor is the generally higher intracellular concentration of glutathione, allowing for the cleavage of a disulfide covalent linker by a disulfide exchange mechanism. Yet another such factor is the presence of lysosomal enzymes such as cathepsin B, which can cleave peptide linkers designed to be preferred substrates (Dubowchik et al. 2002).

Conjugates have been used to deliver enediyne drugs in oncology. Gemtuzumab ozogamicin (Mylotarg®) is a conjugate of an anti-CD33 monoclonal antibody and a derivative of the enediyne calicheamicin. It was approved for treatment of acute

myelogenous leukemia but was later withdrawn from the market. Several other enediyne drugs, especially in the conjugated form, have been the subject of development efforts

If handled carefully, enediynes make powerful cancer drugs.

Inventors N. S. Chowdari, S. Gangwar, and B. Sufi synthesized enediyne compounds with general formula 1 that are based on the natural enediyne uncialamycin (2) scaffold (Figure 1). These compounds, used alone or in conjugates, are potent cytotoxins that may be useful in cancer chemotherapy.

Enediynes are a class of natural antibiotics that are characterized by 9- or 10-membered rings that contain two C≡C bonds separated by a cis (Z)-substituted C=C bond. Enediynes can undergo Bergman cyclization to form 1,4-benzenoid diradicals, which abstract hydrogen atoms from other molecules. When the diradical is generated near DNA, it abstracts hydrogen atoms from the sugar backbone of the DNA molecule and results in single- and double-strand lesions.

The high reactivity of enediynes toward DNA makes them very toxic. Their potent activity may be beneficial, however, if they are used to target the DNA of cancerous tumors. Most enediynes inhibit the proliferation of various cancer cells, including those that resist other chemotherapeutic drugs. Several naturally occurring enediynes are in clinical trials against cancer.

Both epimers at C26 of the natural enediyne uncialamycin are active against several ovarian tumor cell lines, with IC50 values ranging from 9 × 10–12 to 1 × 10–10 M, depending on the epimer and the cell line or subline. The synthetic enediynes described by the inventors are derivatives of uncialamycin.

Using these toxic molecules demands specific delivery systems. Conjugates are innovative drug-delivery systems designed to target tumor cells precisely and minimize the risk of systemic toxicity. Typically, drugs are linked covalently to conjugates that act as targeting moieties, which specifically or preferentially bind to a chemical entity characteristic of the cancer cell.

The covalent linker is designed to be cleaved only by a factor that exists inside a cancer cell and not in plasma, so that the drug remains in an inactive form until it is released from the conjugate. A typical targeting moiety may be a polymer or an antibody. Polymer-conjugated and antibody-linked enediyne drugs such as gemtuzumab ozogamicin (Mylotarg) were used to deliver enediyne drugs to cancer cells. Mylotarg, however, has been withdrawn from the market because of high patient mortality.

General formula of compounds of the invention and natural enediyne uncialamycin

Compounds of structure 1 may be conjugated to a targeted moiety through a chemical bond to substituent R1. Compounds 3 and 4, shown in Figure 2, are examples of the synthetic enediynes with structure 1.

The investors tested the antiproliferative activities of several compounds against cancer cell lines. EC50 data for compounds 3 and 4 against 786-0 renal cancer cells and H226 lung cancer cells are shown in the table:

Example	786-0 cells, EC50 (nM)	H226 cells, EC50 (nM)
3	1.275	0.986
4	0.058	0.873

Representative examples of synthetic enediynes

Several assays were also conducted on conjugates derived from other compounds of formula 1. (Bristol-Myers Squibb [Princeton, NJ]. WIPO Publication 2013122823, Aug 22, 2013;

DAVIES ET AL.: ‘UNCIALAMYCIN, A NEW ENEDIYNE ANTIBIOTIC‘ ORGANIC LETTERS vol. 7, no. 23, 13 October 2005, pages 5233 – 5236

http://pubs.acs.org/doi/abs/10.1021/ol052081f

Production cultures of the producing strain were grown as lawns on solid agar medium (ISP4, 16 L) for 14−21 days at 30 °C. The solid agar cultures were extracted repeatedly with EtOAc. Concentration of the combined EtOAc extracts in vacuo gave a gummy residue that was partitioned between EtOAc and H2O. The EtOAc soluble material was fractionated by sequential application of flash C-18 reversed-phase chromatography (eluent: step gradient from H2O to MeOH) and reversed-phase HPLC (column-Inertsil ODS-2; eluent: CH3CN/H2O 40:60) to give pure uncialamycin (1) (

300 μg) as a bright purple [UV(MeOH): λmaxnm (ε) 206 (25000), 254 (33000), 280 (shoulder), 320 (shoulder), 539 (9400)] optically active ([α]D +3300 (c 0.005, MeOH)) oil.

Uncialamycin (1) gave a [M + Na]+ ion at m/z 462.0956 in the HRESIMS appropriate for a molecular formula of C26H17NO6 (calcd for C26H17NO6Na 462.0954) requiring 19 sites of unsaturation. NMR data for uncialamycin were recorded in DMSO-d6 at 600 MHz using a cryoprobe. The 13C NMR spectrum (Table 1) showed well-resolved resonances for 26 carbon atoms, and the 1H NMR spectrum contained resonances integrating for 17 protons, in agreement with the HRMS data. Inspection of the HSQC data revealed that four of the protons (δ 5.39, 6.66, 10.0, and 13.2) were not attached to carbon atoms. Two major fragments A and B (Figure 1) of uncialamycin could be identified from analysis of the COSY, HSQC, and HMBC data obtained for the molecule.

Table 1. 13C and 1H NMR Assignments for Uncialamycin (1). Data were Recorded in DMSO-d6 at 600 MHz for 1H

position	δ 13C	δ 1H (mult., J (Hz))
1		10.0 (d, 4.6)
2	143.6
3	110.4
4	187.0a
5	134.4b
6	126.1c	8.23 (dd, 1.4, 7.6)c
7	133.6d	7.88 (ddd, 1.4, 7.6, 7.6)d
8	134.9d	7.94 (ddd, 1.4, 7.6, 7.6)d
9	126.6c	8.24 (dd, 1.4, 7.6)c
10	132.2b
11	182.2a
12	112.7
13	154.9
14	129.9	8.51 (s)
15	135.6
16	63.5
17	63.0	5.14 (d, 3.3)
18	100.4
19	89.7
20	123.4	6.05 (dd, 0.8, 10)
21	124.0	5.97 (ddd, 1.4, 1.5, 10)
22	87.4
23	98.9
24	43.2	5.04 (dd, 1.5, 4.6)
25	76.0
26	63.6	4.31 (qd, 6.0, 6.0)
27	22.1	1.30 (d, 6.0)
13-OH		13.2 (brd.s)
17-OH		6.66 (brd.s)
26-OH		5.39 (d,6.0)

a−d May be interchanged.http://pubs.acs.org/doi/suppl/10.1021/ol052081f/suppl_file/ol052081fsi20051004_065853.pdf

……………..

WO2007038868A2

Isolation of Uncialamvcin

[0034] As part of a screening program aimed at discovering new antibiotics active against Bcc, it was found that crude organic extracts of cultures of a previously undescribed Streptomycete showed potent in vitro inhibition of Bcc. Bioassay guided fractionation of the crude extracts led to the identification of uncialamycin (1), a new enediyne antibiotic, as the active component. Bioactivity-guided fractionation involves thin layer chromatography of the extracts and fractions thereof and detection of the activity by overlaying a sensitive tester strain. A zone of inhibition identifies the active fraction containing the active compound.

The producing strain was extracted from the surface of the lichen Cladonia uncialis collected near Pitt River, British Columbia. Characterisation by 16S RNA sequencing showed the strain to be related, but not identical, to Streptomyces cyanogenus. Antibiotic activity of the strain was assayed by cutting plugs from solid agar cultures of the strain and placing them on lawns of tester strains of bacteria. Good inhibitory activity was detected against Gram-positive and Gram-negative bacteria (including Bcc), but not against yeasts.

Production cultures of the producing strain were grown as lawns on solid agar medium ISP4 for 14 to 21 days at room temperature. The solid agar cultures were lyophilized and extracted repeatedly with EtOAc. Concentration of the combined EtOAc extracts in vacuo gave a gummy residue that was partitioned between EtOAc and H₂O. The EtOAc soluble material was fractionated by sequential application of flash C- 18 reversed-phase chromatography (eluent: step gradient from H₂O to MeOH) and reversed-phase HPLC (column-Inertsil ODS-2; eluent: CH₃CN/H₂O 40:60) to give pure uncialamycin (1) (~ 300 μg) as a bright purple [UV(MeOH): λ_maxnm (ε) 206 (25,000), 254 (33,000), 280 (shoulder), 320 (shoulder), 539 (9,400)], optically active ([α]_D +3,300 (c 0.005, MeOH)) oil.

Chemical Characterization of Uncialamycin

Uncialamycin (1) gave a [M + Na]⁺ ion at m/z 462.0956 in the

HRESIMS appropriate for a molecular formula Of C₂₆H₁₇NO₆ (calc’d for C₂₆H₁₇NO₆Na 462.0954) requiring 19 sites of unsaturation. NMR data for uncialamycin was recorded in DMSO-^₆ at 600 MHz using a cryoprobe. The ¹³C NMR spectrum (Table 1) showed well-resolved resonances for 26 carbon atoms and the ¹H NMR spectrum contained resonances integrating for 17 protons in agreement with the HRMS data. Inspection of the HMQC data revealed that four of the protons (δ 5.39, 6.66, 10.0, and 13.2) were not attached to carbon atoms. Two major fragments A and B (Figure 1) of uncialamycin could be identified from analysis of the COSY, HMQC, and HMBC data obtained for the molecule.

Position δ ¹W WH^mult, J(Hz)) ,

1 10.0 (d, 4.6)

2 143.6

3 110.4

4 187.0

5 134.4

6 126.1 8.23 (dd, 1.4, 7.6)

7 133.6 7.88 (ddd, 1.4, 7.6, 7.6)

8 134.9 7.94 (ddd, 1.4, 7.6, 7.6)

9 126.6 8.24 (dd, 1.4, 7.6)

10 132.2

11 182.2

12 112.7

13 154.9

14 129.9 8.51 (s)

15 135.6

16 63.5

17 63.0 5.14 (d, 3.3)

18 100.4

19 89.7

20 123.4 6.05 (dd, 0.8, 10)

21 124.0 5.97 (ddd, 1.4, 1.5, 10)

22 87.4

23 98.9

24 43.2 5.04 (dd, 1.5, 4.6)

25 76

26 63.6 4.31 (qd, 6.0, 6.0)

27 22.1 1.30 (d, 6.0)

13-OH 13.2 (brd.s)

17-OH 6.66 (brd.s)

26-OH 5.39 (d,6.0)

Table 1. C and H NMR assignments for uncialamycin (1). Data were recorded in OMSO-d₆ at 600 MHz for ¹H. [0038] A pair of olefinic resonances at δ 5.97 (H-21 ) and 6.05 (H-20), that were strongly correlated to each other in the COSY spectrum and had a coupling constant of 10 Hz, were assigned to a cis disubsituted olefin. The upfield olefinic resonance at δ 5.97 (H-21) showed strong HMBC correlations to non-protonated carbon resonances at δ 89.7 (C- 19) and 98.9 (C-23), and the downfield olefinic resonance at δ 6.05 (H-20) showed strong correlations to non-protonated carbon resonances at δ 87.4 (C-22) and 100.4 (C- 18). This suite of HMBC correlations identified an enediyne substructure in 1 (see Fragment A in Figure 1). The olefinic resonance at δ 5.97 (H-21) showed a long range COSY correlation to a methine resonance at δ 5.04 (H- 24), indicating that the carbon bearing the methine proton (C-24: δ 43.2) was attached to the C-23 alkyne carbon. A COSY correlation observed between the methine (δ 5.04, H-24) and a broad singlet at 10.0, that was not correlated to a carbon in the HMQC spectrum, and the chemical shift of the methine carbon (C-24, δ 43.2) suggested that C-24 had an NH substituent. HMBC correlations observed between the H-24 methine (δ 5.04) and the two alkyne carbon resonances at δ 87.4 (C-22) and 98.9 (C-23) confirmed the attachment of C-24 to the C-23 alkyne carbon.

A singlet methine resonance at δ 5.14 (H- 17) showed HMBC correlations to the alkyne carbon resonances at δ 89.7 (C- 19) and 100.4 (C- 18), which demonstrated that the methine carbon (C- 17: δ 63.0) was linked to the second alkyne at C-18. Both of the methine resonances at δ 5.04 (H- 24) and 5.14 (H- 17) showed HMBC correlations to a pair of deshielded resonances at δ 63.5 (C- 16) and 76.0 (C-25), assigned to non-protonated oxygen bearing carbons. This set of four HMBC correlations indicated that the two oxygenated carbons bridged the C- 17 and C-24 carbons to form a ten membered ring (C- 16 to C-25) containing the enediyne substructure. A COSY correlation between the methine resonance at δ 5.14 and a broad singlet at 6.66 (17-OH) revealed an alcohol funtionality attached to the methine carbon.

A methyl doublet at δ 1.30 (Me-27, J = 6 Hz) was correlated in the COSY spectrum to a methine at 4.31 (H-26, q, J = 6.0 Hz)) that was further correlated to a broad singlet at 5.39 (OH-26), assigned to an alcohol. The methyl resonance (δ 1.30, Me-27) showed an HMBC correlation to the carbon resonance at 76.0 (C-25), indicating that the hydroxyethyl fragment (C-26 and C-27) was the fourth subsituent on the non-protonated carbon C- 25. Both the NH-I proton (δ 10.0) and the H-17 methine (5.14) were correlated to a carbon at δ 135.6 (C- 15), and the H-24 methine (δ 5.04) was correlated to a carbon at 143.6 (C-2) in the HMBC spectrum indicating that the NH and C- 16 were vicinal substituents on an olefin or aromatic ring. A deshielded singlet at δ 8.51 showed strong HMBC correlations into carbon resonances at δ 63.5 (C-16), 143.6 (C-2), and 112.7 (C- 12) and a weak correlation into the carbon resonance at 154.9 (C- 13). This set of HMBC correlations confirmed that the NH and C-16 were attached to a benzene ring. Based on the assumption that the intense HMBC correlations were through three bonds, these correlations also indicated that the aromatic methine (δ 8.51, H-14) was ortho to C-16 (δ 63.5) and meta to the NH (C-2, δ 143.6). The weak HMBC correlation between δ 8.51 and 154.9 was attributed to a two bond coupling, placing the carbon at 154.9 (C-13) ortho to the methine carbon (C- 14) and its chemical shift required an oxygen substituent. [0041] The second fragment B of uncialamycin contained an isolated

¹H spin system comprised of four contiguous aromatic protons (δ 8.23, dd, J = 1.4, 7.6 Hz H-6; 7.88, ddd, 1.4, 7.6, 7.6 Hz H-7; 7.94, ddd, J = 1.4, 7.6, 7.6 Hz H-8; 8.24, dd, J = 1.4, 7.6 Hz H-9). HMBC correlations observed between the proton resonance at δ 8.23 (H-6) and a carbon resonance at 187.0 (C-4) and between the proton resonance at 8.24 (H-8) and a carbon resonance at 182.2 (C-11) suggested that the other two subsituents on the benzene ring were quinone carbonyls. Fragments A and B shown in Figure 1 accounted for all of the carbon, hydrogen, and nitrogen atoms in the molecular formula of uncialamycin (1), but contained one extra oxygen atom. In order to complete the quinone and satisfy the remaining aromatic valences in Fragment A, the two carbonyl carbons of fragment B (C-4 and C-I l) had to be attached to the two substituted aromatic carbons (C-3 and C- 12) of fragment A. Finally, it was apparent that the two oxygentated carbons C- 16 and C-25 had to be bridged by an epoxide to account for the number of oxygen atoms and sites of unsaturation required by the molecular formula of 1. This implied that the C- 13 oxygen substituent had to be part of a phenol functionality that would engage in intramolecular hydrogen bonding with the C-I l carbonyl consistent with the observed OH chemical shift of δ 13.2.

A ROESY correlation between δ 5.14 (H- 17) and 4.31 (H-26) showed that C-26 and C- 17 were cis oriented about the C-16/C-25 epoxide and also defined the relative stereochemistry of H- 17 as shown. Molecular models revealed that due to steric and bond angle strain the C- 17 to C-23 enediyne containing bridge could only reasonably be cis fused to the piperidine ring. Uncialamycin (1) shares structural features with dynemicin A (2) and deoxydynemicin A (3) isolated from Micromonospora chersina. The H-24 resonance in uncialamycin (1) has a chemical shift of δ 5.04 and a 4.6 Hz coupling to the NH-I proton, which is nearly identical to the chemical shift (δ 5.05) and coupling (J = 4.3 Hz) of the corresponding methine proton (H-2) in dynemicin A (2), in agreement with the relative stereochemical assigment at C-24 in 1. Comparison of the additional NMR assigments reported for dynemicin A (2) and its triacetate derivative provided further strong support for the assigned structure of uncialamycin

…………….

Angewandte Chemie – International Edition, 2008 , vol. 47, 1 p. 185 – 189

http://onlinelibrary.wiley.com/doi/10.1002/anie.200704577/abstract

The highly potent DNA-cleaving molecule uncialamycin (1) was prepared in an asymmetric total synthesis featuring an enantioselective Noyori reduction. Compound 1 and its C26 epimer exhibit impressive broad-spectrum antibacterial properties and highly potent antitumor activities against a variety of cell lines.

LUPIN.. giant leap forward

companies, Uncategorized Comments Off

Mar 022014

Lupin Limited

news

Expanding domestic portfolio, ability to maintain market share in current products and new launches are key growth triggers
The lipid control or cholesterol lowering segment is emerging as a key growth driver for Lupin’s sales in the US. The first among three products is Tricor. The Lupin stock had corrected about five per cent in May on the announcement that Mylan would launch the $1.2 billion (Rs 7,200 crore) drug in tablet form.

Lupin has been able to maintain its generic market share so far with a share of 34 per cent vis-a-vis Mylan (market share of one-two per cent). While Balaji Prasad and Rohit Goel of Barclays estimate the drug will contribute about $29 million to Lupin’s revenues in the June 2014 quarter, Ebitda margins are expected to expand by 210 basis points, feel analysts at Kotak Securities.

Though the launch of generics in capsule form by Mylan has to be watched carefully, Lupin’s Antara, along with authorised generics, continues to dominate with 70 per cent share, observes Hitesh Mahida at Fortune Research.

In addition to Tricor, two other products in the cholesterol lowering segment the company is eyeing are Trilipix and Niaspan. Instead of a launch in January 2014 as was anticipated earlier, the company is now likely to launch the generics version of this $550 million drug Trilipix this month.

While the company says it is mulling options about the launch, Edelweiss analysts believe an early launch is likely to add $11 million in FY14 and about $15 million in FY15 to the company’s revenues. The third drug in this segment is Niaspan, to be launched in March 2014 and expected to add about $35 million to the company’s revenues in FY15. Together, the three cholesterol controlling drugs are expected to contribute $85-90 million to FY15 sales for Lupin.

Lupin Limited is a transnational pharmaceutical company based in Mumbai. It is the 2nd largest Indian pharma company by market capitalization;^[14] the 14th largest generic pharmaceutical company globally^[15] and; the 5th largest generic pharmaceutical company in the US by prescription-led market share.^[16] It has the distinction of being the fastest growing generic pharmaceutical player in the two largest pharmaceutical markets of the world – the US^[17] and Japan;^[18] and is the 5th largest ^[19] and the fastest growing generic pharmaceutical player in South Africa.^[20]

Market Position in the US gradually improving

Lupin 5thlargest generic company in the US in terms of prescriptions

14 products are market leader and 27 (among top 3) out of 30


Type	Public
Traded as	BSE: 500257 NSE: LUPIN
Industry	Pharmaceuticals
Founded	1968 ^[1]
Founder(s)	Dr. Desh Bandhu Gupta^[2]
Headquarters	Mumbai, Maharashtra^[3], India
Key people	Dr. Kamal K Sharma, Vice Chairman;^[4]……………………………………..Vinita Gupta, Chief Executive Officer, Lupin Limited;^[5] …………………. Nilesh Gupta, Managing Director;^[6] …………… Shakti Chakraborty, Group President – India Region Formulations & CIS;^[7] ……………………. Vinod Dhawan, Group President AAMLA & Business Development;^[8] ……………………. Ramesh Swaminathan, President Finance & Planning.^[9] New leadership team at Lupin from September 2013^[10]
Products	Pharmaceuticals, branded andgeneric drugs, biotechnology, Advanced Drug Delivery Systems, New Chemical Entity Research, vaccines, Over-the-Counter drugs
Revenue	9461 crore (US$1.5 billion) (2012-2013)^[11]^[12]
Profit	1314 crore(US$210 million) (2013)
Employees	11355^[13]
Subsidiaries	Lupin Pharmaceuticals, Inc. Kyowa Pharmaceutical Industry Co. Ltd. I’rom Pharmaceutical Co. Ltd. Pharma Dynamics Multicare Pharmaceuticals Generic Health Pte. Ltd. Hormosan Pharma GmbH
Website	www.lupinworld.com

2012 .http://icra.in/Files/ticker/Indian%20Pharmaceutical%20Sector.pdf

History and Evolution

Lupin was founded in 1968 by Dr. Desh Bandhu Gupta,^[21] then an Associate Professor at BITS-Pilani, Rajasthan. Named after the Lupin flowerbecause of its inherent qualities and what it personifies and stands for, the company was created with a vision to fight life threatening infectious diseases and to manufacture drugs of the highest social priority.

Lupin first gained recognition when it became one of the world’s largest manufacturers of tuberculosis drugs.^[22] The company today has a significant market share in key markets in the Cardiovascular (prils and statins), Diabetology, Asthma, Pediatrics, CNS, GI, Anti-Infectives and NSAIDs therapy segments. It also has a global leadership position in the Anti-TB and Cephalosporin segments. The company’s R&D endeavours have resulted in significant progress in its NCE program. Lupin’s foray into Advanced Drug Delivery Systems has resulted in the development of platform technologies that are being used to develop value-added generic pharmaceuticals. Its manufacturing facilities, spread across India and Japan, have played a critical role in enabling the company realize its global aspirations. Benchmarked to International standards, these facilities are approved by international regulatory agencies including the US FDA, UK MHRA, Japan’s MHLW, TGA Australia, WHO, and the MCC South Africa.

Research and Development

Lupin’s Research Program covers the entire pharma value chain. The company’s global R&D program is headquartered out of the Lupin Research Park located near Pune that houses over 1200 scientists. Lupin’s R&D covers:

Generics Research

Process Research
Pharmaceutical Research

Advanced Drug Delivery Systems (ADDS) Research
Intellectual Property Management
Novel Drug Discovery and Development (NDDD)
Biotechnology Research

Differentiation is the heart of our research efforts at Lupin. We have created a truly unique world-class research program, designed to ensure a sustainable pipeline of high-value opportunities to maximise growth.

Research and Development is at the core and is the most critical part of any pharmaceutical company. At Lupin we see R&D differently. It is fundamentally about creativity, originality and being aware of what is really required. At Lupin, our Research & Development program has been the key to our sustained growth over the past ten years; growth that has made us one of the most exciting research driven pharmaceutical companies globally; a hotbed of differentiation and innovation. Today we are building the future by strengthening our research foundation through prudent investments that position us at the cutting-edge of technology, helping us deliver complex products that very few in the world can.

Headquartered at the state-of-the-art Lupin Research Park in Pune, India, the Company’s research program is home to over 1,400 scientists. The Company’s global research operations are spread over multiple research facilities in India and Japan. During FY 2013, the Company invested 7.5% of its net sales in Research & Development and related spends, amounting to 7,098 million. FY 2013 was a record year in terms of progress made all around, be it our pace of filing DMFs (Drug Master Files) and ANDAs (Abbreviated New Drug Applications), progress in our drug discovery and development program, milestones in our drug delivery program and approvals in our biotechnology program.

NOVEL DRUG DISCOVERY & DEVELOPMENT

Long-term, one of the Company’s biggest differentiators will be its Novel Drug Discovery and Development (NDDD) program. The Program focuses on the discovery, development and commercialisation of new drugs that address disease areas with significantly unmet medical need. Lupin’s NDDD efforts are directed towards identifying and developing new therapies for disease areas that include metabolic/endocrine disorders, pain and inflammation, autoimmune diseases, CNS disorders, cancer and infectious diseases.

Scientists at NDDD have been able to create a portfolio of novel compounds that are moving through a robust pipeline from discovery to development. This steady movement will ensure that at least one compound enters the clinical phase in terms of first-in-human studies each year. Lupin has adopted a ‘Quick-win, fail-fast’ cost-efficient development approach, in which novel compounds are filtered at every stage before entering development and differentiated by efficacy with a focus on enhanced safety.
Highlights, FY 2013

Successfully completed Phase I studies in Europe for a program in the CNS area, which is being advanced to Phase II clinical trials in Europe now

Candidates from two programs in the area of endocrine disorders and cancer will enter clinical development in FY 2014
Six other programs in various stages of discovery across different therapy areas
Strong intellectual property creation and management strategy in place, with a total of over 80 patent applications filed to date

Therapeutic Targets

Therapy Area	Differentiated Pipeline
METABOLIC / ENDOCRINE DISEASES	NOVEL MECHANISMS FOR NEW ANTI-DIABETICS
PAIN / INFLAMMATION	HOLY GRAIL OF PAIN REMEDY
AUTO-IMMUNE DISEASES	TARGETED FOR RHEUMATOID ARTHRITIS AND OTHER IMMUNE DISORDERS
CNS DISORDERS	ROBUST TREATMENTS FOR COGNITIVE DEFICITS IN DIFFERENT CNS CONDITIONS
ONCOLOGY	HIGHLY DIFFERENTIATED ANTI-CANCER TREATMENT (HITTING ONLY CANCER CELLS)
INFECTIOUS DISEASES	HIGHLY POTENT ANTI-VIRAL THERAPY

Businesses

Lupin’s businesses encompass the entire pharmaceutical value chain, ranging from branded and generic formulations, APIs, advanced drug deliverysystems to biotechnology. The company’s drugs reach 70 countries^[23] with a footprint that covers Advanced Markets such as USA, Europe, Japan,^[24] Australia as well as Emerging Markets including India,^[25] the Philippines and South Africa to name a few.

KEY MARKETS AND BUSINESSES

USA
Headquartered in Baltimore, Maryland, Lupin Pharmaceuticals, Inc. (LPI), the company’s US subsidiary is a $ 706 million enterprise.^[26] LPI has a presence in the branded and generics markets of the US. In the branded business, Lupin operates in the CVS and Pediatric segments. The company is the market leader in 24 products out of the 46 products marketed in the US generics market, of which it is amongst the Top 3 by market share in 37 of these products (IMS Health, March 2013): Suprax (Cefixime), a paediatric antibiotic, is Lupin’s top-selling product here. The company is also the 5th largest and fastest growing generics player in the US (5.3% market share by prescriptions, IMS Health). Lupin’s US brands business contributed 21% of total US sales whereas the generics business contributed 79% during FY 2012-13.^[26]

In the US market, since December 2012, Lupin has posted a 40-80 per cent growth rate on the back of new launches, as well as growing sales of its existing drugs. For May, the company has posted growth of 50 per cent year-on-year in US sales. The outlook is also good. On the whole, Lupin has one of the strongest pipelines of 18-20 products for the US market over the next 18 months.

Robust US sales continue
Despite the 51 per cent rise in share price over six months, most analysts continue to be bullish on the company due to its strong showing in the US market. Among leading Indian pharma majors, this geography contributes nearly 40 per cent of its revenues, second only to Sun (43 per cent) and a good performance rubs off well on the company’s overall show.

About half its US sales are contributed by Antara, the generic form of the cholesterol lowering drugTricor, antibiotics Suprax and Cefdinir and the generic form of antipsychotic drug Geodon. Among other segments expected to drive growth are oral contraceptives ($100 million estimated sales in FY14), dermatology, ophthalmology and asthama.

India Region Formulations (IRF)
Lupin’s IRF business focuses on lifestyle and chronic therapy segments. The company has emerged as one of the fastest growing players in therapies like Cardiology, Central Nervous System(CNS), Diabetology, Anti-Asthma, Anti-Infective, Gastro Intestinal and Oncology. The IRF business contributed 25% of the company’s overall revenues for FY 2012-13, growing by 24% and recording revenues of 2364 crore (US$380 million) for FY 2012-13 as compared to 1905 crore (US$300 million) for FY 2011-12.^[26]

There are 9 manufacturing plant and 2 Research pant in India, such as Jammu(J&K),Mandideep & Indore(Madhya pradesh), Ankaleswar & Dabasa (Gujarat), Tarapur, Aurangabad and Nagpur (Maharastra) and Goa; where research centre at Pune and Aurangabad.^[27] Among these the baby plant is Nagpur plant which will the the biggest formulation unit for Lupin in coming year.

Lupin is also gradually expanding its domestic portfolio through expansion into more segments and tie-ups. On Thursday, it announced a non-exclusive tie-up with US-basedMerck Sharp and Dohme (MSD) for marketing the latter’s pneumococcal vaccine (preventive care for diabetes and, chronic heart, lung and liver diseases) for adults.

Given its growth, Bank of America Merrill Lynch analysts believe the valuation multiple at 20 times FY15 earnings estimates is likely to expand (closer to larger peers) due to stronger and sustainable growth rates, both on the net profit front (22 per cent annually over the next two years) and return ratios, expected at 30 per cent versus 26 per cent for the peers. Most analysts have a target price of Rs 875-900 for the stock. Though a re-rating could be on the cards, given the surge in share price, investors should look at corrections to add the scrip to their portfolio.

Europe
Lupin’s focus in the European Union encompasses Anti-Infectives, Cardiovascular, and CNS therapy areas, along with niche opportunities in segments like Oral Contraceptives, Dermatology and Ophthalmics. The company’s presence in France is by way of a trade partnership; in Germany, it operates through its acquired entity Hormosan Pharma GmbH (Hormosan);^[28] while the UK business is a direct-to-market initiative.

Japan
Lupin is the fastest-growing Top 10 generic pharmaceuticals player in Japan (IMS). Lupin operates in Japan through its subsidiary, Kyowa Pharmaceutical Industry Co. Ltd. (Kyowa), a company Lupin acquired in 2007,^[29]^[30] and I’rom, Pharmaceutical Co. Ltd (IP), acquired in 2011.^[31]^[32] Kyowa has an active presence in Neurology, Cardiovascular, Gastroenterology and the Respiratory therapy segments. I’rom is a niche injectables company with significant presence in the DPC hospital segment.

South Africa
Lupin’s South African subsidiary, Pharma Dynamics (PD)^[33] is the fastest growing and the 5th largest generic company in the South African market (IMS). The company is a market leader in the Cardiovascular segment and has a growing presence in Neurology, Gastroenterology and the Over the Counter (OTC) segments.

Australia
Lupin entered the Australian market through its subsidiary, Generic Health Pte. Ltd. (GH).^[34] It subsequently acquired the worldwide marketing rights to the over 100 year old Australian brand Goanna,^[35] used for pain management.

Philippines
Lupin’s Philippines subsidiary Multicare Pharmaceuticals (Multicare),^[36] is a branded generic company focused on Women’s Health, Pediatrics, Gastro-Intestinal and Diabetes care. FY 2012 also marked its foray into the Neurology segment when it entered into a strategic marketing partnership with Sanofi.^[37]

API and Global TB

Lupin is one of the most vertically integrated global generic majors and a global leader in Cephalosporins, Cardiovasculars and the anti-TB space. The company is also a strategic supplier of anti-TB products to the Global Drug Facility (GDF), with its formulations being supplied to more than 50 countries through GDF procurement.

Lupin is also a global leader in anti-TB APIs, and is associated with the Revised National TB Program of the Government of India, thereby partnering the Government in its fight against TB in the country. It also supplies to various Government agencies, the Stop TB Partnership and various other international agencies like Pan America Health Organisation (PAHO), Médecins Sans Frontières (MSF) and the Damien Foundation. Ethambutol, Rifampicin and Pyrazinamide are the company’s top selling TB molecules.

Biotechnology Research
The Lupin Biotechnology Research Group, based out of Wakad, near Pune is focussed on developing affordable, high quality biopharmaceuticals with an emphasis on biosimilars. As of May 2013, it has a pipeline of 10 biosimilar products under development, and is close to getting marketing authorization for 2 of its oncology products for the Indian market. Lupin has competencies for the complete development and manufacture of recombinant protein therapeutic products from high yielding and proprietary microbial and mammalian cell culture platforms. The Biotech R&D infrastructure offers a range of product development capabilities ranging from clone development, process optimization, analytical method development, bioassay, formulation, stability studies, non-clinical and clinical studies backed by a sound understanding of regulatory and IP aspects. The company’s biotech development programs are in compliance of and follow ICH, EMEA and Indian Regulatory guidelines.

Corporate Social Responsibility

To further its social responsibility objectives, Lupin established the Lupin Human Welfare & Research Foundation (LHWRF) on 2 October 1988. Its chief objective was to provide an alternative sustainable, replicable and ever evolving model of holistic rural development. LHWRF started with a few small rural development projects covering around 35 villages in Bharatpur District,Rajasthan. Its efforts have touched the lives of over a million people across 2,200 villages in the states of Rajasthan, Madhya Pradesh, Maharashtra and Uttarakhand.

Lupin gets USFDA nod to market HIV drug in US market

ANKLESHWAR GUJARAT PLANT INDIA

LUPIN TARAPUR INDIA

LUPIN GOA INDIA

Oncologyand inflamation

Lupin is building a discovery pipeline with over seven to eight molecules targeting the oncology and inflammation segments.

Lupin is building a discovery pipeline with over seven to eight molecules targeting the oncology and inflammation segments. The company, producing both branded and generic drugs, is also planning an investment of $20 million to expand and build additional facilities and capacities exclusively for biologics in Pune in next two to three years.

“We have over seven to eight molecules in the pipeline. Of these, three are already undergoing clinical trials, while two are in pre-clinical stage. One more molecule will be entering pre-clinical stage soon,” Cyrus Karkaria, Lupin president said, adding that the company was gearing up to launch a new product this year. He also indicated that some of these lead molecules could be potential out-licensing targets at some point of time.

Biosimilar products include recombinant erythropoietin, recombinant granulocyte colony stimulating factor (G-CSF), interferon alpha and beta, human insulin, monoclonal antibodies and human growth hormone. These are used oncology, infectious diseases, chronic autoimmune diseases, growth-related deficiencies and haematology.

As part of its expansion, Lupin will be expanding facilities in Pune, which will be operational in next two to three years. “We currently have a production facility near the lab. We will be building additional facilities with about $20 million investment in next two to three years,” Cyrus explained. Last year, the company announced plans to invest over R450 crore towards capacity expansion and strengthening sales force. Lupin is also gearing up to launch its first biosimilar product in India by early next year besides targeting 5-7% of its business from biosimilar business.

The company had entered into a licensing agreement with Sydney-based NeuClone for cell-line technology which will provide exclusive proprietary cell-line technology to be developed into biosimilar drugs targeting cancer.

Several drug companies, including Dr Reddy’s, Cipla and Biocon, among others, are eying the opportunity in biosimilars. Industry estimates global market for biosimilars or follow-on biologic drugs is about $100 billion and the Indian market is about R2,500 crore.

Meanwhile, the US Food and Drug Administration has issued three guidances on biosimilar product development to assist industry in developing such products in the US. These draft documents are designed to help industry develop

About Lupin Limited
Lupin is an innovation led transnational pharmaceutical company producing and developing a wide range of branded and generic formulations and APIs globally. The Company is a significant player in the Cardiovascular, Diabetology, Asthma, Pediatric, CNS, GI, Anti-Infective and NSAID space and holds global leadership positions in the Anti-TB and Cephalosporin segment.

Lupin is the 5th largest and fastest growing top 5 generics player in the US (5.3% market share by prescriptions, IMS Health) and the 3rd largest Indian pharmaceutical company by revenues. The Company is also the fastest growing top 10 generic pharmaceutical players in Japan and South Africa (IMS).

For the financial year ended March 2013, Lupin’s Consolidated turnover and Profit after Tax were Rs. 94,616 million (USD 1.74 billion) and Rs. 13,142 million (USD 242 million) respectively. Please visithttp://www.lupinpharmaceuticals.com for more information.

Mumbai, February 03, 2014: Pharmaceutical major Lupin Limited announced the acquisition of Nanomi B.V. in the Netherlands today. With this acquisition, Lupin has made its foray into the technology intensive complex injectables space.

Nanomi has patented technology platforms to develop complex injectable products. Nanomi has a rich talent pool of scientists who would be backed by Lupin’s global R&D and manufacturing teams.

Commenting on the acquisition Ms. Vinita Gupta, Chief Executive Officer, Lupin Limited said “We are very pleased with the acquisition of Nanomi. With the use of Nanomi’s proprietary technology platform, Lupin would be able to make significant in-roads into the niche area of complex injectables.”

Mumbai, Baltimore, December 18, 2013: Pharma Major Lupin Limited (Lupin) announced today that its US subsidiary Lupin Pharmaceuticals Inc. has launched its Abacavir Sulfate, Lamivudine, and Zidovudine Tablets, 300 mg (base) / 150 mg / 300 mg in the US after the US District Court for the District of Delaware ruled that the Lupin’s generic version of Trizivir® did not infringe on patents. Lupin had earlier received approval for the same.

Lupin’s Abacavir Sulfate, Lamivudine Zidovudine 300mg (Base)/150mg/300mg Tablets are the AB-rated generic equivalent of ViiV Healthcare’s (ViiV) Trizivir® Tablets, 300 mg (base) / 150 mg / 300mg and are indicated in combination with other antiretrovirals or alone for the treatment of HIV-1 infection.

Lupin is the first applicant to file an ANDA for Trizivir® Tablets and as such is entitled to 180 days of marketing exclusivity.

Trizivir® Tablets, 300 mg (base) / 150 mg / 300mg had annual U.S sales of approximately US$ 111.6 million (IMS MAT Sep, 2013).

Mumbai, Baltimore, December 12, 2013: Pharma Major Lupin Limited (Lupin) announced today that its US subsidiary, Lupin Pharmaceuticals, Inc. (LPI) has launched its Duloxetine Hydrochloride Delayed-release (HCl DR) Capsules 20 mg, 30 mg and 60 mg strengths. The Company received final approval to market its Duloxetine HCl DR Capsules USP, 20 mg, 30 mg, 40 mg and 60 mg strengths from the United States Food and Drugs Administration (FDA) yesterday.

Lupin’s Duloxetine HCl DR Capsules 20 mg, 30 mg and 60 mg strengths are the generic equivalent of Eli Lilly & Company’s (Lilly) Cymbalta® Delayed-release Capsules 20 mg, 30 mg and 60 mg.

Duloxetine HCl DR Capsules are indicated for the treatment of major depressive disorder (MDD), generalized anxiety disorder (GAD) and management of neuropathic pain (DPNP) associated with diabetic peripheral neuropathy.

Cymbalta® Delayed-Release Capsules 20 mg, 30 mg and 60 mg strengths had annual U.S sales of approximately USD 5.43 billion (IMS MAT Sep, 2013).

Lupin launches Generic Trilipix® Delayed – Release Capsules 45 mg & 135 mg in the US

Mumbai, Baltimore, December 06, 2013: Pharma Major Lupin Limited (Lupin) announced today that its US subsidiary, Lupin Pharmaceuticals, Inc. (LPI) has launched its generic Fenofibric Acid Delayed‐Release Capsules 45 mg and 135 mg. Lupin had earlier received final approval from the US FDA for the same.

Lupin�s Fenofibric Acid Delayed‐Release Capsules 45 mg and 135 mg are the generic equivalent of AbbVie Inc.�s Trilipix® Delayed‐Release Capsules 45 mg & 135 mg strengths are indicated as co‐administration therapy with statins for the treatment of mixed dyslipidemia, treatment of severe hypertriglyceridemia and primary hypercholesterolemia or mixed dyslipidemia.

Trilipix® Delayed‐Release Capsules 45 mg & 135 mg strengths had annual U.S sales of approximately US$ 449.5 million (IMS MAT Sep, 2013).

References

PEMETREXED

Uncategorized Comments Off

Mar 022014

Pemetrexed

US 5,344,932 .

(2S)-2-{[4-[2-(2-amino-4-oxo-1,7-dihydro
pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]amino}
pentanedioic acid

N-[4-[2-(2-amino-4,7-dihydro-4-oxo-3H-pyrrolo[2,3-d]-pyrimidin-5-yl)ethyl] benzoyl]-L-glutamic acid or N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1 H-pyrrolo [2,3-d]-pyrimidin-5-yl)ethyl] benzoyl]-L-glutamic acid

GENERIC LICENSING NEWSLETTER TODAY 23 APRIL 2013 REPORTED, SEE LINK BELOW

http://www.leadformix.com/ef1/preview_campaign.php?lf1=775434470d357512625317e6516156

PEMETREXED

Pemetrexed is a chemotherapy drug used in the treatment of pleural mesothelioma as well as non-small cell lung cancer.Used in combination with cisplatin for the treatment of malignant pleural mesothelioma in adults whose disease is unresectable or who otherwise are not candidates for potentially curative surgery. Also used as a monotherapy for the treatment of locally advanced or metastatic non-small cell lung cancer (NSCLC) after prior chemotherapy.Click here to contact Logenex about this product.

Pemetrexed (brand name Alimta) is a chemotherapy drug manufactured and marketed by Eli Lilly and Company. Its indications are the treatment of pleural mesothelioma andnon-small cell lung cancer.

The molecular structure of pemetrexed was developed by Edward C. Taylor at Princeton University and clinically developed by Indianapolis based drug maker, Eli Lilly and Company in 2004.

PEMETREXED

Pemetrexed is chemically similar to folic acid and is in the class of chemotherapy drugs called folate antimetabolites. It works by inhibiting three enzymes used in purine andpyrimidine synthesis—thymidylate synthase (TS), dihydrofolate reductase (DHFR), andglycinamide ribonucleotide formyltransferase^[1]^[2] (GARFT). By inhibiting the formation of precursor purine and pyrimidine nucleotides, pemetrexed prevents the formation of DNAand RNA, which are required for the growth and survival of both normal cells and cancer cells.

Pemetrexed disodium is chemically described as L-Glutamic acid, N-[4-[2- (2-amino-4,7-dihydro-4-oxo-1 H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]- disodium salt heptahydrate, represented by the chemical structure of Formula (I).

Formula I

Pemetrexed is an anti-folate anti-neoplastic agent that exerts its action by disrupting folate-dependent metabolic processes essential for cell replication. It is believed to work by inhibiting three enzymes that are required in purine and pyrimidine biosynthesis — thymidylate synthase (TS), dihydrofolate reductase (DHFR), and glycinamide ribonucleotide formyl transferase (GARFT). Pemetrexed is available in the market under the brand name ALIMTA^®.

Taylor et al., in describe pemetrexed, its related compounds and pharmaceutically acceptable cation. Chelius et al., in WO 01/14379 A2 disclose pemetrexed disodium crystalline hydrate Form I and process for preparation thereof.

Chelius et al., in WO 01/62760 disclose pemetrexed disodium heptahydrate crystalline Form Il and process for the preparation thereof.

Journal of Organic Process Research & Development, Volume 3, 1999, page 184 describes a process for the preparation of pemetrexed diacid. Busolli et al., in WO200802141 1 disclose process for preparation of pharmaceutically acceptable salt of pemetrexed diacid.

Busolli et al., in WO2008021405A1 disclose seven crystalline forms of pemetrexed diacid designated as Form A, B, C, D, E, F, & G and processes for preparation thereof.

In February 2004, the Food and Drug Administration approved pemetrexed for treatment of malignant Pleural Mesothelioma, a type of tumor of the lining of the lung, in combination with cisplatin^[3] for patients whose disease is either unresectable or who are not otherwise candidates for curative surgery.^[4] In September 2008, the FDA granted approval as a first-line treatment, in combination with cisplatin, against locally-advanced and metastatic non-small cell lung cancer (NSCLC) in patients with non-squamous histology. A Phase III study showed benefits of maintenance use of pemetrexed for non-squamous NSCLC.Activity has been shown in malignant peritoneal mesothelioma.Trials are currently testing it against esophagus and other cancers.

MECHANISM

Pemetrexed is also recommended in combination with carboplatin for the first-line treatment of advanced non-small cell lung cancer.However, the relative efficacy or toxicity of pemetrexed-cisplatin versus pemetrexed-carboplatin has not been established beyond what is generally thought about cisplatin or carboplatin doublet drug therapy

In addition to the brand name Alimta, this drug is also marketed in India by Abbott Healthcare as Pleumet and by Cadila Healthcare asPemecad.

Pemetrexed disodium is a multitargeted antifolate agent approved as a single agent for the treatment of non-small cell lung cancer, and in combination with cisplatin for the treatment of patient with malignant pleural mesothelioma, under the trade name Alimta^®.
Pemetrexed disodium is available in a number of crystalline forms.
Barnett et al, Organic Process Research & Development, 1999, 3, 184-188 discloses synthesis and crystallization of pemetrexed disodium from water-ethanol. The product obtained by the process disclosed herein is the 2.5 hydrate of pemetrexed disodium.
United States patent number 7,138,521 discloses a crystalline heptahydrate form of pemetrexed disodium, which has enhanced stability when compared to the known 2.5 hydrate.
To date workers have concentrated on producing stable crystalline forms of pemetrexed disodium and there has been no disclosure of any non-crystalline form of this active.
We have now found a new form of pemetrexed disodium, which is an amorphous form, as characterized by powder X-ray diffraction. Surprisingly, we have found that it is possible to prepare an amorphous form of pemetrexed disodium and that this form is stable. The amorphous form of the invention is stable contrary to expectations. The amorphous form of pemetrexed disodium of the present invention is stable as it retains it’s amorphous character under a variety of storage conditions. The amorphous form of the present invention is particularly advantageously characterized by a bulk density in the range of 0.15 to 0.35 gm/ml.

“Pemetrexed”)

R = H: Pemetrexed; I

R = Na: Pemetrexed Disodium; II is a known compound. Pemetrexed Disodium is an known anticancer agent. It is clinically active in several solid tumors and approved for treatment of malignant pleural mesothelioma (MPM) and metastatic non-small cell lung cancer (NSCLC). Pemetrexed Disodium is supplied as a sterile lyophilized powder for intravenous administration.

The compound of formula I including pharmaceutically salts thereof as well as a process for its preparation is at first and specifically disclosed in EP patent no. 0432677 B1. The preparation and isolation of Pemetrexed (compound of formula I) as its Disodium salt (compound of formula II) was described for the first time in WO patent no. 9916742 A1 and in Drugs of the future 1998, 23(5), 498-507 as well as by Charles J. Barnett et al. in Organic Process Research & Development, 1999, 3, 184-188 and by Peter Norman in Current Opinion in Investigational Drugs 2001 , 2(11 ), 1611-1622.

Detailed information about the crystalline form of Pemetrexed Disodium prepared according to the process as described above were not provided but it is reported by Charles J. Barnett et al. in Organic Process Research & Development, 1999, 3, 184-188 that the disodium salt II was obtained as a hygroscopic solid.

The first crystalline form of Pemetrexed Disodium has been described in WO patent no. 0114379 designated Disodium MTA Hydrate Form I (MTA = multi- targeted antifolate, disodium N-[4-[2-(2-amino-4,7-dihydro-4-oxo-3H- pyrrolo[2,3-d]-pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid salt). The Disodium MTA Hydrate Form I obtained according to the Examples 2, 3 and 4 contained different amounts of water (Example 2: water = 9.1%; Example 3: water = 17.7%; Example 4: water = 11.7%). The Disodium MTA Hydrate Form I has a typical XRD pattern as shown in Figure 4 (the corresponding 2theta values have been calculated from the provided d-spacing values).

An improved crystalline form of Pemetrexed Disodium has been disclosed in WO patent no. 0162760. It is teached that Pemetrexed Disodium can exist in the form of a heptahydrate (Form II; theoretical amount of water: approx 21%) which is much more stable than the previously known 2.5 hydrate (Form I; theoretical amount of water: 8.7%). The Pemetrexed Disodium Heptahydrate Form (Form II) has a typical XRD pattern as shown in Figure 5 (the corresponding 2theta values have been calculated from the provided d- spacing values).

The Chinese patent no. 1778802 describes a hydrate or trihydrate form of Pemetrexed Disodium. The preparation of Pemetrexed Disodium hydrate or trihydrate includes crystallization from water and water soluble solvent. An overview of the X ray powder diffraction data for Pemetrexed Disodium Hydrate provided in Chinese patent no. 1778802 is shown in Figure 6.

The WO patent no. 2008124485 disclose besides crystalline Forms of the diacid Pemetrexed also amorphous Pemetrexed Disodium as well as a crystalline Form III thereof including a composition containing a major amount of amorphous Form and a minor amount of crystalline Form III of Pemetrexed Disodium. An overview of the X ray powder diffraction data for Pemetrexed Disodium crystalline Form 3 is shown in Figure 7.

EP patent application no. 2072518 disclose a stable amorphous form of Pemetrexed Disodium.

According to the more recent US 5,416,211 , which is incorporated herein by reference, pemetrexed can be synthesized from 4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoic acid of formula 1, obtained from simple precursors, in accordance with the following Scheme 1:
This second method seems to be used also for the industrial preparation of the active ingredient. In fact, the same type of synthesis scheme is also described in C. J. Barnett, T. W. Wilson and M. E. Kobierski, Org. Proc. Res. & Develop., 1999, 3, 184-188, in which the experimental examples refer to a scale of the order of tens of kgs.

……………………….

WO2012134392A1

Example 1 Preparation of crude pemetrexed disodium

[0023] N-[4-2-(2-Amino-4, 7-dihydro-4-oxo-

1 H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid Diethyl Ester

4-Methylbenzenesulfonic Acid Salt and purified process water (PPW) (about 10 kg) are charged to a suitable vessel under nitrogen. The reactor is cooled to NMT 10Ό under nitrogen. Pre-cooled sodium hydroxide solution (about 1.5 kg )/PPW (about 11.4 kg) are added and the temperature is maintained at NMT 10Ό. The mixture is stirred at NMT 0 until the solid is dissolved. Pre-cooled isopropanol (about 62.8 kg) is added and the mixture temperature is maintained at NMT 5 . Pre-cooled 1 N hydrochloric acid in isopropanol is added to adjust the pH to 6.5 to 9.5, preferably between pH 7.5 to pH 8.5, at NMT 5 . The mixture is warmed to a room temperature (i.e., 15-30Ό, preferably 20-25″C) and stirred. The solids are filtered and washed with isopropanol/PPW. The wet cake is vacuum dried to provide crude pemetrexed disodium (about 2.30 kg).

Example 2 Purification of crude pemetrexed disodium to pemetrexed disodium

[0024] Crude pemetrexed disodium (about 2.1 kg) and PPW (about 23.3 kg) are charged under nitrogen to a suitable vessel at 15 to 30 . Isopropanol (about 28.3 kg) is added slowly to cloud point and stirred. Isopropanol (up to about 55 kg) is charged and stirred. The solids are filtered arid washed with isopropanol/PPW. The wet cake is vacuum dried to provide pemetrexed disodium (about 1.9 kg) (90% Yiled). 1 H NMR (D₂0): δ 7.51 (2H, d, J=8.0 Hz), 6.98 (2H, d, J=8.0 Hz), 6.12 (1 H, s), 4.26-4.23 (H, m), 3.60-3.54 (4H, m), 2.27-2.23 (2H, m), 2.13-2.08 (1 H, m), 2.00-1.94 (1 H, m)

HPLC

EP2072518A1

Example 7

HPLC Analysis method

Reagent: Water :milliQ,
Sodium perchlorate :AR Grade
Perchloric acid :AR Grade
Acetonitrile :J.T.Baker gradient
Trifluroacetic acid :AR Grade
Buffer solution: 6.1 g of sodium perchlorate into a 1000ml water. Adjust the pH to 3.0 (± 0.1) with perchloric acid.
Mobile phase A:
mixture of buffer and acetonitrile in the proportion of (90:10).
Mobile phase B:
mixture of buffer and acetonitrile in the proportion of (10 : 90).
Diluent -1 : mixture of water and acetonitrile in the ratio of 50 : 50.
Diluent -2: mixture of water and acetonitrile in the ratio of 90 : 10.
Standard Stock Solution:
Transfer accurately weighed 1.5 mg impurity-E RS and into a 200 ml volumetric flask. Dissolve in and dilute upto mark with diluent-1.

Blank solution

Add 10 ml diluent-2 and 50µl of 3% trifluro acetic acid to a 50 ml volumetric flask, and dilute upto mark with diluent-2.

System suitability solution

Transfer about 25 mg accurately weighed pemetrexed disodium sample in to a 50 ml volumetric flask. First add 10ml of diluent-2 and sonicate to dissolve the contents.Then add 50µl of 3% trifluro acetic acid (prepared in water) and add 5 ml of standard stock solution and dilute up to mark with diluent-2.

Sample preparation

Transfer about 25 mg accurately weighed pemetrexed disodium sample in to a 50 ml volumetric flask. First add 10ml of diluent-2 and sonicate to dissolve the contents.Then add 50µl of 3% trifluro acetic acid (prepared in water) and dilute up to mark with diluent-2 (500 µg/ml).

Chromatographic system :

Use a suitable high pressure liquid chromatography system equipped with Column: 250 mm x 4.6mm containing 5µ packing material (suggested column – Inertsil ODS 3V)
Detector: UV detector set to 240 nm
Cooler temp: 5°C.
Flow rate: about 1.5 ml/min.
The system is also equipped to deliver the two phases in a programmed manner as shown in the following table :

Gradient programme :

[0082]

0 92 8

15 85 15

30 65 35

35 65 35

36 92 8

40 92 8

Procedure:

Inject 20µl of blank and system suitability solution into the chromatograph set to above conditions and record the chromatograms up to 40 min.
Calculate the resolution between pemetrexed disodium and impurity-E. The resolution should not be less than 3.0. Calculate the Number of theoretical plate and tailing factor for pemetrexed peak. Number of theoretical plate is NLT 4000 and tailing factor is NMT 2.0.
Inject 20µl of test solution and calculate the chromatographic purity by area normalisation method.

……………………..

US20120329819

Synthetic Route for the Preparation of Pemetrexed Disodium

Starting from commercially available materials a novel synthetic route for the synthesis of Pemetrexed-IM8 (the dimethyl ester of Pemetrexed) was developed which was then used for the preparation of Pemetrexed Disodium (Scheme 1).

The current synthetic route for the preparation of Pemetrexed IM8 starts with an aldol-condensation reaction of Methyl-4-formylbenzoate (SM1) with 1,1-Dimethoxyacetone (SM2) to give Pemetrexed IM1a. As Pemetrexed IM1a irreversibly converts to its aldol-addition product Pemetrexed IM1b under reaction conditions the reaction mixture is directly submitted to hydrogenation (i.e. without isolation of Pemetrexed IM1a) over Pd/C to give Pemetrexed IM2. As under the hydrogenation conditions not only the double-bond of IM1a is hydrogenated but also some amount of Pemetrexed IM2 is converted to Pemetrexed IM3 (hydrogenation of the carbonyl function to the corresponding secondary alcohol) a solution of NaBH4 is added to the reaction mixture to ensure complete conversion to Pemetrexed IM3. The Pd-catalyst is removed by filtration and the reaction mixture is extracted with toluene. The combined organic layers are evaporated to give crude Pemetrexed IM3 as oil. This oil is dissolved in THF and the alcohol functionality is converted to a mesylate using MsCl and NEt3. The salts are removed by filtration, glacial acetic acid is added and THF is removed by distillation. Upon addition of water Pemetrexed IM4 crystallizes and is isolated by filtration. The dried Pemetrexed IM4 is dissolved in glacial acetic acid and gaseous HCl is added to cleave the dimethoxy acetale and liberate the aldehyde functionality of Pemetrexed IM5. Upon complete deprotection a solution of 2,6-diamino-4-hydroxypyrimidine in aq. NaOH and acetonitrile is added. Upon complete conversion the crystallized Pemetrexed IM6 is isolated by filtration. The saponification of the methyl ester of Pemetrexed IM6 to Pemetrexed IM7 is done using aqueous NaOH. Upon addition of aq. HCl first the Na-salt of Pemetrexed IM7 crystallizes from the reaction mixture. The salt is isolated by filtration, purified by slurry in a mixture of MeOH and water and then converted to Pemetrexed IM7 by pH adjustment in water using aq. HCl. Dried Pemetrexed IM7 (water content not more than 6.0%) is dissolved in DMF, activated using 1,1-carbonyldiimidazolide (CDI) and then reacted with dimethyl-L-glutamate hydrochlorid to give, upon addition of water and filtration, crude Pemetrexed IM8. This intermediate is purified by tosylate salt formation, followed by recrystallization and liberation to give pure Pemetrexed IM8. Starting with the saponification of Pemetrexed IM8 the preparation of different solid forms of Pemetrexed Disodium can be achieved.

Methods For Preparing Pemetrexed Disodium Form IV and Investigation of its Stability

An overview on the possible transformations of Pemetrexed IM8 to Pemetrexed Disodium Form IV is shown in FIG. 20.

Description of Possible Routes for the Preparation of Pemetrexed Disodium Form IV Starting from Pemetrexed IM8

All routes start with saponification of Pemetrexed IM8 in water at IT=20° C. to 30° C. using 3.25 eq of NaOH. Upon complete conversion an aqueous solution of Pemetrexed Disodium with a pH of 13.0 to 13.5 is obtained. Starting from this mixture the desired route can be accessed by addition of HCl to adjust the pH to a certain value (depending on the route, FIG. 20).

Structures of Pemetrexed (Compound I), Pemetrexed Disodium (Compound II) and Pemetrexed Monosodium (Compound IV)

Surprisingly we found that the crucial feature of all successful transformations to Pemetrexed Disodium Form IV is the presence of Pemetrexed Monosodium during the transformation. Routes starting from pure Pemetrexed Disodium Heptahydrate, Pemetrexed Disodium 2.5 hydrate or Pemetrexed Disodium Form A in the presence of seeding crystals of Pemetrexed Disodium Form IV were not successful and resulted in isolation of Pemetrexed Disodium Form A. The same transformations, if carried out in the presence of 0.15 eq of Pemetrexed Monosodium were successful and after addition of 0.15 eq NaOH allowed the isolation of pure Pemetrexed Disodium Form IV. The use of 0.15 eq HCl instead of 0.15 eq Pemetrexed Monosodium under the same conditions resulted in isolation of Pemetrexed Disodium Form A without any Pemetrexed Disodium Form IV. Transformations via isolated Pemetrexed Monosodium gave complete transformation to Pemetrexed Disodium Form IV if either 1.0 eq NaOH were added slowly (over a period of several hours) to Pemetrexed Monosodium or if initially only 0.85 eq of NaOH (based on Pemetrexed Monosodium) were added, followed by 0.15 eq once the transformation to Pemetrexed Disodium Form IV was complete. Very fast addition (<1 min) of 1.0 eq NaOH resulted in formation of Pemetrexed Disodium Form A containing traces of Pemetrexed Disodium Heptahydrate.

Starting from Pemetrexed (compound I) the transformation to Pemetrexed Disodium Form IV would be possible if initially 1.85 eq of NaOH were added followed by 0.15 eq once the transformation was complete. Alternatively, 2.0 eq of NaOH could be added over a long period of time (i.e several hours) to achieve formation of Pemetrexed Form IV. Fast addition (<1 min) of 2.0 eq of NaOH is assumed to result in formation of Pemetrexed Disodium Form A. All these experiments show the presence of Pemetrexed Monosodium to be crucial during the transformations. This presence can be achieved by either addition of catalytic amounts of Pemetrexed Monosodium to Pemetrexed Disodium, by slow addition over several hours of NaOH to Pemetrexed Monosodium or by portionwise addition of NaOH to Pemetrexed Monosodium. Addition of catalytic amounts of HCl to Pemetrexed Disodium (in situ preparation of Pemetrexed Monosodium) failed to give Pemetrexed Disodium Form IV.

Fast addition of NaOH to Pemetrexed Monosodium results in fast formation of Pemetrexed Disodium, thereby lacking the necessary catalytic amounts of Pemetrexed Monosodium to catalyze the transformation to Pemetrexed Disodium Form IV. EtOH as solvent and water content of EtOH were found to be crucial parameters for the transformation to Pemetrexed Disodium Form IV. So far the transformation has only been observed in EtOH containing 0-4% water (v/v). A water content>4% (v/v) results in formation of Pemetrexed Disodium Heptahydrate. Under the conditions used (EtOH containing 0-4% water (v/v)) both Pemetrexed Disodium Heptahydrate and Pemetrexed Disodium 2.5 hydrate are transformed to Pemetrexed Form A. Therefore the mechanism of the transformation to Pemetrexed Disodium Form IV is assumed to proceed via Pemetrexed Disodium Form A with Pemetrexed Monosodium acting as catalyst for the transformation.

Preparation of Pemetrexed Disodium Heptahydrate

a) Preparation of Pemetrexed Disodium Heptahydrate Starting from Pemetrexed IM8

Pemetrexed Disodium Heptahydrate was prepared by adjustment of the pH of the Pemetrexed Disodium solution after saponification from pH=13 to pH=8 using HCl followed by addition of EtOH (3 times the volume of water) to achieve crystallization. Precipitated Pemetrexed Disodium Heptahydrate was isolated by filtration, washed with a mixture of EtOH and water (4:1 v/v) followed by EtOH. The wet product was dried in vacuo at 200 mbar at 20° C. to 30° C. until water content of the dried product was 20.1% to 22.1%.

b) Conversion of Pemetrexed Disodium Form A to Pemetrexed Disodium Heptahydrate

To a suspension of Pemetrexed Disodium Form A in EtOH was added water until a mixture of EtOH containing 25% water (v/v) was obtained. The resulting suspension was stirred at 20° C. to 30° C. until conversion was complete according to PXRD. Pemetrexed Disodium Heptahydrate was isolated by filtration, washed with EtOH and dried in vacuo at 200 mbar at 20° C. to 30° C. until water content of the dried product was 20.1% to 22.1%.

………………………..

EP2504341A1

REFERENCES

McLeod, Howard L.; James Cassidy, Robert H. Powrie, David G. Priest, Mark A. Zorbas, Timothy W. Synold, Stephen Shibata, Darcy Spicer, Donald Bissett, Yazdi K. Pithavala, Mary A. Collier, Linda J. Paradiso, John D. Roberts (Jul-2000).“Pharmacokinetic and Pharmacodynamic Evaluation of the Glycinamide Ribonucleotide Formyltransferase Inhibitor AG2034”. Clinical Cancer Research 6 (7): 2677–84.PMID 10914709. More than one of |work= and |journal=specified (help)
Avendano, Carmen; Menendez, J. Carlos (April 2008).Medicinal Chemistry of Anticancer Drugs. Amsterdam:Elsevier. p. 37. ISBN 0-444-52824-5.
Manegold C (August 2003). “Pemetrexed (Alimta, MTA, multitargeted antifolate, LY231514) for malignant pleural mesothelioma”. Semin. Oncol. 30 (4 Suppl 10): 32–6.doi:10.1016/S0093-7754(03)00283-5. PMID 12917819.
National Cancer Institute: FDA Approval for Pemetrexed Disodium

US6090168 *	Oct 6, 1999	Jul 18, 2000	Eli Lilly And Company	Processes and intermediates useful to make antifolates
US7138521	Feb 12, 2001	Nov 21, 2006	Eli Lilly And Company	Crystalline of N-[4-[2-(2-Amino-4,7-dihydro-4oxo-3H-pyrrolo[2,3-D]pyrimidin-5-YL)ethyl]benzoyl]-L-glutamic acid and process therefor
US20030216416 *	Feb 12, 2001	Nov 20, 2003	Chelius Erik Christopher	Novel crystalline of n-[4-[2-(2-amino-4,7-dihydro-4oxo-3h-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-l-glutamic acid and process therefor
WO2001014379A2 *	Aug 15, 2000	Mar 1, 2001	Erik Christopher Chelius	A novel crystalline form of disodium n-[4-[2-(2-amino-4,7-dihydro-4-oxo-3h-pyrrolo[2,3-d]-pyrimidin-5-yl)ethyl]benzoyl]-l-glutamic acid salt and processes therefor
WO2008021405A1 *	Aug 14, 2007	Feb 21, 2008	Sicor Inc	Crystalline forms of pemetrexed diacid and processes for the preparation thereof
WO2008124485A2 *	Apr 3, 2008	Oct 16, 2008	Reddys Lab Ltd Dr	Solid forms of pemetrexed

IBRUTINIB 依鲁替尼 A Btk protein inhibitor.

Uncategorized Comments Off

Feb 272014

IBRUTINIB 依鲁替尼

A Btk protein inhibitor.

1-[(3R)-3-[4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one

1-((R)-3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one

CAS number	936563-96-1

Ibrutinib, PCI 32765, PCI32765, ibrutinibum, IMBRUVICA,

CRA-032765
Ibrutinib
Imbruvica
Pc-32765
PCI 32765
PCI32765
UNII-1X70OSD4VX

Molecular Formula: C₂₅H₂₄N₆O₂

Molecular Weight: 440.49706

Company: Pharmacyclics
Approval Status: Approved February 2014US FDA:link
Treatment Area: chronic lymphocytic leukemia

Bruton’s tyrosine kinase (Btk) inhibitor

U.S. Patent No: 7,514,444 , 7,718,662
patent validity: December 2026

An orally bioavailable small-molecule inhibitor of Bruton’s tyrosine kinase (BTK) with potential antineoplastic activity. Ibrutinib binds to and inhibits BTK activity, preventing B-cell activation and B-cell-mediated signaling and inhibiting the growth of malignant B cells that overexpress BTK. BTK, a member of the src-related BTK/Tec family of cytoplasmic tyrosine kinases, is required for B cell receptor (BCR) signaling, plays a key role in B-cell maturation, and is overexpressed in a number of B-cell malignancies.

Imbruvica (ibrutinib) is an orally available, selective inhibitor of Bruton’s tyrosine kinase (Btk), a gene that is disrupted in the human disease X-linked agammaglobulenemia (XLA). BTK is a signaling molecule of the B-cell antigen receptor (BCR) and cytokine receptor pathways.

Imbruvica is specifically approved for chronic lymphocytic leukemia in patients who have received at least one prior therapy.

Imbruvica (Ibrutinib, previously known as PCI-32765) was approved as a “breakthrough therapy” on November 13, 2013 by the US Food and Drug Administration (FDA) for the treatment of mantle cell lymphoma (MCL), a rare and deadly form of blood cancer.

IBRUTINIB

Ibrutinib, a first in class oral Bruton’s tyrosine kinase (Btk) inhibitor, was launched in the U.S. for the treatment of patients with mantle cell lymphoma in 2013, and for the treatment of chronic lymphocytic leukemia in 2014. In the E.U., the product candidate is awaiting registration for both indications. Additional phase III clinical trials are ongoing for the treatment of these indications in combination with bendamustine and rituximab and for the treatment of relapsed or refractory marginal zone lymphoma (MZL). Janssen and Pharmacyclics are conducting phase II clinical trials for the treatment of refractory follicular lymphoma. Early clinical development is also under way at Pharmacyclics for the treatment of recurrent B-cell lymphoma, relapsed/refractory MCL, and relapsed or relapsed and refractory multiple myeloma. The company filed an IND seeking approval to commence clinical evaluation of ibrutinib for the treatment of autoimmune disease. Preclinical studies had been under way for rheumatoid arthritis; however, no recent development has been reported. Ibrutinib is also active against Lyn and LCK tyrosine kinases.

In 2011, a codevelopment agreement was signed between the National Cancer Institute (NCI) and Pharmacyclics for the treatment of hematologic/blood cancer. Also in 2011, a worldwide codevelopment and comarketing agreement was signed by Janssen and Pharmacyclics for the treatment of cancer. In 2012, orphan drug designation was assigned in the U.S. and the E.U. for the treatment of CLL. This designation was also assigned by the FDA in 2012 for the treatment of mantle cell lymphoma. In 2013, several orphan drug designations were assigned in the U.S.; for the treatment of small lymphocytic lymphoma, for the treatment of Waldenstrom’s macroglobulinemia and for the treatment of diffuse large B-cell lymphoma. For this indication, orphan drug designation was assigned also in the E.U. the same year. In 2012, fast track designation was assigned by the FDA for the treatment of CLL. In 2013, breakthrough therapy designations were assigned to the compound in the U.S.: for the treatment (as monotherapy) of patients with chronic lymphocytic leukemia or small lymphocytic lymphoma, for the treatment of relapsed or refractory mantle cell lymphoma who have received prior therapy and for the treatment of Waldenstrom’s macroglobulinemia.

Imbruvica is supplied as a capsule for oral administration. The recommended dose is 420 mg taken orally once daily (three 140 mg capsules once daily). Capsules should be taken orally with a glass of water. Do not open, break, or chew the capsules.

The FDA approval of Imbruvica for chronic lymphocytic leukemia was based on an open-label, multi-center trial of 48 previously treated patients. Imbruvica was administered orally at 420 mg once daily until disease progression or unacceptable toxicity. The overall response rate (ORR) and duration of response (DOR) were assessed using a modified version of the International Workshop on CLL Criteria by an Independent Review Committee. The ORR was 58.3%, all partial responses. None of the patients achieved a complete response. The DOR ranged from 5.6 to 24.2+ months. The median DOR was not reached.

Imbruvica (ibrutinib) is an orally available, selective inhibitor of Bruton’s tyrosine kinase (Btk). Ibrutinib forms a covalent bond with a cysteine residue in the BTK active site, leading to inhibition of BTK enzymatic activity. BTK is a signaling molecule of the B-cell antigen receptor (BCR) and cytokine receptor pathways. BTK’s crole in signaling through the B-cell surface receptors results in activation of pathways necessary for B-cell trafficking, chemotaxis, and adhesion.

Ibrutinib (USAN,^[1] also known as PCI-32765 and marketed in the U.S. under the name Imbruvica) is an anticancer drug targeting B-cell malignancies. It was approved by the US FDA in November 2013 for the treatment of mantle cell lymphoma^[2] and in February 2014 for the treatment ofchronic lymphocytic leukemia.^[3] It is an orally-administered, selective and covalent inhibitor of the enzyme Bruton’s tyrosine kinase (BTK).^[4]^[5]^[6]Ibrutinib is currently under development by Pharmacyclics, Inc and Johnson & Johnson‘s Janssen Pharmaceutical division for additional B-cell malignancies including diffuse large B-cell lymphoma and multiple myeloma.^[7]^[8]^[9]

Mechanism

In preclinical studies on chronic lymphocytic leukemia (CLL) cells, ibrutinib has been reported to promote apoptosis, inhibit proliferation, and also prevent CLL cells from responding to survival stimuli provided by the microenvironment.^[12] In this study, treatment of activated CLL cells with ibrutinib resulted in inhibition of Btk tyrosine phosphorylation and also effectively abrogated downstream survival pathways activated by this kinase including ERK1/2, PI3K, and NF-κB. Additionally, ibrutinib inhibited proliferation of CLL cells in vitro, effectively blocking survival signals provided externally to CLL cells from the microenvironment including soluble factors (CD40L, BAFF, IL-6, IL-4, and TNF-α), fibronectin engagement and stromal cell contact.

In early clinical studies, the activity of ibrutinib has been described to include a rapid reduction in lymphadenopathy accompanied by a transient lymphocytosis, suggesting that the drug might have direct effects on cell homing or migration to factors in tissue microenvironments.^[13]

Ibrutinib has been reported to reduce CLL cell chemotaxis towards the chemokines CXCL12 and CXCL13, and inhibit cellular adhesion following stimulation at the B cell receptor.^[14]^[15] Together, these data are consistent with a mechanistic model whereby ibrutinib blocks BCR signaling, which drives cells into apoptosis and/or disrupts cell migration and adherence to protective tumour microenvironments.

History

Ibrutinib was first designed and synthesized at Celera Genomics which reported in 2007 a structure-based approach for creating a series of small molecules that inactivate BTK through covalent binding to cysteine-481 near the ATP binding domain of BTK.^[4] These small molecules irreversibly inhibited BTK by using a Michael acceptor for binding to the target cysteine. In April 2006, Pharmacyclics acquired Celera’s small molecule BTK inhibitor discovery program, which included a compound, PCI-32765 that was subsequently chosen for further preclinical development based on the discovery of anti-lymphoma properties in vivo.^[16] Since 2006, Pharmacyclics’ scientists have advanced the molecule into clinical trials and identified specific clinical indications for the drug. It also has potential effects against autoimmune arthritis.^[17] It was approved by the US FDA on November 13, 2013 for the treatment of mantle cell lymphoma.^[2] On Feb. 12, 2014, the U.S. Food and Drug Administration expanded the approved use of the drug ibrutinib to chronic lymphocytic leukemia (CLL). ^[18]

Ibrutinib is an inhibitor of Bruton’s tyrosine kinase (BTK). It is a white to off-white solid with the empirical formula C₂₅H₂₄N₆O₂ and a molecular weight 440.50. Ibrutinib is freely soluble in dimethyl sulfoxide, soluble in methanol and practically insoluble in water.

The chemical name for ibrutinib is 1-[(3R)-3-[4-amino-3-(4-phenoxyphenyl)-1Hpyrazolo[ 3,4-d]pyrimidin-1-yl]-1-piperidinyl]-2-propen-1-one and has the following structure:

IMBRUVICATM (ibrutinib) Structural Formula Illustration

IMBRUVICA (ibrutinib) capsules for oral administration are supplied as white opaque capsules that contain 140 mg ibrutinib as the active ingredient. Each capsule also contains the following inactive ingredients: croscarmellose sodium, magnesium stearate, microcrystalline cellulose, sodium lauryl sulfate. The capsule shell contains gelatin, titanium dioxide and black ink. Each white opaque capsule is marked with “ibr 140 mg” in black ink.

PCI-32765 (ibrutinib) is disclose d in U.S. Patent No. 7,514,444, issued on April 7, 2009, and has the following structur

Ibrutinib is an orally available drug that targets Bruton’s tyrosine kinase (BTK).

Ibrutinib is an irreversible small molecule BTK inhibitor that is in Ph Ib/II of clinical trials in a variety of B-cell malignancies including chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), mantle cell lymphoma (MCL), diffuse large B-cell lymphoma (DLBCL) and multiple myeloma (cancer of plasma cells, a type of white blood cell present in bone marrow). At present ibrutinib is administered orally in clinical trials, via the gastrointestinal tract, at high clinical doses (420 mg/day or 840 mg/day) to patients with CLL and SLL to obtain the desired thereapeutic effect. The need for such high doses of ibrutinib may be due to low bioavailability (the oral bioavailability of ibrutinib is reported to be 22.8% in rats) and may be responsible for the adverse side effects associated with the use of ibrutinib such as nausea or emesis, dizziness and diarrhea. Moreover, low bioavailability results in more variable absorption and potential variability of the desired therapeutic response.

As stated above, at present ibrutinib is administered orally, via the gastrointestinal tract, at high clinical doses (420 mg/day or 840 mg/day) to patients to obtain the desired clinical benefit. It is presently disclosed that when ibrutinib is administered intraduodenally versus via the gastrointestinal tract in rats, the oral bioavailability of ibrutinib unexpectedly increased from 21 % to 100% as determined by AUC.

This unexpected increase in oral bioavailability of ibrutinib can translate into a number of desirable practical benefits. The increase in oral bioavailability should enable administration of ibrutinib at a significantly lower therapeutically effective dose than is currently being used. The lower variability associated with this greater bioavailability should lead to a more reliable therapeutic response as well as more predictable drug absorption.

And avoidance of exposure of Ibtrutinib to the stomach and/or use of lower therapeutically effective dose of ibrutinib can reduce or altogether eliminate potential adverse side effects of this drug such as diahrrea, nausea or emesis, and dizziness. U.S. Patent No. 7,514,444, mentioned above, discloses administration of 0.02-5000 mg/kg andl-1500 mg of ibrutinib/per day and in clinical trials 420 or 840 mg/day of ibrutinib is being administered to the patients with CLL and SLL.

There is no reasonable expectation in the art that ibrutinib can be adminstered orally at lower efficacious doses to the patients with CLL and SLL, particularly as evidenced by the 420 or 840 mg/day of ibrutinib being administered in clinical trials to those patients. Moreover, other than for active agents that are unstable in the stomach or at acidic pH delivery of any active agent with low bioavailability further along in the gastrointestinal tract reduces the path length for drug absorption and would be expected to reduce bioavailability. Therefore, it was unexpected to achieve delivery of ibruntinib directly to the small intestine with greater bioavailability.

PC1-32765 (Ibrutinib), chemical name: 1_ [(3R) _3-[4_-3 – (4 – phenoxy-phenyl)-1H-pyrazolo [3,4-d] pyrimidine – 1 – yl] – 1-piperidinyl]-2 – propen-1 – one, and its structural formula is as follows:

PC1-32765 is an oral medication that inhibits B cell as the main receptor tyrosine kinase signaling and promote cell death process, preventing cell migration and adhesion in malignant B cells.

US20080108636 basic patent has been disclosed a synthetic route:

This synthetic route with 4 – phenoxy-benzoic acid as raw material, after eight-step reaction the final product, the following reaction steps:

The above method has the following disadvantages:

1, eight single-step reaction, long route, the economy is bad; i1, to use synthetic intermediates 4:00 trimethylsilyl diazomethane (TMSCHN2), this material easy to blow up, the risk coefficient is large, so large-scale production greatly reduces the possibility;

ii1, synthetic intermediates 7:00, set out to use polymer-supported triphenylphosphine, non-industrial raw materials used, the price is expensive, the cost of smell;

iv, the final step of acylation, the selectivity is poor, a large amount of negative product, purification is difficult, amplification reaction is difficult.

In summary, the route material is not common, expensive step, high costs, the reaction dangerous side reactions, purification difficult, limiting the possibility of industrial production of the route.

………………………

WO2013184572A1

Polymorphs

EXAMPLES

[00438] The following ingredients, formulations, processes and procedures for practicing the methods disclosed herein correspond to that described above. Example 1; Preparation of Crystalline Forms of l-((R)-3-(4-amino-3-(4-phenoxyphenyl)- lH-pyrazolo[3,4-dlpyrimidin-l-yl)piperidin-l-yl)prop-2-en-l-one (Compound 1)

Form A – Route 1:

[00439] Amorphous Compound 1 (ca. 15 mg) was measured into a vial. Ten volumes (150 μΐ) of solvent [methyl tert-butyl ether (MTBE), diisopropyl ether (DIPE), ethyl acetate, isopropyl acetate, isopropyl alcohol, methyl isobutyl ketone (MIBK), methyl ethyl ketone (MEK), acetone, methanol, nitromethane, 10% aqueous acetone, or 10% aqueous isopropyl alcohol] were added to the vial. The vial was sealed and placed in a shaker at 50 °C for one hour. If a slurry was obtained, an additional thirty volumes (total of 600 μΐ) of solvent was added, then the slurry was returned to 50 °C for another hour. If the sample remained as a slurry at this point, no further solvent was added. The solution/slurry was stirred at 50 °C for one hour, then cooled to 0 °C at 0.1 °C/min, then held at 0 °C overnight. If a slurry was obtained, the solids were filtered under vacuum to provide Compound 1 , Form A; the solution was returned to ambient temperature for slow evaporation through a pin-hole to furnish Compound 1, Form A.

“Compound 1” or “l-((R)-3-(4-amino-3-(4-phenoxyphenyl)-lH-pyrazolo[3,4- d]pyrimidin- 1 -yl)piperidin- 1 -yl)prop-2-en- 1 -one” or “1 – {(3i?)-3-[4-amino-3-(4-phenoxyphenyl)- lH-pyrazolo[3,4-JJpyrimidin-l-yl]piperidin-l-yl}prop-2-en-l-one” or “2-Propen- 1 -one, 1- [(3R)-3-[4-amino-3-(4-phenoxyphenyl)- lH-pyrazolo[3,4-<f]pyrimidin- 1 -yl] – 1 -piperidinyl-” or ibrutinib or any other suitable name refers to the compound with the following structure:

………….

Synthesis

US20080214501

Synthesis of Compound 3—Btk Activity Probe

4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (Intermediate 2) is prepared. Briefly, 4-phenoxybenzoic acid (48 g) is added to thionyl chloride (100 mL) and heated under gentle reflux for 1 hour. Thionyl chloride was removed by distillation, the residual oil was dissolved in toluene and volatile material removed at 80° C./20 mbar. The resulting acid chloride was dissolved in toluene (200 mL) and tetrahydrofuran (35 mL). Malononitrile (14.8 g) was added and the solution and stirred at −10° C. while adding diisopropylethylethylamine (57.9 g) in toluene (150 mL), while maintaining the temperature below 0° C. After 1 hour at 0° C., the mixture was stirred at 20° C. overnight. Amine hydrochloride is removed by filtration and the filtrate evaporated in vacuo. The residue was taken up in ethyl acetate and washed with 1.25 M sulphuric acid, then with brine and dried over sodium sulfate. Evaporation of the solvents gave a semisolid residue which was treated with a portion of ethyl acetate to give 4.1 g of 1,1-dicyano-2-hydroxy-2-(4-phenoxyphenyl)ethene as a white solid (m.p. 160-162° C.). The filtrate on evaporation gave 56.58 (96%) of 1,1-dicyano-2-hydroxy-2-(4-phenoxyphenyl)ethene as a grey-brown solid, which was sufficiently pure for further use.

1,1-Dicyano-2-hydroxy-2-(4-phenoxyphenyl)ethene (56.5 g) in acetonitrile (780 mL) and methanol (85 mL) is stirred under nitrogen at 0° C. while adding diisopropylethylamine (52.5 mL) followed by 2M trimethylsilyldiazomethane (150 mL) in THF. The reaction is stirred for 2 days at 20° C., and then 2 g of silica is added (for chromatography). The brown-red solution is evaporated in vacuo, the residue dissolved in ethyl acetate and washed well with water then brine, dried and evaporated. The residue is extracted with diethyl ether (3×250 mL), decanting from insoluble oil. Evaporation of the ether extracts gives 22.5 g of 1,1-dicyano-2-methoxy-2-(4-phenoxyphenyl)ethene as a pale orange solid. The insoluble oil is purified by flash chromatography to give 15.0 g of a red-orange oil.

1,1-Dicyano-2-methoxy-2-(4-phenoxyphenyl)ethene (22.5 g) and 1,1-dicyano-2-methoxy-2-(4-phenoxyphenyl)ethene oil (15 g) are treated with a solution of hydrazine hydrate (18 mL) in ethanol (25 mL) and heated on the steambath for 1 hour. Ethanol (15 mL) is added followed by water (10 mL). The precipitated solid is collected and washed with ethanol:water (4:1) and then dried in air to give 3-amino-4-cyano-5-(4-phenoxyphenyl)pyrazole as a pale orange solid.

3-Amino-4-cyano-5-(4-phenoxyphenyl)pyrazole (29.5 g) is suspended in formamide (300 mL) and heated under nitrogen at 180° C. for 4 hours. The reaction mixture is cooled to 30° C. and water (300 mL) is added. The solid is collected, washed well with water, then with methanol and dried in air to give of 4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (Intermediate 2).

Synthesis of 1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl (Intermediate 4); a) triphenylphosphine (TPP), diisopropyl diazodicarboxylate (DIAD), tetrahydrofuran (THF); b) TFA/CH₂Cl₂.

To a solution of 1-boc-3-(S)-hydroxypiperidine (3.98 g, 19.8 mmol) and triphenylphosphine (5.19 g, 19.8 mmol) in THF (150 ml) was added DIAD (3.9 ml, 19.8 mmol). The yellow solution was stirred 1 minute then Intermediate 2 (4.0 g, 13.2 mmol) was added and the reaction was heated with a heat gun (3-5 minutes) until the solid had dissolved. After stirring for 1 hour at room temperature, the solvent was removed and the resulting brown oil was subjected to flash chromatography (30% then 50% THF/hexanes) to provide 4.45 g (69%) of Intermediate 3 (trace of triphenylphosphine oxide is present) as a light brown foam.

To a solution of Intermediate 3 (4.4 g, 9.0 mmol) in CH₂Cl₂(20 ml) was added TFA (2.8 ml, 36.2 mmol). After stirring 2 hrs at room temperature, the solvent was removed and the residue was partitioned between ethyl acetate (250 ml) and dilute aq. K₂CO₃. The organic layer was dried (MgSO₄), filtered and concentrated to 70 ml. The resulting solution was stirred and 4.0M HCl in dioxane (4 ml) was added to provide a thick light orange precipitate. The precipitate was collected by filtration and washed with ethyl acetate (50 ml). The material was then partitioned between ethyl acetate (300 ml) and dilute aq. K₂CO₃. The organic layer was dried (MgSO₄), filtered and concentrated to provide 2.78 g (80%) of Intermediate 4 as a light yellow foam.

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SYNTHESIS

US7514444

Compounds described herein may be prepared using the synthetic methods described herein as a single isomer or a mixture of isomers.

A non-limiting example of a synthetic approach towards the preparation of compounds of any of Formula (A), (B), (C) or (D) is shown in Scheme I.

Halogenation of commercially available 1H-pyrazolo[3,4-d]pyrimidin-4-amine provides an entry into the synthesis of compounds of Formula (A), (B), (C) and/or (D). In one embodiment, 1H-pyrazolo[3,4-d]pyrimidin-4-amine is treated with N-iodosuccinamide to give 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine. Metal catalyzed cross coupling reactions are then carried out on 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine. In one embodiment, palladium mediated cross-coupling of a suitably substituted phenyl boronic acid under basic conditions constructs intermediate 2. Intermediate 2 is coupled with N-Boc-3-hydroxypiperidine (as non-limiting example) via Mitsunobu reaction to give the Boc (tert-butyloxycarbonyl) protected intermediate 3. After deprotection with acid, coupling with, but not limited to, an acid chloride, such as, but not limited to, acryloyl chloride, completes the synthesis to give compound 4.

Example 1 Synthesis of Compounds Preparation of 4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (Intermediate 2)

4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (Intermediate 2) is prepared as disclosed in International Patent Publication No. WO 01/019829. Briefly, 4-phenoxybenzoic acid (48 g) is added to thionyl chloride (100 mL) and heated under gentle reflux for 1 hour. Thionyl chloride is removed by distillation, the residual oil dissolved in toluene and volatile material removed at 80° C./20 mbar. The resulting acid chloride is dissolved in toluene (200 mL) and tetrahydrofuran (35 mL). Malononitrile (14.8 g) is added and the solution and stirred at −10° C. while adding diisopropylethylethylamine (57.9 g) in toluene (150 mL), while maintaining the temperature below 0° C. After 1 hour at 0° C., the mixture is stirred at 20° C. overnight. Amine hydrochloride is removed by filtration and the filtrate evaporated in vacuo. The residue is taken up in ethyl acetate and washed with 1.25 M sulphuric acid, then with brine and dried over sodium sulfate. Evaporation of the solvents gives a semisolid residue which is treated with a little ethyl acetate to give 4.1 g of 1,1-dicyano-2-hydroxy-2-(4-phenoxyphenyl)ethene as a white solid (m.p. 160-162° C.). The filtrate on evaporation gives 56.58 (96%) of 1,1-dicyano-2-hydroxy-2-(4-phenoxyphenyl)ethene as a grey-brown solid, which is sufficiently pure for further use.

Example 1a Synthesis of 1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one (Compound 4)

- Synthesis of compound 4; a) polymer-bound triphenylphosphine (TPP), diisopropyl diazodicarboxylate (DIAD), tetrahydrofuran (THF); b) HCl/dioxane; then acryloyl chloride, triethylamine (TEA).

Compounds described herein were synthesized by following the steps outlined in Scheme 1. A detailed illustrative example of the reaction conditions shown in Scheme 1 is described for the synthesis of 1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one (Compound 4).

101 mg of 4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine and 330 mg of polymer-bound triphenylphosphine(TPP) (polymerlab) were mixed together with 5 mL of tetrahydrofuran (THF). tert-Butyl 3-hydroxypiperidine-1-carboxylate (200 mg; 2.0 equivalents) was added to the mixture followed by the addition of diisopropyl diazodicarboxylate (0.099 mL). The reaction mixture was stirred at room temperature overnight. The reaction mixture was filtered to remove the resins and the reaction mixture was concentrated and purified by flash chromatography (pentane/ethyl acetate=1/1) to give intermediate 3 (55 mg).

Intermediate 3 (48.3 mg) was treated with 1 mL of 4N HCl in dioxane for 1 hour and then concentrated to dryness. The residue was dissolved in dichloromethane and triethylamine (0.042 mL) was added followed by acryl chloride (0.010 mL). The reaction was stopped after 2 hours. The reaction mixture washed with 5% by weight aqueous citric acid and then with brine. The organic layer was dried with MgSO₄, and concentrated. Flash chromatography (with CH₂Cl₂/MeOH=25/1) gave 22 mg of compound 4 as a white solid. MS (M+1): 441.2; ¹H-NMR (400 MHz): 8.26, s, 1H, 7.65, m, 2H, 7.42, m, 2H, 7.1-7.2, m, 5H, 6.7-6.9, m, 1H, 6.1, m, 1H, 5.5-5.7, m, 1H, 4.7, m, 1H, 4.54, m, 0.5H, 4.2, m, 1H, 4.1, m, 0.5H, 3.7, m, 0.5H, 3.2, 1,1H, 3.0, m, 0.5H, 2.3, m, 1H, 2.1, m, 1H, 1.9, m, 1H, 1.6, m, 1H.

Example 1b Synthesis of 1-((R)-3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one (Compound 13)

The synthesis of compound 13 was accomplished using a procedure analogous to that described in Example 1a. EM (calc.): 440.2; MS (ESI) m/e (M+1H)⁺: 441.1, (M−1H)⁻: 439.2.

Example 1c Synthesis of 1-((S)-3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one (Compound 14)

The synthesis of compound 14 was accomplished using a procedure analogous to that described for Example 1a. EM (calc.): 440.2; MS (ESI) m/e (M+1H)+: 441.5, (M−1H)−: 439.2.

……………….

US7718662

Synthesis of Compounds Example 1 Preparation of 4-Amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (2a)

Example 1a Synthesis of 1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one (4)

Synthesis of compound 4; a) polymer-bound triphenylphosphine (TPP), diisopropyl diazodicarboxylate (DIAD), tetrahydrofuran (THF); b) HCl/dioxane; then acryloyl chloride, triethylamine (TEA)

Compounds described herein were synthesized by following the steps outlined in Scheme III. A detailed illustrative example of the reaction conditions shown in Scheme II is described for the synthesis of 1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one (Compound 4).

101 mg of 4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine and 330 mg of polymer-bound triphenylphosphine (TPP) (polymerlab) were mixed together with 5 mL of tetrahydrofuran (THF). tert-Butyl 3-hydroxypiperidine-1-carboxylate (200 mg; 2.0 equivalents) was added to the mixture followed by the addition of diisopropyl diazodicarboxylate (0.099 mL). The reaction mixture was stirred at room temperature overnight. The reaction mixture was filtered to remove the resins and the reaction mixture was concentrated and purified by flash chromatography (pentane/ethyl acetate=1/1) to give intermediate 3a (55 mg).

Intermediate 3a (48.3 mg) was treated with 1 mL of 4N HCl in dioxane for 1 hour and then concentrated to dryness. The residue was dissolved in dichloromethane and triethylamine (0.042 mL) was added followed by acryl chloride (0.010 mL). The reaction was stopped after 2 hours. The reaction mixture was washed with 5% by weight aqueous citric acid and then with brine. The organic layer was dried with MgSO₄, and concentrated. Flash chromatography (with CH₂Cl₂/MeOH=25/1) gave 22 mg of compound 4 as a white solid. MS (M+1): 441.2; ¹H-NMR (400 MHz): 8.26, s, 1H, 7.65, m, 2H, 7.42, m, 2H, 7.1-7.2, m, 5H, 6.7-6.9, m, 1H, 6.1, m, 1H, 5.5-5.7, m, 1H, 4.7, m, 1H, 4.54, m, 0.5H, 4.2, m, 1H, 4.1, m, 0.5H, 3.7, m, 0.5H, 3.2, m, 1H, 3.0, m, 0.5H, 2.3, m, 1H, 2.1, m, 1H, 1.9, m, 1H, 1.6, m, 1H.

…………………….

SYNTHESIS

CN 103121999

To solve the above problems, the present invention adopts a technical solution is: to provide a tyrosine kinase inhibitor PC1-32765 synthesis method, the reaction steps are as follows:

The beneficial effect of the present invention: The invention relates to a tyrosine kinase inhibitor synthesis of PC1-32765, as the B cell to inhibit the tyrosine kinase receptor signaling key, not only can inhibit the formation of blood cells and less side effects and mild reaction conditions, simple operation, easy purification, low cost, environmentally friendly, suitable for large-scale production.

A tyrosine kinase inhibitor PC1-32765 synthesis method comprising the steps of:

1, the compound 10 and the coupling reaction of compound 15 to give compound 6;

2, the compound 6 obtained by reacting compound 16 with compound 11 in the process, we have chosen a more perfect catalyst;

3, compound 11 to give compound 12 by protecting;

4, selective deprotection of Compound 12 Compound 13; 5, Compound 13 for Compound 17 only attack only remaining position to obtain a very pure compound 14;

6, take off the protecting group to obtain PC1-32765

Wherein the compound can 10,15,16,17 agent or industrial grade reagent compound or the use of methods and techniques related to synthesis.

Example 1 Preparation of Compound 6

Under nitrogen and the 0.1moL 1.5 equivalents of compound 10 Compound 15 and 800mL of dioxane was added to 2L reaction flask, and then 1.5 equivalents of sodium acetate was added and the catalyst PdC12 (PPh3) 2 0.2 equivalents, 50_60 ° C for 5 hours , filtered hot and the filter residue was washed three times with ethanol, the combined filtrate was concentrated to give a solid, rinsed with ethanol to give the pure product 16.2 g, yield 60%

Example 2 Preparation of Compound 6

Under nitrogen and the 0.1moL 1.5 equivalents of compound 10 Compound 15 and 2L 800mL DMF was added to the reaction flask, and then 1.5 equivalents of sodium acetate was added and the catalyst PdCl2 (PhCN) 2 0.2 equivalents, 50_60 ° C for 5 hours, hot filtered, the filter residue was washed three times with ethanol, the combined filtrate was concentrated to give a solid, which was rinsed with ethanol to give pure product 21.5 g, yield 71%.

Example 3 Preparation of Compound 11

The compound 0.1moL 1.2 equivalent of compound 6 and 16, and 2L IOOOmL THF was added to the reaction flask, 1.5 equivalents of cesium carbonate was added, refluxed for 24 hours, after the reaction, most of the solvent was concentrated and the remaining water was poured into a large, precipitated solid was filtered, washed with water to afford compound 36.9 g compound 11, yield 76%, used without further purification.

Example 4 Preparation of Compound 12

The compound will be to 0.1moL 11 and 1.2 equivalent of compound IOOOmL THF trifluoroacetyl chloride and the reaction was added to 2L flask, then triethylamine was added 2.5 ,30-40 0C for 24 hours, after the reaction, the solvent was concentrated, diluted with water, extracted with ethyl acetate, washed with water, saturated sodium chloride each time, and concentrated to obtain the product 50.1 g of ethyl acrylate, 86% yield, used directly in the next reaction.

Example 5 Preparation of Compound 13

The compound 0.1moL 12 and 500mL of methanol and 50mL 6N hydrochloric acid was added to IL reaction flask, stirred at room temperature for 3 hours to complete the reaction quickly, and a solid precipitates, filtered and the solid was washed several times with ethyl acetate, obtain 38.5 g of pure compound 13 in 80% yield.

Example 6 Preparation of Compound 14 ‘

The 0.1moL compound 13 and 1.2 equivalents of acrylic acid chloride was added to 2L of methylene chloride IL reaction flask ,20-40 ° C was added dropwise 1.2 equivalents of triethylamine was added dropwise, at room temperature for 3 hours after the reaction with two chloride extraction and concentrated to give the product 47.7 g, yield 89%. Without further purification.

Example 7 PC1-32765 Preparation

Compound 14 with the 0.1moL 160mL 800mL of methanol and a saturated solution of sodium carbonate small, 50_60 ° C for 5 hours,

After completion of the reaction was diluted with water, concentrated and then extracted with methylene chloride, concentrated to obtain crude product was recrystallized from toluene to give the final product 28.6 g, yield 65%. HPLC purity 98.6%, ee%> 98%.

The present invention relates to a tyrosine kinase inhibitor of the synthesis of PC1-32765, as the B cell to inhibit the tyrosine kinase receptor signaling key, not only can inhibit the formation of blood cells and less side effects, and the reaction conditions gentle, simple operation, easy purification, low cost, environmentally friendly, suitable for large-scale production.

…

Discovery of selective irreversible inhibitors for Bruton’s tyrosine kinase

ChemMedChem

Volume 2, Issue 1, pages 58–61, January 15, 2007

http://onlinelibrary.wiley.com/doi/10.1002/cmdc.200600221/full

http://www.wiley-vch.de/contents/jc_2452/2007/z600221_s.pdf

SYN OF COMPD 4

To 101 mg of a known intermediate 2 [WO 2001019829] and 330 mg polymer-bound Triphenylphosphine (polymerlab) in 5 ml THF, 200 mg (2.0 eq.) of 3-OH N-Boc piperidine was added followed by 0.099 ml diisopropyl diazodicarboxylate. The reaction mixture stirred at room temperature overnight. After filtered off resins, the reaction mixture was concentrated and purified with flash chromatography (pentane/ethyl acetate = 1/1) to give 55 mg of intermediate 3. This compound (48.3 mg) was treated with 1 ml of 4N HCl in dioxane for 1 hour and concentrated to dryness, which was dissolved in dichloromethane and 0.042 ml of triethylamine, followed by 0.010 ml of acryl chloride. The reaction was stopped after 2 hours. The reaction mixture was washed with 5wt% citric acid (aq.) and brine, dried with MgSO4, and concentrated. Flash chromatography with (CH2Cl2/MeOH = 25/1) gave 22 mg of compound 4 as white solids. MS (M+1): 441.2; 1H-NMR (400MHz): 8.26, s, 1H; 7.65, m, 2H; 7.42, m, 2H; 7.1-7.2, m, 5H; 6.7-6.9, m, 1H; 6.1, m, 1H; 5.5-5.7, m, 1H; 4.7, m, 1H; 4.54, m, 0.5H; 4.2, m, 1H; 4.1, m, 0.5H; 3.7, m, 0.5H; 3.2, m, 1H; 3.0, m, 0.5H; 2.3, m, 1H; 2.1, m, 1H; 1.9, m, 1H; 1.6, m, 1H

……………..

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MORE
1) E · Werner, L · Honeywell Berg, Z · Pan; Bruton tyrosine kinase inhibitor; drugs circulating Company; filing date: 2006.12.28, open) No. (Notice: CN101610676A CN101610676B, CN101805341A, CN101805341B, CN102746305A, CN102887900A

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3) Celera Genomics Announces Sale of Therapeutic Programs to Pharmacyclics , April 10, 2006
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What Does 100% of Your Daily Value of Cholesterol Look Like?

Uncategorized Comments Off

Feb 262014

Healthline just published an interesting infograph that gives a visualization of what your daily value of cholesterol looks like. In the graphic, you can see what 300 mg of cholesterol looks like for 20 high cholesterol foods: http://www.healthline.com/health/high-cholesterol/daily-value

This is a very informative resource as it helps us visualize what their cholesterol intake look like

What Does 100% of Your Daily Value of Cholesterol Look Like?

It’s no secret that eating fatty foods raises your bad cholesterol level, also known as LDL. An elevated LDL clogs up your arteries and makes it difficult for your heart to do its job. Potentially, it could lead to heart disease.

The USDA recommends consuming no more than 300 mg of cholesterol a day. While a deep-fried Twinkie at the county fair is an obvious no-no, other high cholesterol culprits may be sneaking into your diet. Check out what that number looks like in terms of everyday food items.

Warning: you may need to revise your grocery list—and your eating habits!

Fried Chicken:

4 pieces=300mg cholesterol

Croissants:

6 2/3 rolls=300mg cholesterol

Cheddar Cheese:

12 3/4 slices=300mg cholesterol

Prosciutto:

28 slices=300mg cholesterol

Corned Beef:

14 thin slices=300mg cholesterol

Butter:

1 1/5 sticks=300mg cholesterol

read at

http://www.healthline.com/health/high-cholesterol/daily-value

CANGRELOR

Uncategorized Comments Off

Feb 242014

Cangrelor, AR-C69931MX

[dichloro-[[[(2R,3S,4R,5R)-3,4-dihydroxy-5-[6-(2-methylsulfanylethylamino)-2-(3,3,3-trifluoropropylsulfanyl)purin-9-yl]oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-hydroxyphosphoryl]methyl]phosphonic acid

N-[2-(Methylthio)ethyl]-2-[(3,3,3-trifluoropropyl)thio]-5¢-adenylic acid monoanhydride with (dichloromethylene)bis[phosphonic acid]

163706-06-7 cas no

Also known as: AR-C69931XX, 163706-06-7, Cangrelor (USAN/INN), Cangrelor [USAN:INN:BAN], UNII-6AQ1Y404U7, cangrelor (AR-C69931MX),

Molecular Formula: C17H25Cl2F3N5O12P3S2

Molecular Weight: 776.359196

Cangrelor

MAR 09, 2013

The Medicines Company said yesterday it will pursue marketing approvals for its anti-clotting drug candidate Cangrelor after it met its primary efficacy endpoint in a Phase III clinical trial of improvement compared with Plavix (clopidogrel).

The intravenous small molecule antiplatelet agent reduced by 22% the likelihood of patients experiencing death, myocardial infarction, ischemia-driven revascularization, or stent thrombosis within 48 hours of taking it—to 4.7% from 5.9% of subjects randomized during CHAMPION PHOENIX. The Phase III trial compared Cangrelor to oral Plavix in 11,145 patients undergoing percutaneous coronary intervention.

Cangrelor also showed a 38% reduction (0.8% compared with 1.4%) over Plavix in the likelihood of patients experiencing the key secondary endpoint, incidence of stent thrombosis at 48 hours.

Cangrelor is designed to prevent platelet activation and aggregation that leads to thrombosis in acute care settings, including in patients undergoing percutaneous coronary intervention. During CHAMPION PHOENIX, Cangrelor made its best showing in patients with Q-wave myocardial infarction (QMI), lowering by 39% (to 0.2% compared with 0.3%) the incidence of QMI. Cangelor’s most disappoint showing was its inability to lower the odds of death compared with Clopidogrel; both drugs showed a likelihood of 0.3%.

“Our next step is to submit for market approvals in the U.S. and Europe. We anticipate submitting these data for a new drug application to the U.S. Food and Drug Administration in the second quarter with findings of prior trials, including the BRIDGE trial in patients awaiting open heart surgery,” Simona Skerjanec, PharmD, senior vp and innovation leader for antiplatelet therapies at The Medicines Company, said in a statement.

Cangrelor is a P2Y12 inhibitor under investigation as an antiplatelet drug[1] for intravenous application. Some P2Y12 inhibitors are used clinically as effective inhibitors of adenosine diphosphate-mediated platelet activation and aggregation.[1] Unlike clopidogrel (Plavix), which is a prodrug, cangrelor is an active drug not requiring metabolic conversion.

Poor interim results led to the abandonment of the two CHAMPION clinical trials in mid 2009.[2] The BRIDGE study, for short term use prior to surgery, continues.[3] The CHAMPION PHOENIX trial was a randomized study of over 11,000 patients published in 2013. It found usefulness of cangrelor in patients getting cardiac stents. Compared with clopidogrel given around the time of stenting, intravenous ADP-receptor blockade with cangrelor significantly reduced the rate of stent thrombosis and myocardial infarction.[4]Reviewers have questioned the methodology of the trial.[5]

One particularly preferred example of a reversible, short-acting P2Y12 inhibitor is cangrelor. Cangrelor is a potent, direct, and reversible antagonist of the platelet P2Y12 receptor. Cangrelor has a half-life of approximately less than 10 minutes, allowing for a return to normal platelet function in a very short period of time upon discontinuation of the drug. By reducing the need for a compound to be metabolized for activity, and by having a relatively short half-life, reversible, short-acting P2Y12 inhibitors are considered “reversible,” meaning that full platelet functionality may return rather quickly as compared to thienopyridines.

The binding of cangrelor to the P2Y12 receptor inhibits platelet activation as well as aggregation when mediated in whole or in part via this receptor. Cangrelor can be derived completely from synthetic materials, and is an analogue of adenosine triphosphate (ATP). ATP is a natural antagonist of the P2Y12 receptor sites and is found in humans.

The chemical structure for cangrelor is depicted below as Formula I.

Cangrelor is clinically well tolerated and safe and has no drug-drug interaction with aspirin, heparin or nitroglycerin. Unlike orally dosed thienopyridines, cangrelor can be administered intravenously and binds directly to P2Y12 receptor sites of platelets. In each of the embodiments of the present invention, the term “cangrelor” encompasses the compound of Formula I as well as tautomeric, enantiomeric and diastereomeric forms thereof, and racemic mixtures thereof, other chemically active forms thereof, and pharmaceutically acceptable salts of these compounds, including a tetrasodium salt. These alternative forms and salts, processes for their production, and pharmaceutical compositions comprising them, are well known in the art and set forth, for example, in U.S. Pat. No. 5,721,219. Additional disclosure relevant to the production and use of cangrelor may be found in U.S. Pat. Nos. 5,955,447, 6,130,208 and 6,114,313, as well as in U.S. Appln. Publication Nos. 2006/0270607 and 2011/0112030.

Invasive procedures means any technique where entry to a body cavity is required or where the normal function of the body is in some way interrupted by a medical procedure and/or treatment that invades (enters) the body, usually by cutting or puncturing the skin and/or by inserting instruments into the body. Invasive procedures can include coronary artery bypass grafting (CABG), orthopedic surgeries, urological surgeries, percutaneous coronary intervention (PCI), other general invasive procedures, such as endarterectomy, renal dialysis, cardio-pulmonary bypass, endoscopic procedures or any medical, surgical, or dental procedure that could result in excessive bleeding or hemorrhage to the patient.

Perioperative means the period of a patient’s invasive procedure which can occur in hospitals, surgical centers or health care providers’ offices. Perioperative includes admission, anesthesia, surgery, to recovery.

Thrombosis is the formation of a blood clot (thrombus) inside a blood vessel obstructing the flow of blood through the circulatory system. When a blood vessel is injured, the body uses platelets and fibrin to form a blood clot to prevent blood loss. Some examples of the types of thrombosis include venous thrombosis which includes deep vein thrombosis, portal vein thrombosis, renal vein thrombosis, jugular vein thrombosis, Budd-Chiari syndrome, Paget-Schroetter disease, cerebral venous sinus thrombosis, cerebral venous sinus thrombosis and arterial thrombosis which includes stroke and myocardial infarction.

The compound cangrelor from the Medicines Company is represented by the structure

Figure imgf000013_0002

TETRASODIUM SALT

Cangrelor sodium, AR-C69931MX

Cangrelor Tetrasodium [USAN]
RN: 163706-36-3

Platelet P(2T) receptor antagonist.

5′-O-[[[Dichloro(phosphono)methyl](hydroxy)phosphoryloxy](hydroxy)phosphoryl]-N-[2-(methylsulfanyl)ethyl]-2-(3,3,3-trifluoropropylsulfanyl)adenosine tetrasodium salt

C17-H21-Cl2-F3-N5-O12-P3-S2.4-Na,

864.2899

The Medicines Co. (Proprietary), AstraZeneca Charnwood (Originator)

CARDIOVASCULAR DRUGS, Treatment of Disorders of the Coronary Arteries and Atherosclerosis, P2Y12 (P2T) Antagonists

2-Mercaptoadenosine (I) was S-alkylated with 1-chloro-3,3,3-trifluoropropane (II) in the presence of NaH to give trifluoropropyl sulfide (III). Subsequent acetylation of (III) with Ac2O at 80 C provided (IV), which was N-alkylated with methylthioethyl iodide (V) and NaH yielding (VI).

Further hydrolysis of the resulting (VI) with 0.1 M NaOH in refluxing MeOH furnished adenosine derivative (VII). The 5′-hydroxyl group of (VII) was then phosphorylated by reaction with phosphoryl chloride in cold triethyl phosphate followed by aqueous work-up.

The resulting 5′-monophosphate (VIII) was treated with carbonyl diimidazole and tri-n-butylamine to produce the phosphoryl imidazole intermediate (IX), which was finally condensed with dichloromethylenebis(phosphonic acid) (X).

The target compound was isolated as the tetrasodium salt upon treatment with NaI in methanol-acetone.

Alkylation of mercaptopurine (I) with 3-chloro-1,1,1-trifluoropropane (II) in the presence of NaH gave thioether (III).

After protection of the amino group of (III) as the acetamide (IV) by means of Ac2O and NaOAc, N-alkylation with 2-(methylthio)ethyl iodide (V) yielded (VI),

which was deacetylated by hydrolysis with NaOH in refluxing MeOH. Subsequent treatment with POCl3 produced the intermediate phosphoryl chloride (VIII).

Then, condensation of this acid chloride with dichloromethylene bisphosphonic acid (IX) in the presence of tributylamine in triethyl phosphate yielded the title compound, which was isolated as the tetrasodium salt.

Alternatively, hydrolysis of acid chloride (VIII) in the presence of ammonium bicarbonate gave phosphate salt (X), which was treated with carbonyldiimidazole, and the activated intermediate (XI) was then condensed with bisphosphonate (IX) to furnish the target compound.

…………

J. Med. Chem., 1999, 42 (2), pp 213–220

http://pubs.acs.org/doi/full/10.1021/jm981072s

10l (AR-C69931MX)

N6–(2-Methylthioethyl)-2-(3,3,3-trifluoropropylthio)-5‘-adenylic Acid, Monoanhydride withDichloromethylenebis(phosphonic acid) (10l). Prepared as the triammonium salt in 4% yield from 3l: 1H NMR δ(D2O) 8.30 (1H, s, H8), 5.97 (1H, d, J = 5.5 Hz, H1‘), 4.65 (1H, m, H2‘), 4.47 (1H, m, H3‘), 4.28 (1H, m, H4‘), 4.17 (2H, m, H5‘a and H5‘b), 3.67 (br s, NHCH2), 3.21 (2H, t, J = 7.6 Hz, SCH2), 2.72 (2H, t, J = 6.6 Hz, SCH2CH2CF3), 2.58 (2H, m, NCH2CH2), 2.04 (3H, s, SCH3);31P NMR δ(D2O) 8.80 (d, 1P, J = 18.6 Hz, Pγ), 0.42 (dd, 1P, J1 = 18.9 Hz, J2 = 28.9 Hz, Pβ), −9.41 (d, 1P, J = 29.0 Hz, Pα). Anal. (C17H34Cl2F3N8O12P3S2·3H2O) H, N, S; C: calcd, 23.16; found, 23.66.

References

Cangrelor Attenuates Coated-Platelet Formation
CHAMPION Trials With Cangrelor Stopped for Lack of Efficacy
What Cangrelor Failure Means to Medicines
Effect of Platelet Inhibition with Cangrelor during PCI on Ischemic Events (2013) Bhatt, DL etal. New England Journal of Medicine March 10, 2013 DOI: 10.1056/NEJMoa1300815 (published initially online).
The Duel between Dual Antiplatelet Therapies (2013) Lange, RA and Hillis, LD. New England Journal of Medicine March 10, 2013 DOI: 10.1056/NEJMe1302504
15th European Federation for Medicinal Chemistry International Symposium on Medicinal Chemistry (Sept 6 1998, Edinburgh)1998,:Abst P.281
Specific P2Y12 purinoceptor antagonist; inhibits ADP-induced platelet aggregation. Prepn: A. H. Ingall et al., WO 9418216 (1994 to Fisons); eidem, US 5721219 (1998 to Astra); and in vivo antithrombotic activity: idem et al., J. Med. Chem. 42, 213 (1999).
In vivo antithrombotic effects in canine arterial thrombosis: J. Huang et al., J. Pharmacol. Exp. Ther. 295, 492 (2000).
Mechanism of action study: A. Ishii-Watabe et al., Biochem. Pharmacol. 59, 1345 (2000).
Clinical safety assessment and evaluation in acute coronary syndromes: R. F. Storey et al., Thromb. Haemostasis 85, 401 (2001); in angina pectoris and non-Q-wave myocardial infarction: F. Jacobsson et al., Clin. Ther. 24, 752 (2002).
Clinical pharmacodynamics compared with clopidogrel: R. F. Storey et al., Platelets 13, 407 (2002).
Review of clinical development: S. C. Chattaraj, Curr. Opin. Invest. Drugs2, 250-255 (2001).
WO2013/103567 A2,
Journal of Medicinal Chemistry, 1999 , vol. 42, 2 p. 213 – 220

RAMELTEON, TAK 375 ..Melatonin MT1/MT2 receptor agonist

Uncategorized 1 Response »

Feb 232014

RAMELTEON

ACN-S001714, ZINC00007031

HSDB 7787
Ramelteon
Rozerem
TAK-375
UNII-901AS54I69

Molecular Formula: C₁₆H₂₁NO₂ Molecular Weight: 259.34344

CAS number	196597-26-9

	(S)-N-[2-(1,6,7,8-tetrahydro-2H-indeno-[5,4-b]furan-8-yl)ethyl]propionamide

(5)-N-[2-(l,6,7,8-tetrahydro-2H-indeno-[5,4-ό]furan-8- yl)ethyl]propionamide

United States

US 6034239

1999-07-22

expiry 2019-07-22

EP885210B1 , EP1792899A1 and J. Med Chem. 2002, 45, 4222-4239

NMR

[PPTData Supplement – Drug Metabolism and Disposition

dmd.aspetjournals.org/content/suppl/…/Supplemental_Information.pptx‎

May 17, 2010 – Ramelteon NMR Assignments. COSY: Black Arrows. HMBC: Red Arrows. Figure S-1b. 1H NMR Spectrum of Ramelteon. Figure S-1c.

Ramelteon is the first in a new class of sleep agents that selectively binds to the melatonin receptors in the suprachiasmatic nucleus (SCN). It is used for insomnia, particularly delayed sleep onset. Ramelteon has not been shown to produce dependence and has shown no potential for abuse.

Ramelteon, marketed as Rozerem by Takeda Pharmaceuticals North America, is the first in a new class of sleep agents that selectively binds to the MT₁ and MT₂ receptors in the suprachiasmatic nucleus (SCN), instead of binding to GABA _A receptors, such as with drugs like zolpidem,eszopiclone, and zaleplon. Ramelteon is approved by the U.S. Food and Drug Administration (FDA) for long-term use.

Ramelteon does not show any appreciable binding to GABA_A receptors, which are associated with anxiolytic, myorelaxant, and amnesic effects.

Rozerem (ramelteon), FDA Approved 07.04.05, can be used for insomnia, particularly delayed sleep onset. Ramelteon has not been shown to produce dependence and has shown no potential for abuse, and the withdrawal and rebound insomnia that is typical with GABA modulators is not present in ramelteon. Some clinicians also use ramelteon for the treatment of Delayed sleep phase syndrome.

Ramelteon

Mechanism of action

Ramelteon is a melatonin receptor agonist with both high affinity for melatonin MT₁ and MT₂ receptors and selectivity over the MT₃ receptor. Ramelteon demonstrates full agonist activity in vitro in cells expressing human MT₁ or MT₂ receptors, and high selectivity for human MT₁ and MT₂receptors compared to the MT₃ receptor.^[1]

The activity of ramelteon at the MT₁ and MT₂ receptors is believed to contribute to its sleep-promoting properties, as these receptors, acted upon by endogenous melatonin, are thought to be involved in the maintenance of the circadian rhythm underlying the normal sleep-wake cycle. Ramelteon has no appreciable affinity for the GABA receptor complex or for receptors that bind neuropeptides, cytokines, serotonin, dopamine, noradrenaline,acetylcholine, and opiates. Ramelteon also does not interfere with the activity of a number of selected enzymes in a standard panel.

The significance of ramelteon’s lack of affinity for the MT₃ receptor is not clear, despite the manufacturer’s emphasis of this fact in commercial advertisements. The MT₃ receptor appears almost exclusively in the gut and might not have any relationship to sleep or wakefulness.

The major metabolite of ramelteon, M-II, is active and has approximately one tenth and one fifth the binding affinity of the parent molecule for the human MT₁ and MT₂ receptors, respectively, and is 17 – 25-fold less potent than ramelteon in in vitro functional assays. Although the potency of M-II at MT₁ and MT₂ receptors is lower than the parent drug, M-II circulates at higher concentrations than the parent producing 20 – 100 fold greater mean systemic exposure when compared to ramelteon. M-II has weak affinity for the serotonin 5-HT2B receptor, but no appreciable affinity for other receptors or enzymes. Similar to ramelteon, M-II does not interfere with the activity of a number of endogenous enzymes.

All other known metabolites of ramelteon are inactive.

No published studies have indicated whether ramelteon, in humans, is more or less safe or effective than the hormone melatonin which it mimics; melatonin is much less expensive and is widely available over-the-counter in the US and Canada. The biological action of melatonin is similar to that of ramelteon. Ramelteon has been directly compared to melatonin in cats, and Ramelteon had a significant (3x) longer effect and had a more profound effect on the EEG of the sleeping cats.^[2]

Introduction

ROZEREM (ramelteon) is an orally active hypnotic, chemically designated as (S)-N-[2- (l,6,7,8-tetrahydro-2H-indeno-[5,4-b]furan-8-yl)ethyl]propionamide, and contains one chiral center. The compound is produced as the (S)-enantiomer, with an empirical formula of C₁₆H₂₁N0₂, molecular weight of 259.34, and the following chemical structure (I):

(I) -Ramelteon

Ramelteon is used to help patients who have sleep-onset insomnia (difficulty falling asleep) to fall asleep more quickly. It is the first in a new class of sleep agents that selectively binds to the MT] and MT₂ receptors in the suprachiasmatic nucleus (SCN), in a class of medications called melatonin receptor agonists with both high affinity for melatonin MT_! and MT₂ receptors and selectivity over the MT₃ receptor. It works similarly to melatonin, a natural substance in the brain that is needed for sleep.

Ramelteon was first disclosed in European patent EP 885210, which also disclosed a process for synthesizing ramelteon, as shown in scheme 1 : Scheme 1

Ramelteon

The processes of the prior art suffer from many disadvantages, some of which result from the fact that they involve several steps.

For instance, in US patent US 6034239, which is related to EP 885210, there is disclosed a process for preparing an intermediate compound of Formula (IV), which involves conversion of diethylcyano methyl phosphonate in the presence of 60% sodium hydride. Disadvantages of this particular reaction include the need for the highly flammable and corrosive base sodium hydride, the use of toxic triethyl phosphate for the formation of diethylcyano methyl phosphonate (which also has a high boiling point), and low yield of 60%. Such disadvantages mean that the disclosed process is difficult to implement industrially or economically. A further problem associated with prior art preparation techniques is the formation of dimeric impurities at the nitrile reduction stage (i.e. where the intermediate of Formula (IV) is reduced). For instance, US 6034239 discloses reduction of (l,2,6,7-Tetrahydro-8H-indeno-[5,4- b]furan-8-ylidene)-acetonitrile of formula (IV) by means of H₂ over Raney nickel in in a solvent medium of ethanol NH₃ to provide compound of formula (IIA). The reaction is carried out by applying 5 kg of hydrogen pressure, which results in the formation of the byproduct and impurity Dimer A, which in turn affects the yield and purity of the product of formula (IIA).

Dimer A

Similarly, (l,2,6,7-Tetrahydro-8H-indeno-[5,4-b]furan-8-ylidene)-acetonitrile of formula (IV) may be reduced by means of H₂ over Raney cobalt in a solvent medium of ethanol/ NH₃ to afford compound of formula (IIB). The reaction, which is carried out by applying hydrogen pressure, is not selective, and results in the formation of the by-product and impurity Dimer B, which in turn affects the yield and purity of product of formula (IIB).

Dimer B

Repeated purifications are required to remove impurities such as Dimer A and B to obtain ramelteon having the desired purity, which results in the poor yield of ramelteon.

Several other approaches are also described in the literature to make ramelteon and related compounds in WO2006030739, WO208062468, WO2008106179, US 2010152468, WO2009106966 and WO2010/055481. However, all processes of the prior art for the preparation of ramelteon are cumbersome; the processes employ a plurality of reagents and involve multiple steps, which make the overall processes uneconomical. Therefore there is a need for a more economical, efficient and industrially suitable method of making ramelteon, whereby address the problems associated with prior art, some of which are discussed above.

Ramelteon is the active ingredient in trademarked ROZEREM^®, and is approved by the United States Food and Drug Administration for the treatment of insomnia characterized by difficulty with sleep onset.

Different processes for preparing (S)-N-[2-(l,6,7,8-tetrahydro-2H-indeno-[5,4- b]furan-8-yl)ethyl]propionamide, i.e. ramelteon, are disclosed in US 6034239, JP 11080106, JP 11140073 and WO 2006/030739. U.S. Patent No. 6034239 describes the following processes for the preparation of ramelteon:

Japan Patent Publication No. 11080106 reports the following process for the preparation of ramelteon:

Ru(OCOCH₃)[(R)-BI NAP] IOOatm H₂/50 temp

BF₃ DEE Complex

Japan Patent Publication no. 11140073 reports the following process for the preparation of an intermediate of ramelteon:

PCT Publication No. WO/2006/030739 reports the following process for the preparation of ramelteon:

POCI3/DMF (EtO)₂P(O)CH₂CO₂Et Toluene NaH/Toluene

Purification in Ethanol water 95 7%

United States Patent No. 6,034,239 discloses the formation of chiral intermediates (S)-(- )-N-[2-(l,6,7,8,-tetrahydro-2H-indeno[5,4-b]furan-8-yl)ethylamine (sometimes referred to as compound S-2 or intermediate compound S-2) by the catalytic asymmetric hydrogenation of 2- (l,2,6,7,-tetrahydro-8H-indeno[5,4-b]furan-8-ylidene)ethylamine (compound 3 in the reaction scheme shown below) in the presence of a catalytic amount of BINAP-ruthenium complex in approximately 89% e.e. (enantiomeric excess). Following the catalytic reaction, the product is purified by preparing acid salts and acylated with propionyl chloride (compound 4 in the reaction scheme shown below) to obtain ramelteon (compound 1 in the reaction scheme shown below) in its pure (S) isomer form.

|S)*2

An alternate process for preparing ramelteon is disclosed in the Journal of Medicinal Chemistry, Vol. 45, pp. 4222-4239 (2002), wherein the exo double bond of intermediates (A) shown below was asymmetrically reduced using (S)-2, 2′-bis-(diphenylphosphino)-l, 1 ‘- binaphthyl (binap)-Ru complex as the catalyst to obtain the enantiomerically pure compound (B). Compound (B) is subsequently converted to ramelteon (1) through the intermediate steps of Claisen condensation, ozonolysis and cyclization.

m Both of the above processes uses expensive catalyst and give poor enantioselectivity. Additionally, these processes are expensive due to the need to perform multiple purifications steps in order to achieve an enantioselectivity of at least about 99% or greater of the desired isomer.

PCT Patent Publication No. WO 2008/062468 A2 discloses the following process for the preparation of ramelteon:

RAMELTEON

WO 2008/062468 teaches that separation of the enantiomers of intermediate (2) may be accomplished by: i) optical resolution of the racemic amine intermediate (2) by preparing acid salts with chirally pure acids; or ii) chromatographic techniques using chiral and/or achiral stationary phases for batch process, super critical or sub critical chromatography and/or continuous process chromatography. Although WO 2008/062468 mentions the possible use of optical resolution with chirally pure acids, there is no further teaching, discussion or disclosure of this method. WO 2008/062468 does, however, provide detailed descriptions of chromatographic methods for separating the isomers of intermediate compound (2). The disclosed chromatographic process suffers the following disadvantages:

• Preparative chromatography is time consuming & expensive;

• Highly sophisticated instrumentation required; • Not commercially feasible.

PCT Patent Publication No. WO 2008/106179 discloses a process for the preparation of ramelteon that involves the following reaction steps:

wherein X= O-alkyl or NH₂ and chiral reduction of the compound of formula IV in the presence of Ru-BINAP complex under hydrogen atmosphere in an organic solvent.

IV V

The process disclosed in WO 2008/106179 is similar to the process disclosed in United States Patent No. 6,034,239 and the Journal of Medicinal Chemistry, Vol. 45 in that a Ru-BINAP complex is employed.

Resolution of racemic mixtures via reaction with optically active acids and the subsequent crystallization of the resulting salts is preferably employed when the chiral carbon of the racemic compound is an alpha carbon {i.e., one carbon removed) to the functional group forming the acid addition salt. As the distance between the chiral carbon of the racemic compound to the functional group of the racemic compound increases to beta (i.e., two carbon removed) & gamma (i.e., three carbon removed), the resolution of the diastereomeric salt becomes more difficult and not very useful.

Ramelteon has a chiral center at the gamma carbon, which makes the separation of the isomer with an optically active acid quite a daunting task. Similarly, N-[2-(l, 6, 7, 8,- tetrahydro-2H-indeno [5, 4-b]furan-8-yl)]ethylamine (compound T), an intermediate useful in the production of ramelteon has a chiral center at the gamma carbon which would lead a skilled artisan to believe that optical resolution with an optically active acid could prove difficult.

Synthesis

Chilman-Blair, K.; Castañer, J.; Silvestre, J.S.; Bayés, M. (2003). “TAK-375”. Drugs of the Future 28 (10): 950. doi:10.1358/dof.2003.028.10.763214.

………………..

SYNTHESIS

Scheme 1 :

XIV (S)-XII

……………………………………..

SYNTHESIS

WO2012035303A2

Scheme 2

0-30°C

Metal salt Propionyl halide/

Propionc anhydride

Ramelteon (I)

Synthesis of ramelteon

Preparation 1

N-[2-(l,6,7,8-Tetrahydro-2H-indeno[5,4-b]furan-8-yl)-ethyl]-propionamide (2.0 gm) was dissolved in 50.0 ml (n-Hexane:IPA:DEA) (as used herein, “IPA” stands for isopropyl alcohol, and “DEA” stands for diethylamine)

and optically resolved by high performance column chromatography on CHTRAL PACK IA-3 using Mobile phase : n-Hexane:IPA:DEA Flow rate: 1.0ml/min UV:285 nm; at a column temperature of 25°C;sample concentartion: lmg/ml and, eluted with mobile phase. Both the enantiomers were collected separately and after evaporation of solvent under vacuum, enantiomerically pure ramelteon (I) was obtained. Preparation 2- using Supercritical Fluid Chromatography (SFC)

N-[2-(l₅6,7,8-Tetrahydro-2H-indeno[5,4-b]furan-8-yl)-ethyl]-propionamide (2.0 gm) was dissolved in 50.0 ml (n-Hexane:Ethanol:DEA) and optically resolved by Supercritical Fluid Chromatography (SFC) on CHIRAL PACK AD-H using a mobile phase : C02/(Methanol/ Diethylamine[DEA]) and eluted with mobile phase. Both the enantiomers were collected separately and after evaporation of solvent under vacuum, enantiomerically pure S- ramelteon of Formula (I) and R-ramelteon were obtained with isomeric purity>99%.

………………………………

SYNTHESIS

WO2010055481A1

synthesis of ramelteon that comprises the step of separating N-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8- yl)]ethylamine (compound 2) into its isomers using an optically active acid to achieve high enantioselectivity of the desired isomer. This embodiment may further include the step of acylating the substantially pure enantiomer, (S)-N-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8-yl)]ethylamine (compound (S)-2) using a suitable acylating agent, such as propionyl chloride) to provide (S)-7V-[2-(l,6,7,8-tetrahydro-2H-indeno[5,4-b]furan-8-yl]ethyl]propionamide (ramelteon or compound 1) substantially free of the (R)-isomer.

One embodiment of the present invention for the preparation of ramelteon is shown below in Scheme 1.

Example 1

Preparation of (S)-N-2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8-yl) ethylamine (Compound (S)-2)

A solution of N-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8-yl)ethylamine (45 g; 0.22 mol) in methanol (225 ml) is added to a solution of S-(+)-2-(4-isobutylphenyl)propionic acid (41 g; 0.20 mol) in methanol (205 ml) at 25-30⁰C. The reaction mixture is concentrated to dryness under reduced pressure. The crude salt precipitated is recrystallized in methanol to give a diastereomeric salt of (S)-N-2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8-yl) ethylamine with (S)-(+)-2-(4-isobutylphenyl) propionic acid having a chiral purity of greater than 90% enantioselectivity. The product obtained is recrystallized from methanol to give the pure salt having chiral purity of 99% or greater enantioselectivity.

The purified salt is suspended in water and the pH of the suspension is adjusted to 11-12 using aqueous sodium hydroxide. The reaction mixture is extracted with dichloromethane, washed with water and evaporated to give the pure (S)-N-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8-yl)]ethylamine (compound (S)-2), substantially free from its (R) isomer.

Example 2

Preparation of (S)-N-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan-8-yl)ethyl] propionamide (ramelteon)

Triethyl amine (15.15 g, 0.15 mol) and propionyl chloride (13.66 g, 0.15 mol) were added to a solution of S-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5,4-b]furan-8-yl)]ethylamine (25 g, 0.12 mol) (compound (S)-2) (prepared in Example 1) in dichloromethane and stirred at room temperature for 2 hours. 75 mL water was added to the reaction mixture, and the layers were separated. The dichloromethane layer was concentrated under reduced pressure and purified from a mixture of acetone and hexane to give (S)-N-[2-(l, 6, 7, 8-tetrahydro-2H-indeno [5, 4-b] furan- 8-yl) ethyl] propionamide (compound 1) having a chiral purity of 99% or greater enantioselectivity.

…………

INTERMEDIATES

WO2010115897A2

The intermediate compound of formula Vl, 6,7-dihydro-1 H-indeno[5,4-b]furan-8(2H)-one, can then be subjected to further synthesis steps to yield ramelteon by synthesis route known to or readily devisable by a person skilled in the art, suitably involving the introduction of the side chain having chirality and amide function. The documents mentioned infra are incorporated herein by way of reference. For example, the following synthesis route may be applied:

1 ) NaOH

Vl 2) H₂, Ru-BI NAP

3) HCI

4) H₂, Pd/C

Experimental Procedures

Example 1 :

Preparation of 4-(2-chloroethyl)-2,3-dihydrobenzofuran (II)

intermediate FTIR spectra of MeCN (140 ml) was recorded as reference. MeCN was cooled to -20⁰C, oxalyl chloride (16.5 ml) was added at once and waited until temperature re-stabilized at – 20⁰C. DMF (16.6 ml) was then added drop-wise (temperature between -18°C and -22°C, 0.5 ml/min). Reaction was stirred until no oxalyl chloride was visible and DMF level was stable by FTIR. Vilsmeier reagent is thereby formed in situ according to the following reaction:

Product I was then added portion wise (temperature between -18°C and -21⁰C, about 30 min). Formation of intermediate was immediately observed by FTIR. Reaction was stirred for one hour. Et₃N was then added drop-wise (temperature between -18°C and -22°C, 50 ml/h). At the end of addition, reaction was stirred 15 min at -20⁰C and temperature was slowly raised to 50⁰C (within about 15 min). Disappearance of intermediate and formation of DMF and product Il was monitored by FTIR. When reaction looked completed by FTIR (about 2h at 50°C), the reaction was cooled down to 20⁰C and quenched with water (45 ml). Solution was transferred to a round bottom flask and MeCN was removed under reduced pressure. Solution was then diluted with MTBE (100 ml) and water (50 ml). Phases were separated and aqueous phase was re-extracted twice with MTBE (50 ml). Combined organic phases were washed twice with 10% H₃Pθ₄/10% NaCI solution and stored at 4°C until next step.

List of FTIR bands used to follow the reaction (using 2^nd derivative and solvent subtraction): Oxalyl chloride (reactant): Height to two point baseline, peak from 1800 cm^“1 to 1770 cm^“1, baseline 1800 cm^“1 to 1770 cm^“1.

Intermediate: Height to single point baseline, peak from 1722 cm^“1 to 1712 cm^“1, baseline

1722 cm^“1.

Compound Il (product): Area to two point baseline, peak from 993 cm^“1 to 981 cm^“1, baseline 993 cm^“1 to 981 cm^“1.

DMF: Height to single point baseline, peak from 1694 cm^“1 to 1680 cm^“1, baseline 1694 cm^“1.

Example 2:

Preparation of 4-vinyl-2,3-dihydrobenzofuran (III)

M FTIR spectra of MTBE was recorded prior to the reaction as reference. To the solution of 4- (2-chloroethyl)-2,3-dihydrobenzofuran (II) in MTBE (150 ml) obtained at the previous step, was added, water (38 ml), Kl (1.37 g), Bu₄NOH 40% (19 ml) and NaOH 50% solution (66 ml). Reaction was vigorously stirred and heated at 50⁰C until reaction looked completed by FTIR (4 to 5 h). Warm reaction mixture was then transferred into an extraction funnel to give three phases. Water phase (bottom) was removed and did not contain product. Medium phase (colored black) was diluted with water (120 ml) and was extracted three times with MTBE. Combined organic phases were washed twice with water, once with 0.5M NaHSO₃/10% NaCI solution and once with 1 N NaOH/10% NaCI solution. MTBE solution was dried using MgSO₄, filtered, concentrated and used immediately for next step.List of FTIR bands used to follow the reaction (using 2^nd derivative and solvent subtraction) Compound Il (reactant): Area to zero, peak from 1440 cm^“1 to 1437 cm^“1 Compound III (product): Area to zero, peak from 1417 cm^“1 to 1412 cm^“1. Compound III (product): Area to zero, peak from 1565 cm^“1 to 1562 cm^“1.

Example 3:

Preparation of 1-(2,3-dihydrobenzofuran-4-yl)ethanone (V)

4-vinyl-2,3-dihydrobenzofuran (I I I ) (2.4 g) was dissolved in toluene (2 ml) and were successively added (ITC) (51 mg), PdCI₂(30 mg) and H₂O₂ 30% (2 ml). Reaction was vigorously stirred at 55°C until reaction looked completed by FTIR. (for around 24 h). Reaction was cooled down to room temperature, diluted with EtOAc (50 ml) and water (50 ml). Phases were separated and organic phase was washed with 0.5M NaHSO₃/10% NaCI solution and twice with 1 M NaHCO₃, dried over MgSO₄ and concentrated. Purification by flash chromatography gave 1-(2,3-dihydrobenzofuran-4-yl)ethanone (V). ¹H NMR δ (CDCI₃) 7.35 (dd, 1 H, J = 0.8 Hz, J = 7.8 Hz), 7.19 (t, 1 H, J = 7.9 Hz), 6.95 (d, 1 H, J = 8.0 Hz), 4.57 (t, 2H, J = 8.8 Hz), 3.52 (t, 2H, J = 8.8 Hz), 2.57 (s, 3H). ¹³C NMR δ (CDCI₃) 198.8, 161.0, 133.8, 128.2, 127.9, 121.4, 1 13.4, 71.6, 31.0, 27.6.

List of FTIR bands used to follow the reaction (using 2^nd derivative and solvent subtraction) Compound III (reactant): Area to single point baseline, peak from 925 cm^“1 to 915 cm^“1, baseline 915 cm^“1.

Compound V (product): Area to zero, peak from 1730 cm^“1 to 1724 cm^“1.

Example 4:

Preparation of 6,7-dihydro-1 H-indeno[5,4-b]furan-8(2H)-one (Vl)

V Vl1 -(2,3-dihydrobenzofuran-4-yl)ethanone (V) (1 g, 6.2 mmol) was dissolved in dioxane (9 ml). TADCA (dicyclohexylammonium 2,2,2-trifluoroacetate) (1 .82 g, 1 eq) and paraformaldehyde (0.61 1 g, 1.1 eq) were added. The reaction was heated at 100⁰C for 2 h. A second portion of TADCA (0.91 g, 0.5 eq) and paraformaldehyde (0.333 g, 0.6 eq) were added and the reaction was heated at 100⁰C for 2 h. Reaction was partitioned between water (20 ml) and pentane (30 ml). Aqueous phase was re-extracted 4 times with pentane (10 ml). Combined pentane phases were washed with water and brine, dried over MgSO₄. Solution was diluted to 100 ml with pentane. This solution was added dropwise to a pre-heated solution of sulfuric acid at 67°C (10 ml) under nitrogen stream. At the end of addition, the reaction was stirred for 30 min. Reaction was cooled down to room temperature and poured on iced water (50 ml). Solution was extracted 5 times with MTBE. Combined organic phases were washed with water, NaHCO₃ 1 M and brine, dried over MgSO₄ and concentrated. Purification by flash chromatography furnished pure 6,7-dihydro-1 H-indeno[5,4-b]furan-8(2H)-one (Vl). ¹H NMR δ (CDCI₃) 7.21 (dd, 1 H, J = 0.9 Hz, J = 9.0 Hz), 7.02 (d, 1 H, J = 8.2 Hz), 4.66 (t, 2H, J = 8.9 Hz), 3.48 (t, 2H, J = 8.9 Hz), 3.08 (dd, 2H, J = 4.9 Hz, J = 6.0 Hz), 2.69 (m, 2H). ¹³C NMR δ (CDCI₃) 207.5, 160.2, 147.1 , 133.6, 125.6, 123.9, 1 15.6, 72.3, 37.1 , 28.4, 25.4.

…………………………………

SYNTHESIS

Improve the synthesis and flow properties of an insomnia drug.

Ramelteon (1), marketed as Rozerem by Takeda Pharmaceuticals, is used to treat insomnia. V. K. Kansal and co-inventors describe several processes that are used to prepare it, all of which require many steps. The inventors offer no comments about the relative merits of the processes, but they state that a new industrial-scale process is needed. Their main claims are to intermediate acid 2 as a racemic mixture and individual enantiomers; one enantiomer is converted to 1 by the route shown in Figure 1.

The inventors use diastereomeric crystallization to resolve the racemic mixture by forming its (S)-1-phenylethylamine salt. The salt of the (R)-isomer of 2 is recovered first; then the salt of (S)-2 is isolated from the solution and acidified to give the free acid, which is purified by using (R)-1-phenylethylamine. Both enantiomers are isolated with >99.0% purity and >99.0% ee.

The (S)-acid is converted to acid chloride 3 and then to amide 4 by reactions with SOCl2and NH3 gas, respectively. The chloride is not isolated; the amide is recovered in 85–90% yield with 95–98% purity. When aq NH4OH is used instead of NH3 gas, the purity of 4 is slightly lower (93–96%). An alternative method for preparing 4 is to treat 2 with Et3N and ClCO2Et, followed by NH3. This method produces 4 in yields of 80–95% and 97–99% purity.

Amide 4 is reduced to amine 5 with NaBH4 and BF3·Et2O. The amine is purified by forming its chloride or oxalate salt in yields as high as 85% and 96–98% purity. The salts are used to prepare 1 by treating them with EtCOCl in the presence of base: NaOH for the chloride salt and Na2CO3 for the oxalate. In both cases, the yield of 1 is >92%, and the purity is as high as 99.9% after recrystallization from EtOH.

The inventors also recrystallized 1 from toluene to produce what they describe as a “nonelectrostatic” crystalline form, designated as form A. They describe the measurement of the electrostatic charge of the crystals in one of the patent’s examples. The measurements show that the average charge density of form A is ≈15 times lower than crystals obtained from EtOAc. Low electrostatic charge improves the flow characteristics of the solid, which is important in preparing drug formulations.

The inventors report the details of preparing rac-2 by a multistep procedure shown in Figure 2.

In most of the reaction steps, the product is isolated in crude form; the inventors do not indicate whether the product is purified before it is used in the next stage. The synthesis of rac-2 begins with the conversion of benzofuran (6) to aldehyde7 by treatment with POCl3 followed by hydrolysis. The crude product is isolated as a liquid in 85–90% yield and 90–92% purity.

In the next step, 7 is condensed with malonic acid (8) in the presence of piperidine and HOAc; acid 9 is isolated in 92–95% yield and 95% purity. Catalytic hydrogenation of 9produces 10 in 95% yield and 94–96% purity. The hydrogenation also can be carried out in the presence of NaOH and HCO2NH4; the yield and purity of 10 are the same, but the reaction takes 6 h instead of 2 h. [The patent does not state why NaOH and HCO2NH4 would be used.—Ed.] Acid 10 is brominated to produce acid 11, isolated in 50–60% yield and 92–95% purity.

The next stage begins with treating 11 with SOCl2 to activate the carboxyl group by forming acid chloride 12. The chloride is not isolated but cyclized under Friedel–Crafts conditions to give tricyclic compound 13, isolated in yields of 85–92% and 90-95% purity. This reaction also produces two impurities, 14 and 15, but the amounts are not reported. Removing the impurities gives 13 in good yield, but the inventors do not describe how this is done. They do report that the impurities can be isolated, and 1H and 13C NMR data are provided for both.

In the next step, the bromine atoms in 13 are replaced by hydrogen to give 16 in 85–90% yield and 96–97%purity. This reaction produces two impurities, 17 and 18; again, the amounts are not reported, but 1H and 13C NMR data are. After MeOH reflux in the presence active carbon, 16 is isolated in 80–85% yield with 99.3–99.8% purity. It is then converted to ester 20 by treating it with a solution of phosphonate 19 that contains suspended NaH. Crude product 20 is isolated in 80–85% yield and 92–95% purity as a mixture of (E)- and (Z)-isomers. The isomer mixture is hydrogenated, and base hydrolysis gives rac-2 in 90–95% isolated yield and 95–98% purity.

The inventors claim that the overall process is suitable for producing ramelteon on an industrial scale in a crystalline form that has improved flow characteristics. (Teva Pharmaceutical Industries [Petah Tiqva, Israel]. US Patent 8,084,630, Dec. 27, 2011;

RAMELTEON

…………………….

SYNTHESIS

http://sat.ecnu.edu.cn/Uploadnews/20120213113859628.pdf

CHINESE CHEMICAL LETTERS 22, 2011, 264 SEE SYN OF KEY INTERMEDIATE

1:(S)-N-(2-(6-Methoxy-2,3-dihydro-1H-inden-1-yl)ethyl)propionamide 1======KEY INTERMEDIATE

[a]D20 10.0 (c, 0.20, EtOH); mp 76–77 8C;

1H NMR (500 MHz, CDCl3): d1.15 (t, 3H, J = 7.5 Hz), 1.60 (m, 1H), 1.70 (m, 1H), 2.02 (m, 1H), 2.19 (q, 2H, J = 7.5 Hz), 2.32 (m, 1H), 2.76 (m, 1H), 2.85 (m, 1H), 3.11 (m,1H), 3.41 (m, 2H), 3.79 (s, 3H), 5.48 (s, 1H), 6.71 (dd, 1H, J = 2.0 Hz, 8.5 Hz), 6.75 (s, 1H), 7.11 (d, 1H, J = 8.0 Hz).

13C NMR (100 MHz,DMSO–d6): d173.7, 158.7, 148.1, 135.8, 124.9, 112.3, 109.2, 55.5, 42.7, 37.9, 34.8, 32.5, 30.6, 29.8, 9.9. EI-MS: 247 ([M]+); HR-MS 247.1572([M]+
, C15H21NO2; Calcd. 247.1571). The enantiomeric excess of (S)-1 was determined by HPLC as >99.9% [column, CHIRALPAK AS-H
(4.6 mm 250 mm), room temperature; eluent, hexane-2-propanol-triﬂuoroacetic acid (90:10:0.1); ﬂow rate, 1.0 mL/min; detect, 290 nm; tRof (S)-1, 30.7 min; tR of (R)-1 (enantiomer of (S)-1), 37.1 min].

…………………….

NMR

[PPTData Supplement – Drug Metabolism and Disposition

dmd.aspetjournals.org/content/suppl/…/Supplemental_Information.pptx‎

May 17, 2010 – Ramelteon NMR Assignments. COSY: Black Arrows. HMBC: Red Arrows. Figure S-1b. 1H NMR Spectrum of Ramelteon. Figure S-1c.

References

Owen RT (April 2006). “Ramelteon: profile of a new sleep-promoting medication”. Drugs Today 42 (4): 255–63. doi:10.1358/dot.2006.42.4.970842. PMID 16703122.
Miyamoto M, Nishikawa H, Doken Y, Hirai K, Uchikawa O, Ohkawa S (November 2004). “The sleep-promoting action of ramelteon (TAK-375) in freely moving cats”. Sleep 27 (7): 1319–25.PMID 15586784.
Zammit G, Erman M, Wang-Weigand S, Sainati S, Zhang J, Roth T (August 2007). “Evaluation of the Efficacy and Safety of Ramelteon in Subjects with Chronic Insomnia”. J Clin Sleep Med 3 (5): 495–504. PMC 1978328. PMID 17803013.
Daniel F. Kipke, MD |title=Evidence That New Hypnotics Cause Cancer |journal=University of California |date=March 2008 |url=http://escholarship.org/uc/item/12r2f32g#page-2
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WO2008106179A1	Feb 26, 2008	Sep 4, 2008	Teva Pharma	Intermediates and processes for the synthesis of ramelteon
WO2008151170A2	Jun 2, 2008	Dec 11, 2008	Teva Pharma	Process for the synthesis of ramelteon and its intermediates
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EP1792899A1	Sep 12, 2005	Jun 6, 2007	Takeda Pharmaceutical Company Limited	Process for production of optically active amine derivatives
US6034239 *	Mar 6, 1997	Mar 7, 2000	Takeda Chemical Industries, Ltd.	Tricyclic compounds, their production and use
US20010039286 *	Feb 13, 2001	Nov 8, 2001	Kevin Dinnell	2-aryl indole derivatives and their use as therapeutic agents

WO2006030739A1	Sep 12, 2005	Mar 23, 2006	Takeda Pharmaceutical	Process for production of optically active amine derivatives
WO2008062468A2	Oct 15, 2007	May 29, 2008	Cadila Healthcare Ltd	Process for the preparation of optically pure indeno [5,4-b] furan derivatives
WO2008106179A1	Feb 26, 2008	Sep 4, 2008	Teva Pharma	Intermediates and processes for the synthesis of ramelteon
WO2009106966A1	Feb 27, 2009	Sep 3, 2009	Medichem, S.A.	Process for preparing ramelteon.
WO2010055481A1	Nov 12, 2009	May 20, 2010	Watson Pharma Private Limited	Process for the preparation of ramelteon
WO2010092107A1 *	Feb 11, 2010	Aug 19, 2010	Lek Pharmaceuticals D.D.	Synthesis of (s)-n-[2-(1,6,7,8-tetrahydro-2h-indeno-[5,4-b]furan-8-yl)ethyl]propionamide
WO2010103553A1 *	Mar 10, 2009	Sep 16, 2010	Industriale Chimica S.R.L.	Process for the preparation of ramelteon
EP0885210A1	Mar 5, 1997	Dec 23, 1998	Takeda Chemical Industries, Ltd.	Tricyclic compounds, their production and use
US6034239	Mar 6, 1997	Mar 7, 2000	Takeda Chemical Industries, Ltd.	Tricyclic compounds, their production and use
US20100152468	Oct 16, 2009	Jun 17, 2010	Teva Pharmaceutical Industries Ltd.	Process for the synthesis of ramelteon and its intermediates

WO2008062468A2 *	Oct 15, 2007	May 29, 2008	Cadila Healthcare Ltd	Process for the preparation of optically pure indeno [5,4-b] furan derivatives
US5321154 *	Jul 31, 1992	Jun 14, 1994	Nagase & Company, Ltd.	Optical resolution of (.+-.)-2-(4-isobutylphenyl)-propionic acid
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US20080242877 *	Feb 26, 2008	Oct 2, 2008	Vinod Kumar Kansal	Intermediates and processes for the synthesis of Ramelteon

WO2012035303A2

Sep 16, 2011

Mar 22, 2012

Cipla Limited Et Al

A novel process for synthesis of ramelteon, and key intermediates for the synthesis of ramelteon

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CAPREOMYCIN

GENERIC, Uncategorized Comments Off

Feb 082014

Structure

Capreomycin is a peptide antibiotic, commonly grouped with the aminoglycosides, which is given in combination with other antibiotics for MDR-tuberculosis. Adverse effects include nephrotoxicity and 8th cranial auditory vestibular nerve nerve toxicity.

The drug should not be given with streptomycin or other drugs that may damage the auditory vestibular nerve. Patients on this drug will often require audiology tests.

It is a cyclic peptide. Capreomycin is administered intramuscularly and shows bacteriostatic activity.REF 20

Capreomycin is frequently used to treat Mycobacterium tuberculosis infections. Mycobacterium tuberculosis growth has been found to be inhibited at a concentration of 2.5 μg/mL. REF21

This is the basic structure of capreomycin. The table below identifies the various naturally occuring analogues^{12, 14}.

	R₁	R₂
Capreomycin IA	OH	b-Lys
Capreomycin IB	H	b-Lys
Capreomycin IIA	OH	NH₂
Capreomycin IIB	H	NH₂

Introduction

Capreomycin is a metabolite of Streptomyces capreolus, it is an antimycobacterial agent – and a potent tuberlostatic antibiotic. Capreomycin is effective against a number of Gram-positive and Gram-negative organisms, but is primarily active against mycobacteria. It has been used in the treatment of certain resistant strains of Mycobacterium tuberculosis. The drug was first described in 1960 be Herr, and was subsequently found to contain two components (I and II) and later to be comprised of four (IA, IB, IIA, IIB) as shown on thestructure page.

Tuberculosis

Tuberculosis is a disease of the respiratory system, and is airbourne. The bacilli implant themselves in areas such as the lungs, renal cortex and reticuloendothelial system where there is a high partial pressure of oxygen. This is the Primary infection and does not normally affect the person whilst their immune system is intact as the bacteria lie dormant. When the immune system is depressed, the secondary reactivation occurs, and effects of the disease are seen.

This infectious disease has been known since about 1000B.C., and it stills remains the ‘leading cause of death from a single infectious disease agent’⁷. It is estimated that around eight million people contract TB every year, of which 95% are in developing countries. Deaths from the disease is estimated at 3 million people per year by the World Health Organisation. The occurance of the disease is related directly to the economic state of the country. This is because the spread of the disease is greatly assisted by poor public and personal hygiene and by overcrowding. New drugs were develoed about forty years ago allowing tuberculosis to be regarded as a curable disease. This is no longer the case, as many multidrug-resistant strains of the disease have emerged. This is where capreomycin has it uses.

There are three groups of drugs used to treat TB, which vary in their effectiveness and potential side effects.

First line drugs include: isoniazid, rifampicin and pyrazinamide. These are most effective and have the fewest potential side effects.

Second line drugs include: ethambutol, streptomycin and p-amino salicyclic acid. These are less effective and have more toxic effects.

Third line drugs include: Capreomycin, cycloserine, viomycin, kanamycin and amikacin. These are least effective and have the most toxic effects.

The third line drugs have to be used for infections with tubercle bacilli, likely to be resistant to first and second line drugs or when first and second line drugs have been abandoned because of unwanted reactions. To decrease the possibility of resistant organisms from emerging, ‘Compound Drug Therapy’ is used where a concoction of several drugs is administered.

General Physical Data

Molecular Weight	653.70
Molecular Formula	C₂₅H₄₃N₁₃O₈
CAS Registry number	61394-77-2
Beilstein Registry number	876587
Chemical Name	L-3,6-diamino-hexanoyl->-cyclo-[L-2,3-diamino-propionyl->-L-seryl->-L-alanyl->-2-amino-3-ureido -acryloyl->-(S)-amino-((R)-2-amino-1(3),4,5,6-tetrahydro-pyrimidin-4-yl)-acetyl-(1->N%3&)]
Auto name	3,6-diamino-hexanoic acid [12-hydroxymethyl-3-(2-imino-hexahydro-pyrimidin-4-yl)-9-methyl- 2,5,8,11,14-pentaoxo-6-ureidomethylene-1,4,7,10,13-pentaaza-cyclohexadec-15-yl]-amide

		Cpm IA¹⁰	Cpm IB¹⁰	Cpm IIA¹⁴	Cpm IIB¹⁴
m.p. / ^oC		240-5	250-3	250	252
[a]_D / ^o		-22.0	-42.5	+9.3	-24.9
UV / nm	0.1 M HCl	269 (e 23, 400)	268 (22, 000)
	H₂O	268 (23, 200)	268 (21, 900)
	0.1 M NaOH	288 (15, 800)	290 (13, 100)

According to the literature⁹the following applies to naturally occuring capreomycin:
Ratio of IA to IB = 1.16
Capreomycin II = 1.5%

¹³C NMR Data of Cpm IA

Carbon Number	d / ppm
1	51.92
2	40.28
4	172.76
10	176.29
11	54.15
5, 14	55.66
5, 14	56.23
7	168.0
8	105.90
13	172.00
16	176.6
17	135.79
19	155.32
20	18.86
21	68.33
22	49.20
23	23.53
24	49.83
26	157.0 (b)
1′	172.0
2′	36.93
3′	49.26
4′	23.59
5′	29.77
6′	39.77

The included NMR data is taken from tables in the literature^{8, 14}

The ¹³C NMR data is that of Capreomycin IA only, and the carbons are numbered accordingly in red on the structure shown above.

Below are ¹H NMR tables for the four different naturally occurring forms of capreomycin, the NH protons and CH protons are given in different tables. The NH protons are again numbered on the Cpm IA structure above, but this time in blue. The CH protons are numbered according to their postion in the amino acid residue. These are also numbered in pink on the above diagram.

Chemical Shifts of CH protons in Capreomycin Analogues

Position of Amino Acid Residue		Cpm IA	Cpm IB	Cpm IIA	Cpm IIB
1	a-CH₂	2.63 (dd)	2.5 (dd)
		2.85 (dd)	2.81 (dd)
	b-CH₂	3.8 (m)	3.7 (m)
	g-CH₂	1.8 (m)	1.8 (m)
	d-CH₂	1.8 (m)	1.8 (m)
	e-CH₂	3.10 (m)	3.08 (m)
2	a-CH	4.3-3.5 (m)	4.2-4.5 (m)	4.3-4.6 (m)	4.3-4.6 (m)
	b-CH₂	3.3 (m)	3.3 (m)	3.3 (m)	3.3 (m)
		3.8 (m)	3.8 (m)	4.1 (m)	4.1 (m)
3	a-CH	4.86 (t)	4.67 (q)	4.84 (t)	4.68 (q)
	b-CH₂	3.84 (d)		3.95 (d)
	b-CH₃		1.43 (d)		1.45 (d)
4	a-CH	4.3-4.5(m)	4.2-4.5 (m)	4.3-4.5 (m)	4.3-4.5 (m)
	b-CH₂	3.7-4.2 (m)	3.7-4.2 (m)	3.7-4.2 (m)	3.79 (dd)
					3.8-4.2 (m)
5	b-CH	8.04 (s)	8.03 (s)	8.05 (s)	8.04 (s)
6	a-CH	5.01 (d)	4.96 (d)	5.01 (d)	4.95 (d)
	b-CH	4.5 (m)	4.5 (m)	4.5 (m)	4.5 (m)
	g-CH₂	1.6-2.3 (m)	1.6-2.3 (m)	1.6-2.3 (m)	1.6-2.3 (m)
	d-CH₂	3.3 (m)	3.3 (m)	3.3 (m)	3.3 (m)

Chemical Shifts of NH Protons of Capreomycin Analogues

	Cpm IA	Cpm IB	Cpm IIA	Cpm IIB
1	9.33 (d)	9.72(d)	9.60 (d)	9.50 (d)
2	9.24 (d)	9.24 (d)	9.33 (d)	9.30 (d)
3	8.82 (s)	8.76 (s)	9.10 (s)	9.10 (s)
4	8.64 (d)	8.68(d)	8.73 (d)	8.73 (d)
6	8.22 (t)	8.15 (t)
7	8.10 (t)	8.15 (t)	8.19 (t)	8.08 (t)
8	7.61 (d)	7.62 (d)	7.50 (d)	7.49 (d)
9	7.46 (s)	7.42 (s)	7.44 (s)	7.44 (s)
10	7.46 (s)	7.42 s)	7.31 (s)	7.18 (s)
11	6.48 (s)	6.49 (s)	6.43 (s)	6.34 (s)
12	6.29 (s)	6.34 (s)	6.29 (s)	6.27 (s)

Using the program gNMR I attempted to plot the above data. However, this was not successful as this program can only cope with molecules with up to 23 protons. As this molecule has Capreomycin IA has 43 hydogens, the generated ¹H NMR was lacking many essential peaks, and hence was not included.

IR Spectrum of Capreomycin IA

The same process could have done for any of the other three Capreomycin anlogues. The very broad band around 2000 cm^-1 upwards is due to the presence of so many nitrogen and carbonyl groups and hence hydrogen bonding.

Cyclo[3-[[(3S)-3,6-diamino-1-oxohexyl]amino]-L-alanyl-(2Z)-3-[(aminocarbonyl)amino]-2,3-didehydroalanyl-(2S)-2-[(4R)-2-amino-3,4,5,6-tetrahydro-4-pyrimidinyl]glycyl-(2S)-2-amino-b-alanyl-L-seryl]

capreomycinIA;Cyclo[3-[[(3S)-3,6-diamino-1-oxohexyl]amino]-L-alanyl-(2Z)-3-[(aminocarbonyl)amino]-2,3-didehydroalanyl-(2S)-2-[(4R)-2-amino-1,4,5,6-tetrahydro-4-pyrimidinyl]glycyl-(2S)-2-amino-b-alanyl-L-seryl] (9CI);1,4,7,10,13-Pentaazacyclohexadecane, cyclic peptide deriv.

37280-35-6

Formula:	C25H44 N14 O8
Molecular Weight:	668.83

Properties:Crystals. Mp: 246–248^°C.
Synonyms:capreomycin IA;Cyclo[A2pr*-Ser-N3-[(3S)-3,6-diamino-1-oxohexyl]A2pr-2-[(Z)-aminocarbonylaminomethylene]Gly-2-[(4R)-2-iminohexahydropyrimidine-4-yl]Gly-]SynthesisBelow is the peptide synthesis of capreomycin IA and IB. This was taken directly from the literature¹⁰.

No chemical synthesis of capreomycin could be found in any of the literature references. However, below is a synthesis devised from the peptide synthesis shown above. This is colour coded depending on the various amino residues. Each of the amino groups is added to the molecule in sequence linked by a peptide bond to eventually form the cyclo-structure. This was designed with some help from general references^1,2,3.

This synthesis would be identical for capreomycin IB other than the Serine-Bzl is replaced by Alanine and the synthesis works in exactly the same way.

Capreomycin

The individual components of the capreomycin were colour coded as follows:

Red	DEA / UDA	b, b � diethoxyalanine / b – ureidodehydroalanine
Green	A₂pr	a, b � diaminopropionic acid
Turquoise	Ser	Serine
Blue	Cpd	Capreomycidine
Pink	b–Lys	b–Lysine

The black components of the synthesis were the various protecting groups involved:

Boc	tert � butoxycarbonyl
Z	Benzyloxycarbonyl
ONSu	N-hydroxysuccinimide
Nps	o-Nitrophenylsulphenyl
NO₂	Nitro	NO₂
Bzl	Benzene

Abbreviation	Chemical Name
NMM	N-Methylmorpholine
DCC	N,N�-Dicyclohexylcarbodiimine
HOBt	l-Hydroxybenztriazole
HONSu	N-Hydroxysuccinimide
THF	Tetrahydrofuran

This is the synthesis of capreomycin IA. The IB form is produced in an identical fashion except that Ser � Bzl , is replaced with Ala.

……………………..

US8044186

Capastat Sulfate (capreomycin for injection) is a polypeptide antibiotic isolated from Streptomyces capreolus. It is a complex of 4 microbiologically active components which have been characterized in part; however, complete structural determination of all the components has not been established.

Capreomycin is supplied as the disulfate salt and is soluble in water. In complete solution, it is almost colorless.

Each vial contains the equivalent of 1 g capreomycin activity.

The structural formula is as follows:

Capastat Sulfate Structural Formula Illustration

Biological Action

Capreomycin is part of a group of drugs called aminoglycosides. These act to inhibit bacterial protein synthesis. The oxygen-dependent active transport by a polyamine carrier system affects the penetration of the aminoglycosides through the cell membrane of the bacterium. Minimal action on anaerobic organisms is observed. The effect of the aminoglycosides is bactericidal and is enhanced by agents that interfere with cell wall synthesis.

Very little is known about the mechanism of action of capreomycin specifically, but it is thought to inhibit protein synthesis by binding to the 70s ribosomal unit. Other sources⁶support this theory by suggesting that capreomycin “prevents protein biosynthesis by inhibiting group I intron splicing of RNA as well as blocking translation on the bacterial ribosome via inhibition of ribosomal subunits.” It has been reported¹⁴ that the b-amino group of the A₂pr residue promotes biological potency, and that its location within the molecule is of importance.

Side Effects

This powerful antimycobacterial agent can give rise to several side effects, some of which are listed below:

The following Nephrotoxic effects are reversible once treatment is stopped, but capreomycin is not recommended for people with kidney disorders.

Polyuria (excess urination)
Haematuria (red blood cells in the urine)
Proteinuria (protein in the urine)
Nitrogen metabolism
Electrolyte disturbances
Anorexia
Anaemia
Thirst

Capreomycin is also Ototoxic giving the following side effects. The nerve damage is permanent.

Deafness
Loss of vestibular function
Damage to the cranial nerve 8

References:1. An Introduction to Peptide Chemistry – P.D. Bailey
2. Organic Chemistry – Vollhardt and Schore
3. Peptide Synthesis – M. Bodanszky, Y. Klausner and M. Ondetti
4. Pharmacology – H.P. Rand, M.M. Dale and J.M. Ritter
5. http://www.aidsinfonyc.org/network/access/drugs/capr.html
6. http://rwingo1.chm.colostate.edu/group/duane/duane.html
7. http://www.hucmlrc.howard.edu/Pharmacology/handouts/TBRCLSIS.html
8. J. Org.Chem.,1977, 42, 8 – McGahren, Morton, Kunstmann, Ellestad
9. Bull.W.H.O., 1972, 47(3), 343-56 – Lightbrown et al.
10. Tetrahedron, 1978, 34(7), 912-7 – Nomoto, Teshima, Wakamiya, Shiba
11. Tetrahedron Letters, 1976, 43, 3907-10 – Shiba, Nomoto, Teshima, Wakamiya
12. J.Org.Chem., 1992, 57, 5214-5217 – Gould and Minott
13. Tetrahedron Letters, 1969, 30, 2549-41 – Bycroft, Cameron, Hassanali-Walji and Johnson
14. Bull.Chem.Soc.Jpn, 1979, 52(6), 1709-15 – Nomoto and Shiba
15. Experimentia – 1976, 32(9), 1109-11 – Nomoto and Wakamiya
16. Pharmazie – 1970, 25(8), 471-2 – Voigt and Maa Bared
17. Antimicrobial Agents Chemotherapy, 1964, 522-9 – Black, Griffith and Brickler
18. Antimicrobial Agents Chemotherapy, 1962, 201-12 – Herr
19. www2.chemie.uni-erlangen.de/services/telespec
20 “Capreomycin binds across the ribosomal subunit interface using tlyA-encoded 2′-O-methylations in 16S and 23S rRNAs”. Mol. Cell 23 (2): 173–82. July 2006. doi:10.1016/j.molcel.2006.05.044. PMID 16857584
21 http://www.toku-e.com/Assets/MIC/Capreomycin%20sulfate.pdf
CAPREOMYCIN wiki



Systematic (IUPAC) name
(3S)-3,6-diamino-N-[[(2S,5S,8E,11S,15S)-15-amino-11-[(4R)-2-amino-3,4,5,6-tetrahydropyrimidin-4-yl]-8-[(carbamoylamino)methylidene]-2-(hydroxymethyl)-3,6,9,12,16-pentaoxo-1,4,7,10,13-pentazacyclohexadec-5-yl]methyl]hexanamide; (3S)-3,6-diamino-N-[[(2S,5S,8E,11S,15S)-15-amino-11-[(4R)-2-amino-3,4,5,6-tetrahydropyrimidin-4-yl]-8-[(carbamoylamino)methylidene]-2-methyl-3,6,9,12,16-pentaoxo-1,4,7,10,13-pentazacyclohexadec-5-yl]methyl]hexanamide
Clinical data
AHFS/Drugs.com	monograph
MedlinePlus	a682860

Identifiers
CAS number	11003-38-6

Chemical data
Formula	C₂₅H₄₄N₁₄O₈
Mol. mass	668.706 g/mol

Risk Assessment of Potentially Genotoxic Impurities within the Framework of Quality by Design

regulatory, Uncategorized Comments Off

Feb 032014

A strategy for the risk assessment of potentially genotoxic impurities is described that utilizes Quality by Design in an effort to furnish greater process and analytical understanding, ultimately leading to a determination of impurity criticality. By identifying the risks and parameters that most influence those risks, an enhancement of both product and process control is attained that mitigates the potential impact of these impurities. This approach calls for the use of toxicological testing where necessary, chemical fate arguments when possible, multivariate analyses to develop design space, and use of spiking data to support specifications. Strong analytical support, especially with the development of low-level detection methods, is critical. We believe that this strategy not only aids in the development of a robust API process but also delivers on the identification and subsequent mitigation of risks to a class of impurities that are of high interest in the field.