Is it possible to use the results of collaborative trials for analytical methods to prove the laboratory- and product-specific validation of a method? From the perspective of this EMA reflection paper the concrete specifications are missing. These will be developed in the future. Find out more in this news.

http://www.gmp-compliance.org/enews_4415_EMA%20publishes%20Document%20on%20the%20Validation%20of%20analytical%20Methods_8430,8360,8369,Z-QCM_n.html

GMP News: EMA publishes Document on the Validation of analytical Methods

On 26 June 2014, the European Medicines Agency (EMA) published the concept paper “Transferring quality control methods validated in collaborative trials to a product/laboratory specific context”.

To accept a method an authority always requires a scientific validation. The same applies when existing methods are to be replaced, reduced or to be optimized (3R = replacement, reduction, refinement). Many of these new methods principally represent an improvement compared to the old “standard” methods and therefore are acceptable from a regulatory perspective.

The scientific proof of validation also includes the evidence of the concept and the possibility to transfer a method between different laboratories as well as large scale collaborative studies indicating that a method is suitable for the intended purpose. After completing these steps successfully, it can ultimately result in a monograph of the European Pharmacopoeia (Ph. Eur.) or also in a guidance document for the WHO or the EMA.

This method’s validity has to be proven by the laboratory that proposes the new method. Moreover, this validation also needs to be proven specifically for the medicinal products it is supposed to be used for. Laboratories that participated in large scale collaborative studies before, usually already created plenty of data telling something about the function of this method.

This EMA concept paper now suggests that more guidance documents should be developed on this subject: how can these data from large scale collaborative studies be used to easier implement the laboratory- and product-specific validation of 3R methods (3R – see above)? The concrete specifications for this are currently still missing.

The issue is also to introduce an alternative method without necessarily having to show that the new method correlates with the existing Pharmacopoeia method.

To get additional details please see the complete Reflection Paper “Transferring quality control methods validated in collaborative trials to a product/laboratory specific context“.

The deadline for submission of comments is on 31 October 2014.

How can one find a certain GMP requirement in the EU GMP Guide, in the FDA cGMP Guide and in the ISO 9001 without searching for a long time? The Good Practice Guide developed by the ECA has become a standard in many companies and is aimed at providing this information. A 26 pages matrix provides information about where to find a GMP requirement e.g. on Validation, QC Lab testing etc in the three major Guidelines. The comprehensive booklet with 500 pages contains a full text version of all three guidelines. You can find the GMP Matrix here.

http://www.gmp-compliance.org/eca_handbuecher.html

Publications

ECA Good Practice Guide – “GMP Matrix”

“FDA cGMP, EU GMP and ISO 9001 Matrix for a pharmaceutical Quality System – A GMP Roadmap”. (Version 15 of April 2014)
The revised ECA Good Practice Guide is a comprehensive juxtaposition containing the requirements laid down in FDA’s cGMP Guide, the EU GMP Guide and ISO 9001. The updated Matrix now has 26 pages as well as further 500 pages for the following three regulations

• FDA cGMP Guide
• EU GMP Guide Part I, II, and III incl. all Annexes
• ISO 9001 Quality Management Systems

In addition, the Good Practice Guide contains a ISO 9001/ICH10 Matrix and the complete Part III to the EU GMP Guide.

Price*: € 149 Non ECA Members, € 99 ECA Members

ORITAVANCIN

Read more at: http://www.pharmatimes.com/Article/14-08-07/FDA_clears_third_antibacterial_for_skin_infections_this_year.aspx#ixzz39hKIT6nP

OLD ARTICLE CUT PASTE

http://newdrugapprovals.org/2014/02/21/fda-accepts-filing-of-nda-for-iv-antibiotic-oritavancin-with-priority-review/

Oritavancin
(4R)-22-O-(3-Amino-2,3,6-trideoxy-3-C-methyl-alpha-L-arabinohexopyranosyl)-N3-(p-(p-chlorophenyl)benzyl)vancomycin

(3S, 6R, 7R, 22R, 23S, 26S, 36R, 38aR) -22 – (3-Amino-2 ,3,6-trideoxy-3-C-methyl-alpha-L-mannopyranosyloxy) -3 – (carbamoylmethyl ) -10,19-dichloro-44-[2-O-[3 - (4'-chlorobiphenyl-4-ylmethylamino) -2,3,6-trideoxy-3-C-methyl-alpha-L-mannopyranosyl] – beta-D-glucopyranosyloxy] -

Also known as N^DISACC-(4-(4-chlorophenyl)benzyl)A82846B and LY333328,N-(4-(4-chlorophenyl)benzyl)A82846B

Abbott (Supplier), Lilly (Originator), InterMune (Licensee)

The medicines company—

1. the Oritavancin Program Results.pdf

   phx.corporate-ir.net/External.File?item…t=1‎

   Jul 2, 2013 – Inhibits two key steps of cell wall synthesis: – Transglycosylation. – Transpeptidation. • Disrupts bacterial membrane integrity. Differentiated from  …

FDA Accepts Filing of NDA for IV Antibiotic Oritavancin with Priority Review

PARSIPPANY, NJ — (Marketwired) — 02/19/14 — The Medicines Company (NASDAQ: MDCO) today announced that the U.S. Food and Drug Administration (FDA) has accepted the filing of a new drug application (NDA) for oritavancin, an investigational intravenous antibiotic, with priority review. The Medicines Company is seeking approval of oritavancin for the treatment of acute bacterial skin and skin structure infections (ABSSSI) caused by susceptible gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA), administered as a single dose.

In December 2013, the FDA designated oritavancin as a Qualified Infectious Disease Product (QIDP). The QIDP designation provides oritavancin priority review, and an additional five years of exclusivity upon approval of the product for the treatment of ABSSSI. Priority review means the FDA’s goal is to take action on the application within six months, compared to 10 months under standard review. The FDA action date (PDUFA date) for oritavancin is August 6, 2014.
Oritavancin (INN, also known as LY333328) is a novel semi-synthetic glycopeptide antibiotic being developed for the treatment of serious Gram-positive infections. Originally discovered and developed by Eli Lilly, oritavancin was acquired by InterMune in 2001 and then by Targanta Therapeuticsin late 2005.^[1]

In Dec 2008 the FDA declined to approve it, and an EU application was withdrawn.

In 2009 the development rights were acquired by The Medicine Co. who are running clinical trials for a possible new FDA application in 2013.^[2]

Its structure is similar to vancomycin^[3] It is a lipoglycopeptide

About Oritavancin

Oritavancin is an investigational intravenous antibiotic for which The Medicines Company is seeking approval in the treatment of ABSSSI caused by susceptible gram-positive bacteria, including MRSA. In clinical trials, the most frequently reported adverse events associated with oritavancin were nausea, headache, vomiting and diarrhea. Hypersensitivity reactions have been reported with the use of antibacterial agents including oritavancin.

Oritavancin shares certain properties with other members of the glycopeptide class of antibiotics, which includes vancomycin, the current standard of care for serious Gram-positive infections in the United States and Europe.^[4] Data presented at the 47th Annual Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC) in September 2007 demonstrated that oritavancin possesses potent and rapid bactericidal activity in vitro against a broad spectrum of both resistant and susceptible Gram positive bacteria, including Staphylococcus aureus, methicillin-resistant Staphylococcus aureus, Enterococci, and Streptococci.^[5] Two posters presented at the meeting also demonstrated that oritavancin was more active than either metronidazole or vancomycin against strains of Clostridium difficile tested.^[6]

Anthrax : Research presented at the American Society for Microbiology (ASM) 107th Annual General Meeting in May 2007, suggested oritavancin’s potential utility as a therapy for exposure to Bacillus anthracis, the gram-positive bacterium that causes anthrax, having demonstrated efficacy in a mouse model both pre- and post-exposure to the bacterium^[7]

oritavancin

The 4′-chlorobiphenylmethyl group disrupts the cell membrane of gram positive bacteria.^[8] It also acts by inhibition of transglycosylation and inhibition of transpeptidation.^[9]

Results have been presented (in 2003) but possibly not yet published from two pivotal Phase 3 clinical trials testing the efficacy of daily intravenous oritavancin for the treatment of complicated skin and skin-structure infections (cSSSI) caused by Gram-positive bacteria. The primary endpoints of both studies were successfully met, with oritavancin achieving efficacy with fewer days of therapy than the comparator agents (vancomycin followed by cephalexin). In addition, oritavancin showed a significantly improved safety profile with a 19.2 percent relative reduction in the overall incidence of adverse events versus vancomycin/cephalexin (p<0.001) in the second and larger pivotal trial.^[10]

A Phase 2 clinical study was planned to run until May 2008 entitled “Single or Infrequent Doses for the Treatment of Complicated Skin and Skin Structure Infections (SIMPLIFI),” evaluating the efficacy and safety of either a single dose of oritavancin or an infrequent dose of oritavancin compared to the previously studied dosing regimen of 200 mg oritavancin given once daily for 3 to 7 days.^[11] Results published May 2011.^[12]

Regulatory submissions

USA

On February 11, 2008, Targanta submitted a New Drug Application (NDA) to the US FDA seeking approval of oritavancin;^[13] in April 2008, the FDA accepted the NDA submission for standard review.^[14] On 9 Dec 2008 the FDA said insufficient data for approval of oritavancin had been provided and they requested a further phase 3 clinical study to include more patients with MRSA.^[15]

Europe

June 2008, Targanta’s Marketing Authorization Application (MAA) for oritavancin was submitted and accepted for review by the European Medicines Agency (EMEA),^[16] but the company later withdrew the application in Aug 2009.^[17]

About The Medicines Company

The Medicines Company’s purpose is to save lives, alleviate suffering, and contribute to the economics of healthcare by focusing on 3,000 leading acute/intensive care hospitals worldwide. Its vision is to be a leading provider of solutions in three areas: acute cardiovascular care, surgery and perioperative care, and serious infectious disease care. The company operates in the Americas, Europe and the Middle East, and Asia Pacific regions with global centers today in Parsippany, NJ, USA and Zurich, Switzerland.

“We look forward to working with the FDA during the review process, and sharing the knowledge we have gained in our studies of oritavancin,” said Matthew Wikler, MD, Vice President and Medical Director, Infectious Disease Care for The Medicines Company. “We believe that upon approval, oritavancin, administered as a single dose for the treatment of ABSSSI, will offer new options for both physicians and their patients for the treatment of these infections.”

The oritavancin NDA is based on data from two Phase 3 clinical trials, SOLO I and SOLO II, which were conducted under a Special Protocol Assessment (SPA) agreement with the FDA. These Phase 3 trials evaluated the efficacy and safety of a single 1200mg dose of oritavancin compared to 7 to 10 days of twice-daily vancomycin in adults with ABSSSI, including infections caused by MRSA. The combined SOLO studies were conducted in 1,959 patients (modified intent-to -treat population, or mITT), with 405 of the patients suffering from an ABSSSI with a documented MRSA infection.

oritavancin

Drug substance

Oritavancin diphosphate

CLINICAL TRIALS..http://clinicaltrials.gov/search/intervention=oritavancin

• LY 333328 diphosphate
• LY333328 diphosphate
• Oritavancin diphosphate
• UNII-VL1P93MKZN
• 192564-14-0 CAS NO

INTRODUCTION

Oritavancin

Oritavancin inhibits cell wall synthesis by complexing with the terminal D-Ala-D-Ala of a nascent peptidoglycan chain and also to the pentaglycine bridge, thus inhibiting transglyco- sylation and transpeptidation. Unlike other glycopeptides, oritavancin is able to bind to depsipeptides including D-Ala-D-Lac, which fa- cilitates its inhibition of cell wall synthesis even in organisms exhibiting VanA-type resistance. Oritavancin forms homodimers prior to binding to D-Ala-D-Ala or D-Ala-D-Lac, which increases its binding affinity for the target site.The p-chloro-phenylbenzyl side chain of oritavancin interacts with the cell membrane, exerting two beneficial effects. This binding acts to main- tain the antibacterial in a prime position for peptidoglycan interactions and it also imparts oritavancin with the ability to disrupt the bac- terial membrane potential and thus increase membrane permeability.[22,23] Oritavancin has been shown to dissipate membrane potential in both stationary and exponential phase growing bacteria, which is rare and may carry clinical implications in terms of its activity against slowly growing organisms and biofilms. The dual mechanism of action could also theoretically increase effectiveness and reduce the risk of resist- ance selection. In addition to the aforemen- tioned mechanisms, it has also been hypothesized that oritavancin inhibits RNA synthesis.

vancomycin, desmethylvancomycin, eremomycin, teicoplanin (complex of five compounds), dalbavancin, oritavancin, telavancin, and A82846B (LY264826) having structures A, B, C, D, E, F, G and H:

R = B-2-Acetylamido-glucopyraπosyl- Attorney Docket No 33746-704 602

Dalbavancin, oritavancin and telavancin are semisynthetic lipoglycopeptides that demonstrate promise for the treatment of patients with infections caused by multi-drug-resistant Gram-positive pathogens. Each of these agents contains a heptapeptide core, common to all glycopeptides, which enables them to inhibit transglycosylation and transpeptidation (cell wall synthesis). Modifications to the heptapeptide core result in different in vitro activities for the three semisynthetic lipoglycopeptides. All three lipoglycopeptides contain lipophilic side chains, which prolong their half-life, help to anchor the agents to the cell membrane and increase their activity against Gram-positive cocci. In addition to inhibiting cell wall synthesis, telavancin and oritavancin are also able to disrupt bacterial membrane integrity and increase membrane permeability; oritavancin also inhibits RNA synthesis. Enterococci exhibiting the VanA phenotype (resistance to both vancomycin and teicoplanin) are resistant to both dalbavancin and telavancin, whileoritavancin retains activity. Dalbavancin, oritavancin and telavancin exhibit activity against VanB vancomycin-resistant enterococci.

All three lipoglycopeptides demonstrate potent in vitro activity against Staphylococcus aureus and Staphylococcus epidermidis regardless of their susceptibility to meticillin, as well as Streptococcus spp. Both dalbavancin and telavancin are active against vancomycin-intermediate S. aureus (VISA), but display poor activity versus vancomycin-resistant S. aureus (VRSA). Oritavancin is active against both VISA and VRSA. Telavancin displays greater activity against Clostridium spp. than dalbavancin, oritavancin or vancomycin. The half-life of dalbavancin ranges from 147 to 258 hours, which allows for once-weekly dosing, the half-life of oritavancin of 393 hours may allow for one dose per treatment course, while telavancin requires daily administration. Dalbavancin and telavancin exhibit concentration-dependent activity and AUC/MIC (area under the concentration-time curve to minimum inhibitory concentration ratio) is the pharmacodynamic parameter that best describes their activities.

Oritavancin’s activity is also considered concentration-dependent in vitro, while in vivo its activity has been described by both concentration and time-dependent models; however, AUC/MIC is the pharmacodynamic parameter that best describes its activity. Clinical trials involving patients with complicated skin and skin structure infections (cSSSIs) have demonstrated that all three agents are as efficacious as comparators. The most common adverse effects reported with dalbavancin use included nausea, diarrhoea and constipation, while injection site reactions, fever and diarrhoea were commonly observed withoritavancin therapy. Patients administered telavancin frequently reported nausea, taste disturbance and insomnia. To date, no drug-drug interactions have been identified for dalbavancin, oritavancin or telavancin. All three of these agents are promising alternatives for the treatment of cSSSIs in cases where more economical options such as vancomycin have been ineffective, in cases of reduced vancomycin susceptibility or resistance, or where vancomycin use has been associated with adverse events.

Oritavancin diphosphate (oritavancin) is a semi-synthetic lipoglycopeptide derivative of a naturally occurring glycopeptide. Its structure confers potent antibacterial activity against gram-positive bacteria, including vancomycin-resistant enterococci (VRE), methicillin- and vancomycin-resistant staphylococci, and penicillin-resistant streptococci. The rapidity of its bactericidal activity against exponentially-growing S. aureus (≧3-log reduction within 15 minutes to 2 hours against MSSA, MRSA, and VRSA) is one of the features that distinguishes it from the prototypic glycopeptide vancomycin (McKay et al., J Antimicrob Chemother. 63(6):1191-9 (2009), Epub 2009 Apr. 15).

Oritavancin inhibits the synthesis of peptidoglycan, the major structural component of the bacterial cell wall by a mechanism that is shared with glycopeptides, such as vancomycin (Allen et al., Antimicrob Agents Chemother 41(1):66-71 (1997); Cegelski et al., J Mol Biol 357:1253-1262 (2006); Arhin et al., Poster C1-1471: Mechanisms of action of oritavancin in Staphylococcus aureus [poster]. 47th Intersci Conf Antimicro Agents Chemo, Sep. 17-20, 2007; Chicago, Ill.). Oritavancin, like vancomycin, binds to the Acyl-D-Alanyl-D-Alanine terminus of the peptidoglycan precursor, lipid-bound N-acetyl-glucosamine-N-acetyl-muramic acid-pentapeptide (Reynolds, Eur J Clin Microbiol Infect Dis 8(11):943-950 (1989); Nicas and Allen, Resistance and mechanism of action.

In: Nagarajan R, editor. Glycopeptide antibiotics. New York: Marcel Dekker 195-215 (1994); Allen et al., Antimicrob Agents Chemother 40(10):2356-2362 (1996); Allen and Nicas, FEMS Microbiology Reviews 26:511-532 (2003); Kim et al., Biochemistry 45:5235-5250 (2006)). However, oritavancin inhibits cell wall biosynthesis even when the substrate is the altered peptidoglycan precursor that is present in VRE and vancomycin-resistant S. aureus (VRSA). Thus, the spectrum of oritavancin antibacterial activity extends beyond that of vancomycin to include glycopeptide-resistant enterococci and staphylococci (Ward et al., Expert Opin Investig Drugs 15:417-429 (2006); Scheinfeld, J Drugs Dermatol 6:97-103 (2007)). Oritavancin may inhibit resistant bacteria by interacting directly with bacterial proteins in the transglycosylation step of cell wall biosynthesis (Goldman and Gange, Curr Med Chem 7(8):801-820 (2000); Halliday et al., Biochem Pharmacol 71(7):957-967 (2006); Wang et al., Poster C1-1474: Probing the mechanism of inhibition of bacterial peptidoglycan glycotransferases by glycopeptide analogs. 47th Intersci Conf Antimicro Agents Chemo, Sep. 17-20, 2007). Oritavancin also collapses transmembrane potential in gram positive bacteria, leading to rapid killing (McKay et al., Poster C1-682: Oritavancin disrupts transmembrane potential and membrane integrity concomitantly with cell killing in Staphylococcus aureus and vancomycin-resistant Enterococci. 46th Intersci Conf Antimicro Agents Chemo, San Francisco, Calif., Sep. 27-30, 2006). These multiple effects contribute to the rapid bactericidal activity of oritavancin.

Vancomycin (U.S. Patent 3,067,099); A82846A, A82846B, and A82846C (U.S. Patent 5,312,738, European Patent Publication 256,071 A1); PA-42867 factors A, C, and D (U.S. Patent4,946,941 and European Patent Publication 231,111 A2); A83850 (U.S. Patent No. 5,187,082); avoparcm (U.S. Patent 3,338,786 and U.S. Patent 4,322,343); actmoidin, also known as K288 (J. Antibiotics Series A 14:141 (1961); helevecardin (Chem. Abstracts 110:17188 (1989) and Japanese Patent Application 86/157,397); galacardin (Chem. Abstracts 110:17188 (1989) and Japanese Patent Application 89/221,320); and M47767 (European Patent Publication 339,982).

Oritavancin is in clinical development against serious gram-positive infections, where administration of the drug is via intravenous infusion using several dosages administered over a series of days. The development of alternative dosing regimens for the drug could expand treatment options available to physicians. The present invention is directed to novel dosing regimens.

Means for the preparation of the glycopeptide antibiotics, including oritavancin and analogs thereof, may be found, for example, in U.S. Pat. No. 5,840,684,

ORITAVANCIN DIPHOSPHATE

SYNTHESIS

LY-333328 was synthesized by reductocondensation of the glycopeptide antibiotic A82846B (I) with 4′-chlorobiphenyl-4-carboxaldehyde (II) by means of sodium cyanoborohydride in refluxing methanol.

J Antibiot1996, 49, (6) :575-81

(3S,6R,7R,22R,23S,26S,36R,38aR)-3-(Carbamoylmethyl)-10,19-dichloro-7,28,30,32-tetrahydroxy-6-(N-methyl-D-leucylamido)-2,5,24,38,39-pentaoxo-22-(L-vancosaminyloxy)-44-[2-O-(L-vancosaminyl)-beta-D-glucopyranosyloxy]-2,3,4,5,6,7,23,24,25,26,36,37,38,38a-tetradecahydro-1H,22H-8,11:18,21-dietheno-23,36-(iminomethano)-13,16:31,36-dimetheno-[1,6,9]oxadiazacyclohexadecino[4,5-m][10,2,16]benzoxadiazacyclotetracosine-26-carboxylic acid; A82846B (I)
4′-chloro[1,1'-biphenyl]-4-carbaldehyde (II)

LY-333328 was synthesized by reductocondensation of the glycopeptide antibiotic A82846B (I) with 4′-chlorobiphenyl-4-carboxaldehyde (II) by means of sodium cyanoborohydride in refluxing methanol.

…………………..

WO1996030401A1

EXAMPLE 4

Preparation of Compound 229

A three liter 3-necked flask was fitted with a

condenser, nitrogen inlet and overhead mechanical stirring apparatus. The flask was charged with pulverized A82846B acetate salt (20.0 g, 1.21 × 10^-3 mol) and methanol (1000 mL) under a nitrogen atmosphere. 4′-chlorobiphenylcarboxaldehyde (2.88 g, 1.33 × 10^-2 mol, 1.1 eq.) was added to this stirred mixture, followed by methanol (500 mL). Finally, sodium cyanoborohydride (0.84 g, 1.33 × 10^-2 mol, 1.1 eq.) was added followed by methanol (500 mL). The resulting mixture was heated to reflux (about 65°C).

After 1 hour at reflux, the reaction mixture attained homogeneity. After 25 hours ac reflux, the heat source was removed and the clear reaction mixture was measured with a pH meter (6.97 at 58.0°C). 1 N NaOH (22.8 mL) was added

dropwise to adjust the pH to 9.0 (at 54.7°C). The flask was equipped with a distillation head and the mixture was concentrated under partial vacuum to a weight of 322.3 grams while maintaining the pot temperature between 40-45°C.

The distillation head was replaced with an addition funnel containing 500 mL of isopropanol (IPA). The IPA was added dropwise to the room temperature solution over 1 hour. After approximately 1/3 of the IPA was added, a granular precipitate formed. The remaining IPA was added at a faster rate after precipitation had commenced. The flask was weighed and found to hold 714.4 grams of the IPA/methanol slurry.

The flask was re-equipped with a still-head and

distilled under partial vacuum to remove the remaining methanol. The resulting slurry (377.8 g) was allowed to chill in the freezer overnight. The crude product was filtered through a polypropylene pad and rinsed twice with 25 mL of cold IPA. After pulling dry on the funnel for 5 minutes, the material was placed in the vacuum oven to dry at 40°C. A light pink solid (22.87 g (theory = 22.43 g) ) was recovered. HPLC analysis versus a standard indicated 68.0% weight percent of Compound 229 (4- [4-chlorophenyl] benzyl-A82846B] in the crude solid, which translated into a

corrected crude yield of 69.3%.

The products of the reaction were analyzed by reverse-phase HPLC utilizing a Zorbax SB-C18 column with ultraviolet light (UV; 230 nm) detection. A 20 minute gradient solvent system consisting of 95% aqueous buffer/5% CH₃CN at time=0 minutes to 40% aqueous buffer/60% CH₃CN at time=20 minutes was used, where the aqueous buffer was TEAP (5 ml CH₃CN, 3 ml phosphoric acid in 1000 ml water).

………………….

WO2008097364A2

Oritavancin (also termed N-(4-(4-chlorophenyl)benzyl)A82846B and LY333328) has the following Formula III:

References

ABT-414 is in phase I/II clinical development at AbbVie for the treatment of squamous cell carcinoma. The product is also in early clinical development for the treatment of glioblastoma multiforme.

In 2014, orphan drug designation was received in the U.S. and E.U. by AbbVie for the treatment of glioblastoma multiforme.

EGFR antibody-drug conjugate (cancer), Abbott; ABT-414; EGFR antibody-drug conjugate (cancer), AbbVie; EGFR-ADC (cancer), AbbVie; ABT-806-MMAF conjugate; anti-EGFR antibody-MMAF conjugate, AbbVie; EGFR-ADC (cancer), Abbott

AbbVie’s glioblastoma multiforme therapy receives orphan drug designation
AbbVie has obtained orphan drug designation from the European Medicines Agency (EMA) and the US FDA for its anti-epidermal growth factor receptor monoclonal antibody drug conjugate, ABT-414, as a treatment for glioblastoma multiforme.

AbbVie has obtained orphan drug designation from the European Medicines Agency (EMA) and the US FDA for its anti-epidermal growth factor receptor monoclonal antibody drug conjugate, ABT-414, as a treatment for glioblastoma multiforme.

read at

http://www.pharmaceutical-technology.com/news/newsabbvies-glioblastoma-multiforme-therapy-receives-orphan-drug-designation-4335836?WT.mc_id=DN_News

AbbVie’s ABT-414 Receives FDA and EMA Orphan Drug Designation

AbbVie oncology clinical development vice-president Gary Gordon said: “The orphan drug designation is an important regulatory advancement as we further our development in recurrent glioblastoma multiforme, a disease that is uniformly fatal with limited treatment options.

“We are pleased to continue developing ABT-414 in Phase II trials in patients with glioblastoma multiforme based on the results of our Phase I programme.”

AbbVie is currently evaluating the safety and efficacy of ABT-414 in patients with glioblastoma multiforme, the most aggressive type of malignant primary brain tumour.

In May, the company presented results from the Phase I clinical trial evaluating ABT-414 in combination with temozolomide in patients with recurrent or unresectable glioblastoma multiforme.

The Phase I trial was designed to assess the toxicities, pharmacokinetics and recommended Phase II dose of ABT-414 when administered every other week in combination with temozolomide.

Other important assessments included adverse events, pharmacokinetic parameters, objective response and tumour tissue epidermal growth factor receptor biomarkers.

The study results showed four objective responses, including one complete response.

AbbVie has developed ABT-414 with components in-licenced from Life Science Pharmaceuticals and Seattle Genetics.

ABT-414 is also being evaluated in clinical trials for the treatment of patients with squamous cell tumours.

About ABT-414
ABT-414 is an anti-EGFR (epidermal growth factor receptor) monoclonal antibody drug conjugate (ADC). As an ADC, ABT-414 is designed to be stable in the bloodstream and only release the potent cytotoxic agent once inside targeted cancer cells. Developed by AbbVie researchers with components in-licensed from Life Science Pharmaceuticals, ABT-414 is currently being investigated for the treatment of glioblastoma multiforme, the most common and most aggressive malignant primary brain tumor. ABT-414 is also in clinical trials for the treatment of patients with squamous cell tumors. ABT-414 is an investigational compound and its efficacy and safety have not been established by the FDA.

About Glioblastoma Multiforme
Glioblastoma is the most common and most aggressive type of malignant primary brain tumor. Each year in the U.S. and Europe, two to three out of every 100,000 people are diagnosed with glioblastoma, which has a five year survival rate of less than 3 percent. Prior to diagnosis, most patients experience a serious symptom of glioblastoma, such as a seizure. Typically patients succumb to the disease approximately 15 months after diagnosis. Treatment for glioblastoma remains challenging and no long-term treatments are currently available. Standard treatment is surgical resection, radiotherapy and concomitant adjunctive chemotherapy. More than 8,700 patients are enrolled in industry-sponsored clinical studies.

ref………

A phase 1 study evaluating ABT-414 in combination with temozolomide (TMZ) for subjects with recurrent or unresectable glioblastoma (GBM)
50th Annu Meet Am Soc Clin Oncol (ASCO) (May 30-June 3, Chicago) 2014, Abst 2021

ABT-414: An anti-EGFR antibody drug conjugate for the treatment of glioblastoma patients
18th Annu Sci Meet Soc Neuro-Oncol (November 21-24, San Francisco) 2013, Abst ET-079

ABT-414: An anti-EGFR antibody-drug conjugate as a potential therapeutic for the treatment of patients with squamous cell tumors
25th EORTC-NCI-AACR Symp Mol Targets Cancer Ther (October 19-23, Boston) 2013, Abst A250

A Phase I/II Study Evaluating the Safety, Pharmacokinetics and Efficacy of ABT-414 in Subjects With Advanced Solid Tumors Likely to Over-Express the Epidermal Growth Factor Receptor (EGFR) (NCT01741727)
ClinicalTrials.gov Web Site 2012, December 07

UK-414,495

Molecular Formula: C₁₆H₂₅N₃O₃S

Molecular Weight: 339.453

UK 414495

CAS  388630-36-2

OF

(-​)​-​(2R)​-​2-​[[1-​[[(5-​Ethyl-​1,​3,​4-​thiadiazol-​2-​yl)​amino]​carbonyl]​cyclopentyl]​methyl]​pentanoic acid;

AND

Cyclopentanepropanoi​c acid, 1-​[[(5-​ethyl-​1,​3,​4-​thiadiazol-​2-​yl)​amino]​carbonyl]​-​α-​propyl-​, (αR)​-

((R)-2-({1-[(5-ethyl-1,3,4-thiadiazol-2-yl) carbamoyl]cyclopentyl}methyl) valeric acid)

(2R)-2-[(1-{[(5-Ethyl-1,3,4-thiadiazol-2-yl)amino]carbonyl}cyclopentyl) methyl]pentanoic acid

…………………………………………………

Cas 337962-93-3  RACEMIC…………2-​[[1-​[[(5-​Ethyl-​1,​3,​4-​thiadiazol-​2-​yl)​amino]​carbonyl]​cyclopentyl]​methyl]​pentanoic acid  

…………………………………………………………………..

ITS ENANTIOMER

(+)​-​(2S)​-​2-​[[1-​[[(5-​Ethyl-​1,​3,​4-​thiadiazol-​2-​yl)​amino]​carbonyl]​cyclopentyl]​methyl]​pentanoic acid……………337962-74-0

Pfizer Inc.

CAS SUMMARY

desired

UK-414,495 is a drug developed by Pfizer for the treatment of female sexual arousal disorder.^[1] UK-414,495 acts as a potent, selective inhibitor of the enzyme neutral endopeptidase, which normally serves to break down the neuropeptide VIP. The consequent increase in VIP activity alters blood flow to the genital region leading to increased lubrication and muscle relaxation.^[2][3][4]

Female sexual arousal consists of a number of physiological responses resulting from increased genital blood. Vasoactive intestinal peptide (VIP), neuropeptide Y and to a lesser extent nitric oxide are neurotransmitters found in the vasculature of the genitalia. Neutral endopeptidase (NEP) modulates the activity of neuropeptides including VIP.

The aim of this study was to investigate the control of genital blood flow by VIP and endogenous neuropeptides using a selective NEP inhibitor [UK-414,495, ((R)-2-({1-[(5-ethyl-1,3,4-thiadiazol-2-yl) carbamoyl]cyclopentyl}methyl) valeric acid)].

A female equivalent of Viagra could soon be available to help women increase their sexual arousal, scientists claim.

For years they have endeavoured to create an alternative for women that mimics the effects of the male Viagra pill.

Now, the pharmaceutical company behind the original pill has created a prototype which increases blood flow to the genitalia in a similar way to Viagra.

Pfizer have come up with a prototype version of the female equivalent of Viagra

More than half of women experience sexual dysfunction at some point in their lives.

They may suffer a lack of desire, emotional or mental health problems and physical problems that mean they avoid having sex.

Pharmaceutical giant Pfizer has developed a drug, so far called only UK-414,495, which is supposed to increase sexual arousal, but will not affect desire, mood or emotional problems.

Some women take Viagra with mixed results and the drug has been used in fertility treatment to increase blood flow to the pelvis and encourage an embryo to implant in the womb.

But this is the first pill that claims to be an equivalent of the male Viagra.

The research, which involved animals, is published by the British Journal of Pharmacology, though Pfizer say they won’t develop the drug and warn that the chemical may not work the same way in humans, according to the Telegraph.

Chris Wayman, the lead researcher, said: ‘Before this work, we knew surprisingly little about the processes that control all of these changes.

Pfizer claim the tablets may help overcome female sexual arousal disorder

‘Now that we are beginning to establish the pathways involved in sexual arousal, scientists may be able to find ways of helping women who would like to overcome female sexual arousal disorder.

‘While the particular chemical compound in this research did not prove appropriate for further developments, the implications of the research could lead to the development of a product in the future.’

Viagra was originally developed as a treatment for high blood pressure and the heart condition angina, but men who took part in early trials realised the drug had an interesting side effect.

Clinical trials suggested the drug had little effect on angina and instead induced erections in men.

The drug first went on sale in 1998 and has since been prescribed to 25million men, creating a multi-billion pound global market.

The name Viagra has become so associated with men’s erectile problems that many cures are marketed as ‘herbal viagra’.

It is known by many nicknames, including Vitamin V and the Blue Pill.

Read more: http://www.dailymail.co.uk/health/article-1265842/Female-Viagra-help-women-increase-sexual-arousal.html#ixzz39lkmpSik 
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scheme

http://www.google.com/patents/US20020052370

Ex	Prec	n	Y	Data
43	Prep 37	0		1H NMR (CDCl3, 400 MHz) δ: 0.92 (t, 3H), 1.35 (t, 3H), 1.25-1.80 (m, 11H), 2.20-2.50 (m, 4H), 2.95 (q, 2H), 12.10 (bs, 1H); LRMS: m/z 339.8 (MH+) Anal. Found: C, 56.46; H, 7.46; N, 12.36. C16H25N3O3S requires C, 56.62; H, 7.44; N, 12.37%.

Example 29 (2R)-2-[(1-{[(5-Ethyl-1,3,4-thiadiazol-2-yl)amino]carbonyl}cyclopentyl) methyl]pentanoic acid

[0354]

desired

[0355] and

Example 30 (2S)-2-[(1-{[(5-Ethyl-1,3,4-thiadiazol-2-yl)amino]carbonyl}cyclopentyl) methyl]pentanoic acid

[0356]

undesired

[0357] The acid from Example 4 (824 mg) was further purified by HPLC using an AD column and using hexane:iso-propanol:trifluoroacetic acid (85:15:0.2) as eluant to give the title compound of example 29 as a white foam, 400 mg, 99.5% ee, ¹H NMR (CDCl₃, 400 MHz) δ: 0.90 (t, 3H), 1.36 (m, 6H), 1.50-1.80 (m, 9H), 2.19 (m, 1H), 2.30 (m, 1H), 2.44 (m, 1H), 2.60 (m, 1H), 2.98 (q, 2H), 12.10-12.30 (bs, 1H), LRMS: m/z 338 (MH⁻), [α]_D=−9.0°(c=0.1, methanol),

and

the title compound of example 30 as a white foam, 386 mg, 99% ee, ¹H NMR (CDCl₃, 400 MHz) δ: 0.90 (t, 3H), 1.38 (m, 6H), 1.50-1.79 (m, 9H), 2.19 (m, 1H), 2.30 (m, 1H), 2.44 (m, 1H), 2.60 (m, 1H), 2.98 (q, 2H), 12.10-12.27 (bs, 1H);

[0358] LRMS: m/z 338 (MH⁻); and [α]_D=+3.8°(c=0.1, methanol).

[0359] Alternatively, Example 29 may be prepared as follows:

[0360] To a solution of the product from Preparation 51a (574 g, 1.45 mol) in dichloromethane (2.87 L) was added trifluoroacetic acid (1.15 L) over a period of 50 minutes with cooling at 10° C. After addition was complete, the reaction was allowed to warm to ambient temperature with stirring under a nitrogen atmosphere for 24 hours. Deionised water (2.6 L) was then added. The reaction mixture was then washed with deionised water (3×2.6 L). The dichloromethane layer was concentrated to a volume of approximately 1 L to give the crude title compound (439 g, 1.29 mol, 96% yield) as a solution in dichloromethane. A purified sample of the title compound was obtained using the following procedure. To a dichloromethane solution (2.34 L) of the crude product, that had been filtered to remove any particulate contamination, was added isopropyl acetate (1.38 L). The resultant mixture was distilled at atmospheric pressure whilst being simultaneously replaced with isopropyl acetate until the solution temperature reached 87° C. The heating was stopped and the solution was allowed to cool to ambient temperature with stirring for 14 hours to give a cloudy brown solution. The agitation rate was then increased and crystallisation commenced. The suspension was then allowed to granulate for 12 hours at ambient temperature. The resultant suspension was then cooled to 0° C. for 3.5 hours and the solid was then collected by filtration. The filter cake was then washed with isopropyl acetate (2×185 ml, then 2×90 ml) and the solid was dried under vacuum at 40-45° C. for 18 hours to give the title compound (602 g, 0.18 mol, 70% yield) as a cream coloured, crystalline solid;

m.p.: 130-136° C.;

LRMS (negative APCI): m/z [M−H]⁻ 338; 

¹H-NMR (CDCl₃, 300 MHz) δ: 0.92 (t, 3H), 1.27-1.52 (m, 7H), 1.52-1.89 (m, 8H), 2.11-2.27 (m, 1H), 2.27-2.37 (m, 1H), 2.42-2.55 (m, 1H), 2.65 (dd, 2H), 3.00 (q, 2H), 12.25 (bs, 1H).

[0361] Example 29 may be purified as follows:

[0362] The title product from Example 29 was disolved in methanol. To this solution was added sodium methoxide (1 equivalent) in methanol (1 ml/g of Example 29) and the mixture was stirred at room temperature for 20 minutes. The solvent was removed in vacuo and the residue was azeotoped with ethyl acetate to give a brown residue. Ethyl acetate was added and the solution filtered to give a brown solid which was washed with tert-butylmethyl ether to give the crude sodium salt of Example 29. This crude product (35 g) was partitioned between water (200 ml) and ethyl acetate (350 ml). Concentrated hydrochloric acid (˜7 ml) was added until the pH of the aqueous layer was pH2. The aqueous phase was washed with ethyl acetate (2×100 ml). The combined layers were dried using magnesium sulphate. The solvent was removed in vacuo to give a light brown solid (31 g). Ethyl acetate (64 ml, 2 ml/g) and diisopropyl ether (155 ml, 5 ml/g) were added and the mixture heated to 68° C. until a clear solution was obtained (˜30 min). Upon cooling to room temperature, crystallisation of the free acid occurred. After 30 minutes stirring at room temperature the product was collected by filtration and washed with diisopropyl ether. The product was dried in a vacuum oven at 50° C. overnight. (20.2 g, 61% recovery from the sodium salt.); m.p. 135 degC (determined using a Perkin Elmer DSC7 at a heating rate of 20° C./minute).

[0372] The title compound of Example 29 metabolysed to form (2R)-1-(2-{[(5-ethyl-1,3,4-thiadiazol-2-yl)amino]carbonyl}pentyl)cyclopentanecarboxylic acid.

[0373] This compound was prepared as follows:

[0374] The product from Preparation 102 (430 mg, 1 mmol) was taken up in ethanol (5 mls) and methanol (1 ml) and hydrogenated at 30 psi hydrogen pressure at room temperature for 2 h. The mixture was then filtered through a plug of Arbocel®) and evaporated to a yellow oil. This oil was purified by column chromatography using firstly 19:1, then 9:1 DCM:MeOH as eluant to provide the product as a clear oil (120 mg, 35%); ¹HNMR (400 MHz, CDCl₃) 0.88 (t, 3H), 1.20-1.88 (m, 13H), 1.90-2.03 (m, 1H), 2.24-2.38 (m, 1H), 2.43-2.72 (m, 2H), 2.95 (q, 2H); LRMS m/z 340.2 (M+H).

Example 31 (R)-2-{[1-({[2-(Hydroxymethyl)-2,3-dihydro-1H-inden-2-yl]amino}carbonyl)-cyclopentyl]methyl}pentanoic acid

[0375] and

Example 32 (S)-2-{[1-({[2-(Hydroxymethyl)-2.3-dihydro-1H-inden-2-yl]amino}carbonyl)-cyclopentyl]methyl}pentanoic acid

[0376]

[0377] 2-{[1-({[2-(Hydroxymethyl)-2,3-dihydro-1H-inden-2-yl]amino}carbonyl)-cyclopentyl]methyl}pentanoic acid (WO 9110644, Example 8) was further purified by HPLC using an AD column and hexane:isopropanol:trifluoroacetic acid (90:10:0.1) as eluant, to give the title compound of Example 31, 99% ee, [α]_D=+10.40 (c=0.067, ethanol) and the title compound of Example 32, 99% ee, [α]_D=−10.9° (c=0.046, ethanol).

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http://www.google.com/patents/US6734186

Example 7 (+)-2-[(1-{[(5-Ethyl-1,3,4-thiadiazol-2-yl)amino]carbonyl}cyclopentyl)methyl]pentanoic Acid (F63)

The acid from Preparation 18 (18/ex4) (824 mg) was further purified by HPLC using an AD column and using hexane:iso-propanol:trifluoroacetic acid (85:15:0.2) as eluant to give the title compound of example 7 as a white foam, 386 mg, 99% ee,¹H NMR (CDCl₃, 400 MHz) δ: 0.90 (t, 3H), 1.38 (m, 6H), 1.50-1.79 (m, 9H), 2.19 (m, 1H), 2.30 (m, 1H), 2.44 (m, 1H), 2.60 (m, 1H), 2.98 (q, 2H), 12.10-12.27 (bs, 1H); LRMS: m/z 338 (MH-); and [α]_D=+3.80°(c=0.1, methanol)

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Novel selective inhibitors of neutral endopeptidase for the treatment of female sexual arousal disorder. Synthesis and activity of functionalized glutaramides
J Med Chem 2006, 49(14): 4409

Female sexual arousal disorder (FSAD) is a highly prevalent sexual disorder affecting up to 40% of women. We describe herein our efforts to identify a selective neutral endopeptidase (NEP) inhibitor as a potential treatment for FSAD. The rationale for this approach, together with a description of the medicinal chemistry strategy, lead compounds, and SAR investigations are detailed. In particular, the strategy of starting with the clinically precedented selective NEP inhibitor, Candoxatrilat, and targeting low molecular weight and relatively polar mono-carboxylic acids is described. This led ultimately to the prototype development candidate R-13, for which detailed pharmacology and pharmacokinetic parameters are presented.

ACID ENTRY 13

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WO 2002002513

http://www.google.com/patents/WO2002002513A1?cl=en

…………………..

WO 2002003995

http://www.google.com/patents/WO2002003995A2?cl=en

Scheme 12

LiAIHψ THF, 6hr at reflux

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• Novel selective inhibitors of neutral endopeptidase for the treatment of female sexual arousal disorder

  Original Research Article

• Pages 142-159
• David C. Pryde, Andrew S. Cook, Denise J. Burring, Lyn H. Jones, Stephanie Foll, Michelle Y. Platts, Vivienne Sanderson, Martin Corless, Alan Stobie, Donald S. Middleton, Laura Foster, Laura Barker, Piet Van Der Graaf, Peter Stacey, Christopher Kohl, Sara Coggon, Kevin Beaumont
• http://www.sciencedirect.com/science/article/pii/S0968089606008212

• Graphical abstract

References

UK-414,495

Systematic (IUPAC) name
(R)-2-({1-[(5-ethyl-1,3,4-thiadiazol-2-yl)carbamoyl]cyclopentyl}methyl)valeric acid
Clinical data
Legal status	?
Identifiers
CAS number	337962-93-3
ATC code	?
PubChem	CID 9949799
Chemical data
Formula	C16H25N3O3S
Mol. mass	339.452 g/mol

Female Sexual Response

The female sexual response phase of arousal is not easily distinguished from the phase of desire until physiological changes begin to take place in the vagina and clitoris as well as other sexual organs. Sexual excitement and pleasure are accompanied by a combination of vascular and neuromuscular events which lead to engorgement of the clitoris, labia and vaginal wall, increased vaginal lubrication and dilatation of the vaginal lumen (Levin, 1980; Ottesen, 1983; Levin, 1991; Levin, 1992; Sjoberg, 1992; Wagner, 1992; Schiavi et al., 1995; Masters et al., 1996; Berman et al., 1999).

Vaginal engorgement enables transudation to occur and this process is responsible for increased vaginal lubrication. Transudation allows a flow of plasma through the epithelium and onto the vaginal surface, the driving force for which is increased blood flow in the vaginal capillary bed during the aroused state. In addition engorgement leads to an increase in vaginal length and luminal diameter, especially in the distal ⅔ of the vaginal canal. The luminal dilatation of the vagina is due to a combination of smooth muscle relaxation of its wall and skeletal muscle relaxation of the pelvic floor muscles. Some sexual pain disorders such as vaginismus are thought to be due, at least in part, by inadequate relaxation preventing dilatation of the vagina; it has yet to be ascertained if this is primarily a smooth or skeletal muscle problem. (Levin, 1980; Oltesen, 1983; Levin, 1991; Levin, 1992; Sjoberg, 1992; Wagner, 1992; Schiavi et al., 1995; Master et al., 1996; Berman et al., 1999).

The vasculature and micro vasculature of the vagina are innervated by nerves containing neuropeptides and other neurotransmitter candidates. These include calcitonin gene-related peptide (CGRP), neuropeptide Y (NPY; Sequence No. 4), nitric oxide synthase (NOS), substance P and vasoactive intestinal peptide (VIP; Sequence No. 8) (Hoyle et al., 1996). Peptides that are present in the clitoris are discussed infra. Nitric oxide synthase, which is often colocalised with VIP (Sequence No. 8), displays a greater expression, immunologically, in the deep arteries and veins rather than in the blood vessels of the propria (Hoyle et al., 1996).

Female Sexual Dysfunction

It is known that some individuals can suffer from female sexual dysfunction (FSD). FSD is best defined as the difficulty or inability of a woman to find satisfaction in sexual expression. FSD is a collective term for several diverse female sexual disorders (Leiblum, 1998, Berman et al., 1999). The woman may have lack of desire, difficulty with arousal or orgasm, pain with intercourse or a combination of these problems. Several types of disease, medications, injuries or psychological problems can cause FSD.

Studies investigating sexual dysfunction in couples reveals that up to 76% of women have complaints of sexual dysfunction and that 30-50% of women in the USA experience FSD.

Sub-types of FSD include hypoactive sexual desire disorder, female sexual arousal disorder, orgasmic disorder and sexual desire disorder.

Treatments in development are targeted to treat specific subtypes of FSD, predominantly desire and arousal disorders.

The categories of FSD are best defined by contrasting them to the phases of normal female sexual response: desire, arousal and orgasm (Leiblum 1998). Desire or libido is the drive for sexual expression—and manifestations often include sexual thoughts either when in the company of an interested partner or when exposed to other erotic stimuli. In contrast, sexual arousal is the vascular response to sexual stimulation, an important component of which is vaginal lubrication and elongation of the vagina. Thus, sexual arousal, in contrast to sexual desire, is a response relating to genital (e.g. vaginal and clitoral) blood flow and not necessarily sensitivity. Orgasm is the release of sexual tension that has culminated during arousal. Hence, FSD typically occurs when a woman has an inadequate or unsatisfactory response in any of these phases, usually desire, arousal or orgasm. FSD categories include hypoactive sexual desire disorder, sexual arousal disorder, orgasmic disorders and sexual pain disorders.

Hypoactive sexual desire disorder is present if a woman has no or little desire to be sexual, and has no or few sexual thoughts or fantasies. This type of FSD can be caused by low testosterone levels, due either to natural menopause or to surgical menopause. Other causes include illness, medications, fatigue, depression and anxiety.

Female sexual arousal disorder (FSAD) is characterised by inadequate genital response to sexual stimulation. The genitalia (e.g. the vagina and/or the clitoris) do not undergo the engorgement that characterises normal sexual arousal. The vaginal walls are poorly lubricated, so that intercourse is painful. Orgasms may be impeded. Arousal disorder can be caused by reduced oestrogen at menopause or after childbirth and during lactation, as well as by illnesses, with vascular components such as diabetes and atherosclerosis. Other causes result from treatment with diuretics, antihistamines, antidepressants eg SSRIs or antihypertensive agents. FSAD is discussed in more detail infra.

Sexual pain disorders (which include dyspareunia and vaginismus) are characterised by pain resulting from penetration and may be caused by medications which reduce lubrication, endometriosis, pelvic inflammatory disease, inflammatory bowel disease or urinary tract problems.

The prevalence of FSD is difficult to gauge because the term covers several types of problem, some of which are difficult to measure, and because the interest in treating FSD is relatively recent. Many women’s sexual problems are associated either directly with the female ageing process or with chronic illnesses such as diabetes and hypertension.

There are wide variations in the reported incidence and prevalence of FSD, in part explained by the use of differing evaluation criteria, but most investigators report that a significant proportion of otherwise healthy women have symptoms of one or more of the FSD subgroups. By way of example, studies comparing sexual dysfunction in couples reveal that 63% of women had arousal or orgasmic dysfunction compared with 40% of men have erectile or ejaculatory dysfunction (Frank et al., 1978).

However, the prevalence of female sexual arousal disorder varies considerably from survey to survey. In a recent National Health and Social Life Survey 19% of women reported lubrication difficulties whereas 14% of women in an outpatient gynaecological clinic reported similar difficulties with lubrication (Rosen et al., 1993).

Several studies have also reported dysfunction with sexual arousal in diabetic women (up to 47%), this included reduced vaginal lubrication (Wincze et al., 1993). There was no association between neuropathy and sexual dysfunction.

Numerous studies have also shown that between 11-48% of women overall may have reduced sexual desire with age. Similarly, between 11-50% of women report problems with arousal and lubrication, and therefore experience pain with intercourse. Vaginismus is far less common, affecting approximately 1% of women.

Studies of sexually experienced women have detailed that 5-10% have primary anorgasmia. Another 10% have infrequent orgasms and a further 10% experience them inconsistently (Spector et al., 1990).

Because FSD consists of several subtypes that express symptoms in separate phases of the sexual response cycle, there is not a single therapy. Current treatment of FSD focuses principally on psychological or relationship issues. Treatment of FSD is gradually evolving as more clinical and basic science studies are dedicated to the investigation of this medical problem. Female sexual complaints are not all psychological in pathophysiology, especially for those individuals who may have a component of vasculogenic dysfunction (eg FSAD) contributing to the overall female sexual complaint. There are at present no drugs licensed for the treatment of FSD. Empirical drug therapy includes oestrogen administration (topically or as hormone replacement therapy), androgens or mood-altering drugs such as buspirone or trazodone. These treatment options are often unsatisfactory due to low efficacy or unacceptable side effects.

Since interest is relatively recent in treating FSD pharmacologically, therapy consists of the following:- psychological counselling, over-the-counter sexual lubricants, and investigational candidates, including drugs approved for other conditions. These medications consist of hormonal agents, either testosterone or combinations of oestrogen and testosterone and more recently vascular drugs, that have proved effective in male erectile dysfunction. None of these agents has been demonstrated to be very effective in treating FSD.

Female Sexual Arousal Disorder (FSAD)

The sexual arousal response consists of vasocongestion in the pelvis, vaginal lubrication and expansion and swelling of the external genitalia. The disturbance causes marked distress and/or interpersonal difficulty. Studies investigating sexual dysfunction in couples reveals that there is a large number of females who suffer from sexual arousal dysfunction; otherwise known as female sexual arousal disorder (FSAD).

The Diagnostic and Statistical Manual (DSM) IV of the American Psychiatric Association defines Female Sexual Arousal Disorder (FSAD) as being:

“a persistent or recurrent inability to attain or to maintain until completion of the sexual activity adequate lubrication-swelling response of sexual excitement. The disturbance must cause marked distress or interpersonal difficulty.”

FSAD is a highly prevalent sexual disorder affecting pre-, peri- and post menopausal (±HRT) women. It is associated with concomitant disorders such as depression, cardiovascular diseases, diabetes and UG disorders.

The primary consequences of FSAD are lack of engorgement/swelling, lack of lubrication and lack of pleasurable genital sensation. The secondary consequences of FSAD are reduced sexual desire, pain during intercourse and difficulty in achieving an orgasm.

It has recently been hypothesised that there is a vascular basis for at least a proportion of patients with symptoms of FSAD (Goldstein et al., 1998) with animal data supporting this view (Park et al., 1997).

Drug candidates for treating FSAD, which are under investigation for efficacy, are primarily erectile dysfunction therapies that promote circulation to the male genitalia. They consist of two types of formulation, oral or sublingual medications (Apomorphine, Phentolamine, Sildenafil), and prostaglandin (PGE₁-Alprostadil) that are injected or administered transurethrally in men, and topically to the genitalia in women.

The present invention seeks to provide an effective means of treating FSD, and in particular FSAD.

SUMMARY

The present invention is based on findings that FSAD is associated with reduced genital blood flow—in particular reduced blood flow in the vagina and/or the clitoris. Hence, treatment of women with FSAD can be achieved by enhancement of genital blood flow with vasoactive agents. In our studies, we have shown that cAMP mediates vaginal and clitoral vasorelaxation and that genital (e.g. vaginal and clitoral) blood flow can be enhanced/potentiated by elevation of cAMP levels. This is a seminal finding.

In this respect, no one has previously proposed that FSAD can be treated in such a way—i.e. by direct or indirect elevation of cAMP levels. Moreover, there are no teachings in the art to suggest that FSAD was associated with a detrimental modulation of cAMP activity and/or levels or that cAMP is responsible for mediating vaginal and clitoral vasorelaxation. Hence, the present invention is even further surprising.

In addition, we have found that by using agents of the present invention it is possible to increase genital engorgement and treat FSAD—e.g. increased lubrication in the vagina and increased sensitivity in the vagina and clitoris. Thus, in a broad aspect, the present invention relates to the use of a cAMP potentiator to treat FSD, in particular FSAD.

The present invention is advantageous as it provides a means for restoring a normal sexual arousal response—namely increased genital blood flow leading to vaginal, clitoral and labial engorgement. This will result in increased vaginal lubrication via plasma transudation, increased vaginal compliance and increased genital (e.g. vaginal and clitoral) sensitivity. Hence, the present invention provides a means to restore, or potentiate, the normal sexual arousal response.

More particularly, the present invention relates to:

A pharmaceutical composition for use (or when in use) in the treatment of FSD, in particular FSAD; the pharmaceutical composition comprising an agent capable of potentiating cAMP in the sexual genitalia of a female suffering from FSD, in particular FSAD; wherein the agent is optionally admixed with a pharmaceutically acceptable carrier, diluent or excipient.

The use of an agent in the manufacture of a medicament (such as a pharmaceutical composition) for the treatment of FSD, in particular FSAD; wherein the agent is capable of potentiating cAMP in the sexual genitalia of a female suffering from FSD, in particular FSAD.

A method of treating a female suffering from FSD, in particular FSAD; the method comprising delivering to the female an agent that is capable of potentiating cAMP in the sexual genitalia; wherein the agent is in an amount to cause potentiation of cAMP in the sexual genitalia of the female; wherein the agent is optionally admixed with a pharmaceutically acceptable carrier, diluent or excipient.

An assay method for identifying an agent that can be used to treat FSD, in particular FSAD, the assay method comprising: determining whether an agent can directly or indirectly potentiate cAMP; wherein a potentiation of cAMP in the presence of the agent is indicative that the agent may be useful in the treatment of FSD, in particular FSAD.

Cefuroxime Axetil

Cefuroxime Axetil (1-(acetyloxy) ethyl ester of cefuroxime, is (RS)-1-hydroxyethyl (6R,7R)-7-[2-(2-furyl)glyoxyl-amido]-3-(hydroxymethyl)-8-oxo-5-thia-1-azabicyclo[4.2.0]-oct-2-ene-2-carboxylate, 7 ² -(Z)-(O-methyl-oxime), 1-acetate 3-carbamate.

Its molecular formula is C ₂₀ H ₂₂ N ₄ O ₁₀S, and it has a molecular weight of 510.48.

Cefuroxime Axetil is used orally for the treatment of patients with mild-to-moderate infections, caused by susceptible strains of the designated microorganisms.

Cefuroxime axetil is a second generation oral cephalosporin antibiotic. It was discovered by Glaxo now GlaxoSmithKline and introduced in 1987 as Zinnat.^[1] It was approved by FDA on Dec 28, 1987.^[2] It is available by GSK as Ceftin in US^[3] and Ceftum in India.^[4]

It is an acetoxyethyl ester prodrug of cefuroxime which is effective orally.^[5] The activity depends on in vivo hydrolysis and release of cefuroxime.

Cefuroxime is chemically (6R, 7R)-3-carbamoyloxymethyl-7-[(Z)-2-(fur-2-yl)-2-methoxy-iminoacetamido] ceph-3-em-4-carboxylic acid and has the structural Formula II:

Cefuroxime axetil having the structural Formula I:

is the 1-acetoxyethyl ester of cefuroxime, a cephalosporin antibiotic with a broad spectrum of activity against gram-positive and gram negative micro-organisms.

This compound as well as many other esters of cefuroxime, are disclosed and claimed in U.S. Pat. No. 4,267,320. According to this patent, the presence of an appropriate esterifying group, such as the 1-acetoxyethyl group of cefuroxime axetil, enhances absorption of cefuroxime from the gastrointestinal tract, whereupon the esterifying group is hydrolyzed by enzymes present in the human body.

Because of the presence of an asymmetric carbon atom at the 1-position of the 1-acetoxyethyl group, cefuroxime axetil can be produced as R and S diastereoisomers or as a racemic mixture of the R and S diastereoisomers. U.S. Pat. No. 4,267,320 discloses conventional methods for preparing a mixture of the R and S isomers in the crystalline form, as well as for separating the individual R and S diastereoisomers.

The difference in the activity of different polymorphic forms of a given drug has drawn the attention of many workers in recent years to undertake the study on polymorphism. Cefuroxime axetil is the classical example of amorphous form exhibiting higher bioavailability than the crystalline form.

U.S. Pat. No. 4,562,181 and the related U.S. Pat. Nos. 4,820,833; 4,994,567 and 5,013,833, disclose that cefuroxime axetil in amorphous form, essentially free from crystalline material and having a purity of at least 95% aside from residual solvents, has a higher bioavailability than the crystalline form while also having adequate chemical stability.

These patents disclose that highly pure cefuroxime axetil can be recovered in substantially amorphous form from a solution containing cefuroxime axetil by spray drying, roller drying, or solvent precipitation. In each case, crystalline cefuroxime axetil is dissolved in an organic solvent and the cefuroxime axetil is recovered from the solution in a highly pure, substantially amorphous form.

Another U.S. Pat. No. 5,063,224 discloses that crystalline R-cefuroxime axetil which is substantially free of S-isomer is readily absorbed from the stomach and gastrointestinal tract of animals and is therefore ideally suited to oral therapy of bacterial infections.

According to this patent, such selective administration of R-cefuroxime axetil results in surprisingly greater bioavailability ability of cefuroxime, and thus dramatically reduces the amount of unabsorbable cefuroxime remaining in the gut lumen, thereby diminishing adverse side effects attributable to cefuroxime.

British Patent Specification No. 2,145,409 discloses a process for obtaining pure crystalline cefuroxime axetil and is said to be an improvement over British Patent Specification No. 1,571,683. Sodium cefuroxime is used as the starting material in the disclosed specification, which in turn, is prepared from either 3-hydroxy cefuroxime or cefuroxime.

Said process involves an additional step of preparing sodium cefuroxime, and therefore is not economical from commercial point of view.

CEFTIN (cefuroxime axetil) Tablets and CEFTIN (cefuroxime axetil) for Oral Suspension contain cefuroxime as cefuroxime axetil. CEFTIN (cefuroxime axetil) is a semisynthetic, broad-spectrum cephalosporin antibiotic for oral administration.

Chemically, cefuroxime axetil, the 1-(acetyloxy) ethyl ester of cefuroxime, is (RS)-1-hydroxyethyl (6R,7R)-7-[2-(2-furyl)glyoxyl-amido]-3-(hydroxymethyl)-8-oxo-5-thia-1-azabicyclo[4.2.0]-oct-2-ene-2-carboxylate, 7²-(Z)-(O-methyl-oxime), 1-acetate 3-carbamate. Its molecular formula is C₂₀H₂₂N₄O₁₀S, and it has a molecular weight of 510.48.

Cefuroxime axetil is in the amorphous form and has the following structural formula:

CEFTIN (cefuroxime axetil) Tablets are film-coated and contain the equivalent of 250 or 500 mg of cefuroxime as cefuroxime axetil. CEFTIN (cefuroxime axetil) Tablets contain the inactive ingredients colloidal silicon dioxide, croscarmellose sodium, hydrogenated vegetable oil, hypromellose, methylparaben, microcrystalline cellulose, propylene glycol, propylparaben, sodium benzoate, sodium lauryl sulfate, and titanium dioxide.

CEFTIN (cefuroxime axetil) for Oral Suspension, when reconstituted with water, provides the equivalent of 125 mg or 250 mg of cefuroxime (as cefuroxime axetil) per 5 mL of suspension. CEFTIN (cefuroxime axetil) for Oral Suspension contains the inactive ingredients acesulfame potassium, aspartame, povidone K30, stearic acid, sucrose, tutti-frutti flavoring, and xanthan gum.

Cefuroxime axetil

Systematic (IUPAC) name
1-Acetoxyethyl (6R,7R)-3-[(carbamoyloxy)methyl]-7-{[(2Z)-2-(2-furyl)-2-(methoxyimino)acetyl]amino}-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate
Clinical data
Identifiers
PubChem	CID 6321416
ChemSpider	4882027
ChEMBL	CHEMBL1095930
Synonyms	Cefuroxime 1-acetoxyethyl ester
Chemical data
Formula	C20H22N4O10S
Mol. mass	510.475 g/mol

dsc

http://www.google.com/patents/US5013833

http://www.google.com/patents/US6833452

EXAMPLE 1

Dicyclohexylamine (17.2 g) in N,N-dimethylacetamide (50 ml) was added to a solution of cefuroxime acid (42.4 g) in N,N-dimethylacetamide (300 ml) at about −10° C. (R,S)1-Acetoxethylbromide (33.4 g) in N,N-dimethylacetamide (50 ml) was added to the above solution and the reaction mixture was stirred for 45 minutes at about −3 to 0° C. Potassium carbonate (1.1 g) was added to the reaction mixture and it was further stirred at that temperature for about 4 hours. The reaction mixture was worked up by pouring into it ethyl acetate (1.0 It), water (1.2 It) and dilute hydrochloric acid (3.5% w/w, 200 ml). The organic layer was separated and the aqueous layer was again extracted with ethyl acetate. The combined organic extracts were washed with water, dilute sodium bicarbonate solution (1%), sodium chloride solution and evaporated in vacuo to give a residue. Methanol was added to the residue and the crude product was precipitated by adding water.

The resulting precipitate was filtered off and recrystallized from the mixture of ethylacetate, methanol and hexane. The precipitated product was filtered, washed and dried to give pure crystalline cefuroxime axetil (42.5 g).

Assay (by HPLC on anhydrous basis)-98.2% w/w; Diastereoisomer ratio-0.53; Total related substances-0.48% w/w.

…………………………………

http://www.google.com/patents/EP1409492B1?cl=en

• The present invention relates to an improved method for synthesis of cefuroxime axetil of formula (I) in high purity substantially free of the corresponding 2-cephem(Δ²)-ester of formule (II) and other impurities. The compound produced is valuable as a prodrug ester of the corresponding cephalosporin- 4-carboxylic acid derivative i. e. cefuroxime, particularly suitable for oral administration in various animal species and in man for treatment of infections caused by gram-positive and gram-negative bacteria.

BACKGROUND OF THE INVENTION

• [0002]
  One of the ways to improve the absorption of cephalosporin antibiotics which are poorly absorbed through the digestive tract is to prepare and administer the corresponding ester derivatives at the 4-carboxylic acid position. The esters are then readily and completely hydrolysed in vivoby enzymes present in the body to regenerate the active cephalosporin derivative having the free carboxylic acid at the 4-position.
• [0003]
  Among the various ester groups that can be prepared and administered only a selected few are biologically acceptable, in addition to possessing high antibacterial activity and broad antibacterial spectrum. Clinical studies on many such potential “prodrug esters” such as cefcanel daloxate (Kyoto), cefdaloxime pentexil tosilate (Hoechst Marion Roussel) and ceftrazonal bopentil (Roche), to name a few have been discontinued, while ceftizoxime alapivoxil ((Kyoto) in under Phase III clinical studies. The cephalosporin prodrug esters which have been successfully commercialised and marketed include cefcapene pivoxil (Flomox^® , Shionogi), cefditoren pivoxil (Spectracef^®, Meiji Seika), cefetamet pivoxil (Globocef^®, Roche), cefotiam hexetil (Taketiam^®, Takeda), cefpodoxime proxetil (Vantin^®, Sankyo), cefteram pivoxil (Tomiron^®, Toyama) and cefuroxime axetil (Ceftin^® and Zinnat^®, Glaxo Wellcome).
• [0004]
  Typically, such (3,7)-substituted-3-cephem-4-carboxylic acid esters represented by formula (I A) are synthesised by reacting the corresponding (3,7)-substituted-3- cephem-4-carboxylic acid derivative of formula (III A), with the desired haloester compound of formula (IV A) in a suitable organic solvent. The synthesis is summarised in Scheme-I, wherein in compounds of formula (I A), (II A), (III A) and (IV A) the groups R¹ and R² at the 3- and 7-positions of the β-lactam ring are substituents useful in cephalosporin chemistry ; R³ is the addendum which forms the ester function and X is halogen.

• [0005]
  However, the esterification reaction which essentially involves conversion of a polar acid or salt derivative to a neutral ester product invariably produces the corresponding (3,7)-substituted-2-cephem (Δ²)-4-carboxylic acid ester derivative of formula (II A) in varying amounts, arising out of isomerisation of the double bond from the 3-4 position to the 2-3 position as well as other unidentified impurities.
• [0006]
  It has been suggested [D. H. Bentley, et. al., Tetrahedron Lett., 1976, 41, 3739] that the isomerisation results from the ability of the 4-carboxylate anion of the starting carboxylic acid to abstract a proton from the 2-position of the 3-cephem-4-carboxylic acid ester formed, followed by reprotonation at 4-position to give the said Δ²-ester. It has also been suggested [R. B. Morin, et. al., J. Am. Chem. Soc., 1969, 91, 1401 ; R. B. Woodward, et. al., J. Am. Chem. Soc., 1966, 88, 852] that the equilibrium position for isomerisation is largely determined by the size of the ester addendum attached at the 4-carboxylic acid position.
• [0007]
  The 2-cephem-4-carboxylic acid esters of formula (II A) are not only unreactive as antibacterial agents but are undesired by-products. Pharmacopoeias of many countries are very stringent about the presence of the 2-cephem analogues in the finished sample of (3,7)-substituted-3-cephem-4-carboxylic acid esters and set limits for the permissible amounts of these isomers. Due to the structural similarity of the 2-cephem and 3-cephem analogues it is very difficult to separate the two isomers by conventional methods, such as chromatography as well as by fractional crystallisation. In addition to this removal of other unidentified impurities formed in the reaction, entails utilisation of tedious purification methods, thus overall resulting in,

  1. a) considerable loss in yield, increasing the cost of manufacture and
  2. b) a product of quality not conforming to and not easily amenable for upgradation to pharmacopoeial standards.
• [0008]
  Several methods are reported in the prior art for synthesis of cefuroxime axetil of formula (I) and various (3,7)-substituted-3-cephem-4-carboxylic acid esters of formula (I A), with attempts to minimise the unwanted Δ²-isomers formed in such reactions as well as conversion of the Δ²-isomer thus formed back to the desired Δ³- isomer. The prior art methods can be summarised as follows:

  • i) US Patent No, 4 267 320 (Gregson et. al.) describes a method for synthesis of cefuroxime axetil comprising reaction of cefuroxime acid or its alkali metal salts or onium salts with (R,S)-1-acetoxyethyl bromide in an inert organic solvent selected from N,N-dimethylacetamide, N,N-dimethylformamide, dimethyl sulfoxide, acetone, acetonitrile and hexamethylphosphoric triamide at a temperature in the range of -50 to +1150° C. The patent mentions that when alkali metal salts, specially potassium salt of cefuroxime acid are employed the reaction can be carried out in a nitrile solvent in the presence of a crown ether. When cefuroxime acid is employed the reaction is carried out in the presence of a weak inorganic base such as sodium carbonate or potassium carbonate, which is added prior to the addition of the haloester. The patent further mentions that the use of potassium carbonate in conjunction with the haloester, specially the bromo or iodo ester is preferred since it helps to minimise the formation of the Δ²-isomer. Ideally, substantially equivalent amounts of cefuroxime acid and the base is employed.
    The US Patent No. 4 267 320 also describes methods, wherein the said esterification is carried out in the presence of an acid binding agent, which serve to bind hydrogen halide liberated in the reaction, thereby controlling the formation of the Δ²-isomer. The acid binding agents that are utilised include a tertiary amine base such as triethylamine or N, N-dimethylamine ; an inorganic base such as calcium carbonate or sodium bicarbonate and an oxirane compound such as ethylene oxide or propylene oxide.
    However, from the examples provided in the above patent the yield of cefuroxime axetil and other (3,7)-substituted-3-cephem-4-carboxylic acid esters obtained is found to be only of about 50%, implying formation of substantial amounts of impurities in the reaction. Indeed, when cefuroxime acid is reacted with (R,S)-1-acetoxyethyl bromide in the presence of 0.55 molar equivalents of sodium carbonate or potassium carbonate in N,N-dimethylacetamide as solvent, as per the process disclosed in this patent, it is found that substantial amounts of the Δ²-isomer in a proportion ranging from 10-22% is formed, in addition to other unknown impurities. Also, substantial amounts of the starting cefuroxime acid remains unreacted even after 5 hrs of reaction. Isolation of the product generally affords a gummy material, which resists purification even after repeated crystallisations.
    Moreover, the use of the acid binding agents mentioned in the above patent, specially tertiary amines and inorganic bases lead to cleavage of the β-lactam ring and also promote the undesired Δ²-isomerisation, thereby enhancing the level of impurities formed in the reaction.
  • ii) GB Patent No. 2 218 094 describes a method by which the Δ²-isomers formed during esterification can be converted back to the desired Δ³-isomers. The method comprises of oxidation of the dihydrothiazine ring in the mixture of Δ²- and Δ³- cephalosporin acid esters to the corresponding sulfoxide derivatives with suitable oxidising agents, whereby the Δ²-isomer gets isomerised to the corresponding Δ³-isomer during oxidation and the Δ³- cephalosporin acid ester sulfoxide is isolated. The sulfide group is regenerated back by reduction of the sulfoxide function with suitable reducing agents.
    Typically, the oxidation is carried out using m-chloroperbenzoic acid and the reduction achieved by use of an alkali metal halide in presence of acetyl chloride in presence of an inert organic solvent or by use of a phosphorous trihalide.
    Although, this method provides the desired Δ³-isomers in good purity, it cannot be considered as an industrially feasible method since it involves a two step process of oxidation and reduction, isolation of the intermediate products at each stage and necessary purifications, all resulting in considerable loss of the desired product and increase in the cost of manufacture. Moreover, the use of acetyl halide and phosphorous trihalide in the reduction step cannot be applied to cephalosporin derivatives that are sensitive to these reagents.
    A similar method has been reported by Kaiser et. al. in J. Org. Chem., 1970, 35, 2430.
  • (iii)Mobasherry et. al. in J. Org. Chem., 1986, 51, 4723 describe preparation of certain Δ³-cephalosporin-4-carboxylic acid esters by reaction of the corresponding 3-cephem-4-carboxylic acids (in turn prepared form the corresponding carboxylic acid alkali metal salts) with an haloester in presence of 1.1 eq of sodium carbonate in the presence 1.2-1.5 eq of an alkyl halide and in presence of a solvent comprising of a mixture of N,N-dimethylformamide and dioxane. The authors claim that the method provides of Δ³- cephalosporin-4-carboxylic acid esters unaccompanied by the corresponding Δ²-isomer.
    However, the method involves an additional step in that the starting 3-cephem-4-carboxylic acid ester derivatives are obtained from the corresponding alkali metal salts prior to reaction. In addition, longer reaction times of about 24 hrs coupled with the fact that it utilises dioxane, a potent carcinogen, not recommended by International Conference on Harmonisation (ICH) on industrial scale renders the method unattractive commercially.
    Moreover, on duplication of the method exactly as described in the article it is found that about 3-4% of the corresponding Δ²-isomer is indeed formed in the reaction in addition to other unidentified impurities. Also, substantial amounts of the starting cephalosporin carboxylic acid is recovered unreacted.
  • (iv)Shigeto et. al. in Chem. Pharm. Bull., 1995, 43(11), 1998 have carried out the esterification of certain 7-substituted-3-cephem-4-carboxylic acid derivatives with 1-iodoethyl isopropyl carbonate in a solvent system containing a mixture of N, N-dimethylformamide and dioxane in a 3:5 ratio. A conversion to the corresponding 3-cephem- 4-carboxylate ester was achieved in only 34%, out of which the Δ²-isomer amounted to about 8%.
    Esterification of 7-formamido-3-(N,N-dimethylcarbamoyloxy)methyl-3-cephem-4-carboxylic acid sodium salt with a suitable haloester in presence of solvents such as N, N-dimethylacetamide and N, N-dimethylformamide, with formation of about 0.8 to 3.0% of the Δ²-isomer is also reported in the above article by Shigeto et. al. The 7-formamido group was cleaved under acidic conditions to give the corresponding 7-amino derivative contaminated with only about 0.4% of the corresponding Δ²-isomer. The minimisation of the percentage of Δ²-isomer is attributed to the relative unstability of 7-amino-2-cephem-4-carboxylic acid esters in acidic conditions, facilitating isomerisation of the 2-cephem intermediate to the 3-cephem derivative.
    However, the method does not have a general application, especially for synthesis of commercially valuable cephalosporin derivatives containing hydroxyimino or alkoxyimino substituents in the 7-amino side chain addendum, since these oxyimino functions exhibit a tendency to isomerise from the stable (Z)-configuration to the relatively undesirable(E)-configuration under acidic conditions. This would render separation of the two isomers cumbersome. Moreover, longer reaction times of about 18-20 hrs to effect the isomerisation of the double bond from the 2- position to the 3-position and use of toxic dioxane as solvent impose further limitations on the method.
    (v) Demuth et. al. in J. Antibiotics, 1991, 44, 200 have utilised the N, N-dimethylformamide-dioxane system in the coupling of 1-iodocephem-4-nitrobenzyl ester with naldixic acid sodium salt and recommend use of dioxane since it reduces the basicity of the quinolone carboxylate and lowers the polarity of the reaction medium.
    However, low yields of about 35% and use of toxic dioxane makes the method of little industrial application.
  • (vi) Wang et. al. in US Patent No. 5 498 787 claim a method for preparation of certain (3,7)-substituted-3-cephem-4-carboxylic acid prodrug esters, unaccompanied by the analogous 2-cephem esters comprising reaction of the corresponding (3,7)-substituted-3-cephem-4-carboxylic acid alkali metal salts with suitable haloesters in the presence of catalytic amounts of a quaternary ammonium or quarternary phosphonium salt. Among the prodrug esters covered in this patent is cefuroxime axetil.
    US Patent No. 5 498 787 claims that among the quarternary ammonium salts, such salts with acid counter ion, specially tetrabutyl ammonium sulfate (TBA⁺HSO₄ ^-) is the most preferred. When the molar ratio of TBA⁺HSO₄ ^-/cefuroxime sodium was above 0.40 no Δ²-isomer was detected, when the said molar ratio was below 0.40 and near about 0.20 the molar ratio of Δ²/Δ³ isomers formed was about 2.0%. When no TBA⁺HSO₄ ^- was added the molar ratio of Δ²/Δ³ isomers formed was about 10.0%. Examples 1 and 2 of this patent illustrate the esterification of cefuroxime sodium in presence of TBA⁺HSO₄ ^- and indicate that the Δ²-isomer was not detected after 3-12 hours of reaction. The same patent also establishes the superiority of TBA⁺HSO₄ ^- over other salts, specially tetrabutyl ammonium iodide (TBA⁺I^-) since use of the latter salt resulted in considerable isomerisation of the double bond giving the undesired Δ²-isomer in predominant amounts.
    The present inventors have, however, found that when cefuroxime sodium is reacted with (R,S)- 1-acetoxyethyl bromide in the presence of tetrabutylammonium sulfate (TBA⁺HSO₄ ^-) as per the method covered in US Patent No. 5,498 787 the same did not necessarily result in the production of the desired Δ³isomer free of the undesired Δ² isomer and other impurities. Also, such process had limitations in that the reaction could not be completed at times even at the end of 5.0hrs. Moreover, the separation of the impurities; from the product proved cumbersome and could not be removed from the product even after successive crystallisations.
  • (vii) H. W. Lee et. al., Syntheic Communications, 1998, 28(23), 4345-4354 have demonstrated a method essentially similar to that claimed in US Patent No. 5 498 787 . The method of preparation of various esters of cefotaxime consists of reacting cefotaxime sodium with the requisite haloester compound in a suitable solvent and in presence of quarternary ammonium salts as phase transfer catalysts. It is claimed that when no quarternary ammonium salts are added the molar ratio (%) of Δ²/Δ³ isomers formed is about 10%. The formation of Δ²- isomer is minimised when quarternary ammonium salts are added and particularly when the molar ratio of TBA⁺HSO₄ ^-/cefotaxime sodium employed is 0.80 the formation of the Δ²- isomer is completely inhibited.
    However, this method requires long hours (~18-24 hrs) and is carried out at higher temperatures (40-45° C) and as such may not be suitable for cephalosporin derivatives that are sensitive to heat.
  • (viii)H. W. Lee et. al. in Synthetic Communications, 1999, 29(11), 1873-1887 demonstrate a method for preparation of number of (3,7)-substituted-3-cephem-4-carboxylic acid esters comprising reacting the corresponding (3,7)-substituted-3-cephem-4-carboxylic acid derivatives with a base selected form cesium carbonate or cesium bicarbonate either used alone or in combination with potassium carbonate, sodium carbonate, potassium bicarbonate and sodium bicarbonate. The authors established that the formation of Δ²- isomers could be minimised by utilisation of a solvent combination ofN, N-dimethyl formamide and dioxane. The use of the latter mentioned solvent i. e. dioxane was expected to lower polarity of the reaction medium and thereby reduce the basicity of the transient 3-cephem-4-carboxylate anion formed in the reaction and thus preventing the isomerisation of the double bond from the 3-4 position to the 2-3 position.
    The formation of the Δ²- isomer was found to be dependent on the amount of dioxane in the solvent mixture, the more the proportion of dioxane lesser the degree of isomerisation.
    However, yields of representative esters obtained by the method are in the range of 45-85 %, implying that the reaction is accompanied by formation of substantial amounts of impurities and that the isomerisation is dependent on the nature of the substituent at 3α-position of the cephalosporin nucleus as well as on the nature of the haloester employed. Moreover, the method utilises dioxane, not desirable for reasons mentioned herein earlier and expensive cesium salts. This method, therefore, also has limited application.
  • (ix) Y.S. Cho et. al., in Korean J. Med. Chem., 1995, 5(1), 60-63 describe synthesis of several cephalosporin prodrug esters and their efficacy on oral administration. The esters were synthesised by reacting the corresponding cephalosporin-4-carboxylic acid derivative with the respective haloester derivative in presence of cesium carbonate and N, N-dimethylacetamide. The yields of the ester derivatives obtained are in the range of only 25-56%, indicating formation of substantial amounts of impurities in the reaction.

Example – 1

Preparation of (R, S -1-Acetoxyethyl-3-carbamoyloxymethyl-7-[(Z)-2-(fur-2-yl)-2-methoxyiminoacetamido]ceph-3-em-4-carboxylate (Cefuroxime axetil, I) :

Without use of GrouplI

/

II metal phosphate and C_1-4 alcohol

• [0045]
  (R, S)-1-Acetoxyethyl bromide (1.6gms; 0.0094moles) was added to a mixture of cefuroxime acid (2gms; 0.0047moles) and potassium carbonate (0.326gms; 0.00235moles) in N,N-dimethylacetamide (10 ml) at 5°C and stirred at 0 to 20° C for 180 minutes Ethyl acetate was added to the reaction mixture, followed by 3% aqueous sodium bicarbonate solution (15ml). The organic layer containing the title product, Δ² isomer (8.51%) and unidentified impurities (X₁-1.86% and X₂ – 3.54%) was separated and washed with 10% aqueous NaCl solution. The organic solvent was evaporated off under vacuum to give 1.08gms (44.90%) of the title compound as a gummy solid.
• [0046]
  HPLC analysis : Purity (compound I) – 89.11% ; Impurities : Δ² isomer (II) – 8.51%, X₁ – 1.86% and X₂ – 3.54%

………………………………..

The reaction of 3-hydroxymethyl-7- [2- (2-furyl) -2-methoxyiminoacetamido] -3-cephem-4-carboxylic acid (I) with chlorosulfonyl isocyanate (II) and then with sodium 2-ethylhexanoate gives sodium cefuroxime (III), which is then treated with 1-bromoethyl acetate (IV) in DMA.

References

EMA grants orphan drug designations to Alnylam’s ALN-AT3 for haemophilia treatment
Biopharmaceutical company Alnylam Pharmaceuticals has received orphan drug designations for ALN-AT3 from the European Medicines Agency (EMA) Committee to treat haemophilia A and B

SEE

http://www.pharmaceutical-technology.com/news/newsema-grants-orphan-drug-designations-alnylams-aln-at3-hemophilia-treatment-4338904?WT.mc_id=DN_News

 

May 13,2014

Alnylam Pharmaceuticals, Inc., a leading RNAi therapeutics company, announced today positive top-line results from its ongoing Phase 1 trial of ALN-AT3, a subcutaneously administered RNAi therapeutic targeting antithrombin (AT) in development for the treatment of hemophilia and rare bleeding disorders (RBD). These top-line results are being presented at the World Federation of Hemophilia (WFH) 2014 World Congress being held May 11 – 15, 2014 in Melbourne, Australia. In Part A of the Phase 1 study, human volunteer subjects received a single subcutaneous dose of ALN-AT3 and, per protocol, the maximum allowable level of AT knockdown was set at 40%. Initial results show that a single, low subcutaneous dose of ALN-AT3 at 0.03 mg/kg resulted in an up to 28-32% knockdown of AT at nadir that was statistically significant relative to placebo (p < 0.01 by ANOVA). This led to a statistically significant (p < 0.01) increase in peak thrombin generation, that was temporally associated and consistent with the degree of AT knockdown. ALN-AT3 was found to be well tolerated with no significant adverse events reported. With these data, the company has transitioned to the Multiple Ascending Dose (MAD) Part B of the study in moderate-to-severe hemophilia subjects. Consistent with previous guidance, the company plans to present initial clinical results from the Phase 1 study, including results in hemophilia subjects, by the end of the year. These human study results are the first to be reported for Alnylam’s Enhanced Stabilization Chemistry (ESC)-GalNAc conjugate technology, which enables subcutaneous dosing with increased potency, durability, and a wide therapeutic index. Further, these initial clinical results demonstrate a greater than 50-fold potency improvement with ESC-GalNAc conjugates relative to standard template chemistry conjugates.

“We are excited by these initial positive results for ALN-AT3 in the human volunteer ‘Part A’ of our Phase 1 study. Indeed, within the protocol-defined boundaries of single doses that provide no more than a 40% knockdown of AT in normal subjects, we were able to demonstrate a statistically-significant knockdown of AT of up to 28-32% and an associated increase in thrombin generation. Remarkably, this result was achieved at the lowest dose tested of 0.03 mg/kg, demonstrating a high and better than expected level of potency for ALN-AT3, our first ESC-GalNAc conjugate to enter clinical development,” said Akshay Vaishnaw, M.D., Ph.D., Executive Vice President and Chief Medical Officer of Alnylam. “With these results in hand, we are now proceeding to ‘Part B’ of the study, where we will administer multiple ascending doses to up to 18 patients with moderate-to-severe hemophilia A or B. Patients will receive three weekly doses, and we fully expect to achieve robust levels of AT knockdown as we dose escalate. In addition, we will aim to evaluate a once-monthly dosing regimen in future clinical studies, as we believe this could provide a highly attractive prophylactic regimen for patients. We look forward to sharing our detailed Phase 1 results, including data in hemophilia subjects, later this year, consistent with our original guidance.”

“There are several notable implications of these exciting initial results with ALN-AT3. First, ALN-AT3 now becomes the fourth program in our ‘Alnylam 5×15’ pipeline to demonstrate clinical activity. As such, these results increase our confidence level yet further across the entirety of our pipeline efforts, where we remain focused on genetically defined, liver-expressed disease targets with a modular and reproducible delivery platform. Moreover, these results with ALN-AT3 establish human proof of concept for our ESC-GalNAc conjugate technology, extending and broadening the human results we have previously shown with ALN-TTRsc which employs our standard template chemistry. Our ESC-GalNAc conjugate technology enables subcutaneous dosing with increased potency and durability and a wide therapeutic index, and has now become our primary approach for the delivery of RNAi therapeutics,” said John Maraganore, Ph.D., Chief Executive Officer of Alnylam. “Finally, the achievement of target knockdown at such a low dose of 0.03 mg/kg is unprecedented. Based on our evaluation of datasets from non-human primate (NHP) and human studies, these results demonstrate a 10-fold improved potency for ALN-AT3 as compared with NHP and a 50-fold improved potency in humans as compared with ALN-TTRsc. Based on data we announced earlier this week at TIDES, we believe that this increased potency is the combined result of enhanced stability for ESC-GalNAc conjugates and an attenuated nuclease environment in human tissue compared with other species. If these results extend to other ESC-GalNAc-siRNA conjugates, such as those in our complement C5 and PCSK9 programs, we believe we can expect highly potent clinical activities with very durable target knockdown effects.”

The ongoing Phase 1 trial of ALN-AT3 is being conducted in the U.K. as a single- and multi-dose, dose-escalation study comprised of two parts. Part A – which has now been completed – was a randomized, single-blind, placebo-controlled, single-dose, dose-escalation study, intended to enroll up to 24 healthy volunteer subjects. The primary objective of this part of the study was to evaluate the safety and tolerability of a single dose of ALN-AT3, with the potential secondarily to show changes in AT plasma levels at sub-pharmacologic doses. This part of the study evaluated only low doses of ALN-AT3, with a dose-escalation stopping rule at no more than a 40% level of AT knockdown. Based on the pharmacologic response achieved in this part of the study, only the lowest dose cohort (n=4; 3:1 randomization of ALN-AT3:placebo) was enrolled. Part B of the study is an open-label, multi-dose, dose-escalation study enrolling up to 18 people with moderate-to-severe hemophilia A or B. The primary objective of this part of the study is to evaluate the safety and tolerability of multiple doses, specifically three doses, of subcutaneously administered ALN-AT3 in hemophilia subjects. Secondary objectives include assessment of clinical activity as determined by knockdown of circulating AT levels and increase in thrombin generation at pharmacologic doses of ALN-AT3; thrombin generation is known to be a biomarker for bleeding frequency and severity in people with hemophilia (Dargaud, et al., Thromb Haemost; 93, 475-480 (2005)). In this part of the study, dose-escalation will be allowed to proceed beyond the 40% AT knockdown level.

In addition to reporting positive top-line results from the Phase 1 trial with ALN-AT3, Alnylam presented new pre-clinical data with ALN-AT3. First, in a saphenous vein bleeding model performed in hemophilia A (HA) mice, a single subcutaneous dose of ALN-AT3 that resulted in an approximately 70% AT knockdown led to a statistically significant (p < 0.0001) improvement in hemostasis compared to saline-treated HA mice. The improved hemostasis was comparable to that observed in HA mice receiving recombinant factor VIII. These are the first results in what can be considered a genuine bleeding model showing that AT knockdown with ALN-AT3 can control bleeding. Second, a number of in vitro studies were performed in plasma from hemophilia donors. Stepwise AT depletion in these plasma samples was shown to achieve stepwise increases in thrombin generation. Furthermore, it was shown that a 40-60% reduction of AT resulted in peak thrombin levels equivalent to those achieved with 10-15% levels of factor VIII in HA plasma and factor IX in hemophilia B (HB) plasma. These levels of factor VIII or IX are known to significantly reduce bleeding in hemophilia subjects. As such, these results support the hypothesis that a 40-60% knockdown of AT with ALN-AT3 could be fully prophylactic. Finally, a modified Activated Partial Thromboplastin Time (APTT) assay – an ex vivomeasure of blood coagulation that is significantly prolonged in hemophilia – was developed, demonstrating sensitivity to AT levels. Specifically, depletion of AT in HA plasma led to a shortening of modified APTT. This modified APTT assay can be used to routinely and simply monitor functional activity of AT knockdown in further ALN-AT3 clinical studies.

“The unmet need for new therapeutic options to treat hemophilia patients remains very high, particularly in those patients who experience multiple annual bleeds such as patients receiving replacement factor ‘on demand’ or patients who have developed inhibitory antibodies. Indeed, I believe the availability of a safe and effective subcutaneously administered therapeutic with a long duration of action would represent a marked improvement over currently available approaches for prophylaxis,” said Claude Negrier, M.D., head of the Hematology Department and director of the Haemophilia Comprehensive Care Centre at Edouard Herriot University Hospital in Lyon. “I continue to be encouraged by Alnylam’s progress to date with ALN-AT3, including these initial data reported from the Phase 1 trial showing statistically significant knockdown of antithrombin and increased thrombin generation, which has been shown to correlate with bleeding frequency and severity in hemophilia. I look forward to the advancement of this innovative therapeutic candidate in hemophilia subjects.”

About Hemophilia and Rare Bleeding Disorders

Hemophilias are hereditary disorders caused by genetic deficiencies of various blood clotting factors, resulting in recurrent bleeds into joints, muscles, and other major internal organs. Hemophilia A is defined by loss-of-function mutations in Factor VIII, and there are greater than 40,000 registered patients in the U.S. and E.U. Hemophilia B, defined by loss-of-function mutations in Factor IX, affects greater than 9,500 registered patients in the U.S. and E.U. Other Rare Bleeding Disorders (RBD) are defined by congenital deficiencies of other blood coagulation factors, including Factors II, V, VII, X, and XI, and there are about 1,000 patients worldwide with a severe bleeding phenotype. Standard treatment for hemophilia patients involves replacement of the missing clotting factor either as prophylaxis or on-demand therapy. However, as many as one third of people with severe hemophilia A will develop an antibody to their replacement factor – a very serious complication; these ‘inhibitor’ patients become refractory to standard replacement therapy. There exists a small subset of hemophilia patients who have co-inherited a prothrombotic mutation, such as Factor V Leiden, antithrombin deficiency, protein C deficiency, and prothrombin G20210A. Hemophilia patients that have co-inherited these prothrombotic mutations are characterized as having a later onset of disease, lower risk of bleeding, and reduced requirements for Factor VIII or Factor IX treatment as part of their disease management. There exists a significant need for novel therapeutics to treat hemophilia patients.

About Antithrombin (AT)

Antithrombin (AT, also known as “antithrombin III” and “SERPINC1″) is a liver expressed plasma protein and member of the “serpin” family of proteins that acts as an important endogenous anticoagulant by inactivating Factor Xa and thrombin. AT plays a key role in normal hemostasis, which has evolved to balance the need to control blood loss through clotting with the need to prevent pathologic thrombosis through anticoagulation. In hemophilia, the loss of certain procoagulant factors (Factor VIII and Factor IX, in the case of hemophilia A and B, respectively) results in an imbalance of the hemostatic system toward a bleeding phenotype. In contrast, in thrombophilia (e.g., Factor V Leiden, protein C deficiency, antithrombin deficiency, amongst others), certain mutations result in an imbalance in the hemostatic system toward a thrombotic phenotype. Since co-inheritance of prothrombotic mutations may ameliorate the clinical phenotype in hemophilia, inhibition of AT defines a novel strategy for improving hemostasis.

About GalNAc Conjugates and Enhanced Stabilization Chemistry (ESC)-GalNAc Conjugates

GalNAc-siRNA conjugates are a proprietary Alnylam delivery platform and are designed to achieve targeted delivery of RNAi therapeutics to hepatocytes through uptake by the asialoglycoprotein receptor. Alnylam’s Enhanced Stabilization Chemistry (ESC)-GalNAc-conjugate technology enables subcutaneous dosing with increased potency and durability, and a wide therapeutic index. This delivery platform is being employed in several of Alnylam’s genetic medicine programs, including programs in clinical development.

About RNAi

RNAi (RNA interference) is a revolution in biology, representing a breakthrough in understanding how genes are turned on and off in cells, and a completely new approach to drug discovery and development. Its discovery has been heralded as “a major scientific breakthrough that happens once every decade or so,” and represents one of the most promising and rapidly advancing frontiers in biology and drug discovery today which was awarded the 2006 Nobel Prize for Physiology or Medicine. RNAi is a natural process of gene silencing that occurs in organisms ranging from plants to mammals. By harnessing the natural biological process of RNAi occurring in our cells, the creation of a major new class of medicines, known as RNAi therapeutics, is on the horizon. Small interfering RNA (siRNA), the molecules that mediate RNAi and comprise Alnylam’s RNAi therapeutic platform, target the cause of diseases by potently silencing specific mRNAs, thereby preventing disease-causing proteins from being made. RNAi therapeutics have the potential to treat disease and help patients in a fundamentally new way.

About Alnylam Pharmaceuticals

Alnylam is a biopharmaceutical company developing novel therapeutics based on RNA interference, or RNAi. The company is leading the translation of RNAi as a new class of innovative medicines with a core focus on RNAi therapeutics as genetic medicines, including programs as part of the company’s “Alnylam 5x15TM” product strategy. Alnylam’s genetic medicine programs are RNAi therapeutics directed toward genetically defined targets for the treatment of serious, life-threatening diseases with limited treatment options for patients and their caregivers. These include: patisiran (ALN-TTR02), an intravenously delivered RNAi therapeutic targeting transthyretin (TTR) for the treatment of TTR-mediated amyloidosis (ATTR) in patients with familial amyloidotic polyneuropathy (FAP); ALN-TTRsc, a subcutaneously delivered RNAi therapeutic targeting TTR for the treatment of ATTR in patients with TTR cardiac amyloidosis, including familial amyloidotic cardiomyopathy (FAC) and senile systemic amyloidosis (SSA); ALN-AT3, an RNAi therapeutic targeting antithrombin (AT) for the treatment of hemophilia and rare bleeding disorders (RBD); ALN-CC5, an RNAi therapeutic targeting complement component C5 for the treatment of complement-mediated diseases; ALN-AS1, an RNAi therapeutic targeting aminolevulinate synthase-1 (ALAS-1) for the treatment of hepatic porphyrias including acute intermittent porphyria (AIP); ALN-PCS, an RNAi therapeutic targeting PCSK9 for the treatment of hypercholesterolemia; ALN-AAT, an RNAi therapeutic targeting alpha-1 antitrypsin (AAT) for the treatment of AAT deficiency-associated liver disease; ALN-TMP, an RNAi therapeutic targeting TMPRSS6 for the treatment of beta-thalassemia and iron-overload disorders; ALN-ANG, an RNAi therapeutic targeting angiopoietin-like 3 (ANGPTL3) for the treatment of genetic forms of mixed hyperlipidemia and severe hypertriglyceridemia; ALN-AC3, an RNAi therapeutic targeting apolipoprotein C-III (apoCIII) for the treatment of hypertriglyceridemia; and other programs yet to be disclosed. As part of its “Alnylam 5×15” strategy, as updated in early 2014, the company expects to have six to seven genetic medicine product candidates in clinical development – including at least two programs in Phase 3 and five to six programs with human proof of concept – by the end of 2015. Alnylam is also developing ALN-HBV, an RNAi therapeutic targeting the hepatitis B virus (HBV) genome for the treatment of HBV infection. The company’s demonstrated commitment to RNAi therapeutics has enabled it to form major alliances with leading companies including Merck, Medtronic, Novartis, Biogen Idec, Roche, Takeda, Kyowa Hakko Kirin, Cubist, GlaxoSmithKline, Ascletis, Monsanto, The Medicines Company, and Genzyme, a Sanofi company. In March 2014, Alnylam acquired Sirna Therapeutics, a wholly owned subsidiary of Merck. In addition, Alnylam holds an equity position in Regulus Therapeutics Inc., a company focused on discovery, development, and commercialization of microRNA therapeutics. Alnylam scientists and collaborators have published their research on RNAi therapeutics in over 200 peer-reviewed papers, including many in the world’s top scientific journals such as Nature, Nature Medicine, Nature Biotechnology, Cell, the New England Journal of Medicine, and The Lancet. Founded in 2002, Alnylam maintains headquarters in Cambridge, Massachusetts. For more information, please visit www.alnylam.com.

Mocetinostat

SAN DIEGO, Aug. 11, 2014 /PRNewswire/ — Mirati Therapeutics, Inc. (NASDAQ: MRTX) today announced that the U.S. FDA has granted Orphan Drug Designation to mocetinostat, a spectrum selective HDAC inhibitor, for diffuse large B-cell lymphoma (DLBCL). In June, mocetinostat was granted Orphan Drug Designation as a treatment for myelodysplastic syndrome (MDS).  Orphan drug designation is also being sought for bladder cancer patients with specific genetic alterations.

http://www.prnewswire.com/news-releases/mirati-therapeutics-receives-orphan-designation-from-us-food–drug-administration-for-mocetinostat-in-diffuse-large-b-cell-lymphoma-270737161.html

Mocetinostat (MGCD0103) is a benzamide histone deacetylase inhibitor undergoing clinical trials for treatment of various cancers including follicular lymphoma, Hodgkin’s lymphoma and acute myelogenous leukemia.^[1][2][3]

One clinical trial (for refractory follicular lymphoma) was temporarily put on hold due to cardiac problems but resumed recruiting in 2009.^[4]

In 2010 favourable results were announced from the phase II trial for Hodgkin’s lymphoma.^[5]

MGCD0103 has also been used as a research reagent where blockage of members of the HDAC-family of histone deacetylases is required.^[6]

Mechanism of action

It works by inhibiting mainly histone deacetylase 1 (HDAC1), but also HDAC2, HDAC3, and HDAC11.^[7]

About Mocetinostat

Mocetinostat is an orally-bioavailable, spectrum-selective HDAC inhibitor. Mocetinostat is enrolling patients in a Phase 2 dose confirmation study in combination with Vidaza as treatment for intermediate and high-risk MDS. Mirati also plans to initiate Phase 2 studies of mocetinostat as a single agent in patients with mutations in histone acetyl transferases in bladder cancer and DLBCL. Initial data from the Phase 2 studies is expected by the end of 2014. In addition to the ongoing Phase 2 clinical trials, mocetinostat has completed 13 clinical trials in more than 400 patients with a variety of hematologic malignancies and solid tumors.

About Mirati Therapeutics

Mirati Therapeutics is a targeted oncology company developing an advanced pipeline of breakthrough medicines for precisely defined patient populations. Mirati’s approach combines the three most important factors in oncology drug development – drug candidates with complementary and compelling targets, creative and agile clinical development, and a highly accomplished precision medicine leadership team. The Mirati team is using a proven blueprint for developing targeted oncology medicines to advance and maximize the value of its pipeline of drug candidates, including MGCD265 and MGCD516, which are orally bioavailable, multi-targeted kinase inhibitors with distinct target profiles, and mocetinostat, an orally bioavailable, spectrum-selective histone deacetylase inhibitor. More information is available at www.mirati.com.

In eukaryotic cells, nuclear DNA associates with histones to form a compact complex called chromatin. The histones constitute a family of basic proteins which are generally highly conserved across eukaryotic species. The core histones, termed H2A, H2B, H3, and H4, associate to form a protein core. DNA winds around this protein core, with the basic amino acids of the histones interacting with the negatively charged phosphate groups of the DNA. Approximately 146 base pairs of DNA wrap around a histone core to make up a nucleosome particle, the repeating structural motif of chromatin.

Csordas, Biochem. J., 286: 23-38 (1990) teaches that histones are subject to posttranslational acetylation of the α,ε-amino groups of N-terminal lysine residues, a reaction that is catalyzed by histone acetyl transferase (HAT1). Acetylation neutralizes the positive charge of the lysine side chain, and is thought to impact chromatin structure. Indeed, Taunton et al., Science, 272: 408-411 (1996), teaches that access of transcription factors to chromatin templates is enhanced by histone hyperacetylation. Taunton et al. further teaches that an enrichment in underacetylated histone H4 has been found in transcriptionally silent regions of the genome.

Histone acetylation is a reversible modification, with deacetylation being catalyzed by a family of enzymes termed histone deacetylases (HDACs). Grozinger et al., Proc. Natl. Acad. Sci. USA, 96: 4868-4873 (1999), teaches that HDACs are divided into two classes, the first represented by yeast Rpd3-like proteins, and the second represented by yeast Hda1-like proteins. Grozinger et al. also teaches that the human HDAC1, HDAC2, and HDAC3 proteins are members of the first class of HDACs, and discloses new proteins, named HDAC4, HDAC5, and HDAC6, which are members of the second class of HDACs. Kao et al., Genes & Dev., 14: 55-66 (2000), discloses HDAC7, a new member of the second class of HDACs. More recently, Hu et al. J. Bio. Chem. 275:15254-13264 (2000) and Van den Wyngaert, FEBS, 478: 77-83 (2000) disclose HDAC8, a new member of the first class of HDACs.

Richon et al., Proc. Natl. Acad. Sci. USA, 95: 3003-3007 (1998), discloses that HDAC activity is inhibited by trichostatin A (TSA), a natural product isolated from Streptomyces hygroscopicus, and by a synthetic compound, suberoylanilide hydroxamic acid (SAHA). Yoshida and Beppu, Exper. Cell Res., 177: 122-131 (1988), teaches that TSA causes arrest of rat fibroblasts at the G₁ and G₂ phases of the cell cycle, implicating HDAC in cell cycle regulation. Indeed, Finnin et al., Nature, 401: 188-193 (1999), teaches that TSA and SAHA inhibit cell growth, induce terminal differentiation, and prevent the formation of tumors in mice. Suzuki et al., U.S. Pat. No. 6,174,905, EP 0847992, JP 258863/96, and Japanese Application No. 10138957, disclose benzamide derivatives that induce cell differentiation and inhibit HDAC. Delorme et al., WO 01/38322 and PCT/IB01/00683, disclose additional compounds that serve as HDAC inhibitors.

The molecular cloning of gene sequences encoding proteins with HDAC activity has established the existence of a set of discrete HDAC enzyme isoforms. Some isoforms have been shown to possess specific functions, for example, it has been shown that HDAC-6 is involved in modulation of microtubule activity. However, the role of the other individual HDAC enzymes has remained unclear.

These findings suggest that inhibition of HDAC activity represents a novel approach for intervening in cell cycle regulation and that HDAC inhibitors have great therapeutic potential in the treatment of cell proliferative diseases or conditions. To date, few inhibitors of histone deacetylase are known in the art.

………………..

http://www.google.com/patents/WO2011112623A1?cl=en

Mocetinostat (MGCD-0103)

N-(2-aminophenyl)-4-[[(4-pyridin-3-ylpyrimidin-2-yl)amino]methyl^^

…………………………

http://www.google.co.in/patents/US6897220

Example 426 Synthesis of N-(2-Amino-phenyl)-4-[(4-pyridin-3-pyrimidin-2-ylamino)-methyl]-benzamide

Step 1: Synthesis of 4-Guanidinomethyl-benzoic acid methyl ester Intermediate 1

The mixture of 4-Aminomethyl-benzoic acid methyl ester HCl (15.7 g, 77.8 mmol) in DMF (85.6 mL) and DIPEA (29.5 mL, 171.2 mmol) was stirred at rt for 10 min. Pyrazole-1-carboxamidine HCl (12.55 g, 85.6 mmol) was added to the reaction mixture and then stirred at rt for 4 h to give clear solution. The reaction mixture was evaporated to dryness under vacuum. Saturated NaHCO₃ solution (35 mL) was added to give nice suspension. The suspension was filtered and the filter cake was washed with cold water. The mother liquid was evaporated to dryness and then filtered. The two solids were combined and re-suspended over distilled H₂O (50 ml). The filter cake was then washed with minimum quantities of cold H₂O and ether to give 12.32 g white crystalline solid intermediate 1 (77% yield, M+1: 208 on MS).

Step 2: Synthesis of 3-Dimethylamino-1-pyridin-3-yl-propenone Intermediate 2

3-Acetyl-pyridine (30.0 g, 247.6 mmol) and DMF dimethyl acetal (65.8 mL, 495.2 mmol) were mixed together and then heated to reflux for 4 h. The reaction mixture was evaporated to dryness and then 50 mL diethyl ether was added to give brown suspension. The suspension was filtered to give 36.97 g orange color crystalline product (85% yield, M+1: 177 on MS).

Step 3: Synthesis of 4-[(4Pyridin-3-pyrimidin-2-ylamino)-methyl]benzoic acid methyl ester Intermediate 3

Intermediate 1 (0.394 g, 1.9 mmol) and intermediate 2 (0.402 g, 2.3 mmol) and molecular sieves (0.2 g, 4A, powder, >5 micron) were mixed with isopropyl alcohol (3.8 mL). The reaction mixture was heated to reflux for 5 h. MeOH (50 mL) was added and then heated to reflux. The cloudy solution was filtrated over a pad of celite. The mother liquid was evaporated to dryness and the residue was triturated with 3 mL EtOAc. The suspension was filtrated to give 0.317 g white crystalline solid Intermediate 3 (52%, M+1: 321 on MS).

Step 4: Synthesis of N-(2-Amino-phenyl)-4-[(4-pyrymidin-2-ylamino)-methyl]-benzamide

Intermediate 3 (3.68 g, 11.5 mmol) was mixed with THF (23 mL), MeOH (23 mL) and H₂O (11.5 mL) at rt. LiOH (1.06 g, 25.3 mmol) was added to reaction mixture. The resulting reaction mixture was warmed up to 40° C. overnight. HCl solution (12.8 mL, 2N) was added to adjust pH=3 when the mixture was cooled down to rt. The mixture was evaporated to dryness and then the solid was washed with minimum quantity of H₂O upon filtration. The filter cake was dried over freeze dryer to give 3.44 g acid of the title compound (95%, M+1: 307 on MS).

Acid (3.39 g, 11.1 mmol) of the title compound, BOP (5.679 g, 12.84 mmol) and o-Ph(NH₂)₂ (2.314 g, 21.4 mmol) were dissolved in the mixture of DMF (107 mL) and Et₃N (2.98 mL, 21.4 mmol). The reaction mixture was stirred at rt for 5 h and then evaporated to dryness. The residue was purified by flash column (pure EtOAc to 5% MeOH/EtOAc) and then interested fractions were concentrated. The final product was triturated with EtOAc to give 2.80 g of title product

(66%, MS+1: 397 on MS).

 ¹H NMR (400 MHz, DMSO-D₆) δ (ppm): 9.57 (s, 1H), 9.22 (s, 1H), 8.66 (d, J=3.5 Hz, 1H), 8.39 (d, J=5.1 Hz, 2H), 8.00 (t, J=6.5 Hz, 1H), 7.90 (d, J=8.2 Hz, 2H), 7.50 (m, 3H), 7.25 (d, J=5.1 Hz, 1H), 7.12 (d, J=7.4 Hz, 1H), 6.94 (dd, J=7.0, 7.8 Hz, 1H), 6.75 (d, J=8.2 Hz, 1H), 6.57 (dd, J=7.0, 7.8 Hz, 1H), 4.86 (s, 2H), 4.64 (d, J=5.9 Hz, 2H).

References

To start with the simplest one is Quantitative structure-activity relationship (QSAR) which is also referred to as 2D-QSAR sometimes. 3D-QSAR involving Comparative Molecular Field Analysis (CoMFA) and Comparative molecular similarity index analysis (CoMSIA) are extension of QSAR. QSAR is not able to take the three dimensional structure of a molecule into consideration due to absence of three-dimensional parameterization of structures. 3D-QSAR scores over QSAR in this respect. Docking studies throw more light on the binding modes of drugs with their target proteins but it is feasible only when the crystal structure of the target enzyme/protein is known with good resolution. Docking studies are also used for virtual screening of databases. But the ideal technique for virtual screening of compounds is through pharmacophore mapping and screening, especially when the structure of the target is not known. Very large databases can be first screened by pharmacophorebecause the technique is quite fast followed by screening of the positive hits using docking studies. Insilico designing of novel compounds can also be performed using deNovodesigning techniques subject to the condition that the target structure in known.

http://www.japtr.org/article.asp?issn=2231-4040;year=2013;volume=4;issue=1;spage=2;epage=3;aulast=Yadav

• Ketoconazole, 1-acetyl-4- [4-[[2-(2,4-dichlorophenyl)-2-[(1H-imidazol-1-yl)-methyl]-1,3-dioxolan-4-yl] methoxy] phenyl] piperazine, is a racemic mixture of the cis enantiomers (-)-(2S, 4R) and (+)-(2R, 4S) marketed as an anti-fungal agent. Ketoconazole inhibits fungal growth through the inhibition of ergosterol synthesis. Ergosterol is a key component of fungal cell walls.
• More recently, ketoconazole was found to decrease plasma cortisol and to be useful, alone and in combination with other agents, in the treatment of a variety of diseases and conditions, including type 2 diabetes, Metabolic Syndrome (also known as the Insulin Resistance Syndrome, Dysmetabolic Syndrome or Syndrome X), and other medical conditions that are associated with elevated cortisol levels. SeeU.S. Patent Nos. 5,584,790 ; 6,166,017 ; and 6,642,236 , each of which is incorporated herein by reference. Cortisol is a stress-related hormone secreted from the cortex of the adrenal glands. ACTH (adenocorticotropic hormone) increases cortisol secretion. ACTH is secreted by the pituitary gland, a process activated by secretion of corticotropin releasing hormone (CRH) from the hypothalamus.
• Cortisol circulates in the bloodstream and activates specific intracellular receptors, such as the glucocorticoid receptor (GR). Disturbances in cortisol levels, synthetic rates or activity have been shown to be associated with numerous metabolic complications, including insulin resistance, obesity, diabetes and Metabolic Syndrome. Additionally, these metabolic abnormalities are associated with substantially increased risk of cardiovascular disease, a major cause of death in industrialized countries. See Mårin P et al., “Cortisol secretion in relation to body fat distribution in obese premenopausal women.” Metabolism 1992; 41:882-886, Bjorntorp, “Neuroendocrine perturbations as a cause of insulin resistance.” Diabetes Metab Res Rev 1999; 15(6): 427-41, and Rosmond, “Role of stress in the pathogenesis of the metabolic syndrome.” Psychoneuroendocrinology 2005; 30(1): 1-10, each of which is incorporated herein by reference.
• While ketoconazole is known to inhibit some of the enzymatic steps in cortisol synthesis, such as, for example, 17α hydroxylase (Wachall et al., “Imidazole substituted biphenyls: a new class of highly potent and in vivo active inhibitors of P450 17 as potential therapeutics for treatment of prostate cancer.” Bioorg Med Chem 1999; 7(9): 1913-24, incorporated herein by reference) and 11b-hydroxylase (Rotstein et al., “Stereoisomers of ketoconazole: preparation and biological activity.” J Med Chem 1992; 35(15): 2818-25) and 11β-hydroxy steroid dehydrogenase (11β-HSD) (Diederich et al., “In the search for specific inhibitors of human 11β-hydroxysteroid-dehydrogenases (11β-HSDs): chenodeoxycholic acid selectively inhibits 11β-HSD-L” Eur J Endocrinol 2000; 142(2): 200-7, incorporated herein by reference) the mechanisms by which ketoconazole decreases cortisol levels in the plasma have not been reported. For example, there is uncertainty regarding the effect of ketoconazole on the 11β-hydroxy steroid dehydrogenase (11β-HSD) enzymes. There are two 11β-HSD enzymes. One of these, 11β-HSD-I, is primarily a reductase that is highly expressed in the liver and can convert the inactive 11-keto glucocorticoid to the active glucocorticoid (cortisol in humans and corticosterone in rats). In contrast, the other, 11β-HSD-II, is primarily expressed in the kidney and acts primarily as an oxidase that converts active glucocorticoid (cortisol in humans and corticosterone in rats) to inactive 11-keto glucocorticoids. Thus, the plasma concentration of active glucocorticoid is influenced by the rate of synthesis, controlled in part by the activity of adrenal 11β-hydroxylase and by the rate of interconversion, controlled in part by the relative activities of the two 11β-HSD enzymes. Ketoconazole is known to inhibit these three enzymes (Diederich et al., supra) and the 2S,4R enantiomer is more active against the adrenal 11β-hydroxylase enzyme than is the 2R,4S enantiomer (Rotstein et al., supra). However, there are no reports describing the effect of the two ketoconazole enantiomers on either of 11β-HSD-I or 11β-HSD-II, so it is not possible to predict what effects, if any, the two different ketoconazole enantiomers will each have on plasma levels of the active glucocorticoid levels in a mammal.
• Ketoconazole has also been reported to lower cholesterol levels in humans (Sonino et al. (1991). “Ketoconazole treatment in Cushing’s syndrome: experience in 34 patients.” Clin Endocrinol (Oxf). 35(4): 347-52; Gylling et al. (1993). “Effects of ketoconazole on cholesterol precursors and low density lipoprotein kinetics in hypercholesterolemia.” J Lipid Res. 34(1): 59-67) each of which is incorporated herein by reference). The 2S,4R enantiomer is more active against the cholesterol synthetic enzyme 14 αlanosterol demethylase than is the other (2R,4S) enantiomer (Rotstein et al infra). However, because cholesterol level in a human patient is controlled by the rate of metabolism and excretion as well as by the rate of synthesis it is not possible to predict from this whether the 2S,4R enantiomer of ketoconazole will be more effective at lowering cholesterol levels.
• The use of ketoconazole as a therapeutic is complicated by the effect of ketoconazole on the P450 enzymes responsible for drug metabolism. Several of these P450 enzymes are inhibited by ketoconazole (Rotsteinet al., supra). This inhibition leads to an alteration in the clearance of ketoconazole itself (Brass et al., “Disposition of ketoconazole, an oral antifungal, in humans.” Antimicrob Agents Chemother 1982; 21(1): 151-8, incorporated herein by reference) and several other important drugs such as Glivec (Dutreix et al., “Pharmacokinetic interaction between ketoconazole and imatinib mesylate (Glivec) in healthy subjects.” Cancer Chemother Pharmacol 2004; 54(4): 290-4) and methylprednisolone (Glynn et al., “Effects of ketoconazole on methylprednisolone pharmacokinetics and cortisol secretion.” Clin Pharmacol Ther 1986; 39(6): 654-9). As a result, the exposure of a patient to ketoconazole increases with repeated dosing, despite no increase in the amount of drug administered to the patient. This exposure and increase in exposure can be measured and demonstrated using the “Area under the Curve” (AUC) or the product of the concentration of the drug found in the plasma and the time period over which the measurements are made. The AUC for ketoconazole following the first exposure is significantly less than the AUC for ketoconazole after repeated exposures. This increase in drug exposure means that it is difficult to provide an accurate and consistent dose of the drug to a patient. Further, the increase in drug exposure increases the likelihood of adverse side effects associated with ketoconazole use.
• [0008]
  Rotstein et al. (Rotstein et al., supra) have examined the effects of the two ketoconazole cis enantiomers on the principal P450 enzymes responsible for drug metabolism and reported “…almost no selectivity was observed for the ketoconazole isomers” and, referring to drug metabolizing P450 enzymes: “[t]he IC50 values for the cis enantiomers were similar to those previously reported for racemic ketoconazole”. This report indicated that both of the cis enantiomers could contribute significantly to the AUC problem observed with the ketoconazole racemate.
• One of the adverse side effects of ketoconazole administration exacerbated by this AUC problem is liver reactions. Asymptomatic liver reactions can be measured by an increase in the level of liver specific enzymes found in the serum and an increase in these enzymes has been noted in ketoconazole treated patients (Sohn, “Evaluation of ketoconazole.” Clin Pharm 1982; 1(3): 217-24, and Janssen and Symoens, “Hepatic reactions during ketoconazole treatment.” Am J Med 1983; 74(1B): 80-5, each of which is incorporated herein by reference). In addition 1:12,000 patients will have more severe liver failure (Smith and Henry, “Ketoconazole: an orally effective antifungal agent. Mechanism of action, pharmacology, clinical efficacy and adverse effects.” Pharmacotherapy 1984; 4(4): 199-204, incorporated herein by reference). As noted above, the amount of ketoconazole that a patient is exposed to increases with repeated dosing even though the amount of drug taken per day does not increase (the “AUC problem”). The AUC correlates with liver damage in rabbits (Ma et al., “Hepatotoxicity and toxicokinetics of ketoconazole in rabbits.” Acta Pharmacol Sin 2003; 24(8): 778-782 incorporated herein by reference) and increased exposure to the drug is believed to increase the frequency of liver damage reported in ketoconazole treated patients.
• Additionally, U.S. Patent No. 6,040,307 , incorporated herein by reference, reports that the 2S,4R enantiomer is efficacious in treating fungal infections. This same patent application also reports studies on isolated guinea pig hearts that show that the administration of racemic ketoconazole may be associated with an increased risk of cardiac arrhythmia, but provides no data in support of that assertion. However, as disclosed in that patent, arrhythmia had not been previously reported as a side effect of systemic racemic ketoconazole, although a particular subtype of arrhythmia, torsades de pointes, has been reported when racemic ketoconazole was administered concurrently with terfenadine. Furthermore several published reports (for example, Morganroth et al. (1997). “Lack of effect of azelastine and ketoconazole coadministration on electrocardiographic parameters in healthy volunteers.” J Clin Pharmacol. 37(11): 1065-72) have demonstrated that ketoconazole does not increase the QTc interval. This interval is used as a surrogate marker to determine whether drugs have the potential for inducing arrhythmia. US Patent Number 6,040,307 also makes reference to diminished hepatoxicity associated with the 2S,4R enantiomer but provides no data in support of that assertion. The method provided in US Patent Number 6,040,307 does not allow for the assessment of hepatoxicity as the method uses microsomes isolated from frozen tissue.

…………………………

http://www.google.com/patents/EP1853266B1?cl=en

• DIO-902 is the single enantiomer 2S,4R ketoconazole and is derived from racemic ketoconazole. It is formulated using cellulose, lactose, cornstarch, colloidal silicon dioxide and magnesium stearate as an immediate release 200 mg strength tablet. The chemical name is 2S,4R cis-1-acetyl-4-[4-[[2-(2,4-dichlorophenyl)-2-(1H-imidazol-1-ylmethyl)-1,3-dioxolan-4-yl] methoxyl]phenyl] piperazine, the formula is C₂₆H₂₈Cl₂N₄O₄, and the molecular weight is 531.44. The CAS number is 65277-42-1, and the structural formula is provided below. The chiral centers are at the carbon atoms 2 and 4 as marked.
• [0132]
  Ketoconazole is an imidazole-containing fungistatic compound. DIO-902 is an immediate release tablet to be taken orally and formulated as shown in the table below.
  

  Component	Percentage
  2S,4R ketoconazole; DIO-902	50%
  Silicified Microcrystalline Cellulose, NF (Prosolv HD 90)	16.5
  Lactose Monohydrate, NF (316 Fast-Flo)	22.4
  Corn Starch, NF (STA-Rx)	10
  Colloidal Silicon Dioxide, NF (Cab-O-Sil M5P)	0.5
  Magnesium Stearate, NF	0.6

  The drug product may be stored at room temperature and is anticipated to be stable for at least 2 years at 25° C and 50% RH. The drug is packaged in blister packs.

ketoconazole 2S,4R enantiomer

ketoconazole 2S,4S enantiomer

• ketoconazole 2R,4R enantiomer

ketoconazole 2R,4S enantiomer

……………………..

Journal of Medicinal Chemistry (Impact Factor: 5.61). 08/1992; 35(15):2818-25. DOI: 10.1021/jm00093a015

http://pubs.acs.org/doi/abs/10.1021/jm00093a015

…………………….

Enantioselective separation of ketoconazole enantiomers by membrane extraction

http://www.sciencedirect.com/science/article/pii/S1383586611001638

A new process has been developed to separate ketoconazole (KTZ) enantiomers by membrane extraction, with the oppositely preferential recognition of hydrophobic and hydrophilic chiral selectors in organic and aqueous phases, respectively. This system is established by adding hydrophobic l-isopentyl tartrate (l-IPT) in organic strip phase (shell side) and hydrophilic sulfobutylether-β-cyclodextrin (SBE-β-CD) in aqueous feed phase (lumen side), which preferentially recognizes (+)-2R,4S-ketoconazole and (−)-2S,4R-ketoconazole, respectively. The studies performed involve two enantioselective extractions in a biphasic system, where KTZ enantiomers form four complexes with SBE-β-CD in aqueous phase and l-IPT in organic phase, respectively. The membrane is permeable to the KTZ enantiomers but non-permeable to the chiral selector molecules. Fractional chiral extraction theory, mass transfer performance of hollow fiber membrane, enantioselectivity and some experimental conditions are investigated to optimize the separation system. Mathematical model of I/II = 0.893e^0.039NTU for racemic KTZ separation by hollow fiber extraction, is established. The optical purity for KTZ enantiomers is up to 90% when 9 hollow fiber membrane modules of 30 cm in length in series are used.

• I, (−)-2S,4R-ketoconazole;
• II, (+)-2R,4S-ketoconazole;
• CDs, cyclodextrin derivatives;
• l-IPT, l-isopentyl tartrate;
• d-IPT, d-isopentyl tartrate;
• HP-β-CD, hydroxypropyl-β-cyclodextrin;
• Me-β-CD, methyl-β-cyclodextrin;
• β-CD, β-cyclodextrin;
• NTU, number of transfer units;
• HTU, height of a transfer unit;
• PVDF,polyvinylidene fluoride

…………………….

Stereoselective synthesis of both enantiomers of ketoconazole from (R)- and (S)-

• Stereoselective synthesis of both enantiomers of ketoconazole from (R)- and (S)-epichlorohydrin

  Original Research Article

• Pages 1283-1294
• Pelayo Camps, Xavier Farrés, Ma Luisa García, Joan Ginesta, Jaume Pascual, David Mauleón, Germano Carganico

• Bromobenzoates (2R,4R)- and (2S,4S)-18, prepared stereoselectively from (R)- and (S)-epichlorohydrin, were transformed into (2R,4S)-(+)- and (2S,4R)-(−)-Ketoconazole, respectively, following the known synthetic protocols for the racemic mixture.

Tetrahedron Asymmetry 1995, 6(6): 1283

Stereoselective syntheses of both enantiomers of ketoconazole (1) from commercially available (R)- or (S)-epichlorohydrin has been developed. The key-step of these syntheses involves the selective substitution of the methylene chlorine atom by benzoate on a mixture of  and  or of their enantiomers, followed by crystallization of the corresponding cis-benzoates, (2S,4R)-18 or(2S,4S)-18, from which (+)- or (−)-1 were obtained as described for (±)-1. The ee’s of (+)- and (−)-ketoconazole were determined by HPLC on the CSP Chiralcel OD-H.

………………..

WO 1996029325

 http://www.google.com/patents/WO1996029325A1?cl=en

The incidence of fungal infections has considerably increased over the last decades. Notwithstanding the utility of the antifungal compounds commercialized in the last 15 years, the investigation in this field is however very extensive. During this time, compounds belonging to the azole class have beer, commercialized for both the topical and oral administrations, such a class including imidazoles as well as 1,2,4-triazoles. Some of these compounds car. show m some degree a low gastrointestinal tolerance as well as hepatotoxycity.

A large number of pharmaceutically active compounds are commercialized as stereoisomeric mixtures. On the other hand, the case in which only one of said stereoisomers is pharmaceutically active is frequent.

The undesired enantiomer has a lower activity and it sometimes may cause undesired side-effects.

Ketoconazole (1-acetyl-4-[4-[[2-(2,4-dichlorophenyl)-2-[(1H-imidazol-1-yl)methyl]-1,3-dioxolane-4-yl]methoxy]phenyl]piperazine), terconazole (1-[4-[[2(2,4-dichlorophenyl)-2-[(1H-1 , 2 ,4-triazol-1-yl)methyl]-1,3-dioxolane-4-yl]methoxy]phenyl]-4-(1-methylethyl)piperazine) and other related azole antifungal drugs contain in their structure a substituted 1,3-dioxolane ring, in which carbon atoms C2 and C4 are stereogenic centres, therefore four possible stereoisomers are possible. These compounds are commercialized in the form or cis racemates which show a higher antifungal activity than the corresponding trans racemates.

The cis homochiral compounds of the present invention, which are intermediates for the preparation of enantiomerically pure antifungal drugs, have been prepared previously in the racemic form and transformed into the different azole antifungal drugs in the racemic form [J. Heeres et al., J . Med . Chem . , 22 , 1003 (1979). J . Med . Chem . , 26, 611 (1983), J . Med . Chem . , 27 , 894 (1984) and US 4,144,346, 4,223,036, 4,358,449 and 4,335,125].

Scheme 1 shows the synthesis described for racemic ketoconazole [J. Heeres et al., J . Med . Chem . , 22 , 1003 (1979)]. Scheme 1

)

The synthesis of racemic terconazole [J. Heeres et al., J. Med . Chem . , 26 , 611 11983)] is similar. differing in the introduction of a 1 H- 1 , 2,4-triazol-1-yl substituent in place of 1H-imidazol-1-yl and in the nature of the phenol used in the last step of the synthetic sequence, which phenol is 1-methylethyl-4-(4- hydroxyphenyl)piperazme instead of 1-acetyl-4-(4-nydroxyphenyl)piperazine.

The preparation of racemic itraconazole [J. Heeres et al., J. Med . Chem. , 27 , 894 (1984)] is similar to that of terconazole, differing only in the nature of the phenol used in the last step of the synthetic sequence.

In the class of azoles containing a 1,3-dioxolane ring and a piperazine ring and moreover they are pure enantiomers, only the preparation of (+)- and (-)-ketoconazole has been described [D. M. Rotstein et al., J. Med . Chem . , 35, 2818 (1992)] (Scheme 2) starting from the tosylate of (+)- and (-) 2,2-dimethyl-1,3-dioxolane-4-methanol.

Scheme 2

This synthesis suffers from a series of drawbacks, namely: a) the use of expensive, high molecular weight starting products which are available only on a laboratory scale, and b) the need for several chromatographies during the process in order to obtain products of suitable purity, which maKes said synthesis economically unattractive and difficult to apply industrially.

Recently (N. M. Gray, WO 94/14447 and WO 94/14446) the use of (-)-ketoconazole and (+)-ketoconazole as antifungal drugs causing less side-effects than (±)-ketoconazole has been claimed.

The industrial preparation of enantiomerically pure antifungal drugs with a high antifungal activity and less side-effects is however a problem in therapy. The present invention provides novel homochiral compounds which are intermediates for the industrial preparation of already known, enantiomerically pure antifungal drugs such as ketoconazole enantiomers, or of others which have not yet been reported in literature, which are described first in the present invention, such as (+)-terconazole and (-)-terconazoie, which show the cited antifungal action, allowing to attain the same therapeutical effectiveness using lower dosages than those required for racemic terconazole

Example 14 : (2S,4R)-(-)-1-acetyl-4-[4-[ [2-(2,4-dichlorophenyl)-2-[(1H-imidazol-1-yl)-methyl]-1,3-dioxolane-4-yl]methoxy]phenyl]piperazine, (2S,4R) -(- )-ketoconazole.

This compound is prepared following the process described above for (2R,4S)-(+)-ketoconazole. Starting from HNa (60-65% dispersion in paraffin, 32 mg, 0.80 mmol), 1-acetyl-4-(4-hydroxyphenyl)piperazine (153 mg, 0.69 mol) and (2S,4S)-(-)-IV (Ar = 2,4-dichlorophenyl, Y = CH, R = CH₃) (250 mg, 0.61 mmol), upon crystallization from an acetone:ethyl acetate mixture, (2S,4R) -(-)-ketoconazole is obtained [(2S,4R)-V Ar = 2,4-dichlorophenyl, Y = CH, Z = COCH₃] (196 mg, 61% yield) as a solid, m.p. 153-155ºC (lit. 155-157ºC); [α]_D ²⁰ = -10.50 (c = 0.4, CHCl₃) (lit. [α]_D ²⁵ = -10.58. c = 0.4, CHCl₃) with e.e. > 99% (determined by HPLC using the chiral stationary phase CHIRALCEL OD-H and ethanol:hexane 1:1 mixtures containing 0.1 % diethylamine as the eluent).

+ KETOCONAZOLE…. UNDESIRED

Example 7: (2 R ,4S)-(+)-1-acetyl-4-[4-[[2-(2,4-dichlorophenyl)-2-[(1H-imidazol-1-yl)methyl]-1,3-dioxolane-4-yl]methoxy]phenyl]piperazine (22, 4 S)-(+)-ketoconazole.

To a suspension of NaH (dispersed in 60-65% paraffin, 19.2 mg, 0.48 mmol) in anhydrous DMSO (3 ml),

1-acetyl-4-(hydroxyphenyl)piperazine (102 mg, 0.46 mmol) is added and the mixture is stirred for 1 hour at room temperature. Then, a solution of (2R,4R) – (+)-IV (Ar = 2,4-dichlorophenyl, Y = CH, R = CH₃) (160 mg, 0.39 mmol) in anhydrous DMSO (5 ml) is added, and the mixture is heated at 80ºC for 4 hours. The reaction mixture is allowed to cool to room temperature, diluted with water

(20 ml) and extracted with CH₂Cl₂ (3 × 25 ml). The combined organic phases are washed with water (3 × 25), dried with Na₂SO₄ and the solvent is evaporated off under vacuum. The oily residue thus obtained is crystallized from an acetone:ethyl acetate mixture to give (2R,4S)-(+)-ketoconazole ( (2R, 4 S) -V , Ar 2,4-dichlorophenyl, Y = CH , Z = COCH₃ ) ( 110 mg , 5 3 % yie ld ) as a white solid, m.p. 155-156°C (lit. 154-156ºC), [α]_D ²⁰ = + 8.99 (c = 0.4, CHCl₃) (lit. [α]_D ²⁵ = + 8.22, c = 0.4, CHCl₃), with e.e. > 99% (determined by HPLC using the chirai stationary phase CHIRALCEL OD-H and ethanol:hexane 1:1 mixtures containing 0.1% of diethylamine, as the eluent; (+)-Ketoconazole retention time 73,28 min. (-)-Ketoconazole, retention time 79.06 min).

IR (KBr), ʋ : 2875, 1645, 1584, 1511, 1462, 1425, 1250, 103S, 313 cm^-1.

¹H NMR (500 MHz, CDCl₃), δ : 2.12 (s, 3H, COCH₃),

3.02 (m, 2H, 3-H₂), 3.05 (m, 2H, 5-H₂), 3.27 (dd, J= 9.5

Hz, J’=7.0 Hz, 1H) and 3.70 (dd, J=9.5 Hz, J’=5.0 Hz, 1 H) (4″-CH₂), 3.60 (m, 2H, 6-H₂), 3.76 (m, 2H, 2-H₂), 3.73 (dd, J=8.0 Hz, J’=5.0 Hz, 1H) and 3.86 (dd, J=8.0 Hz, J’=6.5 Hz, 1H) (5″-H₂), 4.34 (m, 1H, 4″-H), 4.40 (d, J=15.0 Hz, 1H) and 5.00 (d, J=15.0 Hz, 1H) (CH₂-N), 4.34

(m, 1H, 4″-H), 6.76 [d, J = 9.0 Hz, 2H, 2'(C6' )-H], 6.88

[d, J=9.0 Hz, 2H, C3'(C5)-H], 6.96 (s, 1H, imidazole 5- H), 6.99 (s, 1H, imidazole 4-H), 7.25 (dd, J=8.5 Hz, J’=2.0 Hz, 1H, 5″‘-H), 7.46 (d, J=2.0 Hz, 1H, 3″‘-H),

7.53 (s, 1H, imidazole 2-H), 7.57 (d, J=8.5 Hz, 1H,

6″‘-H).

¹³C NMR (75.4 MHz, CDCI₃), δ : 21.3 (CH₃, COCH₃), 41.4 (CH₂, C₂), 46.3 (CH₂, C6), 50.6 (CH₂, C3), 51.0 (CH₂, C5), 51.2 (CH₂, CH₂-N), 67.6 [CH₂, C5″ and 4″-CH₂), 74.7 (CH, C4″), 108.0 (C, C2″), 115.2 [CH, C2'(6')], 118.8 [CH, C3'(5')], 121.2 (CH, imidazole C5), 127.2 (CH, C5″‘), 128.5 (CH, imidazole C4), 129.5 (CH, C6′”), 131.3 (CH, C3″‘), 133.0 (C, C2″‘), 134.6 (C, C1′”), 135.8 (C, C4″‘), 138.8 (CH, imidazole C2), 145.6 (C, C1′), 152.8 (C, C4′), 168.9 (C, CO).

…………………………

Experimental and theoretical analysis of the interaction of (+/-)-cis-ketoconazole with beta-cyclodextrin in the presence of (+)-L-tartaric acid
J Pharm Sci 1999, 88(6): 599

Experimental and theoretical analysis of the interaction of (±)-cis-ketoconazole with β-cyclodextrin in the presence of (+)-l-Tartaric acid (pages 599–607)

Enrico Redenti, Paolo Ventura, Giovanni Fronza, Antonio Selva, Silvia Rivara, Pier Vincenzo Plazzi and Marco Mor

Article first published online: 12 JUN 2000 | DOI: 10.1021/js980468o

http://onlinelibrary.wiley.com/doi/10.1021/js980468o/pdf

• Abstract
• PDF(154K)

¹H NMR spectroscopy was used for determining the optical purity of cis-ketoconazole enantiomers obtained by fractional crystallization. The chiral analysis was carried out using β-cyclodextrin in the presence of (+)-l-tartaric acid. The mechanism of the chiral discrimination process, the stability of the complexes formed, and their structure in aqueous solution were also investigated by ¹H and ¹³C chemical shift analysis, two-dimensional NOE experiments, relaxation time measurements, and mass spectrometry experiments. Theoretical models of the three-component interaction were built up on the basis of the available NMR data, by performing a conformational analysis on the relevant fragments on ketoconazole and docking studies on the components of the complex. The model derived from a folded conformation of ketoconazole turned out to be fully consistent with the molecular assembly found in aqueous solution, as inferred from NOE experiments. An explanation of the different association constants for the complexes of the two enantiomers is also provided on the basis of the interaction energies.

Aesculus

Aesculus hippocastanum

The genus Aesculus (/ˈɛskjʊləs/^[1] or /ˈaɪskjʊləs/) comprises 13–19 species of trees and shrubs native to the temperate Northern Hemisphere, with 6 species native to North America and 7–13 species native to Eurasia; there are also several hybrids. Aesculus exhibits a classical arcto-Tertiary distribution.^[a] The genus has traditionally been treated in the ditypic family Hippocastanaceae along with Billia,^[3] but recent phylogenetic analysis of morphological^[4] and molecular data^[5] has caused this family, along with the Aceraceae (Maples andDipteronia), to be included in the soapberry family (Sapindaceae).

Linnaeus named the genus Aesculus after the Roman name for an edible acorn. Common names for these trees include “buckeye” and “horse chestnut”. Some are also called white chestnut or red chestnut (as in some of the Bach flower remedies). In Britain, they are sometimes called conker trees because of their link with the game of conkers, played with the seeds, also called conkers. Aesculus seeds were traditionally eaten, after leaching, by the Jōmon people of Japan over about four millennia, until 300 AD.^[6]

Aesculus glabra Ohio buckeye

Flower of Aesculus x carnea, the red Horse Chestnut

Description

Aesculus species have stout shoots with resinous, often sticky, buds; opposite, palmately divided leaves, often very large—to 65 cm (26 in) across in the Japanese horse chestnut Aesculus turbinata. The seeds of the Aesculus are traditionally used in a game called conkers in Europe. Species are deciduous or evergreen. Flowers are showy, insect- or bird-pollinated, with four or five petals fused into a lobedcorolla tube, arranged in a panicle inflorescence. Flowering starts after 80–110 growing degree days. The fruit matures to a capsule, 2–5 cm (²⁵⁄₃₂–1 ³¹⁄₃₂ in) diameter, usually globose, containing one to three seeds (often erroneously called a nut) per capsule. Capsules containing more than one seed result in flatness on one side of the seeds. The point of attachment of the seed in the capsule (hilum) shows as a large circular whitish scar. The capsule epidermis has “spines” (botanically: prickles) in some species, while other capsules are warty or smooth. At maturity, the capsule splits into three sections to release the seeds.^[7][8][9]

The species of Aesculus include:

• Aesculus arguta: Aesculus glabra
• Aesculus californica: California buckeye (western North America)
• Aesculus × carnea: red horse chestnut
• Aesculus chinensis: Chinese horse chestnut (eastern Asia)
• Aesculus chinensis var. wilsonii: Wilson’s horse chestnut (eastern Asia)
• Aesculus flava (A. octandra): yellow buckeye (eastern North America)
• Aesculus glabra: Ohio buckeye (eastern North America)
• Aesculus hippocastanum: common horse chestnut (Europe, native to the Balkans)
• Aesculus indica: Indian horse chestnut (eastern Asia)
• Aesculus neglecta: dwarf buckeye (eastern North America)
• Aesculus parviflora: bottlebrush buckeye (eastern North America)
• Aesculus parryi: Parry’s buckeye (western North America, endemic in Baja California del Norte)
• Aesculus pavia: red buckeye (eastern North America)
• Aesculus pavia var. flavescens: Texas yellow buckeye, yellow woolly buckeye (eastern North America, narrowly endemic in Texas)
• Aesculus sylvatica: painted buckeye (eastern North America)
• Aesculus turbinata: Japanese horse chestnut (eastern Asia, endemic in Japan)
• Aesculus wangii: Aesculus assamica (eastern Asia)

Cultivation

The most familiar member of the genus worldwide is the common horse chestnut Aesculus hippocastanum. The yellow buckeye Aesculus flava (syn. A. octandra) is also a valuable ornamental tree with yellow flowers, but is less widely planted. Among the smaller species, the bottlebrush buckeye Aesculus parviflora also makes a very interesting and unusual flowering shrub. Several other members of the genus are used as ornamentals, and several horticultural hybrids have also been developed, most notably the red horse chestnut Aesculus × carnea, a hybrid between A. hippocastanum and A. pavia.

Use in alternative medicine

Aesculus has been listed as one of the 38 substances used to prepare Bach flower remedies,^[10] a kind of alternative medicine promoted for its effect on health. However according to Cancer Research UK, “there is no scientific evidence to prove that flower remedies can control, cure or prevent any type of disease, including cancer”.^[11]

References

External links

• Germplasm Resources Information Network: Aesculus
• Forest, F., Drouin, J. N., Charest, R., Brouillet, L., & Bruneau A. (2001). A morphological phylogenetic analysis of Aesculus L. and Billia Peyr. (Sapindaceae). Canad. J. Botany79 (2): 154-169. Abstract.
• Aesculus glabra (Ohio buckeye) King’s American Dispensatory
• Winter ID pictures

Aesculus hippocastanum is a large deciduous tree, commonly known as horse-chestnut or conker tree.

Distribution

Aesculus hippocastanum is native to a small area in the Pindus Mountains mixed forests and Balkan mixed forests of South East Europe.^[1]It is widely cultivated in streets and parks throughout the temperate world.

Growth

A. hippocastanum grows to 36 metres (118 ft) tall, with a domed crown of stout branches; on old trees the outer branches often pendulous with curled-up tips. The leaves are opposite and palmately compound, with 5–7 leaflets; each leaflet is 13–30 cm long, making the whole leaf up to 60 cm across, with a 7–20 cm petiole. The leaf scars left on twigs after the leaves have fallen have a distinctive horseshoe shape, complete with seven “nails”. The flowers are usually white with a small red spot; they are produced in spring in erect panicles 10–30 cm tall with about 20–50 flowers on each panicle. Usually only 1–5 fruit develop on each panicle; the shell is a green, spiky capsule containing one (rarely two or three) nut-like seeds called conkers or horse-chestnuts. Each conker is 2–4 cm diameter, glossy nut-brown with a whitish scar at the base.^[2]

Etymology

The common name “horse-chestnut” (often unhyphenated) is reported as having originated from the erroneous belief that the tree was a kind of chestnut (though in fact only distantly related), together with the observation that eating the fruit cured horses of chest complaints^[3] despite this plant being poisonous to horses.

Uses

Cultivation for its spectacular spring flowers is successful in a wide range of temperate climatic conditions provided summers are not too hot, with trees being grown as far north asEdmonton, Alberta, Canada,^[4] the Faroe Islands,^[5] Reykjavík, Iceland and Harstad, Norway.

In Britain and Ireland, the nuts are used for the popular children’s game conkers. During the First World War, there was a campaign to ask for everyone (including children) to collect horse-chestnuts and donate them to the government. The conkers were used as a source of starch for the fermentation via the Clostridium acetobutylicum method devised by Chaim Weizmann to produce acetone. Any starch plant would have done, but they chose to ask for conkers to avoid causing starvation by using food. Weizmann’s process could use any source of starch, but it was never particularly efficient and the factory only produced acetone for three months. The aim was to produce acetone for use as solvent which aided in the production of cordite, which was then used in military armaments.

A selection of fresh conkers from a horse-chestnut

The nuts, especially those that are young and fresh, are slightly poisonous, containing alkaloid saponins and glucosides. Although not dangerous to touch, they cause sickness when eaten; consumed by horses, they can cause tremors and lack of coordination.^[6] Somemammals, notably deer, are able to break down the toxins and eat them safely.^{[citation needed]}

Though the seeds are said to repel spiders there is little evidence to support these claims. The presence of saponin may repel insects but it is not clear whether this is effective on spiders.^[7]

Horse-chestnuts have been threatened by the leaf-mining moth Cameraria ohridella, whose larvae feed on horse chestnut leaves. The moth was described from Macedonia where the species was discovered in 1984 but took 18 years to reach Britain.^[8]

The flower is the symbol of the city of Kiev, capital of Ukraine.^[9] Although the horse-chestnut is sometimes known as the buckeye, this name is generally reserved for the New World members of the Aesculus genus.

Medical uses

The seed extract standardized to around 20 percent aescin (escin) is used for its venotonic effect, vascular protection, anti-inflammatory and free radical scavenging properties.^[10][11] Primary indication is chronic venous insufficiency.^[11][12] A recent Cochrane Review found the evidence suggests that Horse Chestnut Seed Extract is an efficacious and safe short-term treatment for chronic venous insufficiency.^[13]

Aescin reduces fluid leaks to surrounding tissue by reducing both the number and size of membrane pores in the veins.

Safety in medical use

Two preparations are considered; whole horsechestnut extract (whole HCE) and purified β-aescin. Historically, whole HCE has been used both for oral and IV routes (as of year 2001). The rate of adverse effects are low, in a large German study, 0.6%, consisting mainly of gastrointestinal symptoms. Dizziness, headache and itching have been reported. One serious safety issue is rare cases of acute anaphylactic reactions, presumably in a context of whole HCE. Purified β-aescin would be expected to have a better safety profile.

Another is the risk of acute renal failure, “when patients, who had undergone cardiac surgery were given high doses of horse chestnut extract i.v. for postoperative oedema. The phenomenon was dose dependent as no alteration in renal function was recorded with 340 μg kg−1, mild renal function impairment developed with 360 μg kg−1 and acute renal failure with 510 μg kg−1″.^[14] This almost certainly took place in a context of whole HCE.

Three clinical trials were since performed to assess the effects of aescin on renal function. A total of 83 subjects were studied; 18 healthy volunteers given 10 or 20 mg iv. for 6 days, 40 in-patients with normal renal function given 10 mg iv. two times per day (except two children given 0.2 mg/kg), 12 patients with cerebral oedema and normal renal function given a massive iv. dose on the day of surgery (49.2 ± 19.3 mg) and 15.4 ± 9.4 mg daily for the following 10 days and 13 patients with impaired renal function due to glomerulonephritis or pyelonephritis, who were given 20–25 mg iv. daily for 6 days. “In all studies renal function was monitored daily resorting to the usual tests of renal function: BUN, serum creatinine, creatinine clearance, urinalysis. In a selected number of cases paraaminohippurate and labelled EDTA clearance were also measured. No signs of development of renal impairment in the patients with normal renal function or of worsening of renal function in the patients with renal impairment were recorded.” It is concluded that aescin has excellent tolerability in a clinical setting.^[15]

Raw Horse Chestnut seed, leaf, bark and flower are toxic due to the presence of esculin and should not be ingested. Horse chestnut seed is classified by the FDA as an unsafe herb.^[11] The glycoside and saponin constituents are considered toxic.^[11]

Aesculus hippocastanum is used in Bach flower remedies. When the buds are used it is referred to as “chestnut bud” and when the flowers are used it is referred to as “white chestnut”.

Other chemicals

Quercetin 3,4′-diglucoside, a flavonol glycoside can also be found in horse chestnut seeds.^[16] Leucocyanidin, leucodelphinidin and procyanidin A2 can also be found in horse chestnut.

Anne Frank Tree

A famous specimen of the horse-chestnut was the Anne Frank Tree in the centre of Amsterdam, which she mentioned in her diary and which survived until August 2010, when a heavy wind blew it over.^[17][18] Eleven young specimens, sprouted from seeds from this tree, were transported to the United States. After a long quarantine in Indianapolis, each tree was shipped off to a new home at a notable museum or institution in the United States, such as the 9/11 Memorial Park, Central H.S. in Little Rock, and two Holocaust Centers. One of them was planted outdoors in March 2013 in front of the Children’s Museum of Indianapolis, where they were originally quarantined. [1]

Bonsai

The horse-chestnut is a favourite subject for bonsai.^[19]

Diseases

• Bleeding Canker. Half of all horse-chestnuts in Great Britain are now showing symptoms to some degree of this potentially lethal bacterial infection.^[20][21]
• Guignardia leaf blotch, caused by the fungus Guignardia aesculi
• Wood rotting fungi, e.g. such as Armillaria and Ganoderma
• Horse-chestnut scale, caused by the insect Pulvinaria regalis
• Horse-chestnut leaf miner, Cameraria ohridella, a leaf mining moth.^[22] also affecting large numbers of UK trees.^[21]
• Phytophthora bleeding canker, a fungal infection.^[23]

Google+

ANTHONY MELVIN CRASTO

The European Medicines Agency relies on the results of clinical trials carried out by pharmaceutical companies to reach its opinions on the authorisation of medicines. Although the authorisation of clinical trials occurs at Member State level, the Agency plays a key role in ensuring that the standards of good clinical practice (GCP) are applied across the European Economic Area in cooperation with the Member States. It also manages a database of clinical trials carried out in the European Union.

Clinical trials are studies that are intended to discover or verify the effects of one or more investigational medicines. The regulation of clinical trials aims to ensure that the rights, safety and well-being of trial subjects are protected and the results of clinical trials are credible.

Regardless of where they are conducted, all clinical trials included in applications for marketing authorisation for human medicines in the European Economic Area (EEA) must have been carried out in accordance with the requirements set out in Annex 1 ofDirective 2001/83/EC. This means that:

• clinical trials conducted in the EEA have to comply with European Union (EU) clinical-trial legislation (Directive 2001/20/EC);
• clinical trials conducted outside the EEA have to comply with ethical principles equivalent to those set out in the EEA, including adhering to international good clinical practice and the Declaration of Helsinki.

In the EEA, approximately 4,000 clinical trials are authorised each year. This equals approximately 8,000 clinical-trial applications, with each trial involving two Member States on average. Approximately 61% of clinical trials are sponsored by the pharmaceutical industry and 39% by non-commercial sponsors, mainly academia.

Role of the Agency

Clinical-trial data is included in clinical-study reports that form a large part of the application dossiers submitted by pharmaceutical companies applying for a marketing authorisation via the Agency.

The Agency’s Committee for Medicinal Products for Human Use (CHMP) is responsible for conducting the assessment of a human medicine for which an EU-wide marketing authorisation is sought. As part of its scientific evaluation work, the CHMP reviews the clinical-trial data included in the application.

Assessments are based on purely scientific criteria and determine whether or not the medicines concerned meet the necessary quality, safety and efficacy requirements in accordance with EU legislation, particularly Directive 2001/83/EC.

Good clinical practice

The Agency plays a central role in ensuring application of good clinical practice (GCP). GCP is the international ethical and scientific quality standard for designing, recording and reporting clinical trials that involve the participation of human subjects.

The Agency works in cooperation with GCP inspectors from medicines regulatory authorities (‘national competent authorities’) in EEA Member States on the harmonisation and coordination of GCP-related activity at an EEA level.

The Agency does not have a role in the approval of clinical-trial applications in the EEA. The approval of clinical-trial applications is the responsibility of the national competent authorities.

EudraCT database and the EU Clinical Trials Register

The Agency is responsible for the development, maintenance and coordination of the EudraCT database. This is a database used by national competent authorities to enter clinical-trial data from clinical trial sponsors and paediatric-investigation-plan (PIP) addressees.

A subset of this data is made available through the European Union Clinical Trials Register, which the Agency manages on behalf of EU Member States and forms part ofEudraPharm, the EU database of medicines.

Users are able to view:

• the description of phase-II to phase-IV adult clinical trials where the investigator sites are in the EEA;
• the description of any clinical trials in children with investigator sites in the EU and any trials that form part of a PIP including those where the investigator sites are outside the EU.

As of 21 July 2014, it will be mandatory for sponsors to post clinical trial results in the EudraCT database. A subset of the data included in EudraCT is made available to the public in the European Union Clinical Trials Register. The content and level of detail of these summary results is set out in a European Commission guideline and in its technical guidance. A typical set of summary results provides information on the objectives of a given study, explains how it was designed and gives its main results and conclusions.

The Agency is also working towards the proactive publication of data from clinical trials carried out on the medicines that it authorises. For more information, see release of data from clinical trials.

Clinical trials conducted in countries outside the EU

Clinical trials conducted outside the EU but submitted in an application for marketing authorisation in the EU have to follow the principles which are equivalent to the provisions of the Directive 2001/20/EC.

In April 2012, the Agency published the final version of this paper:

• Reflection paper on ethical and GCP aspects of clinical trials of medicinal products for human use conducted outside of the EU / EEA and submitted in marketing-authorisation applications to the EU regulatory authorities.

This paper aims to strengthen existing processes to provide assurance that clinical trials meet the required ethical and GCP standards, no matter where in the world they have been conducted.

The number of clinical trials and clinical-trial subjects outside Western Europe and North America has been increasing for a number of years. More information is available in this document:

• Clinical trials submitted in marketing-authorisation applications to the European Medicines Agency: Overview of patient recruitment and the geographical location of investigator sites

Revision of EU clinical trial legislation

In July 2012, the European Commission published a proposal on a regulation to revise the EU clinical trial legislation.

More information is available at: Revision of the clinical trials directive.

Clinical Trials Facilitation Group

The Clinical Trials Facilitation Group (CTFG) is a working group of the Heads of Medicines Agencies that:

• acts as forum for discussion to agree on common principles and processes to be applied throughout the European medicines regulatory network;
• promotes harmonisation of clinical-trial-assessment decisions and administrative processes by national competent authorities;
• operates the voluntary harmonisation procedure for assessment of clinical-trial applications involving several Member States.

The Group is composed of representatives from the clinical-trial departments of the national competent authorities.

Terconazole

Systematic (IUPAC) name
1-[4-[ [(2S,4S)-2-(2,4-Dichlorophenyl)-2- (1,2,4-triazol-1-ylmethyl)- 1,3-dioxolan-4-yl]methoxy]phenyl]- 4-propan-2-yl-piperazine
Clinical data
Trade names	Terazol
AHFS/Drugs.com	monograph
MedlinePlus	a688022
Legal status	?
Pharmacokinetic data
Protein binding	94.9%
Identifiers
CAS number	67915-31-5
ATC code	G01AG02
PubChem	CID 441383
DrugBank	DB00251
ChemSpider	390122
UNII	0KJ2VE664U
KEGG	D00888
ChEMBL	CHEMBL1306
Chemical data
Formula	C26H31Cl2N5O3
Mol. mass	532.462 g/mol

Terconazole is an anti-fungal medication, primarily used to treat vaginal fungal infections.

The synthesis of racemic terconazole [J. Heeres et al., J. Med . Chem . , 26 , 611 11983)] is similar. differing in the introduction of a 1 H- 1 , 2,4-triazol-1-yl substituent in place of 1H-imidazol-1-yl and in the nature of the phenol used in the last step of the synthetic sequence, which phenol is 1-methylethyl-4-(4- hydroxyphenyl)piperazme instead of 1-acetyl-4-(4-nydroxyphenyl)piperazine.

Example 20: (2S,4R) -(-)-1-[4-[[2-(2,4-dichlorophenyl)-2-[(1H-1,2,4-triazol-1-yl]methyl-1,3-dioxolane-4-yl]methoxy]phenyl]-4-(1-methylethyl)piperazine, (2S,4R) – (-)-terconazole.

This compound is prepared following the process described for (+)-torconazole, starting from (2S,4S)-(-)-IV (Ar = 2,4-dichlorophenyl, Y = N, R = CH₃) (224 mg, 0.55 mmol), 4-(4-hydroxyphenyl)-1-(1-methylethyl)-piperazine (121 mg, 0.55 mmol), NaH (22.4 mg, 0.56 mmol) in 8 ml of DMSO. (2S,4R) -(-(-terconazole ((2S,4R)-V, Ar

= 2,4-dichlorophenyl, Y = N, Z = CH(CH₃)₂) is obtained as a white solid, m.p. 76-78ºC, [α]_D ²⁰= -12.0 (c = 0.4.

CHCl₃).

Example 17 : (2R,4S)-(+)-1-[4-[[2-(2,4-dichlorophenyl)- 2-[(1H-1,2,4-triazol-1-yl]methyl-1,3-dioxolane-4-yl]methyl]phenyl]-4-(1-methylethyl)piperazine, (2R,4S)-(+)-terconazole.

To a suspension of NaH (60-65% dispersion in paraffin, 36 mg, 0.90 mmol) in anhydrous DMSO (8 ml), 4-(4-hydroxyphenyl) -1 – ( 1-methyle thyl ) p iper az ine ( 193 mg , 0 . 88 mmol ) is added and the mixture is stirred for 1 hour at room temperature. Then, (2R,4R)-(+)-IV (Ar = 2,4-dichlorophenyl, Y = N, R = CH₃ ) is added (180 mg, 0.44 mmol) and the mixture is heated at 80°C for 4 hours. The reaction mixture is allowed to cool to room temperature, diluted with water (20 ml) and extraoteo with CH₂Cl₂ (3 × 25 ml). The combined organic phases are washed with 5N NaOH (3 × 25 ml) and water (3 × 25 ml dried with Na₂SO₄ and the solvent is evaporated of: under vacuum. The oily residue thus obtained is crystallized from diisopropyl ether to give (2R,4S)-(+)-terconazole ((2R,4S)-V, Ar = 2,4-cichlorophenyl, Y = N, Z = CH(CH₃)₂) (140 mg, 59 % yield) as a white solid, m.p. 72-74’C, [α]_D ²⁰ = + 11,05 (c = 0.4, CHCl₃).

IR (KBr), ʋ : 1585, 1512, 1454, 1380, 1270, 1239, 1137, 1048, 979, 820, 675 cm^-1.

¹H-NMR (200 MHz, CDCl₃), δ : 1.11 [d, J=6.5 Hz, 5H, (CH₃)₂CH], 2.73 [m, 5H, 3-H₂, 5-H₂ and (CH₃)₂CH], 3.49

(dd, J=9.6 Hz, J’=6.3 Hz, 1H), 3.80 (m, 2H ) and 3.91

(dd, J=8.2 Hz, J’=6.6 Hz, 1H) (4′ ‘-CH₂ and 5′ ‘-H₂), 4.35

(m, 1H, 4′ ‘-H), 4.74 (d, J=14.6 Hz, 1H) and 4.84 (d, J=14.6 Hz, 1H) (CH₂-N), 6.76 [d, J=9.0 Hz, 2H, C2'(6')- H], 6.88 [d, J=9.0 Hz, 2H, C3'(5')-H], 7.24 (dd, J=8.5

Hz, J’=2.0 Hz, 1H, 5”’-H), 7.46 (d, J=2.0 Hz, 1H,

3″‘-H), 7.56 (d, J=8.5 Hz, 1H, 6″‘-H), 7.89 (s, 1 H) and

8.20 (s, 1H) (triazole 3-H and 5-H).

 

Synthesis pathway

Synthesis a)

• DE 2804096 (Janssen; appl. 3.8.1978; prior. 31.1.1978).
• US 4,358,449 (Janssen; 9.11.1982; prior. 21.11.1977).
• US 4,144,346 (Janssen; 13.3.1979; prior. 21.11.1977, 31.1.1977).
• US 4,223,036 (Janssen; 16.9.1980; prior. 8.1.1979, 21.11.1977, 31.1.1977).
• Heeres, J. et al .: J. Med. Chem. (JMCMAR) 26, 611 (1983).

• Information about Terconazole from the United States government

Pantoprazole

Systematic (IUPAC) name
(RS)-6-(Difluoromethoxy)-2-[(3,4-dimethoxypyridin-2-yl)methylsulfinyl]-1H-benzo[d]imidazole
Clinical data
Trade names	Protonix
AHFS/Drugs.com	monograph
MedlinePlus	a601246
Licence data	US FDA:link
Pregnancy cat.	B3 (AU) B (US)
Legal status	℞ Prescription only
Routes	Oral and intravenous
Pharmacokinetic data
Bioavailability	77%
Metabolism	Hepatic (CYP3A4)
Half-life	1 hour
Excretion	Renal
Identifiers
CAS number	102625-70-7
ATC code	A02BC02
PubChem	CID 4679
DrugBank	DB00213
ChemSpider	4517
UNII	D8TST4O562
KEGG	D05353
ChEBI	CHEBI:7915
ChEMBL	CHEMBL1502
Chemical data
Formula	C16H15F2N3O4S
Mol. mass	383.371 g/mol

Pantoprazole is a proton pump inhibitor drug that inhibits gastric acid secretion.

Pantoprazole is a proton pump inhibitor drug used for short-term treatment of erosion and ulceration of the esophagus caused by gastroesophageal reflux disease.

Use

Pantoprazole is used for short-term treatment of erosion and ulceration of the oesophagus caused by gastroesophageal reflux disease. Initial treatment is generally of eight weeks’ duration, after which another eight week course of treatment may be considered if necessary. It can be used as a maintenance therapy for long term use after initial response is obtained.

Adverse effects

Antacid preparations such as pantoprazole work by suppressing the acid-mediated breakdown of proteins. This leads to an elevated risk of developing food and drug allergies due to undigested proteins passing into the gastrointestinal tract where sensitisation occurs. It is unclear whether this risk occurs with short-term or only long-term use.^[1]

Common

• Gastrointestinal: Abdominal pain (3%), diarrhea (4%), flatulence (4%)
• Neurologic: Headache (5%)

Serious

• Gastrointestinal: Atrophic gastritis, clostridium difficile diarrhea
• Hematologic: Thrombocytopenia (less than 1%)
• Immunologic: Stevens-Johnson syndrome, toxic epidermal necrolysis
• Musculoskeletal: Muscle disorders, bone fracture and infection, Clostridium difficile, osteoporosis-related, hip fracture,rhabdomyolysis
• Renal: Interstitial nephritis (rare)
• Nutrition: May reduce the absorption of important nutrients, vitamins and minerals, as well as medications, leaving users at increased risk for pneumonia.^[2]
• Cardiovascular: Increase in a chemical that suppresses the production of nitric oxide by 25% in humans, which have proven to relax and protect arteries and veins. Causes blood vessels to constrict, a development that could lead to a number of cardiovascular problems if continued for a prolonged period of time.^[2]

Pharmacology

Wyeth pantoprazole 20mg.

Pantoprazole is metabolized in the liver by the cytochrome P450 system.^[3] Metabolism mainly consists of demethylation by CYP2C19followed by sulfation. Another metabolic pathway is oxidation by CYP3A4. Pantoprazole metabolites are not thought to have any pharmacological significance. Pantoprazole is relatively free of drug interactions;^[4] however, it may alter the absorption of other medications that depend on the amount of acid in the stomach, such as ketoconazole or digoxin. Generally inactive at acidic pH of stomach, thus it is usually given with a pro kinetic drug. Pantoprazole binds irreversibly to H+K+ATPase (proton pumps) and suppresses the secretion of acid. As it binds irreversibly to the pumps, new pumps have to be made before acid production can be resumed. The drug’s plasma half-life is about 2 hours.^[5]

Pharmacokinetics

Absorption

• Bioavailability: (oral, delayed release tablets), approximately 77%
• Effect of food: (oral, delayed-release tablets), AUC and C_max no effect, T_max variable, absorption delayed, no net effect
• Effect of food: (oral, for-delayed-release suspension), administer 30 minutes before a meal
• Tmax, Oral, delayed-release suspension: 2 to 2.5 h
• Tmax, Oral, delayed-release tablets: 2.5 h
• Tmax, Oral, delayed-release tablets: 1.5 to 2 hours (pediatrics)

Distribution

• Protein binding: about 98% to primarily albumin
• Vd, extensive metabolizers (IV): approximately 11 L to 23.6 L
• Vd, pediatrics (oral): 0.21 to 0.43 L/kg.

Metabolism

• Hepatic; cytochrome P450 CYP2C19; minor metabolism from CYP3A4, 2D6, and 2C9

Excretion

• Fecal: (oral or IV, normal metabolizers), 18%
• Renal: (oral or IV, normal metabolizers), approximately 71%, none as unchanged
• Dialyzable: no (hemodialysis)
• Total body clearance: (IV) 7.6 to 14 L/hour.
• Total body clearance: (oral, pediatrics) 0.18 to 2.08 L/h/kg

Elimination Half Life

• Oral or IV, 1 hour
• Oral or IV, slow metabolizers, 3.5 to 10 hours
• Pediatrics, 0.7 to 5.34 hours

Availability

Pantoprazole was developed by Altana (owned by Nycomed) and was licensed in the USA to Wyeth (which was taken over by Pfizer). It was initially marketed under the brand name Protonix by Wyeth-Ayerst Laboratories and now is available as a generic. It is available by prescription in delayed-release tablets. It is also available for intravenous use.

On 24 December 2007, Teva Pharmaceutical released an AB-rated generic alternative to Protonix.^[6] This was followed by generic equivalents from Sun Pharma and Kudco Pharma. Wyeth sued all three for patent infringement and launched its own generic version of Protonix with Nycomed.^[7][8]

On October 18, 2010 the U.S. Food and Drug Administration (FDA) accepted the filing of an ANDA for a delayed release generic version of Protonix by Canadian companyIntelliPharmaCeutics.^[9]

Brand names

Pantoprazole is available from a range of international suppliers under brand names including Pantazone, Pantop-D, Pantasan, Pantrol, Prazolin, Pantochem, Pansev, Pantec, Somac, API, Tecta, Protium, Pantodac, Perizole, Pansped, Percazole, Astropan, Fenix, Pantecta, Pantoloc, Controloc, Somac, Tecta, Protium, Inipomp, Eupantol, Pantozol, Pantodac, Perizole, Pansped, Zurcazol, Protonex, Pantup,Pantomed, TopZole, Nolpaza, Controloc, UXL-D, Pantid, Pantogen, Pantpas and Prazolin.

Pantoprazole sodium salt

The structural formula

Brief background information

Salt	ATC	Formula	MM	CAS
-	A02BC02 A02BD04	C 16 H 14 F 2 N 3 NaO 4 S	405.36 g / mol	138786-67-1
hydrate	A02BC02 A02BD04	C 16 H 14 F 2 N 3 NaO 4 S · 3 / 2H 2 O	864.76 g / mol	164579-32-2
(+) – Isomer	A02BC02 A02BD04	C 16 H 14 F 2 N 3 NaO 4 S	405.36 g / mol	160098-11-3
(-) – Isomer	A02BC02 A02BD04	C 16 H 14 F 2 N 3 NaO 4 S	405.36 g / mol	160488-53-9
racemate	A02BC02 A02BD04	C 16 H 14 F 2 N 3 NaO 4 S	405.36 g / mol	142678-34-0

Application

• agent for the treatment of gastric ulcer
• inhibitor of gastric H ⁺ / K ⁺ ATPase

Classes of substances

• Benzimidazoles, 2 (alkylsulfinyl) benzimidazoles
  • Fluoro-ethers
    • Pyridines

 

Country	Patent Number	Approved	Expires (estimated)
Canada	2428870	2006-05-23	2021-11-17
Canada	2092694	2005-04-05	2011-09-06
Canada	2341031	2006-04-04	2019-08-12
United States	7544370	2006-12-07	2026-12-07
United States	4758579	1993-07-19	2010-07-19

 

Synthesis pathway

Synthesis a)

 

http://www.google.com/patents/EP1335913A1?cl=en

Pantoprazole is the international non-proprietary name of the chemical product 5-(difluoromethoxy)-2-[[(3,4-dimethoxy-2- pyridinyl)methyl]sulfmyl]-lH-benzimidazole of formula

 

Pantoprazole This product is an active ingredient used in the treatment of gastric ulcers, usually in the form of its sodium salt.

The product was described for the first time in European patent application EP-A-0166287 that also describes several processes for the preparation of products assignable to a general formula among which pantoprazole is to be found. The reaction sequences of these processes, applied precisely to the preparation of pantoprazole, are given in Scheme 1.

Scheme 1

In Scheme 1, the variables Y, Z, Z’ and Z” are leaving groups, for example atoms of halogen, and the variables M and M’ are atoms of alkali metals.

Austrian patent AT-B-394368 discloses another process based on a different route of synthetis, the reaction sequence of which is given in Scheme 2.

 

Pantoprazole Scheme 2

Nevertheless, this process has obvious drawbacks, since the methylation can take place not only in OH in the 4-position of the pyridine ring, but also in the nitrogen linked to a hydrogen of the benzimidazole ring, which can give place to mixtures of the desired product with the two possible methylated isomers of the benzimidazole compounds obtained, 3- methyl or 1 -methyl, which means that additional chromatographic purification steps are needed and the yields obtained are low.

PCT application WO97/29103 discloses another process for the preparation of pantoprazole, the reaction sequence of which is given in Scheme 3.

 

Scheme 3 As may be seen, different synthesis strategies have been proposed for the preparation of pantoprazole, some of them recently, which is an indication that the preparation of the product is still not considered to be sufficiently well developed, whereby there is still a need for developing alternative processes that allow pantoprazole to be prepared by means of simpler techniques and more accessible intermediate compounds and with good chemical yields.

EXAMPLES

Example 1. – Preparation of compound (IX)

 

47.5 ml (0.502 mol) of acetic anhydride were mixed with 1.65 g (0.0135 mol) of 4-dimethylaminopyridine, giving a transparent yellow solution which was heated to 65° – 70°C. This temperature was held by cooling since the reaction is exothermic. 25 g (0.1441 mol) of 2-methyl-3- methoxy-4-chloropyridine N-oxide (X) were added over a period of about 70 minutes. Once the addition was completed, the reaction was held at 65° – 70°C for a further 2h 20 minutes and after this time it was allowed to cool down to below 65°C and 90 ml of methanol were added gradually, while holding the temperature below 65°C. The resulting reaction mass was distilled at reduced pressure in a rotavap to remove the volatile components and the residue containing compound (IX) was used as such for the following reaction. Thin layer chromatography on silica gel 60 F₂₅₄, eluting with CHCl₃/MeOH (15: 1), showed a single spot at Rf – 0.82, indicating that the reaction has been completed.

Example 2. – Preparation of compound fVIII

 

(IX) (VIII)

11.5 ml methanol and 11.5 ml of water were added over the crude residue from Example 1 containing compound (IX), and thereafter, while holding the temperature to between 25° and 30°C with a water bath, the residual acetic acid contained in the crude residue was neutralized by the addition of 33% aqueous NaOH. Once the residual acid had been neutralized, 19 ml (0.2136 mol) of the 33% aqueous NaOH were added over 20 minutes, while holding the temperature to between 25° and 30°C, and, on completion of the addition, the hydrolysis reaction at pH 11.7 – 11.8 was held for 2h 30 minutes, to between 25° and 30°C. On completion of the reaction, the pH was adjusted to 7.0 – 7.5 by the addition of HC1 35%, while holding the temperature to 25°C. Thereafter, 50 ml of methylene chloride were added and, after stirring and allowing to rest, the phases were decanted. A further five extractions were carried out with 30 ml methylene chloride each and the pooled organic phases were dried with anhydrous sodium sulfate, were filtered and washed, and were evaporated at reduced pressure in a rotavap, providing a solid residue having a melting point around 73°C and containing compound (VIII). Thin layer chromatography on silica gel 60 F₂₅₄, eluting with CHCl₃/MeOH (15: 1), gave a main spot at Rf = 0.55, showing that the reaction was complete. The thus obtained crude residue was used as such in the following reaction.

Example 3. – Preparation of compound (VI)

 

24.5 g of the residue obtained in Example 2, containing approximately 0.142 mol of the compound 2-hydroxymethyl-3-methoxy-4-chloropyridine (VIII), were mixed with 0.5 ml of DMF and 300 ml of anhydrous methylene chloride, to give a brown solution which was cooled to 0° – 5°C in an ice water bath. Thereafter, a solution of 11.5 ml (0.1585 mol) of thionyl chloride in 50 ml of anhydrous methylene chloride was added over 20 minutes, while holding the above-mentioned temperature,. Once the addition was complete, the reaction was held at 0° – 5°C for a further 90 minutes and then 120 ml of water and NaOH 33% were added to pH 5 – 6, requiring approximately 29 ml of NaOH. The phases were then decanted and separated. The organic phase was extracted with a further 120 ml of water and the pooled aqueous phases were extracted with a further 4×25 ml of methylene chloride, in order to recover the greatest possible amount of product. The pooled organic phases were dried over anhydrous sodium sulfate, filtered and washed, and evaporated at reduced pressure in a rotavap, to give a residue containing the compound 2-chloromethyl-3- methoxy-4-chloropyridine (VI). Thin layer chromatography on silica gel 60 F₂₅₄, eluting with CHCl₃/MeOH (15:1), showed a main spot at Rf = 0.83, indicating that the reaction was complete. The thus obtained crude residue was used as such in the following reaction. Example 4. – Preparation of compound (III)

 

26.11 g of the residue obtained in the Example 3 containing approximately 0.136 mol of the compound 2-chloromethyl-3-methoxy-4- chloropyridine (VI) were mixed with 370 ml of methylene chloride, to give a brown solution over which were added, at 20° – 25°C, 29.3 g (0.136 mol) of 5-difluoromethoxy-2-mercaptobenzimidazole (VII) and 17.10 ml (0.136 mol) of tetramethylguanidine (TMGH). The mixture was stirred at this temperature for 2 hours, after which 450 ml of water were added, with the pH being held to between 9.5 and 10. Thereafter the phases were decanted and the organic phase was washed 5×50 ml of a IN NaOH aqueous solution and, thereafter, with 2×50 ml of water. The organic phase was treated with 50 ml of water and an amount of HC1 30% sufficient to adjust the pH to between 5 and 6. Thereafter, the phases were decanted, and the organic phase was dried over anhydrous sodium sulfate, was filtered and washed, and evaporated at reduced pressure in a rotavap, to give a solid residue of melting point 64° – 73 °C that contains the compound (III). Thin layer chromatography on silica gel 60 F₂₅₄, eluting with CHCl₃/MeOH (15: 1), presented a main spot at Rf = 0.52. Yield 82%. The thus obtained compound 5-(difluoromethoxy)-2-[[(3-methoxy-4-chlorine-2 pyridinyl)methyl]mercapto]- lH-benzimidazole (III) was used as such in the following reaction Example 5. – Preparation of compound (IV)

 

25.8 g (0.0694 mol) of the compound (III) obtained in the Example 4 were mixed with 88 ml of methanol, to give a brown solution to which 3.7 ml of water, 0.99 g of ammonium molybdate and 0.78 g of sodium carbonate were added. The system was cooled to 0°C – 5°C, 3.4 ml (0.0756 mol) of 60% hydrogen peroxide were added, and the reaction mixture was held at 0°C – 5°C for 1 – 2 days, the end point of the reaction being checked by thin layer chromatography on silica gel 60 F₂₅₄, eluting with CHCl₃/MeOH (15: l).

During the reaction the presence of hydrogen peroxide in the reaction medium was controlled by testing with potassium iodide, water and starch. When effected on a sample containing hydrogen peroxide, it provides a brown-black colour. If the assay is negative before the chromatographic control indicates completion of the reaction, more hydrogen peroxide is added.

On completion of the reaction, 260 ml of water were added, the system was cooled to 0°C – 5°C again and the mixture was stirred for 2 hours at this temperature. The solid precipitate was filtered, washed with abundant water, and dried at a temperature below 60°C, to give 5-(difluoromethoxy)-2-[[(3- methoxy-4-chlorine-2-pyridinyl)methyl]sulfinyl]-lH-benzimidazole (IV), melting point 130° – 136°C, with an 83.5% yield. Thin layer chromatography on silica gel 60 F₂₅₄, eluting with CHCl₃/MeOH (15: 1), gave a main spot at Rf = 0.5.

Compound (IV) can be purified, if desired, by the following crystallization method:

5 g of crude product was suspended in 16 ml of acetone and was heated to boiling until a dark brown solution was obtained. Thereafter the thus obtained solution was allowed to cool down to room temperature and then was then chilled again to -20°C, at which temperature the mixture was held for 23 hours without stirring. Thereafter the solid was filtered and washed with 6×4 ml of acetone chilled to -20°C. Once dry, the resulting white solid weighed 2.73 g, had a point of melting of 142°C and gave a single spot in thin layer chromatography. The IR spectrum of the compound on KBr is given in Figure 1.

The acetonic solution comprising the mother liquors of filtration and the washes was concentrated to a volume of 20 ml and a further 5 g of crude compound were added. The above described crystallization process was repeated to obtain a further 4.11 g of purified product of characteristics similar to the previous one.

The acetonic solution from the previous crystallization was concentrated to a volume of 17 ml and a further 4 g of crude compound were added. The above described crystallization process was repeated to obtain a further 2.91 g of purified product of similar characteristics to the previous ones.

The acetonic solution from the previous crystallization was concentrated to a volume of 15 ml and a further 4 g of crude compound were added. The above described crystallization process was repeated to obtain a further 3.3 g of purified product of similar characteristics to the previous ones.

The acetonic solution from the previous crystallization was concentrated to a volume of 16 ml and a further 4.36 g of crude compound were added. The above described crystallization process was repeated to obtain a further 3.62 g of purified product of similar characteristics to the previous ones.

Finally, the acetonic solution from the previous crystallization was concentrated to a volume of 10 – 12 ml and held at -20°C for two days without stirring. Thereafter, the solid was filtered and washed with 5×3 ml of acetone chilled to -20°C. Once dry, the solid weighed 1.26 g and had similar characteristics to the previous ones.

The total yield of all the crystallizations was 80%.

Example 6. – Preparation of pantoprazole

 

12.95 g (0.0334 mol) of compound (IV) purified by crystallization of Example 5 were mixed with 38 ml of N,N-dimethylacetamide and thereafter 7.03 g (0.1003 mol) of potassium methoxide were added, while holding the temperature to between 20°C and 30°C, whereby a dark brown mixture was obtained. The system was held at approximately 25°C for about 23 hours, after which, once the reaction was complete, the pH was adjusted to 7 with the addition of 3.82 ml of acetic acid. The N,N-dimethylacetamide was removed at reduced pressure at an internal temperature of not more than 75°C. 65 ml of water and 50 ml of methylene chloride were added over the thus obtained residue, followed by decantation of the phases. Once the phases were decanted, the aqueous phase was extracted a with further 3×25 ml of methylene chloride, the organic phases were pooled and the resulting solution dried over anhydrous sodium sulfate, was filtered and washed, and evaporated at reduced pressure in a rotavap, to give a crude residue over which 55 ml of water were added, to give a suspension (if the product does not solidify at this point the water is decanted and a further 55 ml of water are added to remove remains of N,N-dimethylacetamide that hinder the solidification of the product). The solid was filtered and, after drying, 11.61 g of crude pantoprazole of reddish brown colour were obtained (Yield 90%). The thus obtained crude product was decoloured by dissolving the crude product in 150 ml of methanol, whereby a dark brown solution was obtained. 7.5 g of active carbon were added, while maintaining stirring for 45 minutes at 25°C – 30°C, after which the carbon was filtered out and the filter was washed. The methanol was then removed in the rotavap at reduced pressure, a temperature below 40°C. 10.33 g of a solid residue were obtained and were mixed with 14.9 ml of methylethylketone, and the suspension was heated to 45°C for about 10 minutes, after which it was cooled, first to room temperature and then to -20°C. This temperature was held over night and thereafter the solid was filtered, washed with 6×5 ml of methylethylketone chilled to -20°C. Once dry, 7.75 g of a white solid, melting point 140°C – 141 °C, were obtained. Thin layer chromatography on silica gel F₂₅₄, eluting with CHCl₃/MeOH (15: 1), gave a single spot at Rf =

0.41 and a IR spectrum corresponding identically with that of pantoprazole.

The ketonic solution comprising the mother liquors of filtration and the washes, was concentrated to 9.7 ml, was heated to 40°C, was held at this temperature for about five minutes and was then cooled, first to room temperature and then to -20°C, this temperature being held for 4 hours. At the end of this time, the solid was filtered and was washed with 4×2 ml of methylethylketone chilled to -20°C. Once dry, 0.42 g of a white solid of similar characteristics to the previous one was obtained.

The ketone solution from the previous treatment was concentrated to 3.1 ml, was heated to 40°C, was held to this temperature for about five minutes and then was cooled, first to room temperature and then to -20°C, this temperature being held for 4 hours. At the end of this time, the solid was filtered and was washed with 5×3 ml of methylethylketone chilled to – 20°C. Once dry, 0.41 g of a white-beige solid of similar characteristics to the previous one was obtained. The total yield, including purifications, was 67%.

If a whiter solid is desired, one or several washes can be carried with isopropyl acetate as follows: 6.6 g of pantoprazole from the methylethylketone treatment were suspended in 50 ml of isopropyl acetate. The system (white suspension) was stirred for about 30 minutes at 25°C, was then cooled to 0°C – 5°C, was stirred for about 15 minutes at this temperature and the solid was then filtered, was washed with 3×15 ml of isopropyl acetate. Once dry, 6.26 g of a pure white solid were obtained.

 

 

 

Trade Names

Country	Trade name	Manufacturer
Germany	Pantozol	Nycomed
	Rifun	- “-
France	Eupantol	Altana
	Inipomp	Sanofi-Aventis
United Kingdom	Protium	ALTANA
Italy	Pantekta	Abbott
	Pantopan	Pharmacia
	Pantork	Altana
USA	Protonix	Wyeth
Ukraine	Kontrolok	Nycomed Oranienburg GmbH, Germany
	Nolpaza	Krka
	Pultset	Nobel Ilach Sanayi ve Ticaret AS, Turkey
	Proksium	JSC “Lubnyfarm”, Ukraine
	various generic drugs

Formulations

• ampoule 40 mg;
• Tablets 40 mg

UV – spectrum

Conditions : Concentration – 1 mg / 100 ml
Solvent designation schedule	Methanol	Water	0.1 M HCl	0.1M NaOH
The absorption maximum	289 nm	291nm	Observed decay	295 nm
	391	346	-	418
ε	16600	14700	-	17700

IR – spectrum

Wavelength (μm)
Wavenumber (cm -1 )

NMR Spectrum

 will be added

 

 

Links

• EP 134 400 (Byk Gulden Lomberg; appl. 1.5.1984; CH-prior. 3.5.1983).
• US 4,555,518 (Byk Gulden Lomberg; 26.11.1985; appl. 1.5.1984; CH-prior. 3.5.1983).
• US 4,758,579 (Byk Gulden Lomberg; 19.7.1988; appl. 28.4.1987; CH-prior. 16.6.1984).

• UV and IR Spectra. H.-W. Dibbern, RM Muller, E. Wirbitzki, 2002 ECV
• NIST / EPA / NIH Mass Spectral Library 2008
• Handbook of Organic Compounds. NIR, IR, Raman, and UV-Vis Spectra Featuring Polymers and Surfactants, Jr., Jerry Workman.Academic Press, 2000.
• Handbook of ultraviolet and visible absorption spectra of organic compounds, K. Hirayama. Plenum Press Data Division, 1967.

References

1.  Pali-Schöll I, Jensen-Jarolim E (April 2011). “Anti-acid medication as a risk factor for food allergy”. Allergy 66 (4): 469–77. doi:10.1111/j.1398-9995.2010.02511.x. PMID 21121928.
2.  [Dr. John Cooke, chair of Methodist Hospital's cardiovascular services] [Houston Chronicle Health Zone dated Thursday, July 11, 2013 chron.com/refluxmeds] (Journal: Circulation)
3. Jump up^ Meyer, U A (1996). “Metabolic interactions of the proton-pump inhibitors lansoprazole, omeprazole and pantoprazole with other drugs”. European journal of gastroenterology & hepatology8 (Suppl 1): S21–25. doi:10.1097/00042737-199610001-00005.
4.  Steinijans, V. W.; Huber, R.; Hartmann, M.; Zech, K.; Bliesath, H.; Wurst, W.; Radtke, H. W. (1996). “Lack of pantoprazole drug interactions in man: An updated review”. International Journal of Clinical Pharmacology and Therapeutics 34 (6): 243–262. PMID 8793611.
5.  Sachs G, Shin JM, Hunt R (December 2010). “Novel approaches to inhibition of gastric acid secretion”. Curr Gastroenterol Rep 12 (6): 437–47. doi:10.1007/s11894-010-0149-5.PMC 2974194. PMID 20924727.
6.  Teva Announces Launch Of Generic Protonix Tablets
7. Jump up^ Rubenstein, Sarah (29 January 2008). “Wyeth Plans Generic Protonix; Litigation With Teva to Continue”. The Wall Street Journal. p. D9. Retrieved 25 October 2009.
8. Jump up^ “Nycomed and Wyeth announce launch of an own generic version of PROTONIX – lawsuit to defend patent continues”. Retrieved 25 October 2009.^{[dead link]}
9. Jump up^ IntelliPharmaCeutics Press Release

External links

• Protonix (Pantoprazole Sodium) label