A step-wise integrated risk-based approach to determine a control strategy for according to ICH Q3D has to consider data from all kinds of potential sources for elemental impurities in particular from excipients. Read more about the newly created Elemental Impurities Database as a valuable support for performing risk assessments for drug products.

http://www.gmp-compliance.org/eca_mitt_05257_15499_n.html

The new ICH Q3D Guideline on Elemental Impurities strongly advocates the use of risk assessments in order to define a final control strategy. Specific challenges appear when risks associated with production equipment, packaging material and excipients have to be determined, and quantified. In particular the contribution of elemental impurities from excipients is not easy to assess due to their big variety and the lack of information from excipient vendors.

Quite recently a pharma consortium started an initiative which aims to collect and share data from pharmaceutical excipients by establishing a database. This Elemental Impurities (EI) Database provides information required for performing a comprehensive risk assessment of a drug product with respect to elemental impurities. Interested companies can contribute to this database by providing information about excipients and may also benefit from this database by taking out information needed for their risk assessments.

The “Impurities Workshop” from 14-16 June 2016 in Heidelberg, Germany provides a comprehensive and practical oriented review of impurities analysis and characterisation in drug substances and drug products. Part III of the workshop on 16 June 2016 is specifically dedicated to Elemental Impurites. In the subsequent post-Conference Workshop on 17 June 2016 the above mentioned EI Database will be explained. The following questions will be discussed:

• What is the procedure of providing data for the Database?
• How can information be obtained from the Database?
• What has to be considered in terms of confidentiality when data will be received or submitted to the Database?

This post-Conference Workshop is free of charge. It ideally complements the previous parts of the “Impurities Workshop” and can be booked in combination with either Part III or all Parts of the “Impurities Workshop”. As we expect a high interest in this post-Conference Workshop participants joining the “Impurities Workshop” (one day or all three days) will be registered first

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Supplier Qualification is more than auditing. Supplier qualification can be seen as a risk assessment tool. But what are the exact requirements for qualifying suppliers?

http://www.gmp-compliance.org/enews_05231_What-are-the-current-Rules-for-Supplier-Qualification_15159,15099,15179,Z-QAMPP_n.html

Supplier Qualification is more than auditing. Supplier qualification can be seen as a risk assessment tool. It should provide an appropriate level of confidence that suppliers, vendors and contractors are able to supply consistent quality of materials, components and services in compliance with regulatory requirements. An integrated supplier qualification process should also identify and mitigate the associated risks of materials, components and services. But what are the exact requirements?

They are wide-ranging and complex. There are different directives and regulations for medicinal drug products for human or veterinary use and for investigational medicinal drug products. Certain requirements in different directives and the EU-GMP Guidelines define expectations. Here are some examples:

Article 8 of EU-Directive 2001/83/EC
“The application [of a marketing authorization] shall be accompanied […] by […] a written confirmation that the manufacturer of the medicinal product has verified compliance of the manufacturer of active substance with principles and guidelines of good manufacturing practice by conducting audits.”

Article 46 of EU-Directive 2001/83/EC
“The holder of a manufacturing and/or import authorisation shall at least be obliged […] to use only active substances, which have been manufactured in accordance with GMP for active substances and distributed in accordance with GDP for active substances and … to ensure that the excipients are suitable for use in medicinal products by ascertaining what the appropriate GMP is.”

Article 46b of EU-Directive 2001/83/EC
“Active substances shall only be imported if they have been manufactured in accordance with standards of good manufacturing practice at least equivalent to those laid down by the European Union”. This can be shown by a written confirmation, or the exporting country is included in the so called white list, or a waiver has been granted.

EU-GMP Guidelines Chapter 5:
5.25 “The purchase of starting materials is an important operation which should involve staff who have a particular and thorough knowledge of the supplier.”
5.26 “Starting materials should only be purchased from approved suppliers …”
5.40 “…printed packaging materials shall be accorded attention similar to that given to starting materials.”

The revised Chapter 7 of the EU-GMP Guidelines describe the responsibilities of the Contract Giver when it comes to contract manufacturing and testing. He needs to assure the control of the outsourced activities, incorporating quality risk management principles and including continuous reviews of the quality of the Contract Acceptor’s performance. Audits are a helpful tool to asses the “legality, suitability and the competence of the Contract Acceptor“. The new Chapter 7 was obviously designed to intensify the control of Contract Acceptors by the Contract Giver and extend those controls to subcontractors.

The holder of the manufacturing authorisation is responsible for the supplier qualification by law but in fact the supplier qualification is one of the duties of the Qualified Person (which can be delegated) as defined in Annex 16 of the EU-GMP Guidelines. The QP of the marketing authorisation holder is responsible for certifying the drug product for the market place and is now being held accountable to ensure that all aspects of the supply chain have been made under the appropriate GMPs. However, according to Chapter 2 of the EU-GMP Guidelines, the heads of Production, Quality Control and Quality Assurance share the responsibility of approving and monitoring suppliers of materials (2.9).

So how to proceed? At the beginning of a supplier qualification process, the regulatory requirements regarding the type of material, component or service and the type of product (human/veterinary drug product or IMP) should be identified and specified. Audits, if required, should be planned and executed. The compliance of the selected supplier(s) with the requirements and user requirement specification should be demonstrated. The scope of an audit should cover this. But a successful audit is not the end of the qualification process. After finalising the contract, the compliance of the selected supplier(s) with the applicable requirements should be evaluated periodically. Changes at the supplier´s site (for example manufacturing process etc.) that pose a particular risk to the compliance with the requirements should be assessed. There needs to be a mechanism in place so that any change made by the supplier which could have an impact on the GMP status or the production or testing parameters have to be agreed to before any such changes are implemented. A supplier must also notify the contract giver immediately upon discovery of any deviation/non-conformance/complaint that may have an impact on the services provided. Those need to be assessed and respective actions need to be defined.

The use of Brokers:
Some raw materials are only available at reasonable costs if purchased through an intermediary, i.e. a Broker. If the material is critical to the process, e.g. an API or a key excipient this can give an added complexity to the process and this must be fully investigated with the Quality and Regulatory units being involved, before any orders are placed.

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ITI 214

IC200214; ITI-214

(6aR,9aS)-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-3-(phenylamino)-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[1,2-a]pyrazolo[4,3-e]pyrimidin-4-(2H)-one phosphate

(6aR,9aS)-5-methyl-3-(phenylamino)-2-(4-(6-fluoropyridin-2-yl)-benzyl)-5,6a,7,8,9,9a-hexahydrocyclopent[4,5]imidazo[1,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one…BASE

CAS: 1642303-38-5 (phosphate);

1160521-50-5 (free base). 

Chemical Formula: C29H29FN7O5P
Molecular Weight: 605.5672

Takeda Pharmaceutical Company Limited,Intra-Cellular Therapies, Inc.

ITI-214 is an orally active, potent and Selective Inhibitors of Phosphodiesterase 1 for the Treatment of Cognitive Impairment Associated with Neurodegenerative and Neuropsychiatric Diseases. ITI-214 exhibited picomolar inhibitory potency for PDE1, demonstrated excellent selectivity against all other PDE families, and showed good efficacy in vivo. Currently, this investigational new drug is in Phase I clinical development and being considered for the treatment of several indications including cognitive deficits associated with schizophrenia and Alzheimer’s disease, movement disorders, attention deficit and hyperactivity disorders, and other CNS and non-CNS disorders.

• Phase I  Cognition disorders
• • OriginatorIntra-Cellular Therapies
  • ClassAntiparkinsonians; Nootropics; Small molecules
  • Mechanism of ActionType 1 cyclic nucleotide phosphodiesterase inhibitors

• 21 Sep 2015Takeda completes a phase I bioavailability trial in Cognition disorders in Japan
• 21 Sep 2015Takeda completes a phase I trial in Cognition disorders in Japan
• 21 Sep 2015Takeda initiates enrolment in a phase I bioavailability trial for Cognition disorders in Japan before September 2015

Phosphodiesterase-1 (PDE-1) inhibitor

which is a picomolar PDE1 inhibitor with excellent selectivity against other PDE family members and against a panel of enzymes, receptors, transporters, and ion channels.

It is disclosed in WO 2009/075784 (U.S. Pub. No. 2010/0273754). This compound has been found to be a potent and selective phosphodiesterase 1 (PDE 1) inhibitor useful for the treatment or prophylaxis of disorders characterized by low levels of cAMP and/or cGMP in cells expressing PDE1, and/or reduced dopamine Dl receptor signaling activity (e.g., Parkinson’s disease, Tourette’s Syndrome, Autism, fragile X syndrome, ADHD, restless leg syndrome, depression, cognitive impairment of schizophrenia, narcolepsy); and/or any disease or condition that may be ameliorated by the enhancement of progesterone signaling. This list of disorders is exemplary and not intended to be exhaustive.

PATENT

WO 2013192556

http://www.google.com/patents/WO2013192556A2?cl=en

The method of making the Compound (ea^^a^-S^a ^^^a-hexahydro-S- methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)- cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one is generally described in WO 2009/075784, the contents of which are incorporated by reference in their entirety. This compound can also be prepared as summarized or similarly summarized in the following

CMU PCU PHU PPU (SM2)

In particular, (6aR,9aS)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl- 5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)- one may be prepared as described or similarly described below.

PATENT

http://www.google.com/patents/WO2009075784A1?cl=en

EXAMPLE 14

(6aJ?,9aS)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6- fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]iinidazo[l,2-fl]pyrazolo[4,3- e]pyrimidin-4(2//)-one

This compound may be made using similar method as in example 13 wherein 2-(4-(bromomethyl)phenyl)-6-fluoropyridine may be used instead of 2-(4- (dibromomethyl)phenyl)-5-fluoropyridine.

PATENT

WO 2014205354

https://www.google.co.in/patents/WO2014205354A2?cl=en

EXAMPLES

The method of making the Compound (ea^^a^-S^a ^^^a-hexahydro-S-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one is generally described in WO 2009/075784, the contents of which are incorporated by reference in their entirety. This compound can also be prepared as summarized or similarly summarized in the following

CMU PCU PHU PPU (SM2)

In particular, (6aR,9aS)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one (Int-5) may be prepared as described or similarly described below. The free base crystals and the mono-phosphate salt crystals of the invention may be prepared by using the methods described or similarly described in Examples 1-14 below.

Preparation of (6aR,9aS)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one

(4-(6-fluoropyridin-2-yl)phenyl)methanol

The mixture of Na₂C0₃ (121 g), water (500 mL), THF (650 mL), PdCl₂(PPh₃)₂ (997 mg), 2-bromo-6-fluoropyridine (100 g) and 4-(hydroxymethyl)phenylboronic acid (90.7 g) is stirred at 65°C for 4 h under the nitrogen atmosphere. After cooling to room temperature, THF (200 mL) is added. The organic layer is separated and washed with 5% NaCl solution twice. The organic layer is concentrated to 400 mL. After the addition of toluene (100 mL), heptane (500 mL) is added at 55°C. The mixture is cooled to room temperature. The crystals are isolated by filtration, washed with the mixture of toluene (100 mL) and heptane (100 mL) and dried to give (4-(6-fluoropyridin-2-yl)phenyl)methanol (103 g). ^]H NMR (500 MHz, CDC1₃) δ 1.71-1.78 (m, 1H), 4.74-4.79 (m, 2H), 6.84-6.88 (m, 1H), 7.44-7.50 (m, 2H), 7.61-7.65 (m, 1H), 7.80-7.88 (m, 1H), 7.98-8.04 (m, 2H).

2-(4-(chloromethyl)phenyl)-6-fluoropyridine

The solution of thionylchloride (43.1 mL) in AcOEt (200 mL) is added to the mixture of (4-(6-fluoropyridin-2-yl)phenyl)methanol (100 g), DMF (10 mL) and AcOEt (600 mL) at room temperature. The mixture is stirred at room temperature for 1 h. After cooling to 10°C, 15% Na₂C0₃ solution is added. The organic layer is separated and washed with water (500 mL) and 5% NaCl solution (500 mL) twice. The organic layer is concentrated to 500 mL. After the addition of EtOH (500 mL), the mixture is concentrated to 500 mL. After addition of EtOH (500 mL), the mixture is concentrated to 500 mL. After the addition of EtOH (500 mL), the mixture is concentrated to 500 mL. After addition of EtOH (200 mL), water (700 mL) is added at 40°C. The mixture is stirred at room temperature. The crystals are isolated by filtration and dried to give 2-(4-(chloromethyl)phenyl)-6-fluoropyridine (89.5 g). ^]H NMR (500 MHz, CDC1₃) δ 4.64 (s, 2H), 6.86-6.90 (m, 1H), 7.47-7.52 (m, 2H), 7.60-7.65 (m, 1H), 7.82-7.88 (m, 1H), 7.98-8.03 (m, 2H).

6-chloro-l-(4-methoxybenzyl)-3-methylpyrimidine-2,4(lH,3H)-dione

The mixture of 6-chloro-3-methyluracil (100 g), p-methoxybenzylchloride (107 g), K2CO₃ (86.1 g) and DMAc (600 mL) is stirred at 75°C for 4 h. Water (400 mL) is added at 45°C and the mixture is cooled to room temperature. Water (800 mL) is added and the mixture is stirred at room temperature. The crystals are isolated by filtration, washed with the mixture of DMAc and water (1:2, 200mL) and dried to give 6-chloro-l-(4-methoxybenzyl)-3-methylpyrimidine-2,4(lH,3H)-dione (167 g). ^]H NMR (500 MHz, CDC1₃) δ 3.35 (s, 3H), 3.80 (s, 3H), 5.21 (s, 2H), 5.93 (s, 1H), 6.85-6.89 (m, 2H), 7.26-7.32 (m, 2H).

izinyl-l-(4-methoxybenzyl)-3-methylpyrimidine-2,4(lH,3H)-dione

The mixture of 6-chloro-l-(4-methoxybenzyl)-3-methylpyrimidine-2,4(lH,3H)-dione (165 g), IPA (990 mL), water (124 mL) and hydrazine hydrate (62.9 mL) is stirred at room temperature for 1 h. The mixture is warmed to 60°C and stirred at the same temperature for 4 h. Isopropyl acetate (1485 mL) is added at 45°C and the mixture is stirred at the same temperature for 0.5 h. The mixture is cooled at 10°C and stirred for lh. The crystals are isolated by filtration, washed with the mixture of IPA and isopropyl acetate (1:2, 330 mL) and dried to give 6-hydrazinyl-l-(4-methoxybenzyl)-3-methylpyrimidine-2,4(lH,3H)-dione (153 g). ^]H NMR (500 MHz, DMSO-i¾) δ 3.12 (s, 3H), 3.71 (s, 3H), 4.36 (s, 2H), 5.01 (s, 2H), 5.14 (s, 1H), 6.87-6.89 (m, 2H), 7.12-7.17 (m, 2H), 8.04 (s, 1H).

7-(4-methoxybenzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione

To the mixture of DMF (725 mL) and 6-hydrazinyl-l-(4-methoxybenzyl)-3-methylpyrimidine-2,4(lH,3H)-dione (145 g) is added POCI₃ (58.5 mL) at 5°C. The mixture is stirred at room temperature for 1 h. Water (725 mL) is added at 50°C and the mixture is stirred at room temperature for 1 h. The crystals are isolated by filtration, washed with the mixture of DMF and water (1:1, 290 mL) and dried to give 7-(4-methoxybenzyl)-5-methyl-

2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione (145 g). ^]H NMR (500 MHz, DMSO-i¾) δ 3.23 (s, 3H), 3.71 (s, 3H), 5.05 (s, 2H), 6.82-6.90 (m, 2H), 7.28-7.36 (m, 2H), 8.48 (s, IH), 13.51 (br, IH).

2-(4-(6-fluoropyridin-2-yl)benzyl)-7-(4-methoxybenzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione

The mixture of 2-(4-(chloromethyl)phenyl)-6-fluoropyridine (100 g), 7-(4-methoxybenzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione (129 g), K2CO₃(62.3 g) and DMAc (1500 mL) is stirred at 45°C for 5 h. Water (1500 mL) is added at 40°C and the mixture is stirred at room temperature for 1 h. The crystals are isolated by filtration, washed with the mixture of DMAc and water (1:1, 500 mL) and dried to give 2-(4-(6-fluoropyridin-2-yl)benzyl)-7-(4-methoxybenzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione (207 g). ^]H NMR (500 MHz, DMSO- ) δ 3.21 (s, 3H), 3.66 (s, 3H), 4.98 (s, 2H), 5.45 (s, 2H), 6.77-6.82 (m, 2H), 7.13-7.16 (m, IH), 7.25-7.30 (m, 2H), 7.41-7.44 (m, 2H), 7.92-7.96 (m, IH), 8.04-8.11 (m, 3H), 8.68 (s, IH).

2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione

The mixture of 2-(4-(6-fluoropyridin-2-yl)benzyl)-7-(4-methoxybenzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione (105 g), CF₃COOH (300 mL) and

CF₃SO₃H (100 g) is stirred at room temperature for 10 h. Acetonitrile (1000 mL) is added. The mixture is added to the mixture of 25% N¾ (1000 mL) and acetonitrile (500 mL) at 10°C. The mixture is stirred at room temperature for 1 h. The crystals are isolated by filtration, washed with the mixture of acetonitirile and water (1:1, 500 mL) and dried to give the crude product. The mixture of the crude product and AcOEt (1200 mL) is stirred at room temperature for 1 h. The crystals are isolated by filtration, washed with AcOEt (250 mL) and dried to give 2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione (75.3 g). ^]H NMR (500 MHz, DMSO-rf₆) δ 3.16 (s, 3H), 3.50-4.00 (br, 1H), 5.40 (s, 2H), 7.13-7.16 (m, 1H), 7.41-7.44 (m, 2H), 7.91-7.94 (m, 1H), 8.04-8.10 (m, 3H), 8.60 (s, 1H).

2-(4-(6-fluoropyridin-2-yl)benzyl)-6-(((lR,2R)-2-hydroxycyclopentyl)amino)-5-methyl-2H-pyrazolo[3,4-d]pyrimidin-4(5H)-one

The mixture of BOP reagent (126 g), 2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-2H-pyrazolo[3,4-d]pyrimidine-4,6(5H,7H)-dione (80 g), DBU (136 mL) and THF (1120 mL) is stirred at room temperature for 1 h. (lR,2R)-2-Aminocyclopentanol hydrochloride (37.6 g) and THF (80 mL) are added and the mixture is stirred at room temperature for 5 h. After the addition of 5% NaCl (400 mL) and AcOEt (800 mL), the organic layer is separated. The organic layer is washed with 10% NaCl (400 mL), 1M HC1 15% NaCl (400 mL), 5% NaCl (400 mL), 5% NaHC0₃ (400 mL) and 5%NaCl (400 mL) successively. After treatment with active charcoal, the organic layer is concentrated to 400 mL. After the addition of acetonitrile (800 mL), the mixture is concentrated to 400 mL. After the addition of acetonitrile (800 mL), seed crystals are added at 40°C. The mixture is concentrated to 400 mL. Water (800 mL) is added at room temperature and the mixture is stirred for 2 h. The crystals are isolated by filtration, washed with the mixture of acetonitrile and water (1:2, 400 mL) and dried to give 2-(4-(6-fluoropyridin-2-yl)benzyl)-6-(((lR,2R)-2-

hydroxycyclopentyl)amino)-5-methyl-2H-pyrazolo[3,4-d]pyrimidin-4(5H)-one (81.7 g). ^]H NMR (500 MHz, CDC1₃) δ 1.47-1.59 (m, 1H), 1.68-1.93 (m, 3H), 2.02-2.12 (m, 1H), 2.24-2.34 (m, 1H), 3.42 (s, 3H), 3.98-4.12 (m, 2H), 4.68-4.70 (m, 1H), 5.37 (s, 2H), 6.86-6.90 (m, 1H), 7.36-7.42 (m, 2H), 7.58-7.63 (m, 1H), 7.81-7.88 (m, 1H), 7.89 (s, 1H), 7.97-8.01 (m, 2H).

(6aR,9aS)-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one

The mixture of 2-(4-(6-fluoropyridin-2-yl)benzyl)-6-(((lR,2R)-2-hydroxycyclopentyl)amino)-5-methyl-2H-pyrazolo[3,4-d]pyrimidin-4(5H)-one (80 g), p-toluenesulfonylchloride (38.6 g), Et₃N (28.2 mL), N,N-dimethylaminopyridine (24.7 g) and THF (800 mL) is stirred at 50°C for 10 h. To the mixture is added 8M NaOH (11.5 mL) at room temperature and the mixture is stirred for 2 h. After the addition of 5% NaCl (400 mL) and AcOEt (800 mL), the organic layer is separated. The organic layer is washed with 5 NaCl (400 mL) twice. The organic layer is concentrated to 240 mL. After the addition of MeOH (800 mL), the mixture is concentrated to 240 mL. After the addition of MeOH (800 mL), the mixture is concentrated to 240 mL. After the addition of MeOH (160 mL), the mixture is stirred at room temperature for 1 h and at 0°C for 1 h. The crystals are isolated by filtration, washed with cold MeOH (160 mL) and dried to give (6aR,9aS)-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one (55.7 g). ^]H NMR (500 MHz, CDC1₃) δ 1.39-1.54 (m, 1H), 1.58-1.81 (m, 3H), 1.81-1.92 (m, 1H), 2.12-2.22 (m, 1H), 3.28 (s, 3H), 4.61-4.70 (m, 2H), 5.20 (s, 2H), 6.79-6.85 (m, 1H), 7.25-7.32 (m, 2H), 7.53-7.58 (m, 1H), 7.68 (s, 1H), 7.75-7.83 (m, 1H), 7.92-7.98 (m, 2H).

(6aR,9aS)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-

hexahydrocyclopenta[4,5]imi ]pyrimidin-4(2H)-one

The mixture of (6aR,9aS)-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one (50 g) and toluene (1000 mL) is concentrated to 750 mL under the nitrogen atmosphere. Toluene (250 mL) and NCS (24 g) is added. To the mixture is added LiHMDS (1M THF solution, 204 mL) at 0°C and the mixture is stirred for 0.5 h. To the mixture is added 20% NH₄C1 (50 mL) at 5°C. The mixture is concentrated to 250 mL. After the addition of EtOH (250 mL), the mixture is concentrated to 150 mL. After the addition of EtOH (250 mL), the mixture is concentrated to 200 mL. After the addition of EtOH (200 mL), the mixture is warmed to 50°C. Water (300 mL) is added and the mixture is stirred at 50°C for 0.5 h. After stirring at room temperature for 1 h, the crystals are isolated by filtration, washed with the mixture of EtOH and water (1:1, 150 mL) and dried to give (6aR,9aS)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one (51.1 g). ^]H NMR (500 MHz, CDC1₃) δ 1.46-1.61 (m, 1H), 1.67-1.90 (m, 3H), 1.92-2.00 (m, 1H), 2.19-2.27 (m, 1H), 3.37 (s, 3H), 4.66-4.77 (m, 2H), 5.34 (s, 2H), 6.87-6.93 (m, 1H), 7.35-7.41 (m, 2H), 7.59-7.65 (m, 1H), 7.82-7.91 (m, 1H), 7.97-8.05 (m, 2H).

EXAMPLE 1

Crystals of (6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base mono-ethanol solvate

The mixture of (6a/?,9a5′)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one (2.5 g), K₂C0₃ (1.53 g), Pd(OAc)₂ (12.5 mg), Xantphos (32 mg), aniline (0.76 mL), and xylene (12.5 mL) is stirred at 125°C for 7 h under nitrogen atmosphere. After addition of water (12.5 mL), the organic layer is separated. The organic layer is washed with water (12.5 mL) twice. The organic layer is extracted with the mixture of DMAc (6.25 mL) and 0.5N HCl (12.5 mL). The organic layer is extracted with the mixture of DMAc (3.2 mL) and 0.5N HCl (6.25 mL). After addition of DMAc (6.25 mL), xylene (12.5 mL) and 25 wt % aqueous NH₃ solution to the combined aqueous layer, the organic layer is separated. The aqueous layer is extracted with xylene (6.25 mL). The combined organic layer is washed with water (12.5 mL), 2.5 wt % aqueous 1 ,2-cyclohexanediamine solution (12.5 mL) twice and water (12.5 mL) successively. After treatment with active charcoal, the organic layer is concentrated. After addition of EtOH (12.5 mL), the mixture is concentrated. After addition of EtOH (12.5 mL), the mixture is concentrated. After addition of EtOH (12.5 mL), n-heptane (25 mL) is added at 70°C. The mixture is cooled to 5°C and stirred at same temperature. The crystals are isolated by filtration and dried to give (ea^^a^-S^a ^^^a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base mono-ethanol solvate (2.56 g) as crystals.

^]H NMR (500 MHz, DMSO-d₆) δ 0.98-1.13 (m, 3H), 1.34-1.52 (m, 1H), 1.54-1.83 (m, 4H), 2.03-2.17 (m, 1H), 3.11 (s, 3H), 3.39-3.54 (m, 2H), 4.29-4.43 (m, 1H), 4.51-4.60 (m, 1H), 4.60-4.70 (m, 1H), 5.15-5.35 (m, 2H), 6.71-6.88 (m, 3H), 7.05-7.29 (m, 5H), 7.81-7.93 (m, 1H), 7.94-8.11 (m, 3H), 8.67 (s, 1H).

EXAMPLE 4

Crystals of (6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one free

Crystals of (6a«,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base mono-n-propanol solvate (2.0 g) is dissolved with ethanol (10 mL) at 70°C. Isopropyl ether (20 mL) is added and the mixture is cooled to 45°C. Isopropyl ether (10 mL) is added and the mixture is stirred at 40°C. The mixture is cooled to 5°C and stirred at same temperature. The crystals are isolated by filtration and dried to give (ea/^^a^)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base non-solvate (1.7 g) as crystals.

[0082] ^]H NMR (500 MHz, DMSO-d₆) δ 1.32-1.51 (m, 1H), 1.53-1.83 (m, 4H), 1.97-2.20 (m, 1H), 3.11 (s, 3H), 4.49-4.60 (m, 1H), 4.60-4.69 (m, 1H), 5.13-5.37 (m, 2H), 6.70-6.90 (m, 3H), 7.04-7.31 (m, 5H), 7.82-7.93 (m, 1H), 7.93-8.12 (m, 3H), 8.67 (s, 1H).

EXAMPLE 5

Crystals of (6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base non-solvate

The mixture of (6a/?,9a5′)-3-chloro-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one (25 g), K₂C0₃ (15.4 g), Pd(OAc)₂ (125 mg), Xantphos (321 mg), aniline (7.6 mL), DMAc (6.25 mL) and xylene (125 mL) is stirred at 125°C for 6.5 h under nitrogen atmosphere. After addition of water (125 mL) and DMAc (50 mL), the organic layer is separated. The organic layer is washed with the mixture of DMAc (50 mL) and water (125 mL) twice. The organic layer is extracted with the mixture of DMAc (50 mL) and 0.5N HCl (125 mL). The organic layer is extracted with the mixture of DMAc (50 mL) and 0.5N HCl (62.5 mL). After addition of DMAc (50 mL), xylene (125 mL) and 25 wt % aqueous NH₃ solution (25 mL) to the combined aqueous layer, the organic layer is separated. The aqueous layer is extracted with xylene (62.5 mL). The combined organic layer is washed with the mixture of DMAc (50 mL) and water (125 mL), the mixture of DMAc (50 mL) and 2.5 wt % aqueous 1,2-cyclohexanediamine solution (125 mL) twice and the mixture of DMAc (50 mL) and water (125 mL) successively. After treatment with active charcoal (1.25 g), the organic layer is concentrated to 75 mL. After addition of EtOH (125 mL), the mixture is concentrated to 75 mL. After addition of EtOH (125 mL), the mixture is concentrated to 75 mL. After addition of EtOH (125 mL), n-heptane (250 mL) is added at 70°C. After addition of seed crystals of (6a/?,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one non-solvate, the mixture is cooled to room temperature and stirred at room temperature. The crystals are isolated by filtration and dried to give (ea^^a^-S^a ^^^a-hexahydro-S-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo-[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base non-solvate (23.8 g) as crystals.

EXAMPLE 8

(6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt

[0094] Crystals of (6a«,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base non-solvate (20 g) are dissolved in acetonitrile (60 mL) at 50°C. After addition of the active charcoal (1 g), the mixture is stirred at same temperature for 0.5 h. The active charcoal is removed by filtration and washed with acetonitrile (40 mL). The filtrate and the washing are combined and warmed to 50°C. A solution of 85 wt. % phosphoric acid (2.64 mL) in acetonitrile (100 mL) is added. After addition of water (20 mL), the mixture is stirred at 50°C for lh. The crystals are isolated by filtration, washed with acetonitrile (60 mL x 3) and dried to give (6a/?,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt (20.5 g).

EXAMPLE 9

(6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt

[0095] Crystals of (6a«,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base mono-ethanol solvate (4 g) are dissolved in acetonitrile (12 mL) at 50°C. After addition of active charcoal (0.2 g), the mixture is stirred at same temperature for 0.5 h. Active charcoal is removed by filtration and washed with acetonitrile (8 mL). The filtrate and the washing are combined and warmed to 50°C. A solution of 85 wt. % phosphoric acid (0.528 mL) in acetonitrile (20 mL) is added. After addition of water (4 mL), the mixture is stirred at 50°C for lh. The crystals are isolated by filtration, washed with acetonitrile (12 mL x 3) and dried to give (6a/?,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt (4.01 g).

EXAMPLE 10

(6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt

Crystals of (6a«,9a5′)-5,6a,7,8,9,9a-Hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base non-solvate (20 g) are dissolved in acetone (60 mL) at 32°C. After addition of active charcoal (1 g), the mixture is stirred at same temperature for 0.5 h. Active charcoal is removed by filtration and washed with acetone (40 mL). The filtrate and the washing are combined and warmed to 39°C. A solution of 85 wt. % phosphoric acid (2.64 mL) in acetone (100 mL) is added. After addition of water (20 mL), the mixture is stirred at 40°C for lh. The crystals are isolated by filtration, washed with acetone (60 mL x 3) and dried to give (6a/?,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt (22.86 g).

EXAMPLE 11

(6a/f,9a5)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-a]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt

Crystals of (6a«,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one free base mono-ethanol solvate (20 g) are dissolved in acetone (60 mL) at 38°C. After addition of active charcoal (1 g), the mixture is stirred at same temperature for 0.5 h. Active charcoal is removed by filtration and washed with acetone (40 mL). The filtrate and the washing are combined and warmed to 38°C. A solution of 85 wt. % phosphoric acid (2.64 mL) in acetone (100 mL) is added. After addition of water (20 mL), the mixture is stirred at 40°C for lh. The crystals are isolated by filtration, washed with acetone (60 mL x 3) and dried to give (6a/?,9a5′)-5,6a,7,8,9,9a-hexahydro-5-methyl-3-(phenylamino)-2-((4-(6-fluoropyridin-2-yl)phenyl)methyl)-cyclopent[4,5]imidazo[l,2-fl]pyrazolo[4,3-e]pyrimidin-4(2H)-one mono-phosphate salt (23.2 g).

PAPER

A diverse set of 3-aminopyrazolo[3,4-d]pyrimidinones was designed and synthesized. The structure–activity relationships of these polycyclic compounds as phosphodiesterase 1 (PDE1) inhibitors were studied along with their physicochemical and pharmacokinetic properties. Systematic optimizations of this novel scaffold culminated in the identification of a clinical candidate, (6aR,9aS)-2-(4-(6-fluoropyridin-2-yl)benzyl)-5-methyl-3-(phenylamino)-5,6a,7,8,9,9a-hexahydrocyclopenta[4,5]imidazo[1,2-a]pyrazolo[4,3-e]pyrimidin-4-(2H)-one phosphate (ITI-214), which exhibited picomolar inhibitory potency for PDE1, demonstrated excellent selectivity against all other PDE families and showed good efficacy in vivo. Currently, this investigational new drug is in Phase I clinical development and being considered for the treatment of several indications including cognitive deficits associated with schizophrenia and Alzheimer’s disease, movement disorders, attention deficit and hyperactivity disorders, and other central nervous system (CNS) and non-CNS disorders

Discovery of Potent and Selective Inhibitors of Phosphodiesterase 1 for the Treatment of Cognitive Impairment Associated with Neurodegenerative and Neuropsychiatric Diseases

Intra-Cellular Therapies and Takeda Announce Mutual Termination of Collaboration to Develop Phosphodiesterase (PDE1) Inhibitors for CNS Disorders

NEW YORK and OSAKA, Japan, Nov. 3, 2014 (GLOBE NEWSWIRE) — Intra-Cellular Therapies, Inc. (Nasdaq:ITCI) and Takeda Pharmaceutical Company Limited announced today that they have entered into an agreement to mutually terminate the February 2011 license agreement covering Intra-Cellular Therapies’ proprietary compound ITI-214 and related PDE1 inhibitors and to return the rights for these compounds to Intra-Cellular Therapies.

Under the terms of the agreement, Intra-Cellular Therapies has regained all worldwide development and commercialization rights for the compounds previously licensed to Takeda. Takeda will be responsible for transitioning the compounds back toIntra-Cellular Therapies and will not participate in future development or commercialization activities. After transition of the program, Intra-Cellular Therapies plans to continue the clinical development of PDE1 inhibitors for the treatment of central nervous system, cardiovascular and other disorders.

“We are grateful for Takeda’s substantial efforts in advancing this program into clinical development,” said Dr. Sharon Mates, Chairman and CEO of Intra-Cellular Therapies. “This provides us with the opportunity to unify our PDE1 platform and we look forward to continuing the development of ITI-214 and our other PDE1 inhibitors.”

Intra-Cellular Therapies will discuss the PDE1 program in its previously announced earnings call on Monday, November 3, 2014 at 8:30 a.m. Eastern Time. To participate in the conference call, please dial 844-835-6563 (U.S.) or 970-315-3916 (International) five to ten minutes prior to the start of the call. The participant passcode is 25568442.

About PDE1 Inhibitors

PDE1 inhibitors are unique, orally available, investigational drug candidates being developed for the treatment of cognitive impairments accompanying schizophrenia, Alzheimer’s disease and other neuropsychiatric disorders and neurological diseases and may also treat patients with Attention Deficit Hyperactivity Disorder and Parkinson’s disease. These compounds may also have the potential to improve motor dysfunction associated with these conditions and may also have the potential to treat patients with multiple sclerosis and other autoimmune diseases and pulmonary arterial hypertension. These compounds are very selective for the PDE1 subfamily relative to other PDE subfamilies. They have no known significant off target activities at other enzymes, receptors or ion channels.

About Intra-Cellular Therapies

Intra-Cellular Therapies, Inc. (the “Company”) is developing novel drugs for the treatment of neuropsychiatric and neurodegenerative disease and other disorders of the central nervous system (“CNS”). The Company is developing its lead drug candidate, ITI-007, for the treatment of schizophrenia, behavioral disturbances in dementia, bipolar disorder and other neuropsychiatric and neurological disorders. The Company is also utilizing its phosphodiesterase platform and other proprietary chemistry platforms to develop drugs for the treatment of CNS disorders.

About Takeda Pharmaceutical Company Limited

Located in Osaka, Japan, Takeda is a research-based global company with its main focus on pharmaceuticals. As the largest pharmaceutical company in Japan and one of the global leaders of the industry, Takeda is committed to strive towards better health for people worldwide through leading innovation in medicine. Additional information about Takeda is available through its corporate website, www.Takeda.com.

Source: Intra-Cellular Therapies, Inc.; Takeda Pharmaceutical Company Limited

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O=C(C1=C(NC2=CC=CC=C2)N(CC3=CC=C(C4=NC(F)=CC=C4)C=C3)N=C1N56)N(C)C5=N[C@@]7([H])[C@]6([H])CCC7.O=P(O)(O)O

OR

Fc1cccc(n1)c2ccc(cc2)Cn7nc5N3C(=N[C@@H]4CCC[C@H]34)N(C)C(=O)c5c7Nc6ccccc6

Galeterone

SYNTHESIS COMING………..

A SARM potentially for the treatment of prostate cancer.

Research Code, TOK-001; VN; 124; 124-1; 1241

TOK-001; Galeterone; 851983-85-2; VN/124; UNII-WA33E149SW; VN/124-1;

CAS No. 851983-85-2(Galeterone)

(3S,8R,9S,10R,13S,14S)-17-(benzimidazol-1-yl)-10,13-dimethyl-2,3,4,7,8,9,11,12,14,15-decahydro-1H-cyclopenta[a]phenanthren-3-ol

Galeterone (TOK-001 or VN/124-1) is a novel steroidal antiandrogen under development by Tokai Pharmaceuticals for the treatment of prostate cancer. It possesses a unique dual mechanism of action, acting as both an androgen receptor antagonist and an inhibitor of CYP17A1, an enzyme required for the biosynthesis of the androgens.^[1] It shows selectivity for 17,20-lyase over 17-hydroxylase.^[2]

As of 2016, galeterone is being compared to enzalutamide in a phase III clinical trial (ARMOR3-SV) for AR-V7-expressing metastatic castration-resistant prostate cancer.^[3][4]

Specific Androgen Receptor Modulator CYP17 Inhibitor TOK-001 is an orally bioavailable small-molecule androgen receptor modulator and CYP17 lyase inhibitor with potential antiandrogen activity. Galeterone exhibits three distinct mechanisms of action: 1) as an androgen receptor antagonist, 2) as a CYP17 lyase inhibitor and 3) by decreasing overall androgen receptor levels in prostate cancer tumors, all of which may result in a decrease in androgen-dependent growth signaling. Localized to the endoplasmic reticulum (ER), the cytochrome P450 enzyme CYP17 (P450C17 or CYP17A1) exhibits both 17alpha-hydroxylase and 17,20-lyase activities, and plays a key role in the steroidogenic pathway that produces progestins, mineralocorticoids, glucocorticoids, androgens, and estrogens.

References

• 1 Brawer MK (2008). “New treatments for castration-resistant prostate cancer: highlights from the 44th annual meeting of the american society of clinical oncology, may 30-june 3, 2008, chicago, IL”. Rev Urol 10 (4): 294–6. PMC 2615106. PMID 19145273.
• 2
• Yin L, Hu Q (2014). “CYP17 inhibitors–abiraterone, C17,20-lyase inhibitors and multi-targeting agents”. Nat Rev Urol 11 (1): 32–42. doi:10.1038/nrurol.2013.274. PMID 24276076.
• 3
• “A Study of Galeterone Compared to Enzalutamide In Men Expressing Androgen Receptor Splice Variant-7 mRNA (AR-V7) Metastatic CRPC – Full Text View – ClinicalTrials.gov”. clinicaltrials.gov. Retrieved 2016-02-27.
• 4

Silberstein, John L.; Taylor, Maritza N.; Antonarakis, Emmanuel S. (2016-04-01). “Novel Insights into Molecular Indicators of Response and Resistance to Modern Androgen-Axis Therapies in Prostate Cancer”. Current Urology Reports 17 (4): 29. doi:10.1007/s11934-016-0584-4. ISSN 1534-6285. PMID 26902623.

Galeterone

Systematic (IUPAC) name
17-(1H-benzimidazol-1-yl)androsta-5,16-dien-3β-ol
Clinical data
Routes of administration	Oral
Identifiers
CAS Number	851983-85-2
PubChem	CID 11188409
ChemSpider	9363493
KEGG	D10125
Chemical data
Formula	C26H32N2O
Molar mass	388.25

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C[C@]12CC[C@@H](CC1=CC[C@@H]3[C@@H]2CC[C@]4([C@H]3CC=C4N5C=NC6=CC=CC=C65)C)O

CC12CCC(CC1=CCC3C2CCC4(C3CC=C4N5C=NC6=CC=CC=C65)C)O

Apalutamide,, ARN 509

synthesis coming………….

ARN-509;  cas 956104-40-8; ARN 509; UNII-4T36H88UA7;

ARN-509; JNJ-56021927; JNJ-927\

Phase III Prostate cancer

4-(7-(6-CYANO-5-(TRIFLUOROMETHYL)PYRIDIN-3-YL)-8-OXO-6-THIOXO-5,7-DIAZASPIRO[3.4]OCTAN-5-YL)-2-FLUORO-N-METHYLBENZAMIDE;

4-(7-(6-cyano-5-(trifluoroMethyl)pyridin-3-yl)-8-oxo-6-thioxo-5,7-diazaspirooctan-5-yl)-2-fluoro-N-MethylbenzaMide;

• Originator University of California System
• Developer Janssen Research & Development, Aragon Pharmaceuticals, Memorial Sloan Kettering Cancer Center
• Class Antiandrogens; Antihormones; Antineoplastics; Aza compounds; Benzamides; Pyridines; Small molecules; Spiro compounds; Sulfhydryl compounds; Thiohydantoins
• Mechanism of Action Androgen receptor antagonists; Hormone inhibitors
• 03 Nov 2015 Janssen Research & Development plans a drug-interaction and pharmacokinetics phase I trial for Prostate cancer in Moldova (NCT02592317)

• 01 Nov 2015 Phase-III clinical trials in Prostate cancer (Adjunctive treatment) in United Kingdom, Sweden, Poland, Hungary, Australia, Australia, Spain, Canada, Brazil, USA (PO) (NCT02489318; EudraCT2015-000735-32)
• 15 Oct 2015 Aragon plans a phase I cardiac safety trial in patients with Prostate cancer in USA, Canada, the Netherlands and United Kingdom (NCT02578797)

Clinical Information of ARN-509

References on ARN-509

Apalutamide, also known as ARN-509 and JNJ-56021927 , is an androgen receptor antagonist with potential antineoplastic activity. ARN-509 binds to AR in target tissues thereby preventing androgen-induced receptor activation and facilitating the formation of inactive complexes that cannot be translocated to the nucleus. This prevents binding to and transcription of AR-responsive genes. This ultimately inhibits the expression of genes that regulate prostate cancer cell proliferation and may lead to an inhibition of cell growth in AR-expressing tumor cells.

Apalutamide (INN) (developmental code name ARN-509, also JNJ-56021927) is a non-steroidal antiandrogen that is under development for the treatment of prostate cancer.^[1] It is similar to enzalutamide both structurally and pharmacologically,^[2] acting as a selective competitive antagonist of the androgen receptor (AR), but shows some advantages, including greater potency and reduced central nervous system permeation.^[1][3][4] Apalutamide binds weakly to the GABA_A receptor similarly to enzalutamide, but due to its relatively lower central concentrations, may have a lower risk of seizures in comparison.^[1][3][5] The drug has been found to be effective and well-tolerated in clinical trials thus far,^[2][4] with the most common side effects reported including fatigue, nausea, abdominal pain, and diarrhea.^[6][3][5] Apalutamide is currently in phase III clinical trials for castration-resistant prostate cancer.^[7]

Recently, the acquired F876L mutation of the AR identified in advanced prostate cancer cells was found to confer resistance to both enzalutamide and apalutamide.^[8][9] A newer antiandrogen, ODM-201, is not affected by this mutation, nor has it been found to be affected by any other tested/well-known AR mutations.^[10]

Apalutamide may be effective in a subset of prostate cancer patients with acquired resistance to abiraterone acetate.^[2]

The chemical structure of ARN-509 is very similar structure to  that of Enzalutamide (MDV3100) with two minor modifications: (a) two methyl groups in the 5-member ring of MDV3100 is linked by a CH2 group in ARN-509; (b) the carbon atom in the benzene ring of MDV3100 is replaced by a nitrogen atom in ARN-509. ARN-509 is considered as a Me-Too drug of Enzalutamide (MDV3100). ARN-509 was claimed to be more active than Enzalutamide (MDV3100).

ARN-509 is a novel 2nd Generation anti-androgen that is targeted to treat castration resistant prostate cancers where 1st generation anti-androgens fail.  ARN-509 is unique in its action in that it inhibits both AR nuclear translocation and AR binding to androgen response elements in DNA. Importantly, and in contrast to the first-generation anti-androgen bicalutamide, it exhibits no agonist activity in prostate cancer cells that over-express AR. ARN-509 is easily synthesized, and its oral bioavailability and long half-life allow for once-daily oral dosing. In addition, its excellent preclinical safety profile makes it well suited as either a mono- or a combination therapy across the entire spectrum of prostate cancer disease states. (source: http://www.aragonpharm.com/programs/arn509.htm).

ARN-509 is  a competitive AR inhibitor, which is fully antagonistic to AR overexpression, a common and important feature of CRPC. ARN-509 was optimized for inhibition of AR transcriptional activity and prostate cancer cell proliferation, pharmacokinetics and in vivo efficacy. In contrast to bicalutamide, ARN-509 lacked significant agonist activity in preclinical models of CRPC. Moreover, ARN-509 lacked inducing activity for AR nuclear localization or DNA binding. In a clinically valid murine xenograft model of human CRPC, ARN-509 showed greater efficacy than MDV3100. Maximal therapeutic response in this model was achieved at 30 mg/kg/day of ARN-509 , whereas the same response required 100 mg/kg/day of MDV3100 and higher steady-state plasma concentrations. Thus, ARN-509 exhibits characteristics predicting a higher therapeutic index with a greater potential to reach maximally efficacious doses in man than current AR antagonists. Our findings offer preclinical proof of principle for ARN-509 as a promising therapeutic in both castration-sensitive and castration-resistant forms of prostate cancer. (source: Cancer Res. 2012 Jan 20. [Epub ahead of print] )
(source: Cancer Res. 2012 Jan 20. [Epub ahead of print] )

PATENT

US20100190991

Prostate cancer is one of the most common forms of cancer found in Western men and the second leading cause of cancer death in Western men. When prostate cancer is confined locally, the disease can usually be treated by surgery and/or radiation. Advanced disease is frequently treated with anti-androgen therapy, also known as androgen deprivation therapy. Administration of anti-androgens blocks androgen receptor (AR) function by competing for androgen binding; and therefore, anti-androgen therapy reduces AR activity. Frequently, such therapy fails after a time, and the cancer becomes hormone refractory, that is, the prostate cancer no longer responds to hormone therapy and the cancer does not require androgens to progress.

Overexpression of AR has been identified as a cause of hormone refractory prostate cancer (Nat. Med., 10:33-39, 2004; incorporated herein by reference). Overexpression of AR is sufficient to cause progression from hormone sensitive to hormone refractory prostate cancer, suggesting that better AR antagonists than the current drugs may be able to slow the progression of prostate cancer. It has been demonstrated that overexpression of AR converts anti-androgens from antagonists to agonists in hormone refractory prostate cancer. This work explains why anti-androgen therapy fails to prevent the progression of prostate cancer.

The identification of compounds that have a high potency to anatgonize AR activity would overcome the hormone refractory prostate cancer and slowdown the progression of hormone sensitive prostate cancer. Such compounds have been identified by Sayers et al. (WO 2007/126765, published Nov. 8, 2007; which is incorporated herein by reference). One compound is known as A52, a biarylthiohydantoin, and has the chemical structure

• Another compound A51 has the chemical structure:
• Both of these compounds share the same western and central portions. Given the need for larger quantities of pure A51 and A52 for pre-clinical and clinical studies, there remains a need for a more efficient synthesis of the compound from commercially available starting materials.

Convergent Coupling to Yield A52

The final coupling step between intermediates A and B is achieved by microwave irradiation and cyclization to the biarylthiohydantoin A52 (Scheme 6). Although 3 equivalents of A are required for the highest yields in this transformation, the un-reacted amine A can be recovered.

Experimental Section 2-cyano-5-nitro-3-trifluoromethylpyridine

• Zinc cyanide (25 mg, 0.216 mmol, 1.2 eq) is added to the chloride (43 mg, 0.180 mmol) solubilized in DMF (1 ml). The solution is degassed for 10 minutes. Then the ligand dppf (20 mg, 0.036 mmol, 0.2 eq) is added. The solution is degassed again for 5 min. The catalyst Pd₂(dba)₃ (25 mg, 0.027 mmol, 0.15 eq) is added, the solution is degassed for 5 more minutes. The reaction mixture is then heated at 130° C. for 20 min in a microwave. After filtration, the solvent is evaporated and the crude residue is purified by flash chromatography on silica gel (hexane/EtOAc) to afford 16 mg (40%) of the desired product
• ¹H NMR (400 MHz, CDCl₃) δ 8.60 (d, J=2.5, 1H); 9.08 (d, J=2.5, 1H),

5-amino-2-cyano-3-trifluoromethylpyridine

• 2-cyano-5-nitro-3-trifluoromethylpyridine (7 mg, 0.032 mmol) is dissolved in 1:1 EtOAc/AcOH (1 mL) and heated to 65° C. Iron powder (9 mg, 0.161 μmol, 5 eq, 325 mesh) is added and the mixture stirred for 2 hours. The mixture is filtered through celite, and the filtrate is concentrated under vacuo. The crude residue is purified by flash chromatography on silica gel (hexane/EtOAc) to afford 4 mg (67%) of the desired product
• ¹H NMR (400 MHz CDCl3) δ 7.20 (d, J=2.4 Hz, 1H), 8.22 (d, J=2.4 Hz, 1H).

5-iodo-3-trifluoromethyl-2-pyridinol

• 3-trifluoromethyl-2-pyridinol (25 g, 153.3 mmol) is dissolved in anhydrous CH₃CN (150 mL) and DMF (150 mL). N-iodosuccinimide (34.5 g, 153 mmol) is then added. The reaction mixture is stirred at 80° C. for 2 hours and cooled to room temperature. Aqueous 1 M NaHCO₃ (150 mL) is then added to the cooled mixture. After stirring for 5 min, the solvents are evaporated to dryness. Water is added and the aqueous phase is extracted (×2) with dichloromethane. The organic phase is then evaporated and the desired product is recrystallized in water to afford 36.2 g (81%) of a white powder.
• ¹H NMR (500 MHz, CDCl₃) δ 7.85 (d, J=2.3, 1H); 7.98 (d, J=2.3, 1H), 13.41 (br s, 1H); ¹³C NMR (250 MHz CDCl₃) δ 63.0, 121.4 (q, J_C-F=272.3 Hz), 122.2 (q, J_C-F=31.6 Hz), 144.4, 148.1 q, (J_C-F=5.0 Hz), 160.1.

2-chloro-5-iodo-3-trifluoromethylpyridine

• To an ice-cold mixture of POCl₃ (1.60 mL) and DMF (1 mL) in a microwave vial, 5-iodo-3-trifluoromethyl-2-pyridinol (1 g, 3.47 mmol) is added. The vial is sealed and heated 20 min at 110° C. The reaction mixture cooled at room temperature is poured into ice cold water. The product precipitates. The precipitate is filtered, washed with cold water and dried to afford 661 mg (62%) of a light brown powder.
• ¹H NMR (500 MHz CDCl₃) δ 8.32 (d, J=2.0 Hz, 1H), 8.81 (d, J=2.0 Hz, 1H). ¹³C NMR (250 MHz CDCl₃) δ 89.4, 121.2 (q, J_C-F=273.3 Hz), 126.8 (q, J_C-F=33.6 Hz), 144.34, 148.5, 158.7.

2-choro-3-trifluoromethyl-N-paramethoxybenzylpyridin-5-amine

• 2-choro-5-iodo-3-trifluoromethylpyridine is dried under vacuum. To a slurry of chloroiodpyridine (10 g, 32.6 mmol) in toluene (anhydrous) (98 mL) is added sequentially. Pd(OAc)₂ (220 mg, 0.98 mmol, 0.03 eq), rac-BINAP (609 mg, 0.98 mmol, 0.03 eq) solid Cs₂CO₃ (53 g, 163 mmol, 5 eq), paramethoxybenzylamine (4.05 mL, 30.9 mmol, 0.95 eq) and triethylamine (0.41 mL, 2.93 mmol, 0.09 eq). The resulting slurry is degassed (×2) by vacuum/Argon backfills. The mixture is heated to reflux overnight. The mixture is then cooled to room temperature and H₂O is added. The layers are separated and the toluene layer is concentrated under vacuo. The residue is purified by flash chromatography on silica gel (Hexane/EtOac; 95:5 to 30/70) to afford 4 g of white solid desired compound (40%).
• ¹H NMR (500 MHz CDCl₃) δ 3.81 (s, 3H), 4.29 (d, J=5.1 Hz, 2H), 4.32 (br s, 1H), 6.90 (d, J=8.1 Hz, 2H), 7.19 (d, J=2.9 Hz, 1H), 7.26 (d, J=8.1 Hz, 2H), 7.92 (d, J=2.9 Hz, 1H). ¹³C NMR (250 MHz CDCl₃) δ 47.3, 55.4, 114.3, 119.3 (q, J_C-F=5.1 Hz), 122.3 (q, J_C-F=272.9 Hz), 124.80 (q, J_C-F=32.7 Hz), 128.8, 129.1, 135.1, 136.6, 142.9, 159.3.

Alternative Synthesis of Intermediate K:

• A suspension of vacuum dried 2-choro-5-iodo-3-trifluoromethylpyridine (50 g, 163 mmol) in anhydrous toluene (1,500 mL) was treated sequentially with Pd₂(dba)₃ (2.98 g, 3.25 mmol, 0.02 eq), Xantphos (5.65 g, 9.76 mmol, 0.06 eq), solid t-BuONa (23.4 g, 243 mmol, 1.5 eq), and paramethoxybenzylamine (23.2 mL, 179 mmol, 1.1 eq). The resulting slurry is degassed by vacuum/argon backfills for 10 min. The mixture is then quickly brought to reflux by a pre-heated oil bath. After 1.5 hours at this temperature, the mixture was cooled to the ambiant, and the solids were removed by filtration over a packed bed of celite and washed with toluene. The filtrate was then diluted with EtOAc (200 mL), then washed with H₂O. The organic layer was concentrated under reduced pressure gave an oily solid. Crystallization from DCM/Hexane gave (36.6 g, 71%) of B as a light yellow solid.
• Alternatively, smaller scales (5 to 10 gr of A) were purified by column silica gel chromatography using the gradient system Hexane-EtOAc 19-1 to 3-7 (v-v). This gave yields in excess of 85% of B as a white solid.

2-cyano-3-trifluoromethyl-N-paramethoxybenzylpyridin-5-amine

• Zinc cyanide (0.45 g, 3.80 mmol, 1.2 eq) is added to the chloride (1 g, 3.16 mmol) solubilized in DMF (20 ml). The solution is degassed for 10 minutes. Then the ligand dppf (0.35 g, 0.63 mmol, 0.2 eq) is added. The solution is degassed again for 5 min. The catalyst Pd₂(dba)₃ (0.29 g, 0.32 mmol, 0.1 eq) is added, the solution is degassed for 5 more minutes. The reaction mixture is then heated at 150° C. for 10 min. After filtration, the solvent is evaporated and the crude residue is purified by flash chromatography on silica gel (hexane/EtOAc) to afford 900 mg (93%) of a dark yellow oil.
• ¹H NMR (500 MHz CDCl3) δ 3.82 (s, 3H), 4.37 (d, J=5.3 Hz, 2H), 4.93 (br s, 1H), 6.92 (d, J=9.5, 2H), 7.08 (d, J=2.7 Hz, 1H), 7.25 (d, J=9.5, 2H), 8.17 (d, J=2.7 Hz, 1H). ¹³C NMR (250 MHz CDCl3) δ 46.7, 55.4, 113.9, 114.5, 115.9, 116.1, 122.0 (q, J_C-F=274.5 Hz), 128.0, 128.9, 131.4 (q, J_C-F=33.1 Hz), 138.68, 145.9, 159.5.

5-amino-2-cyano-3-trifluoromethylpyridine H

• TFA (1 mL) is added dropwise to a solution of pyridine L (83 mg, 0.27 mmol) in dry DCM (0.5 mL) under argon. The solution is stirred overnight at room temperature. After completion of the reaction, the solvent is evaporated and the residue is purified by flash chromatography on silica gel (Hexane/EtOac) to afford the desired product quantitatively.
• ¹H NMR (500 MHz CDCl3) δ 7.20 (d, J=2.4 Hz, 1H), 8.22 (d, J=2.4 Hz, 1H).

Scale Up and Purification of H

• For the larger scales, an improved process calls for dissolving pyridine L (53 g, 0.172 mol) in TFA/DCM (170 mL, 4:1) at room temperature. Upon reaction completion (approximately 2 hours at room temperature), the volatiles were removed under reduced pressure. The residue is then diluted with EtOAc (800 mL), and washed with saturated aqueous NaHCO₃. Vacuum concentration and precipitation from DCM-Hexane (1-2, v-v) gave a relatively clean product. Further washing with DCM gave pure intermediate H as a white solid (27.43 g, 85%).

Methyl 2,4-difluorobenzylamide

• Methylamine 2M in THF (12.4 mL, 1.1 eq) is added to neat 2,4-difluorobenzoyl chloride (4 g, 22.6 mmol). The reaction mixture is stirred overnight at room temperature. The solvent is evaporated, ethyl acetate is added to solubilize the residue. The organic is washed with aqueous NaHCO₃, dried with Na₂SO₄, filtered and evaporated to afford the quantitatively the desired compound as a white powder.
• ¹H NMR (500 MHz CDCl3) δ 3.00 (d, J=4.8 Hz, 3H), 6.84 (m, J=2.3; 10.3 Hz, 1H), 6.97 (m, J=2.3; 8.2 Hz, 1H), 8.08 (td, J=6.8; 8.9 Hz, 1H)
• ¹³C NMR (100 MHz CDCl3) δ 27.0, 104.3 (d, J=26.0 Hz), 104.6 (d, J=25.9 Hz), 112.4 (dd, J=21.2; 3.1 Hz), 118.1 (dd, J=12.4; 3.8 Hz), 133.7 (dd, J=10.1; 3.9 Hz), 162.9 (dd, J=381.1; 12.3 Hz), 163.5.

Methyl 2-fluoro-4-paramethoxybenzylamine-benzylamide

• Paramethoxybenzylamine (0.069 mL, 0.548 mmol, 2 eq) is added to methyl 2,4-difluorobenzylamide (47 mg, 0.274 mmol) dissolved in dimethylsulfoxide (0.5 mL). The reaction mixture is heated at 190° C. for 20 min in a microwave. After completion the solvent is evaporated and the residue is purified by flash chromatography on silica gel (hexane/ethyl acetate) to give 18 mg (20%) of the desired product.
• ¹H NMR (500 MHz CDCl3) δ 2.98 (d, J=4.5 Hz, 3H), 3.81 (s, 3H), 4.26 (d, J=5.3 Hz, 2H), 4.47 (br s, 1H), 6.23 (dd, J=2.2; 15.1 Hz, 1H), 6.45 (dd, J=2.2; 8.7 Hz, 1H), 6.58 (br s, 1H), 6.89 (d, J=8.7 Hz, 2H), 7.25 (d, J=8.7 Hz, 2H), 7.91 (t, J=9.0 Hz, 1H). ¹³C NMR (500 MHz CDCl3) δ 26.6, 47.3, 55.3, 98.2 (d, J=29.7 Hz), 109.25, 114.4, 128.6, 129.9, 133.1 (d, J=4.5 Hz), 152.3 (d, J=12.5 Hz), 159.1, 161.5, 163.9 (d, J=244 Hz), 164.5.

Methyl 4-amino-2-fluoro-benzylamide

• TFA (1 mL) is added dropwise to a solution of methylamide (60 mg, 0.21 mmol) in dry DCM (0.5 mL) under argon. The solution is stirred overnight at room temperature. After completion of the reaction, the solvent is evaporated and the residue is purified by flash chromatography on silica gel (Hexane/EtOac) to afford the desired product quantitatively.
• ¹H NMR (500 MHz CDCl3) δ 2.98 (d, J=4.8 Hz, 3H), 4.15 (br s, 2H), 6.32 (d, J=14.3 Hz, 1H), 6.48 (d, J=8.2 Hz, 1H), 6.61 (br s, 1H), 7.90 (dd, J=8.6 Hz, 1H), ¹³C NMR (500 MHz CDCl3) δ 26.63, 100.8 (d, J=28.8 Hz), 110.3 (d, J=244.6 Hz), 110.9, 133.3 (d, J=4.3 Hz), 151.4 (d, J=12.5 Hz), 162.2 (d, J=244.6 Hz), 164.3 (d, J=3.5 Hz).

Synthesis of N-methyl-4-[7-(6-cyano-5-trifluoromethylpyridin-2-yl)-8-oxo-6-thioxo-5,7-diazaspiro[3.4]octan-5-yl]-2-fluorobenzamide (A52) One Pot Small Scale (2.8 gr) Thiohydantoin Formation in DMF

• Thiophosgene (1.2 mL, 1.16 eq, 15.6 mmol) is added dropwise to a solution of 5-amino-2-cyano-3-trifluoromethylpyridine (2.8 g, 1.1 eq, 15.0 mmol) and N-methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide (3.35 g, 13.5 mmol) in dry DMF (25 mL) under Argon. The solution is stirred overnight at 60° C. To this mixture were added MeOH (60 mL) and aq. 2M HCl (30 mL), then the mixture was reflux for 2 h. After cooling to rt, the mixture was poured into ice water (100 mL) and extracted with EtOAc (3×60 mL). The organic layer was dried over Mg₂SO₄, concentrated and chromatographed on silica gel using 5% acetone in DCM to yield the desired product (2.65 g, 41%).

Alternative Synthesis of A52

• Thiophosgene (1.23 mL, 16.0 mmol) is added dropwise to a solution of 5-amino-2-cyano-3-trifluoromethylpyridine (3.0 g, 16.0 mmol) and N-methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide (3.96 g, 16.0 mmol) in dry DMA (35 mL) under Argon. The solution is stirred overnight at 60° C. To this mixture were added MeOH (60 mL) and aq. 2M HCl (30 mL), then it was brought to reflux temperature for 2 h. After cooling down to the ambiant, the mixture was poured into ice water (100 mL) and extracted with EtOAc (3×60 mL). The organic layer was dried over Mg₂SO₄, filtered over celite, and concentrated under reduced pressure. Silica gel chromatography using DCM/-acetone 19-1 (v-v) yielded the desired product (5.78 g, 76%).

Scale Up

• Thiophosgene (5.48 mL, 1.05 eq, 70.9 mmol) is added dropwise to a solution of 5-amino-2-cyano-3-trifluoromethylpyridine (13.27 g, 1.05 eq, 70.9 mmol) and N-methyl-4-(1-cyanocyclobutylamino)-2-fluorobenzamide (16.7 g, 67.5 mmol) in dry DMA (110 mL) under Argon at 0° C. After 10 min, the solution was heated up to 60° C. and allowed to stir at that temperature for an overnight period. This was then diluted with MeOH (200 mL) and treated with aq. 2M HCl (140 mL), then the mixture was refluxed for 2 h. After cooling down to RT, the mixture was poured into ice water (500 mL), and filtered over buchner. The solid was recrystallized from DCM/EtOH to get desired product (20.6 g, 64%).

References

• 1 Clegg NJ, Wongvipat J, Joseph JD, Tran C, Ouk S, Dilhas A, Chen Y, Grillot K, Bischoff ED, Cai L, Aparicio A, Dorow S, Arora V, Shao G, Qian J, Zhao H, Yang G, Cao C, Sensintaffar J, Wasielewska T, Herbert MR, Bonnefous C, Darimont B, Scher HI, Smith-Jones P, Klang M, Smith ND, De Stanchina E, Wu N, Ouerfelli O, Rix PJ, Heyman RA, Jung ME, Sawyers CL, Hager JH (2012). “ARN-509: a novel antiandrogen for prostate cancer treatment”. Cancer Res. 72 (6): 1494–503. doi:10.1158/0008-5472.CAN-11-3948. PMC 3306502. PMID 22266222.
• 2
• Patel JC, Maughan BL, Agarwal AM, Batten JA, Zhang TY, Agarwal N (2013). “Emerging molecularly targeted therapies in castration refractory prostate cancer”. Prostate Cancer 2013: 981684. doi:10.1155/2013/981684. PMC 3684034. PMID 23819055.
• 3
• Schweizer MT, Antonarakis ES (2012). “Abiraterone and other novel androgen-directed strategies for the treatment of prostate cancer: a new era of hormonal therapies is born”. Ther Adv Urol 4 (4): 167–78. doi:10.1177/1756287212452196. PMC 3398601. PMID 22852027.
• 4
• Rathkopf D, Scher HI (2013). “Androgen receptor antagonists in castration-resistant prostate cancer”. Cancer J 19 (1): 43–9. doi:10.1097/PPO.0b013e318282635a. PMC 3788593. PMID 23337756.
• 5
• Pinto Á (2014). “Beyond abiraterone: new hormonal therapies for metastatic castration-resistant prostate cancer”. Cancer Biol. Ther. 15 (2): 149–55. doi:10.4161/cbt.26724. PMC 3928129. PMID 24100689.
• 6
• Leibowitz-Amit R, Joshua AM (2012). “Targeting the androgen receptor in the management of castration-resistant prostate cancer: rationale, progress, and future directions”. Curr Oncol 19 (Suppl 3): S22–31. doi:10.3747/co.19.1281. PMC 3553559. PMID 23355790.
• 7
• Agarwal N, Di Lorenzo G, Sonpavde G, Bellmunt J (2014). “New agents for prostate cancer”. Ann. Oncol. 25 (9): 1700–9. doi:10.1093/annonc/mdu038. PMID 24658665.
• 8
• Joseph JD, Lu N, Qian J, Sensintaffar J, Shao G, Brigham D, Moon M, Maneval EC, Chen I, Darimont B, Hager JH (2013). “A clinically relevant androgen receptor mutation confers resistance to second-generation antiandrogens enzalutamide and ARN-509”. Cancer Discov 3 (9): 1020–9. doi:10.1158/2159-8290.CD-13-0226. PMID 23779130.
• 9
• Nelson WG, Yegnasubramanian S (2013). “Resistance emerges to second-generation antiandrogens in prostate cancer”. Cancer Discov 3 (9): 971–4. doi:10.1158/2159-8290.CD-13-0405. PMC 3800038. PMID 24019330.
• 10

Moilanen AM, Riikonen R, Oksala R, Ravanti L, Aho E, Wohlfahrt G, Nykänen PS, Törmäkangas OP, Palvimo JJ, Kallio PJ (2015). “Discovery of ODM-201, a new-generation androgen receptor inhibitor targeting resistance mechanisms to androgen signaling-directed prostate cancer therapies”. Sci Rep 5: 12007. doi:10.1038/srep12007. PMC 4490394. PMID 26137992

11Clegg NJ, Wongvipat J, Tran C, Ouk S, Dilhas A, Joseph J, Chen Y, Grillot K, Bischoff ED, Cai L, Aparicio A, Dorow S, Arora V, Shao G, Qian J, Zhao H, Yang G, Cao C, Sensintaffar J, Wasielewska T, Herbert MR, Bonnefous C, Darimont B, Scher  HI, Smith-Jones PM, Klang M, Smith ND, de Stanchina E, Wu N, Ouerfelli O, Rix P, Heyman R, Jung ME, Sawyers CL, Hager JH. ARN-509: a novel anti-androgen for prostate cancer treatment. Cancer Res. 2012 Mar 15;72(6):1494-1503. Epub 2012 Jan 20.PubMed  PMID: 22266222.

12]. Clegg NJ, Wongvipat J, Joseph JD et al. ARN-509: a novel antiandrogen for prostate cancer treatment. Cancer Res. 2012 Mar 15;72(6):1494-503.

[13]. Courtney KD, Taplin ME. The evolving paradigm of second-line hormonal therapy options for castration-resistant prostate cancer. Curr Opin Oncol. 2012 May;24(3):272-7.

[14]. Schweizer MT, Antonarakis ES. Abiraterone and other novel androgen-directed strategies for the treatment of prostate cancer: a new era of hormonal therapies is born. Ther Adv Urol. 2012 Aug;4(4):167-78.

[15]. Safety, Pharmacokinetic and Proof-of-Concept Study of ARN-509 in Castration-Resistant Prostate Cancer (CRPC)

Apalutamide

Systematic (IUPAC) name
4-[7-[6-Cyano-5-(trifluoromethyl)pyridin-3-yl]-8-oxo-6-sulfanylidene-5,7-diazaspiro[3.4]octan-5-yl]-2-fluoro-N-methylbenzamide
Clinical data
Pregnancy category	X (Contraindicated)
Routes of administration	Oral
Identifiers
CAS Number	956104-40-8
ATC code	None
PubChem	CID 24872560
ChemSpider	28424131
Chemical data
Formula	C21H15F4N5O2S
Molar mass	477.434713 g/mol

////////

CNC(=O)C1=C(C=C(C=C1)N2C(=S)N(C(=O)C23CCC3)C4=CN=C(C(=C4)C(F)(F)F)C#N)F

CNC(=O)C1=C(C=C(C=C1)N2C(=S)N(C(=O)C23CCC3)C4=CN=C(C(=C4)C(F)(F)F)C#N)F

Liarozole fumarate is prepared as shown in Scheme 20970301a. Anisol is reacted with 3-chlorobenzoyl chloride (I) under Friedel-Craft conditions to give (3-chlorophenyl)(4-methoxyphenyl)methanone (II). Nitration of (II) is carried out in dichloromethane at 10 C to yield (III). The methoxy group in (III) is replaced by the amino group by means of NH3 in 2-propanol at 100 C under pressure, giving (IV). By reduction of the keto function of (IV) with sodium borohydride in 2-propanol, the corresponding alcohol (V) is obtained, which upon treatment with 1,1′-carbonyldiimidazole in refluxing dichloromethane yields the imidazolyl compound (VI). Hydrogenation of the nitro group in (VI), followed by cyclization of (VII) in a refluxing mixture of formic acid and 4N hydrochloric acid, gives the benzimidazole derivative (VIII). Finally, the treatment of (VIII) with fumaric acid in ethanol yields liarozole fumarate (IX).

http://www.google.com/patents/WO1995022540A1?cl=en

Liarozole is a racemic mixture, i.e. a mixture of its optical isomers, and is specifically mentioned as compound 28 in EP-0,371,559. Said patent application mentions the use of compounds like liarozole in the treatment of epithelial disorders. EP-0,260,744 describes the use of compounds like liarozole for inhibiting or lowering androgen formation. Whereas EP-0,371,559 and EP-0,260,744 recognize that compounds like liarozole have stereochemically isomeric forms, no example of an enantiomerically pure form is given of liarozole.

Chemically liarozole is (±)-5-[3-chlorophenyl]-lH-imidazol-l-ylmethyl]-lH-benz- imidazole, and is represented by formula (I). As can be seen from the chemical structure, liarozole has one stereogenic center (indicated with an asterisk in formula (I)).

The subject of this invention is the enantiomerically pure dextrorotatory isomer or (+)-isomer of liarozole. Said isomer will hereinafter be referred to as (+)-liarozole. Many organic compounds exist in optically active forms, i.e. they have the ability to rotate the plane of plane-polarized light. In describing an optically active compound, the prefixes D and L or R and S are used to denote the absolute configuration of the molecule about its chiral center(s). The prefixes (+) and (-) or d and 1 are employed to designate the sign of rotation of plane-polarized light by the compound, with (-) or 1 meaning that the compound is iaevorotatory and with (+) or d meaning that the compound is dextrorotatory. For a given chemical structure the optically active isomers having an opposite sign of optical rotation are called enantiomers. Said enantiomers are identical except that they are mirror images of one another. A 1: 1 -mixture of such enantiomers is called a racemic mixture.

General preparation of structures including liarozole have been extensively described in EP-0,371,559 and EP-0,260,744.

Enantiomerically pure (+)-liarozole may be prepared by reacting an enantiomerically pure intermediate diamine of formula (B)-(II) with formic acid or a functional derivative thereof.

Said functional derivative of formic acid is meant to comprise the halide, anhydride, amide and ester, including the ortho and imino ester form thereof. Also methanimidamide or an acid addition salt thereof can be used as cyclizing agent.

The general reaction conditions, work-up procedures and conventional isolation techniques for carrying out the above and following reactions are described in the prior art. When more specific conditions are required they are mentioned hereinunder. The enantiomerically pure intermediate diamine of formula (B)-(II) may be prepared by reducing an intermediate of formula (B)-(iπ) by a standard nitro-to-amine reduction reaction.

The desired enantiomer of the intermediate of formula (B)-(]H) can be prepared by fractional crystallization of a racemic mixture of the intermediate of formula (HI) with an enantiomerically pure chiral acid. Preferred chiral acid for the above fractional crystallization is 7,7-dimethyl-2-oxobicyclo[2.2.1]heptane-l-methanesulfonic acid (i.e. 10-camphorsulfonic acid).

Appropriate solvents for carrying out said fractional crystallization are water, ketones, e.g. 2-propane, 2-butanone; alcohols, e.g. methanol, ethanol, 2-propanol. Mixtures of ketones and water are very suitable for the above fractional crystallization. Preferably a mixture of 2-propanone and water is used.

The ratio of water/2-propanone by volume may vary from 1/10 to 1/2. Preferred range of said ratio is 1/5 to 1/3.

The fractional crystallizations are suitably carried out below room temperature, preferably below 5°C.

It was also found that the subsequent reaction step can be carried out without any appreciable racemization.

Alternatively the (+)-isomer of the compound of formula (I) may be prepared by cyclizing an intermediate of formula (B)-(IV) following procedures as described above for the cyclization of intermediates of formula (B)-(II) and desulfurating the thus obtained intermediate of formula (B)-(V). In formulas (B)-(TV) and (B)-(V) R represents Ci^alkyl, wherein Ci-^alkyl means a straight or branch chained saturated hydrocarbon radicals having 1 to 6 carbon atoms such as, for example, methyl, ethyl, propyl, butyl, pentyl, hexyl. Preferably R is methyl.

The intermediates of formula (B)-(IV) may be prepared by reacting an intermediate of formula (B)-(VI) with a reagent of formula (VII), alkylating the thus formed thiourea derivative of formula (B)-(VIII) subsequently cyclizing the intermediate of formula

(B)-(D ), and reducing the nitro group of the intermediate (B)-(X). In the formulas

(Vπ), (B)-(Vm), (B)-(IX) and (B)-(X) R represents Ci^alkyl as defined hereinabove.

S OR

(B)-(IV)

Experimental part

A. Preparation of the intermediates

Example 1 a) A heterogeneous mixture of (±)-4-[(3-chlorophenyl)-lH-imidazol-l-ylmethyl]-2- nitrobenzenamine (the preparation of which is described in EP-371,559) (500 g) in

2-propanone (2000 ml) and water (100 ml) was stirred at 22°C. (-)-(lR)-7,7-dimethyl- 2-oxo-bicyclo[2.2.1]heptane-l-methanesulfonic acid (353.2 g) was added and the mixture became homogeneous after 10 minutes. The mixture was first stirred for 18 hours at 20°C and then for 3 hours at 0-5°C. The precipitate was filtered off, washed with 2-propanone/water 95/5 (150 ml) and dried, yielding 308.9 g (36.2%) of product A sample (306.7 g) was partitioned between dichloromethane (500 ml) and water (750 ml). Ammonium hydroxide (100 ml) was added. This mixture was stirred for 15 minutes. The aqueous layer was separated and extracted twice with dichloromethane (250 ml each time). The separated organic layer was washed with water (250 ml), dried, filtered and the solvent was evaporated, yielding 179.7 g of (-)-(B)-4-[(3-chlorophenyl)-

20 lH-imidazol-l-ylmethyl]-2-nitrobenzenamine; mp. 89.8°C; [α]_D = -19.80° (c = 0.5% in methanol) (interm. 1). b) A mixture of intermediate (1)(179.7 g) in methanol (656 ml) and a solution of ammonia in methanol (32.7 ml) was hydrogenated at 20-25 °C with platinum on activated carbon (13.1 g) as a catalyst in the presence of thiophene (0.27 g). After uptake of hydrogen (3 eq.) the catalyst was filtered off and washed with 2-propanol (30 ml). A solution of hydrochloric acid in 2-propanol (522 ml) was added to the filtrate at <30°C. The mixture was stirred for 3 hours at 20 °C, then for 3 hours at 0-5 °C. The resulting precipitate was slowly filtered off, washed with methanol (100 ml) and dried

(50 °C), yielding 185.60 g (83.2%) (+)-(B)-4-[(3-chlorophenyl)-lH-imidazol-l-yl-

20 methyl]- 1,2-benzenediamine trihydrochloride; mp. 172.5°C; [α^ = +23.73° (c = 1% in methanol) (interm. 2).

Example 2 a) A mixture of (4-amino-3-nitrophenyl) (3-chlorophenyl)methanone (50 g), formamide (375 ml) and formic acid (63 ml) was stiιτed and refluxed for 17 hours. After cooling, the mixture was poured on ice. The precipitate was filtered off and dried, yielding 55 g (99.4%) of (±)-N-[(4-amino-3-nitrophenyl) (3-chlorophenyl)methyl]formamide (interm. 3). b) A mixture of intermediate (3) (50.7 g), hydrochloric acid 6N (350 ml) and 2-propanol (70 ml) was stirred and refluxed for 17 hours. The yellow precipitate was filtered off and dried in vacuo, yielding 51 g (97.8%) of (±)-4-amino-α-(3-chloro- phenyl)-3-nitrobenzenemethanamine monohydrochloride; mp. 263°C (interm.4). c) To a solution of intermediate (4) (43 g) in tetrahydrofuran (400 ml) at room temperature was added succesively N,N-diethylethanamine (13.8 g) and (R)-(-)-α- hydroxybenzeneacetic acid (20.8 g). Then a solution of 1-hydroxybenzotriazole monohydrate (22.2 g) in tetrahydrofuran (200 ml) was added. After complete addition a solution of N,N’-dicyclohexylcarbodiimide (33.9 g) in dichloromethane (300 ml) was introduced to the mixture. After stirring for 2 hours at room temperature N,N’- dicyclohexylurea was filtered off. The filtrate was washed with a solution of potassium carbonate (10%) and the organic layer was dried to give a mixture of diastereomers (60g) (fraction 1). The same experiment with intermediate (4) (16 g) as starting material resulted in a yield of 26 g of a mixture of diastereomers (fraction 2). Fraction 1 and 2 were combined and purified by HPLC (eluent : CH2θ₂/ethyl acetate 90:10), yielding 30g (32.3%) of (±)-(R,B)-N-[(4-amino-3-nitrophenyl)(3-chlorophenyl)methyl]-α- hydroxybenzeneacetamide (interm.5). d) A mixture of intermediate (5) (30 g), hydrochloric acid 12N (300 ml) and 1-propanol (100 ml) was stirred and refluxed for 17 hours and poured on ice. The mixture was extracted with ethyl acetate. The aqueous phase was basified with ammonium hydroxide and extracted with dichloromethane. The dichloromethane extracts were dried, filtered and evaporated, yielding 7.3 g (36.0%) of (+)-(B)-4-amino-α-(3-chlorophenyl)-3- nitrobenzenemethanamine (interm. 6). e) A mixture of intermediate (6) (7.3 g), 2-isothiocyanato-l,l-dimethoxyethane (4.8 g) and methanol (75 ml) was stirred and refluxed for 2 hours. The mixture was evaporated to an oily residue, yielding 11 g (100%) of (+)-(B)-N-[(4-amino-3-nitrophenyl)(3- chlorophenyl)methyl]-N’-(2,2-dimethoxyethyl)thiourea (interm.7). f) A mixture of intermediate (7) (11 g), iodomethane (2 ml) and potassium carbonate (4.97 g) was stirred at room temperature for 48 hours. The solvent was evaporated and the residue was taken off with dichloromethane and washed with water. The organic layer was dried, filtered and evaporated, yielding 11.4 g of (+)-(S)-methyl (B)-N- [(4-amino-3-nitrophenyl)(3-chlorophenyl)methyl]-N’-(2,2-dimethoxyethyl)carbam- imidothioate as an oily residue (interm. 8). g) To intermediate (8) (11.4 g) at 0°C was added sulfuric acid (100ml) (precooled to 5°C). The mixture was stirred at 5°C until complete dissolution and then was warmed to room temperature. After stirring for 2 hours, the solution was poured on ice and basified with ammonium hydroxide. The aqueous solution was extracted with ethyl acetate. The organic layer was dried, filtered and evaporated. The residue was purified by column chromatography (eluent : CH₂CI₂/CH3OH 98:2). The eluent of the desired fraction was evaporated, yielding 3.7 g (38.0%) of (+)-(B)-4-[(3-chlorophenyl)[2-(methylthio)-lH- imidazol-l-yl]methyl]-2-nitrobenzenamine (interm.9). h) A mixture of intermediate (9) (6.2 g), Raney nickel (6 g) and methanol (100 ml) was hydrogenated for 2 hours at 2 bar and at room temperature. After the calculated amount of hydrogen was taken up, the catalyst was filtered off. The filtrate, (+)-(B)-4-[(3- chlorophenyl)[2-(methylthio)-lH-imidazol-l-yl]methyl]-l,2-benzenediamine (interm. 10), was used for the next step. i) A mixture of intermediate (10) (5.7 g), methanimidamide monoacetate (5.2 g) and methanol (100 ml) was stirred and refluxed for 3 hours. The reaction mixture was evaporated and the residue was taken off in dichloromethane and washed with sodium hydrogen carbonate (10%). The organic layer was dried, filtered and evaporated. The oily residue was purified by column chromatography (eluent : CH2CI2/CH3OH 95:5). The eluent of the desired fraction was evaporated, yielding 4.9 g (83.7%) of (+)-(B)-5-[(3-cWorophenyl)[2-(methylthio)-lH-imidazol-l-yl]methyl]-lH-benzimidazole (interm. 11).

B. Preparation of the final compounds Example 3

A mixture of intermediate (2) (185 g) in water (512 ml) was stirred at 20 °C. Hydrochloric acid (289 ml) was added. Formic acid (85%) (61.17 ml) was added and this mixture was heated to 55°C. The reaction mixture was stirred for 3 hours at 55 °C and then cooled to 20°C. Dichloromethane (1223 ml) was added. Ammonium hydroxide (730 ml) was added dropwise at < 25°C. The separated organic layer was washed with water (500 ml), dried, filtered and the solvent was evaporated, yielding 152.88 g (108.5%) of product. A sample was dried (18 hours at 55 °C), yielding 3.18 g of (+)-(B)-5-[(3-chlorophenyl)-lH-imidazol-l-ylmethyl]-lH-benzimidazole; mp.

20 113.7°C; [αjj = +43.46° (c = 1% in methanol) (comp. 1).

Example 4

A mixture of intermediate (11) (4.9 g), Raney nickel (2 g) and ethanol (100ml) was stirred and refluxed for 5 days, while every day an additional amount of Raney nickel (2 g) was added. The catalyst was filtered off and rinsed with dichloromethane. The filtrate was evaporated and the residue was purified twice by column chromatography (silica gel; CH2CI2/CH3OH 95:5 ; CH2CI2/CH3OH NH4OH 80:20:3). The eluent of the desired fraction was evaporated and the residue was converted into the hydrochloride salt in 2-propanol and ethanol. The salt was recrystallized from 2-butanone, yielding 1.8 g (37.2%) of (+)-(B)-5-[(3-chlorophenyl)(lH-imidazol-l-yl)methyl]-lH-benzimidazole

20 monohydrochloride; mp. 212.1°C; [α]_D = +42.43° (c = 1% in ethanol) (comp. 2)

Example 5

Compound (1) (149.7 g) was dissolved in 2-butanone (2424 ml). A mixture of hydrochloric acid in 2-propanol (82.6 ml) in 2-butanone (727 ml) was added over a 2 hour period at 20 °C. The reaction mixture was stirred for 16 hours at 20 °C. The precipitate was filtered off, washed with 2-butanone (242 ml) and dried (vacuum; 80°C); yielding 147.5 g (99.3%) of (+)-(B)-5-[(3-chlorophenyl)-lH-imidazol-l-ylmethyl]-lH-

20 benzimidazole monohydrochloride; mp. 214.5°C; [α] _j = +36.20° (c = 1% in methanol) (comp. 2). Example 6

A mixture of compound (1) (0.72 g) in ethanol (5.1 ml; denaturated) was stirred at 20 °C until it became homogeneous. (E)-2-butenedioic acid (0.54 g) was added The mixture was stirred for 18 hours at 20 °C and then cooled 0-5 °C and precipitation resulted. More denaturated ethanol (2 ml) was added and the mixture was stirred for 2 hours at 20 °C. The precipitate was filtered off, washed with ethanol (3 ml; denaturated) and dried (vacuum; 50 °C), yielding 0.26 g (23.4%) (B)-5-[(3-chlorophenyl)-lH-imidazol-l-yl- methyl]-lH-benzimidazole (E)-2-butenedioate (2:3).ethanolate (2:1); mp. 111.2°C (comp. 3).

PAPER

Improved synthesis of liarozole

see at..http://lib.syphu.edu.cn/71%E6%A0%A1%E5%86%85%E7%BD%91%E4%B8%93%E7%94%A8/zwlw%E5%85%A8%E6%96%87/60230.pdf

Paper

Conversion of the Laboratory Synthetic Route of the N-Aryl-2-benzothiazolamine R116010 to a Manufacturing Method

1 Vahlquist, A; Blockhuys, S; Steijlen, P; Van Rossem, K; Didona, B; Blanco, D; Traupe, H (2013). “Oral liarozole in the treatment of patients with moderate/severe lamellar ichthyosis: Results of a randomized, double-blind, multinational, placebo-controlled phase II/III trial”. The British journal of dermatology 170 (1): n/a. doi:10.1111/bjd.12626. PMID 24102348.

https://www.researchgate.net/profile/Marc_Le_Borgne/publication/8068537_2-_and_3-%28aryl%29%28azolyl%29methylindoles_as_potential_non-steroidal_aromatase_inhibitors/links/02e7e52fe95f662b24000000.pdf

Literature References: Inhibits cytochrome P450-dependent enzymes involved in steroid biosynthesis and retinoic acid catabolism. Prepn: A. H. M. Raeymaekers et al., EP 260744; eidem, US 4859684 (1988, 1989 both to Janssen). In vivo antitumor activity: R. Van Ginckel et al., Prostate 16, 313 (1990). Pharmacology and effect on steroid synthesis: J. Bruynseels et al., ibid., 345; and effect on retinoic acid: R. De Coster et al., J. Steroid Biochem. Mol. Biol. 43, 197 (1992). Clinical evaluation in prostate cancer: C. Mahler et al., Cancer 71, 1068 (1993); in psoriasis: P. Dockx et al., Br. J. Dermatol. 133, 426 (1995); in combination therapy for malignant brain tumors: M. E. Westarp et al., Onkologie 16, 22 (1993).

Liarozole

Names
IUPAC name 6-[(3-Chlorophenyl)-imidazol-1-ylmethyl]-1H-benzimidazole
Identifiers
CAS Number	115575-11-6
ChemSpider	54664
IUPHAR/BPS	5210
Jmol interactive 3D	Image
PubChem	60652
InChI[show]
SMILES[show]
Properties
Chemical formula	C17H13ClN4
Molar mass	308.77 g·mol−1

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C1=CC(=CC(=C1)Cl)C(C2=CC3=C(C=C2)N=CN3)N4C=CN=C4

Zydus Chairman and Managing Director,Mr. Pankaj R. Patel won the prestigious ‘Gujarat Business Leader of the Year’ award at the CNBC Bajar, Gujarat Ratna Awards 2015-16 from Hon’ble Chief Minister of Gujarat, Smt. Anandiben Patel at a glittering ceremony held at Hyatt, Ahmedabad.

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Lamivudine (2′,3′-dideoxy-3′-thiacytidine, commonly called 3TC) is an antiretroviral medication used to prevent and treat HIV/AIDS and used to treat chronic hepatitis B.^[1]

It is of the nucleoside analog reverse transcriptase inhibitor (NRTI) class. It is marketed in the United States under the tradenames Epivir and Epivir-HBV.

It is on the World Health Organization’s List of Essential Medicines, a list of the most important medication needed in a basic health system.^[2] As of 2015 the cost for a typical month of medication in the United States is more than 200 USD.^[3]

Medical uses

Lamivudine has been used for treatment of chronic hepatitis B at a lower dose than for treatment of HIV/AIDS. It improves the seroconversion of e-antigen positive hepatitis B and also improves histology staging of the liver. Long term use of lamivudine leads to emergence of a resistant hepatitis B virus (YMDD) mutant. Despite this, lamivudine is still used widely as it is well tolerated.

Resistance

In HIV, high level resistance is associated with the M184V/I mutation in the reverse transcriptase gene as reported by Raymond Schinazi’s group at Emory University. GlaxoSmithKline claimed that the M184V mutation reduces “viral fitness”, because of the finding that continued lamivudine treatment causes the HIV viral load to rebound but at a much lower level, and that withdrawal of lamivudine results in a higher viral load rebound with rapid loss of the M184V mutation; GSK therefore argued that there may be benefit in continuing lamivudine treatment even in the presence of high level resistance, because the resistant virus is “less fit”. The COLATE study has suggested that there is no benefit to continuing lamivudine treatment in patients with lamivudine resistance.^[4] A better explanation of the data is that lamivudine continues to have a partial anti-viral effect even in the presence of the M184V mutation.

In hepatitis B, lamivudine resistance was first described in the YMDD (tyrosine–methionine–aspartate-aspartate) locus of the HBV reverse transcriptase gene. The HBV reverse transcriptase gene is 344 amino acids long and occupies codons 349 to 692 on the viral genome. The most commonly encountered resistance mutations are M204V/I/S.^[5] The change in amino acid sequence from YMDD to YIDD results in a 3.2 fold reduction in the error rate of the reverse transcriptase, which correlates with a significant growth disadvantage of the virus. Other resistance mutations are L80V/I, V173L and L180M.^[6]

Mechanism of action

Lamivudine is an analogue of cytidine. It can inhibit both types (1 and 2) of HIV reverse transcriptase and also the reverse transcriptase of hepatitis B virus. It is phosphorylated to active metabolites that compete for incorporation into viral DNA. They inhibit the HIV reverse transcriptase enzyme competitively and act as a chain terminator of DNA synthesis. The lack of a 3′-OH group in the incorporated nucleoside analogue prevents the formation of the 5′ to 3′ phosphodiester linkage essential for DNA chain elongation, and therefore, the viral DNA growth is terminated.

Lamivudine is administered orally, and it is rapidly absorbed with a bio-availability of over 80%. Some research suggests that lamivudine can cross the blood–brain barrier. Lamivudine is often given in combination with zidovudine, with which it is highly synergistic. Lamivudine treatment has been shown to restore zidovudine sensitivity of previously resistant HIV. Lamivudine showed no evidence of carcinogenicity or mutagenicity in in vivo studies in mice and rats at doses from 10 to 58 times those used in humans.^[7]

History

Racemic BCH-189 (the minus form is known as Lamivudine) was invented by Dr. Bernard Belleau while at work at McGill University and Dr Paul Nguyen-Ba at the Montreal-based IAF BioChem International, Inc. laboratories in 1988 and the minus enantiomer isolated in 1989. Samples were first sent to Dr. Yung-Chi Cheng of Yale University for study of its toxicity. When used in combination with AZT, he discovered that Lamivudine’s negative form reduced side effects and increased the drug’s efficiency at inhibiting reverse transcriptase.^[8] The combination of Lamivudine and AZT increased the efficiency at inhibiting an enzyme HIV uses to reproduce its genetic material. As a result, Lamivudine was identified as a less toxic agent to mitochondria DNA than other retroviral drugs.^[9]

Lamivudine was approved by the Food and Drug Administration (FDA) on November 17, 1995 for use with zidovudine (AZT) and again in 2002 as a once-a-day dosed medication. The fifth antiretroviral drug on the market, it was the last NRTI for three years while the approval process switched to protease inhibitors. According to the manufacturer’s 2004 annual report, its patent will expire in the United States in 2010 and in Europe in 2011.

On September 2014, Dr. Gorbee Logan, a Liberian physician, reported positive results while treating Ebola virus disease with Lamivudine. Out of 15 patients treated with the antiviral, 13 (those treated within the third to fifth day of symptoms being manifested) survived the disease and were declared virus-free; the remaining cases (treated from the fifth day or later) died.^[10][11]

Presentation

• Epivir 150 mg or 300 mg tablets (GlaxoSmithKline; US and UK) for the treatment of HIV;
• Epivir-HBV 100 mg tablets (GlaxoSmithKline; US only) for the treatment of hepatitis B;
• Zeffix 100 mg tablets (GlaxoSmithKline; UK only) for the treatment of hepatitis B.
• 3TC 150 mg tablets (GlaxoSmithKline; South Africa) for the treatment of HIV;

Lamivudine is also available in fixed combinations with other HIV drugs:

• Combivir (with zidovudine);
• Epzicom/Kivexa (with abacavir);
• Trizivir (with zidovudine and abacavir)

Lamivudine (I) (CAS No. 134678-17-4) is chemically known as (-)-[2R,5S]-4_T amino- 1 – [2-(hydroxymethyl)- 1 ,3 -oxathiolan-5-yl] -2( 1 H)-pyrimidin-2-one.

Formula (I)

Lamivudine is a reverse transcriptase inhibitor used alone or in combination with other classes of Anti-HIV drugs in the treatment of HIV infection. It is available commercially as a pharmaceutical composition under the brand name EPIVIR^®, marketed by GlaxoSmithKline, and is covered under US 5,047,407.

This molecule has two stereo-centres, thus giving rise to four stereoisomers: (±)- Cis Lamivudine and (±)-Trans Lamivudine. The pharmaceutically active isomer however is the (-)-Cis isomer which has the absolute configuration [2R,5S] as show in Formula (I).

US 5,047,407 discloses the 1,3-oxathiolane derivatives; their geometric (cis/trans) and optical isomers. This patent describes the preparation of Lamivudine as a mixture of cis and trans isomers (shown in scheme I). The diastereomers obtained are converted into N-acetyl derivatives before separation by column chromatography using ethylacetate and methanol (99:1); however, this patent remains silent about further resolution of the cis isomer to the desired (-)- [2R,5S]-Cis-Lamivudine. Secondly, as the ethoxy group is a poor leaving group, the condensation of cytosine with compound VI gives a poor yield, i.e. 30 – 40%, of compound VII. Thirdly, chromatographic separation that has been achieved only after acetylation requires a further step of de-acetylation of the cis-(±)- isomer. Also, separation of large volumes of a compound by column chromatography makes the process undesirable on a commercial scale.

(+/-) Cis (+/-) Cis Lamivudine (VIII)

Scheme – 1 Efforts have been made in the past to overcome the shortcomings of low yield and enantiomeric enrichment, hi general, there have been two approaches to synthesize (— )-[2R,5S]-Cis-Lamivudine. One approach involves stereoselective synthesis, some examples of which are discussed below.

US 5,248,776 describes an asymmetric process for the synthesis of enantiomerically pure β-L-(-)-l,3-oxathiolone-nucleosides starting from optically pure 1,6-thioanhydro-L-gulose, which in turn can be easily prepared from L- Gulose. The condensation of the 1,3-oxathiolane derivative with the heterocyclic base is carried out in the presence of a Lewis acid, most preferably SnCl_4, to give the [2R,5R] and [2R,5S] diastereomers that are then separated chromatographically.

US 5,756,706 relates a process where compound A is esterified and reduced to compound B. The hydroxy group is then converted to a leaving group (like acetyl) and the cis- and trans-2R-tetrahydrofuran derivatives are treated with a pyrimidine base, like N-acetylcytosine, in the presence trimethylsilyl triflate to give compound C in the diastereomeric ratio 4: 1 of cis and trans isomers.

A B C

Z = S₅ CH

Dissolving compound C in a mixture of 3:7 ethyl acetate-hexane separates the cis isomer. The product containing predominantly the cis-2R,5S isomer and some trans-2R,5R compound is reduced with NaBH₄ and subjected to column chromatography (30% MeOH-EtOAc) to yield the below compound.

US 6,175,008 describes the preparation of Lamivudine by reacting mercaptoacetaldehyde dimer with glyoxalate and further with silylated pyrimidine base to give mainly the cis-isomer by using an appropriate Lewis acid, like TMS-

I₅ TMS-Tf, TiCl₄ et cetera. However the stereoselectivity is not absolute and although the cis isomer is obtained in excess, this process still requires its separation from the trans isomer. The separation of the diastereomers Js done by acetylation and chromatographic separation followed by deacetylation. Further separation of the enantiomers of the cis-isomer is not mentioned.

US 6,939,965 discloses the glycosylation of 5-fluoro-cytosine with compound F (configuration: 2R and 2S)

. F

The glycosylation is carried out in the presence of TiCl₃(OiPr) which is stereoselective and the cis-2R,5S-isomer is obtained in excess over the trans- 2S,5S-isomer. These diastereomers are then separated by fractional crystallization.

US 6,600,044 relates a method for converting the undesired trans-l,3-oxathiolane nucleoside to the desired cis isomer by a method of anomerizatioή or transglycosylation and the separation of the hydroxy-protected form of cis-, trans- (-)-nucleosides by fractional crystallization of their hydrochloride, hydrobromide, methanesulfonate salts. However, these cis-trans isomers already bear the [R] configuration at C2 and only differ in their configuration at C5; i.e. the isomers are [2R,5R] and [2R,5S]. Hence diastereomeric separation directly yields the desired [2R, 5S] enantiomer of Lamivudine.

In the second approach to prepare enantiomerically pure Lamivudine the resolution of racemic mixtures of nucleosides is carried out. US 5,728,575 provides one such method by using enzyme-mediated enantioselective hydrolysis of esters of the formula

wherein, ‘R’ is an acyl group and ‘Rl ‘ represents the purine or pyrimidine base.

‘R’ may be alkyl carboxylic, substituted alkyl carboxylic and preferably an acyl group that is significantly electron-withdrawing, eg. α-haloesters. After selective hydrolysis, the process involves further separation of the unhydrolyzed ester from the^‘ enantiomerically pure 1,3-oxathiolane-nucleoside. Three methods are suggested in this patent, which are:

1. Separation of the more lipophilic unhydrolyzed ester by solvent extraction with one of a wide variety of nonpolar organic solvents.

2. Lyophilization followed by extraction into MeOH or EtOH. 3. Using an HPLC column designed for chiral separations.

In another of its aspects, this patent also refers to the use of the enzyme cytidine- deoxycytidine deaminase, which is enantiomer-specific, Λo catalyze the deamination of the cytosine moiety and thereby converting it to uridine. Thus, the enantiomer that remains unreacted is still basic and can be extracted by using an acidic solution.

However, the above methods suffer from the following drawbacks, (a) Enzymatic hydrolysis sets down limitations on choice of solvents: alcohol solvents cannot be used as they denature enzymes. (b) Lyophilization on an industrial scale is tedious, (c) Chiral column chromatographic separations are expensive.

WO 2006/096954 describes the separation of protected or unprotected enantiomers of the cis nucleosides of below formula by using a chiral acid to form diastereomeric salts that are isolated by filtration. Some of the acids used are R-

(-)-Camphorsulfonic acid, L-(-)-Tartaric acid, L-(-)-Malic acid, et cetera.

However, the configuration of these CIS-nucleosides are [2R,4R] and [2S,4S] as the heterocyclic base is attached at the 4 position of the oxathiolane ring and the overall stereo-structure of the molecule changes from that of the 2,5-substituted oxathiolane ring.

Thus various methods are described for the preparation of Lamivudine. However there is no mention in the prior art about the separation of an enantiomeric pair, either cis-(±) or trans-(±), from a mixture containing cis-[2R,5S], [2S,5R] and trans-[2R,5R], [2S,5S] isomers. Further, there also is a need to provide resolution of the cis-(±) isomers to yield the desired enantiomer in high optical purity.

CN 1223262 (Deng et aϊ) teaches the resolution of a certain class of compounds called Prazoles by using chiral host compounds such as dinaphthalenephenols (BINOL), diphenanthrenols or tartaric acid derivatives. The method consists of the formation of a 1:1 complex between the chiral host (BINOL) and one of the enantiomers, the guest molecule. The other enantiomer remains in solution. (S)- Omeprazole, which is pharmaceutically active as a highly potent inhibitor of gastric acid secretion, has been isolated from its racemic mixture in this manner by using S-BINOL.

BINOL is a versatile chiral ligand that has found its uses in various reactions involving asymmetric synthesis (Noyori, R. Asymmetric Catalysis in Organic

Synthesis) and optical resolution (Cram, D. J. et al J. Org. Chem. 1977, 42, 4173-

4184). Some of these reactions include BINOL-mediated oxidation and reduction reactions, C-C bond formation reactions such as Aldol reaction, Michael addition,

Mannich reaction et cetera (Brunei Chem. Rev. 2005 105, 857-897) and kinetic resolution, resolution by inclusion complexation et cetera.

BINOL, or l,l’-bi-2-Naphthol, being an atropoisomer possesses the property of chiral recognition towards appropriate compounds. One of the uses of BINOL in resolution that is known in literature is in Host-Guest complexation. In one such example, 1,1-binaphthyl derivatives have been successfully incorporated into optically active crown ethers for the enantioselective complexation of amino acid esters and chiral primary ammonium ions (Cram, D. J. Ace. Chem. Res. 1978, 11, 8-14). The chiral ‘host’ is thus able to discriminate between enantiomeric compounds by the formation of hydrogen bonds between the ether oxygen and the enantiomers. The complex formed with one of the isomers, the ‘guest’, will be less stable on steric grounds and this forms the basis for its separation.

It is evident from the literature cited that there exists a need to (a) synthesize Lamivudine by a process requiring less expensive, less hazardous and easily available reagents, and (b) achieve good yields with superior quality of product without resorting to column chromatography as a means of separation, thereby making the process of Lamivudine manufacture more acceptable industrially.

CLIP

ideally, the chemical synthesis of APIs begins from simple, inexpensive building blocks or RMs that are used for multiple purposes and are available in the fine chemicals industry, though some require uncommon RMs that contribute significantly to API manufacturing cost. RMs are converted into APIs by multi-step processes of breaking old chemical bonds and making new ones. A synthesis of 3TC is shown in . In the seven-step sequence, six steps involve breaking existing chemical bonds and creating new ones to build the molecular architecture of the API. The final recrystallization of an API is a critical step; at this stage the crystalline form of the API is determined and related substances (impurities) are removed or reduced to acceptable levels. APIs are often milled in a final step so that their particle size distribution (PSD) falls within specified limits. The crystalline form and PSD of an API must be controlled, because these properties are often critical to the formulation, dissolution, absorption and bioavailability of a drug. Bioavailability is the fraction of a drug dose that reaches systemic circulation (that is, is present in blood plasma) after administration. By definition, a drug is 100% bioavailable when administered by injection; drugs for ART are taken every day and administration by injection is not possible.

The cost of ART is absolutely critical to ensuring access in LMICs. The cost of manufacturing an API is dependent upon the cost of RMs, the cost of overheads and labour (OHL) and volume demand for the product. OHL includes the capital investment to build a manufacturing facility and operating costs, including personnel and energy, waste disposal and the eventual cost of decommissioning of the facility. Increased volume demand generally decreases the cost contribution of RM and OHL. Substantial production volumes are required to obtain full economy of scale . Producing 1–5 metric tons per year is substantially more expensive per kilogram than producing 100 metric tons of an API. There is a practical limit of approximately 50–100 metric tons/year beyond which cost reductions are modest with increased volume, but this practical limit refers to the volumes of drug manufactured in any single manufacturing plant. Exceptions to these generalizations do occur, most often when demand exceeds either the existing manufacturing capacity for a specific API or the availability of critical RMs . Exceptions that have occurred include shortages of β-thymidine for producing AZT and a squeeze on the availability and price of adenine as a starting material for TDF. Another contributor to RM and OHL costs is the efficiency of a chemical synthesis. Since operating costs for a manufacturing facility may be USD2,000/h, the number of steps or processing time for a chemical synthesis affects manufacturing cost. The efficiency of a synthesis is often quoted as an E-factor  representing the kilograms of waste produced per kilogram of product manufactured. Waste management is expensive in chemical manufacturing wherever environmental guidelines are both reasonable and followed. From a slightly different perspective, increasing the overall yield of an API synthesis reduces RM use and associated cost for manufacturing.

Jinliang L, Feng LV. inventors; Shanghai Desano Pharmaceutical, assignee. A process for stereoselective synthesis of lamivudine. European Patent Application EP 2161 267 A1. 2007 June 29.

 

Object of the invention

Thus, one object of the present invention is to provide a process for the synthesis of_Lamivudine which is cost effective, uses less hazardous and easily available reagents, yet achieves good yields with superior quality of product without resorting to column chromatography.

A further object of the present invention is to provide an improved process for the synthesis of Lamivudine, by separating the mixture of diastereomers: Cis-[2R,5S], [2S,5R] from Trans-[2R,5R], [2S,5S] and then resolving the Cis isomers using BINOL to obtain (-)-[2R,5S]^Cis-Lamivudine with at least 99% ee.

This 1,3-oxathiolane compound VIII is further condensed with silylated cytosine in the presence of a Lewis acid such as trimethylsilyliodide to get protected 6-amino-3 – {2-hydroxymethyl- 1 ,3 -oxathiolan-5-yl} -3 -hydropyrimidine- 2-one (compound IX). OH

Cis(±)and Trans (±) racemic mixtures

Lamivudine (-)-[2

Compound (IX) is mixture of following optical isomers

SCHEME 2 The separation of the four-component diastereomeric mixture of isomers bearing the following configuration: trans-[2R,5R], [2S.5S] and cis-[2R,5S], [2S,5R] forms the next step. The separation efficiency of the benzoyl-protected compound

Example 9

Preparation of Lamivudine: (-)-[2R,5S]-4-amino-l-[2-(hydroxymethyl)-l,3- oxathiolan-5 -yl] -2(1 H)-pyrimidin-2-one

Compound I 5mL of cone. HCl was slowly added to a solution of 2Og of Lamivudine-BINOL complex in 100ml of ethylacetate and 10OmL of DM water (pH 2-2.5). The layers. were separated and a 10OmL aliquot of ethylacetate was added to the aqueous layer. The layers were separated again and the aqueous layer was neutralized using 1OmL of 10% aqueous NaOH solution. The solvent was recovered under vacuum at 40-45 ⁰C, the product obtained was dissolved in 160 mL of methanol, filtered, the filtrate was concentrated and 32 mL of water-ethanol mixture (3:1) was added to this product, heated to get a clear solution, cooled to 5 – 10 ⁰C and then filtered. The residue was vacuum dried at 45-50 ⁰C. Yield: 4-5g.

Enantiomeric excess = 99.74 % m.p. = 133-135 °C [<X]_D at 25°C = 98.32° (c = 5 water)

¹H NMR (DMSO d₆): 2.99-3.07 (dd, IH), 3.35-3.38 (dd, IH), 3.72-3.74 (m, 2H), 5.14-5.18 (t, IH), 5.32-5.38 (t, IH), 5.71-5.75 (d, IH), 6.16-6.21 (t, IH), 7.22-

7.27 (d, 2H), 7.80-7.83 (d, IH)

Moisture content: 1.67%

IR (in KBr, cm^“1): 3551, 3236, 2927, 1614, 1492, 1404, 1336, 1253, 1146, 1052,

967, 786. MS: M+l =230

XRD [2Θ] (Cu – K_a1=I.54060A, K_a2=1.54443A K_β= 1.39225A; 4OmA, 45kV):

5.08, 9.89, 10.16, 11.40, 11.65, 12.96, 13.23, 15.26, 15.82, 17.74, 18.74, 18.88,

19.67, 20.69, 22.13, 22.88, 23.71, 25.47, 26.07.

PATENT

http://www.google.com/patents/WO2013021290A1?cl=en

EP 0382526; EP 0711771; JP 1996119967; JP 2000143662; US 5047407

 Lamivudine is a nucleoside reverse transcriptase inhibitor, and is a kind of deoxycytidine analogue, which can inhibit the reproduction of Human immunodeficiency virus (HIV) and hepatitis B virus (HBV), whose chemical name is (2R-cis)-4-amino-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-1H-pyrimidin-2-one, and structural formula is as follows:

• In 1990, Belleau et al firstly reported Lamivudine structure, and BioChem Pharma of Canada firstly developed Lamivudine to be used to treat AIDS ( WO91/17159 ) and hepatitis B ( EP0474119 ), and found that it had distinguished therapeutic effect on hepatitis B. Since Lamivudine has two chiral centers, it has 4 stereisomers, among which the 2R,5S (2R–cis)-isomer is the most potent in anti-HIV and anti-HBV activities, and its cytotoxicity on some cells is lower than its enatiomer or racemic body.

• WO94/14802 mentioned two synthetic schemes (see Scheme 1 and Scheme 2):
• In the above two schemes of this process, chirality was not controlled, and the final product was obtained by column chromatography, thus the yield was low and the requirement on the equipment was high, resulting in that the production cost was high and the operation in the production could not be controlled easily.

 The specific reaction scheme is as follows:

 synthetic route is preferably as follows:

. The specific reaction scheme is as follows:

 The specific reaction scheme is as follows:

Example 8 The preparation of (2R,5S)-4-amino-1-(2-hydroxymethyl-1,3-oxathiolane-5-yl) -2(1H)-pyrimidone (Lamivudine)

• The compound of Example 7 (41.0g, 0.1mol) and methanol (250ml) were added to a reaction flask, and then stirred to make the compound dissolved in methanol. The mixture was cooled to 0 °C, and then K₂CO₃ (41.2g, 0.3mol) was added. The mixture was further stirred at room temperature overnight and then was adjusted by 0.1N HCl to a pH of about 7. The mixture was filtered and the solvent was evaporated under reduced pressure from the filtrate, and then to the residue was added 150ml of water. The aqueous layer was extracted by 150ml of toluene (50ml X 3), and then p-nitrobenzoic acid (16.8g, 0.1mol) was added to the aqueous layer and refluxed for 30 minutes, after which, the reaction mixture was cooled and further stirred at 0-5 °C for 2 hours. Then the reaction mixture was filtered and dried to give 31.7g of a white solid.
• The resulting salt and anhydrous ethanol (120ml) were added to a reaction flask, and warmed to 70-75 °C. Triethylamine (12ml) was added dropwise, and the reaction was conducted at that temperature for 2 hours. Then the mixture was cooled to 50 °C, at which point ethyl acetate (150ml) was added dropwsie. After the addition was complete, the mixture was cooled to 10 °C and further stirred for 4 hours. The mixture was filtered to give 15.6g of Lamivudine, and the yield was 68%. 1H-NMR (DMSO-d6) δ: 7.83(dd, 1H), 7.17∼7.23(dd, 2H), 6.21(t, 1H), 5.72 (dd, 1H), 5.29 (t, 2H), 5.16 (t, 1H), 3.70∼3.74 (m, 2H), 3.32∼3.43 (dd, 1H), 3.01∼3.05(dd, 1H); Elemental analysis: C8H11N3O3S found(%): C 41.85, H 4.88 N 18.25, S 13.94; calculated (%) C 41.91, H 4.84, N 18.33, S13.99.

PAPER

http://www.beilstein-journals.org/bjoc/single/articleFullText.htm?publicId=1860-5397-9-265

References

• 1″Lamivudine”. The American Society of Health-System Pharmacists. Retrieved 31 July 2015.
• 2
• “WHO Model List of EssentialMedicines” (PDF). World Health Organization. October 2013. Retrieved 22 April 2014.
• 3
• Hamilton, Richart (2015). Tarascon Pocket Pharmacopoeia 2015 Deluxe Lab-Coat Edition. Jones & Bartlett Learning. p. 65. ISBN 9781284057560.
• 4
• Fox Z, Dragsted UB, Gerstoft J, et al. (2006). “A randomized trial to evaluate continuation versus discontinuation of lamivudine in individuals failing a lamivudine-containing regimen: The COLATE trial”. Antiviral Therapy 11 (6): 761–70. PMID 17310820.
• 5
• http://hivdb.stanford.edu/index.html Stanford University Drug Resistance Database.
• 6
• Koziel MJ, Peters MG (2007). “Viral hepatitis in HIV infection”. N Engl J Med 356 (14): 1445–54. doi:10.1056/NEJMra065142. PMID 17409326.
• 7
• “Epivir package insert” (PDF). GlaxoSmithKline. Retrieved January 20, 2011.
• 8
• Curtis, John (June 20, 1998). “Hunting Down HIV”. Yale Medicine.
• 9
• Soderstrom, E John (July 10, 2003). “National Institutes of Health: Moving Research from the Bench to the Bedside”.
• 10
• Cohen, Elizabeth (September 29, 2014). “Doctor treats Ebola with HIV drug in Liberia — seemingly successfully”. CNN.
• 11 HIV drug may stop Ebola. Operonlabs.com, 27 September 2014

 Literature References: Reverse transcriptase inhibitor. Prepn: J. A. V. Coates et al., WO 9117159 C.A. 117, 111989 (1991). Synthesis of enantiomers: J. W. Beach et al., J. Org. Chem. 57, 2217 (1992); of (-)-enantiomer: D. C. Humber et al., Tetrahedron Lett. 33, 4625 (1992). HPLC determn in urine: D. M. Morris, K. Selinger, J. Pharm. Biomed. Anal. 12, 255 (1994). Clinical trial in hepatitis B: F. Nevens et al., Gastroenterology 113, 1258 (1997). Review of pharmacology and clinical efficacy in HIV infection: C. M. Perry, D. Faulds, Drugs 53, 657-680 (1997).

External links

• Epivir (manufacturer’s website)

Lamivudine

Systematic (IUPAC) name
4-amino-1-[(2R,5S)-2-(hydroxymethyl)-1,3-oxathiolan-5-yl]-1,2-dihydropyrimidin-2-one
Clinical data
Trade names	Epivir
AHFS/Drugs.com	monograph
MedlinePlus	a696011
Pregnancy category	AU: B3 US: C (Risk not ruled out)
Routes of administration	Oral
Legal status
Legal status	UK: POM (Prescription only) US: ℞-only
Pharmacokinetic data
Bioavailability	86%
Protein binding	Less than 36%
Biological half-life	5 to 7 hours
Excretion	Renal (circa 70%)
Identifiers
CAS Number	134678-17-4
ATC code	J05AF05 (WHO)
PubChem	CID 73339
DrugBank	DB00709
ChemSpider	66068
UNII	2T8Q726O95
KEGG	D00353
ChEMBL	CHEMBL141
NIAID ChemDB	000388
Synonyms	L-2′,3′-dideoxy-3′-thiacytidine
PDB ligand ID	3TC (PDBe, RCSB PDB)
Chemical data
Formula	C8H11N3O3S
Molar mass	229.26 g/mol

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BRIVARACETAM, UCB-34714

(2S)-2-[(4R)-2-oxo-4-propylpyrrolidin-1-yl]butanamide

UNII-U863JGG2IA

UCB; For the treatment of partial onset seizures related to epilepsy, Approved February 2016

Brivaracetam, the 4-n-propyl analog of levetiracetam, is a racetam derivative with anticonvulsant properties.^[1][2] Brivaracetam is believed to act by binding to the ubiquitous synaptic vesicle glycoprotein 2A (SV2A).^[3] Phase II clinical trials in adult patients with refractory partial seizures were promising. Positive preliminary results from stage III trials have been recorded,^[4][5] along with evidence that it is around 10 times more potent^[6] for the prevention of certain types of seizure in mouse models than levetiracetam, of which it is an analogue.

On 14 January 2016, the European Commission,^[7] and on 18 February 2016, the USFDA^[8] approved brivaracetam under the trade name Briviact (by UCB). The launch of this anti-epileptic is scheduled for the first quarter of that year. Currently, brivaracetam is still not approved in other countries like Australia, Canada and Switzerland.

Brivaracetam was approved by European Medicine Agency (EMA) on Jan 14, 2016 and approved by the U.S. Food and Drug Administration (FDA) on Feb 18, 2016. It was developed and marketed as Briviact^® by UCB in EU/US.

Brivaracetam is a selective high-affinity synaptic vesicle protein 2A ligand, as an adjunctive therapy in the treatment of partial-onset seizures with or without secondary generalization in adult and adolescent patients from 16 years of age with epilepsy.

Briviact^® is available in three formulations, including film-coated tablets, oral solution and solution for injection/infusion. And it will be available as 10 mg, 25 mg, 50 mg, 75 mg and 100 mg film-coated tablets, a 10 mg/ml oral solution, and a 10 mg/ml solution for injection/infusion. The recommended starting dose is either 25 mg twice a day or 50 mg twice a day, depending on the patient’s condition. The dose can then be adjusted according to the patient’s needs up to a maximum of 100 mg twice a day. Briviact can be given by injection or by infusion (drip) into a vein if it cannot be given by mouth.

European Patent No. 0 162 036 Bl discloses compound (S)-α-ethyl-2-oxo-l- pyrrolidine acetamide, which is known under the International Non-proprietary Name of Levetiracetam.

Levetiracetam

Levetiracetam is disclosed as a protective agent for the treatment and prevention of hypoxic and ischemic type aggressions of the central nervous system in European patent EP 0 162 036 Bl. This compound is also effective in the treatment of epilepsy.

The preparation of Levetiracetam has been disclosed in European Patent No. 0 162 036 and in British Patent No. 2 225 322.

International patent application having publication number WO 01/62726 discloses 2-oxo-l -pyrrolidine derivatives and methods for their preparation. It particularly discloses compound (2S)-2-[(4R)-2-oxo-4-propyl-pyrrolidin-l-yl] butanamide known under the international non propriety name of brivaracetam.

Brivaracetam

International patent application having publication number WO 2005/121082 describes a process of preparation of 2-oxo-l -pyrrolidine derivatives and particularly discloses a process of preparation of (2S)-2-[(4S)-4-(2,2-difluorovinyl)-2-oxo-pyrrolidin-l- yl]butanamide known under the international non propriety name of seletracetam.

Seletracetam

Kenda et al., in J. Med. Chem. 2004, 47, 530-549, describe processes of preparation of 2-oxo-l -pyrrolidine derivatives and particularly discloses compound 1-((1S)-I- carbamoyl-propyl)-2-oxo-pyrrolidone-3-carboxylic acid as a synthetic intermediate.

WO2005028435

CLIPS

The inventors do not comment on the apparent stereoselectivity of the carbonate salts in the reaction of 22 with 23. This is an intriguing finding and worthy of investigation. (UCB S.A. [Brussels]. US Patent 8,338,621

SYNTHESIS

PATENT

WO2005028435

Example 1: Synthesis of (2S)-2-((4R)-2-oxo-4-n-propyl-l-pyrrolidinyl)butanamide 1.1 Synthesis of (2S)-2-aminobutyramide free base

1800 ml of isopropanol are introduced in a 5L reactor. 1800 g of (2S)-2- aminobutyramide tartrate are added under stirring at room temperature. 700 ml of a 25% aqueous solution of ammonium hydroxide are slowly added while maintaining the temperature below 25°C. The mixture is stirred for an additional 3 hours and then the reaction is allowed to complete at 18°C for 1 hour. The ammonium tartrate is filtered. Yield : 86%.

1.2 Synthesis of 5-hydroxy-4-n-propyl-furan-2-one

Heptane (394 ml) and morpholine (127.5 ml) are introduced in a reactor. The mixture is cooled to 0°C and glyoxylic acid (195 g, 150 ml, 50w% in water) is added. The mixture is heated at 20°C during 1 hour, and then valeraldehyde (148.8 ml) is added . The reaction mixture is heated at 43°C during 20 hours. After cooling down to 20^CC, a 37 % aqueous solution of HCl (196.9 ml) is slowly added to the mixture, which is then stirred during 2 hours.

After removal of the heptane phase, the aqueous phase is washed three times with heptane. Diisopropyl ether is added to the aqueous phase. The organic phase is removed, and the aqueous phase further extracted with diisopropyl ether (2x). The diisopropyl ether phases are combined, washed with brine and then dried by azeotropic distillation. After filtration and evaporation of the solvent, 170g of 5- hydroxy-4-n-propyl-furan-2-one are obtained as a brown oil. Yield: 90.8 %

1.3 Synthesis of (2S)-2-((4R)-2-oxo-4-n-propyl-l-pyrrolidinyl)butanamide and (2S)-2-((4S)-2-oxo-4-n-propyl-l-pyrrolidinyl)butanamide

(S, R) (S, S) The (2S)-2-aιninobutyrarnide solution in isopropanol containing 250 g obtained as described here above is dried by azeotropic distillation under vacuum. To the dried (2S)-2-am obutyraιnide solution is added 5-hydroxy-4-n-propyl-furan-2-one (290 g) between 15°C and 25 °C; the mixture is heated to 30 °C and kept for at least 2 hours at that temperature. Acetic acid (1, 18 eq.), Pd/C catalyst (5 w/w%; Johnson Matthey 5% Pd on carbon – type 87L) are then added and hydrogen introduced into the system under pressure. The temperature is kept at 40 °C maximum and the H₂ pressure maintained between 0,2 bar and 0,5 bar followed by stirring for at least 20 hours following the initial reaction. The solution is then cooled to between 15 °C and 25 °C and filtered to remove the catalyst. The solution of product in isopropanol is solvent switched to a solution of product in isopropyl acetate by azeotropic distillation with isopropyl acetate. The organic solution is washed with aqueous sodium bicarbonate followed by a brine wash and then filtered. After recristallisation, 349 g of (2S)-2-((4R)-2- oxo-4-n-propyl-l-pyrrolidinyl)butanamide and (2S)-2-((4S)-2-oxo-4-n-propyl-l- pyιτolidinyl)butanamide are obtained (Yield: 82.5%).

1.4 Preparation of (2S)-2-((4R)-2-oxo-4-n-propyl-l-pyrrolidinyl)butanamide The chromatographic separation of the two diastereoisomers obtained in 1.3 is performed using of (CHIRALPAK AD 20 um) chiral stationary phase and a 45/55 (volume /volume) mixture of n-heptane and ethanol as eluent at a temperature of 25 + 2°C. The crude (2S)-2-((4R)-2-oxo-4-n-propyl-l-pyrrolidinyl)butanamide thus obtained is recristallised in isopropylacetate, yielding pure (2S)-2-((4R)-2-oxo-4-n-propyl-l- pyrrolidinyl)butanamide (Overall yield: 80%) .

Example 2: Synthesis of (2S)-2-((4R)-2-oxo-4-n-propyl-l-pyrrolidinyl)butanamide

Example 1 is repeated except that in step 1.1 a solution of (2S)-2- aminoburyramide.HCl in isopropanol is used (27.72 g, 1.2 equivalent), which is neutralised with a NHs/isopropanol solution (3,4-3,7 mol/L). The resulting ainmonium chloride is removed from this solution by filtration and the solution is directly used for reaction -with 5-hydroxy-4-n-propyl-furan-2-one (23.62 g, 1.0 equivalent) without intermediate drying of the (2S)-2-aminobutyramide solution. Yield after separation of the two diastereoisomers and recristallisation: approximately 84%.

Ref ROUTE1

1. WO0162726A2.

2. WO2005028435A1 / US2007100150A1.

3. J. Med. Chem. 1988, 31, 893-897.

4. J. Org. Chem. 1981, 46, 4889-4894.

PATENT

https://www.google.com/patents/WO2007031263A1?cl=en

Example 3-Synthesis of brivaracetam (I)

3.a. Synthesis of (S) and (R) 2-((R)-2-oxo-4-propyl-pyrrolidin-l-yl)-butyric acid methyl ester fVIaa^*) and (Wlab)

(VIaa) (VIab) A slurry of 60% sodium hydride suspension in mineral oil (0.94g, 23.4 mmol) in tetrahydrofuran (30 mL) is cooled at 0°C under a nitrogen atmosphere. A solution of substantially optically pure (R)-4-propyl-pyrrolidin-2-one (Ilia) (2g, 15.7 mmol) in tetrahydrofuran (2 mL) is added over a 15 minutes period. The reaction mixture is stirred 10 min at 0°C then a solution of methyl-2-bromo-butyric acid methyl ester (V) (3.69g, 20.4 mmol) in tetrahydrofuran (2mL) was added over a 20 minutes period. The reaction mixture is stirred at O⁰C until maximum conversion of starting material and the reaction mixture is then allowed to warm to room temperature and diluted with water (20 mL). Tetrahydrofuran is removed by evaporation and the residue is extracted with isopropyl acetate (20 ml + 10 mL). The combined organic layers are dried on anhydrous magnesium sulfate and evaporated to afford 3g (13.2 mmol, 86 %) of a mixture of epimers of compound (Via), as a mixture respectively of epimer (VIaa) and epimer (VIab). ¹H NMR(400 MHz, CDCI3) of the mixture of epimers (VIaa) and (VIab) : δ = 4.68

(dd, J= 10.8, J= 5.1, 2×1 H) ; 3.71 (s, 2x3H); 3.60 (t app, J= 8.2, IH); 3.42 (t app, J= 8.7, IH); 313 (dd, J= 9.2, J = 6.8, IH); 2.95 (dd, J= 9.2, J= 6.8, IH); 2.56 (dd, J= 16.6, J = 8.7, 2xlH); 2.37 (dm, 2xlH); 2.10 (m, 2xlH); 2.00 (m, 2xlH); 1.68 (m, 2xlH); 1.46 (m, 2x2H); 1.36 (m, 2x2H); 0.92 (m, 2x6H).

¹³C NMR (400 MHz, CDCl₃) of the mixture of epimers (VIaa) and (VIab) : δ =

175.9; 175.2; 171.9; 55.3; 52.4; 49.8; 49.5; 38.0; 37.8; 37.3; 36.9; 32.5; 32.2; 22.6; 22.4; 21.0; 14.4; 11.2; 11.1

HPLC (GRAD 90/10) of the mixture of epimers (VIaa) and (VIab): retention time= 9.84 minutes (100 %)

GC of the mixture of epimers (VIaa) and (VIab): retention time = 13.33 minutes (98.9 %)

MS of the mixture of epimers (VIaa) and (VIab) (ESI) : 228 MH+

3.b. Ammonolysis of compound of the mixture of (VIaa) and (VIab)

(VIaa) (VIab) (I) (VII)

A solution of (VIaa) and (VIab) obtained in previous reaction step (1.46g, 6.4 mmol) in aqueous ammonia 50 % w/w (18 mL) at 0⁰C is stirred at room temperature for 5.5hours. A white precipitate that appears during the reaction, is filtered off, is washed with water and is dried to give 0.77g (3.6 mmol, yield = 56 %) of white solid which is a mixture of brivaracetam (I) and of compound (VII) in a 1 :1 ratio.

¹H NMR of the mixture (I) and (VII) (400 MHz, CDCI3) : δ = 6.36 (s, broad, IH); 5.66 (s, broad, IH); 4.45 (m, IH); 3.53 (ddd, J= 28.8, J= 9.7, J= 8.1, IH); 3.02 (m, IH); 2.55 (m, IH); 2.35 (m, IH); 2.11 (m, IH); 1.96 (m, IH); 1.68 (m, IH); 1.38 (dm, 4H); 0.92 (m, 6H). 13c NMR of the mixture (I) and (VII) (400 MHz, CDCl₃) : δ = 176.0; 175.9; 172.8;

172.5; 56.4; 56.3; 50.0; 49.9; 38.3; 38.1; 37.3; 37.0; 32.3; 32.2; 21.4; 21.3; 21.0; 20.9; 14.4; 10.9; 10.8

HPLC (GRAD 90/10) of the mixture of (I) and (VII) retention time= 7.67 minutes (100 %)

Melting point of the mixture of (I) and (VII) = 104.9⁰C (heat from 40⁰C to 120⁰C at 10°C/min)

Compounds (I) and (VII) are separated according to conventional techniques known to the skilled person in the art. A typical preparative separation is performed on a 11.7g scale of a 1 :1 mixture of compounds (I) and (VII) : DAICEL CHIRALPAK® AD 20 μm, 100*500 mm column at 30⁰C with a 300 mL/minutes debit, 50 % EtOH – 50 % Heptane. The separation affords 5.28g (45 %) of compound (VII), retention time = 14 minutes and 5.2Og (44 %) of compounds (I), retention time = 23 minutes.

¹H NMR of compound (I) (400 MHz, CDCl₃): δ = 6.17 (s, broad, IH); 5.32 (s, broad, IH); 4.43 (dd, J= 8.6, J= 7.1, IH); 3.49 (dd, J= 9.8, J= 8.1, IH); 3.01 (dd, J= 9.8, J= 7.1, IH); 2.59 (dd, J= 16.8, J= 8.7, IH); 2.34 (m, IH); 2.08 (dd, J= 16.8, J= 7.9, IH); 1.95 (m, IH); 1.70 (m, IH); 1.47-1.28 (m, 4H); 0.91 (dt, J= 7.2, J= 2.1, 6H)

HPLC (GRAD 90/10) of compound (I) : retention time = 7.78 minutes

¹H NMR of compound (VII) (400 MHz, CDCl₃): δ = 6.14 (s, broad, IH); 5.27 (s, broad, IH); 4.43 (t app, J = 8.1, IH); 3.53 (t app, J = 9.1, IH); 3.01 (t app, J = 7.8, IH); 2.53 (dd, J = 16.5, J = 8.8, IH); 2.36 (m, IH); 2.14 (dd, J = 16.5, J = 8.1, IH); 1.97 (m, IH); 1.68 (m, IH); 1.43 (m, 2H); 1.34 (m, 2H); 0.92 (m, 6H)

3c. Epimerisation of compound of (2RV2-((R)-2-oxo-4-propyl-pyπOlidin-l-ylV butyramide (VID

Compound (VII) (200 mg, 0.94 mmol) is added to a solution of sodium tert- butoxide (20 mg, 10 % w/w) in isopropanol (2 mL) at room temperature. The reaction mixture is stirred at room temperature for 18h. The solvent is evaporated to afford 200 mg

(0.94 mmol, 100 %) of a white solid. Said white solid is a mixture of brivaracetam (I) and of (VII) in a ratio 49.3 / 50.7.

HPLC (ISO80): retention time= 7.45 min (49.3%) brivaracetam (I); retention time= 8.02 minutes (50.7%) compound (VII).

Route 2

PATENT

http://www.google.com/patents/WO2007065634A1?cl=en

(scheme 3).

Scheme 3

scheme 4.

References

Brivaracetam

Names
IUPAC name (2S)-2-[(4R)-2-oxo- 4-propylpyrrolidin-1-yl] butanamide
Identifiers
CAS Number	357336-20-0
ChEMBL	ChEMBL607400
ChemSpider	8012964
Jmol interactive 3D	Image
PubChem	9837243
UNII	U863JGG2IA
InChI[show]
SMILES[show]
Properties
Chemical formula	C11H20N2O2
Molar mass	212.15 g/mol
Pharmacology
ATC code	N03AX23
Legal status	Investigational
Routes of administration	Oral
Pharmacokinetics:
Bioavailability	Nearly 100%
Protein binding	<20%
Metabolism	Hydrolysis, CYP2C8-mediated hydroxylation
Biological half-life	8 hrs
Excretion	>75% renal
Except where otherwise noted, data are given for materials in their standard state (at 25 °C [77 °F], 100 kPa).

//////BRIVARACETAM, UCB, 2016 FDA, UCB-34714

CCCC1CC(=O)N(C1)C(CC)C(=O)N

,deletion of the 6-OH in the sugar moiety of dapagliflozin led to the discovery of a more potent SGLT2 inhibitor, 6-deoxydapagliflozin (1, ). In an in vitro assay, 1 was a more active SGLT2 inhibitor, with IC ₅₀ = 0.67 nM against human SGLT2 (hSGLT2), as compared with 1.1 nM for dapagliflozin, leading to the identification of 1 as the most active SGLT2 inhibitor discovered so far in this field. Also in an in vivo assay, 1 also introduced more urinary glucose in a rat urinary glucose excretion test (UGE) and exhibited more potent blood glucose inhibitory activity in a rat oral glucose tolerance test (OGTT) than dapagliflozin.

 Tianjin Institute Of Pharmaceutical Research,天津药物研究院

SPECTRAL DATA of Tianagliflozin

1 as a white solid (3.65 g, 93 %). R _f = 0.35 (EtOAc);

m.p.: 148–149 °C;

¹H NMR (400 MHz, DMSO-d ₆): δ = 7.35 (d, 1H, J = 8.4 Hz), 7.25 (s, 1H), 7.18 (d, 1H, J = 8.0 Hz), 7.08 (d, 2H, J = 8.4 Hz), 6.81 (d, 2H, J = 8.4 Hz), 4.95 (d, 1H, J = 5.2 Hz, OH), 4.90 (d, 1H, J = 4.4 Hz, OH), 4.79 (d, 1H, J = 5.6 Hz, OH), 3.92–4.01 (m, 5H), 3.24–3.29 (m, 1H), 3.18–3.22 (m, 1H), 3.09–3.15 (m, 1H), 2.89–2.95 (m, 1H), 1.29 (t, 3H, J = 7.0 Hz, CH₂ CH ₃ ), 1.15 (d, 3H, J = 6.0 Hz, CHCH ₃ ) ppm;

¹³C NMR (100 MHz, DMSO-d ₆): δ = 156.85, 139.65, 137.82, 131.83, 131.16, 130.58, 129.52, 128.65, 127.14, 114.26, 80.71, 77.98, 75.77, 75.51, 74.81, 62.84, 37.55, 18.19, 14.62 ppm;

IR (KBr): v¯¯¯ = 3,564, 3,385 (s), 2,981 (s), 2,899 (s), 2,861 (s), 1,613 (m), 1,512 (s), 1,477 (m), 1,247 (s), 1,102 (s), 1,045 (s), 1,012 (s) cm⁻¹;

HR–MS: calcd for C₂₁H₂₉ClNO₅ ([M + NH₄]⁺) 410.1729, found 410.1724.

PATENT

 CN 103864737

http://www.google.com/patents/CN103864737A?cl=en

PATENT

WO 2014094544

http://www.google.com/patents/WO2014094544A1?cl=en

-27-

1 D1 -6 Optionally, the step (7 ‘) is the step (7’) in place:

LS l- [4 – D (I- Dl- 6)

A.

(DMSO-d _6, 400 MHz), δ 7.35 (d, 1H, J = 8.0 Hz), 7.28 (d, 1H, J ‘. 2.0 Hz), 7.17 (dd, IH, / = 2.0 Hz and 8.4 Hz), 7.05 (d, 2H, _J: 8.8 Hz), 6.79 (d, 2H, 8.8 Hz): 4.924,95 (m, 2H), 4,81 (d, IH, 6,0 Hz), 3.93- 3.99 (m, 5H), 3,85 (d, 1H, J = 10,4 Hz), 3,66 (dd, IH, 5,2 Hz and 11,6 Hz), 3.17-3,28 (m, 3H), 3.02-3.08 _(m: IH), 1.28 (t, 3H, J = 7,0 Hz), 0,80 (s, 9H), -0.05 (s, 3H), -0.09 (s, 3H) .

PATENT

CN 104045614

[0066] The added 100mL dried over anhydrous methanol 0. 5g of sodium metal, nitrogen at room temperature with stirring, until the sodium metal disappeared. Followed by addition of 5. 2g (10mmol) of compound 6, stirring was continued at room temperature for 3 hours. To the reaction system was added 5g strong acid cation exchange resin, stirred at room temperature overnight, the reaction mixture until pH = 7. The resin was removed by suction, and the filtrate evaporated to dryness on a rotary evaporator, the residue was further dried on a vacuum pump to give the product I-D1-6, as a white foamy solid.

PATENT

 WO 2014139447

PATENT related

http://www.google.com/patents/WO2013044608A1?cl=en

http://link.springer.com/article/10.1007%2Fs40242-014-4043-9#/page-1

Design of SGLT2 Inhibitors for the Treatment of Type 2 Diabetes: A History Driven by Biology to Chemistry.

http://www.ncbi.nlm.nih.gov/pubmed/25557661

Paper

Discovery of 6-Deoxydapagliflozin as a Highly Potent Sodium-dependent Glucose Cotransporter 2 (SGLT2) Inhibitor for the Treatment of Type 2 Diabetes

Authors: Zhang, Lingyu; Wang, Yuli; Xu, Huaqiang; Shi, Yongheng; Liu, Bingni; Wei, Qunchao; Xu, Weiren; Tang, Lida; Wang, Jianwu; Zhao, Guilong

Source: Medicinal Chemistry, Volume 10, Number 3, May 2014, pp. 304-317(14)

http://www.ingentaconnect.com/content/ben/mc/2014/00000010/00000003/art00009?crawler=true

CLIP

A facile synthesis of 6-deoxydapagliflozin

The synthetic route to the target compound 1 is shown in Scheme 3. The starting material methyl 2,3,4-tri-O-benzyl-6-deoxy-6-iodo-α–d-glucopyranoside (3) was prepared from commercially available methyl α–d-glucopyranoside (2) according to a known method [5, 6].

Iodide 3 was reductively deiodinated to give 4 in 91 % yield under hydrogenolytic conditions using 10 % Pd/C as catalyst in the presence of Et₃N as base in THF/MeOH at room temperature.

when the iodide 3 was treated with Barton–McCombie reagent (n-Bu₃SnH/AIBN) [7] in toluene at room temperature no reaction occurred; however, when the reaction was carried out at elevated temperatures, such as reflux, a complex mixture formed with only a trace amount (3 %, entry 1) of the desired product 4.

When the iodide 3 was treated with LiAlH₄ in THF at 0 °C to room temperature, another complex mixture was produced with only a trace amount (2 %, entry 2) of 4.

When Pd(OH)₂ was used as the hydrogenolysis catalyst instead of 10 % Pd/C, the desired 4 was indeed formed (14 %, entry 4), but most of the starting material was converted to a few more polar byproducts, which were believed to result from the cleavage of at least one of the benzyl groups.

pdf available

Monatshefte für Chemie – Chemical Monthly

December 2013, Volume 144, Issue 12, pp 1903-1910

http://download.springer.com/static/pdf/721/art%253A10.1007%252Fs00706-013-1053-0.pdf?originUrl=http%3A%2F%2Flink.springer.com%2Farticle%2F10.1007%2Fs00706-013-1053-0&token2=exp=1458808857~acl=%2Fstatic%2Fpdf%2F721%2Fart%25253A10.1007%25252Fs00706-013-1053-0.pdf%3ForiginUrl%3Dhttp%253A%252F%252Flink.springer.com%252Farticle%252F10.1007%252Fs00706-013-1053-0*~hmac=bd1c3c2bdc3712f5540267c99f732b2f7588020a868aa23021792a2a2a58d65e

////////IND Filing, SGLT-2 inhibitor, type 2 diabetes, Tianagliflozin, taigeliejing, 6-deoxydapagliflozin, 1461750-27-5

Clc1c(cc(cc1)C2[C@@H]([C@H]([C@@H]([C@H](O2)C)O)O)O)Cc3ccc(cc3)OCC

TRIM24/BRPF1 bromodomain inhibitor

IACS-9571; IACS 9571; IACS9571.

N-[6-[3-[4-(dimethylamino)butoxy]-5-propoxyphenoxy]-1,3-dimethyl-2-oxobenzimidazol-5-yl]-3,4-dimethoxybenzenesulfonamide

BOARD OF REGENTS, UNIVERSITY OF TEXAS SYSTEM

IACS-9571 is a potent and selective inhibitor TRIM24 and BRPF1. The bromodomain containing proteins TRIM24 (Tripartite motif containing protein 24) and BRPF1 (bromodomain and PHD finger containing protein 1) are involved in the epigenetic regulation of gene expression and have been implicated in human cancer. Overexpression of TRIM24 correlates with poor patient prognosis and BRPF1 is a scaffolding protein required for the assembly of histone acetyltransferase complexes, where the gene of MOZ (monocytic leukemia zinc finger protein) was first identified as a recurrent fusion partner in leukemia patients (8p11 chromosomal rearrangements). IACS-9571 has low nanomolar affinities for TRIM24 and BRPF1 (ITC Kd = 31 nM and 14 nM, respectively). With its excellent cellular potency (EC50 = 50 nM) and favorable pharmacokinetic properties (F = 29%), IACS-9571 is a high-quality chemical probe for the evaluation of TRIM24 and/or BRPF1 bromodomain function in vitro and in vivo. (J Med Chem. 2015 Jun 10. [Epub ahead of print] )

PAPER

http://pubs.acs.org/doi/abs/10.1021/acs.jmedchem.5b00405

Structure-Guided Design of IACS-9571, a Selective High-Affinity Dual TRIM24-BRPF1 Bromodomain Inhibitor

The bromodomain containing proteins TRIM24 (tripartite motif containing protein 24) and BRPF1 (bromodomain and PHD finger containing protein 1) are involved in the epigenetic regulation of gene expression and have been implicated in human cancer. Overexpression of TRIM24 correlates with poor patient prognosis, and BRPF1 is a scaffolding protein required for the assembly of histone acetyltransferase complexes, where the gene of MOZ (monocytic leukemia zinc finger protein) was first identified as a recurrent fusion partner in leukemia patients (8p11 chromosomal rearrangements). Here, we present the structure guided development of a series of N,N-dimethylbenzimidazolone bromodomain inhibitors through the iterative use of X-ray cocrystal structures. A unique binding mode enabled the design of a potent and selective inhibitor 8i (IACS-9571) with low nanomolar affinities for TRIM24 and BRPF1 (ITC K_d = 31 nM and ITC K_d = 14 nM, respectively). With its excellent cellular potency (EC₅₀ = 50 nM) and favorable pharmacokinetic properties (F = 29%), 8i is a high-quality chemical probe for the evaluation of TRIM24 and/or BRPF1 bromodomain function in vitro and in vivo.

TFA salt of 8i (106 mg, 57%) as a white solid. 1H NMR (600 MHz, DMSO-d6) δ 9.46 (s, 1H), 9.30 (br-s, 1H), 7.19 (m, 2H), 7.07 (s, 1H), 6.90 (d, J = 9.0 Hz, 1H), 6.75 (s, 1H), 6.13 (t, J = 2.2 Hz, 1H), 5.71 (t, J = 2.0 Hz, 1H), 5.67 (t, J = 2.0 Hz, 1H), 3.84 (t, J = 5.9 Hz, 2H), 3.77 (m, 5H), 3.62 (s, 3H), 3.29 (s, 3H), 3.20 (s, 3H), 3.12–3.05 (m, 2H), 2.78 (d, J = 4.7 Hz, 6H), 1.77–1.63 (m, 6H), 0.95 (t, J = 7.3 Hz, 3H). 13C NMR (600 MHz, DMSO-d6) δ 160.3, 160.0, 159.3, 154.1, 152.0, 148.4, 143.9, 131.8, 128.2, 126.0, 121.9, 120.5, 110.4, 109.4, 106.4, 100.6, 95.9, 95.8, 95.2, 68.9, 66.7, 56.3, 55.6, 55.4, 42.1, 27.1, 27.0, 25.6, 21.9, 20.7, 10.4. MS (ESI) m/z 644 [M + H]+.

NMR

IACS -9571

CLICK ON IMAGE

ABSTRACT

251st ACS National Meeting & Exposition

MEDI 5

Discovery and development of a potent dual TRIM24/BRPF1 bromodomain inhibitor, IACS -9571, using structure- based drug design Wylie S. Palmer 1 , wpalmer@mdanderson.org, Guillaume Poncet -Montagne 1 , Gang Liu 1 , Alessia Petrocchi 1 , N aphtali Reyna 1 , Govindan Subramanian 1 , Jay Theroff 1 , Maria Kost -Alimova 1 , Jennifer Bardenhagen 1 , Elisabetta Leo 1 , Hannah Sheppard 1 , Trang Tieu 1 , Shi Xi 1 , Yanai Zhan 1 , Shuping Zhao 1 , Michelle Barton 2 , Giulio Draetta 1 , Carlo Toniatti 1 , Philip Jones 1 , Mary Ge ck Do 1 , Jannik Andersen 1 . (1) Institute for Applied Cancer Science, The University of Texas, MD Anderson Cancer Center, Houston, Texas, United States (2) Department of Epigenetics and Molecular Carcinogenesis, The University of Texas, MD Anderson Cancer Center, Houston, Texas, United States

Bromodomains are an important class of chromatin remodeling proteins that recognize acetylated lysine residues on histone tails. As epigenetic targets they regulate gene transcription and offer a new way to treat diseas es, particularly in inflammation and oncology. The bromodomain and extra- terminal (BET) family has emerged as an important and druggable example of this class of proteins with the successful entry of small- molecule inhibitors into the clinic. Other families of bromodomains are only starting to be explored, such as the Tripartite Motif -containing 24 protein (TRIM24) and bromodomain- PHD finger protein 1 (BRPF1). Both proteins contain a dual PHD -bromo motif which have a role in recognizing specific histone mar ks. TRIM24 recognizes the dual histone marks of unmodified H3K4 and acetylated- H3K23 within the same histone tail. TRIM24 is a potent co- activator of ER -alpha and overexpression of TRIM24 has been linked to poor survival rates in breast cancer patients.

This presentation will describe the structure guided development of a series of N,N- dimethyl -benzimidazolones through the iterative use of X -ray cocrystal structures. A unique binding mode enabled the design of a potent and selective inhibitor (IACS -9571) with low nanomolar affinities for TRIM24 and BRPF1 (ITC Kd = 31 nM and ITC Kd = 14 nM, respectively). With its excellent cellular potency (EC 50 = 50 nM) and favorable pharmacokinetic properties, IACS -9571 is a high- quality chemical probe for the evaluation of TRIM24 and/or BRPF1 bromodomain function in vitro and in vivo

PATENT

WO-2016033416-A1

Synthesis of Intermediates:

N-(6-bromo-l ,3-dimethyl-2-oxo-2,3-dihydro-lH-benzo[d]imidazol-5-yl)-2,2,2- trifluoroacetamide (Intermediate 1):

Step 1 : 5-nitro-lH-benzo[d]imidazol-2(3H)-one:

To a 0 °C solution of 4-nitrobenzene- 1 ,2-diamine (44 g, 285 mmol) in 80 mL of DMF was added l, l’-carbonyldiimidazole (70 g, 428 mmol). The reaction mixture was stirred at RT for 4 h, then water (250 mL) was added. The resulting suspension was filtered, and the collected solids were washed with water (200 mL) and dried to give 5-nitro-lH- benzo[d]imidazol-2(3H)-one as a yellow solid (45 g, 88%). MS (ES+) C7H5N3O3 requires: 179, found: 180 [M+H]⁺.

Step 2: l,3-dimethyl-5-nitro-lH-benzo[d]imidazol-2(3H)-one:

To a solution of 5-nitro-lH-benzo[d]imidazol-2(3H)-one (55 g, 309 mmol) in 150 mL of DMF was added K2CO3 (85 g, 618 mmol), the reaction mixture was cooled to 0 °C, then iodomethane (109 g, 772 mmol) was slowly added. The reaction mixture was stirred at RT overnight, then water was added to the reaction mixture. The resulting suspension was filtered and the collected solids were washed with water (200 mL) and dried to give 1,3- dimethyl-5-nitro-lH-benzo[d]imidazol-2(3H)-one as a yellow solid (55 g, 86%). MS (ES+) C9H9N3O3 requires: 207, found: 208 [M+H] ⁺.

Step 3: 5-amino-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one:

 To a solution of l,3-dimethyl-5-nitro-lH-benzo[d]imidazol-2(3H)-one (50 g, 240 mmol) in 200 mL of EtOAc under an inert atmosphere was added 10% palladium on activated carbon (5 g, 24 mmol). The reaction mixture was then charged with hydrogen and stirred at RT under an ¾ atmosphere overnight. The reaction mixture was filtered through a pad of celite then concentrated to give 5-amino-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)- one as a yellow solid (32 g, 68%). MS (ES+) C9H11N3O requires: 177, found: 178 [M+H]⁺.

Step 4: 5-amino-6-bromo-l ,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one:

 To a 0 °C solution of 5-amino-l ,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one (4 g, 22.6 mmol) in 25 mL of CHCI3 and 25 mL of AcOH was slowly added drop wise bromine (3.5 g, 22.6mmol). The mixture was stirred at RT for 30 min, then concentrated and purified by silica gel chromatography (1 : 1 EtOAc/ hexanes) to afford 5-amino-6-bromo-l ,3-dimethyl- lH-benzo[d]imidazol-2(3H)-one as a yellow solid (3.2 g, 69%). MS (ES+) C₉HioBrN₃0 requires: 256, found: 257 [M+H]⁺.

Step 5: N-(6-bromo-l ,3-dimethyl-2-oxo-2,3-dihydro-lH-benzo[d]imidazol-5-yl)-2,2,2- trifluoroacetamide:

To a 0 °C solution of 5-amino-6-bromo-l ,3-dimethyl-lH-benzo[d]imidazol- 2(3H)-one (1.50 g, 5.9 mmol) in DCM (45 ml) was added DMAP (72 mg, 0.59 mmol), triethylamine (1.63 ml, 11.7 mmol) and trifluoroacetic anhydride (0.91 ml, 6.4 mmol). The reaction mixture was stirred for 2 h and warmed to RT. The reaction mixture was then quenched with water and the organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give N-(6-bromo-l,3-dimethyl-2-oxo-2,3-dihydro-lH- benzo[d]imidazol-5-yl)-2,2,2-trifluoroacetamide (Intermediate 1) as a yellow solid (2.20 g, 100%). MS (ES+) CiiH₉BrF₃N302 requires: 352, found 353 [M+H]⁺.

5-amino-6-(3-hydroxyphenoxy)-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one (Intermediate 2, Route A):

To a mixture of 5-amino-6-(3-(benzyloxy)phenoxy)-l,3-dimethyl-lH- benzo[d]imidazol-2(3H)-one (400 mg, 1.07 mmol) in DCM (20 mL) at -78 °C was added tribromoborane (5.3 mL, 5.3 mmol). The mixture was warmed up to room temperature gradually, then quenched by methanol dropwise, concentrated, and purified by column chromatography (20-100% EtOAc/hexanes and then 0-40% methanol/EtOAc) to give 5- amino-6-(3-hydroxyphenoxy)-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one as a solid (240 mg, 79%). MS (ES+) C15H15N3O3 requires: 285, found: 286 [M+H]⁺.

5-amino-6-(3-hydroxyphenoxy)-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one (Intermediate 2, Route B):

Step 2

Step 1: 3-[(ieri-butyldimethylsilyl)oxy]phenol:

A mixture of lH-imidazole (2.25 g, 33.1 mmol), ieri-butylchlorodimethylsilane (3.83 g, 25.4 mmol) and resorcinol (5.6 g, 51 mmol) in THF (30 ml) was stirred at 80 °C for 5 h. The resulting suspension of the cooled reaction mixture was filtered and the collected filtrate was concentrated and purified by silica-gel chromatography (20:80 to 0:100, EtOAc/hexanes) to give 3-((ieri-butyldimethylsilyl)oxy)phenol (2.78 g, 49%). MS (ES+) C12H20O2S1 requires: 224, found 225 [M+H]⁺.

Step 2: 5-amino-6-(3-((ier^butyldimethylsilyl)oxy)phenoxy)-l ,3-dimethyl-lH- benzo[d]imidazol-2(3H)-one:

 A mixture of 3-((ieri-butyldimethylsilyl)oxy)phenol (1.39 g, 6.20 mmol), quinolin-8-ol (79 mg, 0.55 mmol), copper(I) chloride (20 mg, 0.21 mmol), potassium phosphate (526 mg, 2.48 mmol) and 5-amino-6-bromo-l ,3-dimethyl-lH-benzo[d]imidazol- 2(3H)-one (529 mg, 2.07 mmol) in diglyme (20 ml) in a 100 mL round-bottom flask was degassed under a nitrogen atmosphere and heated to 120 °C for 24 h. To the cooled reaction mixture was added silica gel, stirred for 2 min, then the mixture was filtered through a pad of silica gel. The collected filtrate was concentrated and purified by column chromatography (20:80 to 0: 100, EtOAc/hexanesthen 0: 100 to 40:60, MeOH/EtOAc) to give 5-amino-6-(3- ((ieri-butyldimethylsilyl)oxy)phenoxy)-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one (521 mg, 63%). MS (ES+) C21H29N3O3S1 requires: 399, found 400 [M+H]⁺.

Step 3: 5-amino-6-(3-hydroxyphenoxy)-l,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one:

To a 0 °C solution of 5-amino-6-(3-((ieri-butyldimethylsilyl)oxy)phenoxy)-l,3- dimethyl-lH-benzo[d]imidazol-2(3H)-one (623 mg, 1.56 mmol) in THF was added a solution of ieira-butylammonium fluoride (0.90 mL, 3.1 mmol) in THF, the reaction mixture was allowed to warm up to RT and then stirred for 1-2 h. The reaction mixture was quenched with 1 M hydrogen chloride (0.10 mL, 3.1 mmol) and then partitioned between EtOAc and water. The seperated organic layer was washed with water twice, then concentrated and purified by column chromatography (20-80% EtOAc/hexanes and 0-40% MeOH/DCM) to give 5-amino-6-(3-hydroxyphenoxy)-l ,3-dimethyl-lH-benzo[d]imidazol-2(3H)-one (120 mg, 27%) as a solid. MS (ES+) C15H15N3O3 requires: 285, found 286 [M+H]⁺.

EXAMPLE 10: N-(6-(3-(4-(dimethylamino)butoxy)-5-propoxyphenoxy)-l,3-dimethyl-2- oxo-2,3-dihydro-lH-benzo[d]imidazol-5-yl)-3,4-dimethoxybenzenesulfonamide 2,2,2-

To a solution of N-(6-(3-(4-aminobutoxy)-5-propoxyphenoxy)-l ,3-dimethyl-2- oxo-2,3-dihydro-lH-benzo[d]imidazol-5-yl)-3,4-dimethoxybenzenesulfonamide 2,2,2- trifluoroacetate (180 mg, 0.247 mmol) in methanol (3.0 ml) was added triethylamine (0.034 ml, 0.25 mmol), acetic acid (0.028 ml, 0.49 mmol), formaldehyde (0.054 ml, 2.0 mmol), and sodium triacetoxyborohydride (131 mg, 0.618 mmol). The reaction mixture was stirred at room temperature and checked by LCMS every 30 minutes. After 3 h the reaction was complete by LCMS. The reaction was quenched with a few drops of TFA and concentrated under reduced pressure. The residue was purified by prep-HPLC using a gradient of 20-60% ACN/water containing 0.1% TFA to afford N-(6-(3-(4-(dimethylamino)butoxy)-5- propoxyphenoxy)-l,3-dimethyl-2-oxo-2,3-dihydro-lH-benzo[d]imidazol-5-yl)-3,4- dimethoxybenzenesulfonamide 2,2,2-trifluoroacetate (106 mg, 57%) as a white solid. MS (ES+) C32H42N4O8S requires: 642, found 643 [M+H]⁺. ¾ NMR (600 MHz, DMSO-ifc) δ 9.46 (s, 1H), 9.30 (br-s, 1H), 7.19 (m, 2H), 7.07 (s, 1H), 6.90 (d, 7 = 9.0 Hz, 1H), 6.75 (s, 1H), 6.13 (t, 7 = 2.2 Hz, 1H), 5.71 (t, J = 2.0 Hz, 1H), 5.67 (t, J = 2.0 Hz, 1H), 3.84 (t, 7 = 5.9 Hz, 2H), 3.77 (m, 5H), 3.62 (s, 3H), 3.29 (s, 3H), 3.20 (s, 3H), 3.12-3.05 (m, 2H), 2.78 (d, 7 = 4.7 Hz, 6H), 1.77-1.63 (m, 6H), 0.95 (t, 7 = 7.3 Hz, 3H)

References

1: Palmer WS, Poncet-Montange G, Liu G, Petrocchi A, Reyna N, Subramanian G, Theroff J, Yau A, Kost-Alimova M, Bardenhagen JP, Leo E, Shepard HE, Tieu TN, Shi X, Zhan Y, Zhao S, Draetta G, Toniatti C, Jones P, Geck Do M, Andersen JN. Structure-Guided Design of IACS-9571, a Selective High-Affinity Dual TRIM24-BRPF1 Bromodomain Inhibitor. J Med Chem. 2015 Jun 10. [Epub ahead of print] PubMed PMID: 26061247.

US-20160060260-A1

Institute for Applied Cancer Science, The University of Texas, MD Anderson Cancer Center, Houston, Texas, United States

The University of Texas MD Anderson Cancer Center | University of Texas System

The new Institute for Applied Cancer Science will be located at the south campus of M.D.

Draetta arrived at MD Anderson in 2011 to direct the Institute for Applied Cancer Science. He oversees the moon shots platforms

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas, MD Anderson Cancer Center, Houston, Texas, United States

///////IACS-9571, TRIM24, BRPF1 bromodomain inhibitor, IACS-9571,  IACS 9571,  IACS9571, BOARD OF REGENTS, UNIVERSITY OF TEXAS SYSTEM
CAS BASE 1800477-30-8
CAS OF 1:1 TRIFLUOROACETATE 1883598-69-3

c1(cc(cc(c1)OCCC)Oc3cc2N(C(N(c2cc3NS(=O)(=O)c4cc(c(cc4)OC)OC)C)=O)C)OCCCCN(C)C

CCCOC1=CC(=CC(=C1)OC2=C(C=C3C(=C2)N(C(=O)N3C)C)NS(=O)(=O)C4=CC(=C(C=C4)OC)OC)OCCCCN(C)C

TFA salt of 8i (106 mg, 57%) as a white solid. 1H NMR (600 MHz, DMSO-d6) δ 9.46 (s, 1H), 9.30 (br-s, 1H), 7.19 (m, 2H), 7.07 (s, 1H), 6.90 (d, J = 9.0 Hz, 1H), 6.75 (s, 1H), 6.13 (t, J = 2.2 Hz, 1H), 5.71 (t, J = 2.0 Hz, 1H), 5.67 (t, J = 2.0 Hz, 1H), 3.84 (t, J = 5.9 Hz, 2H), 3.77 (m, 5H), 3.62 (s, 3H), 3.29 (s, 3H), 3.20 (s, 3H), 3.12–3.05 (m, 2H), 2.78 (d, J = 4.7 Hz, 6H), 1.77–1.63 (m, 6H), 0.95 (t, J = 7.3 Hz, 3H). 13C NMR (600 MHz, DMSO-d6) δ 160.3, 160.0, 159.3, 154.1, 152.0, 148.4, 143.9, 131.8, 128.2, 126.0, 121.9, 120.5, 110.4, 109.4, 106.4, 100.6, 95.9, 95.8, 95.2, 68.9, 66.7, 56.3, 55.6, 55.4, 42.1, 27.1, 27.0, 25.6, 21.9, 20.7, 10.4. MS (ESI) m/z 644 [M + H]+.

RP 6503

phase 1

RP 6503

Mass: 619.1 (M⁺+l). MP: 175-178° C Specific optical rotation (C=l in chloroform, at 25°C) : [a]_D = + 147.16.

A1

RP 6503

(S)-N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl) ethyl)-lH-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl)methanesulfonamide

(S)-N-[5-[4-amino-1-[1-[5-fluoro-3-(3-fluorophenyl)-4-oxochromen-2-yl]ethyl]pyrazolo[3,4-d]pyrimidin-3-yl]-2-methoxyphenyl]methanesulfonamide

Novartis to develop and commercialize Rhizen’s inhaled dual PI3K-delta gamma inhibitor and related compounds worldwide

The immune pipeline includes ‘dual PI3K inhibitors for various indications’ licensed to Novartis

‘inhaled dual inhibitor’,

Phosphoinositide-3 kinase delta inhibitor; Phosphoinositide-3 kinase gamma inhibitor

WO2011055215A2 and WO2012151525A1 and U.S. Publication Nos. US20110118257 and US20120289496

Rhizen Pharmaceuticals Sa   INNOVATOR

PATENT

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

PATENT

http://www.google.com/patents/WO2012151525A1?cl=en

scheme 1A:

Ste -1

Step-2

Scheme 2

SCHEME 3

SCHEME4

List of Intermediates

Intermediate 27: 2-( l -(4-amino-3-iodo-lH-pyrazolo[3,4-d]pyrimidin- l – yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one: To a solution of 3-iodo- l H- pyrazolo[3,4-d]pyrimidin-4-amine (0.800 g, 2.88 mmol) in DMF (5 ml), potassium carbonate (0.398 g, 2.88 mmol) was added and stirred at RT for 30 min. To this mixture intermediate 22 (0.500 g, 1.44 mmol) was added and stirred for 12h. The reaction mixture was diluted with water and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with methanol: dichloromethane to afford the title compound as a off-white solid (0.300 g, 38%). Ή-NMR (5 ppm, DMSO-d₆₃, 400 MHz): 8.02 (s, 1 H), 7.94 (s, 1 H), 7.84 (dt, J = 8.4,5.7 Hz, 1H), 7.47 (d, 7 = 8.6 Hz, 1H), 7.29 (m, 3H), 7.09 (dt, 7 = 8.8,2.3 Hz, 1 H), 6.87 (s, 2H), 5.88 (q, 7 = 7.0 Hz, 1H), 1.82 (d, 7 = 7.0 Hz, 3H).

SYNTHESIS

MAIN PART

PATENT

http://www.google.com/patents/WO2015198289A1?cl=en

Prashant Kashinath Bhavar, Swaroop Kumar Venkata Satya VAKKALANKA

The present invention relates to a selective dual delta (δ) and gamma (γ) PI3K protein kinase modulator (S)-N-(5-(4-amino-1-(1-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H- chromen-2-yl)ethyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl) methane sulfonamide, methods of preparing them, pharmaceutical compositions containing them and methods of treatment, prevention and/or amelioration of PI3K kinase mediated diseases or disorders with them.

compound of formula (Al):

(Al).

The process comprises the steps of:

(a) subjecting (R)-5-fluoro-3-(3-fluorophenyl)-2-(l-hydroxyethyl)-4H-chromen-4-one:

to a Mitsunobu reaction with 3-(4-methoxy-3-nitrophenyl)-lH-pyrazolo[3,4-d]pyrimidin-4-amine:

(for example, in the presence of triphenylphosphine and diisopropylazodicarboxylate) to give (S)-2-(l-(4-amino-3-(4-methoxy-3-nitrophenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (Intermediate 3):

Intermediate 3;

(b) reducing Intermediate 3, for example with a reducing agent such as Raney Ni, to give (S)-2-(l-(4-amino-3-(3-amino-4-methoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin- l-yl)ethyl)-5-fluoro-3-( -fluorophenyl)-4H-chromen-4-one (Intermediate 4):

Intermediate 4;

The intermediates described herein may be prepared by the methods described in International Publication Nos. WO 11/055215 and WO 12/151525, both of which are hereby incorporated by reference.

Intermediate 1: N-(5-bromo-2-methoxyphenyl)methanesulfonamide:

To a solution of 5-bromo-2-methoxyaniline(1.00 g, 4.94 mmol) in dichloromethane (10 ml), pyridine (0.800 ml, 9.89 mmol) was added and cooled to 0°C. Methane sulphonyl chloride (0.40 ml, 5.19 mmol) was added and stirred for 30 min. The reaction mixture was quenched with water, extracted with ethyl acetate, dried over anhydrous sodium sulphate and concentrated under reduced pressure. The crude product was chromatographed with ethyl acetate : petroleum ether to afford the title compound as a reddish solid (1.20 g, 87%).

Intermediate 2: N-(2-methoxy-5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)phenyl)methanesulfonamide: Potassium acetate (0.841 g, 8.57 mmol) and bis(pinacolato)diboron (1.190 g, 4.71 mmol) were added to a solution of intermediate 1 (1.20 g, 4.28 mmol) in dioxane (17.5 ml) and the solution was degassed for 30 min.[l, -Bis(diphenylphosphino)ferrocene]dichloro palladium(II).CH2Ci2 (0.104 g, 0.128 mmol) was added under nitrogen atmosphere and heated to 80°C. After 2h the

reaction mixture was filtered through celite and concentrated. The crude product was purified by column chromatography with ethyl acetate : petroleum ether to afford the title compound as a yellow solid (1.00 g, 71%).^JH-NMR (δ ppm, CDCb, 400 MHz): 7. 91 (d, / = 1.2Hz, 1H), 7. 62 (dd, / = 8.1, 1.2Hz, 1H), 6. 92 (d, / = 8.1Hz, 1H), 6.73 (s, 1H), 3.91 (s, 3H), 2.98 (s, 3H), 1.32 (s, 12H).

Intermediate 3: (S)-2-(l-(4-amino-3-(4-methoxy-3-nitrophenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one: (S)-2-(l-(4-amino-3-(4-methoxy-3-nitrophenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one: To a solution of (R)-5-fluoro-3-(3-fluorophenyl)-2-(l-hydroxyethyl)-4H-chromen-4-one (0.500 g, 1.64 mmol) in THF (5 ml), 3-(4-methoxy-3-nitrophenyl)-lH-pyrazolo[3,4-d]pyrimidin-4-amine (0.564 g, 1.97 mmol) and triphenylphosphine (0.649 g, 2.47 mmol) were added followed by the addition of diisopropylazodicarboxylate (0.50 ml, 2.47 mmol). ((R)-5-fluoro-3-(3-fluorophenyl)-2-(l-hydroxyethyl)-4H-chromen-4-one can be prepared as described for Intermediates 23, 25, and 26 in International Publication No. WO 2012/0151525.). After 4h at room temperature, the mixture was concentrated and the residue was purified by column chromatography with ethyl acetate : petroleum ether to afford the title compound as a brown solid (0.270 g, 29%). ^JH-NMR (δ ppm, DMSO-d6, 400 MHz): 8.04 (s, 1H), 7.83 (m, 1H), 7.63-7.50 (m, 3H), 7.29 (m, 2H), 7.06 (dt, J = 8.7,2.2Hz, 1H), 6.94 (m, 2H), 6.75 (dd, J = 8.1,2.1Hz, 1H), 5.95 (q, J = 7.0Hz, 1H), 4.98 (s, 2H), 3.81 (s, 3H), 1.86 (d, J = 7.0 Hz, 3H).

[109] Intermediate 4: (S)-2-(l-(4-amino-3-(3-amino-4-methoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one:

(S)-2-(l-(4-amino-3-(3-amino-4-methoxyphenyl)-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one : To a solution of Intermediate 3 (0.260 g, 0.455 mmol) in ethanol (5 ml), Raney Ni (0.130 g) was added and hydrogeneated at 20psi at 50°C for 24h. The reaction mixture was passed through celitepad and concentrated to afford the title compound as a brown solid (0.150 g, 60%). Mass : 540.8 (M⁺).

Example A

N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl)-lH- pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl)methanesulfonamide

To a solution of 2-(l-(4-amino-3-iodo-lH-pyrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one (0.200 g, 0.366 mmol) in DME (2.1 ml) and water (0.67 ml), intermediate 2 (0.179 g, 0.550 mmol) and sodium carbonate (0.116 g, 1.10 mmol) were added and the system was degassed for 30 min. (2-(l-(4-amino-3-iodo-lH^yrazolo[3,4-d]pyrimidin-l-yl)ethyl)-5-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one can be prepared as described for Intermediates 23, 25, and 26 in International Publication No. WO 2012/0151525). Bis(diphenylphosphino) ferrocene]dichloropalladium(II) (0.059 g, 0.075 mmol) was added and kept under microwave irradiation (microwave power = 100W, temperature = 100 °C) for 45 min. The reaction mixture was Celite filtered, concentrated and extracted with ethyl acetate. The organic layer was dried over sodium sulphate and concentrated under reduced pressure. The crude product was purified by column chromatography with methanol: dichloromethane to afford the title compound as a brown solid (0.080 g, 35%). MP: 216-218 °C. ¾-NMR (δ ppm, CDCb, 400 MHz): 8.20 (s, 1H), 7.73 (s, 1H), 7.53 (m, 2H), 7.31 (m, 2H), 7.07-6.73 (m, 6H), 6.07 (q, / = 6.2 Hz, 1H), 3.98 (s, 3H), 3.14 (s, 3H), 2.01 (d, / = 6.0Hz, 3H).

Example Al and A2

Method A

(S)-N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl)- lH-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl)methanesulfonamide

and (R)-N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2- yl)ethyl)-lH-p anesulfonamide

The two enantiomerically pure isomers were separated by preparative SFC (supercritical fluid) conditions from N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl)-lH-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl)methanesulfonamide (0.500 g) on a CHIRALPAK AS-H column (250 x 30 mm; 5μπι) using methanol : CO2 (55:45) as the mobile phase at a flow rate of 80g / min.

Example Al (S-isomer): Brown solid (0.247 g). Enantiomeric excess: 97.4%. Retention time: 2.14 min. Mass: 619.1 (M⁺+l). MP: 156-158° C.

Example A2 (R-isomer): Brown solid (0.182 g). Enantiomeric excess: 99.3%. Retention t: 3.43 min. Mass: 619.1 (M⁺+l). MP: 168-171° C.

Method Al

(S)-N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl)- lH-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl)methanesulfonamide

and (R)-N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2- yl)ethyl)-lH-p anesulfonamide

The two enantiomerically pure isomers were separated by preparative SFC (supercritical fluid) conditions from N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl)-lH-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl) methanesulfonamide (15.0 g) on a CHIRALPAK AS-H column (250 x 20 mm; 5μπι) using methanol : CO2 (45:55) as the mobile phase at a flow rate of 120g / min.

Example Al (S-isomer): Enantiomeric excess: 100 %. Retention time: 2.21 min. Mass: 619.1 (M⁺+l). MP: 175-178° C Specific optical rotation (C=l in chloroform, at 25°C) : [a]_D = + 147.16.

Example A2 (R-isomer): Enantiomeric excess: 99.3%. Retention t: 3.72 min. Mass: 619.1 (M⁺+l). MP: 154-157° C. Specific optical rotation (C=l in chloroform, at 25°C) : [a]_D = – 159.54.

Method B

Example Al

(S)-N-(5-(4-amino-l-(l-(5-fluoro-3-(3-fluorophenyl)-4-oxo-4H-chromen-2-yl)ethyl)- lH-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methoxyphenyl)methanesulfonamide

To a solution of Intermediate 4 (0.500 g, 0.923 mmol) in dichloromethane (5 ml) cooled to 0°C, pyridine (0.200 ml, 1.84 mmol) was added and stirred for 10 min. Methanesulphonyl chloride (0.100 ml, 0.923 mmol) was added stirred for 30 min. The reaction mixture was quenched with water, extracted with dichloromethane and dried over sodium sulphate. The crude product was column chromatographed with methanol : dichloromethane to afford the title compound as an off-white solid (0.240 g, 42%). MP: 211-213°C. ¾-NMR (δ ppm, DMSO-d6, 400 MHz): 9.15 (s, 1H), 8.06 (s, 1H), 7.83 (m, 1H), 7.49 (m, 4H), 7.28 (m, 4H), 7.08 (dt, / = 8.6, 1.7 Hz, 1H), 6.92 (s, 2H), 5.98 (q, / = 6.9 Hz, 1H), 3.88 (s, 3H), 2.99 (s, 3H), 1.88 (d, / = 7.0 Hz, 3H). Enantiomeric excess: 85.4% as determined by HPLC on a chiralpak AS-3R column, enriched in the fast eluting isomer (retention time = 7.46 min.).

CLIPS

La Chaux-de-Fonds, Switzerland, Sept. 6, 2013  — La Chaux-de-Fonds, Switzerland (6 September 2013): Rhizen Pharmaceuticals S.A. announces a scientific poster presentation on the pre-clinical characterization of its lead calcium release activated channel (CRAC) inhibitor, RP3128, for the treatment of respiratory disorders and an oral presentation on the pharmacological profile of its novel, dual Phosphoinositide-3 kinase (PI3K) delta/gamma inhibitor, RP6503, in the pulmonary disease systems, at the European Respiratory Society Annual Congress (ERS), to be held from 7-11 September 2013, at Barcelona, Spain.

RP6503 is a novel, potent and selective inhibitor of the delta and gamma isoforms of PI3K. It is to be delivered via the inhalation route and has a long duration of action along with excellent PI3K isoform selectivity, which is expected to result in better safety. RP3128 has been optimized with high potency for CRAC channel inhibition, selectivity over the other voltage gated channels and excellent oral bioavailability. Rhizen intends to move both these compounds to the clinic in 2014.

Details of the presentations:

1.      Abstract of the Poster Presentation: “Pre-clinical characterization of RP3128, a novel and potent CRAC channel inhibitor for the treatment of respiratory disorders”

Time and Location- 8 September 2013 between 14.45-16.45 in Room 3.6, at Poster Discussion: New drugs in respiratory medicine, at FIRA BARCELONA, Convention Centre de Gran Via, Barcelona, Spain

2.      Abstract of Oral Presentation: “In vitro and in vivo pharmacological profile of RP6503, a novel dual PI3K delta/gamma inhibitor, in pulmonary disease systems”

Time and Location- 11 September 2013 at 8.45 in Room 3.9; Session 8.30-10.30, at the Oral Presentation: Emerging new targets for the treatment of respiratory diseases, at FIRA BARCELONA, Convention Centre de Gran Via, Barcelona, Spain

CLIPS

La Chaux-de-Fonds, Switzerland , Dec. 09, 2015  — Rhizen Pharmaceuticals S.A. announced today that they have entered into an exclusive, worldwide license agreement with Novartis for the development and commercialization of Rhizen’s, inhaled dual PI3K-delta gamma inhibitor and its closely related compounds for various indications.

Under the terms of the agreement, Rhizen will receive an upfront payment and is eligible to receive development, regulatory and sales milestones payments. In addition Rhizen is also eligible to receive tiered royalties on annual nets sales.

The lead compound is a novel, potent, and selective dual PI3K-delta gamma inhibitor with demonstrated anti-inflammatory and immuno-modulatory activity in pre-clinical systems and models representative of respiratory diseases. With a favorable ADME and PK profile and high therapeutic index in animals, the inhaled dual PI3K-delta gamma inhibitor holds promise in the treatment of human airway disorders.

About Rhizen Pharmaceuticals S.A.:

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

SEE

https://newdrugapprovals.org/2015/12/10/alembic-pharma-advances-1-on-rhizen-novartis-license-agreement/

WO-2015181728 

WO-2015001491 

WO-2014072937 

WO-2014006572 

http://www.atsjournals.org/doi/abs/10.1164/ajrccm-conference.2013.187.1_MeetingAbstracts.A3880

///////RP 6503, Novartis, develop, commercialize,  Rhizen, inhaled dual PI3K-delta gamma inhibitor, PHASE 1, RP-6503

c21c(cccc1O/C(=C(\C2=O)c3cc(ccc3)F)C(C)n4c6ncnc(c6c(n4)c5cc(c(cc5)OC)NS(=O)(=O)C)N)F

CC(C1=C(C(=O)C2=C(O1)C=CC=C2F)C3=CC(=CC=C3)F)N4C5=C(C(=N4)C6=CC(=C(C=C6)OC)NS(=O)(=O)C)C(=NC=N5)N

/////

2 (R)-3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

(3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide)

cas 866772-52-3

Novartis Ag

NVP-LBX192

LBX-192

54 Discovery and Evaluation of NVP-LBX192, a Liver Targeted Glucokinase Activator

https://acs.confex.com/acs/nerm09/webprogram/Paper75087.html

Sulfonamide-Thiazolpyridine Derivatives,  Glucokinase Activators, Treatment Of Type 2 Diabetes

2009 52 (19) 6142 – 6152
Investigation of functionally liver selective glucokinase activators for the treatment of type 2 diabetes
Journal of Medicinal Chemistry
Bebernitz GR, Beaulieu V, Dale BA, Deacon R, Duttaroy A, Gao JP, Grondine MS, Gupta RC, Kakmak M, Kavana M, Kirman LC, Liang JS, Maniara WM, Munshi S, Nadkarni SS, Schuster HF, Stams T, Denny IS, Taslimi PM, Vash B, Caplan SL

2010 240th (August 22) Medi-198
Glucokinase activators with improved physicochemicalproperties and off target effects
American Chemical Society National Meeting and Exposition
Kirman LC, Schuster HF, Grondine MS et al

2010 240th (August 22) Medi-197
Investigation of functionally liver selective glucokinase activators
American Chemical Society National Meeting and Exposition
Schuster HF, Kirman LC, Bebernitz GC et al

PATENT

http://www.google.com/patents/US7750020

EXAMPLE 1 3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

A. Phenylacetic Acid Ethyl Ester

A solution of phenylacetic acid (50 g, 0.36 mol) in ethanol (150 mL) is treated with catalytic amount of sulfuric acid (4 mL). The reaction mixture is refluxed for 4 h. The reaction is then concentrated in vacuo. The residue is dissolved in diethyl ether (300 mL) and washed with saturated aqueous sodium bicarbonate solution (2×50 mL) and water (1×100 mL). The organic layer dried over sodium sulfate filtered and concentrated in vacuo to give phenylacetic acid ethyl ester as a colorless oil: ¹H NMR (400 MHz, CDCl₃) δ 1.2 (t, J=7.2, 3H), 3.6 (s, 2H), 4.1 (q, J=7.2, 2H), 7.3 (m, 5H); MS 165 [M+1]⁺.

B. (4-Chlorosulfonyl-phenyl)-acetic acid ethyl ester

To a cooled chlorosulfonic acid (83.83 g, 48 mL, 0.71 mol) under nitrogen is added the title A compound, phenylacetic acid ethyl ester (59 g, 0.35 mol) over a period of 1 h. Reaction temperature is brought to RT (28° C.), then heated to 70° C., maintaining it at this temperature for 1 h while stirring. Reaction is cooled to RT and poured over saturated aqueous sodium chloride solution (200 mL) followed by extraction with DCM (2×200 mL). The organic layer is washed with water (5×100 mL), followed by saturated aqueous sodium chloride solution (1×150 mL). The organic layer dried over sodium sulfate, filtered and concentrated in vacuo to give crude (4-chlorosulfonyl-phenyl)acetic acid ethyl ester. Further column chromatography over silica gel (60-120 mesh), using 100% hexane afforded pure (4-chlorosulfonyl-phenyl)-acetic acid ethyl ester as a colorless oil.

C. [4-(4-Methyl-piperazine-1-sulfonyl)-phenyl]-acetic acid ethyl ester

A solution of N-methylpiperazine (9.23 g, 10.21 ml, 0.092 mol), DIEA (13 g, 17.4 mL, 0.10 mol) and DCM 80 mL is cooled to 0° C., and to this is added a solution of the title B compound, (4-chlorosulfonyl-phenyl)-acetic acid ethyl ester (22 g, 0.083 mol) in 50 mL of DCM within 30 min. Reaction mixture stirred at 0° C. for 2 h, and the reaction mixture is washed with water (100 mL), followed by 0.1 N aqueous hydrochloric acid solution (1×200 mL). The organic layer dried over sodium sulfate, filtered and concentrated under vacuo to give crude [4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-acetic acid ethyl ester. Column chromatography over silicagel (60-120 mesh), using ethyl acetate afforded pure [4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-acetic acid ethyl ester as white crystalline solid: ¹H NMR (400 MHz, CDCl₃) δ 1.3 (t, J=7.4, 3H), 2.3 (s, 3H), 2.5 (m, 4H), 3.0 (br s, 4H), 3.7 (s, 2H), 4.2 (q, J=7.4, 2H), 7.4 (d, J=8.3, 2H), 7.7 (d, J=7.3, 2H); MS 327 [M+1]⁺.

D. 3-Cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid ethyl ester

A solution of the title C compound, [4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-acetic acid ethyl ester (15 g, 0.046 mol) in a mixture of THF (60 mL) and DMTP (10 mL) is cooled to −78° C. under nitrogen. The resulting solution is stirred at −78° C. for 45 min and to this is added LDA (25.6 mL, 6.40 g, 0.059 mol, 25% solution in THF/Hexane). A solution of iodomethylcyclopentane (11.60 g, 0.055 mol) in a mixture of DMTP (12 mL) and THF (20 mL) is added over a period of 15 min at −78° C. and reaction mixture stirred at −78° C. for 3 h further, followed by stirring at 25° C. for 12 h. The reaction mixture is then quenched by the dropwise addition of saturated aqueous ammonium chloride solution (50 mL) and is concentrated in vacuo. The residue is diluted with water (50 mL) and extracted with ethyl acetate (3×100 mL). The organic solution is washed with a saturated aqueous sodium chloride (2×150 mL), dried over sodium sulfate, filtered and concentrated in vacuo. Column chromatography over silica gel (60-120 mesh), using 50% ethyl acetate in hexane as an eluent to afford 3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid ethyl ester as a white solid: ¹H NMR (400 MHz, CDCl₃) δ 0.9-2.1 (m, 11H), 1.2 (t, J=7.1, 3H), 2.3 (s, 3H), 2.5 (br s, 4H), 3.0 (br s, 4H), 3.6 (m, 1H), 4.1 (q, J=7.1, 2H), 7.5 (d, J=8.3, 2H), 7.7 (d, J=8.3, 2H); MS 409 [M+1]⁺.

E. 3-Cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid

A solution of the title D compound, 3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid ethyl ester (14 g, 0.034 mol) in methanol:water (30 mL:10 mL) and sodium hydroxide (4.11 g, 0.10 mol) is stirred at 60° C. for 8 h in an oil bath. The methanol is then removed in vacuo at 45-50° C. The residue is diluted with water (25 mL) and extracted with ether (1×40 mL). The aqueous layer is acidified to pH 5 with 3 N aqueous hydrochloric acid solution. The precipitated solid is collected by vacuum filtration, washed with water (20 mL), followed by isopropyl alcohol (20 mL). Finally, solid cake is washed with 100 mL of hexane and dried under vacuum at 40° C. for 6 h to give 3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid as a white solid: ¹H NMR (400 MHz, CDCl₃) δ 1.1-2.0 (m, 11H), 2.4 (s, 3H), 2.7 (br s, 4H), 3.1 (br s, 4H), 3.6 (m, 1H), 7.5 (d, J=8.3, 2H), 7.6 (d, J=8.3, 2H); MS 381 [M+l]⁺.

F. 5-Methoxy-thiazolo[5,4-b]pyridin-2-ylamine

A solution of 6-methoxy-pyridin-3-ylamine (5.0 g, 0.0403 mol) in 10 mL of acetic acid is added slowly to a solution of potassium thiocyanate (20 g, 0.205 mol) in 100 mL of acetic acid at 0° C. followed by a solution of bromine (2.5 mL, 0.0488 mol) in 5 mL of acetic acid. The reaction is stirred for 2 h at 0° C. and then allowed to warm to RT. The resulting solid is collected by filtration and washed with acetic acid, then partitioned between ethyl acetate and saturated aqueous sodium bicarbonate. The insoluble material is removed by filtration and the organic layer is evaporated and dried to afford 5-methoxy-thiazolo[5,4-b]pyridin-2-ylamine as a tan solid.

G. 3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

A solution of the title E compound, 3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid (5 g, 0.013 mol) in DCM (250 mL) is cooled to 0° C. and then charged HOBt hydrate (2.66 g, 0.019 mol), followed by EDCI hydrochloride (6 g, 0.031 mol). The reaction mixture is stirred at 0° C. for 5 h. After that the solution of the title F compound, 5-methoxy-thiazolo[5,4-b]pyridin-2-ylamine (2.36 g, 0.013 mol) and D1EA (8 mL, 0.046 mol) in a mixture of DCM (60 mL) and DMF (20 mL) is added dropwise over 30 min. Reaction temperature is maintained at 0° C. for 3 h, then at RT (28° C.) for 3 days. Reaction is diluted with (60 mL) of water and the organic layer is separated and washed with saturated sodium bicarbonate solution (2×50 mL) followed by water washing (2×50 mL) and saturated sodium chloride aqueous solution (1×150 mL). Finally the organic layer is dried over sodium sulfate, filtered, and evaporated under vacuo. The crude product is purified using column chromatography over silica gel (60-120 mesh), using 40% ethyl acetate in hexane as an eluent to afford 3-cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide as a white solid: ¹H NMR (400 MHz, CDCl₃) δ 0.9-2.1 (m, 11H), 2.2 (s, 3H), 2.5 (br s, 4H), 3.1 (br s, 4H), 3.7 (m, 1H), 4.0 (s, 3H), 6.8 (d, J=8.8, 1H), 7.5 (d, J=8.3, 2H), 7.7 (d, J=8.3, 2H), 7.8 (d, J=8.8, 1H), 8.6 (s, 1H); MS 617 [M+1]⁺.

H. 3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide dihydrochloride

The title G compound, 3-cyclopentyl-2-(4-methyl piperazinyl sulfonyl)phenyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)propionamide (2.8 g, 0.0051 mol) is added to a cooled solution of 10% hydrochloric acid in isopropanol (3.75 mL). The reaction mixture is stirred at 0° C. for 1 h and then at RT for 2 h. The solid is separated, triturated with 10 mL of isopropanol and collected by vacuum filtration and washed with 50 mL of hexane. The solid is dried at 70° C. for 48 h to afford 3-cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide dihydrochloride as an off white solid.

EXAMPLE 2 (R)-3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

The title compound is obtained analogously to Example 1 by employing the following additional resolution step:

The racemic title E compound of Example 1,3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid (10 g, 0.026 mol) in 1,4-dioxane (500 mL) is treated in a three necked 1 liter flask, equipped with heating mantle, water condenser, calcium chloride guard tube and mechanical stirrer with 3.18 g (0.026 mol) of (R)-(+)-1-phenylethylamine. This reaction mixture is then refluxed at 100° C. for 1 h. The clear reaction solution is cooled to RT (27° C.) and stirred for 10 h. The crystallized salt is collected by filtration under vacuum, washed with 5 mL of hexane and dried under vacuum to afford salt A.

The salt A is dissolved in 1,4-dioxane (500 mL) and heated at 100° C. for 1 h. The clear reaction solution is cooled to RT (27° C.) and stirred for 10 h. The crystallized product is collected by filtration under vacuum, washed with 50 mL of hexane, and dried under vacuum to afford salt B.

The salt B is dissolved in 1,4-dioxane (290 mL) and heated at 100° C. for 1 h. The clear reaction solution is cooled to RT (27° C.) and stirred for 10 h. The crystallized product is collected by filtration under vacuum, washed with 30 mL of hexane, and dried under vacuum to afford salt C.

The salt C is dissolved in 1,4-dioxane (100 mL) and heated at 100° C. for 1 h. The clear reaction solution is cooled to RT (27° C.) and stirred for 10 h. The crystallized product is collected by filtration under vacuum, washed with 30 ml of hexane, and dried under vacuum to afford salt D.

The salt D is treated with aqueous hydrochloric acid solution (20 mL, 1 mL of concentrated hydrochloric acid diluted with 100 mL of water) and stirred for 5 min. The white solid precipitates out and is collected by vacuum filtration, washed with 10 mL of cold water, 5 mL of isopropanol and 20 mL of hexane, and dried under vacuum to yield the hydrochloride salt of (R)-(−)-3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid, salt E.

The salt E is neutralized by stirring with aqueous sodium bicarbonate solution (10 mL, 1 g of sodium bicarbonate dissolved in 120 mL of water) for 5 min. The precipitated solid is collected by filtration, washed with 10 mL of cold water, 100 mL of hexane, and dried to afford (R)-(−)-3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid: m.p. 202.2-203.4° C.

Alternatively, the title compound may be obtained by the resolution of the racemic title compound of Example 1 using the following preparative chiral HPLC method:

• Column: Chiralcel OD-R (250×20 mm) Diacel make, Japan;
• Solvent A: water:methanol:acetonitrile (10:80:10 v/v/v);
• Solvent B: water:methanol:acetonitrile (05:90:05 v/v/v);
• Using gradient elution: gradient program (time, min/% B): 0/0, 20/0, 50/100, 55/0, 70/0;
• Flow rate: 6.0 mL/min; and
• Detection: by UV at 305 nm.

EXAMPLE 3 (S)-3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

The title compound is prepared analogously to Example 2.

J MED CHEM 2009, 52, 6142-52

Investigation of Functionally Liver Selective Glucokinase Activators for the Treatment of Type 2 Diabetes

http://pubs.acs.org/doi/abs/10.1021/jm900839k

Type 2 diabetes is a polygenic disease which afflicts nearly 200 million people worldwide and is expected to increase to near epidemic levels over the next 10−15 years. Glucokinase (GK) activators are currently under investigation by a number of pharmaceutical companies with only a few reaching early clinical evaluation. A GK activator has the promise of potentially affecting both the β-cells of the pancreas, by improving glucose sensitive insulin secretion, as well as the liver, by reducing uncontrolled glucose output and restoring post-prandial glucose uptake and storage as glycogen. Herein, we report our efforts on a sulfonamide chemotype with the aim to generate liver selective GK activators which culminated in the discovery of 3-cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide (17c). This compound activated the GK enzyme (αK_a = 39 nM) in vitro at low nanomolar concentrations and significantly reduced glucose levels during an oral glucose tolerance test in normal mice.

PATENT

EP-1735322-B1

Example 2(R)-3-Cyclopentyl-N-(5-methoxy-thiazolo[5,4-b]pyridin-2-yl)-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionamide

The title compound is obtained analogously to Example 1 by employing the following additional resolution step:

The racemic title E compound of Example 1, 3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid (10 g, 0.026 mol) in 1,4-dioxane (500 mL) is treated in a three necked 1 liter flask, equipped with heating mantle, water condenser, calcium chloride guard tube and mechanical stirrer with 3.18 g (0.026 mol) of (R)-(+)-1-phenylethylamine. This reaction mixture is then refluxed at 100°C for 1 h. The clear reaction solution is cooled to RT (27°C) and stirred for 10 h. The crystallized salt is collected by filtration under vacuum, washed with 5 mL of hexane and dried under vacuum to afford salt A.

The salt A is dissolved in 1,4-dioxane (500 mL) and heated at 100°C for 1 h. The clear reaction solution is cooled to RT (27°C) and stirred for 10 h. The crystallized product is collected by filtration under vacuum, washed with 50 mL of hexane, and dried under vacuum to afford salt B.

The salt B is dissolved in 1,4-dioxane (290 mL) and heated at 100°C for 1 h. The clear reaction solution is cooled to RT (27°C) and stirred for 10 h. The crystallized product is collected by filtration under vacuum, washed with 30 mL of hexane, and dried under vacuum to afford salt C.

The salt C is dissolved in 1,4-dioxane (100 mL) and heated at 100°C for 1 h. The clear reaction solution is cooled to RT (27°C) and stirred for 10 h. The crystallized product is collected by filtration under vacuum, washed with 30ml of hexane, and dried under vacuum to afford salt D.

The salt D is treated with aqueous hydrochloric acid solution (20 mL, 1 mL of concentrated hydrochloric acid diluted with 100 mL of water) and stirred for 5 min. The white solid precipitates out and is collected by vacuum filtration, washed with 10 mL of cold water, 5 mL of isopropanol and 20 mL of hexane, and dried under vacuum to yield the hydrochloride salt of (R)-(-)-3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid, salt E.

The salt E is neutralized by stirring with aqueous sodium bicarbonate solution (10 mL, 1 g of sodium bicarbonate dissolved in 120 mL of water) for 5 min. The precipitated solid is collected by filtration, washed with 10 mL of cold water, 100 mL of hexane, and dried to afford (R)-(-)-3-cyclopentyl-2-[4-(4-methyl-piperazine-1-sulfonyl)-phenyl]-propionic acid: m.p. 202.2-203.4°C.

Alternatively, the title compound may be obtained by the resolution of the racemic title compound of Example 1 using the following preparative chiral HPLC method:

• Column: Chiralcel OD-R (250 x 20 mm) Diacel make, Japan;
• Solvent A: water:methanol:acetonitrile (10:80:10 v/v/v);
• Solvent B: water:methanol:acetonitrile (05:90:05 v/v/v);
• Using gradient elution: gradient program (time, min / %B): 0/0, 20/0, 50/100, 55/0, 70/0;
• Flow rate: 6.0 mL/min; and
• Detection: by UV at 305 nm.

REFERENCES

US 7750020

WO-2005095418-A1

US-20080103167-A1

///NOVARTIS, DIABETES, Sulfonamide-Thiazolpyridine Derivatives,  Glucokinase Activators, Treatment Of Type 2 Diabetes, 866772-52-3, Novartis Molecule, functionally liver selective glucokinase activators, treatment of type 2 diabetes , NVP-LBX192, LBX-192

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