Deuterated dydrogesterone

Abstract

The present invention provides to deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V). The invention further provides commercially viable process for preparation of deuterated Dydrogesterone. The invention specifically provides deuterated dydrogesterone with increased half-life period with therapeutic advantages.

Description

[0001] DEUTERATED DYDROGESTERONE

[0002] FIELD OF THE INVENTION:

[0003] The present invention provides to deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V). The invention further provides commercially viable process for preparation of deuterated Dydrogesterone. The invention specifically provides deuterated dydrogesterone with increased half-life period with therapeutic advantages.

[0004] BACKGROUND OF THE INVENTION:

[0005] Deuterium (2H) is a non-radioactive isotope of hydrogen, which bonds covalently with other atoms in the same manner as hydrogen. However, deuterium being heavier than hydrogen forms strong bonds with carbon and gives rise to a difference in pharmacological properties during the drug’s metabolism in the body. The replacement of hydrogen with deuterium slows down the metabolism, thereby increasing the half-life of the drug in the body. A longer half-life permits a reduced dose amount or dosing frequency and helps to decrease unwanted side effects caused by metabolites.

[0006] Thus, if the Carbon-Hydrogen bond breaking is the rate-determining step in the biotransformation of a drug, the deuterated drug may show improved pharmacokinetic characteristics, such as a longer half-life, so frequent dosing is reduced considerably. If a toxic metabolite or reactive intermediate is formed when the Carbon-Hydrogen bond is broken, then deuteration of the drug may reduce the unwanted side effects of the drugs.

[0007] A carbon-deuterium bond also helps in slowing the undesirable epimerisation of enantiomers in a chiral drug as in the case of the notorious drug, thalidomide. Scientists began incorporating deuterium into potential drugs over 50 years ago, and enthusiasm about using deuterium to modify pharmacological properties has been wavering drastically probably due to incidences of toxicity while evaluating deuterated fludalanine.

[0008] Deutetrabenazine is a deuterated analogue of the old drug tetrabenazine, with the two methoxy groups in the latter being replaced by a pair of trideuteromethoxy groups, thereby altering the rate of metabolism to afford greater tolerability with an improved dosing regimen. Deuterium substitution impedes oxidative metabolism of the methoxy groups, in an excellent demonstration of the primary kinetic isotope effect which is used for probing chemical reaction mechanism.

[0009] Deuruxolitinib (Leqselvi) approved by FDA on July 26, 2024, is the deuterated form of ruxolitinib (Jakafi) that selectively inhibits Janus associated kinases (JAK1 and JAK2). Deuteration allows the drug to circumvent extensive oxidative metabolism around the cyclopentyl ring, which increases the duration of the deuruxolitinib pharmacological activity. Leqselvi is indicated for the treatment of alopecia areata by targeting the yc cytokine and interferon-gamma (IFN-y) signalling pathway to reverse hair loss while Ruxolitinib is an onco drug, indicated for the treatment of intermediate or high-risk myelofibrosis.

[0010] Several other deuterated drugs like lenalidomide, d6-dextrometorphan, ruxolitinib, venlafaxine and its desmethyl derivative, pioglitazone, apremilast and ivacaftor are in various stages of clinical trials.

[0011] Dydrogesterone is a potent, orally active progestogen indicated for a wide variety of gynaecological conditions related to progesterone deficiency like menstrual disorders, endometriosis, infertility, and menopausal symptoms. European Journal of Drug Metabolism and Pharmacokinetics 1980, 5(3), 173-184 mentions the following major metabolites, 20a-hydroxy-9p,10a-pregna-4,6-diene-3-one (52%), 2 l-hydroxy-9p,10a-pregna-4,6-diene-3, 20-dione (18%) and 16a-hydroxy-9p,10a- 4, 6-diene-3, 20-dione (1%) which accounts for 70% of the metabolic products. The 4,6-diene-3-one configuration of the steroid moiety remains intact during the metabolism.

[0012] 20a-hydroxy-9p,10a-pregna-4,6-diene-3-one 21 -hydroxy-9p, 1 Oa- pregna-4,6-diene-

[0013] 16a-hydroxy-9p, 10a-4,6-diene-3 ,20-dione

[0014] Fertility and Sterility 2006, 86(3), 1235-1242 mentions that the major active metabolite of dydrogesterone, 20a-dihydrodydrogesterone (20a-DHD) also has progestogenic activity. But since the metabolite has decreased activity, it is imperative to delay the metabolism of dydrogesterone by selectively deuterating three hydrogen atoms of C-21 of the steroid framework.

[0015] The present inventors after considering the advantages of deuterated derivatives for other drugs and the associated risks have developed deuterated dydrogesterone of formula (I), formula (II), formula (III), formula (IV) and formula (V).

[0016]

[0017] Deuterated Dydrogesterone of formula (I) Deuterated Dydrogesterone of formula (II)

[0018] Deuterated Dydrogesterone of formula (III) Deuterated Dydrogesterone of formula (IV)

[0019] Deuterated Dydrogesterone of formula (V)

[0020] OBJECT OF THE INVENTION An objective of the present invention is to provide deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V). Another objective of the present invention is to provide a commercially viable and cost-effective process for the preparation of deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V).

[0021] Yet another objective of the present invention is to provide a process for preparing deuterated dydrogesterone of formulae (II) and (V) by treating deuterated Dydrogesterone of formula (I) with a deuterating agent.

[0022] A further objective of the present invention is to particularly, provide deuterated dydrogesterone with increased half-life period with therapeutic advantages.

[0023] SUMMARY OF THE PRESENT INVENTION

[0024] An aspect of the present invention is to provide deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V).

[0025] Another aspect of the present invention is to provide a commercially viable and cost-effective process for the preparation of deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V).

[0026] Another aspect of the present invention is to provide deuterated dydrogesterone of formulae (I), (II), (III), (IV) and (V) having increased half-life and therapeutic advantages.

[0027] Yet another aspect of the invention is to provide a process for preparing deuterated dydrogesterone of formula (I) comprising reaction of dydrogesterone with an oxidising agent in an organic solvent to give the conjugated ketone carboxylic acid (DYD-A), which is then converted to Weinreb amide (DYD-B) and reduced to an alcohol (DYD-C), followed by treatment with a deuterated Grignard reagent to get deuterated alcohol (DYD-D) followed by oxidation gives deuterated Dydrogesterone of formula (I)

[0028] Yet another aspect of the invention is to provide a process for preparing deuterated dydrogesterone of formula (I) comprising reaction of dydrogesterone with bromine in presence of aqueous sodium hydroxide in an organic solvent to provide the conjugated ketone carboxylic acid (DYD-A) , which was then treated with N-O- dimethyl hydroxylamine hydrochloride in presence of a base and in an organic solvent to provide Weinreb amide (DYD-B) which is then reduced with sodium borohydride in presence of CeCh to give an alcohol (DYD-C), which on further treatment with a deuterated Grignard reagent CDsMgl gives a deuterated alcohol (DYD-D) followed by oxidation with Dess-Martin periodinane reagent provides deuterated Dydrogesterone of formula (I).

[0029] A further aspect of the invention is to provide a process for preparing deuterated dydrogesterone of formulae (II) and (V) by treating the deuterated Dydrogesterone of formula (I) with a deuterating agent.

[0030] DETAILED DESCRIPTION OF THE INVENTION

[0031] The invention is embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that the disclosures satisfy applicable legal requirements. As used in the specification, and in the appended claims, the singular forms “a,” “an,” “the,” include plural referents unless the context clearly indicates otherwise.

[0032] Abbreviations:

[0033] HLM: Human Liver Microsomal

[0034] RLM: Rat Liver Microsomal

[0035] MLM: Mouse Liver Microsomal

[0036] CLM: Cyano Monkey Liver DLM: Dog liver microsomal CL int: Intrinsic Clearance

[0037] The term solvent used herein, refers to the single solvent or mixture of solvents.

[0038] An embodiment of the present invention is to provide deuterated dydrogesterone of formula (I), (II), (III), (IV) and (V).

[0039] An embodiment of the present invention relates to preparation of deuterated dydrogesterone (I), chemically known as ((8S,9R,10S,13S,14S,17S)-17-(acetyl- d3)-10,13-dimethyl-l,2,8,9,10,ll,12,13,14,15,16,17-dodecahydro-3H- cy clopenta[a] phenanthren- 3 -one) .

[0040] In another embodiment, the present invention provides process for preparing deuterated dydrogesterone of formula (I) comprising reaction of dydrogesterone with an oxidising agent in an organic solvent to give the conjugated ketone carboxylic acid (DYD-A), which is then converted to Weinreb amide (DYD-B) and reduced to an alcohol (DYD-C), followed by treatment with a deuterated Grignard reagent to get deuterated alcohol (DYD-D) followed by oxidation gives deuterated Dydrogesterone of formula (I).

[0041] In a related embodiment, the present invention provides process for preparing deuterated dydrogesterone of formula (I) comprising reaction of dydrogesterone with bromine in presence of aqueous sodium hydroxide in an organic solvent to provide the conjugated ketone carboxylic acid (DYD-A) , which was then treated with N-O-dimethyl hydroxylamine hydrochloride in presence of a base and in an organic solvent to provide Weinreb amide (DYD-B) which is then reduced with sodium borohydride in presence of CeCh to give an alcohol (DYD-C), which on further treatment with a deuterated Grignard reagent CDsMgl gives a deuterated alcohol (DYD-D) followed by oxidation with Dess-Martin periodinane reagent provides deuterated Dydrogesterone of formula (I)

[0042] Deuterated Dydrogesterone formula (I)

[0043] Scheme I: Preparation of Deuterated dydrogesterone (I)

[0044] Deuterated Dydrogesterone formula (I)

[0045] Deuterated Dydrogesterone formula (II) Deuterated Dydrogesterone formula (V) Scheme II: Preparation of Deuterated dydrogesterone (II) and (V)

[0046] The present invention relates to an improved process for the preparation of deuterated dydrogesterone of formulae (II) and (V).

[0047] Deuterated dydrogesterone (I) is synthesized by a route disclosed in Scheme - 1 and involves the preparation of the following compounds: i) DYD-A ((8S,9R,10S,13S,14S,17S)-10,13-dimethyl-3-oxo-

[0048] 2,3,8,9,10,11,12,13,14,15,16, 17-dodecahydro-lH- cyclopenta[a]phenanthrene- 17-carboxylic acid), ii) DYD-B ((8S,9R,10S,13S,14S,17S)-N-methoxy-N,10,13-trimethyl-3-oxo-

[0049] 2,3,8,9,10, 11,12, 13, 14, 15,16, 17-dodecahydro-lH- cy clopenta[a] phenanthrene- 17 -c arboxamide) , iii) DYD-C l-((8S,9R,10S,13S,14S,17S)-3-hydroxy-10,13-dimethyl-

[0050] 2,3,8,9,10,11,12,13,14, 15,16,17-dodecahydro-lH-cyclopenta[a]phenanthren- 17-yl)ethan-l-one-2,2,2-d3 and iv) DYD-D ((8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0051] 1,2,8,9,10,11,12, 13,14,15,16,17-dodecahydro-3H-cyclopenta[a]phenanthren- 3-one).

[0052] Deuterated dydrogesterone of formula (I) is obtained by oxidising Dydrogesterone to a carboxylic acid DYD-A which is then converted to its Weinreb amide DYD- B. The amide DYD-B is reduced to alcohol DYD-C followed by nucleophilic addition of Deuterated Grignard reagent to DYD-C to give DYD-D. Allylic oxidation of DYD-D provided Deuterated Dydrogesterone of formula (I) with desired purity.

[0053] In a related embodiment, the process for preparation of deuterated dydrogesterone of formula (I), comprises oxidation of Dydrogesterone by reaction with sodium hypobromite solution in an organic solvent, and quenching with sodium sulfite solution to give the conjugated ketone carboxylic acid DYD-A. The sodium hypobromite solution was prepared by adding bromine to a cooled solution of sodium hydroxide at -10 to 20°C. The amount of bromine utilized was between 2.0 and 3.0 moles of bromine per mole of dydrogesterone. The cold sodium hypobromite solution was diluted with an organic solvent such as 1,4-dioxane, dichloromethane and added to a stirred mixture of dydrogesterone in aqueous dioxane or dichloromethane at -10 to 20°C. Reaction mixture was quenched with aqueous sodium sulfite solution and the pH adjusted to around 5 to give DYD-A.

[0054] In addition to sodium hypobromite, reagents like sodium hypochlorite (NaOCl) or m-chloroperbenzoic acid can also be used as an oxidizing agent.

[0055] The DYD-A was then converted to its Weinreb amide derivative by reaction with N-O-dimethyl hydroxylamine hydrochloride and isolating after treatment with an aqueous acid to give DYD-B.

[0056] The compound DYD-A was dissolved in an organic solvent like dichloromethane, dimethylformamide (DMF), tetrahydrofuran (THF), 1,4-dioxane or any other solvent in the class of ethers. A mixture of bases selected from diisopropylethylamine and dimethylaminopyridine, in an amount ranging from 0.20 - 1.75 mole equivalent of the base per mole of DYD-A.

[0057] 3-Dimethylaminopropyl)ethylcarbodiimide hydrochloride (1.25-1.75 mole equivalent / mole of DYD-A) was added to the mixture followed by N-O-dimethyl hydroxylamine hydrochloride (1.25 -1.75 mole equivalent / mole of DYD-A) was added. The compound, DYD-B was isolated after quenching with an aqueous base like sodium bicarbonate.

[0058] In addition to CDI, other reagents like thionyl chloride (SOCh), Dicyclohexylcarbodimide (DCC) and acetic anhydride can be used. The amide DYD-B was reduced with sodium borohydride (NaBtU) or any suitable reducing agent in presence of a catalyst which is not limited to CcCh. HiO to give DYD-C.

[0059] DYD-B was dissolved in an organic solvent like methanol and stirred with the catalyst CeCh (2.5-3.5 moles equivalent / mole of DYD-B) at -5 to +5°C. A suitable reducing agent like sodium borohydride (2.5 - 3.5 moles equivalent / mole of DYD- B) was added to the mixture at -5 to +5 °C, and the DYD-C was isolated after quenching with water and dilute HC1.

[0060] The organic solvent is selected from the group comprising of alcohols (ethanol, isopropanol etc.), ether (THF, 1,4-dioxane etc.), aliphatic hydrocarbons (hexane, heptane, methyl cyclohexane etc.) and aromatic hydrocarbons (toluene, xylene etc.)

[0061] The reducing agent can also be Lithium aluminium hydride, Vitride (Metal-hydride reducing reagents), boranes etc.

[0062] Nucleophilic addition of deuterated Grignard reagent (9.5 - 10.5 moles equivalent / mole of DYD-C) like CDsLi or (CDs Mg was added to compound DYD-C dissolved in an organic solvent like tetrahydrofuran (THF) at -10 to +10°C and isolated by quenching with 10% aqueous NH4CI solution, extraction with ethyl acetate followed by evaporation of solvent and purification by column chromatography with ethyl acetate / hexane mixture as eluent to give compound DYD-D.

[0063] DYD-D was then oxidised in dichloromethane or any ether solvent as an organic solvent in presence of oxidising agent, which was not limited to Dess-Martin periodinane reagent (4.0 - 6.0 moles equivalent / mole of DYD-D). Deuterated Dydrogesterone of formula (I) was isolated by quenching with an aqueous solution of an inorganic base like NaHCCL and purification by column chromatography with ethyl acetate / hexane mixture as eluent. In addition to Dess-Martin periodinane, other oxidizing agents like IBX, DMAP, metal catalysed oxidizing reagents can also be employed.

[0064] Similarly, Deuterated dydrogesterone of formulae (II) and (V) are obtained by treating deuterated dydrogesterone of formula (I) with D2O in the presence of an inorganic base in an organic solvent at ambient temperature.

[0065] Deuterated Dydrogesterone of formula (II) chemically known as (8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0066] 1.2.8.9.10.11.12.13.14.15.16.17-dodecahydro-3H-cyclopenta [a]phenanthren-3- one-17-d, Deuterated Dydrogesterone of formula (III) chemically known as (8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0067] 1.2.8.9.10.11.12.13.14.15.16.17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-8-d, Deuterated Dydrogesterone of formula (IV) chemically known as (8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0068] 1,2,8,9,10,11,12, 13, 14, 15, 16, 17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-2,2,4,8,17-d5, and Deuterated Dydrogesterone of formula (V) chemically known as (8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0069] 1,2,8,9,10,11,12, 13, 14, 15,16, 17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-2,2-d2 were prepared by further deuteration of Deuterated Dydrogesterone (I) as disclosed in Scheme - II.

[0070] Deuterated dydrogesterone of formula (I) was dissolved in a solvent like tetrahydrofuran (THF). The amount of solvent is 40-50 volumes per gram of Deuterated dydrogesterone of formula (I).

[0071] D2O (10 volumes per gram of Deuterated dydrogesterone of formula (I)) was added to the mixture in presence of an inorganic base like alkali hydroxide, alkali carbonate or alkali bicarbonate but preferably an alkali carbonate.

[0072] The mixture was stirred at ambient temperature for 24 hours as monitored by LCMS. The deuterated compound (II) or (V) was isolated by concentrating the reaction mixture followed by dilution with a non-polar solvent like a hydrocarbon selected from toluene, xylene, cyclohexane, methyl cyclohexane etc.

[0073] The organic layer / mixture was concentrated under reduced pressure and the residue was diluted with a non-polar solvent like a hydrocarbon to separate out a mixture of compound (II) and (V), which were separated by chromatographic purification.

[0074] PHARMACEUTICAL COMPOSITIONS

[0075] The present invention provides pharmaceutical compositions which include at least one compound described herein and at least one pharmaceutically acceptable excipient. The pharmaceutically acceptable excipient for the purpose of this invention includes but not limited to diluents or carrier, binder, bulking agent. Preferably, the contemplated pharmaceutical compositions include a compound(s) described herein in therapeutically effective amount sufficient to treat conditions related to Dydrogesterone in a subject. The subjects contemplated include, for example, a living cell and a mammal, including human.

[0076] Examples of suitable carriers include, but are not limited to, water, salt solutions, alcohols, polyethylene glycols, polyhydroxyethoxylated castor oil, peanut oil, olive oil, gelatin, lactose, terra alba, sucrose, dextrin, magnesium carbonate, sugar, cyclodextrin, amylose, magnesium stearate, talc, gelatin, agar, pectin, acacia, stearic acid or lower alkyl ethers of cellulose, silicic acid, fatty acids, fatty acid amines, fatty acid monoglycerides and diglycerides, pentaerythritol fatty acid esters, polyoxyethylene, hydroxy methylcellulose and polyvinylpyrrolidone.

[0077] The carrier or diluent may include a sustained release material, such as, for example, glyceryl monostearate or glyceryl distearate, alone or mixed with a wax.

[0078] The pharmaceutical composition may also include one or more pharmaceutically acceptable auxiliary agents, wetting agents, emulsifying agents, suspending agents, preserving agents, salts for influencing osmotic pressure, buffers, sweetening agents, flavoring agents, colorants, or any combination of the foregoing. The pharmaceutical composition of the invention may be formulated to provide quick, sustained, or delayed release of the active ingredient after administration to the subject by employing procedures known in the art.

[0079] The pharmaceutical compositions described herein may be prepared, e.g., as described in Remington: The Science and Practice of Pharmacy, 20th Ed., 2003 (Lippincott Williams & Wilkins). For example, the active compound can be mixed with a carrier, or diluted by a carrier, or enclosed within a carrier, which may be in the form of an ampule, capsule, or sachet. When the carrier serves as a diluent, it may be a solid, semisolid, or liquid material that acts as a vehicle, excipient, or medium for the active compound.

[0080] The pharmaceutical compositions may be, for example, capsules, tablets, aerosols, solutions, suspensions, liquids, gels, or products for topical application.

[0081] The route of administration may be any route which effectively transports the active compound to the appropriate or desired site of action. Suitable routes of administration include, but are not limited to, oral, nasal, pulmonary, buccal, subdermal, intradermal, transdermal, parenteral, rectal, depot, subcutaneous, intravenous, intraurethral, intramuscular, intranasal, ophthalmic (such as with an ophthalmic solution) or topical (such as with a topical ointment). The oral route is preferred.

[0082] Solid oral formulations include, but are not limited to, tablets, capsules (soft or hard gelatin), dragees (containing the active ingredient in powder or pellet form), troches and lozenges. Tablets, dragees, or capsules having talc and / or a carbohydrate carrier or binder or the like are particularly suitable for oral application. Preferable carriers for tablets, dragees, or capsules include lactose, cornstarch, and / or potato starch. A syrup or elixir can be used in cases where a sweetened vehicle can be employed.

[0083] A typical tablet that may be prepared by conventional tableting techniques. Liquid formulations include, but are not limited to, syrups, emulsions, soft gelatin, and sterile injectable liquids, such as aqueous or non-aqueous liquid suspensions or solutions.

[0084] For parenteral application, particularly suitable are injectable solutions or suspensions, preferably aqueous solutions with the active compound dissolved in polyhydroxylated castor oil.

[0085] The advantages of instant invention over the prior art processes are as follows:

[0086] 1. It is expected to increase the half-life period of the drug in the body.

[0087] 2. Increase in the half-life period would reduce the dosage and its frequency.

[0088] 3. Improvement in the Pharmacokinetics of the drug.

[0089] The present invention is explained in detail with reference to the following examples described below, which are given for the purpose of illustration only and are not intended to limit the scope of the invention.

[0090] EXAMPLES

[0091] Example 1: Preparation of DYD-A ((8S,9R,10S,13S,14S,17S)-10,13-dimethyl- 3-oxo-2,3,8,9,10,l l,12,13,14,15,16,17-dodecahydro-lH- cyclopenta[a]phenanthrene- 17-carboxylic acid)

[0092] DYD-A

[0093] Bromine (28.12 g, 0.176 moles) was added slowly to a stirred solution of (20.89 g, 0.522 moles) of sodium hydroxide in 180 ml of water cooled to -5 to 0°C. Dioxane (120 ml) was added and the cold sodium hypobromite solution was added slowly to a stirred solution of dydrogesterone (14gms, 0.44 mol) in dioxane (560 ml) and water (160 ml) at 0°C. After reaction completion, the reaction mixture was quenched with 10% sodium sulfite solution (100 ml) and partially concentrated. The reaction mixture was extracted with ethyl acetate (250 ml) and the aqueous layer was then acidified with cone, hydrochloric acid to pH 5. The product, DYD- A was filtered, washed with water (50 ml) and dried under reduced pressure.

[0094] Yield: 4.0 gms (29 %).

[0095] 'H NMR (400 MHz, CDCh): 6.45-6.20 (m, 2H), 5.68 (s, 1H), 2.63-2.45 (m, 1H), 2.45-2.39 (m, 3H), 2.31-2.21 (m, 1H), 2.21-2.09 (m,lH), 2.09-1.95 (m,2H), 1.94- 1.81 (m, 3H), 1.81-1.60 (m, 3H), 1.58-1.47 (m, 1H), 1.45-1.36 (m, 1H), 1.35-1.28 (m, 1H), 1.27 (s, 3H), 0.82 (s, 3H);

[0096] 13C NMR (100 MHz, CDCh): 199.8, 179.1, 163.3, 140.4, 127.1, 123.8, 54.8, 49.4, 44.4, 39.6, 38.7, 37.2, 36.8, 35.5, 33.9, 25.1, 23.0, 22.2, 20.4, 12.0.

[0097] LC-MS: [M+H]+= 315.1

[0098] Example 2: Preparation of DYD-B (8S,9R,10S,13S,14S,17S)-N-methoxy-

[0099] N,10,13-trimethyl-3-oxo-2,3,8,9,10,l l,12,13,14,15,16,17-dodecahydro-lH- cyclopenta[a]phenanthrene -17-carboxamide)

[0100] DYD-B

[0101] DYD-A (11.11 g, 0.035 moles) was dissolved in dichloromethane (555.5 mL) and cooled to 0°C. Diisopropylethylamine (DIPEA; 9.14ml; 0.052moles) and dimethylaminopyridine (DMAP; 0.855gms, 0.007 moles) followed by 3- (Dimethylamino-propyl)ethyl carbodiimide hydrochloride, (EDC. HC1; lO.Ogms, 0.052moles) were added gradually. The compound, N-O-dimethyl hydroxylamine hydrochloride (5.12 g, 0.052 mol) was then added to the reaction mixture and stirred till completion of reaction as monitored by TLC / HPLC. The reaction mixture was quenched with water (100 ml), the organic layer was washed with 10% sodium bicarbonate solution (200 ml), dried and concentrated under reduced pressure to give DYD-B, which was then passed through column chromatography column using 0-25% Ethyl acetate in hexane as eluent to give DYD-B as a white solid.

[0102] Yield: 5.20gms (41%).

[0103] 'H NMR (400 MHz, CDCh): 6.24-6.13 (m, 2H), 5.67 (s, 1H), 3.64 (s, 3H), 3.20 (s, 3H), 2.87-2.74 (m, 1H), 2.62-2.49 (m, 1H), 2.49-2.41 (m, 2H), 2.31-2.21 (m, 2H), 2.0-1.62 (m, 8H), 1.54-1.40 (m, 2H), 1.30 (s, 3H), 0.82 (s, 3H);

[0104] 13C NMR (100 MHz, CDCh): 199.6, 174.7, 163.3, 140.9, 126.8, 123.7, 60.9, 50.8, 49.7, 45.5, 39.6, 38.8, 37.8, 37.2, 35.5, 33.9, 32.1, 25.5, 24.5, 22.7, 20.6, 12.7.

[0105] LC-MS: [M+H]+= 358.1

[0106] Example 3: Preparation of DYD-C ((8S,9R,10S,13S,14S,17S)-3-hydroxy-N- methoxy-N,10,13-trimethyl-2,3,8,9,10,l l,12,13,14,15,16,17-dodecahydro-lH- cyclopenta[a]-phenanthrene-17-carboxamide)

[0107] DYD-C

[0108] DYD-B (2.0 g, 0.0056 mol) was dissolved in methanol (40 ml) and CeC13.7H2O (6.25 g, 0.0168 moles) was added at room temperature under argon atmosphere. The mixture was cooled to 0°C and NaBH4 (0.63 g, 0.0168 mol) was added gradually at 0°C and stirred for 2 hours. After completion of the reaction as monitored by HPLC / TLC, water (25 ml) was added and concentrated under reduced pressure. The residue was diluted with ethyl acetate (50 ml), the organic layer was separated, and the aqueous layer was further extracted with ethyl acetate (100ml). The combined organic extracts were washed with dil. HC1, dried over anhydrous NaiSCh and concentrated under reduced pressure to afford DYD-C as white solid. Yield: 2 gms (99%).

[0109] 'H NMR (400 MHz, CDCh): (Mixture of Diastereomers); 6.20-6.14 (m, 0.37H), 5.96 (d, J = 9.6 Hz, 1H), 5.71-5.64 (m, 1H), 5.36-5.33 (m, 0.52H), 4.27 (t, J = 8Hz, 0.31H), 4.14-4.08 (m, 0.5H), 3.64 (s, 3H), 3.19 (s, 3H), 2.77 (bs, 1H), 2.6-2.42 (m, 0.6H), 2.40-2.15 (2H), 2.12-1.58 (m, 13H), 1.56-1.35 (m, 3H), 1.35-1.05 (m, 5H), 0.98-0.78 (m, 4H);

[0110] LC-MS: [M+H]+= 360.2

[0111] Example 4: Preparation of DYD-D (l-((8S,9R,10S,13S,14S,17S)-3-hydroxy- 10,13-dimethyl-2,3,8,9,10,l l,12,13,14,15,16,17-dodecahydro-lH- cyclopenta[a]phenanthren-17-yl)ethan-l-one-2,2,2-d3)

[0112] DYD-D

[0113] DYD-C (0.100 g, 0.28 mol) was dissolved in tetrahydrofuran (5 ml) and Grignard reagent CDsMgl (IM in diethyl ether, 2.79 moles) was added under inert atmosphere at 0°C. The reaction mixture was stirred at room temperature for 5 hours till completion of reaction as monitored by HPLC / TLC. The reaction mixture was quenched with 10% aq. NH4CI solution, extracted with ethyl acetate, the organic layer was washed with water, dried on NaiSCU and concentrated to get DYD-D. The compound was passed through a chromatographic column using 0-20% Ethyl acetate in hexane as eluant to give pure DYD-D as a white solid.

[0114] Yield: 0.050 gm, (57%). 'H NMR (400 MHz, CDCh): (Mixture of Diastereomers); 6.17 (s, 0.5 H), 5.96 (d, J = 10 Hz, 1H), 5.69-5.66 (m, 1H), 5.33 (s, 1H), 4.27 (t, J = 7.6 Hz, 1H), 2.60-2.50 (m, 1H), 2.50-2.39 (m, 0.7H), 2.32-2.26 (m, 1H), 2.25-2.15 (m, 2H), 2.25-2.04 (m, 2H), 2.02-1.58 (m, 17H), 1.39-1.22 (m, 6H), 1.2 (s, 3H), 0.70 (s, 0.9H), 0.66 (s, 3H);

[0115] LC-MS: [M-H2O]+= 300.0

[0116] Example 5: Preparation of Deuterated Dydrogesterone of formula (I) ((8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0117] 1,2,8,9,10,11,12, 13, 14, 15,16, 17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one)

[0118] Deuterated Dydrogesterone (I)

[0119] DYD-D (0.045 g, 0.14 mol) was dissolved in dichloromethane (4.5 ml) and stirred with Dess-Martin periodinane reagent (0.71 g, 0.71 mole). The reaction mixture was further stirred at room temperature for 2 hours till reaction completion and then quenched with 10% aqueous NaHCOs (10 ml) solution. The reaction mixture was stirred at room temperature for 10 minutes and extracted with dichloromethane (20 ml). The organic layer was separated, dried on NaiSCh and concentrated under reduced pressure to get deuterated dydrogesterone which was then passed through a chromatographic column and using 0-20% Ethyl acetate in hexane as eluent to give pure deuterated dydrogesterone of formula (I) as an off white solid.

[0120] Yield: (0.025 gm, 56%).

[0121] HPLC purity: 97.81% 'H NMR (400 MHz, CDCh): 6.17 (t, J = 13.6 Hz, 2H), 5.68 (s, 1H), 2.61-2.50 (m, 2H), 2.50-2.39 (m, 2H), 2.31-2.15 (m, 3H), 2.15-2.06 (m, 2H), 2.06-1.93 (m, 3H), 1.90-1.60 (m, 3H) 1.40-1.30 (m, 1H), 1.30 (s, 3H), 0.70 (s, 3H);

[0122] 13C NMR (100 MHz, CDCh): 209.1, 199.5, 163.1, 140.5, 127.1, 123.9, 63.4, 49.8, 44.2, 39.6, 38.6, 37.7, 37.2, 35.5, 33.9, 29.7, 25.1, 22.5, 22.3, 20.5, 12.0.

[0123] LC-MS: [M+H]+= 316.35

[0124] Example 6: Preparation of Deuterated Dydrogesterone of formula (II) (8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0125] 1,2,8,9,10,11,12, 13, 14, 15, 16, 17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-17-d

[0126] Deuterated Dydrogesterone of formula (II)

[0127] Deuterated dydrogesterone of formula I (0.1 g, 0.31 mmoles) was dissolved in THF:D2O (4: 1; 5 mL) and K2CO3 (0.214 g, 1.55 mmol) (dried under vacuum before reaction) was added at room temperature under argon atmosphere. The mixture was stirred for 24 hours. After completion of the reaction as monitored by LCMS, concentrated under reduced pressure. The residue was diluted with toluene (5 mL) and concentrated under reduced pressure to afford Deuterated dydrogesterone of formula II after chromatographic purification.

[0128] LC-MS: [M+H]+= 316.48 Example 7: Preparation of Deuterated Dydrogesterone of formula (V) (8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl-

[0129] 1,2,8,9,10,11,12, 13, 14, 15, 16, 17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-2,2-d2

[0130] Deuterated Dydrogesterone of formula (V)

[0131] Deuterated dydrogesterone of formula I (0.1 g, 0.31 mmoles) was dissolved in THF:D2O (4: 1; 5 mL) and K2CO3 (0.214 g, 1.55 mmol) (dried under vacuum before reaction) was added at room temperature under argon atmosphere. The mixture was stirred for 24 hours. After completion of the reaction as monitored by LCMS, concentrated under reduced pressure. The residue was diluted with toluene (5 ml) and concentrated under reduced pressure to afford Deuterated dydrogesterone of formula V.

[0132] LC-MS: [M+H]+= 318.49

[0133] BIOLOGICAL ASSAYS: In-vitro stability of Dydrogesterone and Deuterated Dydrogesterone of formula (I) in mouse, rat, human, dog, and monkey liver microsomes.

[0134] The test compounds, Dydrogesterone and its deuterated versions were evaluated for their in-vitro stability in various species liver microsomes. A control using Verapamil was also performed to determine the microsomal stability along with test compounds. A. Mouse liver microsomal (MLM) stability i) Both the test compounds, as well as the control compound exhibit a high clearance in MLM. ii) The half-life of the compounds showed 8.68 for Dydrogesterone and 10.63 for Deuterated Dydrogesterone of formula (I). iii) The intrinsic clearance in protein was observed to be >48 pL / min / mg protein which leads to be a high clearance compound.

[0135] B. Rat Liver Microsomal (RLM) stability i) Both the test compounds, as well as the control compound exhibit a high clearance in RLM. ii) The intrinsic clearance for Dydrogesterone and Deuterated Dydrogesterone of formula (I) in protein was observed to be 62.26 and 37.21 pL / min / mg protein which leads to be a medium clearance compound.

[0136] C. Human Liver Microsomal (MLM) stability i) The half-life of the compounds showed 19.73 for Dydrogesterone and 56.87 for Deuterated Dydrogesterone of formula (I). ii) The intrinsic clearance for Dydrogesterone and Deuterated Dydrogesterone of formula (I) in protein was 70.25 and 24.40 pL / min / mg protein, which leads to be a high and medium clearance compounds, respectively.

[0137] D. Dog liver microsomal (DLM) stability i) Both the test compounds, as well as the control compound exhibit a high clearance, ii) The intrinsic clearance for Dydrogesterone and Deuterated Dydrogesterone of formula (I) in protein was observed to be 61.35 and 50.57 pL / min / mg protein which leads to be a high clearance compound. E. Cyano Monkey Liver (CLM) microsomal stability i) Both the test compounds, as well as the control compound exhibit a high clearance, ii) The half-life of the compounds showed <10 minutes.

[0138] 5 iii) The intrinsic clearance for Dydrogesterone and Deuterated Dydrogesterone of formula (I) in protein was observed to be high for both the compounds.

[0139] Table 1: Test results summary of In-vitro stability of Dydrogesterone and Deuterated Dydrogesterone of formula ( I) in mouse, rat, human, and monkey liver

[0140] 10 microsomes.

[0141] Summary: The invitro metabolic stability of test compounds Dydrogesterone and Deuterated Dydrogesterone of formula (I) was evaluated in liver microsomes from various species, including mouse, rat, human, dog, and cynomolgus monkey.

[0142] 15 Verapamil served as a control in all assays, demonstrating the expected high clearance rate. Based on half-life, intrinsic clearance, and hepatic blood flow, Dydrogesterone and Deuterated Dydrogesterone of formula (I) were classified as having high clearance, indicating both compounds are highly unstable in liver microsomes in the presence of a cofactor. Notably, Deuterated Dydrogesterone of

[0143] 20 formula (I) showed slightly greater stability than Dydrogesterone in rat, human, and dog liver microsomes. However, both compounds exhibited rapid clearance in mouse and monkey liver microsomes.

Claims

CLAIMS:1) Deuterated dydrogesterone of formula (I)Deuterated Dydrogesterone of formula (I) ((8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl- 1,2,8,9,10,11,12,13, 14, 15,16, 17-dodecahydro-3H- cy clopenta[a] phenanthren- 3 -one)2) Deuterated Dydrogesterone of formula (II)Deuterated Dydrogesterone of formula (II)(8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl- l,2,8,9,10,l l,12,13,14,15,16,17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-17-d3) Deuterated Dydrogesterone of formula (V)Deuterated Dydrogesterone of formula (V)(8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl- l,2,8,9,10,l l,12,13,14,15,16,17-dodecahydro-3H-cyclopenta[a]phenanthren-3- one-2,2-d24) A process for preparing deuterated dydrogesterone of formula (I) comprising reaction of dydrogesterone with bromine in presence of aqueous sodium hydroxide in an organic solvent to provide the conjugated ketone carboxylic acid (DYD-A), which was then treated with N,O-dimethyl hydroxylamine hydrochloride in presence of a base and in an organic solvent to provide Weinreb amide (DYD-B) which is then reduced with sodium borohydride in presence of CeCh to give an alcohol (DYD-C), which on further treatment with a deuterated Grignard reagent CDsMgl gives a deuterated alcohol (DYD-D) followed by oxidation with Dess-Martin periodinane reagent provides deuterated Dydrogesterone of formula (I).Deuterated Dydrogesterone formula (I)5) A process according to Claim 4, wherein the compound DYD-A is prepared by a process comprising of adding bromine to aqueous sodium hydroxide solution at -10 to 20°C, which is then added to dydrogesterone dissolved in a mixture of dioxane and water at -10 to 20°C to give DYD-A,6) A process according to Claim 4, wherein the compound DYD-B is prepared by a process comprising of dissolving DYD-A in an organic solvent, adding 3-(Dimethylaminopropyl)ethyl carbodiimide hydrochloride in presence ofDiisopropylethylamine and dimethylaminopyridine and treating with N-O- dimethyl hydroxylamine to give DYD-B,7) A process according to Claim 4, wherein the compound DYD-C is prepared by a process comprising of dissolving DYD-B in an organic solvent and adding NaBtC in presence of CeCh at -5 to +5°C to give DYD-C,8) A process according to Claim 4, wherein the compound DYD-D is prepared by a process comprising of dissolving DYD-C in an organic solvent, adding a Grignard reagent CDsMgl at -10 to +10°C to provide DYD-D,9) A process according to Claim 4, wherein deuterated dydrogesterone of formula (I) is obtained by a process comprising of dissolving DYD-D and oxidising with Dess-Martin periodinane reagent at room temperature and isolating deuterated dydrogesterone of formula (I).10) A process for preparing deuterated dydrogesterone of formulae (II) and (V) comprising treating deuterated dydrogesterone of formula (I) with D2O in the presence of an inorganic base in an organic solvent at ambient temperature.11) Deuterated dydrogesterone of formula (I) with improved half-life.Deuterated Dydrogesterone of formula (I) ((8S,9R,10S,13S,14S,17S)-17-(acetyl-d3)-10,13-dimethyl- 1,2,8,9,10,11,12,13, 14, 15,16, 17-dodecahydro-3H- cy clopenta[a] phenanthren- 3 -one)

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