WO2023112055A1

WO2023112055A1 - New synthetic process design for the synthesis of carboxy tormifene and purity analysis to strengthen antidoping testing

Info

Publication number: WO2023112055A1
Application number: PCT/IN2022/051083
Authority: WO
Inventors: Gangasani Jagadeesh Kumar; Sachin Dattram PAWAR; Pullapanthula RADHAKRISHNANAND; Upadhyayula Suryanarayana Murty; Puran Lal SAHU; Sachin DUBEY; Kapendra SAHU; Awanish UPADHYAY; Pramod Kumar
Original assignee: Director, National Institute Of Pharmaceutical Education And Research, Guwahati (Niper-G); Director, National Dope Testing Laboratory (Ndtl)
Priority date: 2021-12-15
Filing date: 2022-12-15
Publication date: 2023-06-22

Abstract

The present invention relates to a process for preparing a compound of formula I: I. The compound of formula I is useful in anti-doping tests.

Description

NEW SYNTHETIC PROCESS DESIGN FOR THE SYNTHESIS OF CARBOXY TORMIFENE AND PURITY ANALYSIS TO STRENGTHEN ANTIDOPING TESTING

RELATED APPLICATION

This application claims the benefit of Indian Complete Patent Application No. 202131058419 filed on December 15, 2021, which is hereby incorporated by reference in its entirety.

TECHNICAL FIELD

The disclosure generally relates to field of organic chemistry, particularly to process to prepare carboxy-toremifene.

BACKGROUND

Selective estrogen receptor modulators (SERMs) being widely treatment of the breast cancer. Currently two classes are approved for clinical use and development 1) triphenylethylene derivatives (tamoxifen and toremifene) and 2) benzothiophene example derivatives (raloxifene)¹. It is vivid from literature that structurally, toremifene differs from tamoxifen only by the substitution of a chloride ion for a hydrogen atom on the ethylene alkyl side-chain.² Abuse of SERMs have been prohibited ‘in’ and ‘out’ of competition since 2005 by the World Anti-Doping Agency (WADA), and they have been included in section S4 ‘agents with anti-oestrogenic activities’ in the WADA list of prohibited substances.^3,4 A total 11 metabolite have been observed in the literature for the toremifene. Toremifene metabolites include N-desmethyl toremifene (TOR Ml), hydroxytoremifene (TO M2a-b),N-hydroxymethyltoremifene (TOR M2c), N- desmethylhydroxytoremifene (TOR-M3a-b), dihydroxytoremifene (TOR M4a-c), N- hydroxymethylhydroxytoremifene (TOR M4d) and carboxy-toremifene (TOR M5).⁵ WADA has prohibited under their list 2021. An in-depth survey revealed that a reference material of carboxy toremifene is not available commercially Hence, it is very essential to have highly purified and characterized carboxy toremifene reference material which will be used in estimation of toremifene metabolite in urine sample during sport doping test.⁴

SUMMARY OF THE INVENTION Accordingly, in one aspect, the present disclosure provides a process to prepare carboxy- toremifene or its salt or stereoisomer thereof. Carboxy-toremifene structural formula is as follows:

I In an aspect, the present invention provides a process to prepare a compound of formula I, or a salt, a solvate or a stereoisomer thereof, comprising hydrolysis of a compound of formula 7 :

7 to provide a compound of formula I; wherein R1 is ester. In another aspect, the present invention provides a process to prepare a compound of formula

7, or a salt, a solvate or a stereoisomer thereof, comprising converting a compound of formula 5:

to a compound of formula 7 ; wherein X is -OR2 or halo; and R2 is hydrogen or a phenol protecting group.

In yet another aspect, the present invention provides a process to prepare a compound of formula 5, or a salt, a solvate or a stereoisomer thereof, comprising reacting a compound of formula 3:

with a compound of formula 4:

to provide compound of formula 5 ; wherein R1 is ester, X is -OR2 or halo; and R2 is hydrogen or alkyl. In yet another aspect, the present invention provides a compound of formula 7, or a salt, or a solvate, or a stereoisomer thereof:

wherein R1 is ester.

In yet another aspect, the present invention provides a compound of formula 5, or a salt, a solvate, or a stereoisomer thereof:

wherein X is -OR2 or halo; and R2 is hydrogen or a phenol protecting group.

BRIEF DESCRIPTION OF ACCOMPANYING DRAWINGS Figure 1 illustrates ¹ H NMR spectrum of carboxy toremifene.

Figure 2 illustrates HRMS (high-resolution mass spectrometry) spectrum of carboxy toremifene.

Figure 3 illustrates FTIR (fourier-transform infrared spectroscopy) spectrum of carboxy toremifene.

Figure 4 illustrates UV-visible spectrum of carboxy toremifene.

Figure 5 illustrates chromatogram of carboxy toremifene.

Figure 6 illustrates overlay chromatogram of carboxy toremifene.

Figure 7 illustrates linearity graph of carboxy toremifene.

Figure 8 illustrates control chart for accuracy (% bias data). DETAILED DESCRIPTION OF THE INVENTION

Before the methods of the present disclosure are described in greater detail, it is to be understood that the methods are not limited to particular embodiments described, as such may, of

5 course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the methods will be limited only by the appended claims.

Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the methods. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges and are also encompassed within the methods, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the methods.

Certain ranges are presented herein with numerical values being preceded by the term “about.” The term “about” is used herein to provide literal support for the exact number that it precedes, as well as a number that is near to or approximately the number that the term precedes. In determining whether a number is near to or approximately a specifically recited number, the near or approximating unrecited number may be a number which, in the context in which it is presented, provides the substantial equivalent of the specifically recited number.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, preferred methods and materials are described. For the purposes of the present invention, the following terms are defined below.

All publications cited in this specification are herein incorporated by reference as if each individual publication was specifically and individually indicated to be incorporated by reference and are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited. The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that the present methods are not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently confirmed.

It is appreciated that certain features of the methods, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the methods, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination. All combinations of the embodiments are specifically embraced by the present invention and are disclosed herein just as if each and every combination was individually and explicitly disclosed, to the extent that such combinations embrace operable processes and/or composites/scaffolds. In addition, all sub-combinations listed in the embodiments describing such variables are also specifically embraced by the present methods and are disclosed herein just as if each and every such sub-combination was individually and explicitly disclosed herein.

As will be apparent to those of skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has discrete components and features which may be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the present methods. Any recited method can be carried out in the order of events recited or in any other order which is logically possible.

The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

The term "alkyl" refers to a straight or branched chain saturated aliphatic hydrocarbon that may be substituted or unsubstituted. In certain embodiments, an alkyl group may have 1 to 20 (i.e., Ci -20 alkyl), 1 to 15 carbon atoms (i.e., Ci-15 alkyl), 1 to 10 carbon atoms (i.e., Ci-10 alkyl) or 1 to 6 carbon atoms (i.e., Ci-6 alkyl). In some instances, the alkyl is Ci-Ce alkyl. Examples of "alkyl" include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, n- pentyl, isobutyl and the like. The alkyl group may be optionally substituted. The term “alkenyl” alone or as part of another group refers to a hydrocarbon radical straight, branched or cyclic containing from 2 to 12 carbon atoms and at least one carbon to carbon double bond. The alkenyl group may be optionally substituted. Examples of "alkeyl" include, but are not limited to propenyl, allyl, methylallyl, butenyl, 2-butenyl, methylbutenyl, pentenyl, 3-methyl-2-pentenyl, hexenyl, and the like.

The term “alkylsulfonyl” refers to a group -S(O2)-alkyl, where alkyl is as defined above. Examples of "alkylsulfonyl" include, but are not limited to, methylsulfonyl, ethylsulfonyl, and propylsulfonyl. The alkynyl group may be futher substituted.

The term "alkoxy" refers to a group -O-alkyl, wherein alkyl is as defined above. Examples include, but are not limited to, methoxy, ethoxy, propoxy, t-butoxy and the like. The alkoxy group may be optionally substituted.

The term “alkoxyalkyl” refers to an alkyl group substituted with one or more alkoxy groups. Examples of “alkoxyalkyl” include, but are not limited to, methoxy methyl, methoxyethyl, 2-ethoxyethyl, tert-butoxy methyl, and the like. The alkoxyalkyl group may be optionally substituted.

The term "alkynyl" refers to an unsaturated hydrocarbon group which is linear or branched and has at least one carbon-carbon triple bond. In certain embodiments, an alkynyl group has 2 to 20 carbon atoms and in other embodiments, has 2 to 6 carbon atoms. An alkynyl group having 2 to 6 carbon atoms may be referred to as a (C2-C6) alkynyl group. The alkynyl group may contain 1, 2 or 3 carbon-carbon triple bonds, or more. In some embodiments, alkynyl groups contain one or two triple bonds, preferably one triple bond. In some instances, alkynyl moiety may be coupled to the remainder of the molecule through an alkyl linkage. Examples of alkynyl include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl or 3-butynyl, 2-pentynyl, 3- pentynyl, 2-hexynyl, 3-hexynyl and the like. The alkynyl group may be further substituted.

As used herein, the term "amino" refers to -NR⁴R⁵, wherein R⁴ and R⁵, independently are hydrogen, alkyl or aryl. The alkyl is same as defined above and the alkyl is same as defined below. The alkynyl group may be optionally substituted. The term "aryl" when used alone or in combination with other terms (such as arylalkyl, arylalkenyl, arylalkynyl, and aralkylalkoxy) refers to an optionally substituted unsaturated or partially saturated aromatic ring. The aryl may be monocyclic, bicyclic, polycyclic, bridged ring or fused ring system. In certain embodiments, aryl as used herein may have 6 to 50 ring carbon atoms (i.e., Ce-so aryl), 6 to 20 ring carbon atoms (i.e., Ce-20 aryl), or 6 to 12 carbon ring atoms (i.e., Ce-i2 aryl). “Optionally substituted aryl” refers to aryl or substituted aryl. Examples of "aryl" include, but are not limited to, phenyl, naphthyl, anthracenyl, azulenyl, indanyl, indenyl, fluorenyl, and biphenyl.

The term "aralkyl" refers to an alkyl group substituted by one or more aryl groups, wherein the alkyl and aryl are same as defined above. Non-limiting examples of the aralkyl group include phenylmethyl, phenylethyl, and the like. Examples of such groups include, but are not limited to, benzyl, 1 -phenylethyl, methylphenylethyl, phenylethyl, phenylpropyl, diphenylmethyl, triphenylmethyl, naphthylmethyl, naphthylethyl, 1,2,3,4-tetrahydronaphtharen-l-yl, pyridylmethyl, pyridylethyl, pyridylpropyl, pyridylbutyl, pyrrolylmethyl, furfuryl, thienylmethyl, triazolylmethyl, and the like. The arylalkyl group may be optionally substituted.

The term "arylalkenyl" refers to an alkenyl group substituted by one or more aryl groups, wherein alkenyl and aryl are same as defined above. The arylalkenyl group may be optionally substituted.

The term "arylalkynyl" refers to an alkynyl group substituted by one or more aryl groups, wherein alkynyl and aryl are same as defined above. The arylalkynyl group may be optionally substituted.

The term "aralkylalkoxy" refers to an alkoxy group substituted by one or more arylalkyl or aralkyl groups, wherein arylalkyl and alkoxy are same as defined above. The aralkylalkoxy group may be optionally substituted.

As used herein, the term “carboxyl” refers to -COOR, wherein R is hydrogen, alkyl or aryl.

As used herein, the term "comprises" or "comprising" is generally used in the sense of include, that is to say permitting the presence of one or more features or components. As used herein, the term “cyano” refers to -CN.

As used herein, the term "cycloalkyl" used herein, either alone or in combination with other radicals, denotes mono, bicyclic or polycyclic saturated, partially saturated hydrocarbon ring system of about 3 to 12 carbon atoms which may be substituted or unsubstituted. Exemplary "cycloalkyl" groups include but are not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl, perhydronapthyl, adamantyl, noradamantyl and spirobicyclic groups such as spiro (4,4)non-2-yl. The cycloalkyl group may be optionally substituted.

The term "ester" refers to -C(O)O-, -C(O)O-R^a-, -R^aC(O)O-R^b-, or -R^aC(O)O-, where O is not bound to hydrogen, and R^a and R^b can independently be selected from alkyl, alkenyl, alkynyl, aryl, aralkyl, heterocyclyl, heteroaryl, alkoxy, aryloxy, amino, amide, cycloalkyl, ether, formyl, haloalkyl, halogen, ketone, phosphate, sulfide, sulfinyl, sulfonyl, sulfonic acid and thioketone. In certain embodiments, the ester may be a cyclic ester, for example the carbon atom and R^a, the oxygen atom and R^b, or R^a and R^b may be joined to form a 3- to 12-membered ring. Esters include, but are not limited to, alkyl esters wherein at least one of R^a or R^b is alkyl, such as -alkyl- C(O)-O-, -C(O)-O-alkyl-, -alkyl-C(O)-O-alkyl-, etc. In certain embodiments, esters also include, but are not limited to, aryl or heteoraryl esters, e.g., wherein at least one of R^a or R^b is an aryl or a heteroaryl group, where aryl and heteroaryl are same as defined herein.

As used herein, the term “halo” or "halogen" refers to fluorine, chlorine, bromine or iodine.

As used herein, the term “haloalkyl” refers to alkyl moiety in which an alkyl hydrogen atom is replaced by one or more halo group. The alkyl and halo are same as defined above. Examples of hydroxyalkyl include, but are not limited to, fluoromethyl, chloromethyl, bromomethyl, iodomethyl, difluoromethyl, trifluoromethyl, chlorofluoromethyl, dichloromethyl, bromofluoromethyl, chlorodifluoromethyl, dichlorofluoromethyl, and the like. The haloalkyl group may be optionally substituted.

As used herein, the term "heteroaryl" refers to monocyclic aromatic ring systems or fused bicyclic aromatic ring systems comprising two or more aromatic rings. These heteroaryl rings contain one or more nitrogen, sulfur and/or oxygen atoms where N-oxides sulfur oxides and dioxides are permissible heteroatom substitutions. The term includes ring(s) optionally substituted with halogens, nitro, amino, alkoxy, alkyl sulfonyl amino, alkylcarbonylamino, carboxy, alkyl carbonyl, hydroxy, and alkyl. Examples of heteroaryl groups include furan, thiophene, pyrrole, imidazole, pyrazole, triazole, tetrazole, thiazole, oxazole, isoxazole, oxadiazole, thiadiazole, isothiazole, pyridine, pyridazine, pyrazine, pyrimidine, quinoline, isoquinoline, benzofuran, benzothiophene, indole, indazole, chromanyl, isochromanyl and the like. The heteroaryl group may be optionally substituted.

The term “heteroaralkyl” or “heteroarylalkyl” refers to an alkyl group substituted by one or more heteroaryl groups, wherein the alkyl and heteroaryl are same as defined above. Examples of heteroaralkyl include, but are not limited to, 4-methoxy-l-pyridin-3-ylmethyl, 2- pyridinylmethyl, 3-pyridinylmethyl, 4-pyridinylmethyl, 3-(2-pyridinyl)-propyl, and thienylmethyl, indolinylalkyl (such as 2-indolinylmethyl, 2-(3-indolinyl)ethyl, l-(4- indolinyl)ethyl, 3-(5-indolinyl)propyl, 4-(6-indolinyl)butyl, 5-(7-indolinyl)pentyl, 6-(l- indolinyl)hexyl, 2-methyl-3-(3-indolinyl)propyl, l,l-dimethyl-2-(2-indolinyl)-ethyl, 3,3- dimethyl-5-indolinylmethyl, 1 ,3,3-trimethyl-5-indolinylmethyl, 1 -ethyl-3,3-dimethyl-5- indolinylmethyl, l-methyl-5-indolinylmethyl, l,3-dimethyl-5-indolinylmethyl, or the like). The heteroarylalkyl group may be optionally substituted.

The term "heterocyclyl" refers to a stable 3 to 15 membered ring that is either saturated or has one or more degrees of unsaturation or unsaturated. These heterocyclic rings contain one or more heteroatoms selected from the group consisting of nitrogen, sulfur and oxygen where N- oxides, sulfur oxides and dioxides are permissible heteroatom substitutions. Such a ring is optionally fused to one or more of another heterocyclic ring(s), aryl ring(s) or cycloalkyl ring(s). Examples of such groups are selected from the group consisting of azetidinyl, acridinyl, pyrazolyl, imidazolyl, triazolyl, pyrrolyl, thiophenyl, thiazolyl, oxazolyl, isoxazolyl, furanyl, pyrazinyl, tetrahydroisoquinolinyl, piperidinyl, piperazinyl, morpholinyl, thiomorphonilyl, pyridazinyl, indolyl, isoindolyl, quinolinyl, chromanyl and the likes thereof. "Heterocyclylalkyl" refers to a heterocyclic ring radical defined above, directly bonded to an alkyl group. The heterocyclylalkyl radical is attached to the main structure at carbon atom in the alkyl group that results in the creation of a stable structure. The heterocyclyl group may be optionally substituted. The term “heterocyclylalkyl” used herein refers to one or more heterocyclyl groups appended to an alkyl radical. Examples of heterocyclylalkyl include, but are not limited to, piperidinylmethyl, piperidinylethyl, morpholinylmethyl, morpholinylethyl, and the like. The heterocyclylalkyl group may be optionally substituted.

As used herein, the term “hydroxy” or "hydroxyl" refers to -OH group.

As used herein, the term "nitro" refers to -NO2 group.

As used herein, the terms "optional" or "optionally" mean that the subsequently described event or circumstance may occur or may not occur, and that the description includes instances where the event or circumstance occurs as well as instances in which it does not. For example, "optionally substituted alkyl" refers to the alkyl may be substituted as well as the event or circumstance where the alkyl is not substituted.

As used herein, the term "oxo" refers to =0 group.

Unless otherwise specified, the term "substituted" as used herein refers to mono, bi, tri or tetra substitution with any one or more combination of the following substituents: hydroxy, halogen, carboxyl, cyano, nitro, oxo (=0), thio (=S), substituted or unsubstituted alkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted cycloalkenyl, substituted or unsubstituted amino, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heterocyclylalkyl ring, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted heterocyclic ring, substituted or unsubstiuted guanidine, -C00R⁶, -C(O) R⁶, - C(S)R⁶, -C(O)N(R⁷)R⁸, -C(O)ON(R⁷)R⁸, -NR⁶CO(R⁷)R⁸, -N(R⁶)SOR⁷, -N(R⁶)SO₂R⁷, -(=N- N(R⁷)R⁸), -NR⁶C(O)OR⁷, -NR⁷R⁸, -NR⁶C(O)R⁷, - NR⁶C(S)R⁷, -NR⁶C(S)NR⁷R⁸, -SONR⁷R⁸, - SO₂NR⁷R⁸, -OR⁶, -OR⁶C(O)NR⁷R⁸, - OR⁶C(O)OR⁷, -0C(0)R⁶, -OC(O)NR⁷R⁸, -R⁶NR⁷C(O)R⁸, - R⁶OR⁷, -R⁶C(O)OR⁷, -R⁶C(O)NR⁷R⁸, -R⁶C(O)R⁷, -R⁶OC(O)R⁷, -SR⁶, -SOR⁶, -SO₂R⁶, and - ONO₂, wherein R⁶, R⁷ and R⁸ are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted cycloalkenyl, substituted or unsubstituted amino, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted heterocyclylalkyl ring, substituted or unsubstituted heteroarylalkyl, or substituted or unsubstituted heterocyclic ring, or substituted or unsubstituted alkylsulfonyl. Alternatively, R⁷ and R⁸ together with the nitrogen they are attached with, form a 4 to 8 membered ring which can be substituted or unsubstituted. In certain embodiments, the substituents in the aforementioned "substituted" groups can be further substituted. In some instances, the substituents in the aforementioned "substituted" groups cannot be further substituted. For example, when the substituent on "substituted alkyl" is "substituted aryl" the substituent on "substituted aryl" cannot be "substituted alkenyl".

Each embodiment is provided by way of explanation of the invention and not by way of limitation of the invention. In fact, it will be apparent to those skilled in the art that various modifications and variations can be made to the compounds, compositions and methods described herein without departing from the scope or spirit of the invention. For instance, features illustrated or described as part of one embodiment can be applied to another embodiment to yield a still further embodiment. Thus, it is intended that the present invention includes such modifications and variations and their equivalents. Other objects, features and aspects of the present invention are disclosed in or are obvious from, the following detailed description. It is to be understood by one of ordinary skill in the art that the present discussion is a description of exemplary embodiments only and is not to be construed as limiting the broader aspects of the present invention.

In an embodiment, the present disclosure provides a process to prepare carboxy- toremifene.

In certain embodiments, the present disclosure provides a process to prepare a compound of formula I, or a salt, or a stereoisomer thereof:

I comprising hydrolysis of a compound of formula 7 :

7 to provide a compound of formula I; wherein R1 is ester.

In a further embodiment, the compound of formula 7 is hydrolyzed using a hydrolyzing agent in the presence of a solvent. In some further embodiments, the hydrolyzing agent is an acid or a base. That means, the hydrolysis may be carried out in an acid or base system. That is, the hydrolysis may be an acidic hydrolysis, or may be an alkaline hydrolysis. In certain embodiments, any acid or base suitable for hydrolysis can be used in the reaction, for example: the acid may be HC1, H2 SO4 , H3 PO4 or AcOH.

In certain embodiments, alkali hydrolysis is carried out in the presence of an alkaline metal hydroxide, an alkaline earth metal hydroxide, an alkaline metal carbonate, an alkaline earth metal carbonate or any combination thereof. In certain embodiments, alkaline metal hydroxide is selected from the group comprising lithium hydroxide, sodium hydroxide, potassium hydroxide, rubidium hydroxide, cesium hydroxide, and any combination thereof. The alkaline earth metal hydroxide is selected from the group comprising beryllium hydroxide, magnesium hydroxide, calcium hydroxide, strontium hydroxide, barium hydroxide, and any combination thereof. The alkaline metal carbonate includes, but not limited to, lithium carbonate, sodium carbonate and potassium carbonate; The alkaline earth metal carbonate includes, but not limited to, magnesium carbonate, calcium carbonate and barium carbonate.

In some embodiments, any solvent suitable for hydrolysis can be used in the reaction. In some instances, the solvent used in the hydrolysis is water, alcohol (such as methanol, ethanol, isopropanol, tert-butanol and a mixture), DCM, DMF, DMSO, THF, or a mixture thereof.

In certain embodiments, the alkali hydrolysis is carried out at an ambient temperature or higher temperature. In some instances, the alkali hydrolysis may be carried out at a reflux temperature of the solvent used. In some embodiments, the reaction may be carried out at a temperature from about 0-100°C. In further embodiments, the hydrolysis temperature can be about 10-100°C, about 10-90°C, about 10-80°C, about 10-70°C, about 10-60°C, about 20-100°C, about 20-90°C, about 20-80°C, or about 20-70°C. After completion of the reaction, the reaction mixture may be concentrated to get a compound of formula I. The product obtained may be further purified.

In certain embodiments, the present disclosure provides a process to prepare a compound of formula 7:

comprising converting a compound of formula 5:

5 to a compound of formula 7 ; wherein X is -OR2 or halo; and R2 is hydrogen or a phenol protecting group.

In a further embodiment, the compound of formula 5 is converted to a compound of formula 7 in the presence of a solvent. Any solvent suitable for reaction can be used in the reaction.

In certain embodiments, the ester in the compound of formula 7 is -COOR’, and R’ is alkyl, aryl, or aralkyl. In some instances, R’ is alkyl.

In a further embodiment, X, in the compound of formula 5, is hydroxyl or halo. In some instances, X is hydroxyl.

In a further embodiment, the compound of formula 5 is converted to a compound of formula 7 using a base in the presence of a solvent. Any base and/or solvent suitable for reaction can be used in the reaction.

In some embodiments, the compound of formula 7 is methyl 4-(4-(2- (dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoate. In some instances, methyl 4-(4-(2- (dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoate is obtained by reacting methyl 4-(4- hydroxyphenyl)-3,4-diphenylbut-3-enoate with 2-chloro-N,N-dimethylethan-l -amine hydrochloride in presence of a base to yield methyl 4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4- diphenylbut-3-enoate. Any base suitable for reaction can be used in the reaction. In some embodiments, the base is a metal carbonate or a metal hydride. In some instances, the metal carbonate is selected from the group comprising Li2CO3, Na2CO3 , K2CO3 and CS2CO3. The metal hydride is NaH or KH. The solvent is selected from the group comprising acetone, ether, THF (tetrahydrofuran), DMF (dimethylformamide), DCM (dichloromethane), ACN (acetonitrile) and any combination thereof.

In certain embodiments, the compound of formula 5 is converted to a compound of formula 7 at an ambient temperature or higher temperature. In some instances, the conversion of 5 to 7 may be carried out at a reflux temperature of the solvent used. In some embodiments, the reaction may be carried out at a temperature from about 0-150°C. In further embodiments, the hydrolysis temperature can be about 10-100°C, about 10-90°C, about 10-80°C, about 10-70°C, about 10-60°C, about 20-100°C, about 20-90°C, about 20-80°C, or about 20-70°C. After completion of the reaction, the reaction mixture may be concentrated to get a compound of formula 7. The product obtained may be directly used in the next step without further purification. In some instances, the product is purified before using in the next step.

In certain embodiments, the present disclosure provides a process to prepare a compound of formula 5:

comprising reacting a compound of formula 3:

with a compound of formula 4:

to provide compound of formula 5 ; wherein R1 is ester, X is -OR2 or halo; and R2 is hydrogen or alkyl.

In a further embodiment, the compound of formula 5 is prepared by reacting a compound of formula 3 with a compound of formula 4 in presence of titanium chloride and a reducing agent, and in a solvent. In some embodiments, titanium chloride is Ti CI3 or TiCU. Any reducing agent and/or solvent suitable for reaction can be used in the reaction. In certain embodiments, the reducing agent is an alkali metal, a metal hydride, zinc dust, zinc-copper couple, zinc-silver couple, magnesium, or magnesium-mercury amalgam. In certain embodiments, the solvent is selected from the comprising tetrahydrofuran, dimethoxye thane, and any combination thereof.

In certain embodiments, the reaction of compound of formula 3 with a compound of formula 4 may be carried out at an ambient temperature or higher temperature. In some instances, the reaction may be carried out at a reflux temperature of the solvent used. In some embodiments, the reaction may be carried out at a temperature from about 0-150°C. In further embodiments, the hydrolysis temperature can be about 10-100°C, about 10-90°C, about 10-80°C, about 10-70°C, about 10-60°C, about 20-100°C, about 20-90°C, about 20-80°C, or about 20-70°C. After completion of the reaction, the reaction mixture may be concentrated to get a compound of formula 5. The product obtained may be directly used in the next step without further purification. In some instances, the product is purified before using in the next step.

In certain embodiments, the compound of formula 3 is prepared by reacting acetophenone with a carbonate 2:

2 wherein each R3 is independently alkyl, aryl, or aralkyl. In a further embodiment, the compound of formula 3 is prepared by reacting acetophenone with a carbonate 2 in presence of base in a solvent. Any base and/or solvent suitable for reaction can be used in the reaction. In some embodiments, the base is selected from a group comprising a metal carbonate or a metal hydride, DIPEA (N,N-diisopropylethylamine), triethylamine. In some instances, the metal carbonate is selected from the group comprising Li2CC>3, Na2COs, K2CO3 and CS2CO3. The metal hydride is NaH or KH. The solvent is selected from the group comprising THF, acetone, ACN, DMF, DMSO, DCM, and any combination thereof.

In certain embodiments, the reaction of acetophenone with the carbonate 2 may be carried out at an ambient temperature or higher temperature. In some instances, the reaction may be carried out at a reflux temperature of the solvent used. In some embodiments, the reaction may be carried out at a temperature from about 0-150°C. In further embodiments, the hydrolysis temperature can be about 10-100°C, about 10-90°C, about 10-80°C, about 10-70°C, about 10- 60°C, about 20-100°C, about 20-90°C, about 20-80°C, or about 20-70°C. After completion of the reaction, the product obtained may optionally be purified using any suitable purification technique, such as, for example, adsorption, and/or extraction, chromatographic technique, recrystallization, filtration, concentration, etc.

The invention will now be illustrated by means of the following examples, it being understood that these are intended to explain the invention, and in no way to limit its scope.

The schemes are given for the purpose of illustrating the invention and are not intended to limit the scope or spirit of the invention. Starting materials shown in the schemes can be obtained from commercial sources or prepared based on procedures described in the literature. Furthermore, in the following schemes, where specific acids, bases, reagents, coupling agents, solvents, etc. are mentioned, it is understood that other suitable acids, bases, reagents, coupling agents etc. may be used and are included within the scope of the present invention. Modifications to reaction conditions, for example, temperature, duration of the reaction or combinations thereof, are envisioned as part of the present invention. All possible stereoisomers are envisioned within the scope of this invention. Scheme I:

The following examples are given by way of illustration of the working of the invention in actual practice and therefore should not be construed to limit the scope of present invention. EXAMPLE 1: SYNTHESIS OF CARBOXY-TOREMIFENE

Carboxy-toremifene was synthesized in four steps as shown in scheme I. In the first step methyl- 3-oxo-3-phenylpropanoate (3) was synthesized by reacting the acetophenone (1, 10 mmol) with dimethyl carbonate (2, 30 mmol) in presence of base sodium hydride (60%, 20 mmol) and solvent THF for 2 h. The reaction mixture was refluxed until TLC indicated the total consumption of the ketones. After cooling, the reaction mixture was quenched by ice water, acidified with 3M HC1 to pH 2-3, and extracted with EtO c (100 ml. x 3). The product was isolated and purified in quantitative yield.

In the second step methyl 4-(4-hydroxyphenyl)-3,4-diphenylbut-3-enoate (5) was synthesized by reacting the methyl-3-oxo-3-phenylpropanoate (3, 3 mmol) with (4- hydroxyphenyl)(phenyl)methanone (4, 1 mmol) in presence of TiClr (4 mmol), Zn (8 mmol) and solvent THF at reflux temperature for 8 h. After completion of the reaction, monitored by TLC, the reaction mixture was quenched with 10% Na2COs, the product was isolated and purified by column chromatography.

In the third step methyl 4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoate (7) was synthesized by reacting the methyl 4-(4-hydroxyphenyl)-3,4-diphenylbut-3-enoate (5, 1 mmol) with 2-chloro-A, /V-di methy lethan- 1 -amine hydrochloride (6, 2 mmol) in presence of base K2CO3 and solvent acetone at reflux temperature for 12 h. The obtained product was isolated and purified by column chromatography.

In the final step, 4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoic acid (8) was synthesized by reacting 4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoate (7, 0.5 mmol) with 15% NaOH (3 ml) in MeOH at reflux condition for 1 h. The obtained product carboxy-toremifene (8) was purified by column chromatography and characterized by

NMR (Figure 1), and ESLHRMS (Figure 2).

Carboxy toremifene was characterized using

NMR and ¹³C NMR, HRMS, and FTIR. spectra were recorded on Bruker 600MHz and 150MHz spectrometers, respectively, in methanol-d4, tetramethyl silane as an internal reference standard. The data are presented as, chemical shift (5), coupling constant J in hertz (Hz). Carboxy toremifene was dissolved in the LC-MS grade methanol and directly infused to the mass detector via autosampler to know the molecular weight of the developed reference material using LC-QTOF-ESI.MS (Model: Agilant 654B LC-QTOF, make: Agilant) and FT-IR was used to determine the functional group of carboxy toremifene. The mixture was compressed using a hydraulic press and converted to KBr thin transparent pellet. This pellet was scanned for the IR region using FT-IR spectrophotometer (Model- ALPHA II, make: Bruker) and functional groups of carboxy toremifene determined.

HPLC method development, optimization, and validation

Preparation of stock solution and working standard

One mg/ml solution of carboxy toremifene was prepared by dissolving Img in 1 mL of Methanol followed by sonicate for some time till all solid dissolved. Stock solution was diluted up to 500pg/ml and filtered through 0.22u syringe filter and injected to HPLC for further analysis. Chromatographic conditions for carboxy toremifene

Chromatographic separation was achieved using Hypersil GOLD™ C8( 150x4.6 mm, 5um) column at the flow rate of 0.5 mL per minute. Injection volume was kept 5pL. Mobile phase was used as 0.2% Formic acid in water and 0.2% Formic acid in methanol {30:70} Column temperature and wavelength were fixed at 30°C and 236/ 278 nm, respectively. The data acquisition was made with control panel Agilent Software. Mobile phase was filtered and sonicated before HPLC analysis.

Purity assessment of carboxy toremifene

The sample purity value was calculated from six replicates of same concentration and mean of relative % peak area was calculated using control panel software.

Results and Discussion

Carboxy toremifene was synthesized and characterized. The isolated product was characterized by ¹ H NMR and ESLHRMS.

Characterization of carboxy toremifene

'H NMR (400 MHz, CDC1₃): 5 7.38 (2H d, J = 8.0 Hz), 7.32 (2H t, J = 7.3 Hz), 7.27-7.24 (1H, m), 7.02 (2H d, J = 8.3 Hz) 6.93 - 6.88 (1H m), 6.84 (4H dd, J = 10.8, 8.0 Hz), 6.57 (2H d, J = 10.1 Hz), 3.97 (2H t), 3.52 (3H s), 2.95 (2H t), 2.38 (6H s); ESLHRMS (M+H): 402.2071. (Figure 1)

HRMS: Q-Exactive Orbitrap high resolution full scan mass spectrum of carboxy toremifene has been observed molecular ion peak (M+H) at 402. 2076, as shown in figure 2.

FT-IR Spectroscopy: FT-IR was used to determine the functional group of carboxy toremifene. The sample was taken with KBr in the ratio of (1:100) and ground well to make a homogenous mixture using ceramic mortar and pestle and result shows that N-H stretching at wavenumber 3425, C-H stretching at 2917 and N-H bending at 1704 wavenumber as shown in figure 3. UV spectroscopy: Maximum absorbance from the obtained UV spectrum for the sample dissolved in methanol using UV spectroscopy was found at wavelength 236 nm and 276 nm as shown in supporting figure 4.

RP-HPLC-DAD method development and optimization for carboxy toremifene Optimized chromatographic conditions have been mentioned in Table 1. Retention time of carboxy toremifene was found to be at 3.7 minutes and it passes all the system suitability parameters, as shown in Tablet. Tailing factor was less than 2 and theoretical plate were observed more than 2000. It indicates that the developed HPLC method passes the system suitability parameters. (Figure 5 and Figure 6) Table 1. HPLC parameters and conditions for carboxy toremifene

Validation of developed HPLC method: Validation of the developed method was carried out as per ICH guidelines.

System suitability: To demonstrate acceptable method performance, different system suitability parameters like theoretical plate, and asymmetric factor should be in limit as shown in table 2. Mean, standard deviation and %RSD were calculated, and values were observed to be less than 2. Table 2: System suitability Parameters

Linearity: Carboxy toremifene linearity curve performed on HPLC is shown in Figure 7. The coefficient of determination (R²), slope, intercept was observed at 0.998, 40.468, 90.68, respectively. The overlay chromatograms of linearity were shown in the Figure 6. It was observed that linearity was found in the concentration window of 35 to 60pg/mL. It may be concluded that developed method will produce the linear response in this range and can be used for routine quantification of carboxy toremifene.

Accuracy: The percentage accuracy for carboxy toremifene was shown good percentage accuracy at each point and percent accuracy was found to be within limit as shown in the Table 3. Control chart is utilized to track and trend the accuracy results as shown in Figure 8. UCL and LCL refer to upper control limit and lower control limit, respectively. Good recovery indicates more accuracy towards the developed method.

Table 3: Spiked sample recovery

Precision: Precision was calculated by studying of six replicates of same concentrations at different intervals on the same day (intraday) and different days (inter day) and their mean, SD and %RSD were calculated and mentioned in Table 4. It was observed that % RSD was below 2%. It may be concluded that the developed method will provide more precise results for routine quality control sample.

Table 4

Robustness: Robustness was performed by changing the flow rate, injection volume and column temperature. It may be concluded that developed method is highly robust even after few deviations. Method robustness has been reported in table 5. Table 5: Results of robustness data

Purity assessment of carboxy toremifene: Developed carboxy toremifene reference material purity was found to be 100% as per the HPLC data.

References

1. Mazzarino, M., Fiacco, I., De La Torre, X. & Botre, F. A mass spectrometric approach for the study of the metabolism of clomiphene, tamoxifen and toremifene by liquid chromatography time-of-flight spectroscopy. Eur. J. Mass Spectrom. 14, 171-180 (2008).

2. Webster, L. K., Crinis, N. A., Stokes, K. H. & Bishop, J. F. High-performance liquid chromatographic method for the determination of toremifene and its major human metabolites. J. Chromatogr. B Biomed. Sci. Appl. 565, 482-487 (1991). 3. Kwok, K. Y., Chan, G. H. M., Kwok, W. H., Wong, J. K. Y. & Wan, T. S. M. In vitro phase I metabolism of selective estrogen receptor modulators in horse using ultra-high performance liquid chromatography-high resolution mass spectrometry. Drug Test. Anal. 9, 1349-1362 (2017). 4. World- Anti Doping Agency. The 2021 WADA Prohibited List. World Anti-Doping 1-24

(2021).

5. Kwok, K. Y., Chan, G. H. M., Kwok, W. H., Wong, J. K. Y. & Wan, T. S. M. In vitro phase I metabolism of selective estrogen receptor modulators in horse using ultra-high performance liquid chromatography-high resolution mass spectrometry. Drug Test. Anal. 9, 1349-1362 (2017).

The Claims:

1. A process to prepare a compound of formula I, or a salt, or a solvate, or a stereoisomer thereof:

comprising hydrolysis of a compound of formula 7 :

to provide a compound of formula I; wherein R1 is ester. 2. A process to prepare a compound of formula 7 or a salt, or a solvate, or a stereoisomer thereof:

comprising converting a compound of formula 5:

5 to a compound of formula 7 ; wherein X is -OR2 or halo; and

R2 is hydrogen or a phenol protecting group.

3. A process to prepare a compound of formula 5, or a salt, or a solvate, or a stereoisomer thereof:

comprising reacting a compound of formula 3:

with a compound of formula 4:

4. The process as claimed in claim 3, wherein the compound of formula 3 is prepared by reacting acetophenone with a carbonate 2:

wherein each R3 is independently alkyl, aryl, or aralkyl.

5. The process as claimed in any one of the claims 1 to 4, wherein ester is -COOR’, and R’ is alkyl, aryl, or aralkyl.

6. The process as claimed in any one of the claims 1 to 5, wherein R’ is alkyl.

7. The process as claimed in any one of the claims 1 to 6, wherein X is hydroxyl or halo.

8. The process as claimed in any one of the claims 1 to 7, wherein R3 is alkyl.

9. The process as claimed in claim 3, wherein the process is carried out in presence of titanium chloride and a reducing agent, and in a solvent.

10. The process as claimed in claim 9, wherein titanium chloride is Ti CI3 or TiCU.

11. The process as claimed in claim 9, wherein the reducing agent is an alkali metal, a metal hydride, zinc dust, zinc-copper couple, zinc-silver couple, magnesium, or magnesium-mercury amalgam.

12. The process as claimed in claim 9, wherein the solvent is tetrahydrofuran, dimethoxyethane, or a combination thereof.

13. The process as claimed in claim 2, comprises reacting methyl 4-(4-hydroxyphenyl)-3,4- diphenylbut-3-enoate with 2-chloro-N,N-dimethylethan-l -amine hydrochloride in presence of a base to yield methyl 4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoate. 14. A compound of formula 7, or a salt, or a stereoisomer thereof:

7; wherein R1 is ester. 15. A compound of formula 5, or a salt, or a stereoisomer thereof:

wherein X is -OR2 or halo; and R2 is hydrogen or a phenol protecting group.

ABSTRACT

The present invention relates to a process for preparing a compound of formula I:

The compound of formula I is useful in anti-doping tests.

Claims

The Claims:

comprising hydrolysis of a compound of formula 7 :

to provide a compound of formula I; wherein R1 is ester.

2. A process to prepare a compound of formula 7 or a salt, or a solvate, or a stereoisomer thereof:

comprising converting a compound of formula 5:

5 to a compound of formula 7 ; wherein X is -OR2 or halo; and

R2 is hydrogen or a phenol protecting group.

comprising reacting a compound of formula 3:

with a compound of formula 4:

wherein each R3 is independently alkyl, aryl, or aralkyl.

8. The process as claimed in any one of the claims 1 to 7, wherein R3 is alkyl.

13. The process as claimed in claim 2, comprises reacting methyl 4-(4-hydroxyphenyl)-3,4- diphenylbut-3-enoate with 2-chloro-N,N-dimethylethan-l -amine hydrochloride in presence of a base to yield methyl 4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-enoate.

14. A compound of formula 7, or a salt, or a stereoisomer thereof:

7; wherein R1 is ester.

15. A compound of formula 5, or a salt, or a stereoisomer thereof:

wherein X is -OR2 or halo; and R2 is hydrogen or a phenol protecting group.