Asymmetric synthesis of compounds and use in the treatment of diseases

Abstract

To provide methods of treating diseases.SOLUTION: The present application discloses, among other things, asymmetric synthesis of a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof. Also provided are methods and compositions for treatment of estrogen deficiency and preventing or mitigating an estrogen deficiency symptom using a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof alone or in combination with at least one additional agent. Further provided are methods and compositions for mitigating a side effect of an additional agent in the context of combination therapy with a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof.SELECTED DRAWING: Figure 6

Description

Technical Field

[0001] Technical Field The present invention relates to the field of asymmetric synthesis of steroid-like compounds, and more particularly, to a method for preparing chiral compounds of anordrin and its analogs. The present invention also relates to the therapeutic use of chiral compounds in the replacement treatment of estrogen deficiency symptoms.

Background Art

[0002] Background A decrease in estrogen production in women with ovariectomy (OVX), postmenopause, or anti-estrogen therapy leads to estrogen deficiency symptoms that can have an adverse impact on the quality of life over decades. Since the 1940s, estrogen replacement therapy (ERT) has been utilized to treat these symptoms.

[0003] Estrogen binds to its receptors in many tissues during mammalian reproduction and development, regulating RNA transcription, stimulating cell proliferation, and modulating metabolic signaling. Three genes for estrogen-binding proteins have been identified, and these genes encode estrogen receptors (ER) α and β, as well as G protein-coupled estrogen receptor 1 (GPER1). ER-α and β have similar structural and functional domains, which include activation function domain 1 (AF-1), DNA-binding domain (DBD), dimerization domain, and activation function domain 2 (AF-2), which is the ligand-binding domain (LBD). Both of them belong to the nuclear superfamily of ligand-dependent transcription factors and have highly conserved DBD and LBD regions. Upon ligand binding, they regulate RNA transcription to yield a ligand-receptor complex, which can dimerize and translocate into the nucleus, where it binds to estrogen response elements (EREs) found in the promoters of estrogen-responsive genes. This type of modulation is typically referred to as the classical estrogen pathway. ER-α and β also regulate diverse biological functions through membrane-initiated estrogen signaling (MIES) and bind to the plasma membrane through their interaction with the ligand-binding domain. The detailed molecular mechanism of signal transduction by membrane-bound ER remains unclear. The modulation effects of estrogen mediated by membrane-bound receptors on cell proliferation, matrix / migration, metabolism, and glucose homeostasis have been outlined (1, 2). Furthermore, studies on ER knockout mice have shown that ER-α is the estrogen receptor that functions predominantly compared to ER-β. Three transcriptional variants (66, 46, and 36) of ER-α have been found. ER-α36 lacks the AF-1 domain and contains a partial ligand-binding domain. It has been found to be localized in the cell membrane and cytosol.ER-α-36 is restricted in modulating MIES and has been found to be constitutively expressed in tamoxifen-resistant cancer cells (such as MDA-MB-231 and Hec1A). Therefore, MIES modulated by membrane-bound ER is considered to be responsible for the resistance to anti-estrogen therapy found by some researchers (3, 4).

[0004] However, studies showing that an increased risk of breast and uterine cancers and the incidence of thromboembolism are associated with ERT have led to a decrease in its use. Administration of a combination of estrogen and progesterone prevents the risks of breast and uterine cancers but causes side effects of progesterone such as dizziness, nausea, vomiting, fatigue, anxiety, depression, and headache. Menopausal symptoms still remain a problem for many elderly women. Therefore, there still remains a continuing need for the development of new replacement therapies for estrogen deficiency symptoms. SUMMARY OF THE INVENTION MEANS FOR SOLVING THE PROBLEM

[0005] SUMMARY OF THE INVENTION In one aspect, the present application provides a method for synthesizing a diastereomeric compound of formula (I) (for example, α-anordrin) or a salt thereof. According to various embodiments described herein, the diastereomeric compound of formula (I) (for example, α-anordrin) or a salt thereof is substantially pure.

[0006] In another aspect, a method of treating estrogen deficiency in an individual or preventing or alleviating estrogen deficiency symptoms including OVX or postmenopausal symptoms, the method comprising administering to the individual: a) an effective amount of a diastereomeric compound of formula (I) (e.g., α-norandrolone) or a salt thereof; and optionally b) an effective amount of at least one additional agent selected from the group consisting of a selective estrogen receptor modulator and an aromatase inhibitor. In some embodiments, the additional agent is a selective estrogen receptor modulator (SERM) selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene. In some embodiments, the additional agent is an aromatase inhibitor selected from the group consisting of anastrozole, letrozole, exemestane, vorozole, formestane, and fadrozole. In some embodiments, the estrogen deficiency symptoms are selected from the group consisting of high liver triglycerides, osteoporosis, vulvovaginal atrophy, high blood triglycerides, high blood glucose, and weight gain.

[0007] In some embodiments, a method for reducing the side effects of at least one additional agent with a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof, the method comprising administering to an individual, in combination with the additional agent, an effective amount of a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof, wherein the additional agent is selected from the group consisting of selective estrogen receptor modulators and aromatase inhibitors, is provided. In some embodiments, the additional agent is tamoxifen. In some embodiments, the additional agent is a selective estrogen receptor modulator (SERM) selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene. In some embodiments, the additional agent is an aromatase inhibitor selected from the group consisting of anastrozole, letrozole, exemestane, vorozole, formestane, and fadrozole.

[0008] In some embodiments, the diastereomeric compound of formula (I) or a salt thereof and the additional agent are administered sequentially. In some embodiments, the diastereomeric compound of formula (I) or a salt thereof and the additional agent are administered simultaneously.

[0009] In some embodiments, the individual is a human.

[0010] In yet another aspect, a pharmaceutical composition comprising a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof and at least one additional agent selected from the group consisting of selective estrogen receptor modulators (SERMs) and aromatase inhibitors is provided. In some embodiments, the additional agent is tamoxifen. In some embodiments, the additional agent is raloxifene or a functional equivalent thereof (e.g., including raloxifene, lasofoxifene, or bazedoxifene). In some embodiments, the additional agent is an aromatase inhibitor, such as anastrozole or is its functional equivalent.

[0011] In some embodiments, the weight ratio of the diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof in the composition to a further drug is from about 1:20 to about 20:1 (e.g., including from about 10:1 to about 1:10 or from about 1:10 to about 1:15).

[0012] The pharmaceutical compositions of the present invention may be present in unit dosage forms, such as oral unit dosage forms, such as capsules, tablets, pills, caplets, gels, liquids (e.g., suspensions, solutions, emulsions), powders or other particulate substances.

[0013] Also provided is a method of using the pharmaceutical compositions described herein to treat estrogen deficiency or to prevent or alleviate estrogen deficiency symptoms including OVX or postmenopausal symptoms, as described herein.

[0014] These and other aspects and advantages of the present invention will become apparent from the detailed description hereinafter and the appended claims. It should be understood that other embodiments of the present invention may be formed by combining one, some or all of the characteristics of the various embodiments described herein.

Brief Description of the Drawings

[0015]

Figure 1

[0016]

Figure 2A

Figure 2B

[0017]

Figure 3A

Figure 3B

Figure 3C

Figure 3D

Figure 3E

[0018]

Figure 4A

Figure 4B

Figure 4C

[0019]

Figure 5A

Figure 5B

Figure 5C

Figure 5D

[0020]

Figure 6

[0021]

Figure 7

[0022]

Figure 8A

Figure 8B

[0023]

Figure 9

[0024]

Figure 10

[0025]

Figure 11

Mode for Carrying Out the Invention

[0026] Detailed Description of the Invention This application provides a method for synthesizing a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof. Also provided is the use of a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof in the treatment of estrogen deficiency in an individual, or in the prevention or alleviation of estrogen deficiency symptoms including OVX or postmenopausal symptoms. In addition, provided is a composition for combination therapy comprising administering a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof in combination with a further agent for the treatment of estrogen deficiency, reduction of side effects, or prevention or alleviation of estrogen deficiency symptoms including OVX or postmenopausal symptoms such as high liver triglycerides, osteoporosis, vulvovaginal atrophy, high blood triglycerides, high blood glucose, and weight gain.

[0027] The present invention is based on the discovery of the unique properties and mechanism of action of diastereomeric compounds of formula (I) (e.g., α-anordrin) or salts thereof. In the present application, it has been recognized that the diastereomeric compounds of formula (I) (e.g., α-anordrin) or salts thereof are more active selective estrogen receptor modulators of membrane-bound estrogen-binding protein compared to other diastereomeric compounds of formula (I) (e.g., β-anordrin) or salts thereof. Beneficial effects of the diastereomeric compounds of formula (I) (e.g., α-anordrin) or salts thereof include i) modulation of estrogen metabolic effects as an agonist leading to prevention or reduction of estrogen deficiency symptoms including OVX or postmenopausal symptoms, such as high liver triglycerides, osteoporosis, vulvovaginal atrophy, high blood triglycerides, high blood glucose and weight gain, ii) neutralization of adverse effects (including osteoporosis, non-alcoholic steatohepatitis (NASH), organ atrophy and endometrial cancer) by drugs such as tamoxifen and anastrozole. Definitions

[0028] As will be recognized by those skilled in the art, in stereochemistry, two or more compounds that differ only in the spatial arrangement of their atoms are each considered diastereomers.

[0029] In one aspect, the compounds prepared by the methods of the present application or salts thereof can be substantially pure diastereomers. Compositions comprising the compounds or salts thereof detailed herein, such as compositions comprising substantially pure diastereomeric compounds, are provided. "Substantially pure diastereomer" is intended to mean a compound containing a small amount of impurities, where the impurities mean diastereomers other than the specific diastereomeric compound. In some embodiments, substantially pure diastereomeric compounds or salts thereof are provided in which the proportion of the diastereomer or its salt is about 98% or more. According to the present application, the proportion can be about 99%, about 99.5% or about 99.9% or more.

[0030] The methods of combination therapy described herein should be understood by those skilled in the art to require the administration of one agent or composition in combination with a further agent. "In combination with" refers to the administration of one treatment modality in addition to another treatment modality, for example, the administration of a diastereomeric compound of formula (I) (e.g., α-anolodrine) or a salt thereof in addition to the administration of a second agent to the same individual. Thus, "in combination with" refers to the administration of one treatment modality before, during, or after the delivery of another treatment modality to an individual.

[0031] The methods described herein are generally useful for the treatment of diseases. As used herein, "treatment" is an approach for obtaining a beneficial or desired clinical outcome. Reduction of the pathological consequences of a proliferative disease is also encompassed by "treatment". The methods of the invention contemplate any one or more of these aspects of treatment.

[0032] The term "effective amount" as used herein refers to an amount of a compound or composition sufficient to treat a particular disorder, condition, or disease, e.g., to ameliorate, alleviate, reduce, and / or delay one or more of its symptoms. With respect to cancer, an effective amount includes an amount sufficient to cause shrinkage of a tumor and / or a decrease in the rate of tumor growth (e.g., to inhibit tumor growth), or to prevent or delay other unwanted cell proliferation.

[0033] The term "individual" is a mammal including a human. Individuals include, but are not limited to, humans, cows, horses, cats, dogs, rodents, or primates. In some embodiments, the individual is a human. In some embodiments, the individual is an animal.

[0034] As used in this application, "alkyl" refers to straight-chain or branched-chain saturated hydrocarbons. In some embodiments, the alkyl group has from 1 to 20 carbon atoms ("C1-C 20"alkyl"). In some embodiments, the alkyl group has from 1 to 12 carbon atoms (i.e., (C1-C 12 alkyl)), or from 1 to 10 carbon atoms (i.e., (C1-C 10 alkyl)), or from 1 to 8 carbon atoms (i.e., (C1-C8 alkyl)), or from 1 to 6 carbon atoms (i.e., (C1-C6 alkyl)), or from 1 to 4 carbon atoms (i.e., (C1-C4 alkyl)). Examples of suitable alkyl groups include, but are not limited to, methyl (Me, -CH3), ethyl (Et, -CH2C H3), 1-propyl (n-Pr, n-propyl, -CH2CH2CH3), 2-propyl (i-Pr, i-propyl, -CH(CH3)2), 1-butyl (n-Bu, n-butyl, -CH2CH2CH2CH3), 2-methyl-1-propyl (i-Bu, i-butyl, -CH2CH(CH3)2), 2-butyl (s-Bu, s-butyl, -CH(CH3)CH2CH3), 2-methyl-2-propyl (t-Bu, t-butyl, -C(CH3)3), 1-pentyl (n-pentyl, -CH2CH2CH2CH2CH3), 2-pentyl (-CH(CH3)CH2CH2CH3), 3-pentyl (-CH(CH2CH3)2), 2-methyl-2-butyl (-C(CH3)2CH2CH3), 3-methyl-2-butyl (-CH(CH3)CH(CH3)2), 3-methyl-1-butyl (-CH2CH2CH(CH3)2), 2-methyl-1-butyl (-CH2CH(CH3)CH2CH3), 1-hexyl (-CH2CH2CH2CH2CH2CH3), 2-hexyl (-CH(CH3)CH2CH2CH2CH3), 3-hexyl (-CH(CH2CH3)(CH2CH2CH3)), 2-methyl-2-pentyl (-C(CH3)2CH2CH2CH3), 3-methyl-2-pentyl (-CH(CH3)CH(CH3)CH2CH3), 4-methyl-2-pentyl (-CH(CH3)CH2CH(CH3)2), 3-methyl-3-pentyl (-C(CH3)(CH2CH3)2), 2-methyl-3-pentyl (-CH(CH2CH3)CH(CH3)2), 2,3-dimethyl-2-butyl (-C(CH3)2CH(CH3)2), 3,3-dimethyl-2-butyl (-CH(CH3)C(CH3)3 and octyl (-(CH2)7CH3) may be mentioned.

[0035] 「-alkyl-」 refers to a divalent radical derived from the above alkyl. In some embodiments, as used herein, -alkyl- has at least 1 carbon atom, at least 2 carbon atoms, at least 3 carbon atoms, at least 4 carbon atoms, at least 5 carbon atoms, at least 6 carbon atoms, at least 10 carbon atoms; at least 12 carbon atoms; at least 20 carbon atoms; or at least 40 carbon atoms; or has 1 to 40 carbon atoms, 1 to 30 carbon atoms, 1 to 25 carbon atoms, 1 to 20 carbon atoms, 5 to 20 carbon atoms, 12 to 20 carbon atoms or 14 to 18 carbon atoms. Examples of -alkyl- include, but are not limited to, groups such as methylene (-CH2-), ethylene (-CH2CH2-), propylene (-CH2CH2CH2-), butylene (-CH2CH2CH2CH2-), etc.

[0036] As used herein, "alkenyl" refers to a straight-chain or branched-chain hydrocarbon having at least one carbon-carbon double bond. In some embodiments, the alkenyl group has 2 to 12 carbon atoms (i.e., C2-C 12 alkenyl), or 2 to 10 carbon atoms (i.e., C2-C 10 alkenyl), or 2 to 8 carbon atoms (i.e., C2-C8 alkenyl), or 2 to 6 carbon atoms (i.e., C2-C6 alkenyl), or 2 to 4 carbon atoms (i.e., C2-C4 alkenyl). Examples of suitable alkenyl groups include, but are not limited to, ethylene or vinyl (-CH=CH2), allyl (-CH2CH=CH2) and 5-hexenyl (-CH2CH2CH2CH2CH=CH2).

[0037] 「-alkenyl-」 refers to a divalent radical derived from the above alkenyl. In some embodiments, as used herein, -alkenyl- has at least 2 carbon atoms, at least 3 carbon atoms, at least 4 carbon atoms, at least 5 carbon atoms, at least 6 carbon atoms, at least 10 carbon atoms; at least 12 carbon atoms; at least 20 carbon atoms; or at least 40 carbon atoms; or has 1 to 40 carbon atoms, 1 to 30 carbon atoms, 1 to 25 carbon atoms, 1 to 20 carbon atoms, 5 to 20 carbon atoms, 12 to 20 carbon atoms or 14 to 18 carbon atoms. By way of example, -alkenyl- can be -CH=CH-.

[0038] As used herein, "alkynyl" refers to an unsaturated straight-chain or branched-chain monovalent hydrocarbon chain or a combination thereof having at least one acetylenic unsaturation site (i.e., having at least one moiety of the formula C≡C). The alkynyl group can have a specified number of carbon atoms (i.e., C2-C 20 means 2 to 20 carbon atoms). In some embodiments, the alkynyl group has 2 to 12 carbon atoms (i.e., "C2-C 12 alkynyl"), has 2 to 10 carbon atoms (i.e., "C2-C 10 alkynyl"), has 2 to 8 carbon atoms (i.e., "C2-C8 alkynyl"), has 2 to 6 carbon atoms (i.e., "C2-C6 alkynyl"), or has 2 to 4 carbon atoms (i.e., "C2-C4 alkynyl"). Examples of alkynyl include, but are not limited to, groups such as ethynyl (or acetylenyl), prop-1-ynyl, prop-2-ynyl (or propargyl), but-1-ynyl, but-2-ynyl, but-3-ynyl, and their homologs and isomers.

[0039] The term "aryl" refers to and includes a polyunsaturated aromatic hydrocarbon group. The aryl may further contain additional fused rings (e.g., 1 to 3 rings) including fused aryl, heteroaryl, cycloalkyl and / or heterocyclyl rings. In one variant form, the aryl group contains 6 to 14 ring carbon atoms. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, biphenyl, and the like.

[0040] The term "cycloalkyl" refers to and includes a cyclic monovalent hydrocarbon structure that may be fully saturated, mono-unsaturated or poly-unsaturated but is non-aromatic and has a specified number of carbon atoms (e.g., C1-C 10 which means 1 to 10 carbons). The cycloalkyl may consist of 1 ring such as cyclohexyl, or multiple rings such as adamantyl, excluding aryl groups. The cycloalkyl containing more than 1 ring may be fused, spiro or bridged or a combination thereof. Preferred cycloalkyls are cyclic hydrocarbons having 3 to 13 ring carbon atoms. More preferred cycloalkyls are cyclic hydrocarbons having 3 to 8 ring carbon atoms ("C3-C8 cycloalkyl"). Examples of cycloalkyls include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, norbornyl, and the like.

[0041] "Halo" or "halogen" refers to Group 17 elements having atomic numbers 9 to 85. Preferred halo groups include fluoro, chloro, bromo and iodo. When a residue is substituted with more than one halogen, it may be designated by using a prefix corresponding to the number of attached halogen moieties. For example, dihaloaryl, dihaloalkyl, trihaloaryl, etc. refer to aryl and alkyl substituted with two ("di") or three ("tri") halo groups (which need not necessarily be the same halo); thus, 4-chloro-3-fluorophenyl is within the scope of dihaloaryl. An alkyl group in which each hydrogen is replaced by a halo group is called "perhaloalkyl". A preferred perhaloalkyl group is trifluoroalkyl (-CF3). Similarly, "perhaloalkoxy" refers to an alkoxy group in which each H in the hydrocarbon constituting the alkyl portion of the alkoxy group is replaced by a halogen. An example of a perhaloalkoxy group is trifluoromethoxy (-OCF3).

[0042] The term "heteroaryl" refers to an unsaturated aromatic cyclic group having from 1 to 10 ring carbon atoms and at least one ring heteroatom including, but not limited to, heteroatoms such as nitrogen, oxygen and sulfur, and includes the same, where the nitrogen and sulfur atoms are optionally oxidized and the nitrogen atoms are optionally quaternized. The heteroaryl group can be attached to the remainder of the molecule at a ring carbon or a ring heteroatom. Heteroaryl can further contain additional fused rings (e.g., 1 to 3 rings) including fused aryl, heteroaryl, cycloalkyl and / or heterocyclyl rings. Examples of heteroaryl groups include, but are not limited to, pyridyl, pyrimidyl, thiophenyl, furanyl, thiazolyl and the like.

[0043] The term "heterocyclic ring" or "heterocyclyl" refers to a saturated or unsaturated non-aromatic group having 1 to 10 ring carbon atoms and 1 to 4 ring heteroatoms such as nitrogen, sulfur or oxygen, where the nitrogen and sulfur atoms are optionally oxidized and the nitrogen atoms are optionally quaternized. The heterocyclyl group may have a monocyclic or multiple fused rings, excluding heteroaryl groups. A heterocyclic ring containing more than one ring can be fused, spiro or bridged or any combination thereof. In a fused ring system, one or more of the fused rings can be aryl or heteroaryl. Examples of heterocyclyl groups include, but are not limited to, tetrahydropyranyl, dihydropyranyl, piperidinyl, piperazinyl, pyrrolidinyl, thiazolinyl, thiazolidinyl, tetrahydrofuranyl, tetrahydrothiophenyl, 2,3-dihydrobenzo[b]thiophen-2-yl, 4-amino-2-oxopyrimidin-1(2H)-yl, and the like.

[0044] Unless otherwise specified, "optionally substituted" means that the group may not be substituted or may be substituted with one or more (e.g., 1, 2, 3, 4 or 5) of the substituents listed for that group, and the substituents may be the same or different. In one embodiment, the optionally substituted group has one substituent. In another embodiment, the optionally substituted group has two substituents. In another embodiment, the group to be optionally substituted has three substituents. In another embodiment, the optionally substituted group has four substituents. In some embodiments, the optionally substituted group has 1 - 2, 2 - 5, 3 - 5, 2 - 3, 2 - 4, 3 - 4, 1 - 3, 1 - 4 or 1 - 5 substituents.

[0045] "Pharmaceutically acceptable carrier" refers to a component in a pharmaceutical formulation other than the active ingredient that is non-toxic to the subject. Pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers or preservatives.

[0046] The aspects and embodiments of the invention described herein are understood to include aspects and embodiments of "consisting of" and / or "consisting essentially of".

[0047] References herein to "about" values or parameters include (and describe) variations that are directed to that value or parameter itself. For example, a description of "about X" includes a description of "X".

[0048] As used herein and in the appended claims, the singular forms "a", "or", and "the" include plural referents unless the context clearly dictates otherwise. Compounds and Synthesis

[0049] The present disclosure includes specific stereochemical forms of the described compounds. In stereochemistry, two or more compounds that differ only in the spatial arrangement of their atoms are each considered diastereomers. The diastereomer compounds detailed herein can be substantially pure.

[0050] Generally speaking, for obtaining specific isomers of a compound, chromatography, recrystallization, and other conventional separation procedures can be used with intermediates or final products. In some embodiments, the diastereomer compounds detailed herein can be prepared by resolution in which the diastereomer compound is separated from a mixture of diastereomers. In other embodiments, the diastereomer compound is prepared by asymmetric synthesis that provides a substantially pure diastereomer compound or a salt thereof.

[0051] In some aspects, the present disclosure describes asymmetric syntheses that result in substantially pure diastereomer compounds or salts thereof. Methods for preparing compounds or salts thereof in substantially pure diastereomeric form are provided herein.

[0052] According to the present invention, the compound is a steroid-like compound detailed herein. For example, anordrin is a steroid-like estrogen. In some embodiments, the diastereomeric compound is α-anordrin, (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane).

[0053] In some embodiments, the diastereomeric compound has the structure of formula (I):

Chemical formula

[0054] In some embodiments, R1 is hydroxyl.

[0055] In some embodiments, R1 is -OC(O)R 1aand wherein R 1a is C1-C6 alkyl. In some embodiments, R 1a is C1-C4 alkyl, such as methyl, ethyl, 1-propyl (n-Pr, n-propyl, -CH2CH2CH3), 2-propyl (i-Pr, i-propyl, -CH(CH3)2), 1-butyl. In some embodiments, R 1a is C1-C3 alkyl or C1-C2 alkyl. In some embodiments, R 1a is ethyl.

[0056] In some embodiments, R1 is -OC(O)R 1b COOH, where R 1b is -C1-C6 alkyl-. In some embodiments, R1 is -OC(O)R 1b COOH, where R 1b is -C2-C6 alkenyl-. In some embodiments, R1 is -OC(O)R 1b COOH, where R 1b is methylene (i.e., -CH2-), and R1 is -OC(O)CH2COOH. As some examples of the salt form, in certain embodiments, R1 is -OC(O)CH2COOLi, -OC(O)CH2COONa or -OC(O)CH2COOK. In some embodiments, R1 is -OC(O)R 1b COOH, where R 1b is ethylene (i.e., -CH2CH2-), and R1 is -OC(O)CH2CH2COOH. In certain embodiments, R1 is -OC(O)CH2CH2COOLi, -OC(O)CH2CH2COONa or -OC(O)CH2CH2COOK. In some embodiments, R1 is -OC(O)R 1b COOH, where R 1b is -CH=CH-, and R1 is -OC(O)CH=CHCOOH. In certain embodiments, R1 is -OC(O)CH=CHCOOLi, -OC(O)CH=CHCOONa or -OC(O)CH=CHCOOK.

[0057] In some embodiments, R4 is hydroxyl.

[0058] In some embodiments, R4 is -OC(O)R 4a wherein R 4a is C1-C6 alkyl. In some embodiments, R 4a is C1-C4 alkyl, such as methyl, ethyl, 1-propyl (n-Pr, n-propyl, -CH2CH2CH3), 2-propyl (i-Pr, i-propyl, -CH(CH3)2), 1-butyl. In some embodiments, R 4a is C1-C3 alkyl or C1-C2 alkyl. In some embodiments, R 4a is ethyl.

[0059] In some embodiments, R4 is -OC(O)R 4b COOH, wherein R 4b is -C1-C6 alkyl-. In some embodiments, R4 is -OC(O)R 4b COOH, wherein R 4b is -C2-C6 alkenyl-. In some embodiments, R4 is -OC(O)R 4b COOH, wherein R 4b is methylene (i.e., -CH2-), and R4 is -OC(O)CH2COOH. As some examples of salt forms, in certain embodiments, R4 is -OC(O)CH2COOLi, -OC(O)CH2COONa or -OC(O)CH2COOK. In some embodiments, R4 is -OC(O)R 4b COOH, wherein R 4b is ethylene (i.e., -CH2CH2-), and R1 is -OC(O)CH2CH2COOH. In certain embodiments, R4 is -OC(O)CH2CH2COOLi, -OC(O)CH2CH2COONa or -OC(O)CH2CH2COOK. In some embodiments, R4 is -OC(O)R 4b COOH, wherein R 4bis -CH=CH-, and R4 is -OC(O)CH=CHCOOH. In certain embodiments, R4 is -OC(O)CH=CHCOOLi, -OC(O)CH=CHCOONa or -OC(O)CH=CHCOOK.

[0060] In some embodiments, R6 is C1-C6 alkyl. In some embodiments, R6 is C2-C6 alkenyl. In some embodiments, R7 is C1-C6 alkyl. In some embodiments, R7 is C2-C6 alkenyl. R6 and R7 may be the same or different.

[0061] In some embodiments, R8 is hydrogen, -OH, -NH2, -NO3, halogen, C1-C6 alkyl or C2-C6 alkenyl. In some embodiments, R8 is hydrogen. In some embodiments, R8 is C1-C6 alkyl. In some embodiments, R9 is hydrogen, -OH, -NH2, -NO3, halogen, C1-C6 alkyl or C2-C6 alkenyl. In some embodiments, R9 is hydrogen. In some embodiments, R9 is C1-C6 alkyl. R8 and R9 may be the same or different.

[0062] The substantially pure diastereomeric compounds or salts thereof described herein are prepared using a method that includes reacting a diketone compound of formula (II) 10 in the presence of an organometallic reagent R

Chemical formula

Chemical formula

[0063] R6, R7, R8, and R9 of the diketone compound of formula (II) are as defined above, and in various embodiments of the compound, it has the structure of formula (I).

[0064] Regarding reaction step III-2 of method II shown in the following examples, an exemplary process for preparing α-anolodrine involves reacting the diketone product (c) with silylacetylene, trimethylsilylacetylene (TMS). In the present application, it has been recognized that silylacetylene can be used for the asymmetric alkynylation of the decarbonyl compounds described herein. Without being bound by any particular theory, it is considered that two terminal alkynes add to the two carbonyl groups of the decarbonyl compound one by one.

[0065] R of the silylacetylene of formula (III) a , R b and R c can be the same or different. In some embodiments, R a , R b and R c are independently selected from the group consisting of hydrogen, -OH, -OH, halogen, or C1-C6 alkyl-substituted C1-C 20 alkyl, -OH, halogen, or C1-C6 alkoxy optionally substituted with C1-C6 alkyl, C3-C6 cycloalkyl, C6-C 14 aryl, 5-6 membered heteroaryl, or 3-6 membered heterocyclyl. In some embodiments, R a , R b and R c are independently -OH, halogen, or C1-C6 alkyl-substituted C1-C 20 alkyl. In some embodiments, R a , R b and R c are independently C1-C6 alkyl, such as methyl, ethyl, 1-propyl ( n-Pr, n-propyl, -CH2CH2CH3), 2-propyl (i-Pr, i-propyl, -CH(CH3)2), 1-butyl. In some embodiments, Ra , R b and R c are methyl, and thus the silylacetylene is trimethylsilylacetylene (TMS). Without being bound by any particular theory, one or more of R a , R b and R c have a large size that creates steric hindrance, which is thought to result in the preferential formation of one diastereomer over another.

[0066] The organometallic reagent used according to this method is defined as R 10 -M, where M is a metal. By way of example, R 10 -M is n-butyllithium.

[0067] In some embodiments, M is Li, Na, or K. In some embodiments, M is Li.

[0068] In some embodiments, R 10 is C1-C 20 alkyl optionally substituted with -OH, halogen, or C1-C6 alkyl. In some embodiments, R 10 is C1-C6 alkyl. In some embodiments, R 10 is n-butyl.

[0069] In some embodiments, tetramethylethylenediamine (TEMED) is added together with the organometallic reagent.

[0070] In various embodiments, the methods provided herein are performed in the presence of an organic solvent. A non-limiting list of suitable organic solvents includes, but is not limited to, dichloromethane, chloroform, acetonitrile, dichloroethane, tetrahydrofuran (THF), dimethyl sulfoxide, and toluene.

[0071] The methods provided herein are carried out at relatively low reaction temperatures. The reaction temperature can be in the range of about 0 °C to about -100 °C. In some embodiments, the reaction temperature is from about -20 °C to about -90 °C. In some embodiments, the reaction temperature is from about -40 °C to about -80 °C. In some embodiments, the reaction temperature is from about -30 °C to about -50 °C.

[0072] According to some embodiments of the present invention, a method for stereospecifically preparing a substantially pure diastereomeric compound or a salt thereof comprises a silyl group of formula (IV):

Chemical formula

[0073] In some embodiments, the removal step is carried out by contacting with a deprotecting agent. A non-limiting list of suitable deprotecting agents includes, but is not limited to, tetrabutylammonium fluoride (TBAF), hydrofluoric acid, and potassium fluoride. Pharmaceutical Compositions and Formulations

[0074] The present disclosure includes specific diastereomeric forms of the compounds described herein. Also provided are salts of the compounds referred to herein, such as pharmaceutically acceptable salts. All forms of the diastereomeric compounds, such as crystalline or amorphous forms of the diastereomeric compounds, are also specifically included in the present invention. Compositions comprising the diastereomeric compounds of the present invention or salts thereof, such as compositions of substantially pure diastereomeric compounds or salts thereof, are also contemplated.

[0075] Any pharmaceutical composition of any of the diastereomeric compounds detailed herein is encompassed by the present disclosure. The present disclosure includes pharmaceutical compositions comprising a substantially pure diastereomeric compound of formula (I) or a salt thereof and a pharmaceutically acceptable carrier or excipient. In one aspect, the salt is a pharmaceutically acceptable salt. For example, the salt is an acid addition salt, such as a salt formed with an inorganic or organic acid. The pharmaceutical composition can take a suitable form for oral administration, buccal administration, parenteral administration, nasal administration, topical administration or rectal administration, or a form suitable for administration by inhalation.

[0076] In one variant, the substantially pure diastereomeric compound or a salt thereof is a synthetic product prepared for administration to an individual. In another variant, a composition containing the substantially pure diastereomeric compound or a salt thereof is provided. In another variant, the present disclosure encompasses pharmaceutical compositions comprising a compound detailed herein and a pharmaceutically acceptable carrier.

[0077] The diastereomeric compounds or salts thereof detailed herein can be formulated for any available route of delivery including oral delivery, mucosal (e.g., nasal, sublingual, vaginal, buccal or rectal) delivery, parenteral (e.g., intramuscular, subcutaneous or intravenous) delivery, topical delivery or transdermal delivery. The diastereomeric compounds or salts thereof can be formulated with suitable carriers for providing delivery forms including, but not limited to, tablets, caplets, capsules (e.g., hard gelatin capsules or soft gelatin capsules), cachets, troches, lozenges, gums, dispersions, suppositories, ointments, cataplasms (poultices), pastes, powders, dressings, creams, solutions, patches, aerosols (e.g., nasal sprays or inhalers), gels, suspensions (e.g., aqueous or non-aqueous liquid suspensions, oil-in-water emulsions or water-in-oil emulsions), solutions and elixirs.

[0078] The diastereomeric compounds or salts thereof described herein can be used in the preparation of formulations such as pharmaceutical preparations by combining the compound or salt thereof as an active ingredient with a pharmaceutically acceptable carrier, such as those described above. Depending on the form of treatment of the system (e.g., transdermal patch vs. oral tablet), the carrier can be in various forms. In addition, the pharmaceutical preparation can contain preservatives, solubilizers, stabilizers, rewetting agents, emulsifiers, sweeteners, pigments, regulators and salts for adjusting osmotic pressure, buffers, coating agents or antioxidants. The formulation containing the compound can also contain other substances having beneficial therapeutic properties. The pharmaceutical preparation can be prepared by known pharmaceutical methods. Suitable formulations can be found, for example, in Remington’s Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, PA, 20 th ed. (2000) (which is incorporated herein by reference).

[0079] In some embodiments, the diastereomeric compounds described herein can be administered to an individual in the form of generally acceptable oral compositions, such as tablets, coated tablets and gel capsules in hard shell or soft shell, emulsion or suspension. Examples of carriers that can be used in the preparation of such compositions are lactose, corn starch or its derivatives, talc, stearate or its salts, etc. Carriers acceptable for gel capsules having a soft shell are, for example, vegetable oils, waxes, fats, semi-solids and liquid polyols, etc. In addition, the pharmaceutical preparation can contain preservatives, solubilizers, stabilizers, rewetting agents, emulsifiers, sweeteners, pigments, regulators and salts for adjusting osmotic pressure, buffers, coating agents or antioxidants.

[0080] Any of the diastereomeric compounds or salts thereof described herein can be formulated into tablets of any of the dosage forms described, for example, the compounds or pharmaceutically acceptable salts thereof described herein can be formulated as 10 mg tablets. Method of Use

[0081] The compounds and compositions described in detail herein, e.g., pharmaceutical compositions comprising a diastereomeric compound of formula (I) provided herein or a salt thereof and a pharmaceutically acceptable carrier or excipient, can be used in the methods of administration and treatment provided herein.

[0082] Estrogen deficiency is associated with the development of many health conditions such as infertility, premature aging, osteoporosis, and cardiovascular disease. Ovariectomized (OVX) or postmenopausal individuals typically suffer from estrogen deficiency. Estrogen deficiency symptoms described herein include, but are not limited to, osteoporosis, fatty liver, weight gain, high blood triglycerides and glucose, and organ atrophy (e.g., atrophy of the kidney, muscle, and vulvovagina).

[0083] In some embodiments, provided is a method of treating estrogen deficiency in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof.

[0084] In some embodiments, provided is a method of preventing or alleviating estrogen deficiency symptoms in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof. In some embodiments, provided is a method of preventing or alleviating estrogen deficiency symptoms selected from the group consisting of high liver triglycerides, osteoporosis, vulvovaginal atrophy, high blood triglycerides, high blood glucose, and weight gain.

[0085] While not bound by any particular theory, it has been observed that the diastereomeric compounds of formula (I) or salts thereof, as detailed herein, have a therapeutic effect not possessed by other stereochemical forms. Thus, the diastereomeric compounds of formula (I), compositions and methods thereof, as detailed in this application, may have advantageous therapeutic effects compared to other stereochemical forms or stereochemical mixtures of the compounds of formula (I). As one specific example, two chiral anordrin compounds named 2 alpha-(α-) and 2 beta-(β-) anordrin have been found (18, 19). According to some embodiments of the present invention, α-anordrin has a therapeutic effect in the treatment of estrogen deficiency and in the prevention or alleviation of estrogen deficiency symptoms, while β-anordrin has no minimal therapeutic effect in these treatments.

[0086] In some embodiments, co-administration (which may be separate or simultaneous) of a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof with one or more additional agents is provided herein. For example, the diastereomeric compound of formula (I) or a salt thereof and one or more additional agents are co-administered (e.g., in a single composition such as a pharmaceutical composition described herein). Additionally or alternatively, the diastereomeric compound of formula (I) or a salt thereof and one or more additional agents may be administered sequentially, e.g., one or more additional agents are administered first and the diastereomeric compound of formula (I) or a salt thereof is administered next, or the diastereomeric compound of formula (I) or a salt thereof is administered first and one or more additional agents are administered next.

[0087] The present disclosure is a method for treating estrogen deficiency and for preventing or alleviating estrogen deficiency symptoms, comprising administering a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof in combination with at least one additional agent It includes a method. Further agents can be selective estrogen receptor modulators (SERMs) and aromatase inhibitors.

[0088] Currently, SERMs are used for the treatment of breast cancer, osteoporosis and postmenopausal symptoms, because these drugs have the characteristic that they can act as agonists and antagonists of estrogen depending on the target tissue. According to the present invention, suitable SERMs include, but are not limited to, tamoxifen, raloxifene, toremifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene and levormeloxifene. In some embodiments, the SERM used in combination with the diastereomeric compound or its salt detailed herein is tamoxifen. In some embodiments, the SERM used in combination with the diastereomeric compound or its salt detailed herein is raloxifene or a functional equivalent thereof. As used herein, "a functional equivalent thereof" refers to a compound that functions through the same mechanism as raloxifene. For example, functional equivalents of raloxifene include, but are not limited to, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene and levormeloxifene.

[0089] In some embodiments, the method for treating estrogen deficiency comprises administering a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof in combination with at least one SERM. In some embodiments, the method for preventing or alleviating estrogen deficiency symptoms comprises administering a diastereomeric compound (e.g., α-anordrin) or a salt thereof in combination with at least one SERM.

[0090] "Aromatase inhibitor" refers to a class of drugs that inhibit aromatase activity. Aromatase inhibitors are used in the treatment of breast cancer and ovarian cancer in postmenopausal women to reduce the increased estrogen conversion during cycles using exogenous testosterone. Suitable aromatase inhibitors include, but are not limited to, anastrozole (Arimidex), letrozole (Femara), exemestane (Aromasin), vorozole (Rivizor), formestane (Lentaron), and fadrozole (Afema). In some embodiments, the aromatase inhibitor used in combination with the diastereomeric compound or its salt detailed herein is anastrozole.

[0091] In some embodiments, a method for treating estrogen deficiency comprises administering a diastereomeric compound of formula (I) (e.g., α-anordrin) or a salt thereof in combination with at least one aromatase inhibitor. In some embodiments, a method for preventing or alleviating estrogen deficiency symptoms comprises administering a diastereomeric compound (e.g., α-anordrin) or a salt thereof in combination with at least one aromatase inhibitor.

[0092] In some embodiments, there is provided a method for preventing or alleviating osteoporosis in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof.

[0093] In some embodiments, there is provided a method for preventing or alleviating osteoporosis in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is raloxifene or a functional equivalent thereof. In some embodiments, the additional agent is raloxifene.

[0094] In some embodiments, a method for preventing or reducing osteoporosis in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is an aromatase inhibitor, is provided. In some embodiments, the additional agent is anastrozole.

[0095] In some embodiments, the above method for preventing or reducing osteoporosis further comprises administering to the individual an effective amount of calcium. Suitable amounts of calcium include, but are not limited to, about 0.25 to about 500 mg / day, such as about 10 to about 200 mg / day, about 50 to about 1500 mg / day. In some embodiments, the method further comprises administering to the individual an effective amount of vitamin D. Suitable amounts of vitamin D include, but are not limited to, about 400 to about 800 IU / day, such as about 500 to about 600 IU / day.

[0096] In some embodiments, a method for preventing or reducing high liver triglycerides in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof is provided.

[0097] In some embodiments, a method for preventing or reducing high liver triglycerides in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is raloxifene or a functional equivalent thereof, is provided. In some embodiments, the additional agent is raloxifene. In some embodiments, the additional agent is selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene.

[0098] In some embodiments, provided is a method for preventing or reducing hyperglycemia in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof.

[0099] In some embodiments, provided is a method for preventing or reducing hyperglycemia in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is raloxifene or a functional equivalent thereof. In some embodiments, the additional agent is raloxifene. In some embodiments, the additional agent is selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene.

[0100] In some embodiments, provided is a method for preventing or reducing vulvovaginal atrophy in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof.

[0101] In some embodiments, provided is a method for preventing or reducing vulvovaginal atrophy in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is raloxifene or a functional equivalent thereof. In some embodiments, the additional agent is raloxifene. In some embodiments, the additional agent is selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene selected from the group consisting of.

[0102] In some embodiments, a method for preventing or reducing vulvovaginal atrophy in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is an aromatase inhibitor, is provided. In some embodiments, the additional agent is selected from the group consisting of, but not limited to, anastrozole or a functional equivalent thereof.

[0103] In some embodiments, a method for preventing or reducing weight gain in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof is provided.

[0104] In some embodiments, a method for preventing or reducing weight gain in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is raloxifene or a functional equivalent thereof, is provided. In some embodiments, the additional agent is raloxifene. In some embodiments, the additional agent is selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene.

[0105] In some embodiments, a method for preventing or reducing hypertriglyceridemia in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof is provided.

[0106] In some embodiments, a method for preventing or reducing hypertriglyceridemia in an individual, the method comprising administering to the individual a) an effective amount of a diastereomeric compound of formula (I) or a salt thereof; and b) an effective amount of at least one additional agent, wherein the additional agent is raloxifene or a functional equivalent thereof, is provided. In some embodiments, the additional agent is raloxifene. In some embodiments, the additional agent is selected from the group consisting of tamoxifen, raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene.

[0107] In certain embodiments, a method for preventing or reducing high liver triglycerides in an individual, the method comprising administering to the individual an effective amount of a substantially pure diastereomeric compound of (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstan (α-norandrolone) of formula (Ia)

Chemical formula

[0108] In certain embodiments, a method for preventing or reducing hypertriglyceridemia in an individual, the method comprising administering to the individual an effective amount of a substantially pure diastereomeric compound of (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstan (α-norandrolone) of formula (Ia)

Chemical formula

[0109] In certain embodiments, a method for preventing or reducing osteoporosis in an individual, the method comprising administering to the individual an effective amount of a substantially pure diastereomeric compound of (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstan (α-norandrolone) of formula (Ia) [Chemical formula] Provided herein is a method comprising administering an effective amount of a substantially pure diastereomeric compound that is.

[0110] In certain embodiments, a method for preventing or reducing vulvovaginal atrophy in an individual, the method comprising administering to the individual (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstan (α-nordrolone) of formula (Ia) [Chemical formula] Provided herein is a method comprising administering an effective amount of a substantially pure diastereomeric compound that is.

[0111] In certain embodiments, a method for preventing or reducing hyperglycemia in an individual, the method comprising administering to the individual (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstan (α-nordrolone) of formula (Ia) [Chemical formula] Provided herein is a method comprising administering an effective amount of a substantially pure diastereomeric compound that is.

[0112] In certain embodiments, a method for preventing or reducing weight gain in an individual, the method comprising administering to the individual (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstan (α-nordrolone) of formula (Ia) [Chemical formula] Provided herein is a method comprising administering an effective amount of a substantially pure diastereomeric compound that is.

[0113] In the above method, the substantially pure diastereomeric compound has a diastereomeric excess (de) of about 98% or more.

[0114] In the above method, the substantially pure diastereomeric compound has a diastereomeric excess (de) of about 99% or more, about 99.5% or more, or about 99.9% or more.

[0115] However, the side effects seen in traditional estrogen replacement therapy (ERT), aromatase inhibitors, and SERMs remain a concern for some researchers (12). Tamoxifen is marketed as an antagonist of the classical estrogen pathway for treating breast cancer patients and has also been reported as an agonist of ER-α36, potentially causing anti-estrogen treatment resistance while stimulating the growth of endometrial epithelial cells, resulting in endometrial cancer (7, 8). Raloxifene is marketed as an upgraded version of tamoxifen, with fewer side effects and the advantage of preventing postmenopausal symptoms such as osteoporosis. However, raloxifene can still cause serious side effects common to tamoxifen treatment, such as thromboembolism and non-alcoholic steatohepatitis (NASH) (9, 10). The detailed mechanisms involved in the side effects caused by either raloxifene or tamoxifen remain unclear. Ipriflavone is a derivative of phytohormones, and its metabolites bind to ER-αLBD with lower affinity than E2 and exhibit low estrogen-like effects. The metabolites of ipriflavone and isoflavones show binding affinity and activity for ER-β and E2, and these metabolites are used as pharmaceuticals for preventing osteoporosis in some countries. Their effectiveness has not been supported in at least one clinical trial (11). Also, for ER-positive breast cancer in postmenopausal women, aromatase inhibitors have been marketed. It also inhibits the synthesis of estrogen in bone, resulting in osteoporosis.

[0116] A method for reducing the side effects of at least one additional agent in an individual, the method comprising administering to the individual an effective amount of a diastereomeric compound of formula (I) or a salt thereof when used in combination with said at least one additional agent is provided herein. In some embodiments, the diastereomeric compound of formula (I) or a salt thereof used in combination with tamoxifen can reduce the side effects of tamoxifen. For example, in the present application, it has been observed that α-anordrin used in combination with tamoxifen reduces the side effects of tamoxifen and provides an effective combination therapy for the treatment of estrogen deficiency or the prevention or reduction of estrogen deficiency symptoms. In some embodiments, the diastereomeric compound of formula (I) or a salt thereof used in combination with at least one aromatase inhibitor can reduce the side effects of the aromatase inhibitor.

[0117] In certain embodiments, a method for reducing the side effects of tamoxifen in an individual, the method comprising administering to the individual, in combination with tamoxifen, an effective amount of a substantially pure diastereomeric compound of (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-anordrin) of formula (Ia)

Chemical formula

[0118] In certain embodiments, a method for reducing the side effects of at least one aromatase inhibitor in an individual, the method comprising administering to the individual, in combination with the aromatase inhibitor, an effective amount of a substantially pure diastereomeric compound of (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-anordrin) of formula (Ia)

Chemical formula

[0119] In the above method, the substantially pure diastereomeric compound has a diastereomeric excess (de) of about 98% or more.

[0120] In the above method, the substantially pure diastereomeric compound has a diastereomeric excess (de) of about 99% or more, about 99.5% or more, or about 99.9% or more. Mode of administration

[0121] In the context of combination therapy, a composition comprising a diastereomeric compound of formula (I) or a salt thereof and a further agent can be administered simultaneously (i.e., co - administered) and / or sequentially (i.e., sequential administration).

[0122] In some embodiments, a diastereomeric compound of formula (I) or a salt thereof and a further agent (including the specific agents described herein) are co - administered. As used herein, the term "co - administered" means that the diastereomeric compound of formula (I) or a salt thereof and the further agent are administered at time intervals of about 15 minutes or less (e.g., any of about 10 minutes or less, about 5 minutes or less, or about 1 minute or less). When the drugs are co - administered , the diastereomeric compound of formula (I) or a salt thereof and the further agent can be included in the same composition (e.g., a composition comprising both the diastereomeric compound of formula (I) or a salt thereof and the further agent, e.g., a pharmaceutical composition included herein) or in separate compositions (e.g., the diastereomeric compound of formula (I) or a salt thereof and the further agent are included in separate compositions).

[0123] In some embodiments, the diastereomeric compound of formula (I) or a salt thereof and the additional agent are administered sequentially. The term "sequential administration" as used herein means that the diastereomeric compound of formula (I) or a salt thereof and the additional agent are administered at time intervals greater than about 15 minutes (e.g., greater than any of about 20 minutes, about 30 minutes, about 40 minutes, about 50 minutes, about 60 minutes or longer). First, either the diastereomeric compound of formula (I) or a salt thereof or the additional agent may be administered. The diastereomeric compound of formula (I) or a salt thereof and the additional agent are contained in separate compositions, and these may be placed in the same or different packages.

[0124] In some embodiments, the administration of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is simultaneous, i.e., the administration periods of the diastereomeric compound of formula (I) or a salt thereof and the additional agent overlap with each other. In some embodiments, the diastereomeric compound of formula (I) or a salt thereof is administered in at least 1 cycle (e.g., any one of at least 2, 3, or 4 cycles) prior to the administration of the additional agent. In some embodiments, the additional agent is administered over any one of at least 1, 2, 3, or 4 weeks. In some embodiments, the administration of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is started substantially simultaneously (e.g., within any one period of 1, 2, 3, 4, 5, 6, or 7 days). In some embodiments, the administration of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is ended substantially simultaneously (e.g., within any one period of 1, 2, 3, 4, 5, 6, or 7 days). In some embodiments, the administration of the additional agent is continued after the completion of the administration of the diastereomeric compound of formula (I) or a salt thereof (e.g., over any one period of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months). In some embodiments, the administration of the additional agent is started after the start of the administration of the diastereomeric compound of formula (I) or a salt thereof (e.g., after any one period of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 months or we months). In some embodiments, the administration of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is started and ended substantially simultaneously. In some embodiments, the administration of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is started substantially simultaneously, and the administration of the additional agent is continued after the completion of the administration of the diastereomeric compound of formula (I) or a salt thereof (e.g., over any one period of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months).In some embodiments, administration of the diastereomeric compound of formula (I) or a salt thereof and of a further agent is stopped substantially simultaneously, and administration of the further agent is started after the start of administration of the diastereomeric compound of formula (I) or a salt thereof (e.g., after any one period of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 months or 12 months).

[0125] The dosing frequency of the diastereomeric compound of formula (I) or a salt thereof and / or of a further agent can be adjusted during the course of treatment based on the judgment of the physician administering it. When administered separately, the diastereomeric compound of formula (I) or a salt thereof and the further agent can be administered at different dosing frequencies or intervals. For example, the diastereomeric compound of formula (I) or a salt thereof can be administered weekly while the further agent can be administered at a higher or lower frequency. A variety of formulations and devices for achieving sustained release are known in the art. Exemplary dosing frequencies are further provided herein.

[0126] The diastereomeric compound of formula (I) or a salt thereof and a further agent can be administered using the same or different routes of administration. Exemplary routes of administration are further provided herein. In some embodiments (for both co - administration and sequential administration), the diastereomeric compound of formula (I) or a salt thereof and a further agent are administered in a predetermined ratio. For example, in some embodiments, the weight ratio of the diastereomeric compound of formula (I) or a salt thereof and a further agent is about 1 to 1. In some embodiments, the weight ratio can be from about 0.001 to about 1 to about 1000 to about 1 or from about 0.01 to about 1 to 100 to about 1. In some embodiments, the weight ratio of the diastereomeric compound of formula (I) or a salt thereof and a further agent is less than any of about 100:1, about 50:1, about 30:1, about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1 and about 1:1. In some embodiments, the weight ratio of the diastereomeric compound of formula (I) or a salt thereof and a further agent is greater than any of about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, about 30:1, about 50:1, about 100:1. Other ratios are contemplated.

[0127] The dosage required for the diastereomeric compound of formula (I) or a salt thereof and / or a further agent may be lower (but not necessarily) than the dosage normally required when each agent is administered alone. Thus, in some embodiments, a sub - therapeutic amount of the diastereomeric compound of formula (I) or a salt thereof and / or a further agent is administered. "Sub - therapeutic amount" or "sub - therapeutic level" refers to an amount that is less than the therapeutic amount (i.e., less than the amount normally used when the diastereomeric compound of formula (I) or a salt thereof and / or a further agent is administered alone). The decrease can be reflected in terms of the amount administered per given dose and / or the amount administered over a given period (decrease in frequency).

[0128] In some embodiments, sufficient additional agent is administered to allow for a decrease of at least about 5%, about 10%, about 20%, about 30%, about 50%, about 60%, about 70%, about 80%, about 90% or more of the normal dosage of the diastereomeric compound of formula (I) or a salt thereof to achieve the same degree of treatment. In some embodiments, sufficient diastereomeric compound of formula (I) or a salt thereof is administered to allow for a decrease of at least about 5%, about 10%, about 20%, about 30%, about 50%, about 60%, about 70%, about 80%, about 90% or more of the normal dosage of the additional agent to achieve the same degree of treatment.

[0129] In some embodiments, the dosages of both the diastereomeric compound of formula (I) or a salt thereof and the additional agent are decreased as compared to the respective normal dosages when administered alone. In some embodiments, both the diastereomeric compound of formula (I) or a salt thereof and the additional agent are administered at sub-therapeutic levels (i.e., decreased levels). In some embodiments, the dosages of the diastereomeric compound or a salt thereof and / or the additional agent of formula (I) are substantially lower than the established maximum toxic dose (MTD). For example, the dosages of the diastereomeric compound or a salt thereof and / or the additional agent of formula (I) are less than about 50%, less than about 40%, less than about 30%, less than about 20% or less than about 10% of the MTD.

[0130] In some embodiments, the dosage of the diastereomeric compound of formula (I) or a salt thereof and / or the dosage of the additional agent are higher than the dosages normally required when each agent is administered alone. For example, in some embodiments, the dosages of the diastereomeric compound of formula (I) or a salt thereof and / or the additional agent are substantially higher than the established maximum toxic dose (MTD). For example, the dosages of the diastereomeric compound of formula (I) or a salt thereof and / or the additional agent are about 50%, about 40% greater than, about 30%, about 20% or greater than about 10% of the MTD of the agent when administered alone.

[0131] In some embodiments, the amount of the diastereomeric compound of formula (I) or a salt thereof (alone or in combination with a further agent) is included in any of the following ranges: about 0.1 to about 0.5 mg, about 0.5 to about 5 mg, about 5 to about 10 mg, about 10 to about 15 mg, about 15 to about 20 mg, about 20 to about 25 mg, about 20 to about 50 mg, about 25 to about 50 mg, about 50 to about 75 mg, about 50 to about 100 mg, about 75 to about 100 mg, about 100 to about 125 mg, about 125 to about 150 mg, about 150 to about 175 mg, about 175 to about 200 mg, about 200 to about 225 mg, about 225 to about 250 mg, about 250 to about 300 mg, about 300 to about 350 mg, about 350 to about 400 mg, about 400 to about 450 mg or about 450 to about 500 mg. In some embodiments, the amount of the diastereomeric compound of formula (I) or a salt thereof (e.g., unit dosage form) is in the range of about 5 mg to about 500 mg, such as about 30 mg to about 300 mg or about 50 mg to about 200 mg.

[0132] In some embodiments, the amount of the diastereomeric compound of formula (I) or a salt thereof (alone or in combination with a further agent) includes any of at least about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 0.25 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2.5 mg / kg, about 3.5 mg / kg, about 5 mg / kg, about 6.5 mg / kg, about 7.5 mg / kg, about 10 mg / kg, about 15 mg / kg or about 20 mg / kg. In some embodiments, the amount of the diastereomeric compound of formula (I) or a salt thereof (alone or in combination with a further agent) includes any of at least about 0.01 mg / kg / day, about 0.05 mg / kg / day, about 0.1 mg / kg / day, about 0.25 mg / kg / day, about 0.5 mg / kg / day, about 1 mg / kg / day, about 2.5 mg / kg / day, about 3.5 mg / kg / day, about 5 mg / kg / day, about 6.5 mg / kg / day, about 7.5 mg / kg / day, about 10 mg / kg / day, about 15 mg / kg / day or about 20 mg / kg / day.

[0133] In some embodiments, the amount of the additional agent comprises any of at least about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 0.25 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2.5 mg / kg, about 3.5 mg / kg, about 5 mg / kg, about 6.5 mg / kg, about 7.5 mg / kg, about 10 mg / kg, about 15 mg / kg or about 20 mg / kg. In some embodiments, the amount of the diastereomeric compound of formula (I) or a salt thereof (alone or in combination with an additional agent) is any of at least about 0.01 mg / kg / day, about 0.05 mg / kg / day, about 0.1 mg / kg / day, about 0.25 mg / kg / day, about 0.5 mg / kg / day, about 1 mg / kg / day, about 2.5 mg / kg / day, about 3.5 mg / kg / day, about 5 mg / kg / day, about 6.5 mg / kg / day, about 7.5 mg / kg / day, about 10 mg / kg / day, about 15 mg / kg / day or about 20 mg / kg / day.

[0134] Exemplary dosing frequencies of the diastereomeric compound of formula (I) or a salt thereof (and additional agent) include, but are not limited to, once every two weeks, once every three weeks, once every four weeks, once every six weeks or once every eight weeks. In some embodiments, the composition is administered at any of about once, about twice, about three times, about four times, about five times, about six times or about seven times (i.e., daily), about three times daily or about twice daily per week at least. In some embodiments, the interval between each administration is shorter than any of about six months, about three months, about one month, about twenty days, about fifteen days, about twelve days, about ten days, about nine days, about eight days, about seven days, about six days, about five days, about four days, about three days, about two days or about one day. In some embodiments, the interval between each administration is longer than any of about one month, about two months, about three months, about four months, about five months, about six months, about eight months or about twelve months. In some embodiments, there is no rest period in the dosing schedule. In some embodiments the interval between each administration is about one week or less.

[0135] Administration of the diastereomeric compound of formula (I) or a salt thereof (and additional agent) can be extended over a long period, for example, from about 1 month to up to about 7 years. In some embodiments, the composition is administered for any period of at least about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 18, about 24, about 30, about 36, about 48, about 60, about 72 or about 84 months.

[0136] In some embodiments, the individual is treated with any of at least about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9 or about 10 treatment cycles.

[0137] The dosing frequency of the additional agent may be the same as or different from the dosing frequency of the diastereomeric compound of formula (I) or a salt thereof. Exemplary frequencies are provided above.

[0138] The diastereomeric compound of formula (I) or a salt thereof (and additional agent) described herein can be administered to an individual (e.g., a human) via various routes including, for example, oral, intravenous, intraarterial, intraperitoneal, intralung, inhalation, intracapsular, intramuscular, intratracheal, subcutaneous, intraocular, intrathecal, transmucosal and transdermal. In some embodiments, a sustained release formulation of the composition can be used.

[0139] Combinations of the dosing regimens described herein can be used. The combination therapies described herein can be carried out alone or in combination with another treatment, such as surgery, irradiation, chemotherapy, immunotherapy and gene therapy, etc. In addition, those having a higher risk of developing a proliferative disease can receive treatment to inhibit and / or delay the onset of the disease.

[0140] As will be understood by those skilled in the art, the appropriate dosage of the additional agent is close to that already used in clinical treatments where the additional agent is administered alone or in combination with additional agents. Variations in dosage may occur depending on the condition being treated. As noted above, in some embodiments, the additional agent may be administered at a reduced level. Kit and medicament

[0141] The present invention also provides medicaments, kits and unit doses useful in the methods described herein. Any use described herein is provided, regardless of the context of use as a medicament and / or for the manufacture of a medicament.

[0142] In some embodiments, the composition may be in unit dosage form (such as an oral unit dosage form). Suitable unit dosage forms include, but are not limited to, capsules, tablets, pills, caplets, gels, liquids (such as suspensions, solutions, emulsions), powders or other particulate substances.

[0143] In some embodiments, the weight ratio of the diastereomeric compound of formula (I) or a salt thereof and at least one additional agent in the composition is about 1:1. In some embodiments, the weight ratio is from about 0.001 to about 1 to about 1000 to about 1 or from about 0.01 to about 1 to 100 to about 1. In some embodiments, the weight ratio of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is less than any of about 100:1, about 50:1, about 30:1, about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1 and about 1:1. In some embodiments, the weight ratio of the diastereomeric compound of formula (I) or a salt thereof and the additional agent is greater than any of about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, about 30:1, about 50:1, about 100:1. In some embodiments, the formula in the composition The weight ratio of the diastereomeric compound of (I) or a salt thereof to the additional agent is from about 1:20 to about 20:1 (including, for example, from about 10:1 to about 1:10 or from about 1:10 to about 1:15).

[0144] In another aspect, a kit is provided that includes the diastereomeric compound of formula (I) or a salt thereof and an additional agent, either in separate containers or in the same container. The kits of the invention include one or more containers comprising a diastereomeric compound of formula (I) or a salt thereof (or unit dosage form and / or product) and / or at least one additional agent, and in some embodiments, further include instructions for use according to any of the methods described herein. The kit may further include instructions regarding the selection of an appropriate individual for treatment. The instructions provided with the kits of the invention are typically instructions on a label or package insert (e.g., a sheet included in the kit), but machine-readable instructions (e.g., instructions on a magnetic storage disk or an optical storage disk) are also acceptable.

[0145] In some embodiments, the kit includes a) an effective amount of the diastereomeric compound of formula (I) or a salt thereof and b) an effective amount of at least one additional agent selected from the group consisting of tamoxifen, raloxifene or a functional equivalent thereof, and an aromatase inhibitor. In some embodiments, the kit includes a) an effective amount of the diastereomeric compound of formula (I) or a salt thereof, b) an effective amount of at least one additional agent selected from the group consisting of tamoxifen, raloxifene or a functional equivalent thereof, and an aromatase inhibitor, and c) instructions for co-administering, sequentially or together, the diastereomeric compound of formula (I) or a salt thereof and the additional agent for the treatment of cancer (or other uses described herein).

[0146] The diastereomeric compound of formula (I) or a salt thereof and the additional agent may be present in separate containers or in a single container. The kit may comprise one separate composition, or it is understood that one composition contains the diastereomeric compound of formula (I) or a salt thereof and one or more than two compositions contain an additional agent.

[0147] The kits of the invention are appropriately packaged. Suitable packagings include, but are not limited to, vials, bottles, jars, and flexible packagings (e.g., sealed Mylar or plastic bags). The kits may optionally provide additional components such as buffer solutions and interpretive information. Accordingly, the present application also provides products including vials (e.g., sealed vials), bottles, jars, and flexible packagings.

[0148] Instructions for the use of the diastereomeric compound of formula (I) or a salt thereof generally include information regarding dosage, dosing schedule, and route of administration for the intended treatment. The container can be a unit dose, a bulk package (e.g., a multiple-dose package), or less than a unit dose. For example, kits containing an amount of the diastereomeric compound of formula (I) or a salt thereof disclosed herein sufficient to provide effective treatment of an individual over an extended period (e.g., any of 1 week, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks, 3 months, 4 months, 5 months, 7 months, 8 months, 9 months, or longer) can be provided. The kit can also include multiple unit doses of the diastereomeric compound of formula (I) or a salt thereof, a pharmaceutical composition, and instructions for use, and can be packaged in an amount sufficient for storage and use in pharmacies, e.g., hospital pharmacies and dispensing pharmacies.

[0149] Those skilled in the art will recognize that several embodiments are possible within the scope and spirit of the present invention. The present invention will be described in more detail with reference to the following non-limiting examples. The following examples further illustrate the present invention, but of course do not limit its scope in any way. should not be construed as

Example

[0150] Although the present invention has been described and illustrated in some detail, the present disclosure has been made by way of example only, and it is understood that numerous changes in combinations and permutations of parts may be made without departing from the spirit and scope of the invention as defined in the claims. Example 1: Synthesis Example

[0151] The chemical synthesis of α-anordrin is detailed in Method I below (shown in Scheme I incorporated herein) or Method II (Scheme II) by reference to the publications of U.S. Patent No. 5,001,120 and Organic Letters, 2007, 9(9); 1643-6 and can be prepared as described using the procedures.

Chemical formula

[0152] Reaction Step I: Preparation of Jones reagent: 57 g of CrO3 was dissolved in 400 ml of H2O, and then 85 ml of H2SO4 was slowly added. The resulting Jones reagent was then cooled to 40 °C. 50 g of 5α-androstane and 200 ml of acetic acid were added to a 1 L three-necked flask. The mixture was warmed to 55 - 60 °C, stirred, and dissolved under a nitrogen atmosphere. The Jones reagent was added dropwise within 1 hour. The reaction temperature was raised to 90 °C and reacted for 1 hour, and acetic acid was distilled off under reduced pressure. The residue was filtered, washed with H2O, and dried to obtain 50 g of the diacid product (b).

[0153] Reaction Step I: Preparation of Jones reagent: 57 g of CrO3 was dissolved in 400 ml of H2O , Next, 85 ml of H2SO4 was slowly added. Then, the resulting Jones reagent was cooled to 40 °C. 50 g of 5α-androstane and 200 ml of acetic acid were added to a 1 L three-necked flask. The mixture was warmed to 55 - 60 °C, stirred, and dissolved under a nitrogen atmosphere. The Jones reagent was added dropwise within 1 hour. The reaction temperature was raised to 90 °C and reacted for 1 hour, and acetic acid was distilled under reduced pressure. The residue was filtered, washed with H2O, and dried to obtain 50 g of the diacid product (b).

[0154] Reaction Step II: 30 g of the diacid product (b) and 600 mL of acetic anhydride were dissolved in a 1 L three-necked flask, added to 24 g of sodium acetate, stirred, and refluxed for 4 hours. The residue was filtered and dried to obtain 18 g of the diketone product (c).

[0155] Reaction Steps III-1 and IV-1: Preparation of the potassium acetylide reagent: 15 g of a KOH solution (20% w / w) was cooled to 4 °C in a 250 mL three-necked flask, and acetylene was injected to a stable mass. 6 g of the diketone product (c) was dissolved in 42 mL of THF and then added to the three-necked flask at 25 °C over 2 hours. The product was diluted with H2O and acidified with HCl. The residue was filtered, washed with H2O, and dried to obtain 5.9 g of the 2,17α-diethynyl-A-nor-5a-androstane-2,17β-diol product (e).

[0156] Reaction Step V: 2.8 g of the 2,17α-diethynyl-A-nor-5a-androstane-2,17β-diol product (e), 4.5 mL of propionic acid, and 7.8 mL of propionic anhydride were mixed in a 100 mL flask and then stirred for 3 hours. 30 mL of H2O and 40 mL of ethyl acetate were added to the flask, stirred, and the water and organic phases were separated. The organic phase was washed 5 times with water, and 10 mL of n-hexane was added. The residue was filtered and dried to obtain a 2.0 g mixture of the products α-nor-drin and β-nor-drin (fα and fβ, respectively).

[0157] According to some embodiments of the present invention, the asymmetric synthesis of α-anordrin can be prepared as described using the steps detailed in Method II below (as shown in Scheme II).

Chemical formula

[0158] Similar to Method I, the diketone product (c) was prepared using the above Reaction Steps I and II.

[0159] Reaction Step III-2: 6 mL of dry THF and 9.2 mL of trimethylsilylacetylene (TMS) were added to a dry 250 mL three-necked flask, then cooled to -40 to -80 °C, and 20 mL of n-buLi was added to the solution and stirred for 30 minutes. 7 mL of tetramethylethylenediamine (TEMED) and 6 g of the diketone product (c) from Reaction Step II of Method I were dissolved in 42 mL of THF, then cooled to -40 to -80 °C. The two solutions were mixed together and reacted overnight. The reaction was stopped with an NH4Cl solution. 100 ml of ethyl acetate was added to the flask and stirred for 5 minutes. The organic phase was dried to obtain 4.8 g of product (d-2).

[0160] Reaction Step IV-2: 4.8 g of product (d-2) was dissolved in a mixture of 15 ml of THF and 17 ml of tetrabutylammonium fluoride (TBAF) solution and reacted for 3 hours. THF was distilled off. The 2α,17α-diethynyl-A-nor-5a-androstane-2β,17β-diol product α-anordiol (e-2) was dissolved in ethyl acetate. The organic solution was washed 3 times with distilled water. After removing the ethyl acetate, 3.5 g of the 2α,17α-diethynyl-A-nor-5a-androstane-2β,17β-diol product, α-anordiol (e-2) was obtained.

[0161] Reaction Engineering V: 2.8 g of 2α,17α - diethynyl - A - nor - 5α - androstane - 2β,17β - diol product (e - 2), 4.5 mL of propionic acid, and 7.8 mL of propionic anhydride were mixed in a 100 mL flask and then stirred for 3 hours. 30 mL of H2O and 40 mL of ethyl acetate were added to the flask, stirred, and the water and organic phases were separated. The organic phase was washed 5 times with distilled water, and 10 mL of n - hexane was added. The residue was filtered and dried to obtain 2.0 g of a mixture of the product α - nor - drolone (fα).

[0162] Silica gel chromatography analysis showed that only one diastereomeric compound was synthesized using Method II (Figure 1B). In contrast, when Method I was used, a mixture of diastereomeric compounds was synthesized (Figure 1A).

[0163] NMR Analysis: Fractions 1 (F1) and 2 (F2) were collected and dissolved in CCl4 (50 mg / mL) for NMR analysis. Figures 2A and 2B showed α - nor - drolone from fraction 1 (F1) and β - nor - drolone from fraction 2 (F2), respectively. Example 2: Effects of α - nor - drolone or β - nor - drolone

[0164] In this example, the estrogenic effects of α - and β - nor - drolone on estrogen - modulated metabolic signaling and tissue atrophy have been shown by research. The ovaries of 7 - week - old mice were surgically removed. One week after the surgery, the mice were given isoflavones, α - nor - drolone, or β - nor - drolone in their food. Body weight, blood glucose, and food intake were measured monthly. After 3 months, the mice were sacrificed. The legs and vertebrae, liver, uterus, and vagina were collected.

[0165] The wet weights of the uterus and vagina and H&E staining showed that α-androline, rather than β-androline, prevented vulvovaginal atrophy (Figure 4). α-Androline-treated OVX mice showed milder uterine atrophy symptoms compared to mice treated with β-androline and the blank group (Figure 3). α-Androline showed lower blood glucose, TG, and body weight compared to β-androline (Figure 5A and Figure 5B), respectively, but there were no significant differences in the initial body weight and food intake (Figure 5C and Figure 5D), respectively. These data indicated that the differences were due to changes in energy consumption. Liver H&E sections and statistical analysis showed that α-androline, rather than β-androline, decreased the amount of liver TG in OVX mice (Figure 6). Micro-CT results showed that α-androline, rather than β-androline, prevented osteoporosis in OVX mice (Figure 7).

[0166] Tamoxifen was the first FDA-approved drug for breast cancer patients with positive expression of ER. However, tamoxifen also induces side effects such as endometrial cancer and NASH. Importantly, in the case of women with ER-positive (ER + ) cancer, continuing tamoxifen treatment for up to 10 years instead of stopping it after 5 years results in a further reduction in recurrence and mortality, especially after 10 years. The inventors found that androline can eliminate tamoxifen-induced endometrial epithelial cell (EEC) mitosis. The inventors further tested whether α-androline or β-androline can eliminate tamoxifen-induced EEC mitosis. Foods containing isoflavone (blank), α-androline (a-ANO), β-androline (b-ANO), α-androline + tamoxifen (TAM + a-ANO), β-androline + tamoxifen (TAM + b-ANO), or tamoxifen (TAM) were given to 7-week-old mice for 6 months. Then, the mice were sacrificed and the uterus and liver were recovered. The uterus and liver tissues were fixed.

[0167] By H&E staining sections, EECs in the TAM + b-ANO and TAM groups were rough and mitotic compared to the smooth monolayer EECs in the TAM + a-ANO and sham groups (Figure 8A), and lipid depositions in hepatocytes of the TAM + b-ANO and TAM groups were shown to be higher compared to hepatocytes of the TAM + a-ANO and sham groups (Figure 9A). Statistical analysis of H&E staining sections showed that α-anordrin, rather than β-anordrin, inhibited tamoxifen-induced EEC mitosis (Figure 8B) and NASH (Figure 9B).

[0168] An aromatase inhibitor was administered to ER-positive breasts in postmenopausal women. It also induced side effects such as tissue atrophy and osteoporosis. Anastrozole is an aromatase inhibitor. It was administered to ER-positive breast cancer in postmenopausal women. Anastrozole (ANA) also induced vaginal atrophy and osteoporosis. Vulvovaginal atrophy usually results in vaginal and urinary tract infections. As an example, the inventors tested whether α-anordrin or β-anordrin eliminates anastrozole-induced uterine and vulvovaginal atrophy and osteoporosis. Foods containing isoflavone (blank), α-anordrin (a-ANO), β-anordrin (b-ANO), α-anordrin + anastrozole (ANA + a-ANO), β-anordrin + anastrozole (ANA + b-ANO) or anastrozole (ANA) were given to 7-week-old mice for 6 months. Then the mice were sacrificed and the vagina, legs and vertebrae were collected. The tissues were fixed with paraformaldehyde. Micro-CT results showed that α-anordrin, rather than β-anordrin, inhibited anastrozole-induced osteoporosis (Figures 10A, 10B and 10C). Statistical analysis of the wet weight of the vagina and H&E staining sections of the vulvovagina showed that α-anordrin, rather than β-anordrin, inhibited anastrozole-induced vaginal atrophy (Figures 11A, 11B and 11C). Example 3: Materials and Methods for Preparing Materials

[0169] Tamoxifen and anastrozole (MPG USP grade) were provided by Okahata (Shanghai) Trading Co., Ltd. Epiandrostenestone was purchased from Jiangsu Jiaerke Pharmaceuticals Group, LTD. All other compounds were purchased from Sigma or Aladdin. Mouse food was prepared by Trophic Animal Feed High-tech Co., Ltd, Nantong, China. Mice were purchased from BK animal Inc. Animal experiments were carried out at the Southern Center of Pesticide Research, Shanghai.

[0170] Glucose concentration assay: The glucose concentration in the medium or whole blood from mouse tails was measured using a glucose assay kit according to the manufacturer's instructions (Yicheng, Beijing).

[0171] Construction of ovariectomized (OVX) mouse model and drug administration: The ovaries of 6-week-old mice were surgically removed. Three days after the surgery, the drug was administered by daily gavage or mixed with food.

[0172] Preparation of paraffin sections and HE staining: Mouse tissues were excised surgically and fixed using 4% paraformaldehyde (Beijing Solar Bioscience & Technology co., Ltd) in 1×DPBS. Preparation of paraffin sections and HE staining were carried out at the GLP laboratory of BK animal model, Inc.

[0173] Measurement of TG in mouse liver: 30 - 50 mg of mouse liver was excised surgically and 1 Homogenized in 1 ml of chloroform: methanol (2:1) mixture and extracted with 0.5 ml of ddH2O. The organic phase was transferred to a new tube and air-dried. The amount of TG was measured using a kit according to the manufacturer's instructions. The error was corrected using an internal standard control.

[0174] Measurement of the height of the EEC, the diameter of the uterus, and the thickness of the circular muscle: The height of the EEC was measured from EEC images at magnifications of 20 - 200 times of paraffin-embedded H&E mouse uterine sections.

[0175] Measurement of the thickness of the vulvovaginal wall: The vagina was cut at the center. For the preparation of H&E sections, both the uterine site and the vulvovaginal site were embedded in paraffin in the same direction. The thickness of the vulvovaginal wall was measured from an image at a magnification of 40 times of paraffin-embedded H&E mouse vaginal sections.

[0176] Bone density assay using micro-CT: The femurs were fixed in 1×DPBS containing 3% formaldehyde for 2 weeks. After 1 week, the fixative was changed. The density of the femurs was measured by Siemens Inveon micro-CT. The HU2000 value was analyzed using Inveon Research Workplace (IRW) under the following measurement conditions: 80 KVP, 500 mA, exposure time 1500 milliseconds; CCD readout settings: axial 2048, binning 2048; FOV transaxial: 19.03 mm, axial: 19.03 mm, pixel size: 9.29 μm.

[0177] Statistical analysis: In tables and figures, the results were presented as mean + STDEV. Asterisks indicate statistically significant differences calculated using a two-sided Student's t-test. References: 1. Boonyaratanakornkit, V., Steriods. 76, 877 - 884, (2011). 2. Mauvais-jarvis, F., Clegg, DJ., and Hevener, AI., Endocrine Reviews. 34(3), 309 - 338 (2013) 3. Kuang, LG., Zhang, XT., Xie, Y., et al., Molecular endocrinology. 24(4), 709 - 721 (2010) 4. Rao, J., Jiang, XM., Wang, Y., and Chen, B.. Journal of Steroid Biochemistry and Molecular Biology. 127, 231 - 237 (2011) 5. Revankar, CM., Cimno D., Sklar, LA., Arterburn, JB., and Prossnitz, ER.. Science. 307(11),1625 - 1630(2005) 6. Nilsson, BO., Olde, B., and Leeb - Lundberg, LF.. British Journal of Pharmacology. 163. 1131 - 1139 (2011) 7. Zhang, XT., Ding, L., Kang, LG., and Wang, Y.. PLos one. 7(1), e30174 (2012) 8. O’Brien, JE., Peterson, TJ., Tong, MH., et al. J. of Biol. Chem. 281(36). 26683 - 26692 (2006) 9. Takamura, T., shimizu, A., Kumura,T., Ando, K Zen, Y., et al. Internal Med. 579 - 581 (2007) 10. Barrett - Connor, E., Mosca, L., Collins, P., et al.. N Engl. J. Med.. 355. 125 - 137 (2006) 11. Alexanderson, P., Toussaint, A., Christiansen, C., et al.. JAMA. 285(11). 1482-8 (2001) 12. Hershberger, PA., Stabile, LP., Kanterewicz, B., et al.. J. of Steroid Biochem. & Mol. Biol. 16. 102-9 (2009) 13. Bank, U.K. and Pincus, U., Proc. Soc. Expt. Biol. Med. 111, 595 (1962). 14. Pincus, U. and Gordon, H.L., Steroids, 5, 193, (1965). 15. Xu, B., Zhou, P.Q., and Yu, W.J., Tumor, 9, 197 (1989). 16. Ma, Z.C., Lou, L.G., Zhang, Z., and Bin, X, Acta Pharmacol. Sin, 21, 939 (2000). 17. Li R.L., Lee, D.Y., Cheng, Q.L., US Patent 5001120 (1991). 18. Mehta, R.R., Jenco, J.M., and Chatterton, R.T., Steroids, 38, 679 (1981). 19. Li ZH, Li L. X-ray diffraction studies on the absolute configuration of alpha- and beta-anordrins. Steroids. 1990 Dec;55(12):565-70. 20. Nehra, R., Riggins, RB., Shajahan, AN et al.. FASEB J. 24(6). 2040-55 (2010). 21. Gu W, Xu W, Sun X, Zeng B, Wang S, Dong N, Zhang X, Chen C, Yang L, Chen G, Xin A, Ni Z, Wang J, Yang J. Anordrin Eliminates Tamoxifen Side Effects without Changing Its Antitumor Activity. Sci Rep. 2017 Mar 7;7:43940.

[0178] According to a preferred embodiment of the present invention, for example, the following is provided. (Item 1) Formula (I):

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Chem.

Claims

1. A pharmaceutical composition comprising a substantially pure diastereomeric compound of formula (I): 【Chemical 24】 or a salt thereof, wherein in the formula R １ is -OH or -OC(O)-R １ａ wherein R ４ is -OH or -OC(O)-R ４ａ wherein, R １ａ and R ４ａ are ethyl; R ２ and R ５ is -C≡CH; R ６ and R ７ are independently C １ -C ６ alkyl, C ２ -C ６ alkenyl, C ２ -C ６ alkynyl or C ３ -C ６ cycloalkyl; R ８ and R ９ are each independently hydrogen, -OH, -NH ２ , -NO ３ , halogen, C １ -C ６ alkyl, C ２ -C ６ alkenyl, C ２ -C ６ alkynyl, C ３ -C ６ cycloalkyl, C ６ -C １４ aryl, 5- to 6-membered heteroaryl or 3- to 6-membered heterocyclyl, wherein said substantially pure diastereomeric compound has a diastereomeric excess (de) of 99.9% or more. Pharmaceutical composition.

2. The pharmaceutical composition according to claim 1, further comprising at least one additional agent, wherein said additional agent is selected from the group consisting of selective estrogen receptor modulators and aromatase inhibitors.

3. The pharmaceutical composition according to claim 2, wherein said additional agent is tamoxifen.

4. The pharmaceutical composition according to claim 2, wherein said additional agent is selected from the group consisting of raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene.

5. The pharmaceutical composition according to claim 2, wherein said additional agent is an aromatase inhibitor.

6. The pharmaceutical composition according to claim 5, wherein said aromatase inhibitor is anastrozole.

7. The pharmaceutical composition according to claim 5, wherein said aromatase inhibitor is selected from the group consisting of anastrozole, letrozole, exemestane, vorozole, formestane, and fadrozole.

8. The pharmaceutical composition according to any one of claims 2 to 7, wherein the weight ratio of the substantially pure diastereomeric compound of formula (I) or a salt thereof to said additional agent in said composition is from 20:1 to 1:

20.

9. Said substantially pure diastereomeric compound is of formula (Ia) 【Chemical 25】 (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone). The pharmaceutical composition according to any one of claims 2 to 8.

10. A composition for the treatment of estrogen deficiency in an individual, comprising the pharmaceutical composition according to claim 1.

11. A composition for preventing or alleviating estrogen deficiency symptoms in an individual, comprising the pharmaceutical composition according to claim 1.

12. The composition according to claim 11, wherein the estrogen deficiency symptoms are selected from the group consisting of high liver triglycerides, osteoporosis, vulvovaginal atrophy, high blood triglycerides, high blood glucose, and weight gain.

13. The composition according to claim 11, wherein at least one additional agent is further administered to the individual, and the additional agent is selected from the group consisting of a selective estrogen receptor modulator and an aromatase inhibitor.

14. The composition according to claim 13, wherein the additional agent is tamoxifen.

15. The composition according to claim 13, wherein the additional agent is selected from the group consisting of raloxifene, lasofoxifene, bazedoxifene, arzoxifene, ormeloxifene, ospemifene, and levormeloxifene.

16. The composition according to any one of claims 13 to 15, wherein the estrogen deficiency symptom is selected from the group consisting of high liver triglycerides, osteoporosis, vulvovaginal atrophy, high blood triglycerides, high blood glucose, and weight gain.

17. The composition according to any one of claims 13 to 16, wherein the composition and the additional agent are administered sequentially.

18. The composition according to any one of claims 13 to 16, wherein the composition and the additional agent are administered simultaneously.

19. A composition for reducing the side effects of at least one additional agent in an individual, comprising the pharmaceutical composition according to claim 1, wherein the composition is administered in combination with the additional agent, and the additional agent is selected from the group consisting of a selective estrogen receptor modulator and an aromatase inhibitor.

20. The composition according to claim 19, wherein the additional agent is tamoxifen.

21. The composition according to any one of claims 10 to 20, wherein the individual is a human.

22. R ６ and R ７ The composition according to any one of claims 10 to 21, wherein both are methyl.

23. R ８ and R ９ are each independently hydrogen or C １ -C ６ alkyl, the composition according to any one of claims 10 to 22.

24. R ８ and R ９ The composition according to any one of claims 10 to 23, wherein both are hydrogen.

25. The substantially pure diastereomeric compound is of formula (Ia) 【Chemical 23】 (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone) of the composition according to any one of claims 10 to 20.

26. The estrogen deficiency symptom is high liver triglycerides, and the substantially pure diastereomeric compound is of formula (Ia) 【Chemical 26】 The composition according to claim 11, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

27. The estrogen deficiency symptom is high blood triglyceride, and the substantially pure diastereomer compound is of formula (Ia) 【Chemical 27】 The composition according to claim 11, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

28. The estrogen deficiency symptom is osteoporosis, and the substantially pure diastereomer compound is of formula (Ia) 【Chemical 28】 The composition according to claim 11, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

29. The estrogen deficiency symptom is vulvovaginal atrophy, and the substantially pure diastereomer compound is of formula (Ia) 【Chemical 29】 The composition according to claim 11, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

30. The estrogen deficiency symptom is high blood glucose, and the substantially pure diastereomer compound is of formula (Ia) 【Chemical Formula 30】 The composition according to claim 11, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

31. The estrogen deficiency symptom is weight gain, and the substantially pure diastereomer compound is of formula (Ia) 【Chemical 31】 The composition according to claim 11, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

32. The further agent is tamoxifen, and the substantially pure diastereomer compound is of formula (Ia) combined with tamoxifen 【Chemical 32】 The composition according to claim 19, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

33. The further agent is an aromatase inhibitor, and the substantially pure diastereomer compound is of formula (Ia) combined with the aromatase inhibitor 【Chemical 33】 The composition according to claim 19, which is (2α,17α)-diethynyl-(2β,17β)-diol-dipropionate-A-nor-5α-androstane (α-nordrolone).

Images