Phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid derivatives for use as 5-alpha reductase antagonists in therapeutic methods - Patent Application 20070122997

Abstract

The present invention provides compounds that are phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid derivatives for use as 5-alpha reductase antagonists, particularly antagonists of type 1 and type 2. The present invention further provides pharmaceutical compositions comprising these compounds and the use of these compounds and compositions in the treatment of diseases or disorders, particularly, but not limited to, the treatment of benign prostatic hyperplasia and androgenetic alopecia.

Description

[Technical Field]

[0001] introduction The present invention relates to compounds that are phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid derivatives for use as 5-alpha reductase antagonists. The present invention further provides pharmaceutical compositions containing these compounds and the use of these compounds and compositions in the treatment of diseases or disorders, particularly, but not limited to, androgenetic alopecia and benign prostatic hyperplasia. [Background technology]

[0002] Androgenetic alopecia ("AGA"), commonly known as male pattern baldness, is a highly prevalent condition across all populations, primarily affecting men and, to a lesser extent, women. AGA is non-scarring and mediated by the sensitivity of hair follicles to androgens and an individual's genetic predisposition. While not fatal, it has potentially adverse psychosocial sequelae that can reduce quality of life.

[0003] AGA is mediated by the conversion of testosterone to dihydrotestosterone (DHT) by 5-alpha reductase type 2 (SRD5A2). Increased DHT regulation leads to the perifollicular miniaturization, inflammation, and fibrosis characteristic of AGA, ultimately resulting in hair loss in AGA-susceptible areas of the scalp. In addition to its role in AGA, the predominance of DHT in the prostate derived from 5-alpha reductase type 1 (SRD5A1) may play a beneficial role in the development of benign prostatic hyperplasia, which may later lead to prostate cancer. The literature suggests that SRD5A1 and SRD5A2 are responsible for the production of one-third and two-thirds of circulating DHT, respectively.

[0004] Current treatment strategies for these DHT-mediated diseases include three Food and Drug Administration (FDA)-approved medications. AGA is treated with the vasodilator minoxidil and the 5-alpha reductase type 2 (SRD5A2) inhibitor finasteride. Benign prostatic hyperplasia (BPH) is treated with the 5-alpha reductase type 1 (SRD5A1) inhibitor dutasteride and finasteride, which also inhibits SRD5A1. However, finasteride is 100 times more selective for SRD5A2 than SRD5A1. Furthermore, minoxidil and finasteride are thought to be more effective when used in combination, and long-term use, with a minimum of 4–6 months of treatment required to see substantial improvement. While long-term drug administration is essential for effective treatment, it often leads to poor adherence and other side effects that persist even after discontinuation of the medication. Therefore, the need for alternative treatments for AGA, especially improved drug compounds targeting SRD5A2, necessitates further research into novel effective drug molecules with fewer drawbacks and side effects.

[0005] SRD5A2 is a G protein-coupled receptor ("GPCR") whose structure, consisting of a typical seven transmembrane (TM) domain helix, has recently been elucidated. GPCRs are the largest family of proteins encoded by the human genome and are the primary therapeutic targets for over 50% of currently marketed drugs. These molecules bind to GPCRs as either agonists or antagonists. Agonists are drugs that bind to receptors and elicit a response similar to that of the natural substrate, upregulating cellular responses. Antagonists bind to receptors at orthosteric or allosteric sites, inhibiting receptor responses.

[0006] The active site of SRD5A2 contains six cytoplasmic loops (L) and two independent tunnel-like pockets for NADP and DHF. The entrance to the ligand binding site is between TM1 and TM4. Loop 1 (L1) controls NADPH / NADP+ exchange from the cytoplasm. The active site contains two catalytic residues, E57 and E67. TM2 and Y91 TM3These residues convert NADPH to testosterone (E57 TM2 and Y91 TM3 ) promotes hydride transfer to the C-5 atom of testosterone to form an enolized intermediate, followed by the Δ 4,5 The bond is reduced, releasing DHT and NADP+.

[0007] The mechanism of inhibition of SRD5A2 is the conversion of NADPH (4-pro-(R)-hydride) to finasteride's Δ 1,2 This involves the transfer of a hydride to the bond, forming an enolized adduct of NADPH and finasteride (NADP-DHF). This covalent bond between NADP and DHF inhibits the transfer of hydride from NADPH to testosterone, preventing the production of DHT. The dissociation constant of the covalently bound NADP-DHF adduct is Ki = koff / kon ≤ 1 x 10 -13 M is one of the most potent non-covalent inhibitors known for any enzyme. The protein interacts with the adduct through three key residues: E57, R114, and F118. TM2 interacts with finasteride, and R114 and F118 interact with the NADP moiety of NADP-DHF.

[0008] Finasteride acts as a competitive and specific inhibitor of SRD5A1 and 2, preventing the conversion of testosterone to its more active metabolite, 5-alpha-dihydrotestosterone. However, systemic finasteride use is associated with serious side effects.

[0009] It is therefore an object of the present invention to provide alternative therapeutic agents that inhibit 5-alpha reductase with improved selectivity and specificity.

[0010] It is a further object of the present invention to provide novel therapeutic agents with improved selectivity for the inhibition of 5-alpha reductase, particularly 5-alpha reductase type 2. Summary of the Invention

[0011] According to a first aspect to the present invention, there is a pharmaceutical composition comprising a compound of formula (I), or a pharmaceutically acceptable salt, stereoisomer, hydrate or solvate thereof, and a pharmaceutically acceptable carrier.

[0012] [ka]

[0013] During the ceremony, R 1 , R 1’ , R 2 , R 2’ , and R 3 are each independently H, halogen, or -OR 5 is selected from the group consisting of R 5 is selected from substituted or unsubstituted C1-C2 alkyl, substituted or unsubstituted linear or branched C3-C6 alkyl; R 4 is selected from substituted or unsubstituted C1-C2 alkyl, substituted or unsubstituted, linear or branched C3-C6 alkyl, and substituted or unsubstituted C3-C6 cycloalkyl.

[0014] In one embodiment, R 1 , R 1’ , R 2 and R 2’ is H and R 3 -OR 5 is.

[0015] In one embodiment, R 1 or R 1’ One of the is -OR 5 and R 2 , R 2’ and R 3 is H.

[0016] In one embodiment, R 2 or R 2’ One of them is -OR 5 and R1 , R 1’ , and R 3 is H.

[0017] In one embodiment, R 1 , R 1’ , R 2 , R 2’ , and R 3 At least one of the groups is a halogen.

[0018] In one embodiment, R 1 , R 1’ , R 2 , R 2’ and R 3 One of them is a halogen and the rest are H.

[0019] In a further embodiment of the present invention, R 4 is selected from unsubstituted C1-C2 alkyl, unsubstituted straight or branched C3-C6 alkyl, and unsubstituted C3-C6 cycloalkyl.

[0020] According to a second aspect to the present invention there is provided the use of a compound or pharmaceutical composition of the present invention as a medicament.

[0021] According to a third aspect to the present invention, there is provided for the use of a compound or pharmaceutical composition of the present invention in a method of treating a disease or disorder, the method comprising administering to a subject in need thereof a pharmaceutically effective amount of the compound or composition.

[0022] According to a further aspect of the present invention there is provided the use of a compound of the present invention in the preparation of a medicament for treating a disease or disorder, said treatment comprising administering a pharmaceutically effective amount of said medicament to a subject in need thereof.

[0023] According to a further aspect of the present invention, there is provided a method of treating a disease or disorder in a patient, the method comprising administering to a subject in need thereof a compound or pharmaceutical composition of the present invention.

[0024] In one embodiment, the disease or disorder is a disease or disorder mediated by high levels of dihydrotestosterone.

[0025] In one embodiment, the disease or disorder is benign prostatic hyperplasia or androgenetic alopecia.

[0026] In a preferred embodiment, the disease or disorder is benign prostatic hyperplasia.

[0027] In a preferred embodiment, the disease or disorder is androgenetic alopecia.

[0028] In a preferred embodiment, the subject is a human.

[0029] The invention will now be described in more detail with reference to the following non-limiting embodiments and figures. [Brief explanation of the drawings]

[0030] [Figure 1] FIG. 1 shows the pose and intermolecular interactions of compound Bsy1. [Figure 2] FIG. 2 shows the relative expression of the SRD5A gene (relative to HaCaT) in various cell lines (the SRD5A2 gene is expressed in all cell lines; SRD5A2 is highly expressed in HepG2, HE293, and DU-145). [Figure 3] Figure 3 shows the IC50 hyperbola of DU-145-drugs. IC50 values: finasteride (0.72 ± 1.09 nM); dutasteride (1.67 ± 1.85 μM); Bsy1 (0.01 ± 0.015 nM). [Figure 4] Figure 4 shows the HEK293-drug IC50 hyperbolic curves. IC50 values: finasteride (1.24 ± 1.43 nM); dutasteride (1.21 ± 1.37 μM); Bsy1 (0.02 ± 0.03 nM). [Figure 5]FIG. 5 shows the relative expression of the SRD5A gene after treatment of the DU-145 cell line with IC50 values ​​of finasteride (0.9 nM), dutasteride (1.6 μM) and Bsy1 (0.0054 nM). [Figure 6] FIG. 6 shows the relative expression of the SRD5A gene after treatment of the HEK293 cell line with IC50 values ​​of finasteride (1.34 nM), dutasteride (1.29 μM) and Bsy1 (0.02 nM). DETAILED DESCRIPTION OF THE INVENTION

[0031] The present invention will now be described more fully hereinafter with reference to the accompanying drawings, which show some non-limiting embodiments of the invention.

[0032] The invention described below should not be construed as limited to the particular embodiments disclosed, as it includes certain modifications and other embodiments that are intended to be included within the scope of the invention.

[0033] Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.

[0034] As used herein, throughout this specification and in the claims that follow, the singular forms "a," "an," and "the" include the plural forms unless the context clearly dictates otherwise.

[0035] The terms and phrases used herein are for purposes of description and should not be regarded as limiting. As used herein, the use of the terms "comprising," "containing," "having," "including," and variations thereof, is meant to encompass the items listed thereafter, and equivalents thereof, as well as additional items.

[0036] When describing the present invention, including pharmaceutical compositions containing particular compounds, and methods of using such compounds and compositions, the following terms, when present, have the following meanings unless otherwise indicated. It should also be understood that, as described herein, any of the moieties defined below may be substituted with various substituents, and that each definition is intended to include such substituted moieties within its scope as defined below. In this regard, unless otherwise indicated, the term "substituted" is defined as defined below. It should further be understood that the terms "group" and "radical" are considered interchangeable when used herein.

[0037] "Alkyl" means a straight or branched chain aliphatic hydrocarbon group having the specified number of carbon atoms. Particular alkyl groups have 1 to 6 carbon atoms. More particular are lower alkyls having 1 to 4 carbon atoms. Even more particular are groups having 1 to 3 carbon atoms. Exemplary straight chain groups include methyl, ethyl, n-propyl, n-butyl, and the like. Branched chain means that one or more lower alkyl groups, such as methyl, ethyl, propyl, butyl, and the like, are attached to a linear alkyl chain. Exemplary branched chain groups include isopropyl, isobutyl, and the like.

[0038] "Alkoxy" refers to, for example, the group -OR 5 where R 5 is alkyl having the specified number of carbon atoms. Particular alkoxy groups are methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, n-hexoxy, and 1,2-dimethylbutoxy. Particular alkoxy groups are lower alkoxy, i.e., having 1 to 6 carbon atoms. Further particular alkoxy groups have 1 to 4 carbon atoms.

[0039] "Alkylene" refers to a divalent alkene radical group having the specified number of carbon atoms, particularly 1 to 6 carbon atoms, more particularly 1 to 4 carbon atoms, which may be straight or branched. This term is exemplified by groups such as methylene (-CH-), ethylene (-CH-CH-), or -CH(CH)-.

[0040] "Alkenyl" refers to a monovalent olefinically unsaturated hydrocarbon group having the specified number of carbon atoms. Particular alkenyls have from 2 to 8 carbon atoms, more particularly from 2 to 6 carbon atoms, may be straight-chained or branched, and have at least one site of olefinic unsaturation, especially one or two sites. Particular alkenyl groups include ethenyl (-CH=CH), n-propenyl (-CHCH=CH), isopropenyl (-C(CH)=CH), and the like.

[0041] "Aryl" refers to a monovalent aromatic hydrocarbon group derived by removing one hydrogen atom from one carbon atom of a parent aromatic ring system. In particular, aryl refers to a monocyclic or polycyclic aromatic ring structure, where the number of ring atoms is specified. Specifically, the term includes groups containing 6 to 10 ring members. When the aryl group is a monocyclic ring system, it preferably contains 6 carbon atoms. Particular aryl groups include phenyl, naphthyl, and the like. The terms "phenyl" and "Ph" are used interchangeably herein unless otherwise indicated.

[0042] "Cycloalkyl" refers to a monocyclic or polycyclic non-aromatic hydrocarbyl ring structure, specifying the number of ring atoms. Cycloalkyls may have from 3 to 10 carbon atoms, specifically from 3 to 6 carbon atoms. Such cycloalkyl groups include, by way of example, single ring structures such as cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0043] "Halo" or "halogen" refers to fluoro (F), chloro (Cl), bromo (Br), and iodo (I). Specifically, a halo group is fluoro, bromo, or chloro.

[0044] "Hetero," when used to describe a compound or a group present on a compound, means that one or more carbon atoms in the compound or group are replaced with a nitrogen, oxygen, or sulfur heteroatom. Hetero can apply to any of the above hydrocarbyl groups, such as alkyl, e.g., heteroalkyl, cycloalkyl, e.g., heterocycloalkyl, and aryl, e.g., heteroaryl, having 1 to 4, particularly 1, 2, or 3, heteroatoms, more typically 1 or 2, heteroatoms, e.g., a single heteroatom.

[0045] "Heteroaryl" refers to a monocyclic or fused polycyclic aromatic ring system containing one or more heteroatoms independently selected from O, N, and S and a specified number of ring atoms. In particular, the aromatic ring system can have 5 to 9 ring members. A heteroaryl group can be, for example, a 5- or 6-membered monocyclic ring, or a fused bicyclic system formed from a fused 5-membered ring and a 6-membered ring, or two fused 6-membered rings, or, as a further example, two fused 5-membered rings. Each ring can contain up to four heteroatoms, typically selected from nitrogen, sulfur, and oxygen. Typically, a heteroaryl ring contains up to four heteroatoms, more typically up to three heteroatoms, and more usually up to two heteroatoms, e.g., a single heteroatom. In one embodiment, a heteroaryl ring contains at least one ring nitrogen atom. The nitrogen atom of a heteroaryl ring can be basic, as in the case of imidazole or pyridine, or essentially non-basic, as in the case of the nitrogen of indole or pyrrole. The nitrogen atoms in the heteroaryl ring can be basic, as in the case of an imidazole or pyridine, or essentially non-basic, as in the case of an indole or pyrrole nitrogen. Generally, the number of basic nitrogen atoms present in the heteroaryl group, including any amino group substituents on the ring, will be fewer than 5.

[0046] As used herein, the term "heterocycloalkyl" refers to a stable, monocyclic or polycyclic, non-aromatic ring structure containing one or more heteroatoms independently selected from O, N, and S. The non-aromatic ring structure can have 4 to 10 ring members, particularly 4 to 6 ring members. A fused heterocyclic ring system can contain a carbocyclic ring or may contain one heterocyclic ring. As used herein, the term "heterocycloalkenyl" refers to a "heterocycloalkyl" in which one bond in the ring is reduced, so that the ring consists of a double bond.

[0047] "Substituted" refers to a group in which one or more hydrogen atoms are each independently replaced with the same or different substituent(s).

[0048] As used herein, the term "substituted with one or more" refers to 1 to 4 substituents. In one embodiment, it refers to 1 to 3 substituents. In a further embodiment, it refers to 1 or 2 substituents. In a further embodiment, it refers to 1 substituent.

[0049] "Pharmaceutically acceptable" means approved or approvable by a regulatory body, such as the U.S. Food and Drug Administration, or a similar body in a country other than the United States, or listed in a generally recognized pharmacopoeia, such as the United States Pharmacopoeia, for use in animals, and more particularly in humans.

[0050] "Pharmaceutically acceptable salts" refer to salts of the compounds of the present invention that are pharmaceutically acceptable and possess the desired pharmacological activity of the parent compound. In particular, such salts are non-toxic and may be inorganic or organic acid addition salts and base addition salts. Specific examples include the following salts: (1) acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid; or acid addition salts formed with organic acids such as acetic acid and propionic acid: acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, and 4-toluenesulfonic acid. or (2) salts formed when an acidic proton present in the parent compound is replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth metal ion, or aluminum, or coordinates with an organic base such as ethanolamine, diethanolamine, triethanolamine, and N-methylglucamine. Salts further include, by way of example, sodium, potassium, calcium, magnesium, ammonium, and tetraalkylammonium; and, if the compound contains a basic functional group, salts of non-toxic organic or inorganic acids, including, by way of example, hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, and oxalate. The term "pharmaceutically acceptable cation" refers to an acceptable cationic counterion of an acidic functional group, examples of which include sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium cations, and the like.

[0051] "Pharmaceutically acceptable vehicle" refers to a diluent, adjuvant, excipient, or carrier with which a compound of the invention is administered.

[0052] "Prodrug" refers to a compound containing a derivative of the compound of the present invention that has a cleavable group and becomes a pharmaceutically active compound of the present invention in vivo by solvation or under physiological conditions. Examples of such a compound include, but are not limited to, choline ester derivatives, N-alkylmorpholine esters, etc.

[0053] "Solvate" usually refers to a form of a compound associated with a solvent through solvation. This physical association involves hydrogen bonding. Conventional solvents include, for example, water, ethanol, acetic acid, etc. The compound of the present invention may be prepared, for example, in a crystalline form, and then solvated or hydrated. Suitable solvates include pharmaceutically acceptable solvates, such as hydrates, and further include both stoichiometric and non-stoichiometric solvates. In certain embodiments, the solvate will be isolatable, for example, when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid. "Solvate" encompasses both solution-phase and isolatable solvates. Representative solvates include hydrates, ethanolates, methanolates, etc.

[0054] A "subject" includes a human. The terms "human," "patient," and "subject" are used interchangeably herein.

[0055] An "effective amount" means the amount of a compound of the present invention that, when administered to a subject for treating a disease, is sufficient to effectively effect such treatment for the disease. The "effective amount" can vary depending on the compound, the disease and its severity, and the age, weight, etc., of the subject being treated.

[0056] "Treating" or "treatment" of any disease or disorder includes ameliorating the disease or disorder, i.e., halting the disease or reducing the onset, extent, or severity of at least one of its clinical symptoms. In another embodiment, "treating" or "treatment" refers to improving at least one physical parameter, which may not be discernible by the subject. In yet another embodiment, "treating" or "treatment" refers to modulating the disease or disorder, either physically (e.g., stabilization of discernible symptoms), physiologically (e.g., stabilization of physical parameters), or both. In a further embodiment, "treating" or "treatment" relates to slowing the progression of the disease.

[0057] In this specification, for example, C 1-4 When a range is referred to, such as alkyl, the recitation of the range should be considered a representation of each member of the range.

[0058] It should also be understood that compounds that have the same molecular formula but differ in the nature or sequence of bonding of their atoms or the arrangement of their atoms in space are termed "isomers." Isomers that differ in the arrangement of their atoms in space are termed "stereoisomers."

[0059] "Stereoisomers" that are not mirror images of one another are called "diastereomers," and those that are not mirror images of one another are called "enantiomers." When a compound has an asymmetric center, for example, if it is bonded to four different groups, a pair of enantiomers is possible. Enantiomers are characterized by the absolute configuration of their asymmetric center and are described by the R- and S-sequencing rules of Cahn and Prelog, or by the way the molecule rotates the plane of polarized light and are designated as dextrotrotatory or leborotrotatory (i.e., as (+)- or (-)-isomers, respectively). Chiral compounds can exist as individual enantiomers or as mixtures thereof. A mixture containing equal proportions of enantiomers is called a "racemic mixture."

[0060] "Tautomers" are interchangeable forms of a particular compound structure, differing in the displacement of hydrogen atoms and electrons. Thus, two structures may be in equilibrium due to the shifting of π electrons and atoms (usually H). For example, enols and ketones are tautomers, rapidly interconverting upon treatment with acid or base. Another example of a tautomer is the nitro and acetylphenylnitromethane forms, which are also formed upon treatment with acid or base. Tautomers can be relevant to achieving optimal chemical reactivity and biological activity of a desired compound.

[0061] The compounds of the present invention may have one or more asymmetric centers. Thus, such compounds can be prepared as individual (R)- or (S)-stereoisomers, or as mixtures thereof. Unless otherwise indicated, the description or naming of a particular compound in the specification and claims is intended to include both individual enantiomers and mixtures, racemic or otherwise. Methods for the determination of stereochemistry and the separation of stereoisomers are well known in the art.

[0062] It will be appreciated by those skilled in the art that compounds of the present invention can be metabolized to yield biologically active metabolites.

[0063] The present invention provides compounds that are phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid derivatives for use as 5-alpha reductase antagonists, particularly antagonists of type 1 and type 2. The present invention further provides pharmaceutical compositions comprising these compounds and the use of these compounds and compositions in the treatment of diseases or disorders, particularly, but not limited to, the treatment of benign prostatic hyperplasia and androgenetic alopecia.

[0064] According to a first aspect to the present invention, there is a pharmaceutical composition comprising a compound of formula (I), or a pharmaceutically acceptable salt, stereoisomer, hydrate or solvate thereof, and a pharmaceutically acceptable carrier.

[0065] [ka]

[0066] During the ceremony, R 1 , R 1’ , R 2 , R 2’ , and R 3 are each independently H, halogen, or -OR 5 selected from the group consisting of R 5 is selected from substituted or unsubstituted C1-C2 alkyl, substituted or unsubstituted linear or branched C3-C6 alkyl; R 4 is selected from substituted or unsubstituted C1-C2 alkyl, substituted or unsubstituted linear or branched C3-C6 alkyl, and substituted or unsubstituted C3-C6 cycloalkyl.

[0067] In one embodiment of the present invention, R 1 , R 1’ , R 2 , and R 2’ is H and R 3 HA-OR 5 is.

[0068] In one embodiment, R 1 or R 1’ One of them is -OR 5 and R 2 , R 2’ and R 3 is H.

[0069] In one embodiment, R 2 or R 2’ One of them is -OR 5 and R 1 , R 1’ , and R 3 is H.

[0070] In one embodiment, R 1 , R 1’ , R 2 , R 2’ , and R 3At least one of is a halogen.

[0071] In one embodiment, R 1 , R 1’ , R 2 , R 2’ , and R 3 One of them is a halogen and the rest are H.

[0072] In a further embodiment of the present invention, R 4 is selected from unsubstituted C1-C2 alkyl, unsubstituted straight or branched C3-C6 alkyl, and unsubstituted C3-C6 cycloalkyl.

[0073] Exemplary compounds included in the pharmaceutical compositions of the present invention can be represented by the chemical structures provided in Table 1 below.

[0074] [Table 1-1]

[0075] [Table 1-2]

[0076] [Table 1-3]

[0077] 1. In silico target identification, ligand preparation, docking, and toxicity screening 1.1 Target Identification and Validation The recently solved structure of SRD5A2 was co-crystallized with a dual steric ligand modeled as the adduct of finasteride and NADP (NADP-DHF). Analysis of this protein structure guided us in identifying and selecting compounds with potential for further testing and analysis.

[0078] 1.2 Ligand preparation and modeling According to the binding mechanism of finasteride, the finasteride-binding domain was analyzed using ChemDraw Professional software (Chemdraw RRID:SCR Using the NMR spectroscopy (DEM) algorithm, the structures of selected compounds were drawn in covalent complex with NADPH, and these compounds were docked as actual ligands. The mechanism of inhibition of SRD5A2 by finasteride is that SRD5A2 catalyzes hydride transfer from NADPH to the C-5 atom of finasteride, resulting in the formation of a covalent bond between NADPH and finasteride.

[0079] Modeled compounds were prepared for computational studies at physiological pH using the LigPrep module in Schrodinger suite v12.3 (Schrodinger, LLC, NY, USA, 2020). Ligprep generated multiple structures with various ionization states, tautomers, stereochemistries, and ring structures from each input structure. The OPLS-4 force field was used to minimize ligand geometry optimization and generate low-energy conformers of the ligands: prior to screening, 3D geometries were created, appropriate bond orders were assigned, and accessible tautomers and ionization states were generated. Epik in the LigPrep module was used to generate ionization / tautomerization states and calculate plausible ligand states.

[0080] 1.3 Preparation of target protein The coordinates and crystal structure of SRD5A2 were retrieved from the RCSB protein data bank (https: / / www.rcsb.org / ) under accession code 7BW1 at 2.8 Å X-ray resolution. The three-dimensional structure of SRD5A2 was generated using the Protein Preparation Workflow Wizard (PPWW) in Maestro v 12.3 (Schrodinger, LLC, NY, USA, 2020). Protein minimization restrained all atoms with a termination criterion based on the root mean square deviation of heavy atoms compared to their initial positions (RMSD = 0.3 Å). Water molecules outside a 5 Å radius from the ligand (hets) were removed.

[0081] 1.4 Generating the receptor grid The receptor grid was generated using the Ligand Docking with Energetics (Glide) receptor grid generation wizard with default settings (van der Waals radius scaling 1.0 Å, partial charge cutoff 0.25). The active site of the protein was identified as the center of mass of the workspace ligand preselected when defining the receptor. No constraints were defined during receptor grid generation, and only ligands with sizes similar to the workspace ligands were docked. Receptor grids for plant compounds were generated slightly differently from those for synthetic compounds. The first model treated NADPH in the substrate-binding cavity as part of the protein, while the second model removed NADPH from the binding cavity.

[0082] 1.5 Docking verification The docking model was validated by redocking the NADP-DHF adduct into the active site and ensuring that key interactions between the receptor and ligand were maintained. NADPH was fixed with the "CORE constraint." The pose of the redocked ligand was superimposed with the co-crystallized ligand.

[0083] 1.6 Molecular docking and compound screening The selected compounds and two positive control compounds (finasteride and dutasteride) were docked into the SRD5A2 binding site using the previously created receptor grid. Molecular docking was performed using the Glide docking module in Maestro, with the OPLS-4 force field and a curated library. The Glide docking constraint was 0.3:1, and the "core" constraint was set when docking compounds, fixing the NADP moiety of the adduct so that the docking pose had NADP in a fixed position. The Glide score (G score) was read using the "Standard Precision" (SP-visualizer module), and compounds were ranked based on the docking score, which represents the binding energy.

[0084] 1.7 Binding energy prediction Prime MM-GBSA (Prime model of the Schrodinger suite 2021-4) was used to predict the binding energies of the bound ligands by rescoring a priori ensembles of SRD5A2 poses generated from docking. MM-GBSA incorporates the OPLS-4 power field and a new energy dissolution model (VSGB) to perform calculations, incorporating a generalized surface area implicit model. The Prime MM-GBSA module ranks and displays the binding free energies of the docked compounds.

[0085] From the 58 compounds selected for docking experiments, eight compounds with favorable poses and conformations were obtained by Glide docking and MM-GBSA binding. Of these compounds, one compound, referred to herein as Bsy1, was selected from the eight compounds for further evaluation, along with phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid derivatives of Bsy1, referred to herein as Bsy2–Bsy10 (see Table 1).

[0086] 1.8 Toxicity prediction ProTox-II(http: / / tox.charite.de / protox II) was used to predict the toxicity and drug-like properties of the docked compounds.

[0087] The predicted toxicity profiles and drug-like properties of the selected compounds are shown in Table 2. According to the toxicity predictions of these compounds, the compounds belong to class 4 and have the LD of Bsy1. 50The bioavailability score was 418 mg / kg. Class 4 is considered harmful if swallowed according to GHS criteria and should be administered in the smallest possible amount. The Lipinski "rule of 5" (a set of physicochemical parameters that indicate a compound is likely to be a good oral drug) was used to describe the drug-like properties of compounds. Compound Bsy1 did not violate the Lipinski rules and had a bioavailability score of 0.56, making it a promising candidate for a therapeutic drug (bioavailability was measured by SwissADME; http: / / www.swissadme.ch / ).

[0088] [Table 2]

[0089] 1.9 Docking results and binding free energy of synthetic compounds Molecular docking of the initial 58 selected compounds resulted in 8 compounds with a Glide score below the cutoff value of -17 kcal / mol. Among the promising compounds, compound Bsy1 was selected for further evaluation and development at this time.

[0090] 1.10 Protein-Ligand Complexes After docking, refinement and rescoring were performed using Prime MMGBSA dG to describe the relative binding energies of each molecule (Table 3). The MM-GBSA dG binding scores of the compounds ranged from -128.45 kcal / mol to -157.73 kcal / mol.

[0091] The binding mode and residue interaction pattern of compound Bsy1 are shown in Figure 1. Figure 1A shows the binding pose and protein-ligand interaction (H-bond) between residues E57 and Y91, and the ππ interaction between F118 and the p-fluorophenyl group. Figure 1B shows the two-dimensional intermolecular interactions. Figure 1C shows the two-dimensional chemical structure of compound Bsy1. Bsy1 shows the interaction between residues E57 and Y91 (phenol group) via the carboxylate functional group of the reactive head. The binding strength is O カルボキシル -H...O=C カルボキシレート and Oフェニル -H...O カルボキシレート Furthermore, Bsy1 has ππ stacking interactions with the p-fluorophenyl group (Fig. 1B).

[0092] [Table 3]

[0093] 2. In-vitro screening and validation of Bsy1 and Bsy1 derivatives as SRD5A2 inhibitors 2.1 Evaluation of relative expression levels of SRD5A2 Seven tissue- or organ-related cell lines expressing endogenous SRD5A2 were selected and cultured to approximately 90% confluence for RNA extraction. The relative endogenous SRD5A2 gene expression levels in these cell lines were quantified using quantitative polymerase chain reaction (qPCR): (i) C4-2, a human prostate cancer cell line; (ii) HepG2, a human liver cancer cell line; (iii) A549, an adenocarcinoma human alveolar basal epithelial cell line; (iv) PC-3 and (v) DU-145, both human prostate cancer cell lines; (vi) HEK293, a human embryonic kidney cell line commonly used for recombinant expression; and (vii) HaCaTs, an immortalized human keratinocyte cell line.

[0094] The relative expression of the three SRD5A genes was compared by qPCR, and the SRD5A2 gene was approximately twice as expressed in all cell lines. The control cell line was HaCaT. The SRD5A2 gene was highly expressed in HepG2 (liver), DU-145 (prostate), and HEK293 (human embryo) (Figure 2). Statistical analysis using the Mann-Whitney t-test compared the expression levels of two cancer cell lines (HepG2 and DU-145). There was no statistically significant difference in SRD5A2 expression between HepG2 and DU-145 (p = 0.6667). Therefore, in this study, we chose the DU-145 cell line based on extensive reports of SRD5A2 cell-based assays. For selectivity purposes, we used a cancer cell line (DU-145) and a non-cancer cell line (HEK293) to screen for compounds that inhibit SRD5A2.

[0095] 2.2 Evaluation of the relative expression level of SRD5A2 after treatment with test compounds DU-145 and HEK293 cell lines were grown in DMEM supplemented with 10% (v / v) fetal bovine serum and 1% antibiotics (100 μg / mL streptomycin and 100 units / mL penicillin (P / S)) at 37°C in a 5% CO environment until 70% confluence.

[0096] For selected cell lines, half-maximal inhibitory concentrations (IC 50 ) was established. Using cell lines previously identified as having high endogenous expression levels of SRD5A2, the in vitro cytotoxic effects of Bsy1 on DU-145 and HEK293 cells were measured. Results were compared with those of the comparator compounds finasteride and dutasteride. The cytotoxic concentrations and cell viability of these agents were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, with the comparator compounds used as positive controls.

[0097] 1 x 10 cells in a 96-well plate 4Cells were seeded at a density of 100 cells / well and cultured for 24 hours in a 37°C, 5% CO2 incubator to allow cell attachment. 1% (v / v) DMSO / methanol in DMEM served as the control, and DMEM alone served as the blank. Cells were stained with 100 μl of 0.5 mg / ml MTT solution and incubated for 4 hours in a 37°C incubator under a 5% CO2 atmosphere. The medium was aspirated, and 100 μl of absolute ethanol was added to each well. Total proliferation and chemosensitivity were measured by the formation of tetrazolium salts. Absorbance was detected spectrophotometrically at 570 nm with a reference wavelength of 630 nm. Experiments were performed in triplicate.

[0098] IC of these cells 50 To determine values, dose responses (inhibitor vs. normalized response - variable slope curves) from non-linear curve plots were used using GraphPad Prism V 9.3.1.

[0099] The selectivity index showing the selectivity between normal cells and cancer cells is the IC 50 It was calculated from the ratio of the values ​​(Equation 1).

[0100]

number

[0101] Dose-response curves in the DU-145 cell line showed that Bsy1 showed antagonistic activity against SRD5A2 at 0.0054 nM (5.4 pM), compared with finasteride at 0.9 nM and dutasteride at 1.6 μM. In this study, nonlinear curve fitting with a hyperbolic model was used to determine the IC 50 The IC values ​​of the standard drugs were determined (Figure 3). 50 The IC values ​​obtained in other studies 50 These values ​​are comparable to those of the β-actin-dependent inhibitors of Bsy1, demonstrating that this assay can accurately predict the inhibitory kinetics and IC 50 It was suggested that this method is suitable for determining the value.

[0102] However, the IC of Bsy1 in HEK293 cell lines 50The antagonistic activity of Bsy1 against SRD5A2 was 0.02 nM, compared with 1.34 nM for finasteride and 1.29 μM for dutasteride (Figure 4).

[0103] IC of Bsy1 in DU-145 cancer cell line 50 The IC values ​​are for the non-cancer HEK293 cell line. 50 The HEK293 cell line was more resistant to Bsy1-containing compounds and showed lower efficacy than the DU-145 cell line.

[0104] The estimated selectivity index (Table 3, Equation 1) indicates a differential activity of compound Bsy1 of 3.7 units, which is significantly higher than that of finasteride (1.49 units) and dutasteride (0.81 units). It is generally accepted that values ​​greater than 1 indicate desirable selectivity toward cancer cells and may exhibit a better toxicity profile than values ​​less than 1.

[0105] [Table 4]

[0106] 2.3 Relative expression of the SRD5A gene in the DU-145 cell line The androgen-independent DU-145 cell line was treated with finasteride (0.9 nM), dutasteride (1.6 μM), and Bsy1 (0.0054 nM) for 24 hours, and the regulation of SRD5A gene expression was measured using qPCR. The test compounds induced a statistically significant downregulation of SRD5A2 (p ≤ 0.001) in DU-145 cells compared with the control (untreated) cells. The relative expression of SRD5A2 was reduced approximately three-fold by finasteride and dutasteride, and approximately two-fold by Bsy1 (Figure 5A). However, it should be noted that the concentration of finasteride was 100-fold higher than that of Bsy1, and that of dutasteride was 1,000-fold higher. Also, as can be seen in Figure 5A, the inhibition of SRD5A1 by finasteride and Bsy1 was comparable, even at a 100-fold higher concentration of finasteride than Bsy1. The level of inhibitory effect on SRD5A1 and SRD5A2 is surprising, as phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid Bsy1 and its derivatives, as described herein, would be expected to show similar inhibitory results. [Figure 5A - IC values ​​of finasteride (0.9 nM), dutasteride (1.6 μM), and Bsy1 (0.0054 nM) in the DU-145 cell line. 50 Relative expression of the SRD5A gene after treatment with values. ns' - Expression levels not statistically different between finasteride and compound Bsy1, and dutasteride and compound Bsy1. Experiments were repeated three times.]

[0107] To compare the gene suppression levels of each drug treatment relative to the control (untreated), a two-way analysis of variance (ANOVA) statistical analysis was performed with Dunnett's correction test (DCT) (Figure 5B). [Figure 5B-IC] 50 Two-way ANOVA Dunnett's test for control vs. treatment on values. Comparison test of control vs. drug treatment at p≦0.05: SRD5A1 (ns); SRD5A2**; SRD5A3 (ns). Data are expressed as mean ± standard error of the mean for each bar. ***P ≤ 0.001, *P ≤ 0.001. Significantly downregulated genes should be to the right of the zero margin with positive values ​​on the graph and not on the "zero" line.

[0108] 2.4 Relative expression of the SRD5A gene in HEK293 cell lines We measured SRD5A gene regulation using qPCR in HEK293 cells treated with finasteride (1.34 nM), dutasteride (1.29 μM), and compound Bsy1 (0.02 nM) for 24 hours (Figure 6A). Compared to the control, the test compounds induced significant upregulation of SRD5A2 in the HEK293 cell line (p<0.0001). Similar results were obtained for SRD5A1 and SRD5A3. The relative expression of the SRD5A2 gene increased approximately 2-fold with compounds Bsy1 and dutasteride, and 0.75-fold with finasteride. [Figure 6A - IC of finasteride (1.34 nM), dutasteride (1.29 μM), and Bsy1 (0.02 nM)] 50 Relative expression of the SRD5A gene after treatment of HEK293 cell lines with the values. "ns" - expression levels with no statistical difference between finasteride and compound Bsy1, and dutasteride and compound Bsy1. Experiments were repeated three times.

[0109] Two-way ANOVA was used to compare the expression levels of the SRD5A gene in HEK293 cell lines with those of different drugs (Figure 6B). [Figure 6B - IC in HEK293 cell lines] 50 Two-way ANOVA Dunnett's test for control vs. treatment on values. Comparison test of control vs. drug treatment at p≦0.05: SRD5A1 (ns); SRD5A2 excluding finasteride **** Vs. untreated (ns); SRD5A3 (ns). Data are the mean ± standard error of the mean for each bar. **** P < 0.0001. Significantly upregulated genes do not touch the zero line of the graph but lie to the left of the zero margin.

[0110] The compound Bsy1 and reference drugs (finasteride and dutasteride) were screened for SRD5A2 and SRD5A1 inhibition in DU-145 and HEK293 cell lines. Prior to screening, cells were treated with various concentrations of the drugs to determine IC 50 The IC value was determined. 50 The IC values ​​ranged from 5.4 pM (0.0054 nM) to 1.2 μM. The IC values ​​of Bsy1 among the tested compounds were 50 The IC50 values ​​were lowest for DU-145 (5.4 pM) and HEK293 (0.02 nM), with selectivity indices of 3.7. The reference drugs had higher IC50 values ​​than Bsy1 in cancer cell lines compared to normal cell lines. 50 Based on the in silico studies and the in vitro results obtained with Bsy1, it would be reasonable to expect that phenyl-pyrazolo[3,4-b]pyridine-4-carboxylic acid derivatives of Bsy1, including analogs Bsy2 through Bsy10, would show similar results and therefore be viable as therapeutic agents for androgen-dependent diseases, benign prostatic hyperplasia, and androgenetic alopecia.

[0111] In the experiment, cells were treated with IC for 24 hours. 50 We treated the cells with three different compounds and measured the gene regulation levels of SRD5A2 and SRD5A1. The three compounds suppressed the SRD5A2 and SRD5A1 genes in the DU-145 cancer cell line, but upregulated them in HEK293 normal cells. The relative expression of SRD5A2 was downregulated approximately three-fold by finasteride and dutasteride, and two-fold by Bsy1, relative to SRD5A2 in DU-145 cells, at concentrations 100-fold higher than finasteride and Bsy1. However, in HEK293 cells, it was elevated approximately two-fold by Bsy1 and dutasteride, and 0.75-fold by finasteride.

[0112] 3. In Vitro Drug Kinetic Profiling Solubility (Bsy1-6 and finasteride) and hepatocellular stability (Bsy1 and finasteride) were tested. Drug solubility is an important parameter for achieving the desired concentration of a drug in the systemic circulation to achieve the desired pharmacological response. Drugs with poor water solubility often require high doses to reach therapeutic plasma concentrations after oral administration.

[0113] Hepatocellular stability studies are important for measuring the hepatic metabolism and clearance of drugs absorbed from the gastrointestinal tract. In vitro metabolic stability is a good indicator of the in vivo half-life and clearance of a test compound.

[0114] 3.1 Evaluation of solubility and hepatocyte stability of Bsy1-6 and finasteride Solubility was measured at pH 6.5 using a miniature shake flask method adapted for 96-well plates. Briefly, 4 μl of a 10 mM stock in DMSO was added to a 96-well plate and evaporated using a GeneVac system. Phosphate buffer, pH 6.5, was then added to the wells, and the plate was incubated at 25°C for 24 h with shaking. At the end of this incubation, the samples were centrifuged at 3500 g for 15 min and transferred to an analysis plate. A calibration curve in DMSO for each sample ranging from 10 to 220 μM was generated and included on the analysis plate. Analysis was then performed by LC-MS (Agilent Rapid Resolution HPLC, AB SCIEX 4500 MS), and the solubility of each sample was determined from the corresponding calibration curve.

[0115] Hepatocyte stability was assessed using cryopreserved hepatocytes in suspension using a five-point assay design. Briefly, compounds (1 μM) were incubated in human (Hepatosure 100-donor pool, XenoTech LLC) and mouse (Cryostax mouse CD1, XenoTech LLC) hepatocytes (500,000 cells / ml) at 37°C for 120 minutes at various time points. The reaction was quenched by adding ice-cold acetonitrile containing an internal standard. Samples were then centrifuged and analyzed for parent compound elimination by LC-MS / MS (Agilent Rapid Resolution HPLC, AB SCIEX 4500 MS). Half-life, clearance, and hepatic excretion ratios were determined using standard equations.

[0116] Plasma protein binding was measured by ultracentrifugation. Briefly, pooled human plasma was spiked with test compound (10 μM) from a 10 mM DMSO stock. Aliquots were immediately removed, quenched with ice-cold acetonitrile containing an internal standard (carbamazepine, 0.0236 μg / mL), and placed in a freezer. This represented the full-concentration sample. After preincubation (37°C, 1 h), duplicate aliquots of the spiked plasma were transferred to ultracentrifuge tubes and ultracentrifuged (42,000 rpm, 37°C, Beckman Optima L-80XP) for 4 h. Aliquots were also taken after preincubation, quenched, and stored at 4°C. All samples were analyzed by LC-MS / MS (Agilent Rapid Resolution HPLC, AB SCIEX 4500 MS). Protein binding was then calculated by comparing the analyte:peak area ratio of the ultracentrifuged sample to the analyte:peak area ratio of the total concentration sample. Percent degradation was calculated by comparing the analyte:peak area ratio of the preincubation to the total concentration sample. Plasma binding results were failed if degradation exceeded 15%.

[0117] 3.2 Solubility test results The high solubility of Bsy1-6 (Table 4) under aqueous conditions at pH 7.4 provides confidence that other assay measurements performed under similar conditions will yield reproducible data because the compound is in solution. This solubility should facilitate a more reproducible oral absorption process in in vivo experiments. However, once in vivo evaluation plans are underway, thermodynamic solubility testing, which requires solid samples of the compound, would be recommended.

[0118] [Table 5]

[0119] 3.2 Hepatocyte stability Hepatocyte intrinsic clearance (CL) int,appは CL is an indicator of the rate at which a compound is metabolized in liver cells. A low value like this suggests that the compound is metabolized very slowly, which is ideal. int,app These values ​​can be used to calculate predicted in vivo values, taking into account scaling factors (number of hepatocytes per gram of liver and grams of liver per kg of body weight) and binding (measured plasma protein binding and estimated incubation binding). The predicted clearance values ​​for mice and humans represent the expected total hepatic clearance of these compounds in vivo. Bsy1 and finasteride are classified as low hepatic clearance compounds, as their predicted clearances are less than 30% of the hepatic blood flow in each species (mouse hepatic blood flow = 126 ml / min / kg, human hepatic blood flow = 20.7 ml / min / kg) (Table 5).

[0120] [Table 6]

[0121] Although some exemplary embodiments of the present invention have been described above to illustrate how the invention may be made and practiced, those skilled in the art will recognize that various details may be modified to arrive at further embodiments, many of which remain within the scope of the present invention.

Claims

1. Formula (I): 【Chemistry 1】 [During the ceremony, R １ , R １’ , R ２ , R ２’ , and R ３ are each independently H, halogen, or —OR ５ selected from the group consisting of R ５ is a substituted or unsubstituted C １ -C ２ Alkyl, substituted or unsubstituted, linear or branched C ３ -C ６ alkyl, R ４ is a substituted or unsubstituted C １ -C ２ Alkyl, substituted or unsubstituted, linear or branched C ３ -C ６ Alkyl, and substituted or unsubstituted C ３ -C ６ cycloalkyl] or a pharmaceutically acceptable salt, stereoisomer, hydrate or solvate thereof, and a pharmaceutically acceptable carrier.

2. R １ , R １’ , R ２ and R ２’ is H and R ３ Ga-OR ５ The pharmaceutical composition of claim 1, wherein

3. R １ or R １’ One of the two is -OR ５ and R ２ , R ２’ and R ３ The pharmaceutical composition of claim 1 , wherein is H.

4. R ２ or R ２’ One of the two is -OR ５ and R １ , R １’ and R ３ The pharmaceutical composition of claim 1 , wherein is H.

5. R １ , R １’ , R ２ , R ２’ and R ３ The pharmaceutical composition of claim 1 , wherein at least one of is a halogen.

6. R １ , R １’ , R ２ , R ２’ , and R ３ 2. The pharmaceutical composition of claim 1, wherein one of is halogen and the rest are H.

7. 2. The pharmaceutical composition of claim 1, wherein the halogen is selected from Cl, Br and F.

8. R ４ is unsubstituted C １ -C ２ Alkyl, unsubstituted linear or branched C ３ -C ６ Alkyl and unsubstituted C ３ -C ６ 2. The pharmaceutical composition of claim 1, wherein the aryl group is selected from cycloalkyl.

9. R ４ The pharmaceutical composition of claim 1 , wherein is methyl, ethyl, or cyclopropyl.

10. -OR ５ Ga-OCH ３ The pharmaceutical composition of claim 1, wherein

11. The compound of formula (I) Table 1-1 Table 1-2 Table 1-3 The pharmaceutical composition of claim 1 , wherein the pharmaceutical composition is selected from the group consisting of:

12. A compound of formula (I): for use in a method for treating a disease or disorder in a patient, the method comprising administering to a subject in need thereof a compound or pharmaceutical composition comprising: 【Chemistry 2】 [During the ceremony, R １ , R １’ , R ２ , R ２’ , and R ３ are each independently H, halogen, or —OR ５ selected from the group consisting of R ５ is a substituted or unsubstituted C １ -C ２ Alkyl, substituted or unsubstituted, linear or branched C ３ -C ６ alkyl, R ４ is a substituted or unsubstituted C １ -C ２ Alkyl, substituted or unsubstituted, linear or branched C ３ -C ６ Alkyl, and substituted or unsubstituted C ３ -C ６ cycloalkyl] or a pharmaceutically acceptable salt, stereoisomer, hydrate, or solvate thereof, or a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt, stereoisomer, hydrate, or solvate thereof.

13. R １ , R １’ , R ２ and R ２’ is H and R ３ Ga-OR ５ 13. The compound or pharmaceutical composition for use according to claim 12, wherein:

14. R １ or R １’ One of the two is -OR ５ and R ２ , R ２’ and R ３ 13. The compound or pharmaceutical composition for use according to claim 12, wherein is H.

15. R ２ or R ２’ One of the two is -OR ５ and R １ , R １’ and R ３ 13. The compound or pharmaceutical composition for use according to claim 12, wherein is H.

16. R １ , R １’ , R ２ , R ２’ , and R ３ 13. The compound or pharmaceutical composition for use according to claim 12, wherein at least one of is a halogen.

17. R １ , R １’ , R ２ , R ２’ and R ３ 13. The compound or pharmaceutical composition for use according to claim 12, wherein one of is halogen and the rest are H.

18. 13. The compound or pharmaceutical composition for use according to claim 12, wherein the halogen is selected from Cl, Br and F.

19. R ４ is unsubstituted C １ -C ２ Alkyl, unsubstituted linear or branched C ３ -C ６ Alkyl and unsubstituted C ３ -C ６ 13. The compound or pharmaceutical composition for use according to claim 12, wherein said alkyl is selected from cycloalkyl.

20. R ４ 13. The compound or pharmaceutical composition for use according to claim 12, wherein is methyl, ethyl or cyclopropyl.

21. -OR ５ Ga-OCH ３ 13. The compound or pharmaceutical composition for use according to claim 12, wherein:

22. The compound of formula (I) Table 2-1 Table 2-2 Table 2-3 13. The compound or pharmaceutical composition for use according to claim 12, selected from:

23. 23. The compound or pharmaceutical composition for use according to any one of claims 12 to 22, wherein the disease or disorder is a disease or disorder mediated by high levels of dihydrotestosterone.

24. 24. The compound or pharmaceutical composition for use according to claim 23, wherein the disease or disorder is benign prostatic hyperplasia.

25. 24. The compound or pharmaceutical composition for use according to claim 23, wherein the disease or disorder is androgenetic alopecia.

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