A steroid compound, its use, preparation method and pharmaceutical composition comprising the steroid compound

Abstract

The present application relates to the field of compounds, discloses a kind of steroid compound with general formula I structure and its purposes, preparation method and the pharmaceutical composition comprising the steroid compound, to GABA A Receptor has strong positive regulation, can be used for preparing oral anti-severe depression and postpartum depression drug, with the characteristics of high efficiency.

Description

Technical Field

[0001] This invention relates to the field of compounds, and more particularly to novel steroidal compounds, methods for their preparation, and the use of the steroidal compounds in the preparation of pharmaceuticals, as well as pharmaceutical compositions containing the steroidal compounds and the use of the pharmaceutical compositions in the preparation of pharmaceuticals. Background Technology

[0002] Depression is a mood disorder characterized by low mood as the primary clinical manifestation. It is characterized by three lows: low mood, decreased willpower, and slowed thinking. Symptoms may include pessimism, hopelessness, delusions, loss of appetite, functional decline, severe suicidal attempts, and even suicidal behavior. It poses a serious threat to health and must be taken very seriously, requiring timely treatment.

[0003] Currently, medications used to treat depression can be classified into five main categories based on their mechanism of action: A. Tricyclic antidepressants; B. Monoamine oxidase inhibitors (MAOIs); C. Selective serotonin reuptake inhibitors (SSRIs); D. Selective norepinephrine (NA) reuptake inhibitors; and E. Serotonin and norepinephrine reuptake inhibitors. Traditional antidepressants have not been effective in treating major depressive disorder and postpartum depression, and instead have significant and varying degrees of toxic side effects, such as anorexia, diarrhea, and nausea. They may also cause headaches, fatigue, insomnia, anxiety, tremors, sexual dysfunction (including impotence and decreased libido), and drowsiness.

[0004] On March 19, 2019, the U.S. FDA approved Brexanolone (brand name Zulresso) injection for the treatment of postpartum depression in adult women. This was the first drug specifically approved for postpartum depression (PPD). Brexanolone is a synthetic tetrahydroprogesterone analogue that enhances the expression of recombinant GABA. A The receptor is a GABA-mediated current in mammalian cells. However, brinolone is administered via continuous intravenous infusion for more than 60 hours. During the two and a half days of brinol infusion, medical personnel must be on-site to continuously monitor the patient, which is particularly inconvenient.

[0005] Therefore, given the low oral bioavailability of existing drugs such as brinolone, there is an urgent need to develop next-generation synthetic oral tetrahydroprogesterone analogs for the treatment of major depressive disorder and postpartum depression. Summary of the Invention

[0006] To address the aforementioned problems, this invention provides a steroidal compound having the general formula I structure, its stereoisomers, pharmaceutically acceptable salts, hydrates, solvates, or isotopic compounds.

[0007]

[0008] in,

[0009] R1, R2, R3, R4, R5, R x The groups are selected from hydrogen, fluorine, chlorine, bromine, iodine, carbon, nitrogen, oxygen, sulfur, methyl, monofluoromethyl, difluoromethyl, trifluoromethyl, cyano, acyl, formaldehyde oxime, formaldehyde oxime ether, nitro, and carboxyl, wherein the acyl group is selected from: alkanoyl and its halogenated derivatives, enoyl and its halogenated derivatives, alkynyl and its halogenated derivatives, alkoxycarbonyl and its halogenated derivatives, alkynyloxycarbonyl and its halogenated derivatives, alkynyloxycarbonyl and its halogenated derivatives, aryloxycarbonyl and its halogenated derivatives, heteroaryloxycarbonyl and its halogenated derivatives, alkylamine carbonyl and its halogenated derivatives, alkynamine carbonyl and its halogenated derivatives, alkynamine carbonyl and its halogenated derivatives, arylamine carbonyl and its halogenated derivatives, and heteroarylamine carbonyl and its halogenated derivatives.

[0010] C1-C6 alkanes and their halogenated derivatives, C1-C6 ethers and their halogenated derivatives, C1-C6 alkoxy aliphatic chains and their halogenated derivatives, C1-C6 alkylthiol aliphatic chains and their halogenated derivatives, C1-C6 alkylamine aliphatic chains and their halogenated derivatives, C2-C6 alkenes and their halogenated derivatives, and C2-C6 alkynes and their halogenated derivatives.

[0011] Five-membered ring aromatic hydrocarbon group, six-membered ring aromatic hydrocarbon group, C3-C7 cycloalkane group;

[0012] Five-membered heterocyclic aromatic hydrocarbon groups containing nitrogen, oxygen, or sulfur;

[0013] Six-membered nitrogen heterocyclic aromatic group;

[0014] Heterocyclic alkane groups containing nitrogen, oxygen, or sulfur;

[0015] Hydroxyl, amino, and thiol groups;

[0016] The linkages include hydroxyl, amino, and mercapto groups of five-membered aromatic hydrocarbons, six-membered aromatic hydrocarbons, C3-C7 cycloalkanes, five-membered heterocyclic aromatic hydrocarbons containing nitrogen, oxygen, or sulfur, six-membered nitrogen heterocyclic aromatic hydrocarbons, and heterocyclic alkanes containing nitrogen, oxygen, or sulfur.

[0017] Protecting groups such as esters and amides formed by inorganic acids or C1-C6 organic acids with hydroxyl and amino groups;

[0018] M is selected from carbon, nitrogen, oxygen, sulfur, and phosphorus;

[0019] When M is nitrogen, the absence of either R2 or R3 can lead to the formation of nitrogen oxides; or the presence of both R2 and R3 can lead to the formation of quaternary ammonium salts.

[0020] When M is oxygen, R2 or R3 is not present;

[0021] When M is sulfur, both R2 and R3 are present, forming a sulfone;

[0022] Alternatively, if either R2 or R3 is absent, a sulfoxide is formed.

[0023] Alternatively, neither R2 nor R3 exists.

[0024] According to one embodiment of the present invention, steroidal compounds having the general formula I structure and their stereoisomers, pharmaceutically acceptable salts, hydrates, solvates or isotopic compounds, R x Oriented inwards. A plane refers to a plane or approximate plane containing heterocyclic rings M and N.

[0025] According to one embodiment of the present invention, steroidal compounds having the general formula I structure and their stereoisomers, pharmaceutically acceptable salts, hydrates, solvates or isotopic compounds, R x Facing outwards.

[0026] According to one embodiment of the present invention, a steroidal compound having the general formula I structure and its stereoisomers, pharmaceutically acceptable salts, hydrates, solvates, or isotopic compounds, wherein R2 and R3 are linked together to form a C8-C12 spirocycle, a C8-C12 azaspirocycle, or a C8-C12 oxaspirocycle, or R2 and R3 linked together. x The C8-C12 bridged ring, C8-C12 nitrogen-bridged ring, and C8-C12 oxygen-bridged ring are formed by connecting them together.

[0027] According to one embodiment of the present invention, a steroidal compound having the general formula I structure and its stereoisomers, pharmaceutically acceptable salts, hydrates, solvates or isotopic compounds, wherein R1, R2, R3, R4 and R5 are not hydrogen.

[0028] According to one embodiment of the present invention, a steroidal compound having the general formula I structure and its stereoisomers, pharmaceutically acceptable salts, hydrates, solvates or isotopic compounds, wherein R1, R2, R3, R4, R5 has only one hydrogen atom; or R1, R2, R3, R4, R5 has only two hydrogen atom; or R1, R2, R3, R4, R5 has only three hydrogen atom; or R1, R2, R3, R4, R5 has only four hydrogen atom; or R1, R2, R3, R4, R5 are all hydrogen atom.

[0029] According to one embodiment of the present invention, a steroid compound having the general formula I structure is synthesized via the following route:

[0030] Compound B1 was subjected to catalytic hydrogenation to obtain compound B2;

[0031] Methylation of compound B2 yields compound B3;

[0032] Compound B3 was subjected to a Ylide reaction to yield compound B4;

[0033] Compound B4 was subjected to a hydroboration oxidation reaction to obtain compound B5;

[0034] Compound B5 was oxidized to obtain compound B6;

[0035] Compound B6 was subjected to a haloform reaction to yield compound B;

[0036] Compound B was condensed with fragment Q to yield a steroidal compound with general formula I.

[0037] Among them, compound B1 is 19-nor-4-androstenedione, compound B is (3R,5R,8R,9R,10S,13S,14S,17S)-3-hydroxy-3,13-dimethylestradiol-17-acid, and compound Q is a para-M-substituted nitrogen-containing six-membered heterocycle.

[0038]

[0039] As used herein, the terms "heteroaryl" and "heterocyclic aromatic hydrocarbon group" refer to an aromatic cyclic group comprising a monocyclic or multicyclic fused (fused) ring (e.g., containing two or three rings) with 5 to 14 ring atoms, wherein, in addition to a carbon atom, the ring atoms also contain at least one heteroatom selected from oxygen, nitrogen, and / or sulfur. If the ring is aromatic, the sulfur and nitrogen atoms may also be present in oxidized forms. Multicyclic fused (fused) heteroaryl groups are formed by fusing a monocyclic heteroaryl group as defined above with one or more rings selected from the following to form a multicyclic fused ring system: heteroaryl (to form, for example, naphthidyl, such as 1,8-naphthidyl), heterocycle (e.g., to form 1,2,3,4-tetrahydronaphthidyl, such as 1,2,3,4-tetrahydro-1,8-naphthidyl), carbocyclic (to form, for example, 5,6,7,8-tetrahydroquinolinyl), and aryl (to form, for example, indazole). Such multicyclic fused ring systems may optionally have one or more (e.g., 1, 2, 3, or 4) oxo groups substituted on the carbocyclic or heterocyclic portion of the fused ring. When valence requirements permit, the rings of a multicyclic fused ring system can be interconnected by fusion, spirocyclic, and bridging bonds. It should be understood that the individual rings of a multicyclic fused ring system can be connected relative to each other in any order. It should also be understood that the connection points of a multicyclic fused ring system can be at any location within the system, including the heteroaryl, heterocyclic, aryl, or carbocyclic portion of a multicyclic fused system. It should also be understood that the connection points of heteroaryl groups can be on any suitable atom of the heteroaryl group, including carbon atoms and heteroatoms (e.g., nitrogen). Exemplary heteroaryl groups include, but are not limited to: pyridyl, pyrroloyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrazolyl, thiophenyl, indolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, furanyl, oxadiazolyl, thiazolyl, quinolinyl, isoquinolinyl, benzothiazolyl, benzooxazolyl, inzolyl, quinoxolinyl, quinazolinyl, 5,6,7,8-tetrahydroisoquinolinyl, benzofuranyl, benzoimidazolyl, thiaindyl, pyrrolo[2,3-b]pyridyl, quinazolinyl-4(3H)-one, triazolyl, 4,5,6,7-tetrahydro-1H-inzolyl and 3b,4,4a,5-tetrahydro-1H-cyclopropane[3,4]cyclopentane[1,2-c]pyrazolyl.

[0040] As used herein, the terms "heterocyclic group" and "heterocyclic alkane group" refer to a monocyclic saturated or partially unsaturated group having a 3- to 8-membered monocyclic ring or multiple fused (fused) rings or bridged rings having 3 to 14 ring atoms within the ring, wherein, in addition to carbon atoms, the ring atoms also contain at least one heteroatom selected from oxygen, nitrogen, and / or sulfur. Examples of heterocyclic groups include, but are not limited to, tetrahydrofuranyl, morpholinyl, piperidinyl, piperazineyl, dihydropyridinyl, 4,5,6,7-tetrahydro-1H-benzo[d]imidazolyl, benzo[d]imidazolyl, 4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridinyl, azircyclic propane, azircyclic butane, tetrahydropyrroleyl, azircyclic heptane, azircyclic octane, etc. Oxycyclopropane, oxycyclobutane, tetrahydrofuranyl, tetrahydropyranyl, oxycycloheptane, oxycyclooctane, thiocyclopropane, thiocyclobutane, tetrahydrothiophene, tetrahydrothioranyl, thiocycloheptane, thiocyclooctane, tetrahydroimidazolyl, tetrahydropyrazolyl, tetrahydrooxazolyl, tetrahydroisooxazolyl, tetrahydrothiazolyl, tetrahydroisothiazolyl, dioxane, thiazolyl, dithiazolyl, etc.

[0041] As used herein, the term "stereoisomer" refers to compounds that have the same chemical composition and connectivity, but whose atoms have different spatial orientations that cannot be interchanged by single bond rotation. "Stereoisomer" includes both "diastereomers" and "enantiomers." A "diastereomer" is a stereoisomer with two or more chiral centers whose molecules are not mirror images of each other. Diastereomers have different physical properties, such as melting point, boiling point, spectral characteristics, and reactivity. Mixtures of diastereomers can be separated using high-resolution analytical procedures such as crystallization, electrophoresis, and chromatography. An "enantiomer" is a stereoisomer of a compound that is a non-overlapping mirror image of another compound.

[0042] As used in this article, the term "tautomer" refers to the coexistence of two (or more) compounds that differ only in the position and electron distribution of one (or more) active atoms, such as keto-enol tautomers.

[0043] As used herein, the term "pharmaceutically acceptable" means that the substance or composition must be chemically and / or toxicologically compatible with other components of the formulation and / or mammals treated with it.

[0044] Any general formula or structure given herein, including general formula I or any general formula disclosed herein, is also intended to represent both unlabeled and isotopically labeled forms of the compound. These forms of the compound may also be referred to as “isotopically labeled” or “isotopically enriched analogs.” Isotopically labeled compounds have the structures depicted herein, except that one or more atoms are replaced by atoms having a selected atomic mass or mass number. Examples of isotopes that can be incorporated into the compounds of the present invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine, chlorine, and iodine, for example, but not limited to… 2 H (deuterium, D) 3 H (tritium) 11 C 13 C 14 C 13 N、 15 N、 15 O、 17 O、 18 O、 31 P, 32 P, 35 S, 18 F, 36 Cl、 123 I and 125 I. These compounds can be synthesized using methods well-known in the field, such as by using one or more starting materials in which hydrogen has been replaced by deuterium.

[0045] As used herein, the terms "solvent" and "solvent complex" refer to an association or complex of one or more solvent molecules with the compounds of this invention. Examples of solvents that form solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine. The term "hydrate" refers to a complex in which the solvent molecules are water.

[0046] As used herein, the term "pharmaceutically acceptable salt" means that, to the extent of reliable medical judgment, it is suitable for contact with the tissues of humans and lower animals without excessive toxicity, irritation, allergic reactions, etc., and in proportion to a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences (1977) 66:1 19. Pharmaceutically acceptable salts of the compounds of this invention include salts derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable non-toxic acid addition salts are salts formed by amino groups with inorganic acids, such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid; or salts formed with organic acids, such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid; or salts formed using methods used in the art, such as ion exchange methods. Other pharmaceutically acceptable salts include: adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentylpropionate, diglucuronate, dodecyl sulfate, ethanesulfonate, formate, fumarate, gluconate, glyceryl phosphate, glucuronate, hemisulfate, heptahydrate, hexanoate, hydroiodate, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, dihydroxynaphthalate, pectin ester, persulfate, 3-phenylpropionate, phosphate, picrate, pentanoate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, etc. Pharmaceutically usable salts derived from suitable bases include alkali metals, alkaline earth metals, ammonium, and nitrogen. + (C1-C4 alkyl)4 salts. Representative alkali metal or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, etc. Further pharmaceutically acceptable salts, where appropriate, include non-toxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxyl, sulfate, phosphate, nitrate, lower alkyl sulfonate, and aryl sulfonate.

[0047] According to another aspect of the present invention, a pharmaceutical composition is provided comprising a steroidal compound having the general formula I structure and its stereoisomers, a pharmaceutically acceptable salt, hydrate, solvate or isotopic compound as an active ingredient.

[0048] According to one embodiment of the present invention, the pharmaceutical composition further includes a pharmaceutical carrier or diluent.

[0049] According to another aspect of the present invention, the aforementioned pharmaceutical composition is used as a medicament for the preparation of a treatment for central nervous system-related disorders, including: postpartum depression, clinical depression, atypical depression, severe depressive disorder, catatonic depression, dysphoric mood, dual depression, depressive personality disorder, recurrent transient depression, mild depressive disorder, bipolar disorder, manic-depressive disorder, mood disorders, anxiety, post-traumatic stress disorder (PTSD), premenstrual anxiety disorder (PMDD), depression caused by chronic medical conditions, and other related conditions. Treatment for depression, suicidal ideation or behavior, premenstrual syndrome, generalized anxiety disorder, seasonal affective disorder (SAD), social anxiety disorder, memory loss, poor stress tolerance, Niemann-Pick II type C disease or related neurological or physical symptoms, epilepsy, essential tremor, epileptiform disorder, NMDA insufficiency, migraine, status epilepticus, sleep disorders, fragile X syndrome, 5α-reductase inhibitor-induced depression, PCDH19 female childhood epilepsy, sexual dysfunction, cognitive impairment, Parkinson's disease or Alzheimer's disease.

[0050] According to another aspect of the present invention, the aforementioned steroidal compounds having the general formula I structure and their stereoisomers, pharmaceutically acceptable salts, hydrates, solvates, or isotopic compounds are used as medicaments for the preparation of treatments for central nervous system-related disorders, including: postpartum depression, clinical depression, atypical depression, severe depressive disorder, catatonic depression, dysphoric mood, dual depression, depressive personality disorder, recurrent transient depression, mild depressive disorder, bipolar disorder, manic-depressive disorder, mood disorders, anxiety, post-traumatic stress disorder (PTSD), and premenstrual anxiety disorder. (PMDD), depression due to chronic medical conditions, intractable depression, suicidal ideation or behavior, premenstrual syndrome, generalized anxiety disorder, seasonal affective disorder (SAD), social anxiety disorder, memory loss, poor stress tolerance, Niemann-Pick II type C disease or related neurological or physical symptoms, epilepsy, essential tremor, epileptiform disorder, NMDA insufficiency, migraine, status epilepticus, sleep disorders, fragile X syndrome, depression induced by 5α-reductase inhibitors, epilepsy in female children with PCDH19, sexual dysfunction, cognitive impairment, Parkinson's disease or Alzheimer's disease.

[0051] Pharmaceutical composition and administration

[0052] Compounds provided by the present invention, or pharmaceutically acceptable salts, stereoisomers, tautomers, prodrugs, solvates, and pharmaceutically acceptable salts thereof, are typically administered in the form of pharmaceutical compositions. The present invention provides pharmaceutical compositions comprising, as an active ingredient, a compound provided by the present invention, or pharmaceutically acceptable salts, stereoisomers, tautomers, prodrugs, solvates, and pharmaceutically acceptable salts thereof, and one or more pharmaceutically acceptable carriers. The pharmaceutical compositions may be administered alone or in combination with other therapeutic agents. Such compositions are prepared in a manner well known in the art, for example, Reminton's Pharmaceutical Sciences, Mace Publishing Co., Philadelphia, PA 17th Ed. (1985); and Modern Pharmaceutics, Marcel Dekker, Inc. 3rd Ed. (GS Banker & C.T. Rhodes, Eds.).

[0053] Pharmaceutically acceptable carriers can be solid or liquid. Solid carriers can be one or more substances used as excipients, diluents, sweeteners, solubilizers, lubricants, binders, tablet disintegrants, stabilizers, preservatives, or encapsulating materials. Liquid carriers can be solvents or liquid dispersion media. Suitable solid carriers include, but are not limited to, cellulose, glucose, lactose, mannitol, magnesium stearate, magnesium carbonate, sodium carbonate, sodium saccharin, sucrose, dextrin, talc, starch, pectin, gelatin, astragalus gum, gum arabic, sodium alginate, parabens, methylcellulose, sodium carboxymethyl cellulose, low-melting-point waxes, cocoa butter, etc. Suitable liquid carriers include, but are not limited to, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.), vegetable oils (e.g., peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil), glycerides, agar, pyrogen-free water, isotonic saline, Ringer's solution, and mixtures thereof.

[0054] The pharmaceutical compositions according to the invention can be in any form suitable for the intended method of administration. For example, when for oral use, they can be prepared as tablets, lozenges, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, syrups or elixirs, solutions, or sprays. Compositions for oral use can be prepared according to any method known in the art for manufacturing pharmaceutical compositions. The pharmaceutical compositions of the invention can also be prepared as formulations suitable for intrapulmonary or intranasal administration, such as aerosols or dry powder formulations, nasal drops, or nasal sprays. The pharmaceutical compositions of the invention can also be prepared as suppositories suitable for rectal administration. The pharmaceutical compositions of the invention can also be prepared as transdermal formulations for topical administration or eye drops suitable for ocular administration.

[0055] The pharmaceutical compositions of the present invention can be administered via parenteral, intramuscular, subcutaneous, oral, or local single or multiple doses.

[0056] The effective dose of the compounds of the present invention depends at least on the nature of the condition being treated, its toxicity, the method of delivery, and the pharmaceutical formulation, and will be determined by clinicians using conventional dose-escalation studies. A dose of about 0.0001 to about 100 mg per kilogram of body weight per day is anticipated; typically about 0.01 to about 10 mg per kilogram of body weight per day; more typically about 0.01 to about 5 mg per kilogram of body weight per day; and most typically about 0.05 to about 0.5 mg per kilogram of body weight per day. For example, a candidate daily dose for an adult weighing about 70 kg would be in the range of 1 mg to 1000 mg, preferably in the range of 5 mg to 500 mg, and could be in the form of a single dose or multiple doses.

[0057] In this invention, steroidal compounds, stereoisomers, pharmaceutically acceptable salts, hydrates, solvates, or isotopic compounds of general formula I can simultaneously act on multiple GABAs on the synaptic surface through allosteric regulation. A Ion channel receptors facilitate the influx of chloride ions into cells along the electrochemical gradient of the GABA receptor complex (GRC). Increased intracellular levels of this anion lead to hyperpolarization of the transmembrane potential, making neurons less sensitive to excitatory input, i.e., reducing brain excitability. The compounds of this invention not only have a highly efficient action but also solve the inconvenience caused to patients by prolonged intravenous infusion. Attached Figure Description

[0058] Figure 1 The remaining percentage trend for each compound is shown. Detailed Implementation

[0059] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0060] It should be understood that the terms "comprising" and "including" as used in the specification and claims of this invention indicate the presence of the described features, integrals, steps, operations, elements and / or components, but do not exclude the presence or addition of one or more other features, integrals, steps, operations, elements, components and / or collections thereof.

[0061] It should also be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the invention. As used in this specification and claims, the singular forms “a,” “an,” and “the” are intended to include the plural forms unless the context clearly indicates otherwise. It should also be understood that the term “and / or” as used in this specification and claims refers to any combination and all possible combinations of one or more of the associated listed items, and includes such combinations.

[0062] Table 1 shows the preferred compounds of the present invention.

[0063]

[0064]

[0065]

[0066]

[0067]

[0068]

[0069] Before synthesizing the above compounds, intermediates need to be prepared first.

[0070] Preparation and synthesis of intermediate B: Compound B1 is 19-nor-4-androstenedione, which is currently available in the market.

[0071]

[0072] in,

[0073] The first step is the synthesis of compound B2.

[0074] In a three-necked flask, compound B1 was added and dissolved in tetrahydrofuran. Then, 10% palladium on carbon and hydrobromic acid (48% content) were added. Nitrogen was used for purging, followed by hydrogen purging. The mixture was stirred overnight at room temperature under hydrogen atmosphere. The reaction was monitored by TLC until it was complete. The mixture was filtered through diatomaceous earth, concentrated under reduced pressure to remove the solvent, crystallized in acetone, and filtered to obtain B2.

[0075] The second step involves the synthesis of compound B3.

[0076] In a three-necked flask, 2,6-di-tert-butyl-p-cresol was added and dissolved in tetrahydrofuran. The mixture was cooled to -78°C, and then 2M trimethylaluminum toluene solution was added. After the addition was complete, the reaction mixture was brought to room temperature and reacted for 1.5 hours under nitrogen protection. The reaction mixture was then cooled to -78°C, and a toluene solution of compound B2 was added. The mixture was kept at this temperature for 1.5 hours, and then 3M methylmagnesium bromide tetrahydrofuran solution was added dropwise while maintaining the temperature below -75°C. After the addition was complete, the mixture was kept at this temperature for 1.5 hours, brought to room temperature, and stirred overnight. Under TLC monitoring, saturated ammonium chloride solution was added dropwise while maintaining the temperature below 5°C. Bubbles were generated in the reaction mixture, and a large amount of white solid precipitated. The mixture was filtered, and the mother liquor was collected, washed with water, dried, concentrated under reduced pressure, and purified by column chromatography (petroleum ether: ethyl acetate = 1:1, v / v) to obtain the oily product B3.

[0077] The third step involves the synthesis of compound B4.

[0078] Ethyltriphenylphosphine bromide was added to a three-necked flask and dissolved in tetrahydrofuran. The mixture was cooled to 0°C, then potassium tert-butoxide dissolved in tetrahydrofuran was added to the reaction solution. After completion, the temperature was raised to 60°C, and a tetrahydrofuran solution of B3 was added. The reaction was carried out at 60°C overnight under TLC monitoring. The mixture was then cooled to room temperature, and a saturated ammonium chloride solution was added. The mixture was concentrated under reduced pressure, and ethyl acetate was added. The layers were separated, and the organic phase was separated and washed with water. The organic phase was distilled under reduced pressure, and the residue was separated by column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to obtain product B4.

[0079] Step 4: Synthesis of compound B5

[0080] Compound B4 was added to a three-necked flask and dissolved in tetrahydrofuran. The mixture was cooled to 0°C and, under nitrogen protection, a 10M solution of a borane dimethyl sulfide complex was added, maintaining the temperature below 10°C. After the addition was complete, the mixture was allowed to react at room temperature for 3 hours, resulting in a white precipitate. TLC monitoring was performed, and an aqueous sodium hydroxide solution was added. Once the reaction solution became translucent, a 30% hydrogen peroxide solution was added, again producing a white precipitate. The temperature was maintained below 10°C throughout the reaction. After the reaction was complete, a sodium thiosulfate solution was added to quench the reaction. Ethyl acetate was then added, the mixture was stirred, and the layers were separated. The organic phase was washed with water and concentrated under reduced pressure to obtain product B5.

[0081] Step 5: Synthesis of compound B6

[0082] In a 1L three-necked flask, compound B5 was added and dissolved in dichloromethane. Pyridinium chlorochromate was weighed and added to the reaction solution. The mixture was stirred at room temperature and reacted overnight. TLC was monitored, and sodium sulfite solution was added to quench the reaction. The mixture was separated, retaining the organic phase. The aqueous phase was extracted with dichloromethane, washed with water, and purified by vacuum distillation and column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to obtain product B6.

[0083] Synthesis of intermediate B

[0084] In a three-necked flask, sodium hydroxide solution was added, and the temperature was lowered to 0°C. Liquid bromine was added dropwise until completely dissolved. Dioxane was then added, and the mixture was stirred at room temperature for 2 hours. Compound B6 was dissolved in dioxane and water, and added to the reaction solution. The mixture was stirred at room temperature overnight. TLC monitoring was performed, and sodium sulfite solution was added to quench the reaction. The temperature was raised to 80°C, and the reaction was carried out for 3 hours. The pH was adjusted to 2-3 with 1M dilute sulfuric acid aqueous solution at room temperature. A white solid precipitated out. The product B was obtained by filtration and drying.

[0085] Example 1

[0086] Synthesis of Compound 1

[0087]

[0088] Compound B was added to a reaction flask and dissolved in DMF. Then, 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU) was added, and the mixture was stirred at room temperature for 20 min. Methylpiperazine was then added, and the reaction was allowed to proceed overnight at room temperature. After the reaction was complete, water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to give an off-white solid 1.

[0089] 1 H NMR (400MHz, chloroform-d) δ3.77-3.68(m,4H),2.71-2.62(m,5H),2.48(s,3H),2.38-2.28(m,2H),1.85-1.7 6(m,4H),1.71-1.61(m,3H),1.49-1.40(m,5H),1.34-1.23(m,10H),1.13-1.04(m,3H),0.71(s,3H).

[0090] MS: m / z[M+H] + 403.35.

[0091] Example 2

[0092] Synthesis of Compound 4

[0093]

[0094] Compound B and DMF were dissolved in the reaction flask, followed by the addition of 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The mixture was stirred at room temperature for 20 min, then morpholine was added, and the reaction was allowed to proceed overnight at room temperature. After the reaction was complete, water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to give a white solid 4.

[0095] 1 H NMR (400MHz, chloroform-d) δ3.63-3.58(m,8H), 2.69-2.65(t,J=8.0Hz,1H), 2.33(q,J=2 0.0,12.0Hz,1H),1.85-1.77(m,4H),1.70-1.62(m,4H),1.49-1.40(m,5H),1.35 -1.26(m,10H),1.13 -1.07(m,3H),0.73(s,3H).

[0096] MS: m / z[M+H] + 390.5.

[0097] Example 3

[0098] Synthesis of Compound 5

[0099]

[0100] Compound B and DMF were dissolved in the reaction flask, followed by the addition of 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The mixture was stirred at room temperature for 20 min, then (S)-2-methylmorpholine and N,N-diisopropylethylamine (DIEA) were added, and the mixture was reacted overnight at room temperature. After the reaction was completed, water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to give a white solid 5.

[0101] 1 ¹H NMR (400MHz, chloroform-d) δ 3.89 (s, 1H), 3.50–3.45 (m, 4H), 2.98 (s, 3H), 2.68 (m, 1H), 2.35 (q, J = 12.0, 9.0 Hz, 1H), 1.86–1.68 (m, 8H), 1.49–1.45 (m, 5H), 1.42–1.39 (m, 10H), 1.27–1.25 (m, 3H), 1.19–1.05 (m, 2H), 0.73 (s, 3H).

[0102] MS: m / z[M+H] + 404.30.

[0103] Example 4

[0104] Synthesis of Compound 6

[0105]

[0106] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then (R)-2-methylmorpholine was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to give an off-white solid 6.

[0107] 1 H NMR (400 MHz, chloroform-d) δ4.14-4.09(m,1H),3.92-3.80(m,4H),3.53-3.42(m,2H),3.06(s,3H),2.68(br,1H),2.35(q,J=20.0,12.0 Hz,1H),1.86-1.61(m,8H),1.49(s,1H),1.45-1.41(m,4H),1.39-1.31(m,7H),1.27-1.25(m,3H),1.20-1.05(m,2H),0.68(s,3H).

[0108] MS: m / z[M+H] + 404.2.

[0109] Example 5

[0110] Synthesis of Compound 10

[0111]

[0112] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then compound 10a was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to give an off-white solid 10.

[0113] 1 ¹H NMR (400 MHz, chloroform-d) δ 3.97 (d, 1H), 3.56–3.43 (m, 4H), 3.39 (s, 3H), 3.00–2.92 (m, 1H), 2.71–2.67 (m, 1H), 2.54–2.35 (m, 2H), 2.32–2.27 (m, 1H), 1.86–1.68 (m, 8H), 1.45–1.27 (m, 18H), 0.72 (s, 3H).

[0114] MS: m / z[M+H] + 434.4

[0115] Example 6

[0116] Synthesis of Compound 11

[0117]

[0118] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then compound 11a was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 1:1, v / v) to give an off-white solid 11.

[0119] 1 ¹H NMR (400MHz, chloroform-d) δ 3.98 (d, 1H), 3.57–3.43 (m, 4H), 3.39 (s, 3H), 3.00–2.92 (m, 1H), 2.71–2.67 (m, 1H), 2.54–2.35 (m, 2H), 2.32–2.27 (m, 1H), 1.86–1.68 (m, 8H), 1.46–1.27 (m, 18H), 0.71 (s, 3H).

[0120] MS: m / z[M+H] + 434.4

[0121] Example 7

[0122] Synthesis of Compound 22

[0123]

[0124] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then compound 22a was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 4:1, v / v) to give an off-white solid 22.

[0125] 1H NMR (400MHz, chloroform-d) δ4.21(s,1H),2.90-2.75(m,1H),2.55-2.45(m,1H),2.40-2.23(m,1H),1.89-1.75(m,6H),1.73 -1.63(m,6H),1.47-1.37(m,6H),1.35-1.31(m,2H),1.29-1.25(m,9H),1.15-1.05(m,4H),0.93(d,3H)0.76(s,3H).

[0126] MS: m / z[M+H] + 402.4

[0127] Example 8

[0128] Synthesis of Compound 29

[0129]

[0130] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then compound 29a was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 2:1, v / v) to give an off-white solid 29.

[0131] 1 H NMR (400MHz, DMSO-d6) δ4.21-4.31(m,1H), δ4.18-4.07(m,1H), δ3.96-3.75 (m,2H),δ3.72-3.63(m,3H),δ3.55-3.40(m,2H),δ3.35-3.20(m,2H),2.90-2 .80(m,1H),2.16-2.07(m,1H),1.74-1.68(m,3H),1.65-1.57(m,4H),1.42-1 .34(m,4H),1.30-1.22(m,8H),1.12(s,4H),1.05-0.98(m,2H),0.67(s,3H).

[0132] MS: m / z[M+H] + 448.4

[0133] Example 9

[0134] Synthesis of Compound 41

[0135]

[0136] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then compound 41a was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 4:1, v / v) to give an off-white solid 41.

[0137] 1 H NMR (400MHz, DMSO-d6) δ5.15-4.40(m,4H), δ4.09(t,1H), δ3.95-3.65(m,2H), δ3.44-3.33(m,1H),2.81-2.69(m,1 H), 2.38-2.26(m,4H),2.22-2.10(m,4H),1.98-1.76(m,10H),1.74-1.65(m,5H),1.63-1.50(m,3H),1.16(s,3H).

[0138] MS: m / z[M+H] + 458.2

[0139] Example 10

[0140] Synthesis of Compound 44

[0141]

[0142] Synthesis of intermediate 44a

[0143]

[0144] Material A was weighed into a reaction flask, dissolved in dichloromethane, and then triethylamine and di-tert-butyl dicarbonate were added. The mixture was stirred at room temperature for 12 hours. TLC analysis showed that the reactants had reacted completely, and the reaction was terminated. Water was added to the reaction system, and the mixture was extracted three times with dichloromethane. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain crude product 44a.

[0145] Synthesis of intermediate 44b

[0146]

[0147] Crude product 44a was weighed into a reaction flask, dissolved in dichloromethane, and the flask was purged with nitrogen. The mixture was then stirred in an ice bath. Dess-Martin reagent was added, and the mixture was slowly heated to room temperature and stirred for 4 hours. TLC analysis showed that the reactants had reacted completely, and the reaction was terminated. Saturated sodium bicarbonate solution was added to the reaction system and stirred to quench the reaction. The mixture was extracted three times with dichloromethane, and the organic phase was backwashed with purified water. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain crude product 44b.

[0148] Synthesis of intermediate 44c

[0149]

[0150] Crude product 44b was weighed into a reaction flask, dissolved in dichloromethane, and the flask was purged with nitrogen. The mixture was then stirred in an ice bath. Diethylaminosulfur trifluoride (DAST) was added, and the mixture was slowly heated to room temperature and stirred for 12 hours. TLC analysis showed that the reactants had reacted completely, and the reaction was terminated. Purified water was added to the reaction system, and the mixture was extracted three times with dichloromethane. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain crude product 44c.

[0151] Synthesis of intermediate 44d

[0152]

[0153] Crude product 44c was weighed into a reaction flask, and 4M HCl 1,4-dioxane solution was added. The mixture was stirred at room temperature for 2 hours. TLC analysis showed that the reaction proceeded completely, and the reaction was terminated. The reaction mixture was then concentrated under reduced pressure to obtain crude product 44d.

[0154] Synthesis of Compound 44

[0155]

[0156] Compound B and DMF were added to a reaction flask, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The reaction was carried out at room temperature for 20 min, then compound 44d was added, and the reaction was carried out at room temperature overnight. Water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 5:1, v / v) to give an off-white solid 44.

[0157] 1 H NMR (400MHz, chloroform-d) δ5.90-5.59(m,1H), δ4.95-4.38(m,1H), δ4.13-3.87(m,2H), δ3.70-3.48(m,2H), δ3.30-3.05(m,1H), 2.85-2.65(m,1H), 2.38-2.20(m,1H),1.87-1.77(m,4H),1.75-1.63(m,4H),1.57(m,5H), 1.47-1.39(m,4H),1.31-1.24(m,7H),1.14-0.86(m,3H),0.73(s,3H).

[0158] MS: m / z[M+H] + 458.2

[0159] Example 11

[0160] Synthesis of Compound 49

[0161]

[0162] Compound B and DMF were dissolved in a reaction flask, followed by the addition of 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The mixture was stirred at room temperature for 20 min, then (S)-2-ethylmorpholine and N,N-diisopropylethylamine (DIEA) were added, and the mixture was allowed to react overnight at room temperature. After the reaction was complete, water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 3:1, v / v) to give 49 g of an off-white solid.

[0163] 1 ¹H NMR (400MHz, chloroform-d) δ 3.89 (s, 1H), 3.50–3.45 (m, 4H), 2.98–2.78 (m, 2H), 2.68 (m, 1H), 2.35 (q, J = 12.0, 9.0 Hz, 1H), 1.86–1.68 (m, 8H), 1.49–1.45 (m, 5H), 1.42–1.39 (m, 13H), 1.27–1.25 (m, 3H), 1.19–1.05 (m, 2H), 0.73 (s, 3H).

[0164] MS: m / z[M+H] + 428.3

[0165] Example 12

[0166] Synthesis of compounds 52 and 53

[0167]

[0168] Compound B and DMF were dissolved in the reaction flask, followed by the addition of 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU). The mixture was stirred at room temperature for 20 min, then 2-cyclopropylmorpholine and N,N-diisopropylethylamine (DIEA) were added, and the mixture was allowed to react overnight at room temperature. After the reaction was complete, water and ethyl acetate were added, and the mixture was stirred to separate the layers. The organic phase was washed with water, distilled under reduced pressure, and subjected to column chromatography (petroleum ether:ethyl acetate = 2:1, v / v) to obtain an off-white solid 52a. This was then separated into white solids 52 and 53 by chiral column chromatography (mobile phase: acetonitrile: 5% trifluoroacetic acid aqueous solution = 60:40, isocratic elution, flow rate 1 ml / min, wavelength 200 nm).

[0169] Structural characterization of compound 52:

[0170] 1 H NMR (400MHz, DMSO-d6) δ4.39-4.23(m,1H),4.01(d,J=13.2Hz,1H),3.82(q,J= 13.9,11.6Hz,2H),3.35-3.15(m,2H),3.10-2.84(m,2H),2.76-2.54(m,2H),2. 49-2.43(m,1H),2.22-2.05(m,1H),1.76-1.53(m,8H),1.44-0.94(m,18H),0. 84(dd,J=13.0,5.7Hz,2H),0.59(s,3H),0.49-0.40(m,2H),0.36-0.17(m,2H).

[0171] MS: m / z[M+H] + 430.3

[0172] Structural characterization of compound 53:

[0173] 1 H NMR (400MHz, DMSO-d6) δ4.36(d,J=10.4Hz,1H),4.18(d,J=13.4Hz,1H),3.94(d,J=12.5Hz,1H),3 .88-3.72(m,1H),3.35-3.07(m,2H),2.98-2.77(m,2H),2.62(td,J=14.7,12.8,8.8Hz,2H),2.20 -2.09(m,1H),1.81-1.53(m,8H),1.47-0.94(m,19H),0.85(q,J=7.4,6.9Hz,2H),0.59(s,3H),0.54-0.42(m,2H),0.40-0.08(m,2H).

[0174] MS: m / z[M+H] + 430.3

[0175] Although the synthesis of compounds 1, 4, 5, 6, 10, 11, 22, 29, 41, 44, 49, 52, and 53 is used as an example in this invention, those skilled in the art will understand that all compounds listed in the general formula of this invention can be prepared by referring to the above synthetic route.

[0176] Bioevaluation

[0177] Electrophysiological experiments:

[0178] Sulfanolone (SAGE-217) is a novel, synthetic, approved neuroactive steroid GABA.A This receptor-positive allosteric modulator exhibits favorable oral pharmacokinetic properties, supporting daily oral doses. This invention employs the same testing method, using SAGE-217 as a control to test the activity of the compound.

[0179] Assay Methods: Using a CHO cell line transiently expressing the GABA(α4β3δ) receptor, the effects of compounds on the GABA(α4β3δ) receptor were investigated, and the half-maximal effective concentration (Cmax) of the compounds was tested. 50 (Value).

[0180] The experiment used a manual patch-clamp system, a HEKA EPC 10 signal amplifier, and a digital conversion system to record the chloride ion current of the GABA channel in the whole cell. All experiments were conducted at normal room temperature.

[0181] After whole-cell sealing was achieved, the cell membrane voltage was clamped at -70mV. In Gap-free mode, the peak current was recorded after sequentially spraying the cell surface with 5nM GABA, a mixture of the present invention's compound and 5nM GABA at low to high concentrations, and 10μM GABA.

[0182] Administration of the test substance: The mixture of GABA5nM and each concentration of the compound of the present invention was administered 1-2 times, followed by rinsing with extracellular fluid for 1 min before measuring the next concentration. Experimental data were acquired using an EPC 10 amplifier (HEKA) and stored in PatchMaster (HEKA) software.

[0183] Current recording: During drug administration, a coverslip containing cells was placed in a recording bath under an inverted microscope. The blank control solution and the working solution of the compound of this invention were sequentially flowed through the recording bath from low to high concentration using gravity perfusion, thereby acting on the cells. During recording, a peristaltic pump was used for fluid exchange. Each concentration was measured independently twice using at least two cells.

[0184] The peak current of the GABA(α4β3δ) receptor after reacting each concentration of the compound of the present invention with the mixture of GABAECx GABAECX+Compound ) and GABA ECx-induced receptor current peak GABA ECX Normalization was performed to calculate the PAM effect (%Normalized to GABA EC) of the compounds of this invention. X ),

[0185] Right now, by This serves as the quality control standard. Simultaneously, the mean, standard deviation (SD), and standard error (SE) are calculated for each concentration-effect ratio, with data expressed as mean ± SE.

[0186] Y=Bottom+(Top-Bottom) / (1+10^((LogEC 50 -X)×HillSlope))

[0187] The EC values ​​of the compounds of this invention can be calculated using the above equations. 50 The values ​​were calculated, and a nonlinear fit was performed on the dose-dependent effect, where EC 50 This is the half-maximal effect concentration. EC 50 The calculations and curve fitting were performed using GraphPad Prism software.

[0188] The test results are shown in Table 2.

[0189] Table 2.

[0190]

[0191] EC 50 (nM) range:

[0192] A:EC 50 <300nM;

[0193] B: 300nM≤EC 50 <500nM;

[0194] C:500nM≤EC 50 <1000nM;

[0195] D:EC 50 ≥1000nM.

[0196] Table 2 shows that the compound activity (EC) 50 (The smaller the value, the stronger the activity) 2>1>3, 5>4>6, 8>7>9, 10>11, 12>13, 14>15, 16>17, 18>19, 20>21, that is, the bioactivity data proves that among the compounds selected in this invention, R x Compounds facing inward (e.g., compounds 2, 5, 8) are more reactive than chiral substituted compounds (e.g., compounds 1, 4, 7), and significantly more reactive than R. x The out-of-plane reactivity of compounds (e.g., compounds 3, 6, and 9) demonstrates in-plane reactivity towards R. x In GABA A They play a crucial role in receptor regulation. Furthermore, compounds 5, 10, 14, 45, 52, and 53 of this invention have effects on GABA(α4β3δ) active EC.50 All values ​​are less than 300 nM, indicating that the compounds of this invention have a high affinity for GABA. A The receptor has a high positive regulatory efficiency.

[0197] Liver microsomal stability test:

[0198] Liver microsomal metabolic stability assays are the most widely used in vitro model for studying the metabolic transformation of drugs under the action of phase I metabolic enzymes (such as cytochrome P450 enzymes). Because drug metabolism varies across species, liver microsomal metabolic stability assays are typically performed in multiple species. The most commonly used species include humans for predicting clinical doses, rats for toxicity studies, mice for specific pharmacodynamic models, and dogs and monkeys for large animal toxicology studies.

[0199] This study used a commercially available female SD rat liver phase I metabolic stability kit (containing liver microsomes and reagents used in the study) to conduct in vitro metabolic research.

[0200] Store microparticles in a refrigerator (below -60.0°C) before use. See the table below for detailed microparticle information.

[0201] Species strain gender Number of donors source Item number batch number rats SD female 300 IPHASE 0121D1.01 23F013

[0202] Reagents and consumables:

[0203]

[0204]

[0205] instrument:

[0206] Water bath (HH-42, Guohua, Changzhou, China)

[0207] Pipetting workstation (iPPA2-96-500(E), Apricot, Covina, USA)

[0208] Pipetting workstation (iPPA2-96-125(E), Apricot, Covina, USA)

[0209] Centrifuge (5910R, Eppendorf, Hamburg, Germany)

[0210] Experimental steps:

[0211] Prepare eight 96-well incubation plates, named T0, T5, T15, T30, T45, T60, Blank60, and NCF60. The reaction time points for the first six incubation plates are 0, 5, 15, 30, 45, and 60 min, respectively. Do not add the test or control compound to the Blank60 plate; sample after 60 min of incubation. In the NCF60 plate, use potassium phosphate buffer instead of the NADPH regeneration solution for 60 min of incubation. Samples at all time points are in triplicate.

[0212] Add 2.00 μL of the test sample or control working solution and 100 μL of microsomal working solution (liver microsomal protein concentration of 1.00 mg / mL) to the T0, T5, T15, T30, T45, T60 and NCF60 plates respectively. Add only microsomal working solution to the Blank60 plate. Then, place the incubation plates (Blank60, T5, T15, T30, T45 and T60, except for T0 and NCF60) in a 37.0℃ water bath for pre-incubation for about 10 min.

[0213] For T0 plates, first add 600 μL of stop solution (the stop solution for both test and control samples is an acetonitrile solution containing 250 nM tolbutamide) followed by the NADPH regeneration working solution. Add 98.0 μL of potassium phosphate buffer to each well of the NCF60 plate and incubate for 60 min. After pre-incubation at Blank 60, T5, T15, T30, T45, and T60 plates, add 98.0 μL of NADPH regeneration working solution to each sample well to initiate the reaction.

[0214] After incubation for appropriate times (e.g., 5, 15, 30, 45, and 60 min), 600 μL of stop solution was added to each well of the test sample and the reference sample on Blank 60, T5, T15, T30, T45, T60, and NCF60 plates to terminate the reaction. All sample plates were shaken well and centrifuged at 3220 × g for 20 min. 100 μL of the test sample supernatant was diluted in 300 μL of pure water containing 0.3% formic acid and 50% acetonitrile for LC-MS / MS analysis, and 100 μL of the reference sample supernatant was diluted in 300 μL of pure water for LC-MS / MS analysis. Sample analysis was performed using liquid chromatography-tandem mass spectrometry (LC-MS / MS). Retention times of analytes and internal standards, chromatographic acquisition, and chromatographic integration were processed using Analyst software (Sciex, Framingham, MA, USA).

[0215] Data Analysis

[0216] The in vitro elimination rate constant k of the test and control compounds can be obtained by converting the ratio of the peak area of ​​the compound to that of the internal standard into the residual percentage using the following formula.e :

[0217]

[0218]

[0219] when

[0220]

[0221] via k e Calculation of in vitro intrinsic clearance of liver microsomes (CL) int(mic) ) and hepatic intrinsic clearance (CL) (liver) ).

[0222] CL int(mic) =0.693 / T 1 / 2 / Microsomal protein content (microsomal concentration during incubation, mg / mL).

[0223] CL int(liver) =CL int(mic) × Microsomal protein content in the liver (mg / g) × Liver weight to body weight ratio

[0224] Based on the well-stirred model, the in vivo liver clearance rate (CL) was estimated. (liver) ):

[0225] CL (liver) =(CL) int(liver) ×f u ×Q h ) / (CL int(liver) ×f u +Q h ), default f u The free fraction in blood equals 1. The parameters used in the formula are as follows:

[0226]

[0227] The results of the experiment, including the concentration data of the six compounds and the positive control drug SAGE-217 (Female Rat) at various time points, are summarized in Table 3.

[0228] Table 3.

[0229]

[0230]

[0231] The remaining percentage trends of each compound in Table 3 are as follows: Figure 1 As shown, the horizontal axis represents time, and the vertical axis represents the remaining percentage.

[0232] Table 3 summarizes the in vitro half-life and intrinsic clearance rate of each compound, as shown in Table 4.

[0233] Table 4.

[0234]

[0235] Data Analysis and Discussion:

[0236] From Table 3, Figure 1 As shown in Table 4, compound 10 has significantly better liver microsomal stability than SAGE-217, compound 52 has similar liver microsomal stability to SAGE-217, and compound 45 has slightly worse liver microsomal stability than SAGE-217.

[0237] In vivo pharmacokinetic studies:

[0238] Study Objective: To investigate the pharmacokinetics of the target compound in female SD rats after single oral gavage and single tail vein administration, with a focus on determining its C60-62% pharmacokinetic profile. max T 1 / 2 Information such as AUC was recorded; the status and behavior of the animals at different time points after administration were recorded in detail; and the oral bioavailability was calculated based on the pharmacokinetic data of tail vein administration.

[0239] Experimental drug:

[0240] Compounds 10, 45, and 52: 0.6 mg / ml aqueous solution (orally administered using 30% SBECD as a solvent);

[0241] Compounds 10, 45, and 52: 0.2 mg / ml aqueous solution (intravenously administered using 30% SBECD as solvent).

[0242] Test method:

[0243] Oral administration to rats: Three female SD rats were administered compound X orally once by gavage at a dose of 6 mg / kg (approximately 2 ml of drug solution). Blood samples were collected from the orbital venous plexus at 30 min, 1 h, 1.5 h, 2 h, 3 h, 4 h, 6 h, 8 h, 12 h, and 24 h post-administration. After heparin-anticoagulated centrifugation, the plasma was separated for analysis.

[0244] Tail vein administration in rats: Three female SD rats were administered compound X once via tail vein at a dose of 2 mg / kg (approximately 2 ml of drug solution). Blood samples were collected from the orbital venous plexus at 5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, 12 h, and 24 h post-administration. After heparin-anticoagulated centrifugation, plasma was separated for analysis.

[0245] Plasma sample processing: Accurately measure 50 μL of plasma sample, add 150 μL of protein precipitant (acetonitrile solution of 5 ng / mL testosterone-d3, 5 ng / mL terfenadine, and 5 ng / mL Buspirone), vortex for 1 min, centrifuge (4000 rpm, 4℃) for 10 min, take 120 μL of supernatant and add 40 μL of 50% methanol aqueous solution, vortex for 1 min, and detect by UPLC-MS / MS.

[0246] Data calculation and statistics:

[0247] Drug concentrations were directly read from UPLC-MS / MS, and the mean, standard deviation, and coefficient of variation were calculated using Excel. max C max Pharmacokinetic parameters such as AUC were calculated using DAS2.0 software.

[0248] The pharmacokinetic parameters of compounds 10, 45, and 52 are shown in Table 5.

[0249] Table 5.

[0250]

[0251] Data Analysis:

[0252] PK parameters in female SD rats showed that compounds 10 and 52 had good metabolic stability, good oral absorption, high blood drug concentrations and exposure, and high bioavailability.

[0253] This invention provides a novel, highly effective, orally administered antidepressant for major depressive disorder and postpartum depression. Through allosteric regulation, it can simultaneously act on multiple GABA groups on the synaptic surface. A Ion channel receptors facilitate the influx of chloride ions into cells along the electrochemical gradient of the GABA receptor complex (GRC). Increased intracellular levels of this anion lead to hyperpolarization of the transmembrane potential, making neurons less sensitive to excitatory input, i.e., reducing brain excitability. The compounds of this invention target GABA... A Receptors have a strong positive regulatory effect, which is beneficial to improving the bioavailability of drugs.

[0254] While numerous embodiments of the invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. Many modifications, alterations, and alternatives will occur to those skilled in the art without departing from the spirit and intent of the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in the practice of the invention. The appended claims are intended to define the scope of protection of the invention and therefore cover equivalents or alternatives within the scope of these claims.

Claims

1. A steroidal compound having the structure of general formula I and a pharmaceutically acceptable salt thereof, characterized in that, （Ⅰ） in, R1, R4, and R5 are all hydrogen; R2 and R3 are selected from hydrogen, C1-C6 alkane groups and their halogenated derivatives, respectively. R x Selected from: nitro, carboxyl, C1-C6 alkane groups and their halogenated derivatives, C1-C6 aliphatic chain substituents containing oxygen heteroatoms and their halogenated derivatives, C1-C6 aliphatic chain substituents containing sulfur heteroatoms and their halogenated derivatives, C1-C6 aliphatic chain substituents containing nitrogen heteroatoms and their halogenated derivatives, six-membered ring aromatic hydrocarbon groups, C3-C7 cycloalkane groups, hydroxyl, amino, and mercapto groups; Or, either R2 or R3 is related to R. x C8-C12 nitrogen-bridged rings formed by phase connection; in, M is selected from nitrogen and oxygen. When M is nitrogen, either R2 or R3 is absent; When M is oxygen, R2 or R3 does not exist; R x Facing inwards.

2. The steroidal compound having the structure of general formula I and its pharmaceutically acceptable salt according to claim 1, characterized in that, R2 and R3 are selected from: hydrogen, methyl, monofluoromethyl, difluoromethyl, and trifluoromethyl, respectively.

3. The steroidal compound having the structure of general formula I according to any one of claims 1 to 2, and its pharmaceutically acceptable salt, characterized in that, R1, R2, R3, R4, and R5 have exactly three hydrogen atoms; or, R1, R2, R3, R4, and R5 are exactly four hydrogen atoms; or, R1, R2, R3, R4, and R5 are all hydrogen.

4. A steroidal compound and its pharmaceutically acceptable salt, characterized in that, The steroidal compounds are selected from: 。 5. A pharmaceutical composition, characterized in that, The active ingredient comprises any one of the steroidal compounds of claims 1 to 4 and their pharmaceutically acceptable salts.

6. The pharmaceutical composition according to claim 5, characterized in that, It also includes pharmaceutical carriers or diluents.

7. The use of the pharmaceutical composition according to claim 5 as a medicament for the preparation of a treatment for central nervous system-related disorders. The central nervous system-related conditions include: Postpartum depression, clinical depression, atypical depression, severe depressive disorder, catatonic depression, dysphoric mood, dual depression, depressive personality disorder, recurrent transient depression, mild depressive disorder, bipolar disorder, manic-depressive disorder, mood disorder, anxiety, post-traumatic stress disorder (PTSD), premenstrual anxiety disorder (PMDD), depression due to chronic medical conditions, treatment-resistant depression, suicidal ideation or behavior, premenstrual syndrome, generalized anxiety disorder, seasonal affective disorder (SAD), social anxiety disorder, memory loss, poor stress tolerance, Niemann-Pick II type C disease or related neurological or physical symptoms, epilepsy, essential tremor, epileptiform disorder, NMDA insufficiency, migraine, status epilepticus, sleep disorder, fragile X syndrome, 5α-reductase inhibitor-induced depression, PCDH19 female childhood epilepsy, sexual dysfunction, cognitive impairment, Parkinson's disease or Alzheimer's disease.

8. The use of any one of the steroidal compounds of claims 1 to 4 and their pharmaceutically acceptable salts as a medicament for the preparation of a treatment for central nervous system-related disorders. The central nervous system-related conditions include: Postpartum depression, clinical depression, atypical depression, severe depressive disorder, catatonic depression, dysphoric mood, dual depression, depressive personality disorder, recurrent transient depression, mild depressive disorder, bipolar disorder, manic-depressive disorder, mood disorder, anxiety, post-traumatic stress disorder (PTSD), premenstrual anxiety disorder (PMDD), depression due to chronic medical conditions, treatment-resistant depression, suicidal ideation or behavior, premenstrual syndrome, generalized anxiety disorder, seasonal affective disorder (SAD), social anxiety disorder, memory loss, poor stress tolerance, Niemann-Pick II type C disease or related neurological or physical symptoms, epilepsy, essential tremor, epileptiform disorder, NMDA insufficiency, migraine, status epilepticus, sleep disorder, fragile X syndrome, 5α-reductase inhibitor-induced depression, PCDH19 female childhood epilepsy, sexual dysfunction, cognitive impairment, Parkinson's disease or Alzheimer's disease.

9. A method for preparing a steroidal compound having the structure of general formula I according to any one of claims 1 to 3, characterized in that, The preparation route is as follows: Among them, R1~R x The definitions of R1~R are the same as those for steroidal compounds having the general formula I structure as described in any one of claims 1-3. x .

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