CN115650928B - Polycyclic thyroid hormone beta receptor agonist and application thereof - Google Patents
Polycyclic thyroid hormone beta receptor agonist and application thereof Download PDFInfo
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Abstract
The invention relates to a polycyclic thyroid hormone beta receptor agonist and application thereof, in particular to a compound shown as a formula (1) or a pharmaceutically acceptable form thereof, a pharmaceutical composition containing the compound, and a preparation method and application thereof. The compound or the pharmaceutical composition can be used for preparing medicines for preventing, treating or alleviating diseases modulated by thyroid hormone beta receptors.
Description
Technical Field
The invention belongs to the field of medicinal chemistry, and relates to polycyclic compounds serving as thyroid hormone beta receptor agonists, a pharmaceutical composition containing the polycyclic compounds, a preparation method of the polycyclic compounds, and application of the polycyclic compounds in preparing medicines for preventing, treating or relieving diseases regulated by thyroid hormone beta receptors.
Background
Thyroid Hormone (TH) is synthesized in the thyroid gland in response to Thyroid Stimulating Hormone (TSH) secreted by the pituitary. Thyroxine plays a very important role in regulating body growth, development, metabolism, and matrix balance. Thyroid hormones function by binding to Thyroid Hormone Receptors (THR). The THRs belong to the nuclear receptor superfamily, which forms heterodimers with their common ligand, retinoid X receptors, and function as ligand-induced transcription factors. Like other nuclear receptors, THRs have a ligand binding domain and a DNA binding domain, and regulate gene expression through ligand-dependent interaction with DNA response elements (thyroid response elements, THREs).
Currently, there are two subtypes of THR: THR α and THR β. THR alpha is mainly distributed in heart tissues and plays an important role in regulating and controlling the functions of the heart. THR β is expressed mainly in the liver and pituitary, regulates the metabolism of fatty acids and cholesterol, and regulates thyroid stimulating hormone secretion. THR α and THR β are both expressed in Brown Adipose Tissue (BAT) and play an important role in regulating basal oxygen consumption, fat storage, lipogenesis and lipolysis (Oppenheimer et al, J.Clin. Invest.87 (1): 125-32 (1991)).
THR agonists increase metabolic rate, oxygen consumption and thermogenesis, promote cholesterol metabolism to bile acids, and, in addition, lower the lipoprotein levels associated with atherosclerosis. The liver and heart are the major target organs for THR agonists. Genes involved in the synthesis and metabolism of fatty acids and cholesterol are mainly regulated in the liver and affect carbohydrates by increasing glycogenolysis and gluconeogenesis and decreasing the action of insulin. In the heart, systemic vascular resistance can be reduced, blood volume increased and inotropic and chronotropic effects produced.
The THR beta agonist can also improve the metabolism of cell lipid and play the role of reducing cholesterol and blood fat. Therefore, it is of great interest to research and develop THR β agonists for the treatment and/or prevention of diseases modulated by thyroid hormone receptors.
Disclosure of Invention
Through a large number of researches, a series of polycyclic compounds serving as thyroid hormone beta receptor agonists are discovered, and the polycyclic compounds have potential values of preventing and/or treating diseases regulated by thyroid hormone beta receptors.
In a first aspect, the present invention provides a compound having the structure of formula (1) or a pharmaceutically acceptable form thereof:
wherein the content of the first and second substances,
R 1 selected from H, halogen, -CN, -NH 2 、-NO 2 -OH or C 1-6 Alkyl radical, said C 1-6 Alkyl is optionally substituted by one or more groups independently selected from halogen, -CN, -NH 2 、-NO 2 or-OH;
R 2 and R 3 Independently selected from H, halogen, -CN, -NH 2 、-NO 2 -OH or C 1-6 Alkyl radical, said C 1-6 Alkyl is optionally substituted by one or more groups independently selected from halogen, -CN, -NH 2 、-NO 2 or-OH;
R 4 is selected from-O- (C) 1-6 Alkyl), -NH- (C) 1-6 Alkyl), -S- (C) 1-6 Alkyl), -S (= O) - (C) 1-6 Alkyl), -S (= O) 2 -(C 1-6 Alkyl), -C (= O) - (C) 1-6 Alkyl), -C (= O) - (C) 3-8 Cycloalkyl), C 1-6 Alkyl or-NHC (= O) - (C) 1-6 Alkyl group), said C 1-6 Alkyl is optionally substituted by one or more groups independently selected from halogen, -O- (C) 1-6 Alkyl), -CN, -NH 2 、-NO 2 or-OH;
the pharmaceutically acceptable form is selected from the group consisting of pharmaceutically acceptable salts, esters, stereoisomers, tautomers, solvates, nitrogen oxides, isotopic labels, metabolites and prodrugs.
In some embodiments, R 1 Selected from H, F, cl, br, -CN, -NH 2 Or C 1-4 Alkyl radical, said C 1-4 Alkyl is optionally substituted by one or more groups independently selected from F, cl, br, -CN, -NH 2 or-OH.
In some preferred embodiments, R 1 Selected from H, -CN, -NH 2 、-CH 3 、-CH 2 F、-CHF 2 or-CF 3 (ii) a More preferably, R 1 Selected from H or-CHF 2 。
In some embodiments, R 2 And R 3 Independently selected from H, F, cl, br, -CN, -NH 2 Or C 1-4 Alkyl radical, said C 1-4 Alkyl is optionally substituted by one or more groups independently selected from F, cl, br, -CN, -NH 2 、-NO 2 or-OH.
In some preferred embodiments, R 2 And R 3 Independently selected from H, F, cl, br or-CH 3 (ii) a More preferably, R 2 And R 3 Independently selected from Cl or Br.
In some embodiments, R 4 Is selected from-O- (C) 1-4 Alkyl), -NH- (C) 1-4 Alkyl), -S- (C) 1-4 Alkyl), -S (= O) - (C) 1-4 Alkyl), -S (= O) 2 -(C 1-4 Alkyl), -C (= O) - (C) 1-4 Alkyl), -C (= O) - (C) 3-6 Cycloalkyl), C 1-4 Alkyl or-NHC (= O) - (C) 1-4 Alkyl group), said C 1-4 Alkyl is optionally substituted by one or more groups independently selected from halogen, -O- (C) 1-4 Alkyl), -CN, -NH 2 or-OH.
In some preferred embodiments, R 4 Is selected from-OCH 3 、-NHCH 3 、-NHC(=O)CH 3 、-SCH 3 、-S(=O)CH 3 、-S(=O) 2 CH 3 、-C(=O)CH 3 、-C(=O)CH 2 CH 3 、-C(=O)CH 2 CH 2 CH 3 、
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It will be understood by those skilled in the art that the present invention encompasses compounds resulting from any combination of the various embodiments. Embodiments resulting from a combination of technical features or preferred technical features in one embodiment with technical features or preferred technical features in another embodiment are also included in the scope of the present invention.
In a second aspect, the present invention also provides a compound selected from the group consisting of:
in a third aspect, the present invention provides a method for producing a compound represented by formula (1), comprising the steps of:
step 1: synthesis of intermediate M1
(a) Reacting a compound shown in a general formula I serving as an initial raw material under the action of N-bromosuccinimide and a free radical initiator to obtain a compound shown in a general formula M1;
in some embodiments, step 1 above (a) is carried out in the presence of a free radical initiator selected from the group consisting of azobisisobutyronitrile, azobisisoheptonitrile, cumene hydroperoxide, tert-butyl hydroperoxide, p-menthane hydroperoxide, dibenzoyl peroxide, lauroyl peroxide, di-tert-butyl peroxide, dicumyl peroxide, preferably azobisisobutyronitrile.
And 2, step: synthesis of Compound represented by the formula (1)
(b) Reacting a compound shown in a general formula M1 serving as a starting material with a compound shown in a general formula II under the action of alkali to obtain a compound shown in a general formula III;
(c) Reacting the compound shown in the general formula III and the compound shown in the general formula IV with a copper catalyst under the action of alkali to obtain a compound shown in a formula (1);
in some embodiments, step 2 (b) above is performed in the presence of a base selected from triethylamine, N-diisopropylethylamine, pyridine, imidazole, 1, 8-diazabicycloundecen-7-ene, sodium carbonate, potassium carbonate, cesium carbonate, lithium hydroxide, sodium hydroxide, potassium hydroxide, sodium methoxide, sodium ethoxide, potassium acetate, sodium acetate, preferably potassium carbonate.
In some embodiments, step 2 (c) above is performed in the presence of a base selected from triethylamine, N-diisopropylethylamine, pyridine, imidazole, 1, 8-diazabicycloundecen-7-ene, sodium carbonate, potassium carbonate, cesium carbonate, lithium hydroxide, sodium hydroxide, potassium phosphate, potassium hydroxide, sodium methoxide, sodium ethoxide, potassium ethoxide, preferably potassium phosphate and potassium carbonate.
In some embodiments, step 2 (c) above is carried out in the presence of a copper catalyst selected from cuprous oxide, cuprous chloride, cuprous iodide, cuprous thiocyanate, cupric acetate, cuprous bromide, copper, cupric oxide, cupric chloride, cupric bromide, cupric iodide, preferably cuprous iodide.
In a fourth aspect, the present invention provides a pharmaceutical composition comprising at least one compound of formula (1) as described above, or a pharmaceutically acceptable form thereof, and one or more pharmaceutically acceptable carriers.
In a fifth aspect, the present invention provides a compound of formula (1) as described above or a pharmaceutically acceptable form thereof, or a pharmaceutical composition as described above, for use as a thyroid hormone beta receptor agonist, for the prevention and/or treatment of a disease or condition mediated at least in part by the thyroid hormone beta receptor.
In a sixth aspect, the present invention provides the use of a compound of formula (1) as described above or a pharmaceutically acceptable form thereof, or a pharmaceutical composition as described above, in the manufacture of a medicament for the prevention and/or treatment of a disease or condition mediated at least in part by the thyroid hormone β receptor (e.g. a metabolic disease such as non-alcoholic fatty liver disease, dyslipidaemia, atherosclerosis or hypothyroidism).
In a seventh aspect, the present invention provides a method for the prevention and/or treatment of a disease or condition mediated at least in part by thyroid hormone beta receptor, comprising the steps of: administering to a subject in need thereof a prophylactically and/or therapeutically effective amount of a compound of formula (1) or a pharmaceutically acceptable form thereof as described above, or a pharmaceutical composition as described above.
The present invention is not limited to the particular embodiments described herein; it is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.
Definition of terms
The following terms have the following meanings in the present invention unless otherwise specified.
The terms "comprises," "comprising," "includes," "including," "has," "having" or "containing," or any other variation thereof, are intended to cover a non-exclusive or open-ended inclusion. For example, a composition, method, or apparatus that comprises a list of elements is not necessarily limited to only those elements explicitly listed, but may include other elements not explicitly listed or inherent to such composition, method, or apparatus.
When the lower and upper limits of a range of values are disclosed, any value or any sub-range falling within the range is specifically disclosed. In particular, each numerical range of parameters disclosed herein (e.g., in the form of "about a to b," or equivalently "about a-b") is to be understood to encompass each number and subrange therein. For example, "C 1-4 "is to be understood to cover any subrange therein as well as each point value, e.g. C 2-4 、C 3-4 、C 1-2 、C 1-3 、C 1-4 Etc. and C 1 、C 2 、C 3 、C 4 And the like. Also for example, "5-10 way" should be understood to encompass any subrange therein as well as each point value, e.g., 5-6 way, 5-7 way, 5-8 way, 5-9 way, 6-7 way, 6-8 way, etc., as well as 5, 6, 7, 8, 9, 10 way, etc.
The term "substituted" and other variations thereof herein mean that one or more (e.g., 1,2, 3, or 4) atoms or groups of atoms (e.g., hydrogen atoms) on the designated atom is replaced with other equivalents, provided that the designated atom or group of atoms does not exceed the normal valence under the current circumstances and that a stable compound can be formed. If an atom or group of atoms is described as "optionally substituted with" \8230; "substituted", it may be substituted or unsubstituted. Unless otherwise indicated, the attachment site of a substituent herein may be from any suitable position of the substituent. When a bond in a substituent is shown as crossing a chemical bond between two atoms attached to each other in a ring system, it means that the substituent may be attached to any one of the ring system atoms.
The term "pharmaceutical composition" refers to a composition that can be used as a medicament, comprising a pharmaceutically active ingredient (or therapeutic agent) and optionally one or more pharmaceutically acceptable carriers. The term "pharmaceutically acceptable carrier" refers to an excipient that is administered with a therapeutic agent, and which is, within the scope of sound medical judgment, suitable for contact with the tissues of humans and/or other animals without excessive toxicity, irritation, allergic response, or other problem or complication commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable carriers that may be used in the present invention include, but are not limited to: a) A diluent; b) A lubricant; c) A binder; d) A disintegrant; e) Absorbents, coloring, flavoring and/or sweetening agents; f) An emulsifier or dispersant; and/or g) substances that enhance the absorption of the compounds, and the like.
The pharmaceutical compositions described above may act systemically and/or locally. For this purpose, they may be administered by a suitable route, for example by parenteral, topical, intravenous, oral, subcutaneous, intraarterial, intradermal, transdermal, rectal, intracranial, intraperitoneal, intranasal, intramuscular routes or as an inhalant.
The above administration route can be achieved by a suitable dosage form. Dosage forms that may be used in the present invention include, but are not limited to: tablets, capsules, troches, hard candies, powders, sprays, creams, ointments, suppositories, gels, pastes, lotions, ointments, aqueous suspensions, injectable solutions, elixirs, syrups, and the like.
When administered orally, the above pharmaceutical compositions may be formulated into any orally acceptable dosage form, including, but not limited to, tablets, capsules, aqueous solutions, aqueous suspensions, and the like.
The pharmaceutical compositions described above may also be administered in the form of sterile injectable preparations, including sterile injectable aqueous or oleaginous suspensions, or sterile injectable aqueous or oleaginous solutions. Among the carriers that can be used are, but not limited to: water, ringer's solution and isotonic sodium chloride solution. In addition, the sterilized fixed oil may also be employed as a solvent or suspending medium, such as a monoglyceride or diglyceride.
The above pharmaceutical composition may comprise 0.01mg to 1000mg of at least one compound of the above formulae (1) to (3) or a pharmaceutically acceptable form thereof.
The term "a disease or condition mediated at least in part by the thyroid hormone beta receptor" refers to a disease that involves at least a portion of the factors associated with the thyroid hormone beta receptor in its pathogenesis, such as metabolic diseases, e.g., non-alcoholic fatty liver disease, dyslipidemia, atherosclerosis, or hypothyroidism.
The term "effective amount" refers to a dose that is capable of inducing a biological or medical response in a cell, tissue, organ or organism (e.g., an individual) and is sufficient to achieve a desired prophylactic and/or therapeutic effect.
The dosing regimen may be adjusted to provide the best desired response. For example, it may be administered in a single dose, may be administered in divided doses over time, or may be administered after proportionally decreasing or increasing the dose as the case may be. It will be appreciated that for any particular individual, the specific dosage regimen will be adjusted as needed and as the professional judgment of the person administering the composition or supervising it.
The term "in need thereof" refers to a judgment by a physician or other caregiver that an individual needs or will benefit from a prophylactic and/or therapeutic procedure, the judgment being made based on various factors of the physician or other caregiver in their area of expertise.
The term "individual" (or subject) refers to a human or non-human animal. The subject of the invention includes both subjects (patients) suffering from a disease and/or disorder and normal subjects. Non-human animals of the invention include all vertebrates, e.g., non-mammals, such as birds, amphibians, reptiles, and the like, and mammals, e.g., non-human primates, livestock, and/or domesticated animals (e.g., sheep, dogs, cats, cows, pigs, and the like).
The term "treating" refers to alleviating or eliminating the disease or disorder in question. A subject is successfully "treated" if the subject receives a therapeutic amount of a compound of the invention or a pharmaceutically acceptable form thereof, or a pharmaceutical composition of the invention, and the subject exhibits an observable and/or detectable remission and/or improvement of at least one of the indications and symptoms. It is understood that treatment includes not only complete treatment, but also less than complete treatment, but achieves some biologically or medically relevant result. In particular, "treatment" means that the compound of the invention or a pharmaceutically acceptable form thereof, or the pharmaceutical composition of the invention, can achieve at least one of the following effects, for example: (1) Preventing disease from occurring in an animal that may be predisposed to the disease but has not yet experienced or exhibited disease pathology or symptomology; (2) Inhibiting disease in an animal experiencing or exhibiting disease pathology or symptomatology (i.e., arresting further development of pathology and/or symptomatology); (3) Ameliorating the disease (i.e., reversing pathology and/or symptomatology) in an animal experiencing or exhibiting disease pathology or symptomatology.
The term "pharmaceutically acceptable salt" refers to salts of the compounds of the present invention that are substantially non-toxic to organisms. Pharmaceutically acceptable salts generally include, but are not limited to, salts formed by reacting a compound of the invention with a pharmaceutically acceptable inorganic/organic acid or inorganic/organic base, such salts also being referred to as acid addition salts or base addition salts. For a review of suitable salts see, for example, jusiak, soczewinski,et al., Remington’s Pharmaceutical Sciences [M],Mack Publishing Company2005 and Stahl, wermuth, handbook of Pharmaceutical Salts: properties, selection, and Use [ M],Wiley-VCH,2002. Methods for preparing pharmaceutically acceptable salts of the compounds of the present invention are known to those skilled in the art.
The term "pharmaceutically acceptable ester" refers to an ester that is substantially non-toxic to an organism and that hydrolyzes in vivo to a compound of the invention or a salt thereof. Pharmaceutically acceptable esters generally include, but are not limited to, esters of the compounds of the present invention with pharmaceutically acceptable carboxylic or sulfonic acids, such esters also being referred to as carboxylic or sulfonic esters.
The term "isomers" refers to compounds having the same molecular weight, but differing in the spatial arrangement or configuration of the atoms, due to the same number and type of atoms.
The term "stereoisomer" (or "optical isomer") refers to a stable isomer having a perpendicular plane of asymmetry due to having at least one chiral factor (including chiral center, chiral axis, chiral plane, etc.) that enables rotation of plane polarized light. Since the compounds of the present invention contain asymmetric centers as well as other chemical structures that may lead to stereoisomers, the present invention also includes such stereoisomers and mixtures thereof. Unless otherwise indicated, all stereoisomeric forms of the compounds of the present invention are within the scope of the present invention.
The term "tautomer" (or "tautomeric form") refers to structural isomers having different energies that can interconvert through a low energy barrier. If tautomerism is possible (e.g., in solution), then the chemical equilibrium of the tautomer can be reached. For example, proton tautomers (or proton transfer tautomers) include, but are not limited to, interconversions by proton transfer, such as keto-enol isomerization, imine-enamine isomerization, amide-iminoalcohol isomerization, and the like. Unless otherwise indicated, all tautomeric forms of the compounds of the invention are within the scope of the invention.
The term "solvate" refers to a substance formed by the binding of a compound of the present invention (or a pharmaceutically acceptable salt thereof) to at least one solvent molecule by non-covalent intermolecular forces. For example, solvates include, but are not limited to, hydrates (including hemihydrate, monohydrate, dihydrate, trihydrate, and the like), ethanolates, acetonates, and the like.
The term "nitroxide" refers to a compound formed by oxidation of a nitrogen atom in the structure of a tertiary amine or nitrogen (aryl) containing heterocyclic compound. For example, the nitrogen atom in the parent nucleus of a compound of formula I may form the corresponding nitroxide.
The term "isotopic label" refers to a derivatized compound formed by replacing a particular atom in a compound of the invention with its isotopic atom. Unless otherwise indicated, the compounds of the present invention include various isotopes of H, C, N, O, F, P, S, cl, such as, but not limited to 2 H(D)、 3 H(T)、 13 C、 14 C、 15 N、 17 O、 18 O、 18 F、 31 P、 32 P、 35 S、 36 S and 37 Cl。
the term "metabolite" refers to a derivative compound formed after the compounds of the present invention are metabolized. Further information on metabolism can be found in Goodman and Gilman's: the pharmaceutical Basis of Therapeutics (9) th ed.) [M]McGraw-hillInternational proportions, 1996. The present invention encompasses all possible metabolite forms of the compounds of the invention, i.e. substances formed in the body of the individual to whom the compounds of the invention are administered. Metabolites of a compound can be identified by techniques well known in the art, and their activity can be characterized by assays.
The term "prodrug" refers to a derivative compound that is capable of providing, directly or indirectly, a compound of the invention upon administration to a subject. Particularly preferred derivative compounds or prodrugs are those that, when administered to a subject, increase the bioavailability of the compounds of the invention (e.g., more readily absorbed into the blood), or facilitate delivery of the parent compound to the site of action (e.g., the lymphatic system). Unless otherwise indicated, all prodrug forms of the compounds of the present invention are within the scope of the present invention, and various prodrug forms are known in the art, see, e.g., T. Higuchi, V. Stella, pro-drugs as Novel Drug Delivery Systems [ J],American Chemical SocietyVol. 14,1975. Furthermore, the present invention also encompasses compounds of the present invention containing protecting groups. In any process for the preparation of the compounds of the present invention, it may be necessary and/or desirable to protect sensitive or reactive groups on any of the molecules concerned, thereby forming a chemically protected form of the compounds of the present invention. This can be achieved by conventional protecting Groups, for example as described in T.W. Greene, P.G.M. Wuts, protective Groups in Organic Synthesis [ M],John Wiley&SonsProtecting groups as described in 2006. These protecting groups may be removed at a suitable subsequent stage using methods known in the art.
The term "independently" means that at least two groups (or ring systems) present in a structure that are the same or similar in value can have the same or different meaning in a particular instance. For example, substituent X and substituent Y are independently hydrogen, halogen, hydroxy, cyano, alkyl or aryl, and when substituent X is hydrogen, substituent Y may be either hydrogen, halogen, hydroxy, cyano, alkyl or aryl; similarly, when the substituent Y is hydrogen, the substituent X may be hydrogen, or may be halogen, hydroxy, cyano, alkyl or aryl.
The term "halogen", when used herein alone or in combination with other groups, refers to fluorine (F), chlorine (Cl), bromine (Br), and iodine (I).
The term "alkyl", when used herein alone or in combination with other groups, refers to a straight or branched chain aliphatic hydrocarbon group. For example, the term "C" as used in the present invention 1-6 Alkyl "refers to an alkyl group having 1 to 6 carbon atoms. For example, the alkyl group may be a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, a sec-butyl group, an isobutyl group, a tert-butyl group or the like.
The term "cycloalkyl", as used herein alone or in combination with other groups, refers to a saturated monocyclic or polycyclic (such as bicyclic, e.g., fused, bridged, or spiro) nonaromatic hydrocarbon group. For example, the term "C" as used in the present invention 3-6 Cycloalkyl "refers to cycloalkyl groups having 3 to 6 carbon atoms. For example, cycloalkyl can be cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl or bicyclo [2.2.1 ]]Heptyl, and the like. Cycloalkyl groups in the present invention are optionally substituted with one or more substituents described herein.
Detailed Description
In order to make the objects and technical solutions of the present invention clearer, embodiments of the present invention will be described in detail below with reference to examples. It will be understood by those skilled in the art that the following examples are illustrative of the present invention only and should not be taken as limiting the scope of the invention.
The reagents or instruments used in the examples are all conventional products which are commercially available. Those who do not have specific conditions noted are conducted under conventional conditions or conditions recommended by the manufacturer. The term "room temperature" used in the present invention means 20 ℃. + -. 5 ℃. As used herein, the term "about" when used in reference to a value or range of values is intended to encompass the value or range of values and tolerances acceptable to those skilled in the art for that value or range of values, e.g., within ± 10%, ± 5%, ± 4%, ± 3%, ± 2%, ± 1%, ± 0.5%, etc.
The structures of the compounds described in the following examples were determined by Nuclear Magnetic Resonance (NMR) and/or Mass Spectrometry (MS).
Nuclear Magnetic Resonance (NMR) measuring apparatus Bruker 400 MHz NMR was used, and deuterated methanol (CD) was used as a measuring solvent 3 OD), deuterated chloroform (CDCl) 3 ) Hexadeuterio dimethyl sulfoxide (DMSO-d) 6 ) The internal standard substance is Tetramethylsilane (TMS). In that 1 In H NMR, part of hydrogen may not be peaked due to interference by a salt or a solvent.
Abbreviations in the Nuclear Magnetic Resonance (NMR) data in the following examples represent the following meanings:
s: singlet, d: doublet, t: triplet, q: quartet, dd: doublet, qd: quartet, ddd: double doublet, ddt: double triplet, dddd: double doublet, m: multiplet, br: broad peak, J: coupling constant, hz: hertz, δ: chemical shift.
All chemical shift (δ) values are given in parts per million (ppm).
Mass Spectrometry (MS) was performed using an Agilent 6120B mass spectrometer with an electrospray ion source (ESI).
HPLC was performed using an Agilent 1200DAD high pressure liquid chromatograph (Sunfirc C18, 150X 4.6mm,5 μm column) and a Waters 2695-2996 high pressure liquid chromatograph (Gimini C18, 150X 4.6mm,5 μm column).
The thin layer chromatography silica gel plate is Qingdao sea GF254 silica gel plate, the specification of the silica gel plate used by Thin Layer Chromatography (TLC) is 0.15mm-0.2mm, and the specification of the thin layer chromatography separation and purification product is 0.4mm-0.5 mm.
Column chromatography is carried out by using Qingdao sea silica gel of 200-300 meshes as carrier.
The progress of the reaction in the examples was monitored by Thin Layer Chromatography (TLC) using a system of developing reagents, A: dichloromethane and methanol systems; b: petroleum ether and ethyl acetate, the volume ratio of the solvent is adjusted according to the polarity of the compound.
The system of eluents for column chromatography and developing agents for thin layer chromatography used for purifying compounds include a: dichloromethane and methanol systems; b: the volume ratio of the solvent in the petroleum ether and ethyl acetate system is adjusted according to different polarities of the compounds, and a small amount of triethylamine, an acidic or basic reagent and the like can be added for adjustment.
Synthesis of Compounds
Synthesis example 1: synthesis of intermediate M1 a:
step a: ia (2.5g, 10.4 mmol), N-bromosuccinimide (5.56g, 32.1mmol) and azobisisobutyronitrile (854mg, 5.21mmol) were added to carbon tetrachloride (40 mL), heated to 80 ℃ and stirred overnight. After the reaction is finished, the crude product is obtained by direct concentration. Then, compound M1a (3.3 g, yield 99%) was obtained as a white solid by column chromatography.
1 H NMR (400 MHz, DMSO-d6) δ 7.86 (s, 2H), 4.72 (s, 2H)。
Synthesis example 2: synthesis of intermediate M1 b:
step b: ib (5 g,15.2 mmol), N-bromosuccinimide (8.1g, 45.7 mmol) and azobisisobutyronitrile (1.5 g, 9.1mmol) were added to carbon tetrachloride (40 mL) and heated to 80 ℃ and stirred overnight. After the reaction is finished, the crude product is obtained by direct concentration. Then, compound M1b (5.3 g, yield 85.5%) was obtained as a white solid by column chromatography.
Example 1: synthesis of Compound 1
Step b: compound IIa (155mg, 1.25mmol) and potassium carbonate (3)46mg, 2.51mmol) was added to N, N-dimethylformamide (5 mL), and then M1a (400mg, 1.25mmol) was added to the reaction solution and stirred at room temperature for one hour. After completion of the reaction, ethyl acetate (100 mL) was added, the organic phases were washed with saturated brine, and the combined organic phases were concentrated to give a crude product. Then, compound IIIa (420 mg, yield 92.5%) was obtained as a yellow solid by column chromatography. MS (ESI, m/z): 361[ 2 ], [ M ] +H] + 。
Step c: compounds IIIa (100mg, 0.28mmol) and IVa (31mg, 0.28mmol) were dissolved in N, N-dimethylformamide (3 mL), followed by addition of cuprous iodide (52mg, 0.28mmol), potassium phosphate (117mg, 0.55mmol) and N, N' -dimethylethylenediamine (24mg, 0.28mmol), heating to 120 ℃ under nitrogen for two hours, cooling the reaction to room temperature, and isolation directly by column chromatography gave compound 1 as a white solid (4.8 mg, 4.4% yield). MS (ESI, m/z): 394[ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.76 (s, 2H), 7.68 (s, 1H), 7.13 (d,J= 7.6 Hz, 1H), 7.01 - 6.90 (m, 3H), 5.22 (s, 2H), 3.73 (s, 3H)。
Example 2: synthesis of Compound 2
Step d: compound IIb (154mg, 1.25mmol) and di-tert-butyl dicarbonate (545mg, 2.5mmol) were added to acetonitrile (15 mL), and then 4-dimethylaminopyridine (381mg, 3.13mmol) was added to the reaction solution and stirred at room temperature for one hour. After completion of the reaction, ethyl acetate (100 mL) was added, the organic phases were washed with saturated brine, and the combined organic phases were concentrated to give a crude product. Then, compound IIc (210 mg, yield 75.3%) was obtained as a white solid by column chromatography. MS (ESI, m/z): 224[ 2 ] M + H] + 。
And b and c: the synthesis method of reference example 1 gave compound va as a white solid (30 mg, yield 56.6%). MS (ESI, m/z): 493[ M ] C + H] + 。
Step e: compound Va (30mg, 0.061mmol) is added to the dichloro-benzeneTo methane (5 mL), a hydrochloric acid-dioxane solution (4 mL) was added to the reaction solution and stirred at room temperature for four hours. After completion of the reaction, the crude product was directly concentrated and then isolated by column chromatography to obtain compound 2 (18.3 mg, yield 76.3%) as a yellow solid. MS (ESI, m/z): 2, 2M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.48 (s, 1H), 7.75 (s, 2H), 7.71 (s, 1H), 6.99 (d,J= 8.0 Hz, 1H), 6.84 (t,J= 7.6 Hz, 1H), 6.56 (d,J= 7.6 Hz, 1H), 6.48 (d,J= 8.0 Hz, 1H), 4.79 - 4.70 (m, 1H), 5.21 (s, 2H), 2.66 (s, 3H)。
Example 3: synthesis of Compound 3
After substituting the starting material IIa for IId, compound 3 (700 mg, yield 68.0%) as a white solid was obtained according to the synthesis method of example 1, MS (ESI, m/z): 410[ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.50 (s, 1H), 7.77 (s, 2H), 7.73 (s, 1H), 7.20 - 7.16 (m, 3H), 7.05 - 7.01 (m, 1H), 5.28 (s, 2H), 2.31 (s, 3H)。
Example 4: synthesis of Compound 4
Step f: compound 3 (300mg, 0.733mmol) and m-chloroperoxybenzoic acid (91mg, 0.53mmol) were added to 1, 2-dichloroethane (5 mL) and stirred at room temperature for one day. After the reaction is finished, the crude product is obtained by direct concentration. Then, compound 4 (10 mg, yield 3.2%) as a white solid was isolated by column chromatography, MS (ESI, m/z): 426[ deg. ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.80 (s, 2H), 7.73 (s, 1H), 7.67 (d,J= 7.6 Hz, 1H), 7.59 (t,J= 8.0 Hz, 1H), 7.42 (d,J= 8.0 Hz, 1H), 7.29 (s, 1H), 5.36 (s, 2H), 2.64 (s, 3H)。
Example 5: synthesis of Compound 5
Step f: compound 3 (200mg, 0.49mmol) and m-chloroperoxybenzoic acid (337mg, 1.96mmol) were added to a mixed solution of 1, 2-dichloroethane (10 mL) and dichloromethane (5 mL) and stirred at room temperature for two hours. After the reaction was complete, the crude product was obtained by filtration. Then, compound 5 (90 mg, yield 42.0%) was obtained as a white solid by column chromatography, MS (ESI, m/z): 442[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.51 (s, 1H), 7.91 - 7.68 (m, 5H), 7.57 (d,J= 8.4 Hz, 1H), 7.24 (t,J= 7.6 Hz, 1H), 5.44 (s, 2H), 3.13 (s, 3H)。
Example 6: synthesis of Compound 6
The synthetic route is as follows:
after substituting the starting material IIa for IIe, compound 6 (120 mg, yield 21.3%) was obtained as a yellow solid by referring to the synthesis method of example 1, MS (ESI, m/z): 406[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.80 (s, 2H), 7.67 (s, 1H), 7.60 (d,J= 7.6 Hz, 2H), 7.43 (d,J= 8.0 Hz, 1H), 7.10 (t,J= 7.6 Hz, 1H), 5.38 (s, 2H), 2.36 (s, 3H)。
Example 7: synthesis of Compound 7
Replacing raw material IIa with IIf, and then adding ginsengCompound 7 (5 mg, yield 16.7%) was obtained as a white solid according to the synthetic method of example 1, MS (ESI, m/z): 420[ alpha ], [ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.79 (s, 2H), 7.66 (s, 1H), 7.56 (t,J= 7.6 Hz, 1H), 7.52 (d,J= 8.0 Hz, 1H), 7.40 (d,J= 8.4 Hz, 1H), 7.08 (t,J= 7.6 Hz, 1H), 5.35 (s, 2H), 2.73 (q,J= 7.2 Hz, 2H), 0.87 (t,J= 7.2 Hz, 3H)。
Example 8: synthesis of Compound 8
After substituting the starting material IIa for IIg, the synthesis method of example 1 was referenced to give compound 8 (7 mg, yield 22.6%) as a white solid, MS (ESI, m/z): 434[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.80 (s, 2H), 7.59 - 7.55 (m, 2H), 7.50 (d,J= 7.6 Hz, 1H), 7.39 (d,J= 8.0 Hz, 1H), 7.07 (t,J= 7.2 Hz, 1H), 5.35 (s, 2H), 2.67 (t,J= 7.2 Hz, 2H), 1.40 (q,J= 7.2 Hz, 2H), 0.61 (t,J= 7.2 Hz, 3H)。
Example 9: synthesis of Compound 9
The synthesis route is as follows:
after substituting the starting material IIa for IIh, the synthesis method of example 1 was referenced to give compound 9 (10 mg, yield 32.3%) as a white solid, MS (ESI, m/z): 432[ C ], [ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.77 (s, 2H), 7.65 (s, 1H), 7.57 (t,J= 7.6 Hz, 1H), 7.41 (d,J= 8.0 Hz, 2H), 7.08 (t,J= 7.6 Hz, 1H), 5.37 (s, 2H), 2.56 - 2.52 (m, 1H), 0.88 - 0.84 (m, 2H), 0.68 - 0.64 (m, 2H)。
Example 10: synthesis of Compound 10
Step g: compound II i (1g, 7.35mmol) was added to tetrahydrofuran (10 mL) then at 0 o Isopropyl magnesium bromide (8 mL, 8.09mmol) was added to the reaction solution at 0 deg.C o Stirring for one hour under C. After the reaction was completed, a saturated ammonium chloride solution was added to quench the reaction system, then ethyl acetate (100 mL) was added, the organic phases were washed with a saturated saline solution, and the combined organic phases were concentrated to obtain a crude product. Then, the compound IIj was isolated by column chromatography as a colorless oil (634 mg, yield 48.0%). MS (ESI, m/z): 181[ 2 ] M + H] + 。
Step h: compound IIj (634mg, 3.52mmol) was added to dichloromethane (10 mL) and then dess-martin oxidant (1.79g, 4.23mmol) was added to the reaction and stirred at room temperature for one hour. After the reaction was completed, the reaction system was quenched by adding a saturated sodium bicarbonate solution, then dichloromethane (100 mL) was added, the organic phases were washed with a saturated saline solution, and the combined organic phases were concentrated to give a crude product. Then, the compound IIk was isolated by column chromatography to give a colorless oily compound (483 mg, yield 77.0%). MS (ESI, m/z): 179[ M ] C + H] + 。
Step i: compound IIk (483mg, 2.71mmol) was added to dichloromethane (10 mL) then at 0 o Boron tribromide (3.3mL, 3.3mmol) was added to the reaction at 0 deg.C o Stirring for one hour under C. After the reaction is finished, methanol is added to quench the reaction system, and the crude product is obtained by direct concentration. Then, compound II l (504 mg, yield 99%) was obtained as a yellow oily substance by column chromatography. MS (ESI, m/z): 165 2[ M + H ]] + 。
After replacing the starting material IIa by IIl, the synthesis method of example 1 was referenced to give compound 10 (210 mg, yield 33.9%) as a white solid, MS (ESI, m/z): 434[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.47 (s, 1H), 7.79 (s, 2H), 7.71 (s, 1H), 7.56 (t,J= 7.6 Hz, 1H), 7.40 (d,J= 8.0 Hz, 2H), 7.08 (t,J= 7.6 Hz, 1H), 5.36 (s, 2H), 3.26 - 3.20 (m, 1H), 0.88 (d,J= 6.8 Hz, 6H)。
Example 11: synthesis of Compound 11
The synthetic route is as follows:
step j: compound 6 (60mg, 0.148mmol) was added to methanol (1 mL) and tetrahydrofuran (5 mL), then sodium borohydride (28mg, 0.741mmol) was slowly added to the reaction solution and stirred for one hour under ice bath, after the reaction was completed, saturated ammonium chloride solution was added to quench the excess reducing agent, then ethyl acetate (100 mL) was added, the organic phases were washed with saturated saline, and the combined organic phases were concentrated to give a crude product. Then, compound 11 (11.1 mg, yield 18.2%) was obtained as a white solid by column chromatography. MS (ESI, m/z): 408[ 2 ], [ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.78 (s, 2H), 7.68 (s, 1H), 7.46 (d,J= 7.6 Hz, 1H), 7.25 (t,J= 7.6 Hz, 1H), 7.17 (d,J= 8.4 Hz, 1H), 7.00 (t,J= 7.6 Hz, 1H), 5.24 (s, 2H), 4.95 (d,J= 4.4 Hz, 1H), 4.91 - 4.88 (m, 1H), 1.19 (d,J= 6.0 Hz, 3H)。
Example 12: synthesis of Compound 12
Reference to the synthesis of example 11 gave compound 12 (28 mg, yield 41.2%) as a white solid, MS (ESI, m/z): 422[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.78 (s, 2H), 7.42 (d,J= 7.2 Hz, 1H), 7.24 (t,J= 7.6 Hz, 1H), 7.18 (d,J= 8.0 Hz, 2H), 6.99 (t,J= 7.2 Hz, 1H), 5.24 (s, 2H), 4.87 (d,J= 4.4 Hz, 1H), 4.75 - 4.71 (m, 1H), 1.60 - 1.51 (m, 1H), 1.47 - 1.40 (m, 1H), 0.73 (t,J= 7.2 Hz, 3H)。
Example 13: synthesis of Compound 13
Reference to the synthesis of example 11 gave compound 13 (36 mg, yield 35.6%) as a white solid, MS (ESI, m/z): 436[ 2 ], M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.78 (s, 2H), 7.72 (s, 1H), 7.38 (d,J= 7.6 Hz, 1H), 7.24 (t,J= 7.2 Hz, 1H), 7.18 (d,J= 7.6 Hz, 1H), 6.99 (t,J= 7.2 Hz, 1H), 5.22 (s, 2H), 4.82 (d,J= 4.8 Hz, 1H), 4.60 (t,J= 4.8 Hz, 1H), 1.78 - 1.74 (m, 1H), 0.72 (d,J= 6.4 Hz, 6H)。
Example 14: synthesis of Compound 14
Step k: compound IIId (365mg, 0.98mmol) was added to a mixed solution of tetrahydrofuran (5 mL) and methanol (5 mL) and then added at 0 o Sodium borohydride (41mg, 1.07mmol) was added slowly at C and stirred at room temperature for thirty minutes. After the reaction was completed, the system was quenched by addition of saturated ammonium chloride solution, ethyl acetate (200 mL) was added, the organic phases were washed with saturated brine, and the combined organic phases were concentrated to give Iii (300 mg, yield 81.7%) as a yellow solid.
Step l: compound IIIi (115mg, 0.31mmol) was added to N, N-dimethylformamide (5 mL) and then at 0 o Sodium hydride (18mg, 0.46mmol) was added slowly under C and stirred at room temperature for thirty minutes. Methyl iodide (52mg, 0.37mmol) was added to the reaction mixture at 80 deg.C o C stirring overnight, and reactingAfter completion of the reaction, ethyl acetate (100 mL) was added, the organic phase was washed with saturated brine, and the combined organic phases were concentrated to give the crude product. Then, it was isolated by column chromatography to give IIIj (47 mg, yield 39.5%) as a yellow oil.
Step c: the synthesis procedure of reference example 1 gave compound 14 (5.7 mg, yield 11.2%) as a yellow solid, MS (ESI, m/z): 422[ 2 ], [ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.78 (s, 2H), 7.71 (s, 1H), 7.31 - 7.28 (m, 2H), 7.24 (d,J= 7.6 Hz, 1H), 7.04 (t,J= 7.6 Hz, 1H), 5.26 (s, 2H), 4.54 (q,J= 6.4 Hz, 1H), 3.08 (s, 3H), 1.21 (d,J= 6.4 Hz, 3H)。
Example 15: synthesis of Compound 15
Step m: compound IIe (1.17g, 8.60mmol) was added to tetrahydrofuran (15 mL) then at 0 o Methyllithium (8mL, 12.9mmol) was added slowly under C and stirred at room temperature overnight. After the reaction was completed, a saturated ammonium chloride solution was added to quench the system, ethyl acetate (200 mL) was added, the organic phases were washed with a saturated saline solution, and the combined organic phases were concentrated to give a crude product. Then, it was isolated by column chromatography to give IIm (1.4 g, yield 99%) as a yellow oil.
1 H NMR (400 MHz, DMSO-d6) δ 9.75 (s, 1H), 7.25 (d,J= 7.6 Hz, 1H), 7.05 (t,J= 7.6 Hz, 1H), 6.76 - 6.71 (m, 2H), 5.86 (s, 1H), 1.51 (s, 6H)。
Step b and step c: reference to the synthetic procedure of example 1 gave compound 15 (34.3 mg, yield 21.2%) as a yellow solid, MS (ESI, m/z): 422[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.45 (s, 1H), 7.78 (s, 2H), 7.72 (s, 1H), 7.60 (dd,J= 7.6 Hz, 1.2 Hz, 1H), 7.24 (t,J= 8.4 Hz, 1H), 7.17 (d,J= 7.6 Hz, 1H), 6.95 (t,J= 7.6 Hz, 1H), 5.23 (s, 2H), 4.88 (s, 1H), 1.35 (s, 6H)。
Example 16: synthesis of Compound 16
After substituting the starting material IIa for IIn, the synthesis method of example 1 was referenced to give 16 (4.8 mg, yield 26.7%) as a white solid, MS (ESI, m/z): 421[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.47 (s, 1H), 8.92 (s, 1H), 7.90 (d,J= 7.2 Hz, 1H), 7.76 (s, 2H), 7.70 (s, 1H), 7.27 (d,J= 7.2 Hz, 1H), 7.10 (t,J= 7.6 Hz, 1H), 6.97 (t,J= 7.6 Hz, 1H), 5.28 (s, 2H), 2.02 (s, 3H)。
Example 17: synthesis of Compound 17
After substituting the starting material M1a for M1b and IIa for IIe, the synthesis method of example 1 was referenced to give compound 17 (19.2 mg, yield 18.8%) as a white solid, MS (ESI, M/z): 496 2[ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 8.00 (s, 2H), 7.64 - 7.57 (m, 2H), 7.48 (s, 1H), 7.42 (d,J= 8.8 Hz, 1H), 7.09 (t,J= 7.2 Hz, 1H), 5.40 (s, 2H), 2.37 (s, 3H)。
Example 18: synthesis of Compound 18
The synthetic route is as follows:
step b: compound IIo (178mg, 1.61mmol) and potassium carbonate (334mg, 2.42mmol) were added to N, N-dimethylformamide (10 mL), followed by addition of M1a (C), (D), (E) and510mg, 1.61mmol) was added to the reaction solution and stirred at room temperature for one hour. After the reaction was completed, ethyl acetate (100 mL) was added, the organic phase was washed with saturated brine, and the combined organic phases were concentrated to give a crude product. Then, compound IIIn (360 mg, yield 64.1%) was obtained as a yellow solid by column chromatography. MS (ESI, m/z): 347[ 2 ] M + H] + 。
Step n: compound IIIn (360mg, 1.04mmol), compound VIa (327mg, 1.14mmol) and caesium carbonate (509mg, 1.56mmol) were added to N, N-dimethylformamide (10 mL) and the mixture was washed with brine o Stir overnight at C. After completion of the reaction, ethyl acetate (100 mL) was added, the organic phases were washed with saturated brine, and the combined organic phases were concentrated to give a crude product. Then, compound IIIo (180 mg, yield 37.4%) was obtained as a yellow solid by column chromatography. MS (ESI, m/z): 461[ 2 ] M + H] + 。
Step c: the compounds IIIo (180mg, 0.39mmol) and IVa (48mg, 0.43mmol) were dissolved in a solution of N, N-dimethylformamide (10 mL), and then cuprous iodide (74mg, 0.39mmol), potassium phosphate (165mg, 0.78mmol) and N, N' -dimethylethylenediamine (34mg, 0.39mmol) were added, heated to 120 ℃ under nitrogen protection for three hours, the reaction solution was cooled to room temperature, and directly isolated by column chromatography to give a white solid compound VIIa (77 mg, yield 40.1%). MS (ESI, m/z): 494[ M ] +H] + 。
Step o: compound VIIa (77mg, 0.16mmol) was added to a mixed solution of acetic acid (4 mL) and water (2 mL), heated to 70 ℃ for reaction for one hour, the reaction mixture was cooled to room temperature, and compound 18 (43.3 mg, yield 61.2%) was obtained as a white solid by direct column chromatography. MS (ESI, m/z): 454[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.75 (s, 2H), 7.71 (s, 1H), 7.09 (d,J= 7.6 Hz, 1H), 7.03 (d,J= 8.0 Hz, 1H), 6.97 (t,J= 7.6 Hz, 1H), 6.90 (t,J= 7.6 Hz, 1H), 5.25 (s, 2H), 4.85 (s, 1H), 4.58 (s, 1H), 3.99 - 3.95 (m, 1H), 3.90 - 3.86 (m, 1H), 3.81 - 3.75 (m, 1H), 3.45 - 3.41 (m, 2H)。
Example 19: synthesis of Compound 19
Step p: compound IVa (2g, 17.7mmol) and sodium difluoromethylsulfinate (4.9g, 35.4mmol) were added to dimethyl sulfoxide (80 mL), and acid Red 94 (360mg, 0.35mmol) was added to the reaction solution and stirred at green for ten hours at room temperature. After completion of the reaction, ethyl acetate (500 mL) was added, the organic phases were washed with saturated brine, and the combined organic phases were concentrated to give a crude product. Then, by column chromatography, IVb (680 mg, yield 23.6%) as a red oily substance, MS (ESI, m/z): 164[ 2 ] M + H] + 。
The starting material IVa was replaced with IVb, and then, referring to the synthesis method of example 1, compound 19 (100 mg, yield 24.2%) was obtained as a yellow solid, MS (ESI, m/z): 460[ 2 ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.64 (s, 2H), 7.18 - 7.10 (m, 2H), 7.05 - 6.95 (m, 2H), 6.60 (t,J= 52.4 Hz, 1H), 5.31 (s, 2H), 2.35 (s, 3H)。
Example 20: synthesis of Compound 20
The synthesis method according to example 4 gave compound 20 (20 mg, yield 40.0%) as a yellow solid, MS (ESI, m/z): 476[ n ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.80 (s, 2H), 7.68 (d,J= 7.6 Hz, 1H), 7.59 (t,J= 8.0 Hz, 1H), 7.43 (d,J= 8.0 Hz, 1H), 7.30 (d,J= 7.6 Hz, 1H), 6.92 (t,J= 52.4 Hz, 1H), 5.37 (s, 2H), 2.64 (s, 3H)。
Example 21: synthesis of Compound 21
The synthesis method of reference example 1 gave compound 21 (38.3 mg, yield 2) as a yellow solid7.4%)，MS(ESI, m/z)：456[M+H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 12.86 (s, 1H), 7.80 (s, 2H), 7.61 (d,J= 7.2 Hz, 2H), 7.44 (d, J = 8.4 Hz, 1H), 7.11 (t,J= 7.2 Hz, 1H), 6.92 (t,J= 52.4 Hz, 1H), 5.40 (s, 2H), 2.37 (s, 3H)。
Example 22: synthesis of Compound 22
The synthetic route is as follows:
the synthesis method of reference example 1 gave compound 22 (80 mg, yield 24.0%) as a white solid, MS (ESI, m/z): 470[ deg. ] M + H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.79 (s, 2H), 7.58 (t,J= 8.0 Hz, 1H), 7.53 (d,J= 8.0 Hz, 1H), 7.41 (d,J= 7.6 Hz, 1H), 7.09 (t,J= 7.6 Hz, 1H), 6.92 (t,J= 52.4 Hz, 1H), 5.37 (s, 2H), 2.74 (q,J= 6.8 Hz, 2H), 0.88 (t,J= 6.8 Hz, 3H)。
Example 23: synthesis of Compound 23
Reference to the synthetic method of example 11 gave compound 23 (28 mg, yield 41.2%) as a white solid, MS (ESI, m/z): 472[ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.78 (s, 2H), 7.42 (d,J= 7.6 Hz, 1H), 7.25 (t,J= 8.0 Hz, 1H), 7.18 (d,J= 7.2 Hz, 1H), 7.00 (t,J= 7.2 Hz, 1H), 6.96 (t,J= 52.4 Hz, 1H), 5.25 (s, 2H), 4.88 (d,J= 4.8 Hz, 1H), 4.77 - 4.70 (m, 1H), 1.60 - 1.50 (m, 1H), 1.49 - 1.39 (m, 1H),0.73 (t,J= 6.8 Hz, 3H)。
Example 24: synthesis of Compound 24
The synthesis method of reference example 11 gave compound 24 (20 mg, yield 16.9%) as a white solid, MS (ESI, m/z): 546[ 2 ], [ M ] +H] + 。
1 H NMR (400 MHz, DMSO-d6) δ 7.98 (s, 2H), 7.65 - 7.58 (m, 1H), 7.43 (d,J= 8.8 Hz, 1H), 7.11 (t,J= 7.6 Hz, 1H), 6.96 (t,J= 52.4 Hz, 1H), 5.43 (s, 2H), 4.88 (d,J= 4.8 Hz, 1H), 2.37 (s, 3H)。
Biological activity assay
Experimental example 1: method for detecting THR alpha and THR beta agonistic activity of compound based on reporter gene activity detection
1. Method for producing a composite material
1.1 construction and preparation of plasmids pGAL4-THR-LBD and pG5-Luc
pGAL4-THR alpha-LBD and pGAL4-THR beta-LBD plasmids used by the reporter gene detection system are constructed according to a conventional molecular cloning method. The method mainly comprises the following steps: inserting cDNA sequences of THR alpha (NM _ 003250) and THR beta (NM _ 000461) corresponding to the amino acid sequences of THR alpha (163-407 AA) and THR beta (217-461 AA) into BamHI and NotI enzyme cutting sites of a pGAL4 vector by utilizing a PCR technology to obtain pGAL4-THR alpha-LBD and pGAL4-THR beta-LBD plasmids; pG5-Luc (# E249A) and pRL-TK (# E2241) plasmids were purchased from Promega; using CaCl 2 The method transforms the plasmid into DH5 alpha colibacillus, and after further culture and amplification, corresponding plasmid DNA is obtained by purification with a plasmid extraction kit (TIANGEN, # D107).
Plasmid co-transfection of HEK293T cells and compound treatment
HEK293T cells were transfected at 1X 10 the day before plasmid transfection 4 Density per well was seeded in 96-well plates. Cells were transfected according to the instructions for the transfection reagent FuGENE HD (Promega, # E2311). The method mainly comprises the following steps: taking one well as an example, plasmid pGAL4-THR α -LBD or pGAL4-THR beta-LBD, pG5-Luc and pRL-TK were added to 10 uL of Opti-MEM I medium (Gibco, # 11058021) in the ratio of 20 ng, 50 ng and 5 ng and mixed; then adding 0.25 uL of FuGENE HD, uniformly mixing, and then standing at room temperature for 5 min; this 10 uL mixture was then added to the wells containing 100 uL of medium. After 6 h of cell cotransfection, the compound is diluted by dimethyl sulfoxide in 3-fold gradient with the highest concentration of 1 uM, and is added into a cell culture solution in 10 concentrations for treatment for 24 h, and the total of 2 multiple wells are divided, and triiodothyronine (T3) is used as a positive control.
Detection of
After 24 h of compound treatment, the cells were tested according to the instructions of Dual-Glo Luciferase Assay System (Promega, # E2940). The method mainly comprises the following steps: absorbing and discarding 50 uL of culture solution in each hole, then adding 50 uLDual-Glo Luciferase reagent, and oscillating for 10 min at room temperature; taking 80 uL of the cracking reaction solution to a white opaque optiPlate-96 pore plate, and detecting a luminescence signal value (Firefly-Luc) of Firefly luciferase (Firefly luciferase) by using an MD i3x multifunctional microplate reader; then adding 40 uLDual-Glo Stop and Glo reagents, and oscillating for 10 min at room temperature; the luminescence signal value (Renilla-Luc) of Renilla luciferase (Renilla luciferase) was detected using an MD i3x multifunctional microplate reader. The activation activity of the compounds on THR was normalized to the ratio of Firefly-Luc/Renilla-Luc and to the ratio of the solvent DMSO groups, and EC50 values were calculated using GraphPadPrism6.0 software to fit the dose-response curves with four parameters.
2. As a result, the
Resimuliprole (MGL-3196) is an oral, liver-targeted and highly selective THR- β agonist, and therefore, MGL-3196 is used herein as a control compound to demonstrate the biological activity of the compounds of the present application.
Experimental data show that the compound has stronger THR beta agonistic activity and certain THR alpha/beta selectivity. The specific data are shown in Table 1.
TABLE 1
Experimental example 2: method for detecting agonistic activity of compound on THR alpha/beta based on time-resolved fluorescence resonance energy transfer (THR-FRET)
1. Construction of THR alpha/beta overexpression vector
THR alpha/beta LBD domain (domain) sequences were found by consulting NCBI, pET21-His-GST-dLBT-THR alpha LBD and pET21-His-GST-dLBT-THR beta LBD overexpression vectors were constructed by the fusion (fusion) method, and sequencing confirmed the accuracy of the sequences.
Prokaryotic expression recombinant protein of escherichia coli
Correctly sequenced THR α LBD and THR β LBD overexpression vectors were transferred into e.coli cells BL21 (DE 3) and plated on agar plates with ampicillin resistance, and single clones were picked up for amplification in LB medium and mass-cultured in 1L of LB at a ratio of 1. When the OD was 0.8-1.2, 0.5mM isopropyl-beta-D-thiogalactopyranoside (IPTG) was added and induction was carried out overnight at 18 ℃. Collecting bacteria, crushing, and purifying by GST column and molecular sieve to obtain His-GST-dLBT-THR alpha LBD and His-GST-dLBT-THR beta LBD proteins. The raw Bradford protein quantification kit measured protein concentrations of 24. Mu.M and 23. Mu.M, respectively.
Compound preparation and reaction system preparation
The protein was removed from the-80 ℃ freezer, in which the protein with the GST-tagged THR α/β LBD domain and the Eu-labeled GST antibody were thawed slowly on ice, and a detection buffer containing Dithiothreitol (DTT) at a final concentration of 5mM was prepared.
Preparation of Compounds
The starting concentration of compound was 100 μ M in DMSO, and the compound (100 μ M in DMSO) was diluted 3-fold with an equal gradient of DMSO to give 11 equal gradients, which were then further diluted 50-fold with assay containing 5mM DTT.
Preparation of reaction System
The final concentrations of all components were calculated according to a system with a final volume of 20. Mu.L per well, and 18. Mu.L of protein, polypeptide and antibody reaction mixture per well was prepared by adding GST-tagged THR α/β protein, SRC2 (LKEKIHLHRLLQDSSSSPV) polypeptide, XL665 (Cisobio, #610 SAXLB) and Eu-labeled GST antibody to 18. Mu.L of detection buffer containing 5mM DTT at final concentrations of 2nM, 200nM, 0.05nM and 7.6nM, respectively.
mu.L of the reaction mixture and 2. Mu.L of the diluted compound were added to the optiplate-384 well plate and reacted at room temperature for 24 hours.
Reading board
And reading the plate by using an MD i3X multifunctional microplate reader, wherein the wavelengths of excitation light and emission light are 340nm and 665nm respectively. The method is characterized in that 616nm wavelength light intensity generated by exciting europium by using 340nm wavelength light of an MD i3X multifunctional microplate reader is used as a background, according to the fact that different compounds have different activation degrees on THR alpha and THR beta, the intensity of 665nm wavelength emission light generated by exciting XL665 nm wavelength light by using 616nm wavelength excitation light is different, the intensity ratio of the two wavelengths (665 nm and 616 nm) is used as the activation activity of the compounds on the THR alpha or THR beta, normalization treatment is carried out according to the ratio of a solvent DMSO group, a dose-reaction curve is fitted by using GraphPad Prism6.0 software according to four parameters, and the EC50 value is calculated.
Results
Experimental data show that the compound has stronger THR beta agonistic activity and certain THR alpha/beta selectivity. The specific data are shown in Table 2.
TABLE 2
Claims (6)
1. A compound having the structure of formula (1) or a pharmaceutically acceptable salt thereof:
wherein the content of the first and second substances,
R 1 selected from H or-CHF 2 ；
R 2 And R 3 Independently selected from Cl or Br;
R 4 is selected from-OCH 3 、-NHCH 3 、-NHC(＝O)CH 3 、-SCH 3 、-S(＝O)CH 3 、-S(＝O) 2 CH 3 、-C(＝O)CH 3 、
-C(＝O)CH 2 CH 3 、-C(＝O)CH 2 CH 2 CH 3 、
2. A compound, or a pharmaceutically acceptable salt thereof, selected from:
3. a pharmaceutical composition comprising a compound according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers.
4. Use of a compound according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 3, in the manufacture of a medicament for the prevention and/or treatment of a disease or condition mediated by the thyroid hormone β receptor.
5. The use according to claim 4, wherein the disease is a metabolic disease.
6. Use according to claim 5, said disease being selected from non-alcoholic fatty liver disease, dyslipidemia, atherosclerosis or hypothyroidism.
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