CN116178340B - PROTAC compound, preparation method and pharmaceutical composition - Google Patents

PROTAC compound, preparation method and pharmaceutical composition Download PDF

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CN116178340B
CN116178340B CN202310473755.2A CN202310473755A CN116178340B CN 116178340 B CN116178340 B CN 116178340B CN 202310473755 A CN202310473755 A CN 202310473755A CN 116178340 B CN116178340 B CN 116178340B
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compound
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CN116178340A (en
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孟帅
张霖
任秀宝
郝希山
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Cell Ecology Haihe Laboratory
Tianjin Medical University Cancer Institute and Hospital
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Cell Ecology Haihe Laboratory
Tianjin Medical University Cancer Institute and Hospital
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/07—Tetrapeptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/02—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing at least one abnormal peptide link
    • C07K5/021—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing at least one abnormal peptide link containing the structure -NH-(X)n-C(=0)-, n being 5 or 6; for n > 6, classification in C07K5/06 - C07K5/10, according to the moiety having normal peptide bonds
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Abstract

The invention relates to the technical field of medicine synthesis and application, and provides a PROTAC compound, a preparation method and a medicine composition, wherein the PROTAC compound comprises a pharmacologically or physiologically acceptable salt of the PROTAC compound, and the PROTAC compound is prepared by connecting a Sirt2 inhibitor and an E3 ligase ligand by using a linker, so that Sirt2 protein can be degraded, the growth of tumor cells is inhibited, a HOOK effect is not observed at a high concentration, and a novel method is provided for treating Sirt 2-mediated malignant tumors.

Description

PROTAC compound, preparation method and pharmaceutical composition
Technical Field
The invention relates to the technical field of medicine synthesis and application, in particular to a PROTAC compound, a preparation method and a medicine composition.
Background
Protein cleavage targeting chimeras (Proteolysistargeting chimeras, PROTACs) are a class of bifunctional small molecule compounds based on ubiquitin-proteasome systems that chemically induce polyubiquitination and degradation of target proteins. The structure contains two ligands: one end is a ligand which can recruit a protein degradation system (such as E3 ubiquitin ligase), the other end is a ligand which is combined with target protein in cells, and the two ligands are connected through a connecting chain (linker). After entering cells, the ligands at the two ends of the PROTACs are specifically combined with corresponding proteins, and the target proteins and E3 ubiquitin ligase in the cells are pulled up to form a 'target protein-PROTAC-E3' ternary complex, wherein the E3 ubiquitin ligase can mediate ubiquitin-conjugating enzyme E2 to the target proteins, after the ternary complex is dissociated, the dissociated PROTACs can be combined with the proteins again, and the target proteins marked by ubiquitin are recognized and degraded by the proteasome, so that the level of the target proteins in the cells is selectively reduced.
Sirtuin 2 (Sirt 2) is the only deacetylase protein in the Sirtuin family, whose chemical structure consists of 14 alpha helices and 9 beta sheets, forming a smaller zinc ion binding domain and Rossmann folding domain. At the same time, the "loop" linking these two domains forms the catalytically active site of the Sirt2 protein, including the substrate binding site and NAD + Binding sites. Sirt2 deacetylates histones and many non-histones, and in neoplastic diseases Sirt2 plays an important regulatory role in the development and progression of tumors, thus the development of inhibitors or degradants against Sirt2 provides a new approach to the treatment of malignant tumors.
The Hong JY et al synthesizes a degradation agent Sirt2-PROTAC aiming at a Sirt2 target spot, and in vitro screening (MCF-7 and BT549 cells) finds that the compound TM-P4-THAL can degrade Sirt2 and has an inhibition effect on tumor cells MCF-7 and MDA-MB-231. However, the compound concentration at 10-25 mu M produces a HOOK effect, which results in a gradual decrease in degradation rate (from 82% to 37%); and its killing effect on MCF-7 is about 75% at 20. Mu.M, the difference in killing effect on MCF-7 between TM and TM-P4-THAL becomes smaller with increasing concentration.
Therefore, the degradation agent Sirt2-PROTAC synthesized aiming at the Sirt2 target point can degrade Sirt2 and inhibit tumor cells, but the HOOK effect easily occurs, so that the degradation rate of Sirt2 is gradually reduced, and the killing effect on the tumor cells is gradually reduced.
Disclosure of Invention
The present invention is directed to solving at least one of the technical problems existing in the related art. Therefore, the invention provides a PROTAC compound, a preparation method and a pharmaceutical composition, wherein the PROTAC compound comprises a pharmacologically or physiologically acceptable salt of the PROTAC compound, and the pharmacologically or physiologically acceptable salt of the PROTAC compound is prepared by connecting a Sirt2 inhibitor and an E3 ligase ligand by a linker, so that the PROTAC compound not only can degrade Sirt2 protein, but also has an inhibition effect on the growth of tumor cells, and no HOOK effect is observed at high concentration, thereby providing a novel method for treating Sirt 2-mediated malignant tumors.
The invention provides a PROTAC compound, comprising a pharmacologically or physiologically acceptable salt of the PROTAC compound, wherein the formula I is the structural formula of the pharmacologically or physiologically acceptable salt of the PROTAC compound:
i is a kind of
Wherein X is O or NH;
is a Sirt2 inhibitor;
The E3 ligase ligand refers to a ligand molecule which binds to E3 ligase and Cereblon ligand, and the structural formula of the E3 ligase ligand comprises:
、/>or->One of the following;
linker is used to link the X and the E3 ligase ligand.
According to the PROTAC compound provided by the invention, the structural formula of the linker comprises:
、/>、/>、/>、or->One of them.
According to the present invention there is provided a PROTAC compound comprising, in pharmaceutically or physiologically acceptable salts:
、
、
、
、
、
、
、
、
、
、
、
or (b)
One of them.
The invention also provides a preparation method of the PROTAC compound, which comprises the following steps:
s1: preparation of N 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine;
s2: preparation of N from the product of step S1 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine;
s3: e3 ligase ligand reacts with linker to obtain intermediate compound;
s4: the Sirt2-PROTAC compound is prepared from the step S2 product and the step S3 product.
According to the preparation method of the PROTAC compound provided by the invention, the step S1 product is used for preparing N 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine comprising the steps of:
s21: the N is set to 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]Adding L-lysine into a methanol solution, stirring to obtain a first mixed solution, adding acetyl chloride into the methanol solution to obtain a second mixed solution, dripping the first mixed solution into the second mixed solution, stirring for 12h to obtain a first reaction solution, spinning and purifyingThe first reaction liquid is used for obtaining a first intermediate product;
s22: adding the first intermediate product and the Lawson reagent into tetrahydrofuran solution to obtain a third mixed solution, introducing inert gas flow into the third mixed solution, stirring for 12 hours to obtain a second reaction solution, and spin-drying and purifying the second reaction solution to obtain a second intermediate product;
s23: dissolving the second intermediate in THF/H 2 In O, a fourth mixed solution is obtained, lithium hydroxide is added into the fourth mixed solution, stirring is carried out, a third reaction solution is obtained, THF in the third reaction solution is dried in a spinning mode, a fourth reaction solution is obtained, HCl is added into the fourth reaction solution to adjust the pH value to 2-3, and the fourth reaction solution is filtered, so that N is obtained 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine.
According to the preparation method of the PROTAC compound provided by the invention, the E3 ligase ligand reacts with the linker to obtain an intermediate compound, which comprises the following steps:
S31: adding the E3 ligase ligand, DIPEA and the linker into NMP for reaction to obtain a fifth reaction solution, purifying the fifth reaction solution to obtain a first purified product, adding the first purified product into DCM/TFA, stirring to obtain a fifth mixed solution, spin-drying the fifth mixed solution to obtain a first oily product, adding anhydrous diethyl ether into the first oily product to precipitate solids to obtain a first filtrate, and filtering the first filtrate to obtain a first group of compounds;
s32: adding HATU, DIPEA and the linker into NMP, stirring to obtain a sixth mixed solution, adding the E3 ligase ligand into the sixth mixed solution, stirring for 12h to obtain a sixth reaction solution, extracting the sixth reaction solution with water and DCM to obtain a first organic layer solution, spin-drying and purifying the first organic layer solution to obtain a second purified product, adding the second purified product into DCM/TFA to obtain a seventh mixed solution, stirring, spin-drying the seventh mixed solution to obtain a second oily product, adding anhydrous diethyl ether into the second oily product to obtain a second filtrate, and filtering the second filtrate to obtain a second group of compounds;
S33: adding the E3 ligase ligand, DIPEA and the linker into DMF, stirring for 12 hours at 90 ℃ to obtain a seventh reaction solution, cooling, adding water and ethyl acetate into the seventh reaction solution for extraction to obtain a second organic layer solution, spin-drying and purifying the second organic layer solution to obtain a third intermediate product, adding the third intermediate product into DCM/TFA, stirring to obtain an eighth mixed solution, spin-drying the eighth mixed solution to obtain a third oily product, adding anhydrous diethyl ether into the third oily product, separating out solids to obtain a third filtrate, and filtering the third filtrate to obtain a first compound;
s34: adding the linker, HATU and DIPEA into DMF, stirring to obtain a ninth mixed solution, cooling the ninth mixed solution to 0 ℃, dropwise adding the E3 ligase ligand into the ninth mixed solution, stirring to obtain an eighth reaction solution, extracting the eighth reaction solution with ethyl acetate to obtain a third organic layer solution, spin-drying and purifying the third organic layer solution to obtain a fourth intermediate product, adding the fourth intermediate product into DCM/TFA, stirring to obtain a tenth mixed solution, spin-drying the tenth mixed solution to obtain a fourth oily product, adding anhydrous diethyl ether into the fourth oily product, separating out solids to obtain a fourth separated-out solution, and filtering the fourth separated-out solution to obtain a second compound.
According to the preparation method of the PROTAC compound provided by the invention, the E3 ligase ligand comprises the following components: lenalidomide, 2- (2, 6-dioxo-piperidin-3-yl) -4-fluoro-isoindole-1, 3-dione or (2S, 4 r) -1- ((S) -2-amino-3, 3-dimethylbutyryl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide hydrochloride.
According to the preparation method of the PROTAC compound provided by the invention, the linker comprises、/>One of 1-tert-butoxycarbonyl-1, 8-diaminooctane or N-tert-butoxycarbonyl-8-aminocaprylic acid.
According to the preparation method of the PROTAC compound provided by the invention, the method for preparing the Sirt2-PROTAC compound by using the product of the step S2 and the product of the step S3 comprises the following steps:
the N is set to 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-adding L-lysine, HATU and DIPEA to DMF, stirring to obtain an eleventh mixed solution, adding the first group of compounds, the second group of compounds, the first compound and the second compound to the eleventh mixed solution, stirring for 20h to obtain a ninth reaction solution, adding water and DCM to the ninth reaction solution, extracting to obtain a fourth organic layer solution, spin-drying and purifying the fourth organic layer solution to obtain the Sirt2-PROTAC compound.
The invention also provides a pharmaceutical composition comprising a PROTAC compound as described above, which pharmaceutical composition further comprises one or more combinations of pharmaceutically acceptable carriers, excipients, diluents, adjuvants, vehicles for the PROTAC compound.
The above technical solutions in the embodiments of the present invention have at least one of the following technical effects:
according to the invention, the ligand of the ubiquitination ligase E3 is introduced from different sites of the Sirt2 inhibitor, and the novel Sirt2-PROTAC compound is synthesized and screened through the change of different linker, compared with the existing PROTACs, the activity is higher, the Sirt2 is more degraded, the tumor cells are more killed, the HOOK effect is not observed under high concentration, the degradation effect of the Sirt2 is not influenced, and a novel method is provided for treating Sirt 2-mediated malignant tumors.
Additional aspects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention.
Drawings
In order to more clearly illustrate the invention or the technical solutions of the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described, and it is obvious that the drawings in the description below are some embodiments of the invention, and other drawings can be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C1 provided by the invention.
Fig. 2 is a nuclear magnetism hydrogen spectrum of the Sirt2-PROTAC compound C2 provided by the invention.
FIG. 3 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C3 provided by the invention.
Fig. 4 is a nuclear magnetic resonance hydrogen spectrum of the Sirt2-PROTAC compound C4 provided by the invention.
FIG. 5 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C5 provided by the invention.
FIG. 6 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C6 provided by the invention.
FIG. 7 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C7.
FIG. 8 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C8 provided by the invention.
FIG. 9 is a nuclear magnetic resonance hydrogen spectrum of the degradation agent Sirt2-PROTAC compound C9 synthesized by Hong JY et al aiming at the Sirt2 target.
Fig. 10 is a nuclear magnetic resonance hydrogen spectrum of the Sirt2-PROTAC compound C10 provided by the invention.
FIG. 11 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C11.
FIG. 12 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C12 provided by the invention.
FIG. 13 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C13.
Fig. 14 is a nuclear magnetic resonance hydrogen spectrum of Sirt2-PROTAC compound C14 provided by the present invention.
FIG. 15 is a graph showing the residual rate of the compounds C1-C12 in example 1 on tumor cells after Sirt2 degradation.
FIG. 16 is a graph showing the residual rate of Sirt 2-degraded tumor cells by the compounds C1-C12 in example 1.
FIG. 17 is a graph showing the comparative degradation effect of compound C8 and compound C9 on Sirt2 in example 1.
FIG. 18 is a graph showing the inhibitory activity of compounds C1-C7 on Sirt2 in example 2.
FIG. 19 is a graph showing the inhibitory activity of compounds C8-C14 on Sirt2 in example 2.
FIG. 20 is a graph showing the inhibitory activity of Compound C8 of example 3 on various tumor cell lines.
Detailed Description
For the purpose of making the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings, and it is apparent that the described embodiments are some embodiments of the present invention, not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention. The following examples are illustrative of the invention but are not intended to limit the scope of the invention.
In the description of the embodiments of the present invention, it should be noted that the terms "center", "longitudinal", "lateral", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", etc. indicate orientations or positional relationships based on the orientations or positional relationships shown in the drawings, are merely for convenience in describing the embodiments of the present invention and simplifying the description, and do not indicate or imply that the apparatus or elements referred to must have a specific orientation, be configured and operated in a specific orientation, and thus should not be construed as limiting the embodiments of the present invention. Furthermore, the terms "first," "second," and "third" are used for descriptive purposes only and are not to be construed as indicating or implying relative importance.
In the description of the present specification, a description referring to terms "one embodiment," "some embodiments," "examples," "specific examples," or "some examples," etc., means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the embodiments of the present invention. In this specification, schematic representations of the above terms are not necessarily directed to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, the different embodiments or examples described in this specification and the features of the different embodiments or examples may be combined and combined by those skilled in the art without contradiction.
The following describes a PROTAC compound, a preparation method and a pharmaceutical composition provided by the present invention with reference to fig. 1 to 20:
the invention provides a PROTAC compound, comprising a pharmacologically or physiologically acceptable salt of the PROTAC compound, wherein the formula I is the structural formula of the pharmacologically or physiologically acceptable salt of the PROTAC compound:
I is a kind of
Wherein X is O or NH;
is a Sirt2 inhibitor;
the E3 ligase ligand refers to a ligand molecule which binds to E3 ligase and Cereblon ligand, and the structural formula of the E3 ligase ligand comprises:
、/>or->One of the following;
linker is used to link the X and the E3 ligase ligand.
According to the PROTAC compound provided by the invention, the structural formula of the linker comprises:
、/>、/>、/>、/>or->One of them.
According to the present invention there is provided a PROTAC compound comprising, in pharmaceutically or physiologically acceptable salts:
、
、
、
、
、
、
、
、
、
、
、
or (b)
One of them.
The invention also provides a preparation method of the PROTAC compound, which comprises the following steps:
s1: preparation of N 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine;
s2: preparation of N from the product of step S1 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine;
s3: e3 ligase ligand reacts with linker to obtain intermediate compound;
s4: the Sirt2-PROTAC compound is prepared from the step S2 product and the step S3 product.
Wherein the preparation method comprises the steps of 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]The method for L-lysine is as follows: adding tetradecanoic acid into anhydrous DMF, andstirring to obtain a mixed solution, adding NHS and DCC into the mixed solution, reacting for 2 hours at room temperature, filtering to obtain a filtrate, adding the filtrate, Z-Lys-OH and DIPEA into DMF, stirring for 12 hours at room temperature to obtain a reaction solution, adding water and 1M HCl into the reaction solution to adjust the pH value to 2-3 to obtain an acidic mixed solution, extracting the acidic mixed solution with ethyl acetate for multiple times to obtain an organic layer solution, washing with water and spin-drying the organic layer solution to obtain N 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine.
Further, under the action of N-hydroxysuccinimide (NHS), N' -Dicyclohexylcarbodiimide (DCC) and N, N-Diisopropylethylamine (DIPEA), tetradecanoic acid and Z-Lys-OH) Condensation reaction is carried out to obtain N 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine of the formula +.>。
Further, washing with water and drying the organic layer solution means extracting the organic layer solution with saturated saline multiple times, collecting the organic layer solution extracted multiple times, adding anhydrous sodium sulfate to the collected organic layer solution, absorbing the residual moisture in the organic solution, and drying the organic layer solution.
Further, spin drying refers to spin drying ethyl acetate and DMF in the organic layer solution.
Wherein N prepared in step S2 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]L-lysine is an inhibitor of Sirt 2.
Wherein the intermediate compounds include a first group of compounds, a second group of compounds, a first compound, and a second compound.
According to the preparation method of the PROTAC compound provided by the invention, the step S1 product is used for preparing N 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine comprising the steps of:
s21: the N is set to 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]Adding L-lysine into a methanol solution, stirring to obtain a first mixed solution, adding acetyl chloride into the methanol solution to obtain a second mixed solution, dripping the first mixed solution into the second mixed solution, stirring for 12 hours to obtain a first reaction solution, spinning and purifying the first reaction solution to obtain a first intermediate product.
Wherein N is 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]Adding L-lysine into methanol solution, adding acetyl chloride for catalysis, stirring at room temperature for 12h, and performing esterification reaction to obtain a first reaction solution.
Further, spin-drying the first reaction solution refers to spin-drying the methanol and acetyl chloride solution in the first reaction solution.
Further, purification refers to purification by column chromatography of the product obtained after spin-drying the first reaction solution.
Further, the first intermediate product has the structural formula of。
S22: and adding the first intermediate product and the Lawson reagent into tetrahydrofuran solution to obtain a third mixed solution, introducing inert gas flow into the third mixed solution, stirring for 12 hours to obtain a second reaction solution, spin-drying and purifying the second reaction solution to obtain a second intermediate product.
Under the action of Lawson reagent, stirring at room temperature for 12h, and then performing oxygen-sulfur exchange on the first intermediate product to obtain a second intermediate product.
Further, the inert gas flow can be nitrogen, argon and the like, so that the reaction is under an anaerobic reaction condition, and the reaction is facilitated.
Further, spin-drying the second reaction solution refers to spin-drying the tetrahydrofuran solution in the second reaction solution.
Further, purification refers to column chromatography purification of the product obtained after spin-drying.
Further, the second intermediate product has the structural formula of。
S23: dissolving the second intermediate in THF/H 2 In O, a fourth mixed solution is obtained, lithium hydroxide is added into the fourth mixed solution, stirring is carried out, a third reaction solution is obtained, THF in the third reaction solution is dried in a spinning mode, a fourth reaction solution is obtained, HCl is added into the fourth reaction solution to adjust the pH value to 2-3, and the fourth reaction solution is filtered, so that N is obtained 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine.
Wherein, after adding lithium hydroxide, stirring for 3 hours at room temperature, the second intermediate product undergoes hydrolysis reaction to obtain N 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine.
Further, the pH-adjusted HCl was 1M HCl, THF/H 2 O is 1/1 of the mixed solution.
Further, N obtained after filtration 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]L-lysine as white solid of formula。
According to the preparation method of the PROTAC compound provided by the invention, the E3 ligase ligand reacts with the linker to obtain an intermediate compound, which comprises the following steps:
s31: adding the E3 ligase ligand, DIPEA and the linker into NMP for reaction to obtain a fifth reaction solution, purifying the fifth reaction solution to obtain a first purified product, adding the first purified product into DCM/TFA, stirring to obtain a fifth mixed solution, spin-drying the fifth mixed solution to obtain a first oily product, adding anhydrous diethyl ether into the first oily product to precipitate solids to obtain a first filtrate, and filtering the first filtrate to obtain a first group of compounds;
wherein the E3 ligase ligand is lenalinaDomide, the linker is。/>
Further, future nadir, DIPEA andNMP was added thereto, and the reaction was carried out at room temperature for 12 hours to obtain a fifth reaction solution.
Wherein the first purified product was added to DCM/TFA and stirred at room temperature for 1h to obtain a fifth mixed solution.
Further, purification refers to column chromatography separation and purification of the fifth reaction solution, and the first purified product refers to a product protected by Boc, and has the structural formula of。
Further, spin drying refers to spin drying NMP, DCM and TFA in the fifth mixed solution;
further, the first group of compounds are respectively、、/>And。
s32: adding HATU, DIPEA and the linker into NMP, stirring to obtain a sixth mixed solution, adding the E3 ligase ligand into the sixth mixed solution, stirring for 12h to obtain a sixth reaction solution, extracting the sixth reaction solution with water and DCM to obtain a first organic layer solution, spin-drying and purifying the first organic layer solution to obtain a second purified product, adding the second purified product into DCM/TFA to obtain a seventh mixed solution, stirring, spin-drying the seventh mixed solution to obtain a second oily product, adding anhydrous diethyl ether into the second oily product to obtain a second filtrate, and filtering the second filtrate to obtain a second group of compounds after solid precipitation.
Wherein the E3 ligase ligand is lenalidomide and the linker isIs a t-butoxycarbonyl protected linker.
Wherein DCM/TFA is 1/1 of the mixed solution.
Wherein HATU, DIPEA and To NMP was added and stirred at room temperature for 30min to obtain a sixth mixed solution.
Wherein lenalidomide is added into the sixth mixed solution, and stirring is carried out for 12 hours at room temperature, thus obtaining a sixth reaction solution.
Further, the first organic layer solution was dried over anhydrous sodium sulfate, and DCM, NMP and HATU remaining from the reaction were dried.
Further, purification refers to column chromatography purification, and the structural formula of the second purified product is。
Spin-drying the seventh mixed solution refers to spin-drying DCM and TFA in the seventh mixed solution.
Further, the structural formulas of the second group of compounds are respectively:、、/>、、/>、、/>and。
s33: adding the E3 ligase ligand, DIPEA and the linker into DMF, stirring at 90 ℃ for 12 hours to obtain a seventh reaction liquid, cooling, adding water and ethyl acetate into the seventh reaction liquid for extraction to obtain a second organic layer solution, spin-drying and purifying the second organic layer solution to obtain a third intermediate product, adding the third intermediate product into DCM/TFA, stirring to obtain an eighth mixed solution, spin-drying the eighth mixed solution to obtain a third oily product, adding anhydrous diethyl ether into the third oily product to obtain a third filtrate after solid precipitation, and filtering the third filtrate to obtain the first compound.
Wherein the E3 ligase ligand is 2- (2, 6-dioxo-piperidine-3-yl) -4-fluoro-isoindole-1, 3-dione, and the linker is 1-tert-butoxycarbonyl-1, 8-diaminooctane, and is Boc-protected linker.
The second organic layer solution is dried and purified by spinning, namely, the second organic layer solution is extracted by saturated saline water, the extracted organic layer solution is collected and dried by anhydrous sodium sulfate to absorb redundant water; further, spin drying refers to spin drying DMF and ethyl acetate in the organic layer solution; purification refers to the column chromatography purification of the spin-dried product.
The third intermediate product has the structural formula of。
Wherein the third intermediate was added to DCM/TFA and stirred at room temperature for 1h to obtain a seventh mixed solution.
Further, the DCM/TFA was 1/1 of the mixed solution.
The eighth mixed solution is spin-dried, namely, DCM and TFA in the ninth mixed solution are spin-dried.
Wherein the structural formula of the first compound is。
S34: adding the linker, HATU and DIPEA into DMF, stirring to obtain a ninth mixed solution, cooling the ninth mixed solution to 0 ℃, dropwise adding the E3 ligase ligand into the ninth mixed solution, stirring to obtain an eighth reaction solution, extracting the eighth reaction solution with ethyl acetate to obtain a third organic layer solution, spin-drying and purifying the third organic layer solution to obtain a fourth intermediate product, adding the fourth intermediate product into DCM/TFA, stirring to obtain a tenth mixed solution, spin-drying the tenth mixed solution to obtain a fourth oily product, adding anhydrous diethyl ether into the fourth oily product, separating out solids to obtain a fourth separated-out solution, and filtering the fourth separated-out solution to obtain a second compound.
Wherein the E3 ligase ligand is (2S, 4R) -1- ((S) -2-amino-3, 3-dimethylbutyryl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide hydrochloride, and the linker is N-tert-butoxycarbonyl-8-aminocaprylic acid and is Boc-protected linker.
Wherein N-t-butoxycarbonyl-8-aminocaprylic acid, HATU and DIPEA were added to DMF and stirred at room temperature for 30min to obtain a ninth mixed solution.
Wherein (2S, 4 r) -1- ((S) -2-amino-3, 3-dimethylbutyryl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide hydrochloride was added dropwise to the ninth mixed solution at 0 ℃ and then stirred at room temperature for 16 hours to obtain an eighth reaction solution.
Wherein, the third organic layer solution is washed with water and dried before spin-drying and purifying, saturated NaHCO 3 The third organic layer solution was extracted with saturated brine, and dried over anhydrous sodium sulfate.
Further, spin drying refers to spin drying ethyl acetate, DMF and unreacted HATU in the third organic layer solution.
Further, purification meansPurifying the product obtained after spin drying by column chromatography to obtain a fourth intermediate product with a structural formula of。
Wherein the fourth intermediate was added to DCM/TFA and stirred at room temperature for 1h to give a ninth mixed solution.
Further, spin drying the tenth mixed solution refers to spin drying DCM and TFA in the tenth mixed solution.
Further, the second compound has the structural formula。
According to the preparation method of the PROTAC compound provided by the invention, the E3 ligase ligand comprises the following components: lenalidomide, 2- (2, 6-dioxo-piperidin-3-yl) -4-fluoro-isoindole-1, 3-dione or (2S, 4 r) -1- ((S) -2-amino-3, 3-dimethylbutyryl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide hydrochloride.
According to the preparation method of the PROTAC compound provided by the invention, the linker comprises、/>One of 1-tert-butoxycarbonyl-1, 8-diaminooctane or N-tert-butoxycarbonyl-8-aminocaprylic acid.
According to the preparation method of the PROTAC compound provided by the invention, the method for preparing the Sirt2-PROTAC compound by using the product of the step S2 and the product of the step S3 comprises the following steps:
the N is set to 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]Adding L-lysine, HATU and DIPEA into DMF, stirring to obtain an eleventh mixed solution, adding the first group of compounds, the second group of compounds, the first compound and the second compound into the eleventh mixed solution, stirring for 20h to obtain a ninth reaction solution, and adding the ninth reaction solution into the ninth reaction solution Adding water and DCM, extracting to obtain a fourth organic layer solution, spin-drying and purifying the fourth organic layer solution to obtain the Sirt2-PROTAC compound.
Wherein N is 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]L-lysine, HATU and DIPEA were added to DMF and stirred at room temperature for 30min.
Wherein washing with water and drying the fourth organic layer solution means extracting the fourth organic layer solution with saturated saline to obtain an organic layer solution, and drying the organic layer solution with anhydrous sodium sulfate to absorb excessive moisture.
Further, spin-drying refers to spin-drying DCM, DMF and unreacted HATU in the organic layer solution.
Further, purification refers to column chromatography purification of the product obtained after spin-drying.
The invention also provides a pharmaceutical composition comprising a PROTAC compound as described above, which pharmaceutical composition further comprises one or more combinations of pharmaceutically acceptable carriers, excipients, diluents, adjuvants, vehicles for the PROTAC compound.
The medicine composition provided by the invention is applied to preparation of medicines for treating or preventing Sirt2 related diseases.
Further, the Sirt2 related disease is a tumor, specifically breast cancer, leukemia, lung cancer, liver cancer, esophageal cancer, pancreatic cancer, colorectal cancer, gastric cancer, cervical cancer, brain cancer, prostate cancer and the like.
In addition, the control compounds in examples 1 to 3 refer to degradation agents Sirt2-PROTAC synthesized by Hong JY and the like for the Sirt2 target, and the structural formula thereof is as follows:the nuclear magnetic hydrogen spectrum of the sample is shown as a figure 9, and the reference number is C9.
In examples 1 to 3, for convenience of description, the protoc compounds prepared in the present invention are respectively labeled as follows:
C1、
C2、
C3、
C4、
C5、
C6、/>
C7、
C8、
C10、
C11、
C12、
C13/>
and C14。
Nuclear magnetic hydrogen spectrograms of the compounds C1-C8 are shown in figures 1-8; the nuclear magnetic hydrogen spectra of the compounds C10 to C14 are shown in FIGS. 10 to 14.
Example 1 degradation of Sirt2 protein by the PROTAC compound:
in this example, sirt2 protein is mainly the MCF-7 protein of breast cancer cells.
Firstly, culturing MCF-7 cells (breast cancer cell line), performing digestion count, inoculating 100k cells in each well of a 6-well plate, and adding Sirt2-PROTAC compounds C1-C12, a control drug thiomyristyl (TM, a commercially purchased product) and DMSO into each well respectively until the final concentration of the compounds C1-C12 is 100 nM; DMSO alone and control Thiomyristoyl (TM) were added to control wells and DMSO alone was added to background wells in order to demonstrate whether DMSO had an effect on Sirt2 degradation. At 37 ℃,5% CO 2 After incubation of the cultured cells for 48h, removing the supernatant, extracting whole cell proteins from the cells in a protein lysis buffer with 1% Triton X-100,0.1% SDS and 0.1M Tris-HCl (pH 7.0), quantifying the proteins by BCA method, separating protein samples on SDS-PAGE gel, transferring to NC membrane (Millipore) after separation, blocking with 10% defatted dry milk at room temperature for 1 h, incubating the treated NC membrane with primary antibodies at 4 ℃, incubating the NC membrane incubated with primary antibodies with secondary antibodies for 1 h, and finally displaying signals by ECL detection reagent (Pierce). The exposed bands were analyzed using ImageJ software (NIH, bethesda, MD, USA) to calculate Sirt2 residue. The calculation formula is as follows:
Sirt2 residual Rate= (Sirt 2) Experimental group /β-actin Experimental group ）/（Sirt2 DMSO group /β-actin DMSO group ）x100%。
Wherein, the liquid crystal display device comprises a liquid crystal display device,
Sirt2 experimental group : gray values expressed by Sirt2 of each dosing group;
β-actin experimental group : gray values expressed by beta-actin of each dosing group;
Sirt2 DMSO group : gray values expressed by DMSO group Sirt 2;
β-actin DMSO group : gray scale values of beta-actin expression in DMSO group;
degradation rate = 100% -Sirt2 residual rate.
The ability of part of Sirt2-PROTAC compounds to degrade Sirt2 was evaluated and screened by Western immunoblotting (Western Blot), and the specific results are shown in Table 1:
TABLE 1 degradation of Sirt2 by Sirt2-PROTAC Compounds
As shown in fig. 15, the residual ratios of the compounds C1 to C12 to Sirt2 show that the residual ratio of the compound C8 is the lowest, and the degradation ratio of the compound C8 is the highest as shown in the above-described calculation formula of the degradation ratio.
Further, FIG. 16 is a graph showing the residual ratio of Sirt2 after Sirt2-PROTAC compounds C1-C12 degrade Sirt2, the experimental group normalized the gray scale of the exposed band of beta-actin, and then unified with Sirt2 DMSO group And beta-actin DMSO group And comparing to obtain Sirt2 residual rates of different dosing groups. The lower Sirt2 residual rate indicates a higher degradation of Sirt2 by the Sirt2-PROTAC compound.
As shown in FIG. 16, the residual rate of the control drug TM is not changed basically, which indicates that the control drug TM has no degradation effect on Sirt2 basically, and the compounds C1 to C12 synthesized by the method have different degrees of degradation on Sirt2, and the degradation rates of C1 to C12 are respectively 9 to 65 percent when the concentration of the compound is 100 nM, wherein the residual rate of the compound C8 is the lowest, which indicates that the degradation degree of the compound C8 on Sirt2 is the highest.
Wherein, the compound C9 is a degradation agent Sirt2-PROTAC synthesized by Hong JY and the like aiming at Sirt2 targets, and is used as a control compound of the embodiment. Further, as shown in FIG. 17, there is shown a graph showing experimental comparison of residual rates of Sirt2 after degradation of compound C8 and compound C9 at concentrations of 0.001uM, 0.01uM, 0.1uM, 1uM, 10uM and 20uM, respectively, wherein DMSO is used as a control, indicating that DMSO does not degrade Sirt2 and therefore does not affect the residual rate of Sirt 2. As can be seen from fig. 17, at the concentrations of 0.001uM, 0.01uM, 0.1uM, 1uM, 10uM and 20uM, the residual rate of compound C8 is significantly smaller than that of compound C9, which means that the degradation rate of compound C8 to Sirt2 is significantly better than that of compound C9, and at the concentrations of 0.001uM to 1uM, the residual rate of compound C9 is gradually decreased, but at the concentrations of 1uM to 20uM, the residual rate of compound C9 is gradually increased because the HOOK effect occurs to compound C9, which affects the residual rate of compound C9; in contrast, the residual rate of the compound C8 gradually decreased with the increase in concentration, and the HOOK effect did not occur.
Example 2 determination of antitumor Activity of Compounds C1 to C14:
MCF-7 cells in logarithmic growth phase were inoculated into 96-well plates (1X 104 cells/well), 10% fetal bovine serum medium (100. Mu.L) was added, and the 96-well plates were placed at 37℃and 5% CO 2 Is cultured in a cell culture box for 24 hours; 200. Mu.L of a drug-containing medium, i.e., diluted drug medium containing Sirt2-PROTAC compounds C1 to C14, was added to the test wells to give final drug concentrations of 40. Mu.M, 20. Mu.M, 10. Mu.M, 5. Mu.M, 2.5. Mu.M, 1.25. Mu.M, respectively, and 3 parallel test wells were set. 96-well plates were placed at 37℃with 5% CO 2 Culturing in incubator for 72 hr, sucking out the culture medium containing medicine in the medicine-containing well, washing with PBS once, removing PBS, adding 100 μl of culture medium containing 10% CCK-8 reagent into each well, standing at 37deg.C, and 5% CO 2 The cells were incubated in an incubator for 2 hours, and finally, the OD value (OD value means the optical density absorbed by the compounds C1 to C14) was measured at a wavelength of 450nM using an enzyme-labeled instrument. Numerical value processing: the OD value of each test well was subtracted from the control OD value (blank), and the OD value of each parallel experimental test well was taken as the average OD value and OD standard deviation. The antitumor activities of compounds C1 to C14 are shown in Table 2:
TABLE 2 antitumor Activity of Sirt2-PROTAC Compounds
Wherein the IC 50 At 50% inhibition concentration, i.e., the ratio of OD of the dosing cells to OD of the control cells is 50%The concentration at which half inhibition is measured is used to measure the antitumor activity of the Sirt2-PROTAC compound, the lower the half inhibition concentration, the better the antitumor activity of the Sirt2-PROTAC compound.
As can be seen from Table 2, compound C8 shows a better antitumor activity, IC 50 The value was 7.93. Mu.M.
Cell viability% = (dosed cell OD/control cell OD) ×100% and then plotted by GraphPad against compound concentration and corresponding viability to represent the antitumor activity of compounds C1-C14, as shown in fig. 18 and 19. FIG. 18 shows the survival rate of MCF-7 cells after degradation of tumor cells MCF-7 by compounds C1-C7; FIG. 19 shows the survival rate of MCF-7 cells after degradation of tumor cells MCF-7 by compounds C8-C14; as is evident from fig. 18 and 19, compound C8 showed the lowest survival rate of MCF-7 cells after degradation of MCF-7 cells, indicating that compound C8 had the best degradation efficiency on MCF-7 cells.
Example 3 determination of the Activity of Compound C8 against different tumor cell lines:
From examples 1 and 2, it was found that compound C8 exhibited a good antitumor activity against MCF-7 tumor cells, and thus, according to the experimental methods of the above examples, the antitumor activity of compound C8 against human non-small cell lung cancer cell line A549, liver cancer cell line HepG2, human renal cell adenocarcinoma cell line CRL1611, human colon cancer cell line SW480, human gastric cancer cell line MGC-803, cervical cancer cell line HELA, human pancreatic cancer cell line SW1990, human breast cancer cell line MCF-7 and human esophageal cancer cell line TE-2 was measured, respectively. The inhibitory activity of compound C8 at a concentration of 40uM against different tumor cell lines is shown in table 3:
TABLE 3 inhibitory Activity of C8 against different tumor cell lines
As can be seen from Table 3, compound C8 exhibited better antitumor activity against HepG2, CRL1611, MGC-803, HELA and MCF-7.
FIG. 20 shows the inhibitory activity of compound C8 at concentrations of 0uM, 5uM, 10uM, 20uM and 40uM, respectively, on different tumor cell lines, and it is apparent from FIG. 20 that the lower the cell viability of each tumor cell line with increasing concentration of compound C8, the better the inhibitory activity of compound C8 on each tumor cell line with increasing concentration of compound C8. Further, it can be seen from FIG. 20 that the cell viability of the compound C8 at 40uM was the lowest, indicating that the compound C8 had better antitumor activity against HepG2, CRL1611, MGC-803, HELA and MCF-7 at 40 uM.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and are not limiting; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims (11)

1. A PROTAC compound characterized by the structure:
、
、
、
、
、
、
、
、
、
、
、
or (b)
One of them.
2. A process for the preparation of a PROTAC compound as claimed in claim 1, comprising the steps of:
s1: preparing N6- (1-oxo-tetradecyl) -N2- [ (phenylmethoxy) carbonyl ] -L-lysine;
s2: preparing N6- (1-thiotetradecyl) -N2- [ (phenylmethoxy) carbonyl ] -L-lysine from the product of the step S1;
s3: reacting the E3 ligase ligand with a linker to obtain a first group of compounds, a second group of compounds,
And
；
s4: preparing Sirt2-PROTAC compound by using the product of the step S2 and the product of the step S3;
wherein the E3 ligase ligand is selected from one of lenalidomide, 2- (2, 6-dioxo-piperidin-3-yl) -4-fluoro-isoindole-1, 3-dione or (2S, 4R) -1- ((S) -2-amino-3, 3-dimethylbutyryl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide hydrochloride;
The linker is selected from、/>One of 1-tert-butoxycarbonyl-1, 8-diaminooctane or N-tert-butoxycarbonyl-8-aminocaprylic acid;
the first group of compounds is:
、
、
or (b)
One of the following;
the second group of compounds is:
、
、
、
、
、
、
or (b)
One of them.
3. The method for producing a PROTAC compound according to claim 2, wherein S2 comprises the steps of:
s21: the N is set to 6 - (1-oxo-tetradecyl) -N 2 - [ (benzyloxy) carbonyl group]Adding L-lysine into a methanol solution, stirring to obtain a first mixed solution, adding acetyl chloride into the methanol solution to obtain a second mixed solution, dripping the first mixed solution into the second mixed solution, stirring for 12h to obtain a first reaction solution, spinning and dryingPurifying the first reaction solution to obtain；
S22: will beAnd Lawson reagent is added into tetrahydrofuran solution to obtain a third mixed solution, inert gas flow is introduced into the third mixed solution, after stirring is carried out for 12 hours, a second reaction solution is obtained, and the second reaction solution is spin-dried and purified to obtain +.>；
S23: will beDissolved in THF/H 2 In O, a fourth mixed solution is obtained, lithium hydroxide is added into the fourth mixed solution, stirring is carried out, a third reaction solution is obtained, THF in the third reaction solution is dried in a spinning mode, a fourth reaction solution is obtained, HCl is added into the fourth reaction solution to adjust the pH value to 2-3, and the fourth reaction solution is filtered, so that N is obtained 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-L-lysine.
4. The method for producing a PROTAC compound according to claim 2, wherein S3 comprises the steps of:
s31: future nadir, DIPEA andadding NMP to react to obtain a fifth reaction solution, and purifying the fifth reaction solution to obtain +.>Will->Adding inStirring in DCM/TFA to obtain a fifth mixed solution, spin-drying the fifth mixed solution to obtain a first oily product, adding anhydrous diethyl ether into the first oily product, separating out solids to obtain a first filtrate, and filtering the first filtrate to obtain a first group of compounds;
s32, HATU, DIPEA andadding NMP, stirring to obtain a sixth mixed solution, adding lenalidomide into the sixth mixed solution, stirring for 12h to obtain a sixth reaction solution, extracting the sixth reaction solution with water and DCM to obtain a first organic layer solution, spin-drying and purifying the first organic layer solution to obtainWill->Adding the mixture into DCM/TFA to obtain a seventh mixed solution, stirring, spin-drying the seventh mixed solution to obtain a second oily product, adding anhydrous diethyl ether into the second oily product, separating out solids to obtain a second filtrate, and filtering the second filtrate to obtain a second group of compounds;
S33 adding 2- (2, 6-dioxo-piperidin-3-yl) -4-fluoro-isoindole-1, 3-dione, DIPEA and 1-tert-butoxycarbonyl-1, 8-diaminooctane into DMF, stirring at 90deg.C for 12h to obtain a seventh reaction solution, cooling, adding water and ethyl acetate to the seventh reaction solution, extracting to obtain a second organic layer solution, spin-drying and purifying the second organic layer solution to obtainWill->Adding the mixture into DCM/TFA, stirring to obtain an eighth mixed solution, spin-drying the eighth mixed solution to obtain a third oily product, and adding the third oily product into the eighth mixed solutionAdding anhydrous diethyl ether into the three oily products, separating out solid to obtain a third filtrate, and filtering the third filtrate to obtain +.>；
S34: adding N-tert-butoxycarbonyl-8-aminocaprylic acid, HATU and DIPEA into DMF, stirring to obtain a ninth mixed solution, cooling the ninth mixed solution to 0 ℃, dropwise adding (2S, 4R) -1- ((S) -2-amino-3, 3-dimethylbutyryl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide hydrochloride into the ninth mixed solution, stirring to obtain an eighth reaction solution, extracting the eighth reaction solution with ethyl acetate to obtain a third organic layer solution, spin-drying and purifying the third organic layer solution to obtain Will beAdding DCM/TFA, stirring to obtain tenth mixed solution, spin-drying the tenth mixed solution to obtain fourth oily product, adding anhydrous diethyl ether into the fourth oily product, separating out solid to obtain fourth filtrate, and filtering the fourth filtrate to obtain DOUQI>。
5. The method for producing a PROTAC compound according to claim 2, wherein S4 comprises the steps of:
the N is set to 6 - (1-thiotetradecyl) -N 2 - [ (benzyloxy) carbonyl group]-adding L-lysine, HATU and DIPEA to DMF, stirring to obtain an eleventh mixed solution, adding said first group of compounds, said second group of compounds, and,Andstirring for 20h to obtain a ninth reaction solution, adding water and DCM into the ninth reaction solution, extracting to obtain a fourth organic layer solution, spin-drying and purifying the fourth organic layer solution to obtain the Sirt2-PROTAC compound.
6. A pharmaceutical composition comprising the PROTAC compound of claim 1, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier for the PROTAC compound.
7. A pharmaceutical composition comprising the PROTAC compound of claim 1, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable excipient for the PROTAC compound.
8. A pharmaceutical composition comprising the PROTAC compound of claim 1, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable diluent for the PROTAC compound.
9. A pharmaceutical composition comprising the PROTAC compound of claim 1, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable vehicle for the PROTAC compound.
10. The pharmaceutical composition according to any one of claims 6-9, for use in the manufacture of a medicament for the treatment of Sirt 2-related diseases.
11. The pharmaceutical composition of claim 10, wherein the Sirt 2-related disorder is a tumor, the tumor being one of breast cancer, lung cancer, liver cancer, esophageal cancer, pancreatic cancer, colon cancer, stomach cancer, cervical cancer, or renal cell adenocarcinoma.
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Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103974934A (en) * 2011-10-07 2014-08-06 康奈尔大学 Methods of treatment using modulators of sirt2

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103974934A (en) * 2011-10-07 2014-08-06 康奈尔大学 Methods of treatment using modulators of sirt2

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
Phthalimide conjugations for the degradation of oncogenic PI3K;Li, Wenlu等;European Journal of Medicinal Chemistry;第151卷;237-247 *

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