CN106674209B - Programmed death receptor 1 gene inhibitor and preparation method and application thereof - Google Patents
Programmed death receptor 1 gene inhibitor and preparation method and application thereof Download PDFInfo
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Abstract
The invention provides a programmed death receptor 1 gene inhibitor and a preparation method and application thereof. In particular to a compound for inhibiting the expression of a programmed death receptor 1 and pharmaceutically acceptable salt thereof, which has a structure shown in a formula I
Wherein A is selected from C or N; r1Selected from H, CH3、(CH2)3‑N(CH3)2、(CH2)2‑N(CH3)2、CH2‑N(CH3)2、(CH2)3‑N(CH3)CH2CH2NHR3；R2Selected from H, CH3、
R3Selected from H, CH3、
The polyamide molecule disclosed by the invention belongs to polypeptide micromolecules, can realize chemical synthesis, is favorable for large-scale production, and provides more methods for inhibiting the function of PD 1.
Description
Technical Field
The invention relates to the field of medicines, and in particular relates to an expression inhibitor of a programmed death receptor 1.
Background
PD-1 (programmed death receptor 1) is an immune checkpoint protein, is a type I transmembrane protein, consists of 268 amino acids encoded by a PDCD1 gene, and is an important member in a CD28/CTLA4 family. The main function of PD-1 is to suppress the activity of T cells when an immune response occurs. The protein structure is divided into three parts: extracellular immunoglobulin variable regions, transmembrane domains, and intracellular domains; two motifs of ITSM (immunoreceptor type rosine-based switch motif) and ITIM (immunoreceptor type rosine-based inhibitor motif) in an intracellular domain, wherein the two motifs contain phosphorylation sites, and the ITSM motif is related to the inhibition function of PD-1. After binding of PD-1 molecules on the surface of T cells to PDL-1/PDL-2 receptors on the surface of antigen presenting cells, ITSM is phosphorylated, and then phosphatase SHP-1 and SHP-2 bind to ITSM, directly resulting in dephosphorylation of signal molecules downstream of the TCR complex, thereby inhibiting activation of T cells. In a tumor microenvironment, a large amount of PDL-1/PDL-2 molecules are expressed on the surface of tumor cells, when T cells infiltrate into tumor tissues, the PD-1 molecules on the surface of the T cells can recognize PDL-1/PDL-2, and the T cells cannot be activated by tumor antigens and cannot play a role in killing the tumor cells.
The basic research of the transcription control element of the PDCD1 gene promoter region provides a target point basis for the design of the patent. In 1997, Finger et al studied the structure of the human PDCD1 gene and predicted a number of cis-acting elements in the promoter region such as Sp1, PU.1box, AP-2, E-box MYC site, etc. Oestreich et al, 2008, performed on mouse EL4 cell line and mouse naive CD8+Two tissue-specific regulatory sites, namely CR-B (Conserved Region-B) and CR-C (Conserved Region-C), are found in a promoter Region of a PDCD1 gene in a T cell model, while an N1NFAT cis-Element site located in a CR-C Region is a regulatory Element necessary for transcription of PDCD 1. Cho et al in the same year found that an IFN- α Response Element ISRE (IFN-Sensitive Response Element) is involved in expression of PDCD1 in Response to IFN- α in a mouse macrophage line RAW264.7, and subsequently Terawaki et al in T cells also obtained similar results in 2011, and treatment with IFN- α, combined with blocking and tumor-bearing mice of PD-1, is effective in treating tumor-bearing miceThe expression of the PDCD1 gene is regulated by a series of cis-acting elements in the promoter region, and some elements exist in a range of about 10-50kb upstream of the promoter region, such as STAT3, STAT4, NFATc1, CTCF and the like.
The PD1 monoclonal antibody blocks the binding of PD1 protein and its receptor PDL1/PDL2 by the principle of antibody-antigen binding, thereby blocking the inhibition of T cell immune activation. Many monoclonal antibody drugs are successfully marketed.
The production process of the antibody medicament is complex, the yield is low, and the large-scale production is not facilitated; the antibody medicine belongs to the protein with poor stability and high preservation condition, and is not beneficial to transportation and preservation.
Disclosure of Invention
The purpose of the invention is to provide a plurality of compounds, which can block the expression of PDCD1 gene from the gene transcription level through pyrrole imidazole polyamide molecule, thereby realizing the inhibition effect of PD 1.
The polypeptide small molecules used in the patent are pyrrole-imidazole polyamide polypeptides (PIPs) and have strong DNA sequence recognition specificity. It is composed of N-methylpyrrole (Im) and N-methylimidazole (Py). The Im/Py pair is capable of recognizing the G: C base pair; Py/Py pairs recognize the A: T and T: A base pairs; the Hp (hydroxypyrole)/Py pair can recognize the T: A base pair; the N-terminal Ct (3-chlorothiophene)/Py pair binds more readily to the T: A base pair. According to the combination rule, the PIPS molecules can recognize specific DNA sequences and are combined in the minor groove of the double helix structure. The pyrrole-imidazole polyamide polypeptide molecule is designed and synthesized according to a cis-acting element DNA sequence of a PDCD1 gene promoter region. The polypeptide molecule and transcription factors are combined with the cis-acting elements in a competitive way, so that the transcriptional activation of the PDCD1 gene is blocked, and the expression of PD1 is inhibited from the gene transcription level.
The invention discloses a pyrrole-imidazole polyamide polypeptide molecule for inhibiting the transcription of a programmed death receptor 1 gene. The sequence of this molecule is: PyPyIm-gamma-PyPyIm; PyPyIm- γ -PyPyIm. Im is methylimidazole, Py is methylpyrrole, and gamma is gamma-aminobutyric acid. 1A, 1B, 1C, 2A, 2B, 2C and 2D are different modifications at 1 and 2, respectively.
One aspect of the present invention provides a compound that inhibits the expression of programmed death receptor 1 and pharmaceutically acceptable salts thereof, having the structure shown in formula I
Wherein A is selected from C or N;
R1selected from H, CH3、(CH2)3-N(CH3)2、(CH2)2-N(CH3)2、CH2-N(CH3)2、(CH2)3-N(CH3)CH2CH2NHR3；
R2Selected from H, CH3、
R3Selected from H, CH3、
In one embodiment of the present invention, there is provided a compound for inhibiting the expression of programmed death receptor 1 and pharmaceutically acceptable salts thereof, having the structure shown in formula I
Wherein A is C;
R1selected from H, CH3、(CH2)3-N(CH3)2、(CH2)2-N(CH3)2、CH2-N(CH3)2(ii) a And R is2Is selected from
Or
R1Is selected from (CH)2)3-N(CH3)CH2CH2NHR3(ii) a And R is2Selected from H, CH3；R3Selected from H, CH3、
The technical scheme of the invention provides a compound for inhibiting expression of a programmed death receptor 1 and pharmaceutically acceptable salt thereof, wherein the compound has a structure shown in a formula I
Wherein A is N;
R1selected from H, CH3、(CH2)3-N(CH3)2、(CH2)2-N(CH3)2、CH2-N(CH3)2(ii) a And R is2Is selected from
Or
R1Is selected from (CH)2)3-N(CH3)CH2CH2NHR3(ii) a And R is2Selected from H, CH3；R3Selected from H, CH3、
In a particular embodiment, the structure of formula I is selected from one of 1A, 1B, 1C, 2A, 2B, 2C, or 2D.
In another aspect, the invention provides the use of the above compound and its pharmaceutically acceptable salts in the preparation of a medicament for inhibiting the expression of PD-1 on the cell surface.
Another aspect of the present invention provides a method of modulating the expression of the PD1 gene in a cell in vitro, the method comprising the step of administering to the cell to be treated a compound of the present invention or a pharmaceutically acceptable salt thereof.
In another aspect, the invention provides a use for preparing a medicament for treating a disease with abnormal expression or regulation of PD-1 gene, preferably, the disease with abnormal expression or regulation of PD-1 gene is selected from cancer, virus infection or autoimmune disease.
Cancer is selected from diseases in which there is an unlimited proliferation of cells in an organ or body tissue. Preferred are ovarian cancer, leukemia, lung cancer, colorectal/colon cancer, CNS cancer, melanoma, renal cell carcinoma, plasmacytoma/myeloma, prostate cancer, breast cancer, and the like.
The autoimmune disease is selected from a disease in which the subject exerts a destructive immune response to its own tissues. Preferred are Hashimoto's thyroiditis, systemic lupus erythematosus, Sjogren's sjordrome, Graves ' disease, scleroderma, rheumatoid arthritis, multiple sclerosis, myasthenia gravis and diabetes.
In a further aspect, the invention provides a pharmaceutical composition comprising a therapeutically effective amount of a compound of the invention, or a pharmaceutically acceptable salt thereof.
In a further aspect, the present invention provides a process for the preparation of the aforementioned compound, comprising the steps of:
1) sequentially coupling 2 4-amino-1-methyl-2-carboxy-pyrroles and 2 4-amino-1-methyl-2-carboxy-imidazoles on a solid phase resin; or
1) Sequentially coupling 3 4-amino-1-methyl-2-carboxy-pyrroles and 1 4-amino-1-methyl-2-carboxy-imidazole on a solid phase resin;
2) coupling carboxyl of 2, 4-diaminobutyric acid on amino at 4-position of imidazolyl of the intermediate obtained in the step 1);
3) sequentially coupling 2 4-amino-1-methyl-2-carboxy-pyrroles, 1 4-amino-1-methyl-2-carboxy-imidazole and one 1-methyl-2-carboxy-imidazole to the 4-amino group of 2, 4-diaminobutyric acid of the intermediate obtained in step 2);
4) cracking the resin, and modifying isophthalic acid, a herstonic acid derivative Ht-1, a herstonic acid derivative Ht-2 or folic acid FA on the intermediate obtained in the step 3) to obtain a final product; or
4) Modifying isophthalic acid, a hoechst acid derivative Ht-1, a hoechst acid derivative Ht-2 or folic acid FA on the intermediate obtained in the step 3), and cracking the resin to obtain the final product.
Wherein the step 1) is as follows:
removing the protective group from the solid-phase resin phenylhydrazine, coupling 2 4-amino-1-methyl-2-carboxy-pyrroles with 2 4-amino-1-methyl-2-carboxy-imidazoles, or
Removing a protecting group on the solid-phase resin phenylhydrazine, and coupling 3 4-amino-1-methyl-2-carboxyl-pyrrole and 1 4-amino-1-methyl-2-carboxyl-imidazole;
preferably, the step of coupling 4-amino-1-methyl-2-carboxy-pyrrole in step 1) or 3) is:
activating carboxyl on 4-tert-butyloxycarbonylamino-1-methyl-1H-pyrrole-2-carboxylic acid, and coupling with amino on aniline solid-phase synthetic resin or intermediate with a removed protecting group; removing the amino protecting group of the tert-butyloxycarbonyl group;
more preferably, the step of coupling 4-amino-1-methyl-2-carboxy-pyrrole in step 1) or 3) is:
dissolving 4-tert-butyloxycarbonylamino-1-methyl-1H-pyrrole-2-carboxylic acid and triphosgene in an organic solvent together, dropwise adding collidine, adding an alkaline agent after the reaction is completed, coupling and mixing the mixture with amino on the protecting group removing intermediate, and reacting completely in an inert atmosphere; TFA/phenol/H2And removing the tert-butyloxycarbonyl protecting group from the O (v: v: v ═ 92:5:2.5) mixed solution.
The step of coupling 4-amino-1-methyl-2-carboxy-imidazole in step 1) or 3) is:
activating carboxyl on 4-tert-butyloxycarbonylamino-1-methyl-1H-imidazole-2-carboxylic acid, and coupling with amino on aniline solid-phase synthetic resin or intermediate with a removed protecting group; removing the amino protecting group of the tert-butyloxycarbonyl group;
more preferably, the step of coupling 4-amino-1-methyl-2-carboxy-imidazole in step 1) or 3) is:
dissolving 4-tert-butyloxycarbonylamino-1-methyl-1H-pyrrole-2-carboxylic acid and triphosgene in an organic solvent together, dropwise adding collidine, after the reaction is completed, adding an alkaline agent, coupling and mixing with amino on aniline solid-phase synthetic resin or an intermediate with a protective group removed, and reacting completely in an inert atmosphere; TFA/phenol/H2And removing the tert-butyloxycarbonyl protecting group from the O (v: v: v ═ 92:5:2.5) mixed solution.
In the technical scheme of the invention, step 2) is to activate R-2- (9-fluorenylmethoxycarbonylamino) -4-tert-butyloxycarbonylaminobutyric acid and perform amino coupling on the intermediate obtained in step 1); removing the tert-butyloxycarbonyl protecting group;
preferably, the step 2) is to dissolve R-2- (9-fluorenylmethoxycarbonylamino) -4-tert-butoxycarbonylaminobutyric acid and triphosgene into an organic solvent together, dropwise add collidine, after the reaction is completed, add an alkaline agent and a condensing agent until the reaction is completed, perform coupling mixing with the amino on the protecting group-removed intermediate, and perform reaction completely in an inert atmosphere; TFA/phenol/H2And removing the tert-butyloxycarbonyl protecting group from the O (v: v: v ═ 92:5:2.5) mixed solution.
In the technical scheme of the invention, the step of coupling 1-methyl-2-carboxyl-imidazole in the step 3) comprises the following steps:
activating carboxyl on the 1-methyl-1H-imidazole-2-carboxylic acid, and coupling with amino on the intermediate obtained in the previous step of removing the protecting group; the Fmoc protecting group is subsequently removed.
Preferably, 1-methyl-1H-imidazole-2-carboxylic acid and a condensing agent are dissolved in an organic solvent, an alkaline agent is added for reaction till completion, the mixture is added into the intermediate obtained in the previous step, and the condensation reaction is carried out till completion in an inert atmosphere; the Fmoc protecting group was removed with 20% piperidine/DMF.
The condensing agent used in the invention is selected from one or more of HATU, HOBt, HCTU, DCC, DIC, EDC, HBTU, HOAt, PyBOP, PyAOP, BOP-Cl, SOCl2 and oxalyl chloride.
The alkaline agent refers to a compound capable of providing an alkaline environment, and is selected from one or more of DIEA, collidine and triethylamine
In the technical scheme of the invention, step 4) is to protect 2-amino butyric acid in the product obtained in step 3) by using tert-butyloxycarbonyl; then adding N, N-bis (3-aminopropyl) methylamine, reacting completely, and purifying; adding the compound A, a condensing agent and an alkaline agent into the obtained product, reacting completely, and purifying to obtain a final product; the compound A is selected from isophthalic acid, folic acid, a Herster acid derivative Ht-2 or a Herster acid derivative Ht-1.
In the present inventionIn the technical scheme, the step 4) is to activate the compound A with a condensing agent and an alkaline agent, then react with the product obtained in the step 3) to the completion, and react with dimethylaminopropylamine and Cu (OAc)2Reacting completely, and purifying to obtain a final product; the compound A is selected from isophthalic acid, folic acid, a Herster acid derivative Ht-2 or a Herster acid derivative Ht-1.
Advantageous effects
The polyamide molecule disclosed by the invention belongs to polypeptide micromolecules, can realize chemical synthesis, is favorable for large-scale production, and provides more methods for inhibiting the function of PD 1.
Drawings
FIG. 1 is a HRMS spectrum of a compound of formula 1A.
FIG. 2 is a HRMS spectrum of the compound of formula 1B
FIG. 3 is a HRMS spectrum of the compound of formula 1C.
FIG. 4 is a HRMS spectrum of the compound of formula 2A.
FIG. 5 is a HRMS spectrum of the compound of formula 2B
FIG. 6 is a HRMS spectrum of compound of formula 2C.
Figure 7 is a HRMS spectrum of a compound of formula 2D.
FIG. 8 is a HNMR spectrum of Ht-2.
FIG. 9 is a graph showing the results of experiments on the inhibition of apoptosis receptor 1 gene expression by the compounds of the present invention.
Detailed Description
EXAMPLE 1 preparation of Compound 1A
(a) Swelling resin: a10 mL solid phase reactor was charged with 400mg of Fmoc-protected phenylhydrazine resin (0.66mmol/g, 0.264mmol) and 3mL of CH2Cl2Swelling the resin for 30min, removing CH2Cl2And is ready for use;
(b) removing Fmoc protecting groups: adding 3mL of 20% piperidine/DMF solution to the swollen resin from step (a), N2Bubbling and mixing evenly, after 10min, removing the solvent, and then adding 3mL of 20% piperidine/DMF solution, N2Bubbling and uniformly mixing, washing the resin with DMF (4X 3mL) after 10min, and then washing the resin with 3mL of anhydrous DMF for later use;
(c) amino acid condensation: dissolving 4-tert-butyloxycarbonylamino-1-methyl-1H-pyrrole-2-carboxylic acid (254mg, 1.056mmol) and triphosgene (BTC, 128mg, 0.433mmol) in 2mL of anhydrous THF, slowly adding collidine (488. mu.L, 3.696mmol) dropwise into the solution, reacting to generate a large amount of white precipitate immediately, adding the reaction for 3min, adding 2mL of DIEA/DMF solution (5%, v/v), completely removing the white precipitate, transferring the reaction solution to the phenylhydrazine resin without protecting group in step (b), and removing N2Bubbling and uniformly mixing, carrying out condensation reaction for 0.5-1 h, pumping out reaction liquid, and washing resin with DMF (4X 3mL) for later use;
(d) removing the tert-butyloxycarbonyl protecting group: by CH2Cl2(2X 3mL) washing, CH extraction2Cl23.0mL of TFA/phenol/H was added2Removing the tert-butoxycarbonyl protecting group on the condensation product obtained in the step (b) by using a mixed solution of O (v: v: v ═ 92:5:2.5), removing the solvent after 2min, and adding 3.0mL of TFA/phenol/H again2The mixed solution of O (v: v: v ═ 92:5:2.5) was reacted for 20min with CH2Cl2(2X 3mL) and DMF (4X 3mL) and then 3mL of anhydrous DMF;
repeating the condensation and deprotection steps (c) and (d) until the synthesis of the peptide represented by the formula (1) supported on the phenylhydrazine resin is completed;
(e) amino acid condensation: dissolving 4-tert-butyloxycarbonylamino-1-methyl-1H-imidazole-2-carboxylic acid (254mg, 1.056mmol) and triphosgene (BTC, 128mg, 0.433mmol) in 2mL of anhydrous THF, slowly adding collidine (488. mu.L, 3.696mmol) dropwise into the solution, reacting to generate a large amount of white precipitate immediately, adding the reaction for 3min, adding 2mL of DIEA/DMF solution (5%, v/v), completely removing the white precipitate, transferring the reaction solution to the phenylhydrazine resin with the protective groups removed in the step (b), and removing N from the reaction solution2Bubbling and uniformly mixing, carrying out condensation reaction for 0.5-1 h, pumping out reaction liquid, and washing resin with DMF (4X 3mL) to obtain the peptide shown as a formula (2) loaded on phenylhydrazine resin;
(g) condensation of γ -amino acids: dissolving R-2- (9-fluorenylmethoxycarbonylamino) -4-tert-butoxycarbonylaminobutyric acid (465mg, 1.056mmol) and triphosgene (128mg, 0.433mmol) in 2mL of anhydrous THF, slowly adding collidine (488 uL, 3.696mmol) dropwise into the solution to react to generate a large amount of white precipitate immediately, adding HOAt (144mg, 1.056mmol), adding 2mL of DIEA/DMF solution (5%, v/v), reacting for 5min, completely removing the white precipitate, and transferring the reaction solution to the straight-chain peptide (NH) loaded on phenylhydrazine resin shown in the formula (1)2-Im-Py-phenylhydrazine resin), N2Bubbling and uniformly mixing, carrying out condensation reaction for 0.5-1 h, pumping out reaction liquid, and washing resin with DMF (4X 3mL) to obtain the peptide shown as a formula (3) loaded on the phenylhydrazine resin;
(h) repeating the deprotection and condensation protection steps (d) and (c), wherein the synthesis of the peptide loaded on the phenylhydrazine resin shown in the formula (4) is completed;
(i) repeating the deprotecting and condensation protecting steps (d) and (e) wherein until completion to obtain formula (5)
Synthesizing the peptide loaded on the phenylhydrazine resin;
(j) condensation of the terminal amino acids: 1-methyl-1H-imidazole-2-carboxylic acid (132mg, 1.056mmol) and PyBOP (550mg, 1.056mmol) were dissolved in 3mL of anhydrous DMF, DIEA (350. mu.L, 2.112mmol) was added, reaction was carried out for 5min, and the reaction solution was transferred to the peptide of formula (4) obtained in step (H) supported on phenylhydrazine resin, N2Bubbling, mixing, condensation reacting for 2 hr, pumping out reaction liquid, and adding DMF (4 extract)3mL) of the washing resin, and removing the Fmoc protecting group in the peptide loaded on the phenylhydrazine resin shown in the formula (5) by adopting the method of the step (b) to obtain the peptide loaded on the phenylhydrazine resin shown in the formula (6);
(k) reacting Boc2O (243. mu.L, 1.056mmol) was dissolved in 3mL of anhydrous DMF, DIEA (350. mu.L, 2.112mmol) was added, and the reaction solution was transferred to Fmoc-free peptide of formula (6) supported on phenylhydrazine resin, N2Bubbling, mixing uniformly, and carrying out condensation reaction for 20 min. The reaction solution was aspirated off, and the resin was washed with DMF (4X 3mL) to obtain Boc-protected peptide represented by formula (7) supported on phenylhydrazine resin;
taking out the resin obtained above, adding 1mL of DMF, 200 mu L N, N-bis (3-aminopropyl) methylamine, shaking at 90 ℃ for reaction for 1h, cooling to room temperature, filtering the resin, and adding 20mL of CH2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 16min, and freeze-drying to obtain a light yellow solid for later use;
4mg (3. mu. mol) of the above solid was dissolved in 0.5mL of anhydrous DMF, and isophthalic acid IPA (0.99mg, 6. mu. mol), PyBOP (3.1mg, 6. mu. mol) and DIEA (5. mu.L, 30. mu. mol) were added, reacted for 2h at room temperature with shaking, and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 18min, and freeze-drying to obtain a solid; this solid was dissolved in 1mL CH2Cl21mL TFA was added under ice bath, reacted at 0 ℃ for 1h, 20mL cold ether was added, and the precipitate was collected by centrifugation and purified by semi-preparative HPLC: 10% acetonitrile-H2O (containing 1% of TF)A) Gradient eluting for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 15min, lyophilized to give compound 1A as a pale yellow solid,
the HRMS is shown in FIG. 1.
HRMS(ESI)m/z:[M+H]+1358.6041。
EXAMPLE 2 preparation of Compound 1B
The peptide represented by the formula (7) supported on a phenylhydrazine resin was obtained in the same manner as in example 1,
taking out the resin obtained above, adding 1mL of DMF, 200 mu L N, N-bis (3-aminopropyl) methylamine, shaking at 90 ℃ for reaction for 1h, cooling to room temperature, filtering the resin, and adding 20mL of CH2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 16min, and freeze-drying to obtain a light yellow solid for later use;
4mg (3. mu. mol) of the above solid was dissolved in 0.5mL of anhydrous DMF, and the Herstellic acid derivative Ht-2(2.7mg, 6. mu. mol), PyBOP (3.1mg, 6. mu. mol) and DIEA (5. mu.L, 30. mu. mol) were added, reacted with shaking at room temperature for 2h, and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 18min, and freeze-drying to obtain a solid; this solid was dissolved in 1mL CH2Cl21mL TFA was added under ice bath, reacted at 0 ℃ for 1h, 20mL cold ether was added, and the precipitate was collected by centrifugation and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient of O (containing 1% TFA) etcEluting for 5min, 10-100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 17min, lyophilized to give compound 1B as a pale yellow solid,
the HRMS is shown in fig. 2.
HRMS(ESI)m/z:[M+H]+：1644.7748
The structure and the synthetic route of the herceptic acid derivative Ht-2 are as follows:
Ht-2-B (11.4g,44.2mmol) and 4- (5- (4-methylpiperazin-1-yl) -1H-benzo [ d ] imidazol-2-yl) benzene-1, 2-diamine (Ht-2-A, 10g,31.0mmol, prepared as described in Inorg. chem.1998,37, 6018. sup. 6022) were dissolved in acetic acid (100mL) and reacted in an oil bath under heating and refluxing for 4 hours. And (4) distilling off acetic acid under reduced pressure, cooling to room temperature, and purifying by column chromatography to obtain a grass green solid. Dissolving the obtained product in methanol (100ml), magnetically stirring, protecting with nitrogen, cooling in ice water bath, slowly adding dropwise 50ml aqueous solution of 3.1g sodium hydroxide, reacting at room temperature for 8h, detecting by TLC to complete the reaction, evaporating to remove methanol, adjusting pH with dilute hydrochloric acid to acidity, separating out solid, filtering, and drying to obtain yellowish green solid Ht-2, wherein HNMR is shown in FIG. 8.
HNMR(DMSO-d6，400MHz)：2.44(s,3H),2.79(s,2H),3.23(s,2H),6.96(m,1H),7.05(br s,1H),7.47(m,1H),7.69-7.73(m,2H),8.08(m,1H),8.33-8.49(m,2H),8.83(s,1H)，12.73(br s,1H),13.41(br s,1H)。
EXAMPLE 3 preparation of Compound 1C
The peptide represented by the formula (7) supported on a phenylhydrazine resin was obtained in the same manner as in example 1,
will be described in detailThe resulting resin was removed, 1mL DMF, 200. mu. L N, N-bis (3-aminopropyl) methylamine was added, the reaction was shaken at 90 ℃ for 1h, cooled to room temperature, the resin was filtered off and 20mL CH was used2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 16min, and freeze-drying to obtain a light yellow solid for later use;
4mg (3. mu. mol) of the above solid was dissolved in 0.5mL of anhydrous DMF, and folic acid FA (1.8mg, 6. mu. mol), PyBOP (3.1mg, 6. mu. mol) and DIEA (5. mu.L, 30. mu. mol) were added, reacted at room temperature with shaking for 2h, and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 18min, and freeze-drying to obtain a solid; this solid was dissolved in 1mL CH2Cl21mL of THF was added under ice-bath, reacted at 0 ℃ for 1h, 20mL of cold ether was added, and the precipitate was collected by centrifugation and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 17min, lyophilized to give compound 1C as a pale yellow solid,
the HRMS is shown in fig. 3.
HRMS(ESI)m/z:[M+H]+:1633.7277。
Wherein the structure of folic acid FA:
EXAMPLE 4 preparation of Compound 2A
(a) Swelling resin: 400mg of Fmoc-protected phenylhydrazine resin (f) was added to a 10mL solid phase reactor0.66mmol/g, 0.264mmol) and 3mL CH2Cl2Swelling the resin for 30min, removing CH2Cl2And is ready for use;
(b) removing Fmoc protecting groups: adding 3mL of 20% piperidine/DMF solution to the swollen resin from step (a), N2Bubbling and mixing evenly, after 10min, removing the solvent, and then adding 3mL of 20% piperidine/DMF solution, N2Bubbling and uniformly mixing, washing the resin with DMF (4X 3mL) after 10min, and then washing the resin with 3mL of anhydrous DMF for later use;
(c) amino acid condensation: dissolving 4-tert-butyloxycarbonylamino-1-methyl-1H-pyrrole-2-carboxylic acid (254mg, 1.056mmol) and triphosgene (BTC, 128mg, 0.433mmol) in 2mL of anhydrous THF, slowly adding collidine (488. mu.L, 3.696mmol) dropwise into the solution, reacting to generate a large amount of white precipitate immediately, adding the reaction for 3min, adding 2mL of DIEA/DMF solution (5%, v/v), completely removing the white precipitate, transferring the reaction solution to the phenylhydrazine resin without protecting group in step (b), and removing N2Bubbling and uniformly mixing, carrying out condensation reaction for 0.5-1 h, pumping out reaction liquid, and washing resin with DMF (4X 3mL) for later use;
(d) removing the tert-butyloxycarbonyl protecting group: by CH2Cl2(2X 3mL) washing, CH extraction2Cl23.0mL of TFA/phenol/H was added2Removing the tert-butoxycarbonyl protecting group on the condensation product obtained in the step (b) by using a mixed solution of O (v: v: v ═ 92:5:2.5), removing the solvent after 2min, and adding 3.0mL of TFA/phenol/H again2The mixed solution of O (v: v: v ═ 92:5:2.5) was reacted for 20min with CH2Cl2(2X 3mL) and DMF (4X 3mL) and then 3mL of anhydrous DMF;
repeating the condensation and deprotection steps (c) and (d) until the synthesis of the peptide represented by the formula (8) supported on the phenylhydrazine resin is completed;
(e) amino acid condensation: 4-tert-Butoxycarbonylamino-1-methyl-1H-imidazole-2-carboxylic acid (254mg, 1.056mmol) and triphosgene (BTC, 128mg, 0.433mmol) were dissolved in 2mL of anhydrous THF, to whichSlowly dripping collidine (488 mu L, 3.696mmol) into the solution, immediately generating a large amount of white precipitate after the reaction, finishing the reaction for 3min, then adding 2mL DIEA/DMF solution (5%, v/v), completely eliminating the white precipitate, transferring the reaction solution into the phenylhydrazine resin for removing the protecting group in the step (b), and adding N into the solution2Bubbling and uniformly mixing, carrying out condensation reaction for 0.5-1 h, pumping out reaction liquid, and washing resin with DMF (4X 3mL) for later use;
repeating the condensation and deprotection steps (e) and (d) until the synthesis of the peptide represented by the formula (9) supported on the phenylhydrazine resin is completed;
(g) condensation of γ -amino acids: dissolving R-2- (9-fluorenylmethoxycarbonylamino) -4-tert-butoxycarbonylaminobutyric acid (465mg, 1.056mmol) and triphosgene (128mg, 0.433mmol) in 2mL of anhydrous THF, slowly adding collidine (488 uL, 3.696mmol) dropwise to the solution to react to generate a large amount of white precipitate immediately, adding HOAt (144mg, 1.056mmol), adding 2mL of DIEA/DMF solution (5%, v/v), reacting for 5min to completely remove the white precipitate, and transferring the reaction solution to the straight-chain peptide (NH) loaded on phenylhydrazine resin shown in the formula (9)2-Im-Py-phenylhydrazine resin), N2Bubbling and uniformly mixing, carrying out condensation reaction for 0.5-1 h, pumping out reaction liquid, and washing resin with DMF (4X 3mL) to obtain the peptide shown as a formula (10) loaded on the phenylhydrazine resin;
(h) repeating the deprotection and condensation protection steps (d) and (c), wherein the synthesis of the peptide loaded on the phenylhydrazine resin shown in the formula (11) is completed;
(i) repeating the deprotection and condensation protection steps (d) and (e), wherein the synthesis of the peptide loaded on the phenylhydrazine resin shown in the formula (12) is completed;
(j) condensation of the terminal amino acids: 1-methyl-1H-imidazole-2-carboxylic acid (132mg, 1.056mmol) and PyBOP (550mg, 1.056mmol) were dissolved in 3mL of anhydrous DMF, DIEA (350. mu.L, 2.112mmol) was added, reaction was carried out for 5min, and the reaction solution was transferred to the peptide of formula (12) obtained in step (H) supported on phenylhydrazine resin, N2Bubbling, uniformly mixing, carrying out condensation reaction for 2h, pumping out reaction liquid, and washing the resin with DMF (4X 3 mL); removing the Fmoc protecting group in the peptide loaded on the phenylhydrazine resin shown in the formula (12) in the step (b) in the embodiment 1 to obtain the peptide loaded on the phenylhydrazine resin shown in the formula (13);
reacting Boc2O (243. mu.L, 1.056mmol) was dissolved in 3mL of anhydrous DMF, DIEA (350. mu.L, 2.112mmol) was added, and the reaction solution was transferred to Fmoc-free peptide of formula (13) supported on phenylhydrazine resin, N2Bubbling, mixing uniformly, and carrying out condensation reaction for 20 min. The reaction solution was aspirated off, and the resin was washed with DMF (4X 3mL) to obtain Boc-protected peptide represented by formula (14) supported on phenylhydrazine resin;
the peptide of the above formula (14) supported on phenylhydrazine resin was removed, 1mL of DMF, 200. mu. L N, N-bis (3-aminopropyl) methylamine were added, the mixture was shaken at 90 ℃ for 1 hour, cooled to room temperature, the resin was filtered off, and 20mL of CH was used2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 16min, and freeze-drying to obtain a light yellow solid for later use;
4mg (3. mu. mol) of the above solid was dissolved in 0.5mL of anhydrous DMF, and isophthalic acid IPA (0.99mg, 6. mu. mol), PyBOP (3.1mg, 6. mu. mol) and DIEA (5. mu.L, 30. mu. mol) were added, reacted for 2h at room temperature with shaking, and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 18min, and freeze-drying to obtain a solid; this solid was dissolved in 1mL CH2Cl21mL TFA was added under ice bath, reacted at 0 ℃ for 1h, 20mL cold ether was added, and the precipitate was collected by centrifugation and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 15min, lyophilized to give compound 2A as a pale yellow solid,
the HRMS is shown in fig. 4.
HRMS(ESI)m/z:[M+H]+：1359.5992。
EXAMPLE 5 preparation of Compound 2B
The peptide represented by the formula (14) supported on a phenylhydrazine resin was obtained in the same manner as in example 4.
The peptide of the formula (14) obtained above, which is supported on phenylhydrazine resin, is removed, 1mL of DMF, 200. mu. L N, N-bis (3-aminopropyl) methylamine are added, the mixture is shaken at 90 ℃ for reaction for 1 hour, the mixture is cooled to room temperature, the resin is filtered off, and 20mL of CH is used2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 16min, and freeze-drying to obtain a light yellow solid for later use;
4mg (3. mu. mol) of the above solid was dissolved in 0.5mL of anhydrous DMF, and the Herstellic acid derivative Ht-2(2.7mg, 6. mu. mol), PyBOP (3.1mg, 6. mu. mol) and DIEA (5. mu.L, 30. mu. mol) were added, reacted with shaking at room temperature for 2h, and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 18min, and freeze-drying to obtain a solid; this solid was dissolved in 1mL CH2Cl21mL TFA was added under ice bath, reacted at 0 ℃ for 1h, 20mL cold ether was added, and the precipitate was collected by centrifugation and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 17min, lyophilized to give compound 2B as a pale yellow solid,
the HRMS is shown in fig. 5.
HRMS(ESI)m/z:[M+H]+：1645.7797.
The structure and the synthetic route of the herceptic acid derivative Ht-2 are as follows:
ht-2(11.4g,44.2mmol) and 4- (5- (4-methylpiperazin-1-yl) -1H-benzo [ d ] imidazol-2-yl) benzene-1, 2-diamine (Ht-2-A, 10g,31.0mmol, prepared as described in Inorg. chem.1998,37, 6018. 6022) were dissolved in acetic acid (100mL) and heated to reflux for 4 hours in an oil bath. And (4) distilling off acetic acid under reduced pressure, cooling to room temperature, and purifying by column chromatography to obtain a grass green solid. Dissolving the obtained product in methanol (100ml), magnetically stirring, protecting with nitrogen, cooling in ice water bath, slowly adding 3.1g sodium hydroxide 50ml water solution dropwise, reacting at room temperature for 8h, detecting by TLC to completely react, evaporating to remove methanol, adjusting pH with dilute hydrochloric acid, precipitating solid, filtering, and drying to obtain yellowish green solid Ht-2, wherein HNMR is shown in FIG. 8.
HNMR(DMSO-d6，400MHz)：2.44(s,3H),2.79(s,2H),3.23(s,2H),6.96(m,1H),7.05(br s,1H),7.47(m,1H),7.69-7.73(m,2H),8.08(m,1H),8.33-8.49(m,2H),8.83(s,1H)，12.73(br s,1H),13.41(br s,1H)。
EXAMPLE 6 preparation of Compound 2C
The peptide represented by the formula (14) supported on a phenylhydrazine resin was obtained in the same manner as in example 4;
the peptide of the above formula (14) supported on phenylhydrazine resin was removed, 1mL of DMF, 200. mu. L N, N-bis (3-aminopropyl) methylamine were added, the mixture was shaken at 90 ℃ for 1 hour, cooled to room temperature, the resin was filtered off, and 20mL of CH was used2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 16min, and freeze-drying to obtain a light yellow solid for later use;
4mg (3. mu. mol) of the above solid was dissolved in 0.5mL of anhydrous DMF, and folic acid FA (1.8mg, 6. mu. mol), PyBOP (3.1mg, 6. mu. mol) and DIEA (5. mu.L, 30. mu. mol) were added, reacted at room temperature with shaking for 2h, and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRCollecting the product for 18min, and freeze-drying to obtain a solid; this solid was dissolved in 1mL CH2Cl21mL of THF was added under ice-bath, reacted at 0 ℃ for 1h, 20mL of cold ether was added, and the precipitate was collected by centrifugation and purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 17min, lyophilized to give compound 2C as a pale yellow solid,
the HRMS is shown in fig. 6.
HRMS(ESI)m/z:[M+H]+：1633.7277。
Structure of folate FA
EXAMPLE 7 preparation of Compound 2D
The peptide represented by the formula (13) supported on a phenylhydrazine resin was obtained in the same manner as in example 4.
(k) The herceptic acid derivative Ht-1(539mg, 1.056mmol) and PyBOP (550mg, 1.056mmol) were dissolved in 3mL of anhydrous DMF, DIEA (350. mu.L, 2.112mmol) was added thereto, reaction was carried out for 5min, and the reaction solution was transferred to the peptide represented by formula (13) supported on phenylhydrazine resin, N2Bubbling, uniformly mixing, carrying out condensation reaction for 1h, and pumping out reaction liquid to obtain the peptide loaded on the phenylhydrazine resin in the formula (15).
The structure of the herceptic acid derivative Ht-1 is as follows (see J.AM. CHEM. SOC.2004,126, 3736-3747):
(l) The resin of formula (15) is removed and 1mL of DMF, 200. mu.L of dimethylaminopropylamine and 10mg of Cu (OAc) are added2The reaction was shaken at room temperature for 12h, the resin filtered off and charged with 20mL CH2Cl2Washing the resin; the organic phase was concentrated and the residue was purified by semi-preparative HPLC: 10% acetonitrile-H2Gradient elution of O (1% TFA) for 5min, 10% to 100% acetonitrile-H2Gradient elution with O (1% TFA) for 25min, retention time TRThe product was collected for 15min, lyophilized to give compound 2D as a pale yellow solid,
the HRMS is shown in fig. 7.
HRMS (ESI) m/z theoretical calculation C82H94N29O11[M+H]+1660.7682 found 1660.7652.
EXAMPLE 8 inhibition of apoptosis receptor 1 Gene expression assay by Compounds
1) Separating peripheral blood mononuclear cells from fresh blood, and culturing by using Takara + 10% FBS + IL 2;
2) peripheral blood mononuclear cells are inoculated into a 96-well plate, wherein each well is inoculated with 50 ten thousand cells, and the volume is 150 ul; adding compound with the concentration of 20uM, and adding CD3 antibody for stimulation;
3) after culturing for 48 hours, dividing the cells in the 72h group into two groups, adding the same compound again for culturing one group, adding no compound in the other group, and adding a new compound in the other group according to the drug concentration of 20uM and the total volume of 200 ul;
4) cell samples were collected at 0, 24, 48, and 72 hours, stained with PE-PD1 antibody and FITC-CD3 antibody, and subjected to flow assay, see fig. 9. From fig. 9, it can be found that the inhibition effect of the two sequences 2A and 2D is better, the inhibition rate of 2A on PD1 in 48 hours of treatment reaches 40%, and the inhibition rate of 2D approaches 50%.
Claims (11)
1. A compound that inhibits the expression of programmed death receptor 1, the compound having the structure set forth in the following table:
2. use of a compound according to claim 1 and pharmaceutically acceptable salts thereof in the manufacture of a medicament for inhibiting the expression of PD-1 on the surface of a cell.
3. Use of a compound according to claim 1 for the manufacture of a medicament for the treatment of a disease in which PD-1 gene expression or regulation is abnormal; the disease with abnormal PD-1 gene expression or regulation is selected from cancer, virus infection or autoimmune disease;
cancer is selected from diseases in which there is unlimited proliferation of cells in an organ or body tissue;
the autoimmune disease is selected from a disease in which the subject exerts a destructive immune response to its own tissues.
4. The use according to claim 3, wherein the cancer is selected from ovarian cancer, leukemia, lung cancer, colorectal/colon cancer, CNS cancer, melanoma, renal cell carcinoma, plasmacytoma/myeloma, prostate cancer, breast cancer.
5. Use according to claim 3, wherein the autoimmune disease is selected from hashimoto's thyroiditis, systemic lupus erythematosus, sjogren's syndrome, Graves ' disease, scleroderma, rheumatoid arthritis, multiple sclerosis, myasthenia gravis and diabetes.
6. A pharmaceutical composition comprising a therapeutically effective amount of a compound of claim 1 or a pharmaceutically acceptable salt thereof.
7. A process for the preparation of a compound according to claim 1, or a pharmaceutically acceptable salt thereof, comprising the steps of:
1) sequentially coupling 2 4-amino-1-methyl-2-carboxy-pyrroles and 2 4-amino-1-methyl-2-carboxy-imidazoles on a solid phase resin; or
Sequentially coupling 3 4-amino-1-methyl-2-carboxy-pyrroles and 1 4-amino-1-methyl-2-carboxy-imidazole on a solid phase resin;
2) coupling carboxyl of 2, 4-diaminobutyric acid on amino at 4-position of imidazolyl of the intermediate obtained in the step 1);
3) sequentially coupling 2 4-amino-1-methyl-2-carboxy-pyrroles, 1 4-amino-1-methyl-2-carboxy-imidazole and one 1-methyl-2-carboxy-imidazole to the 4-amino group of 2, 4-diaminobutyric acid of the intermediate obtained in step 2);
4) cracking the resin, and modifying isophthalic acid, a herstonic acid derivative Ht-1, a herstonic acid derivative Ht-2 or folic acid FA on the intermediate obtained in the step 3) to obtain a final product; or
Modifying isophthalic acid, a hoechst acid derivative Ht-1, a hoechst acid derivative Ht-2 or folic acid FA on the intermediate obtained in the step 3), and cracking the resin to obtain the final product.
8. The method of claim 7, wherein step 1) is:
removing the protective group from the solid-phase resin phenylhydrazine, coupling 2 4-amino-1-methyl-2-carboxy-pyrroles with 2 4-amino-1-methyl-2-carboxy-imidazoles, or
Removing the protective group on the solid-phase resin phenylhydrazine, and coupling 3 4-amino-1-methyl-2-carboxyl-pyrrole and 1 4-amino-1-methyl-2-carboxyl-imidazole.
9. The method according to claim 8, wherein the step of coupling 4-amino-1-methyl-2-carboxy-pyrrole in step 1) or 3) is:
activating carboxyl on 4-tert-butyloxycarbonylamino-1-methyl-1H-pyrrole-2-carboxylic acid, and coupling with amino on aniline solid-phase synthetic resin or intermediate with a removed protecting group; removing the amino protecting group of the tert-butyloxycarbonyl group;
the step of coupling 4-amino-1-methyl-2-carboxy-imidazole in step 1) or 3) is:
activating carboxyl on 4-tert-butyloxycarbonylamino-1-methyl-1H-imidazole-2-carboxylic acid, and coupling with amino on aniline solid-phase synthetic resin or intermediate with a removed protecting group; and removing the amino protecting group of the tert-butyloxycarbonyl group.
10. The preparation method according to claim 9, wherein step 2) is activating R-2- (9-fluorenylmethoxycarbonylamino) -4-tert-butoxycarbonylaminobutyric acid, and coupling with an amino group on the intermediate obtained in step 1); and removing the amino protecting group of the tert-butyloxycarbonyl group.
11. The method of claim 10, wherein the step of coupling 1-methyl-2-carboxy-imidazole in step 3) is:
activating carboxyl on the 1-methyl-1H-imidazole-2-carboxylic acid, and coupling with amino on the intermediate obtained in the previous step of removing the protecting group.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004078943A2 (en) * | 2003-03-04 | 2004-09-16 | California Institute Of Technology | Alternative heterocycles for dna recognition |
| EP1491534A1 (en) * | 2002-03-08 | 2004-12-29 | Japan Science and Technology Agency | Novel hairpin polyamide |
| WO2008066890A2 (en) * | 2006-11-29 | 2008-06-05 | California Institute Of Technology | Polyamides with tail structures |
| CN106068275A (en) * | 2014-01-23 | 2016-11-02 | 瑞泽恩制药公司 | People's antibody of anti-PD 1 |
-
2016
- 2016-12-23 CN CN201611205967.9A patent/CN106674209B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1491534A1 (en) * | 2002-03-08 | 2004-12-29 | Japan Science and Technology Agency | Novel hairpin polyamide |
| WO2004078943A2 (en) * | 2003-03-04 | 2004-09-16 | California Institute Of Technology | Alternative heterocycles for dna recognition |
| WO2008066890A2 (en) * | 2006-11-29 | 2008-06-05 | California Institute Of Technology | Polyamides with tail structures |
| CN106068275A (en) * | 2014-01-23 | 2016-11-02 | 瑞泽恩制药公司 | People's antibody of anti-PD 1 |
Non-Patent Citations (4)
| Title |
|---|
| Activity of a Py–Im Polyamide Targeted to the Estrogen Response Element;Nicholas G. Nickols;《 Mol Cancer Ther.》;20130226;第12卷(第5期);参见图1及图S1，化合物3，第680页第2栏第1-3段，第678页第2栏第3段，第676页第1栏第3段 * |
| Inhibition of DNA Binding by NF-κB with Pyrrole-Imidazole Polyamides;Nicholas R. Wurtz;《Biochemistry》;20020516;第41卷;第7606页图3化合物3 * |
| Microwave assisted synthesis of Py-Im polyamides;Puckett JW;《Org Lett.》;20120511;第14卷;第2777页第1栏第1段，Supporting Firgure 1 * |
| Modifications at the C-Terminus To Improve Pyrrole-Imidazole Polyamide Activity in Cell Culture;Claire S. Jacobs;《J. Med. Chem.》;20090702;第52卷;第7380页第2栏第1段 * |
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