Small-molecule coupling compound of integrin receptor antagonist c (RGDfK) and application of small-molecule coupling compound

Abstract

The invention relates to the field of medicines, in particular to a small-molecule coupling compound of an integrin receptor antagonist c (RGDfK) and application of the small-molecule coupling compound. The structural formula of the small-molecule coupling compound is # imgabs0 #, a coupling technology is utilized, an integrin receptor antagonist c (RGDfK) and a coupling bifunctional molecule with anti-tumor activity small molecules are provided, and the efficient anti-tumor effect is achieved by utilizing the targeting property of the small-molecule coupling compound on an integrin receptor and generating a synergistic effect.

Description

Technical Field

[0001] The present invention relates to the field of medicine, and in particular to a small molecule coupling compound of an integrin receptor antagonist c (RGDfK) and applications thereof. Background Art

[0002] Traditional combination drug delivery faces challenges such as pharmacokinetic mismatch, systemic toxicity superposition, and insufficient local drug concentration in tumors. In recent years, antibody-drug conjugates (ADCs) have enhanced their efficacy through targeted delivery and controlled release mechanisms. However, their design often relies on large molecular carriers (such as antibodies) and faces numerous limitations, such as long circulation time in the blood, low penetration into solid tumors, high synthesis costs, and potential immunogenicity. In contrast, small molecule targeting ligands in small molecule drug conjugates (SMDCs) offer a promising alternative to overcome these challenges due to their inherent non-immunogenicity, in vitro and in vivo stability, low molecular weight, and overall greater cost-effectiveness.

[0003] Integrin receptors are a class of transmembrane glycoproteins on the cell surface. Multiple integrin isoforms (such as αvβ3, αvβ5, α5β1, and α6β4) are significantly upregulated in tumor cells or tumor vascular endothelial cells, and their expression levels correlate with tumor stage, metastatic potential, and poor prognosis. When cells come into contact with the extracellular matrix, integrin receptors aggregate and bind to extracellular matrix ligands, triggering conformational changes in the integrin receptors and subsequently recruiting FAK to focal adhesion sites. FAK interacts with the intracellular domain of the integrin receptor via its N-terminal FERM domain, leading to autophosphorylation of FAK and the formation of phosphotyrosine sites. These phosphotyrosine sites recruit SH2 domain-containing proteins, such as the Src family kinases, thereby activating Src kinase activity. Activated Src, in turn, further phosphorylates additional tyrosine residues on FAK, forming a positive feedback loop that amplifies signaling and contributes to tumor cell proliferation, metastasis, adhesion, and invasion.

[0004] The cyclic peptide c(RGDfK) is a widely studied integrin receptor antagonist. Compared with the linear RGD peptide, c(RGDfK) provides structural rigidity, reduces sensitivity to chemical degradation, and enhances stability and specificity. However, the anti-tumor activity of c(RGDfK) is low and does not meet clinical needs. Summary of the Invention

[0005] Based on the above content, the present invention provides a small molecule coupling compound of integrin receptor antagonist c(RGDfK) and its application.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] The present invention provides a compound represented by formula (I) (i.e., a small molecule coupled compound of integrin receptor antagonist c(RGDfK)), a stereoisomer thereof, or a pharmaceutically acceptable salt thereof;

[0008]

[0009] In formula (I), L is selected from methylene, ethylene or

[0010] X is selected from CH or N;

[0011] R 1 is selected from hydrogen, deuterium, fluorine, chlorine, nitro, cyano, C1-C3 alkyl or C1-C3 haloalkyl;

[0012] R 2 is selected from hydrogen, deuterium, C1-C3 alkyl, C1-C3 deuterated alkyl or C3-C6 cycloalkyl;

[0013] R 3 is selected from hydrogen, deuterium, halogen, nitro, cyano, C1-C3 alkyl, C1-C3 haloalkyl, C1-C3 deuterated alkyl, C1-C3 alkoxy, C1-C3 haloalkoxy or C1-C3 deuterated alkoxy.

[0014] In a preferred embodiment of the present invention, the compound represented by formula (I) is one of the following compounds:

[0015]

[0016]

[0017]

[0018]

[0019] The present invention also provides use of the above-mentioned compound, its stereoisomer or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating integrin receptor-mediated diseases.

[0020] In a preferred embodiment of the present invention, the integrin receptor-mediated related disease is ovarian cancer, breast cancer, hepatocellular carcinoma, bladder cancer, colorectal cancer, lung cancer, thyroid cancer, prostate cancer, melanoma, osteosarcoma or glioblastoma.

[0021] The present invention also provides a drug for treating integrin receptor-mediated diseases, wherein the active ingredient comprises the above-mentioned compound, its stereoisomer or a pharmaceutically acceptable salt thereof.

[0022] In a preferred embodiment of the present invention, the drug further comprises a pharmaceutically acceptable excipient.

[0023] The synthetic route of the compound represented by formula (I) of the present invention is as follows:

[0024]

[0025] The preparation method of the compound represented by formula (I) is as follows:

[0026] Step I: Compound A is treated in a piperidine-containing N,N-dimethylformamide solution to remove the fluorenylmethoxycarbonyl group (Fmoc group) to obtain compound B with an exposed amino group;

[0027] Step II: Compound B and 1-2 equivalents of activated spacer C are reacted in a suitable solvent and catalyzed by a basic reagent to obtain compound D;

[0028] Wherein, L is methylene, ethylene, or The solvent is selected from N,N-dimethylformamide, tetrahydrofuran, dioxane, etc.; the alkaline reagent is selected from pyridine, triethylamine, N,N-diisopropylethylamine, potassium carbonate, cesium carbonate, etc.

[0029] Step III: Compound D is condensed with the integrin receptor antagonist cyclic peptide c (RGDfK) in an appropriate solvent and under the catalysis of an alkaline reagent to obtain a small molecule conjugate of formula (I); the solvent is selected from N,N-dimethylformamide, tetrahydrofuran, dioxane, etc.; the alkaline reagent is selected from pyridine, triethylamine, N,N-diisopropylethylamine, potassium carbonate, cesium carbonate, etc.

[0030] The present invention discloses the following technical effects:

[0031] The present invention utilizes coupling technology to provide a class of integrin receptor antagonist c (RGDfK) coupled bifunctional molecules with small molecules having anti-tumor activity. By utilizing the targeting of this class of small molecule conjugates to integrin receptors, a synergistic effect is generated to achieve a highly effective anti-tumor effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0033] Figure 1 This is the dissociation curve of the small molecule coupled compound (Compound of Example 1, Compound of Example 2 and Compound of Example 3) under the action of cathepsin B. DETAILED DESCRIPTION

[0034] Unless otherwise specified, the technical solutions described in the present invention are all conventional solutions in the field, and the reagents or raw materials used, unless otherwise specified, are purchased from commercial channels or have been disclosed.

[0035] The technical solutions provided by the present invention are described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0036] Unless otherwise stated, the terms used in the specification and claims have the following meanings.

[0037] The term "alkyl" refers to a saturated aliphatic hydrocarbon group, which is a straight or branched chain group containing 1 to 4 carbon atoms, preferably an alkyl group containing 1 to 3 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, and sec-butyl.

[0038] The term "cycloalkyl" refers to a saturated monocyclic or polycyclic hydrocarbon, the cycloalkyl ring containing 3 to 8 carbon atoms, preferably 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl.

[0039] The term "heterocycloalkyl" refers to a saturated monocyclic or polycyclic hydrocarbon substituent containing 3 to 8 ring atoms, wherein one or more ring atoms is selected from nitrogen, oxygen, C(O), S(O)(=NH) or S(O) m (wherein m is an integer from 0 to 2), but excluding the ring portion of -OO-, -OS- or -SS-, the remaining ring atoms are carbon.

[0040] The term "alkoxy" refers to an -O-(alkyl) group, wherein alkyl is as defined above.

[0041] The term "halogen" refers to fluorine, chlorine, bromine or iodine.

[0042] The term "deuterated alkyl" refers to an alkyl group substituted with one or more deuterium groups, wherein alkyl is as defined above.

[0043] The term "haloalkyl" refers to an alkyl group substituted with one or more halogens, wherein alkyl is as defined above.

[0044] The term "haloalkoxy" refers to an alkoxy group substituted with one or more halogens, wherein alkoxy is as defined above.

[0045] The term "deuterated alkoxy" refers to an alkoxy group substituted with one or more deuterium groups, wherein alkoxy is as defined above.

[0046] The term "hydroxyalkyl" refers to an alkyl group substituted with one or more hydroxy groups, wherein alkyl is as defined above.

[0047] Different expressions such as “X is selected from A, B, or C”, “X is selected from A, B and C”, “X is A, B or C”, and “X is A, B and C” all express the same meaning, that is, X can be any one or more of A, B, and C.

[0048] "Optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not. For example, "a heterocyclic group optionally substituted with an alkyl group" means that the alkyl group may but need not be present, and that the description includes instances where the heterocyclic group is substituted with an alkyl group and instances where the heterocyclic group is not substituted with an alkyl group.

[0049] "Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms, in a group are replaced independently of one another by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and a person skilled in the art can determine (by experiment or theory) which substitutions are possible or impossible without undue effort. For example, an amino or hydroxyl group with free hydrogen may be unstable when combined with a carbon atom with an unsaturated (e.g., olefinic) bond.

[0050] A "pharmaceutical composition" refers to a mixture containing one or more compounds described herein, or their physiologically / pharmaceutically acceptable salts or prodrugs, together with other chemical components, as well as other components such as physiologically / pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitating absorption of the active ingredient and thereby exerting its biological activity.

[0051] "Pharmaceutically acceptable salts" refer to salts of the compounds of the present invention that are safe and effective when used in mammals and have the desired biological activity.

[0052] Example 1

[0053]

[0054] Compound A-1 (110 mg, 0.105 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 77 mg of compound B-1, an 89% yield. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.67(d,J=4.5Hz,1H),8. 20(s,1H),7.84(d,J=8.5Hz,1H),7.72(d,J=7.7Hz,1H),7.57(d,J=8.5Hz,2H),7.52–7.43(m,2H),7.30(d,J=8.4Hz,2H),7.19(t ,J=7.9Hz,1H),7.08(t,J=7.5Hz,1H),6.87(s,1H),6.77(d,J=7.6Hz,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.67 (s,2H),3.49(s,4H),3.35(s,4H),2.81(d,J=4.5Hz,3H),1.96–1.89(m,1H),1.30(d,J=6.8Hz,3H),0.88–0.77(m,6H).ESI-MSm / z 822.2[M+H] + .

[0055] Compound B-1 (77 mg, 0.09 mmol, 1 eq) and compound C-1 (56 mg, 0.18 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-1. Compound G cyclic peptide c(RGDfK) (54 mg, 0.09 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid separation (acetonitrile 30% → 40%) was performed to obtain 13 mg of Example 1, a 10% yield. 1HNMR(400MHz,DMSO-d6)δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2H z,1H),8.70(q,J=4.0Hz,1H),8.42(d,J=6.9Hz,1H),8.31–8.07(m,5H),8.03(d,J=7.7Hz,1H),7 .94(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.76–7.72(m,1H),7.72–7.68(m,1H),7.62(d,J=8. 5Hz,2H),7.52–7.45(m,2H),7.29(d,J=8.5Hz,2H),7.27–7.00(m,10H),6.87(t,J=2.3Hz,1H),6 .76(d,J=7.7Hz,1H),5.01(s,2H),4.64(td,J=8.9,6.1Hz,1H),4.51(d,J=6.7Hz,1H),4.42–4.2 7(m,2H),4.25–4.08(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,6H),3.09(q,J=7.0,6. 5Hz,2H),2.95(s,2H),2.81(d,J=4.5Hz,3H),2.77–2.57(m,3H),2.39(d,J=7.2Hz,1H),2.12–1. 98(m,2H),1.70–1.43(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.93–0.79(m,6H).ESI-MSm / z 1507.7[M+H] + .

[0056] Example 2

[0057]

[0058] Compound B-1 (50 mg, 0.06 mmol, 1 eq) and compound C-2 (36 mg 0.12 mmol, 2 eq) obtained in Example 1 were dissolved in 2 ml of N,N-dimethylformamide solution, and triethylamine (0.5 ml) was added dropwise. After stirring at room temperature for 12 h, the product D-2 was directly added with cyclic peptide c (RGDfK) (36 mg, 0.06 mmol, 1 eq) without separation. Stirring was continued for 12 h, and the solution was evaporated under reduced pressure. Semi-preparative liquid separation (acetonitrile 30% → 40%) was performed to obtain 10 mg of the compound of Example 2 in a yield of 10%. 1HNMR(400MHz,DMSO-d6)11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2Hz ,1H),8.70(q,J=4.0Hz,1H),8.42(d,J=6.9Hz,1H),8.31–8.07(m,5H),8.03(d,J=7.7Hz,1H),7. 94(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.76–7.72(m,1H),7.72–7.68(m,1H),7.62(d,J=8.5 Hz,2H),7.51–7.44(m,2H),7.29(d,J=8.5Hz,2H),7.26–7.01(m,10H),6.87(t,J=2.3Hz,1H),6.7 6(d,J=7.7Hz,1H),5.01(s,2H),4.63(td,J=8.9,6.1Hz,1H),4.51(d,J=6.7Hz,1H),4.42–4.27( m,2H),4.24–4.06(m,3H),3.66(s,2H),3.49(s,4H),3.40(s,2H),3.32(s,4H),3.09(q,J=7.0,6 .5Hz,2H),2.94(s,2H),2.81(d,J=4.5Hz,3H),2.78–2.56(m,3H),2.39(d,J=7.2Hz,1H),2.10–1 .96(m,2H),1.71–1.44(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.94–0.79(m,6H).ESI-MSm / z 1493.7[M+H] + .

[0059] Example 3

[0060]

[0061] Compound B-1 (50 mg, 0.06 mmol, 1 eq) and compound C-3 (43 mg 0.12 mmol, 2 eq) obtained in Example 1 were dissolved in 2 ml of N,N-dimethylformamide solution, and triethylamine (0.5 ml) was added dropwise. After stirring at room temperature for 12 h, the product D-3 was directly added with cyclic peptide c (RGDfK) (36 mg, 0.06 mmol, 1 eq) without separation. Stirring was continued for 12 h, and the solution was evaporated under reduced pressure. Semi-preparative liquid separation (acetonitrile 30% → 40%) was performed to obtain 13 mg of the compound of Example 3, with a yield of 14%. 1HNMR(400MHz,DMSO-d6)δ11.48(s,1H),11.08(s,1H),9.96(s,1H),9.06(s,1H),9.01(d,J=8.4Hz ,1H),8.67(q,J=4.3Hz,1H),8.49(d,J=8.9Hz,1H),8.31–8.13(m,5H),7.89(d,J=7.5Hz,2H),7.8 3(d,J=8.4Hz,1H),7.71(t,J=8.9Hz,2H),7.58(d,J=8.4Hz,2H),7.50–7.42(m,2H),7.29(d,J=8. 3Hz,2H),7.23–7.01(m,10H),6.87(s,1H),6.76(d,J=7.5Hz,1H),5.00(s,2H),4.65(td,J=9.0,8. 5,5.6Hz,1H),4.60–4.47(m,1H),4.40–4.30(m,2H),4.22–4.10(m,3H),3.67(s,2H),3.58(q,J=6 .1Hz,4H),3.48(s,4H),3.35(s,4H),3.09(q,J=6.9Hz,2H),3.03–2.97(m,2H),2.81(d,J=4.4Hz,3 H),2.76–2.54(m,3H),2.54(s,2H),2.42–2.36(m,1H),2.30(t,J=6.6Hz,2H),2.07–1.90(m,2H), 1.70–1.42(m,6H),1.37(s,2H),1.30(d,J=7.0Hz,3H),1.23(s,2H),0.93–0.79(m,6H).ESI-MSm / z 1551.7[M+H] + .

[0062] Example 4

[0063]

[0064] Compound B-1 (50 mg, 0.06 mmol, 1 eq) and compound C-4 (48 mg, 0.12 mmol, 2 eq) obtained in Example 1 were dissolved in 2 ml of N,N-dimethylformamide solution, and triethylamine (0.5 ml) was added dropwise. After stirring at room temperature for 12 h, the product D-4 was directly added with cyclic peptide c (RGDfK) (36 mg, 0.06 mmol, 1 eq) without separation. Stirring was continued for 12 h, and the solution was evaporated under reduced pressure. Semi-preparative liquid separation (acetonitrile 30% → 40%) was performed to obtain 17 mg of the compound of Example 4, with a yield of 17%. 11H NMR (400 MHz, DMSO-d6) 1 1H NMR (400 MHz, DMSO-d6) δ 11.48 (s, 1H), 11.08 (s, 1H), 9.96 (s, 1H), 9.06 (s, 1H), 9.01 (d, J = 8.4 Hz, 1H), 8.68 (q, J = 4.3 Hz, 1H), 8.49 (d, J = 8.9 Hz, 1H), 8.30–8.12 (m, 5H), 7.88 (d, J = 7.5 Hz, 2H), 7.84 (d, J = 8.4 Hz, 1H), 7.71 (t, J = 8.9 Hz, 2H), 7.56 (d, J = 8.4 Hz, 2H), 7.51–7.43 (m, 2H), 7.29 (d, J = 8.3 Hz, 2H), 7.23–7.00 (m, 10H), 6.87 (s, 1H), 6.76 (d, J = 7.5 Hz, 1H), 5.00 (s, 2H), 4.65 (td, J = 9.0, 8.5, 5.6 Hz, 1H), 4.60–4.47 (m, 1H), 4.41–4.32 (m, 2H), 4.24–4.12 (m, 3H), 3.67 (s, 2H), 3.58 (q, J = 6.1 Hz, 4H), 3.47 (s, 8H), 3.35 (s, 4H), 3.09 (q, J = 6.9 Hz, 2H), 3.03–2.97 (m, 2H), 2.81 (d, J = 4.4 Hz, 3H), 2.78–2.56 (m, 3H), 2.54 (s, 2H), 2.43–2.36 (m, 1H), 2.30 (t, J = 6.6 Hz, 2H), 2.08–1.91 (m, 2H), 1.70–1.43 (m, 6H), 1.37 (s, 2H), 1.30 (d, J = 7.0 Hz, 3H), 1.23 (s, 2H), 0.93–0.79 (m, 6H). ESI-MS m / z 1595.7 [M+H] + .

[0065] Example 5

[0066]

[0067] Compounds B-1 (77 mg, 0.09 mmol, 1 eq) and C-5 (80 mg 0.18 mmol, 2 eq) obtained in Example 1 were dissolved in 2 ml of N,N-dimethylformamide solution, and triethylamine (0.5 ml) was added dropwise. After stirring at room temperature for 12 h, compound D-5 was directly added without separation, and cyclic peptide c(RGDfK) (54 mg, 0.18 mmol, 1 eq) was added. Stirring was continued for 12 h, and the solution was evaporated under reduced pressure. Semi-preparative liquid separation (acetonitrile 30% → 40%) was performed to obtain 51 mg of the compound of Example 5 with a yield of 37%. 1 HNMR(400MHz,DMSO-d6)δ11.48(s,1H),11.08(s,1H),9.96(s,1H),9.06(s,1H),9.01(d,J=8.4Hz ,1H),8.69(q,J=4.3Hz,1H),8.49(d,J=8.9Hz,1H),8.31–8.13(m,5H),7.89(d,J=7.5Hz,2H),7.84 (d,J=8.4Hz,1H),7.71(t,J=8.9Hz,2H),7.58(d,J=8.4Hz,2H),7.51–7.44(m,2H),7.29(d,J=8.3 Hz,2H),7.23–7.01(m,10H),6.87(s,1H),6.76(d,J=7.5Hz,1H),5.00(s,2H),4.65(td,J=9.0,8.5 ,5.6Hz,1H),4.60–4.47(m,1H),4.41–4.32(m,2H),4.24–4.12(m,3H),3.67(s,2H),3.58(q,J=6. 1Hz,4H),3.47(s,12H),3.35(s,4H),3.09(q,J=6.9Hz,2H),3.03–2.97(m,2H),2.81(d,J=4.4Hz,3 H),2.78–2.56(m,3H),2.54(s,2H),2.43–2.36(m,1H),2.30(t,J=6.6Hz,2H),2.08–1.91(m,2H), 1.70–1.43(m,6H),1.37(s,2H),1.30(d,J=7.0Hz,3H),1.23(s,2H),0.93–0.79(m,6H).ESI-MSm / z 1639.8[M+H] + .

[0068] Example 6

[0069]

[0070] Compounds B-1 (45 mg, 0.055 mmol, 1 eq) and C-6 (54 mg 0.11 mmol, 2 eq) obtained in Example 1 were dissolved in 2 ml of N,N-dimethylformamide solution, and triethylamine (0.5 ml) was added dropwise. After stirring at room temperature for 12 h, the product D-6 was directly added without separation, and cyclic peptide c(RGDfK) (33 mg, 0.055 mmol, 1 eq) was added. Stirring was continued for 12 h, and the solution was evaporated under reduced pressure. Semi-preparative liquid separation (acetonitrile 30% → 40%) was performed to obtain 13 mg of the compound of Example 6, with a yield of 14%. 1 HNMR(400MHz,DMSO-d6)δ11.48(s,1H),11.08(s,1H),9.96(s,1H),9.07(s,1H),9.01(d,J=9.2Hz,1H ),8.69(s,1H),8.50(d,J=9.1Hz,1H),8.33–8.11(m,5H),7.89(d,J=8.2Hz,2H),7.84(d,J=9.6Hz,1H ),7.72(d,J=7.3Hz,1H),7.68(s,1H),7.58(d,J=8.0Hz,2H),7.49(d,J=7.7Hz,2H),7.29(d,J=7.7Hz ,2H),7.23–7.14(m,8H),7.10(s,1H),7.07(d,J=8.1Hz,1H),6.87(s,1H),6.76(d,J=8.7Hz,1H),5.00 (s,2H),4.70–4.61(m,1H),4.60–4.51(m,1H),4.42–4.33(m,2H),4.23–4.11(m,3H),3.67(s,2H),3. 58(q,J=6.0Hz,4H),3.48(s,16H),3.36(s,4H),3.14–3.05(m,2H),3.03–2.94(m,2H),2.80(d,J=3.5H z,3H),2.76–2.61(m,3H),2.60(s,2H),2.38(d,J=8.0Hz,1H),2.30(t,J=6.4Hz,2H),2.07–1.90(m,2 H),1.73–1.45(m,6H),1.39(s,2H),1.29(d,J=7.0Hz,3H),1.23(s,2H),0.93–0.79(m,6H).ESI-MSm / z 1683.8[M+H] + .

[0071] Example 7

[0072]

[0073] Compound A-2 (109 mg, 0.103 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 60 mg of compound B-2, a 70% yield. 1 H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.05(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.67(d,J=4.5Hz,1H),8. 20(s,1H),7.83(d,J=8.5Hz,1H),7.70(d,J=7.7Hz,1H),7.55(d,J=8.5Hz,2H),7.52–7.43(m,2H),7.30(d,J=8.4Hz,2H),7.19(t ,J=7.9Hz,1H),7.04(t,J=7.5Hz,1H),6.87(s,1H),6.75(d,J=7.6Hz,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.67 (s,2H),3.49(s,4H),3.35(s,4H),2.81(d,J=4.5Hz,3H),1.96–1.89(m,1H),1.30(d,J=6.8Hz,3H),0.88–0.77(m,6H).ESI-MSm / z 838.3[M+H] + .

[0074] Compound B-2 (60 mg, 0.07 mmol, 1 eq) and compound C-1 (43 mg, 0.14 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-7. Cyclic peptide c(RGDfK) (50 mg, 0.07 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 16 mg of Example 7, a yield of 15%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2H z,1H),8.70(q,J=4.0Hz,1H),8.41(d,J=6.9Hz,1H),8.29–8.05(m,5H),8.03(d,J=7.7Hz,1H),7 .92(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.76–7.72(m,1H),7.70–7.66(m,1H),7.61(d,J=8. 5Hz,2H),7.50–7.43(m,2H),7.27(d,J=8.5Hz,2H),7.27–7.01(m,10H),6.87(t,J=2.3Hz,1H),6 .76(d,J=7.7Hz,1H),5.01(s,2H),4.64(td,J=8.9,6.1Hz,1H),4.50(d,J=6.7Hz,1H),4.42–4.2 6(m,2H),4.26–4.07(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,6H),3.09(q,J=7.0,6. 5Hz,2H),2.95(s,2H),2.80(d,J=4.5Hz,3H),2.77–2.57(m,3H),2.39(d,J=7.2Hz,1H),2.12–1. 98(m,2H),1.70–1.43(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.93–0.79(m,6H).ESI-MSm / z 1523.7[M+H] + .

[0075] Example 8

[0076]

[0077] Compound A-3 (125 mg, 0.118 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 78 mg of compound B-3, an 80% yield. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.66(d,J=4.5Hz,1H),8. 20(s,1H),7.84(d,J=8.5Hz,1H),7.70(d,J=7.7Hz,1H),7.57(d,J=8.5Hz,2H),7.50–7.40(m,2H),7.30(d,J=8.4Hz,2H),7.19(t ,J=7.9Hz,1H),7.08(t,J=7.5Hz,1H),6.87(s,1H),6.76(d,J=7.6Hz,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.67 (s,2H),3.48(s,4H),3.35(s,4H),2.81(d,J=4.5Hz,3H),1.94–1.87(m,1H),1.30(d,J=6.8Hz,3H),0.86–0.74(m,6H).ESI-MSm / z 829.4[M+H] + .

[0078] Compound B-3 (78 mg, 0.09 mmol, 1 eq) and compound C-1 (55 mg, 0.19 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-8. Cyclic peptide c(RGDfK) (60 mg, 0.09 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 27 mg of Example 8, a yield of 19%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.03(d,J=8.2H z,1H),8.70(q,J=4.0Hz,1H),8.41(d,J=6.9Hz,1H),8.28–8.04(m,5H),8.03(d,J=7.7Hz,1H),7 .92(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.74–7.70(m,1H),7.70–7.66(m,1H),7.61(d,J=8. 5Hz,2H),7.51–7.44(m,2H),7.27(d,J=8.5Hz,2H),7.27–7.01(m,10H),6.87(t,J=2.3Hz,1H),6 .76(d,J=7.7Hz,1H),5.01(s,2H),4.63(td,J=8.8,6.0Hz,1H),4.50(d,J=6.7Hz,1H),4.43–4.2 5(m,2H),4.26–4.07(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,6H),3.09(q,J=7.0,6. 5Hz,2H),2.95(s,2H),2.82(d,J=4.5Hz,3H),2.76–2.55(m,3H),2.39(d,J=7.2Hz,1H),2.12–1. 98(m,2H),1.71–1.42(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.93–0.79(m,6H).ESI-MSm / z 1514.7[M+H] + .

[0079] Example 9

[0080]

[0081] Compound A-4 (101 mg, 0.09 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 61 mg of compound B-4, a 78% yield. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.67(d,J=4.5Hz,1H),8. 20(s,1H),7.84(d,J=8.5Hz,1H),7.71(d,J=7.6Hz,1H),7.57(d,J=8.5Hz,2H),7.51–7.42(m,2H),7.29(d,J=8.4Hz,2H),7.19(t ,J=7.9Hz,1H),7.06(t,J=7.3Hz,1H),6.87(s,1H),6.78(d,J=7.4Hz,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.67 (s,2H),3.49(s,4H),3.34(s,4H),2.81(d,J=4.5Hz,3H),1.96–1.89(m,1H),1.30(d,J=6.8Hz,3H),0.89–0.78(m,6H).ESI-MSm / z 872.3[M+H] + .

[0082] Compound B-4 (61 mg, 0.07 mmol, 1 eq) and compound C-1 (43 mg, 0.14 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-9. Cyclic peptide c(RGDfK) (50 mg, 0.07 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 19 mg of Example 9, with a yield of 18%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2H z,1H),8.70(q,J=4.0Hz,1H),8.41(d,J=6.9Hz,1H),8.29–8.05(m,5H),8.03(d,J=7.7Hz,1H),7 .92(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.75–7.70(m,1H),7.68–7.63(m,1H),7.61(d,J=8. 5Hz,2H),7.51–7.44(m,2H),7.27(d,J=8.5Hz,2H),7.26–7.00(m,10H),6.86(t,J=2.3Hz,1H),6 .76(d,J=7.7Hz,1H),5.01(s,2H),4.64(td,J=8.9,6.1Hz,1H),4.50(d,J=6.7Hz,1H),4.41–4.2 5(m,2H),4.26–4.07(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,6H),3.08(q,J=7.0,6. 5Hz,2H),2.95(s,2H),2.80(d,J=4.5Hz,3H),2.75–2.56(m,3H),2.38(d,J=7.2Hz,1H),2.11–1. 98(m,2H),1.69–1.42(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.93–0.79(m,6H).ESI-MSm / z 1557.7[M+H] + .

[0083] Example 10

[0084]

[0085] Compound A-5 (111 mg, 0.105 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 72 mg of compound B-5, with a yield of 83%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.66(d,J=4.5Hz,1H),8. 20(s,1H),7.84(d,J=8.5Hz,1H),7.71(d,J=7.7Hz,1H),7.57(d,J=8.5Hz,2H),7.53–7.42(m,2H),7.30(d,J=8.4Hz,2H),7.19(t ,J=7.8Hz,1H),7.08(t,J=7.5Hz,1H),6.86(s,1H),6.77(d,J=7.6Hz,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.67 (s,2H),3.48(s,4H),3.35(s,4H),2.81(d,J=4.5Hz,3H),1.96–1.89(m,1H),1.30(d,J=6.8Hz,3H),0.90–0.76(m,6H).ESI-MSm / z 836.4[M+H] + .

[0086] Compound B-5 (72 mg, 0.09 mmol, 1 eq) and compound C-1 (56 mg, 0.18 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-10. Cyclic peptide c(RGDfK) (54 mg, 0.09 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 27 mg of Example 10, with a yield of 20%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2H z,1H),8.70(q,J=4.0Hz,1H),8.40(d,J=6.9Hz,1H),8.29–8.05(m,5H),8.03(d,J=7.7Hz,1H),7 .91(t,J=5.4Hz,1H),7.83(d,J=8.1Hz,1H),7.76–7.71(m,1H),7.70–7.66(m,1H),7.61(d,J=8. 5Hz,2H),7.50–7.43(m,2H),7.27(d,J=8.5Hz,2H),7.27–7.01(m,10H),6.87(t,J=2.3Hz,1H),6 .76(d,J=7.7Hz,1H),5.01(s,2H),4.64(td,J=8.9,6.1Hz,1H),4.50(d,J=6.7Hz,1H),4.42–4.2 6(m,2H),4.23–4.06(m,3H),3.67(s,2H),3.48(d,J=9.9Hz,6H),3.32(s,6H),3.08(q,J=7.0,6. 5Hz,2H),2.94(s,2H),2.80(d,J=4.5Hz,3H),2.77–2.57(m,3H),2.39(d,J=7.2Hz,1H),2.11–1. 98(m,2H),1.70–1.43(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.92–0.79(m,6H).ESI-MSm / z 1521.7[M+H] + .

[0087] Example 11

[0088]

[0089] Compound A-6 (113 mg, 0.106 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 72 mg of compound B-6, an 80% yield. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.67(d,J=4.5Hz,1 H),8.20(s,1H),7.85(d,J=8.5Hz,1H),7.73(d,J=7.7Hz,1H),7.57(d,J=8.5Hz,2H),7.53–7.45(m,2H),7.30(d,J=8.4Hz,2 H),7.19(t,J=7.9Hz,1H),7.09(t,J=7.5Hz,1H),6.87(s,1H),5.02(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.68(s,2H ),3.49(s,4H),3.35(s,4H),2.81(d,J=4.5Hz,3H),1.97–1.88(m,1H),1.30(d,J=6.8Hz,3H),0.89–0.77(m,6H).ESI-MSm / z 856.3[M+H] + .

[0090] Compound B-6 (72 mg, 0.08 mmol, 1 eq) and compound C-1 (54 mg, 0.16 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-11. Cyclic peptide c(RGDfK) (54 mg, 0.08 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 22 mg of Example 11, with a yield of 18%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.09(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8. 2Hz,1H),8.70(q,J=4.0Hz,1H),8.41(d,J=6.9Hz,1H),8.32–8.08(m,5H),8.04(d,J=7.7Hz,1 H),7.91(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.76–7.70(m,1H),7.70–7.66(m,1H),7.61 (d,J=8.5Hz,2H),7.50–7.43(m,2H),7.27(d,J=8.5Hz,2H),7.27–7.01(m,10H),6.87(t,J=2. 3Hz,1H),5.01(s,2H),4.63(td,J=8.9,6.1Hz,1H),4.50(d,J=6.7Hz,1H),4.42–4.26(m,2H) ,4.26–4.06(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,6H),3.08(q,J=7.0,6.3Hz,2 H),2.95(s,2H),2.80(d,J=4.5Hz,3H),2.77–2.56(m,3H),2.39(d,J=7.2Hz,1H),2.12–1.98 (m,2H),1.70–1.43(m,6H),1.32(d,J=7.1Hz,3H),1.23(s,4H),0.92–0.79(m,6H).ESI-MSm / z 1541.6[M+H] + .

[0091] Example 12

[0092]

[0093] Compound A-7 (98 mg, 0.09 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 65 mg of compound B-7, an 85% yield. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.07(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.67(d,J=4.5Hz,1H), 8.20(s,1H),7.84(d,J=8.5Hz,1H),7.72(d,J=7.7Hz,1H),7.57(d,J=8.5Hz,2H),7.51–7.42(m,2H),7.30(d,J=8.4Hz,2H),7. 18(t,J=7.9Hz,1H),7.08(t,J=7.5Hz,1H),6.87(s,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.66(s,2H),3.49(s ,4H),3.35(s,4H),2.81(d,J=4.5Hz,3H),2.30(s,3H),1.96–1.89(m,1H),1.30(d,J=6.8Hz,3H),0.88–0.77(m,6H).ESI-MSm / z 836.4[M+H] + .

[0094] Compound B-7 (65 mg, 0.08 mmol, 1 eq) and compound C-1 (54 mg, 0.16 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-12. Cyclic peptide c(RGDfK) (54 mg, 0.08 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 16 mg of Example 12, with a yield of 14%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2 Hz,1H),8.70(q,J=4.0Hz,1H),8.40(d,J=6.9Hz,1H),8.29–8.05(m,5H),8.03(d,J=7.7Hz,1H) ,7.92(t,J=5.5Hz,1H),7.83(d,J=8.0Hz,1H),7.75–7.72(m,1H),7.71–7.67(m,1H),7.61(d,J =8.5Hz,2H),7.50–7.43(m,2H),7.27(d,J=8.5Hz,2H),7.28–7.04(m,10H),6.87(t,J=2.3Hz,1 H),5.01(s,2H),4.60(td,J=8.9,6.1Hz,1H),4.50(d,J=6.6Hz,1H),4.44–4.28(m,2H),4.26–4 .07(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,6H),3.08(q,J=7.0,6.5Hz,2H),2.95( s,2H),2.80(d,J=4.5Hz,3H),2.76–2.58(m,3H),2.39(d,J=7.2Hz,1H),2.29(s,3H),2.12–1.9 8(m,2H),1.70–1.44(m,6H),1.34(d,J=7.2Hz,3H),1.23(s,4H),0.95–0.79(m,6H).ESI-MSm / z 1521.7[M+H] + .

[0095] Example 13

[0096]

[0097] Compound A-8 (113 mg, 0.108 mmol, 1 eq) was dissolved in 2 ml of 20% piperidine in N,N-dimethylformamide and stirred at room temperature. The reaction was terminated after 3 h, concentrated under reduced pressure, extracted with dichloromethane and saturated brine, and dried over anhydrous sodium sulfate. Column chromatography (dichloromethane:methanol = 20:1) afforded 72 mg of compound B-8, an 81% yield. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.06(s,1H),10.06(s,1H),9.05(s,1H),9.01(d,J=8.4Hz,1H),8.67(d,J=4.5Hz,1H),8. 21(s,1H),7.83(d,J=8.5Hz,1H),7.72(d,J=7.6Hz,1H),7.56(d,J=8.5Hz,2H),7.52–7.44(m,2H),7.30(d,J=8.4Hz,2H),7.18(t ,J=7.9Hz,1H),7.08(t,J=7.5Hz,1H),6.86(s,1H),6.76(d,J=7.6Hz,1H),5.01(s,2H),4.46(s,1H),3.97(t,J=6.7Hz,1H),3.66 (s,2H),3.49(s,4H),3.35(s,5H),2.81(d,J=4.5Hz,3H),1.96–1.89(m,1H),1.30(d,J=6.8Hz,3H),0.86–0.76(m,6H).ESI-MSm / z 821.4[M+H] + .

[0098] Compound B-8 (72 mg, 0.09 mmol, 1 eq) and compound C-1 (57 mg, 0.18 mmol, 2 eq) were dissolved in 2 ml of N,N-dimethylformamide solution. Triethylamine (0.5 ml) was added dropwise and stirred at room temperature for 12 h to obtain compound D-13. Cyclic peptide c(RGDfK) (59 mg, 0.09 mmol, 1 eq) was added directly without separation and stirring was continued for 12 h. The solution was evaporated under reduced pressure and semi-preparative liquid phase separation (acetonitrile 30% → 40%) was performed to obtain 22 mg of Example 13, with a yield of 17%. 1H NMR (400MHz, DMSO-d6) δ11.48(s,1H),11.08(s,1H),9.69(s,1H),9.07(s,1H),9.01(d,J=8.2H z,1H),8.70(q,J=4.0Hz,1H),8.41(d,J=6.9Hz,1H),8.28–8.04(m,5H),8.03(d,J=7.7Hz,1H),7 .92(t,J=5.5Hz,1H),7.84(d,J=8.1Hz,1H),7.75–7.71(m,1H),7.70–7.66(m,1H),7.61(d,J=8. 5Hz,2H),7.51–7.42(m,2H),7.27(d,J=8.5Hz,2H),7.27–7.01(m,10H),6.86(t,J=2.2Hz,1H),6 .76(d,J=7.7Hz,1H),5.01(s,2H),4.64(td,J=8.7,6.1Hz,1H),4.50(d,J=6.7Hz,1H),4.41–4.2 5(m,2H),4.26–4.07(m,3H),3.67(s,2H),3.49(d,J=9.9Hz,6H),3.32(s,7H),3.09(q,J=7.0,6. 5Hz,2H),2.95(s,2H),2.80(d,J=4.5Hz,3H),2.76–2.56(m,3H),2.39(d,J=7.2Hz,1H),2.12–1. 98(m,2H),1.70–1.42(m,6H),1.33(d,J=7.1Hz,3H),1.23(s,4H),0.93–0.79(m,6H).ESI-MSm / z 1506.7[M+H] + .

[0099] Effect example 1: in vitro enzymatic release experiment

[0100] Experimental method: (1) Drawing of standard curve of test compound: accurately weigh about 1 mg of the standard of test compound, dissolve it in DMSO to prepare a 20 mg / mL solution, and then dilute it with chromatographic grade methanol to 6 concentrations (200 μg / mL, 100 μg / mL, 50 μg / mL, 25 μg / mL, 12.5 μg / mL, 6.25 μg / mL). After filtering with a 0.22 μm ultrafiltration membrane, the solution was tested on an instrument. The area under the curve (Area) was used as the abscissa, represented by x, and the concentration of the test compound (μg / mL) was used as the ordinate, represented by y, to draw the standard curve of each compound; (2) weigh an appropriate amount of the compound of Example 1, the compound of Example 2, and the compound of Example 3 in 50 μl of DMSO to obtain a 10 mM stock solution respectively; (3) prepare 10 ml of buffer solution, the components of which are: 50 mM sodium acetate, 100 mM sodium chloride, 4 mM EDTA, 8 mM L-cysteine, the pH after preparation is 5-5.4; (4) Take an appropriate amount of prepared buffer and dissolve cathepsin B to obtain a 5U / ml cathepsin B buffer solution, incubate and activate at 37℃ for 15 minutes for use; (5) Taking Example 1 as an example: take 5μl of the mother solution + 45μl of the cathepsin B buffer solution and mix them to obtain a test solution with a final concentration of 1mM, incubate at 37℃. This experiment was carried out in parallel for 8 groups, and 100μL of 1% HCOOH methanol solution was added at 0, 0.25, 0.5, 1, 2, 4, 6, and 24 hours, and placed in a -20℃ refrigerator to quench the reaction; (6) The sample was centrifuged at 12000rpm at 4℃ for 10 minutes, filtered through a mixed cellulose filter, and analyzed by HPLC. The compound content at each time point was obtained by referring to the standard curve. (7) Data was collected and the curve was generated by Graphpad.

[0101] The experimental results showed that the tested compounds could be effectively released (e.g. Figure 1 shown)

[0102] Effect Example 2: Tumor Cell Growth Inhibition Experiment

[0103] Experimental Methods: Tumor cells were routinely cultured in a sterile incubator at 37°C and 5% CO2 saturated humidity using DMEM supplemented with 10% fetal bovine serum (FBS). After the experiment began, cells in the logarithmic growth phase were digested with 0.25% trypsin, centrifuged, and culture medium was added to adjust the cell concentration to 50,000 cells / mL. The cells were then inoculated into a 96-well culture plate with 100 μL of cell suspension per well and cultured overnight in an incubator to allow the cells to adhere. The drug to be tested was then added to the wells at different concentrations, with a concentration gradient of 100 μM, 50 μM, 20 μM, 10 μM, 1 μM, and 0.1 μM, with four replicates per well. A negative control without drug addition was also included. The drug-treated cells were cultured in an incubator at 37°C and 5% CO2 saturated humidity. After 48 h, 10 μL of a 5 mg / mL MTT solution was added to each well and cultured for another 4 h. After the reaction is complete and formazan crystals are generated, the culture medium is aspirated and 150 μL of DMSO is added to each well to dissolve the formazan. After thorough mixing, the absorbance OD value of each well at a wavelength of 490 nM is measured using a microplate reader. The cell proliferation inhibition rate (%) is calculated according to the formula [(OD 阴性 -OD 给药 ) / (OD 阴性 -OD 调零 )]*100% to calculate the proliferation inhibition rate of each concentration, and then processed with Graphpad Prism 5 software to obtain the IC 50 value.

[0104] The experimental results showed that the compound significantly inhibited the growth of tumor cells (as shown in Table 1)

[0105] Table 1. Half-maximal inhibitory concentration (IC50) of the compounds of Examples 1-13 for inhibiting tumor cell proliferation

[0106]

[0107] The present invention also conducted an inhibition experiment on MDA-MB-231 tumor cells on the compounds of Example 1 and Example 2. The results showed that both the compounds of Example 1 and Example 2 showed significant tumor growth inhibition activity in mice. At a concentration of 10 mg / kg, the tumor inhibition effect of the compound of Example 1 was superior to that of the positive control paclitaxel at the same dosage, and no obvious toxicity was observed.

[0108] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art should fall within the scope of protection defined by the claims of the present invention.

Claims

1. A compound represented by formula (I), a stereoisomer thereof, or a pharmaceutically acceptable salt thereof; In formula (I), L is selected from methylene, ethylene or X is selected from CH or N; R 1 is selected from hydrogen, deuterium, fluorine, chlorine, nitro, cyano, C1-C3 alkyl or C1-C3 haloalkyl; R 2 is selected from hydrogen, deuterium, C1-C3 alkyl, C1-C3 deuterated alkyl or C3-C6 cycloalkyl; R 3 is selected from hydrogen, deuterium, halogen, nitro, cyano, C1-C3 alkyl, C1-C3 haloalkyl, C1-C3 deuterated alkyl, C1-C3 alkoxy, C1-C3 haloalkoxy or C1-C3 deuterated alkoxy.

2. The compound according to claim 1, wherein One of the following compounds:

3. Use of the compound according to claim 1 or 2, its stereoisomer or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating integrin receptor-mediated diseases.

4. The use according to claim 3, characterized in that The integrin receptor-mediated related diseases are ovarian cancer, breast cancer, hepatocellular carcinoma, bladder cancer, colorectal cancer, lung cancer, thyroid cancer, prostate cancer, melanoma, osteosarcoma or glioblastoma.

5. A drug for treating integrin receptor-mediated diseases, characterized in that: The active ingredient includes the compound according to claim 1 or 2, its stereoisomer or a pharmaceutically acceptable salt thereof.

6. The drug for treating integrin receptor-mediated diseases according to claim 5, characterized in that: Pharmaceutically acceptable excipients are also included.

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