Tumor targeting polypeptide, conjugate containing same, composition and application

Abstract

The present disclosure relates to tumor targeting polypeptides, conjugates and compositions comprising the same for use in treating tumors or reducing tumor volume.

Description

Technical Field

[0001] The present disclosure relates to the fields of medicinal chemistry and disease treatment. In particular, the present disclosure relates to tumor targeting polypeptides, conjugates, compositions and uses thereof. Background Art

[0002] SSTR2 (Somatostatin Receptor 2) is a somatostatin receptor that is overexpressed or highly expressed in many solid tumors, such as neuroendocrine tumors (NET), small cell lung cancer, glioma, pancreatic cancer, breast cancer, etc. At present, the treatment methods for these solid tumors are mainly surgical resection combined with radiotherapy and chemotherapy, the clinically available drugs are limited, and secondary drug resistance is very likely to occur, and recurrence is easy.

[0003] Kerry et al. disclosed in "Targeting the Somatostatin Receptor 2 with the Miniaturized Drug Conjugate, PEN-221: A Potent and Novel Therapeutic for the Treatment of Small Cell Lung Cancer", Molecular Cancer Therapeutics, 18(11), November 2019, pp. 1926-1936, that the peptide-drug conjugate (PDC) PEN-221 is used to treat tumors overexpressing SSTR2. PEN-221 is obtained by conjugating octreotide (OCT) to maytansine DM1 (Mertansine) via a linker. Octreotide is a growth inhibitor that can specifically target SSTR2 and induce endocytosis, thereby acting as a homing peptide. Maytansine DM1 is a highly cytotoxic compound with good stability and water solubility. It can inhibit the depolymerization of microtubules by affecting microtubules and tubulin, thereby blocking the formation of spindles during mitosis and achieving the effect of inhibiting cell proliferation.

[0004] However, there is still a need for alternative drugs that can specifically recognize and kill tumor cells with high SSTR2 expression. Summary of the invention

[0005] In order to solve the above problems, the inventors conducted in-depth research and proposed the technical solution of the present disclosure. Specifically, the inventors synthesized and screened a specific OCT analog, connected it to Maytansine DM1 through a specific linker, and the obtained PDC conjugate can specifically recognize tumor cells with high or overexpression of SSTR2 and enter the tumor cells through endocytosis. In other words, the PDC conjugate according to the present disclosure can specifically recognize and target the elimination of tumor cells with high or overexpression of SSTR2. At the same time, the PDC conjugate according to the present disclosure has excellent plasma stability.

[0006] In one aspect, the present disclosure provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of Formula I:

[0007]

[0008] Among them, R 1 To R 7 , and R 15 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkylcarbonyl, halogen, cyano, hydroxyl, carboxyl, -C(O)NR 10 R 11 , phenyl, naphthyl, biphenyl, Polyethylene glycol, and C5-C containing one or more heteroatoms selected from N, S and O 10 Aromatic heterocyclic ring; or R 15 Condensed with adjacent phenyl groups to form naphthyl groups; these groups are optionally substituted by one or more substituents R 12 replace;

[0009] R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and

[0010] R 9 With R 16 are the same or different and are independently selected from S, -CH2- and -C(CH3)2-; or R 9 With R 16 Together they represent -SS-, -S-CH2-, -S-CH(CH3)-, -SC(CH3)2-, -CH2-S-, -CH(CH3)-S-, -C(CH3)2-S-, -CH2-CH=CH-CH2- and as well as

[0011] R 10 To R 14 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, halogen, cyano, hydroxy, carboxyl, phenyl, and naphthyl;

[0012] R 17 express or biphenyl; or R 4 With R 17 Together with the -N-CH-CH2- in between, it forms

[0013] p represents an integer selected from 1 to 4; and

[0014] * indicates the point of attachment of the identified group to the rest of Formula I.

[0015] In another aspect, the present disclosure provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of Formula II:

[0016]

[0017] Among them, the group R 1 To R17 , and p has the same meaning as given above for the compounds of formula I,

[0018] X represents a linking moiety and is selected from one of the following groups or any combination thereof:

[0019] -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20 Cycloalkyl and its isomers, phenyl, naphthyl, maleimide groups and their hydrides, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl,

[0020] ** indicates that the identified group is connected to a leaving group.

[0021] In another aspect, the present disclosure provides a conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate has a structure of Formula III:

[0022]

[0023] Among them, the group R 1 To R 17 , and p has the same definition as given above for the compound of general formula I, X represents a linking portion and is selected from one of the following groups or any combination thereof:

[0024] -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20Cycloalkyl and its isomers, phenyl, naphthyl, maleimide group and its hydride, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine, histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl; and

[0025] Y represents an active moiety and is maytansine (DM1),

[0026] ** indicates the connection point between the identified group and Y.

[0027] In another aspect, the present disclosure provides a pharmaceutical composition comprising the conjugate according to the present disclosure or a pharmaceutically acceptable salt thereof, and

[0028] A pharmaceutically acceptable carrier, diluent or excipient.

[0029] In yet another aspect, the present disclosure provides a method for treating a tumor or reducing tumor volume, comprising administering an effective amount of the pharmaceutical composition according to the present disclosure, or the conjugate according to the present disclosure or a pharmaceutically acceptable salt thereof to an individual in need thereof. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1A-1 is a mass spectrum of compound A-1 according to the present disclosure.

[0031] Figure 1A-2 is a mass spectrum of compound A-2 according to the present disclosure.

[0032] Figure 1A-3 is a mass spectrum of compound A-3 according to the present disclosure.

[0033] Figure 1A-4 is a mass spectrum of compound A-4 according to the present disclosure.

[0034] Figure 1A-5 is a mass spectrum of compound A-5 according to the present disclosure.

[0035] Figure 1A-6 is a mass spectrum of compound A-6 according to the present disclosure.

[0036] Figure 1A-7 is a mass spectrum of compound A-7 according to the present disclosure.

[0037] Figure 1A-8 is a mass spectrum of compound A-8 according to the present disclosure.

[0038] Figure 1A-9is a mass spectrum of compound A-9 according to the present disclosure.

[0039] Figure 1A-10 is a mass spectrum of compound A-11 according to the present disclosure.

[0040] Figure 1A-11 is a mass spectrum of compound A-12 according to the present disclosure.

[0041] Figure 1A-12 is a mass spectrum of compound A-13 according to the present disclosure.

[0042] Figure 1A-13 is a mass spectrum of compound A-14 according to the present disclosure.

[0043] Figure 1A-14 is a mass spectrum of compound A-15 according to the present disclosure.

[0044] Figure 1A-15 is a mass spectrum of compound A-17 according to the present disclosure.

[0045] Figure 1A-16 is a mass spectrum of compound A-18 according to the present disclosure.

[0046] Figure 1B-1 is a mass spectrum of compound B-1 according to the present disclosure.

[0047] Figure 1B-2 is a mass spectrum of compound B-2 according to the present disclosure.

[0048] Figure 1B-3 is a mass spectrum of compound B-3 according to the present disclosure.

[0049] Figure 1B-4 is a mass spectrum of compound B-4 according to the present disclosure.

[0050] Figure 1B-5 is a mass spectrum of compound B-5 according to the present disclosure.

[0051] Figure 1B-6 is a mass spectrum of compound B-6 according to the present disclosure.

[0052] Figure 1B-7 is a mass spectrum of compound B-7 according to the present disclosure.

[0053] Figure 1B-8 is a mass spectrum of compound B-8 according to the present disclosure.

[0054] Figure 1B-9 is a mass spectrum of compound B-9 according to the present disclosure.

[0055] Figure 1B-10 is a mass spectrum of compound B-10 according to the present disclosure.

[0056] Figure 1B-11 is a mass spectrum of compound B-11 according to the present disclosure.

[0057] Figure 1B-12 is a mass spectrum of compound B-12 according to the present disclosure.

[0058] Figure 1B-13 is a mass spectrum of compound B-13 according to the present disclosure.

[0059] Figure 1B-14 is a mass spectrum of compound B-14 according to the present disclosure.

[0060] Figure 1B-15 is a mass spectrum of compound B-15 according to the present disclosure.

[0061] Figure 1B-16 is a mass spectrum of compound B-16 according to the present disclosure.

[0062] Figure 1B-17 is a mass spectrum of compound B-17 according to the present disclosure.

[0063] Figure 1B-18 is a mass spectrum of compound B-18 according to the present disclosure.

[0064] Figure 1B-19 is a mass spectrum of compound B-19 according to the present disclosure.

[0065] Figure 1B-20 is a mass spectrum of compound B-20 according to the present disclosure.

[0066] Figure 1B-21 is a mass spectrum of compound B-21 according to the present disclosure.

[0067] Figure 1B-22 is a mass spectrum of compound B-22 according to the present disclosure.

[0068] Figure 1B-23 is a mass spectrum of compound B-23 according to the present disclosure.

[0069] Figure 1C-1 is a mass spectrum of the conjugate DM1-A-1 according to the present disclosure.

[0070] Figure 1C-2 is a mass spectrum of the conjugate DM1-A-2 according to the present disclosure.

[0071] Figure 1C-3 is a mass spectrum of the conjugate DM1-A-3 according to the present disclosure.

[0072] Figure 1C-4 is a mass spectrum of the conjugate DM1-A-4 according to the present disclosure.

[0073] Figure 1C-5 is a mass spectrum of the conjugate DM1-A-5 according to the present disclosure.

[0074] Figure 1C-6 is a mass spectrum of the conjugate DM1-A-6 according to the present disclosure.

[0075] Figure 1C-7 is a mass spectrum of the conjugate DM1-A-7 according to the present disclosure.

[0076] Figure 1C-8 is a mass spectrum of the conjugate DM1-A-8 according to the present disclosure.

[0077] Figure 1C-9 is a mass spectrum of the conjugate DM1-A-9 according to the present disclosure.

[0078] Figure 1C-10 is a mass spectrum of the conjugate DM1-A-11 according to the present disclosure.

[0079] Figure 1C-11 is a mass spectrum of the conjugate DM1-A-15 according to the present disclosure.

[0080] Figure 1C-12 is a mass spectrum of the conjugate DM1-A-17 according to the present disclosure.

[0081] Figure 1C-13 is a mass spectrum of the conjugate DM1-A-18 according to the present disclosure.

[0082] Figure 1C-14 is a mass spectrum of the conjugate DM1-B-1 according to the present disclosure.

[0083] Figure 1C-15 is a mass spectrum of the conjugate DM1-B-2 according to the present disclosure.

[0084] Figure 1C-16 is a mass spectrum of the conjugate DM1-B-3 according to the present disclosure.

[0085] Figure 1C-17 is a mass spectrum of the conjugate DM1-B-5 according to the present disclosure.

[0086] Figure 1C-18 is a mass spectrum of the conjugate DM1-B-6 according to the present disclosure.

[0087] Figure 1C-19 is a mass spectrum of the conjugate DM1-B-7 according to the present disclosure.

[0088] Figure 2A-1 is a high performance liquid chromatogram of compound A-1 according to the present disclosure.

[0089] Figure 2A-2 is a high performance liquid chromatogram of compound A-2 according to the present disclosure.

[0090] Figure 2A-3 is a high performance liquid chromatogram of compound A-3 according to the present disclosure.

[0091] Figure 2A-4 is a high performance liquid chromatogram of compound A-4 according to the present disclosure.

[0092] Figure 2A-5 is a high performance liquid chromatogram of compound A-5 according to the present disclosure.

[0093] Figure 2A-6 is a high performance liquid chromatogram of compound A-6 according to the present disclosure.

[0094] Figure 2A-7 is a high performance liquid chromatogram of compound A-7 according to the present disclosure.

[0095] Figure 2A-8 is a high performance liquid chromatogram of compound A-8 according to the present disclosure.

[0096] Figure 2A-9 is a high performance liquid chromatogram of compound A-9 according to the present disclosure.

[0097] Figure 2A-10 is a high performance liquid chromatogram of compound A-11 according to the present disclosure.

[0098] Figure 2A-11 is a high performance liquid chromatogram of compound A-12 according to the present disclosure.

[0099] Figure 2A-12 is a high performance liquid chromatogram of compound A-13 according to the present disclosure.

[0100] Figure 2A-13 is a high performance liquid chromatogram of compound A-14 according to the present disclosure.

[0101] Figure 2A-14 is a high performance liquid chromatogram of compound A-15 according to the present disclosure.

[0102] Figure 2A-15 is a high performance liquid chromatogram of compound A-17 according to the present disclosure.

[0103] Figure 2A-16 is a high performance liquid chromatogram of compound A-18 according to the present disclosure.

[0104] Figure 2B-1 is a high performance liquid chromatogram of compound B-1 according to the present disclosure.

[0105] Figure 2B-2 is a high performance liquid chromatogram of compound B-2 according to the present disclosure.

[0106] Figure 2B-3 is a high performance liquid chromatogram of compound B-3 according to the present disclosure.

[0107] Figure 2B-4 is a high performance liquid chromatogram of compound B-4 according to the present disclosure.

[0108] Figure 2B-5 is a high performance liquid chromatogram of compound B-5 according to the present disclosure.

[0109] Figure 2B-6 is a high performance liquid chromatogram of compound B-6 according to the present disclosure.

[0110] Figure 2B-7 is a high performance liquid chromatogram of compound B-7 according to the present disclosure.

[0111] Figure 2B-8 is a high performance liquid chromatogram of compound B-8 according to the present disclosure.

[0112] Figure 2B-9 is a high performance liquid chromatogram of compound B-9 according to the present disclosure.

[0113] Figure 2B-10 is a high performance liquid chromatogram of compound B-10 according to the present disclosure.

[0114] Figure 2B-11 is a high performance liquid chromatogram of compound B-11 according to the present disclosure.

[0115] Figure 2B-12 is a high performance liquid chromatogram of compound B-12 according to the present disclosure.

[0116] Figure 2B-13 is a high performance liquid chromatogram of compound B-13 according to the present disclosure.

[0117] Figure 2B-14 is a high performance liquid chromatogram of compound B-14 according to the present disclosure.

[0118] Figure 2B-15 is a high performance liquid chromatogram of compound B-15 according to the present disclosure.

[0119] Figure 2B-16 is a high performance liquid chromatogram of compound B-16 according to the present disclosure.

[0120] Figure 2B-17 is a high performance liquid chromatogram of compound B-17 according to the present disclosure.

[0121] Figure 2B-18is a high performance liquid chromatogram of compound B-18 according to the present disclosure.

[0122] Figure 2B-19 is a high performance liquid chromatogram of compound B-19 according to the present disclosure.

[0123] Figure 2B-20 is a high performance liquid chromatogram of compound B-20 according to the present disclosure.

[0124] Figure 2B-21 is a high performance liquid chromatogram of compound B-21 according to the present disclosure.

[0125] Figure 2B-22 is a high performance liquid chromatogram of compound B-22 according to the present disclosure.

[0126] Figure 2B-23 is a high performance liquid chromatogram of compound B-23 according to the present disclosure.

[0127] Figure 2C-1 is a high performance liquid chromatogram of the conjugate DM1-A-1 according to the present disclosure.

[0128] Figure 2C-2 is a high performance liquid chromatogram of the conjugate DM1-A-2 according to the present disclosure.

[0129] Figure 2C-3 is a high performance liquid chromatogram of the conjugate DM1-A-3 according to the present disclosure.

[0130] Figure 2C-4 is a high performance liquid chromatogram of the conjugate DM1-A-4 according to the present disclosure.

[0131] Figure 2C-5 is a high performance liquid chromatogram of the conjugate DM1-A-5 according to the present disclosure.

[0132] Figure 2C-6 is a high performance liquid chromatogram of the conjugate DM1-A-6 according to the present disclosure.

[0133] Figure 2C-7 is a high performance liquid chromatogram of the conjugate DM1-A-7 according to the present disclosure.

[0134] Figure 2C-8 is a high performance liquid chromatogram of the conjugate DM1-A-8 according to the present disclosure.

[0135] Figure 2C-9 is a high performance liquid chromatogram of the conjugate DM1-A-9 according to the present disclosure.

[0136] Figure 2C-10 is a high performance liquid chromatogram of the conjugate DM1-A-11 according to the present disclosure.

[0137] Figure 2C-11 is a high performance liquid chromatogram of the conjugate DM1-A-15 according to the present disclosure.

[0138] Figure 2C-12 is a high performance liquid chromatogram of the conjugate DM1-A-17 according to the present disclosure.

[0139] Figure 2C-13 is a high performance liquid chromatogram of the conjugate DM1-A-18 according to the present disclosure.

[0140] Figure 2C-14 is a high performance liquid chromatogram of the conjugate DM1-B-1 according to the present disclosure.

[0141] Figure 2C-15 is a high performance liquid chromatogram of the conjugate DM1-B-2 according to the present disclosure.

[0142] Figure 2C-16 is a high performance liquid chromatogram of the conjugate DM1-B-3 according to the present disclosure.

[0143] Figure 2C-17 is a high performance liquid chromatogram of the conjugate DM1-B-5 according to the present disclosure.

[0144] Figure 2C-18 is a high performance liquid chromatogram of the conjugate DM1-B-6 according to the present disclosure.

[0145] Figure 2C-19 is a high performance liquid chromatogram of the conjugate DM1-B-7 according to the present disclosure.

[0146] Figure 3A-1 1 is a compound efficacy-concentration change curve of compound A-1 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0147] Figure 3A-2 3 is a compound efficacy-concentration change curve of compound A-2 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0148] Figure 3A-3 3 is a compound efficacy-concentration change curve of compound A-3 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0149] Figure 3A-4 3 is a compound efficacy-concentration change curve of compound A-4 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0150] Figure 3A-53 is a compound efficacy-concentration change curve of compound A-5 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0151] Figure 3A-6 3 is a compound efficacy-concentration change curve of compound A-6 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0152] Figure 3A-7 3 is a compound efficacy-concentration change curve of compound A-7 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0153] Figure 3A-8 3 is a compound efficacy-concentration change curve of compound A-8 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0154] Figure 3A-9 3 is a compound efficacy-concentration change curve of compound A-9 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0155] Figure 3A-10 3 is a compound efficacy-concentration change curve of compound A-11 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0156] Figure 3A-11 3 is a compound efficacy-concentration change curve of compound A-12 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0157] Figure 3A-12 1 is a compound efficacy-concentration change curve of compound A-13 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0158] Figure 3A-13 3 is a compound efficacy-concentration change curve of compound A-14 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0159] Figure 3A-14 3 is a compound efficacy-concentration change curve of compound A-15 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0160] Figure 3A-15 1 is a compound efficacy-concentration change curve of compound A-17 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0161] Figure 3A-16 3 is a compound efficacy-concentration change curve of compound A-18 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0162] Figure 3B-1 3 is a compound efficacy-concentration change curve of compound B-1 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0163] Figure 3B-2 3 is a compound efficacy-concentration change curve of compound B-2 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0164] Figure 3B-3 3 is a compound efficacy-concentration change curve of compound B-3 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0165] Figure 3B-4 3 is a compound efficacy-concentration change curve of compound B-4 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0166] Figure 3B-5 3 is a compound efficacy-concentration change curve of compound B-5 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0167] Figure 3B-6 3 is a compound efficacy-concentration change curve of compound B-6 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0168] Figure 3B-7 3 is a compound efficacy-concentration change curve of compound B-7 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0169] Figure 3B-8 3 is a compound efficacy-concentration change curve of compound B-8 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0170] Figure 3B-9 3 is a compound efficacy-concentration change curve of compound B-9 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0171] Figure 3B-10 3 is a compound efficacy-concentration change curve of compound B-10 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0172] Figure 3B-11 3 is a compound efficacy-concentration change curve of compound B-11 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0173] Figure 3B-123 is a compound efficacy-concentration change curve of compound B-12 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0174] Figure 3B-13 3 is a compound efficacy-concentration change curve of compound B-13 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0175] Figure 3B-14 3 is a compound efficacy-concentration change curve of compound B-14 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0176] Figure 3B-15 3 is a compound efficacy-concentration change curve of compound B-15 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0177] Figure 3B-16 3 is a compound efficacy-concentration change curve of compound B-16 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0178] Figure 3B-17 3 is a compound efficacy-concentration change curve of compound B-17 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0179] Figure 3B-18 3 is a compound efficacy-concentration change curve of compound B-18 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0180] Figure 3B-19 3 is a compound efficacy-concentration change curve of compound B-19 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0181] Figure 3B-20 3 is a compound efficacy-concentration change curve of compound B-20 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0182] Figure 3B-21 3 is a compound efficacy-concentration change curve of compound B-21 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0183] Figure 3B-22 3 is a compound efficacy-concentration change curve of compound B-22 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0184] Figure 3B-233 is a compound efficacy-concentration change curve of compound B-23 according to the present disclosure and OCT as a control in the FLIPR calcium flux experiment.

[0185] Figure 4A-1 It is the compound efficacy-concentration change curve of compound A-2 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0186] Figure 4A-2 The compound efficacy-concentration change curve of compound A-3 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0187] Figure 4A-3 The compound efficacy-concentration change curve of compound A-4 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0188] Figure 4A-4 It is the compound efficacy-concentration change curve of compound A-5 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0189] Figure 4A-5 The compound efficacy-concentration change curve of compound A-6 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0190] Figure 4A-6 The compound efficacy-concentration change curve of compound A-7 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0191] Figure 4A-7 The compound efficacy-concentration change curve of compound A-8 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0192] Figure 4A-8 The compound efficacy-concentration change curve of compound A-9 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0193] Figure 4A-9 The compound efficacy-concentration change curve of the compound A-11 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0194] Figure 4A-10 The compound efficacy-concentration change curve of the compound A-12 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0195] Figure 4A-11 The compound efficacy-concentration change curve of the compound A-13 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0196] Figure 4A-12 The compound efficacy-concentration change curve of compound A-14 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0197] Figure 4A-13 The compound efficacy-concentration change curve of the compound A-15 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0198] Figure 4A-14 The compound efficacy-concentration change curve of the compound A-17 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0199] Figure 4A-15 The compound efficacy-concentration change curve of the compound A-18 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0200] Figure 4B-1 The compound efficacy-concentration change curve of the compound B-3 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0201] Figure 4B-2 The compound efficacy-concentration change curve of the compound B-4 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0202] Figure 4B-3 It is the compound efficacy-concentration change curve of compound B-5 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0203] Figure 4B-4 The compound efficacy-concentration change curve of the compound B-6 disclosed herein and OCT as a control in the Nanobit experiment.

[0204] Figure 4B-5 The compound efficacy-concentration change curve of the compound B-7 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0205] Figure 4B-6 The compound efficacy-concentration change curve of the compound B-8 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0206] Figure 4B-7 The compound efficacy-concentration change curve of the compound B-9 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0207] Figure 4B-8The compound efficacy-concentration change curve of the compound B-10 disclosed herein and OCT as a control in the Nanobit experiment.

[0208] Figure 4B-9 The compound efficacy-concentration change curve of the compound B-11 disclosed herein and OCT as a control in the Nanobit experiment.

[0209] Figure 4B-10 The compound efficacy-concentration change curve of the compound B-12 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0210] Figure 4B-11 The compound efficacy-concentration change curve of the compound B-13 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0211] Figure 4B-12 The compound efficacy-concentration change curve of the compound B-14 disclosed herein and OCT as a control in the Nanobit experiment.

[0212] Figure 4B-13 The compound efficacy-concentration change curve of the compound B-15 disclosed herein and OCT as a control in the Nanobit experiment.

[0213] Figure 4B-14 The compound efficacy-concentration change curve of the compound B-16 disclosed herein and OCT as a control in the Nanobit experiment.

[0214] Figure 4B-15 The compound efficacy-concentration change curve of the compound B-17 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0215] Figure 4B-16 The compound efficacy-concentration change curve of the compound B-18 disclosed herein and OCT as a control in the Nanobit experiment.

[0216] Figure 4B-17 The compound potency-concentration change curve of the compound B-19 disclosed herein and OCT as a control in the Nanobit experiment.

[0217] Figure 4B-18 The compound efficacy-concentration change curve of the compound B-20 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0218] Figure 4B-19The compound efficacy-concentration change curve of the compound B-21 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0219] Figure 4B-20 The compound efficacy-concentration change curve of the compound B-22 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0220] Figure 4B-21 The compound efficacy-concentration change curve of the compound B-23 according to the present disclosure and OCT as a control in the Nanobit experiment.

[0221] Figure 5 Western Blot experiments verified the high expression of SSTR2 in small cell lung cancer cells H524, H69 and H2171, glioma cells A172 and U87, and SSTR2 negative cells A549.

[0222] Fig. 6A The uptake intensities of compounds A-1, A-2, A-3, A-4, A-5, A-6, A-7, A-8, A-9 according to the present disclosure and blank (Blank), negative peptide 106 and OCT as controls in H524 cells and A549 cells.

[0223] Figure 6B is the uptake intensity of compounds A-12, A-13, A-15 and A-17 according to the present disclosure in H524 cells and A549 cells.

[0224] Figure 6C is the uptake intensity of compounds B-1, B-2, B-3, B-4, B-5, B-6, B-7, B-8, B-9 and B-17 according to the present disclosure in H524 cells and A549 cells.

[0225] Fig.6D is the uptake intensity of compounds B-18 and B-20 according to the present disclosure in H524 cells and A549 cells.

[0226] Fig. 7A This is the stability-time curve of OCT in rat plasma as a control.

[0227] Figure 7B is the stability-time curve of compound A-1 in rat plasma according to the present disclosure.

[0228] Figure 7C is the stability-time curve of compound A-2 in rat plasma according to the present disclosure.

[0229] Fig.7Dis the stability-time curve of compound B-1 in rat plasma according to the present disclosure.

[0230] Fig. 7E is the stability-time curve of compound B-2 in rat plasma according to the present disclosure.

[0231] Figure 8A-1 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-1 according to the present disclosure and DM1 as a control in a killing experiment on A549 cells.

[0232] Figure 8A-2 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-2 according to the present disclosure and DM1 as a control in a killing experiment on A549 cells.

[0233] Figure 8A-3 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-1 according to the present disclosure and DM1 as a control in a killing experiment on A549 cells.

[0234] Figure 8A-4 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-2 according to the present disclosure and DM1 as a control in a killing experiment on A549 cells.

[0235] Figure 8B-1 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-1 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0236] Figure 8B-2 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-2 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0237] Figure 8B-3 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-3 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0238] Figure 8B-4 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-4 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0239] Figure 8B-5 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-5 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0240] Figure 8B-6The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-6 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0241] Figure 8B-7 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-7 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0242] Figure 8B-8 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-8 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0243] Figure 8B-9 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-9 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0244] Figure 8B-10 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-11 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0245] Figure 8B-11 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-15 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0246] Figure 8B-12 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-17 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0247] Figure 8B-13 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-18 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0248] Figure 8B-14 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-1 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0249] Figure 8B-15 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-2 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0250] Figure 8B-16 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-3 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0251] Figure 8B-17 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-5 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0252] Figure 8B-18 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-6 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0253] Figure 8B-19 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-7 according to the present disclosure and DM1 as a control in a killing experiment on H524 cells.

[0254] Figure 8C-1 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-1 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0255] Figure 8C-2 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-2 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0256] Figure 8C-3 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-3 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0257] Figure 8C-4 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-4 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0258] Figure 8C-5 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-5 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0259] Figure 8C-6 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-6 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0260] Figure 8C-7 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-7 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0261] Figure 8C-8 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-8 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0262] Figure 8C-9 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-9 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0263] Figure 8C-10 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-11 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0264] Figure 8C-11 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-15 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0265] Figure 8C-12 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-17 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0266] Figure 8C-13 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-18 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0267] Figure 8C-14 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-1 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0268] Figure 8C-15 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-2 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0269] Figure 8C-16 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-3 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0270] Figure 8C-17 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-5 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0271] Figure 8C-18 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-6 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0272] Figure 8C-19The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-7 according to the present disclosure and DM1 as a control in a killing experiment on H69 cells.

[0273] Figure 8D-1 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-1 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0274] Figure 8D-2 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-2 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0275] Figure 8D-3 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-3 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0276] Figure 8D-4 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-4 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0277] Figure 8D-5 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-6 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0278] Figure 8D-6 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-7 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0279] Figure 8D-7 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-8 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0280] Figure 8D-8 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-9 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0281] Figure 8D-9 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-17 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0282] Figure 8D-10 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-1 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0283] Figure 8D-11 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-2 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0284] Figure 8D-12 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-3 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0285] Figure 8D-13 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-5 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0286] Figure 8D-14 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-6 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0287] Figure 8D-15 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-7 according to the present disclosure and DM1 as a control in a killing experiment on A172 cells.

[0288] Figure 8E-1 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-1 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0289] Figure 8E-2 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-2 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0290] Figure 8E-3 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-3 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0291] Figure 8E-4 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-4 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0292] Figure 8E-5 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-6 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0293] Figure 8E-6The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-7 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0294] Figure 8E-7 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-8 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0295] Figure 8E-8 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-9 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0296] Figure 8E-9 The figure is a cell survival rate-concentration change curve of the conjugate DM1-A-17 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0297] Figure 8E-10 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-1 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0298] Figure 8E-11 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-2 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0299] Figure 8E-12 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-3 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0300] Figure 8E-13 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-5 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0301] Figure 8E-14 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-6 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0302] Figure 8E-15 The figure is a cell survival rate-concentration change curve of the conjugate DM1-B-7 according to the present disclosure and DM1 as a control in a killing experiment on U87 cells.

[0303] Fig.9A The figure is a curve of body weight change versus days after administration of the conjugates DM1-A-1, DM1-A-2, DM1-B-2 and DM1 according to the present disclosure and a vehicle control in a drug efficacy test in a mouse H524 xenograft model.

[0304] Fig. 9B The figure is a curve of body weight change rate versus days after administration of the conjugates DM1-A-1, DM1-A-2, DM1-B-2 and DM1 according to the present disclosure and a vehicle control in a drug efficacy test in a mouse H524 xenograft model.

[0305] Fig. 10A The graph is a curve of the change of tumor volume versus days after administration in the efficacy test of the conjugates DM1-A-1, DM1-A-2, DM1-B-2 and DM1 according to the present disclosure and the vehicle control in the mouse H524 xenograft model.

[0306] Fig. 10B The graph is a curve of the change rate of tumor volume versus days after administration in a drug efficacy test of the conjugates DM1-A-1, DM1-A-2, DM1-B-2 and DM1 according to the present disclosure and a vehicle control in a mouse H524 xenograft model.

[0307] Fig.11 It is a tumor measurement observation diagram of the conjugates DM1-A-1, DM1-A-2, DM1-B-2 and DM1 according to the present disclosure and the vehicle control after the efficacy test in the mouse H524 xenograft model.

[0308] Fig. 12A The figure is a curve of body weight change over days after administration of the conjugates DM1-A-3, DM1-A-2, DM1-A-9, DM1-A-17, DM1-A-6 and DM1-A-7 according to the present disclosure and the vehicle control in the efficacy test in the mouse H524 xenograft model.

[0309] Fig. 12B The figure is a curve of body weight change rate versus days after administration of the conjugates DM1-A-3, DM1-A-2, DM1-A-9, DM1-A-17, DM1-A-6 and DM1-A-7 according to the present disclosure and the vehicle control in a drug efficacy test in a mouse H524 xenograft model.

[0310] Fig.13A The graph is a curve of the change in tumor volume versus days after administration in the efficacy test of the conjugates DM1-A-3, DM1-A-2, DM1-A-9, DM1-A-17, DM1-A-6 and DM1-A-7 according to the present disclosure and the vehicle control in the mouse H524 xenograft model.

[0311] Fig. 13BThe figure is a curve of the change rate of tumor volume versus days after administration in a drug efficacy test of the conjugates DM1-A-3, DM1-A-2, DM1-A-9, DM1-A-17, DM1-A-6 and DM1-A-7 according to the present disclosure and the vehicle control in a mouse H524 xenograft model.

[0312] Fig.14 It is a tumor measurement observation diagram of the conjugates DM1-A-3, DM1-A-2, DM1-A-9, DM1-A-17, DM1-A-6 and DM1-A-7 according to the present disclosure and the vehicle control after the efficacy test in the mouse H524 xenograft model.

[0313] Fig.15A The figure is a curve of body weight change in days after administration of the conjugates DM-A-11, DM-A-15, DM-A-8 and DM-A-18 according to the present disclosure and DM1 and a vehicle control in a drug efficacy test in a mouse H69 xenograft model.

[0314] Fig. 15B The figure is a curve of body weight change rate versus days after administration of the conjugates DM-A-11, DM-A-15, DM-A-8 and DM-A-18 according to the present disclosure and DM1 and a vehicle control in a drug efficacy test in a mouse H69 xenograft model.

[0315] Fig. 15C The figure is the curve of body weight change in days after administration of the conjugates DM1-A-3 and DM-A-9 according to the present disclosure and DM1 and vehicle control in the efficacy test of the mouse H69 xenograft model.

[0316] Fig.15D The figure is a curve of body weight change rate versus days after administration of the conjugates DM1-A-3 and DM-A-9 according to the present disclosure and DM1 and vehicle control in a drug efficacy test in a mouse H69 xenograft model.

[0317] Fig.16A The graph is a curve of the change in tumor volume versus days after administration of the conjugates DM-A-11, DM-A-15, DM-A-8 and DM-A-18 according to the present disclosure and DM1 and a vehicle control in a drug efficacy test in a mouse H69 xenograft model.

[0318] Fig. 16B The figure is a curve of the change rate of tumor volume versus days after administration in the efficacy test of the conjugates DM-A-11, DM-A-15, DM-A-8 and DM-A-18 disclosed herein and DM1 and the vehicle control in the mouse H69 xenograft model.

[0319] Fig. 16CThe graph is a curve of the change of tumor volume versus days after administration in the efficacy test of the conjugates DM1-A-3 and DM-A-9 according to the present disclosure and DM1 and vehicle control in the mouse H69 xenograft model.

[0320] Fig.16D The graph is a curve of the change rate of tumor volume versus days after administration in the efficacy test of the conjugates DM1-A-3 and DM-A-9 according to the present disclosure and DM1 and vehicle control in the mouse H69 xenograft model.

[0321] Fig.17A The figure is a tumor observation diagram after the efficacy test of the conjugates DM-A-11, DM-A-15, DM-A-8 and DM-A-18 according to the present disclosure and the vehicle control in the mouse H69 xenograft model.

[0322] Fig. 17B This is a tumor measurement observation diagram of the conjugate DM-A-9 according to the present disclosure and the vehicle control after the efficacy test in the mouse H69 xenograft model.

[0323] Fig. 17C This is a tumor observation diagram of the conjugate DM1-A-3 according to the present disclosure and the vehicle control after the efficacy test in the mouse H69 xenograft model.

[0324] Fig.18A The figure is the curve of body weight change versus days after administration of the conjugate DM1-A-3 according to the present disclosure and DM1 and vehicle control in the efficacy test of the mouse U87 xenograft model.

[0325] Fig.18B The weight change rate-days-after-dose curve of the conjugate DM1-A-3 and DM1 and vehicle control in the efficacy test of the mouse U87 xenograft model according to the present disclosure.

[0326] Fig.19A The graph is a curve of the change of tumor volume-days after administration in the efficacy test of the conjugate DM1-A-3 according to the present disclosure and DM1 and vehicle control in the mouse U87 xenograft model.

[0327] Fig.19B The graph is a curve of the change rate of tumor volume versus days after administration in a drug efficacy test of the conjugate DM1-A-3 according to the present disclosure and DM1 and a vehicle control in a mouse U87 xenograft model.

[0328] Fig. 20 It is a tumor measurement observation diagram after the efficacy test of the conjugate DM1-A-3 according to the present disclosure and DM1 and vehicle control in the mouse U87 xenograft model. DETAILED DESCRIPTION

[0329] [definition]

[0330] As used herein, "compounds" and "conjugates" may exist as one or more stereoisomers. Stereoisomers are isomers that have identical constitution but differ in the arrangement of their atoms in space, and include enantiomers, diastereomers, cis-trans isomers (also called geometric isomers), and atropisomers.

[0331] As used herein, "polypeptide", "compound", "polypeptide compound", "octreotide analogue" and "OCT analogue" have the same meaning and can be used interchangeably.

[0332] As used herein, "conjugate", "PDC" and "PDC conjugate" have the same meaning and can be used interchangeably.

[0333] Two configurational isomers that are mirror images of each other and cannot overlap are called enantiomers.

[0334] The present disclosure includes substantially pure enantiomers of compounds or conjugates; racemic mixtures, i.e., mixtures of equal amounts of two enantiomers; and enriched enantiomers, i.e., one enantiomer is present in a greater amount than the other enantiomer. Those skilled in the art will appreciate that when one stereoisomer is enriched relative to one or more other stereoisomers, or when one stereoisomer is separated from one or more other stereoisomers, it may be more active and / or may exhibit beneficial effects. In addition, those skilled in the art know how to separate, enrich, and / or selectively prepare the stereoisomers.

[0335] As used herein, the sequence or structural formula of a compound or conjugate contains the standard single-letter or three-letter codes for the 20 natural amino acids that make up proteins (also called protein amino acids or coded amino acids). Except for proline (Pro), the amino and carboxyl groups of the remaining 19 protein amino acids are all connected to the α carbon atom, also known as α-amino acids. For example, α-Ala represents α-alanine, with the structure CH3CH(NH2)COOH, wherein the amino group is connected to the α carbon atom. β-Ala represents β-alanine, with the structural formula NH2CH2CH2COOH, wherein the amino group is connected to the β carbon atom. As used herein, each amino acid residue can be independently of each other in the L-configuration or the D-configuration, and can also independently of each other have side substituents on the carbon atom.

[0336] As used herein, nal represents naphthyl alanine. α-nal represents α-naphthyl alanine, the structure of

[0337] , where the naphthyl group is attached to the α carbon atom. β-nal represents β-naphthylalanine, with the structure

[0338] , wherein the naphthyl group is connected to the β carbon atom, and the floating bond indicates that the alanine moiety can be connected to any carbon atom of the naphthyl ring except the fused bond.

[0339] In the context of this article, unless otherwise specified or clearly contradictory, the position number of the amino acid is counted from the leftmost N-terminus of the peptide chain or structural formula of the compound. Taking octreotide as an example, octreotide, also known as somatostatin octapeptide, is composed of 8 amino acids and has the following structure:

[0340]

[0341] Expanding the amino acids in the three-letter code above, octreotide has the following molecular structure:

[0342]

[0343] (Octreotide, OCT).

[0344] As used herein, "individual in need" means a mammal, preferably a human, having a condition, disease, disorder or symptom that needs treatment or prevention, but may also be a non-human animal, including non-human primates (e.g., cynomolgus monkeys), pets (e.g., cats, dogs, etc.), livestock (e.g., cows, sheep, pigs, horses, etc.), and rodents (e.g., rats, mice, guinea pigs, etc.).

[0345] As used herein, "effective amount" means the amount, concentration or dosage of one or more conjugates of the present disclosure or a pharmaceutically acceptable salt thereof that provides the desired effect (i.e., can produce a clinically measurable difference in the individual's condition, such as, for example, reduction or disappearance of a tumor) in the individual being diagnosed or treated after a single or multiple doses are administered to an individual in need thereof. An effective amount is readily determined by one skilled in the art by using known techniques and by observing the results obtained under similar circumstances. In determining an effective amount for an individual, many factors are considered, including but not limited to the species, size, age, general health of the mammal, the specific disease or condition involved, the severity of the disease or condition, the response of the individual, the specific compound administered, the mode of administration, the bioavailability characteristics of the administered formulation, the selected dosage regimen, the use of concomitant drug therapy, and other relevant circumstances.

[0346] As used herein, the term "treat," ...

[0347] As used herein, the term "plasma half-life" or "half-life" refers to the time required for half of the compound or conjugate of interest to be cleared from plasma.

[0348] As used herein, "in vitro activity" is an indicator of a peptide's ability to activate SSTR2 in a cell-based assay.

[0349] [Targeting ligand]

[0350] The present disclosure provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of general formula I:

[0351]

[0352] Among them, R 1 To R 7 , and R 15 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkylcarbonyl, halogen, cyano, hydroxyl, carboxyl, -C(O)NR 10 R 11 , phenyl, naphthyl, biphenyl, Polyethylene glycol, and C5-C containing one or more heteroatoms selected from N, S and O 10 Aromatic heterocyclic ring; or R 15 Condensed with adjacent phenyl groups to form naphthyl groups; these groups are optionally substituted by one or more substituents R 12 replace;

[0353] R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and

[0354] R 9 With R16 are the same or different and are independently selected from S, -CH2- and -C(CH3)2-; or R 9 With R 16 Together they represent -SS-, -S-CH2-, -S-CH(CH3)-, -SC(CH3)2-, -CH2-S-, -CH(CH3)-S-, -C(CH3)2-S-, -CH2-CH=CH-CH2- and as well as

[0355] R 10 To R 14 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, halogen, cyano, hydroxy, carboxyl, phenyl, and naphthyl;

[0356] R 17 express or biphenyl; or R 4 With R 17 Together with the -N-CH-CH2- in between, it forms

[0357] p represents an integer selected from 1 to 4; and

[0358] * indicates the point of attachment of the identified group to the rest of Formula I.

[0359] In some embodiments, in the compound of Formula I, R 1 is selected from H and phenyl;

[0360] R 2 is selected from phenyl, hydroxyphenyl and naphthyl;

[0361] R 3 is selected from H and -C(O)-CH3;

[0362] R 4H, or R 4 With R 17 Together with the -N-CH-CH2- in between, it forms

[0363] R 5 is selected from hydroxyl and methyl;

[0364] R 6 and R 7 are different and are independently selected from methyl, hydroxy, H and naphthyl;

[0365] R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and

[0366] R 9 With R 16 are the same or different and are independently selected from S and -CH2-; and p is 1 or 2.

[0367] In some examples, the present disclosure provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of Formula I-1:

[0368]

[0369] Among them, R 1 To R 7 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, halogen, cyano, hydroxyl, carboxyl, -C(O)NR 10 R 11 , phenyl, naphthyl, and C5-C6 containing one or more heteroatoms selected from N, S and O 10 Aromatic heterocyclic rings, these groups are optionally substituted by one or more substituents R 12 replace; or

[0370] R 4 With adjacent Connected via -CH2- and together form

[0371] R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and

[0372] R 9 selected from S and C; and

[0373] R 10 To R 14 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, halogen, cyano, hydroxy, carboxyl, phenyl, and naphthyl;

[0374] * indicates the connection point between the identified group and the rest of the general formula I-1.

[0375] In some examples, in the compound of formula I-1,

[0376] R 1 is selected from H and phenyl;

[0377] R 2 is selected from phenyl, hydroxyphenyl and naphthyl;

[0378] R 3 is selected from H and -C(O)-CH3;

[0379] R 4 H, or R 4 With adjacent Connected via -CH2-, forming

[0380] R 5 is selected from hydroxyl and methyl;

[0381] R 6 and R 7 are different and are independently selected from methyl, hydroxy, H and naphthyl;

[0382] R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and

[0383] R 9 is selected from S and -CH2-; and

[0384] p is 1 or 2.

[0385] In some embodiments, the compound of Formula I or Formula 1-1 is not octreotide.

[0386] The tumor-targeting polypeptide compound of general formula I or general formula I-1 according to the present invention is an octreotide analog. Compared with octreotide, the compound of general formula I or general formula I-1 of the present invention has a transformation such as substitution, deletion, addition, conversion and / or modification at one or more amino acid sites. By modifying octreotide, the obtained octreotide analog of general formula I or general formula I-1 maintains specific targeting of SSTR2 and induces endocytosis, has good drug targeting and delivery functions, can preferentially deliver drugs to tumor sites with high or overexpression of SSTR2 that need to be treated; has excellent plasma stability; and the presence of the tumor-targeting polypeptide compound of general formula I or general formula I-1 does not substantially interfere with the efficacy of the drug part with cytotoxicity.

[0387] In some examples, the compound of Formula I or Formula I-1 has a structure selected from any one of the following:

[0388]

[0389]

[0390]

[0391]

[0392] *: PEG20K represents a polyethylene glycol unit having a molecular weight of about 20K.

[0393] In some examples, the compound of Formula I or Formula I-1 has a structure selected from any one of the following: A-1, A-2, A-3, A-4, A-5, A-6, A-7, A-8, A-9, A-11, A-12, A-13, A-15, A-17, A-18, B-1, B-2, B-3, B-5, B-6 and B-7. These compounds can specifically target SSTR2 and induce endocytosis, have good drug targeting and delivery functions, have excellent plasma stability, and do not substantially interfere with the efficacy of the active drug portion to be delivered.

[0394] In some examples, the compound of Formula I or Formula I-1 has a structure selected from any one of the following:

[0395]

[0396]

[0397]

[0398]

[0399] The present disclosure also provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of general formula II:

[0400]

[0401] Among them, the group R 1 To R 17 , and p has the same definition as given above for the compound of general formula I, X represents a linking portion and is selected from one of the following groups or any combination thereof:

[0402] -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20 Cycloalkyl and its isomers, phenyl, naphthyl, maleimide groups and their hydrides, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl,

[0403] **Indicates that the identified group is attached to a leaving group or an active moiety.

[0404] In some embodiments, the present invention provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of Formula II-1:

[0405]

[0406] Among them, the group R 1 To R 9 has the same definitions as given above for the compounds of formula I-1,

[0407] X represents a linking moiety and is selected from one of the following groups or any combination thereof:

[0408] -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl,

[0409] **Indicates that the identified group is attached to a leaving group or an active moiety.

[0410] In some embodiments, the leaving group is H, or a C5-C ... 10 Aromatic heterocyclic rings, such as pyridyl, are optionally substituted by one or more substituents R as defined above for formula I. 12 replace.

[0411] In some examples, the linking moiety is

[0412] In some examples, the compound of formula II or II-1 is any one of compounds selected from A-1 to A-18, and B-1 to B-23, connected to the following linking moiety:

[0413]

[0414] As used herein, a linking moiety can be hydrolyzed in vivo, or comprises a cleavable moiety that can be hydrolyzed in vivo.

[0415] The compounds of Formula II or II-1 can be used as intermediates for synthesizing PDC conjugates according to the present disclosure.

[0416] [Conjugate]

[0417] As used herein, the term "conjugate" includes a targeting ligand and an active substance connected by a linking moiety. A targeting ligand can be conjugated to one or more active substances. An active substance can also be conjugated to one or more active substances.

[0418] In some examples, the targeting ligand is a compound of Formula I or I-1 as described above.

[0419] In some embodiments, the active substance is maytansine (DM1). Maytansine (DM1) has the following structure:

[0420]

[0421] The present disclosure provides a conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate has a structure of general formula III:

[0422]

[0423] Among them, the group R 1 To R 17 , and p has the same definition as in claim 1,

[0424] X represents a linking moiety and is selected from one of the following groups or any combination thereof:

[0425] -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20 Cycloalkyl and its isomers, phenyl, naphthyl, maleimide groups and their hydrides, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl; and

[0426] Y represents an active moiety and is maytansine (DM1),

[0427] ** indicates the connection point between the identified group and Y.

[0428] In some examples, the present disclosure provides a conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate has a structure of Formula III-1:

[0429]

[0430] Among them, the group R 1 To R 9 has the same definitions as given above for the compounds of formula I-1,

[0431] X represents a linking moiety and is selected from one of the following groups or any combination thereof:

[0432] -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20 Cycloalkyl and its isomers, phenyl, naphthyl, maleimide groups and their hydrides, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl; and

[0433] Y represents an active moiety and is maytansine (DM1),

[0434] ** indicates the connection point between the identified group and Y.

[0435] The inventors synthesized and screened out specific OCT analogs of general formula I or I-1, and connected them to maytansine DM1 through a specific linker. The obtained PDC conjugate of general formula III or III-1 can specifically recognize tumor cells with high or overexpression of SSTR2 and enter the tumor cells through endocytosis. In other words, the PDC conjugate according to the present disclosure can specifically recognize and target tumor cells with high or overexpression of SSTR2. At the same time, the PDC conjugate according to the present disclosure has excellent plasma stability.

[0436] In some examples, the conjugate of Formula III or III-1 has a structure selected from the following:

[0437]

[0438]

[0439]

[0440]

[0441]

[0442]

[0443]

[0444]

[0445]

[0446]

[0447]

[0448]

[0449]

[0450]

[0451]

[0452] In some examples, the conjugate of Formula III or Formula III-1 has a structure selected from any one of the following: DM1-A-1, DM1-A-2, DM1-A-3, DM1-A-4, DM1-A-5, DM1-A-6, DM1-A-7, DM1-A-8, DM1-A-9, DM1-A-11, DM1-A-12, DM1-A-13, DM1-A-15, DM1-A-17, DM1-A-18, DM1-B-1, DM1-B-2, DM1-B-3, DM1-B-5, DM1-B-6 and DM1-B-7. In some examples, the conjugate of Formula III or Formula III-1 has a structure selected from any one of the following: DM1-A-2, DM1-A-3, DM1-A-7, DM1-A-8, DM1-A-9, DM1-A-11, DM1-A-15, DM1-A-17 and DM1-A-18. Such PDC conjugates can specifically recognize and target tumor cells with high or overexpression of SSTR2, and have excellent plasma stability.

[0453] [Methods for preparing compounds and conjugates]

[0454] [Method for preparing the compound of formula I or I-1]

[0455] Rink-amide-AM resin (Tianjin Nankai Hecheng Technology Co., Ltd.) was used as a synthetic carrier to prepare the compounds according to the present disclosure via a solid phase synthesis method. The amino groups of each amino acid raw material used in the synthesis process were protected by an Fmoc group (9-fluorenyl-methyloxy carbonyl, fluorenyl-methyloxy carbonyl, Fmoc). For neutral polar amino acids, acidic amino acids and basic amino acids, the following protecting groups were selected to protect the polar side chains of these amino acid raw materials according to the different side chain functional groups. For example, the side chain thiol of cysteine ​​(Cys) is protected by Trt (trityl); the side chain indole of tryptophan (Trp) and the side chain amino group of lysine (Lys) are protected by Boc (tert-butoxycarbonyl protecting group, tert-butoxycarbonyl); the side chain hydroxyl of threonine (Thr) and the side chain phenol group of tyrosine (Tyr) are protected by tBu (tert-butyl, tert-butyl).

[0456] In some examples, the compounds according to the present disclosure are prepared by a method comprising the following steps:

[0457] i) Swelling and deprotection of resin support

[0458] The Rink-Amide-AM resin previously protected by Fmoc was swollen, and then the Fmoc protecting group of the Rink-Amide-AM resin was removed using a N,N-dimethylformamide (DMF) solution containing 20% ​​piperidine.

[0459] ii) Formation of polypeptides

[0460] Starting from the C-terminus of the compound structure, toward the N-terminus, amino acids are linked one by one in the following manner to form a polypeptide, wherein an amide bond is formed by a condensation reaction of the carboxyl group of the amino acid whose amino group is protected by Fmoc:

[0461] First, the carboxyl group of the first amino acid with Fmoc protection (starting from the C-terminus) is condensed in the form of an amide bond onto the swollen and deprotected Rink-amide-AM resin using a condensation agent 6-chlorobenzotriazole-1,1,3,3-tetramethyluronium hexafluorophosphate (O-(1H-6-Chlorobenzotriazole-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate, HCTU), and then the Fmoc protecting group on the amino group is removed using an N,N-dimethylformamide (DMF) solution containing 20% ​​piperidine, followed by washing.

[0462] Then, in a similar manner, using the condensing agent HCTU and the amino acids at positions 2 to 8 (starting from the C-terminus) protected by Fmoc, the above coupling reaction of forming an amide bond, deprotection of removing the Fmoc protecting group and washing cycles were repeated in sequence.

[0463] iii) Cyclization

[0464] After the amino acids are sequentially condensed to the first amino acid at the N-terminus, 1 equivalent (relative to the amount of the peptide on the resin) of iodine / DMF is used to directly oxidize the two Trt-protected cysteines (the 2nd and 7th positions from the N-terminus) on the resin, thereby condensing and cyclizing to form a disulfide bond.

[0465] iv) Peptide cleavage and characterization

[0466] A mixture of trifluoroacetic acid (TFA, trifluoroacetic acid) / water / phenol / triisopropyl silane (Tips, Triisopropyl silane) is used as a cleavage reagent, and the cleavage reagent reacts with Rink-Amide-AM resin to cleave the polypeptide from the resin carrier. The polypeptide solid crude product is obtained by precipitation with frozen ice ether. The polypeptide solid crude product is centrifuged, dried, and purified by semi-preparative high performance liquid chromatography to obtain a pure peptide, i.e., an octreotide analog of general formula I or I-1.

[0467] The purified peptides were analyzed for purity by analytical HPLC and electrospray ionization mass spectrometry (ESI-MS) was performed to confirm the accuracy of the molecular weight.

[0468] [Method for preparing the intermediate compound of formula II or II-1]

[0469] The preparation process of the intermediate compound of general formula II or II-1 is similar to that of the compound of general formula I or I-1, except that general formula II uses 2-Chlorotrityl chloride resin, and the first amino acid at the C-terminus does not require a condensing agent and is directly connected to the resin in the presence of a base of N,N-diisopropylethylamine (DIEA); and after the completion of polypeptide synthesis and cyclization, the polypeptide resin is cut using 20% ​​hexafluoroisopropanol (HFIP) / dichloromethane, and the fully protected crude peptide is obtained after spin drying, ether precipitation, and centrifugation. The fully protected crude peptide is subjected to a condensation reaction with S-trityl-L-cysteine ​​amide, and the obtained condensation product is added to a deprotection base liquid containing trifluoroacetic acid (TFA) to obtain a polypeptide with a connecting portion, and after purification using semi-preparative high performance liquid chromatography, the polypeptide, i.e., the intermediate compound of general formula II or II-1, is obtained.

[0470] The purified peptides were analyzed for purity by analytical HPLC and electrospray ionization mass spectrometry (ESI-MS) was performed to confirm the accuracy of the molecular weight.

[0471] [Method for preparing the conjugate of formula III or III-1]

[0472] The intermediate compound of general formula II or II-1 obtained above is condensed with dithiodipyridine, and the obtained condensation product is condensed with maytansine DM1 to obtain a polypeptide connected by a linking part and DM1. After purification using semi-preparative HPLC, a PDC compound of general formula III or III-1 is obtained.

[0473] The purified PDC compound was analyzed by analytical HPLC to determine its purity and electrospray ionization mass spectrometry (ESI-MS) to confirm the accuracy of the molecular weight.

[0474] [Drug composition]

[0475] The present disclosure provides a pharmaceutical composition comprising:

[0476] The conjugate according to the present disclosure or a pharmaceutically acceptable salt thereof, and

[0477] A pharmaceutically acceptable carrier, diluent or excipient.

[0478] In addition to one or more compounds according to the present disclosure or pharmaceutically acceptable salts thereof, the pharmaceutical composition may further comprise other components, such as physiologically / pharmaceutically acceptable carriers, diluents and excipients, to facilitate administration to an individual, facilitate absorption of the active ingredient or pharmaceutically acceptable salt thereof, and thereby exert biological activity.

[0479] "Pharmaceutically acceptable salts" refers to salts of compounds according to the present disclosure that are safe and effective for use in a subject and that possess biological activity.

[0480] The compounds according to the present disclosure can react with a variety of inorganic acids and organic acids to form pharmaceutically acceptable salts. Pharmaceutically acceptable salts and common methods for preparing them are well known in the art. See, for example, P. Stahl et al., Handbook of Pharmaceutical Salts: Properties, Selection and Use, Second Revised Edition (Wiley-VCH, 2011); SM Berge et al., "Pharmaceutical Salts", Journal of Pharmaceutical Sciences, Vol. 66, No. 1, January 1977.

[0481] In some examples, the conjugates according to the present disclosure or pharmaceutically acceptable salts thereof can be formulated into pharmaceutical compositions for administration by parenteral routes (e.g., subcutaneous, intravenous, intraperitoneal, intramuscular or transdermal). Such pharmaceutical compositions and methods for preparing them are well known in the art. See, for example, Remington: The Science and Practice of Pharmacy (DB Troy ed., 21st ed., Lippincott, Williams & Wilkins, 2006).

[0482] Pharmaceutically acceptable salts according to the present disclosure include, but are not limited to, trifluoroacetate, hydrochloride, and acetate.

[0483] [use]

[0484] The present disclosure provides a use of a pharmaceutical composition according to the present disclosure, or a conjugate according to the present disclosure or a pharmaceutically acceptable salt thereof, or a compound according to the present disclosure for preparing a drug for treating a tumor or reducing tumor volume;

[0485] In particular, the tumor expresses the somatostatin receptor (SSTR2);

[0486] In particular, the tumor is a neuroendocrine cancer;

[0487] In particular, the tumor is selected from lung cancer (eg small cell lung cancer), glioma, pancreatic cancer, breast cancer, colorectal cancer, ovarian cancer, colorectal cancer, bladder cancer, prostate cancer, cervical cancer, kidney cancer, leukemia, central nervous system cancer, myeloma and melanoma.

[0488] [Methods for treating tumors or reducing tumor size]

[0489] The present disclosure provides a method for treating a tumor or reducing the volume of a tumor, comprising administering an effective amount of the pharmaceutical composition according to the present disclosure, or the conjugate according to the present disclosure or a pharmaceutically acceptable salt thereof to an individual in need thereof.

[0490] In some examples, the pharmaceutical composition, the conjugate, or a pharmaceutically acceptable salt thereof is administered to the individual by subcutaneous injection.

[0491] In some examples, the pharmaceutical composition, the conjugate, or a pharmaceutically acceptable salt thereof is administered to the individual once a week.

[0492] In some examples, the pharmaceutical composition, the conjugate, or a pharmaceutically acceptable salt thereof is administered to the individual once a week by subcutaneous injection.

[0493] [Example]

[0494] [Preparation Example]

[0495] Experimental reagents:

[0496] Unless otherwise specified, all reagents used herein are commercially available. Unless otherwise specified, N,N-dimethylformamide (DMF) and dichloromethane (DCM) used herein are common reagents with a purity of 99.7%, and the excipients are all 0.05% NaHCO3 solution. 20% piperidine / DMF solution is prepared by the following method: 100 mL of piperidine is measured with a measuring cylinder, and DMF is added to the measuring cylinder mark of 500 mL to obtain a 20% piperidine / DMF solution.

[0497] Unless otherwise stated, in the embodiments of the present disclosure, draining the solvent means using an air pump to draw the solvent in the polypeptide synthesis tube into a suction filtration bottle.

[0498] Unless otherwise stated, in the examples of the present disclosure, vehicle control refers to citrate buffer.

[0499] [Preparation of Compound A-1]

[0500] (1) Resin swelling

[0501] a) Weigh 0.4 mmol (1000 mg) of Fmoc-protected Rink-Amide resin and place it into a peptide synthesis tube.

[0502] b) Add 5 mL of DMF (N,N-dimethylformamide) and 5 mL of DCM (dichloromethane) to the peptide synthesis tube for swelling and leave at room temperature for 30 min.

[0503] c) Use an air pump to drain the solvent.

[0504] d) Rinse with 10 mL of DMF and drain the solvent.

[0505] (2) Resin deprotection

[0506] a) Add 5 mL of 20% piperidine / DMF solution to the peptide synthesis tube to submerge the swollen resin obtained in step (1), and transfer to a 33°C constant temperature shaker for 5 min.

[0507] b) Take the peptide synthesis tube out of the shaker.

[0508] c) Cleaning: First, rinse the resin three times with DMF (5 mL each time), and drain the solvent; then rinse the resin three times with DCM (5 mL each time), and drain the solvent; finally, rinse the resin three times with DMF (5 mL each time), and drain the solvent.

[0509] d) Deprotection: Add 5 mL of 20% piperidine / DMF solution to the peptide synthesis tube, shake in a constant temperature shaker at 33° C. for 10 min, and take out the peptide synthesis tube.

[0510] e) Repeat the washing step c) in step (2) "Resin deprotection".

[0511] Then, the polypeptide is formed in the following manner: starting from the first amino acid at the rightmost C-terminus of the polypeptide chain, the amino acids are connected one by one towards the leftmost N-terminus to obtain the following polypeptide structure:

[0512]

[0513] (3) Connect (starting from the rightmost C-terminus) to the first amino acid

[0514] a) Weigh 0.4 mmol (636 mg) of Fmoc-Thr(Trt)-OH (threonine) and 0.8 mmol (314 mg) of condensing agent HCTU (6-chlorobenzotriazole-1,1,3,3-tetramethyluronium hexafluorophosphate) into a 10 mL EP tube, add 4 mL of DMF to the EP tube to dissolve, shake well, and then add 265 μL (1.6 mmol) of DIEA to the EP tube to obtain a mixed solution.

[0515] b) The mixed solution was transferred to a peptide synthesis tube, and then the peptide synthesis tube was transferred to a 33°C constant temperature shaker for 1 hour and then taken out.

[0516] c) Washing: Repeat the washing step c) in step (2) "Resin deprotection".

[0517] d) Deprotection: Add 5 mL of 20% piperidine / DMF solution to the peptide synthesis tube, shake in a constant temperature shaker at 33° C. for 10 min, and take out the peptide synthesis tube.

[0518] (4) Connect (starting from the rightmost C-terminus) the 2nd to 8th amino acids

[0519] Similar to the connection of the first amino acid (starting from the rightmost C-terminus) in step (3), the second to eighth amino acids (starting from the rightmost C-terminus) are connected in sequence, except that Fmoc-Cys(Trt)-OH (cysteine), Fmoc-Val-OH (valine), Fmoc-Lys(Boc)-OH (lysine), Fmoc-D-Trp(Boc)-OH (tryptophan), Fmoc-Phe-OH (phenylalanine), Fmoc-Cys(Trt)-OH (cysteine) and Fmoc-D-β-nal(Boc)-OH (β-naphthylalanine) are used as raw materials for coupling (starting from the rightmost C-terminus) the second to eighth amino acids, respectively. The difference also lies in that different condensation agents are used according to different amino acid raw materials. For 0.4 mmol (210 mg) of Fmoc-D-Trp-OH (tryptophan) and 0.4 mmol (154 mg) of Fmoc-Dβnal-OH (D-β-naphthylalanine), 0.4 mmol (144 mg) of HCTU and 0.4 mmol (51 mg) of HOAT (N-hydroxy-7-azobenzotriazole) are used instead of 0.8 mmol (314 mg) of HCTU as condensation agents.

[0520] (5) Cyclization

[0521] a) Cyclization: Weigh 0.4 mmol (101 mg) of iodine into a 10 mL EP tube, add 2 mL of DMF to the EP tube to dissolve the iodine, shake well, and transfer to a peptide synthesis tube. Transfer the peptide synthesis tube to a 33°C constant temperature shaker for 2 h, take out the peptide synthesis tube, drain the solvent, and then wash it three times with DMF.

[0522] b) After the cyclization is completed, the peptide synthesis tube is taken out and deprotection is performed: 5 mL of 20% piperidine / DMF solution is added to the peptide synthesis tube, and the tube is shaken in a constant temperature shaker at 33° C. for 10 minutes, and the peptide synthesis tube is taken out.

[0523] c) The resin was rinsed three times with DMF (5 mL each time), and the solvent was drained after each rinse. Then, the resin was rinsed three times with methanol (5 mL each time), and the solvent was drained after each rinse (drained until the resin was in a dry powder state).

[0524] (6) Cleavage of crude peptide

[0525] After drying, prepare a cleavage reagent of TFA / H2O / phenol / Tips (triisopropylsilane) (volume ratio of 88:4:4:4) in a 50mL EP tube. Transfer the cleavage reagent to the above-mentioned peptide synthesis tube, place it in a 26°C constant temperature shaker for 2 hours, take out the peptide synthesis tube, and the solution in the tube is the peptide chain cleavage solution.

[0526] (7) Post-treatment of peptides

[0527] a) Use an ear bulb to transfer 10 mL of peptide chain lysate into a 50 mL centrifuge tube and blow the lysate dry to less than 5 mL using nitrogen at room temperature.

[0528] b) Ice ether precipitation and centrifugation: Add 40 mL of ice ether to a 50 mL centrifuge tube. After shaking the tube appropriately, place the tube in a centrifuge at 3500 rpm for 3 min. After centrifugation, discard the supernatant.

[0529] c) Repeat the above-mentioned ether precipitation and centrifugation operations, discard the supernatant, and the obtained precipitate is the crude peptide.

[0530] d) Dry at room temperature and crush to obtain crude peptide.

[0531] The purified peptides were separated using Shimadzu semi-preparative liquid chromatography. Figure 1A-1 This is the ESI-MS (electrospray ionization mass spectrometry) characterization of the purified polypeptide compound A-1. The peaks in the figure represent molecular weights with different mass-to-charge ratios. Figure 2A-1 is a high performance liquid chromatogram, which shows that the synthesized compound A-1 has a purity of 95.28%.

[0532] [Preparation of Compounds A-2 to A-18, and B-1 to B-23]

[0533] Compounds A-2 to A-18 and compounds B-1 to B-23 were prepared in a similar manner to the preparation of compound A-1, except that the Fmoc-protected amino acid raw materials and condensing agents were changed according to the structural formula of each compound. Condensing agents include but are not limited to HATU, HBTU, HCTU, HOAT and PyAOP.

[0534] [N-terminal acylation]

[0535] In the case of acylation of the N-terminal amino acid, taking compound A-8 as an example, acetic anhydride is used as the acetylation agent. In other acylation cases, the corresponding acylating agents can be used instead of acetic anhydride.

[0536] In a similar manner to the preparation of compound A-1, 8 amino acids were sequentially connected in the order from Thr at the C-terminus to D-Phe at the N-terminus to obtain a polypeptide with the following structure.

[0537]

[0538] Prior to cyclization, N-terminal acetylation was performed as follows:

[0539] ① Add 10mL of acetic anhydride / DIEA / DMF (volume ratio of 1:1:8) to the peptide synthesis tube; transfer the peptide synthesis tube to a 33°C constant temperature shaker and shake for 10 minutes, then take out the peptide synthesis tube, drain the solvent, and wash it three times with DMF;

[0540] ② Repeat step ①, take out the peptide synthesis tube and drain the solvent, wash it with DMF five times, and drain the solvent.

[0541] Then, cyclization, cleavage of the crude peptide and post-treatment of the polypeptide were performed sequentially as described above for the preparation of compound A-1, except that no deprotection operation was performed after the cyclization process.

[0542] [C-terminal acylation]

[0543] When modifying the C-terminus, taking B-22 as an example, the first amino acid (starting from the rightmost C-terminus) uses Fmoc-dap(alloc)-OH. After eight amino acids are connected, the protecting group allyloxycarbonyl (alloc) is removed, and then the palmitoyl group (pamitoyl) is connected.

[0544] ① Add 10mL of BOC anhydride / DIEA / DMF (1:1:8, volume ratio) reagent to the peptide synthesis tube; transfer the peptide synthesis tube to a 33°C constant temperature shaker, shake for 10 minutes, then take out the peptide synthesis tube; drain the solvent and wash it three times with DMF;

[0545] ②Repeat operation ①;

[0546] ③ Weigh 232 mg of tetrakistriphenylphosphine palladium and put it into a 10 mL EP tube. Add 4 mL

[0547] Dissolve DMF and 4mL DCM, shake well, and then add 248μL of phenylsilane to the EP tube; mix the above mixture evenly and transfer it to the peptide synthesis tube, transfer the peptide synthesis tube to a 33℃ constant temperature shaker, shake for 2h, and then take out the peptide synthesis tube; after draining the solvent, wash it with DMF 5-6 times;

[0548] ④ Repeat ③ steps, drain the solvent and wash with DMF 5-6 times until the solution is colorless;

[0549] ⑤ Weigh 1.6 mmol (409 mg) of palmitic acid and 1.52 mmol (628 mg) of condensing agent (HCTU) into a 10 mL EP tube, add 8 mL of DMF to dissolve, shake well, and then add 3.2 mmol (530 μL) of DIEA to the EP tube;

[0550] ⑥ Transfer the above mixed solution to a peptide synthesis tube, and then transfer the peptide synthesis tube to a 33°C constant temperature shaker. After shaking for 1 hour, take out the peptide synthesis tube; drain the solvent and wash it with DMF 3 times.

[0551] According to HPLC analysis, the purity of compounds A-2 to A-18 and B-1 to B-23 were all ≥95%.

[0552] [Preparation of Peptide-Fluorescein A-1-FITC]

[0553] (1) Cysteine

[0554]

[0555] The side chain fully protected A-1 (220 mg, 0.14 mmol, 1 eq.) and HATU (65 mg, 0.17 mmol, 1.2 eq.) were added to 5 mL of dichloromethane, stirred to dissolve, and DIPEA (N,N-diisopropylethylamine, 36 mg, 0.28 mmol, 2 eq.) was added dropwise. After the addition was complete, the mixture was stirred at room temperature for half an hour, and then S-trityl-L-cysteineamide (HL-Cys(Trt)-NH2, 51 mg, 0.14 mmol, 1 eq.) was added and reacted at room temperature for 2 hours.

[0556] Dichloromethane (50 mL) and water (50 mL) were added to the reaction mixture, and the layers were separated by extraction. The aqueous phase was extracted once with 30 mL of dichloromethane. The organic layers were combined, washed with 10 mL of 1 mol / L dilute hydrochloric acid, and then washed with a saturated sodium bicarbonate aqueous solution. Finally, the organic layer was dried and concentrated to obtain 185 mg of a light yellow solid (fully protected A-1-Cys (Trt), crude yield 70%).

[0557] (2) Deprotection

[0558]

[0559] The fully protected A-1-Cys (Trt) (231 mg, 0.135 mmol) was added to the deprotection base solution (mainly containing TFA, 4 mL), stirred at room temperature for 2 hours, and sent to MS for detection, and the reaction was completed.

[0560] Ether (30 mL) was added dropwise under ice bath, and a large amount of white solid precipitated.

[0561] After centrifugation and washing with ether three times, the residue was blown dry with nitrogen to obtain a crude product (A-1-Cys, 172 mg, crude yield>100%).

[0562]

[0563] (3) Fluorescein

[0564] The previously prepared A-1-Cys (100 mg, 0.08 mmol, 1 eq.) and maleimide fluorescein (34.2 mg, 0.08 mmol, 1 eq.) were added to DMF (2 mL) and stirred to dissolve. pH 5.6 acetate buffer (1.5 mL) was added dropwise and stirred at room temperature for 2 hours, and then sent for MS detection.

[0565] The reaction solution was concentrated using an oil pump and purified by HPLC to obtain the product (A-1-FITC, 8 mg) with a yield of 6%.

[0566] In a similar manner, conjugates of other octreotide analogs A-2 to A-18, and B-1 to B-23 with fluorescein FITC were prepared.

[0567] [Preparation of PDC conjugate DM1-A-1]

[0568] (1) Formation of disulfide bond (-SS-) linking part

[0569]

[0570] The previously prepared A-1-Cys (100 mg, 0.08 mmol, 1 eq.) and disulfide dipyridine (73 mg, 0.33 mmol, 4 eq.) were added to DMF (1.8 mL) and stirred to dissolve. Acetic acid (0.12 mL) and .2 M sodium acetate aqueous solution (1.2 mL) were added dropwise and stirred at room temperature overnight. The mixture was sent for MS detection.

[0571] The reaction solution was concentrated to dryness using an oil pump and purified by HPLC to obtain the product (A-1-SS-Pyd, 13 mg) with a yield of 12%.

[0572] (2) Connect to DM1

[0573]

[0574] The previously prepared A-1-SS-Pyd (13 mg, 0.01 mmol, 1 eq.) and DM1 (7.4 mg, 0.01 mmol, 1 eq.) were added to DMF (2 mL) and stirred to dissolve. pH 4.6 phosphate buffer (2 mL) was added dropwise and stirred at room temperature for 2 hours. The mixture was then sent for MS detection.

[0575] The reaction solution was concentrated to dryness using an oil pump and purified by HPLC to obtain the product (DM1-SSA-1, abbreviated as DM1-A-1, 12 mg) with a yield of 63%.

[0576] In a similar manner, PDC conjugates of other octreotide analogs A-2 to A-18, and B-1 to B-23 with DM1 were prepared.

[0577] [In vitro experiments]

[0578] [FLIPR calcium flux experiment]

[0579] (1) Preparation of HEK293T cells

[0580] (a) HEK293T cell preparation and transfection

[0581] HEK293T cells were grown in medium dishes (35 mm culture dishes) to 60%-70% confluency and transfected with 4 μg SSTR2 + 2 μg Gqi9 plasmid;

[0582] (b) Transfer of business

[0583] 24 hours after transfection, the cells were plated in the following manner: 100 μL / well poly-lysine (P2636-1G, purchased from Sigma) was used to coat a black 96-well plate, and then the transfected cells were evenly blown with culture medium and counted. The cell density was adjusted with culture medium to 7×10 4 Cells / 100 μL / well were plated into 96-well plates;

[0584] (c) Cultivation

[0585] The cells were cultured overnight in a 37°C carbon dioxide incubator (HERAcell vios 160i, Thermo Fisher Scientific) to prepare for subsequent experiments.

[0586] (2) Preparation of calcium flux reagent

[0587] (a) Preparation of fresh calcium flux buffer

[0588] 2.5 mL of probenecid (purchased from Sigma, 250 mM, 71 mg) was dissolved in 1 mL of 1 M NaOH.

[0589] The following substances were added to HBSS balanced salt (Hanks' Balanced Salt Solution, BL561A, purchased from Biosharp): 500 μL of the NaOH solution of probenecid prepared above, 50 mg of bovine serum albumin powder (BSA, B2064-50G, purchased from Sigma), and 1 mL of 1M 4-hydroxyethylpiperazineethanesulfonic acid (HEPES, H3357, purchased from Sigma). Then, the pH was adjusted to 7.4 with hydrochloric acid, and the volume was fixed to 50 mL with HBSS balanced salt, that is, diluted 100 times, and the concentrations of the substances in the resulting solution were: 2.5 mM probenecid, 20 mM HEPES, and 0.1 mass % BSA.

[0590] (b) 20% Pluronic acid preparation

[0591] 20 mg of Pluronic F-127 was weighed and dissolved in 100 μL of DMSO to obtain a 20 mass % Pluronic F-127 solution.

[0592] (c) Fluo4-AM dye solution configuration

[0593] Immediately before use, fluo4-AM was taken out from the -20°C freezer, diluted to 1000 times the volume with the calcium flux buffer prepared above, and 1 / 2000 volume of 20% Pluronic F-127 solution was added.

[0594] (d) Ligand gradient configuration

[0595] The ligand dye fluo4-AM was diluted to 2×10 -4 ~2×10 -11 M, 3 wells for each concentration, each well volume 100μL, actual filling volume is 50μL / well.

[0596] (3) Calcium flux experiment

[0597] (a) The culture medium of the HEK293T cells transfected and plated above in the 96-well plate was discarded, and the cells were washed once with 100 μL / well of fresh calcium flux buffer;

[0598] (b) Add 50 μL / well of the above-prepared Fluo 4-AM and incubate at 37°C for 45 min;

[0599] (c) Discard Fluo 4-AM, wash once with 100 μL / well of calcium flow buffer, and then add 50 μL / well of calcium flow buffer;

[0600] (d) A high-throughput CCD fluorescence detection and analysis system (Molecular Devices, FLIPR Penta) was used to perform a calcium flux experiment on the octreotide analogs prepared above. The results are shown in Tables 1 and Figure 3A-1 to Figure 3B-23 shown.

[0601] Table 1 Results of calcium flux experiments of OCT analogs (EC 50 / M)

[0602]

[0603]

[0604] From the above table 1 and Figure 3A-1 to Figure 3B-23 It can be seen that compared with OCT, compounds A-1, A-2, A-3, A-4, A-5, A-6, A-7, A-8, A-9, A-11, A-12, A-13, A-15, A-17, A-18, B-1, B-2, B-3, B-5, B-6, and B-7 have good downstream signal responses.

[0605] [Nanobit Experiment]

[0606] (1) Plasmid construction

[0607] Human SSTR2 and βarrestin2 were cloned into pBiT1.1-C[TK / LgBiT] (model C7238HA060-1, purchased from Jinruis) and pBiT2.1-N[TK / SmBiT] vectors (purchased from University of Science and Technology of China), respectively. LgBiT and SmBiT are Two subunits of luciferase.

[0608] (2) Transfection

[0609] Using Lipofectamin3000 Transfection kit (model 2296643, purchased from Invitroge), 3 μg pBiT1.1-SSTR2-LgBiT and 3 μg pBiT2.1-SmBiT-βarrestin2 were co-transfected into HEK293T cells cultured in 35 mm culture dishes and cultured in a carbon dioxide incubator (model 3131, Thermo Fisher) for 24 hours.

[0610] (3) Plating and drug administration

[0611] Use OMEM to dilute the transfected HEK293T cells to 4 × 10 5 100 μL of diluted cells were plated in a 96-well plate, and 25 μL of 250 μM luminescent substrate furimazine (model HY-111497, purchased from MCE) dissolved in OMEM was added, and 10 μL of ten-fold serial dilutions (1.35×10 -4 , 1.35×10 -5 , ···, 1.35×10 - 13 M) the octreotide analogue prepared as described above.

[0612] (4) Detection

[0613] The chemiluminescent signal was continuously detected for 1 h using a TECAN microplate reader (model INFINITE 200PRO, purchased from Tecan). The experimental results are recorded in Table 2 and Figure 4A-1 to Figure 4B-21 middle.

[0614] Table 2 Nanobit experimental results of OCT analogs (EC 50 / M)

[0615] Compound No. <![CDATA[EC 50 / M]]> Compound No. <![CDATA[EC 50 / M]]> OCT <![CDATA[7.0×10 -9 ]]> A-11 <![CDATA[2.8×10 -7 ]]> A-2 <![CDATA[1.7×10 -8 ]]> A-12 <![CDATA[6.6×10 -7 ]]> A-3 <![CDATA[3.7×10 -7 ]]> A-13 Weak response A-4 <![CDATA[9.0×10 -8 ]]> A-14 No response A-5 <![CDATA[7.2×10 -7 ]]> A-15 Weak response A-6 Weak response A-17 <![CDATA[3.4×10 -7 ]]> A-7 Weak response A-18 <![CDATA[3.3×10 -7 ]]> A-8 <![CDATA[5.3×10 -8 ]]> A-9 <![CDATA[7.6×10 -8 ]]> B-3 <![CDATA[7.2×10 -7 ]]> B-4 <![CDATA[1.0×10 -6 ]]> B-5 <![CDATA[3.0×10 -8 ]]> B-6 <![CDATA[1.1×10 -6 ]]> B-7 <![CDATA[1.1×10 -6 ]]> B-8 No response B-9 No response B-10 <![CDATA[1.8×10 -6 ]]> B-11 No response B-12 No response B-13 Weak response B-14 No response B-15 <![CDATA[1.4×10 -6 ]]> B-16 No response B-17 Weak response B-18 Weak response B-19 Weak response B-20 No response B-21 No response B-22 <![CDATA[1.8×10 -6 ]]> B-23 <![CDATA[2.9×10 -6 ]]>

[0616] From Table 2 and Figure 4A-1 to Figure 4B-21 It can be seen that A-2, A-3, A-4, A-5, A-8, A-9, A-11, A-12, A-17, A-18, B-3, and B-5 have good βArrestin2 recruitment capabilities, thereby increasing the endocytosis of SSTR2, reducing the amount of SSTR2 on the tumor cell membrane, and enhancing the ability to specifically identify and target the elimination of tumor cells with high or overexpression of SSTR2.

[0617] [Cell selection]

[0618] Western Blot experiments were performed to confirm positive cells with high expression of SSTR2 and negative cells with no or low expression of SSTR2. The cells to be tested were: non-small cell lung cancer cells A549; small cell lung cancer cells H524, H2171 and H69; or glioma cells A172 and U87. The experimental steps are as follows:

[0619] (1) Sample preparation

[0620] Take out the cells to be tested from the incubator (SERIES II WATER JACKET, purchased from Thermo Fisher), wash them with PBS, add trypsin to digest, and then terminate the digestion with culture medium and count. Take 2 million cells, centrifuge at 500g for 5 minutes, discard the culture medium supernatant, add 200μL phosphate buffer (PBS, phosphate buffered solution, catalog number 10010-031, purchased from GIBCO) to resuspend the cell pellet, centrifuge at 500g for 5 minutes and discard the supernatant. Dilute 10% SDS solution (sodium dodecyl sulfate, sodium dodecyl sulfate, model B548118, purchased from Biotech) with PBS to a concentration of 1%, add 150μL 1% SDS solution to resuspend the cell pellet, and then place it in a metal bath at 100℃ and boil for 15-20 minutes until the cell solution is no longer viscous. After boiling, the cell samples were placed on ice, and after cooling, protease inhibitors (model K1007-1ML, purchased from APExBIO) were added. After centrifugation at 11000 rpm for 10 minutes, 150 μL 2×Loading was added to the supernatant and mixed evenly, and placed in a -80°C refrigerator for use.

[0621] (2) Glue running

[0622] Take 4-20% precast gel, take out WB (Western Blot) sample and thaw it, then load 10 μL WB sample and 5 μL protein marker (model: 26619, purchased from Invitrogen) respectively. Run at 80V for 30 minutes, then run at 140V for about 1 hour until loading is reached.

[0623] (3) Transfer

[0624] After running, take out the gel and place it in the order of sponge-filter paper-gel-cellulose membrane-filter paper-sponge. Transfer the membrane on ice and transfer the membrane at 300mA for 1.5 hours.

[0625] (4) Closed

[0626] After the transfer, the cellulose membrane was removed and placed in 5% skim milk powder (model 232100, purchased from BD) for blocking at room temperature for 1 hour. After the blocking was completed, the membrane was washed 5 times with TBST (TBS+Tween) solution, each time for 5 minutes.

[0627] (5) Incubation with primary antibody

[0628] After washing, the membrane was incubated with Anti-SSTR2 (Rb mAb, model Ab134152, purchased from abcam) and Anti-GAPDH (Mouse mAb, GAPDH (7E4) Mouse mAb, model 200306-7E4, purchased from ZENBIO) antibodies overnight at 4° C. After incubation, the membrane was washed 5 times with TBST solution, each time for 5 minutes.

[0629] (6) Incubation with secondary antibody

[0630] Mouse-HRP (Anti-mouse IgG, HRP-linked Antibody, Model 7076S, purchased from Cell Signaling) and Rabbit-HRP (Goat anti-rabbit IgG, Model XB-2301, purchased from Zhongshan Jinqiao) secondary antibodies were incubated at room temperature for 1 hour. After incubation, the membranes were washed 5 times with TBST solution, 5 minutes each time.

[0631] (7) Color exposure

[0632] Mix the A solution and B solution of ECL luminescent solution (model C510043, purchased from Bio-Tech) in a ratio of 1:1 (prepare and use immediately) to obtain fresh ECL working solution. After taking out the cellulose membrane from the TBST washing solution, use filter paper to absorb the water (generally use clean tweezers to clamp the cellulose membrane and let one corner of the membrane touch the filter paper); place the cellulose membrane on the imager, evenly add fresh ECL working solution, remove bubbles, and start exposure.

[0633] The experimental results are as follows Figure 5 As shown, the SSTR2 positive cells are H524, A172, H2171, U87, and H69 according to their expression levels from high to low, and the negative cells are A549.

[0634] [Cellular uptake experiment]

[0635] (1) Cell counting

[0636] The suspended H524 cells were blown evenly, counted, and 300,000 cells were taken from each tube. The culture medium of the adherent A549 cells was discarded, washed once with phosphate buffered saline (PBS) (Cat. No. 10010-031, purchased from GIBCO), and then digested with trypsin for two minutes, then the trypsin was discarded, and F12K complete medium Ham's F12K (Cat. No. 21127030, purchased from GIBCO) was added to blow the cells into a uniform suspension, counted, and 300,000 cells were taken from each tube.

[0637] (2) Intake

[0638] After centrifuging the cells at 500g for 5 minutes, discard the culture supernatant, and add 200μL PBS to resuspend the cell pellet. Centrifuge again for 5 minutes, discard the supernatant, add 150μL of culture medium containing 10μM of the above-prepared polypeptide compound and FITC conjugate ("polypeptide-fluorescein"), and incubate in a 37°C incubator for 30 minutes. Centrifuge again for 5 minutes, discard the culture supernatant containing the polypeptide, resuspend in 200μL PBS, and centrifuge at 500g for 5 minutes. Repeat this step twice, that is, wash the cells twice with PBS and resuspend in 200μL PBS, and blow evenly.

[0639] (3) Detection

[0640] A flow cytometer (model CytoFLEX, purchased from Beckman) was used to select the FITC channel to detect the fluorescence intensity. The detection results are shown in Table 3 and FIG. 6A to FIG. 6D shown.

[0641] Table 3 Cellular uptake ratio of peptide-fluorescein conjugates

[0642]

[0643]

[0644] From the above table 3 and FIG. 6A to FIG. 6D It can be seen that the polypeptide compounds OCT, A-1, A-2, A-3, A-4, A-5, A-6, A-7, A-8, A-9, A-12, A-13, A-15, A-17, B-1, B-2, B-3, B-4, B-5, B-6, B-7, B-8, B-9, and B-17 have specific uptake effects in the cell line H524 with high expression of SSTR2.

[0645] [Stability test under rat plasma conditions]

[0646] (1) Storage fluid preparation

[0647] Compounds OCT, A-1, A-2, B-1 and B-2 were prepared as 10 mM stock solutions.

[0648] (2) Obtaining plasma samples

[0649] Ethylenediaminetetraacetic acid dipotassium (EDTA-2K) was added to rat plasma, and compounds OCT, A-1, A-2, B-1, and B-2 were diluted to 1 μM using rat plasma. Duplicate samples were prepared and placed in a 37° C. water bath.

[0650] (3) Detection

[0651] At time 0, take out an appropriate amount of sample, add 15 times the volume of the incubation system of methanol as a precipitant to precipitate the protein, then centrifuge at 15700 rpm at 6°C for 10 min, take 150 μL of the supernatant, add 150 μL of water, vortex mix, and analyze by LC-MS / MS.

[0652] Repeat the above test at 5, 15, 30, and 60 minutes.

[0653] The obtained rat plasma stability curve is as follows 7A to 7E The remaining amount of the original drug (%) was calculated using the ratio of the mean of the peak area ratios at different incubation times to the initial time, and the calculation results are shown in Table 4 below.

[0654] Table 4 Rat plasma stability

[0655]

[0656] Depend on 7A to 7E As can be seen from Table 4 above, compounds OCT, A-1, A-2, B-1, and B-2 can remain undegraded in plasma for more than 3 hours, and their stability allows them to be used as targeting peptides to construct PDC products.

[0657] [In vitro killing experiment]

[0658] (1) Cell culture

[0659] The following cells were revived and cultured according to experimental requirements: small cell lung cancer cells H524 and H69, glioma cells U87 and A172, and non-small cell lung cancer A549 cells for control.

[0660] (2) Cell plating

[0661] After digestion, adherent cells were centrifuged and resuspended, counted, and diluted to 3-5×10 4 / mL, and then plated in a 96-well plate, with 100 μL per well in columns 2 to 12. The suspended cells were directly centrifuged and resuspended, then counted and diluted to 1 to 2 × 10 5 After the concentration reached 1 / ml, the cells were plated in a 96-well V-shaped plate, with 100 μL per well in columns 2 to 12. The cells were cultured in an incubator for 24 h.

[0662] (3) Conjugate gradient configuration

[0663] The conjugate was weighed and dissolved in dimethyl sulfoxide (DMSO) to a 20 mM system, and diluted 1000× with the corresponding cell culture medium to obtain a maximum concentration of 20 μM, and then diluted 4-fold in a culture medium containing 1‰ DMSO. Ten gradients of conjugates with a 4-fold dilution starting at 20 μM were obtained.

[0664] (4) Conjugate treatment

[0665] 100 μL of conjugates of different concentrations were added to columns 3 to 12 of the 96-well plate, and 100 μL of 1‰ DMSO culture medium was added to column 2. The plates were incubated in an incubator for 6 h.

[0666] (5) Removal of conjugates

[0667] Directly remove the medium containing the conjugate from the adherent cells, add 100 μL of the normal corresponding medium, and repeat the operation twice to ensure that there is no free conjugate left. Suspended cells need to be centrifuged at 1000 rpm for 2 minutes to allow the cells to sink, and then perform the same washing operation. After washing, transfer the cells in the 96-well V-plate to a 96-well plate. Finally, add 100 μL of medium to the first column. Continue to culture in the incubator for 72 hours.

[0668] (6) CCK8 detection

[0669] 10 μL CCK8 reagent was added to each well, incubated in a dark incubator for 2 to 6 hours, and detected by an enzyme reader (model INFINITE200PRO, purchased from Tecan). The experimental results were recorded in Figures 8A-1 to 8E-15 and Table 5 below.

[0670] Table 5 In vitro killing test results of PDC conjugates

[0671]

[0672]

[0673] Depend on Figures 8A-1 to 8E-15As can be seen from Table 5 above, for the high-expressing cell line H524 (small cell lung cancer cells), the in vitro killing effect of PDC is more significant, especially DM1-A-1, DM1-A-2, DM1-A-5, DM1-A-6, DM1-A-7, DM1-A-8, DM1-A-11, DM1-A-17, DM1-A-18, DM1-B-1, and DM1-B-2. The in vitro killing effect on H69 (small cell lung cancer cells) and U87 (glioma cells) was second only to H524, and the overall killing effect on H69 cells was good. In U87 cells, DM1-A-1, DM1-A-2, DM1-A-4, DM1-A-6, DM1-A-7, DM1-A-8, DM1-A-9, DM1-A-17, DM1-B-2, DM1-B-3, DM1-B-5, DM1-B-6 and DM1-B-7 had significant in vitro killing effects. The killing effect on A172 (glioma cells) was weaker than that of the other three cells (H524, H69 and U87), but DM1-A-1, DM1-A-6, DM1-A-7, DM1-A-8 and DM1-A-17 also had significant in vitro killing effects on A172.

[0674] [Tumor cell xenografts]

[0675] (1) Animal husbandry

[0676] Balb / c nude female mice aged 6-7 weeks, weighing 16-18 grams, were provided by Weitong Lihua Laboratory Animal Co., Ltd. After the mice arrived, they were kept in the experimental environment for 3-7 days before the experiment began. The experimental environment was an SPF animal room, and IVC (independent ventilation system) cages were used for feeding (5 mice per cage). The cages, feed and drinking water were changed twice a week. The breeding environment was 20-26℃ temperature and 40-70% humidity, and the light cycle was 12 hours of light and 12 hours of no light.

[0677] (2) H524, H69, and U87 cell culture

[0678] H524, H69: Add 10% fetal bovine serum, 100U / mL penicillin and 100μg / mL streptomycin to RPMI-1640 medium and culture in a 37℃ 5% CO2 incubator. Centrifuge and resuspend at a 1:3 ratio for passage two to three times a week. When the cell number reaches the required level, harvest the cells and adjust the cell concentration for experimental inoculation.

[0679] U87: DMEM high-glucose medium with 10% fetal bovine serum, 100U / mL penicillin and 100μg / mL streptomycin, cultured in a 37℃ 5% CO2 incubator. Routine treatment and subculture once or twice a week. When the cell saturation is 80%-90% and the number reaches the requirement, collect the cell count and adjust the cells to the appropriate concentration for animal inoculation.

[0680] (3) Tumor inoculation

[0681] 0.2 mL of each of the above cells was subcutaneously inoculated on the right back of each mouse. When the tumor grew to 150-200 mm 3 The animals were randomly divided into groups and given medication according to tumor volume and animal body weight.

[0682] (4) Screening of xenotransplantation conditions

[0683] 5×10 6 or 10×10 6 / mouse, and the axillary tumors were implanted. The time and size of tumor formation were used to determine the subsequent detection of H524 and H69. 6 / mouse, U87 used 1×10 5 / mouse was implanted with tumor.

[0684] (5) MTD detection of test samples: Perform MTD detection according to the dosage of the experimental plan to ensure that the mice can tolerate the dosage.

[0685] (6) Pharmacodynamic testing

[0686] a) Tumor Implantation

[0687] Tumors were implanted in 1.5 times the number of mice required for the experiment. One week later, the tumors generally grew to 150-200 mm. 3 The number of mice required for the experiment was selected according to the tumor volume and animal weight, and the mice were randomly divided into groups.

[0688] b) Administration

[0689] Drugs were administered separately, and the drugs were dissolved in 10% propylene glycol + 90% 5mM citric acid buffer (pH = 4). On the day of grouping, intravenous administration was performed, and the frequency of administration was designed according to the current round of experiments.

[0690] c) Observation

[0691] The health status and mortality of the animals were monitored daily. Routine examinations included observing tumor growth and the effect of drug treatment, measuring the long and short diameters of the tumors with a vernier caliper, and calculating the tumor volume (V = 0.5a × b 2, a and b are the long diameter and short diameter, respectively), to examine whether the tumor growth is inhibited, delayed or cured. Adjust the dosage according to the efficacy. At the same time, observe the impact on the daily behavior of the animals, such as behavioral activities, food and water intake, weight changes (weight is measured at the same time as each tumor volume test), appearance signs or other abnormal conditions. Based on the number of animals in each group, record the number of deaths and side effects of animals in the group.

[0692] (d) Termination of the experiment

[0693] The average tumor volume in the group exceeded 2,000 mm 3 Or the volume of a single tumor exceeds 3,000mm 3 , or have serious illness or pain, must be euthanized.

[0694] The first round of pharmacodynamic testing was to test the efficacy of DM1-A-1, DM1-A-2, and DM1-B-2 in H524 xenografts. Among the 40 tumor-bearing mice, 25 were selected based on tumor volume and animal weight and randomly divided into 5 groups, with 5 mice in each group (N=5). The grouping and dosing are shown in Table 6. The weight changes and tumor growth of the mice were recorded during the experiment, as shown in Tables 7 to 8, and Fig.9A , Fig. 9B , Fig. 10A and Fig. 10B As shown, the inhibitory effect of the conjugate on tumors was calculated and presented, as shown in Table 9 and Fig.11 .

[0695] Table 6 Drug administration in mice

[0696]

[0697] Table 7 Changes in mouse body weight during the experiment (body weight and body weight change rate compared to Day 0)

[0698]

[0699]

[0700] Table 8 Tumor growth in mice during the experiment (tumor size, growth rate compared to Day 0 and final tumor mass)

[0701]

[0702]

[0703] Table 9 Inhibition of H524 xenograft tumors by the conjugates in mice

[0704] Experimental Group T / C(%) Vehicle control group (D0, D8) 100.00 DM1 (0.75 mg / kg, D0, D8) 27.35 DM1-A-1 (1.96 mg / kg, D0, D8) 90.75 DM1-A-2 (1.96 mg / kg, D0, D11) 39.29 DM1-B-2 (1.94 mg / kg, D0, D8) 74.87

[0705] In this experiment, DM1-A-2 had a relatively good inhibitory effect on tumors, close to that of DM1, but it also caused weight loss when administered in the early stages.

[0706] The second round of pharmacodynamic testing was a H524 xenograft repeat test to test the efficacy of DM1-A-2, screen the efficacy of DM1-A-6, DM1-A-7, DM1-A-9, and DM1-A-17, and screen the efficacy and dosage of DM1-A-3. Among the 45 tumor-implanted mice, 32 were selected based on tumor volume and animal weight and randomly divided into 8 groups, with 4 mice in each group (N=4). The grouping and dosing are shown in Table 10. The weight changes and tumor growth of the mice were recorded during the experiment, as shown in Tables 11 to 12, and Fig. 12A , Fig. 12B , Fig.13A and Fig. 13B As shown in Table 13, the inhibitory effect of the conjugate on tumors was calculated and presented. Fig.14 .

[0707] Table 10: Drug administration in mice

[0708]

[0709]

[0710] Table 11 Changes in mouse body weight during the experiment (body weight and body weight change rate compared to Day 0)

[0711]

[0712]

[0713] Table 12 Tumor growth in mice during the experiment (tumor size, growth rate compared to Day 0 and final tumor mass)

[0714]

[0715]

[0716] Table 13 Inhibition of H524 xenograft tumors by the conjugates in mice

[0717] Experimental Group T / C(%) Vehicle control group (D0) 100.00 DM1-A-3 (3.90 mg / kg, D0) 0.00 DM1-A-3 (1.95 mg / kg, D0, 8) 52.36 DM1-A-2 (1.96 mg / kg, D0, 8) 47.42 DM1-A-9 (2.037 mg / kg, D0) 4.03 DM1-A-17 (2.03 mg / kg, D0, 8) 49.92 DM1-A-6 (2.00 mg / kg, D0) 90.50 DM1-A-7 (1.90 mg / kg, D0, 8) 40.62

[0718] In this experiment, the efficacy of DM1-A-2 was repeated, and the inhibitory effect on tumors was relatively good, close to the effect of the first round, but no weight loss occurred during administration. DM1-A-6 had almost no inhibitory effect, DM1-A-7, DM1-A-17 and DM1-A-3 had good tumor inhibitory effects, and DM1-A-9 was able to almost completely inhibit tumors, and there was no obvious weight loss. DM1-A-3 at twice the dose was able to quickly and completely inhibit tumors, but it also caused weight loss during the early administration.

[0719] The third round of pharmacodynamic testing was to test the efficacy of DM1-A-3, DM1-A-9, DM1-A-11, DM1-A-15, DM1-A-8, and DM1-A-18 in H69 xenografts, and the dose screening was performed on the efficacy window of DM1-A-3 and DM1-A-9. Among the 105 tumor-implanted mice, 70 were selected based on tumor volume and animal weight and randomly divided into 14 groups, with 5 mice in each group (N=5). The grouping and dosing are shown in Table 14. The weight changes and tumor growth of the mice were recorded during the experiment, as shown in Tables 15 to 16, and FIG. 15A to FIG. 15D ,as well as FIG. 16A to FIG. 16D As shown in Table 17, the inhibitory effect of the conjugate on tumors was calculated and presented. FIG. 17A to FIG. 17C shown.

[0720] Table 14 Drug administration in mice

[0721]

[0722]

[0723]

[0724]

[0725]

[0726]

[0727]

[0728] Table 17 Inhibition of H69 xenograft tumors by the conjugates in mice

[0729] Experimental Group T / C(%) Vehicle control group (QW) 100.00 DM1 (0.75 mg / kg, D0) die DM1 (0.375 mg / kg, QW) 70.36 DM1 (0.1875 mg / kg, QW) 102.54 DM1-A-3 (1.95 mg / kg, QW) 9.13 DM1-A-3 (0.975 mg / kg, QW) 70.53 DM1-A-3 (0.4875 mg / kg, QW) 88.79 DM1-A-9 (2.037 mg / kg, QW) 4.96 DM1-A-9 (1.0185 mg / kg, QW) 54.96 DM1-A-9 (0.50925 mg / kg, QW) 88.50 DM1-A-11 (1.95 mg / kg, QW) 11.72 DM1-A-15 (2.02 mg / kg, QW) 48.22 DM1-A-8 (2.04 mg / kg, QW) 1.43 DM1-A-18 (2.12 mg / kg, QW) 64.08

[0730] In this experiment, except for 0.75 mg / kg of DM1, which caused mice to lose weight rapidly and die, there was no obvious weight loss in mice at other doses or under the action of PDC conjugates. Among them, DM1-A-8 has the best efficacy and can almost completely inhibit tumors. DM1-A-3, DM1-A-9, and DM1-A11 also have very good inhibitory effects on tumors. DM1-A-15 and DM1-A-18 have certain tumor inhibitory effects. DM1-A-3 and DM1-A-9 also underwent dose escalation to explore the efficacy window. It was found that there was almost no efficacy under the 1 / 4 dose condition, and there was a certain efficacy under the 1 / 2 dose condition. Except for DM1-A-3 at the 1 / 2 dose, the efficacy was similar to the equimolar DM1 dose group. The efficacy of 1 / 2 dose DM1-A-9, 1 / 4 dose DM1-A-3 and DM1-A-9 were all stronger than the equimolar DM1 dose group.

[0731] The fourth round of pharmacodynamics testing was to test the efficacy of DM1-A-3 in U87 xenografts. 20 mice were selected from 30 tumor-bearing mice based on tumor volume and animal weight, and randomly divided into 4 groups, with 5 mice in each group (N=5). The grouping and dosing are shown in Table 18. The weight changes and tumor growth of the mice were recorded during the experiment, as shown in Tables 19 to 20, and Fig.18A and Fig.18B ,as well as Fig.19A and Fig.19B As shown in Table 21, the inhibitory effect of the conjugate on tumors was calculated and presented. Fig. 20 shown.

[0732] Table 18: Drug administration in mice

[0733]

[0734] Table 19 Changes in mouse body weight during the experiment (body weight and body weight change rate compared to Day 0)

[0735]

[0736]

[0737] Table 20 Tumor growth in mice during the experiment (tumor size, growth rate compared to Day 0 and final tumor mass)

[0738]

[0739]

[0740] Table 21 Inhibition of U87 xenograft tumors by the conjugates in mice

[0741] Experimental Group T / C(%) Vehicle control group (D0) 100.00 DM1 (0.75 mg / kg, D0) 22.00 DM1-A-3 (3.90 mg / kg, D0) 0.75 DM1-A-3 (1.95 mg / kg, D0) 34.52

[0742] In this experiment, two doses of DM1-A-3 were tested for efficacy in U87 xenograft tumors. When the dose was equal to that of DM1, the effect of DM1-A-3 was close to that of DM1, and the weight changes of mice after administration were also similar. The double dose of DM1-A-3 showed very good efficacy and could quickly and almost completely inhibit tumors, but the early administration also caused weight loss.

Claims

1. A compound or a pharmaceutically acceptable salt thereof, wherein the compound has a structure of general formula I: in, R 1 To R 7 , and R 15 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkylcarbonyl, halogen, cyano, hydroxyl, carboxyl, -C(O)NR 10 R 11 , phenyl, naphthyl, biphenyl, Polyethylene glycol, and C5-C containing one or more heteroatoms selected from N, S and O 10 Aromatic heterocyclic ring; or R 15 Condensed with adjacent phenyl groups to form naphthyl groups; these groups are optionally substituted by one or more substituents R 12 replace; R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and R 9 With R 16 are the same or different and are independently selected from S, -CH2- and -C(CH3)2-; or R 9 With R 16 Together they represent -SS-, -S-CH2-, -S-CH(CH3)-, -SC(CH3)2-, -CH2-S-, -CH(CH3)-S-, -C(CH3)2-S-, -CH2-CH=CH-CH2- and as well as R 10 To R 14 are the same or different and are independently selected from H, C1-C 20 Alkyl, C2-C 20 Alkenyl, C2-C 20 Alkynyl, C1-C 20 Halogenated alkyl, C1-C 20 Cyanoalkyl, C1-C 20 Hydroxyalkyl, C1-C 20 Alkoxy, C1-C 20 Alkylthio, C1-C 20 Halogenated alkoxy, C1-C 20 Alkoxyalkyl, C3-C 20 Cycloalkyl, C3-C 20 Cycloalkylalkyl, C3-C 20 Halogenated cycloalkyl, C1-C 20 Alkoxycarbonyl, halogen, cyano, hydroxy, carboxyl, phenyl, and p represents an integer selected from 1 to 4; and * indicates the point of attachment of the identified group to the rest of Formula I.

2. The compound according to claim 1, wherein R 1 is selected from H and phenyl; R 2 is selected from phenyl, hydroxyphenyl and naphthyl; R 3 is selected from H and -C(O)-CH3; R 4 H, or R 4 With R 17 Together with the -N-CH-CH2- in between, it forms R 5 is selected from hydroxyl and methyl; R 6 and R 7 are different and are independently selected from methyl, hydroxy, H and naphthyl; R 8 Selected from -CH2OH, -C(O)-NR 13 R 14 and R 9 With R 16 are the same or different and are independently selected from S and -CH2-; and p is 1 or 2.

3. The compound according to claim 1 or 2, having a structure selected from any one of the following:

4. A compound or a pharmaceutically acceptable salt thereof, wherein the compound has the structure of general formula II: in, Group R 1 To R 17 , and p has the same definition as in claim 1, X represents a linking moiety and is selected from one of the following groups or any combination thereof: -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20 Cycloalkyl and its isomers, phenyl, naphthyl, maleimide groups and their hydrides, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl, ** indicates that the identified group is connected to a leaving group.

5. The compound according to claim 4, wherein The connecting part is 6. A conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate has a structure of general formula III: in, Group R 1 To R 17 , and p has the same definition as in claim 1, X represents a linking moiety and is selected from one of the following groups or any combination thereof: -S-**, -SS-**, -C(O)-O-**, -C(O)-NH-**, -NH-**, -O-**, -OC(O)-O-**, -NH-CR 19 R 20 -NH-**, -NH-C(O)-NH-, -OCH2-CH2-**, C1-C 10 Alkyl and its isomers, C3-C 20 Cycloalkyl and its isomers, phenyl, naphthyl, maleimide groups and their hydrides, glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, wherein R 19 and R 20 are the same or different and are independently selected from H and C1-C6 alkyl; and Y represents an active moiety and is maytansine (DM1), ** indicates the connection point between the identified group and Y.

7. The conjugate according to claim 6, which has a structure selected from any one of the following:

8. A pharmaceutical composition comprising the conjugate according to claim 6 or 7 or a pharmaceutically acceptable salt thereof, and A pharmaceutically acceptable carrier, diluent or excipient.

9. Use of the pharmaceutical composition according to claim 8, or the conjugate according to claim 6 or 7 or a pharmaceutically acceptable salt thereof, or the compound according to any one of claims 1 to 5 for preparing a medicament for treating a tumor or reducing tumor volume; In particular, the tumor expresses the somatostatin receptor (SSTR2); In particular, the tumor is a neuroendocrine cancer; In particular, the tumor is selected from lung cancer (eg small cell lung cancer), glioma, pancreatic cancer, breast cancer, colorectal cancer, ovarian cancer, colorectal cancer, bladder cancer, prostate cancer, cervical cancer, kidney cancer, leukemia, central nervous system cancer, myeloma and melanoma.

10. A method for treating a tumor or reducing tumor volume, comprising administering an effective amount of the pharmaceutical composition according to claim 8, or the conjugate according to claim 6 or 7, or a pharmaceutically acceptable salt thereof, to an individual in need thereof.

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