Somatostatin peptide analogue, chelate and use thereof

Abstract

A somatostatin peptide analogue. The somatostatin peptide analogue has a cyclic or non-cyclic hexapeptide unit, and the residue sequence at positions 2 to 5 in the hexapeptide unit is: -X1-(D / L-)Trp-X2-X3-; or the somatostatin peptide analogue has a cyclic or non-cyclic octapeptide unit, and the residue sequence at positions 2 to 7 in the octapeptide unit is: -Cys-X1-(D / L-)Trp-X2-X3-Cys-; wherein X1 is an α-amino acid residue having an aryl group on a Cα side chain, and X3 is an α-amino acid residue, and X2 is a Lys-variant residue: formula (I). The somatostatin peptide analogue breaks the "taboo" that Lys9 residues cannot be substituted, otherwise, the biological activity will be greatly reduced. By varying the Lys9 residues in the somatostatin peptide analogue, the Lys9 residues are varied, which enables the somatostatin peptide analogue to exhibit higher (at least similar) binding affinity to SSTR2.

Description

Somatostatin peptide analogs, chelates and their applications Technical Field

[0001] The present invention belongs to the technical field of anti-tumor compound synthesis, and in particular relates to a somatostatin peptide analog, a chelate and applications thereof. Background Art

[0002] Somatostatin (SST) is a cyclic polypeptide composed of 14 or 28 amino acids (SST-14 or SST-28). Its primary biological effect is through targeted binding to somatostatin receptors (SSTRs), inhibiting the secretion of other hormones in the body, such as insulin, glucagon, thyroid-stimulating hormone, growth hormone, prolactin, and cholecystokinin, playing a key role in endocrine regulation. SSTRs are G protein-coupled receptors (GPCRs) that inhibit the Gi / o family of G proteins to signal and prevent the release of hormone-secreting vesicles. The SSTR family includes five distinct subtypes: SSTR1, SSTR2, SSTR3, SSTR4, and SSTR5. Tissue expression profiles of different receptors vary. Neuroendocrine tumors (NETs) often overexpress SSTR2, making this subtype a primary target for somatostatin agonists. These properties make agonists clinically useful both as tracers for diagnostic imaging of diseases and for the treatment of conditions caused by hormonal disturbances, including carcinoid syndrome, gigantism, acromegaly, hyperthyroidism, and Cushing's disease.

[0003] Currently, all FDA-approved somatostatin agonists are peptide analogs of SST-14, including octreotide and lanreotide, which target SSTR2, and pasireotide, which targets SSTR1, SSTR2, SSTR3, and SSTR5. Numerous studies have shown that different SSTR subtypes play important roles in a variety of tumor types beyond NETs, ​​necessitating the use of more selective pharmacological molecules to probe subtype-specific functions.

[0004] In this patent application, natural somatostatin is referred to as a tetradecapeptide (Formula V) having the following structure:

[0005] As used in this patent application, somatostatin peptide analogs refer to linear or cyclic peptide derivatives of naturally occurring SST-14, i.e., wherein one or more residues of the sequence of Formula V are deleted or replaced with one or more other amino acids, or one or more functional groups are deleted or replaced with one or more other functional groups. Generally speaking, the term encompasses all modified derivatives of naturally occurring SST-14 comprising the sequence of Formula V, which have high affinity for at least one somatostatin receptor subtype and reach nmol levels.

[0006] The extremely short half-life of natural somatostatin has limited its clinical application. To address this issue, a large number of somatostatin analogs were synthesized starting in the 1970s. These studies not only revealed the role of key residues Trp8-Lys9 but also laid the foundation for the clinical application of somatostatin analogs. The Sandoz team used an approach that systematically extended the N- and C-termini of the hexapeptide lead structure, Cys-Phe-DTrp-Lys-Thr-Cys. In 1980, they successfully synthesized and characterized a very stable and highly active analog, octreotide, with the following structure: H-DPhe-Cys-Phe-DTrp-Lys-Thr-Cys-Thr-Ol. This compound inhibited growth hormone secretion more effectively than the two natural somatostatins and exhibited sufficient stability in vivo, resulting in its selection for clinical studies. Octreotide was first approved by the FDA in 1988 for the treatment of acromegaly and carcinoid tumors. In addition, oxyoctreotide (an octreotide analogue) is coupled with positron-emitting isotope metal ions to form a complex that can be used for tumor focused scanning imaging to detect tumor cells that highly express somatostatin receptors. Oxyoctreotide can also be coupled with beta-ray-emitting isotope metal ions to form a complex to kill tumor cells that highly express somatostatin receptors.

[0007] Other somatostatin peptide analogs are also on the market, including lanreotide, vapreotide, and pasireotide. Numerous studies have demonstrated that the core binding site of SST-14 to its receptor is composed of two residues, Trp8 and Lys9. These two key binding residues are crucial and indispensable for its biological activity and were subsequently demonstrated to be the core pharmacophore for somatostatin peptide analogs binding to the somatostatin receptor. In fact, numerous studies examining Lys9 residue substitutions have failed to improve receptor binding affinity and bioactivity. In fact, most studies have shown that substitutions at Lys9 have resulted in a loss of receptor binding affinity or bioactivity. These studies have led to a "taboo" conclusion: "Lys9 cannot be substituted, even with closely related residues such as Arg or Orn. ​​Once substituted, its bioactivity is significantly reduced."

[0008] Summary of the Invention

[0009] A series of studies have shown that the SSTR2 protein structure reveals that the SSTR2 pocket bound to Lys9 still has ample room for other atoms. By modifying the Lys9 group, it is possible to increase steric tension and limit the number of possible side chain conformations, thereby facilitating more effective binding of drug molecules to SSTR2.

[0010] Therefore, the object of the present invention is to provide a somatostatin peptide analog, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable complex thereof, or a pharmaceutically acceptable chelate thereof,

[0011] The somatostatin peptide analogues have a cyclic or non-cyclic hexapeptide unit, and the residue sequence at positions 2 to 5 in the hexapeptide unit is: -X1-(D / L-)Trp-X2-X3-;

[0012] Alternatively, the somatostatin peptide analogue has a cyclic or non-cyclic octapeptide unit, and the residue sequence at positions 2 to 7 in the octapeptide unit is: -Cys-X1-(D / L-)Trp-X2-X3-Cys-;

[0013] wherein X1 is an α-amino acid residue having an aromatic group on the Cα side chain, X3 is an α-amino acid residue, and X2 is a Lys-variant residue:

[0014] n is 0 or 1;

[0015] A1, A2, A3 are independently selected from C(R)2, O, S, NR, C=O, C=S, C=NR and C=C(R)2;

[0016] R is independently selected from R1, -R'OR1, -R'SR1, -R'N(R1)2, -R'C(O)R1, -R'S(O)R1, -R'S(O)2R1, -R'C(O)N(R1)2, -R'N(R1)C(O)R1, -R'C(O)OR1 and -R'OC(O)R1;

[0017] or independently, R within C(R)2 at position A2 is connected to R within C(R)2 at position A1 or position A3 to form a double bond or triple bond;

[0018] Alternatively, independently, R within C(R)2 forms a 3-8 membered carbocyclic ring with its own R within C(R)2, or with an adjacent R within C(R)2, or with an R within a C(R)2 separated by one C(R)2, or forms a 3-8 membered heterocyclic ring containing 1-3 heteroatoms selected from N, O, and S;

[0019] R' is a linker, a C1-C6 alkylene group, a C2-C6 alkenylene group, a C2-C6 alkynylene group, a C3-C8 cycloalkylene group, a C3-C8 cycloalkenylene group, a halogenated C1-C6 alkylene group, a halogenated C2-C6 alkenylene group, a halogenated C2-C6 alkynylene group, a halogenated C3-C8 cycloalkylene group, a halogenated C3-C8 cycloalkenylene group, a (C6-C8)arylene group, a 3-8 membered heterocyclylene group having 1 to 3 heteroatoms selected from N, O, and S, or a 3-8 membered heteroarylene group having 1 to 3 heteroatoms selected from N, O, and S;

[0020] R1 is independently selected from H, halogen, CN, NO2, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, halogenated C1-C6 alkyl, halogenated C2-C6 alkenyl, halogenated C2-C6 alkynyl, halogenated C3-C8 cycloalkyl, halogenated C3-C8 cycloalkenyl, (C6-C8)aryl, 3-8 membered heterocyclyl having 1 to 3 heteroatoms selected from N, O, and S, and 3-8 membered heteroaryl having 1 to 3 heteroatoms selected from N, O, and S;

[0021] When n is 0, R in C(R)2 at the A1 position, A2 position, and A3 position are not H at the same time.

[0022] In the present invention, "R within C(R)2 forms a 3- to 8-membered carbocyclic ring together with R within itself, or with R within an adjacent C(R)2, or with R within a C(R)2 separated by one C(R)2, or forms a 3- to 8-membered heterocyclic ring containing 1 to 3 heteroatoms selected from N, O, and S", which includes the following four forms:

[0023] Independently, the two Rs in C(R)2 at the A1 position, A2 position, or A3 position together form a 3- to 8-membered carbocyclic ring, or a 3- to 8-membered heterocyclic ring containing 1 to 3 heteroatoms selected from N, O, and S;

[0024] Alternatively, independently, the R within the C(R)2 at position A1 and the R within the C(R)2 at position A3 are linked together to form a ring such that A1, A2, and A3 form a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring containing 1 to 3 heteroatoms selected from N, O, and S;

[0025] Alternatively, independently, the R within the C(R)2 at position A1 and the R within the C(R)2 at position A2 are linked together to form a ring such that A1 and A2 form a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring containing 1 to 3 heteroatoms selected from N, O, and S;

[0026] Alternatively, independently, R within C(R)2 at position A2 and R within C(R)2 at position A3 are connected together to form a ring so that A2 and A3 form a 3-8 membered carbocyclic ring or a 3-8 membered heterocyclic ring containing 1-3 heteroatoms selected from N, O, and S.

[0027] In a preferred example,

[0028] The somatostatin peptide analogues have a cyclic or non-cyclic hexapeptide unit, and the residue sequence at positions 1 to 6 in the hexapeptide unit is: Xa-X1-(D / L-)Trp-X2-X3-Xb;

[0029] The somatostatin peptide analogues have a cyclic or non-cyclic octapeptide unit, and the residue sequence at positions 1 to 8 in the octapeptide unit is: X0-Cys-X1-(D / L-)Trp-X2-X3-Cys-Xb;

[0030] Wherein Xa and Xb are amino acid residues; X0 is an α-amino acid residue having an aromatic group on the Cα side chain.

[0031] In a preferred example,

[0032] X1 is a Phe, Tyr, Trp or Phg (phenylglycine, phenylglycinamide) residue; X3 is a Thr, Ser, Cys or tyrosylbenzyl residue.

[0033] In a preferred example, the somatostatin peptide analog has a cyclic octapeptide unit structure represented by the following general formula (I):

[0034] n is 0 or 1;

[0035] A1, A2, A3 are independently selected from C(R)2, O, S, NR, C=O, C=S, C=NR and C=C(R)2;

[0036] R is independently selected from R1, -R'OR1, -R'SR1, -R'N(R1)2, -R'C(O)R1, -R'S(O)R1, -R'S(O)2R1, -R'C(O)N(R1)2, -R'N(R1)C(O)R1, -R'C(O)OR1 and -R'OC(O)R1;

[0037] or independently, R within C(R)2 at position A2 is connected to R within C(R)2 at position A1 or position A3 to form a double bond or triple bond;

[0038] Alternatively, independently, R within C(R)2 forms a 3-8 membered carbocyclic ring with its own R within C(R)2, or with an adjacent R within C(R)2, or with an R within a C(R)2 separated by one C(R)2, or forms a 3-8 membered heterocyclic ring containing 1-3 heteroatoms selected from N, O, and S;

[0039] R' is a linker, a C1-C6 alkylene group, a C2-C6 alkenylene group, a C2-C6 alkynylene group, a C3-C8 cycloalkylene group, a C3-C8 cycloalkenylene group, a halogenated C1-C6 alkylene group, a halogenated C2-C6 alkenylene group, a halogenated C2-C6 alkynylene group, a halogenated C3-C8 cycloalkylene group, a halogenated C3-C8 cycloalkenylene group, a (C6-C8)arylene group, a 3-8 membered heterocyclylene group having 1 to 3 heteroatoms selected from N, O, and S, or a 3-8 membered heteroarylene group having 1 to 3 heteroatoms selected from N, O, and S;

[0040] R1 is independently selected from H, halogen, CN, NO2, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, halogenated C1-C6 alkyl, halogenated C2-C6 alkenyl, halogenated C2-C6 alkynyl, halogenated C3-C8 cycloalkyl, halogenated C3-C8 cycloalkenyl, (C6-C8)aryl, 3-8 membered heterocyclyl having 1 to 3 heteroatoms selected from N, O, and S, and 3-8 membered heteroaryl having 1 to 3 heteroatoms selected from N, O, and S;

[0041] When n is 0, R in C(R)2 at the A1 position, A2 position, and A3 position are not H at the same time.

[0042] In a preferred example,

[0043] The R within the C(R)2 at position A1 and the R within the C(R)2 at position A3 are linked together to form a ring such that A1, A2, and A3 form a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring containing 1 to 3 heteroatoms selected from N, O, and S;

[0044] Preferably,

[0045] The R within C(R)2 at position A1 and the R within C(R)2 at position A3 are connected together to form a ring so that A1, A2, and A3 form a 5- to 7-membered carbocyclic ring or a 5- to 7-membered heterocyclic ring containing 1 to 2 heteroatoms selected from N, O, and S.

[0046] In a preferred example,

[0047] The R within C(R)2 at the A2 position is connected to the R within C(R)2 at the A1 position or the A3 position to form a double bond.

[0048] In a preferred example,

[0049] R' is a linker, a C1-C4 alkylene group, a C2-C4 alkenylene group, a C2-C4 alkynylene group, a C3-C7 cycloalkylene group, a C3-C7 cycloalkenylene group, a halogenated C1-C4 alkylene group, a halogenated C2-C4 alkenylene group, a halogenated C2-C4 alkynylene group, a halogenated C3-C7 cycloalkylene group, a halogenated C3-C7 cycloalkenylene group, a (C6-C8)arylene group, a 3-7 membered heterocyclylene group having 1 to 3 heteroatoms selected from N, O, and S, or a 3-7 membered heteroarylene group having 1 to 3 heteroatoms selected from N, O, and S;

[0050] R1 is independently selected from H, halogen, CN, NO2, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C7 cycloalkyl, C3-C7 cycloalkenyl, halogenated C1-C4 alkyl, halogenated C2-C4 alkenyl, halogenated C2-C4 alkynyl, halogenated C3-C7 cycloalkyl, halogenated C3-C7 cycloalkenyl, (C6-C8)aryl, 3-7 membered heterocyclyl having 1 to 3 heteroatoms selected from N, O, and S, and 3-7 membered heteroaryl having 1 to 3 heteroatoms selected from N, O, and S;

[0051] Preferably,

[0052] R' is a linker, a C1-C3 alkylene group, a C2-C3 alkenylene group, a C2-C3 alkynylene group, a C5-C6 cycloalkylene group, a C5-C6 cycloalkenylene group, a halogenated C1-C3 alkylene group, a halogenated C2-C3 alkenylene group, a halogenated C2-C3 alkynylene group, a halogenated C5-C6 cycloalkylene group, a halogenated C5-C6 cycloalkenylene group, a (C6-C8)arylene group, a 5-6 membered heterocyclylene group having 1 to 2 heteroatoms selected from N, O, and S, or a 5-6 membered heteroarylene group having 1 to 2 heteroatoms selected from N, O, and S;

[0053] R1 is independently selected from H, halogen, CN, NO2, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl, C5-C6 cycloalkenyl, halogenated C1-C3 alkyl, halogenated C2-C3 alkenyl, halogenated C2-C3 alkynyl, halogenated C5-C6 cycloalkyl, halogenated C5-C6 cycloalkenyl, (C6-C8)aryl, 5-6 membered heterocyclic group having 1 to 2 heteroatoms selected from N, O, and S, and 5-6 membered heteroaryl having 1 to 2 heteroatoms selected from N, O, and S.

[0054] Preferably, X2 is a Lys-variant residue:

[0055] Selected from:

[0056] Among them, Lys(c), Lys(d), Lys(f), Lys(h), Lys(j), Lys(l), Lys(m), Lys(r), and Lys(s) can also be any of their side chain stereoisomers. For example, Lys(d) can be any of the following two stereoisomers, namely:

[0057] Lys(j) can be any of the following four stereoisomers:

[0058] In a preferred embodiment of the present invention, the somatostatin peptide analogs are specifically:

[0059] Core amino acid sequence Phe-DTrp-Lys(a)-Thr,

[0060] Core amino acid sequence Phe-DTrp-Lys(b)-Thr,

[0061] Core amino acid sequence Phe-DTrp-Lys(d)-Thr,

[0062] Core amino acid sequence Phe-DTrp-Lys(j)-Thr,

[0063] The core amino acid sequence is Phe-DTrp-Lys(n)-Thr.

[0064] Another object of the present invention is to provide a somatostatin chelate, which is formed by removing the hydrogen atom from the N-terminal NH2 substituent of a somatostatin peptide analog of the invention's general formula (I) and then directly linking it to a chelating group or indirectly linking it via a spacer. The structural formula of the chelate of the present invention is: chelating group-(spacer)-dehydrogenation formula (I).

[0065] The chelating group includes any feasible chelating group mentioned in the prior art. Preferred chelating groups are DTPA, DOTA, TETA or substituted EDTA or DTPA chelating groups. More preferred chelating groups are DTPA or DOTA.

[0066] Suitable spacers include those disclosed in German Patent GB-A-2,225,579, for example a divalent residue of an aminocarboxylic acid, such as β-Ala or a divalent residue derived from 6-aminocaprioic acid.

[0067] The structural formulas of DTPA (diethylenetriaminepentaacetic acid), DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), TETA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid) and EDTA (ethylenediaminetetraacetic acid) are as follows:

[0068] The structural formulas of β-Ala and 6-aminohexanoic acid are as follows:

[0069] One structure of the somatostatin chelate of the present invention is, for example:

[0070] The chelating group can be bound to a detectable element. Suitable detectable elements include, for example, 19F, heavy metal elements or rare earth ions, such as paramagnetic ions used in CAT scans (computed axial tomography), such as Gd 3+ 、Fe 3+ 、Mn 2+ and Cr 2+ , fluorescent metal ions, such as Eu 3+ and radionuclides, such as radioactive lanthanides, in particular gamma-emitting radionuclides, beta-emitting radionuclides, alpha-emitting radionuclides, Auger-e - Emitting radionuclides, positron-emitting radionuclides e.g. 68 Ga.

[0071] Suitable gamma-emitting radionuclides include those useful in diagnostic techniques, such as those derived from gallium, indium, technetium, ytterbium, rhenium, terbium, thallium, and samarium, e.g. 67 Ga, 111 In, 99m Tc, 161 Tb, 169 Yb and 186 Re.

[0072] Suitable beta-emitting radionuclides include those useful in therapeutic applications, e.g. 90 Y. 67 Cu, 177 Lu, 186 Re、 188 Re、 169 Second, 121 Sn, 127 Te, 143 Pr, 198 Au, 109 Pd, 165 Dy, 32 P. 142 Pr and 156 Sm.

[0073] Suitable alpha-emitting radionuclides are those used in therapy, e.g. 211 At 212 Bi, 225Ac, or 201 Tl.

[0074] The compounds of the present invention may exist in free or salt form.

[0075] Another object of the present invention is to provide the use of somatostatin peptide analogs of general formula (I) of the present invention in the preparation of anti-tumor drugs, preferably for neuroendocrine tumors, more preferably acromegaly and carcinoid tumors. For example, the anti-tumor drugs are used to treat, prevent, and / or diagnose conditions, disorders, and / or pathologies that express SSTR1, SSTR2, SSTR3, SSTR4, and / or SSTR5 somatostatin receptors, the etiologies of which include or are related to excessive secretion of growth hormone, gastrointestinal diseases, malignant cell proliferative diseases, angiogenesis, or to prevent or combat transplant vascular disease, restenosis, and vascular occlusion following vascular injury. The method comprises administering a therapeutically effective amount of a somatostatin analog or a chelate thereof, or a pharmaceutically acceptable salt or complex with a detectable element, to a subject in need of such treatment.

[0076] Another object of the present invention is to provide salts, prodrugs, deuterated substances, solvates or hydrates of the somatostatin peptide analogs of general formula (I) of the present invention.

[0077] The positive progress of the present invention is that: the reference compound of the present invention is octreotide, and its molecular structure is as follows (VI):

[0078] The core amino acid sequence of octreotide is Phe-DTrp-Lys-Thr.

[0079] The present invention breaks the taboo that Lys9 residues cannot be substituted, otherwise biological activity will be greatly reduced. By altering the Lys9 residue in somatostatin peptide analogs, the present invention modifies and mutates the Lys9 residue. This modification enables the somatostatin peptide analogs of the present invention to exhibit higher (or at least similar) binding affinity to SSTR2. BRIEF DESCRIPTION OF THE DRAWINGS

[0080] FIG1 is a graph showing the effect of the concentration of a reference compound (octreotide) on the binding affinity of SSTR 2;

[0081] FIG2 is a graph showing the effect of the concentration of target compound 1 on the binding affinity of SSTR 2;

[0082] FIG3 is a graph showing the effect of the concentration of target compound 2 on the binding affinity of SSTR 2;

[0083] FIG4 is a graph showing the effect of the concentration of target compound 3 on the binding affinity of SSTR 2;

[0084] FIG5 is a graph showing the effect of the concentration of target compound 4 on the binding affinity of SSTR 2;

[0085] FIG6 is a graph showing the effect of the concentration of target compound 5 on the binding affinity of SSTR 2;

[0086] FIG7 is a graph showing the effect of the concentration of target compound 6 on the binding affinity of SSTR 2;

[0087] FIG8 is a graph showing the effect of the concentration of target compound 7p2 on the binding affinity of SSTR 2;

[0088] Figure 9 is a chromatogram showing the separation of compound 7h into 7h-p1+p2 and 7h-p3+p4;

[0089] Figure 10 is a chromatogram showing the separation of compound 7h-p1+p2 into 7h-p1 and 7h-p2. DETAILED DESCRIPTION

[0090] Some abbreviations used in the present invention are as follows:

[0091] RT is room temperature; MS is mass spectrometry; Min is minute; HPLC is high performance liquid chromatography; UPLC is ultra-performance liquid chromatography; LC-MS is liquid chromatography-mass spectrometry; MW is molecular weight; N2 is nitrogen; I2 is iodine; VC is vitamin C; EC50 is half maximal effect concentration;

[0092] DMF = dimethylformamide; TFA = trifluoroacetic acid; BOC = tert-butyloxycarbonyl; Fmoc = 9-fluorenylmethoxycarbonyl; HOBT = hydroxybenzotriazole; DODT = 3,6-dioxa-1,8-octanedithiol; AcOH = acetate acid; DCM = dichloromethane; ACN = acetonitrile; DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid; TETA = 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid acid (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid); EDTA = ethylenediaminetetraacetic acid; DTPA = diethylenetriaminepentaacetic acid; EA = ethyl acetate;

[0093] Cys is cysteine; Trp is tryptophan; Lys is lysine; Tyr is tyrosine; Ser is serine; Ala is alanine; Asp is aspartic acid; Glu is glutamic acid; Gly is glycine; His is histidine; Ile is isoleucine; Leu is leucine; Met is methionine; Asn is asparagine; Pro is phenylalanine; Pro is proline; Gln is glutamine; Arg is arginine; Thr is threonine; Val is valine; Phg is phenylglycine; Orn is ornithine; DTrp = (D-)Trp; DPhe = (D-)Phe. Unless otherwise specified, amino acid residues are generally in the L-form.

[0094] The synthesis process of the somatostatin peptide analogs of the general formula (I) of the present invention is as follows:

[0095] Step S1: Swelling of the resin

[0096] Chloro-2-chloro-trityl resin (1 g, degree of substitution: 0.7 mmol per gram) was added with DCM (dichloromethane, 20 ml) and swelled under nitrogen for half an hour. The DCM was then removed and washed three times with DMF (N,N-dimethylformamide solution).

[0097] Step S2: Removal of Fmoc protecting group

[0098] To the product of step S1, Fmoc-Thr(OtBu)-OH (2.1 mmol) and diisopropylethylamine (DIPEA, 8.4 mmol) were added. After reacting for two hours, the mixture was filtered and washed with DMF six times to obtain Fmoc-Thr(OtBu)-O-2-chloro-trityl resin.

[0099] Step S3: Condensation reaction of amino acids

[0100] To the product from step S2, add the Fmoc-protected amino acid (2.1 mmol), HOBt (2.1 mmol), HBTU (2.1 mmol), diisopropylethylamine (DIPEA, 4.2 mmol), and N,N-dimethylformamide (15 ml). After reacting for 1 hour, filter. The resulting Fmoc-protected amino acid-Thr(OtBu)-O-2-chloro-trityl resin is washed three times with N,N-dimethylformamide (two minutes each time).

[0101] Step S4

[0102] Repeat steps S2 and S3 until all designed polypeptide units are assembled.

[0103] Step S5: Resin shrinkage

[0104] In the product obtained in step S4, the Fmoc protecting group of the last amino acid was removed according to the method of step S2, and then the deprotection solution was extracted. Finally, it was washed three times with DMF, DCM, and methanol respectively, and dried to obtain a resin polypeptide.

[0105] Step S5: Formation of disulfide bonds using the iodine-methanol method

[0106] The resin polypeptide was dissolved in H2O / acetonitrile to a concentration of approximately 0.001 mol / L. AcOH (acetic acid) was added to adjust the pH to 4-5. Iodine solution (0.04 mol / L) (approximately 1 g of elemental iodine in 100 ml of methanol) was slowly added dropwise until the solution no longer turned colorless. The reaction was allowed to proceed for 30 minutes. After the reaction was complete, VC (ascorbic acid) was added to remove excess iodine and the reaction was continued for 30 seconds until the solution turned colorless. The resulting white solid was separated by high-performance liquid chromatography (prep-HPLC) and lyophilized to obtain the product.

[0107] Example 1 Synthesis of Compound 1 (QHJ01-V001)

[0108] Lys(a) is a commercially available raw material. The synthesis of compound 1 was carried out according to the general synthesis process of somatostatin peptide analogs described above, and the specific steps are:

[0109] Compound 1 is a white powder with an average molecular weight of 1033.3 g / mol. LC-MS determinations (M+H)+: 1034.4 and (M+2H)2+: 517.3.

[0110] Purity determined by UPLC (214 nm): >99%

[0111] Example 2 Compound 2 (QHJ01-V017)

[0112] The synthesis steps of the Lys(b) variant residue in compound 2 are as follows:

[0113] Synthesis of compound 2b

[0114] Isobutyronitrile 2a (2.0 g) was dissolved in THF (20 mL), the temperature was lowered to -78°C, and LDA (15.9 mL, 2 M in THF) was added dropwise. The mixture was stirred at -78°C for 45 min, followed by the dropwise addition of 3-bromopropene (7.1 g). The reaction mixture was slowly warmed to room temperature and allowed to react at room temperature for 3.5 h. The reaction mixture was quenched with water (50 mL), the aqueous phase was extracted with n-hexane (3 × 70 mL), and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to afford compound 1 (3.12 g) as a yellow oil.

[0115] 1 H NMR (400MHz, DMSO) δ5.91–5.75(m,1H),5.25–5.12(m,2H),2.30-2.26(m,2H),1.29(s,6H).

[0116] Synthesis of compound 2c

[0117] Compound 2b (2.55 g) was dissolved in MTBE (30 mL). LAH (3.56 g) was added portionwise at 0°C. The mixture was slowly warmed to room temperature and allowed to react overnight. The reaction solution was quenched with 6M NaOH (20 mL, aq.) in an ice bath. The aqueous phase was extracted with MTBE (3 × 40 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to afford compound 2c (2.64 g) as a light yellow viscous oil.

[0118] 1 H NMR (400MHz, DMSO) δ5.88–5.71(m,1H),5.07–4.95(m,2H),2.28(s,2H),1.94-1.89(m,2H),0.77(s,6H).

[0119] Synthesis of compound 2d

[0120] Compound 2c (2.64 g) was dissolved in MTBE (20 mL), and (Boc)2O (7.01 g) and TEA (6 mL) were added. The mixture was reacted at room temperature for 4 h. The reaction solution was diluted with water (20 mL), and the aqueous phase was extracted with EA (3 × 40 mL). The organic phase was washed with saturated aqueous sodium chloride (1 × 50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5 / 1) to obtain compound 2d (2.08 g).

[0121] 1 H NMR(400MHz,DMSO)δ6.79(t,J=6.1Hz,1H),5.87-5.74(m,1H),5.06–4.93(m, 2H), 2.76 (d, J = 6.4Hz, 2H), 1.88 (d, J = 7.5Hz, 2H), 1.38 (s, 9H), 0.76 (s, 6H).

[0122] Synthesis of compound 2e

[0123] Compound 2d (1.0 g) was dissolved in a mixture of MeOH (20 mL) and DCM (20 mL). After the temperature was lowered to -78°C, O3 was continuously introduced until the reaction solution turned blue. The reaction solution was concentrated and dried by rotary evaporation and used directly in the next reaction.

[0124] 1 H NMR (400MHz, DMSO) δ11.51 (s, 1H), 6.67 (t, J = 6.2Hz, 1H), 2.79-2.75 (m, 2H), 1.46-1.41 (m, 2H), 1.38 (s, 9H), 0.82 (s, 6H).

[0125] Synthesis of compound 2f

[0126] Compound 2e (921.1 mg) was dissolved in DCM (10 mL), and compound 2w (1.463 g) and DBU (758 mg) were added. The mixture was allowed to react at room temperature overnight. The reaction solution was diluted with water (20 mL), and the aqueous phase was extracted with DCM (3 × 40 mL). The organic phase was washed with water (1 × 50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography to obtain compound 2f (550 mg).

[0127] 1H NMR(400MHz,DMSO)δ8.32(br,1H),6.80(t,J=6.1Hz,1H),6.35(br,1H),3.66(s ,3H),2.77(d,J=6.4Hz,2H),2.01(d,J=7.8Hz,2H),1.38(s,18H),0.78(s,6H).

[0128] Synthesis of compound 2g

[0129] Compound 2f (550 mg) was dissolved in MeOH (20 mL), and (S,S)-Et-DuPHOS-Rh (13 mg) was added. The mixture was reacted at room temperature for 3 h under a hydrogen atmosphere (400 psi). The solid was filtered off, and the filtrate was concentrated to afford compound 2g (630 mg, crude).

[0130] 1 H NMR (400MHz, DMSO) δ7.11(d,J=7.5Hz,1H),6.78(t,J=6.2Hz,1H),3.87-3.79(m,1H),3.61(s,3H),2.77- 2.68(m,2H),1.64–1.44(m,2H),1.41–1.34(m,18H),1.24-1.19(m,1H),1.12-1.02(m,1H),0.74(s,6H).

[0131] Synthesis of compound 2h

[0132] Compound 2g (630 mg, crude) was dissolved in a mixture of THF (10 mL) and H₂O (10 mL). LiOH.H₂O (138 mg) was added, and the mixture was allowed to react overnight at room temperature. After the reaction solution was concentrated, the pH was adjusted to approximately 3. The aqueous phase was extracted with EA (3 × 30 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness to afford compound 2h (450 mg).

[0133] Synthesis of compound 2i

[0134] Compound 2h (210 mg) was dissolved in 2M NaHCO₃ (2.4 mL, 420 mg), followed by a solution of CuSO₄.5H₂O (300 mg) in H₂O (2.40 mL). NaHCO₃ (100 mg) was then added, followed by (Boc)₂O (414 mg) dissolved in 3 mL of acetone. The mixture was stirred for 24 hours. Methanol (1 mL) was added to the solution, and stirring continued for 14 hours. H₂O (2 mL) was added to the mixture, and the product was filtered, washed with H₂O (3 x 2 mL), and dried to yield a light blue solid. To remove the copper, the light blue Cu-chelate was suspended in H₂O (27 mL). 8-quinolinol (234 mg) was then added, and the mixture was stirred for 18 hours (a light salad green precipitate formed). The suspension was filtered, the precipitate was washed with H₂O (3 x 7 mL), and the filtrate was washed with ethyl acetate (3 x 15 mL). The aqueous layer was lyophilized to give 120 mg of a white solid. LC-MS: [M+H] + =275.2

[0135] Synthesis of compound 2j

[0136] Compound 2i (120 mg) was dissolved in a mixture of H2O (5 mL), and a solution of Fmoc-OSu (168 mg) in ACN (5 mL) was added. The mixture was stirred at room temperature for 3 hours. The mixture was extracted with EA (3 × 6 mL), and the organic phase was concentrated to obtain a residue which was purified by flash chromatography (C18 column) to afford compound 2j (150 mg) as a white solid.

[0137] LC-MS: [M+H] + =497.4

[0138] 1H NMR (400MHz, DMSO-d6) δ12.38(br,1H),7.82(dd,J=8.2,1.3Hz,2H),7.62(dd,J=7.5,1.4 Hz,2H),7.50(td,J=7.7,1.2Hz,2H),7.41(td,J=7.9,1.5Hz,2H),6.23(d,J=8.4Hz,1H), 5.45–5.38(m,1H),4.78(t,J=5.1Hz,1H),4.42(d,J=4.9Hz,2H),4.24(dt,J=8.6,6.2Hz, 1H),3.15–3.01(m,2H),1.94–1.74(m,2H),1.42(s,9H),1.40–1.25(m,2H),0.85(s,6H).

[0139] Synthesis of compound 2

[0140] According to the general synthesis process of somatostatin peptide analogs described above, the specific steps are:

[0141] Compound 2 is a white powder with an average molecular weight of 1047.3 g / mol and an average molecular weight of 524.3 g / mol as determined by LC-MS (M+2H). The purity of the compound (214 nm) is >99%.

[0142] Example 3 Compound 3 (QHJ01-V002)

[0143] The synthesis steps of the Lys(d) variant residue in compound 3 are as follows:

[0144] Synthesis of compound 3b

[0145] Compound 3a (7.98 g) was dissolved in THF (100 mL), and TEA (11.35 g) and (Boc)2O (24.50 g) were added. The reaction mixture was stirred at room temperature overnight, diluted with water (50 mL), and the aqueous phase was extracted with EA (3 x 50 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 9 / 1) to afford compound 3b (13.4 g).

[0146] Synthesis of compound 3d

[0147] Compound 3c (13.4 g) was dissolved in aqueous NaHCO₃ (29.36 g, 30 mL), and CbzCl (29.82 g) was added dropwise at room temperature. The mixture was allowed to react overnight. The reaction mixture was adjusted to pH 2-3 with 1 M HCl, and the aqueous phase was extracted with EA (3 × 100 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated to yield the amide intermediate (40.5 g, crude product). This amide intermediate and benzyl alcohol (12.58 g) were dissolved in DCM (100 mL). DCC (26.40 g) and DMAP (1.42 g) were added at 0°C. The reaction mixture was stirred at room temperature for 5 h. The white solid was removed by filtration, and the filtrate was concentrated. The resulting residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 7 / 3) to yield compound 3d (6.7 g).

[0148] Synthesis of compound 3e

[0149] Compound 3d (5.64 g) and compound 3 (8.53 g) were dissolved in DCM (50 mL), and 2nd Grubbs catalyst (833 mg) was added. The mixture was stirred at 60°C overnight. The reaction mixture was cooled and diluted with water (50 mL). The aqueous phase was extracted with DCM (3 x 50 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting residue was purified by silica gel column chromatography (petroleum ether / dichloromethane = 1 / 1) to afford compound 3e (4.44 g).

[0150] Synthesis of compound 3g

[0151] Compound 3e (4.0 g, 8.30 mmol) was dissolved in methanol (30 mL). After replacing the nitrogen atmosphere, 10% Pd / C (500 mg) was added to replace the hydrogen atmosphere. The mixture was reacted at room temperature overnight under a hydrogen atmosphere. The solid was filtered off, and the filtrate was dried by rotary evaporation. The resulting residue (compound 3f) was used directly in the next step.

[0152] Compound 3f (1.7 g) and NaCO₃ (1.3 g) were dissolved in dixoane (20 mL) and H₂O (6 mL). Fmoc-Cl (1.68 g) was added at 0°C. The mixture was stirred at room temperature for 30 min and quenched with water (20 mL). The aqueous phase was extracted with EA (3 × 40 mL). The combined organic phases were washed with saturated aqueous sodium chloride (1 × 60 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by Prep-HPLC (acetonitrile / water (containing 0.5% formic acid)) to afford compound 3g (300 mg).

[0153] Molecular weight (calculated): 482.6; LCMS: [MH] - =481.2, Purity:97.39%, Rt=1.814min;

[0154] 1 H NMR (400MHz, DMSO) δ7.897-7.878(m,2H),7.678-7.661(m,2H),7.429-7.307(m,4H),6.741-6.726(m,1H),6.420-6.408(m,1H),4.256- 4.212(m,3H),3.515-3.499(m,1H),2.822-2.626(m,2H),1.598-1.459(m,3H),1.409(s,9H),1.235-1.193(m,2H),0.786-0.744(m,3H).

[0155] Synthesis of compound 3

[0156] According to the general synthesis process of somatostatin peptide analogs described above, the specific steps are:

[0157] Compound 3 is a white powder with an average molecular weight of 1033.3 g / mol and an average molecular weight of 517.4 g / mol.

[0158] Purity determined by UPLC (214 nm): >99%

[0159] Example 4 Compound 4 (QHJ01-V029)

[0160] The synthesis steps of the Lys(j) variant residue in compound 4 are as follows:

[0161] Synthesis of compound 4b

[0162] Compound 4a (10 g) was dissolved in DCM (60 mL), and O-Allyl-N-[(9-anthracenyl)methyl]cinchonidium bromide (2.05 g) and CsOH.H2O (57 g) were added. After the mixture was cooled to -78°C, a solution of 2-cyclohexen-1-one (10.4 g) in DCM (20 ml) was slowly added dropwise. The reaction mixture was kept at -78°C and stirred for 6 h. The reaction mixture was poured into water (50 mL) to quench the reaction. The aqueous phase was extracted with EA (3 × 50 mL), and the organic phase was washed with saturated aqueous sodium chloride (1 × 70 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1) to obtain compound 4b (7.2 g) as a white solid.

[0163] LC-MS: [M+H] + =392.4

[0164] 1 H NMR (400MHz, CDCl3) δ7.70–7.68(m,2H),7.53–7.33(m,6H),7.19-7.14(m,2H),3.92(d,J=4.3Hz,1H),2.70( t,J=13.1Hz,1H),2.58–2.36(m,3H),2.33–2.21(m,1H),2.07–2.01(m,1H),1.75–1.55(m,3H),1.48(s,9H).

[0165] Synthesis of compound 4c

[0166] Trimethyl phosphonoacetate (6.7 g) was dissolved in DMF (300 mL). Under nitrogen, NaH (1.8 g) was added portionwise at 0°C. The mixture was stirred at 0°C for 20 min, followed by the addition of compound 4b (7.2 g). The reaction mixture was stirred at room temperature for 1.5 h. The reaction mixture was poured into water (500 mL) to quench the mixture. The aqueous phase was extracted with EA (3 × 500 mL). The organic phase was washed with saturated aqueous sodium chloride solution (1 × 700 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1) to obtain compound 4c (7.2 g).

[0167] LC-MS: [M+H] + =448.4

[0168] Synthesis of compound 4d

[0169] Compound 4c (5 g) was dissolved in MeOH (100 ml), and Pd / C was added. The mixture was stirred at room temperature under hydrogen atmosphere for 3 h. The solid was filtered off and the filtrate was dried to obtain compound 4d, which was used directly in the next reaction.

[0170] LC-MS: [M+H] + =286.3

[0171] Synthesis of compound 4e

[0172] Compound 4d (3.19 g) was dissolved in a mixture of MeOH and H₂O (100 ml), and LiOH (1.4 g) was added. The mixture was stirred at room temperature for 2 h. The reaction solution was concentrated. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 5 / 1) to afford compound 4e (3 g) as a colorless oil.

[0173] LC-MS: [M+H] + =272.2

[0174] Synthesis of compound 4f

[0175] Compound 4e (3 g) was dissolved in DMF (50 mL), and FmocOSu (11.2 g) and DIEA (4.3 g) were added. The mixture was stirred at room temperature for 5 h. The reaction solution was diluted with water (80 mL), and the aqueous phase was extracted with EA (3 × 100 mL). The organic phase was washed with saturated aqueous sodium chloride (1 × 150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 1) to obtain compound 4f (3 g).

[0176] LC-MS: [M+H] + =494.3

[0177] 1 H NMR (400MHz, CDCl3) δ7.76(d,J=7.5Hz,2H),7.61(d,J=7.4Hz,2H),7.39(t,J=7.4Hz,2H),7.33-7.28(m,2H),5.39-5.33(m,1H),4 .42-4.37(m,2H),4.25-4.21(m,2H),2.36(br,1H),2.24-2.22(m,1H),1.98–1.41(m,16H),1.38-1.25(m,1H),1.12–0.78(m,2H).

[0178] Synthesis of compound 4g

[0179] Compound 4f (2.9 g) was dissolved in DMF (20 mL), DPPA (1.94 g) and DIEA (2.3 g) were added, and the mixture was stirred at room temperature for 3 h. The reaction solution was diluted with water (50 mL), the aqueous phase was extracted with EA (3 × 70 mL), and the organic phase was washed with saturated aqueous sodium chloride (1 × 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5 / 1) to obtain compound 4g (1.13 g) as a white solid.

[0180] LC-MS: [M+H] + =519.2

[0181] 1 H NMR (400MHz, CDCl3) δ7.77(d,J=7.5Hz,2H),7.60(d,J=7.3Hz,2H),7.40(t,J=7.4Hz,2H),7.32-7.29(m,2H),5.30-5.21(m, 1H),4.43-4.40(m,2H),4.23(t,J=6.8Hz,2H),2.47–2.13(m,2H),1.95–1.63(m,4H),1.63-1.48(m,13H),1.13–0.78(m,3H).

[0182] Synthesis of compound 4h

[0183] Compound 4g (600 mg) was dissolved in t-BuOH (5 ml), and the mixture was reacted at 100° C. for 2 h under microwave. The reaction solution was concentrated, and the residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 7 / 1) to obtain compound 4h (600 mg).

[0184] LC-MS: [M+H] + =565.4

[0185] 1 H NMR (400MHz, CDCl3) δ7.77(d,J=7.6Hz,2H),7.61(d,J=7.3Hz,2H),7.40(t,J=7.4Hz,2H),7.36–7.29(m,2H),5.32-5.29(m,1H),4.40- 4.38(m,2H),4.25-4.21(m,2H),3.17-2.88(m,2H),1.84-1.70(m,3H),1.65-1.52(m,4H),1.39(s,9H),1.36(s,9H),1.13–0.67(m,3H).

[0186] Synthesis of compound 4j

[0187] Compound 4h (600 mg) was dissolved in a mixture of TFA (10 ml) and DCM (10 ml), and the mixture was reacted at room temperature for 30 min. The reaction solution was spin-dried to obtain compound 4i, which was used directly in the next reaction.

[0188] Compound 4i (600 mg) was dissolved in DCM (20 ml), and (Boc)2O (267 mg) and DIEA (569 mg) were added. The mixture was reacted at room temperature for 1 h. The reaction solution was concentrated, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 10 / 1) to obtain compound 4j (1 g).

[0189] LC-MS: [M+H] + =509.4

[0190] 1H NMR (400MHz, DMSO) δ7.90(d,J=7.5Hz,2H),7.76-7.63(m,2H),7.50-7.45(m,1H),7.42(t,J=7.4Hz,2H),7.33(t,J=7.4Hz,2H),6.81-6.7 7(m,1H),4.26–4.22(m,3H),3.96–3.80(m,1H),2.90–2.71(m,2H),1.72-1.54(m,4H),1.37(s,9H),1.36-1.30(m,2H),1.15–0.63(m,4H).

[0191] Synthesis of compound 4:

[0192] The synthesis of compound 4 was carried out according to the general synthesis process of somatostatin peptide analogs described above, and the specific steps are:

[0193] Compound 4 is a white powder with an average molecular weight of 1059.3 g / mol and an average molecular weight of 530.4 as determined by LC-MS (M+2H). The purity determined by UPLC (214 nm) is 93%.

[0194] Example 5 Compound 5 (QHJ01-V003)

[0195] Lys(n) residue was a commercially available raw material. The synthesis of compound 5 was carried out according to the general synthesis process of somatostatin peptide analogs described above. The specific steps are:

[0196] Compound 5 is a white powder with an average molecular weight of 1067.3 g / mol. LC-MS analysis revealed (M+H)+ values ​​of 1068.4 and (M+2H)2+ ​​values ​​of 534.4. UPLC analysis revealed a purity of 99% (214 nm).

[0197] Example 6 Compound 6 (QHJ01-V059)

[0198] The specific steps for the synthesis of compound 6 are:

[0199] Compound 6a was synthesized according to the general synthesis process of somatostatin peptide analogs mentioned above.

[0200] Compound 6a is a white powder with the chemical formula: C 67 H 94 N 14 O 19S2; molecular weight (average): calculated value 1463.69 g / mol; LC-MS measured value (M+H)+: 1463.6; (M+H)+: 1485.6; UPLC measured purity (214 nm): 99%.

[0201] Compound 6a (5 mg) and 0.1 M sodium acetate (1 mL, pH = 5.5) were mixed, and LuCl3·6H2O (5.6 mg) was slowly added. The mixture was stirred at room temperature for 12 hours, then separated by high performance liquid chromatography (prep-HPLC) and lyophilized to obtain compound 6. Chemical formula: C 67 H 91 LuN 14 O 19 S2; Molecular weight (average): calculated value 1635.6 g / mol; LC-MS measured value (M+H)+: 1635.7; HPLC determination purity (214 nm): >99%

[0202] Example 7 Compound 7p2 (QHJ01-V29p2)

[0203] The synthesis steps of the Lys variant residue in compound 7p2 are as follows:

[0204] Synthesis of compound 7b

[0205] Nitromethane (19.05 g) was dissolved in toluene (70.00 mL), and (1S,2S)-(+)-N-(p-toluenesulfonyl)-1,2-diphenylethylenediamine (9.53 g), AcOH (0.45 mL), and H2O (0.25 mL) were added. After stirring, a solution of 2-cyclohexen-1-one (25.00 g) in toluene (60 mL) was slowly added dropwise. The reaction mixture was incubated at room temperature for 8 h. The reaction mixture was quenched by pouring into water (100.00 mL), and the aqueous phase was extracted with EA (3 × 50 mL). The organic phase was washed with saturated sodium chloride solution (1 × 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 15 / 1) to obtain compound 7b (20.00 g).

[0206] Synthesis of compound 7c

[0207] N-CBZ-(diethoxyphosphate)amino acid methyl ester (54.87 g) was dissolved in DMF (500 mL). Under nitrogen, DBU (22.80 mL) was added dropwise at 0°C. The mixture was stirred at 0°C for 20 min, and then compound 7b (20.00 g) was slowly added. The reaction mixture was stirred at room temperature for 5 h. The reaction solution was poured into water (1000.00 mL) to quench the mixture. The aqueous phase was extracted with EA (3 × 500 mL). The organic phase was washed with saturated sodium chloride solution (1 × 700 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1) to obtain compound 7c (29.3 g).

[0208] Synthesis of compound 7d

[0209] At 0°C under nitrogen, compound 7c (29.30 g) was stirred in AcOH (400 mL). Zinc powder (37.00 g) was then added portionwise and stirred for 4 h. The reaction solution was filtered through celite and concentrated by evaporation to afford crude product 7d (17.70 g), which was used directly in the next step without purification.

[0210] Synthesis of compound 7e

[0211] The crude compound 7d (17.70 g) and (Boc)2O (14.65 mL) were dissolved in DCM (50 mL), and TEA (8.85 mL) was slowly added. The mixture was reacted at room temperature for 3 h. The reaction solution was concentrated, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 15 / 1) to obtain compound 7e (8.30 g).

[0212] Synthesis of compound 7f

[0213] Compound 7e (8.30 g) was dissolved in a mixture of MeOH and H2O (100 ml), and LiOH (1.61 g) was added. The mixture was stirred at room temperature for 4 h. The reaction solution was concentrated, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 10 / 1) to obtain compound 7f (4.70 g).

[0214] Synthesis of compound 7g

[0215] Compound 7f (4.70 g) was dissolved in MeOH (70 ml), Pd / C was added, and the mixture was stirred at room temperature under hydrogen atmosphere for 6 h. The solid was filtered off and the filtrate was dried to obtain crude compound 7g (3.00 g), which was used directly in the next reaction.

[0216] Synthesis of compound 7h

[0217] The crude product of compound 7g (3.00g) was dissolved in DMF (50mL), FmocOSu (3.89g) and DIEA (2.1mL) were added, and the mixture was stirred at room temperature for 5h. The reaction solution was diluted with water (80mL), the aqueous phase was extracted with EA (3×100mL), and the organic phase was washed with saturated aqueous sodium chloride solution (1×150mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue obtained by concentration was purified by silica gel column chromatography (dichloromethane / methanol = 10 / 1) to obtain compound 7h (0.65g).

[0218] LC-MS: [M+H] + =509.4

[0219] 1H NMR (400MHz, DMSO-d6) δ7.89(d,J=7.5Hz,2H),7.75(d,J=7.5Hz,2H),7.42(t,J=7.4Hz,2H),7.33(t,J=7.4Hz,2H),6.79(d,J=6.0Hz,1H),4. 24(dd,J=8.2,4.4Hz,3H),3.85(s,1H),2.84–2.65(m,2H),1.82–1.52(m,5H),1.37(s,9H),1.25–0.91(m,1H),0.74(dt,J=22.9,11.9Hz,2H).

[0220] Chiral resolution of compound 7h

[0221] Compound 7h was subjected to a first chiral separation using liquid chromatography to obtain 7h-p1+p2 and 7h-p3+p4. A second chiral separation was then performed on 7h-p1+p2 and 7h-p3+p4, respectively. The chiral separation method is as follows:

[0222] Table 1. Chromatographic method for the separation of 7h into 7h-p1+p2 and 7h-p3+p4

[0223] The chromatogram of 7h split into 7h-p1+p2 and 7h-p3+p4 is shown in Figure 9. The retention time of 7h-p1+p2 is 7.826 min, and the retention time of 7h-peak3+4 is 9.492 min.

[0224] Table 2. Chromatographic method for the separation of 7h-p1+p2 into 7h-p1 and 7h-p2

[0225] The chromatogram of 7h-peak 1+2, which was resolved into 7h-p1 and 7h-p2, is shown in Figure 10. The retention time of 7h-p1 was 7.666 min, and the retention time of 7h-p2 was 8.420 min. After resolution, 42 mg of 7h-p1 and 65 mg of 7h-p2 were obtained.

[0226] The specific steps for the synthesis of compound 7p2 are:

[0227] Compound 7p2 was synthesized according to the general synthesis process of somatostatin peptide analogs mentioned above.

[0228] Compound 7p2 is a white powder with an average molecular weight of 1059.3 g / mol and an average molecular weight of 530.3 g / mol as determined by LC-MS (M+2H). The purity determined by UPLC (214 nm) is 96%.

[0229] Effect Example Determination of Competitive Affinity of Radioligands

[0230] SSTR2 overexpressing CHO-K1 cells (Hunter express SSTR2 CHO-K1: Discover X) were used to 125 I-TYR 11 - Radioligand competition binding assay for Somatostatin-14 (Perkin Elmer).

[0231] Step S1: Extraction of cell membrane

[0232] SSTR2-overexpressing CHO-K1 cells (from Discover X) were cultured in complete culture medium (F12 minimal medium supplemented with 10% FBS and 1% P / S (100 U / ml penicillin and 100 μg / ml streptomycin)) in a 37°C incubator with 5% CO2. When cell confluency reached 80-90%, the cells were washed twice with pre-chilled PBS. The cells were then scraped off with a cell scraper and transferred to a 50 mL centrifuge tube. Centrifugation was performed at 500 × g and 4°C for 10 minutes. The supernatant was discarded and the cell pellet retained. Cell membrane lysis buffer (containing 25 mM HEPES, 1 mM EDTA, 2 mM MgCl2, and 1× protease inhibitor, pH = 7.4) was added at a ratio of 4E+07 cells / 10 mL and incubated on ice for 10 minutes. The cells were then disrupted using an IKA disruptor. The operation procedure is 15,000 rpm, 10 seconds, twice. The disrupted cell suspension is centrifuged at 4°C and 650×g for 5 minutes. The supernatant is transferred to a new centrifuge tube and centrifuged at 4°C and 13,000 rpm for 30 minutes. The supernatant is discarded and the cell membrane pellet is resuspended in 1.6 mL of resuspension buffer, which is a cell membrane lysis buffer with 10% sucrose added. The cell membrane lysis buffer is repeatedly aspirated 4 times with a 27-gauge needle to break the DNA. The protein concentration of the cell membrane extract is determined by the BCA method and the aliquots are frozen in a -80°C refrigerator.

[0233] Step S2: Cell membrane binding

[0234] (1) Compound Dilution: The assay buffer contained 50 mM Hepes, 5 mM MgCl2, 1 mM CaCl2, and 0.5% BSA, pH = 7.4. The buffer was filtered. The assay buffer was maintained at 4°C. Serial dilutions of the ligand solution were performed using an Agilent Bravo. Each test compound was serially diluted 3× eight times. The eight binding concentrations of a typical compound were: 50 nM, 12.5 nM, 3.13 nM, 0.78 nM, 0.20 nM, 0.05 nM, 12.2 pM, and 3.05 pM. Affinity measurements were performed in duplicate for each concentration.

[0235] Transfer 1 μL of each diluted compound to the corresponding wells of a 96-well V-bottom plate.

[0236] (2) 125 Preparation of I-SST:

[0237] The storage solution 125I-SST was diluted to 3 nM (10×) with binding buffer (containing 50 mM Hepes, 5 mM MgCl2, 1 mM CaCl2 and 0.5% BSA, pH=7.4, stored at 4°C) and 10 μL was transferred to the corresponding compound wells.

[0238] (3) Combined experiment:

[0239] The extracted cell membrane extract was diluted to 2.5 μg / 90 μL with binding buffer and added to the corresponding wells. Incubate at 25°C on a shaker at 500 rpm for 2 hours.

[0240] Membrane complexes were harvested using a Cell Harvester onto GF / B plates pre-coated with 0.5% PEI. The plates were rinsed three times with 200 μL per well of 4°C pre-chilled buffer (the same buffer used to dilute the compounds). The plates were then dried at 37°C for 2 hours. After 30 minutes, 50 μL of liquid scintillation fluid (MicroScint 20, Perkin Elmer) was added to each well and the plates were incubated for 30 minutes. The radioactivity was then read on a Topcount plate reader (30 seconds per well).

[0241] Step S3 Curve fitting

[0242] For each compound, the 8-point reading data (2× of 8 readings) were imported into X Lfit software to fit the sigmoidal curve. The EC value of the compound on SSTR2 was 50 It is calculated based on the mathematically optimal Sigmoidal dose-response model equation:

[0243] Where A is the bottom of the curve, B is the top of the curve, and C is the relative EC 50 , D is the Hill slope, y is the % effect value (%Effect) of the curve fitting, and x is the molar concentration.

[0244] Where A, B, C, and D have the same meaning, y is the % Effect reading, and x is the molar concentration calculated by curve fitting. When y is 50%, the calculated x is the absolute EC 50 The values ​​are shown in Table 4 and Figures 1 to 8.

[0245] Table 4 Binding affinity of target compounds 1, 2, 3, 4, and 5 to SSTR2

[0246] in conclusion:

[0247] By changing the Lys9 group, appropriately increasing the spatial tension and limiting the number of possible conformations of the side chain, it is possible to facilitate more effective binding between the drug molecule and SSTR2, thus breaking the "taboo" that the Lys9 residue cannot be replaced, otherwise the biological activity will be greatly reduced.

Claims

1. A somatostatin peptide analog, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable complex thereof, or a pharmaceutically acceptable chelate thereof, The somatostatin peptide analogs have a cyclic or non-cyclic hexapeptide unit, and the residue sequence at positions numbered 2 to 5 in the hexapeptide unit is: -X1-(D / L-)Trp-X2-X3-; Alternatively, the somatostatin peptide analogue has a cyclic or non-cyclic octapeptide unit, and the residue sequence at positions numbered 2 to 7 in the octapeptide unit is: -Cys-X1-(D / L-)Trp-X2-X3-Cys-; wherein X1 is an α-amino acid residue having an aromatic group on the Cα side chain, and X3 is an α-amino acid residue characterized in that X2 is a Lys-variant residue: n is 0 or 1; A1, A2, A3 are independently selected from C(R)2, O, S, NR, C=O, C=S, C=NR and C=C(R)2; R is independently selected from R1, -R'OR1, -R'SR1, -R'N(R1)2, -R'C(O)R1, -R'S(O)R1, -R'S(O)2R1, -R'C(O)N(R1)2, -R'N(R1)C(O)R1, -R'C(O)OR1 and -R'OC(O)R1; or independently, the R in C(R)2 at position A2 is connected to the R in C(R)2 at position A1 or position A3 to form a double bond or a triple bond; or independently, R in C(R)2 forms a 3-8 membered carbocyclic ring with R in its own C(R)2, or with R in an adjacent C(R)2, or with R in a C(R)2 separated by one C(R)2, or forms a 3-8 membered heterocyclic ring containing 1-3 heteroatoms selected from N, O, and S; R' is a linking bond, a C1-C6 alkylene group, a C2-C6 alkenylene group, a C2-C6 alkynylene group, a C3-C8 cycloalkylene group, a C3-C8 cycloalkenylene group, a halogenated C1-C6 alkylene group, a halogenated C2-C6 alkenylene group, a halogenated C2-C6 alkynylene group, a halogenated C3-C8 cycloalkylene group, a halogenated C3-C8 cycloalkenylene group, a (C6-C8)arylene group, a 3-8 membered heterocyclylene group having 1 to 3 heteroatoms selected from N, O, and S, and a 3-8 membered heteroarylene group having 1 to 3 heteroatoms selected from N, O, and S; R1 is independently selected from H, halogen, CN, NO2, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, halogenated C1-C6 alkyl, halogenated C2-C6 alkenyl, halogenated C2-C6 alkynyl, halogenated C3-C8 cycloalkyl, halogenated C3-C8 cycloalkenyl, (C6-C8)aryl, 3-8 membered heterocyclic group having 1-3 heteroatoms selected from N, O, S, 1-3 heteroatoms selected from N, O, S A 3- to 8-membered heteroaryl group; When n is 0, R in C(R)2 at the A1 position, A2 position and A3 position are not H at the same time.

2. The somatostatin peptide analogue according to claim 1, characterized in that The somatostatin peptide analogs have a cyclic or non-cyclic hexapeptide unit, and the residue sequence at positions numbered 1 to 6 in the hexapeptide unit is: Xa-X1-(D / L-)Trp-X2-X3-Xb; The somatostatin peptide analogs have a cyclic or non-cyclic octapeptide unit, and the residue sequence at positions numbered 1 to 8 in the octapeptide unit is: X0-Cys-X1-(D / L-)Trp-X2-X3-Cys-Xb; Wherein Xa and Xb are amino acid residues; X0 is an α-amino acid residue having an aromatic group on the Cα side chain.

3. The somatostatin peptide analogue according to claim 1 or 2, characterized in that X1 is a Phe, Tyr, Trp or Phg residue; X3 is a Thr, Ser, Cys or tyrosylbenzyl residue.

4. The somatostatin peptide analogue according to claim 1, characterized in that The somatostatin peptide analogs have a cyclic octapeptide unit structure as shown in the following general formula (I): n is 0 or 1; A1, A2, A3 are independently selected from C(R)2, O, S, NR, C=O, C=S, C=NR and C=C(R)2; R is independently selected from R1, -R'OR1, -R'SR1, -R'N(R1)2, -R'C(O)R1, -R'S(O)R1, -R'S(O)2R1, -R'C(O)N(R1)2, -R'N(R1)C(O)R1, -R'C(O)OR1 and -R'OC(O)R1; or independently, the R in C(R)2 at position A2 is connected to the R in C(R)2 at position A1 or position A3 to form a double bond or a triple bond; or independently, R in C(R)2 forms a 3-8 membered carbocyclic ring with R in its own C(R)2, or with R in an adjacent C(R)2, or with R in a C(R)2 separated by one C(R)2, or forms a 3-8 membered heterocyclic ring containing 1-3 heteroatoms selected from N, O, and S; R' is a linking bond, a C1-C6 alkylene group, a C2-C6 alkenylene group, a C2-C6 alkynylene group, a C3-C8 cycloalkylene group, a C3-C8 cycloalkenylene group, a halogenated C1-C6 alkylene group, a halogenated C2-C6 alkenylene group, a halogenated C2-C6 alkynylene group, a halogenated C3-C8 cycloalkylene group, a halogenated C3-C8 cycloalkenylene group, a (C6-C8)arylene group, a 3-8 membered heterocyclylene group having 1 to 3 heteroatoms selected from N, O, and S, and a 3-8 membered heteroarylene group having 1 to 3 heteroatoms selected from N, O, and S; R1 is independently selected from H, halogen, CN, NO2, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, halogenated C1-C6 alkyl, halogenated C2-C6 alkenyl, halogenated C2-C6 alkynyl, halogenated C3-C8 cycloalkyl, halogenated C3-C8 cycloalkenyl, (C6-C8)aryl, 3-8 membered heterocyclyl having 1 to 3 heteroatoms selected from N, O, and S, and 3-8 membered heteroaryl having 1 to 3 heteroatoms selected from N, O, and S; When n is 0, R in C(R)2 at the A1 position, A2 position and A3 position are not H at the same time.

5. The somatostatin peptide analogue according to claim 1 or 4, characterized in that The R in C(R)2 at position A1 and the R in C(R)2 at position A3 are connected together to form a ring so that A1, A2, and A3 form a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring having 1 to 3 heteroatoms selected from N, O, and S; Preferably, The R in C(R)2 at position A1 and the R in C(R)2 at position A3 are connected together to form a ring so that A1, A2, and A3 form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring having 1-2 heteroatoms selected from N, O, and S.

6. The somatostatin peptide analogue according to claim 1 or 4, characterized in that The R in C(R)2 at the A2 position is connected to the R in C(R)2 at the A1 position or the A3 position to form a double bond.

7. The somatostatin peptide analogue according to claim 1 or 4, characterized in that R' is a connecting bond, C1-C4 alkylene, C2-C4 alkenylene, C2-C4 alkynylene, C3-C7 cycloalkylene, C3-C7 cycloalkenylene, halogenated C1-C4 alkylene, halogenated C2-C4 alkenylene, halogenated C2-C4 alkynylene, halogenated C3-C7 cycloalkylene, halogenated C3-C7 cycloalkenylene, (C6-C8)arylene, 3-7 membered heterocyclylene having 1 to 3 heteroatoms selected from N, O, and S, and 3-7 membered heteroarylene having 1 to 3 heteroatoms selected from N, O, and S; R1 is independently selected from H, halogen, CN, NO2, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C7 cycloalkyl, C3-C7 cycloalkenyl, halogenated C1-C4 alkyl, halogenated C2-C4 alkenyl, halogenated C2-C4 alkynyl, halogenated C3-C7 cycloalkyl, halogenated C3-C7 cycloalkenyl, (C6-C8)aryl, 3-7 membered heterocyclyl having 1 to 3 heteroatoms selected from N, O, and S, and 3-7 membered heteroaryl having 1 to 3 heteroatoms selected from N, O, and S; Preferably, R' is a linking bond, C1-C3 alkylene, C2-C3 alkenylene, C2-C3 alkynylene, C5-C6 cycloalkylene, C5-C6 cycloalkenylene, halogenated C1-C3 alkylene, halogenated C2-C3 alkenylene, halogenated C2-C3 alkynylene, halogenated C5-C6 cycloalkylene, halogenated C5-C6 cycloalkenylene, (C6-C8)arylene, 5-6 membered heterocyclylene having 1-2 heteroatoms selected from N, O, and S, and 5-6 membered heteroarylene having 1-2 heteroatoms selected from N, O, and S; R1 is independently selected from H, halogen, CN, NO2, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl, C5-C6 cycloalkenyl, halogenated C1-C3 alkyl, halogenated C2-C3 alkenyl, halogenated C2-C3 alkynyl, halogenated C5-C6 cycloalkyl, halogenated C5-C6 cycloalkenyl, (C6-C8)aryl, 5-6 membered heterocyclic group having 1 to 2 heteroatoms selected from N, O, and S, and 5-6 membered heteroaryl having 1 to 2 heteroatoms selected from N, O, and S.

8. The somatostatin peptide analogue according to claim 1 or 4, characterized in that X2 is a Lys-variant residue: Selected from:

9. The somatostatin peptide analogue according to claim 1 or 4, characterized in that Somatostatin peptide analogs are specifically:

10. A somatostatin chelate, characterized in that The somatostatin chelate is a chelate formed by removing the H atom from the N-terminal NH2 substituent of the somatostatin peptide analogue described in claim 1 or 4 and then directly connecting it to the chelating group or indirectly connecting it through a spacer.

11. The somatostatin chelate according to claim 10, characterized in that The chelating group is a chelating group of DTPA, DOTA, TETA or substituted EDTA or DTPA, and the spacer is a divalent residue of an aminocarboxylic acid, such as a β-Ala residue or a divalent residue derived from 6-aminocaprioic acid.

12. Use of the somatostatin peptide analogs according to claim 1 or 4 in the preparation of anti-tumor drugs, preferably for neuroendocrine tumors, acromegaly and carcinoid tumors.

13. The salt, prodrug, deuterated substance, solvate or hydrate of the somatostatin peptide analogue according to claim 1 or 4.

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