Radioelement-labeled products and their use

Abstract

The present application relates to radiohalogen labeled linkers and uses thereof, and specifically provides a linker radiohalogen conjugate comprising a linker and a radiohalogen covalently linked, the linker comprising a structure represented by Formula I or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

Description

[0001] This application claims priority to the following prior patent applications:

[0002] The applicant filed an earlier application with the China National Intellectual Property Administration on April 30, 2025, with patent application number 202510563090.3 and title "Products with Radioactive Element Labelling and Their Applications".

[0003] The full text of the aforementioned prior patent applications is incorporated herein by reference. Technical Field

[0004] This invention belongs to the field of radiopharmaceuticals and relates to a product that can rapidly label halogen radioactive elements and its pharmaceutical uses. Background Technology

[0005] Radiopharmaceuticals utilize radionuclides or their labeled compounds for disease diagnosis and treatment, representing a new approach to precision oncology and holding promise as a major direction in future cancer treatment. The emergence of this new technology has spurred innovative research into radiopharmaceuticals, leading to their increasing adoption in frontline diagnosis and treatment for patients with complex and rare diseases. Over the past few decades, new diagnostic / prognostic tools, especially imaging examinations, have been introduced into clinical practice to better support cancer patient care. Furthermore, Lu... 177 Oxyoctreotide, Lu 177 The approval of drugs such as terciviride has spurred a surge in radiopharmaceutical research and development. Both of these marketed radioconjugates use beta-isotopes, while alpha-isotopes theoretically possess greater killing power and safety in tumor treatment compared to beta-isotopes. This is because alpha-isotopes kill tumors with over 100 times the energy of beta-isotopes, and their shorter range results in less impact on adjacent normal tissues, making them safer, especially for small tumors and micrometastases.

[0006] Currently, alpha nuclides that have entered the clinical trial stage include metal nuclides. 212 Pb, 225 Ac; Halogens 211 At. Both Pb and Ac are metallic nuclides that are chelated with their respective ligands through non-covalent bonds, and their decay pathways are relatively complex. During the decay process, radioactive elements may escape due to the recoil of α nuclides. 211 At, as a halogen, is covalently bonded in radiocoupled drugs. In terms of the production pathway, an alpha beam cyclotron is used... 209 Bi(α,2n) 211 At nuclear reactions produce astatine. 211 At), the target material is an inexpensive, natural metal. 209Bi can be produced in batches with a reasonably high yield. In terms of separation and purification, it can be separated from irradiated targets using methods such as dry distillation or wet chemistry. 211 At the same time, this method is sustainable for commercial mass production. 211 The decay process of At is very simple; each decay produces only one α ion. 211 At has excellent decay characteristics. 211 At, as an alpha-ray emitting nuclide, has a half-life of only 7.2 hours, therefore... 211 At-labeled nuclides have short durations of action, allowing for outpatient treatment. Furthermore, as short-lived nuclides, they offer the advantage of a lower risk of prolonged side effects, making their efficacy as novel anticancer drugs highly anticipated. However, the short half-life is a double-edged sword; behind these numerous advantages lie stringent requirements for the labeling efficiency, labeling time, and stability of the linker.

[0007] Therefore, linkers that can rapidly label halogen radioactive elements require further research. Summary of the Invention

[0008] Radioactive halogens often require the protection of sensitive groups during labeling. This invention discloses a series of bifunctional linkers for radioactive halogen labeling. These linkers eliminate the need for protection and deprotection reactions during halogen labeling, significantly shortening the labeling reaction time and improving labeling efficiency. This is particularly beneficial for radioactive halogens with short half-lives, especially... 211 At has significant advantages, and the introduction of electron-donating groups or PEG chains onto the aromatic groups of the linker increases... 211 At labeling improves stability and reduces the dehalogenation effect of radioactive halogens in vivo.

[0009] Therefore, in one aspect of the invention, a linker-radiohalogen conjugate is provided, comprising a linker and a radioactive halogen, said linker and said radioactive halogen being covalently linked, said linker comprising the structure of Formula I or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0010] Formula I

[0011] in: R A Selected from hydrogen, cyano, nitro, halogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, C2-C6 alkenyl, C2-C6 alkynyl, R a -L a -; L a Selected from -(CH2)m1 -、-O-、-S-、-NH-; R a Selected from guanidinyl, protonated guanidinyl, ; m1 is an integer selected from 1 to 5; m2 is an integer selected from 1 to 20; L 1 Selected from carbonyl, -(CH2) n1 -C(=O)-、-L 11 -L 12 -; L 11 Selected from carbonyl, -(CH2) n1 -C(=O)-; L 12 The amino acid residues are short peptides formed from 2 to 10 amino acid residues or isopeptides formed from 2 to 10 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid. n1 is an integer selected from 1 to 5; L 2 Selected from , , , ; L 21 Selected from direct bonds, -(CH2) n21 -、-O-、-S-; L 22 Selected from polyethylene glycol chains and polysarcosine chains; L 23 Selected from direct bonds, -(CH2) n23 -、-O-、-S-; R 1 Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, ; R 2 Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, ; n21 is an integer selected from 1 to 5; n23 is an integer selected from 1 to 5; n25 is an integer selected from 1 to 20; L 3 Selected from direct bond, carbonyl group, , , , , , ; R3 Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, C2-C6 alkenyl, and C2-C6 alkynyl; n31, n32, n33, n34, n35, and n36 are each independently selected from integers from 1 to 5; R B Selected from , , , , ; R b1 Selected from C1-C6 alkyl groups; R b2 R b3 Each is independently selected from hydrogen, cyano, nitro, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 alkylthio; R b4 R b5 R b6 Each is independently selected from hydrogen, cyano, nitro, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 alkylthio; Y is selected from halogens; q is an integer selected from 1 to 5; And, when R B Selected from R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0012] In some implementation schemes, R A Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, R a -L a -

[0013] In some implementation schemes, R A Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, R a -L a -

[0014] In some implementation schemes, R A Selected from hydrogen, methyl, methoxy, , , .

[0015] In some implementations, L aSelected from -(CH2) m1 -、-O-、-S-.

[0016] In some implementations, L a Selected from -(CH2) m1 -、-O-.

[0017] In some implementations, L a Selected from -CH2- and -O-.

[0018] In some implementation schemes, R a Selected from , , .

[0019] In some implementation schemes, R a Selected from , , .

[0020] In some implementations, m1 is an integer selected from 1 to 3, preferably 1.

[0021] In some implementations, m2 is an integer selected from 1 to 10, preferably an integer selected from 1 to 5, and more preferably 3.

[0022] In some implementations, L 1 Selected from carbonyl, -CH2-C(=O)-, -L 11 -L 12 -

[0023] In some implementations, L 1 Selected from carbonyl, -CH2-C(=O)-, , , , , .

[0024] In some implementations, L 11 Selected from carbonyl, -CH2-C(=O)-.

[0025] In some implementations, L 12 The amino acid residues are short peptides formed from 2-3 amino acid residues or isopeptides formed from 2-3 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid.

[0026] In some implementations, L 12 It is a short peptide formed from 2-3 amino acid residues, wherein the amino acids are natural or non-natural amino acids.

[0027] In some embodiments, the amino acid is selected from glutamic acid, valine, methionine, lysine, and... .

[0028] In some embodiments, the amino acid is selected from L-glutamic acid, L-valine, L-methionine, L-lysine, .

[0029] In some implementations, L 12 Selected from , , , , .

[0030] In some implementations, n1 is an integer selected from 1 to 3, preferably 1.

[0031] In some implementations, L 2 Selected from , , , , n22 is an integer selected from 1 to 20. n24 is an integer selected from 1 to 20.

[0032] In some implementations, L 2 Selected from , , , , , , , , , , , .

[0033] In some implementations, L 21 Selected from direct bonds, -(CH2) n21 -

[0034] In some implementations, L 23 Selected from direct bonds, -(CH2) n23 -

[0035] In some implementation schemes, R 1 Selected from hydrogen, C1-C6 alkyl, .

[0036] In some implementation schemes, R 1 Selected from hydrogen, .

[0037] In some implementation schemes, R 2 Selected from hydrogen, C1-C6 alkyl, .

[0038] In some implementation schemes, R 2 Selected from hydrogen, .

[0039] In some implementations, n21 is an integer selected from 1 to 3, preferably 1 or 2.

[0040] In some implementations, n22 is an integer selected from 1 to 10, preferably an integer selected from 2 to 8, and more preferably 2, 3, 4 or 8.

[0041] In some implementations, n23 is an integer selected from 1 to 3, preferably 1 or 2.

[0042] In some implementations, n24 is an integer selected from 1 to 10, preferably an integer selected from 5 to 10, and more preferably 6 or 7.

[0043] In some implementations, n25 is an integer selected from 1 to 10, preferably an integer selected from 1 to 5, and more preferably 4.

[0044] In some implementations, L 3 Selected from direct bond, carbonyl group, , , , , , , .

[0045] In some implementation schemes, R 3 Selected from hydrogen and C1-C6 alkyl groups.

[0046] In some implementation schemes, R 3 Selected from hydrogen and methyl.

[0047] In some implementations, n31 is an integer selected from 1 to 3, preferably 3.

[0048] In some implementations, n32 is an integer selected from 1 to 3, preferably 1.

[0049] In some implementations, n33 is an integer selected from 1 to 3, preferably 2.

[0050] In some implementations, n34 is an integer selected from 1 to 3, preferably 2.

[0051] In some implementations, n35 is an integer selected from 1 to 3, preferably 2.

[0052] In some implementations, n36 is an integer selected from 1 to 3, preferably 2.

[0053] In some implementation schemes, R b1 It is a methyl group.

[0054] In some implementation schemes, R b2 R b3 Each is independently selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 alkylthio.

[0055] In some implementation schemes, R b2 R b3 Each is independently selected from hydrogen and C1-C6 alkoxy groups.

[0056] In some implementation schemes, R b2 R b3 In the given information, one of the components is hydrogen, and the other is selected from hydrogen or C1-C6 alkoxy groups.

[0057] In some implementation schemes, R b2 R b3 Each is independently selected from hydrogen and methoxy groups.

[0058] In some implementation schemes, R b2 R b3 In this formula, one of the components is hydrogen, and the other is selected from hydrogen or methoxy.

[0059] In some implementation schemes, R b4 R b5 R b6 Each is independently selected from hydrogen, cyano, nitro, and C1-C6 alkyl groups.

[0060] In some implementation schemes, R b4 R b5 R b6 Each is independently selected from hydrogen and cyano groups.

[0061] In some implementation schemes, R b4 R b5 R b6 In this group, any one of them is a cyano group, and the rest are hydrogen.

[0062] In some implementations, Y is -Br.

[0063] In some implementations, q is an integer selected from 1 to 3, preferably 1.

[0064] In some implementation schemes, R B Selected from , , , , , .

[0065] In some embodiments, the linker comprises the structure shown in Formula Ia or Formula Ib, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0066] Formula Ia Formula Ib

[0067] in: L 1 L 2 L 3 R A R B Each is independent as described above.

[0068] In some embodiments, the linker comprises the structure shown in Formula I-1a or Formula I-1b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0069] Formula I-1a Formula I-1b

[0070] in: R B1 Selected from , , , ; L 1 L 2 L 3 R A R b1 R b2 R b3 R b4 R b5 R b6 Y and q are as described above, each independently.

[0071] In some embodiments, the linker comprises the structure shown in Formula I-2a or Formula I-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0072] Equation I-2a Equation I-2b

[0073] in: L 1 L 2 L3 R A R b1 Each is independent as described above; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0074] In some embodiments, the linker comprises a structure selected from the following or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof. .

[0075] In some embodiments, the radioactive halogen conjugate is a compound having the structure shown in Formula II or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0076] Formula II

[0077] in: X is a radioactive halogen; Preferably, X is selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At; More preferably, X is selected from125 I, 211 At; L 1 L 2 L 3 R A R B Each is independent as described above; And, when R B Selected from R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0078] In some embodiments, the radioactive halogen conjugate is a compound having the structure shown in Formula II-a or Formula II-b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0079] Formula II-a Formula II-b

[0080] in: L 1 L 2 L 3 R A R B X and X are as described above, each independently.

[0081] In some embodiments, the radioactive halogen conjugate is a compound having the structure shown in Formula II-1a or Formula II-1b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0082] Formula II-1a Formula II-1b

[0083] in: L 1 L 2 L 3 R A X and X are as described above, each independently; R B1 As mentioned above.

[0084] In some embodiments, the radioactive halogen conjugate is a compound having the structure shown in Formula II-2a or Formula II-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0085] Formula II-2a Formula II-2b

[0086] in: L 1 L 2 L 3 R A X and X are as described above, each independently; R b1 As mentioned above; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0087] In some embodiments, the radioactive halogen conjugate is selected from compounds with the following structures or their pharmaceutically acceptable salts, tautomers, enantiomers, or stereoisomers. .

[0088] In another aspect, the present invention provides compounds of Formula III or pharmaceutically acceptable salts, tautomers, enantiomers, or stereoisomers thereof.

[0089] Formula III

[0090] in: X 1 The solvent is F, Cl, Br or I, preferably I; L 1 L 2 L 3 R A R B Each is independent as described above; And, when R B Selected from R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0091] In some embodiments, the compound has the structure shown in Formula III-a or Formula III-b.

[0092] Formula III-a Formula III-b

[0093] in: L 1 L 2 L 3 R A R B X 1 Each is independent as described above.

[0094] In some embodiments, the compound has the structure shown in Formula III-1a or Formula III-1b.

[0095] Formula III-1a Formula III-1b

[0096] in: L 1 L 2 L 3 R A X 1 Each is independent as described above; R B1 As mentioned above.

[0097] In some embodiments, the compound has the structure shown in formula III-2a or III-2b.

[0098] Formula III-2a Formula III-2b

[0099] in: L 1 L 2 L 3 R A X 1 Each is independent as described above; R b1 As mentioned above; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0100] In some embodiments, the compound is selected from the following: .

[0101] In another aspect, the present invention provides a linker radioactive halogen conjugate precursor compound or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof, wherein the precursor compound has the structure shown in Formula IV.

[0102] Formula IV

[0103] in: X 2 It is a leaving group; Preferably, X 2 Selected from -SnR c 3, -SiR c 3. -B(OH)2; R c Selected from C1-C6 alkyl groups; Preferably, R c It is methyl; R A’ Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, C2-C6 alkenyl, C2-C6 alkynyl, R a’ -L a -; Preferably, R A’ Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, R a’ -L a -; More preferably, R A’ Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, R a’ -L a -; Most preferably, R A’ Selected from hydrogen, methyl, methoxy, , , , ; R a’ Selected from guanidinyl, protonated guanidinyl, guanidinyl with one or more amino protecting groups, and protonated guanidinyl with one or more amino protecting groups. ; Preferably, R a’ Selected from guanidinyl, protonated guanidinyl, guanidinyl with one or more amino protecting groups, ; More preferably, R a’ Selected from , , , ; Most preferably, R a’ Selected from , , , ; L 1’ Selected from carbonyl, -(CH2) n1 -C(=O)-、-L 11 -L 12’ -; Preferably, L 1’ Selected from carbonyl, -CH2-C(=O)-, -L 11 -L 12’ -; More preferably, L 1’ Selected from carbonyl, -CH2-C(=O)-, , , , , , , , , ; L 12’ The amino acid residue is a short peptide formed from 2 to 10 amino acid residues or an isopeptide formed from 2 to 10 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group. Preferably, L 12’ The amino acid residue is a short peptide formed by 2-3 amino acid residues or an isopeptide formed by 2-3 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group. More preferably, L 12’ It is a short peptide formed from 2-3 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group; Preferably, the amino acid is selected from glutamic acid, valine, methionine, lysine, and so on. And the lysine, Each of them may independently and optionally have a carboxyl protecting group; Preferably, the amino acid is selected from L-glutamic acid, L-valine, L-methionine, and L-lysine. , , ; Most preferably, L 12’ Selected from , , , , , , , , ; L 2 L 3 R B L a m2, L 11 n1 and n2 are as described above, each independently; And, when R B Selected from R a’ Selected from guanidino or guanidino groups having one or more amino protecting groups, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1’ It is not a carbonyl group.

[0104] In some embodiments, the precursor compound has the structure shown in formula IV-a or IV-b.

[0105] Formula IV-a Formula IV-b

[0106] in: L 1’ L 2 L 3 R A’ R B X 2 Each is independent as described above.

[0107] In some embodiments, the precursor compound has the structure shown in formula IV-1a or IV-1b.

[0108] Formula IV-1a Formula IV-1b

[0109] in: L 1’ L 2 L 3 R A’ X 2 Each is independent as described above; R B1 As mentioned above.

[0110] In some embodiments, the precursor compound has the structure shown in formula IV-2a or IV-2b.

[0111] Formula IV-2a Formula IV-2b

[0112] in: L 1’ L 2 L 3 R A’ X 2 Each is independent as described above; R b1 As mentioned above; And, when R a’ Selected from guanidino or guanidino groups having one or more amino protecting groups, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1’ It is not a carbonyl group.

[0113] In some embodiments, the precursor compound is selected from the following: .

[0114] In another aspect of the invention, the invention provides a single-domain antibody or antigen-binding fragment thereof capable of specifically binding to FRα, comprising: CDR1 or a variant thereof contained in the heavy chain variable region (VHH) shown in SEQ ID NO: 4, CDR2 or a variant thereof contained in the heavy chain variable region (VHH) shown in SEQ ID NO: 4, and CDR3 or a variant thereof contained in the heavy chain variable region (VHH) shown in SEQ ID NO: 4; The variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, or 3 amino acids) compared to the sequence from which it originates; preferably, the substitutions are conservative substitutions.

[0115] In some embodiments, the single-domain antibody or its antigen-binding fragment comprises CDR1, CDR2, and CDR3 contained in the heavy chain variable region (VHH) as shown in SEQ ID NO: 4.

[0116] In some implementations, the three CDRs contained in the VHH are defined by the Kabat, IMGT, or Chothia numbering system.

[0117] In some embodiments, the single-domain antibody or its antigen-binding fragment comprises: CDR1 as shown in SEQ ID NO: 1, CDR2 as shown in SEQ ID NO: 2, and CDR3 as shown in SEQ ID NO: 3.

[0118] In some implementations, the three CDRs contained in the VHH are defined by the Kabat numbering system.

[0119] In some embodiments, the single-domain antibody or its antigen-binding fragment comprises a heavy chain framework region sequence of an immunoglobulin (e.g., a heavy chain framework region sequence of a human or camel-derived immunoglobulin).

[0120] In some embodiments, the single-domain antibody or its antigen-binding fragment comprises the heavy chain framework region of a human immunoglobulin.

[0121] In some embodiments, the single-domain antibody or its antigen-binding fragment comprises a heavy chain framework region contained in the amino acid sequence encoded by a human heavy chain germline gene; preferably, the heavy chain framework region optionally comprises one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) reversion mutations from human residues to camel residues.

[0122] In some embodiments, the single-domain antibody or its antigen-binding fragment comprises an amino acid sequence selected from the following: (i) A sequence as shown in SEQ ID NO: 4; (ii) A sequence having one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to the sequence shown in SEQ ID NO: 4; or (iii) A sequence having at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence shown in SEQ ID NO: 4; Preferably, the substitution is a conservative substitution.

[0123] In another aspect of the invention, the invention provides the use of the aforementioned linker radiohalogen conjugate in the preparation of radiolabeled proteins / peptides.

[0124] In another aspect, the present invention provides a radionuclide-coupled drug comprising a linker radiohalogen coupling unit and a targeting ligand, wherein the linker radiohalogen coupling unit is covalently linked to the targeting ligand, and the linker radiohalogen coupling unit comprises a structure of Formula V or a pharmaceutically acceptable salt thereof, a tautomer, an enantiomer, or a stereoisomer.

[0125] Formula V

[0126] in: X is a radioactive halogen; Preferably, X is selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At; More preferably, X is selected from 125 I, 211 At; R B’ Selected from , , , , Among them, site 1 and L 3 Connected; L 1 L 2 L 3 R A R b2 R b3 R b4 R b5 R b6 q and q are as described above, each independently; And, when R B’ for R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0127] In some embodiments, the linker radiohalogen coupling unit comprises a structure represented by formula Va or Vb or a pharmaceutically acceptable salt thereof, a tautomer, an enantiomer, or a stereoisomer.

[0128] Formula Va Formula Vb

[0129] in: L 1 L 2 L 3 R A R B’ X and X are as described above, each independently.

[0130] In some embodiments, the linker radiohalogen coupling unit comprises the structure shown in formula V-1a or V-1b or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0131] Formula V-1a Formula V-1b

[0132] in: R B1’ Selected from , , , Among them, site 1 and L 3 Connected; L 1 L 2 L 3 R A X, R b2 R b3 R b4 R b5 R b6 q and q are as described above, each independently.

[0133] In some embodiments, the linker radiohalogen coupling unit comprises the structure shown in formula V-2a or V-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0134] Formula V-2a Formula V-2b

[0135] in: L 1 L 2 L 3 RA X and X are as described above, each independently; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0136] In some embodiments, the linker radiohalogen coupling unit comprises a structure selected from the following or a pharmaceutically acceptable salt thereof, a tautomer, an enantiomer, or a stereoisomer. ; X is a radioactive halogen; Preferably, each X is independently selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At; More preferably, each X is independently selected 125 I, 211 At.

[0137] In some embodiments, the radionuclide-coupled drug is a compound having the structure shown in Formula VI or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0138] Style VI

[0139] in: Ab is the target ligand, preferably a protein / peptide; Preferably, Ab is an antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment that can specifically bind to FRα; Preferably, the single-domain antibody or its antigen-binding fragment capable of specifically binding to FRα is as described above; R B’ Selected from , , , , Among them, site 1 and L 3 The two sites are connected; site 2 is connected to Ab. L 1 L 2 L 3 R A X, R b2 R b3 R b4 R b5 R b6 q and q are as described above, each independently; And, when R B’ for R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0140] In some embodiments, the radionuclide-coupled drug is a compound having the structure shown in Formula VI-a or Formula VI-b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0141] Formula VI-a Formula VI-b

[0142] in: L 1 L 2 L 3 R A X, R B’ Ab are as described above, each independently.

[0143] In some embodiments, the radionuclide conjugate is a compound having the structure shown in Formula VI-1a or Formula VI-1b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0144] Formula VI-1a Formula VI-1b

[0145] in: R B1’ Selected from , , , Among them, site 1 and L 3 The two sites are connected; site 2 is connected to Ab. L 1 L 2 L 3 R A X, R b2 R b3 R b4 R b5 R b6 , q, and Ab are as described above independently.

[0146] In some embodiments, the radionuclide conjugate is a compound having the structure shown in Formula VI-2a or Formula VI-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof.

[0147] Formula VI-2a Formula VI-2b

[0148] in: L 1 L 2 L 3 R A X and Ab are as described above independently; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

[0149] In some embodiments, the radionuclide-coupled drug is selected from substances with the following structures or pharmaceutically acceptable salts, tautomers, enantiomers, or stereoisomers thereof. , in: X is a radioactive halogen; Preferably, each X is independently selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At; More preferably, each X is independently selected 125 I, 211 At; Ab is the target ligand, preferably a protein / peptide; Preferably, Ab is an antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment that can specifically bind to FRα; Preferably, the single-domain antibody or its antigen-binding fragment capable of specifically binding to FRα is as described above.

[0150] In another aspect of the invention, the invention provides a pharmaceutical composition comprising the aforementioned linker radiohalogen conjugate or the aforementioned radionuclide conjugate.

[0151] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier and / or excipient.

[0152] In another aspect of the invention, a kit is provided that comprises the aforementioned linker radiohalogen conjugate or the aforementioned radionuclide conjugate drug.

[0153] In another aspect of the invention, the invention provides the use of the aforementioned linker radiohalogen conjugate, or the aforementioned radionuclide conjugate drug, or the aforementioned pharmaceutical composition in the preparation of a medicament for treating and / or preventing diseases.

[0154] In another aspect of the invention, the invention provides the aforementioned linker radiohalogen conjugate, the aforementioned radionuclide conjugate drug, or the aforementioned drug composition for the treatment and / or prevention of disease.

[0155] In another aspect of the invention, the invention provides a method for treating and / or preventing disease, comprising administering to a subject in need an effective amount of the aforementioned linker radiohalogen conjugate or the aforementioned radionuclide conjugate drug or the aforementioned pharmaceutical composition.

[0156] In some implementations, the disease is a disease related to FRα.

[0157] In some implementations, the disease is selected from ovarian cancer, breast cancer (such as triple-negative breast cancer), endometrial cancer, mesothelioma, lung cancer, cervical cancer, colorectal cancer, head and neck squamous cell carcinoma, and prostate cancer.

[0158] In another aspect of the invention, the invention provides the use of the aforementioned linker radiohalogen conjugate or the aforementioned radionuclide conjugate in the preparation of imaging diagnostic agents.

[0159] In some implementations, the imaging is SPECT imaging, PET imaging, or CT imaging.

[0160] In some implementations, the imaging diagnostic agent is used to diagnose diseases.

[0161] In another aspect of the invention, the invention provides the aforementioned linker radiohalogen conjugate or the aforementioned radionuclide conjugate drug for the diagnosis of diseases.

[0162] In another aspect of the invention, the invention provides a method for diagnosing a disease, comprising administering an effective amount of the aforementioned linker radiohalogen conjugate or the aforementioned radionuclide conjugate drug to a subject in need.

[0163] In some implementations, the diagnosis is an imaging diagnosis.

[0164] In some implementations, the imaging is SPECT imaging, PET imaging, or CT imaging.

[0165] In some implementations, the disease is a disease related to FRα.

[0166] In some implementations, the disease is selected from ovarian cancer, breast cancer (such as triple-negative breast cancer), endometrial cancer, mesothelioma, lung cancer, cervical cancer, colorectal cancer, head and neck squamous cell carcinoma, and prostate cancer.

[0167] Terminology Definition

[0168] Unless specifically defined, the nomenclature, methods, and techniques used in combination with those of molecular biology, analytical chemistry, synthetic organic chemistry, and medical and medicinal chemistry described herein are those well-known and commonly used in the art. Standard techniques are applicable to chemical synthesis and chemical analysis. Some of these techniques and methods can be found, for example, in “Carbohydrate Modifications in Antisense Research” edited by Sangvi and Cook, American Chemical Society, Washington, 1994; “Remington’s Pharmaceutical Sciences,” Mack Publishing Co., Easton, Pa., 18th ed., 1990; and “Antisense Drug Technology, Principles, Strategies, and Applications” edited by Stanley T. Crooke, CRC Press, Boca Raton, Fla.; and Sambrook et al., “Molecular Cloning, A Laboratory Manual,” 2nd ed., Cold Spring Harbor Laboratory Press, 1989, which are incorporated herein by reference for any purpose. Where permitted, all patents, applications, publications and other publications and data cited throughout this disclosure are incorporated herein by reference in their entirety.

[0169] As used herein, the term "antibody" is used in the broadest sense to encompass a wide variety of antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, provided they exhibit the desired antigen-binding activity. For example, an immunoglobulin molecule can consist of two pairs of polypeptide chains, each pair having one light chain (LC) and one heavy chain (HC). Antibody light chains can be classified as κ (kappa) and λ (lambda) light chains. Heavy chains can be classified as μ, δ, γ, α, or ε, and antibody isotypes are defined as IgM, IgD, IgG, IgA, and IgE, respectively. Within both the light and heavy chains, variable and constant regions are linked by a "J" region of approximately 12 or more amino acids, and the heavy chain also contains a "D" region of approximately 3 or more amino acids. Each heavy chain consists of a heavy chain variable region (VH) and a heavy chain constant region (CH). The heavy chain constant region consists of three domains (CH1, CH2, and CH3). Each light chain consists of a variable region (VL) and a constant region (CL). The constant region consists of a single CL domain. While not directly involved in antibody-antigen binding, the constant domain exhibits various effector functions, such as mediating the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system. The VH and VL regions can be further subdivided into highly degenerated regions (called complementarity-determining regions (CDRs)) interspersed with more conserved regions called framework regions (FRs). Each VH and VL consists of three CDRs and four FRs arranged in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4, from the amino terminus to the carboxyl terminus. The variable regions (VH and VL) of each heavy / light chain pair form the antigen-binding sites. The distribution of amino acids in different regions or domains can follow the definitions in Kabat, Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), or Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883.

[0170] In this article, unless the context clearly indicates otherwise, when referring to the term "antibody," it includes not only the complete antibody but also the antigen-binding fragment of the antibody.

[0171] As used herein, the term "single-domain antibody" (sdAb) has the meaning commonly understood by those skilled in the art, referring to an antibody fragment consisting of a single monomeric variable antibody domain (e.g., a single heavy chain variable region, VHH), typically derived from the variable region of a heavy chain antibody (e.g., a camel antibody or a shark antibody). Single-domain antibodies are also called nanobodies, and the terms are used interchangeably. Typically, a nanobody consists of four framework regions and three complementarity-determining regions, having a structure of FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. Nanobodies can be truncated at the N-terminus or C-terminus to contain only a portion of FR1 and / or FR4, or to lack one or both of those framework regions, as long as they substantially maintain antigen binding and specificity.

[0172] As used herein, the term “complementarity-determining region” or “CDR” refers to the amino acid residues in the variable region of an antibody responsible for antigen binding. The precise boundaries of these amino acid residues can be defined according to various numbering systems known in the art, such as the Kabat numbering system (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991), the Chothia numbering system (Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883), or the IMGT numbering system (Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003). For a given antibody, those skilled in the art will readily identify the CDR as defined by each numbering system. Furthermore, the correspondence between different numbering systems is well known to those skilled in the art (see, for example, Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003).

[0173] In this invention, the CDR contained in the antibody or antigen-binding fragment thereof can be determined according to various numbering systems known in the art. In some embodiments, the CDR contained in the antibody or antigen-binding fragment thereof is preferably determined by the IMGT, Kabat, or Chothia numbering system.

[0174] As used herein, the term “framework region” or “FR” residues refer to those amino acid residues in the antibody variable region other than the CDR residues as defined above.

[0175] As used herein, the term “antigen-binding fragment” of an antibody refers to a molecule other than the intact antibody that includes a portion of the intact antibody that binds to the antigen to which the intact antibody binds. For example, an antigen-binding fragment can be a polypeptide fragment of a full-length antibody that retains the ability to specifically bind to the same antigen bound by the full-length antibody, and / or competes with the full-length antibody for specific binding to the antigen; this is also referred to as an “antigen-binding moiety.” See also Fundamental Immunology, Ch. 7 (Paul, W., ed., 2nd ed., Raven Press, NY (1989), which is incorporated herein by reference in its entirety for all purposes. Antigen-binding fragments of antibodies can be generated by recombinant DNA technology or by enzymatic or chemical cleavage of the intact antibody. An antigen-binding fragment contains at least a portion of an antibody sufficient to confer specific antigen-binding ability to the polypeptide. A review of engineered antibody variants is found in Holliger et al., 2005; Nat Biotechnol, 23:1126-1136.

[0176] As used herein, the term "specific binding" refers to a non-random binding reaction between two molecules, such as the reaction between an antibody and its target antigen. The strength or affinity of a specific binding interaction can be measured by the equilibrium dissociation constant (KD) or half-maximal effective concentration (EC50) of the interaction. 50 )express.

[0177] The specific binding properties between two molecules can be determined using methods known in the art. One method involves measuring the rate of formation and dissociation of the antigen binding site / antigen complex. Both the “binding rate constant” (ka or kon) and the “dissociation rate constant” (kdis or koff) can be calculated from the concentration and the actual rates of association and dissociation (see Malmqvist M, Nature, 1993, 361:186-187). The ratio of kdis / kon is equal to the dissociation constant KD (see Davies et al., Annual Rev Biochem, 1990; 59:439-473). The values ​​of KD, kon, and kdis can be measured using any effective method. In some embodiments, the dissociation constant can be measured using bioluminescent interferometry (e.g., the ForteBio Octet method). Alternatively, surface plasmon resonance techniques (e.g., Biacore) or Kinexa can be used to measure the dissociation constant.

[0178] As used herein, the term "identity" refers to the sequence matching between two polypeptides or two nucleic acids. Two compared sequences are identical at a position when the same base or amino acid monomeric subunit occupies the same location (e.g., a position in each of two DNA molecules is occupied by adenine, or a position in each of two polypeptides is occupied by lysine). The "percentage identity" between two sequences is a function of the number of matching positions shared by the two sequences divided by the number of positions compared × 100. For example, if six out of ten positions in two sequences match, then the two sequences have 60% identity. For example, the DNA sequences CTGACT and CAGGTT have 50% identity (three out of six positions match). Typically, two sequences are compared to produce the maximum identity. Such comparisons can be made using methods readily available, for example, computer programs such as the Align program (DNAstar, Inc.) Needleman et al. (1970) J. Mol. Biol. 48:443-453. The percentage identity between two amino acid sequences can also be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl Biosci., 4:11-17 (1988)) integrated into the ALIGN program (version 2.0), which uses a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. Alternatively, the percentage identity between two amino acid sequences can be determined using the Needleman and Wunsch algorithm (J MoI Biol. 48:444-453 (1970)) in the GAP program integrated into the GCG software package (available at www.gcg.com), which uses a Blossum 62 matrix or a PAM250 matrix, along with gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6.

[0179] As used herein, the term "conservative substitution" means an amino acid substitution that does not adversely affect or alter the intended properties of a protein / peptide containing an amino acid sequence. For example, conservative substitutions can be introduced using standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include substitutions of amino acid residues with amino acid residues having similar side chains, such as substitutions with residues that are physically or functionally similar to the corresponding amino acid residues (e.g., having similar size, shape, charge, chemical properties, including the ability to form covalent or hydrogen bonds). Families of amino acid residues with similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid and glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, and tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, and methionine), β-branched side chains (e.g., threonine, valine, and isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, and histidine). Therefore, it is preferable to replace the corresponding amino acid residue with another amino acid residue from the same side chain family. Methods for identifying conserved amino acid substitutions are well known in the art (see, for example, Brummell et al., Biochem. 32:1180-1187 (1993); Kobayashi et al., Protein Eng. 12(10):879-884 (1999); and Burks et al., Proc. Natl Acad. Set USA 94:412-417 (1997), which are incorporated herein by reference).

[0180] The twenty common amino acids referred to herein are written in accordance with conventional usage. See, for example, Immunology-ASynthesis (2nd Edition, ES Golub and DR Gren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. In this invention, the terms “polypeptide” and “protein” have the same meaning and are used interchangeably. Furthermore, in this invention, amino acids are generally represented by single-letter and three-letter abbreviations known in the art. For example, alanine can be represented by A or Ala.

[0181] In this invention, unless otherwise explicitly stated, the descriptive phrase “...each independently selected” used throughout this document can mean either that the specific options expressed by the same or different symbols in different groups do not affect each other, or that the specific options expressed by the same or different symbols in the same group do not affect each other.

[0182] The substituents in the compounds of this invention are disclosed according to the type or range of groups. In particular, this invention includes every independent sub-combination of the members of these types and ranges. For example, the term "C1-C6 alkyl" specifically refers to the independently disclosed methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl groups.

[0183] The term "C1-C6 alkyl" refers to straight-chain and branched alkyl groups having 1 to 6 carbon atoms, preferably "C1-C4 alkyl", more preferably "C1-C3 alkyl", and most preferably "C1-C2 alkyl". Examples of "C1-C6 alkyl" include, but are not limited to, methyl, ethyl, propyl (e.g., n-propyl, isopropyl), butyl (e.g., n-butyl, isobutyl, tert-butyl), and pentyl (e.g., n-pentyl, isopentyl, neopentyl). Examples of "C1-C4 alkyl" include, but are not limited to, methyl, ethyl, propyl (e.g., n-propyl, isopropyl), and butyl (e.g., n-butyl, isobutyl, tert-butyl). Examples of "C1-C3 alkyl" include methyl, ethyl, and propyl (e.g., n-propyl, isopropyl). Examples of "C1-C2 alkyl" include methyl and ethyl.

[0184] The term “C1-C6 alkoxy” refers to any “C1-C6 alkyl” as defined above, which is attached to the rest of the molecule by an oxygen atom (-O-), preferably C1-C4 alkoxy, C1-C3 alkoxy or C1-C2 alkoxy, examples of which include methoxy, ethoxy, isopropoxy and the like.

[0185] The term “C1-C6 alkylthio” refers to any “C1-C6 alkyl” as defined above, which is attached to the rest of the molecule by a sulfur atom (-S-), preferably C1-C4 alkylthio, C1-C3 alkylthio or C1-C2 alkylthio, such as methylthio (i.e. CH3-S-).

[0186] The term "C2-C6 alkenyl" refers to a straight-chain or branched unsaturated hydrocarbon group consisting of 2 to 6 carbon atoms, which contains one or more carbon-carbon double bonds (C=C) and the double bond positions can form different isomers (such as 1-propenyl, 2-butenyl, etc.).

[0187] The term "C2-C6 ynyl" refers to a straight-chain or branched unsaturated hydrocarbon group consisting of 2 to 6 carbon atoms, which contains one or more carbon-carbon triple bonds (C≡C) and the triple bond positions can form isomers (such as 1-propynyl, 2-butynyl, etc.).

[0188] In this invention, unless otherwise specified, "halogen" refers to conventional non-radioactive halogens, such as F, Cl, Br, and I. "Radioactive halogen" can be... 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At, etc.

[0189] The term "direct bond" refers to a bond in which the groups at both ends are directly connected. For example, if L... 2 for And L 21 and L 23 If it is a direct key, then L 2 for For other similar descriptions, please refer to the definitions above for understanding.

[0190] In this invention, "the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group" means that the carboxyl group in the amino acid is protected by a carboxyl protecting group. For example, if the amino acid is When the carboxyl protecting group is tert-butyl, the structural formula of the amino acid with one carboxyl protecting group is: Please refer to this definition for understanding other similar descriptions.

[0191] Carboxyl protecting groups include, but are not limited to, methyl, ethyl, tert-butyl, trimethylsilylethyl (TMSE), trimethylsilyl (TMS), triethylsilyl (TES), tert-butyldimethylsilyl (TBDMS), tert-butyldiphenylsilyl (TBDPS), triisopropylsilyl (TIPS), etc.

[0192] The term "amino acid residue" refers to a structural unit that integrates into a protein, polypeptide chain, or other structure after a single amino acid molecule forms a peptide bond (amide bond) through condensation. It can be a unit where the α-amino and α-carboxyl groups are covalently linked, losing their complete independence while retaining the chemical characteristics of the side chain (R group). The amino acid can be either natural or non-natural. For example, the amino acid residue can be... , , , , As mentioned above, if the amino acid has a carboxyl protecting group, the amino acid residue may be, for example, a carboxyl protecting group. , wait.

[0193] The term "short peptide composed of 2-10 amino acid residues" refers to an oligopeptide formed by the linear tandem condensation of 2-10 (preferably 2-5, 2-4, or 2-3) of the aforementioned amino acid residues through a typical peptide bond (an amide bond formed by the condensation of α-amino and α-carboxyl groups). For example, it could be... , or wait.

[0194] The term "isopeptide formed of 2-10 amino acid residues" refers to an oligopeptide formed by the linear tandem condensation of 2-10 (preferably 2-5, 2-4 or 2-3) of the aforementioned amino acid residues via peptide bonds, and the peptide bonds are not all typical peptide bonds (α-amino and α-carboxyl groups condensed together to form amide bonds), but contain at least one (e.g., 1, 2 or more) isopeptide bonds.

[0195] The term "isopeptide bond" refers to an amide bond formed by the condensation of an α-carboxyl group of one amino acid with a side-chain amino group of another amino acid, or by the condensation of a side-chain carboxyl group of one amino acid with an α-amino group of another amino acid. Unlike typical peptide bonds (where α-amino and α-carboxyl groups condense to form an amide bond), isopeptide bonds involve the participation of side-chain functional groups.

[0196] In this invention, "multiple" refers to two, three, four, five or more, preferably two to three, and more preferably two.

[0197] In this invention, "guanidinium" refers to .

[0198] In this invention, "protonated guanidine group" refers to .

[0199] In this invention, "guanidinium group having one or more amino protecting groups" means that the amino group in the aforementioned guanidinium group is protected by one or more (preferably two) amino protecting groups, for example, it can be... Please refer to this definition for understanding other similar descriptions.

[0200] Amino protecting groups include, but are not limited to, Boc (tert-butyloxycarbonyl), Fmoc (9-fluorenyloxycarbonyl), Cbz (benzyloxycarbonyl), Alloc (allyloxycarbonyl), etc.

[0201] In this invention, a variable is an integer selected from 1 to n (n>1), meaning that the variable can be any integer point value or any integer range value from 1 to n, and the endpoints 1 and n are also included. For example, m1 is an integer selected from 1 to 5, meaning that m1 can be 1, 2, 3, 4, or 5, or any integer selected from 1-2, 1-3, 1-4, 1-5, 2-3, 2-4, 2-5, 3-4, 3-5, or 4-5. Other similar descriptions are understood with reference to this definition.

[0202] The term "polyethylene glycol chain (PEG)" refers to a polymer chain consisting of repeating ethylene glycol units (-O-CH2-CH2- or -CH2-O-CH2-).

[0203] The term "polysarcosine chain" refers to a polymer chain formed by the repeated polymerization of N-methylglycine (sarcosine) through amide bonds.

[0204] The term "pharmaceutically acceptable" means that a carrier, delivery substance, diluent, excipient, and / or the salt formed therefrom is generally chemically or physically compatible with other components constituting a drug dosage form and physiologically compatible with receptors.

[0205] Tautomerism refers to the phenomenon where a functional group in a compound changes its structure to become an isomer of another functional group, and these two isomers can rapidly interconvert. This rapid and reversible conversion process allows the two isomers to coexist in a certain proportion under given conditions, forming a dynamic equilibrium state, such as enol-ketone tautomerism. Therefore, those skilled in the art will understand that tautomers that can rapidly interconvert to reach a certain equilibrium essentially represent the same compound. In other words, for a compound existing in different tautomeric forms, the compound is not limited to any specific tautomer, but rather aims to encompass all tautomeric forms.

[0206] The term "stereoisomer" refers to a compound with the same molecular structure as the compound of the present invention but a different spatial configuration. The compounds of the present invention may include one or more asymmetric centers and therefore may exist in various stereoisomeric forms, such as enantiomers and / or diastereomers. For example, the compounds of the present invention may be individual enantiomers, diastereomers, or geometric isomers (e.g., cis and trans isomers), or may be in the form of mixtures of stereoisomers, including racemic mixtures and mixtures rich in one or more stereoisomers. The isolation of individual isomers or the selective synthesis of individual isomers is achieved by applying various methods well known to those skilled in the art. For example, isomers may be isolated from mixtures by methods known to those skilled in the art, including but not limited to: chiral high-performance liquid chromatography (HPLC) and the formation and crystallization of chiral salts; or preferred isomers may be prepared by asymmetric synthesis. Unless otherwise stated, all such isomers and mixtures thereof are included within the scope of the compounds disclosed herein. Unless otherwise stated, the structures described herein also imply all stereochemical forms encompassing that structure; i.e., the (R) and (S) configurations of each asymmetric center. Therefore, those skilled in the art will generally consider single stereochemical isomers of the compounds of the present invention, as well as mixtures of enantiomers and diastereomers, to be stable within the scope of this disclosure.

[0207] The term "salt" refers to (i) the salt formed by an acidic functional group (e.g., -COOH) present in the compounds provided by this invention and a suitable inorganic or organic cation (base), including but not limited to, alkali metal salts such as sodium salts, potassium salts, lithium salts, etc.; alkaline earth metal salts such as calcium salts, magnesium salts, etc.; other metal salts such as aluminum salts, iron salts, zinc salts, copper salts, nickel salts, cobalt salts, etc.; inorganic base salts such as ammonium salts; organic base salts such as tert-octylamine salts, dibenzylamine salts, morpholine salts, glucosamine salts, phenylglycine alkyl ester salts, ethylenediamine salts, N-methylglucosamine salts, guanidine salts, diethylamine salts, triethylamine salts, dicyclohexylamine salts, N,N'-dibenzylethylenediamine salts, chloroprocaine salts, procaine salts, diethanolamine salts, N-benzyl-phenylethylamine salts, piperazine salts, tetramethylamine salts, and tris(hydroxymethyl)aminomethane salts. And (ii) the salts formed by the basic functional group (e.g., -NH2) present in the compounds provided by the present invention and suitable inorganic or organic anions (acids), including but not limited to, hydrohalides, such as hydrofluoric acid salts, hydrochlorides, hydrobromic acid salts, hydroiodides, etc.; inorganic acid salts, such as nitrates, perchlorates, sulfates, phosphates, etc.; lower alkyl sulfonates, such as methanesulfonates, trifluoromethanesulfonates, ethanesulfonates, etc.; aryl sulfonates, such as benzenesulfonates, p-benzenesulfonates, etc.; organic acid salts, such as acetates, malates, fumarates, succinates, citrates, tartrates, oxalates, maleates, trifluoroacetates, etc.; amino acid salts, such as glycine salts, trimethylglycine salts, arginine salts, ornithine salts, glutamate salts, aspartate salts, etc.

[0208] The term "pharmaceutically acceptable carrier and / or excipient" refers to a carrier, delivery carrier, diluent, excipient, and / or the salts / esters / hydrates formed therefrom that are generally chemically or physically compatible with other components constituting a pharmaceutical dosage form (such as the siRNA of this invention) and physiologically compatible with the subject. "Pharmaceutically acceptable carriers and / or excipients" do not exert or are not intended to exert a therapeutic effect at the intended dosage. Such components may serve to: a) assist in the processing of the drug delivery system during manufacturing; b) protect, support, or enhance the stability, bioavailability, or patient acceptability of the active ingredient; c) assist in product identification; and / or d) enhance the overall safety, efficacy, delivery, or any other property of the active ingredient during storage and use. "Pharmaceutically acceptable carriers and / or excipients" include, but are not limited to: viruses, liposomes, nanoparticles, bacteria, lipid nanoparticles (LNP), neutral liposomes (NL), polymer nanoparticles, double-stranded RNA binding motifs (dsRBMs), pH adjusters, surfactants, adjuvants, ionic strength enhancers, diluents, agents for maintaining osmotic pressure, agents for delaying absorption, and preservatives. For example, viruses include, but are not limited to, retroviruses, adenoviruses, lentiviruses, baculoviruses, and AAV. Liposomes include, but are not limited to, Lipofectamine, cationic DOTAP, and neutral DOPC. Nanoparticles include, but are not limited to, cationic polymers and PEI. Bacteria include, but are not limited to, tkRNAi. Polymer nanoparticles include, but are not limited to, low-molecular-weight polymers or high-molecular-weight polymers. For example, pH adjusters include, but are not limited to, phosphate buffers. Surfactants include, but are not limited to, cationic, anionic, or nonionic surfactants, such as Tween-80. Ionic strength enhancers include, but are not limited to, sodium chloride. Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, etc. Reagents that maintain osmotic pressure include, but are not limited to, sugars, NaCl, and their analogues. Reagents that delay absorption include, but are not limited to, monostearate and gelatin. Diluents include, but are not limited to, water, aqueous buffers (such as buffered saline), alcohols, and polyols (such as glycerol).

[0209] The term "prevention" refers to a method implemented to prevent or delay the occurrence of a disease, condition, or symptom in a subject; the term "treatment" refers to a method implemented to obtain a beneficial or desired clinical outcome. For the purposes of this invention, a beneficial or desired clinical outcome includes, but is not limited to, alleviating symptoms, reducing the extent of the disease, stabilizing (i.e., no longer worsening) the state of the disease, delaying or slowing the progression of the disease, improving or alleviating the state of the disease, and relieving symptoms (whether partial or complete), whether detectable or undetectable. Furthermore, "treatment" can also refer to prolonged survival compared to the expected survival (if no treatment was received).

[0210] "Subject" refers to a vertebrate. In some embodiments, vertebrate refers to a mammal. Mammals include, but are not limited to, livestock (such as cattle), pets (such as cats, dogs, and horses), primates, mice, and rats. In some embodiments, mammal refers to a human.

[0211] "Effective amount" refers to the amount that effectively achieves the desired therapeutic, preventive, or diagnostic effect at the necessary dose and time. The "therapeutic effective amount" of the substance / molecule of the present invention may vary depending on factors such as an individual's disease state, age, sex, weight, and the ability of the substance / molecule to elicit the desired response in the individual. Therapeutic effective amount also encompasses the amount in which the beneficial therapeutic effect of the substance / molecule outweighs any toxic or harmful consequences. "Prophylactic effective amount" refers to the amount that effectively achieves the desired preventive effect at the necessary dose and time. Typically, but not necessarily, the preventive effective amount will be lower than the therapeutic effective amount because the preventive dose is administered to the subject before the onset of the disease or in the early stages of the disease. In the case of cancer, the therapeutically effective amount of the drug may reduce the number of cancer cells; shrink the tumor volume; inhibit (i.e., slow down, preferably stop) the infiltration of cancer cells into surrounding organs; inhibit (i.e., slow down, preferably stop) tumor metastasis; inhibit tumor growth to some extent; and / or alleviate one or more symptoms associated with cancer to some extent.

[0212] Beneficial effects

[0213] This invention provides a linker for rapidly labeling radioactive halogen elements. This type of linker eliminates the need for protection and deprotection reactions during halogen labeling, significantly shortening the labeling reaction time and improving labeling efficiency. It enables rapid radioactive labeling of the linker, reducing the amount of radioactive halogens, especially... 211 At, the decay of short-half-life halogen nuclides during the labeling process, and the relatively charge-rich electronic properties of the linker at the halogen labeling site or the introduction of the PEG chain can also play a certain stabilizing role for radioactive halogen elements, reducing the occurrence of dehalogenation side reactions. Attached Figure Description

[0214] Figure 1 Showing 125 I. Experimental results of tissue distribution of radiolabeled conjugate RDC-11A in a mouse model of human oral epithelioid carcinoma bearing KB cells. Detailed Implementation

[0215] The technical solution of the present invention will be further described in detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanatory of the present invention, and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are covered within the scope of protection intended by the present invention.

[0216] Example 1

[0217] Synthesis of N-(14-[(3-(aminomethamidomethyl)-5-iodophenyl)formamido]-3,6,9,12-tetraoxatetradecane-1-yl)-6-(2-methanesulfonylpyrimidin-5-yl)hex-5-acetylamine (compound 01)

[0218] Step 1: At room temperature, TCFH (124.58 mg, 0.44 mmol) was added to a solution of 6-(2-methanesulfonylpyrimidin-5-yl)hex-5-ethynic acid (100 mg, 0.37 mmol), N-(14-amino-3,6,9,12-tetraoxane-1-yl)carbamate tert-butyl ester (136.92 mg, 0.41 mmol), and 1-methyl-1H-imidazolium (63.79 mg, 0.78 mmol) in acetonitrile (2 mL). The reaction mixture was reacted at 30 °C for 1 h under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated, diluted with water (10 mL), extracted with ethyl acetate (3 mL x 3), the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by Prep-TLC (ethyl acetate) to give a yellow solid N-(14-(6-(2-methanesulfonylpyrimidin-5-yl)hex-5-ynylamino)-3,6,9,12-tetraoxatetradecane-1-yl)carbamate tert-butyl ester 01-c 200 mg, yield 91.46%. LCMS (ESI) m / z = 587.4 [M+H] + .

[0219] Step 2: At room temperature, 1 mL of TFA was added to a 1 mL solution of compound 01-c (200 mg, 0.34 mmol) in DCM. The reaction mixture was allowed to react at room temperature for 30 minutes. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated to give 160 mg of a yellow oily substance, N-(14-amino-3,6,9,12-tetraoxane-1-yl)-6-(2-methanesulfonylpyrimidin-5-yl)hex-5-acetylenamine 01-d, in 96.46% yield. LCMS (ESI) m / z = 487.3 [M+H] + .

[0220] Step 3: At room temperature, TCFH (53.87 mg, 0.19 mmol) was added to an acetonitrile solution of compound 01-d (80 mg, 0.16 mmol), 3-({[(1Z)-{[(tert-butoxy)carbonyl]amino}({[[(tert-butoxy)carbonyl]imino})methyl]amino}methyl)-5-iodobenzoic acid 01-e (91.40 mg, 0.18 mmol), and 1-methyl-1H-imidazole (27.59 mg, 0.34 mmol). The reaction mixture was reacted at 30 °C for 1 h under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated, diluted with water (10 mL), extracted with ethyl acetate (3 mL x 3), and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC to give 110 mg of white solid 01-f, in 61.85% yield. LCMS(ESI)m / z=988.3 [M+H] + .

[0221] Step 4: At room temperature, 1 mL of TFA was added to a 1 mL solution of compound 01-f (110 mg, 0.11 mmol) in DCM. The reaction mixture was allowed to react at room temperature for 30 minutes. The reaction was monitored by LCMS until completion, and the reaction mixture was concentrated. The residue was purified by high performance liquid chromatography (0.1% TFA) to give 30.73 mg of compound 01, with a yield of 35.04%.

[0222] LCMS(ESI)m / z=787.9 [M+H] + .

[0223] 1 H NMR (400 MHz, DMSO- d 6): δ 9.11(s, 2H), 8.65(t, J = 5.2 Hz, 1H), 8.14(s, 1H), 8.07(t, J = 6.0 Hz, 1H), 7.93(t, J = 6.4 Hz, 1H), 7.80 (d, J = 5.2 Hz, 2H), 7.31(brs, 3H), 4.39(d, J = 6.0 Hz, 2H), 3.53-3.49(m, 14H), 3.41-3.38(m, 7H), 3.22-3.18(m, 2H), 2.55(t, J = 6.8 Hz, 2H), 2.26(t, J= 7.6 Hz, 2H), 1.84-1.77(m, 2H).

[0224] Example 2 Synthesis of Compound 02

[0225] Step 1: In a 250 mL single-necked flask, 5-bromo-2-chloronicotinonitrile (2 g, 9.20 mmol) and DME (50 mL) were added separately. NaSMe (0.64 g, 9.2 mmol) was added with stirring at 0 °C. The reaction system was reacted at 0 °C for 2 h. Then, the reaction was continued at room temperature for another 2 h. The reaction was monitored by LCMS to indicate completion. The reaction was quenched with saturated ammonium chloride solution, extracted three times with ethyl acetate (100 mL), and the organic phases were combined. The mixture was washed twice with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated to dryness to obtain a yellow solid crude product. The crude product was purified by silica gel column chromatography (EA / PE: 10 / 1) to obtain a white solid 5-bromo-2-(methylthioalkyl)pyridine-3-onitrile O2-b 1.9 g, with a yield of 90.1%. LCMS calculation for C7H6BrN2S (M+H) + m / z = 228.9, found: 228.9; 1 H NMR (400 MHz, CDCl3) δ 8.90-8.60 (m, 1H), 7.93-7.85 (m, 1H), 2.62 (s, 3H).

[0226] Step 2: At room temperature, 5-bromo-2-(methylthioalkyl)pyridine-3-onitrile (1.8 g, 7.86 mmol), methyl 5-hexynate (1.29 g, 10.22 mmol), palladium dichloride (275.85 mg, 0.39 mmol), and CuI (149.69 mg, 0.79 mmol) were added sequentially to a dry 100 mL single-necked flask. Then, N,N-dimethylformamide (30 mL) and TEA (7.95 g, 78.60 mmol) were added. The reaction mixture was incubated at 80 °C for 5 hours under argon protection. The reaction was monitored by LCMS until completion. Dilute with water (60 mL), extract three times with ethyl acetate (80 mL), combine the organic phases, wash with saturated brine, dry with anhydrous sodium sulfate, and concentrate the filtrate under reduced pressure to obtain the crude product. Then, purify by reversed-phase chromatography (ACN / H2O: 5.0%~95%) to obtain 1.3 g of methyl 6-(5-cyano-6-(methylthioalkyl)pyridin-3-yl)hexyl-5-acetylacetic acid 02-d, a yellow solid, with a yield of 60.3%. 14 H 15 N₂O₂S (M+H) + m / z = 275.1, found: 275.2.

[0227] Step 3: At room temperature, methyl 6-(5-cyano-6-(methylthioalkyl)pyridin-3-yl)hexyl-5-acetylic acid (1.2 g, 4.37 mmol) and tetrahydrofuran (12 mL) were added to a 100 mL single-necked flask, followed by a 3 mL solution of LiOH (0.21 g, 8.74 mmol). The system was reacted at room temperature for 2 h. The reaction was monitored by LCMS. The system was adjusted to pH 6 with 1 M HCl, and extracted three times with ethyl acetate (80 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and the filtrate was filtered and concentrated under reduced pressure to obtain the crude product. The crude product was then purified by reverse-phase chromatography (ACN / H2O: 5.0%~95%) to obtain 1 g of compound 6-(5-cyano-6-(methylthioalkyl)pyridin-3-yl)hexyl-5-acetylic acid O2-e, as a yellow solid, with a yield of 87.8%. LCMS calcld for C13H13N2O2S (M+H)+ m / z =261.0, found:261.1.

[0228] Step 4: At room temperature, potassium peroxide (2.27 g, 3.69 mmol) was added to a solution of 6-(5-cyano-6-(methylthio)pyridin-3-yl)hexyl-5-ethynic acid (320 mg, 1.23 mmol) in tetrahydrofuran (10 mL) and water (5 mL). The reaction mixture was reacted at 80 °C for 6 hours under argon protection. LCMS monitoring showed that the reaction was complete. The mixture was diluted with water (30 mL) and extracted three times with ethyl acetate (50 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was then purified by reverse-phase chromatography (ACN / H2O: 5.0%–95%) to obtain 6-(5-cyano-6-methanesulfonylpyridin-3-yl)hexyl-5-ethynic acid 02-f 260 mg, with a yield of 72.3%. 13 H 13 N₂O₄S (M+H) + m / z = 293.1, found: 293.2.

[0229] Step 5: At room temperature, NMI (292.2 mg, 3.56 mmol) was added to a solution of 6-(5-cyano-6-methanesulfonylpyridin-3-yl)hexyl-5-alkynic acid (260 mg, 0.89 mmol), N-(14-amino-3,6,9,12-tetraoxotetradecane-1-yl)carbamate (359.30 mg, 1.07 mmol) and N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (374.5 mg, 1.33 mmol) in ACN (6 mL). The reaction mixture was reacted at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The mixture was diluted with water (20 mL) and extracted three times with ethyl acetate (30 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure after filtration to obtain the crude product. The compound N-(14-(6-(5-cyano-6-methanesulfonylpyridin-3-yl)hexyl-5-amido)-3,6,9,12-tetraoxotetradecane-1-yl)carbamate tert-butyl ester was then purified by reverse-phase chromatography (ACN / H2O: 5.0%~95%) to obtain 180 mg of N-(14-(6-(5-cyano-6-methanesulfonylpyridin-3-yl)hexyl-5-amido)-3,6,9,12-tetraoxotetradecane-1-yl)carbamate as a yellow solid, with a yield of 33.1%. LCMS calculation for C 28 H 43 N4O9S (M+H) + m / z = 611.3, found: 611.3.

[0230] Step 6: At room temperature, compound O2-h (180 mg, 0.29 mmol) was dissolved in MeOH (1 mL), and 4 M HCl / EtOAc (5 mL) was added. The mixture was stirred at room temperature for 30 minutes. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated under reduced pressure to give compound N-(14-amino-3,6,9,12-tetraoxotetradecane-1-yl)-6-(5-cyano-6-methanesulfonylpyridin-3-yl)hex-5-amide O2-i 150 mg, as a yellow oil, with a yield of 99.6%. LCMS calculation for C 23 H 35 N4O7S (M+H) + m / z = 511.3, found: 511.3.

[0231] Step 7: At room temperature, DIEA (0.19 g, 1.45 mmol) was added to a solution of compound 02-i (150 mg, 0.29 mmol), compound 01-e (180 mg, 0.35 mmol), and HATU (0.17 g, 0.43 mmol) in N,N-dimethylformamide (5 mL). The reaction mixture was reacted at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The mixture was diluted with water (20 mL) and extracted three times with ethyl acetate (20 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to obtain the crude product. The compound tert-butylN-[(1Z)-{[(tert-butoxy)carbonyl]imino}({[(3-[(14-(6-(5-cyano-6-methanesulfonylpyridin-3-yl)hexyl-5-amido)-3,6,9,12-tetraoxotetradecano-1-yl)carbamoyl]-5-iodophenyl)methyl]amino})methyl]carbamic acid 02-k 180 mg, was obtained as a yellow solid with a yield of 60.5%. LCMS calcld for C 42 H 59 IN7O 12 S (M+H) + m / z = 1012.3, found: 1012.3.

[0232] Step 8: At room temperature, compound O2-k (180 mg, 0.18 mmol) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (3.06 g, 26.8 mmol) was added. The mixture was stirred at room temperature for 30 minutes. The reaction was monitored by LCMS to indicate completion. The reaction mixture was concentrated, and the crude product was purified by high-performance liquid chromatography to give compound O2 30.8 mg, with a yield of 21.3%.

[0233] LCMS calcld for C 32 H 43 IN7O8S (M+H) + m / z = 812.2, found: 812.0.

[0234] 1 H NMR (400 MHz, DMSO- d 6) δ 9.02-8.95 (m, 2H), 8.76-8.61 (m, 2H), 8.14(s, 1H), 8.10-8.01 (m, 1H), 7.98-7.88 (m, 1H), 7.85-7.76 (m, 1H), 7.32 (brs,3H), 4.44-4.35 (m, 2H), 3.57-3.35 (m, 21H), 3.26-3.15 (m, 2H), 2.60-2.50 (m,2H), 2.32-2.22 (m, 2H), 1.85-1.75 (m, 2H).

[0235] Example 3 Synthesis of Compound 03

[0236] Step 1: In a 250 mL single-necked flask, add 2 g (8.35 mmol) of 5-bromo-4-chloro-2-(methylthioalkyl)pyrimidine and 20 mL of methanol, respectively. Add sodium methoxide (0.59 g, 10.86 mmol) with stirring at 0 °C. Raise to room temperature and react for 8 hours. LCMS analysis indicates the reaction is complete. Quench with saturated ammonium chloride solution, extract three times with ethyl acetate, combine the organic phases, wash twice with saturated brine, dry with anhydrous sodium sulfate, filter, and evaporate the solvent to obtain 1.8 g of the white solid crude product 5-bromo-4-methoxy-2-(methylthioalkyl)pyrimidine 03-b, yield 91.69%. The crude product does not require purification and can be directly added to the next reaction. LCMS (ESI) m / z = 234.9 [M+H] + . 1H NMR (400 MHz, CDCl3): δ 8.35 (s, 1H), 4.06 (s, 3H), 2.55 (s, 3H).

[0237] Step 2: At room temperature, methyl 5-acetylacetate (0.91 g, 7.19 mmol), CuI (105.32 mg, 0.55 mmol), and triethylamine (5.60 g, 55.30 mmol) were added to a solution of 5-bromo-2-(methylthioalkyl)pyridin-4-onitrile (1.3 g, 5.67 mmol) in N,N-dimethylformamide (15 mL). Then, under argon protection, bis(triphenylphosphine)palladium(II) chloride (194.08 mg, 0.28 mmol) was added. The reaction solution was heated to 80 °C. o The reaction was carried out at C for 5 hours. The reaction was monitored by LCMS until completion. The reaction solution was diluted with water (100 mL) and extracted three times with ethyl acetate (20 mL). The organic phases were combined, washed with saturated brine (20 mL x 4), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (5-95% ACN in H2O) to give methyl 6-(5-cyano-6-(methylthioalkyl)pyridin-3-yl)hex-5-ynyl ester 03-c 1.32 g, as a yellow oil, with a yield of 84.7%. LCMS (ESI) m / z = 281.0 [M+H] + .

[0238] Step 3: At room temperature, lithium hydroxide (225.61 mg, 9.42 mmol) was added to a solution of methyl 6-(4-methoxy-2-(methylthio)pyrimidin-5-yl)hexyl-5-acetylic acid (1.32 g, 4.71 mmol) in tetrahydrofuran (10 mL) and water (2 mL). The reaction solution was reacted at 30°C for 3 hours under argon protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated, diluted with water (20 mL), and extracted three times with ethyl acetate (10 mL). The resulting aqueous solution was adjusted to pH 3-4 with 2N hydrochloric acid solution and extracted three times with ethyl acetate (10 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 1.1 g of 6-(4-methoxy-2-(methylthio)pyrimidin-5-yl)hexyl-5-acetylic acid O3-d as a white solid, with a yield of 87.7%. LCMS(ESI)m / z=267.2 [M+H] + .

[0239] Step 4: At room temperature, potassium persulfate (1383 mg, 2.25 mmol) was added to a solution of 6-(4-methoxy-2-(methylthio)pyrimidin-5-yl)hexyl-5-ethynic acid (200 mg, 0.75 mmol) in tetrahydrofuran (10 mL) and water (5 mL). The reaction mixture was allowed to react at room temperature for 24 hours, and the reaction was monitored by LCMS until completion. The reaction mixture was concentrated, diluted with water (10 mL), and extracted three times with ethyl acetate (3 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 202 mg of 6-(2-methylsulfonyl-4-methoxypyrimidin-5-yl)hexyl-5-ethynic acid 03-e as a white solid, with a yield of 90.1%. LCMS (ESI) m / z = 298.9 [M+H] + .

[0240] Step 5: At room temperature, add [chloro(dimethylamino)methylene]-dimethylazonium hexafluorophosphate (228.95 mg, 0.82 mmol) to a solution of 6-(2-methylsulfonyl-4-methoxypyrimidin-5-yl)hexyl-5-alkynic acid (202 mg, 0.68 mmol), N-(14-amino-3,6,9,12-tetraoxotetradec-1-yl)carbamate (251.64 mg, 0.75 mmol), and 1-methyl-1H-imidazolium (117.24 mg, 1.43 mmol) in acetonitrile (4 mL). The reaction mixture was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated, diluted with water (10 mL), and extracted three times with ethyl acetate (3 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC (ethyl acetate) to give N-(14-(6-(2-methylsulfonyl-4-methoxypyrimidin-5-yl)hexyl-5-amido)-3,6,9,12-tetraoxotetradec-1-yl)tert-butyl carbamate 03-f 396 mg, as a white solid, in a yield of 94.83%. LCMS (ESI) m / z = 617.4 [M+H] + .

[0241] Step 6: At room temperature, a solution of trifluoroacetic acid (1 mL) was added to a dichloromethane (1 mL) solution of compound 03-f (396 mg, 0.64 mmol), and the reaction was carried out for 30 minutes at room temperature. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated under reduced pressure to give N-(14-amino-3,6,9,12-tetraoxotetradecane-1-yl)-6-(2-methylsulfonyl-4-methoxypyrimidin-5-yl)hex-5-amide 03-g 320 mg, as a yellow oil, with a yield of 96.4%. LCMS (ESI) m / z = 517.2 [M+H] + .

[0242] Step 7: At room temperature, add tert-butylN-[(1Z)-{[(tert-butoxy)carbonyl]imino}({[(3-iodo-5-[(14-(2-methylsulfonyl-4-methoxypyrimidin-5-yl)hexyl-5-carbamate)-3,6,9,12-tetraoxotetradecane-1-yl)carbamoyl]phenyl)methyl]amino})methyl]carbamate (160 mg, 0.16 mmol), 3-({[(1Z)-{[(tert-butoxy)carbonyl]amino}({[[(tert-butoxy)carbonyl]imino})methyl]amino}methyl)-5-iodobenzoic acid (91.40 mg, 0.18 mmol) and 1-methyl-1H-imidazolium (27.91 mg, 0.34 mmol) to acetonitrile (3 mL). [Chloro(dimethylamino)methylene]-dimethylazonium hexafluorophosphate (53.87 mg, 0.19 mmol) was added to the solution. The reaction mixture was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated, diluted with water (10 mL), and extracted three times with ethyl acetate (3 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC (ethyl acetate) to give tert-butyl N-[(1Z)-{[(tert-butoxy)carbonyl]imino}({[(3-iodo-5-[(14-(2-methylsulfonyl-4-methoxypyrimidin-5-yl)hexyl-5-carbamate)-3,6,9,12-tetraoxotetradecane-1-yl)carbamoyl]phenyl)methyl]amino}methyl]carbamate 0.3-h 112 mg, as a white solid, yield 35.5%. LCMS (ESI) m / z = 1018.4 [M+H] + .

[0243] Step 8: At room temperature, trifluoroacetic acid (1 mL) was added to a solution of tert-butyl compound O3-h (112 mg, 0.11 mmol) in dichloromethane (1 mL). The reaction mixture was allowed to react at room temperature for 30 minutes. The reaction was monitored by LCMS until completion. The residue was purified by high performance liquid chromatography (0.1% trifluoroacetic acid) to give compound O3 30.51 mg, yield 33.9%.

[0244] LCMS(ESI)m / z=818.3 [M+H] + .

[0245] 1 H NMR (400 MHz, DMSO- d 6): δ 8.81(s, 1H), 8.65(t, J = 5.2 Hz, 1H), 8.15(s, 1H), 8.01(t, J = 6.0 Hz, 1H), 7.91(t, J = 5.2 Hz, 1H), 7.81(t, J = 5.2Hz, 2H), 7.24(brs, 3H), 4.39(d, J = 6.4 Hz, 2H), 4.08(s, 3H), 3.52-3.49(m,14H), 3.41-3.38(m, 9H), 3.22-3.18(m, 2H), 2.24(t, J =7.2 Hz, 2H), 1.82-1.76(m, 2H).

[0246] Example 4

[0247] Preparation of 3-(guanidinomethyl)-5-iodo-N-methyl-N-(3,6,9,12,5-pentamethyl-17-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-2,5,8,11,14,17-hexaoxo-3,6,9,12,15-pentazaheptadecyl)benzamide (compound 04)

[0248] Step 1: 10.0 g of 2-chlorotriphenylmethyl resin (10 mmol Cl / g) was swollen in a solid-phase synthesis tube with 100 mL of dry DCM for 1 h. After removing the solvent, 3 eq of Fmoc-sarcosine (9.3 g, 30 mmol), 3 eq of DIEA (5.2 mL, 30 mmol), and 100 mL of DMF were added to the resin, and the mixture was bubbled under nitrogen at room temperature for 2 h. After removing the solvent, 100 mL of methanol was added, and the reaction was continued for 1 h to block the active end. The resin was washed three times alternately with DCM and DMF. Then, 3 eq of Fmoc-sarcosine (× 2) was coupled to the resin in DMF with PyBop (15.6 g, 30 mmol) and DIEA (5.2 mL, 30 mmol), and deprotected for 15 min with a 20% piperidine DMF solution after each coupling. Subsequently, 10 eq of bromoacetic acid, 13 eq of diisopropylcarbodiimide, and 100 mL of DMF were added to the washed resin, and the mixture was bubbled under nitrogen at room temperature for 1 hour. After the reaction was complete, the resin was washed three times alternately with DCM and DMF. 40 mL of 40% (wt) methylamine aqueous solution was added to the bromoacetylated resin, and the mixture was incubated on a shaker for 30 minutes, followed by washing three times alternately with DCM and DMF. This process was repeated three times. After the reaction was complete, the resin was cleaved for 2 h with 95% TFA, 2.5% TIPS, and 2.5% H2O. After filtering the mixture, the filtrate was evaporated to dryness and purified by reversed-phase preparative chromatography to obtain 2 g of compound O4-f as a colorless oil. LCMS (ESI) M / Z: 445.3 [M+H] + .

[0249] Step 2: At room temperature, 2 g of O4-f was dissolved in 40 mL of water in a single-necked flask, followed by the addition of TEA (1.9 mL, 13.5 mmol) and (Boc)2O (1 mL, 4.5 mmol). The mixture was stirred at room temperature for 3 h. The reaction was monitored by LCMS to indicate completion. The reaction solution was diluted with 50 mL of water, extracted twice with 50 mL of EA, and the aqueous phase was lyophilized to obtain the crude product. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 50%) to obtain 1.5 g of compound O4-g as a colorless oily liquid, with a yield of 62%. LCMS (ESI) M / Z: 545.5 [M+H] + .

[0250] Step 3: At room temperature, in a single-necked flask, 04-g (500 mg, 0.92 mmol) was dissolved in 20 mL of acetonitrile, followed by the addition of 04-h (191 mg, 0.92 mmol), TCFH (515 mg, 1.84 mmol), and NMI (0.35 mL, 3.68 mmol). The system was stirred at room temperature for 2 h. The reaction was monitored by LCMS to indicate completion. The reaction solution was evaporated to dryness to obtain the crude product of compound 04-i. The crude product did not require purification and proceeded directly to the next step. LCMS (ESI) M / Z: 734.3 [M+H] + .

[0251] Step 4: At room temperature, in a single-necked flask, dissolve 500 mg of crude 04-i in 10 mL of DCM, add 3 mL of TFA, and stir the system at room temperature for 2 h. Monitor the reaction completion by LCMS. Dry the reaction mixture to obtain the crude product. Purify the crude product by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 50%) to obtain 300 mg of compound 04-j as a yellow solid. LCMS (ESI) M / Z: 634.4 [M+H] + .

[0252] Step 5: At room temperature, in a single-necked flask, compound 04-j (300 mg, 0.47 mmol) was dissolved in 5 mL of acetonitrile. Then, compound 01-e (243 mg, 0.47 mmol), TCFH (263 mg, 0.94 mmol), and NMI (170 μL, 1.88 mmol) were added, and the system was stirred at room temperature for 2 h. The reaction was monitored by LCMS to indicate completion. The reaction solution was evaporated to dryness to obtain the crude product. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 100%) to obtain compound 04-k 200 mg as a yellow solid. LCMS (ESI) M / Z: 1135.2 [M+H] + .

[0253] Step 6: At room temperature, in a single-necked flask, compound 04-k (200 mg, 0.18 mmol) was dissolved in 10 mL of THF / H2O (1 / 1, v / v), followed by the addition of potassium peroxymonosulfonate (622 mg, 1.8 mmol). The system was heated at 30 °C and stirred for 2 h. The reaction was monitored by LCMS to indicate completion. The reaction solution was diluted with water (30 mL), extracted three times with 50 mL of DCM / MeOH (10:1), and the organic phases were combined. After washing with saturated brine, the solution was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude compound 04-k. The crude product did not require purification and proceeded directly to the next step. LCMS (ESI) M / Z: 1167.4 [M+H] + .

[0254] Step 7: At room temperature, in a single-necked flask, compound 04-l (200 mg, crude product) was dissolved in 4 mL of DCM, and 2 mL of TFA was added. The system was stirred at room temperature for 4 h. The reaction was monitored by LCMS until completion. The reaction solution was evaporated to dryness to obtain the crude product. The crude product was purified by Prep HPLC to obtain 04 4 mg of trifluoroacetate of compound 3-(guanidinyl)-5-iodo-N-methyl-N-(3,6,9,12,5-pentamethyl-17-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-2,5,8,11,14,17-hexaoxo-3,6,9,12,15-pentazocephaloyl)benzamide.

[0255] LCMS (ESI) M / Z: 967.1 [M+H] + ;

[0256] 1 H NMR (400 MHz, DMSO- d 6) δ 10.78-10.34 (m, 1H), 8.31-7.47 (m, 6H), 7.41-6.83 (m, 5H), 4.49-3.88 (m, 14H), 3.70 (s, 3H), 3.26-2.56 (m, 18H).

[0257] Example 5

[0258] Preparation of (4S)-4-{[(1S)-3-carboxy-1-[(2-(2-(3-[(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propamido)ethoxy)ethyl)carbamoyl]propyl]carbamoyl}-4-(2-(4-iodophenyl)acetamamido)butyric acid (compound 05)

[0259] Step 1: At room temperature, N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (3.96 g, 14.1 mmol) was added to a solution of Fmoc-O-tert-butyl-L-glutamic acid (5 g, 11.75 mmol), methanol (1129.41 mg, 35.25 mmol), and N-methylimidazole (2.03 g, 24.68 mmol) in acetonitrile (50 mL). The reaction solution was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was concentrated, diluted with water (100 mL), and extracted three times with ethyl acetate (30 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 0.5-b4.96 g of compound 5-tert-butyl-1-methyl(2S)-2-({[(9H-fluorene-9-yl)methoxy]carbonyl}amino)glutarate, a yellow oil with a yield of 96.03%. LCMS (ESI) m / z = 462.2 [M+Na] + .

[0260] Step 2: At room temperature, diethylamine (10 mL) was added to a tetrahydrofuran (50 mL) solution of 5-tert-butyl-1-methyl(2S)-2-({[(9H-fluoren-9-yl)methoxy]carbonyl}amino)glutarate (4.96 g, 11.29 mmol)). The reaction mixture was allowed to react at room temperature for 2 hours. The reaction was monitored by LCMS to indicate completion. The residue was purified by silica gel column chromatography (0-10% methanol in dichloromethane) to give 1.2 g of compound 1-methyl(2S)-2-aminoglutarate 5-tert-butyl 05-c, as a yellow oil, with a yield of 48.9%. LCMS (ESI) m / z = 218.1 [M+H] + .

[0261] Step 3: At room temperature, N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (1535.33 mg, 5.47 mmol) was added to a solution of 1-methyl(2S)-2-aminopentanoic acid 5-tert-butyl ester (1.1 g, 4.56 mmol, 90% purity), 2-(4-iodophenyl)acetic acid (1.19 g, 4.56 mmol), and N-methylimidazolium (786.19 mg, 9.58 mmol) in acetonitrile (40 mL). The reaction solution was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was concentrated, diluted with water (50 mL), and ethyl acetate (25 mL) was added. 3) Extracted three times, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-30% ethyl acetate in petroleum ether) to give compound 1-methyl(2S)-2-(2-(4-iodophenyl)acetamitano)glutaric acid 5-tert-butyl ester (0.5-e 720 mg, yellow oil, yield 34.2%). LCMS (ESI) m / z = 462.1 [MH] + .

[0262] Step 4: At room temperature, lithium hydroxide (74.72 mg, 3.12 mmol) was added to a tetrahydrofuran (8 mL) and water (2 mL) solution of compound 05-e (720 mg, 1.56 mmol). The reaction mixture was reacted at room temperature under argon protection for 2 hours. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (0-40% acetonitrile in water) to give compound (2S)-5-(tert-butoxy)-2-(2-(4-iodophenyl)acetamide)-5-oxopentanoic acid 05-f 660 mg as a white solid, yield 94.5%. LCMS (ESI) m / z = 446.1 [MH] + .

[0263] Step 5: At room temperature, N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (498.3 mg, 1.78 mmol) was added to a solution of (2S)-5-(tert-butoxy)-2-(2-(4-iodophenyl)acetamide)-5-oxopentanoic acid (660 mg, 1.48 mmol), 5-tert-butyl 1-methyl (2S)-2-aminopentanedioate (0.35 g, 1.63 mmol), and N-methylimidazole (255.1 mg, 3.11 mmol) in acetonitrile (15 mL). The reaction solution was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was concentrated, diluted with water (30 mL), and ethyl acetate (15 mL) was added. 3) Extraction was performed three times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:2) to obtain compound 5-tert-butyl-1-methyl(2S)-2-[(2S)-5-(tert-butoxy)-2-(2-(4-iodophenyl)acetamido)-5-oxopentanamido]glutarate, 0.5-g to 910 mg, as a yellow oil, with a yield of 95.4%. LCMS (ESI) m / z = 647.3 [M+H] + .

[0264] Step 6: At room temperature, lithium hydroxide (62.7 mg, 2.62 mmol) was added to a tetrahydrofuran (8 mL) and water (2 mL) solution of 5-tert-butyl-1-methyl(2S)-2-[(2S)-5-(tert-butoxy)-2-(2-(4-iodophenyl)acetamyl)-5-oxopentamido]glutarate (850 mg, 1.31 mmol) (850 mg, 1.31 mmol). The reaction solution was reacted at room temperature under argon protection for 2 hours. The reaction was monitored by LCMS until completion. The reaction solution was concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (0-40% acetonitrile in water) to obtain (2S)-5-(tert-butoxy)-2-[(2S)-5-(tert-butoxy)-2-(2-(4-iodophenyl)acetamyl)-5-oxopentamido]-5-oxopentamidoic acid (0.5-h 630 mg), as a white solid, with a yield of 75.7%. LCMS(ESI)m / z=633.3 [M+H] + .

[0265] Step 7: At room temperature, N,N,N',N'-tetramethylchloromethanesulfonate (323.2 mg, 1.15 mmol) was added to a solution of 0.5-h (610 mg, 0.96 mmol), N-(2-(2-aminoethoxy)ethoxyethyl)carbamate (391.1 mg, 1.06 mmol), and N-methylimidazole (165.5 mg, 2.02 mmol) in acetonitrile (10 mL). The reaction solution was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was concentrated, diluted with water (30 mL), and ethyl acetate (15 mL) was added. 3) Extracted three times, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-50% ethyl acetate in petroleum ether) to give compound (4S)-4-[(2S)-5-(tert-butoxy)-2-(2-(4-iodophenyl)acetamityl)-5-oxopentanamido]-4-[(2-({[(9H-fluorene-9-yl)methoxy]carbonyl}amino)ethoxy)ethoxyoxy)ethyl)carbamoyl]tert-butyl butyrate 0.5-j 750 mg, as a yellow oil, yield 78.9%. LCMS (ESI) m / z = 1007.5 [M+Na] + .

[0266] Step 8: At room temperature, diethylamine (2 mL) was added to a tetrahydrofuran (4 mL) solution of compound 05-j (750 mg, 0.76 mmol). The reaction mixture was reacted at room temperature for 2 hours under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-10% methanol in dichloromethane) to give compound (4S)-4-{[(1S)-1-[(2-(2-aminoethoxy)ethoxyethyl)carbamoyl]-4-(tert-butoxy)-4-oxobutyl]carbamoyl}-4-(2-(4-iodophenyl)acetamido)butyrate tert-butyl ester 05-k 368 mg, as a yellow oil, with a yield of 63.3%. LCMS (ESI) m / z = 763.2 [M+H] + .

[0267] Step 9: At room temperature, HATU (59.3 mg, 0.16 mmol) was added to a solution of 3-[(4-(5-methylsulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propionic acid 05-l (48.5 mg, 0.14 mmol) in N,N-dimethylformamide (2 mL). The reaction mixture was reacted at room temperature under argon protection for 30 min. Then, compound 05-k (100 mg, 0.13 mmol) and diisopropylethylamine (50.40 mg, 0.39 mmol) were added to the above solution, and the reaction mixture was reacted at room temperature under argon protection for 16 hours. The reaction was monitored by LCMS to indicate completion. The reaction mixture was diluted with water (10 mL) and ethyl acetate (5 mL) was added. 3) Extract three times, combine the organic phases, and rinse with saturated saline (5 mL). 4) After washing, the mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC (PE / EtOAc = 1 / 2) to give tert-butyl(4S)-4-{[(1S)-4-(tert-butoxy)-1-[(2-(2-(3-[(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propamido)ethoxy(ethoxy)ethyl)carbamoyl]-4-oxobutyl]carbamoyl}-4-(2-(4-iodophenyl)acetamamido)butyrate 0.5-m 90 mg, as a yellow solid, with a yield of 63.3%. LCMS (ESI) m / z = 1084.4 [M+H] + .

[0268] Step 10: At room temperature, trifluoroacetic acid (0.5 mL) was added to a solution of compound 05-m (90 mg, 0.083 mmol) in dichloromethane (0.5 mL). The reaction mixture was allowed to react at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated, and the residue was purified by high performance liquid chromatography (0.1% TFA) to give compound (4S)-4-{[(1S)-3-carboxy-1-[(2-(2-(3-[(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propamido)ethoxy)ethyl)carbamoyl]propyl]carbamoyl}-4-(2-(4-iodophenyl)acetamamido)butyric acid 05 20.50 mg, yield 25.4%.

[0269] LCMS(ESI)m / z = 972.3 [M+H] + .

[0270] 1 H NMR (400 MHz, DMSO-d6): δ 12.11(brs, 2H), 10.39(s, 1H), 8.36-8.18(m, 1H), 8.04(d, J = 8.8 Hz, 2H), 7.99-7.84(m, 5H), 7.63(d, J = 8.0 Hz, 2H), 7.08-7.05(m, 2H), 4.25-4.17(m, 2H), 3.69(s, 3H), 3.42-3.36(m, 10H), 3.24-3.13(m, 4H), 2.60(t, J =6.8 Hz, 2H), 2.44(t, J = 6.8 Hz, 2H), 2.24-2.17(m, 4H), 1.91-1.68(m, 4H).

[0271] Example 6 Synthesis of Compound 06

[0272] Step 1: At room temperature, NMI (577.98 mg, 7.04 mmol) was added to a solution of 2-(methylthioalkyl)pyrimidine-5-carboxylic acid (300 mg, 1.76 mmol), N-(14-amino-3,6,9,12-tetraoxotetradec-1-yl)carbamate tert-butyl ester (710.52 mg, 2.11 mmol), and TCFH (740.73 mg, 2.64 mmol) in ACN (2 mL). The reaction mixture was reacted at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The mixture was diluted with water (20 mL) and extracted three times with ethyl acetate (30 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to give N-(14-[(2-(methylthioalkyl)pyrimidin-5-yl)carbamate]-3,6,9,12-tetraoxotetradecane-1-yl)carbamate tert-butyl 06-c 500 mg, as a yellow solid, with a yield of 58.1%. LCMS calculation for C 21 H 36 N4O7S (M+H) + m / z = 489.3, found: 489.3.

[0273] Step 2: At room temperature, Oxone (0.50 g, 0.82 mmol) was added to a solution of N-(14-[(2-(methylthioalkyl)pyrimidin-5-yl)formamido]-3,6,9,12-tetraoxotetradecane-1-yl)carbamate (200 mg, 0.41 mmol) in THF (6 mL) and water (3 mL). The reaction solution was reacted at 25 °C for 16 hours under argon protection. LCMS monitoring showed that the reaction was complete. The solution was diluted with water (20 mL) and extracted three times with ethyl acetate (30 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give N-(14-[(2-methylsulfonylpyrimidin-5-yl)formamido]-3,6,9,12-tetraoxotetradecane-1-yl)carbamate 06-d 210 mg, as a yellow solid, in yield 98.5%. LCMS calculation for C 21 H 36 N4O9S (M+H) + m / z = 521.3, found: 521.3.

[0274] Step 3: At room temperature, N-(14-[(2-methylsulfonylpyrimidin-5-yl)carbamate]-3,6,9,12-tetraoxotetradecane-1-yl)carbamate tert-butyl ester (170 mg, 0.33 mmol) was dissolved in MeOH (1 mL), and 4 M HCl / EtOAc (4 mL) was added. The mixture was stirred at room temperature for 30 minutes. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated under reduced pressure to give N-(14-amino-3,6,9,12-tetraoxalate-tradecan-1-yl)-2-methanesulfonylpyrimidin-5-carbamate 06-e 130 mg, which was a yellow oil. LCMS calcld for C 16 H 28 N4O7S (M+H) + m / z = 421.3, found:421.3.

[0275] Step 4: At room temperature, DIEA (0.20 g, 1.55 mmol) was added to a DMF (5 mL) solution of N-(14-amino-3,6,9,12-tetraoxotetradec-1-yl)-2-methylsulfonylpyrimidine-5-carboxamide (130 mg, 0.31 mmol), 3-({[(1Z)-{[(tert-butoxy)carbonyl]amino}({[[(tert-butoxy)carbonyl]imino})methyl]amino}methyl)-5-iodobenzoic acid (193.2 mg, 0.37 mmol), and HATU (0.18 g, 0.46 mmol). The reaction mixture was reacted at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The mixture was diluted with water (20 mL), extracted three times with ethyl acetate (20 mL), the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse-phase chromatography to give tert-butylN-[(1Z)-{[(tert-butoxy)carbonyl]imino}({[(3-iodo-5-[(14-[(2-methylsulfonylpyrimidin-5-yl)carbamate]-3,6,9,12-tetraoxotetradecane-1-yl)carbamoyl]phenyl)methyl]amino})methyl]carbamate 0.6-g 20 mg, as a yellow solid. LCMS calcld for C 35 H 52 IN7O 12 S (M+H)+ m / z =922.2, found:922.2.

[0276] Step 5: At room temperature, compound 06-g (20 mg, 0.022 mmol) was dissolved in DCM (2 mL), and TFA (2 mL) was added. The mixture was stirred at room temperature for 30 minutes. The reaction was monitored by LCMS until completion. The reaction system was concentrated, and the crude product was purified by high performance liquid chromatography to obtain N-(14-[(3-(carbamoylaminomethyl)-5-iodophenyl)carbamoyl]-3,6,9,12-tetraoxotetradecane-1-yl)-2-methanesulfonylpyrimidine-5-carboxamide trifluoroacetate 06 10.61 mg.

[0277] LCMS calcld for C 25 H 36 IN7O8S (M+H) + m / z = 722.1, found: 722.1;

[0278] 1 H NMR (400 MHz, DMSO- d 6 ) δ 9.37 (s, 2H), 9.17-9.08 (m, 1H), 8.69-8.61(m, 1H), 8.14 (s, 1H), 8.07-8.00 (m, 1H), 7.84-7.97 (m, 2H), 7.28 (brs, 3H), 4.43-4.35 (m, 2H), 3.62-3.42 (m, 19H), 3.40-3.30 (m, 4H).

[0279] Example 7 Synthesis of Compound 07

[0280] Step 1: At room temperature, triethylamine (6 mL) and palladium dichloride dichloride (342.53 mg, 0.49 mmol) were added to a DMF (1 mL) solution of 5-bromo-2-methylmercaptopyrimidine (1 g, 4.88 mmol), trimethylethynylsilane (2.40 g, 24.4 mmol), and cuprous iodide (92.94 mg, 0.49 mmol). The reaction solution was heated at 120 °C under argon protection. oThe reaction mixture was sealed in a tube at C for 16 hours. After cooling to room temperature, TLC showed the formation of new spots. The reaction mixture was diluted with water (20 mL), extracted three times with dichloromethane (5 mL), and the organic phases were combined. The mixture was washed four times with saturated brine (5 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at low temperature. The crude product was purified by silica gel column chromatography (0-5% ethyl acetate / petroleum ether) to give compound 2-(methylthioalkyl)-5-(2-(trimethylsilyl)ethynyl)pyrimidine 07-b 730 mg, as a yellow oil, with a yield of 67.3%. 1 H NMR (400 MHz, CDCl3): δ 8.55 (s, 2H), 2.57 (s, 3H), 0.26 (s, 9H).

[0281] Step 2: At room temperature, potassium carbonate (559.7 mg, 4.05 mmol) was added to a methanol (3 mL) solution of 2-(methylthioalkyl)-5-(2-(trimethylsilyl)ethynyl)pyrimidine 07-b (300 mg, 1.35 mmol). The reaction mixture was allowed to react at room temperature for 30 minutes. The reaction was monitored by LCMS to indicate completion. The reaction mixture was concentrated, diluted with water (20 mL), and extracted three times with dichloromethane (5 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at low temperature to obtain 200 mg of compound 5-(2-hydroxyethylidene)-2-(methylthioalkyl)pyrimidine 07-c as a yellow oil, with a yield of 98.7%. LCMS (ESI) m / z = 151.1 [M+H] + .

[0282] Step 3: At room temperature, Oxone (2.45 g, 3.99 mmol) was added to a mixture of tetrahydrofuran (10 mL) and water (5 mL) of 5-(2-hydroxyethylidene)-2-(methylthioalkyl)pyrimidine 07-c (200 mg, 1.33 mmol). The reaction mixture was allowed to react at room temperature for 2 hours. The reaction was monitored by LCMS to indicate completion. The reaction mixture was diluted with water (50 mL), extracted three times with dichloromethane (10 mL), and the organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at low temperature to give compound 5-ethynyl-2-methylsulfonylpyrimidine 07-d 200 mg, as a yellow solid (yield 82.4%). LCMS (ESI) m / z = 183.1 [M+H] + .

[0283] Step 4: At 0°C, a boronane dimethyl sulfide complex (4.10 mL, 8.22 mmol, 2 M in THF) was slowly added to a tetrahydrofuran (15 mL) solution of 1-{[(tert-butoxy)carbonyl]amino}-3,6,9,12-tetraoxopentadecane-15-acid 07-e (1 g, 2.74 mmol). The reaction mixture was incubated at room temperature for 16 hours under argon protection. The reaction was monitored by LCMS to indicate completion. The reaction mixture was cooled to 0 °C, and the reaction was quenched by slowly adding methanol (20 mL). The mixture was then stirred at room temperature for 30 minutes. The reaction mixture was concentrated under reduced pressure to give 1.1 g of N-(15-hydroxy-3,6,9,12-tetraoxopentadecane-1-yl)carbamate tert-butyl 07-f as a white solid, with a yield of 97.2%. LCMS (ESI) m / z = 352.3 [M+H] + .

[0284] Step 5: At 0°C, 2-methylpropane-2-sulfonyl chloride (0.74 g, 4.70 mmol) was slowly added to a solution of N-(15-hydroxy-3,6,9,12-tetraoxopentadecane-1-yl)carbamate tert-butyl 07-f (1.10 g, 3.13 mmol) and triethylamine (0.95 g, 9.39 mmol) in dichloromethane (10 mL). The reaction mixture was incubated at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was diluted with water (30 mL), and extracted three times with dichloromethane (10 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 0.7 g to 1.2 g of N-(15-(methanesulfonyloxy)-3,6,9,12-tetraoxopentadecane-1-yl)carbamate, as a pale yellow oil, with a yield of 89.2%. LCMS (ESI) m / z = 452.3 [M+Na] + .

[0285] Step 6: At room temperature, sodium azide (1.81 g, 27.9 mmol) was added to a DMF (10 mL) solution of N-(15-(methanesulfonyloxy)-3,6,9,12-tetraoxopentadecane-1-yl)carbamate 0.7-g (1.2 g, 2.79 mmol). The reaction solution was reacted at 80°C for 16 hours under argon protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was diluted with water (50 mL), extracted three times with ethyl acetate (10 mL), the organic phases were combined, washed four times with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain N-(15-azido-3,6,9,12-tetraoxopentadecane-1-yl)carbamate 0.7-h 1 g, as a yellow oil, with a yield of 95.1%. LCMS(ESI)m / z=399.2 [M+Na] + .

[0286] Step 7: At room temperature, 5-ethynyl-2-methylsulfonylpyrimidine 07-d (80.17 mg, 0.44 mmol), copper sulfate (6.38 mg, 0.040 mmol), and sodium vitamin C (0.016 g, 0.080 mmol) were added to a solution of N-(15-azido-3,6,9,12-tetraoxopentacapram-1-yl)carbamate 07-h (150 mg, 0.40 mmol) in water (2 mL) and tert-butanol (2 mL). The reaction mixture was reacted at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was diluted with water (15 mL), extracted three times with ethyl acetate (5 mL), and the organic phases were combined. The mixture was washed with saturated brine (15 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 210 mg of N-(15-(4-(2-methanesulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentapentadecane-1-yl)carbamate, as a pale yellow oil, with a yield of 94.3%. LCMS (ESI) m / z = 581.4 [M+Na] + .

[0287] Step 8: At room temperature, trifluoroacetic acid (1 mL) was added to a solution of N-(15-(4-(2-methanesulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentadecane-1-yl)carbamate tert-butyl ester 07-i (210 mg, 0.38 mmol) in dichloromethane (1 mL). The reaction mixture was allowed to react at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated under reduced pressure to give 170 mg of 15-(4-(2-methylmethanesulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentadecane-1-amine 07-j as a white solid, with a yield of 98.6%. LCMS (ESI) m / z = 459.1 [M+H] + .

[0288] Step 9: At room temperature, N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (50.50 mg, 0.18 mmol) was added to a solution of 15-(4-(2-methylmethanesulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentadecane-1-amine 07-j (70 mg, 0.15 mmol), 3-({[(1E)-{[(tert-butoxy)carbonyl]amino}({[[(tert-butoxy)carbonyl]imino})methyl]amino}methyl)-5-iodobenzoic acid 01-e (85.69 mg, 0.17 mmol), and 1-methyl-1H-imidazolium (25.86 mg, 0.32 mmol). The reaction solution was reacted at 30°C for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The reaction solution was concentrated, diluted with water (10 mL), and extracted three times with ethyl acetate (5 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC (PE / EtOAc = 1 / 2) to give tert-butyl N-[(1E)-{[(tert-butoxy)carbonyl]imino}({[(3-iodo-5-[(15-(4-(2-methylsulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentadecane-1-yl)carbamoyl]phenyl)methyl]amino})methyl]carbamate 07-k 68 mg, as a yellow oil, with a yield of 46.4%. LCMS (ESI) m / z = 960.1 [M+H] + .

[0289] Step 10: At room temperature, add 0.5 mL of trifluoroacetic acid to a solution of tert-butyl N-[(1E)-{[(tert-butoxy)carbonyl]imino}({[(3-iodo-5-[(15-(4-(2-methylsulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentadecane-1-yl)carbamoyl]phenyl)methyl]amino})methyl]carbamate 07-k (68 mg, 0.071 mmol) in dichloromethane. The reaction mixture was allowed to react at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction solution was concentrated, and the resulting residue was purified by high performance liquid chromatography (0.1% TFA) to give 6.07 mg of compound 3-(carbamoylaminomethyl)-5-iodo-N-(15-(4-(2-methanesulfonylpyrimidin-5-yl)-1H-1,2,3-triazol-1-yl)-3,6,9,12-tetraoxopentadecane-1-yl)benzamide, with a yield of 11.3%.

[0290] LCMS(ESI)m / z=760.2 [M+H] + ;

[0291] 1 H NMR (400 MHz, DMSO- d 6): δ 9.48(s, 2H), 8.94(s, 1H), 8.64(t, J = 5.2 Hz, 1H), 8.14 (s, 1H), 7.98 (t, J = 6.0 Hz, 1H), 7.80 (d, J = 6.8 Hz, 2H), 7.24(brs, 3H), 4.54(t, J = 7.2 Hz, 2H), 4.39 (d, J = 6.4 Hz, 2H), 3.51-3.49(m, 14H), 3.44(s, 3H), 3.43-3.36(m, 4H), 2.15-2.09(m, 2H).

[0292] Example 8 Synthesis of Compound 09

[0293] Step 1: At room temperature, NMI (0.95 g, 11.56 mmol) was added to a solution of 3-({[(1Z)-{[(tert-butoxy)carbonyl]amino}({[[(tert-butoxy)carbonyl]imino})methyl]amino}methyl)-5-iodobenzoic acid O1-e (1.50 g, 2.89 mmol), (2S)-2-amino-4-(methylthioalkyl)butyrate methyl ester (0.52 g, 3.18 mmol), and TCFH (0.97 g, 3.47 mmol) in ACN (20 mL). The reaction mixture was reacted at room temperature for 1 hour under argon protection. The reaction was monitored by LCMS until completion. The mixture was diluted with 50 mL of water and ethyl acetate (50 mL) was added. 3) Extracted three times, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse-phase chromatography to give methyl (2S)-2-[(3-({[(1E)-{[(tert-butoxy)carbonyl]amino}({[[(tert-butoxy)carbonyl]imino})methyl]amino}methyl)-5-iodophenyl)carbamate]-4-(methylthioalkyl)butyrate 09-b 1.0 g, as a yellow solid, with a yield of 46.9%. LCMS calculation for C 25 H 38 IN4O7S (M+H)+ m / z = 665.2, found: 665.2.

[0294] Step 2: At room temperature, compound 09-b (1 g, 1.50 mmol) and THF (15 mL) were added, followed by 3 mL of an aqueous solution of LiOH (107 mg, 4.5 mmol). The reaction system was allowed to react at room temperature for 2 h. The reaction was monitored by LCMS. 1 M HCl was added to adjust the pH to 6, and the system was concentrated to obtain the crude product. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to obtain compound (E)-(3-((2,3-bis(tert-butyloxycarbonyl)guanidinyl)methyl)-5-iodobenzoyl)-L-methionine 09-c 920 mg, as a yellow solid, with a yield of 94%. LCMS calculation for C 24 H 36 IN4O7S (M+H)+ m / z = 651.1, found: 651.1.

[0295] Step 3: At room temperature, compound 09-c (920 mg, 1.41 mmol) and L-valine methyl ester (204 mg, 1.55 mmol) were dissolved in 20 mL of anhydrous DCM, and EEDQ (417 mg, 1.69 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The mixture was then diluted with 30 mL of water and 40 mL of DCM was added. 3) Extraction was performed three times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluted with PE in EA from 0% to 30%) to obtain compound (3-(((E)-2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzoyl)-L-methionyl-L-valine methyl ester 09-d 0.9 g, as a yellow solid, with a yield of 83%. LCMS calculation for C 30 H 47 IN5O8S (M+H) + m / z = 764.2, found: 764.2.

[0296] Step 4: At room temperature, compound 09-d (0.9 g, 1.18 mmol) was dissolved in 10 mL of THF, and then 3 mL of an aqueous solution of LiOH (85 mg, 3.54 mmol) was added. The reaction system was then reacted at room temperature for 2 h. The reaction was monitored by LCMS. 1 M HCl was added to adjust the pH of the system to 6, and the system was concentrated to obtain the crude product. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to obtain compound (3-(((E)-2,3-bis(tert-butyloxycarbonyl)guanidin)methyl)-5-iodobenzoyl)-L-methionyl-L-valine 09-e 700 mg, as a yellow solid, with a yield of 79%. LCMS calculation for C 29 H 45 IN5O8S (M+H) + m / z = 750.2, found: 750.2

[0297] Step 5: At room temperature, compound 09-e (700 mg, 0.93 mmol) and (S)-3-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-2-aminopropionate tert-butyl ester 09-f (392 mg, 1.02 mmol) were dissolved in 10 mL of anhydrous DCM, and EEDQ (275 mg, 1.12 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The mixture was diluted with 30 mL of water and 40 mL of DCM was added. 3) Extracted three times, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to obtain tert-butyl(S)-3-((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-2-((S)-2-(S)-2-(3-((((E)-2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzamido)-4-(methylthio)butamido)-3-methylbutamido 0.9-g 550 mg, as a yellow solid, yield 53%. LCMS calculation for C 51 H 69 IN7O 11 S (M+H) + m / z = 1114.3, found: 1114.4.

[0298] Step 6: At room temperature, DEA (3.55 g, 48.54 mmol) was added to a DCM (5 mL) solution of compound 09-g (200 mg, 0.18 mmol). The reaction solution was reacted at room temperature under argon protection for 16 hours. The reaction was monitored by LCMS. The reaction solution was concentrated, and the crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to give compound (S)-3-amino-2-((S)-2-((R)-2-(3-(((E)-2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzamido)-4-(methylthio)butamido)-3-methylbutamido 09-h 100 mg, as a yellow solid, with a yield of 62%. LCMS calculation for C 36 H 59 IN7O9S (M+H) + m / z = 892.3, found: 892.3.

[0299] Step 7: At room temperature, HATU (59.3 mg, 0.16 mmol) was added to a solution of 3-[(4-(5-methylsulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propionic acid 05-L (48.5 mg, 0.14 mmol) in N,N-dimethylformamide (2 mL). The reaction mixture was reacted at room temperature under argon protection for 30 min. Then, 3-(2-(2-(3-aminopropoxy)ethoxy)ethoxy)propionic acid (32.9 mg, 0.14 mmol) and diisopropylethylamine (50.40 mg, 0.39 mmol) were added to the above solution, and the reaction mixture was reacted at room temperature under argon protection for 16 hours. The reaction was monitored by LCMS to ensure completion. The reaction mixture was diluted with water (10 mL) and ethyl acetate (5 mL) was added. 3) Extract three times, combine the organic phases, and rinse with saturated saline (5 mL). 4) After washing, the mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC (PE / EtOAc = 1 / 2) to give compound 3-(2-(2-(3-(3-[(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propamido)propoxy)ethoxy)methoxy)propionic acid 09-i 37.4 mg, as a yellow solid, with a yield of 46%. LCMS calculation for C 23 H 32 N4O 10 S (M+H) + m / z = 557.18, found: 557.2.

[0300] Step 8: At room temperature, compound 09-h (50 mg, 0.056 mmol), 3-(2-(2-(3-(3-[(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propamido)propoxy)ethoxy)methoxy)propionic acid 09-i (37.4 mg, 0.067 mmol), and EEDQ (27.7 mg, 0.11 mmol) were dissolved in DCM (2 mL). The reaction mixture was reacted at room temperature for 2 hours under argon protection. The reaction was monitored by LCMS until completion. The system was concentrated to obtain a crude product, which was then purified by reverse chromatographic column chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to give tert-butyl(S)-2-((S)-2-[(R)-2-(3-((((E)-2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzamido)-4-(methylthio)butamido)-3-methylbutamido]-22-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-5,19,22-trioxo-8,11,14-trioxo-4,18-diazacosanoate 09-j 20 mg, as a yellow solid, with a yield of 25%. LCMS calculation for C 59 H 89 IN 11 O 18 S2 (M+H) + m / z = 1430.4, found:1430.5.

[0301] Step 9: At room temperature, compound 09-j (20 mg, 0.014 mmol) was dissolved in DCM (S0, 2 mL), and TFA (3.06 g, 26.8 mmol) was added. The mixture was stirred at room temperature for 30 minutes. The reaction was monitored by LCMS to indicate completion. The system was concentrated to obtain a crude product, which was then purified by high performance liquid chromatography (0.1% FA) to give compound (S)-2-((S)-2-(S)-2-[(3-(guanidinyl)-5-iodobenzamido)-4-(methylthio)butamido)-3-methylbutamido]-22-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-5,19,22-trioxo-8,11,14-trioxo-4,18-diazacosanoic acid 09 8.10 mg, yield 49%.

[0302] LCMS calcld for C 45 H 65 IN 11 O14 S2 (M+H) + m / z = 1174.3, found: 1174.0;

[0303] 1 H NMR (400 MHz, DMSO- d 6 ) δ 10.45 (s, 1H), 10.00 (brs, 1H), 8.64-8.55(m, 1H), 8.12 (s, 1H), 8.08-7.97 (m, 3H), 7.94-7.82 (m, 5H), 7.67-7.22 (m,4H), 4.59-4.40 (m, 2H), 4.33-4.21 (m, 1H), 4.15-4.06 (m, 1H), 4.02-3.94 (m,1H), 3.70 (s, 3H), 3.62-3.36 (m, 16H), 3.14-3.03 (m, 2H), 2.66-2.52 (m, 4H),2.47-2.38 (m, 2H), 2.33-2.23 (m, 2H), 2.12-1.94 (m, 6H), 1.68-1.55 (m, 2H),0.88-0.76 (m, 6H).

[0304] Example 9

[0305] Preparation of (S)-24-((S)-2-((S)-2-(3-(guanidinyl)-5-iodobenzoylamino)-3-methylbutyramide)-4-(methylthio)butyramide)-1-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-1,4,18-trioxo-9,12,15-trioxy-5,19-diazapentane-25 carboxylic acid (compound 10)

[0306] Step 1: At room temperature, (Z)-3-((2,3-bis(tert-butyloxycarbonyl)guanidin)methyl)-5-iodobenzoic acid O1-e (2 g, 3.8 mmol) and L-valine methyl ester (547.6 mg, 4.1 mmol) were dissolved in 40 mL of anhydrous DCM, and EEDQ (1.13 g, 4.56 mmol) was added. The reaction solution was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The solution was diluted with 30 mL of water, and 40 mL of DCM was added. 3) Extraction was performed three times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give compound (Z)-(3-((2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzoyl)-L-valine methyl ester 10-a 2 g, as a yellow solid, with a yield of 82.3%. LCMS calculation for C 25 H 38 IN4O7 (M+H) + m / z = 633.2, found: 633.3.

[0307] Step 2: Under ice bath conditions, compound 10-a (2 g, 3.16 mmol) was dissolved in 15 mL of THF, followed by the addition of 6 mL of an aqueous solution of lithium hydroxide (227 mg, 9.48 mmol). The reaction was allowed to proceed at room temperature for 4 h. The reaction was monitored by LCMS. The pH was adjusted to 6 with 1 M HCl, and the system was concentrated to obtain the crude product. The crude product was purified by reverse-phase chromatography (using acetonitrile and water as the mobile phase, with acetonitrile ranging from 5.0% to 95%) to obtain 1.6 g of compound (Z)-(3-((2,3-bis(tert-butyloxycarbonyl)guanidinyl)methyl)-5-iodobenzoyl)-L-valine 10-b, with a yield of 82.1%. LCMS calculation for C 24 H 36 IN4O7 (M+H) + m / z = 619.2, found: 619.2

[0308] Step 3: At room temperature, compound 10-b (1.6 g, 2.58 mmol) and L-methyl methyl sulfate (462 mg, 2.84 mmol) were dissolved in 20 mL of anhydrous DCM, and EEDQ (767 mg, 3.11 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS. The mixture was diluted with water (30 mL), extracted three times with DCM (40 mL x 3), and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluted with PE in EA from 0% to 50%) to give compound (3-(((Z)-2,3-bis(tert-butyloxycarbonyl)guanidine)methyl)-5-iodobenzoyl)-L-valine-L-methionine methyl ester 10-c 1.4 g, as a yellow solid, with a yield of 71%. LCMS calculation for C 30 H 47 IN5O8S (M+H)+ m / z = 764.2, found: 764.2.

[0309] Step 4: Under ice bath conditions, compound 10-c (1.4 g, 1.8 mmol) was dissolved in 15 mL of THF, followed by the addition of Lithium Hydroxide (129 mg, 5.4 mmol) dissolved in 6 mL of Water. The reaction was then carried out at room temperature for 4 h. LCMS was used to monitor the reaction completion. The system was adjusted to pH 6 with 1 M HCl, and the mixture was concentrated to obtain the crude product. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to obtain 1.1 g of compound (3-(((Z)-2,3-bis(tert-butyloxycarbonyl)guanidinyl)methyl)-5-iodobenzoyl)-L-valine-L-methionine 10-d, as a yellow solid, with a yield of 80.3%. LCMS calculation for C 29 H 45 IN5O8S (M+H) + m / z = 750.2, found: 750.2

[0310] Step 5: At room temperature, compound 10-d (1.1 g, 1.47 mmol) and N6-(((9H-fluorene-9-yl)methoxy)carbonyl)-L-lysine tert-butyl ester 10-e (684 mg, 1.61 mmol) were dissolved in 10 mL of anhydrous DCM, and EEDQ (435 mg, 1.76 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The mixture was diluted with 30 mL of water and 40 mL of DCM was added. 3) Extracted three times, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to obtain compound N6-(((9H-fluorene-9-yl)methoxy)carbonyl)-N2-(3-((((E)-2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzoyl)-L-valine-L-methionine-L-lysine tert-butyl ester 10-f 850 mg, as a yellow solid, yield 50%. LCMS calculation for C 54 H 75 IN7O 11 S (M+H)+ m / z = 1156.4, found:1156.5.

[0311] Step 6: At room temperature, compound 10-f (850 mg, 0.74 mmol) was dissolved in 8 mL of DCM, and 8 mL of diethylamine was added. The reaction solution was reacted under argon protection for 3 hours. The reaction was monitored by LCMS to ensure completion. The reaction solution was concentrated, and the crude product was purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to give 10-g (450 mg) of tert-butyl(3-((((E)-2,3-bis(tert-butyloxycarbonyl)guanidin)methyl)-5-iodobenzoyl)-L-valine-L-methoxy-L-lysine as a yellow solid, with a yield of 65.6%. LCMS calculation for C 39 H 65 IN7O9S (M+H) + m / z = 934.4, found: 934.3.

[0312] Step 7: At room temperature, 10-g (450 mg, 0.48 mmol) of compound 1, 294 mg (0.53 mmol) of 18-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-15,18-dioxo-4,7,10-trioxa-14-earoctadecanoic acid 09-i) and TCFH (201 mg, 0.72 mmol) were dissolved in 5 mL of acetonitrile, and NMI (114 μL, 1.44 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated to obtain a crude product, which was then purified by reverse-phase chromatography (eluted with CH3CN in H2O from 5.0% to 95%) to obtain compound (S)-24-((S)-2-(S)-2--(3-(((Z)-2,3-bis(tert-butoxycarbonyl)guanidine)methyl)-5-iodobenzamido)-3-methylbutamido)-4-(methylthio)butamido-1-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-1,4,18-trioxo-9,12,15-trioxy-5,19-diazapentazolidine-25-carboxylic acid tert-butyl ester 10-i 300 mg, yield 42%. LCMS calculation for C 62 H 95 IN 11 O 18 S2 (M+H) + m / z = 1472.5, found: 1472.5.

[0313] Step 8: At room temperature, 10-i (100 mg, 0.07 mmol) was dissolved in 2 mL of DCM, and 2 mL of TFA was added. The mixture was stirred at room temperature for 2 h. The reaction was monitored by LCMS to indicate completion. The system was concentrated to obtain a crude product, which was then purified by high performance liquid chromatography (0.1% FA) to obtain 10-30 mg of compound (S)-24-((S)-2-((S)-2-(3-(guanidinyl)-5-iodobenzoylamino)-3-methylbutyramide)-4-(methylthio)butyramide)-1-((4-(5-(methylsulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-1,4,18-trioxo-9,12,15-trioxy-5,19-diazapentane-25 carboxylic acid compound.

[0314] LCMS calcld for C 48 H 71 IN 11 O 14 S2 (M+H) + m / z = 1216.4, found:1216.4;

[0315] 1 H NMR (400 MHz, DMSO- d 6 ) δ 10.88 (s, 1H), 10.45 (s, 1H), 8.38 (d, J =9.6 Hz, 2H), 8.28 (d, J = 8.5 Hz, 1H), 8.04 (d, J = 8.9 Hz, 3H), 7.97 (s,2H), 7.92-7.82 (m, 3H), 7.77 (t, J = 5.3 Hz, 1H), 7.29 (d, J = 5.9 Hz, 2H),7.24-7.08 (m, 1H), 5.75 (s, 1H), 4.49-4.39 (m, 1H), 4.31-4.24 (m, 1H), 4.22-4.15 (m, 2H), 3.82-3.75 (m, 2H), 3.69 (s, 3H), 3.57 (t, J = 6.5 Hz, 4H),3.12-3.06 (m, 4H), 3.00-2.92 (m, 3H), 2.61 (t, J = 6.9 Hz, 4H), 2.42 (t,J =7.0 Hz, 4H), 2.34-2.22 (m, 4H), 2.03 (s, 3H), 1.95-1.84 (m, 2H), 1.77-1.64(m, 2H), 1.63-1.50 (m, 4H), 1.37-1.25 (m, 3H), 0.91 (dd, J = 6.5, 3.3 Hz, 6H).

[0316] Example 10

[0317] Synthesis of tert-butyl(S)-2-((S)-2-((S)-2-(3-(((E)-2,3-bis(tert-butoxycarbonyl)guanidinyl)methyl)-5-(trimethylsteryl)benzamido)-4-(methylthio)butamido)-3-methylbutamido)-22-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)--5,19,22-trioxo-8,11,14-trioxa-4,18-diazaeicosanoate (compound Sn-10)

[0318] Step 1: Methyl 3-(aminomethyl)-5-iodobenzoate Sn-10-a (1 g, 3.44 mmol) was dissolved in 10 mL of anhydrous toluene. Pd(PPh3)4 (400 mg, 0.34 mmol) and hexamethyldistin (1.69 g, 5.16 mmol) were added, and the mixture was microwaved at 130 °C for 20 min. After the reaction was monitored by LCMS, the insoluble matter was filtered off, and the crude product was obtained by rotary evaporation. The crude product was purified by silica gel chromatography to obtain methyl 3-(aminomethyl)-5-(trimethylmethylenetinyl)benzoate Sn-10-b 1 g, with a yield of 88%. LCMS calcld for C 12 H 20 NO2Sn (M+H) + m / z = 327.0, found: 327.1.

[0319] Step 2: Methyl 3-(aminomethyl)-5-(trimethyltinyl)benzoate Sn-10-b (1 g, 3.05 mmol) was dissolved in 10 mL of anhydrous DCM. N-[(1E)-{[(tert-butoxy)carbonyl]imino}(trifluoromethanesulfonyl)methyl]carbamate tert-butyl ester (1.43 g, 3.66 mmol) and triethylamine (1.27 mL, 9.15 mmol) were added. The mixture was stirred for 2 h under argon protection. After the reaction was complete as monitored by LCMS, water (30 mL) was added to the reaction solution for extraction, followed by washing with brine (30 mL). The organic phase was dried over Na₂SO₄, filtered, and evaporated to dryness to obtain the crude product. The crude product was purified by silica gel chromatography (eluted with PE in EA from 0% to 100%) to give methyl (Z)-3-((2,3-bis(tert-butoxycarbonyl)guanidinyl)methyl)-5-(trimethylmethylenetinyl)benzoate Sn-10-c 1 g, yield 57%. LCMS calculation for C 23 H 38 N3O6Sn (M+H) + m / z = 572.2, found: 572.3.

[0320] Step 3: At room temperature, methyl (Z)-3-((2,3-bis(tert-butoxycarbonyl)guanidinyl)methyl)-5-(trimethylsilyl)benzoate (1 g, 1.7 mmol) was dissolved in a mixture of 6 mL THF and 2 mL H2O, and LiOH (310 mg, 13 mmol) was added. The reaction mixture was stirred at room temperature for 16 h under nitrogen protection. The reaction was monitored by LCMS until completion. The mixture was extracted with EA (50 mL) and water (30 mL), and then washed with brine (30 mL). The organic phase was dried over Na2SO4, filtered, and evaporated to dryness to obtain the crude product. The crude product was purified by reversed-phase chromatography (eluted with ACN in H2O from 0% to 100%) to obtain a white solid product (Z)-3-((2,3-bis(tert-butoxycarbonyl)guanidinyl)methyl)-5-(trimethylmethylenetinyl)benzoic acid Sn-10-d 500 mg, yield 51%. LCMS calculation for C 22 H 36 N3O6Sn (M+H) + m / z = 557.2, found: 557.3.

[0321] Step 4: At room temperature, L-valine (10 g, 67 mmol) was dissolved in a mixture of 100 mL THF and 20 mL H2O, followed by the addition of NaHCO3 (11.2 g, 134 mmol) and allyl chloroformate (12 g, 100 mmol). The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. After the reaction was completed as monitored by LCMS, it was diluted with 50 mL of water, and DCM (200 mL) was added. 3) Extraction was performed three times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by reversed-phase chromatography (ELECTRIC) to obtain 8 g of colorless oily ((allyloxy)carbonyl)-L-valine Sn-10-f, with a yield of 51%. 16 NO4S (M+H) + m / z = 234.1, found: 234.1.

[0322] Step 5: At room temperature, ((allyloxy)carbonyl)-L-valine Sn-10-f (8 g, 34 mmol) and L-methionine tert-butyl hydrochloride (7.1 g, 41 mmol) were dissolved in 50 mL of anhydrous DCM, and EEDQ (12.6 g, 51 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS until completion. The mixture was diluted with 50 mL of water, and 80 mL of DCM was added. 3) Extraction was performed three times. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluted with PE in EA from 0% to 30%) to obtain tert-butyl((allyloxy)carbonyl)-L-valine-L-methionine ester Sn-10-g 6 g, as a colorless oil, with a yield of 46%. LCMS calculation for C 18 H 33 N₂O₅S (M+H) + m / z = 389.2, found: 389.3.

[0323] Step 6: Dissolve tert-butyl((allyloxy)carbonyl)-L-valine-L-methionine ester Sn-10-g (6 g, 15 mmol) in 30 mL of TFA. The reaction mixture was stirred at 50 °C for 16 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction mixture was concentrated to obtain a crude product, which was then purified by reversed-phase chromatography to give 2 g of compound ((allyloxy)carbonyl)-L-valine-L-methionine Sn-10-h, with a yield of 39%. LCMS calculation for C 14 H 25 N₂O₅S (M+H) + m / z = 333.1, found: 333.1.

[0324] Step 7: At room temperature, ((allyloxy)carbonyl)-L-valine-L-methionine Sn-10-h (2 g, 6 mmol) and (S)-3-(((((9H-fluorene-9-yl)methoxy)carbonyl)amino)-2-aminopropionate tert-butyl ester (2.7 g, 7.2 mmol) were dissolved in 30 mL of anhydrous DCM, and EEDQ (2.2 g, 9 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The mixture was diluted with 50 mL of water and 60 mL of DCM was added. 3) Extracted three times, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluted with PE in EA from 0% to 100%) to obtain 2 g of compound (2S,5S,8S)-2-(((((9H-fluorene-9-yl)methoxy)carbonyl)amino)methyl)-5-(2-(methylthio)ethyl)-8-isopropyl-4,7,10-trioxo-11-oxa-3,6,9-triazatetradec-13-enoic acid tert-butyl ester Sn-10-i, as a colorless oil, with a yield of 48%. LCMS calculation for C 36 H 49 N4O8S (M+H) + m / z = 697.3, found: 697.3.

[0325] Step 8: At room temperature, Sn-10-i (500 mg, 0.72 mmol) was dissolved in 5 mL of chloroform, and Pd(PPh3)4 (81 mg, 0.07 mmol) and PhSiH3 (90 μL, 0.72 mmol) were added. The reaction solution was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS until completion. The insoluble matter in the reaction solution was filtered off, and the crude product was concentrated. The crude product was purified by reversed-phase chromatography to obtain 250 mg of tert-butyl(S)-3-((((9h-fluorene-9-yl(methoxy)carbonyl)amino)-2-((S)-2-((S)-2-amino-3-methylbutanamide)-4-(methylthio)butanamide)propionate Sn-10-j, as a white solid, with a yield of 57%. LCMS calculation for C 32 H 45 N4O6S (M+H) + m / z = 613.3, found: 613.4.

[0326] Step 9: At room temperature, tert-butyl(S)-3-((((9h-fluorene-9-yl(methoxy)carbonyl)amino)-2-((S)-2-((S)-2-amino-3-methylbutanamide)-4-(methylthio)butanamide)propionate Sn-10-j (100 mg, 0.16 mmol), (Z)-3-((2,3-bis(tert-butoxycarbonyl)guanidinyl)methyl)-5-(trimethylmethylenetinyl)benzoic acid Sn-10-d (110 mg, 0.20 mmol), and TCFH (68 mg, 0.24 mmol) were dissolved in 2 mL of acetonitrile, and NMI (26 μL, 0.32 mmol) was added. The reaction solution was stirred at room temperature for 1 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated to obtain the crude product, which was then passed through a reverse-phase chromatography column (eluted with CH3CN in H2O from The compound tert-butyl(S)-3-((((9h-fluorene-9-yl(methoxy)carbonyl)amino)-2-((S)-2-((S)-2-(3-(((E)-2,3-bis(tert-butylcarbonyl)guanidine)methyl)-5-(trimethyltinyl)benzamide)-3-methylbutyramide)-4-(methylthio)butyramide)propionate Sn-10-k 50 mg was isolated and purified to give Sn-10-k 50 mg as a white solid in 27% yield. LCMS calcld for C 54 H 78 N7O 11 SSn (M+H) + m / z = 1152.4, found: 1152.9.

[0327] Step 10: At room temperature, tert-butyl(S)-3-((((9h-fluorene-9-yl(methoxy)carbonyl)amino)-2-((S)-2-((S)-2-(3-(((E)-2,3-bis(tert-butylcarbonyl)guanidine)methyl)-5-(trimethyltinyl)benzamide)-3-methylbutyramide)-4-(methylthio)butyramide)propionate Sn-10-k (50 mg, 0.04 mmol) was dissolved in 2 mL DCM, and 2 mL diethylamine was added. The reaction solution was reacted under argon protection for 2 hours. The reaction was monitored by LCMS to ensure completion. The reaction solution was concentrated, and the crude product was subjected to silica gel column chromatography (eluted with MeOH in DCM from 0% to...). 30%) The compound (S)-3-amino-2-((S)-2-((S)-2-(3-(((E)-2,3-bis(tert-butoxycarbonyl)guanidinyl)methyl)-5-(trimethylsteryl)benzamido)-4-(methylthio)butamido)-3-methylbutamido)tert-butyl propionate Sn-10-l 25 mg was isolated and purified as a pale yellow oil with a yield of 63%. LCMS calculation for C 39 H 68 N7O9SSn (M+H) + m / z = 930.4, found: 930.3.

[0328] Step 11: At room temperature, tert-butyl(S)-3-amino-2-((S)-2-(S)-2-(3)-((E)-2,3-bis(tert-butylcarbonyl)guanidinyl)methyl)-5-(trimethyltinyl)benzamide)-3-methylbutyramide)-4-(methylthio)butyramide)propane ester Sn-10-l (25 mg, 0.03 mmol), 18-((4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)amino)-15,18-dioxo-4,7,10-trioxa-14-azaoctadecanoic acid 09-i (22 mg, 0.04 mmol) and TCFH (17 mg, 0.06 mmol) were dissolved in 2 mL of acetonitrile, and NMI (10 μL, 0.12 mmol) was added. The reaction solution was stirred at room temperature for 1 h under nitrogen protection. The reaction was monitored for completion by LCMS. The reaction solution was concentrated to obtain a crude product, which was then purified by preparative chromatography (mobile phase: acetonitrile and 0.1% ammonium bicarbonate aqueous solution, with the acetonitrile ratio increasing from 5% to 100% within 20 minutes) to obtain 6 mg of compound Sn-10, with a yield of 15%.

[0329] LCMS calcld for C 62 H 98 N11 O 18 S2Sn (M+H) + m / z = 1468.6, found: 1468.7;

[0330] 1 H NMR (400 MHz, DMSO- d 6) δ 11.29 (s, 1H), 10.18 (s, 1H), 8.54 (t, J =5.7 Hz, 1H), 8.40-8.32 (m, 1H), 8.08-8.00 (t, J = 7.0 Hz, 1H), 7.81 (d, J =8.8 Hz, 2H), 7.63 (d, J = 8.9 Hz, 4H), 7.50-7.41 (m, 2H), 4.43-4.28 (m, 3H), 4.08-3.93 (m, 2H), 3.47 (s, 3H), 3.38-3.32 (m, 2H), 3.30-3.19 (m, 10H), 3.19-3.15 (m, 5H), 2.91-2.81 (m, 2H), 2.41-2.36 (m, 2H), 2.23-2.17 (m, 2H), 2.11-2.02 (m, 2H), 1.84-1.82 (m, 3H), 1.80-1.73 (m, 2H), 1.53 (s, 1H), 1.42-1.36(m, 2H), 1.25 (s, 9H), 1.18-1.11 (m, 18H), 1.01 (s, 1H), 0.66-0.58 (m, 6H), 0.14-0.01 (m, 9H).

[0331] Example 11

[0332] Synthetic N 1 -(1-(3-iodophenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N 4 Synthesis of 4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)butyramide (compound 11)

[0333] Step 1: At room temperature, HATU (59.3 mg, 0.16 mmol) was added to a solution of 3-[(4-(5-methylsulfonyl-1,3,4-oxadiazol-2-yl)phenyl)carbamoyl]propionic acid 05-I (48.5 mg, 0.14 mmol) in N,N-dimethylformamide (2 mL). The reaction solution was reacted at room temperature for 30 min under argon protection. Then, 3-(2-(2-(3-aminopropoxy)ethoxy)ethoxy)propylamine (30.8 mg, 0.14 mmol) and diisopropylethylamine (50.40 mg, 0.39 mmol) were added to the above solution, and the reaction solution was reacted at room temperature for 16 hours under argon protection. The reaction was monitored by LCMS to ensure completion. The reaction solution was diluted with water (10 mL) and ethyl acetate (5 mL) was added. 3) Extract three times, combine the organic phases, and rinse with saturated saline (5 mL). 4) After washing, the mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by Prep-TLC (PE / EtOAc = 1 / 2) to obtain compound N. 1 -(3-(2-(2-(3-aminopropoxy)ethoxy)ethoxy)propyl)-N 4 -(4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)succinamide 11-a 34 mg, yield 45%. LCMS calcld for C 23 H 35 N5O8S(M+H) + m / z = 542.22, found: 542.2.

[0334] Step 2: At room temperature, compound 11-a (200 mg, 0.37 mmol), m-iodophenylacetic acid (116 mg, 0.44 mmol), and TCFH (154 mg, 0.55 mmol) were dissolved in 4 mL of acetonitrile, and NMI (100 µL, 1.2 mmol) was added. The reaction solution was stirred at room temperature for 1 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated to obtain a crude product, which was then purified by high performance liquid chromatography (0.1% NH4HCO3) to obtain product N. 1 -(1-(3-iodophenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N 4 -(4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)butadiamide 11 25 mg.

[0335] LCMS calcld for C 31 H41 IN5O9S (M+H) + m / z = 786.2, found: 786.3.

[0336] 1 H NMR (400 MHz, DMSO- d 6 ) δ 10.40 (s, 1H), 8.09-7.99 (m, 3H), 7.88-7.81 (m, 3H), 7.66-7.55 (m, 2H), 7.26 (d, J = 7.8 Hz, 1H), 7.10 (t, J = 7.6Hz, 1H), 3.69 (s, 3H), 3.52-3.48 (m, 4H), 3.47-3.44 (m, 4H), 3.40-3.36 (m,6H), 3.12-3.05 (m, 4H), 2.61 (t, J = 7.0 Hz, 2H), 2.42 (t, J = 7.0 Hz, 2H),1.64-1.57 (m, 4H).

[0337] Example 12

[0338] N 1 -(4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)-N 4 Synthesis of 2-oxo-1-(3-(trimethylsteryl)phenyl)-7,10,13-trioxa-3-azahexadecane-16-yl)succinamide (compound Sn-11)

[0339] Step 1: Dissolve m-iodophenylacetic acid (500 mg, 1.91 mmol) in 10 mL of anhydrous toluene, add Pd(PPh3)4 (220 mg, 0.19 mmol) and hexamethyldistin (0.59 mL, 2.86 mmol), and incubate overnight at 90 °C. After the reaction is complete as monitored by LCMS, filter off the insoluble matter and evaporate to dryness to obtain the crude product. The crude product is purified by silica gel chromatography (mobile phase: PE:EA = 10:1) to give compound 2-(3-(trimethylsilyl)phenyl)acetic acid Sn-11-a 300 mg, yield 53%. 1H NMR (400 MHz, DMSO-d6) δ 7.33-7.10 (m, 4H), 3.42-3.36 (m, 2H), 0.41-0.23 (m, 9H).

[0340] Step 2: At room temperature, di-tert-butyl((S)-6-((S)-2-(2-(2-(2-aminoethoxy)ethoxy)acetamyl)-N-(2-(2-(tert-butoxy)-2-oxoethyl)-3-(4-iodophenyl)propamido)-1-(tert-butoxy)-1-oxohexyl-2-yl)carbamoyl)-L-glutamic acid 11-a (80 mg, 0.15 mmol), 2-(3-(trimethylsilyl)phenyl)acetic acid Sn-11-a (54 mg, 0.18 mmol), and TCFH (63 mg, 0.23 mmol) were dissolved in 2 mL of acetonitrile, and NMI (24 μL, 0.3 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated to obtain a crude product, which was then purified by high-performance liquid chromatography (0.1% NH4HCO3) to obtain N. 1 -(4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)-N 4 -(2-oxo-1-(3-(trimethylsteryl)phenyl)-7,10,13-trioxa-3-azahexadecane-16-yl)succinamide Sn-11 6 mg, yield 5%.

[0341] LCMS calcld for C 34 H 50 N5O9SSn (M+H) + m / z = 824.2, found: 823.9.

[0342] Example 13 Synthesis of Compound 12

[0343] Step 1: Under an argon atmosphere at room temperature, DIEA (0.25 g) was added to a DMF (5 mL) solution of compound 12-a (500 mg, 0.96 mmol) and PyBOP (0.65 g, 1.25 mmol). After stirring at room temperature for 10 min, compound 01-e (0.42 g, 0.96 mmol) was added. The mixture was stirred at room temperature for 1 h. The reaction solution was concentrated, and the crude product was purified by silica gel column chromatography (EA / PE: 1 / 3) to obtain compound 12-b 528 mg. LCMS (ESI) m / z = 966.2 [M+H]+ .

[0344] Step 2: Under an argon atmosphere at room temperature, compound 12-b (500 mg, 0.51 mmol) was dissolved in a mixed solvent of DCM (3 mL) and DEA (1 mL) and stirred overnight at room temperature. The solution was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (MeOH / DCM: 1 / 10) to give compound 12-c 243 mg, yield 62.9%. LCMS (ESI) m / z = 722.2 [M+H] + .

[0345] Step 3: 0 o At C, under an argon atmosphere, a solution of 2-bromoacetyl chloride (0.076 g, 0.48 mmol) in acetone was slowly added dropwise to an aqueous solution of compound 12-c (230 mg, 0.32 mmol). o The reaction was carried out at C for 1 h. Water (30 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 2). The organic phases were combined, dried, and concentrated. The residue was purified by high performance liquid chromatography (0.1% trifluoroacetic acid) to give compound 12-d 110 mg as a colorless oil, with a yield of 40.9%.

[0346] Step 4: Compound 12-d (110 mg, 0.13 mmol) was dissolved in TFA / DCM (2 mL / 2 mL) at room temperature and stirred for 6 h. The reaction was monitored by LCMS until completion. The residue was purified by high performance liquid chromatography (0.1% trifluoroacetic acid) to obtain compound N-(1-bromo-2-oxo-7,10,13-trioxo-3-azaoctaoctaanoic-16-yl)-3-(guanidinylmethyl)-5-iodobenzamide 12-13 mg.

[0347] LCMS (ESI) m / z= 642.0; 644.0 [M+H] + ;

[0348] 1 H NMR (400 MHz, CD3OD) δ 8.11 (s, 1H), 7.86 (s, 1H), 7.77 (s, 1H), 4.43 (s, 2H), 3.66-3.53 (m, 12H), 3.50-3.44 (m, 4H), 3.30-3.25 (m, 2H), 1.88-1.25 (m, 2H), 1.76-1.72 (m, 2H).

[0349] Example 14

[0350] Synthesis of (3-(17-(2,5-dioxy-2,5-dihydro-1h-pyrrolo-1-yl)-15-oxy-4,7,10-trioxy-14-azaheptyl)carbamoyl)-5-(guanidinomethyl)phenyl)boronic acid (compound B(OH)2-13).

[0351]

[0352] Step 1: Under an argon atmosphere, at room temperature, DIEA (0.24 g, 1.86 mmol) was added to a DCM (5 mL) solution of compound B(OH)₂-13-a (0.2 g, 0.62 mmol), compound B(OH)₂-13-f (0.10 g, 0.62 mmol%), and PyBOP (0.48 g, 0.93 mmol). The mixture was stirred at room temperature for 1 h. The solution was concentrated under reduced pressure, and the residue was purified by column chromatography (EA / PE: 1 / 3) to give compound B(OH)₂-13-b 0.18 g. LCMS (ESI) m / z = 472.2. [M+H] +

[0353] Step 2: Under an argon atmosphere, compound B(OH)₂-13-b (180 mg, 0.38 mmol) was dissolved in TFA / DCM (2 mL / 2 mL) and stirred at room temperature for 2 h. The solution was concentrated under reduced pressure to obtain 190 mg of crude compound B(OH)₂-13-c, which was used directly in the next reaction. LCMS (ESI) m / z = 371.5. [M+H] +

[0354] Step 3: Under an argon atmosphere, DIEA (0.081 g, 0.63 mmol) was added to a DCM (1 mL) solution of crude compound B(OH)2-13-c (90 mg), B(OH)2-13-d (78 mg, 0.21 mmol), and PyBOP (0.14 g, 0.27 mmol). The reaction was allowed to proceed at room temperature for 2 h. The mixture was concentrated under reduced pressure, and the crude product was purified by preparative HPCL to obtain 30 mg of compound B(OH)2-13-e, with a yield of 18.3%. LCMS (ESI) m / z = 691.4. [M+H-Boc] +

[0355] Step 4: Under an argon atmosphere at room temperature, compound B(OH)2-13-e (30 mg, 0.038 mmol) was dissolved in DCM (1 mL), and TFA (1 mL) was added. The mixture was stirred at room temperature for 2 h. The reaction was monitored by LCMS. The reaction solution was concentrated, and the residue was purified by high performance liquid chromatography (0.1% trifluoroacetic acid) to obtain compound (3-(17-(2,5-dioxy-2,5-dihydro-1h-pyrrolo-1-yl)-15-oxy-4,7,10-trioxy-14-azaheptyl)carbamoyl)-5-(guanidinomethyl)phenyl)boronic acid B(OH)2-13 8 mg, yield 31%.

[0356] LCMS(ESI)m / z = 591.2 [M+H] + .

[0357] Example 15

[0358] Synthesis of N-(17-(2,5-dioxy-2,5-dihydro-1h-pyrrolo-1-yl)-15-oxy-4,7,10-trioxy-14-azaheptyl)-3-(guanidinylmethyl)-5-iodobenzamide (compound 13)

[0359] Compound B(OH)₂-13 (3 mg, 0.005 mmol) was dissolved in 0.5 mL of acetonitrile under an argon atmosphere at room temperature. 0.5 mL of 7% NaHCO₃ aqueous solution, NBS (2.65 mg, 0.015 mmol), and NaI (1 mg, 0.0065 mmol) were added. The reaction mixture was stirred at room temperature for 30 minutes. The reaction was monitored by LCMS. The reaction mixture was concentrated, and the residue was purified by high-performance liquid chromatography (0.1% trifluoroacetic acid) to obtain 1 mg of compound B(OH)₂-13. LCMS (ESI) m / z = 673.2 [M+H] + .

[0360] Example 16

[0361] Synthesis of N-(1-(4-(3-(2,5-dioxo-2,5-dihydro-1H-pyrrolo-1-yl)propamido)phenyl)-1,5-dioxo-10,13,16-trioxa-2,6-diazadodecane-19-yl)-3-(guanidinomethyl)-5-iodobenzamide (compound 13-Ph)

[0362] Step 1: Under nitrogen protection, 13-Ph-a (200 mg, 0.39 mmol), O1-e (202.5 mg, 0.39 mmol), TCFH (218.8 mg, 0.78 mmol), and N-methylimidazole (128.0 mg, 1.56 mmol) were added sequentially to acetonitrile and stirred at room temperature for 30 min. The reaction solution was directly concentrated and purified by normal-phase silica gel column chromatography (PE:EtOAc = 6:1). Compound 13-Ph-b 200 mg was obtained as a colorless oil, with a yield of 50.6%. LCMS (ESI) m / z = 1015.0 [M+H] + .

[0363] Step 2: Under nitrogen protection, 13-Ph-b (200 mg, 0.20 mmol) was added to a mixed solvent of DCM (1 mL) and ethylenediamine (1 mL), and stirred at room temperature for 0.5 h. After concentration, the crude product was purified by reversed-phase silica gel column chromatography (mobile phase: acetonitrile and water, acetonitrile from 5.0% to 50%) to obtain 80 mg of compound 13-Ph-c as a colorless oil, with a yield of 51.21%. LCMS (ESI) m / z = 793.3 [M+H] + .

[0364] Step 3: Under nitrogen protection, 13-Ph-c (55 mg, 0.069 mmol), 13-Ph-d (23.87 mg, 0.083 mmol), TCFH (31.48 mg, 0.083 mmol), and N-methylimidazole (11.5 mg, 0.14 mmol) were added sequentially to DCM (1 mL), and stirred at room temperature for 30 min. The reaction solution was purified by reversed-phase chromatography (mobile phase: acetonitrile and water, with the acetonitrile ratio increasing from 5.0% to 55% over 20 minutes) to obtain 20 mg of 13-Ph-e as a yellow oil, with a yield of 27.12%. LCMS (ESI) m / z = 1063.3 [M+H] + .

[0365] Step 4: Under nitrogen protection, 13-Ph-e (20 mg, 0.019 mmol) was placed in a single-necked flask, and DCM (0.2 mL) and TFA (0.2 mL) were added. The mixture was then reacted at room temperature for 1 hour. The reaction solution was concentrated and purified by prep-HPLC (mobile phase: acetonitrile and 0.1% TFA aqueous solution, with the acetonitrile concentration increasing from 5.0% to 55% over 20 minutes). The product 13-Ph was obtained as 5.65 mg of white solid with a purity of 99.3%, yielding 34.56%.

[0366] LCMS(ESI)m / z=863.1 [M+H] + ;

[0367] 1 H NMR (400 MHz, CD3OD) δ 8.10 (s, 1H), 7.84 (s, 1H), 7.78-7.73 (m,3H), 7.60 (d, J = 8.7 Hz, 2H), 6.81 (s, 2H), 4.44 (s, 2H), 3.86 (t, J = 7.0Hz, 2H), 3.66-3.57 (m, 10H), 3.50-3.41 (m, 6H), 3.23 (t, J = 6.8 Hz, 2H), 2.68 (t, J = 7.2 Hz, 2H), 2.49 (t, J = 6.8 Hz, 2H), 1.86 (p, J = 6.4 Hz, 2H), 1.70 (p, J = 6.4 Hz, 2H).

[0368] Example 17

[0369] Synthesis of N1-(1-(3-iodo-5-methoxyphenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N4-(4-(5-(methylsulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)succinamide (compound 14)

[0370] Step 1: Under an argon atmosphere, DIEA (129 mg, 1 mmol) and 14-d-1 (106 mg, 0.51 mmol) were added to a DMF (1 mL) solution of succinic acid (60 mg, 0.51 mmol) and HATU (232 mg, 0.61 mmol), and the mixture was stirred at room temperature for 1 h. The reaction solution was concentrated, and the crude product was purified by column chromatography to give 97 mg of compound 14-d-2. LCMS (ESI) m / z = 308.1 [M+H] + .

[0371] Step 2: At room temperature and under an argon atmosphere, DIEA (77 mg, 0.60 mmol) and 14-d-3 (64 mg, 0.29 mmol) were added to a DMF (1 mL) solution of 14-d-2 (90 mg, 0.29 mmol) and HATU (132 mg, 0.35 mmol), and the mixture was stirred at room temperature for 1 h. The reaction solution was concentrated, and the crude product was purified by column chromatography to give 63 mg of compound 14-d. LCMS (ESI) m / z = 510.2 [M+H] + .

[0372] Step 3: At room temperature and under an argon atmosphere, add CuI (0.11 g, 0.55 mmol) to a solution of 1,4-dioxane (10 mL) containing 14-a (4.14 g, 27.65 mmol) and N,N'-dimethyl-1,2-ethylenediamine (97 mg, 1.11 mmol). Seal the tube and heat to 110°C. o The reaction was carried out overnight at C. The mixture was cooled to room temperature, diluted with 30 mL of water, extracted with ethyl acetate (30 mL x 2), the organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the residue was purified by column chromatography to give compound 14-b 1.5 g as a colorless oil. 1 H NMR (400 MHz, CD3OD) δ 7.33-7.27 (m,1H), 7.26-7.21 (m, 1H), 6.91 (s, 1H), 3.85 (s, 2H), 3.79 (s, 3H).

[0373] Step 4: At room temperature and under an argon atmosphere, dissolve 14-b (1 g, 3.66 mmol) in a mixed solution of sodium hydroxide (10 mmol, 2 mol / L aqueous solution) and EtOH (5 mL). Seal the tube and heat to 95°C. oIncubate overnight at room temperature. Neutralize to pH 3 with 1N HCl (25 mL) and stir at room temperature for 2 h. Filter and collect the solid as the target product 14-c 600 mg. LCMS (ESI) m / z = 583.0 [2M-H] - .

[0374] Step 5: At room temperature and under an argon atmosphere, DIEA (0.071 g, 0.55 mmol) and 14-d (0.11 g, 0.21 mmol) were added to a DMF (1 mL) solution of 14-c (60 mg, 0.21 mmol) and HATU (0.10 g, 0.27 mmol), and the mixture was stirred at room temperature for 1 h. The reaction solution was concentrated, and the crude product was purified by column chromatography to give 30 mg of compound 14-e. LCMS (ESI) m / z = 784.1 [M+H] + .

[0375] Step 6: At room temperature and under an argon atmosphere, mCPBA (0.11 g, 0.66 mmol) was added to a DCM (3 mL) solution of 14-e (30 mg, 0.039 mmol), and the reaction was carried out overnight in the dark. The mixture was filtered, the filtrate was concentrated, and the crude product was purified by preparative HPLC to give 14-e 3 mg.

[0376] LCMS (ESI) m / z = 816.0 [M+H] + .

[0377] Example 18

[0378] N 1 -(1-(3-methoxy-5-(trimethylstanyl)phenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N 4 Synthesis of 4-(5-(methanesulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)butyramide (compound Sn-14)

[0379] Compound 11-a (120 mg, 0.22 mmol), compound Sn-14-a (89 mg, 0.27 mmol), and TCFH (93 mg, 0.33 mmol) were dissolved in 2 mL of acetonitrile at room temperature, and N-methylimidazole (53 μL, 0.66 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction mixture was concentrated to obtain a crude product, which was then purified by high-performance liquid chromatography (0.1% NH4HCO3) to obtain 18 mg of compound Sn-14, with a yield of 9.5%.

[0380] LCMS calcld for C 35 H 52 N5O 10 SSn (M+H) + m / z = 854.2, found: 854.2;

[0381] 1 H NMR (400 MHz, DMSO- d 6) δ 10.40 (s, 1H), 8.04 (d, J = 8.8 Hz, 2H), 7.99 (t, J = 5.4 Hz, 1H), 7.90-7.83 (m, 3H), 6.94-6.83 (m, 2H), 6.80-6.74 (m,1H), 3.73 (s, 3H), 3.70 (s, 3H), 3.54-3.42 (m, 9H), 3.42-3.34 (m, 5H), 3.13-3.03 (m, 4H), 2.61 (t, J = 7.0 Hz, 2H), 2.42 (t, J = 7.0 Hz, 2H), 1.66-1.57(m, 4H), 0.33-0.17 (m, 9H).

[0382] Example 19

[0383] Synthesis of N-(1-(3-iodo-5-methoxyphenyl)-2-oxo-6,9,12,15-tetraoxa-3-azaheptadecane-17-yl)-2-(methanesulfonyl)pyrimidine-5-carboxamide (compound 15)

[0384] Step 1: At room temperature, 15-a (1 g, 5.9 mmol) of compound 2-(methylthio)pyrimidine-5-carboxylic acid, tert-butyl (14-amino-3,6,9,12-tetraoxatetradecyl)carbamate (2.17 g, 6.4 mmol), and TCFH (2.5 g, 8.8 mmol) were dissolved in 10 mL of acetonitrile, and NMI (1.5 mL, 17.6 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction mixture was concentrated to obtain a crude product, which was purified by silica gel chromatography (EA:PE = 1:5) to give 1.1 g of compound 15-b as a white solid, with a yield of 38%. LCMS calculation for C 21 H 37 N4O7S (M+H) + m / z = 489.2, found: 489.2.

[0385] Step 2: At room temperature, compound (1-(2-(methylthio)pyrimidin-5-yl)-1-oxo-5,8,11,14-tetraoxa-2-azahexadecane-16-yl)carbamate tert-butyl ester 15-b (1 g, 2.04 mmol) was dissolved in 10 mL of anhydrous DCM, and 5 mL of TFA was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored for completion by LCMS. After the reaction was completed as monitored by LCMS, the reaction mixture was evaporated to dryness to obtain the crude product of compound 15-c. The crude product did not require purification and proceeded directly to the next step.

[0386] Step 3: At room temperature, N-(14-amino-3,6,9,12-tetraoxotetradecyl)-2-(methylthio)pyrimidine-5-carboxamide 15-c (200 mg, 0.51 mmol), 2-(3-iodo-5-methoxyphenyl)acetic acid 14-c (165 mg, 0.57 mmol), and TCFH (22 mg, 0.77 mmol) were dissolved in 2 mL of acetonitrile, and N-methylimidazole (41 µL, 1.54 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction mixture was concentrated to obtain a crude product, which was purified by silica gel chromatography (EA:PE = 1:5) to give compound 15-d 150 mg as a white solid, with a yield of 44%. LCMS calcld for C 25 H 36 IN4O7S (M+H) + m / z = 663.1, found: 663.2.

[0387] Step 4: At room temperature, compound N-(1-(3-iodo-5-methoxyphenyl)-2-oxo-6,9,12,15-tetraoxa-3-azaheptadecane-17-yl)-2-(methylthio)pyrimidine-5-carboxamide 15-d (50 mg, 0.075 mmol) was dissolved in 5 mL of anhydrous DCM, and mCPBA (130 mg, 0.75 mmol) was added. The reaction mixture was stirred at room temperature for 16 h under nitrogen protection. The reaction was monitored by LCMS to ensure completion. After the reaction was completed by LCMS, saturated NaHCO3 (10 mL) was added to quench the reaction mixture, and then it was washed with saturated brine (20 mL). The organic phase was dried over Na2SO4, filtered, and evaporated to dryness to obtain the crude product. The crude product was purified by preparative chromatography (0.1% NH4HCO3) to obtain compound 15 4.7 mg, with a yield of 10%.

[0388] LCMS calcld for C 25 H 36 IN4O9S (M+H) + m / z = 695.1, found: 695.1.

[0389] Example 20

[0390] Synthesis of N-(1-(3-methoxy-5-(trimethylmethylenetinyl)phenyl)-2-oxo-6,9,12,15-tetraoxa-3-azaheptadecane-17-yl)-2-(methanesulfonyl)pyrimidine-5-carboxamide (compound Sn-15)

[0391] Step 1: At room temperature, N-(14-amino-3,6,9,12-tetraoxotetradecyl)-2-(methylthio)pyrimidine-5-carboxamide 15-c (300 mg, 0.77 mmol), 2-(3-methoxy-5-(trimethylstanyl)phenyl)acetic acid Sn-14-a (278 mg, 0.93 mmol), and TCFH (324 mg, 1.16 mmol) were dissolved in 2 mL of acetonitrile, and N-methylimidazole (186 μL, 2.3 mmol) was added. The reaction mixture was stirred at room temperature for 2 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. The reaction solution was concentrated to obtain a crude product. The crude product was then purified by reversed-phase chromatography (mobile phase: acetonitrile and water; the acetonitrile percentage increased from 0% to 100% within 20 minutes) to obtain compound N-(1-(3-methoxy-5-(trimethylsilyl)phenyl)-2-oxo-6,9,12,15-tetraoxa-3-azaheptadecane-17-yl)-2-(methylthio)pyrimidine-5-carboxamide Sn-15-a 100 mg, as a white solid, with a yield of 18%. LCMS calculation for C 28 H 45 N4O8SSn (M+H) + m / z = 701.2, found:701.2.

[0392] Step 2: At room temperature, compound N-(1-(3-methoxy-5-(trimethylmethylenetinyl)phenyl)-2-oxo-6,9,12,15-tetraoxa-3-azaheptadecane-17-yl)-2-(methylthio)pyrimidine-5-carboxamide Sn-15-a (100 mg, 0.14 mmol) was dissolved in 5 mL of anhydrous DCM, and mCPBA (245 mg, 1.4 mmol) was added. The reaction mixture was stirred at room temperature for 16 h under nitrogen protection. The reaction was monitored by LCMS to indicate completion. After the reaction was completed by LCMS, saturated NaHCO3 (10 mL) was added to quench the reaction mixture, and then it was washed with saturated brine (20 mL). The organic phase was dried over Na2SO4, filtered, and evaporated to dryness to obtain the crude product. The crude product was purified by preparative chromatography (0.1% NH4HCO3) to give 10 mg of compound N-(1-(3-methoxy-5-(trimethylsilyl)phenyl)-2-oxo-6,9,12,15-tetraoxa-3-azaheptadecane-17-yl)-2-(methanesulfonyl)pyrimidine-5-carboxamide Sn-15, with a yield of 10%.

[0393] LCMS calcld for C 28 H 45 N4O9SSn (M+H) +m / z = 733.2, found: 733.0;

[0394] 1 H NMR (400 MHz, DMSO- d 6) δ 9.37 (s, 2H), 9.19-9.09 (m, 1H), 8.11-8.04(m, 1H), 6.92-6.90 (m, 1H), 6.88-6.85 (m, 1H), 6.79-6.76 (m, 1H), 3.73 (s,3H), 3.60-3.56 (m, 2H), 3.55-3.52 (m, 14H), 3.45 (s, 3H), 3.42-3.40 (m, 4H), 3.22-3.17 (m, 2H), 0.25 (s, 9H).

[0395] Example 21

[0396] Synthesis of N1-(1-(3-iodo-5-methoxyphenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N4-(4-(5-(methylsulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)succinamide (compound 16)

[0397] Step 1: Under an argon atmosphere, add AIBN (0.071 g, 0.43 mmol) to a 20 mL solution of compound 16-a (2.0 g, 8.62 mmol) and NBS (1.53 g, 8.62 mmol) in carbon tetrachloride, and heat to 85°C. o The reaction was carried out overnight at C. The mixture was cooled to room temperature and concentrated under reduced pressure. It was diluted with water (30 mL), extracted with ethyl acetate (30 mL x 2), the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the residue was purified by column chromatography to give 1.4 g of compound 16-b. 1 H NMR (400 MHz, CD3OD) δ 7.55 (s, 1H), 7.49-7.46 (m, 1H), 7.20 (s, 1H), 4.43 (s, 2H), 2.27 (s, 3H).

[0398] Step 2: 16-b (900 mg, 2.89 mmol) and 15-crown ether (0.64 g, 2.89 mmol) were dissolved in DMF (5 mL), and NaCN (0.28 g, 5.78 mmol) was added. The mixture was stirred overnight under an argon atmosphere. The solution was diluted with water (30 mL), extracted with ethyl acetate (30 mL x 2), and the organic phases were combined. The mixture was washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the residue was purified by column chromatography to give compound 16-c 0.52 g. LCMS (ESI) m / z = 280.3 [M+Na] + .

[0399] Step 3: Dissolve 16-c (0.42 g, 2.02 mmol) in NaOH (10 mmol, 2 mol / L aqueous solution) / EtOH (5 mL), and heat at 95 °C for 2 h under an argon atmosphere. Add 1 N HCl (25 mL) and stir at room temperature for 2 h. This yields compound 16-d (0.42 g), a pale yellow solid. LCMS (ESI) m / z = 293.0 [M+H] + .

[0400] Step 4: Under an argon atmosphere, N-methylimidazole (0.024 g, 0.29 mmol) was added to a 2 mL solution of 16-d (20 mg, 0.072 mmol), TCFH (0.040 g, 0.14 mmol), and compound 11-a (0.039 g, 0.072 mmol) in ACN. The mixture was stirred at room temperature for 0.5 h. The solution was concentrated under reduced pressure, and the crude product was purified by preparative HPLC to obtain 12 mg of compound 16.

[0401] LCMS (ESI) m / z = 800.0 [M+H] + ;

[0402] 1 H NMR (400 MHz, CD3OD) δ 8.11-7.97 (m, 2H), 7.85-7.80 (m, 2H), 7.44(d, J = 8.1 Hz, 2H), 7.09 (s, 1H), 3.62-3.58 (m, 4H), 3.56-3.52 (m, 6H), 3.51-3.44 (m, 4H), 3.38 (s, 2H), 3.29-3.22 (m, 5H), 2.71 (t, J= 6.8 Hz, 2H), 2.55 (t, J = 6.8 Hz, 2H), 2.26 (s, 3H), 1.78-1.69 (m, 4H).

[0403] Example 22

[0404] Synthesis of N1-(1-(3-methyl-5-(trimethyltinyl)phenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N4-(4-(5-(methylsulfonyl)-1,3,4-oxadiazol-2-yl)phenyl(5-(methylsulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)succinamide (compound Sn-16)

[0405] Step 1: At room temperature and under an argon atmosphere, compound 16-d (1 g, 3.44 mmol), tetra(triphenylphosphine)palladium (125.9 mg, 0.11 mmol), and hexamethyldistin (0.54 g, 1.64 mmol) were dissolved in toluene (10 mL), and the mixture was sealed and heated to 130 °C for 2 h. The reaction solution was concentrated, and the crude product was purified by silica gel column chromatography to give Sn-16-a 450 mg as a white solid. 1 H NMR (400 MHz, CDCl3) δ 7.24-7.05 (m, 3H), 3.62-3.59 (m, 2H), 2.34 (s, 3H), 0.34-0.20 (m, 9H).

[0406] Step 2: At room temperature and under an argon atmosphere, N-methylimidazole (0.062 g, 0.76 mmol) was added to a solution of Sn-16-a (60 mg, 0.19 mmol), TCFH (0.11 g, 0.38 mmol), and compound 11-a (0.10 g, 0.19 mmol) in ACN (2 mL). The mixture was stirred at room temperature for 0.5 h and concentrated under reduced pressure. The crude product was purified by preparative HPLC to obtain 13 mg of compound Sn-16.

[0407] LCMS (ESI) m / z = 800.0 [M+H] + ;

[0408] 1 H NMR (400 MHz, CD3OD) δ 8.05 (d, J = 8.8 Hz, 2H), 7.82 (d, J= 8.8Hz, 2H), 7.25-6.97 (m, 3H), 3.61-3.44 (m, 15H), 3.42 (s, 2H), 3.27-3.22 (m,4H), 2.70 (t, J = 6.8 Hz, 2H), 2.55 (t, J = 6.8 Hz, 2H), 2.29 (s, 3H), 1.80-1.66 (m, 4H), 0.35-0.14 (m, 9H).

[0409] Example 23

[0410] Synthesis of N1-(1-(3-methyl-5-(trimethylsilyl)phenyl)-2-oxo-7,10,13-trioxa-3-azahexadecane-16-yl)-N4-(4-(5-(methylsulfonyl)-1,3,4-oxadiazol-2-yl)phenyl)succinamide (compound Si-16)

[0411] Step 1: At room temperature, trimethylcycloboroxane (40.75 g, 162.3 mmol), cesium carbonate (18.51 g, 56.80 mmol), and [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloromethane complex (0.13 g, 0.16 mmol) were added to a 1,4-dioxane (50 mL) solution of compound 3,5-dibromo-1-trimethylsilylbenzene Si-16-a (5 g, 16.23 mmol) and 1,4-dioxane (50 mL). The reaction mixture was reacted at 110 °C for 16 hours under argon protection. The reaction was monitored by LCMS. The reaction mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (mobile phase: pure petroleum ether) to obtain 2.2 g of compound (3,5-dimethylphenyl)trimethylsilane Si-16-b as a yellow oil, with a yield of 76.0%. 1 H NMR (400MHz, CDCl3): δ 7.16 (s, 2H), 7.03 (s, 1H), 2.35 (s, 6H), 0.28 (s, 9H).

[0412] Step 2: At room temperature, azobisisobutyronitrile (59.85 mg, 0.36 mmol) was added to a carbon tetrachloride (10 mL) solution of compound (3,5-dimethylphenyl)trimethylsilane Si-16-b (1.3 g, 7.29 mmol) and N-bromosuccinimide (1.30 g, 7.29 mmol). The reaction mixture was reacted at 80°C for 16 hours under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (mobile phase: pure petroleum ether) to obtain 1 g of compound (3-(bromomethyl)-5-methylphenyl)trimethylsilane Si-16-c, with a yield of 53.3%. 1 H NMR (400 MHz, CDCl3): δ 7.33(s, 1H), 7.23(s, 1H), 7.15(s, 1H), 4.50(s, 2H), 2.37(s, 3H), 0.28(s, 9H).

[0413] Step 3: At room temperature, sodium cyanide (0.38 g, 7.78 mmol) was added to a solution of compound (3-(bromomethyl)-5-methylphenyl)trimethylsilane Si-16-c (1 g, 3.89 mmol) and 15-crown ether-5 (1.71 g, 7.78 mmol) in N,N-dimethylformamide (10 mL). The reaction mixture was reacted at room temperature under argon protection for 16 hours. The reaction was monitored by LCMS until completion. The reaction mixture was quenched with water and extracted with ethyl acetate. The combined organic phase solutions were washed with brine, dried over sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether = 1 / 7) to obtain compound 2-(3-methyl-5-(trimethylsilyl)phenyl)acetonitrile Si-16-d 710 mg, as a yellow oil, with a yield of 89.8%. 1 H NMR (400 MHz, CDCl3): δ 7.29(s, 1H), 7.23(s, 1H), 7.16(s, 1H), 3.73(s, 2H), 2.38(s, 3H), 0.28(s, 9H).

[0414] Step 4: At room temperature, 6M NaOH (5 mL) was added to an ethanol (5 mL) solution of Si-16-d (610 mg, 3.00 mmol). The reaction mixture was reacted at 95°C for 2 hours under argon protection. The reaction was monitored by LCMS until completion. The reaction mixture was concentrated under reduced pressure. The residue was dissolved in water (15 mL) and extracted with ethyl acetate (10 mL x 2). The resulting aqueous solution was adjusted to pH=2 with 1M hydrochloric acid and extracted with ethyl acetate (10 mL x 3). The combined organic phases were washed with brine, dried over sodium sulfate, filtered, and concentrated to give 420 mg of compound 2-(3-methyl-5-(trimethylsilyl)phenyl)acetic acid Si-16-e as a yellow oil, with a yield of 62.9%. 1 H NMR (400 MHz, CDCl3): δ 7.28(s, 1H), 7.24(s, 1H), 7.14(s, 1H), 3.65(s, 2H), 2.38(s, 3H), 0.29(s, 9H).

[0415] Step 5: At room temperature, N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (22.56 mg, 0.080 mmol) was added to a solution of 2-(3-methyl-5-(trimethylsilyl)phenyl)acetic acid Si-16-e (15 mg, 0.067 mmol), N-(3-(2-(3-aminopropoxy)ethoxy)propyl)-N'-(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)succinamide 11-a (36.2 mg, 0.067 mol), and N-methylimidazolium (27.50 mg, 0.34 mmol) in acetonitrile (1 mL). The reaction mixture was reacted at room temperature under argon protection for 30 minutes. The reaction was monitored by LCMS until completion. The reaction mixture was then concentrated under reduced pressure. The residue was separated and purified by high performance liquid chromatography (0.1% trifluoroacetic acid) to obtain compound N-(4-(5-methanesulfonyl-1,3,4-oxadiazol-2-yl)phenyl)-N'-(3-(2-(3-methyl-5-(trimethylsilyl)phenyl)acetamido)propoxy)ethoxy)propyl)succinamide Si-16 28.73 mg, yield 57.0%.

[0416] LCMS(ESI) m / z = 746.1 [M+H] + ;

[0417] 1 H NMR (400 MHz, DMSO-) d 6): δ 10.19(s, 1H), 7.83-7.77(m, 3H), 7.65-7.63(m, 3H), 6.95(d, J = 4.0 Hz, 2H), 6.83(s, 1H), 3.48(s, 3H), 3.29-3.27(m, 4H), 3.25-3.21(m, 4H), 3.19-3.14(m, 4H), 3.12(s, 2H), 2.86(q, J = 6.4, 12.8 Hz, 4H), 2.41-2.38(m, 2H), 2.22-2.18(m, 2H), 2.00(s, 3H), 1.43-1.36(m, 4H), 0.00(s, 9H).

[0418] Preparation Example A-1

[0419] I. Compounds 125 Preparation of I-11

[0420] 1. To prepare a 1 mg / mL Sn-11 solution: Weigh 0.1 mg Sn-11 (Fw: 822.56), add 100 μL of methanol solution containing 1% acetic acid, and dissolve thoroughly.

[0421] 2. To prepare a 0.4 mg / mL NCS solution: Weigh 0.1 mg NCS, add 250 μL of methanol solution containing 1% acetic acid, and dissolve thoroughly.

[0422] 3. Take one 1.5 mL centrifuge tube as the reaction tube, and add the following ingredients sequentially: 10 mCi of Na 125 Solution I; 100 μL of a 1 mg / mL precursor solution (i.e., Sn-11 solution); 100 μL of 0.4 mg / mL NCS solution; 2 μL glacial acetic acid; The compound was obtained by reacting the compound at room temperature with low-speed shaking for at least 20 minutes. 125 I-11 9.6 mCi, radiochemical purity 91.1%.

[0423] 4. After shaking, dry the labeled product with nitrogen gas, and then reconstitute it with 0.5 mL of 40% acetonitrile solution.

[0424] 5. Purify using a purification instrument: Column: Waters Symmetry® C18 5μm 4.6×250mm Mobile phase A: an aqueous solution containing 0.1% TFA; Mobile phase B: an acetonitrile solution containing 0.1% TFA.

[0425] 6. Take a small glass vial and combine the eluents from 15 to 18 minutes.

[0426] 7. The purified product was dried under nitrogen gas to obtain the compound. 125 I-11 9.6 mCi, HPLC radiochemical purity 99%; stored in a refrigerator at 2-8℃, reconstituted with DMA before experiment.

[0427] Preparation Example A-2

[0428] The compound was prepared according to the method of Preparation Example A-1. 125 I-14; 9.3 mCi, HPLC radiochemical purity result was 99%.

[0429] Preparation Example A-3

[0430] The compound was prepared according to the method of Preparation Example A-1. 125 I-16; 9.1 mCi, HPLC radiochemical purity result was 99%.

[0431] Preparation Example B

[0432] Preparation method of nanobody Ab-1 (FRα antibody): Camel-derived antibody discovery: A 100-fold volume of the natural alpaca VHH antibody library was revived, cultured, and panned. The eluted phages were re-infected with TG1 cells for amplification of the elution products. The phages were purified by PEG / NaCl precipitation for the next round of screening. Four rounds of phage library enrichment and screening were performed, with progressively decreasing antigen concentrations and progressively increasing washing intensities. The titers of the elution products from each round were determined. The bacterial cultures from rounds 1-4 were limitedly diluted and plated, incubated overnight, and then single clones were picked and cultured overnight in 96-well deep-well plates containing 0.5 mL / well 2YT-AG medium for induced expression, ELISA detection, and sequencing to obtain the camel-derived molecules.

[0433] Humanization and affinity maturation of camel-derived antibodies: Humanization of natural alpaca VHH antibodies was achieved using CDR transplantation technology. The variable region of the natural alpaca VHH antibody was sequence-aligned with human GermLine (IMGT database). Based on sequence consistency, human GermLine V and J region templates with high sequence identity were selected. Simultaneously, CDRs were annotated using the Kabat method based on the natural alpaca VHH antibody sequence. The CDRs of the natural alpaca VHH antibody were then transplanted into their corresponding positions in each human GermLine V and J region template. Reverse mutation was then used to retain the FR region amino acids, which are crucial for antigen binding in the framework region of the natural alpaca VHH antibody. Finally, the humanized molecule was humanized. Affinity maturation modification was performed on the humanized molecule to obtain molecule Ab-1.

[0434] The amino acid sequence of nanobody Ab-1 is shown below (CDR is divided according to Kabat encoding):

[0435] Preparation Example C:

[0436] Preparation of nanobody conjugates

[0437] 1. Preparation of 10mM TCEP: Weigh 1 mg of TCEP, add 349 μL of pure water, and dissolve completely.

[0438] 2. Pre-dilution: Take nanobody Ab-1 and add 1×PBS (pH 7-7.5) to dilute to about 0.15 mg / mL (about 10 nmol / mL); 3. Reduction: Take a 1.5 mL centrifuge tube and add 1000 μL of 10 nmol / mL Ab-1 nanobody (10 nmol), 10 μL of 10 mM TCEP (100 nmol), and 50 μL of 0.1 M EDTA (5 μmol) sequentially. After thorough mixing, react at 22℃ and 650 rpm for at least 2 h.

[0439] 4. Buffer Change: 1) Use a 3kD ultrafiltration tube to change the buffer of the nanobody Ab-1 with PB buffer (50mM, pH 8.0). Repeat ultrafiltration 4 times; 2) Replace the nanobody Ab-1 with PB buffer (50mM, pH 8.0) using a PD10 column and collect the first 3 mL of eluent; 3) Concentrate 3 mL of eluent using a 3kD ultrafiltration tube. Finally, bring the volume to 1 mL with PB buffer (50 mM, pH 8.0).

[0440] 5. Reconstitute with 100 μL DMA 125 I-11 9.6 mCi.

[0441] 6. Conjugation: Take a 1.5 mL centrifuge tube and add 1000 μL of reduced nanobody Ab-1 and approximately 3 mCi of [unclear - possibly a specific ingredient or solution]. 125 I-11. After thorough mixing, react at 22℃ and 650 rpm for 30 min.

[0442] 7. Desalting and purification: The conjugate was desalted using a PD10 column by replacing the medium with 1×PBS (pH 7-7.5). One fraction was collected for every 0.5 mL of the column, and the fractions with high radioactivity were combined. The final product RDC-11A was obtained at 5.3 mCi, with a yield of 55% and an HPLC radiochemical purity of 100%.

[0443] The above liquid chromatography methods used for measuring radiochemical purity are as follows:

[0444] Test Example 1: Tissue Distribution Experiment of Radiolabeled Conjugates Prepared from the Compounds of the Invention

[0445] I. Experimental Materials and Instruments

[0446] 1. Cell line (human oral epithelioid carcinoma KB cells)

[0447] 2. Mice (BALB / c Nude, male, 8-10 weeks old, Zhejiang Vital River Pharmaceutical Co., Ltd.)

[0448] 3. HPLC (Shimadzu, LC-20AD)

[0449] 4. γ-counter (PerkinElmer, 3470)

[0450] II. Experimental Procedure

[0451] Male BALB / c Nude mice aged approximately 8-10 weeks were subcutaneously inoculated with 5 × 10⁸ g of insecticide. 6 Human oral epithelioid carcinoma KB cells. The tumor grows to approximately 100-400 mm. 3 When the size is, 125 Taking the radiolabeled conjugate RDC-11A as an example, 125I. The radiolabeled conjugate RDC-11A was injected intravenously into mice (approximately 100 μCi / mouse). Animals were euthanized at 1, 4, and 24 h post-administration via carbon dioxide inhalation. After euthanasia, serum, urine, brain, heart, liver, lungs, kidneys (cortex of the right kidney), muscle, spleen, salivary glands, bones, duodenum, rectum, pancreas, stomach, and tumors were collected. The %ID / g (per gram of tissue radioactivity count relative to the total injected radioactivity count) was calculated for each organ.

[0452] III. Experimental Results

[0453] The tissue distribution results of RDC-11A in a human oral epithelioid carcinoma KB cell-bearing mouse model are shown in Figure 1.

[0454] IV. Experimental Conclusions

[0455] Experimental results showed that compound RDC-11A achieved tumor uptake of 5.2% ID / g at 1 hour, while renal uptake was only 19% ID / g and was rapidly metabolized. At 4 hours, tumor uptake was 3%, while renal uptake had decreased to 0.6% ID / g. This demonstrates that the radiolabeled conjugate prepared from the compounds of this invention exhibits high and long-lasting tumor targeting, extremely low levels in most normal tissues, good tissue selectivity for the kidney / tumor, relatively low renal uptake, and rapid clearance.

Claims

1. A linker-radiohalogen conjugate comprising a linker and a radioactive halogen, said linker and said radioactive halogen being covalently linked, said linker comprising the structure of Formula I or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof. Formula I in: R A Selected from hydrogen, cyano, nitro, halogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, C2-C6 alkenyl, C2-C6 alkynyl, R a -L a -; L a Selected from -(CH2) m1 -、-O-、-S-、-NH-; R a Selected from guanidinyl, protonated guanidinyl, ; m1 is an integer selected from 1 to 5; m2 is an integer selected from 1 to 20; L 1 Selected from carbonyl, -(CH2) n1 -C(=O)-、-L 11 -L 12 -; L 11 Selected from carbonyl, -(CH2) n1 -C(=O)-; L 12 The amino acid residues are short peptides formed from 2 to 10 amino acid residues or isopeptides formed from 2 to 10 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid. n1 is an integer selected from 1 to 5; L 2 Selected from , , , ; L 21 Selected from direct bonds, -(CH2) n21 -、-O-、-S-; L 22 Selected from polyethylene glycol chains and polysarcosine chains; L 23 Selected from direct bonds, -(CH2) n23 -、-O-、-S-; R 1 Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, ; R 2 Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, ; n21 is an integer selected from 1 to 5; n23 is an integer selected from 1 to 5; n25 is an integer selected from 1 to 20; L 3 Selected from direct bond, carbonyl group, , , , , , ; R 3 Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, C2-C6 alkenyl, and C2-C6 alkynyl; n31, n32, n33, n34, n35, and n36 are each independently selected from integers from 1 to 5; R B Selected from , , , , ; R b1 Selected from C1-C6 alkyl groups; R b2 R b3 Each is independently selected from hydrogen, cyano, nitro, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 alkylthio; R b4 R b5 R b6 Each is independently selected from hydrogen, cyano, nitro, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 alkylthio; Y is selected from halogens; q is an integer selected from 1 to 5; And, when R B Selected from R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

2. The linker radioactive halogen conjugate according to claim 1, wherein, R A Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, R a -L a -; Preferably, R A Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, R a -L a -; More preferably, R A Selected from hydrogen, methyl, methoxy, , , ; Or, L a Selected from -(CH2) m1 -、-O-、-S-; Preferably, L a Selected from -(CH2) m1 -、-O-; More preferably, L a Selected from -CH2- and -O-; Or, R a Selected from , , ; Preferably, R a Selected from , , ; Alternatively, m1 can be an integer selected from 1 to 3, preferably 1; Alternatively, m2 can be an integer selected from 1 to 10, preferably an integer selected from 1 to 5, and more preferably 3.

3. The linker radioactive halogen conjugate according to claim 1 or 2, wherein, L 1 Selected from carbonyl, -CH2-C(=O)-, -L 11 -L 12 -; Preferably, L 1 Selected from carbonyl, -CH2-C(=O)-, , , , , ; Or, L 11 Selected from carbonyl, -CH2-C(=O)-; Or, L 12 The amino acid residues are short peptides formed from 2-3 amino acid residues or isopeptides formed from 2-3 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid; Preferably, L 12 It is a short peptide formed from 2-3 amino acid residues, wherein the amino acids are natural amino acids or non-natural amino acids; Preferably, the amino acid is selected from glutamic acid, valine, methionine, lysine, and so on. ; Preferably, the amino acid is selected from L-glutamic acid, L-valine, L-methionine, and L-lysine. ; More preferably, L 12 Selected from , , , , ; Alternatively, n1 can be an integer selected from 1 to 3, preferably 1; Preferably, L 2 Selected from , , , , n22 is an integer selected from 1 to 20. n24 is an integer selected from 1 to 20; Preferably, L 2 Selected from , , , , , , , , , , , ; Or, L 21 Selected from direct bonds, -(CH2) n21 -; Or, L 23 Selected from direct bonds, -(CH2) n23 -; Or, R 1 Selected from hydrogen, C1-C6 alkyl, ; Preferably, R 1 Selected from hydrogen, ; Or, R 2 Selected from hydrogen, C1-C6 alkyl, ; Preferably, R 2 Selected from hydrogen, ; Alternatively, n21 can be an integer selected from 1 to 3, preferably 1 or 2; Alternatively, n22 is an integer selected from 1 to 10, preferably an integer selected from 2 to 8, and more preferably 2, 3, 4 or 8; Alternatively, n23 is an integer selected from 1 to 3, preferably 1 or 2; Alternatively, n24 is an integer selected from 1 to 10, preferably an integer selected from 5 to 10, and more preferably 6 or 7; Alternatively, n25 is an integer selected from 1 to 10, preferably an integer selected from 1 to 5, and more preferably 4; Preferably, L 3 Selected from direct bond, carbonyl group, , , , , , , ; Or, R 3 Selected from hydrogen and C1-C6 alkyl groups; Preferably, R 3 Selected from hydrogen and methyl; Alternatively, n31 can be an integer selected from 1 to 3, preferably 3; Alternatively, n32 can be an integer selected from 1 to 3, preferably 1; Alternatively, n33 can be an integer selected from 1 to 3, preferably 2; Alternatively, n34 can be an integer selected from 1 to 3, preferably 2; Alternatively, n35 can be an integer selected from 1 to 3, preferably 2; Alternatively, n36 can be an integer selected from 1 to 3, preferably 2; Preferably, R b1 It is methyl; or, R b2 R b3 Each is independently selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 alkylthio; Preferably, R b2 R b3 Each is independently selected from hydrogen and C1-C6 alkoxy groups; Preferably, R b2 R b3 In the above, one of them is hydrogen, and the other is selected from hydrogen or C1-C6 alkoxy groups; More preferably, R b2 R b3 Each is independently selected from hydrogen and methoxy groups; Most preferably, R b2 R b3 In this composition, one of the components is hydrogen, and the other is selected from hydrogen or methoxy groups. Or, R b4 R b5 R b6 Each is independently selected from hydrogen, cyano, nitro, and C1-C6 alkyl groups; Preferably, R b4 R b5 R b6 Each is independently selected from hydrogen and cyano groups; More preferably, R b4 R b5 R b6 In this group, any one of them is a cyano group, and the rest are hydrogen. Alternatively, Y can be -Br; Alternatively, q can be an integer selected from 1 to 3, preferably 1; Or, R B Selected from , , , , , .

4. The linker radioactive halogen conjugate according to any one of claims 1-3, wherein, The linker comprises the structure shown in Formula Ia, Formula Ib, Formula I-1a, Formula I-1b, Formula I-2a or Formula I-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer or stereoisomer thereof. Formula Ia Formula Ib Where: L 1 L 2 L 3 R A R B Each is independently defined as described in any one of claims 1-3; Formula I-1a Formula I-1b Where: R B1 Selected from , , , ; L 1 L 2 L 3 R A R b1 R b2 R b3 R b4 R b5 R b6 Y and q are each independently defined as described in any one of claims 1-3; Equation I-2a Equation I-2b Where: L 1 L 2 L 3 R A R b1 Each is independently defined as described in any one of claims 1-3; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 Not carbonyl; Preferably, the linker comprises a structure selected from the following or a pharmaceutically acceptable salt thereof, a tautomer, an enantiomer, or a stereoisomer. 。 5. The linker radioactive halogen conjugate according to claim 1, wherein, The radioactive halogen conjugate is a compound having the structure shown in Formula II or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof. Formula II in: X is a radioactive halogen; Preferably, X is selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br, 76 Br, 77 Br, 80m Br, 211 At; More preferably, X is selected from 125 I, 211 At; L 1 L 2 L 3 R A R B Each is independently defined as described in any one of claims 1-3; And, when R B Selected from R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

6. The linker radioactive halogen conjugate according to claim 5, wherein, The radioactive halogen conjugate is a compound having the structure shown in Formula II-a, Formula II-b, Formula II-1a, Formula II-1b, Formula II-2a or Formula II-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer or stereoisomer thereof. Formula II-a Formula II-b Where: L 1 L 2 L 3 R A R B X and X are each independently defined as described in claim 5; Formula II-1a Formula II-1b Where: L 1 L 2 L 3 R A X and X are each independently defined as described in claim 5; R B1 As defined in claim 4; Formula II-2a Formula II-2b Where: L 1 L 2 L 3 R A X and X are each independently defined as described in claim 5; R b1 As defined in claim 4; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 Not carbonyl; Preferably, the radioactive halogen conjugate is selected from compounds with the following structures or their pharmaceutically acceptable salts, tautomers, enantiomers, or stereoisomers. 。 7. The compound represented by Formula III or its pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer, Formula III in: X 1 The solvent is F, Cl, Br or I, preferably I; L 1 L 2 L 3 R A R B Each is independently defined as described in any one of claims 1-3; And, when R B Selected from R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

8. The compound according to claim 7, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof, wherein, The compound has the structure shown in formula III-a or III-b, formula III-1a, formula III-1b, formula III-2a or III-2b. Formula III-a Formula III-b Where: L 1 L 2 L 3 R A R B X 1 Each is independently defined as described in claim 7; Formula III-1a Formula III-1b Where: L 1 L 2 L 3 R A X 1 Each is independently defined as described in claim 7; R B1 As defined in claim 4; Formula III-2a Formula III-2b Where: L 1 L 2 L 3 R A X 1 Each is independently defined as described in claim 7; R b1 As defined in claim 4; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 Not carbonyl; Preferably, the compound is selected from the following: 。 9. A linker radioactive halogen conjugate precursor compound or a pharmaceutically acceptable salt thereof, tautomer, enantiomer, or stereoisomer, wherein, The precursor compound has the structure shown in Formula IV. Formula IV in: X 2 It is a leaving group; Preferably, X 2 Selected from -SnR c 3, -SiR c 3. -B(OH)2; R c Selected from C1-C6 alkyl groups; Preferably, R c It is methyl; R A’ Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, C2-C6 alkenyl, C2-C6 alkynyl, R a’ -L a -; Preferably, R A’ Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylthio, R a’ -L a -; More preferably, R A’ Selected from hydrogen, C1-C6 alkyl, C1-C6 alkoxy, R a’ -L a -; Most preferably, R A’ Selected from hydrogen, methyl, methoxy, , , , ; R a’ Selected from guanidinyl, protonated guanidinyl, guanidinyl with one or more amino protecting groups, and protonated guanidinyl with one or more amino protecting groups. ; Preferably, R a’ Selected from guanidinyl, protonated guanidinyl, guanidinyl with one or more amino protecting groups, ; More preferably, R a’ Selected from , , , ; Most preferably, R a’ Selected from , , , ; L 1’ Selected from carbonyl, -(CH2) n1 -C(=O)-、-L 11 -L 12’ -; Preferably, L 1’ Selected from carbonyl, -CH2-C(=O)-, -L 11 -L 12’ -; More preferably, L 1’ Selected from carbonyl, -CH2-C(=O)-, , , , , , , , , ; L 12’ The amino acid residue is a short peptide formed from 2 to 10 amino acid residues or an isopeptide formed from 2 to 10 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group. Preferably, L 12’ The amino acid residue is a short peptide formed by 2-3 amino acid residues or an isopeptide formed by 2-3 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group. More preferably, L 12’ It is a short peptide formed from 2-3 amino acid residues, wherein the amino acid is a natural amino acid or a non-natural amino acid, and the amino acid optionally has a carboxyl protecting group; Preferably, the amino acid is selected from glutamic acid, valine, methionine, lysine, and so on. And the lysine, Each of them may independently and optionally have a carboxyl protecting group; Preferably, the amino acid is selected from L-glutamic acid, L-valine, L-methionine, and L-lysine. , , ; Most preferably, L 12’ Selected from , , , , , , , , ; L 2 L 3 R B L a m2, L 11 n1 and n2 are each independently defined as described in any one of claims 1-3; And, when R B Selected from R a’ Selected from guanidino or guanidino groups having one or more amino protecting groups, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1’ It is not a carbonyl group.

10. The precursor compound according to claim 9, or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof, wherein, The precursor compound has the structure shown in formula IV-a, IV-b, IV-1a, IV-1b, IV-2a, or IV-2b. Formula IV-a Formula IV-b Where: L 1’ L 2 L 3 R A’ R B X 2 Each is independently defined as described in claim 9; Formula IV-1a Formula IV-1b Where: L 1’ L 2 L 3 R A’ X 2 Each is independently defined as described in claim 9; R B1 As defined in claim 4; Formula IV-2a Formula IV-2b Where: L 1’ L 2 L 3 R A’ X 2 Each is independently defined as described in claim 9; R b1 As defined in claim 4; And, when R a’ Selected from guanidino or guanidino groups having one or more amino protecting groups, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1’ Not carbonyl; More preferably, the precursor compound is selected from the following: 。 11. A single-domain antibody or its antigen-binding fragment capable of specifically binding to FRα, comprising: CDR1 or a variant thereof contained in the heavy chain variable region (VHH) shown in SEQ ID NO: 4, CDR2 or a variant thereof contained in the heavy chain variable region (VHH) shown in SEQ ID NO: 4, and CDR3 or a variant thereof contained in the heavy chain variable region (VHH) shown in SEQ ID NO: 4; in, The variant has one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, or 3 amino acids) compared to its source sequence; preferably, the substitutions are conservative substitutions. Preferably, the single-domain antibody or its antigen-binding fragment comprises CDR1, CDR2 and CDR3 contained in the heavy chain variable region (VHH) as shown in SEQ ID NO: 4; Preferably, the single-domain antibody or its antigen-binding fragment comprises: CDR1 as shown in SEQ ID NO: 1, CDR2 as shown in SEQ ID NO: 2, and CDR3 as shown in SEQ ID NO:

3.

12. The single-domain antibody or its antigen-binding fragment according to claim 11, wherein, The single-domain antibody or its antigen-binding fragment contains a heavy chain framework region sequence of an immunoglobulin (e.g., a heavy chain framework region sequence of a human or camel immunoglobulin). Preferably, the single-domain antibody or its antigen-binding fragment comprises the heavy chain framework region of human immunoglobulin; Preferably, the single-domain antibody or its antigen-binding fragment comprises a heavy chain framework region contained in the amino acid sequence encoded by the human heavy chain germline gene; preferably, the heavy chain framework region optionally comprises one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) reversion mutations from human residues to camel residues; More preferably, the single-domain antibody or its antigen-binding fragment comprises an amino acid sequence selected from the following: (i) A sequence as shown in SEQ ID NO: 4; (ii) A sequence having one or more amino acid substitutions, deletions, or additions (e.g., substitutions, deletions, or additions of 1, 2, 3, 4, or 5 amino acids) compared to the sequence shown in SEQ ID NO: 4; or (iii) A sequence having at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence shown in SEQ ID NO: 4; Preferably, the substitution is a conservative substitution.

13. Use of the linker radiohalogen conjugate according to any one of claims 1-6 in the preparation of radiolabeled proteins / peptides.

14. A radionuclide-coupled drug comprising a linker radiohalogen coupling unit and a targeting ligand, wherein the linker radiohalogen coupling unit is covalently linked to the targeting ligand, and the linker radiohalogen coupling unit comprises a structure of Formula V or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof. Formula V in: X is a radioactive halogen; Preferably, X is selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br, 76 Br, 77 Br, 80m Br, 211 At; More preferably, X is selected from 125 I, 211 At; R B’ Selected from , , , , Among them, site 1 and L 3 Connected; L 1 L 2 L 3 R A R b2 R b3 R b4 R b5 R b6 q are each independently defined as described in any one of claims 1-3; And, when R B’ for R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

15. The radionuclide conjugate drug according to claim 14, wherein, The linker radioactive halogen coupling unit comprises a structure represented by formula Va, Vb, V-1a, V-1b, V-2a, or V-2b, or a pharmaceutically acceptable salt thereof, a tautomer, enantiomer, or stereoisomer. Formula Va Formula Vb Where: L 1 L 2 L 3 R A R B’ X and X are each independently defined as described in claim 14; Formula V-1a Formula V-1b Where: R B1’ Selected from , , , Among them, site 1 and L 3 Connected; L 1 L 2 L 3 R A X, R b2 R b3 R b4 R b5 R b6 q and q are each independently defined as described in claim 14; Formula V-2a Formula V-2b Where: L 1 L 2 L 3 R A X and X are each independently defined as described in claim 14; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 Not carbonyl; More preferably, the linker radioactive halogen coupling unit comprises a structure selected from the following or a pharmaceutically acceptable salt thereof, a tautomer, an enantiomer, or a stereoisomer. ； X is a radioactive halogen; Preferably, each X is independently selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At; More preferably, each X is independently selected 125 I, 211 At.

16. The radionuclide conjugate drug according to claim 14, wherein, The radionuclide-coupled drug is a compound having the structure shown in Formula VI or a pharmaceutically acceptable salt, tautomer, enantiomer, or stereoisomer thereof. Style VI in: Ab is the target ligand, preferably a protein / peptide; Preferably, Ab is an antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment that can specifically bind to FRα; Preferably, the single-domain antibody or its antigen-binding fragment capable of specifically binding to FRα is as defined in claim 11 or 12; R B’ Selected from , , , , Among them, site 1 and L 3 The two sites are connected; site 2 is connected to Ab. L 1 L 2 L 3 R A X, R b2 R b3 R b4 R b5 R b6 q and q are each independently defined as described in claim 14; And, when R B’ for R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 It is not a carbonyl group.

17. The radionuclide conjugate drug according to claim 16, wherein, The radionuclide conjugate is a compound having the structure shown in Formula VI-a, Formula VI-b, Formula VI-1a, Formula VI-1b, Formula VI-2a or Formula VI-2b, or a pharmaceutically acceptable salt, tautomer, enantiomer or stereoisomer thereof. Formula VI-a Formula VI-b in: L 1 L 2 L 3 R A X, R B’ Each of Ab is independently defined as described in claim 16; Formula VI-1a Formula VI-1b Where: R B1’ Selected from , , , Among them, site 1 and L 3 The two sites are connected; site 2 is connected to Ab. L 1 L 2 L 3 R A X, R b2 R b3 R b4 R b5 R b6 , q, and Ab are each independently defined as in claim 16; Formula VI-2a Formula VI-2b Where: L 1 L 2 L 3 R A X, Ab, and each of them are independently defined as described in claim 16; And, when R a It is a guanidinyl group, L 2 Selected from And L 22 It is a polyethylene glycol chain, R 1 For hydrogen, R 2 When it is hydrogen, L 1 Not carbonyl; More preferably, the radionuclide-coupled drug is selected from substances with the following structures or their pharmaceutically acceptable salts, tautomers, enantiomers, or stereoisomers. ， in: X is a radioactive halogen; Preferably, each X is independently selected from 18 F, 122 I, 123 I, 124 I, 125 I, 131 I, 75 Br、 76 Br、 77 Br、 80m Br、 211 At; More preferably, each X is independently selected 125 I, 211 At; Ab is the target ligand, preferably a protein / peptide; Preferably, Ab is an antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment; Preferably, Ab is a single-domain antibody or its antigen-binding fragment that can specifically bind to FRα; Preferably, the single-domain antibody or its antigen-binding fragment capable of specifically binding to FRα is as defined in claim 11 or 12.

18. A pharmaceutical composition comprising a linker radiohalogen conjugate as described in any one of claims 1-6, or a radionuclide conjugate as described in any one of claims 14-17; Optionally, the pharmaceutical composition may further comprise a pharmaceutically acceptable carrier and / or excipient.

19. A kit comprising a linker radiohalogen conjugate as described in any one of claims 1-6, or a radionuclide conjugate as described in any one of claims 14-17.

20. Use of the linker radiohalogen conjugate according to any one of claims 1-6, or the radionuclide conjugate drug according to any one of claims 14-17, or the pharmaceutical composition according to claim 18, in the preparation of a medicament for treating and / or preventing disease; Preferably, the disease is a disease related to FRα; Preferably, the disease is selected from ovarian cancer, breast cancer (such as triple-negative breast cancer), endometrial cancer, mesothelioma, lung cancer, cervical cancer, colorectal cancer, head and neck squamous cell carcinoma, and prostate cancer.

21. Use of the linker radiohalogen conjugate according to any one of claims 1-6 or the radionuclide conjugate according to any one of claims 14-17 in the preparation of imaging diagnostic agents; Preferably, the imaging is SPECT imaging, PET imaging, or CT imaging; Preferably, the imaging diagnostic agent is used to diagnose diseases; Preferably, the disease is a disease related to FRα; Preferably, the disease is selected from ovarian cancer, breast cancer (such as triple-negative breast cancer), endometrial cancer, mesothelioma, lung cancer, cervical cancer, colorectal cancer, and head and neck squamous cell carcinoma.

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