Polypeptide diagnostic and therapeutic agent targeting grpr

Abstract

This invention belongs to the field of biomedicine and discloses a peptide-based therapeutic agent targeting GRPR. Specifically, it discloses a peptide-based therapeutic agent with high affinity for gastrin-releasing peptide receptor (GRPR), its pharmaceutically acceptable salt, and pharmaceutical compositions containing the same, as well as the use of the peptide-based therapeutic agent and pharmaceutical compositions in the preparation of diagnoses, preventions, and / or treatments of diseases overexpressing GRPR.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine, specifically relating to a dual-modal imaging peptide-based drug for PET / CT, fluorescence imaging, and photodynamic therapy targeting GRPR, as well as its preparation method and application. Background Technology

[0002] Gastrin-releasing peptide (GRP) is a regulatory peptide that modulates the physiological functions of various organs through its receptor (GRPR). GRPR has gained favor in the fields of oncology and nuclear medicine due to its high expression in a variety of cancers, including prostate cancer, breast cancer, pancreatic cancer, and small cell lung cancer.

[0003] Currently, GRPR-based diagnosis and treatment mainly focus on utilizing radiolabeled GRPR ligands. Toad protein, the amphibian counterpart of mammalian gastrin-releasing peptide, has been widely used in the development of GRPR imaging molecular probes; specifically, the fragment peptide BBN(7-14) has been extensively applied. It is worth noting that GRPR antagonists have a significant advantage in terms of biocompatibility compared to agonists. This is because antagonists do not induce pharmacological effects after binding to the receptor, making them more suitable for intravenous administration.

[0004] Mansi et al. previously studied the GRPR antagonist RM1. 111 In / 68 By comparing the effects of Ga labeling and studying the biodistribution and PET / CT imaging of PC-3 tumor-bearing mice, researchers found... 111 In / 68 Ga-RM1 has shown strong targeting ability and tumor retention ability (Clin. Cancer Res. 2009 15(16):5240-9.).

[0005] In the field of GRPR-based therapy, Moody et al. found that camptothecin-bufotoxin (6-14) conjugates can significantly inhibit the growth of several human tumors (J. Biol. Chem. 2004 279(22):23580-9.). Furthermore, although antagonists have difficulty entering the cell membrane via receptor-mediated endocytosis, their accumulation on the tumor cell membrane, simply as an accumulation of antitumor drugs on the tumor cell membrane, can still result in stronger toxicity than without a carrier. Engel et al., using AN-125, a conjugate of bufotoxin analogue and doxorubicin, demonstrated that GRPR antagonists, as carriers of antitumor drugs, also possess tumor-suppressive properties (Clin. Cancer Res. 2005 11(6):2408-15). Summary of the Invention

[0006] The purpose of this invention is to provide a therapeutic peptide targeting GRPR and its preparation method for application. The therapeutic peptide has high affinity for the target, good selectivity, high stability, and excellent pharmacokinetics, and has important clinical value in the integrated diagnosis and treatment of cancer, inflammatory diseases, cardiovascular diseases, and nervous system diseases with high GRPR expression.

[0007] Specifically, the first technical solution is a polypeptide-based therapeutic drug or its pharmaceutically acceptable salt as shown in formula (Ⅰ), wherein: P is a GRPR-targeting polypeptide, C is a linker, T is a photodynamic group with fluorescent function, and D is a radionuclide chelating group.

[0008]

[0009] The aforementioned formula (I) is a therapeutic polypeptide or a pharmaceutically acceptable salt thereof, wherein the amino acid sequence of the GRPR-targeting polypeptide is: (D-Phe)-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2.

[0010]

[0011] Or a polypeptide sequence or polypeptide derivative that has 1 to 3 amino acids replaced, deleted, or inserted on (D-Phe)-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2.

[0012] The second technical solution of the present invention, wherein the formula (I) polypeptide therapeutic drug or its pharmaceutically acceptable salt, wherein the linker is selected from one of the following compounds: unmodified or modified monoamino acids, unmodified or modified amino acid chains of different lengths, such as glycine, alanine, valine, leucine, isoleucine, phenylalanine, tryptophan, tyrosine, aspartic acid, histidine, asparagine, glutamic acid, lycine, glutamine, methionine, arginine, serine, threonine, cysteine, proline, selenocysteine, pyrrolidone, hydroxyproline, sarcosine, guanidinine, homocysteine, β-alanine, and combinations thereof; alkyl chains containing or without O and N heteroatoms, such as:

[0013]

[0014] m, n, and p are each independently selected from integers between 0 and 20;

[0015] t are each independently selected from integers between 1 and 150;

[0016] r is an integer independently selected from 0 to 150.

[0017] The third technical solution of the present invention is that the formula (I) polypeptide therapeutic drug or its pharmaceutically acceptable salt, wherein the photodynamic group with fluorescent function is selected from one of IR-780, IR-783, IR-808, IR-825, indocyanine green, protoporphyrin IX, and rose red.

[0018] The fourth technical solution of the present invention, wherein the photodynamic group with fluorescence function described in the third technical solution is characterized in that IR-780, IR-783, IR-808, IR-825, and indocyanine green are subjected to photodynamic therapy under an 808nm laser, protoporphyrin IX is subjected to photodynamic therapy under a 630-635nm laser, and rose red is subjected to photodynamic therapy under a 560nm laser.

[0019] The fifth technical solution of the present invention, wherein the formula (I) polypeptide therapeutic drug or its pharmaceutically acceptable salt, wherein the radioactive nuclide chelating group is selected from one of the following:

[0020]

[0021] This invention provides a method for preparing a polypeptide therapeutic agent or a pharmaceutically acceptable salt thereof as described in any of the foregoing technical solutions, characterized by comprising the following steps:

[0022] Synthesize the polypeptide compound;

[0023] The polypeptide compound is coupled with the linker arm, the photodynamic group with fluorescent function, and the chelating group to obtain the crude product of the polypeptide-based diagnostic and therapeutic drug.

[0024] The crude product of the labeled precursor is purified to obtain the polypeptide-based therapeutic drug.

[0025] The sixth technical solution of the present invention is a radiopeptide-based integrated diagnostic and therapeutic drug of formula (I) obtained by chelating it with a radionuclide, or a pharmaceutically acceptable salt thereof, wherein the radionuclide is selected from... 18 F, 64 Cu、 67 Cu、 68 Ga、 89 Zr、 99m Tc, 111 In、 177 Lu、 186 Re、 188 Re、 203 Pb, 212 Pb, 213 Bi、 225 Ac、 227 Th.

[0026] This invention provides a method for preparing the integrated therapeutic peptide drug or its pharmaceutically acceptable salt as described in the sixth technical solution. The method comprises the following steps: mixing and incubating the labeled precursor with a radionuclide solution to obtain the radioactive integrated therapeutic peptide drug; the incubation conditions are 20-110°C for 5-60 minutes. Preferably, before mixing the integrated therapeutic peptide drug or its pharmaceutically acceptable salt with the radionuclide solution, the integrated therapeutic peptide drug or its pharmaceutically acceptable salt is first dissolved in a buffer solution, wherein the buffer solution is one or a mixture of sodium acetate, water, ethanol, phosphate buffer solution, or dimethyl sulfoxide, with a pH value of 4.0-10.0.

[0027] The present invention also relates to a pharmaceutical composition or pharmaceutical formulation comprising the (radioactive) therapeutic polypeptide drug or its pharmaceutically acceptable salt as described in any of the foregoing technical solutions, and a pharmaceutically acceptable carrier and / or excipient. Further, the pharmaceutical composition or pharmaceutical formulation comprises 1-1500 mg of the (radioactive) therapeutic polypeptide drug or its pharmaceutically acceptable salt as described in any of the foregoing technical solutions, and a pharmaceutically acceptable carrier and / or excipient.

[0028] This invention also relates to a use of the (radioactive) polypeptide therapeutic agent or its pharmaceutically acceptable salt, or pharmaceutical composition or pharmaceutical preparation described in any of the foregoing technical solutions, in the diagnosis, prevention and / or treatment of diseases overexpressing GRPR, preferably diseases overexpressing GRPR selected from cancers, and examples of diagnosable cancers (and their benign counterparts) include, but are not limited to, tumors of epithelial origin (adenomas and various types of cancer, including adenocarcinoma, squamous cell carcinoma, transitional cell carcinoma and other cancers), such as bladder and urinary tract cancers, breast cancers, gastrointestinal cancers (including esophageal cancer, gastric cancer, small bowel cancer, colon cancer, rectal cancer and anal cancer), liver cancer (hepatocellular carcinoma), gallbladder and biliary system cancers, exocrine pancreatic cancers, kidney cancers, lung cancers (e.g. Cancers including adenocarcinoma, small cell lung cancer, non-small cell lung cancer, bronchoalveolar carcinoma, and mesothelioma; head and neck cancers (e.g., tongue cancer, buccal carcinoma, laryngeal cancer, pharyngeal cancer, nasopharyngeal carcinoma, tonsil cancer, salivary gland cancer, nasal cavity cancer, and paranasal sinus cancer); ovarian cancer, fallopian tube cancer, peritoneal cancer, vaginal cancer, vulvar cancer, penile cancer, cervical cancer, uterine fibroids, endometrial cancer; thyroid cancer (e.g., follicular thyroid carcinoma); kidney cancer; prostate cancer; skin and appendage cancers (e.g., melanoma, basal cell carcinoma, squamous cell carcinoma, keratoacanthoma, and dysplastic nevus); hematologic malignancies (i.e., leukemia, lymphoma) and pre-existing hematologic disorders as well as borderline malignancies, including hematologic malignancies and lymphoid spectrum disorders (e.g., acute lymphoblastic leukemia [A]). [LL], chronic lymphocytic leukemia [CLL], B-cell lymphomas such as diffuse large B-cell lymphoma [DLBCL], follicular lymphoma, Burkitt's lymphoma, mantle cell lymphoma, T-cell lymphoma and leukemia, natural killer [NK] cell lymphoma, Hodgkin's lymphoma, hairy cell leukemia, monoclonal gammopathy of indeterminate significance, plasmacytoma, multiple myeloma and post-transplant lymphoproliferative disorders, and hematologic malignancies and myeloid-related conditions (e.g., acute myeloid leukemia [AML], chronic myeloid leukemia [CML], chronic myelomonocytic leukemia [CMML], eosinophilic syndrome, myeloproliferative disorders such as polycythemia vera, essential thrombocytosis Multiple diseases and primary myelofibrosis, myelodysplastic syndrome, myelodysplastic syndrome, and promyelocytic leukemia; tumors of mesenchymal origin, such as soft tissue sarcomas, bone or chondrosarcomas, such as osteosarcoma, fibrosarcoma, chondrosarcoma, rhabdomyosarcoma, leiomyosarcoma, liposarcoma, angiosarcoma, Kaposi's sarcoma, Ewing's sarcoma, synovial sarcoma, epithelioid sarcoma, gastrointestinal stromal tumors, benign and malignant tissue sarcomas, and dermofibrosarcoma protuberans; tumors of the central or peripheral nervous system (such as astrocytomas, gliomas and glioblastomas, meningiomas, ependymomas, pineal tumors, and schwannomas); endocrine tumors (such as pituitary adenomas, adrenal tumors, islet cell tumors, parathyroid tumors, carcinoid tumors, and medullary thyroid carcinomas);Tumors of the eye and adnexa (e.g., retinoblastoma); germ cell and trophoblastic tumors (e.g., teratoma, seminoma, dysgerminoma, hydatidiform mole, and choriocarcinoma); pediatric and embryonic tumors (e.g., medulloblastoma, neuroblastoma, Wilms' tumor, and primitive neuroectodermal tumor); or congenital or other forms of syndrome that predispose a patient to malignancy (e.g., xeroderma pigmentosum). More preferably, the aforementioned diseases are breast cancer, uterine cancer, cervical cancer, ovarian cancer, or prostate cancer.

[0029] This invention relates to a kit that may comprise a single-dose or multi-dose composition comprising the present invention’s (radioactive) therapeutic peptide medicament or a pharmaceutically acceptable salt thereof, wherein the amount of the present invention’s (radioactive) therapeutic peptide medicament or the pharmaceutically acceptable salt thereof is the same as the amount in the aforementioned pharmaceutical composition.

[0030] The amount of (radioactive) polypeptide therapeutic agent or its pharmaceutically acceptable salt described in this invention is converted in the form of free base in each case.

[0031] The carbon, hydrogen, oxygen, sulfur, and nitrogen involved in the groups and compounds described in this invention all include their isotopes, and the carbon, hydrogen, oxygen, sulfur, and nitrogen involved in the groups and compounds described in this invention may optionally be further replaced by one or more of their corresponding isotopes, wherein the isotopes of carbon include 12 C 13 C and 14 C, the isotopes of hydrogen include protium (H), deuterium (D, also known as heavy hydrogen), and tritium (T, also known as superheavy hydrogen), and the isotopes of oxygen include... 16 O、 17 O and 18 O, isotopes of sulfur include 32 S, 33 S, 34 S and 36 S, nitrogen isotopes include 14 N and 15 N.

[0032] "Pharmaceutically acceptable salt" means that the compound of the present invention retains the bioavailability and properties of a free acid or a free base, wherein the free acid is obtained by reacting with a non-toxic inorganic or organic base, and the free base is obtained by reacting with a non-toxic inorganic or organic acid.

[0033] "Pharmaceutical composition" means one or more compounds described herein or their pharmaceutically acceptable salts, mixed with other components, wherein the other components contain physiologically / pharmaceuticalally acceptable carriers and / or excipients.

[0034] "Carrier" refers to a system that does not cause significant stimulation to the organism and does not eliminate the biological activity and properties of the given compound, and can change the way the drug enters the human body and its distribution in the body, control the release rate of the drug, and deliver the drug to the target organ. Non-limiting examples include microcapsules and microspheres, nanoparticles, liposomes, etc.

[0035] "Excipient" refers to an agent that is not itself a therapeutic agent but is used as a diluent, excipient, binder, and / or medium to be added to a pharmaceutical composition to improve its disposal or storage properties or to allow or promote the formation of a unit dosage form of the compound or pharmaceutical composition for administration. As is known to those skilled in the art, pharmaceutical excipients can provide a variety of functions and can be described as wetting agents, buffers, suspending agents, lubricants, emulsifiers, disintegrants, absorbents, preservatives, surfactants, colorants, flavoring agents, and sweeteners. Examples of pharmaceutical excipients include, but are not limited to: (1) sugars, such as lactose, glucose, and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose, cellulose acetate, hydroxypropyl methyl cellulose, hydroxypropyl cellulose, microcrystalline cellulose, and croscarmellose (e.g., sodium croscarmellose); (4) tragacanth gum powder; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter. (9) Oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) Diols, such as propylene glycol; (11) Polyols, such as glycerol, sorbitol, mannitol and polyethylene glycol; (12) Esters, such as ethyl oleate and ethyl laurate; (13) Agar; (14) Buffers, such as magnesium hydroxide and aluminum hydroxide; (15) Alginate; (16) Atherless water; (17) Isotonic saline; (18) Ringer's solution; (19) Ethanol; (20) pH buffer solution; (21) Polyesters, polycarbonates and / or polyanhydrides; and (22) Other non-toxic compatible substances used in pharmaceutical preparations. Attached Figure Description

[0036] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0037] Figure 1 The results are from HPLC and mass spectrometry analysis of the polypeptide precursor P1.

[0038] Figure 2 The results of HPLC and mass spectrometry analysis of the therapeutic drug PXC-110;

[0039] Figure 3 for[ 64 RP-HPLC analysis results of Cu]PXC-110;

[0040] Figure 4 The fluorescence imaging results of PXC-110 at different time points in the PC-3 tumor-bearing NCG mouse model;

[0041] Figure 5 The results show the fluorescence biodistribution of PXC-110 in major organs 6 hours after drug injection in a PC-3 tumor-bearing NCG mouse model.

[0042] Figure 6 for[ 64 PET / CT imaging results of Cu]PXC-110 at different time points in PC-3 tumor-bearing NCG mouse model;

[0043] Figure 7 The tumor-suppressive activity of PXC-110 in the PC-3 tumor-bearing NCG mouse model;

[0044] Figure 8 The effect of PXC-110 on mouse body weight in PC-3 tumor-bearing NCG mouse model. Detailed Implementation

[0045] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0046] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0047] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0048] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be readily apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0049] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0050] This invention uses the human prostate cancer PC-3 cell line as a cell model. The culture medium is Ham's F-12K (Kaighn's) medium + 10% fetal bovine serum + 1% penicillin / streptomycin. The incubator is set at 37°C and 5% CO2. The medium is changed once a day, and passages are performed every two days.

[0051] All animal experiments used in this invention were approved by the Animal Ethics Committee of the Institute of Materia Medica, Chinese Academy of Medical Sciences. All animal experiments adhered to the 3R principle and were conducted under the guidance of the Laboratory Animal Ethics Committee. The PC-3 mouse model was established via subcutaneous injection, with each mouse receiving a unilateral axillary injection of 1×10⁻⁶ ppm. 6 One tumor was constructed per mouse using individual cells. After 3 weeks, when the tumor tissue grew to approximately 200-300 mm... 3 At that time, mice were given the drug and PET / CT imaging studies were performed.

[0052] The English and Chinese meanings of the abbreviations in this invention are as follows:

[0053] Fmoc: 9-Shuthylmethoxycarbonyl;

[0054] DMF: N,N-dimethylformamide;

[0055] TIS: Triisopropylsilane;

[0056] DOTA: 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid;

[0057] Rink Amide-MBHA Resin: 4-Methyldiphenylmethylamine resin hydrochloride;

[0058] HCTU: 6-Chlorobenzotriazole-1,1,3,3-Tetramethylurea hexafluorophosphate;

[0059] DIPEA: N,N-diisopropylethylamine;

[0060] Example

[0061] Example 1: PXC-110, a peptide-based therapeutic drug targeting GRPR, and its radiolabeled counterpart [ 64 Cu]PXC- Synthesis of 110

[0062] Using an Fmoc-based solid-phase peptide synthesis method, the sequence (D-Phe)-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2 was synthesized on Rink Amide MBHA resin. This was then coupled with the amino acid (β-Ala)-Lys-(β-Ala), which acts as a linker, followed by coupling with the chelating group DOTA. Deprotection was then performed, and the crude product was cleaved. Intermediate P1 was obtained by HPLC purification. The specific synthetic route is as follows:

[0063] (1) Resin swelling: Add 500 mg of Rink Amide MBHA resin with a degree of substitution of 0.35 mmol / g to the reaction tube, then add 10 mL of DMF, place on a shaker and shake at room temperature for 30 minutes, then discard the solution.

[0064] (2) Removal of resin protecting group Fmoc: Add 10 mL of 50% morpholine / DMF solution (morpholine:DMF = 1:1, v:v), place on a shaker and shake at room temperature for 60 minutes, then discard the solution.

[0065] (3) Resin cleaning: Add 10 mL of DMF to the tube, place it on a shaker and shake at room temperature for 2 minutes, discard the solution, and repeat this step 4 times.

[0066] (4) Linkage of the first amino acid: Add Fmoc-Leu-OH (4 eq, 247.4 mg) and HCTU (4 eq, 289.6 mg), then add DMF to dissolve, and finally add DIPEA (8 eq, 243.8 μL). Place on a shaker and shake at room temperature for 120 minutes, then discard the solution.

[0067] (5) Resin cleaning: Add 10 mL of DMF to the tube, place it on a shaker and shake at room temperature for 2 minutes, discard the solution, and repeat this step 4 times.

[0068] (6) Removal of amino acid protecting group Fmoc: Add 10 mL of 50% morpholine / DMF solution (morpholine:DMF = 1:1, v:v), place on a shaker and shake at room temperature for 60 minutes, then discard the solution.

[0069] (7) Resin cleaning: Add 10 mL of DMF to the tube, place it on a shaker and shake at room temperature for 2 minutes, discard the solution, and repeat this step 4 times.

[0070] (8) Second amino acid linkage: Add Fmoc-Sta-OH (4 eq, 247.4 mg) and HCTU (4 eq, 289.6 mg), then add DMF to dissolve, and finally add DIPEA (8 eq, 243.8 μL). Place on a shaker and shake at room temperature for 120 minutes, then discard the solution.

[0071] Repeat the above steps, sequentially connecting: Fmoc-His(Trt)-OH (52.67 mg), Fmoc-Gly-OH (38.05 mg), Fmoc-Val-OH (83.04 mg), Fmoc-Ala-OH (38.05 mg), Fmoc-Trp(Boc)-OH (49.08 mg), Fmoc-Gln(Trt)-OH (38.05 mg), Fmoc-D-Phe-OH (49.08 mg), Fmoc-β-Ala-OH (38.05 mg), Fmoc-Lys(Boc)-OH (49.08 mg), and Fmoc-β-Ala-OH (38.05 mg), with the total amount of each substance being 4 eq.

[0072] (9) Resin cleaning: Add 10 mL of DMF to the tube, place it on a shaker and shake at room temperature for 2 minutes, discard the solution, and repeat this step 4 times.

[0073] (10) Removal of amino acid protecting group Fmoc: Add 10 mL of 50% morpholine / DMF solution (morpholine:DMF = 1:1, v:v), place on a shaker and shake at room temperature for 60 minutes, then discard the solution.

[0074] (11) Resin cleaning: Add 10 mL of DMF to the tube, place it on a shaker and shake at room temperature for 2 minutes, discard the solution, and repeat this step 4 times.

[0075] (12) Coupling of DOTA: Add DOTA-tris(tBuester) (4eq, 400.9mg) and HCTU (4eq, 289.6mg), then add DMF to dissolve, and finally add DIPEA (8eq, 243.8μL). Place on a shaker and shake at room temperature for 120 minutes, then discard the solution.

[0076] (13) Peptide shearing: Add 10 mL of Cocktail shearing solution (TFA:TIPS:H2O = 95:2.5:2.5, v:v:v) to the tube, place it in a shaker and shake at room temperature for 2 hours, filter, collect the filtrate, and then blow it dry with nitrogen to obtain solid crude peptide.

[0077] (14) Peptide purification: A cold mixture of diethyl ether and n-hexane (ether:n-hexane = 1:1, v:v) was added to the crude peptide solid. After a white precipitate was formed, the mixture was sonicated for 30 seconds, centrifuged at 1500 rpm for 5 minutes, the supernatant was discarded, and the precipitate was dried with nitrogen. The precipitate was dissolved in 20% acetonitrile / water, purified by HPLC, and then freeze-dried to obtain a white powdery peptide precursor P1.

[0078] HPLC and mass spectra of peptide precursor P1 are shown below. Figure 1 As shown in A and 1B, the structure of the synthesized polypeptide precursor P1 is shown below:

[0079]

[0080] (15) Synthesis of the therapeutic drug PXC-110:

[0081]

[0082] The polypeptide precursor P1 (1 eq, 8.85 mg) was dissolved in 200 μL DMSO, and ICG-NHS (1 eq, 4.14 mg) and DIPEA (2 eq, 1.74 μL) were added. The mixture was reacted in the dark for 24 h. After purification by HPLC, the solid was freeze-dried to obtain a green powder, PXC-110.

[0083] HPLC and mass spectra of the therapeutic drug PXC-110 are shown below. Figure 2 As shown in A and 2B.

[0084] (16) Integrated diagnosis and treatment drugs [ 64 Preparation of Cu]PXC-110: 200 μg of peptide was dissolved in 200 μL of DMSO to prepare the stock solution, and then 20 μL of the stock solution was diluted with 80 μL of acetate-sodium acetate buffer (0.1 M, pH = 4.1). The diluted solution was then mixed with 100 μL of... 64 The CuCl2 solution (350-400 MBq, acetate-sodium acetate buffer, 0.1 M, pH 4.1) was mixed and reacted at 80 °C for 15 min. The radiolabeled peptide was purified using a Sep Pak C18 solid-phase extraction column. Its radiochemical purity was analyzed by radioactive RP-HPLC.

[0085] Integrated diagnosis and treatment drugs 64 The RP-HPLC analysis results of Cu]PXC-110 are as follows: Figure 3 As shown.

[0086] Example 2: Fluorescence imaging results of the therapeutic drug PXC-110 in a PC-3 tumor-bearing NCG mouse model

[0087] Severely immunodeficient mice (NCG mice) were used as the recipients of cancer cell transplantation. Mice were inoculated with PC-3 cells starting at 4-6 weeks of age. The specific procedure was as follows: Cells were prepared, and Ham's F12K medium:Matrix gel = 1:1 was added, mixed well, and placed on ice. The cells were then inoculated at a rate of 1.0 × 10⁻⁶ cells / year. 7 200 μL / cell per mouse was injected subcutaneously into the axillary region of mice, resulting in an average tumor volume of 300-400 mm². 3 Fluorescence imaging experiments were performed at that time.

[0088] PXC-110 was dissolved in PBS and administered intravenously at a dose of 100 μg / 200 μL per animal. Fluorescence imaging was performed at 1, 2, 4, 6, 8, 12, and 24 hours post-administration. Images were processed using in vivo fluorescence software. The results are shown below. Figure 4 As shown in the figure. Six hours after drug injection, mice were euthanized by cervical dislocation under anesthesia. Dissection was performed, and tumor tissue, heart, liver, spleen, lungs, kidneys, pancreas, small intestine, muscle, and brain were collected. In vivo fluorescence imaging was used for fluorescence imaging, and fluorescence quantification was performed on the major organs. The results are shown in the figure. Figure 5 As shown.

[0089] Example 3: [ 64 PET / CT imaging of Cu]PXC-110 in PC-3 tumor-bearing NCG mouse model

[0090] Will[ 64 Cu]PXC-110 was administered intravenously to PC-3 tumor-bearing NCG mouse models at a dose of 7.4 MBq / 200 μL / mouse. PET / CT imaging was performed at 1, 4, 8, and 12 hours post-administration. The PET / CT imaging results are as follows: Figure 6 As shown. The results indicate that [ 64 Cu]PXC-110 exhibits significant tumor uptake in the PC-3 tumor-bearing NCG mouse model.

[0091] Example 4: Photodynamic therapy with PXC-110 in a PC-3 tumor-bearing NCG mouse model

[0092] PC-3 tumor-bearing NCG mouse models were administered PBS (200 μL / mouse), ICG (33 μg / mouse), and PXC-110 (100 μg / mouse) intravenously. Six hours after administration, the tumor sites of the mice were irradiated with an 808 nm laser for 8 minutes at a power of 1.5 W / cm². 2 The medication was administered every three days for a total of five times. Tumor volume and body weight were measured every two days. The mouse tumor volume was calculated using the formula: length * width * width / 2, and plotted as a line graph. The results are shown below. Figure 7 and Figure 8 As shown in the results, PXC-110 exhibits significant tumor growth inhibition under 808nm laser irradiation and demonstrates good biocompatibility.

Claims

1. A therapeutic polypeptide as shown in formula (Ⅰ) or a pharmaceutically acceptable salt thereof, wherein: P is the GRPR targeting peptide, C is the linker, T is the photodynamic group with fluorescence function, and D is the radionuclide chelating group; The amino acid sequence of the GRPR-targeting peptide is: (D-Phe)-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2. Or a polypeptide sequence or polypeptide derivative that has 1 to 3 amino acids replaced, deleted, or inserted on (D-Phe)-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2.

2. The polypeptide-based therapeutic agent according to claim 1, or a pharmaceutically acceptable salt thereof, wherein the linker is selected from one of the following compounds: Unmodified or modified monoamino acids, unmodified or modified amino acid chains of different lengths, and alkyl chains with or without O and N heteroatoms, such as: m, n, and p are each independently selected from integers between 0 and 20; t are each independently selected from integers between 1 and 150; r is an integer independently selected from 0 to 150.

3. The polypeptide therapeutic agent according to claim 1 or its pharmaceutically acceptable salt, wherein the photodynamic group with fluorescent function is selected from one of IR-780, IR-783, IR-808, IR-825, indocyanine green, protoporphyrin IX, and rose red.

4. The photodynamic group with fluorescent function according to claim 3, characterized in that... IR-780, IR-783, IR-808, IR-825, and indocyanine green were used for photodynamic therapy under an 808nm laser; protoporphyrin IX was used for photodynamic therapy under a 630-635nm laser; and rose red was used for photodynamic therapy under a 560nm laser.

5. The polypeptide-based therapeutic agent according to claim 1, or a pharmaceutically acceptable salt thereof, wherein the radionuclide chelating group is selected from one of the following compounds:

6. A method for preparing a polypeptide-based therapeutic drug or a pharmaceutically acceptable salt thereof as described in any one of claims 1-5, characterized in that, Includes the following steps: Synthesize the polypeptide compound; The polypeptide compound is coupled with the linker arm, the photodynamic group with fluorescent function, and the chelating group to obtain the crude product of the polypeptide-based diagnostic and therapeutic drug. The crude product of the labeled precursor is purified to obtain the polypeptide-based therapeutic drug.

7. The radiopeptide-based therapeutic drug obtained by chelating the peptide-based therapeutic drug according to any one of claims 1-6 with a radionuclide, or a pharmaceutically acceptable salt thereof, wherein the radionuclide is selected from... 18 F, 64 Cu、 67 Cu、 68 Ga、 89 Zr、 99m Tc, 111 In、 177 Lu、 186 Re、 188 Re、 203 Pb, 212 Pb, 213 Bi、 225 Ac、 227 Th.

8. A method for preparing a radiopeptide-based therapeutic drug as described in claim 7, or a pharmaceutically acceptable salt thereof, characterized in that, The preparation method includes the following steps: The labeled precursor is mixed with a radionuclide solution and incubated to prepare the radiopeptide-based diagnostic and therapeutic drug; the incubation conditions are 20-110℃ for 5-60 minutes.

9. A pharmaceutical composition or pharmaceutical preparation comprising a polypeptide-based therapeutic drug according to any one of claims 1-6 or a radioactive polypeptide-based therapeutic drug according to any one of claims 7 and 8, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier and / or excipient.

10. The use of the polypeptide-based therapeutic medicine according to any one of claims 1-6, or the radiopeptide-based therapeutic medicine according to any one of claims 7 and 8, or a pharmaceutically acceptable salt thereof, or the composition according to claim 8, in the diagnosis, prevention, and / or treatment of diseases that overexpress GRPR, preferably the diseases that overexpress GRPR are selected from cancers, more preferably breast cancer, uterine cancer, cervical cancer, ovarian cancer, or prostate cancer.

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