Functional polypeptide with ovarian cancer metastasis resisting effect and preparation method
By using iTRAQ-8 labeling combined with LC-MS/MS peptomics to screen and validate the peptide pAHSG, the lack of targeted drugs for omental metastasis of ovarian cancer was solved. This enabled efficient screening and validation of peptides with anti-ovarian cancer metastasis effects, which have significant anti-proliferation and anti-invasive capabilities and are suitable for high-throughput peptide drug screening systems.
Patent Information
- Application Number
- CN202510904874.8
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-07-01
- Publication Date
- 2025-12-05
AI Technical Summary
Current technologies lack effective peptide drugs that target omental metastasis in ovarian cancer, and the identification and intervention mechanisms of peptide drugs in the process of ovarian cancer metastasis are unclear, resulting in limited treatment efficacy.
The peptide pAHSG was screened using iTRAQ-8 labeling combined with LC-MS/MS peptidomics. The peptide pAHSG was synthesized and its anti-proliferation and anti-invasion abilities in ovarian cancer cells were verified. It was combined with 40 target protein pathways. The preparation methods included solid-phase synthesis, and the purity was higher than 95%, which is suitable for drug translation research.
It significantly improves the scope and efficiency of peptide screening. pAHSG at a concentration of 100 μM significantly reduces the activity of ovarian cancer cells and inhibits cell invasion, demonstrating clear functional advantages and mechanistic basis, and is suitable for high-throughput peptide drug screening systems.
Smart Images
Figure CN121064288A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the technical field of tumor biology and peptide drug research, and particularly relates to a functional polypeptide with anti-ovarian cancer metastasis effect and a preparation method. BACKGROUND
[0002] At present, the conventional treatment methods for ovarian cancer mainly include surgical operation, radiotherapy and platinum-based chemotherapy regimen. These methods have certain curative effects in the early stage of the disease, but for patients with advanced or metastatic ovarian cancer, the treatment effect is often limited. Among them, the gradual development of chemotherapy resistance has become one of the important reasons for treatment failure. Studies have shown that tumor cells can escape from drugs by activating alternative signaling pathways after long-term chemotherapy, thereby leading to decreased efficacy or even recurrence. In addition, the traditional treatment process has a long cycle, large toxic and side effects, and lacks selectivity for normal cells, which further limits its wide application.
[0003] In recent years, with the progress of peptide drug synthesis and delivery technology, researchers have gradually paid attention to the potential value of peptides in tumor treatment. As a class of highly biocompatible and targeted therapeutic agents, peptide molecules have shown good performance in tumor-specific recognition, cell pathway regulation, etc. A variety of peptide-drug conjugates have entered clinical trials, trying to optimize the pharmacological properties of traditional drugs by improving targeting and metabolic stability.
[0004] However, most of the existing research on peptide anti-tumor drugs is focused on the intervention of primary lesions, and the mechanism of peptide molecular recognition and intervention for tumor metastasis is still unclear. In particular, in the process of ovarian cancer peritoneal metastasis, there is still a lack of systematic polypeptide analysis and functional verification of ovarian cancer omentum metastasis, resulting in the absence of a clear peptide drug targeting the metastasis process in clinical practice. This situation is mainly due to the complexity of omental metastasis mechanism, multi-factor regulation and the difficulty of identifying related molecular markers in metastatic tissues. Therefore, how to screen and verify functional peptides with regulatory potential from metastatic lesions has become a key problem that needs to be broken through in the existing technology.
[0005] The current treatment for ovarian cancer mainly focuses on primary tumor control, lacks effective prevention and inhibition means for peritoneal cavity metastasis such as omentum, especially lacks molecular tools targeting metastasis mechanism, and although peptide drugs have shown good therapeutic potential in other cancers, functional peptides for omentum metastasis of ovarian cancer have not been systematically identified and developed, thereby limiting the application of peptides in this field, and the prior art lacks a peptideomics-based systematic screening platform, which is difficult to identify differentially expressed polypeptides with intervention significance from metastatic tissues, and the existing polypeptide function research also generally lacks evidence of the functional mechanism of the candidate polypeptides in tumor metastasis core links such as cell cycle regulation, migration and invasion, which cannot support the feasibility of the polypeptides as therapeutic targets.
[0006] Therefore, in order to solve the above problems, the present application provides a functional polypeptide with anti-ovarian cancer metastasis effect and a preparation method. SUMMARY
[0007] In order to overcome the problem that the peptide molecular recognition and intervention mechanism of the existing tumor metastasis link is still unclear, the present application provides a functional polypeptide with anti-ovarian cancer metastasis effect and a preparation method.
[0008] The technical scheme of the present application is: a functional polypeptide with anti-ovarian cancer metastasis effect, comprising: the polypeptide is a synthetic polypeptide pAHSG, the amino acid sequence of the synthetic polypeptide is MGVVSLGSPSGEVSHPRKT, the molecular weight is 1925.19 g / mol, and the isoelectric point is 10.043.
[0009] As a preferred, the in vitro half-life of the polypeptide in mammalian reticulocytes is 30 hours, the in vivo half-life in yeast cells is greater than 20 hours, and the in vivo half-life in Escherichia coli is greater than 10 hours.
[0010] As a preferred, a preparation method of a functional polypeptide with anti-ovarian cancer metastasis effect, comprising the following steps:
[0011] S1, weigh 3g of resin in a 150ml reactor, soak with 50ml of dichloromethane;
[0012] S2, wash the resin with 3 times the volume of the resin nitrogen-dimethylformamide, then dry, repeat four times, and dry the resin before use;
[0013] S3, weigh the first amino acid threonine Thr at the C-terminal and 1-hydroxy-benzotriazole in a 50ml centrifuge tube, dissolve in 20ml of DMF, then add 3ml of N,N-diisopropyl carbodiimide, shake well for 1 minute, then add to the reactor, and add 3 times the molar amount of DMAP, and place the reactor in a 30℃ shaking bed for reaction;
[0014] S4, four hours later, acetic anhydride was added to seal the head for half an hour, then washed with 3 times the volume of DMF four times, and dried for use;
[0015] S5, 20% piperidine was added to the reactor to remove the Fmoc protecting group, and the reactor was placed on a decolorizing shaker for 20 min. After the Fmoc protecting group on the resin was removed, the resin was washed with DMF four times, and then dried;
[0016] S6, a small amount of resin was taken for ninhydrin detection, and the resin had color, indicating that the deprotection was successful;
[0017] S7, the second amino acid lysine Lys and HOBT were weighed into a 50 ml centrifuge tube, 25 ml of DMF was added to dissolve them, then 2.5 ml of DIC was added and shaken for 1 min, and after the solution was clear, it was added to the reactor, and then the reactor was placed in a 30°C shaker for reaction;
[0018] S8, after 1 hour, a small amount of resin was taken for detection, and the resin was colorless, indicating that the reaction was complete; if the resin had color, the condensation was not complete, and the reaction was continued;
[0019] S9, after the reaction was complete, the resin was washed with DMF four times, then dried, a certain amount of 20% piperidine was added to the reactor, placed on a decolorizing shaker for 20 min, the Fmoc protecting group on the resin was removed, washed with DMF four times, then dried, and the protection was detected to see if it was removed;
[0020] S10, the following amino acids RPHSVEGSPSGLSVVGM were sequentially coupled according to steps S7-S9;
[0021] S11, after the Fmoc of Arg was removed, the resin was washed with DMF four times, then dried, the polypeptide was cut from the resin with 95 cleavage solution, and the polypeptide was centrifuged with ice ethyl ether four times, finally purified by HPLC, and then freeze-dried to obtain a polypeptide with a certain purity.
[0022] As a preferred, the soaking time of 50 ml of dichloromethane in step S1 is 2 hours.
[0023] As a preferred, the acetic anhydride seal head consists of the following mass percentages: acetic anhydride: DIEA: DCM = 1:1:2.
[0024] As a preferred, the piperidine consists of the following mass percentages: piperidine: DMF = 1:4.
[0025] As a preferred, the 95 cleavage solution in step S11 consists of the following mass percentages: trifluoroacetic acid: ethanedithiol: isopropylsilane: water = 95:2:2:1.
[0026] As preferred, the amount of the cutting solution in step S11 is 10 ml per gram of resin.
[0027] As preferred, the ninhydrin detection step comprises: placing two drops of the ninhydrin solution on the resin, and reacting for 1 min in an environment of 100°C.
[0028] As preferred, the diethyl ether in step S11 is composed of the following mass percentage: cutting solution: diethyl ether = 1:9.
[0029] The beneficial effects of the present application are:
[0030] 1. The present application adopts iTRAQ-8 labeling combined with LC-MS / MS polypeptidomics method, which has the advantages of higher throughput and stronger quantitative precision than the traditional proteomics or single gene screening strategy. In one experiment, more than 2500 polypeptides can be identified, thereby significantly improving the screening range and efficiency of candidate functional molecules. Compared with 1-2 molecules analyzed by traditional Western blot or immunohistochemical analysis, the present method can screen hundreds of polypeptides with significant differences from the primary lesion at one time, and more than 98% of the polypeptide molecules have a molecular weight of less than 4 kDa, which is more suitable for development as drugs.
[0031] 2. The present application not only stops at the screening level, but also further synthesizes and verifies that the polypeptide pAHSG has significant anti-proliferation and anti-invasion ability in ovarian cancer cells. pAHSG can significantly reduce the activity of SKOV3 and HeyA8 cells at a concentration of 100 μM, and concentration-dependently inhibit cell invasion, having a clear functional advantage.
[0032] 3. pAHSG binds to 40 target proteins, involving pathways such as aminoacyl tRNA synthesis, protein transport, and mRNA metabolism, and has a clear mechanism basis.
[0033] 4. The polypeptide sequence is based on patient tissue sample screening, has a clear source and biological relevance, is prepared by solid-phase synthesis method, and has a synthesis purity higher than 95%, which is suitable for subsequent drug conversion research.
[0034] 5. The present application adopts standardized processes from tissue processing, peptide extraction, purification to mass spectrometry analysis, which can be repeatedly applied in clinical samples; the cell experiment system is complete, which is convenient for efficacy verification, and the overall process is suitable for establishing a technical platform, which is suitable for high-throughput polypeptide drug screening system. BRIEF DESCRIPTION OF DRAWINGS
[0035] Figure 1 A 2D structure schematic diagram of the synthetic polypeptide pAHSG of the present application is shown.
[0036] Figure 2The functional experiment schematic diagram of the synthetic polypeptide pAHSG of the application inhibiting proliferation of ovarian cancer cells is shown.
[0037] Figure 3 The functional experiment schematic diagram of the synthetic polypeptide pAHSG of the application inhibiting invasion of ovarian cancer cells is shown.
[0038] Figure 4 The functional experiment schematic diagram of the synthetic polypeptide pAHSG of the application regulating cell cycle of ovarian cancer cells is shown. DETAILED DESCRIPTION
[0039] In order to make the objects, technical solutions and advantages of the embodiments of the application clearer, the technical solutions in the embodiments of the application will be described clearly and completely below with reference to the drawings in the embodiments of the application. Obviously, the described embodiments are some but not all of the embodiments of the application. Based on the embodiments in the application, all other embodiments obtained by a person of ordinary skill in the art without creative work fall within the protection scope of the application.
[0040] Please refer to Figure 1 The application provides an embodiment of a functional polypeptide with anti-ovarian cancer metastasis effect, which comprises that the polypeptide is a synthetic polypeptide pAHSG, the amino acid sequence of the synthetic polypeptide is MGVVSLGSPSGEVSHPRKT, the molecular weight is 1925.19 g / mol, and the isoelectric point is 10.043.
[0041] Further, the application provides an embodiment of a preparation method of the functional polypeptide with anti-ovarian cancer metastasis effect, which specifically comprises the following steps.
[0042] Firstly, 3 g of resin (degree of substitution 0.3 mmol / g) is weighed in a 150 ml reactor and soaked with 50 ml of dichloromethane;
[0043] After 2 hours, the resin is washed with 3 times the volume of nitrogen-dimethylformamide (DMF) and then dried, and the process is repeated four times; after the resin is dried, it is used;
[0044] The C-terminal first amino acid threonine Thr and 1-hydroxy-benzotriazole (HOBT) are weighed in a 50 ml centrifuge tube, 20 ml of DMF is added for dissolution, 3 ml of N,N-diisopropylcarbodiimide (DIC) is added, oscillation is uniformly shaken for 1 minute, after clarification, the reactor is added, and 3 times the molar amount of DMAP is added, and the reactor is placed in a 30℃ shaking bed for reaction;
[0045] After four hours, the head is sealed with acetic anhydride (acetic anhydride: DIEA: DCM = 1:1:2) for half an hour, then washed with 3 times the volume of DMF four times, and dried for use;
[0046] A certain amount of 20% piperidine (piperidine / DMF = 1:4) was added to the reactor, and it was shaken on the decolorizing shaker for 20 min to remove the Fmoc protecting group on the resin. After deprotection, the resin was washed four times with DMF, and then dried by suction;
[0047] A small amount of resin was taken for ninhydrin (Kikuo hydrindrin) detection, and the resin had color, indicating that the deprotection was successful;
[0048] The second amino acid lysine Lys and HOBT were weighed into a 50 ml centrifuge tube, 25 ml of DMF was added to dissolve them, then 2.5 ml of DIC was added and shaken uniformly for 1 min, and after the solution was clear, it was added to the reactor, and then the reactor was placed in a 30℃ shaker for reaction;
[0049] After 1 hour, a small amount of resin was taken for detection, and the ninhydrin method was used for detection. If the resin is colorless, it indicates that the reaction is complete; if the resin has color, it indicates that the condensation is not complete, and the reaction continues;
[0050] After the reaction was complete, the resin was washed four times with DMF, and then dried by suction. A certain amount of 20% piperidine (piperidine / DMF = 1:4) was added to the reactor, and it was shaken on the decolorizing shaker for 20 min to remove the Fmoc protecting group on the resin. After deprotection, the resin was washed four times with DMF, and then dried by suction to detect whether the protection was removed;
[0051] According to steps S7-S9, the subsequent amino acids RPHSVEGSPSGLSVVGM were sequentially connected;
[0052] After removing the Fmoc of Arg, the resin was washed four times with DMF, then dried by suction with methanol, the polypeptide was cut from the resin with 95 cleavage solution (trifluoroacetic acid: ethanedithiol: isopropylsilane: water = 95:2:2:1), and the polypeptide was centrifuged four times with ice ethyl ether (cleavage solution: ethyl ether = 1:9). Finally, HPLC separation and purification were performed, and then freeze-drying was performed to obtain a polypeptide with a certain purity.
[0053] Further, the present application provides an embodiment, screening of a polypeptide:
[0054] This example aims to screen differentially expressed polypeptide molecules from primary cancer tissues, omental metastasis tissues and normal omentum tissues of ovarian cancer patients, and the experimental steps are as follows:
[0055] First, 3 patients diagnosed as high-grade serous ovarian cancer by pathology were selected, and after approval by the ethics committee (number: 2022KY-051) and obtaining informed consent, their primary cancer tissues, omental metastasis tissues and normal omentum tissues were collected, and the samples were immediately frozen and stored at -80℃ for standby.
[0056] The tissue sample was ground into powder in liquid nitrogen, added with lysis solution for ultrasonic disruption, and after centrifugation, the supernatant was taken, impurities were removed by acetonitrile precipitation method, and after freeze-drying, the sample was dialyzed using a 10 kDa ultrafiltration tube to obtain a crude peptide sample.
[0057] After the crude peptide sample was purified by a C18 column, the iTRAQ-8 labeling technology was used to label the peptide segments to distinguish the tissue samples of different sources (primary focus, metastatic focus, and normal tissue), and after labeling, the peptide segments were fractionated by high performance liquid chromatography (HPLC), desalted and concentrated.
[0058] The TripleTOF 5600+ mass spectrometer combined with a nanoliter level liquid phase system was used to perform LC-MS / MS analysis on the sample to obtain polypeptide sequences and relative quantitative information, and the ProteinPilot V4.5 software was used to identify proteins and peptide segments, and finally, the differentially expressed polypeptides significantly up-regulated or down-regulated in the greater omentum metastatic tissue were screened out.
[0059] The test results show that through mass spectrometric analysis, the expression level of the polypeptide pAHSG (sequence: MGVVSLGSPSGEVSHPRKT) in the greater omentum metastatic tissue is significantly lower than that in the primary cancer focus and normal tissue as shown in Table 1, which is specifically manifested as:
[0060] The ratio of the greater omentum metastatic cancer focus to the primary cancer focus is 0.198 (*p* = 0.001), indicating that pAHSG is significantly down-regulated in the metastatic focus.
[0061] The ratio of the greater omentum metastatic cancer focus to the normal tissue is 0.098 (*p* = 0.001), further verifying the significance of the expression difference.
[0062] Table 1 Down-regulated expression level of pAHSG polypeptide in greater omentum metastatic tissue of ovarian cancer
[0063]
[0064] Further, the present application provides an embodiment, the anti-tumor proliferation ability of the synthesized polypeptide pAHSG:
[0065] SKOV3 and HeyA8 ovarian cancer cells were inoculated in a 96-well plate at a density of 1 × 10 3 μM, 50 μM and 100 μM, respectively, and after 24 hours of treatment, the absorbance value at 450 nm was measured by an enzyme marker at 0, 24, 48 and 72 hours, and the cell proliferation ability was evaluated by CCK-8 method and EdU method.
[0066] Please refer to Figure 2The experimental results show that the pAHSG can significantly inhibit the proliferation of ovarian cancer cells in a dose-dependent manner in a certain concentration range, indicating that the polypeptide has potential anti-tumor activity.
[0067] Further, the present application provides an embodiment of the anti-tumor invasion ability of the synthetic polypeptide pAHSG.
[0068] First, the Transwell membrane is coated with Matrigel to simulate the extracellular matrix barrier, and SKOV3 and HeyA8 ovarian cancer cells are inoculated on the upper chamber of the Transwell at a density of 30,000 cells per well, and the final concentration of the synthetic polypeptide pAHSG is added to 10 μM, 50 μM and 100 μM. After the cells pass through the membrane to a certain number, 4% paraformaldehyde is used for fixation, then 0.5% crystal violet solution is used for staining, and finally, 5 fields of view are randomly selected under an optical microscope for counting to count the migration and invasion of cells.
[0069] Please refer to Figure 3 The experimental results show that in the Transwell chamber experiment, the final concentration of the candidate synthetic polypeptide pAHSG is added to 10 μM, 50 μM and 100 μM in the upper chamber to detect the invasion ability of ovarian cancer cells, and the experimental results show that with the increase of the concentration of pAHSG, the number of invasive cells crossing the Transwell membrane shows a significant decreasing trend, showing a typical dose-dependent inhibition effect. The above results show that pAHSG can stably inhibit the membrane crossing ability of ovarian cancer cells in an effective concentration range, suggesting that it has the potential to inhibit tumor cell invasion and metastasis, and provides experimental basis for its use as an anti-ovarian cancer metastasis candidate polypeptide.
[0070] Further, the present application provides an embodiment of the tumor cell cycle regulation ability of the synthetic polypeptide pAHSG.
[0071] After treating ovarian cancer cells with different concentrations of polypeptide pAHSG for 24 hours, the cells are digested using trypsin without ethylenediaminetetraacetic acid (EDTA) to avoid damage to the integrity of the cell membrane. Subsequently, the cells are stained with PI and Annexin V-FITC, and after treatment, the flow cytometry is used to detect the apoptosis and cell cycle of the cells.
[0072] Please refer to Figure 4 The flow cytometry results show that pAHSG has a significant effect on the cell cycle after treatment with different concentrations of pAHSG, the proportion of G1 phase cells is significantly increased, and the proportion of S phase cells is significantly decreased, and shows a dose-dependent trend, which indicates that pAHSG can inhibit DNA synthesis and proliferation of ovarian cancer cells by blocking G1 phase, and further verifies its anti-tumor activity.
[0073] Further, the application provides an embodiment of the mechanism of synthesizing the polypeptide pAHSG:
[0074] The pull-down system is constructed by using the biotin-labeled pAHSG and streptavidin magnetic beads, and after incubation with the ovarian cancer cell lysate and washing, the silver staining method is used to identify the protein band, and mass spectrometry is performed on the target band to identify the protein.
[0075] The experimental results show that 40 proteins interacting with pAHSG are confirmed, and the GO function annotation analysis of the 40 proteins shows that the molecular functions thereof are mainly related to tRNA aminoacylation, intracellular protein transport, and mRNA metabolic process. In terms of cell components, these proteins are mainly located in lysosomal membranes, organellar outer membranes, and transport complexes. In terms of biological processes, they are related to ligase activity, GTP binding, proton transmembrane transport activity, etc. KEGG pathway analysis further shows that these proteins are mainly involved in aminoacyl-tRNA biosynthesis and various cancer-related signaling pathways.
[0076] Through the above steps, the iTRAQ-8 labeling combined with LC-MS / MS polypeptidomics method is used to identify more than 2500 polypeptides in one experiment, thereby significantly improving the screening range and efficiency of candidate functional molecules. The method can screen hundreds of polypeptides with significant differences from the primary lesion at one time, and more than 98% of the polypeptide molecules have a molecular weight of less than 4 kDa, which is more suitable for development as drugs to solve the problem that the peptide molecular recognition and intervention mechanism in the tumor metastasis link is still unclear.
[0077] The above is only a preferred embodiment of the present application, and is not intended to limit the application in other forms. Any person skilled in the art can use the disclosed technical content to make changes or modifications to equivalent embodiments applied to other fields, but any simple modification, equivalent change and modification made in accordance with the technical essence of the present application to the above embodiments shall still fall within the protection scope of the present application.
Claims
1. A functional polypeptide having an anti-metastatic effect on ovarian cancer, characterized in that, The polypeptide is a synthetic polypeptide pAHSG, the amino acid sequence of the synthetic polypeptide is MGVVSLGSPSGEVSHPRKT, the molecular weight is 1925.19 g / mol, and the isoelectric point is 10.
043.
2. The functional polypeptide with anti-ovarian cancer metastasis effect according to claim 1, characterized in that: The in-vitro half-life of the polypeptide in mammalian reticulocytes is 30 hours, the in-vivo half-life in yeast cells is greater than 20 hours, and the in-vivo half-life in Escherichia coli is greater than 10 hours.
3. A method for preparing a functional polypeptide having an anti-metastatic effect on ovarian cancer, characterized by The method comprises the following steps: S1, 3g of resin is weighed in a 150ml reactor, soaked with 50ml of dichloromethane; S2, the resin is washed with 3 times the volume of nitrogen-dimethylformamide, then dried, repeated four times, and the resin is dried and used; S3, the C-terminal first amino acid threonine Thr and 1-hydroxy-benzotriazole are weighed in a 50ml centrifuge tube, dissolved in 20ml of DMF, then 3ml of N,N-diisopropyl carbodiimide is added, shaken for 1 minute, then added to the reactor, and 3 times the molar amount of DMAP is added, and the reactor is placed in a 30℃ shaking bed for reaction; S4, after four hours, acetic anhydride is added for half an hour, then washed with 3 times the volume of DMF for four times, and dried for use; S5, 20% piperidine is added to the reactor to remove the Fmoc protecting group, placed on a decolorizing shaking bed for 20min, after the Fmoc protecting group on the resin is removed, washed with DMF for four times, and then dried; S6, a small amount of resin is taken for detection by the ninhydrin method, and the resin has color, indicating that the deprotection is successful; S7, the second amino acid lysine Lys and HOBT are weighed in a 50ml centrifuge tube, dissolved in 25ml of DMF, then 2.5ml of DIC is added and shaken for 1min, then added to the reactor, and the reactor is placed in a 30℃ shaking bed for reaction; S8, after 1 hour, a small amount of resin is taken for detection by the ninhydrin method, if the resin is colorless, it indicates that the reaction is complete; if the resin has color, it indicates that the condensation is not complete, and the reaction continues; S9, after the reaction is complete, the resin is washed with DMF for four times, then dried, a certain amount of 20% piperidine is added to the reactor, placed on a decolorizing shaking bed for 20min, after the Fmoc protecting group on the resin is removed, washed with DMF for four times, and then dried to detect whether the protection is removed; S10, the following amino acids RPHSVEGSPSGLSVVGM are sequentially connected according to steps S7-S9; S11, after the Fmoc of Arg is removed, washed with DMF for four times, then the resin is dried with methanol, the polypeptide is cut from the resin with 95 cleavage solution, and the polypeptide is centrifuged and precipitated four times with ice ethyl ether, finally separated and purified by HPLC, then freeze-dried to obtain a polypeptide with a certain purity.
4. The method for preparing a functional polypeptide with anti-ovarian cancer metastasis effect according to claim 3, characterized in that: The soaking time of 50ml of dichloromethane in step S1 is 2 hours.
5. The method for preparing a functional polypeptide with anti-ovarian cancer metastasis effect according to claim 3, characterized in that, The acetic anhydride cap is composed of the following mass percentages: acetic anhydride: DIEA: DCM = 1:1:
2.
6. The method for preparing a functional polypeptide with anti-ovarian cancer metastasis effect according to claim 3, characterized in that, The piperidine is composed of the following mass percentages: piperidine: DMF = 1:
4.
7. The method for preparing a functional polypeptide with anti-ovarian cancer metastasis effect according to claim 3, characterized in that, The cutting solution in step S11 is composed of the following mass percentages: trifluoroacetic acid: ethanedithiol: isopropylsilane: water = 95:2:2:
1.
8. The method for preparing a functional polypeptide with anti-ovarian cancer metastasis effect according to claim 3, characterized in that: The amount of the cutting solution in step S11 is 10 ml per gram of resin.
9. The method for preparing a functional polypeptide with anti-ovarian cancer metastasis effect according to claim 3, characterized in that, The ninhydrin detection step includes: placing two drops of ninhydrin solution on the resin, and reacting at 100°C for 1 min.
10. A method for preparing a functional polypeptide with anti-ovarian cancer metastasis activity according to claim 3, characterized in that, The ethyl ether in step S11 is composed of the following mass percentages: cutting solution: ethyl ether = 1:9.