Active peptide for senescent cells and method for preparing the same

Abstract

The application belongs to the technical field of functional polypeptides, and particularly relates to an active peptide for senescent cells and a preparation method thereof, wherein the active peptide is obtained by molecular docking simulation screening and optimization with a helical structure segment in an IL-6R protein combined with a gp130 protein as a polypeptide template; the active peptide after functional optimization contains an FF unit driving nano self-assembly, a GFLG unit capable of being specifically cut by a lysosome protease highly expressed in a microenvironment of senescent cells, and an RYFQKWEKLRNQ unit generating high affinity with an IL-6 / IL-6R / gp130 complex; the designed active peptide itself has anti-aging activity and can also be used for synergistic delivery of drugs, and has a broad application prospect.

Description

Technical Field

[0001] This invention belongs to the field of functional polypeptide technology, specifically relating to an active peptide for senescent cells and its preparation method. Background Technology

[0002] Cellular senescence is a stable state of cell cycle arrest that occurs when cells, under the influence of factors such as stress, damage, or replication exhaustion, are accompanied by phenotypic changes such as metabolic reprogramming, persistent activation of DNA damage responses, mitochondrial dysfunction, and enhanced inflammatory signaling. Senescent cells secrete various pro-inflammatory cytokines, chemokines, and proteases, forming SASP (senescence-associated secretory phenotype), including but not limited to IL-6, IL-8, CCL2, and MMPs. SASP can amplify inflammation through autocrine and paracrine processes, induce cellular senescence, disrupt tissue homeostasis, and promote fibrosis, metabolic disorders, and the development of various age-related diseases. Therefore, precise intervention targeting key nodes in the formation, maintenance, and amplification of senescent cells is an important strategy for delaying aging and improving cellular degeneration caused by pathological factors such as stroke.

[0003] Current anti-aging interventions mainly focus on eliminating senescent cells or inhibiting SASP (Sensitive Aging Pathogens). While small-molecule drugs widely used in clinical practice are convenient to administer, they suffer from issues such as insufficient targeting, systemic side effects, and long-term safety concerns. Antibody drugs have high targeting specificity, but their preparation costs are high, and their in vivo distribution and tissue penetration are limited. Peptides, on the other hand, possess advantages such as designable structures, precisely regulated mechanisms of action, and relatively good tissue penetration, making them promising for specific antagonism of aging maintenance pathways. Furthermore, due to the presence of non-covalent interactions such as hydrophobicity, hydrogen bonding, and conjugation, peptides also have the potential for self-assembly and drug loading, exhibiting both activity and functionality. This could provide a novel and industrially viable technological pathway for delaying aging and intervening in related diseases. Summary of the Invention

[0004] In response to the above situation, the present invention provides an active peptide for senescent cells and a method for preparing the same. The method selects protein targets by taking the key signal axis of SASP self-maintenance in senescent cells as the starting point, obtains antagonistic active peptides that can specifically bind to receptor proteins through molecular docking simulation, and optimizes their sequence and improves their functionality, thereby achieving the comprehensive effects of delaying cell senescence, improving the tissue microenvironment and enhancing repair capabilities.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0006] This invention provides an active peptide for senescent cells, the active peptide comprising the following raw materials: RinkAmide AM resin, Phe (phenylalanine F), Gly (glycine G), Leu (leucine L), Arg (arginine R), Gln (glutamine Q), Lys (lysine K), Tyr (tyrosine Y), Trp (tryptophan W), Glu (glutamate E), and Asn (asparagine N).

[0007] Furthermore, the mass ratio of Rink Amide AM resin, Phe, Gly, Leu, Arg, Gln, Lys, Tyr, Trp, Glu and Asn is 10:24.8:9.5:11.3:20.7:19.5:15:7.4:8.4:6.8:9.5.

[0008] Furthermore, the amino acid sequence of the active peptide is H2N-FFGFLGRYFQKWEKLRNQ, as shown in SEQ ID NO.9.

[0009] Furthermore, the active peptide was obtained by molecular docking simulation screening and optimization using the helical segment of IL-6R protein that binds to gp130 protein as a polypeptide template.

[0010] Furthermore, the specific methods for screening and optimizing the active peptides through molecular docking simulation are as follows:

[0011] S1: Based on the SASP conduction mechanism, the IL-6 / IL-6R / gp130 complex was selected as the intervention target. Its high-resolution crystal structure (PDB ID: 1P9M) was obtained by searching the protein structure database. The binding interface between IL-6 and the effector protein gp130, i.e. the Site II region, was analyzed. Molecular dynamics simulation and SASA (solvent accessible surface area) analysis were used to determine the hot spot region, i.e. the peptide template.

[0012] S2: Based on the polypeptide template selected in step S1, 8 candidate peptides were designed and numbered P1-P8 respectively, and their amino acid sequences are shown in SEQ ID NO.1-8 respectively;

[0013] S3: Bioinformatics tools were used to evaluate the physicochemical properties and predict the secondary structure of candidate peptides P1-P8, and the α-helix formation tendency of each candidate peptide in aqueous solution was analyzed. Candidate peptides P1, P2 and P6 with good amphiphilicity, strong stability and high helical tendency were screened out.

[0014] S4: The selected candidate peptides P1, P2 and P6 were subjected to computer molecular docking simulation with 1P9M to evaluate the binding conformation and binding ability of each candidate peptide with the IL-6 / IL-6R / gp130 complex, calculate the binding free energy, and screen again to obtain the candidate peptide P1 with the strongest binding force and the most stable conformation.

[0015] S5: The candidate peptide P1 obtained from screening was functionally sequenced and modified with the FFGFLG peptide at its N-terminus. The π-π stacking and hydrophobic interaction between the benzene rings in FF facilitates the self-assembly of the peptide into stable nanofibers or particles under physiological conditions, which can stabilize the conformation and properties of the peptide molecules in vivo and in vitro. The GFLG peptide can be specifically recognized and cleaved by lysosomal proteases highly expressed in the microenvironment of senescent cells, realizing the targeted and efficient release of active peptides or loaded drugs. The amino acid sequence of the active peptide for senescent cells was designed as H2N-FFGFLGRYFQKWEKLRNQ, as shown in SEQ ID NO.9.

[0016] Furthermore, the amino acid sequence of the candidate peptide P1 is H2N-RYFQKWEKLRNQ, as shown in SEQ ID NO.1.

[0017] Furthermore, the amino acid sequence of the candidate peptide P2 is H2N-WKRLYEHFQKRN, as shown in SEQ ID NO.2.

[0018] Furthermore, the amino acid sequence of the candidate peptide P3 is H2N-KWEYRFQNRLKK, as shown in SEQ ID NO.3.

[0019] Furthermore, the amino acid sequence of the candidate peptide P4 is H2N-RLFWKQEYNRKK, as shown in SEQ ID NO.4.

[0020] Furthermore, the amino acid sequence of the candidate peptide P5 is H2N-YWKRQEFNRLKK, as shown in SEQ ID NO.5.

[0021] Furthermore, the amino acid sequence of the candidate peptide P6 is H2N-KRWYQEFKLRNQ, as shown in SEQ ID NO.6.

[0022] Furthermore, the amino acid sequence of the candidate peptide P7 is H2N-CRYFQKWEKLRNC, as shown in SEQ ID NO.7, wherein the cysteine ​​C residues at both ends of the candidate peptide P7 can form disulfide bonds and thus undergo cyclization.

[0023] Furthermore, the amino acid sequence of the candidate peptide P8 is H2N-CWKRLYEHFQKRNC, as shown in SEQ ID NO.8.

[0024] This invention also provides a method for preparing bioactive peptides for senescent cells, the specific steps of which are as follows:

[0025] Step 1: Place Rink Amide AM resin in a peptide synthesis reactor, add DMF (N,N-dimethylformamide) to swell, filter to remove solvent, and then deprotect and wash with DMF to obtain deprotected resin;

[0026] Step 2: Following the amino acid sequence from C-terminus to N-terminus of the active peptide sequence H2N-FFGFLGRYFQKWEKLRNQ-COOH, the first amino acid Gln at the C-terminus is activated and added to the deprotected resin. The reaction is carried out with shaking to graft Gln(Q) into the deprotected resin. The reaction endpoint is determined by the ninhydrin method until no color development occurs, indicating successful grafting of the first amino acid. After deprotection and washing with DMF, the grafting reaction of the second amino acid is carried out. The remaining amino acids Phe, Gly, Leu, Arg, Lys, Tyr, Trp, Glu, and Asn are taken according to the active peptide sequence and the above activation, shaking reaction, ninhydrin color development, deprotection, and DMF washing process is repeated until the last amino acid Phe(F) is grafted, resulting in peptide resin.

[0027] Step 3: Wash the peptide resin alternately with DMF and DCM (dichloromethane) to carry out the cleavage reaction, cut the peptide off the resin, and remove the protecting groups of the amino acid side chains to obtain crude peptide.

[0028] Step 4: Precipitate the crude polypeptide using ice-cold ether at 4°C, centrifuge, discard the supernatant, and obtain the precipitate. The precipitate is then vacuum-dried to obtain the active peptide for senescent cells.

[0029] Furthermore, the deprotection process uses a DMF solution containing 20% ​​piperidine by volume.

[0030] Furthermore, the activation process in step 2 uses an activation solution, which is prepared by HBTU (O-benzotriazole-tetramethylurea hexafluorophosphate), DIPEA (N,N-diisopropylethylamine) and DMF in a mass ratio of 10.4:7.4:2.3.

[0031] Furthermore, the cutting reaction process described in step 3 uses a cutting fluid, which is prepared by mixing TFA (trifluoroacetic acid), Tis (triisopropylsilane), and deionized water in a mass ratio of 9.5:0.25:0.25.

[0032] The beneficial effects achieved by this invention are as follows:

[0033] The bioactive peptide prepared in this invention for senescent cells targets the SASP-self-maintained IL-6 / IL-6R / gp130 complex. By binding to the Site II hotspot region where IL-6R and gp130 bind, an antagonistic peptide P1 that mimics the helical structure of IL-6R is designed. This peptide specifically occupies the binding site of the gp130 receptor, competitively blocking downstream signal transduction induced by IL-6R, thereby achieving the reversal regulation of apoptosis and senescence. This not only effectively inhibits the secretion of pro-inflammatory factors by senescent cells but also prevents the spread of senescent phenotypes to surrounding healthy tissues, thus achieving the therapeutic effects of delaying organ aging and promoting damage repair.

[0034] Based on the sequence of the antagonistic peptide P1 obtained through screening, a functionalized bioactive peptide was further constructed. The introduction of bisphenylalanine (FF) and a cleavage linker arm (GFLG) enabled the bioactive peptide to undergo self-assembly, forming a stable nanostructure. This significantly reduced the risk of the peptide drug being easily cleaved by enzymes and having a short half-life in vivo, achieving long-term delivery. Furthermore, the bioactive peptide with the nanostructure can also serve as a carrier for co-delivery of drugs. Simultaneously, the GFLG peptide acts as a smart "switch," specifically recognizing and cleaving lysosomal proteases such as cathepsin B, which are highly expressed in the microenvironment of senescent cells, thus improving the efficiency of in-situ release of the bioactive peptide from senescent cells. Attached Figure Description

[0035] Figure 1 This is a schematic diagram of the structure of the active peptide for senescent cells in this invention;

[0036] Figure 2 The molecular structures of candidate peptides P1-P8 designed for Example 2;

[0037] Figure 3 The results of molecular docking simulations of candidate peptides P1, P2, P6, and the active peptide with the IL-6 / IL-6R / gp130 complex are shown.

[0038] Figure 4 The results of the microstructure analysis of the bioactive peptides prepared for senescent cells in Example 6;

[0039] Figure 5 The results of the anti-aging activity study of the active peptides prepared for senescent cells in Example 6 are presented. Detailed Implementation

[0040] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0041] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those familiar to those skilled in the art. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to this invention. The preferred embodiments and materials described herein are for illustrative purposes only and do not limit the scope of this application.

[0042] In the following embodiments, the structure of the active peptide for senescent cells is as follows: Figure 1 Unless otherwise specified, all methods are conventional; all parts are by weight; unless otherwise specified, all materials used in the following examples are new materials purchased from the market; all amino acids used are amino acids with the N-terminus protected by the Fmoc (9-fluorenylmethoxycarbonyl) group, commercially available products; the degree of substitution of the Rink Amide AM resin used is 0.44 mmol / g.

[0043] Example 1 (Target Analysis and Peptide Template Determination): Based on the core role of senescence-associated secretory phenotype (SASP) in maintaining cellular senescence, the IL-6 / IL-6R / gp130 complex was selected as the intervention target. By analyzing the crystal structure numbered 1P9M in the Protein Structure Database (PDB), the D-helix region of the IL-6R protein was identified as Site II at the interface with the extracellular domain of the effector protein gp130. Using molecular dynamics simulations and solvent accessibility surface area (SASA) analysis, it was determined that the core binding pocket on gp130 is mainly composed of residues such as Phe191, Val189, and Tyr190. Therefore, the corresponding D-helix segment in the IL-6R sequence was extracted as the original peptide template, and a candidate peptide library was designed through amino acid substitution.

[0044] Example 2 (Construction of Candidate Peptide Library): Based on the peptide template selected in Example 1, eight candidate peptides were designed, as follows: P1: H2N-RYFQKWEKLRNQ, as shown in SEQ ID NO.1; P2: H2N-WKRLYEHFQKRN, as shown in SEQ ID NO.2; P3: H2N-KWEYRFQNRLKK, as shown in SEQ ID NO.3; P4: H2N-RLFWKQEYNRKK, as shown in SEQ ID NO.4; P5: H2N-YWKRQEFNRLKK, as shown in SEQ ID NO.5; P6: H2N-KRWYQEFKLRNQ, as shown in SEQ ID NO.6; P7: H2N-CRYFQKWEKLRNC, as shown in SEQ ID NO.7; P8: H2N-CWKRLYEHFQKRNC, as shown in SEQ ID NO.4. As shown in IDNO.8, the cysteine ​​C residues at both ends of candidate peptides P7 and P8 can form disulfide bonds and are prone to cyclization, which is used to examine the binding affinity of cyclized peptides in the IL-6 / IL-6R / gp130 complex.

[0045] Example 3 (Initial Screening of Candidate Peptides): The physicochemical properties and secondary structures of candidate peptides P1-P8 were predicted using ExPASy ProtParam and PEP-FOLD3 tools. Since the 1P9M binding interface is a typical helical-groove binding mode, a higher helical percentage can significantly reduce entropy loss during binding, thereby improving affinity. The molecular weight, isoelectric point, GRAVY (average hydrophobicity), and α-helical tendency of the candidate peptides were obtained, and the results are shown in Table 1. Based on the peptide property prediction results, candidate peptides P1, P2, and P6 with isoelectric points in the non-physiological pH range, good hydrophilicity, and high degree of α-helicalization were screened.

[0046] Example 4 (Molecular docking simulation and binding ability evaluation): Using the AutoDock Vina tool, the candidate peptides P1, P2 and P6 obtained from the initial screening were subjected to molecular docking simulation with the IL-6 / IL-6R / gp130 complex, respectively. The binding free energy ΔG, van der Waals force energy, electrostatic interaction, number of hydrogen bonds and interfacial contact area were recorded. The results are shown in Table 2. The candidate peptide P1 with the lowest binding free energy, i.e. the most stable energy and the largest interfacial contact area, was selected as the template for the anti-aging active peptide sequence.

[0047] Example 5 (Functional Sequence Optimization and Self-Assembly Evaluation): To address the shortcomings of candidate peptide P1, such as easy degradation and difficulty in local enrichment under complex physiological environments, this example fused an FF-GFLG module peptide to the N-terminus of P1 to obtain the active peptide H2N-FFGFLGRYFQKWEKLRNQ, as shown in SEQ ID NO.9. Its binding ability, self-assembly ability, enzyme responsiveness, and physiological environment stability were evaluated simultaneously.

[0048] Example 6: This example provides an active peptide for senescent cells, the active peptide comprising the following raw materials in parts by weight: 1 part Rink Amide AM resin, 2.48 parts Phe, 0.95 parts Gly, 1.13 parts Leu, 2.07 parts Arg, 1.95 parts Gln, 1.5 parts Lys, 0.74 parts Tyr, 0.84 parts Trp, 0.68 parts Glu, and 0.95 parts Asn;

[0049] The Phe, Gly, Leu, Arg, Gln, Lys, Tyr, Trp, Glu, and Asn used are all amino acids protected by an N-terminal Fmoc group. Some of these amino acids have protecting groups on their side chains, specifically Fmoc-Phe-OH, Fmoc-Gly-OH, Fmoc-Leu-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Gln(Trt)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Trp(Boc)-OH, Fmoc-Glu(OtBu)-OH, and Fmoc-Asn(Trt)-OH.

[0050] The activation solution used was prepared from 62.4 parts HBTU, 44.4 parts DIPEA and 13.8 parts DMF;

[0051] The cutting fluid used was prepared from 9.5 parts TFA, 0.25 parts TiS and 0.25 parts deionized water;

[0052] This embodiment also provides a method for preparing active peptides for senescent cells, the specific steps of which are as follows:

[0053] Step 1: Weigh one part of Rink Amide AM resin and place it in a peptide synthesis reactor. Add 5 mL of DMF to swell for 20 min, filter to remove the solvent, then deprotect it with 5 mL of 20% piperidine DMF solution for 10 min, wash with 20 mL of DMF, and blow dry with a nitrogen pump to obtain the deprotected resin.

[0054] Step 2: Following the amino acid sequence from C-terminus to N-terminus of the active peptide sequence H2N-FFGFLGRYFQKWEKLRNQ-COOH, 0.975 parts of the first amino acid Gln from the C-terminus were dissolved in 5 mL of activation solution, activated for 2 min, and then added to the deprotected resin. The reaction was carried out with shaking at 100 rpm for 1.5 h. The reaction endpoint was determined by the ninhydrin method. If no color development was observed, the first amino acid Q was successfully grafted. The residue was then deprotected with 5 mL of 20% piperidine DMF solution for 10 min and 20%... After washing with mL of DMF, the second amino acid grafting reaction was carried out. Following the order of the active peptide amino acids, 0.95 parts of Asn, 1.035 parts of Arg, 0.565 parts of Leu, 0.75 parts of Lys, 0.68 parts of Glu, 0.84 parts of Trp, 0.75 parts of Lys, 0.975 parts of Gln, 0.62 parts of Phe, 0.74 parts of Tyr, 1.035 parts of Arg, 0.475 parts of Gly, 0.565 parts of Leu, 0.62 parts of Phe, 0.475 parts of Gly, 0.62 parts of Phe, and 0.62 parts of Phe were dissolved in 5 mL of activating solution, activated for 2 min, and then added to the deprotected resin. The above shaking reaction, ninhydrin color development, deprotection, and DMF washing process were repeated until the last amino acid, Phe(F), was grafted and deprotected, resulting in the peptide resin.

[0055] Step 3: Wash the peptide resin three times each with 5 mL DMF and 5 mL DCM, dry it with a nitrogen pump, add 10 parts of cutting solution and perform a cutting reaction for 30 min to cut the peptide off the resin and remove the protecting groups of the amino acid side chains to obtain crude peptide.

[0056] Step 4: Precipitate the crude peptide using ice-cold ether pre-cooled at 4°C. The volume ratio of ice-cold ether to crude peptide is 10:1. Centrifuge, discard the supernatant, and obtain the precipitate. The precipitate is then vacuum dried to obtain the active peptide for senescent cells.

[0057] Table 1 Physicochemical properties and α-helix structure of candidate peptides

[0058]

[0059] Table 2. Molecular docking simulation results parameters for candidate peptides P1, P2, and P6.

[0060]

[0061] Molecular structure analysis of candidate peptides: The secondary structures of candidate peptides P1-P8 were analyzed using the PEP-FOLD3 tool in the RPBS Web Portal. Results are shown below. Figure 2 .

[0062] Molecular docking simulation: The screened candidate peptides P1, P2, P6 and the active peptide were docked with the IL-6 / IL-6R / gp130 complex to investigate their structure and binding ability in protein binding. The results are shown in [Figure number missing]. Figure 3 .

[0063] Microscopic morphology characterization: 1 mg of the active peptide prepared in Example 6 was dissolved in 1 mL of deionized water, and its microscopic morphology was examined using transmission electron microscopy. The results are shown in the figure. Figure 4 .

[0064] Anti-aging activity assessment: A stroke-induced senescent cell model was constructed. SH-SY5Y cells were used as the in vitro efficacy evaluation subject. Cells were incubated with glucose-free Earle's balanced salt solution for 4 hours to achieve oxygen and glucose deprivation. Then, simulating clinical stroke ischemia-reperfusion, the oxygen-glucose-deprived cells were removed and replaced with normal culture medium for incubation to achieve oxygen and glucose enrichment. Culture was continued for 5 days to promote the senescence phenotype in SH-SY5Y cells. After the stroke-induced senescent cell model was constructed, 100 nM of the active peptide prepared in Example 6 and 100 nM of rapamycin (positive control drug) were administered as the active peptide group and positive control group, respectively. The control group and model group were treated with normal cells + normal culture medium and SH-SY5Y senescent cell model + normal culture medium, respectively. Cell apoptosis in each group was examined by flow cytometry. The results are shown in […]. Figure 5 .

[0065] The results in Tables 1 and 2 show that candidate peptide P1 has good water solubility, high helicity, and good affinity binding to the IL-6 / IL-6R / gp130 complex. Therefore, P1 was selected as the template peptide.

[0066] Figure 2 The results showed that candidate peptides P1-P6 exhibited a helical structure, which met the expected design goals. P7 showed a tendency to form a ring, and the thiol groups in the cysteine ​​residues at both ends of P7 were easily covalently linked to form a cyclic peptide. Although P8 also had cysteine ​​C residues at both ends, its helical conformation was more stable and its ring formation was relatively weak.

[0067] Figure 3 Molecular docking simulation results show that P1, P2 and P6 can form receptor-ligand structures at the protein-binding interface in the IL-6 / IL-6R / gp130 complex, all exhibiting a helical tendency. P1 has the highest degree of helicity and the largest binding area with the complex protein. The active peptide obtained after P1 sequence optimization also retains high binding capacity.

[0068] Figure 4 The results showed that the active peptides exhibited a long nanofiber morphology in aqueous solution, indicating that they had strong self-assembly capabilities and that the interior of the fibers could be used for drug loading to achieve synergistic delivery.

[0069] Figure 5 The results showed that the bioactive peptides had a good therapeutic effect on senescent cells caused by ischemia-reperfusion of nerve cells in stroke patients, and could significantly inhibit cell apoptosis and reverse the pathological cell senescence process.

[0070] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

[0071] The present invention and its embodiments have been described above. This description is not restrictive, and the accompanying drawings are only one embodiment of the present invention. The actual application is not limited to this. In conclusion, if those skilled in the art are inspired by this description and design similar methods and embodiments without departing from the spirit of the present invention, they should all fall within the protection scope of the present invention.

Claims

1. An active peptide for senescent cells, characterized in that, The active peptide comprises raw materials in the following mass ratio: Rink Amide AM resin: Phe: Gly: Leu: Arg: Gln: Lys: Tyr: Trp: Glu: Asn = 10: 24.8: 9.5: 11.3: 20.7: 19.5: 15: 7.4: 8.4: 6.8: 9.5; The amino acid sequence of the active peptide is H2N-FFGFLGRYFQKWEKLRNQ, as shown in SEQ ID NO.9; The active peptide was obtained by molecular docking simulation screening and optimization using the helical segment of IL-6R protein that binds to gp130 protein as a peptide template. The specific method is as follows: S1: The IL-6 / IL-6R / gp130 complex was used as the intervention target. Its high-resolution crystal structure was retrieved from the protein structure database and was numbered PDB ID: 1P9M. The polypeptide template was selected. S2: Eight candidate peptides were designed based on peptide templates and numbered P1-P8 in sequence; S3: Perform property and structural evaluation analysis on candidate peptides P1-P8, and screen out candidate peptides P1, P2 and P6; S4: Molecular docking simulations were performed between candidate peptides P1, P2 and P6 and 1P9M to screen out candidate peptide P1. S5: Functional sequence optimization of candidate peptide P1 was performed to design an active peptide for senescent cells; The amino acid sequence of the candidate peptide P1 is H2N-RYFQKWEKLRNQ, as shown in SEQ ID NO.1; The amino acid sequence of the candidate peptide P2 is H2N-WKRLYEHFQKRN, as shown in SEQ ID NO.2; The amino acid sequence of the candidate peptide P3 is H2N-KWEYRFQNRLKK, as shown in SEQ ID NO.3; The amino acid sequence of the candidate peptide P4 is H2N-RLFWKQEYNRKK, as shown in SEQ ID NO.4; The amino acid sequence of the candidate peptide P5 is H2N-YWKRQEFNRLKK, as shown in SEQ ID NO.5; The amino acid sequence of the candidate peptide P6 is H2N-KRWYQEFKLRNQ, as shown in SEQ ID NO.6; The amino acid sequence of the candidate peptide P7 is H2N-CRYFQKWEKLRNC, as shown in SEQ ID NO.7; The amino acid sequence of the candidate peptide P8 is H2N-CWKRLYEHFQKRNC, as shown in SEQ ID NO.

8.

2. A method for preparing an active peptide for senescent cells according to claim 1, characterized in that, The specific steps are as follows: Step 1: Take Rink Amide AM resin, swell it, filter it, remove the protection, and wash it to obtain deprotected resin; Step 2: Amino acid grafting was performed starting from the C-terminus of the active peptide sequence. Phe, Gly, Leu, Arg, Gln, Lys, Tyr, Trp, Glu and Asn were taken for activation, grafting reaction, identification by ninhydrin method, deprotection and washing to obtain peptide resin. Step 3: Cut the peptide resin to obtain crude peptide; Step 4: Precipitate the crude polypeptide using ice-cold ether, centrifuge, discard the supernatant, and obtain the precipitate. The precipitate is then vacuum dried to obtain the active peptide for senescent cells.

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