A polypeptide or polypeptide composition and uses thereof

Abstract

The application discloses a polypeptide or polypeptide composition based on AAV2 capsid protein design, and uses a linear polypeptide or polypeptide composition derived from an exposed region of AAV2 capsid protein to constitute a decoy composition, which can be combined with neutralizing antibodies (Nab) with high affinity, so as to selectively neutralize anti-AAV2 Nab without destroying the whole immune system, and effectively improve the transduction efficiency of an AAV2 gene vector in a neutralizing antibody positive individual.

Description

Technical Field

[0001] This invention belongs to the field of bioengineering technology, specifically relating to a polypeptide or polypeptide composition and its application. Background Technology

[0002] Adeno-associated virus type 2 (AAV2) has become one of the preferred vectors for in vivo gene therapy due to its non-pathogenicity, low immunogenicity, and persistent gene expression capacity. However, the AAV2 capsid protein can induce the production of neutralizing antibodies (Nab) after natural infection or vaccination. These antibodies significantly reduce gene transduction efficiency by blocking the binding of the virus to cell receptors.

[0003] Epidemiological studies show that approximately 30% to 80% of the global population possesses Nab against AAV2, with titers and distribution varying by region, age, and individual immune status. Furthermore, patients who have received AAV2 gene therapy produce higher Nab titers after treatment, further limiting the possibility of repeat administration. Therefore, effectively overcoming the influence of Nab has become a key challenge in improving the success rate of AAV2 gene therapy.

[0004] Currently, the main strategies for dealing with AAV2 Nab include the following:

[0005] (1) Plasmapheresis: This method removes antibodies from plasma through physical means, reducing Nab levels in the short term. However, this method is complex to operate, costly, and antibody levels may rebound rapidly, limiting its widespread application.

[0006] (2) Combination therapy with immunosuppressants: glucocorticoids or other immunosuppressants are used to reduce antibody production, but long-term use may cause side effects such as infection, and the effect on existing Nab is limited.

[0007] (3) Use of immunoglobulins (such as IdeS): These specifically cleave IgG antibodies, rapidly reducing Nab levels. Although this method has shown good results in some studies, its safety and long-term effects still require further evaluation.

[0008] (4) Developing low immunogenic AAV variants: By engineering the capsid protein, the binding to Nab can be reduced. However, the transduction efficiency and tissue specificity of the variants may be affected, and they may still trigger new immune responses.

[0009] (5) Using empty viral particles as decoys: By binding to Nab, their neutralizing effect on therapeutic viruses is reduced. However, this method may require a large number of empty particles and carries the risk of inducing an immune response. Although the above methods alleviate the effects of Nab to some extent, they still have many limitations, such as complex operation, high cost, poor specificity, and potential side effects. Therefore, developing a safe, efficient, highly specific, and easy-to-operate Nab neutralization strategy has become a current research hotspot. Summary of the Invention

[0010] The purpose of this invention is to overcome the shortcomings of the prior art. This invention provides a polypeptide or polypeptide composition designed based on AAV2 capsid protein, which can specifically neutralize Nab, improve the success rate of gene therapy, and has important clinical application value.

[0011] This invention is achieved through the following technical solution:

[0012] In a first aspect, the present invention provides a polypeptide, wherein the polypeptide is one of the amino acid sequences shown in SEQ ID NO: 1 to 20.

[0013] Optionally, the polypeptide is an amino acid sequence obtained by substituting, deleting, adding or inserting at least one amino acid residue into one of the amino acid sequences described in SEQ ID NO: 1 to 20.

[0014] Optionally, the polypeptide is a derived polypeptide obtained by mutating some amino acid residues of one of the amino acid sequences shown in SEQ ID NO: 1 to 20;

[0015] Optionally, the polypeptide is an amino acid sequence having at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% sequence identity with one of the amino acid sequences shown in SEQ ID NO: 1-20.

[0016] In a second aspect, the present invention provides a polypeptide composition, said polypeptide composition being selected from the group consisting of or a combination thereof:

[0017] (1) The polypeptide composition comprises at least one of the amino acid sequences shown in SEQ ID NO: 1 to 20;

[0018] (2) The polypeptide composition comprises at least one of the amino acid sequences obtained by substituting, deleting, adding or inserting at least one amino acid residue of one of the amino acid sequences of SEQ ID NO: 1 to 20;

[0019] (3) The polypeptide composition comprises at least one of the derivative polypeptides obtained by mutating some amino acid residues of one of the amino acid sequences shown in SEQ ID NO: 1 to 20;

[0020] (4) The polypeptide composition comprises at least one of the amino acid sequences having at least 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95% sequence identity with one of the amino acid sequences shown in SEQ ID NO: 1 to 20.

[0021] Optionally, the polypeptide or polypeptide composition includes, but is not limited to, one or more of the amino acid sequences shown in SEQ ID NO: 1 to 20.

[0022] Optionally, the polypeptide originates from the surface exposed region of the AAV2 capsid protein, specifically:

[0023] SEQ ID NO:1: WALPTYNNHLYKQISSQSGA;

[0024] SEQ ID NO:2: QISSQSGASNDNHYFGYSTP;

[0025] SEQ ID NO:3:KEVTQNDGTTTIANNLTSTV;

[0026] SEQ ID NO:4: SAHQGCLPPFPADVFMVPQY;

[0027] SEQ ID NO: 5: DVFMVPQYGYLTLNNGSQAV;

[0028] SEQ ID NO:6: PLIDQYLYYLSRTNTPSGTT;

[0029] SEQ ID NO:7: RTNTPSGTTTQSRLQFSQAG;

[0030] SEQ ID NO:8: QFSQAGASDIRDQSRNWLPG;

[0031] SEQ ID NO:9:PGPCYRQQRVSKTSADNNNS;

[0032] SEQ ID NO:10: SKTSADNNNSEYSWTGATKY;

[0033] SEQ ID NO: 11: EYSWTGATKYHLNGRDSLVN;

[0034] SEQ ID NO:12: NPGPAMASHKDDEEKFFPQS;

[0035] SEQ ID NO:13: GKQGSEKTNVDIEKVMITDE;

[0036] SEQ ID NO:14:SEKTNVDIEKVMITDEEEIR;

[0037] SEQ ID NO:15: TEQYGSVSTNLQRGNRQAAT;

[0038] SEQ ID NO: 16: STNLQRGNRQAATADVNTQG;

[0039] SEQ ID NO:17:VPANPSTTFSAAKFASFITQ;

[0040] SEQ ID NO:18: KRWNPEIQYTSNYNKSVNVD;

[0041] SEQ ID NO:19: TSNYNKSVNVDFTVDTNGVY;

[0042] SEQ ID NO:20: FTVDTNGVYSEPRPIGTRYL.

[0043] The polypeptide has the following characteristics:

[0044] (1) Derived from the surface region of the three-dimensional structure of the AAV2 capsid protein;

[0045] (2) It has good hydrophilicity, with a GRAVY value of less than -1.0; it has good water solubility, is easy to synthesize, and is easy to bind to antibodies;

[0046] (3) It has high affinity for human immunoglobulins (such as IVIG) and anti-AAV2 neutralizing antibodies in the plasma of immunized mice in in vitro experiments;

[0047] (4) It can significantly block the neutralizing effect of neutralizing antibodies on AAV2 vector and improve gene expression efficiency;

[0048] (5) The polypeptide itself does not induce significant immunogenicity or inflammatory response.

[0049] Optionally, the polypeptide composition comprises one or more of the amino acid sequences shown in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 14, and SEQ ID NO: 16.

[0050] Optionally, the polypeptide composition includes at least: SEQ ID NO: 14 and SEQ ID NO: 16.

[0051] Thirdly, the present invention provides the use of the polypeptide or polypeptide composition described in the above technical solution in the preparation of a neutralizing antibody drug against AAV2.

[0052] Optionally, the AAV2 neutralizing antibody drug described above is further defined as a drug for improving the efficacy of AAV2-mediated in vivo gene therapy, especially in cases of repeated administration of AAV2-mediated in vivo gene therapy drugs.

[0053] Fourthly, the present invention provides a pharmaceutically acceptable salt of a polypeptide or polypeptide composition, wherein the polypeptide is selected from the polypeptides or polypeptide compositions described above.

[0054] Fifthly, this application provides a pharmaceutical composition comprising a polypeptide or polypeptide composition thereof, a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the polypeptide or polypeptide composition is selected from the polypeptide or polypeptide composition described above, and the pharmaceutically acceptable salt thereof is the polypeptide or polypeptide composition described above.

[0055] Sixthly, this application provides an in vitro binding reagent, which comprises the polypeptide or polypeptide composition described in the above-mentioned technical solutions.

[0056] Optionally, the in vitro binding reagent is used for Nab level screening and / or diagnosis.

[0057] In a seventh aspect, the present invention provides a method for evaluating the neutralizing effect of AAV2 Nab in animals, the method comprising:

[0058] (1) Establish an AAV2-immunized mouse model or a human venous blood immunoglobulin IVIG model;

[0059] (2) Give the polypeptide or polypeptide composition as described above;

[0060] (3) Detect AAV2-mediated reporter gene expression (such as luciferase activity or the proportion of EGFP-positive cells) as an evaluation indicator;

[0061] (4) Compare the expression differences between the groups with and without peptide treatment.

[0062] Optionally, this method can be used to screen for novel Nab-neutralizing peptides or to evaluate enhancement strategies for existing compositions.

[0063] The advantages and beneficial effects of this invention are as follows:

[0064] (1) This invention proposes for the first time to use linear polypeptides or polypeptide compositions derived from the exposed region of AAV2 capsid protein to form a bait composition, which can bind with high affinity to neutralizing antibodies (Nab), thereby selectively neutralizing anti-AAV2 Nab without damaging the overall immune system, effectively improving the transduction efficiency of AAV2 gene vector in neutralizing antibody-positive individuals;

[0065] (2) The polypeptides or polypeptide compositions described in this invention are short-chain synthetic peptides, which are rationally designed, easy to chemically synthesize and mass-produce, have strong stability, simple structure, are easy to store and transport, and have good potential for pharmaceutical development and industrial transformation.

[0066] (3) By screening multiple AAV2-derived peptides with complementary structural features, this invention has the ability to cover antibody epitopes of different conformations, and is applicable to different titers and lineages of Nab in a wide range of populations, thereby improving the broad spectrum and adaptability of neutralization.

[0067] (4) Validation in animal models showed that the polypeptide or polypeptide composition of the present invention can improve the efficiency of AAV2-mediated gene expression without inducing abnormal activation of the immune system, and no increase in anti-polypeptide antibodies or inflammatory factors was detected, showing good biocompatibility and safety.

[0068] (5) The polypeptide composition described in this invention can be flexibly applied to a variety of clinical scenarios. It can be used to clear the basic Nab of an individual before the first AAV2 gene therapy, or it can be used in combination with enzymes (such as IdeS) before repeated AAV2 administration, providing a practical solution to the problem of “non-repeatable drug administration” in AAV2 gene therapy. Attached Figure Description

[0069] Figure 1 (A) Spatial distribution characteristics of neutralizing peptides in the three-dimensional structure of the AAV2 capsid; (B) Distribution characteristics of neutralizing peptides in the amino acid sequence of the AAV2 capsid protein.

[0070] Figure 2 (A) One valid assay is missing (1 out of 20) to assess the importance of detecting AAV2 Nab in human IVIG using peptide compositions; (B) One valid assay is missing (1 out of 20) to assess the importance of detecting AAV2 Nab in mouse serum using peptide compositions.

[0071] Figure 3 (A) The peptide composition improves the gene delivery efficiency of AAV2-Luc in an AAV2-immunized mouse model; (B) The peptide composition improves the gene delivery efficiency of AAV2-Luc in an IVIG passive immunization model.

[0072] Figure 4Safety assessment results after peptide injection: (A) Whether the peptide composition injection induces the production of specific antibodies against the peptide; (B) Whether the peptide composition injection induces an increase in inflammatory factors in mice.

[0073] For those skilled in the art, other related figures can be obtained from the above figures without any creative effort. Detailed Implementation

[0074] To enable those skilled in the art to better understand the present invention, the technical solution of the present invention will be further described below with reference to specific embodiments.

[0075] Where specific techniques or conditions are not specified in the examples, they shall be performed in accordance with the techniques or conditions described in the literature in this field, or in accordance with the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels.

[0076] Example 1: Screening Method for Neutralizing Antibody-Decoy Peptides

[0077] This embodiment aims to illustrate the screening method for neutralizing antibody decoy peptides described in this invention and their spatial distribution characteristics in the three-dimensional structure of the AAV2 capsid.

[0078] First, the amino acid sequence of the AAV2 capsid protein VP1 (Uniprot: P03135-1), consisting of 735 amino acid residues, was obtained. This sequence was then processed using a sliding window technique, with each window containing 20 amino acids and a step size of 1, resulting in 716 candidate polypeptide sequences.

[0079] Next, multiple rounds of screening will be conducted based on the following criteria:

[0080] (1) Three-dimensional structural exposure analysis: Using the resolved crystal structure of the AAV2 capsid protein trimer (PDB ID: 6IH9), PyMOL and structural visualization algorithms were used to identify all exposed residues on the surface of amino acids and map them to each polypeptide fragment, and fragments with an exposure ratio of more than 70% were screened.

[0081] (2) Homology exclusion: Remove sequences that are highly homologous to human proteins to avoid potential cross-reactions or immune tolerance;

[0082] (3) Functional region priority: Prioritize the retention of sequences distributed in known antibody binding regions of VP3 (such as VR-I, VR-IV, VR-V) to improve decoy efficiency.

[0083] Finally, 20 polypeptide fragments that met the above criteria were selected, which are the polypeptides of the present invention (see SEQ ID NO:1 to SEQ ID NO:20 for details). All selected polypeptides are 20 amino acids in length, distributed in different regions of the VP3 sequence, and extensively cover the surface of the AAV2 capsid.

[0084] In structural distribution analysis ( Figure 1 A) Using PyMOL software, the spatial locations of 20 peptides were visualized in three dimensions. The results showed that all peptides were distributed on the outer surface of the AAV2 capsid, located in multiple pentamer-trimer junction regions; some peptides were located in known antibody binding hotspots such as the B / C ring and GH ring; the overlap between peptide-covered regions was less than 5 amino acid residues, and after binding, they could structurally and synergistically shield multiple key antibody binding sites, exhibiting a potential synergistic neutralization effect. Meanwhile, in Figure 1 B indicates the region of the polypeptide composition covered in the amino acid sequence of the AAV2 capsid protein.

[0085] The results in summary indicate that the 20 linear peptides obtained through systematic screening in this invention can efficiently cover the exposed surface of the AAV2 capsid protein, possess a good structural distribution basis and functional synergy, and are ideal candidate sequences for neutralizing antibody decoys.

[0086] Example 2: Neutralization of human IVIG and mouse serum Nab by peptide composition

[0087] This embodiment aims to evaluate the neutralizing effect of the peptide composition on human immunoglobulin (IVIG) and AAV2 neutralizing antibody (Nab) in immunized mouse plasma, and to verify the synergistic contribution of each peptide in the peptide composition.

[0088] Materials and Methods

[0089] 1. IVIG source: Commercial human intravenous immunoglobulin (IVIG) purchased from Sigma, concentration 10 mg / mL, known to contain medium to high titers (>1:256) of anti-AAV2 Nab.

[0090] 2. Mouse plasma preparation: C57 mice were injected with AAV2 empty vector (1×10⁻⁶). 12 Two weeks later, serum was collected and confirmed to have a moderate titer (>1:256) of anti-AAV2 Nab.

[0091] 3. Peptide composition 0: Composed of 20 20-aa peptides selected in Example 1 “Peptide Screening Process” (see SEQ ID NO:1 to SEQ ID NO:20), each peptide is mixed at an equimolar concentration, and the final concentration is 1 μM per peptide.

[0092] 4. Peptide compositions 1-20: Composed of 19 of the 20 20-aa peptides selected in Example 1 “Peptide Screening Procedure” (LOO strategy);

[0093] The polypeptide composition 1 is missing SEQ ID NO:1, and the remaining SEQ ID NO:2 to SEQ ID NO:20 are mixed at an equimolar concentration, with a final concentration of 1 μM per polypeptide; for single peptides, the final concentration of each single peptide is 50 nM.

[0094] In this peptide composition 2, SEQ ID NO:1 is missing, and the remaining SEQ ID NO:1 and SEQ ID NO:3 to SEQ ID NO:20 are mixed at equimolar concentrations, with a final concentration of 1 μM per peptide; for single peptides, the final concentration of each single peptide is 50 nM.

[0095] And so on;

[0096] ...

[0097] The polypeptide composition 20 is the one lacking SEQ ID NO:20, and the remaining SEQ ID NO:1 to SEQ ID NO:19 are mixed at an equimolar concentration, with a final concentration of 1 μM per polypeptide; for single peptides, the final concentration of each single peptide is 50 nM.

[0098] Neutralization experiment procedure:

[0099] 1. IVIG or mouse plasma were added to different treatment groups (① no peptide control; ② peptide composition 0 (20 peptides in total); ③ peptide compositions 1-20 were added respectively (each peptide was deleted one by one, and each was a mixture of 19 peptides); pre-mixed and incubated for 30 minutes;

[0100] 2. Add the AAV2-Luc virus (MOI=5000);

[0101] 3. Add HEK293T cells to the mixture and culture for 48 hours;

[0102] 4. The expression level of AAV2-mediated Luc (luciferase) was detected using the luciferase method.

[0103] Experimental results

[0104] 1. Mouse plasma experiment results: Luc expression decreased to approximately 12% in the group with added plasma but without peptides; expression recovered to approximately 68% after adding the complete peptide composition; LOO strategy results showed that the neutralizing antibodies produced after AAV2 immunization of mice could recognize multiple peptides, and the absence of any one of the key peptides (SEQ ID NO:1, NO:3, NO:14, NO:16, or NO:19) could lead to a significant decrease in AAV2-Luc infection efficiency. Figure 2 A) indicates that blocking all neutralizing antibodies requires a mixture of multiple peptides representing different epitopes.

[0105] 2. Human IVIG Experiment Results: Luc expression in the group without peptides decreased to baseline (<10%); after adding the complete 20-peptide combination, Luc expression recovered to approximately 75% (compared to the Nab negative control); LOO analysis showed that the most significant decrease in Luc expression (to 45%–55%) occurred when SEQ ID NO:1, NO:3, NO:14, NO:16, or NO:19 was removed, indicating that the neutralizing antibody could not effectively block these peptides, leading to a significant reduction in AAV-Luc gene delivery efficiency due to neutralizing antibody binding during cell infection, suggesting that these are key neutralizing regions; similarly, the removal of other sequences had a smaller impact on expression recovery (>65%), suggesting that these are minor key neutralizing regions.

[0106] in conclusion

[0107] This experiment verified that the peptide composition consisting of the 20 peptides can significantly neutralize anti-AAV2 Nab in human IVIG and mouse immune serum, effectively restoring AAV2-mediated gene expression. The LOO strategy results further confirmed the existence of key active sequences within the peptide composition and the synergistic effect between the peptides. This neutralization strategy demonstrated broad adaptability in plasmas from different species and sources, showing good versatility and clinical application potential.

[0108] Example 3: Enhancement of AAV2-Luc expression using the peptide composition in an animal model.

[0109] Experimental Design

[0110] To evaluate the inhibitory effect of the peptide composition on AAV neutralizing antibody (Nab) in vivo, the following animal models were established:

[0111] AAV2-immunized mouse model: C57BL / 6 mice were intravenously injected with AAV2-GFP (5×10⁻⁶). 10 (vg / animal), after 7 days, the neutralizing peptide composition was injected intravenously or plasma was collected to detect Nab levels.

[0112] Passive immunization model: Mice were intravenously injected with human IVIG (0.5 mg / mouse), and Nab levels were measured 7 days later.

[0113] A gene therapy-enhanced model was constructed based on the AAV2-immunized mouse model and the passive immunization model:

[0114] Pretreatment group: Based on the AAV2 immunized mouse model and the passive immunization model, the peptide composition (50 μL, 10 μM peptide composition containing SEQ ID NO: 1-20 in equimolar ratio, with a final concentration of 0.5 μM per peptide) was injected intraperitoneally. Four hours later, AAV2-Luc (i.e., the aforementioned AAV2-GFP or human IVIG, at a dose of 1×10⁻⁶) was injected again. 11 vg / each);

[0115] Control group: Mouse models immunized with AAV2 or passively immunized only received AAV2-Luc (1×10⁻⁶). 11 vg / each).

[0116] Gene therapy based on non-AAV2 immunized mouse models and passive immunization models:

[0117] AAV2 group: Mice were not pre-immunized with AAV2 (i.e., they were not injected with the aforementioned AAV2-GFP or human IVIG) but were directly injected with AAV2-Luc virus.

[0118] Monitoring time points: Liver luciferase (Luc) expression was detected at 5, 15, and 25 days later.

[0119] Experimental results

[0120] 1. The peptide composition significantly improved the gene delivery efficiency of AAV2-Luc in an AAV2-immunized mouse model.

[0121] Luciferase activity assay: Results are shown in [link to results]. Figure 3 A

[0122] Pretreatment group (peptide composition + AAV2-Luc): After 25 days, the liver Luc activity reached 80% of that in the AAV2 group (AAV2 group, mice were not pre-immunized with AAV2 and were directly injected with AAV2-Luc virus) (expression level without Nab interference).

[0123] Control group (AAV2-Luc only, control mice in the figure were pre-immunized, without peptide injection pretreatment, and injected with AAV2-Luc virus): Due to the presence of Nab, Luc activity was only <15%.

[0124] 2. The peptide composition significantly improved the gene delivery efficiency of AAV2-Luc in the IVIG passive immunization model.

[0125] Luciferase activity assay: Results are shown in [link to results]. Figure 3 B

[0126] Pretreatment group (peptide composition + AAV2-Luc): After 25 days, liver Luc activity reached 80% of that in the AAV2 group (expression level without Nab interference).

[0127] Control group (AAV2-Luc only, control in the figure, mice pre-injected with IVIG, no peptide injection, injected with AAV2-Luc): Due to the presence of Nab, Luc activity was only <15%.

[0128] Example 4: Safety assessment results after peptide injection

[0129] Experimental Design

[0130] To ensure the safety of peptide mixtures in clinical applications, a systematic evaluation was conducted, including:

[0131] 1. Immunogenicity test: to assess whether the peptide induces antibody production.

[0132] 2. Monitoring of inflammatory response: Detecting the levels of key pro-inflammatory / anti-inflammatory cytokines.

[0133] 3. Physiological index analysis: Observe the behavior, metabolism and organ function of mice.

[0134] Experimental Groups

[0135] 1. PBS control group: injected with phosphate-buffered saline (PBS).

[0136] 2. Peptide composition group: Intraperitoneal injection peptide composition (100 μg / animal, the ip peptide composition contains SEQ ID NO: 1 to 20 in equimolar ratio).

[0137] 3. Positive control group: injected with ovalbumin (OVA, 100 μg / animal, ip, known immunogenic protein).

[0138] Experimental results

[0139] 1. Immunogenicity analysis ( Figure 4 A)

[0140] Antibody production detection (direct ELISA):

[0141] Peptide composition group: 7 days after injection, no specific IgG antibody against AAV peptide was detected in plasma (OD450<0.1, no difference from PBS group).

[0142] OVA positive control group: High levels of anti-OVA antibodies were induced (OD450>1.5, p<0.001), serving as a positive control for mouse immunization and demonstrating normal mouse immunity.

[0143] Conclusion: The AAV peptide mixture has no significant immunogenicity and does not induce an adaptive immune response.

[0144] 2. Assessment of inflammatory response ( Figure 4 B)

[0145] Cytokine assay (CBA method): 24 hours after injection, there was no significant difference in the levels of key inflammatory factors in plasma between the peptide group and the PBS injection group.

[0146] Conclusion: The peptide composition did not induce a cytokine storm or systemic inflammation.

[0147] 3. Monitoring of physiological indicators

[0148] All peptide groups showed a score of 0 in mice, with no neurotoxicity, motor dysfunction, or metabolic abnormalities.

[0149] The applicant declares that the detailed method of the present invention is illustrated by the above embodiments, but the present invention is not limited to the above detailed method, that is, it does not mean that the present invention must rely on the above detailed method to be implemented. Those skilled in the art should understand that any improvements to the present invention, equivalent substitutions of the raw materials of the product of the present invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

Claims

1. A polypeptide or polypeptide composition, characterized in that, The polypeptide or polypeptide mixture comprises one or more of the amino acid sequences shown in SEQ ID NO: 1 to 20.

2. The polypeptide composition according to claim 1, characterized in that, The polypeptide composition comprises one or more of the amino acid sequences shown in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 14, and SEQ ID NO:

16.

3. The polypeptide composition according to claim 1, characterized in that, The polypeptide composition comprises at least: SEQ ID NO: 14 and SEQ ID NO:

16.

4. The use of a polypeptide or polypeptide composition according to any one of claims 1 to 3 in the preparation of an antibody drug against AAV2.

5. The application according to claim 4, characterized in that, The aforementioned AAV2 neutralizing antibody drug is a drug used to improve the efficacy of AAV2-mediated in vivo gene therapy.

6. The application according to claim 4, characterized in that, The aforementioned AAV2 neutralizing antibody drug is a drug used to improve the efficacy of AAV2-mediated in vivo gene therapy in the context of repeated administration of AAV2-mediated in vivo gene therapy drugs.

7. A pharmaceutically acceptable salt of a polypeptide or polypeptide composition, wherein the polypeptide is selected from the polypeptides described in any one of claims 1 to 3; The polypeptide composition is selected from the polypeptide compositions described in any one of claims 1 to 3.

8. A pharmaceutical composition, characterized in that, Includes polypeptides or polypeptide compositions thereof, pharmaceutically acceptable salts thereof, and pharmaceutical carriers thereof, wherein the polypeptides are selected from the polypeptides described in any one of claims 1 to 3; The polypeptide composition is selected from the polypeptide compositions described in any one of claims 1 to 3; The pharmaceutically acceptable salt of the polypeptide or polypeptide composition is the pharmaceutically acceptable salt of the polypeptide or polypeptide composition as described in claim 7.

9. An in vitro binding reagent, characterized in that, The in vitro binding reagent comprises a polypeptide or polypeptide composition as described in any one of claims 1 to 3; Preferably, the in vitro binding reagent is used for Nab level screening and / or diagnosis.

10. A method for evaluating the neutralizing effect of AAV2 on Nab in animals, characterized in that, The method includes: (1) Establish an AAV2-immunized mouse model or a human venous blood immunoglobulin IVIG model; (2) The polypeptide or polypeptide composition as described in any one of claims 1 to 3 is given; (3) Detect AAV2-mediated reporter gene expression (such as luciferase activity or the proportion of EGFP-positive cells) as an evaluation indicator; (4) Compare the expression differences between the groups with and without peptide treatment; Preferably, this method is used to screen for novel Nab-neutralizing peptides or to evaluate enhancement strategies for existing compositions.

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