Neoantigen epitope peptide fragment presented based on MHC-II, vaccine, pharmaceutical composition and application

Abstract

The invention relates to the technical field of biological medicines, and particularly discloses a neoantigen epitope peptide fragment presented based on MHC-II, a vaccine, a pharmaceutical composition and application. Wherein the epitope key sequence of the peptide fragment is MVMGVLDQAFDVLVL, and the epitope key sequence is MVMGVLDQAFDVLVL; a KVMGVLDQAFDVLVL is used as a model; kVMGVLKQAFDVLVL is used as a model; kVLGVLKQAFDVLVL is used as a model; and KVKGVLKQAFDVLVL can be selected from any one of KVKGVLKQAFDVLVL. The new antigen with the epitope peptide fragment can be recognized by CD4 + T cells, so that the CD4 + T cells can provide necessary auxiliary signals to enhance the immune response of the CD8 + T cells, and finally the CD8 + T cells are helped to kill tumor cells such as pancreatic cancer. The discovery of the peptide fragment can be beneficial to popularization and application of vaccine therapy in the field of tumor diseases such as pancreatic cancer.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to antigenic epitope peptides for treating tumor diseases, vaccines and pharmaceutical compositions made from these peptides, and the application of the aforementioned materials in the field of tumor treatment. Background Technology

[0002] Malignant tumors such as pancreatic cancer seriously threaten human quality of life and physical health. Conventional clinical treatments such as surgery, radiotherapy and chemotherapy cannot effectively curb the development of these tumors due to their large side effects and high recurrence rate. Immunotherapy, especially tumor vaccines, has attracted widespread attention from clinicians and researchers because it can induce the body to produce a specific immune response that kills tumor cells.

[0003] Neoantigens used in tumor vaccines originate from non-synonymous somatic cell mutations in tumor cells. These neoantigens are not present in normal cells and therefore can elicit an immune response unaffected by central and peripheral tolerance. The immunogenicity of a neoantigen depends on its ability to be effectively presented to T cells by MHC molecules. Specifically, MHC molecules can bind to the cell surface to form different types of peptide-MHC complexes, generally classified as MHC class I and MHC class II. These complexes can be recognized by different T cell receptors. CD8+ T cells can recognize the peptide-MHC class I complex corresponding to the neoantigen, and upon recognition, they are activated, proliferate, and ultimately kill tumor cells expressing these neoantigens. CD4+ T cells can recognize the neoantigen peptides presented by MHC class II molecules and, upon recognition, provide necessary auxiliary signals to enhance the immune response of CD8+ T cells, thereby helping to kill the corresponding tumor cells.

[0004] As research into immunotherapy continues to deepen, it has been discovered that traditional neoantigen strategies targeting MHC-I cells suffer from a high defect rate. Meanwhile, MHC-II neoantigens, due to their advantages such as easier neoantigen binding, lower defect rate, and crucial role in activating CD4+ T cells, are becoming an important emerging target for tumor immunotherapy. However, the discovery of immunogenic neoantigens has become a core issue hindering the development of neoantigen therapy.

[0005] Pancreatic cancer, a digestive system tumor, is a common type of tumor that can cause great harm to human health. However, current research on neoantigens for this type of tumor is still insufficient. Therefore, there is an urgent need for a neoantigen to effectively treat or prevent digestive system tumors. Summary of the Invention

[0006] One of the objectives of this invention is to provide a novel antigenic epitope peptide based on MHC-II presentation that is suitable for tumor treatment;

[0007] A second objective of this invention is to provide a tumor vaccine comprising the aforementioned peptides;

[0008] A third objective of this invention is to provide an application of the above-mentioned peptide or vaccine in a drug for treating tumor diseases;

[0009] The fourth objective of this invention is to provide a combination drug for the treatment of tumor diseases, comprising the aforementioned peptides or vaccines.

[0010] This invention is achieved through the following technical solution:

[0011] In a first aspect, the present invention provides a novel antigenic epitope peptide based on MHC-II presentation, wherein the key epitope sequence of the peptide is any one of: MVMGVLDQAFDVLVL; KVMGVLDQAFDVLVL; KVMGVLKQAFDVLVL; KVLGVLKQAFDVLVL; KVKGVLKQAFDVLVL.

[0012] As a further improvement of this invention, the key epitope sequence of the peptide is MVMGVLDQAFDVLVL. The inventors have found that using this key epitope sequence results in better therapeutic effects for tumor diseases.

[0013] As a further improvement of the present invention, the full sequence of the new antigenic epitope peptide is: MDAMKRGLCCVLLLCGAVFVSPS-GS-AKFVAAWTLKAAA-EAAAK-(α)-GPGPG-VVVGAVGVGKSALTI-GGGS-HHHHHH-GYQTI-RKRSHAGYQTI, where α is any one of the key epitope sequences of the peptide.

[0014] Secondly, the present invention provides a tumor vaccine comprising any of the aforementioned peptides and pharmaceutically acceptable adjuvant components.

[0015] As a further improvement of the present invention, the auxiliary component includes a vaccine adjuvant, which is any one of poly(I:C), AS04, MF59, and CFA.

[0016] As a further improvement of the present invention, the mass ratio of the peptide to the vaccine adjuvant is 1:(1~3).

[0017] Thirdly, the present invention provides the use of any of the above-mentioned peptides or any of the above-mentioned tumor vaccines in the preparation of medicaments for the prevention and / or treatment of tumor diseases.

[0018] As a further improvement of the present invention, the drug is an injectable preparation, and the injection route is at least one of intramuscular injection, intravenous injection, subcutaneous injection, and intradermal injection.

[0019] As a further improvement of the present invention, the tumor diseases treated with the drug include at least one of pancreatic cancer, intestinal cancer, liver cancer, and stomach cancer.

[0020] Fourthly, the present invention provides a combination drug for the treatment of tumors, comprising any of the aforementioned peptides or tumor vaccines administered separately or simultaneously, and other drugs for treating tumor diseases.

[0021] The beneficial effects of this invention lie in providing a novel antigenic epitope peptide. This novel antigen, possessing this epitope peptide, can be recognized by CD4+ T cells, thereby enabling CD4+ T cells to provide necessary auxiliary signals to enhance the immune response of CD8+ T cells, ultimately helping CD8+ T cells kill the corresponding tumor cells. Furthermore, the discovery of this peptide can facilitate the widespread application of vaccine therapy in the field of tumor diseases such as pancreatic cancer. In practical use, the inventors combine the aforementioned peptide with adjuvants to formulate a tumor vaccine. This tumor vaccine can effectively inhibit the growth of tumors such as pancreatic cancer, thereby achieving the effect of treating tumor diseases. Attached Figure Description

[0022] The accompanying drawings are provided below to illustrate the preferred embodiments of the invention and to aid in understanding the objectives and advantages of the invention, wherein:

[0023] Figure 1 A summary chart of the affinity analysis results of tumor vaccines for immune cells (BMDC);

[0024] Figure 2 This is a summary chart of the analysis results of tumor cell apoptosis induced by tumor vaccines in Examples 1-5;

[0025] Figure 3 This is a summary chart of the results of tumor vaccine induction in Examples 1-5, which increased the levels of key indicators in the serum of diseased mice.

[0026] Figure 4 This is a summary chart of the test results of the inhibitory effect of the tumor vaccines in Examples 1-5 on pancreatic tumors.

[0027] In the accompanying figures, the tumor vaccines of Examples 1 to 5 are marked as P1 to P5 respectively. The blank group is the Blank group, the control group is the Control group and the C group, and the model group is the Model group and the M group.

[0028] in, Figure 1A shows the flow cytometry analysis of the binding of the tumor vaccine to BMDC cells after co-culturing for 0h, 0.5h, 1h, 2h, 6h, 12h, and 24h; B shows the quantitative analysis of the average FITC fluorescence intensity of cells after co-culturing for 0h, 0.5h, 1h, 2h, 6h, 12h, and 24h (2h, 6h vs 0h: p < 0.0001); C shows the laser confocal scanning microscopy observation of the binding of the tumor vaccine to BMDC cells after co-culturing for 0h, 0.5h, 1h, 2h, 6h, 12h, and 24h (BMDC cells were modified with DAPI, and the tumor vaccine was modified with FITC).

[0029] Figure 2 A shows the apoptosis of tumor cells after co-culturing with activated lymphocytes using flow cytometry, as detected by the tumor vaccines in Examples 1-5; B shows the quantitative analysis of tumor cell apoptosis rate (P1 vs M: p < 0.0001; P2, P3 vs M: p < 0.001); C shows the inhibitory effect of CCK8 assay on tumor cells after co-culturing with activated lymphocytes, with P1 activated lymphocytes showing the highest tumor inhibition rate.

[0030] Figure 3 A shows the IFN-γ level in mouse serum after the tumor vaccines in Examples 1-5 were used on pancreatic cancer mice, analyzed by ELISA (C, P1, P2, P3 vs M: p < 0.0001); B shows the TNF-α level in mouse serum after ELISA (C, P1, P2, P3 vs M: p < 0.0001; P4 vs M: p < 0.001; P5 vs M: p < 0.01); C shows the IL-2 level in mouse serum after ELISA (C, P1, P2, P3 vs M: p < 0.0001; P4 vs M: p < 0.001; P5 vs M: p < 0.001); D shows the Granzyme B level in mouse serum after ELISA (C vs M: p < 0.05; P1, P3 vs M: p < 0.0001; P2 vs M: p < 0.01).

[0031] Figure 4 A shows the trend of tumor volume change in pancreatic cancer mice in each group from 0 to 22 days using the tumor vaccine in Examples 1-5 (C, P1, P2, P3 vs M: p < 0.0001); B shows the change in body weight of mice in each group from 0 to 22 days (no significant difference); C shows the tumors of mice in each group; D shows the quantitative analysis and comparison of tumor quality in each group of mice (C, P1, P2, P3 vs M: p < 0.05). Detailed Implementation

[0032] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments.

[0033] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without inventive effort are within the scope of protection of this invention.

[0034] Example 1:

[0035] This embodiment provides a novel antigenic epitope peptide P1MHC based on MHC-II presentation. The key epitope sequence of this peptide is MVMGVLDQAFDVLVL, and its full sequence is MDAMKRGLCCVLLLCGAVFVSPS-GS-AKFVAAWTLKAAA-EAAAK-MVMGVLDQAFDVLVL-GPGPG-VVVGAVGVGKSALTI-GGGS-HHHHHH-GYQTI-RKRSHAGYQTI.

[0036] In this embodiment, the above-mentioned peptides were obtained by conventional Fmoc solid-phase synthesis. The specific reagents used, etc., are described in the following steps:

[0037] In this embodiment, the polypeptide is synthesized from the C-terminus to the N-terminus.

[0038] S1. Weigh 3g of RINK resin (degree of substitution 0.3mmol / g) into a 150ml reactor and soak it in 50ml of dichloromethane (hereinafter referred to as DCM);

[0039] After soaking for 2 hours, the resin was washed with 3 times the volume of nitrogen-dimethylformamide (hereinafter referred to as DMF), then dried, and the washing and drying process was repeated four times. Finally, the resin was dried and ready for use.

[0040] S2. Add 20% piperidine (piperidine / DMF=1:4) to the reactor and shake it on a color shaker for 20 minutes to remove the Fmoc protecting groups on the resin. After deprotection, wash four times with DMF at a volume of 3 times the resin volume, and then dry it for later use.

[0041] Take a small amount of resin and test it using the ninhydrin (Ninhydrin hydrate) method (two drops each of test A and test B, react at 100℃ for 1 min). If the resin is colored, it indicates that the deprotection was successful and the subsequent preparation steps can be carried out.

[0042] S3. Weigh an appropriate amount of the first amino acid at the C-terminus and an appropriate amount of 1-hydroxy-benzotriazole (HOBT) into a 50ml centrifuge tube, add 20ml of DMF to dissolve them, then add 3ml of N,N-diisopropylcarbodiimide (hereinafter referred to as DIC) and shake well for 1min. After the solution is clear, add it to the reactor and then place the reactor in a shaker at 30℃ to react.

[0043] After the reaction continued for 2 hours, the head was capped with acetic anhydride (where acetic anhydride:DIEA:DCM=1:1:2) for half an hour, then washed four times with 3 times the volume of DMF, and dried for later use.

[0044] S4. Add 20% piperidine (piperidine / DMF = 1:4) to the reactor, and shake on a decolorizing shaker for 20 minutes to remove the Fmoc protecting groups from the resin. After deprotection, wash four times with 3 times the volume of DMF, and then dry for later use.

[0045] Take a small amount of resin and test it using the ninhydrin (Ninhydrin hydrate) method (two drops each of test A and test B, react at 100℃ for 1 min). If the resin is colored, it indicates that the deprotection was successful.

[0046] S5. Weigh the second amino acid and HOBT into a 50ml centrifuge tube, add 25ml of DMF to dissolve them, then add 2.5ml of DIC and shake for 1min. After the solution is clear, add it to the reactor and then place the reactor in a shaker at 30℃ to react.

[0047] After the reaction has continued for 1 hour, a small amount of resin is taken for testing using the ninhydrin method (two drops each of test A and test B, reacted at 100℃ for 1 minute). If the resin is colorless, the reaction is complete; if the resin is colored, the condensation is incomplete, and the reaction should continue.

[0048] S6. After the reaction is complete, wash the resin four times with 3 times the resin volume of DMF, then dry it under vacuum. Add 20% piperidine (piperidine / DMF = 1:4) to the reactor and shake it on a decolorizing shaker for 20 minutes to remove the Fmoc protecting group from the resin. After deprotection, wash the resin four times with 3 times the resin volume of DMF, then dry it under vacuum to check if the protection has been removed.

[0049] S7. Follow steps S5 to S6 to connect the following amino acids in sequence.

[0050] S8. After the last amino acid is attached, the protection is removed, and the mixture is washed four times with 3 times the volume of DMF. Then, the resin is dried with methanol. The peptide is then cleaved from the resin using 95% cleavage buffer (trifluoroacetic acid: 1,2-ethylenedithiol: 3, isopropylsilane: water = 95:2:2:1) (10 ml of cleavage buffer is added per gram of resin, depending on the resin weight). The peptide is then centrifuged four times with ice-cold ether (cleavage buffer: ether = 1:9). Finally, the peptide is separated and purified by HPLC and then lyophilized to obtain the purified peptide.

[0051] S9. The peptide is purified using conventional techniques, wherein the purification conditions are as follows:

[0052] Stationary phase: C18; Mobile phase: Pump A: V(tfa) / V(water) = 1 / 1000, Pump B: V(tfa) / V(acetonitrile) = 1 / 1000; Flow rate: 10 ml / min; Retention time: 20-30 min. The purification process was repeated twice to obtain the peptides in this example.

[0053] Example 2:

[0054] This embodiment provides a novel antigenic epitope peptide P2MHC based on MHC-II presentation. The difference between this peptide and that in Example 1 is that the key epitope sequence of the peptide in this embodiment is KVMGVLDQAFDVLVL.

[0055] Example 3:

[0056] This embodiment provides a novel antigenic epitope peptide P3MHC based on MHC-II presentation. The difference between this peptide and that in Example 1 is that the key epitope sequence of the peptide in this embodiment is KVMGVLKQAFDVLVL.

[0057] Example 4:

[0058] This embodiment provides a novel antigenic epitope peptide P4MHC based on MHC-II presentation. The difference between this peptide and that in Example 1 is that the key epitope sequence of the peptide in this embodiment is KVLGVLKQAFDVLVL.

[0059] Example 5:

[0060] This embodiment provides a novel antigenic epitope peptide P5MHC based on MHC-II presentation. The difference between this peptide and that in Example 1 is that the key epitope sequence of the peptide in this embodiment is KVKGVLKQAFDVLVL.

[0061] Performance testing:

[0062] In this embodiment, the peptides from Examples 1-5 were combined with the adjuvant poly(I:C) at a mass ratio of 1:1 to form a tumor vaccine, and various properties of the tumor vaccine were tested, including the affinity analysis of P1MHC tumor vaccine for BMDC, the in vitro test of the inhibitory effect of P1MHC~P5MHC on tumor cells, the in vitro test of the immune response of P1MHC~P5MHC in the serum of diseased mice, and the in vivo test of the inhibitory effect of P1MHC~P5MHC on tumor cells.

[0063] For experimental methods that do not specify conditions, the determination is usually carried out according to national standards. If there is no corresponding national standard, then the generally accepted international standards, standard conditions, or conditions recommended by the manufacturer shall be followed.

[0064] In this embodiment, the tumor targeted is mainly pancreatic cancer; the immune response in the serum of diseased mice is mainly determined by detecting changes in the levels of IFN-γ, TNF-α, IL-2 and Granzyme B in the mouse serum.

[0065] The main steps and results of each performance test are as follows:

[0066] (1) Affinity analysis of tumor vaccines to BMDC

[0067] First, mouse bone marrow-derived dendritic cells need to be counted after acquisition. Then, the cell concentration is adjusted to 1 × 10⁻⁶ cells using RPMI-1640 complete medium. 6 Cells were cultured at a density of 10 cells / ml, with 20 ng / ml of IL-4 and GM-CSF added. On days 2 and 4, half the medium was replaced. On days 6-8, suspended cells and loosely adherent cells were collected and plated (BMDC) at a density of approximately 50%. 2 μM of FITC-labeled tumor vaccine was added using a countdown method in the dark. After the incubation period, the culture medium was discarded, and the cells were washed three times with PBS. Simultaneously, 250 μl of 4% paraformaldehyde was added to each well for fixation for 20 min, followed by two washes with PBS in the dark. DAPI was then diluted with sterile water to a concentration of approximately 0.5 μg / mL, and 250 μl of DAPI was added to each well. After incubation at room temperature for 20 min, the cells were washed three times with PBS in the dark. Finally, 1 mL of PBS was added, and the cells were analyzed using a microarray, keeping the microarray dark throughout the analysis.

[0068] The results of flow cytometry analysis show that, for example Figure 1As shown, the affinity of the tumor vaccine to BMDC cells reached its optimal level when co-cultured for 2 hours and 6 hours. Laser confocal scanning microscopy revealed that the fluorescence colocalization of the tumor vaccine and BMDC cells was most obvious at the 6th hour of co-culture, and the tumor vaccine clearly entered the cytoplasm of BMDC cells, indicating that the affinity of the tumor vaccine to BMDC cells reached its optimal level when co-cultured for 6 hours.

[0069] The above results also demonstrate that the tumor vaccine in this embodiment is relatively successful in terms of antigen uptake and presentation, and has the function of initiating an immune response.

[0070] (2) In vitro testing of the inhibitory effect of tumor vaccines on tumor cells

[0071] In this embodiment, a model group (Model group, M group), a drug administration group (P1~P5 group) and a positive control group (Control group, C group) were set up.

[0072] In the treatment group, BMDCs fully combined with the tumor vaccine were co-cultured with mouse primary T lymphocytes to activate T cells. After 3 days, the activated T lymphocytes were co-cultured with Panc02 pancreatic cancer cells in vitro for 3 days. The model group used BMDCs without the tumor vaccine, and all other experimental conditions were the same as those in the treatment group. The positive control group used mouse primary T lymphocytes stimulated by CD3 / CD28, and all other experimental conditions were the same as those in the treatment group.

[0073] After culture, tumor cells in the logarithmic growth phase were collected, treated under different conditions, and then digested with EDTA-free trypsin. The cells were resuspended in pre-chilled PBS and the cell concentration was adjusted to approximately 1 × 10⁻⁶ cells / day. 6 Then take 100 μL of cell suspension (approximately 1 × 10⁻⁶ cells / mL). 5 Add 5 μL of Annexin V-FITC staining solution to a flow cytometry tube, mix gently, and incubate at room temperature in the dark for 15 minutes. Finally, add 5 μL of PI staining solution, mix gently, and place on ice in the dark. Perform flow cytometry analysis within 1 hour.

[0074] During flow cytometry analysis, the excitation and emission wavelengths of Annexin V-FITC were set to 488 nm and 525 nm (typically the FITC channel); the excitation and emission wavelengths of PI were set to 488 nm and 617 nm (typically the PE channel).

[0075] After obtaining the detection data, software such as FlowJo was used to analyze the data to distinguish between live cells (Annexin V⁻ / PI⁻), early apoptotic cells (Annexin V⁺ / PI⁻), and late apoptotic / necrotic cells (Annexin V⁺ / PI⁺), and to calculate the total apoptosis rate, which is the sum of the percentages of early apoptotic and late apoptotic cells.

[0076] Then, the Annexin V / PI method was used to detect tumor cell apoptosis, such as... Figure 2 As shown in the flow cytometry analysis, the tumor vaccines P1MHC~P5MHC (labeled P1~P5 in the figure) can all induce apoptosis in pancreatic cancer cells to a certain extent. In the treatment group P1, the apoptosis rate of Panc02 pancreatic cancer cells was the highest. Similarly, the CCK8 assay results show that the activated lymphocytes in the treatment group P1 exhibited the highest inhibition rate against Panc02 pancreatic cancer cells. Therefore, it is evident that the tumor vaccine described in this example can successfully kill and inhibit the growth of pancreatic cancer cells, and the tumor vaccine in Example 1 demonstrates relatively better performance.

[0077] (3) In vitro examination of the immune response of P1MHC~P5MHC in the serum of diseased mice

[0078] In this embodiment, the main focus is on detecting the levels of IFN-γ, TNF-α, IL-2, and Granzyme B in the serum of diseased mice. Elevated IFN-γ levels indicate a strong type I immune response, particularly effective activation of cellular immunity. Elevated TNF-α levels indicate a strong inflammatory response and the initiation of direct cytotoxic activity. Elevated IL-2 levels indicate that T cells are fully activated and have entered the proliferation and amplification phase. Elevated Granzyme B levels demonstrate that cytotoxic T cells and NK cells are activated and performing cytotoxic functions.

[0079] In this embodiment, a double-antibody sandwich ELISA method was used to detect the IFN-γ content in serum samples. Specifically, anti-IFN-γ antibody was first pre-coated onto the wells of a 96-well plate, and then a standard or sample and HRP-labeled detection antibody were added to form an immune complex. After washing, TMB substrate was added, producing a color change proportional to the IFN-γ concentration. Immediately after the color development was complete, 50 μL of stop solution was added, and the OD value was measured at 450 nm using a microplate reader. The concentration was determined using a standard curve, and the actual sample value was calculated using an appropriate dilution. The determination methods for the remaining indicators were the same as above.

[0080] In this embodiment, the test results of various indicators of the vaccines prepared in Examples 1 to 5 (labeled as P1 to P5 in the figure) are as follows: Figure 3As shown, the IFN-γ levels increased most significantly in the control group (C group), P1, P2, and P3 groups; the TNF-α levels were highest in the control group, P1, P2, and P3 groups, followed by P4 and P5; the IL-2 levels were highest in the control group, P1, P2, and P3 groups, followed by P4 and P5; the Granzyme B level was highest in the control group, followed by P1 and P3 groups, while the relative differences in P2, P4, and P5 were relatively low.

[0081] Based on the above test results, it can be seen that all five key neoantigen peptides protected by this invention have a significant inhibitory effect on tumor cell growth, with P1 showing the best inhibitory effect.

[0082] (4) In vivo testing of the inhibitory effect of P1MHC~P5MHC on tumor cells

[0083] In this embodiment, male C57BL / 6 mice aged 6-8 weeks were selected and divided into 8 groups of 6 mice each. These groups were labeled as the Blank group, Control group (C group), Model group (M group), and P1, P2, P3, P4, and P5 groups, as arranged according to Examples 1-5. The Control group received a vaccine containing CD3 / CD28-stimulated mouse primary T lymphocytes. Except for the Blank group, each mouse in each group received a subcutaneous injection of 5 × 10⁵ mmol / L of vaccine into the right axilla. 6 A single Panc02 pancreatic cancer cell can grow to 100 mm in size in about a week. 3 about.

[0084] In this embodiment, the tumor vaccine is administered via subcutaneous injection using an insulin needle, twice a week, at a dose of 20ug / 100ul / time. The detection standard is to measure the tumor volume every other day, using calipers to measure the longest and shortest points of the tumor, V= (where a is the major axis and b is the minor axis), and the mice were weighed every other day.

[0085] The results in this embodiment are as follows: Figure 4 As shown, compared with the model group, the control group, P1, P2, and P3 groups all exhibited extremely strong tumor growth inhibition capabilities. Throughout the observation period of 0-22 days, the growth of tumor volume in mice was significantly inhibited. At the end of the experiment, the tumor photographs and quantitative analysis of tumor mass further confirmed the above results, with the tumor mass of the control group, P1, P2, and P3 groups all being significantly lower than that of the model group.

[0086] In summary, the above results indicate that the control group, P1, P2, and P3 treatments all effectively inhibited tumor growth without causing a decrease in mouse body weight. The P1 and P3 groups also showed excellent immunomodulatory effects in previous immunological index analyses, suggesting that their anti-tumor effect is related to the activation of the body's immune response. Overall, the test results in this embodiment demonstrate that the peptides and vaccines composed of these peptides provided by this invention can effectively inhibit tumors, thus potentially enabling the treatment of cancer.

[0087] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A novel antigenic epitope peptide based on MHC-II presentation, characterized in that, The key epitope sequence of the peptide is: MVMGVLDQAFDVLVL; KVMGVLDQAFDVLVL; KVMGVLKQAFDVLVL; KVLGVLKQAFDVLVL; Any one of KVKGVLKQAFDVLVL.

2. The novel antigenic epitope peptide based on MHC-II presentation according to claim 1, characterized in that, The key epitope sequence of the peptide is MVMGVLDQAFDVLVL.

3. The novel antigenic epitope peptide based on MHC-II presentation according to claim 1, characterized in that, The full sequence of the neoantigen epitope peptide is as follows: MDAMKRGLCCVLLLCGAVFVSPS-GS-AKFVAAWTLKAAA-EAAAK-(α)-GPGPG-VVVGAVGVGKSALTI-GGGS-HHHHHH-GYQTI-RKRSHAGYQTI, where α is any one of the key epitope sequences of the peptide.

4. A tumor vaccine, characterized in that, It comprises the peptide as described in any one of claims 1 to 3 and a pharmaceutically acceptable auxiliary component.

5. A tumor vaccine according to claim 4, characterized in that, The auxiliary component includes a vaccine adjuvant, which is any one of poly(I:C), AS04, MF59, and CFA.

6. A tumor vaccine according to claim 5, characterized in that, The mass ratio of the peptide to the vaccine adjuvant is 1:(1~3).

7. The use of the peptide according to any one of claims 1 to 3 or the tumor vaccine according to any one of claims 4 to 6 in the preparation of a medicament for the prevention and / or treatment of tumor diseases.

8. The application according to claim 7, characterized in that, The drug is an injectable preparation, and the injection route is at least one of intramuscular injection, intravenous injection, subcutaneous injection, and intradermal injection.

9. The application according to claim 7, characterized in that, Drug treatment is available for at least one of the following cancers: pancreatic cancer, colorectal cancer, liver cancer, and stomach cancer.

10. A combination therapy for tumor treatment, characterized in that, The drug comprises a peptide as described in any one of claims 1 to 3 or a tumor vaccine as described in any one of claims 4 to 6, administered separately or simultaneously, and other drugs for treating tumor diseases.

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