CN112142824B

CN112142824B - Polypeptide with cytotoxic T lymphocyte inducing ability and application thereof

Info

Publication number: CN112142824B
Application number: CN201910565831.6A
Authority: CN
Inventors: 温瞳; 吕贵亮; 韩新峰
Original assignee: Shenzhen Donggui Bokang Biochemical Technology Co ltd
Current assignee: Shenzhen Qianyue Biotechnology Co ltd
Priority date: 2019-06-27
Filing date: 2019-06-27
Publication date: 2022-03-11
Anticipated expiration: 2039-06-27
Also published as: CN112142824A

Abstract

The invention relates to the technical field of antitumor immune drugs, and particularly provides a polypeptide with cytotoxic T lymphocyte inducing capacity and application thereof. The polypeptide is at least one of polypeptides with amino acid sequences of SEQ ID NO. 1-SEQ ID NO. 8. The polypeptide provided by the invention has a T cell mediated immunity function, can specifically kill cancer cells closely related to the overexpression of an estrogen receptor, greatly reduces the recurrence rate of cancers related to the overexpression of the estrogen receptor, and improves the cure rate.

Description

Polypeptide with cytotoxic T lymphocyte inducing ability and application thereof

Technical Field

The invention belongs to the technical field of antitumor immune drugs, and particularly relates to a polypeptide with cytotoxic T lymphocyte inducing capacity and application thereof.

Background

Breast cancer is the first most common cancer in women, and the number of patients has a tendency to climb year after year, making treatment of breast cancer a great challenge.

Estrogen receptor, progesterone receptor and epithelial growth factor receptor 2 are indispensable identification factors for internationally recognized use in the formulation of clinical classification, diagnosis and treatment regimens for breast cancer. According to the tissue and clinical characteristics of breast cancer growth, breast cancer is divided into luminal breast cancer, invasive breast cancer and triple negative breast cancer, wherein luminal breast cancer is clinically characterized by estrogen receptor overexpression and/or progestogen receptor overexpression. Clinical statistics data show that more than 70% of breast cancers are closely related to the overexpression of estrogen receptors, the estrogen receptors are large proteins encoded by estrogen genes and play important roles in starting the downstream signal transduction in cells, regulating the growth, proliferation and differentiation of the cells and the like, and the overexpression of the estrogen receptors enables the transcriptional and translational overactivation of the cells, so that malignant proliferation of cancer cells such as the breast cancers is caused.

With the development of molecular biology and the gradual improvement of gene and second-generation sequencing technologies, more and more clinical researches have found that estrogen receptor blockers, especially Aromatase inhibitor (Aromatase I), used in the treatment of breast cancer cause multiple genetic variations of ligand binding domains in estrogen receptors. Clinical studies have found that about 53% of breast cancer patients have genetic variation of tyrosine kinase. The mutant gene causes the drug resistance and cure rate of breast cancer patients to be reduced, primary breast cancer is easy to relapse, and the risk of transferring the breast cancer to bones and internal organs is increased. Therefore, it is imperative to find new methods or drugs for treating breast cancer, especially immunotherapy against the overexpression and gene variation region of estrogen receptor.

Disclosure of Invention

Aiming at the problems that the prior estrogen receptor gene is over-expressed in cancer and an aromatase inhibitor used in clinical treatment is easy to cause gene variation of a ligand binding domain in the estrogen receptor, and the like, the invention provides a polypeptide with the capability of inducing cytotoxic T lymphocytes and application thereof.

In order to achieve the purpose, the technical scheme of the invention is as follows:

a polypeptide with the ability of inducing cytotoxic T lymphocytes, wherein the polypeptide is at least one of polypeptides with amino acid sequences of SEQ ID NO. 1-SEQ ID NO. 8;

wherein,

SEQ ID NO.1LLQCAWLEI；

SEQ ID No.2YTFLPSTLK；

SEQ ID No.3YLSGLLLEM；

SEQ ID No.4VVPLSGLLL；

SEQ ID No.5YLSDLLLEV；

SEQ ID No.6VYTFLPSTL；

SEQ ID No.7ARHGGWTTK；

SEQ ID No.8LYGLLLEML。

further, the use of the polypeptide having an ability to induce cytotoxic T lymphocytes as described above for the preparation of a pharmaceutical composition or a preparation for preventing or treating cancer.

A pharmaceutical composition or formulation for the prevention or treatment of cancer, comprising an active pharmaceutical ingredient comprising a polypeptide having the ability to induce cytotoxic T lymphocytes as described above, and a pharmaceutically acceptable carrier.

A cancer vaccine comprising an antigen for inducing a human body to produce a corresponding antibody, the antigen for inducing a human body to produce a corresponding antibody comprising a polypeptide having an ability to induce cytotoxic T lymphocytes as described above.

The invention has the technical effects that:

compared with the prior art, the polypeptide with the ability of inducing cytotoxic T lymphocytes provided by the invention can be recognized by a T cell receptor, has the function of T cell mediated immunity, can specifically kill cancer cells closely related to the overexpression of the estrogen receptor, greatly reduces the recurrence rate of cancers related to the overexpression of the estrogen receptor, and improves the cure rate.

The polypeptide with the ability of inducing cytotoxic T lymphocytes is applied to the preparation of the pharmaceutical composition or preparation for preventing or treating cancers, mainly because the polypeptide has the function of T cell mediated immunity, can specifically kill cancer cells closely related to the overexpression of estrogen receptors, greatly reduces the recurrence rate of the cancers, improves the cure rate, and is prepared into the pharmaceutical composition or preparation for preventing the cancers or the pharmaceutical composition or preparation for treating the cancers, thereby having specific treatment effect.

The active pharmaceutical components of the pharmaceutical composition or the preparation for preventing or treating the cancer provided by the invention comprise the polypeptide with the ability of inducing cytotoxic T lymphocytes, so the pharmaceutical composition or the preparation has the function of T cell mediated immunity, can specifically kill cancer cells closely related to the overexpression of estrogen receptors, greatly reduces the recurrence rate of the cancer and improves the cure rate.

The antigen of the cancer vaccine provided by the invention contains polypeptide with the ability of inducing cytotoxic T lymphocyte, so that the cancer vaccine has good immune response effect on cancer closely related to estrogen receptor overexpression.

Drawings

In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art that other drawings can be obtained according to these drawings without creative efforts.

FIG. 1 is a schematic representation of the data of a polypeptide activity scan during the screening of a polypeptide of the present invention;

FIG. 2 is a schematic diagram showing the specific T cell killing activity of the polypeptides screened by the present invention according to the combination of single peptide, double peptide and tripeptide;

FIG. 3 is a schematic diagram showing the killing activity of the polypeptide of the present invention in combination with specific T cells;

FIG. 4 is a schematic diagram of the anti-cancer effect of the polypeptide combination 1 and the blank control group 1 in an extraneous tumor animal model;

FIG. 5 is a schematic diagram showing the anti-cancer effect of the polypeptide combination 2 and the blank control group 2 in an extraneous tumor animal model;

FIG. 6 is a schematic diagram of the anti-cancer effect of the polypeptide combination 3 and the blank control group 3 in the extraneous tumor animal model.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below 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 are not intended to limit the invention.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The terms "polypeptide", "peptide" and "protein" are used interchangeably herein to refer to a polymer of amino acid residues, and are applicable not only to naturally occurring amino acid polymers, but also to amino acid polymers in which one or more amino acid residues are modified or non-naturally occurring residues, such as artificial chemical mimetics of the corresponding naturally occurring amino acids, and the like.

As used herein, the term "amino acid" refers to both naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function similarly to naturally occurring amino acids. Naturally occurring amino acids refer to amino acids encoded by the genetic code, as well as amino acids that are post-translationally modified in a cell.

Amino acids may be referred to herein by their commonly known three-letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical nomenclature Commission.

Unless otherwise defined, the term "cancer" refers to cancers that overexpress the estrogen receptor gene, including, e.g., breast, ovarian, prostate, liver cancers.

The terms "cytotoxic T lymphocyte," "cytotoxic T cell," and "CTL" are used interchangeably herein and refer to a subpopulation of T lymphocytes that are capable of recognizing non-self cells (e.g., tumor cells, virally infected cells) and inducing death of such cells.

In one aspect of the invention, a polypeptide having the ability to induce cytotoxic T lymphocytes is provided.

The polypeptide is selected from at least one of amino acid sequences SEQ ID NO. 1-SEQ ID NO. 8; wherein,

SEQ ID NO.1LLQCAWLEI；

SEQ ID No.2YTFLPSTLK；

SEQ ID No.3YLSGLLLEM；

SEQ ID No.4VVPLSGLLL；

SEQ ID No.5YLSDLLLEV；

SEQ ID No.6VYTFLPSTL；

SEQ ID No.7ARHGGWTTK；

SEQ ID No.8LYGLLLEML。

the technical solution of the present invention is explained in further detail below.

The above-mentioned polypeptides of SEQ ID NO.1 to SEQ ID NO.8 of the present invention can be obtained by a chemical synthesis method based on the selected amino acid sequence, or can be obtained by applying any known genetic engineering peptide production method. The amino acid sequence of each polypeptide can replace, delete or add one, two or three amino acids or add modifying groups, the amino acid sequence obtained by replacing, deleting or adding one, two or three amino acids or modifying groups, and the processed amino acid sequence can retain the original function of recognizing ability by a T cell receptor, and belongs to the polypeptide with the ability of inducing cytotoxic T lymphocytes. In addition, the polypeptide with 8 amino acid sequences has specific adaptability to immune complexes of main tissues of Asian ethnic group.

The synthesized polypeptide has a modified N-terminal and a modified C-terminal, and the amino acids in the synthesized polypeptide series comprise various biological modifications, such as glycosylation modification, methylation modification, basic modification, aldehyde modification, sulfhydrylation modification, phosphorylation modification and the like. The modifying group may be specifically anywhere in the polypeptide, including the backbone, amino acids, side chains and amino or carboxyl termini, etc., and may be chemically modified, such as chemically modified from the group consisting of pseudouridine, N1-methylpseudouridine, N1-ethylpseudouridine, 2-thiouridine, 4' -thiouridine, 5-methylcytosine, 2-thio-1-methyl-1-deaza-pseudouridine, 2-thio-1-methyl-pseudouridine, 2-thio-5-aza-uridine, 2-thio-dihydropseudouridine, 2-thio-dihydrouridine, 2-thio-pseudouridine, 4-methoxy-pseudouridine, N-acetyluridine, etc, 4-thio-1-methyl-pseudouridine, 4-thio-pseudouridine, 5-aza-uridine, dihydropseudouridine, 5-methyluridine, 5-methoxyuridine and 2' -O-methyluridine.

As a preferred embodiment, the polypeptides of SEQ ID NO. 1-8 are combined to form a polypeptide combination, which can effectively avoid the immune escape of a single polypeptide in T cell mediated immunotherapy. Such as the amino acid sequences of SEQ ID NO.1, SEQ ID NO.3 and SEQ ID NO.5 are combined to form a polypeptide combination 1; combining the amino acid sequences of SEQ ID No.2, SEQ ID No.4 and SEQ ID No.7 to form polypeptide combination 2; the amino acid sequences of SEQ ID NO.6 and SEQ ID NO.8 are combined to form the polypeptide combination 3, which has excellent immunotherapy relative to a single polypeptide and solves the problem of immune escape of the single polypeptide. Especially has good treatment effect on metastatic breast cancer and recurrent breast cancer, greatly reduces the recurrence rate of the breast cancer and improves the cure rate. The polypeptide combination 1, the polypeptide combination 2 and the polypeptide combination 3 are all obtained by physically mixing a plurality of polypeptides, but a plurality of polypeptides are not connected into a single polypeptide.

The polypeptide is matched with the content of main immune tissues of Asian race, particularly Chinese race, so that the polypeptide can specifically act on the Asian race, particularly Chinese race, and T cell mediated immunotherapy for the Asian race, particularly Chinese race is realized.

Since the polypeptide having an ability to induce cytotoxic T lymphocytes provided by the present invention has an ability to induce highly cytotoxic T lymphocytes, it can be contacted with antigen presenting cells to induce highly cytotoxic T lymphocytes. Specifically, at least one polypeptide with the amino acid sequence of SEQ ID NO. 1-SEQ ID NO.8 is subjected to immunization, and T lymphocytes with high cytotoxicity can be induced, so that the overexpressed estrogen receptor tumor cells can be specifically killed.

Preferably, the antigen-presenting cell is contacted with any one of the above-mentioned polypeptide combination 1, polypeptide combination 2, and polypeptide combination 3.

The polypeptide with the ability of inducing the cytotoxic T lymphocyte provided by the invention has the ability of inducing the high cytotoxic T lymphocyte, so that the polypeptide can be conjugated (conjugated) or connected (attached) with an anti-cancer substance, can be combined with a detected marker, and can also be combined with the anti-cancer substance and the marker. Such as in combination with chemotherapeutic drugs to form conjugates. The marker combination with the assay may be a direct or indirect marker, including but not limited to fluorescent markers, bioluminescent markers, visible light markers, enzyme markers, radioactive markers, affinity agent markers, and the like. Therefore, the polypeptide with the ability of inducing cytotoxic T lymphocytes provided by the invention can be applied to the preparation of cancer prevention pharmaceutical compositions, cancer prevention preparations, cancer treatment pharmaceutical compositions and cancer treatment preparations.

The present invention thus also provides any of a pharmaceutical composition for preventing cancer, an agent for preventing cancer, a pharmaceutical composition for treating cancer, and an agent for treating cancer, which comprise the above-described polypeptide having an ability to induce cytotoxic T lymphocytes as a pharmaceutically active ingredient. The pharmaceutical composition for preventing cancer, the agent for preventing cancer, the pharmaceutical composition for treating cancer and the agent for treating cancer have good effects on the prevention and treatment of cancers or tumors closely related to the overexpression of estrogen receptors.

Generally, cancers or tumors that are closely related to the overexpression of estrogen receptors, such as breast cancer, ovarian cancer, prostate cancer, liver cancer, and the like. Therefore, the cancer prevention pharmaceutical composition or the preparation may be any one of a pharmaceutical composition for preventing breast cancer, a preparation for preventing breast cancer, a pharmaceutical composition for preventing ovarian cancer, a preparation for preventing ovarian cancer, a pharmaceutical composition for preventing prostate cancer, a preparation for preventing prostate cancer, a pharmaceutical composition for preventing liver cancer, and a preparation for preventing liver cancer.

The pharmaceutical composition or preparation for treating cancer can be any one of a pharmaceutical composition for treating breast cancer, a preparation for treating breast cancer, a pharmaceutical composition for treating ovarian cancer, a preparation for treating ovarian cancer, a pharmaceutical composition for treating prostate cancer, a preparation for treating prostate cancer, a pharmaceutical composition for treating liver cancer and a preparation for treating liver cancer.

The pharmaceutical composition or preparation for preventing cancer and the pharmaceutical composition or preparation for treating cancer may further comprise a pharmaceutically acceptable carrier, and other polypeptides having the ability to induce cytotoxic T cells against cancer cells, other polynucleotides encoding the other polypeptides, and the like. In the pharmaceutical composition or preparation for preventing cancer and the pharmaceutical composition or preparation for treating cancer, the concentration of the polypeptide (as a pharmaceutically active ingredient) having the ability to induce cytotoxic T lymphocytes is (1-1000) μmol/L, and the concentration can be adjusted adaptively according to diseases, age, weight and the like of a patient during specific use.

The method of administration may be oral, intradermal, subcutaneous, intravenous injection, etc., systemic administration or local administration to the vicinity of the target site. Administration may be effected by a single administration, or may be enhanced by multiple administrations.

In addition, the pharmaceutical composition or preparation for preventing cancer, or the pharmaceutical composition or preparation for treating cancer may further comprise an adjuvant known to be effective in establishing cellular immunity. Adjuvants may be cytokines that stimulate myeloid stem cells to form granulocytes, macrophage colonies, interleukin 2, Toll-like receptor series, aluminum phosphate, aluminum hydroxide, alum, cholera toxin, salmonella toxin, and the like.

The polypeptide with the ability of inducing the cytotoxic T lymphocyte provided by the invention can be used as a vaccine, particularly a cancer vaccine, because the polypeptide has the ability of inducing the high cytotoxic T lymphocyte. Such a cancer vaccine comprises an antigen for inducing a human body to produce a corresponding antibody, which comprises a polypeptide having an ability to induce cytotoxic T lymphocytes as described above.

The cancer vaccine has specific immune response effect on cancers or tumors targeting human estrogen receptor gene variation regions and tyrosine kinase variation gene regions.

Preferably, the active ingredient of the cancer vaccine is any one of polypeptide combination 1, polypeptide combination 2, and polypeptide combination 3.

The cancer vaccine has specific immunity and therapeutic effect especially on the immunity of the luminal breast cancer.

In order to more effectively explain the technical solution of the present invention, the technical solution of the present invention is explained below by a plurality of specific examples.

Example 1

A method for screening a polypeptide having an ability to induce cytotoxic T lymphocytes, comprising the steps of:

(a) designing and synthesizing 39 polypeptides by using a conventional genetic engineering peptide production method, wherein the polypeptides comprise polypeptides with the numbers of A1-A37, negative control group polypeptides and positive control group polypeptides.

(b) Will T&B hybrid cells (Shanghai and Yuan biol Ltd.) were plated at 0.5X 10⁶The above polypeptides were added to each cell in a 24-well plate at 100. mu. mol/L, and cultured at 37 ℃ for 4 hours in a 5% carbon dioxide incubator.

(c) The cultured cells were collected, washed with physiological saline, and stained with an anti-human tissue-compatible (MHC) class I antibody for 30 min. As a negative control to which no polypeptide was added, a polypeptide (YLLPAIVHI, from Cytometry 41:271-278, 2000) with a known positive reaction was set as a positive control. After staining, the cells were washed 2 times, and fluorescence intensity data were read by a flow cytometer (BD Caliber), and the results are shown in FIG. 1.

As can be seen from FIG. 1, the binding activity of different polypeptide sequences to T & B hybrid cells is significantly different. The polypeptides having fluorescence intensity lower than or close to that of the negative control, such as A5, A6, A12, A19, A21, A27, A31, A34 and A37 in FIG. 1, show low ability of the polypeptide to form a complex with an antigen-presenting cell and low binding activity, and are not in a preferred range because these polypeptide sequences are not presented as epitope sequences recognizable by T cells by the antigen-presenting cell. Other polypeptide sequences basically show strong binding activity, and eight polypeptides with amino acid sequences of SEQ ID NO. 1-SEQ ID NO.8 are finally screened by combining the stability of the polypeptides, the killing property on targeted cancer cells and other factors.

Wherein, SEQ ID NO. 1LLQCAWLEI; SEQ ID No. 2YTFLPSTLK; SEQ ID No.3 YLSGLLEM; SEQ ID No.4 VVPSLLL; SEQ ID No. 5YLSDLLEV; SEQ ID No.6 VYTFLPSTL; SEQ ID No. 7ARHGGWTTK; SEQ ID No.8 LYGLLLEML.

Example 2

Experiment on killing activity of eight polypeptide specificity CD 8T killing cells with amino acid sequences of SEQ ID No. 1-SEQ ID No.8 on cancer cells

(1) Enriching dendritic cells expressing CD14, CD11c from healthy human peripheral blood cells

Human peripheral blood cells are placed in an incubator at 37 ℃ and are kept still for 1-2h, adherent cells are used as dendritic cell precursors, suspension cells are used as T cell aggregates, and stimulation is carried out for 5d by using 10ng/mL of interleukin-7 (IL7), interleukin-2 (IL2) and interleukin-15 (IL 15).

(2) Stimulation of effector cells by polypeptide combinations

Five days later the cells were mixed and stimulated with single polypeptide SEQ ID No.1, single polypeptide SEQ ID No.3, single polypeptide SEQ ID No.5, a dipeptide combination (i.e. SEQ ID No.1 and SEQ ID No.3, or SEQ ID No.1 and SEQ ID No.5), polypeptide combination 1 (i.e. the combination of SEQ ID No.1, SEQ ID No.3 and SEQ ID No.5), polypeptide combination 2 (i.e. the combination of SEQ ID No.2, SEQ ID No.4 and SEQ ID No. 7), polypeptide combination 3 (i.e. the combination of SEQ ID No.6 and SEQ ID No. 8) at a concentration of 100. mu. mol/L for 4h, respectively, after which the culture broth was changed every two days. After 10d, cells were collected and counted by washing with physiological saline, and the cells were used as effector cells.

(3) Mixing of effector cells with target cells

Taking the breast cancer cell MCF7, the ovarian cancer cell OVCAR3 and the melanoma cell A375 as target cells, and enabling the effector cells (T killer cells) obtained in the step (2) and the target cells to be 20: 1. 10: 1. 5: 1 to obtain mixed cells of effector cells and breast cancer cells MCF7, effector cells and ovarian cancer cells OVCAR3 and effector cells and melanoma cells A375, staining the mixed cells of each group by fluorescence, and reading the fluorescence intensity by a fluorescence spectrophotometer.

Melanoma cells A375 as target cells in the experiment are cells which basically do not express estrogen receptors, and therefore have no relevant negative control in the experiment, and specific results are shown in FIGS. 2 and 3.

As can be seen from FIG. 2, the killing activity of the polypeptide combination 1 targeting the CD 8T cells of MCF7 is obviously better than that of the single polypeptide SEQ ID No.1, the single polypeptide SEQ ID No.3, the single polypeptide SEQ ID No.5 and the dipeptide combination (i.e. SEQ ID No.1 and SEQ ID No.3 or SEQ ID No.1 and SEQ ID No.5) targeting the CD 8T cells of MCF 7.

As can be seen from fig. 3, for polypeptide combination 1 (i.e., the combination of SEQ ID No.1, SEQ ID No.3 and SEQ ID No.5), it had significant CD 8T cell killing activity and the ratio of effector cells to target cells was 20: 1, the CD 8T killer cells killed about 45% of the breast cancer cells MCF7 and about 30.1% of the ovarian cancer cells OVCAR3, while melanoma cells a375 were only killed by about 9.8% due to very low estrogen receptor expression. From the killing effect on breast cancer cells MCF7, the ratio of effector cells to target cells was from 5: 1 to 20: 1, the killing rate of CD 8T cells increased from 23% to 45%.

It also showed significant CD 8T cell killing activity for polypeptide combination 2 (i.e., the combination of SEQ ID No.2, SEQ ID No.4 and SEQ ID No. 7). In the ratio of effector cells to target cells of 20: 1, the breast cancer cell MCF7 underwent 40% killing, the ovarian cancer cell OVCAR3 was also killed by 23%, and the melanoma cell A375, which has very low estrogen receptor expression, had a killing rate of only 8.5%.

For polypeptide combination 3 (i.e., the combination of SEQ ID No.6 and SEQ ID No. 8), it showed a CD 8T cell killing activity that was substantially similar to that of polypeptide combination 1, in that the ratio of effector cells to target cells was 20: 1, can kill 48% of breast cancer cells MCF7 and about 28% of ovarian cancer cells OVCAR 3.

The results of the three groups of polypeptide combinations show that the killing rate of CD 8T cells is increased in proportion with the increase of effector cells. The data show that the polypeptide combination has higher killing activity of CD 8T cells compared with single peptide, and can be recognized by T cells as epitope sequences, and the screened polypeptide combination has higher killing activity of CD 8T cells and has more obvious curative effect, so the polypeptide combination is more suitable for being used as a medicament or a vaccine. Since breast cancer cells are over-expressed with the strongest estrogen receptors compared to ovarian cancer and melanoma, polypeptide-specific CD 8T killer cells also showed the highest killing activity.

Example 3

Anticancer effect experiment of external tumor animal model

(1) 8 weeks old immune cells T&SCID female mice with double B-cell depletion (supplied by Shanghai and Yuan biol Ltd.) and mammary gland region at 10⁶Individual cells were seeded with estrogen receptor positive, previously bioluminescent labeled breast cancer cells MCF 7. After culturing for about 1 week to 10 days, the mammary part has accessible tumor, and the initial tumor size is recorded by a bioluminescence instrument.

(2) Polypeptide combination 1 (i.e., the combination of SEQ ID No.1, SEQ ID No.3 and SEQ ID No.5), polypeptide combination 2 (i.e., the combination of SEQ ID No.2, SEQ ID No.4 and SEQ ID No. 7), and polypeptide combination 3 (i.e., the combination of SEQ ID No.6 and SEQ ID No. 8), each dissolved in physiological saline, were administered by intraperitoneal Injection (IP) at a concentration of 100 μmol/L2 times a week for a total of four weeks, with a corresponding blank control set for each group, and physiological saline in each of blank controls 1, 2 and 3. Tumor size was recorded weekly using a bioluminescence instrument.

Fig. 4 shows bioluminescence absorbance pictures a1 and a2 for tumor sizes of polypeptide combination 1 and placebo 1;

fig. 5 shows bioluminescence absorbance pictures B1 and B2 of tumor size for polypeptide combination 2 and placebo 2;

fig. 6 shows bioluminescence absorbance pictures C1 and C2 of tumor size for polypeptide combination 3 and placebo 3.

Performing density analysis on the bioluminescence absorption of the tumors in fig. 4-6, and comparing the bioluminescence absorption with a physiological saline blank control group, wherein the polypeptide combination group shows a significant inhibition effect on the growth of the animal tumor, for example, the tumor of the polypeptide combination 1 is reduced to 62% of that of the blank control group 1, and the P value is 0.02; the tumor of polypeptide combination 2 decreased to 76% of that of placebo 2, with a P value of 0.05; the tumor of the polypeptide combination 3 is reduced to 60 percent of that of the physiological saline blank control group 3, the P value is 0.0061, and the statistical significance is remarkable. The number of CD 8T cells positive in the tumor was analyzed, and the polypeptide combination group had a significant increase in human CD45, CD3, CD8 positive cells over the saline blank control group (data not shown).

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the present invention. Any modification, equivalent replacement, and improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A polypeptide with the ability of inducing cytotoxic T lymphocytes is characterized in that the polypeptide is at least one selected from the polypeptides with amino acid sequences of SEQ ID NO. 1-SEQ ID NO. 8;

wherein,

SEQ ID NO.1 LLQCAWLEI；

SEQ ID No.2 YTFLPSTLK；

SEQ ID No.3 YLSGLLLEM；

SEQ ID No.4 VVPLSGLLL；

SEQ ID No.5 YLSDLLLEV；

SEQ ID No.6 VYTFLPSTL；

SEQ ID No.7 ARHGGWTTK；

SEQ ID No.8 LYGLLLEML。

2. the polypeptide having the ability to induce cytotoxic T lymphocytes according to claim 1, wherein said polypeptide is polypeptide combination 1 comprising SEQ ID No.1, SEQ ID No.3 and SEQ ID No. 5; or a polypeptide combination 2 comprising SEQ ID NO.2, SEQ ID NO.4 and SEQ ID NO. 7; or a polypeptide combination 3 comprising SEQ ID NO.6 and SEQ ID NO. 8.

3. Use of the polypeptide having an ability to induce cytotoxic T lymphocytes according to claim 2 for the preparation of a pharmaceutical composition or a preparation for the treatment of cancer, which is breast cancer or ovarian cancer.

4. A pharmaceutical composition or formulation for the treatment of cancer, whether breast or ovarian, comprising an active pharmaceutical ingredient comprising a polypeptide having the ability to induce cytotoxic T lymphocytes of claim 2 and a pharmaceutically acceptable carrier.

5. A vaccine for cancer, wherein the cancer is breast cancer or ovarian cancer, comprising an antigen for inducing production of a corresponding antibody in a human, wherein the antigen for inducing production of a corresponding antibody in a human comprises the polypeptide of claim 2 having the ability to induce cytotoxic T lymphocytes.