CN109608536B

CN109608536B - Epitope polypeptide CD44-P3 based on prostate cancer stem cell marker CD44 and application thereof

Info

Publication number: CN109608536B
Application number: CN201811521662.8A
Authority: CN
Inventors: 王铸; 梁辉; 邓琼; 张颖; 张建文; 植凡; 胡七一; 张圣平
Original assignee: Shenzhen Longhua Peoples Hospital
Current assignee: Shenzhen Longhua Peoples Hospital
Priority date: 2018-12-12
Filing date: 2018-12-12
Publication date: 2020-09-15
Anticipated expiration: 2038-12-12
Also published as: CN109608536A

Abstract

The invention provides an epitope polypeptide CD44-P3 based on a prostate cancer stem cell marker CD44 and application thereof, relates to the technical field of biomedical engineering, and provides an epitope polypeptide based on a prostate cancer stem cell marker CD44 and application thereof.

Description

Epitope polypeptide CD44-P3 based on prostate cancer stem cell marker CD44 and application thereof

Technical Field

The invention relates to the technical field of biomedical engineering, in particular to an epitope polypeptide CD44-P3 based on a prostate cancer stem cell marker CD44 and application thereof.

Background

At present, the incidence and mortality of prostate cancer are on the rising trend year by year, and the prostate cancer is the first killer seriously threatening the health of human beings. Despite the ongoing advances in surgical, chemotherapeutic, radiotherapeutic and molecular targeted therapies for prostate cancer, effective treatment of prostate cancer remains a critical issue that remains urgently to be addressed.

In recent years, with the continuous disclosure of molecular mechanism and immune mechanism of prostate cancer, prostate cancer specific immunotherapy is increasingly emphasized, and the key to tumor immunotherapy is to obtain ideal tumor antigen. With the rapid development of the immunological theory method and the molecular biology method, a great amount of tumor antigens capable of inducing the immune response of organisms are discovered, and a new era is opened up for the tumor immunotherapy.

The theory of "tumor stem cells" is that a group of cells exists in tumor tissues, which have the ability of self-renewal and can generate heterogeneous tumor cell groups, and are closely related to the proliferation, metastasis, recurrence and insensitivity to radiotherapy and chemotherapy of tumors, and are ideal targets for tumor-specific immunotherapy. In recent years, CD133 has been used⁺/CD44⁺After a phenotypic prostate cancer stem cell is discovered, its characteristics of stem cell function have been demonstrated in several ways.

In recent years, as the molecular mechanism of immune response has been studied, it has been gradually recognized that immune cells of the body do not correspond to the whole molecules of various pathogens or natural antigens, but are directed to epitopes of various antigen molecules, i.e., protein antigens, by which the immunospecificity is expressed. Research shows that tumor antigen recognized by CD8+ T cell needs to be treated by antigen presenting cell and then presented on the surface of antigen presenting cell or target cell in the form of "antigen peptide-MHC-I molecule" complex, and the corresponding antigen peptide combined with MHC-I molecule is CTL epitope. The existing anti-tumor treatment technology has low specificity, limited killing effect and easy tumor recurrence.

In view of the above, the present invention is particularly proposed.

Disclosure of Invention

The invention aims to solve the problems in the prior art and provides an epitope polypeptide CD44-P3 based on a prostate cancer stem cell marker CD44 and application thereof.

In order to solve the above problems, the technical solution adopted by the present invention is as follows:

the invention provides an isolated epitope polypeptide CD44-P3, wherein the epitope polypeptide CD44-P3 is selected from the following (a) or (b):

(a) 1, polypeptide consisting of an amino acid sequence shown in SEQ ID NO;

(b) 1 through substitution and/or deletion and/or addition of one or more amino acid residues, and has the functions of the epitope of the prostate cancer stem cell marker CD 44.

Further, the epitope polypeptide CD44-P3 has the molecular weight of 1000-1300.

Further, the epitope polypeptide CD44-P3 is obtained by expression and purification of prokaryotic cells or eukaryotic cells; or the like, or, alternatively,

the epitope polypeptide CD44-P3 is synthesized artificially.

The invention also provides a nucleic acid, which encodes the epitope polypeptide CD 44-P3.

Further, the nucleic acid is selected from any one of the following (A) to (C):

(A) a nucleotide sequence represented by SEQ ID NO. 2;

(B) 1, a polynucleotide encoding the polypeptide shown in SEQ ID NO;

(C) nucleotide which has more than 80 percent of homology with the nucleotide sequence shown in SEQ ID NO. 2 and codes polypeptide with prostate cancer stem cell marker CD44 epitope function.

The invention also provides a vector comprising the nucleic acid as described above.

The invention also provides a host cell comprising a nucleic acid or vector as described above.

In addition, the invention also provides application of the epitope polypeptide CD44-P3 in preparation of products for preventing and/or treating prostate cancer.

Further, the product is a vaccine or a medicament.

Further, the epitope polypeptide CD44-P3 can prevent and/or treat prostatic cancer by killing LNCAP-multicellular spheroids; and/or the presence of a gas in the gas,

the epitope polypeptide CD44-P3 prevents and/or treats prostate cancer by killing VCaP-multicellular spheroids.

The separated epitope polypeptide CD44-P3 provided by the invention is an epitope polypeptide based on a prostate cancer stem cell marker CD 44. The epitope polypeptide CD44-P3 based on the prostate cancer stem cell surface marker CD44 is designed and synthesized through a bioinformatics approach, the epitope polypeptide CD44-P3 has good immunogenicity, can specifically activate cytotoxic T lymphocytes, has a good killing effect on the prostate cancer stem cell, and lays a good foundation for the research of the epitope of the prostate cancer stem cell surface marker.

The epitope polypeptide CD44-P3 based on the prostate cancer stem cell marker CD44 provided by the invention can stimulate an organism to generate protective immune response against prostate cancer stem cells, so that the inhibition and the clear effect on the prostate cancer stem cells are enhanced.

Drawings

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

FIG. 1 is a diagram of mass spectrometry analysis of epitope polypeptide CD44-P3 provided in example 1 of the present invention;

FIG. 2 is a high performance liquid chromatography analysis chart of epitope polypeptide CD44-P3 provided in example 1 of the present invention;

FIG. 3A is a graph showing the results of the action of the antigen-epitope polypeptide CD44-P3 on LNCAP-multicellular spheres in example 2 of the present invention;

FIG. 3B is a graph showing the killing effect of antigen-epitope polypeptide CD44-P3 on LNCAP-multicellular spheroids in example 2 of the present invention;

FIG. 4A is a schematic diagram showing the results of the action of the antigen-epitope polypeptide CD44-P3 on VCaP-multicellular spheres in example 3 of the present invention;

FIG. 4B is a graph showing the killing effect of the antigen-epitope polypeptide CD44-P3 on VCaP-multicellular spheroids in example 3 of the present invention.

Detailed Description

The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments, and it should be understood that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

It should be noted that:

in the present invention, all the embodiments and preferred methods mentioned herein can be combined with each other to form a new technical solution, if not specifically stated.

In the present invention, all the technical features mentioned herein and preferred features may be combined with each other to form a new technical solution, if not specifically stated.

In the present invention, unless otherwise specified, the individual reactions or operation steps may be performed sequentially or may be performed in sequence. Preferably, the reaction processes herein are carried out sequentially.

Unless otherwise defined, technical and scientific terms used herein have the same meaning as is familiar to those skilled in the art. In addition, any methods or materials similar or equivalent to those described herein can also be used in the present invention.

According to one aspect of the present invention, there is provided an isolated epitope polypeptide CD44-P3, wherein the epitope polypeptide CD44-P3 is selected from the group consisting of (a) or (b):

(a) 1, polypeptide consisting of an amino acid sequence shown in SEQ ID NO;

Wherein SEQ ID NO. 1 is Asp-Gln-Phe-Met-Thr-Ala-Asp-Glu-Thr-Arg-Asn.

In the present application, the term "epitope polypeptide" refers to any polypeptide fragment that can be recognized by a specific antibody.

The human CD44 gene is located in the short arm of chromosome 11, has a total length of about 50kb, and consists of 20 highly conserved exons, each exon has a length of 70-210bp, and is separated by introns with different lengths. CD44 is a cell surface molecule, plays a functional role in cell adhesion and signal transduction, is used as a tumor marker of the prostate cancer stem cell, has a very strong epitope, and the epitope polypeptide based on the prostate cancer stem cell marker CD44 provided by the invention has stronger immunogenicity and killing effect on the prostate cancer stem cell.

The substitution and/or deletion and/or addition of one or more amino acid residues can be the substitution and/or deletion and/or addition of 1-10 amino acid residues; the substitution and/or deletion and/or addition of one or several amino acid residues may occur outside the above-mentioned domains.

In some preferred embodiments, the epitope polypeptide CD44-P3 has a molecular weight of 1000-1300, and can be, for example, but not limited to 1202.35, 1208.26 or 1084.18, preferably 1084.18.

In some preferred embodiments, the epitope polypeptide CD44-P3 is obtained by expression and purification in prokaryotic cells or eukaryotic cells; or the like, or, alternatively,

the epitope polypeptide CD44-P3 is synthesized artificially.

In some embodiments, the invention also provides a nucleic acid encoding the epitope polypeptide CD44-P3 described above.

Herein, a nucleic acid sequence comprises conservatively substituted variants thereof (e.g., substitution of degenerate codons) and complementary sequences. The terms "nucleic acid" and "polynucleotide" are synonymous and encompass genes, cDNA molecules, mRNA molecules, and fragments thereof such as oligonucleotides.

In some preferred embodiments, the nucleic acid is selected from any one of the following (a) - (C):

(A) a nucleotide sequence represented by SEQ ID NO. 2;

(B) 1, a polynucleotide encoding the polypeptide shown in SEQ ID NO;

(C) and the nucleotide has more than 80 percent of homology with the nucleotide sequence shown in SEQ ID NO. 2 and codes the polypeptide with the prostate cancer stem cell marker CD44 epitope function.

SEQ ID NO. 2 of the sequence Listing is gaccagtttatgacagctgatgagacaaggaac.

In some embodiments, the invention also provides an expression vector comprising a nucleic acid sequence encoding the epitope polypeptide CD 44-P3.

In the present invention, a vector may refer to a molecule or an agent comprising a nucleic acid of the present invention or a fragment thereof, capable of carrying genetic information and capable of delivering the genetic information into a cell. Typical vectors include plasmids, viruses, bacteriophages, cosmids and minichromosomes. The vector may be a cloning vector (i.e., a vector for transferring genetic information into a cell, which may be propagated and in which the genetic information may be present or absent) or an expression vector (i.e., a vector which comprises the necessary genetic elements to permit expression of the genetic information of the vector in a cell). Thus, a cloning vector may contain a selectable marker, as well as an origin of replication compatible with the cell type specified by the cloning vector, while an expression vector contains the regulatory elements necessary to effect expression in a specified target cell.

The nucleic acid of the invention or fragments thereof may be inserted into a suitable vector to form a cloning or expression vector carrying the nucleic acid fragment of the invention. Such novel vectors are also part of the present invention. The vector may comprise a plasmid, phage, cosmid, minichromosome, or virus, as well as naked DNA that is transiently expressed only in a particular cell. The cloning and expression vectors of the invention are capable of autonomous replication and thus provide high copy numbers for high level expression or high level replication purposes for subsequent cloning. The expression vector may comprise a promoter for driving expression of the nucleic acid fragment of the invention, optionally a nucleic acid sequence encoding a signal peptide for secretion or integration of the peptide expression product into a membrane, a nucleic acid fragment of the invention, and optionally a nucleic acid sequence encoding a terminator. When the expression vector is manipulated in a production strain or cell line, the vector, when introduced into a host cell, may or may not be integrated into the genome of the host cell. Vectors typically carry a replication site, as well as a marker sequence capable of providing phenotypic selection in transformed cells.

The expression vectors of the invention are useful for transforming host cells. Such transformed cells are also part of the invention and may be cultured cells or cell lines for propagation of the nucleic acid fragments and vectors of the invention, or for recombinant production of the polypeptides of the invention. The transformed cells of the present invention include microorganisms such as bacteria (e.g., Escherichia coli, Bacillus spp., etc.). Host cells also include cells from multicellular organisms such as fungi, insect cells, plant cells, or mammalian cells.

In some preferred embodiments, the product is a vaccine or a medicament.

In some embodiments, the vaccine further comprises an adjuvant and the medicament further comprises a pharmaceutically acceptable carrier.

In some preferred embodiments, the epitope polypeptide CD44-P3 prevents and/or treats prostate cancer by killing LNCAP-multicellular spheroids; and/or the presence of a gas in the gas,

To facilitate a clearer understanding of the contents of the present invention, reference will now be made in detail to the following specific embodiments.

Unless otherwise specified, the sources of experimental animals, drugs and reagents used in the examples of the present invention are normal and readily available:

lncap cells and VCaP cells were purchased from American Type Culture Collection (ATCC);

b. erythrocyte lysate, IFN-gamma, IL1 alpha, anti-CD 3 monoclonal antibody, anti-CD 28 monoclonal antibody, IL-2, AIM-V culture solution, IL4, TNF beta and GM-CSF factor are all purchased from Beijing Hokkiso Hi Biotech Co., Ltd;

c. the flow cytometer used LSRFortasa Cell Analyzer available from BD bioscience;

d. cell culture dishes and flasks were purchased from Corning, usa.

Example 1 design and Synthesis of short peptide of CD44 epitope

1. The complete amino acid sequence (NP-000601.3) of human CD44 protein is searched from the international open shared gene bank NCBI Genbank for 742 amino acids. The amino acid sequence is as follows:

MDKFWWHAAWGLCLVPLSLAQIDLNITCRFAGVFHVEKNGRYSISRTEAADLCKAFNSTLPTMAQMEKALSIGFETCRYGFIEGHVVIPRIHPNSICAANNTGVYILTSNTSQYDTYCFNASAPPEEDCTSVTDLPNAFDGPITITIVNRDGTRYVQKGEYRTNPEDIYPSNPTDDDVSSGSSSERSSTSGGYIFYTFSTVHPIPDEDSPWITDSTDRIPATTLMSTSATATETATKRQETWDWFSWLFLPSESKNHLHTTTQMAGTSSNTISAGWEPNEENEDERDRHLSFSGSGIDDDEDFISSTISTTPRAFDHTKQNQDWTQWNPSHSNPEVLLQTTTRMTDVDRNGTTAYEGNWNPEAHPPLIHHEHHEEEETPHSTSTIQATPSSTTEETATQKEQWFGNRWHEGYRQTPKEDSHSTTGTAAASAHTSHPMQGRTTPSPEDSSWTDFFNPISHPMGRGHQAGRRMDMDSSHSITLQPTANPNTGLVEDLDRTGPLSMTTQQSNSQSFSTSHEGLEEDKDHPTTSTLTSSNRNDVTGGRRDPNHSEGSTTLLEGYTSHYPHTKESRTFIPVTSAKTGSFGVTAVTVGDSNSNVNRSLSGDQDTFHPSGGSHTTHGSESDGHSHGSQEGGANTTSGPIRTPQIPEWLIILASLLALALILAVCIAVNSRRRCGQKKKLVINSGNGAVEDRKPSGLNGEASKSQEMVHLVNKESSETPDQFMTADETRNLQNVDMKIGV

2. this example predicts the location of the antigenic short peptide by DNA Star (Protean) software based on a comprehensive analysis of the hydrophilicity, antigenic index and surface accessibility of the amino acid sequence.

3. Epitope polypeptides comprising epitopes of CD44 protein were designed in conjunction with the Immune Epitope Database (IEDB, http:// www.iedb.org /) and were designated CD 44-P3.

As shown in figure 1, the molecular weight of the epitope polypeptide CD44-P3 is: 1084.18, the desolventizing temperature is: the flow rate of the desolventizing agent is 350L/h at 350 ℃.

As shown in FIG. 2, the HPLC analysis flow rate of the epitope polypeptide CD44-P3 is: 1.0ml/min, wavelength: 220nm, loading amount: 10 μ l, the specific results are shown in Table 1:

TABLE 1 high performance liquid chromatography analysis results of epitope polypeptide CD44-P3

Example 2

The epitope polypeptide CD44-P3 of the CD44 protein epitope provided by the embodiment 1 of the invention is adopted to research the killing effect of effector T cells induced by CD44 on human prostate cancer cell LNCAP-multicellular cells. The method specifically comprises the following steps:

1. separation and preparation of peripheral blood mononuclear cells: collecting 10mL of peripheral blood of the collected HLA-A2 positive healthy volunteers into 2 centrifugal tubes, centrifuging at 2000r/min for 5min, discarding the supernatant, mixing the precipitated cells, adding physiological saline to 25mL to fully suspend the precipitated cells to form a blood cell suspension. And adding 20mL of lymphocyte separation liquid into 1 centrifugal tube, and slowly transferring the blood cell suspension to the surface of the lymphocyte separation liquid by using a dropper to form a clear interface between the two.

2. Centrifuging the centrifuge tube with clear interface at 2000r/min for 20min, and separating into 4 layers from tube bottom to liquid surface, which are red blood cell and granulocyte layer, lymphocyte separation liquid layer, and peripheral blood mononuclear cell layer (PBMC layer and plasma layer); the peripheral blood mononuclear cell layer was aspirated by a pipette and transferred to another centrifuge tube for use.

3. Adding physiological saline into a centrifuge tube filled with a peripheral blood mononuclear cell layer to 40mL, centrifuging at 1500r/min for 5min, discarding supernatant after finishing, adding physiological saline to 40mL to suspend precipitated cells, fully and uniformly mixing, and centrifuging at 1500r/min for 5 min; the total centrifugation was performed 3 times.

4. After the final centrifugation, the supernatant was discarded, the pelleted cells were transferred to a culture flask and cultured with 40mL of AIM-V medium and labeled A.

5. After 2h of culture, the cultured cells were removed and the lymphocytes suspended in A were transferred to another 1 flask, labeled B.

Adding IFN-gamma, IL1 alpha and anti-CD 3 and anti-CD 28 monoclonal antibodies into a B bottle containing lymphocytes, adding IL-2 on the next day, and inducing and culturing CIK cells; to A flask containing adherent cells, 40mL of AIM-V culture medium and IL4 were added, and TNF β was added on the third day, and GM-CSF factor was added on the fifth day, to induce culture of DC cells.

And 6, in the process of DC cell culture, if yellowing occurs, replacing the solution, and supplementing IL-4, TNF beta and GM-CSF factors.

And 7, in the CIK cell culture process, if the culture solution turns yellow, replacing the culture solution, and supplementing IL-2 after replacing the culture solution. And if the cell mass is found to be in poor floccule shape, collecting the cell mass by using a centrifugal tube, standing and settling for about 5min, and carefully pouring the supernatant cell suspension into the culture bottle after the floccule is precipitated.

And 8, adding the CD44 epitope polypeptide CD44-P3 into the culture flask A when the DC cells are cultured to the 7 th day, shoveling the DC cells at the 8 th day, collecting the DC cells, mixing the collected DC cells with the corresponding CIK cells in the B cells for co-culture, and discarding the culture flask of the DC cells.

If the total cell amount is too large, the DC cells and CIK cells may be mixed and divided into the A and B flasks to continue the culture. T cells are induced into cytotoxic T Cells (CTLs), i.e., effector T cells.

9. Co-cultured DC-CIK cells were subjected to cell killing experiments at day 14.

10. Preparation of target cells: recovering normal culture and passage of prostate cancer cells LNCAP. Washing and resuspending the mixture by using a phosphate buffer solution after the digestion by pancreatin; labeling target cells with 10mM Calcein-AM, washing, resuspending, and co-culturing at 37 deg.C for 4h according to the effective target ratio of 5:1 and 10:1 between effector cells and tumor stem cells; the culture medium was collected, centrifuged at L000rpm for 5min, and 100. mu.L of the supernatant was taken to measure the average fluorescence intensity, with the target cells alone as the spontaneous release amount and the target cells alone plus detergent as the maximum release amount.

As shown in fig. 3A and fig. 3B, the result of analyzing the killing effect of the prostate cancer stem cell by using the CD44 epitope polypeptide shows that the CIK cell activated by the CD44 epitope polypeptide has a significant killing effect on the prostate cancer stem cell. The CD44 epitope polypeptide CD44-P3 can specifically induce the effect of cell killing, and under the condition that the effective-target ratio is 5:1, the average killing effect is improved to 26.9% from 4.7% of the background; at an effective target ratio of 10:1, the average killing effect increased from 6.2% to 22.7% of background, where killing is expressed as a kill (%).

Example 3

The epitope polypeptide CD44-P3 of the CD44 protein epitope provided by the embodiment 1 of the invention is adopted to research the killing effect of CD44 induced effector T cells on human prostate cancer cells VCaP-multicellular balls.

1. The method for separating peripheral blood mononuclear cells was the same as in example 2.

The DC-CIK cells were cultured in the same manner as in example 2.

3. The preparation of target cells VCaP-multicellular spherocytes and the detection of the killing effect were carried out in the same manner as in example 2.

As shown in fig. 4A and 4B, CIK cells induced by CD44-P3 epitope peptide have significant killing effect on human prostate cancer cell stem cell-enriched VCaP-tumor multicellular spheroids, and the average killing effect is improved from 9.8% of background to 27.2% under the condition that the effective-target ratio is 5: 1; at an effective target ratio of 10:1, the average killing effect increased from 4.4% to 31.5% of background, where killing is expressed as a kill (%).

Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and not to limit the same; while the invention has been described in detail and with reference to the foregoing embodiments, it will be understood by those skilled in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.

SEQUENCE LISTING

<110> Shenzhen Shenzhong Longhua region people hospital

<120> epitope polypeptide CD44-P3 based on prostate cancer stem cell marker CD44 and application thereof

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<170>PatentIn version 3.5

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Asp Gln Phe Met Thr Ala Asp Glu Thr Arg Asn

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gaccagttta tgacagctga tgagacaagg aac 33

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Met Asp Lys Phe Trp Trp His Ala Ala Trp Gly Leu Cys Leu Val Pro

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Val Phe His Val Glu Lys Asn Gly Arg Tyr Ser Ile Ser Arg Thr Glu

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Phe Ile Glu Gly His Val Val Ile Pro Arg Ile His Pro Asn Ser Ile

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Gln Tyr Asp Thr Tyr Cys Phe Asn Ala Ser Ala Pro Pro Glu Glu Asp

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Cys Thr Ser Val Thr Asp Leu Pro Asn Ala Phe Asp Gly Pro Ile Thr

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Asp Val Ser Ser Gly Ser Ser Ser Glu Arg Ser Ser Thr Ser Gly Gly

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Met Ser Thr Ser Ala Thr Ala Thr Glu Thr Ala Thr Lys Arg Gln Glu

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Thr Trp Asp Trp Phe Ser Trp LeuPhe Leu Pro Ser Glu Ser Lys Asn

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His Leu His Thr Thr Thr Gln Met Ala Gly Thr Ser Ser Asn Thr Ile

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Ser Ala Gly Trp Glu Pro Asn Glu Glu Asn Glu Asp Glu Arg Asp Arg

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Thr Ala Tyr Glu Gly Asn Trp Asn Pro Glu Ala His Pro Pro Leu Ile

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His His Glu His His Glu Glu Glu Glu Thr Pro His Ser Thr Ser Thr

370 375 380

Ile Gln Ala Thr Pro Ser Ser Thr Thr Glu Glu Thr Ala Thr Gln Lys

385 390 395 400

Glu Gln Trp Phe Gly Asn Arg Trp His GluGly Tyr Arg Gln Thr Pro

405 410 415

Lys Glu Asp Ser His Ser Thr Thr Gly Thr Ala Ala Ala Ser Ala His

420 425 430

Thr Ser His Pro Met Gln Gly Arg Thr Thr Pro Ser Pro Glu Asp Ser

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Ser Trp Thr Asp Phe Phe Asn Pro Ile Ser His Pro Met Gly Arg Gly

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His Gln Ala Gly Arg Arg Met Asp Met Asp Ser Ser His Ser Ile Thr

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Arg Thr Gly Pro Leu Ser Met Thr Thr Gln Gln Ser Asn Ser Gln Ser

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Phe Ser Thr Ser His Glu Gly Leu Glu Glu Asp Lys Asp His Pro Thr

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Thr Ser Thr Leu Thr Ser Ser Asn Arg Asn Asp Val Thr Gly Gly Arg

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Arg Asp Pro Asn His Ser Glu Gly Ser Thr Thr Leu Leu Glu Gly Tyr

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Thr Ser His Tyr Pro His Thr Lys Glu Ser Arg ThrPhe Ile Pro Val

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Thr Ser Ala Lys Thr Gly Ser Phe Gly Val Thr Ala Val Thr Val Gly

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Asp Ser Asn Ser Asn Val Asn Arg Ser Leu Ser Gly Asp Gln Asp Thr

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Phe His Pro Ser Gly Gly Ser His Thr Thr His Gly Ser Glu Ser Asp

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Gly His Ser His Gly Ser Gln Glu Gly Gly Ala Asn Thr Thr Ser Gly

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Pro Ile Arg Thr Pro Gln Ile Pro Glu Trp Leu Ile Ile Leu Ala Ser

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Arg Arg Arg Cys Gly Gln Lys Lys Lys Leu Val Ile Asn Ser Gly Asn

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Gly Ala Val Glu Asp Arg Lys Pro Ser Gly Leu Asn Gly Glu Ala Ser

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Lys Ser Gln Glu Met Val His Leu Val Asn Lys Glu Ser Ser Glu Thr

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725 730 735

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740

Claims

1. The application of the epitope polypeptide CD44-P3 in the preparation of products for preventing and/or treating prostatic cancer is characterized in that the amino acid sequence of the epitope polypeptide CD44-P3 is Asp-Gln-Phe-Met-Thr-Ala-Asp-Glu-Thr-Arg-Asn; the molecular weight is 1084.18.

2. The use according to claim 1, wherein the epitope polypeptide CD44-P3 is obtained by expression and purification in prokaryotic cells or eukaryotic cells; or the like, or, alternatively,

the epitope polypeptide CD44-P3 is synthesized artificially.

3. Use according to claim 1, wherein the product is a vaccine or a medicament.

4. The use according to claim 1, wherein the epitope polypeptide CD44-P3 prevents and/or treats prostate cancer by inducing T cell killing of LNCAP-multicellular spheroids; and/or the presence of a gas in the gas,

the epitope polypeptide CD44-P3 can prevent and/or treat prostatic cancer by inducing T cells to kill VCaP-multicellular spheres.

5. Use of a nucleic acid encoding the epitope polypeptide CD44-P3 of claim 1 for the preparation of a product for the prevention and/or treatment of prostate cancer.

6. Use of a vector comprising a nucleic acid according to claim 5 for the preparation of a product for the prevention and/or treatment of prostate cancer.

7. Use of a host cell comprising a nucleic acid according to claim 5 or a vector according to claim 6 for the preparation of a product for the prevention and/or treatment of prostate cancer.