FOXM1-derived peptides and vaccines containing the same

Abstract

The present invention provides epitope peptides derived from FOXM1 having the ability to induce cytotoxic T cells. The present invention also provides polynucleotides encoding the peptides, antigen presenting cells presenting the peptides, and cytotoxic T cells targeting the peptides, as well as methods for inducing antigen presenting cells or CTLs. The present invention also provides compositions and pharmaceutical compositions containing them as active ingredients. In addition, the present invention provides methods for treating and / or preventing cancer, and / or preventing its postoperative recurrence using the peptides, polynucleotides, antigen presenting cells, cytotoxic T cells or pharmaceutical compositions of the present invention. A method for inducing an immune response against cancer is also provided.

Description

[0001] This application is a divisional application of the Chinese invention patent application with application date of October 6, 2016, application number 201680057758.6, and invention name “FOXM1-derived peptides and vaccines containing the same”. Technical Field

[0002] The invention relates to the field of biological sciences, and more specifically to the field of cancer treatment. Specifically, the invention relates to novel peptides effective as cancer vaccines, methods for treating and / or preventing tumors using the peptides, and pharmaceutical compositions containing the peptides. Background Art

[0003] It is known that CD8-positive cytotoxic T cells (CTLs) recognize epitope peptides derived from tumor-associated antigens (TAAs) presented on major histocompatibility complex (MHC) class I molecules expressed on the cell surface, and then kill tumor cells. Since the discovery of the melanoma antigen (MAGE) family, many TAAs have been found by immunological methods (NPL1: Boon T, Int J Cancer 1993, 54 (2): 177-80; NPL2: Boon T & van der Bruggen P, J Exp Med 1996, 183 (3): 725-9). Some of these TAAs are currently being clinically developed as immunotherapy targets.

[0004] In several of these TAAs, epitope peptides that can be recognized by CTLs have been identified, and their application in immunotherapy for various types of cancer is expected (NPL3: Harris CC, J Natl Cancer Inst 1996, 88(20):1442-55; NPL4: Butterfield LH et al., Cancer Res 1999, 59(13):3134-42; NPL5: Vissers JL et al., Cancer Res 1999, 59(21):5554-9; NPL6: van der Burg SH et al., J Immunol 1996, 156(9):3308-14; NPL7: Tanaka F et al., Cancer Res 1997, 57(20):4465-8; NPL8: Fujie T et al., Int J Cancer 1999, 80 (2): 169-72; NPL9: Kikuchi M et al., Int J Cancer 1999, 81 (3): 459-66; NPL10: Oiso M et al., Int J Cancer 1999, 81 (3): 387-94). So far, several clinical trials using these TAA-derived epitope peptides have been reported. Unfortunately, however, the response rates in many clinical trials were not high (NPL11: Belli F et al., J Clin Oncol 2002, 20 (20): 4169-80; NPL12: Coulie PG et al., Immunol Rev 2002, 188: 33-42; NPL13: Rosenberg SA et al., Nat Med 2004, 10 (9): 909-15). Therefore, there is still a need to identify new CTL epitope peptides that can be used for cancer immunotherapy.

[0005] Results from genome-wide expression profiling using cDNA microarrays targeting 27,648 genes identified and reported FOXM1 (GenBank accession number: NM_202003; Forkhead box M1) as a gene upregulated in intrahepatic cholangiocarcinoma, non-small cell lung cancer, and esophageal cancer tissues (NPL14: Obama K et al., Hepatology 2005, 41(6):1339-48; NPL15: Yokomine K et al., Int J Cancer 2010, 126(9):2153-63; PTL1: WO2005 / 090603). On the other hand, compared with these cancer tissues, the expression level of FOXM1 in normal tissues is very low. In addition, when FOXM1 expression is inhibited, the proliferation of medulloblastoma cells is inhibited (NPL16: Priller M et al., Clin Cancer Res 2011, 17(21): 6791-801), and FOXM1 is considered to be a gene involved in regulating cancer cell proliferation. More specifically, FOXM1 is a TAA that repeatedly proliferates in many cancers, and therefore epitope peptides derived from FOXM1 are considered to be suitable for immunotherapy targeting cancer patients.

[0006] Recently, FOXM1-derived HLA-A02-restricted epitope peptides (PTL2: WO2009 / 025196) and HLA-A24-restricted epitope peptides (PTL3: WO2010 / 095428) have been identified. The therapeutic effects of these peptides in cancer patients with HLA-A02 or HLA-A24 types can be expected, but not for other cancer patients.

[0007] [Citation List]

[0008] [Patent Document]

[0009] [PTL 1] WO2005 / 090603

[0010] [PTL 2] WO2009 / 025196

[0011] [PTL 3] WO2010 / 095428

[0012] [Non Patent Literature]

[0013] [NPL 1]Boon T,Int J Cancer 1993,54(2):177-80

[0014] [NPL 2]Boon T&van der Bruggen P,J Exp Med 1996,183(3):725-9

[0015] [NPL 3]Harris CC,J Natl Cancer Inst 1996,88(20):1442-55

[0016] [NPL 4]Butterfield LH et al.,Cancer Res 1999,59(13):3134-42

[0017] [NPL 5]Vissers JL et al.,Cancer Res 1999,59(21):5554-9

[0018] [NPL 6]van der Burg SH et al.,J Immunol 1996,156(9):3308-14

[0019] [NPL 7]Tanaka F et al.,Cancer Res 1997,57(20):4465-8

[0020] [NPL 8]Fujie T et al.,Int J Cancer 1999,80(2):169-72

[0021] [NPL 9]Kikuchi M et al.,Int J Cancer 1999,81(3):459-66

[0022] [NPL 10]Oiso M et al.,Int J Cancer 1999,81(3):387-94

[0023] [NPL 11]Belli F et al.,J Clin Oncol 2002,20(20):4169-80

[0024] [NPL 12]Coulie PG et al.,Immunol Rev 2002,188:33-42

[0025] [NPL 13]Rosenberg SA et al.,Nat Med 2004,10(9):909-15

[0026] [NPL 14]Obama K et al.,Hepatology 2005,41(6):1339-48

[0027] [NPL 15]Yokomine K et al.,Int J Cancer 2010,126(9):2153-63

[0028] [NPL 16]Priller M et al.,Clin Cancer Res 2011,17(21):6791-801 Summary of the invention

[0029] The present invention relates to peptides that can induce CTLs that specifically react with cells expressing FOXM1. When these peptides form a complex with human leukocyte antigen (HLA) and are presented to CD8-positive T cells by antigen presenting cells (APCs) (which present the complex on their surface), CTLs that show peptide-specific cytotoxic activity are induced. The FOXM1-derived peptides with CTL induction ability (CTL induction power) identified so far are HLA-A02 restricted peptides and HLA-A24 restricted peptides, and CTLs cannot be induced when antigen presenting cells do not express these HLAs. Therefore, conventional peptides are not suitable for immunotherapy in subjects who do not have these HLAs. HLA-A33 is a common allele in Asians, and HLA-A01 is a common allele in Caucasians (Cao K, et al., Hum Immunol 2001, 62 (9): 1009-30). It is desirable to administer HLA-A33-restricted peptides to HLA-A33-positive subjects, and to administer HLA-A01-restricted peptides to HLA-A01-positive subjects. Therefore, the present invention relates to FOXM1-derived peptides having CTL inducibility restricted to HLA-A33 or HLA-A01. Based on the results disclosed herein, it has been demonstrated that the peptides of the present invention are epitope peptides capable of inducing a strong and specific immune response against cells expressing FOXM1 and HLA-A33 or HLA-A01.

[0030] Therefore, one of the objects of the present invention is to provide FOXM1-derived peptides capable of inducing CTLs in an HLA-A33- or HLA-A01-restricted manner. These peptides can be used to induce CTLs in vitro, ex vivo or in vivo, or can be used for administration to a subject for the purpose of inducing an immune response against cancer cells expressing FOXM1. Preferred peptides are peptides comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60, and 61; more preferred peptides are nonapeptides or decapeptides; and even more preferred peptides are peptides selected from the group consisting of the amino acid sequences of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60, and 61.

[0031] The peptides of the present invention include peptides in which one, two or more amino acids are substituted, deleted, inserted and / or added, as long as the resulting modified peptide retains the CTL inducibility of the original peptide.

[0032] The present invention also provides isolated polynucleotides encoding any of the peptides of the present invention. Similar to the peptides of the present invention, these polynucleotides can be used to induce APCs with CTL inducibility and can be administered to a subject to induce an immune response against cancer cells expressing FOXM1.

[0033] The present invention also provides a composition comprising one or more types of peptides of the present invention, one or more types of polynucleotides encoding one or more types of peptides of the present invention, APCs of the present invention, exosomes presenting the peptides of the present invention, and / or CTLs of the present invention. The composition of the present invention is preferably a pharmaceutical composition. The pharmaceutical composition of the present invention can be used to treat and / or prevent cancer, and to prevent its postoperative recurrence. They can also be used to induce an immune response against cancer. When administered to a subject, the peptides of the present invention are presented on the surface of the APCs, and as a result, CTLs targeting the peptides are induced. Therefore, another object of the present invention is to provide a composition for inducing CTLs, wherein the composition comprises one or more types of peptides of the present invention, one or more types of polynucleotides encoding one or more types of peptides of the present invention, APCs of the present invention, and / or exosomes presenting the peptides of the present invention.

[0034] Another object of the present invention is to provide a method for inducing APCs having CTL inducibility, wherein the method comprises the step of contacting one or more types of peptides of the present invention with APCs, or the step of introducing a polynucleotide encoding any one of the peptides of the present invention into APCs.

[0035] The present invention also provides a method for inducing CTL, comprising the steps of co-culturing CD8-positive T cells with APCs presenting a complex of an HLA antigen and the peptide of the present invention on its surface, co-culturing CD8-positive T cells with exosomes presenting a complex of an HLA antigen and the peptide of the present invention on its surface, or introducing a vector into CD8-positive T cells, the vector comprising a polynucleotide encoding each subunit of a T cell receptor (TCR), the TCR being capable of binding to the peptide of the present invention presented by the HLA antigen on the cell surface. The preferred HLA antigen in the present invention is HLA-A33 or HLA-A01.

[0036] Another object of the present invention is to provide an isolated APC presenting a complex of an HLA antigen and a peptide of the present invention on its surface. The present invention also provides an isolated CTL targeting the peptide of the present invention. These APCs and CTLs can be used for immunotherapy of cancers expressing FOXM1. In the present invention, the cancer to be immunotherapy is, for example, a cancer present in a homozygous or heterozygous patient with HLA-A33 or HLA-A01. Therefore, the APC or CTL is also a cell homozygous or heterozygous with HLA-A33 or HLA-A01. That is, the present invention provides an immunotherapy for cancers expressing FOXM1 and at least one HLA antigen selected from HLA-A33 and HLA-A01.

[0037] Another object of the present invention is to provide a method for inducing an immune response against cancer in a subject, wherein the method comprises the step of administering to the subject a composition comprising a peptide of the present invention or a polynucleotide encoding the peptide, an APC of the present invention, an exosome presenting the peptide of the present invention, and / or a CTL of the present invention. Another object of the present invention is to provide a method for treating and / or preventing cancer and preventing its postoperative recurrence in a subject, wherein the method comprises the step of administering to the subject a peptide of the present invention, a polynucleotide encoding the peptide, an APC of the present invention, an exosome presenting the peptide of the present invention, and / or a CTL of the present invention.

[0038] In addition to the above, other objects and features of the present invention will become more apparent when the following detailed description is read in conjunction with the accompanying drawings and examples. However, it should be understood that the foregoing overview of the present invention and the following detailed description are exemplary embodiments, and are not restrictive to the present invention or other alternative embodiments of the present invention. In particular, although the present invention is described herein with reference to a plurality of specific embodiments, it should be understood that the description is an illustration of the present invention and does not constitute a limitation of the present invention. Various modifications and applications may be conceived by those skilled in the art without departing from the spirit and scope of the present invention as described in the appended claims. Similarly, other objects, features, benefits and advantages of the present invention will become apparent from this overview and certain embodiments described below, and will be apparent to those skilled in the art. Such objects, features, benefits and advantages will be apparent from the above in conjunction with the attached examples, data, drawings and all reasonable inferences drawn therefrom, considered alone or together with the references incorporated herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1-1 Composed of photos (a) to (v), it shows the results of interferon (IFN)-gamma enzyme-linked immunosorbent assay (ELISPOT) using cells induced by peptides derived from FOXM1. In these figures, "+" shows IFN-gamma production for target cells impacted with the peptide of interest; and "-" shows IFN-gamma production for target cells not impacted with any peptide (negative control). By comparison with the negative control, it can be seen that peptide-specific IFN-gamma production was observed in the following wells:

[0040] Well #6(a) using FOXM1-A33-9-180 (SEQ ID NO: 1),

[0041] Well #3(b) using FOXM1-A33-9-308 (SEQ ID NO: 2),

[0042] Well #4(c) using FOXM1-A33-9-693 (SEQ ID NO: 3),

[0043] Well #3(d) using FOXM1-A33-9-516 (SEQ ID NO:6),

[0044] Well #5(e) using FOXM1-A33-9-146 (SEQ ID NO:7),

[0045] Well #6(f) using FOXM1-A33-9-289 (SEQ ID NO: 11),

[0046] Well #6(g) using FOXM1-A33-9-228 (SEQ ID NO: 12),

[0047] Well #4 (h) using FOXM1-A33-9-502 (SEQ ID NO: 17),

[0048] Well #2(i) using FOXM1-A33-9-321 (SEQ ID NO: 18),

[0049] Well #6(j) using FOXM1-A33-9-341 (SEQ ID NO:20),

[0050] Well #8(k) using FOXM1-A33-10-514 (SEQ ID NO:22),

[0051] Well #6(l) using FOXM1-A33-10-179 (SEQ ID NO:24),

[0052] Well #5(m) using FOXM1-A33-10-501 (SEQ ID NO:26),

[0053] Well #5(n) using FOXM1-A33-10-124 (SEQ ID NO:32),

[0054] Well #3(o) using FOXM1-A33-10-595 (SEQ ID NO:33),

[0055] Well #5(p) using FOXM1-A33-10-546 (SEQ ID NO:36),

[0056] Well #6(q) using FOXM1-A33-10-391 (SEQ ID NO:39),

[0057] Well #3 (r) using FOXM1-A33-10-607 (SEQ ID NO: 41),

[0058] Well #2(s) using FOXM1-A33-10-265 (SEQ ID NO:42),

[0059] Well #6(t) using FOXM1-A33-10-4 (SEQ ID NO:45), and

[0060] Well #5 (u) using FOXM1-A33-10-388 (SEQ ID NO: 46). The cells circled in the photo that showed a reaction proliferated to establish a CTL cell line. Meanwhile, FOXM1-A33-10-288 (SEQ ID NO: 25) (v) is shown as an example of typical negative data in which peptide-specific IFN-gamma production was not observed.

[0061] Figure 1-2 Continue to show Figure 1-1 .

[0062] Figure 2 Consists of line graphs (a) to (d), which show the measurement results of IFN-gamma produced by CTL lines stimulated with FOXM1-A33-9-308 (SEQ ID NO: 2) (a), FOXM1-A33-9-146 (SEQ ID NO: 7) (b), FOXM1-A33-10-391 (SEQ ID NO: 39) (c) or FOXM1-A33-10-265 (SEQ ID NO: 42) (d) using IFN-gamma enzyme-linked immunosorbent assay (ELISA). These results show that after induction with each peptide, CTL cell lines that produce IFN-gamma in a peptide-specific manner were established. In the figure, "+" shows IFN-gamma production by CTL lines against target cells pulsed with the peptide of interest; and "-" shows IFN-gamma production by CTL lines against target cells not pulsed with any peptide. The R / S ratio represents the ratio of the cell number of the CTL line (responder cells) to the cell number of the target cells that stimulated them (stimulator cells).

[0063] Figure 3 Consists of a series of line graphs (a) to (b), which show IFN-gamma production in CTL clones established by the limiting dilution method after induction with FOXM1-A33-9-308 (SEQ ID NO: 2) (a) or FOXM1-A33-9-146 (SEQ ID NO: 7) (b). These results show peptide-specific IFN-gamma production by CTL clones. In the figure, "+" shows IFN-gamma production by CTL clones against target cells pulsed with the peptide of interest; and "-" shows IFN-gamma production by CTL clones against target cells not pulsed with any peptide. The R / S ratio represents the ratio of the number of cells of the CTL clones (responding cells) to the number of cells of the target cells (stimulating cells) that stimulate them.

[0064] Figure 4The line graph shows the IFN-gamma production of CTL clones against target cells expressing both FOXM1 and HLA-A*33:03. Target cells into which HLA-A*33:03 or full-length FOXM1 gene was introduced were used as negative controls. CTL clones established using FOXM1-A33-9-308 (SEQ ID NO:2) induced IFN-gamma production (black diamonds) against COS7 into which both FOXM1 and HLA-A*33:03 were introduced. On the other hand, no significant IFN-gamma production was shown for COS7 cells into which either HLA-A*33:03 (white triangles) or FOXM1 (white circles) was introduced.

[0065] Figure 5-1 Composed of photos (a) to (n), it shows the results of IFN-gamma enzyme-linked immunosorbent assay (ELISPOT) using cells induced with peptides derived from FOXM1. In the figure, "+" shows IFN-gamma production against target cells punched with the peptide of interest; and "-" shows IFN-gamma production against target cells punched with no peptide (negative control).

[0066] As can be seen by comparison with the negative control, peptide-specific IFN-gamma production was observed in the following wells:

[0067] In well #3 (a) using FOXM1-A01-9-233 (SEQ ID NO: 48),

[0068] In well #3 (b) using FOXM1-A01-9-539 (SEQ ID NO: 49),

[0069] In well #3 (c) using FOXM1-A01-9-631 (SEQ ID NO: 50),

[0070] In well #2 (d) using FOXM1-A01-9-231 (SEQ ID NO: 52),

[0071] In well #2 (e) using FOXM1-A01-9-663 (SEQ ID NO: 53),

[0072] In well #5 (f) using FOXM1-A01-9-494 (SEQ ID NO: 55),

[0073] In well #2 (g) using FOXM1-A01-9-341 (SEQ ID NO: 20),

[0074] In well #1 using FOXM1-A01-10-566 (SEQ ID NO: 56) (h),

[0075] In well #2 (i) using FOXM1-A01-10-263 (SEQ ID NO: 57),

[0076] In well #4 (j) using FOXM1-A01-10-308 (SEQ ID NO: 58),

[0077] In well #6 (k) using FOXM1-A01-10-232 (SEQ ID NO: 59),

[0078] Well #6 (1) and

[0079] Well #6 (m) using FOXM1-A01-10-341 (SEQ ID NO: 61).

[0080] Cells that responded to the display of the frame in the photograph proliferated to establish CTL lines. Meanwhile, FOXM1-A01-10-265 (SEQ ID NO: 42) (n) is shown as an example of typical negative data in which peptide-specific IFN-gamma production was not observed.

[0081] Figure 5-2 Continue to show Figure 5-1 .

[0082] Figure 6 The line graph shows the results of IFN-gamma produced by CTL lines stimulated with FOXM1-A01-10-566 (SEQ ID NO: 56) by ELISA. The results show that after induction with peptides, CTL lines that produce IFN-gamma in a peptide-specific manner were established. In the figure, "+" shows IFN-gamma production of CTL lines against target cells impacted with the peptide of interest; and "-" shows IFN-gamma production of CTL lines against target cells not impacted with any peptide. The R / S ratio represents the ratio of the number of cells of the CTL line (responding cells) to the number of cells of the target cells (stimulating cells) that stimulate them.

[0083] Figure 7It consists of a series of line graphs (a) to (b), which show the IFN-gamma production in CTL clones established by the limiting dilution method after induction with FOXM1-A01-9-233 (SEQ ID NO: 48) or FOXM1-A01-10-566 (SEQ ID NO: 56). These results show the peptide-specific IFN-gamma production of CTL clones. In the figure, "+" shows the IFN-gamma production of CTL lines against target cells pulsed with the peptide of interest; and "-" shows the IFN-gamma production of CTL lines against target cells not pulsed with any peptide. The R / S ratio represents the ratio of the cell number of CTL clones (responding cells) to the cell number of target cells (stimulating cells) that stimulate them.

[0084] Figure 8 The line graph shows the IFN-gamma production of CTL clones against target cells expressing both FOXM1 and HLA-A*01:01. Target cells into which HLA-A*01:01 or full-length FOXM1 gene was introduced were used as negative controls. CTL clones established using FOXM1-A01-10-566 (SEQ ID NO:56) induced IFN-gamma production (black diamonds) against COS7 into which both FOXM1 and HLA-A*01:01 were introduced. On the other hand, no significant IFN-gamma production was shown for COS7 cells into which either HLA-A*01:01 (white triangles) or FOXM1 (white circles) was introduced. DETAILED DESCRIPTION

[0085] Description of Specific Embodiments

[0086] Although any methods and materials similar or equivalent to the methods and materials described herein can be used in implementing or testing embodiments of the present invention, preferred methods, devices and materials are now described. However, before describing the materials and methods of the present invention, it should be understood that the present invention is not limited to the specific sizes, shapes, scales, materials, methodology, schemes, etc. described herein, because these can be changed according to routine experiments and optimization. It should also be understood that the terms used in this specification are only used for the purpose of describing specific versions or embodiments, and are not intended to limit the scope of the present invention, which will only be limited by the appended claims.

[0087] I. Definitions

[0088] As used herein, the terms "a", "an", "the", and "said" mean "at least one", unless expressly stated otherwise.

[0089] The terms "isolated" and "purified", when used in connection with a substance (e.g., a peptide, an antibody, a polynucleotide, etc.), refer to a substance that is substantially free of at least one substance that may be included in a natural source. Thus, an isolated or purified peptide refers to a peptide that is substantially free of another cellular material, such as carbohydrates, lipids, or other contaminating proteins from the cell or tissue source of the peptide. When the peptide is chemically synthesized, an isolated or purified peptide refers to a peptide that is substantially free of a precursor substance or another chemical substance. The phrase "substantially free of cellular material" includes peptide preparations, in which the peptide is separated from the cellular components of the cell from which the peptide is isolated or recombinantly produced. Thus, peptides that are substantially free of cellular material include peptide preparations that contain less than about 30%, 20%, 10%, or 5%, 3%, 2% or 1% (by dry weight) of other cellular material.

[0090] When the peptide is recombinantly produced, the isolated or purified peptide is substantially free of culture medium, and the substantially culture medium-free peptide comprises a peptide preparation that contains less than about 20%, 10%, or 5%, or 3%, 2% or 1% (by dry weight) of culture medium by volume of the peptide preparation.

[0091] Alternatively, when the peptide is chemically synthesized, the isolated or purified peptide is substantially free of precursors or other chemicals, and the peptide substantially free of precursors or other chemicals contains a peptide preparation that contains less than about 30%, 20%, 10%, 5%, 3%, 2% or 1% (by dry weight) of the peptide preparation volume of precursors or other chemicals. The peptide preparation can be confirmed to be isolated or purified by, for example, the appearance of a single band after sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and Coomassie brilliant blue staining or such gels. In preferred embodiments, the peptides and polynucleotides of the present invention are isolated or purified.

[0092] The terms "polypeptide," "peptide," and "protein" are used interchangeably herein and refer to a polymer of amino acid residues. In addition to naturally occurring amino acid polymers, these terms also apply to non-naturally occurring amino acid polymers that contain one or more non-naturally occurring amino acid residues. Non-naturally occurring amino acids include amino acid analogs, amino acid mimetics, and the like.

[0093] The term "amino acid" as used herein refers to naturally occurring amino acids, as well as amino acid analogs and amino acid mimetics that perform similar functions to naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code in cells, as well as those that are post-translationally modified (e.g., hydroxyproline, gamma-carboxyglutamate, and O-phosphoserine, etc.). The phrase "amino acid analog" refers to a chemical compound having the same basic chemical structure (alpha carbon bound to hydrogen, carboxyl group, amino group, and R group) as a naturally occurring amino acid but having a modified R group or a modified backbone (e.g., homoserine, norleucine, methionine sulfoxide, methionine methylsulfonium, etc.). The phrase "amino acid mimetics" refers to a compound that has a different structure from general amino acids but has similar functions to amino acids. Amino acids can be L-amino acids or D-amino acids, and the peptides of the present invention are preferably L-amino acid polymers.

[0094] The terms "polynucleotide," "oligonucleotide," and "nucleic acid" are used interchangeably herein and refer to a polymer of nucleotides.

[0095] The term "composition" as used in this specification is intended to include products containing specified ingredients in specified amounts, as well as any products directly or indirectly produced by the combination of specified ingredients in specified amounts. When the composition is a pharmaceutical composition, the term "composition" is intended to include products of active ingredients and inert ingredients, as well as any products directly or indirectly produced from the combination, complexation or aggregation of any two or more ingredients, from the dissociation of one or more ingredients, or from other types of reactions or interactions of one or more ingredients. Therefore, the pharmaceutical composition of the present invention includes any composition prepared by mixing the compound or cell of the present invention with a pharmaceutically or physiologically acceptable carrier. Without being limited thereto, the term "pharmaceutically acceptable carrier" or "physiologically acceptable carrier" used in this specification includes liquid or solid fillers, diluents, excipients, solvents and encapsulating materials; and refers to pharmaceutically or physiologically acceptable materials, compositions, substances or media.

[0096] Unless otherwise indicated, the term "cancer" refers to cancers that overexpress the FOXM1 gene; and examples thereof include acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc., but are not limited thereto. In an exemplary embodiment, "cancer" is a cancer that expresses FOXM1 and HLA-A33 and / or HLA-A01.

[0097] Unless otherwise indicated, the terms "cytotoxic T lymphocytes" and "cytotoxic T cells" and "CTLs" are used interchangeably herein. Unless otherwise specifically indicated, they refer to a subset of T lymphocytes that can recognize non-self cells (e.g., tumor / cancer cells, virus-infected cells) and induce the death of such cells.

[0098] Unless otherwise indicated, the term "HLA-A33" refers to a subset of * 33:03,HLA-A * 33:01, and HLA-A * 33:04 subtype of HLA-A33.

[0099] Unless otherwise indicated, the term "HLA-A01" refers to a subset of * 01:01,HLA-A * 01:03, and HLA-A * 01:04 subtype of HLA-A01.

[0100] In the context of a subject or patient, as used herein, the phrase "the HLA antigen of the subject (or patient) is HLA-A33" means that the subject or patient has a homozygous or heterozygous HLA-A33 antigen gene as an MHC (major histocompatibility complex) class I molecule, and the HLA-A33 antigen is expressed as an HLA antigen in the cells of the subject or patient. Similarly, the phrase "the HLA antigen of the subject (or patient) is HLA-A01" used herein means that the subject or patient has a homozygous or heterozygous HLA-A01 antigen gene as an MHC (major histocompatibility complex) class I molecule, and the HLA-A01 antigen is expressed as an HLA antigen in the cells of the subject or patient.

[0101] As long as the methods and compositions of the present invention are useful in the context of "treatment" of cancer, the treatment is considered "effective" when it achieves a clinical advantage, for example, reducing the size, spread or metastatic ability of the cancer, delaying cancer progression, alleviating clinical symptoms of cancer, prolonging survival, inhibiting postoperative recurrence in the subject. When the treatment is administered prophylactically, "effective" means that the treatment delays or prevents the formation of cancer, or prevents or alleviates clinical symptoms of cancer. Effectiveness is determined relative to any known method for diagnosing or treating a particular tumor type.

[0102] As long as the methods and compositions of the present invention are used in the context of cancer "prevention", the term "prevention" herein includes any work that reduces the burden of mortality or morbidity associated with cancer. Prevention can be carried out at the "primary, secondary and tertiary prevention levels". Although primary prevention avoids the development of the disease, prevention at the secondary and tertiary levels includes preventing the progression of the disease and the appearance of symptoms, as well as aiming to reduce the adverse effects of existing diseases by restoring function and reducing disease-related complications. Alternatively, prevention can include reducing the severity of a particular condition, such as a broad range of preventive treatments aimed at reducing tumor growth and metastasis.

[0103] In the context of the present invention, the treatment and / or prevention (prevention) of cancer and / or the prevention (prevention) of its postoperative recurrence includes any event, such as inhibition of cancer cell proliferation, tumor regression or regression, induction of remission and inhibition of cancer development, tumor regression, and reduction or inhibition of metastasis, inhibition of postoperative recurrence of cancer and prolonged survival. Effective treatment and / or prevention (prevention) of cancer reduces mortality, improves the prognosis of individuals with cancer, reduces blood levels of tumor markers, and alleviates detectable symptoms associated with cancer. For example, alleviation or improvement of symptoms constitutes effective treatment and / or prevention (prevention), and includes patients in which symptoms are stabilized or alleviated by 10%, 20%, 30% or more.

[0104] In the context of the present invention, the term "antibody" refers to an immunoglobulin and a fragment thereof that specifically reacts with a specified protein or its peptide. Antibodies can include human antibodies, primatized antibodies, chimeric antibodies, bispecific antibodies, humanized antibodies, antibodies fused with other proteins or radioactive markers, and antibody fragments. In addition, "antibody" herein is used in the broadest sense, and specifically encompasses complete monoclonal antibodies, polyclonal antibodies, multispecific antibodies, multispecific antibodies (such as bispecific antibodies) formed by two or more complete antibodies, and antibody fragments, as long as they exhibit the desired biological activity. "Antibody" can be antibodies of all classes (such as IgA, IgD, IgE, IgG and IgM).

[0105] Unless otherwise defined, 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.

[0106] II. Peptides

[0107] HLA-A33 is a common allele in Asians, and HLA-A01 is a common allele in Caucasians (Cao et al., Hum Immunol 2001; 62(9): 1009-30). Therefore, for a large population of Asians or Caucasians, an effective method for treating cancers expressing FOXM1 can be provided by providing FOXM1-derived CTL-inducing peptides restricted to HLA-A33 or HLA-A01. Therefore, the present invention provides FOXM1-derived peptides that can induce CTLs in an HLA-A33- or HLA-A01-restricted manner.

[0108] The peptide of the present invention is a FOXM1-derived peptide that can induce CTL in an HLA-A33- or HLA-A01-restricted manner. Peptides that can induce CTL in an HLA-A33-restricted manner include peptides having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, and 46. Peptides that can induce CTL in an HLA-A01-restricted manner include peptides having an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60, and 61.

[0109] CTLs having cytotoxic activity specific to these peptides can be established by in vitro stimulation of T cells with dendritic cells (DCs) pulsed with these peptides. The established CTLs show specific cytotoxic activity against target cells pulsed with each peptide.

[0110] The FOXM1 gene is overexpressed in cancer cells, such as acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc., but is not expressed in most normal organs. Therefore, it is an excellent target for immunotherapy. Therefore, the peptide of the present invention can be appropriately used for cancer immunotherapy. Preferred peptides are nonapeptides (peptides consisting of 9 amino acid residues) or decapeptides (peptides consisting of 10 amino acid residues), and more preferably peptides selected from the group consisting of the following amino acid sequences: SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61. For example, a peptide having the amino acid sequence of SEQ ID NO: 2 is suitable for inducing CTLs that show specific cytotoxic activity against cells expressing HLA-A33 and FOXM1, and can be suitable for cancer immunotherapy of HLA-A33-positive patients. In a more preferred embodiment, the peptide of the present invention is a peptide consisting of the amino acid sequence of SEQ ID NO: 2. In addition, for example, a peptide having the amino acid sequence of SEQ ID NO: 56 is suitable for inducing CTLs that exhibit specific cytotoxic activity against cells expressing HLA-A01 and FOXM1, and can be used for cancer immunotherapy for HLA-A01-positive patients. In a more preferred embodiment, the peptide of the present invention is a peptide consisting of the amino acid sequence of SEQ ID NO: 56.

[0111] For the peptides of the present invention, additional amino acid residues can be prepared to add (adjoin) the amino acid sequence of the peptides of the present invention, as long as the resulting peptide retains the CTL inducibility of the original peptide. The additional amino acid residues can be composed of any type of amino acids, as long as they do not impair the CTL inducibility of the original peptide. Therefore, the peptides of the present invention include peptides with CTL inducibility, which include an amino acid sequence selected from the following: SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61. Such peptides are, for example, less than about 40 amino acids, in many cases less than about 20 amino acids, and usually less than about 15 amino acids. Thus, if the original peptide is a nonapeptide, the peptides of the present invention include peptides of 10 amino acids in length or 11-40 amino acids in length, which are produced by appending additional amino acids to the peptide. In addition, if the original peptide is a decapeptide, the peptides of the present invention include peptides of 11-40 amino acids in length. Such peptides may be, for example, peptides of 11-20 amino acids in length or peptides of 11-15 amino acids in length. Preferred examples of additional amino acid residues are amino acid residues (e.g., SEQ ID NO: 67, 69, 71, 73 or 75) adjacent to the amino acid sequence of the peptide of the present invention in the full-length amino acid sequence of FOXM1. Therefore, the peptides of the present invention include peptides containing an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61, and wherein the peptide is a peptide fragment of FOXM1 and has CTL inducibility.

[0112] Generally speaking, the modification of one, two or more amino acids in certain peptides will not affect the function of the peptide, or in some cases will even enhance the desired function of the original protein. In fact, it is known that modified peptides (i.e., peptides consisting of an amino acid sequence in which one, two or several amino acid residues are modified (i.e., substituted, added, deleted and / or inserted) compared to the original reference sequence) retain the biological activity of the original peptide (Mark et al., Proc Natl Acad Sci USA 1984, 81: 5662-6; Zoller and Smith, Nucleic Acids Res 1982, 10: 6487-500; Dalbadie-McFarland et al., Proc Natl Acad Sci USA 1982, 79: 6409-13). Therefore, in one embodiment, the peptide of the present invention may be a peptide comprising an amino acid sequence in which one, two or several amino acids are substituted, deleted, inserted and / or added to an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61, and has CTL inducibility.

[0113] Those skilled in the art will recognize that individual substitutions to an amino acid sequence (which change a single amino acid or a small percentage of amino acids) tend to result in the retention of the properties of the original amino acid side chains. This is often referred to as "conservative substitutions" or "conservative modifications"; and modification of a protein by "conservative substitutions" or "conservative modifications" can result in a modified protein having a function similar to the original protein. Conservative substitution tables that provide functionally similar amino acids are well known in the art. Examples of functionally similar amino acid side chain features include, for example, hydrophobic amino acids (A, I, L, M, F, P, W, Y, V), hydrophilic amino acids (R, D, N, C, E, Q, G, H, K, S, T), and side chains with the following common functional groups or features: aliphatic side chains (G, A, V, L, I, P); hydroxyl-containing side chains (S, T, Y); sulfur-containing side chains (C, M); carboxylic acid and amide-containing side chains (D, N, E, Q); basic side chains (R, K, H); and aromatic side chains (H, F, Y, W). In addition, the following eight groups each contain amino acids that are mutually conservatively substituted as recognized in the art:

[0114] 1) Alanine (A), Glycine (G);

[0115] 2) Aspartic acid (D), Glutamic acid (E);

[0116] 3) Asparagine (N), glutamine (Q);

[0117] 4) Arginine (R), Lysine (K);

[0118] 5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V);

[0119] 6) Phenylalanine (F), tyrosine (Y), tryptophan (W);

[0120] 7) serine (S), threonine (T); and

[0121] 8) Cysteine ​​(C), methionine (M) (see, for example, Creighton, Proteins 1984).

[0122] Such conservatively modified peptides are also included in the peptides of the present invention. However, the peptides of the present invention are not limited thereto and may include non-conservative modifications as long as the peptide retains the CTL inducibility of the original peptide. In addition, the modified peptides do not exclude CTL-inducible peptides derived from polymorphic variants, interspecies homologs, and alleles of FOXM1.

[0123] As long as the peptide retains the CTL inducibility of the original peptide, a small number (e.g., 1, 2 or several) or a small percentage of amino acids may be modified (i.e., substituted, deleted, inserted and / or added). Here, the term "several" means 5 or less amino acids, such as 4 or 3 or less. The percentage of modified amino acids is preferably 20% or less, more preferably 15% or less, further more preferably 10% or less, or 1 to 5%.

[0124] When used in the context of immunotherapy, the peptides of the present invention should be presented on the surface of cells or exosomes, preferably as a complex with HLA antigens. Therefore, it is preferred that the peptides of the present invention have a high binding affinity to HLA antigens. For this purpose, the peptides can be modified by substitution, deletion, insertion and / or addition of amino acid residues to produce modified peptides with improved binding affinity. Since the sequence regularity of the peptides displayed by binding to HLA antigens is known (Falk, et al., Immunogenetics 1994 40 232-41; Chujoh, et al., Tissue Antigens 1998:52:501-9; Takiguchi, et al., Tissue Antigens 2000:55:296-302.), modifications based on such regularities can be introduced into the peptides of the present invention.

[0125] For example, in a peptide having binding affinity to HLA class I molecules, the second amino acid from the N-terminus and the C-terminal amino acid are generally anchor residues involved in binding to HLA class I (Rammensee HG, et al., Immunogenetics. 1995; 41(4): 178-228). For example, in HLA-A33, phenylalanine, tyrosine, alanine, isoleucine, leucine, and valine, the second amino acid from the N-terminus, and arginine and lysine, the C-terminal amino acid, are known as anchor residues having high binding affinity to HLA-A33 (Falk, et al., Immunogenetics 1994, 40: 232-41; Takiguchi, et al., Tissue Antigens 2000, 55: 296-302).

[0126] In addition, in HLA-A33, the first amino acid residue from the N-terminus is known to function as an anchor residue, and aspartic acid and glutamic acid are known to be preferred as the first amino acid from the N-terminus (Falk, et al., Immunogenetics 1994, 40: 232-41; Takiguchi, et al., Tissue Antigens 2000: 55: 296-302). Therefore, in order to maintain or enhance HLA-A33 binding affinity, it may be desirable to replace the first amino acid from the N-terminus with aspartic acid or glutamic acid, and replace the second amino acid from the N-terminus with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine, and / or replace the C-terminal amino acid with arginine or lysine.

[0127] Therefore, peptides comprising the following amino acid sequences having CTL inducibility are included in the peptides of the present invention, wherein in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, the first amino acid from the N-terminus is substituted with aspartic acid or glutamic acid, the second amino acid from the N-terminus is substituted with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine, and / or the C-terminal amino acid is substituted with arginine or lysine.

[0128] In a preferred embodiment, the peptide of the present invention may be a peptide consisting of an amino acid sequence having CTL inducibility, wherein, in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, the first amino acid from the N-terminus is substituted with aspartic acid or glutamic acid, the second amino acid from the N-terminus is substituted with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine, and / or the C-terminal amino acid is substituted with arginine or lysine.

[0129] That is, the peptides of the present invention include peptides having CTL inducibility comprising an amino acid sequence, which is contained in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, having one or more substituted amino acid sequences selected from the following (a) to (c):

[0130] (a) the first amino acid from the N-terminus is substituted with aspartic acid or glutamic acid;

[0131] (b) the second amino acid from the N-terminus is substituted with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine; and

[0132] (c) The C-terminal amino acid is substituted with arginine or lysine.

[0133] In a preferred embodiment, the peptide of the present invention may be a peptide consisting of an amino acid sequence having CTL inducibility, wherein one or more substitutions selected from (a) to (c) above are introduced into an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46. In the present invention, the preferred number of substitutions is 1, 2 or 3 substitutions selected from (a) to (c) above.

[0134] Furthermore, the peptide of the present invention may be a peptide having CTL inducibility comprising an amino acid sequence, wherein in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, the second amino acid from the N-terminus is substituted with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine, and / or the C-terminal amino acid is substituted with arginine or lysine. Preferably, the peptide of the present invention may be a peptide having CTL inducibility consisting of an amino acid sequence, wherein in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, the second amino acid from the N-terminus is substituted with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine, and / or the C-terminal amino acid is substituted with arginine or lysine. That is, the peptide of the present invention may be a peptide having CTL inducibility, comprising an amino acid sequence having one or more substitutions selected from the following (a) and (b) in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46:

[0135] (a) the second amino acid from the N-terminus is substituted with phenylalanine, tyrosine, alanine, isoleucine, leucine or valine;

[0136] (b) The C-terminal amino acid is substituted with arginine or lysine.

[0137] In a preferred embodiment, the peptide of the present invention may be a peptide having CTL inducibility, which consists of an amino acid sequence having one or more substitutions selected from the above (a) and (b) in an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46. In a more preferred embodiment, the second amino acid from the N-terminus is substituted with phenylalanine or tyrosine.

[0138] In HLA-A01, aspartic acid and glutamic acid, which are the third amino acid from the N-terminus, and tyrosine, which is the amino acid at the C-terminus, are known to be anchor residues having high binding affinity to HLA-A01. In addition, it is known that there is an auxiliary anchor residue at the second position from the N-terminus of HLA-A01, and it is known that threonine and serine are preferred as the second amino acid from the N-terminus (Kubo, RT Journal of Immunology 1994, 152: 3913; Gambacorti-Passerini, C. Clinical Cancer Research 1997, 3: 675-83; Falk, K. Immunogenetics 1994, 40: 238-41).

[0139] Therefore, in order to maintain or enhance HLA-A01 binding affinity, it may be necessary to replace the third amino acid from the N-terminus with aspartic acid or glutamic acid, and / or replace the C-terminal amino acid with tyrosine. Another possibility is that the second amino acid from the N-terminus needs to be replaced with threonine or serine. Therefore, peptides comprising the following amino acid sequences having CTL inducibility are included in the peptides of the present invention, wherein in the amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, the second amino acid from the N-terminus is substituted with threonine or serine, the third amino acid from the N-terminus is substituted with aspartic acid or glutamic acid, and / or the C-terminal amino acid is substituted with tyrosine.

[0140] In a preferred embodiment, the peptide of the present invention may be a peptide consisting of an amino acid sequence having CTL inducibility, wherein in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, the second amino acid from the N-terminus is substituted with threonine or serine, the third amino acid from the N-terminus is substituted with aspartic acid or glutamic acid, and / or the C-terminal amino acid is substituted with tyrosine.

[0141] That is, the peptides of the present invention include peptides having CTL inducibility comprising an amino acid sequence, which is contained in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, having one or more substituted amino acid sequences selected from the following (a) to (c):

[0142] (a) the second amino acid from the N-terminus is substituted with threonine or serine;

[0143] (b) the third amino acid from the N-terminus is substituted with aspartic acid or glutamic acid;

[0144] (c) The C-terminal amino acid was substituted with tyrosine.

[0145] In a preferred embodiment, the peptide of the present invention may be a peptide consisting of an amino acid sequence having CTL inducibility, wherein the amino acid sequence has one or more substitutions selected from the above (a) to (c) in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61. In the present invention, the preferred number of substitutions is 1, 2 or 3 substitutions selected from the above (a) to (c).

[0146] Furthermore, the peptide of the present invention may be a peptide having CTL inducibility comprising an amino acid sequence, wherein the third amino acid from the N-terminus is substituted with aspartic acid or glutamic acid, and / or the C-terminal amino acid is substituted with tyrosine in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61. Preferably, the peptide of the present invention may be a peptide having CTL inducibility consisting of an amino acid sequence, wherein the third amino acid from the N-terminus is substituted with aspartic acid or glutamic acid, and / or the C-terminal amino acid is substituted with tyrosine in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61. That is, the peptides of the present invention include peptides having CTL inducibility comprising an amino acid sequence, wherein the amino acid sequence has one or more substitutions selected from the following (a) and (b) in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61:

[0147] (a) the third amino acid from the N-terminus is substituted with aspartic acid or glutamic acid; and

[0148] (b) The C-terminal amino acid was substituted with tyrosine.

[0149] In a preferred embodiment, the peptide of the present invention may be a peptide consisting of an amino acid sequence having CTL inducibility, wherein the amino acid sequence has one or more substitutions selected from the above (a) to (b) in an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61.

[0150] Amino acid substitutions can be introduced not only at the anchoring site, but also at the potential T cell receptor (TCR) recognition site of the peptide. Several studies have demonstrated that peptides with amino acid substitutions can function equivalent to or better than the original function, such as CAP1, p53 (264-272) 、Her-2 / neu (369-377) or gp100 (209-217) (Zaremba et al. Cancer Res. 57, 4570-4577, 1997, TK Hoffmann et al. J Immunol. (2002); 168(3): 1338-47., SODionne et al. Cancer Immunol immunother. (2003) 52: 199-206 and SODionne et al. Cancer Immunology, Immunotherapy (2004) 53, 307-314).

[0151] The present invention also contemplates that one, two or several amino acids may be added to the N-terminus and / or C-terminus of the peptides of the present invention (e.g., peptides consisting of amino acid sequences selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61). More specifically, the present invention provides peptides consisting of amino acids, wherein one, two or several amino acids are added to both or one of the N-terminus and C-terminus of the amino acid sequence shown in each SEQ ID NO. Such modified peptides that retain CTL inducibility are also included in the present invention. For example, when the peptide is contacted with APC (when one, two or several amino acids are added to the N-terminus and / or C-terminus of the peptide consisting of the amino acid sequence of SEQ ID NO: 2 or 56), it is incorporated into the APC and processed into a peptide consisting of the amino acid sequence of SEQ ID NOs: 2 or 56. It can then induce CTLs on the cell surface of APC by presenting via an antigen presentation pathway. More specifically, the peptide of the present invention may be a peptide in which one, two or several amino acids are added to either or both of the N-terminus and the C-terminus.

[0152] In addition, in another embodiment of the present invention, a peptide consisting of an amino acid sequence is provided, wherein the amino acid sequence contains one, two or several amino acid substitutions in the amino acid sequence shown in each SEQ ID NOs, and wherein one, two or several amino acids are added to one or both of the N-terminus and C-terminus of these substituted amino acid sequences.

[0153] When the peptide of the present invention contains an amino acid substitution, the desired substitution position can be, for example, selected from the first position from the N-terminus, the second position from the N-terminus, and one, two or three positions at the C-terminus in the amino acid sequence shown in SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, or selected from the second position from the N-terminus, the third position from the N-terminus, and one, two or three positions at the C-terminus in the amino acid sequence shown in SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61.

[0154] However, when the amino acid sequence of a peptide is identical to a portion of the amino acid sequence of an endogenous or exogenous protein having a different function, side effects such as autoimmune diseases and / or allergic symptoms against a specific substance may be induced. Therefore, it is preferred to perform a homology search using an available database to avoid a situation where the amino acid sequence of a peptide matches the amino acid sequence of another protein. When it is clear from a homology search that there are no peptides that differ from the target peptide by only 1 or 2 amino acids, the target peptide may be modified to increase its binding affinity to HLA antigens and / or to increase its CTL inducibility without the risk of such side effects.

[0155] It is predicted that a peptide in which one, two or several amino acids of the peptide of the present invention are modified can retain the CTL inducibility of the original peptide; however, it is preferred to verify the CTL inducibility of the modified peptide. Herein, "a peptide having CTL inducibility (CTL inducibility)" refers to a peptide that induces CTLs by APCs stimulated with the peptide.

[0156] "CTL induction" includes induction of differentiation into CTLs, induction of CTL activation, induction of CTL proliferation, induction of CTL cytotoxic activity, induction of CTL-mediated target cell lysis, and induction of increased IFN-gamma production by CTLs.

[0157] The CTL inducibility can be confirmed by stimulating APCs (e.g., B lymphocytes, macrophages, or dendritic cells) expressing the HLA antigen of interest with peptides and mixing them with CD8-positive cells; then measuring IFN-gamma released by CTLs against target cells. For APCs, human peripheral blood mononuclear cell-derived dendritic cells can be preferably used. As a reaction system, transgenic animals that have been made to express HLA antigens can be used. Alternatively, for example, 51Cr or such radioactive labeling target cells, and the cytotoxic activity of peptide-induced CTL can be calculated from the radioactivity released from the target cells. Alternatively, in the presence of peptide-stimulated APCs, CTL inducibility may be assessed by measuring IFN-gamma produced and released by CTLs and visualizing the inhibition zone on the culture medium using an anti-IFN-gamma monoclonal antibody.

[0158] In addition to the above modifications, the peptides of the present invention can also be connected to other peptides, as long as the resulting connected peptide retains CTL inducibility. Examples of suitable peptides connected to the peptides of the present invention include CTL inducing peptides derived from other TAAs. In addition, the peptides of the present invention can also be connected to each other. Suitable linkers for connecting peptides are known in the art, and for example, linkers such as AAY (PM Daftarian et al., J Trans Med 2007, 5: 26), AAA, NKRK (SEQ ID NO: 62) (RPMSutmuller et al., J Immunol. 2000, 165: 7308-15) or K (S. Ota et al., Can Res. 62, 1471-6; KSKawamura et al., J Immunol. 2002, 168: 5709-15) can be used. The peptides can be connected in various arrangements (e.g., chain, repeat, etc.), and three or more peptides can also be connected.

[0159] The peptides of the present invention may also be linked to other substances, as long as the resulting linked peptide retains CTL inducibility. Examples of suitable substances linked to the peptides of the present invention include, for example, peptides, lipids, sugars or sugar chains, acetyl groups, and natural or synthetic polymers. The peptides of the present invention may be modified as follows: glycosylation, side chain oxidation, or phosphorylation, etc., as long as their CTL inducibility is not destroyed. This type of modification may also be implemented to impart additional functions (e.g., targeting functions and delivery functions) or to stabilize the peptides.

[0160] For example, to improve the in vivo stability of peptides, it is known in the art to introduce D-amino acids, amino acid mimetics or unnatural amino acids; this concept can also be applied to the peptides of the present invention. The stability of peptides can be determined in a variety of ways. For example, stability can be tested by using peptidases and various biological media such as human plasma and serum (see, for example, Verhoef et al., Eur J Drug Metab Pharmacokin 1986, 11: 291-302).

[0161] In addition, as described above, among the modified peptides substituted, deleted, inserted and / or added with 1, 2 or several amino acid residues, those having the same or higher activity compared with the original peptide can be screened or selected. Therefore, the present invention also provides a method for screening or selecting modified peptides having the same or higher activity compared with the original peptide. Specifically, the present invention provides a method for screening peptides having CTL inducibility, wherein the method comprises the following steps:

[0162] (a) generating a candidate sequence consisting of an amino acid sequence in which one, two or several amino acid residues are substituted, deleted, inserted and / or added to an original amino acid sequence, wherein the original amino acid sequence consists of an amino acid sequence selected from the group consisting of SEQ ID Nos: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61;

[0163] (b) selecting, from the candidate sequences generated in (a), candidate sequences that have no significant homology (or sequence homology) to any peptide of a known human gene product other than FOXM1;

[0164] (c) contacting the peptide consisting of the candidate sequence selected in (b) with APC;

[0165] (d) contacting the APC of (c) with CD8-positive T cells; and

[0166] (e) Selecting a peptide having a CTL inducibility equal to or higher than that of a peptide consisting of the original amino acid sequence.

[0167] The peptides of the present invention are also described herein as "FOXM1 peptides".

[0168] III. Preparation of the Peptides of the Invention

[0169] The peptides of the present invention can be prepared using known techniques. For example, the peptides of the present invention can be prepared using recombinant DNA technology or chemical synthesis. The peptides of the present invention can be synthesized independently, or synthesized as longer polypeptides comprising two or more peptides. The peptides of the present invention can be separated from host cells or synthetic reaction products after producing them in host cells using recombinant DNA technology or chemically synthesizing them. That is, the peptides of the present invention can be purified or isolated so that they are substantially free of other host cell proteins and fragments thereof, or any other chemical substances.

[0170] The peptides of the present invention may contain modifications, such as glycosylation, side chain oxidation, or phosphorylation, etc., provided that the modifications do not destroy the biological activity of the original peptide. Other exemplary modifications include incorporating one or more D-amino acids or other available amino acid mimetics, such as to increase the serum half-life of the peptide.

[0171] The peptide of the present invention can be obtained by chemical synthesis according to the selected amino acid sequence. Examples of conventional peptide synthesis methods that can be suitable for synthesis include the methods described in the following documents:

[0172] (i) Peptide Synthesis, Interscience, New York, 1966;

[0173] (ii)The Proteins,Vol.2,Academic Press,New York,1976;

[0174] (iii) "Peptide Synthesis" (in Japanese), Maruzen Co., 1975;

[0175] (iv) "Basics and Experiment of Peptide Synthesis" (in Japanese), MaruzenCo., 1985;

[0176] (v) "Development of Pharmaceuticals" (in Japanese), Continued Vol.14 (peptide synthesis), Hirokawa, 1991;

[0177] (vi)WO99 / 67288; and

[0178] (vii) Barany G.&Merrifield RB, Peptides Vol.2, Solid Phase PeptideSynthesis, Academic Press, New York, 1980, 100-118.

[0179] Alternatively, any known genetic engineering peptide production method can be applied to obtain the peptide of the present invention (e.g. Morrison J, J Bacteriology 1977, 132: 349-51; Clark-Curtiss & Curtiss, Methods in Enzymology (eds. Wu et al.) 1983, 101: 347-62). For example, first, a suitable vector containing a polynucleotide encoding a target peptide in an expressible form (e.g., downstream of a regulatory sequence equivalent to a promoter sequence) is prepared and transformed into a suitable host cell. The present invention also provides such vectors and host cells. The host cells are then cultured to produce the peptide of interest. The peptide of the present invention can also be produced in vitro using an in vitro translation system.

[0180] IV. Polynucleotides

[0181] The present invention also provides polynucleotides encoding any of the peptides of the present invention. These include polynucleotides derived from naturally occurring FOXM1 genes (e.g., GenBank Accession Nos. NM_202003 (SEQ ID NO: 52), NM_001243088 (SEQ ID NO: 54), NM_001243089 (SEQ ID NO: 56), NM_021953 (SEQ ID NO: 58) or NM202002 (SEQ ID NO: 60)) and polynucleotides having conservatively modified nucleotide sequences thereof. Herein, the phrase "conservatively modified nucleotide sequence" refers to a sequence encoding the same or substantially the same amino acid sequence. Due to the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For example, the codons GCA, GCC, GCG, and GCU all encode the amino acid alanine. Therefore, at any position specified by a codon as alanine, the codon can be changed to any corresponding codon without changing the encoded polypeptide. Such nucleic acid variations are "silent variations", which are a type of conservatively modified variations. Each nucleic acid sequence encoding a peptide herein also describes each possible silent variation of the nucleic acid. One of ordinary skill in the art will recognize that each codon in a nucleic acid (except AUG and TGG, which are normally the only codon for methionine and TGG, which are normally the only codon for tryptophan) can be modified to produce a functionally identical molecule. Therefore, each silent variation of a nucleic acid encoding a peptide is implicitly described in each disclosed sequence.

[0182] The polynucleotide of the present invention can be composed of DNA, RNA, and derivatives thereof. DNA is appropriately composed of bases such as A, T, C, and G, and T is replaced by U in RNA.

[0183] The polynucleotides of the present invention can encode a plurality of peptides of the present invention, with or without intervening amino acid sequences. For example, the intervening amino acid sequence can provide a cleavage site (e.g., an enzyme recognition sequence) for the polynucleotide or the translated peptide. In addition, the polynucleotide can include any additional sequence of the coding sequence encoding the peptide of the present invention. For example, the polynucleotide can be a recombinant polynucleotide including the regulatory sequences required for expressing the peptide, or can be an expression vector (e.g., a plasmid) with a marker gene or the like. In general, such recombinant polynucleotides can be prepared by manipulating the polynucleotides using conventional recombinant techniques, such as by using a polymerase and an endonuclease.

[0184] Both recombinant and chemical synthesis techniques can be used to generate the polynucleotides of the present invention. For example, the polynucleotides of the present invention can be generated by inserting a suitable vector that can be expressed after transfection into competent cells. Alternatively, polynucleotides can be amplified using PCR technology or expressed in a suitable host (see, for example, Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, New York, 1989). Alternatively, solid phase techniques can be used to synthesize polynucleotides, such as Beaucage SL & Iyer RP, Tetrahedron 1992, 48: 2223-311; Matthes et al., EMBO J1984, 3: 801-5. The connection products of several peptides that can be obtained in this way can be purified as needed and applied in the connected form. In this case, the connected peptides are processed to produce peptides for antigen presentation and induce the CTL induction activity of each peptide. Therefore, when peptides are connected, it is preferred to have a combination of peptides with the same HLA restriction. Or the peptides can be applied as a mixture of single peptides by cutting the connecting portion.

[0185] V. Exosomes

[0186] The present invention further provides intracellular vesicles called exosomes, which present complexes formed between the peptides of the present invention and HLA antigens on their surfaces. Exosomes can be prepared by methods described in detail, for example, in JPH11-510507 and WO99 / 03499, and can be prepared using APCs obtained from patients who are the subject of treatment and / or prevention. The exosomes of the present invention can be used as vaccines to be inoculated in a similar manner to the peptides of the present invention.

[0187] The type of HLA antigen contained in the above complex must match the type of the subject in need of treatment and / or prevention (prevention).* For example, HLA-A01 (e.g., HLA-A03:03) is an allele widely and commonly seen in Asians, and this HLA antigen type is considered suitable for treatment in Asian patients. In addition, HLA-A01 (e.g., HLA-A*01:01) is an allele widely and commonly seen in Caucasian populations, and these HLA antigen types are considered suitable for treatment in Caucasian patients. Typically, in the clinic, by preliminarily studying the HLA antigen type of the patient in need of treatment, a suitable peptide can be selected that has a high level of binding affinity for a specific HLA antigen, or CTL inducibility through antigen presentation mediated by a specific HLA antigen.

[0188] The exosomes of the present invention present a complex of the peptide of the present invention and HLA-A33 or HLA-A01 on their surface. When the HLA that forms a complex with the peptide of the present invention is HLA-A33, the peptide of the present invention is preferably a peptide having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, or a modified peptide thereof, and more preferably a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, or a modified peptide thereof. Furthermore, when the HLA that forms a complex with the peptide of the present invention is HLA-A01, the peptide of the present invention is preferably a peptide having an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, or a modified peptide thereof, and more preferably a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, or a modified peptide thereof.

[0189] VI. Antigen Presenting Cells (APC)

[0190] The present invention also provides an APC that presents a complex formed between an HLA antigen and a peptide of the present invention on its surface. Or the present invention provides an APC having a complex formed between an HLA antigen and a peptide of the present invention on its surface. The APC of the present invention may be an isolated APC. When used in the context of cells (APCs, CTLs, etc.), the term "isolated" means that the cell is separated from another type of cell. The APC of the present invention may be an APC induced from an APC derived from a patient to be treated and / or prevented (prevented), and may be administered as a vaccine itself or in combination with other drugs (including peptides of the present invention, exosomes or CTLs).

[0191] The APCs of the present invention are not limited to specific types of cells, and may be cells that inhibit the presentation of protein antigens on their cell surfaces for recognition by lymphocytes (e.g., dendritic cells (DC), Langerhans cells, macrophages, B cells, and activated T cells). Since DC is a representative APC with the strongest CTL induction activity among APCs, DC may be preferably used as the APCs of the present invention. In the present invention, preferred DCs are isolated DCs derived from humans. In addition, the APCs of the present invention may also be a mixture of various types of cells having antigen presenting function, and may be a mixture of APCs, wherein each APC presents different types of peptides of the present invention.

[0192] For example, the APCs of the present invention can be obtained by isolating DCs from peripheral blood mononuclear cells and then stimulating them with the peptides of the present invention in vitro. When the peptides of the present invention are administered to a subject, APCs that present the peptides of the present invention are induced in the subject's body. Therefore, after administering the peptides of the present invention to the subject, the APCs of the present invention can be obtained by collecting APCs from the subject. Alternatively, the APCs of the present invention can be obtained by contacting APCs collected from a subject with the peptides of the present invention.

[0193] In order to induce an immune response against cancer cells expressing FOXM1 in a subject, the APC of the present invention can be administered to the subject by itself or in combination with other drugs (including the peptides, exosomes or CTLs of the present invention). For example, ex vivo administration may include the following steps:

[0194] (a) collecting APCs from a first subject;

[0195] (b) contacting the APC of step (a) with the peptide of the present invention; and

[0196] (c) administering the APCs of step (b) to a second subject.

[0197] The first subject and the second subject may be the same individual, or may be different individuals. When the first subject and the second subject are different individuals, preferably the HLA of the first subject and the second subject are of the same type. The APC obtained from the above step (b) may be a vaccine for cancer treatment and / or prevention.

[0198] The APCs of the present invention obtained by the method described above have CTL inducibility. The term "CTL inducibility (CTL inducibility)" used in the context of APC refers to the ability of APC to induce CTL when in contact with CD8 positive cells. In addition, "CTL inducibility (CTL inducibility)" includes the ability of APC to induce CTL activation, the ability of APC to induce CTL proliferation, the ability of APC to promote CTL-mediated lysis of target cells, and the ability of APC to increase CTL-mediated IFN-gamma production. The CTL induced by the APC of the present invention is a CTL specific for FOXM1 and exhibits specific cytotoxic activity against cells expressing FOXM1.

[0199] In addition to the above methods, the APCs of the present invention can be prepared by introducing a polynucleotide encoding the peptide of the present invention into APCs in vitro. The polynucleotide to be introduced can be in the form of DNA or RNA. The method of introduction is not particularly limited, and examples thereof include various methods routinely practiced in the art, such as liposome transfection, electroporation, and calcium phosphate method. More specifically, the methods described in Cancer Res 1996, 56: 5672-7; J Immunol 1998, 161: 5607-13; J ExpMed 1996, 184: 465-72, and JP2000-509281 can be used. By introducing a polynucleotide encoding the peptide of the present invention into APC, the polynucleotide is transcribed and translated in the cell, and then the produced peptide is processed by MHC class I and presented on the cell surface of the APC through a presentation pathway.

[0200] In a preferred embodiment, the APCs of the present invention present the peptides of the present invention and HLA-A11 (more preferably HLA-A * 33:03) or HLA-A01 (more preferably HLA-A *When the HLA that forms a complex with the peptide of the present invention is HLA-A33, the peptide of the present invention is preferably a peptide having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, or a modified peptide thereof, and more preferably a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46. When the HLA that forms a complex with the peptide of the present invention is HLA-A01, the peptide of the present invention is preferably a peptide having an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, or a modified peptide thereof, and more preferably a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61.

[0201] The APCs of the present invention are preferably APCs induced by a method comprising the following steps (a) or (b):

[0202] (a) contacting APCs expressing at least one HLA selected from HLA-A33 (more preferably HLA-A*33:03) and HLA-A01 (more preferably HLA-A*01:01) with the peptide of the present invention; or

[0203] (b) introducing a polynucleotide encoding the peptide of the present invention into APCs expressing at least one HLA selected from HLA-A33 (more preferably HLA-A*33:03) and HLA-A01 (more preferably HLA-A*01:01).

[0204] The peptide of the present invention to be contacted with APC expressing HLA-A33 is preferably a peptide having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, or a modified peptide thereof, and more preferably a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46.

[0205] The peptide of the present invention that is contacted with the APC expressing HLA-A01 is preferably a peptide having an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, or a modified peptide thereof, and more preferably a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61.

[0206] The present invention provides the use of the peptide of the present invention for preparing a pharmaceutical composition for inducing APCs with CTL inducibility. In addition, the present invention provides a method or process for preparing a pharmaceutical composition for inducing APCs with CTL inducibility. Further, the present invention provides the peptide of the present invention for inducing APCs with CTL inducibility.

[0207] VII. Cytotoxic T lymphocytes (CTL)

[0208] The CTL induced by the peptide of the present invention can be used as a vaccine in a similar manner to the peptide of the present invention for enhancing the immune response of cancer cells targeting FOXM1 expression in vivo. Therefore, the present invention provides CTLs, which are induced or activated by the peptide of the present invention. The CTL of the present invention is a CTL that targets the peptide of the present invention and is capable of binding to a complex of the peptide of the present invention and an HLA antigen. The binding of CTL to the complex is mediated by the T cell receptor (TCR) present on the cell surface of the CTL. The CTL of the present invention may be an isolated CTL. Preferred CTLs are isolated human CTLs. The CTL of the present invention may also be a mixture of CTLs, each of which targets a different type of peptide of the present invention.

[0209] The CTL of the present invention can be obtained by (1) administering the peptide of the present invention to a subject, (2) stimulating APCs and CD8-positive T cells or peripheral blood mononuclear cells (PBMCs) derived from a subject with the peptide of the present invention in vitro, (3) contacting CD8-positive T cells or PBMCs with APCs or exosomes in vitro, the APCs or exosomes presenting a complex of HLA antigens and the peptide of the present invention on their surfaces, or (4) introducing a vector containing a polynucleotide encoding each subunit of a T cell receptor (TCR) capable of binding to the peptide of the present invention presented by the HLA antigen on the cell surface into CD8-positive T cells. The exosomes and APCs used in the method of (2) or (3) above can be prepared by the methods described in the sections "V. Exosomes" and "VI. Antigen presenting cells (APCs)", respectively, and the details of the method of (4) above will be described in the section "VIII. T cell receptor (TCR)".

[0210] The CTL of the present invention can be administered alone to a patient undergoing treatment and / or prevention (prevention), or in combination with other drugs (including peptides of the present invention, APCs or exosomes) for the purpose of regulating the effect. Further, the CTL of the present invention can be a CTL induced from CD8 positive T cells, which are derived from patients undergoing administration of CTL. The CTL of the present invention specifically acts on target cells presenting the peptide of the present invention, for example, the same peptide used to induce the CTL of the present invention. The target cell can be a cell endogenously expressing FOXM1, such as a cancer cell, or a cell transfected with the FOXM1 gene. Cells presenting the peptide of the present invention on the cell surface due to the stimulation of the peptide can also become the attack target of the CTL of the present invention. The cells targeted by the CTL of the present invention are preferably cells that are positive for at least one of HLA-A33 (more preferably HLA-A*33:03) and HLA-A01 (more preferably HLA-A*01:01).

[0211] In a preferred embodiment, the CTL of the present invention specifically targets cells expressing FOXM1 and at least one HLA selected from HLA-A33 (more preferably HLA-A*33:03) and HLA-A01 (more preferably HLA-A*01:01). In the present invention, the cell targeted by CTL may be a cell homozygous or heterozygous for any one of the alleles of HLA-A33 and HLA-A01.

[0212] Herein, CTL "targeting" cells refers to the recognition of CTL on cells presenting a complex of HLA and the peptide of the present invention on its cell surface, and the expression of cytotoxic activity on the cells. Further, "specific targeting" means that CTL exhibits cytotoxic activity on these cells, but does not show destructive activity on other cells. The expression "recognizing cells" used in the context of CTL refers to binding to a complex of HLA and the peptide of the present invention presented on the cell surface via its TCR, and exhibiting specific cytotoxic activity on the cell. Therefore, the CTL of the present invention is preferably capable of binding to a complex of the peptide of the present invention and HLA-A presented on the cell surface via TCR. * 33:03) or HLA-A01 (more preferably HLA-A * 01:01) between CTLs that form a complex.

[0213] Furthermore, the CTL of the present invention is preferably a CTL induced by a method comprising the steps described in the following (a) or (b):

[0214] (a) contacting CD8-positive T cells with APCs or exosomes presenting a complex of the peptide of the present invention and HLA-A33 (more preferably HLA-A*33:03) or HLA-A01 (more preferably HLA-A*01:01) on its surface in vitro; or

[0215] (b) A polynucleotide containing a subunit encoding each TCR capable of binding to the peptide of the present invention presented on the cell surface by HLA-A33 (more preferably HLA-A*33:03) or HLA-A01 (more preferably HLA-A*01:01) is introduced into CD8-positive T cells. The CTLs induced by this method have TCRs that specifically recognize the complex of the peptide for induction and the HLA antigen. Therefore, they are cells with a structure different from other CTLs having different reaction specificities due to differences in TCR structure.

[0216] VIII. T cell receptor (TCR)

[0217] The present invention also provides a composition and a method for using the composition, wherein the composition comprises a polynucleotide encoding each TCR subunit, and the TCR is capable of binding to the peptide of the present invention presented on the cell surface by HLA antigen. The polynucleotide confers specificity to CD8 positive T cells for cancer cells expressing FOXM1, which is achieved by expressing a TCR that can bind to the peptide of the present invention presented on the target cell surface of CD8 positive T cells by HLA antigen. By using methods known in the art, polynucleotides encoding alpha and beta chains of TCR subunits in CTLs induced by the peptide of the present invention can be identified (WO2007 / 032255 and Morgan et al., J Immunol, 171, 3288 (2003)). For example, preferably PCR methods are used for TCR analysis. Without being limited thereto, the PCR primers used for analysis can be, for example, a primer set for amplification by combining the following 5' side primers and 3' side primers:

[0218] 5' primer:

[0219] 5'-R primer (5'-gtctaccaggcattcgcttcat-3') (SEQ ID NO: 62)

[0220] 3' primer:

[0221] TCR-alpha-chain C-region-specificity

[0222] 3-TRa-C primer (5'-tcagctggaccacagccgcagcgt-3') (SEQ ID NO: 63)

[0223] TCR-beta-chain C1-region-specificity

[0224] 3-TRb-C1 primer (5'-tcagaaatcctttctcttgac-3') (SEQ ID NO: 64) or

[0225] TCR-beta-chain C2-region-specificity

[0226] 3-TR-beta-C2 primer (5'-ctagcctctggaatcctttctctt-3') (SEQ ID NO: 65)

[0227] TCRs formed by introducing the identified polynucleotides into CD8-positive T cells are able to bind with high affinity to target cells presenting the peptides of the present invention and mediate efficient killing of target cells presenting the peptides of the present invention in vivo and in vitro.

[0228] The polynucleotides encoding each TCR subunit can be incorporated into a suitable vector, such as a retroviral vector. These vectors are well known in the art. The polynucleotides or vectors comprising them can be introduced into CD8 positive T cells, such as CD8 positive T cells derived from patients. The present invention provides a ready-to-use composition that allows rapid modification of the patient's own T cells (or T cells derived from other subjects) to quickly and easily generate modified T cells with excellent cancer cell killing properties.

[0229] Herein, specific TCR is when TCR is presented on the surface of CD8 positive T cells, can be presented on the target cell surface by specific recognition of the peptide of the present invention and the complex of HLA antigen, give TCR for target cell specific cytotoxic activity.The specific recognition of the above-mentioned complex can be confirmed by any known method, and its preferred example includes HLA polymer staining analysis using HLA molecules and peptides of the present invention, and ELISPOT assay method.By implementing ELISPOT assay method, can confirm by the T cell of T cell and intracellular signal transduction of the above-mentioned polynucleotide specific TCR mediated recognition of target cells.When the above-mentioned TCR is presented on the surface of CD8 positive T cells, it is also possible to confirm by known methods whether TCR can give target cell specific cytotoxic activity for CD8 positive T cells.Preferred method includes, for example, measuring the cytotoxic activity for HLA positive target cells by chromium release assay method etc.

[0230] In the context of HLA-A33, the present invention provides CTLs prepared by transforming CD8-positive T cells with a polynucleotide encoding each TCR subunit (which, for example, binds to a complex formed by a peptide having an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46 and an HLA-A33 antigen).

[0231] In the context of HLA-A01, the present invention provides CTLs prepared by transforming CD8-positive T cells with a polynucleotide encoding each TCR subunit (which, for example, binds to a complex formed by a peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 and an HLA-A01 antigen).

[0232] The transformed CTL can be homed in vivo (translocation from blood to lymphatic tissue), and can be expanded using known in vitro culture methods (e.g., Kawakami et al., J Immunol., 1989, 142: 3252-61). The CTL of the present invention can be used to form an immunogenic composition, which is useful for the treatment or prevention of diseases in patients in need of treatment and / or prevention (prevention) (for reference WO2006 / 031221, the contents of which are incorporated herein by reference).

[0233] IX. Pharmaceutical Compositions

[0234] The present invention also provides a composition or pharmaceutical composition comprising at least one active ingredient selected from the following:

[0235] (a) a peptide of the present invention;

[0236] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0237] (c) APC of the present invention;

[0238] (d) exosomes of the present invention; and

[0239] (e) CTL of the present invention.

[0240] In addition to the above-mentioned active ingredients, the pharmaceutical composition of the present invention may contain carriers, excipients, etc. commonly used in medicines as needed without particular limitation. Examples of carriers that can be used in the pharmaceutical composition of the present invention include sterile water, physiological saline, phosphate buffer, culture fluid, etc. Therefore, the present invention also provides a pharmaceutical composition comprising at least one active ingredient selected from the following (a) to (e) and a pharmaceutically acceptable carrier:

[0241] (a) a peptide of the present invention;

[0242] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0243] (c) APC of the present invention;

[0244] (d) exosomes of the present invention; and

[0245] (e) CTL of the present invention.

[0246] In addition, the pharmaceutical composition of the present invention may contain stabilizers, suspending agents, preservatives, surfactants, solubilizers, pH regulators, aggregation inhibitors, etc. as needed.

[0247] Compared with normal tissues, FOXM1 expression is significantly upregulated in cancer cells. Therefore, the peptides of the present invention or the polynucleotides encoding the peptides can be used to treat and / or prevent cancer, and / or prevent its postoperative recurrence. Therefore, the present invention provides a pharmaceutical composition for treating and / or preventing cancer, and / or preventing its postoperative recurrence, which contains one or more types of peptides or polynucleotides of the present invention as active ingredients. Alternatively, the peptides of the present invention can be prepared for presentation on the surface of exosomes or APCs for use as a pharmaceutical composition. In addition, the CTL of the present invention targeting any of the peptides of the present invention can also be used as an active ingredient of the pharmaceutical composition of the present invention. The pharmaceutical composition of the present invention may contain a therapeutically effective amount or a pharmaceutically effective amount of the above-mentioned active ingredient.

[0248] The pharmaceutical composition of the present invention can also be used as a vaccine. In the context of the present invention, the term "vaccine" (also referred to as "immunogenic composition") refers to a composition that has the function of inducing an immune response that leads to an anti-tumor effect when inoculated into an animal. Therefore, the pharmaceutical composition of the present invention can be used to induce an immune response that leads to an anti-tumor effect. The immune response induced by the peptides, polynucleotides, APCs, CTLs and pharmaceutical compositions of the present invention is not particularly limited as long as it is an immune response that leads to an anti-tumor effect, and examples include inducing cancer cell-specific CTLs and inducing cancer cell-specific cytotoxic activity.

[0249] The pharmaceutical composition of the present invention can be used for treating and / or preventing cancer in human subjects or patients, and / or preventing its postoperative recurrence. The pharmaceutical composition of the present invention can be preferably used for subjects who are positive for at least one HLA selected from HLA-A33 and HLA-A01. In addition, the pharmaceutical composition of the present invention can be preferably used for treating and / or preventing cancer expressing FOXM1 and at least one HLA selected from HLA-A33 and HLA-A01 and / or preventing its postoperative recurrence.

[0250] In another embodiment, the present invention provides the use of an active ingredient selected from the following in the preparation of a pharmaceutical composition for treating or preventing cancer:

[0251] (a) a peptide of the present invention;

[0252] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0253] (c) APCs presenting the peptides of the present invention on their surface;

[0254] (d) an exosome presenting a peptide of the present invention on its surface; and

[0255] (e) CTL of the present invention.

[0256] Alternatively, the present invention further provides an active ingredient selected from the following for use in treating or preventing cancer:

[0257] (a) a peptide of the present invention;

[0258] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0259] (c) APCs presenting the peptides of the present invention on their surface;

[0260] (d) an exosome presenting a peptide of the present invention on its surface; and

[0261] (e) CTL of the present invention.

[0262] Alternatively, the present invention also provides a method or process for preparing a pharmaceutical composition for treating or preventing cancer, wherein the method or process comprises the step of formulating at least one active ingredient selected from the following with a pharmaceutically or physiologically acceptable carrier:

[0263] (a) a peptide of the present invention;

[0264] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0265] (c) APCs presenting the peptides of the present invention on their surface;

[0266] (d) an exosome presenting a peptide of the present invention on its surface; and

[0267] (e) CTL of the present invention.

[0268] In another embodiment, the present invention also provides a method or process for preparing a pharmaceutical composition for treating or preventing cancer, wherein the method or process comprises the step of mixing an active ingredient selected from the following with a pharmaceutically or physiologically acceptable carrier:

[0269] (a) a peptide of the present invention;

[0270] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0271] (c) APCs presenting the peptides of the present invention on their surface;

[0272] (d) an exosome presenting a peptide of the present invention on its surface; and

[0273] (e) CTL of the present invention.

[0274] In another embodiment, the present invention further provides a method for treating or preventing cancer, comprising the step of administering to a subject at least one active ingredient selected from the following:

[0275] (a) a peptide of the present invention;

[0276] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0277] (c) APCs presenting the peptides of the present invention on their surface;

[0278] (d) an exosome presenting a peptide of the present invention on its surface; and

[0279] (e) CTL of the present invention.

[0280] In the present invention, peptides having an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46 are identified as HLA-A33 restricted epitope peptides that can induce a potent and specific immune response. Therefore, the pharmaceutical composition of the present invention comprising at least one peptide having an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46 is particularly suitable for administration to a subject having HLA-A33 (e.g., HLA-A*33:03) as an HLA antigen. The same applies to pharmaceutical compositions comprising a polynucleotide encoding any of these peptides (i.e., a polynucleotide of the present invention), an APC or exosome presenting these peptides (i.e., an APC or exosome of the present invention), or a CTL targeting these peptides (i.e., a CTL of the present invention). That is, a pharmaceutical composition comprising an active ingredient associated with a peptide having an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, and 46 is suitable for administration to a subject having HLA-A33 (i.e., an HLA-A33-positive subject). In a more preferred embodiment, the pharmaceutical composition of the present invention is a pharmaceutical composition comprising a peptide having an amino acid sequence of SEQ ID NO: 2.

[0281] Similarly, in the present invention, peptides having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 are identified as HLA-A01-restricted epitope peptides that can induce potent and specific immune responses. Therefore, the pharmaceutical composition of the present invention comprising at least one peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 is particularly suitable for administration to a subject having HLA-A01 (e.g., HLA-A*01:01) as an HLA antigen. The same applies to pharmaceutical compositions comprising: a polynucleotide encoding any one of these peptides (i.e., a polynucleotide of the present invention). APCs or exosomes presenting these peptides (i.e., an APC or exosome of the present invention), or CTLs targeting these peptides (i.e., a CTL of the present invention). That is, the pharmaceutical composition comprising an active ingredient associated with a peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 is suitable for administration to a subject having HLA-A01 (i.e., an HLA-A01 positive subject). In a more preferred embodiment, the pharmaceutical composition of the present invention is a pharmaceutical composition comprising a peptide having an amino acid sequence of SEQ ID NO: 56.

[0282] Cancers to be treated and / or prevented by the pharmaceutical composition of the present invention are not particularly limited as long as they are cancers expressing FOXM1, and include various cancers such as acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc. In addition, the pharmaceutical composition of the present invention can be preferably used for subjects homozygous or heterozygously having an HLA allele selected from HLA-A33 and HLA-A01.

[0283] In addition to the above-mentioned active ingredients, the pharmaceutical composition of the present invention may contain other peptides having the ability to induce CTLs against cancer cells (e.g., other TAA-derived CTL-inducing peptides), other polynucleotides encoding other peptides, other cells presenting other peptides, etc.

[0284] The pharmaceutical composition of the present invention may also optionally include other therapeutic substances as active ingredients, as long as they do not inhibit the anti-tumor effect of the above-mentioned active ingredients (e.g., peptides of the present invention). For example, the pharmaceutical composition of the present invention may optionally include anti-inflammatory compositions, analgesics, chemotherapeutic agents, etc. In addition to including other therapeutic substances in the pharmaceutical composition of the present invention itself, the pharmaceutical composition of the present invention may also be administered sequentially or simultaneously with one or more other pharmaceutical compositions. The dosage of the pharmaceutical composition of the present invention and other pharmaceutical compositions depends on, for example, the type of pharmaceutical composition used and the disease being treated, as well as the scheduling of administration and the route of administration.

[0285] It should be understood that the pharmaceutical compositions of this invention may contain, in addition to the ingredients specifically mentioned herein, other ingredients conventional in the art having regard to the type of formulation.

[0286] The present invention also provides an article or kit comprising the pharmaceutical composition of the present invention. The article or kit of the present invention may include a container for holding the pharmaceutical composition of the present invention. Examples of suitable containers include, but are not limited to, bottles, vials or test tubes. The container may be made of a variety of materials, such as glass or plastic. A label may be attached to the container, and the disease or disease state to which the pharmaceutical composition of the present invention is applied may be described in the label. The label may also indicate instructions for administration, etc.

[0287] In addition to the container for holding the pharmaceutical composition of the present invention, preferably the article of manufacture or kit of the present invention may also optionally contain a second container for holding a pharmaceutically acceptable diluent. The article of manufacture or kit of the present invention may also include other materials required from a commercial standpoint and a user's perspective, such as other buffers, diluents, filters, injection needles, syringes, and package inserts with instructions for use.

[0288] As required, the pharmaceutical composition of the present invention can be packaged in a medicine bag or a dispensing device, which can be equipped with one or more unit dosage forms containing active ingredients. For example, the medicine bag can include, for example, metal or plastic foil, such as a blister pack. The medicine bag or the dispensing device can be accompanied by instructions for use.

[0289] (1) Pharmaceutical compositions containing peptides as active ingredients

[0290] The pharmaceutical composition comprising the peptide of the present invention can be formulated by conventional preparation methods as needed. In addition to the peptide of the present invention, the pharmaceutical composition of the present invention may contain carriers, excipients, etc. commonly used in drugs without particular limitation. Examples of carriers that can be used in the pharmaceutical composition of the present invention include sterile water (e.g., water for injection), physiological saline, phosphate buffer, phosphate-buffered saline, Tris-buffered saline, 0.3% glycine, culture medium, and the like. In addition, the pharmaceutical composition of the present invention may contain stabilizers, suspending agents, preservatives, surfactants, solubilizers, pH regulators, aggregation inhibitors, and the like as needed. The pharmaceutical composition of the present invention can induce specific immunity against cancer cells expressing FOXM1, and can therefore be used for the purpose of cancer treatment or prevention (prevention).

[0291] For example, the pharmaceutical composition of the present invention can be prepared by dissolving in a pharmaceutically or physiologically acceptable water-soluble carrier, such as sterile water (e.g., water for injection), physiological saline, phosphate buffer, phosphate buffered saline, and Tris buffered saline, and adding stabilizers, suspending agents, preservatives, surfactants, solubilizers, pH regulators, aggregation inhibitors, etc. as needed, and then sterilizing the peptide solution. The method for sterilizing the peptide solution is not particularly limited, and is preferably performed by filtration sterilization. Filtration sterilization can be performed using, for example, a filtration sterilization filter with a pore size of 0.22 μm or less. The filter-sterilized peptide solution can be administered to a subject, for example, as an injection, but is not limited thereto.

[0292] The pharmaceutical composition of the present invention can be prepared as a freeze-dried preparation by freeze-drying the above-mentioned peptide solution. The freeze-dried preparation can be prepared by the following: the peptide solution prepared as described above is loaded into an appropriate container, such as an ampoule, a vial or a plastic container, followed by freeze drying and encapsulation in a container after pressure recovery with a washed and sterilized rubber stopper, etc. The freeze-dried preparation can be administered to a subject after it is redissolved in a pharmaceutically or physiologically acceptable water-soluble carrier, such as sterile water (e.g., water for injection), saline, phosphate buffer, phosphate-buffered saline, and Tris-buffered saline, etc. Preferred examples of the pharmaceutical composition of the present invention include injections of such filtered sterilized peptide solutions and freeze-dried preparations produced by freeze-drying the peptide solution. The present invention also includes a kit containing such freeze-dried preparations and a re-dissolved solution. The present invention also includes a kit containing a container for accommodating a freeze-dried preparation (which is a pharmaceutical composition of the present invention) and a container for accommodating a re-dissolved solution.

[0293] The pharmaceutical composition of the present invention may comprise a combination of two or more types of peptides of the present invention. The combination of peptides may take the form of a cocktail of mixed peptides, or may be conjugated to each other using standard techniques. For example, the peptides may be chemically linked or expressed as a single fusion polypeptide. By administering the peptides of the present invention, the peptides are displayed on APCs at high density by HLA antigens, and then CTLs that specifically react with the complex formed between the displayed peptides and HLA antigens are induced. Alternatively, APCs (e.g., DCs) may be isolated from a subject, and then stimulated with the peptides of the present invention to obtain APCs that display any of the peptides of the present invention on their cell surfaces. These APCs are re-administered to the subject to induce CTLs in the subject, and as a result, the aggressiveness against cancer cells expressing FOXM1 may be increased.

[0294] The pharmaceutical composition of the present invention may also include adjuvants known to be effective in establishing cellular immunity. An adjuvant refers to a compound that enhances the immune response to an antigen with immunological activity when administered together (or continuously) with an antigen. Known adjuvants described in the literature may be used, such as Clin Microbiol Rev 1994, 7: 277-89. Examples of suitable adjuvants include aluminum salts (aluminum phosphate, aluminum hydroxide, aluminum oxyhydroxide, etc.), alum, cholera toxin, salmonella toxin, IFA (incomplete Freund's adjuvant), CFA (complete Freund's adjuvant), ISCOMatrix, GM-CSF and other immunostimulatory cytokines, oligodeoxynucleotides containing CpG motifs (CpG7909, etc.), oil-in-water emulsions, saponin or its derivatives (QS21, etc.), lipopolysaccharides such as lipid A or its derivatives (MPL, RC529, GLA, E6020, etc.), lipopeptides, lactoferrin, flagellin, double-stranded RNA or its derivatives (poli IC, etc.), bacterial DNA, imidazoquinoline (Imiquimod, R848, etc.), C-type lectin ligands (trehalose-6,6'-dibehenic acid / ester (TDB), etc.), CD1d ligands (alpha-galactosylceramide, etc.), squalene emulsions (MF59, AS03, AF03, etc.), PLGA, etc., but are not limited thereto. In the kit comprising the pharmaceutical composition of the present invention, the adjuvant may be contained in another container separate from the pharmaceutical composition comprising the peptide of the present invention. In this case, the adjuvant and the pharmaceutical composition can be administered to the subject continuously, or mixed together immediately before administration to the subject. The present invention also provides such a kit comprising a pharmaceutical composition of the peptide of the present invention and an adjuvant. When the pharmaceutical composition of the present invention is a freeze-dried preparation, the kit may also include a reconstitution solution. In addition, the present invention provides a kit comprising a container for holding the pharmaceutical composition of the present invention and a container for storing the adjuvant. The kit may further include a container for storing the reconstitution solution as needed.

[0295] When an oil adjuvant is used as an adjuvant, the pharmaceutical composition of the present invention can be prepared as an emulsion. For example, an emulsion can be prepared by mixing and stirring the peptide solution and the oil adjuvant prepared as described above. The peptide solution can be a solution that is redissolved after freeze drying. The emulsion can be a W / O type emulsion and an O / W type emulsion, and the W / O type emulsion is preferably used to obtain a high immune response enhancement effect. IFA can be preferably used as an oil adjuvant, but is not limited thereto. The preparation of the emulsion can be carried out immediately before administering the subject, and in this case, the pharmaceutical composition of the present invention can be provided as a kit comprising the peptide solution of the present invention and an oily adjuvant. When the pharmaceutical composition of the present invention is a freeze-dried preparation, the kit can further include a redissolved solution.

[0296] Furthermore, the pharmaceutical composition of the present invention may be a liposome preparation in which the peptide of the present invention is encapsulated, a particle formulation in which the peptide is bound to beads having a diameter of several microns, or a formulation in which lipids are bound to the peptide.

[0297] In another embodiment of the present invention, the peptide of the present invention can also be administered in the form of a pharmaceutically acceptable salt. Examples of preferred salts include salts with alkali metals (lithium, potassium, sodium, etc.), salts with alkaline earth metals (calcium, magnesium, etc.), salts with other metals (copper, iron, zinc, manganese, etc.), salts with organic bases, salts with amines, salts with organic acids (e.g., acetic acid, formic acid, propionic acid, fumaric acid, maleic acid, succinic acid, tartaric acid, citric acid, malic acid, oxalic acid, benzoic acid, methanesulfonic acid, etc.), and salts with inorganic acids (e.g., hydrochloric acid, phosphoric acid, hydrobromic acid, sulfuric acid, nitric acid, etc.). The phrase "pharmaceutically acceptable salt" used herein refers to salts that retain the biological, physiological, pharmacological, and pharmaceutical effectiveness and properties of the compound. Therefore, pharmaceutical compositions comprising pharmaceutically acceptable salts of the peptide of the present invention are also included in the present invention. In addition, in addition to free peptides, "peptides of the present invention" also include pharmaceutically acceptable salts thereof.

[0298] In some embodiments, the pharmaceutical composition of the present invention may further include a component that primes CTL. It has been confirmed that lipid is a component that can prime CTL against viral antigens in vivo. For example, palmitic acid residues can be connected to the ε- and α-amino groups of lysine residues and then connected to the peptide of the present invention. The lipidated peptide can then be directly applied in micelles or particles, incorporated into liposomes, or emulsified in adjuvants. As other examples of lipid-induced CTL responses, Escherichia coli lipoproteins, such as tripalmitoyl-S-glyceryl cysteinylseryl-serine (P3CSS), when covalently linked to suitable peptides, can be used to prime CTL (see, for example, Deres et al., Nature 1989, 342:561-4).

[0299] Examples of suitable methods for administering the peptide or pharmaceutical composition of the present invention include oral, intradermal, subcutaneous, intramuscular, intraosseous, intraperitoneal and intravenous injections, as well as systemic administration or local administration near the target site, but are not limited thereto. Preferred methods of administration include subcutaneous injection near lymph nodes such as the axilla or groin. More preferably (for example), when the pharmaceutical composition of the present invention comprises a peptide or exosome as an active ingredient, subcutaneous administration is preferred. Alternatively, a composition having an APC or CTL as an active ingredient may be administered by intravenous injection or the like. Administration may be performed by a single administration or by multiple administrations to enhance administration.

[0300] The peptides of the present invention can be administered to a subject in a therapeutically or pharmaceutically effective amount to treat cancer, or in a therapeutically or pharmaceutically effective amount to induce immunity (more specifically CTL) against cancer cells expressing FOXM1. The dosage of the peptides of the present invention can be appropriately adjusted according to the disease to be treated, the age, weight of the patient, the method of administration, and the like. For each peptide of the present invention, the dosage is generally 0.001 mg-1000 mg, such as 0.01 mg-100 mg, such as 0.1 mg-30 mg, such as 0.1 mg-10 mg, such as 0.5 mg-5 mg. The dosing interval can be once every few days to several months, and, for example, can be administered at intervals of once a week. A person skilled in the art can appropriately select a suitable dose (dose).

[0301] In a preferred embodiment, the pharmaceutical composition of the present invention comprises a therapeutically effective amount of the peptide of the present invention and a pharmaceutically or physiologically acceptable carrier. In another embodiment, the pharmaceutical composition of the present invention comprises a therapeutically effective amount of the peptide of the present invention, a pharmaceutically or physiologically acceptable carrier and an adjuvant. The pharmaceutical composition of the present invention may comprise 0.001mg-1000mg, preferably 0.01mg-100mg, more preferably 0.1mg-30mg, even more preferably 0.1mg-10mg, such as 0.5mg-5mg of the peptide of the present invention. When the pharmaceutical composition of the present invention is an injection, it may comprise the following concentrations of the peptide of the present invention: 0.001mg / ml-1000mg / ml, preferably 0.01mg / ml-100mg / ml, more preferably 0.1mg / ml-30mg / ml, even more preferably 0.1mg / ml-10mg / ml, such as 0.5mg / ml-5mg / ml. In this case, for example, 0.1 to 5ml, preferably 0.5ml to 2ml of the pharmaceutical composition of the present invention may be administered to a subject by injection.

[0302] In addition, the present invention provides a method for treating and / or preventing cancer and / or preventing its postoperative recurrence, which includes administering a therapeutically effective amount of the peptide of the present invention or the pharmaceutical composition of the present invention to a subject. As described above, the peptide of the present invention can be administered to a subject in a single dose generally in the following amounts: 0.001 mg-1000 mg, for example, 0.01 mg-100 mg, for example, 0.1 mg-30 mg, for example, 0.1 mg-10 mg, or for example, 0.5 mg-5 mg. In a preferred embodiment, the peptide of the present invention is administered to a subject together with an adjuvant. In addition, the dosing interval can be once every few days to several months, preferably once every few days to once a month, for example once a week or once every two weeks.

[0303] (2) Pharmaceutical compositions containing polynucleotides as active ingredients

[0304] The pharmaceutical composition of the present invention may also contain a polynucleotide encoding a peptide disclosed in the present invention in an expressible form. In this article, the phrase "in an expressible form" means that the polynucleotide will be expressed as a peptide of the present invention when introduced into a cell. In an exemplary embodiment, the sequence of the polynucleotide of the present invention includes regulatory elements necessary for the expression of the peptide of the present invention. The polynucleotide of the present invention may have a sequence required for stable insertion into the genome of the target cell (for a description of homologous recombination cassette vectors, see, for example, Thomas KR & Capecchi MR, Cell 1987, 51: 503-12). See, for example, Wolff et al., Science 1990, 247: 1465-8; U.S. Patent Nos. 5,580,859, 5,589,466, 5,804,566, 5,739,118, 5,736,524, 5,679,647; and WO98 / 04720. Examples of DNA-based delivery technologies include "naked DNA," facilitated (bupivacaine, polymer, peptide-mediated) delivery, cationic lipid complexes, and particle-mediated ("gene gun") or pressure-mediated delivery (see, e.g., U.S. Pat. No. 5,922,687).

[0305] The peptides of the present invention can also be expressed with viral or bacterial vectors. Examples of expression vectors include attenuated viral hosts, such as vaccinia or fowlpox. For example, vaccinia virus can be used as a vector for expressing the peptides of the present invention. After introduction into the host, the recombinant vaccinia virus expresses the immunogenic peptide, and thereby induces an immune response. Vaccinia vectors and methods that can be used for immunization protocols are described in, for example, U.S. Patent No. 4,722,848. Another vector is BCG (Bacille Calmette Guerin). BCG vectors are described in Stover et al., Nature 1991, 351: 456-60. Various other vectors that can be used for therapeutic administration or immunization are obvious, such as adenovirus and adeno-associated virus vectors, retroviral vectors, Salmonella typhi vectors, detoxified anthrax toxin vectors, etc. See, for example, Shata et al., Mol Med Today 2000, 6:66-71; Shedlock et al., J Leukoc Biol 2000, 68:793-806; Hipp et al., In Vivo 2000, 14:571-85.

[0306] Delivery of the polynucleotides of the invention to a patient can be direct, in which case the patient can be directly exposed to a vector carrying the polynucleotides of the invention, or indirect, in which case cells are first transformed in vitro with a vector carrying the polynucleotides of the invention and then transplanted into the patient. These two approaches are referred to as in vivo and ex vivo gene therapy, respectively.

[0307] For a general review of methods of gene therapy, see Goldspiel et al., Clinical Pharmacy 1993, 12: 488-505; Wu and Wu, Biotherapy 1991, 3: 87-95; Tolstoshev, Ann Rev Pharmacol Toxicol 1993, 33: 573-96; Mulligan, Science 1993, 260: 926-32; Morgan & Anderson, Ann Rev Biochem 1993, 62: 191-217; Trends in Biotechnology 1993, 11 (5): 155-215. Well-known methods in the field of recombinant DNA technology that can also be applied to the present invention are described in Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, NY, 1993; and Krieger, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY, 1990.

[0308] Similar to the administration of peptides, the administration of polynucleotides can be carried out by oral, intradermal, subcutaneous, intravenous, intramuscular, intraosseous, or intraperitoneal injection, etc. The administration of polynucleotides can be systemic administration or local administration near the target site. Administration can be implemented by a single administration, or enhanced by multiple administrations. The polynucleotides of the present invention can be administered to a subject in a therapeutic or pharmaceutically effective amount for inducing immunity (more specifically, CTL) against cancer cells expressing FOXM1, or in a therapeutic or pharmaceutically effective amount for treating cancer. The dose of the polynucleotide in a suitable carrier or the dose of the polynucleotide in a cell transformed with a polynucleotide encoding a peptide of the present invention can be appropriately adjusted according to the disease to be treated, the age, weight, method of administration, etc. of the patient, and is generally 0.001 mg-1000 mg, for example, 0.01 mg-100 mg, for example, 0.1 mg-30 mg, for example, 0.1 mg-10 mg, or for example 0.5 mg-5 mg. The dosing interval can be once every few days to several months, for example, administration can be performed at intervals of once a week. A person skilled in the art can appropriately select a suitable dose (dosage).

[0309] X. Methods Using Peptides, Exosomes, APCs, and CTLs

[0310] The peptides and polynucleotides of the present invention can be used to induce APCs and CTLs. The exosomes and APCs of the present invention can also be used to induce CTLs. The peptides, polynucleotides, exosomes and APCs can be used in combination with any other compound as long as their CTL induction ability is not inhibited. Therefore, a pharmaceutical composition comprising any peptide, polynucleotide, APC and exosome of the present invention can be used to induce the CTL of the present invention. In addition, a pharmaceutical composition comprising the peptide or polynucleotide of the present invention can be used to induce the APC of the present invention.

[0311] (1) Methods for inducing APCs

[0312] The present invention provides a method for inducing APCs having CTL inducibility using the peptides or polynucleotides of the present invention.

[0313] The method of the present invention includes the step of contacting APC with the peptide of the present invention in vitro, ex vivo or in vivo. For example, the method of contacting APC with the peptide in vitro may include the following steps:

[0314] (a) collecting APCs from a subject; and

[0315] (b) contacting the APCs of step (a) with the peptide of the present invention.

[0316] The above-mentioned APC is not limited to a specific type of cell, and cells known to present protein antigens on their cell surfaces to be recognized by lymphocytes, such as DC, Langerhans cells, macrophages, B cells, and activated T cells can be used. DC has the strongest CTL induction ability among APCs, and therefore DC is preferably used. Any peptide of the present invention can be used by itself or in combination with other peptides of the present invention. In addition, the peptide of the present invention can be used in combination with other CTL inducing peptides (e.g., CTL inducing peptides derived from other TAAs).

[0317] Meanwhile, when the peptide of the present invention is administered to a subject, APCs come into contact with the peptide in vivo, and as a result, APCs with high CTL inducibility are induced in the body of the subject. Therefore, the method of the present invention may include the step of administering the peptide of the present invention to a subject. Similarly, when the polynucleotide of the present invention in an expressible form is administered to a subject, the peptide of the present invention is expressed in vivo, and the expressed peptide comes into contact with APCs in vivo, and as a result, APCs with high CTL inducibility are induced in the body of the subject. Therefore, the present invention may also include the step of administering the polynucleotide of the present invention to a subject.

[0318] In order to induce APCs with CTL inducibility, the present invention also includes the step of introducing the polynucleotide of the present invention into APCs. For example, the method may include the following steps:

[0319] (a) collecting APCs from a subject; and

[0320] (b) introducing a polynucleotide encoding the peptide of the present invention into the APCs of step (a).

[0321] Step (b) can be performed as described above in the section "VI. Antigen presenting cells (APC)".

[0322] Therefore, in one embodiment, the present invention provides a method for inducing APCs with CTL inducibility, comprising the following steps (a) or (b):

[0323] (a) contacting APC with the peptide of the present invention; and

[0324] (b) A polynucleotide encoding the peptide of the present invention is introduced into APCs.

[0325] In addition, the present invention provides a method for preparing APCs having CTL inducibility, comprising the following steps (a) or (b):

[0326] (a) contacting APC with the peptide of the present invention; or

[0327] (b) A polynucleotide encoding the peptide of the present invention is introduced into APCs.

[0328] The above methods can be implemented in vitro, ex vivo or in vivo, and preferably they are implemented in vitro or ex vivo. The APCs used in the above methods can be derived from the subject to which the induced APCs are planned to be administered, or they can be derived from different subjects. When using APCs from a subject (donor) different from the subject planned to be administered, the administered subject and the donor must have the same HLA type.

[0329] In the method of the present invention, when a peptide having an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46 or a modified peptide thereof is used as the peptide of the present invention, HLA in both the administration subject and the donor is preferably HLA-A33 (more preferably HLA-A*33:03). Alternatively, the APC used in the above method is preferably an APC expressing HLA-A33 (more preferably HLA-A*33:03).

[0330] Similarly, when a peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 or a modified peptide thereof is used as the peptide of the present invention, HLA in the administration subject and the donor is preferably HLA-A01 (more preferably HLA-A*01:01). Alternatively, the APC used in the above method is preferably an APC expressing HLA-A01 (more preferably HLA-A*01:01). After separating PBMC from blood collected from a donor by specific gravity centrifugation or the like, APC can be prepared from PBMC using a known method.

[0331] In another embodiment, the present invention also provides a pharmaceutical composition comprising the peptide of the present invention or a polynucleotide encoding the peptide, which is used for inducing APCs having CTL inducibility.

[0332] Alternatively, the present invention also provides use of the peptide of the present invention or a polynucleotide encoding the peptide in the preparation of a pharmaceutical composition for inducing APCs with CTL inducibility.

[0333] Alternatively, the present invention also provides the peptide of the present invention or a polynucleotide encoding the peptide for use in inducing APCs having CTL inducibility.

[0334] Alternatively, the present invention further provides a method or process for preparing a pharmaceutical composition for inducing APCs, wherein the method or process comprises the step of formulating the peptide of the present invention or the polynucleotide of the present invention with a pharmaceutically or physiologically acceptable carrier.

[0335] In another embodiment, the present invention further provides a method or process for preparing a pharmaceutical composition for inducing APCs with CTL inducibility, wherein the method or process comprises the step of mixing the peptide of the present invention or the polynucleotide of the present invention with a pharmaceutically or physiologically acceptable carrier.

[0336] APCs induced by the method of the present invention can induce CTLs specific for FOXM1 (ie, CTLs of the present invention).

[0337] (2) Methods of inducing CTL

[0338] The present invention also provides a method for inducing CTL using the peptides, polynucleotides, exosomes or APCs of the present invention. The present invention also provides a method for inducing CTL using one or more polynucleotides encoding polypeptides capable of forming a T cell receptor (TCR) (i.e., TCR subunits), wherein the TCR is capable of recognizing a complex of the peptides of the present invention and HLA antigens. Preferably, the method for inducing CTL comprises at least one step selected from the following:

[0339] (a) contacting CD8-positive T cells with antigen-presenting cells presenting a complex of an HLA antigen and the peptide of the present invention on its surface;

[0340] (b) contacting CD8-positive T cells with exosomes presenting a complex of an HLA antigen and the peptide of the present invention on its surface; and

[0341] (c) introducing one or more polynucleotides encoding polypeptides capable of forming TCR into CD8-positive T cells, wherein the TCR is capable of recognizing a complex of the peptide of the present invention and an HLA antigen.

[0342] When the peptide, polynucleotide, APC or exosome of the present invention is administered to a subject, CTL is induced in the subject's body, and the strength of the immune response targeting cancer cells expressing FOXM1 is enhanced. Therefore, the method of the present invention may include the step of administering the peptide, polynucleotide, APC or exosome of the present invention to a subject.

[0343] Alternatively, they can also be used in vitro or ex vivo to induce CTL. For example, the method of the present invention can include the following steps:

[0344] (a) collecting APCs from a subject;

[0345] (b) contacting the APC of step (a) with a peptide of the present invention; and

[0346] (c) co-culturing the APCs from step (b) with CD8-positive T cells.

[0347] The induced CTLs can then be returned to the subject.

[0348] The APCs to be co-cultured with CD8-positive T cells in step (c) above can also be prepared by introducing a polynucleotide encoding the peptide of the present invention into the APCs, as described in the section "VI. Antigen presenting cells (APCs)" above. However, the APCs used in the method of the present invention are not limited thereto, and any APCs that present a complex of an HLA antigen and the peptide of the present invention on its surface can be used.

[0349] In the method of the present invention, in addition to such APCs, exosomes presenting a complex formed by an HLA antigen and the peptide of the present invention on its surface may also be used. That is, the method of the present invention may include a step of co-culturing with exosomes presenting a complex formed by an HLA antigen and the peptide of the present invention on its surface. Such exosomes can be prepared by the method described in the above section "V. Exosomes".

[0350] In addition, CTL can be induced by introducing a vector comprising a polynucleotide encoding each subunit of TCR into CD8 positive T cells, and the TCR is capable of binding to the peptide of the present invention presented by the HLA antigen on the cell surface. Such conversion can be implemented as described in the "VIII. T cell receptor (TCR)" section above.

[0351] Therefore, in one embodiment, the present invention provides a method for inducing CTLs, comprising a step selected from the following:

[0352] (a) co-culturing CD8-positive T cells with APCs, wherein the APCs present a complex of an HLA antigen and the peptide of the present invention on their surface;

[0353] (b) co-culturing CD8-positive T cells with exosomes presenting a complex of an HLA antigen and the peptide of the present invention on its surface; and

[0354] (c) introducing into CD8-positive T cells a vector comprising a polynucleotide encoding each subunit of TCR capable of binding to the peptide of the present invention presented by the HLA antigen on the cell surface.

[0355] The above method can be implemented in vitro, in vitro or in vivo, and is preferably implemented in vitro or in vitro. The APCs or exosomes and CD8 positive T cells used in the above method can be derived from the subject of the CTL induced by the plan, or they can be derived from different subjects. When using APCs or exosomes and CD8 positive T cells from a subject (donor) different from the subject to be administered, the subject and the donor must have the same HLA type. For example, when there is a peptide with SEQ ID NOs:1,2,3,6,7,11,12,17,18,20,22,24,26,32,33,36,39,41,42,45 and 46 amino acid sequence or its modified peptide is used as the peptide of the present invention, in the subject and the donor, HLA type is preferably HLA-A33 (more preferably HLA-A*33:03). Alternatively, the APC or exosome used in the above method is preferably an APC or exosome that presents a complex of HLA-A33 (more preferably HLA-A*33:03) and the peptide of the present invention (peptides having the amino acid sequences of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46 or modified peptides thereof) on its surface. In this case, the induced CTL exhibits cytotoxic activity against cells presenting a complex of HLA-A33 and the peptide of the present invention (e.g., HLA-A33-positive cells expressing FOXM1).

[0356] Alternatively, for example, when a peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 or a modified peptide thereof is used as the peptide of the present invention, the HLA type in both the subject and the donor to be administered is preferably HLA-A01 (more preferably HLA-A*01:01). Alternatively, the APCs and exosomes used in the above method are preferably APCs and exosomes that present a complex of HLA-A01 (more preferably HLA-A*01:01) and the peptide of the present invention (a peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61 or a modified peptide thereof) on their surface. In this case, the induced CTLs show cytotoxic activity against cells presenting a complex of HLA-A01 and the peptide of the present invention (eg, HLA-A01-positive cells expressing FOXM1).

[0357] In another embodiment, the present invention also provides a composition or pharmaceutical composition for inducing CTL, comprising at least one active ingredient selected from the following:

[0358] (a) a peptide of the present invention;

[0359] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0360] (c) APCs presenting the peptide of the present invention on their surface; and

[0361] (d) Exosomes presenting the peptides of the present invention on their surface.

[0362] In another embodiment, the present invention also provides use of an active ingredient selected from the following in the preparation of a composition or pharmaceutical composition for inducing CTL:

[0363] (a) a peptide of the present invention;

[0364] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0365] (c) APCs presenting the peptide of the present invention on their surface; and

[0366] (d) Exosomes presenting the peptides of the present invention on their surface.

[0367] Alternatively, the present invention further provides an active ingredient selected from the following for inducing CTL:

[0368] (a) a peptide of the present invention;

[0369] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0370] (c) APCs presenting the peptide of the present invention on their surface; and

[0371] (d) Exosomes presenting the peptides of the present invention on their surface.

[0372] Alternatively, the present invention further provides a method or process for preparing a composition or pharmaceutical composition for inducing CTL, which is a method or process comprising the steps of formulating an active ingredient selected from the following with a pharmaceutically or physiologically acceptable carrier:

[0373] (a) a peptide of the present invention;

[0374] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0375] (c) APCs presenting the peptide of the present invention on their surface; and

[0376] (d) Exosomes presenting the peptides of the present invention on their surface.

[0377] In another embodiment, the present invention further provides a method or process for preparing a composition or pharmaceutical composition for inducing CTL, which comprises the step of mixing an active ingredient selected from the following with a pharmaceutically or physiologically acceptable carrier:

[0378] (a) a peptide of the present invention;

[0379] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0380] (c) APCs presenting the peptide of the present invention on their surface; and

[0381] (d) Exosomes presenting the peptides of the present invention on their surface.

[0382] XI. Methods of Inducing an Immune Response

[0383] The present invention also provides a method for inducing an immune response against a cancer expressing FOXM1. Applicable cancers include acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc., but are not limited thereto. Preferably, the cancer expresses at least one HLA selected from HLA-A33 and HLA-A01.

[0384] The present invention also provides a method for inducing an immune response against cancer cells expressing FOXM1. FOXM1 is believed to be overexpressed in the various types of cancers described above. Therefore, when an immune response is induced against cancer cells expressing FOXM1, the proliferation of cancer cells is inhibited as a result. Therefore, the present invention also provides a method for inhibiting the proliferation of cancer cells expressing FOXM1. The method of the present invention is particularly suitable for inhibiting the proliferation of cancer cells expressing FOXM1 and at least one HLA selected from HLA-A33 and HLA-A01.

[0385] The method of the present invention may include the step of administering a composition comprising any peptide of the present invention or a polynucleotide encoding the peptide. The method of the present invention also encompasses administering an APC or exosome presenting any peptide of the present invention. For details, see the "IX. Pharmaceutical Composition" section, particularly the section describing the use of the pharmaceutical composition of the present invention as a vaccine. In addition, exosomes and APCs that can be used in the method of inducing an immune response of the present invention are described in detail in the above "V. Exosomes", "VI. Antigen Presenting Cells (APCs)", and "X. Methods Using Peptides, Exosomes, APCs and CTLs" Items (1) and (2).

[0386] In another embodiment, the present invention provides a pharmaceutical composition or vaccine for inducing an immune response against a cancer expressing FOXM1, wherein the pharmaceutical composition or vaccine comprises an active ingredient selected from the group consisting of:

[0387] (a) a peptide of the present invention;

[0388] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0389] (c) APCs presenting the peptides of the present invention on their surface;

[0390] (d) an exosome presenting a peptide of the present invention on its surface; and

[0391] (e) CTL of the present invention.

[0392] Alternatively, the present invention also provides a pharmaceutical composition or vaccine for inducing an immune response against cancer cells expressing FOXM1, wherein the pharmaceutical composition or vaccine comprises an active ingredient selected from the following:

[0393] (a) a peptide of the present invention;

[0394] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0395] (c) APCs presenting the peptides of the present invention on their surface;

[0396] (d) an exosome presenting a peptide of the present invention on its surface; and

[0397] (e) CTL of the present invention.

[0398] Alternatively, the present invention also provides a pharmaceutical composition or vaccine for inhibiting the proliferation of cancer cells expressing FOXM1, wherein the pharmaceutical composition or vaccine comprises an active ingredient selected from the following:

[0399] (a) a peptide of the present invention;

[0400] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0401] (c) APCs presenting the peptides of the present invention on their surface;

[0402] (d) an exosome presenting a peptide of the present invention on its surface; and

[0403] (e) CTL of the present invention.

[0404] In another embodiment, the present invention provides the use of an active ingredient selected from the following in the preparation of a pharmaceutical composition or vaccine for inducing an immune response against a cancer expressing FOXM1:

[0405] (a) a peptide of the present invention;

[0406] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0407] (c) APCs presenting the peptides of the present invention on their surface;

[0408] (d) an exosome presenting a peptide of the present invention on its surface; and

[0409] (e) CTL of the present invention.

[0410] Alternatively, the present invention also provides use of an active ingredient selected from the following in the preparation of a pharmaceutical composition or vaccine for inducing an immune response against cancer cells expressing FOXM1:

[0411] (a) a peptide of the present invention;

[0412] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0413] (c) APCs presenting the peptides of the present invention on their surface;

[0414] (d) an exosome presenting a peptide of the present invention on its surface; and

[0415] (e) CTL of the present invention.

[0416] Alternatively, the present invention further provides use of an active ingredient selected from the following in the preparation of a pharmaceutical composition or vaccine for inhibiting the proliferation of cancer cells expressing FOXM1:

[0417] (a) a peptide of the present invention;

[0418] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0419] (c) APCs presenting the peptides of the present invention on their surface;

[0420] (d) an exosome presenting a peptide of the present invention on its surface; and

[0421] (e) CTL of the present invention.

[0422] The present invention also provides a method or process for preparing a pharmaceutical composition for inducing an immune response against cancer expressing FOXM1, wherein the method may include the step of mixing or formulating the peptide or polynucleotide of the present invention with a pharmaceutically acceptable carrier.

[0423] Alternatively, the present invention provides a method for inhibiting proliferation of cancer cells expressing FOXM1 or a method for inducing an immune response against cancer, comprising the step of administering to a subject a vaccine or a pharmaceutical composition comprising an active ingredient selected from the following:

[0424] (a) a peptide of the present invention;

[0425] (b) a polynucleotide encoding the peptide of the present invention in an expressible form;

[0426] (c) APCs presenting the peptides of the present invention on their surface;

[0427] (d) an exosome presenting a peptide of the present invention on its surface; and

[0428] (e) CTL of the present invention.

[0429] In the context of the present invention, cancers expressing FOXM1 can be treated by administering the peptides, polynucleotides, APCs, exosomes and / or CTLs of the present invention. Alternatively, an immune response to cancers expressing FOXM1 can be induced by administering the peptides, polynucleotides, APCs, exosomes and / or CTLs of the present invention. Examples of such cancers include acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc., but are not limited thereto. In addition, an immune response to cancers expressing FOXM1 can be induced by administering the peptides, polynucleotides, APCs, exosomes and / or CTLs of the present invention. Therefore, before administering a vaccine or pharmaceutical composition comprising the above-mentioned active ingredient, it is preferably confirmed whether the expression level of FOXM1 in the diseased part of the subject to be treated is increased.

[0430] Thus, in one embodiment, the present invention provides a method of treating a cancer expressing FOXM1 in a patient in need of cancer treatment, wherein the method comprises the steps of:

[0431] (i) measuring the level of FOXM1 expression in a biological sample collected from a diseased site of a subject suffering from cancer;

[0432] (ii) identifying a subject having a cancer expressing FOXM1 based on the level of FOXM1 expression measured in (i); and

[0433] (iii) administering at least one component selected from the group consisting of (a) to (e) above to a patient suffering from cancer that overexpresses FOXM1 compared to a normal control.

[0434] Alternatively, the present invention further provides a vaccine and a pharmaceutical composition comprising at least one active ingredient selected from the group consisting of (a) to (e) above for administration to a cancer patient expressing FOXM1. The present invention also provides a method for identifying or selecting a subject to be treated with at least one active ingredient selected from (a) to (e) above, wherein the method comprises the following steps:

[0435] (i) measuring the level of FOXM1 expression in a biological sample collected from a diseased site of a subject suffering from cancer;

[0436] (ii) identifying a subject having a cancer expressing FOXM1 based on the level of FOXM1 expression measured in (i); and

[0437] (iii) identifying or selecting the subject identified in (ii) as a subject that can be treated with at least one active ingredient selected from the group consisting of (a) to (e).

[0438] The biological sample collected from the subject for measuring the expression level of FOXM1 in the above method is not particularly limited, and for example, a tissue sample containing cancer cells collected by biopsy or the like can be preferably used. The expression level of FOXM1 in the biological sample can be measured by a known method, for example, a method for detecting the transcription product of the FOXM1 gene by a probe or PCR method (e.g., cDNA microarray method, Northern blotting method, RT-PCR method, etc.), a method for detecting the translation product of the FOXM1 gene by an antibody or the like (e.g., Western blotting method, immunostaining method, etc.), and the like can be used. In addition, the biological sample may be a blood sample, and in this case, the blood level of an antibody against FOXM1 or a fragment thereof is measured, and the expression level of FOXM1 in the diseased site can be evaluated based on the blood level. The blood level of an antibody against FOXM1 can be measured using a known method, for example, an enzyme immunoassay (EIA), an enzyme-linked immunosorbent assay (ELISA), a radioimmunoassay (RIA), etc. using the FOXM1 protein or the peptide of the present invention as an antigen can be used.

[0439] Generally, in tissues and cells that do not express FOXM1, FOXM1 transcription products and translation products are hardly detected. Therefore, when FOXM1 transcription products or translation products are detected in cancer cells or tissue samples containing cancer cells collected from a subject, it can be determined that the subject's cancer expresses FOXM1. In blood samples of subjects with cancer that does not express FOXM1, antibodies against FOXM1 or fragments thereof are hardly detected. Therefore, when antibodies against FOXM1 or fragments thereof are detected in blood samples collected from a subject, it can be determined that the subject's cancer expresses FOXM1.

[0440] Whether the subject's cancer expresses FOXM1 can also be compared with the measurement results of the same type of biological material collected from a non-cancer site of the subject or compared with the same type of biological material collected from a subject who does not have cancer (normal control sample). That is, when the level in the biological sample of the test subject is increased in comparison with the measured target level in the normal control sample (normal control level), the subject's cancer is assessed to express FOXM1. For example, when the amount of the measured object detected increases by at least 10% or more compared to the normal control level, the subject's cancer can be assessed to express FOXM1. It is expected that the amount of the measured target detected is preferably 25% or more, more preferably 50% or more, higher than the normal control level. In addition, the amount of the transcription product or translation product of FOXM1 detected can be evaluated by normalization relative to the amount of a known housekeeping gene detected (e.g., β-actin, glyceraldehyde-3-phosphate dehydrogenase, or ribosomal protein P1).

[0441] In a preferred embodiment, it is preferred to confirm the HLA type of the subject before administering at least one active ingredient selected from the group consisting of (a) to (e) above. For example, for a subject to be administered with an active ingredient related to a peptide having an amino acid sequence selected from SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46, it is preferred to select an HLA-A33 positive subject. For a subject to be administered with an active ingredient related to a peptide having an amino acid sequence selected from SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61, it is preferred to select an HLA-A01 positive subject.

[0442] The present invention also provides a complex of the peptide of the present invention and HLA. The complex of the present invention can be a monomer or a polymer. When the complex of the present invention is a polymer, the number of polymerization is not particularly limited, and it can be a polymer of any polymerization number. Examples include, but are not limited to, tetramers, pentamers, hexamers, etc. The multimers of the present invention also include dextramers (WO2002 / 072631) and streptamers (Knabel M et al., Nat Med. 2002 Jun; 8 (6): 631-7.). The complex of the peptide of the present invention and HLA can be prepared according to known methods (e.g., Altman JD et al., Science. 1996, 274 (5284): 94-6; WO2002 / 072631; WO2009 / 003492; Knabel M, Nat Med. 2002 Jun; 8 (6): 631-7, etc.).

[0443] For example, the complex of the present invention can be used to quantify CTLs specific for the peptide of the present invention. For example, a blood sample is collected from a subject to whom the pharmaceutical composition of the present invention is administered, and after isolating PBMCs, CD4 negative cells are prepared and contacted with the fluorescent dye-conjugated complex of the present invention. Then, the percentage of CTLs specific for the peptide of the present invention can be measured by flow cytometry analysis. For example, the immune response induction effect of the pharmaceutical composition of the present invention can be monitored by measuring the specific CTLs for the peptide of the present invention before, during and / or after the administration of the pharmaceutical composition of the present invention.

[0444] XII. Antibodies

[0445] The present invention further provides antibodies that bind to the peptides of the present invention. Preferred antibodies specifically bind to the peptides of the present invention and will not bind (or will weakly bind) to peptides that are not of the present invention. In another embodiment, such antibodies may include antibodies that recognize peptides in the context of HLA molecules, i.e., antibodies that bind to peptide-MHC complexes. The binding specificity of the antibody can be confirmed by inhibition assays. That is, if the binding between the antibody to be analyzed and the full-length FOXM1 polypeptide is inhibited in the presence of the peptides of the present invention, the antibody shows specific binding to the peptides of the present invention. Antibodies against the peptides of the present invention can be used for determination of disease diagnosis and prognosis, as well as for selecting subjects to administer the pharmaceutical compositions of the present invention and monitoring the pharmaceutical compositions of the present invention.

[0446] The present invention also provides various immunoassays for detecting and / or quantifying the peptides of the present invention or fragments thereof. Such immunoassays include, but are not limited to, radioimmunoassays, immunochromatography, enzyme-linked immunosorbent assays (ELISA), enzyme-linked immunofluorescence assays (ELIFA), etc., and are performed within the scope of various immunoassay formats well known in the art.

[0447] The antibodies of the present invention can be used in immunological imaging methods that can detect cancers expressing FOXM1, and examples thereof include, but are not limited to, radioscintigraphic imaging using the labeled antibodies of the present invention. Such assays are clinically used to detect, monitor, and prognose cancers expressing FOXM1; and examples of such cancers include acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc., but are not limited thereto.

[0448] The antibodies of the present invention may be used in any arbitrary form, such as monoclonal or polyclonal antibodies, and may also include antisera obtained by immunizing animals (such as rabbits) with the peptides of the present invention, all kinds of polyclonal and monoclonal antibodies, human antibodies, and chimeric antibodies and humanized antibodies produced by genetic recombination.

[0449] The peptide of the present invention or a fragment thereof used as an antigen for obtaining an antibody can be obtained by chemical synthesis or genetic engineering technology based on the amino acid sequence disclosed herein.

[0450] The peptide used as an immunizing antigen may be a peptide of the present invention or a fragment of a peptide of the present invention. In addition, the peptide may be bound to a carrier or conjugated to a carrier to increase immunogenicity. Keyhole limpet hemocyanin (KLH) is well known as a carrier. Methods for binding KLH to a peptide are also well known in the art.

[0451] Any mammal can be immunized with the above antigens, and when producing monoclonal antibodies, compatibility with the parental cells used for cell fusion is preferably considered. Generally, animals of the order Rodentia, Lagomorpha or Primate can be used. Rodents include, for example, mice, rats and hamsters. Lagomorpha include, for example, rabbits. Primates include, for example, pygmy monkeys (old world monkeys) such as Macaca fascicularis, rhesus monkeys, baboons and chimpanzees.

[0452] Methods for immunizing animals with antigens are known in the art. Intraperitoneal and subcutaneous injections of antigens are standard methods for immunizing mammals. More specifically, the antigen can be diluted and suspended in an appropriate amount of phosphate buffered saline (PBS), saline, etc. If necessary, the antigen suspension can be mixed with an appropriate amount of standard adjuvant such as Freund's complete adjuvant, made into an emulsion, and then applied to mammals. Then, it is preferred to use an antigen mixed with an appropriate amount of Freund's incomplete adjuvant several times every 4 to 21 days. Suitable carriers can be used for immunization. After immunization as described above, the increase in the amount of the desired antibody in the serum can be tested by standard methods.

[0453] Polyclonal antibodies to the peptides of the present invention can be prepared as follows: blood is collected from a mammal (in which an increase in the serum level of the desired antibody has been confirmed after immunization) and serum is separated from the blood by any conventional method. The polyclonal antibody can be a serum containing the polyclonal antibody, or a fraction containing the polyclonal antibody that can be separated from the serum. Immunoglobulin G or M can be prepared from a fraction that only recognizes the peptides of the present invention using, for example, an affinity column coupled with the peptides of the present invention, and the fraction is further purified using a protein A or protein G column.

[0454] In order to prepare monoclonal antibodies, when the serum level of the desired antibody is confirmed to increase after immunization, immune cells are collected from mammals and cell fusion is performed. The immune cells used for cell fusion can preferably be obtained from the spleen. For other parent cells fused with the above-mentioned immune cells, for example, mammalian myeloma cells can be used, and more preferably myeloma cells that have obtained the drug selection properties of the fused cells.

[0455] The above immune cells can be fused with myeloma cells according to known methods, for example, the method of Milstein et al. (Galfre and Milstein, Methods Enzymol, 1981, 73: 3-46).

[0456] For the hybridoma obtained by cell fusion, it can be selected by cultivating in standard selection medium such as HAT medium (medium containing hypoxanthine, aminopterin and thymidine). Usually in HAT medium, continue cell culture enough time (for example, several days to several weeks), to allow all other cells (non-fused cells) except required hybridoma to die. Then, by standard limiting dilution (standard limiting dilution), screen and clone the hybridoma cell of required antibody.

[0457] In addition to the above-mentioned method of preparing hybridomas by antigen immunization of non-human animals, it is also possible to immunize human lymphocytes, such as those infected with Epstein-Barr virus, with peptides, cells expressing peptides or their lysates in vitro. Then, immune lymphocytes can be fused with immortalized myeloma cells derived from humans such as U266 to produce hybridomas (JPS63-17688) that generate human antibodies that can be combined with the peptides.

[0458] The obtained hybridoma is then transplanted into the peritoneal cavity of mice, and the ascites is extracted. The obtained monoclonal antibody can be purified by, for example, ammonium sulfate precipitation, protein A or protein G column, DEAE ion exchange chromatography, or affinity column coupled with the peptide of the present invention.

[0459] Alternatively, antibody producing immune cells (eg, immune lymphocytes) can be immortalized by oncogenes and used to prepare monoclonal antibodies.

[0460] The monoclonal antibodies thus obtained can also be recombinantly prepared using genetic engineering techniques (see, for example, Borrebaeck and Larrick, Therapeutic Monoclonal Antibodies, published in the UK by MacMillan Publishers LTD (1990)). For example, DNA encoding antibodies can be cloned from immune cells such as hybridomas or immune lymphocytes, the DNA can be inserted into a suitable vector, and introduced into a host cell to prepare recombinant antibodies. The present invention also provides recombinant antibodies prepared as described above.

[0461] In addition, the antibody of the present invention may be an antibody fragment or a modified antibody, as long as it binds to the peptide of the present invention. For example, it is desirable that the antibody fragment contains the antigen binding site of the antibody. Specifically, the antibody fragment may be Fab, F(ab')2, Fv, or a single-chain Fv (scFv) (Huston et al., Proc Natl Acad Sci USA, 1988, 85: 5879-83) formed by connecting Fv fragments from H chain and L chain through a suitable linker. More specifically, antibody fragments may be generated by treating the antibody with an enzyme such as papain or pepsin. Alternatively, a gene encoding an antibody fragment can be constructed, inserted into an expression vector, and expressed in an appropriate host cell (see, e.g., Co et al., J Immunol, 1994, 152:2968-76; Better and Horwitz, Methods Enzymol, 1989, 178:476-96; Pluckthun and Skerra, Methods Enzymol, 1989, 178:497-515; Lamoyi, Methods Enzymol, 1986, 121:652-63; Rousseaux et al., Methods Enzymol, 1986, 121:663-9; Bird and Walker, Trends Biotechnol, 1991, 9:132-7)).

[0462] Antibodies can be modified by conjugation with various molecules such as polyethylene glycol (PEG). The present invention provides antibodies that have undergone such modifications. Modified antibodies can be obtained by chemically modifying antibodies. These modification methods are routine in the art.

[0463] Alternatively, the antibodies of the present invention can be obtained in the form of chimeric antibodies between the variable regions derived from non-human antibodies and the constant regions derived from human antibodies, or humanized antibodies comprising complementary determining regions (CDRs) derived from non-human antibodies, framework regions (FRs) derived from human antibodies, and constant regions. Such antibodies can be prepared according to known techniques. Humanization can be performed by replacing the corresponding sequences of human antibodies with non-human antibody CDR sequences (see, e.g., Verhoeyen et al., Science, 1988, 239: 1534-6). Thus, such humanized antibodies are chimeric antibodies in which a small portion of the human variable domain has been replaced by the corresponding sequence from a non-human species.

[0464] Completely human antibodies can also be used, such antibodies also contain human variable regions in addition to human framework regions and constant regions. Such antibodies can be prepared using a variety of techniques known in the art. For example, in vitro methods include the use of recombinant libraries of human antibody fragments displayed on phage (e.g., Hoogenboom & Winter, J. Mol. Biol., 1991, 227: 381). Similarly, human antibodies can be generated by introducing human immunoglobulin loci into transgenic animals, such as mice in which endogenous immunoglobulin genes are partially or completely inactivated. This method is described in, for example, U.S. Patent Nos. 6,150,584; 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425 and 5,661,016.

[0465] The antibody obtained as above can be purified to homogeneity. For example, the separation and purification of the antibody can be carried out according to the separation and purification method for general proteins. For example, the antibody can be separated and purified by appropriately selecting and combining column chromatography such as affinity chromatography, filtration, ultrafiltration, salting out, dialysis, SDS polyacrylamide gel electrophoresis and isoelectric focusing (Antibodies: A Laboratory Manual. Ed Harlow and David Lane, Cold Spring Harbor Laboratory (1988)), but is not limited thereto. Protein A columns and protein G columns can be used as affinity columns. Exemplary usable protein A columns include, for example, Hyper D, POROS and Sepharose FF (Pharmacia).

[0466] In addition to affinity chromatography, exemplary chromatography includes, for example, ion exchange chromatography, hydrophobic chromatography, gel filtration, reverse phase chromatography, adsorption chromatography, and the like (Strategies for Protein Purification and Characterization: A Laboratory Course Manual. Ed Daniel R. Marshak et al., Cold Spring Harbor Laboratory Press (1996)). Chromatographic procedures can be performed by liquid chromatography, such as HPLC and FPLC.

[0467] For example, the antigen-binding activity of the antibody of the present invention can be measured by using absorbance measurement, enzyme-linked immunosorbent assay (ELISA), enzyme immunoassay (EIA), radioimmunoassay (RIA) and / or immunofluorescence (IF). In ELISA, the antibody of the present invention is immobilized on a plate, the peptide of the present invention is applied to the plate, and then a sample containing the desired antibody, such as a culture supernatant of a cell that generates the antibody or a purified antibody, is applied. Then a second antibody labeled with an enzyme (such as alkaline phosphatase) that recognizes the first antibody is applied, and the plate is incubated. Then, after washing, an enzyme substrate, such as p-nitrophenyl phosphate, is added to the plate, and the antigen-binding activity of the sample is evaluated by measuring absorbance. In order to evaluate the binding activity of the antibody, a peptide fragment such as a C-terminal or N-terminal fragment can be used as an antigen. BIAcore (Pharmacia) can be used to evaluate the activity of the antibody of the present invention.

[0468] The peptide of the present invention using the above method can be detected or measured by exposing the antibody of the present invention to a sample assumed to contain the peptide of the present invention, and detecting or measuring the immune complex formed between the antibody and the peptide.

[0469] For example, the antibodies of the present invention can be used to detect the peptides of the present invention present in a blood sample (e.g., a serum sample) of a subject. Alternatively, the peptides of the present invention can also be used to detect the antibodies of the present invention present in a blood sample (e.g., a serum sample) of a subject. The results of measuring the peptides of the present invention or the antibodies of the present invention in a blood sample of a subject can be used to select a subject for administering a pharmaceutical composition of the present invention or to monitor the efficacy of a pharmaceutical composition. In addition, it has been reported that patients with antibodies (which are directed to peptides administered as vaccines) may have high responsiveness to vaccines. Therefore, when the peptides of the present invention as a vaccine are administered using the patient's antibodies as an index, the peptides of the present invention can be used as immunoassay antigens for selecting patients with high responsiveness.

[0470] XII. Vectors and Host Cells

[0471] The present invention provides a vector comprising a polynucleotide encoding a peptide of the present invention and a host cell into which the vector is introduced. The vector of the present invention can be used to maintain the polynucleotide of the present invention in a host cell, to express the peptide of the present invention in the host cell, or to administer the polynucleotide of the present invention for gene therapy.

[0472] When E. coli is a host cell and a vector is amplified and generated in large quantities in E. coli (e.g., JM109, DH5-alpha, HB101, or XL1-Blue), the vector needs to have a "replication origin" for amplification in E. coli and a marker gene for selecting transformed E. coli (e.g., a drug resistance gene selected by drugs such as ampicillin, tetracycline, kanamycin, chloramphenicol, etc.). For example, M13 series vectors, pUC series vectors, pBR322, pBluescript, pCR-Script, etc. can be used. In addition, pGEM-T, pDIRECT, and pT7 can be used for cloning as well as the above vectors. When the vector is used to produce the peptide of the present invention, an expression vector can be used. For example, an expression vector to be expressed in E. coli should have the above-mentioned characteristics of amplification in E. coli. When using E. coli such as JM109, DH5-alpha, HB101 or XL1-Blue as a host cell, the vector should have a promoter that can efficiently express the desired gene in E. coli, such as lacZ promoter (Ward et al., Nature 341: 544-6 (1989); FASEB J 6: 2422-7 (1989)), araB promoter (Better et al., Science 240: 1041-3 (1988)), T7 promoter, etc. In this regard, for example, pGEX-5X-1 (Pharmacia), "QIAexpress system" (Qiagen), pEGFP and pET (in this case, the host is preferably BL21 expressing T7 RNA polymerase) can be used to replace the above vectors. In addition, the vector may also contain a signal sequence for peptide secretion. An exemplary signal sequence directing secretion of a peptide to the periplasm of E. coli is the pelB signal sequence (Lei et al., J Bacteriol, 1987, 169:4379). Means for introducing a vector into a target host cell include, for example, the calcium chloride method and electroporation.

[0473] In addition to E. coli, the polypeptides of the present invention can also be produced using, for example, expression vectors derived from mammals (e.g., pcDNA3 (Invitrogen) and pEGF-BOS (Nucleic Acids Res 18(17):5322(1990)), pEF, pCDM8), expression vectors derived from insect cells (e.g., "Bac-to-BAC baculovirus expression system" (GIBCO BRL), pBacPAK8), expression vectors derived from plants (e.g., pMH1, pMH2), expression vectors derived from animal viruses (e.g., pHSV, pMV, pAdexLcw), expression vectors derived from retroviruses (e.g., pZIpneo), expression vectors derived from yeast (e.g., "Pichia expression kit" (Invitrogen), pNV11, SP-Q01), and expression vectors derived from Bacillus subtilis (e.g., pPL608, pKTH50).

[0474] In order to express the vector in animal cells such as CHO, COS or NIH3T3 cells, the vector should have a promoter necessary for expression in the cells, such as SV40 promoter (Mulligan et al., Nature, 1979, 277: 108), MMLV-LTR promoter, EF1-alpha promoter (Mizushima et al., Nucleic Acids Res, 1990, 18: 5322), CMV promoter, etc., and preferably a marker gene for selecting transformants (e.g., a drug resistance gene selected by drugs (e.g., neomycin, G418)). Examples of known vectors having these characteristics include, for example, pMAM, pDR2, pBK-RSV, pBK-CMV, pOPRSV and pOP13.

[0475] Embodiments of the present invention are exemplified below based on the above description; however, the present invention is not limited to these embodiments.

[0476] [1] A peptide of less than 15 amino acids having cytotoxic T cell (CTL) inducibility, comprising an amino acid sequence selected from the following group:

[0477] (a) an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60, and 61; and

[0478] (b) an amino acid sequence in which 1, 2 or several amino acids are substituted, deleted, inserted and / or added in an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61.

[0479] [2] The peptide of [1] is selected from the following groups (i) to (ii):

[0480] (i) a peptide comprising an amino acid sequence comprising one or more substitutions selected from the following groups (a) to (c) introduced into an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46:

[0481] (a) replacing the first amino acid from the N-terminus with an amino acid selected from the group consisting of aspartic acid and glutamic acid;

[0482] (b) substituting the second amino acid from the N-terminus with an amino acid selected from the group consisting of phenylalanine, tyrosine, alanine, isoleucine, leucine and valine; and

[0483] (c) substituting the amino acid at the C-terminus with an amino acid selected from the group consisting of arginine and lysine;

[0484] (ii) a peptide comprising an amino acid sequence comprising one or more substitutions selected from the following groups (a) to (c) introduced into an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61:

[0485] (a) substituting the second amino acid from the N-terminus with an amino acid selected from the group consisting of threonine and serine;

[0486] (b) substituting the third amino acid from the N-terminus with an amino acid selected from the group consisting of aspartic acid and glutamic acid; and

[0487] (c) Substitution of the C-terminal amino acid with tyrosine.

[0488] [3] The peptide of claim [1], which consists of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61.

[0489] [4] A polynucleotide encoding the peptide of any one of [1] to [3].

[0490] [5] A composition comprising a pharmaceutically acceptable carrier and at least one component selected from the following groups (a) to (e):

[0491] (a) one or more types of peptides according to any one of [1] to [3];

[0492] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0493] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on its cell surface;

[0494] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on its cell surface; and

[0495] (e) CTLs targeting the peptide of any one of [1] to [3].

[0496] [6] The composition of [5], which is a composition for inducing CTL, wherein the component is at least one component selected from the following groups (a) to (d):

[0497] (a) one or more types of peptides according to any one of [1] to [3];

[0498] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0499] (c) an antigen presenting cell (APC) that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on its cell surface; and

[0500] (d) An exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on its cell surface.

[0501] [7] The composition of [5] is a pharmaceutical composition.

[0502] [8] The composition of [7], which is used for one or more uses selected from the group consisting of: (i) cancer treatment, (ii) cancer prevention (prophylaxis) and (iii) prevention (prophylaxis) of cancer recurrence after surgery.

[0503] [9] The composition of [7] for inducing an immune response against cancer.

[0504]

[10] The composition of [8] or [9], wherein the cancer is selected from the group consisting of acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, small cell lung cancer (SCLC), soft tissue tumors, and testicular tumors.

[0505]

[11] The composition of any one of [5] to

[10] , formulated for administration to a subject positive for at least one HLA selected from the group consisting of HLA-A33 and HLA-A01.

[0506]

[12] A method for inducing APCs with CTL inducibility, comprising a step selected from the group consisting of:

[0507] (a) contacting APC with the peptide of any one of [1] to [3] in vitro, ex vivo or in vivo; and

[0508] (b) introducing a polynucleotide encoding the peptide of any one of [1] to [3] into APC.

[0509]

[13] A method for inducing CTL, comprising steps selected from the following groups (a) to (c):

[0510] (a) co-culturing CD8-positive T cells with APCs presenting a complex of an HLA antigen and the peptide of any one of [1] to [3] on its surface;

[0511] (b) co-culturing CD8-positive T cells with exosomes presenting a complex of an HLA antigen and the peptide of any one of [1] to [3] on its surface;

[0512] (c) introducing a polynucleotide encoding each subunit of a T cell receptor (TCR) capable of binding to a peptide of any one of [1] to [3] presented by an HLA antigen on the cell surface into a CD8-positive T cell.

[0513]

[14] APC that presents a complex of an HLA antigen and the peptide of any one of [1] to [3] on its surface.

[0514]

[15]

[14] APCs, which were induced by the method of

[12] .

[0515]

[16] CTLs targeting the peptide of any one of [1] to [3].

[0516]

[17]

[16] CTL, which was induced by the method of

[13] .

[0517] 18. A method for inducing an immune response against cancer, comprising administering to a subject at least one component selected from the following groups (a) to (e):

[0518] (a) one or more types of peptides according to any one of [1] to [3];

[0519] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0520] (c) APCs that present a complex of the peptide of any one of [1] to [3] and an HLA antigen on their cell surface;

[0521] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on its cell surface; and

[0522] (e) CTLs targeting the peptide of any one of [1] to [3].

[0523] 19. A method for treating and / or preventing cancer, and / or preventing its post-operative recurrence, the method comprising administering to a subject at least one component selected from the following groups (a) to (e):

[0524] (a) one or more types of peptides according to any one of [1] to [3];

[0525] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0526] (c) APCs that present a complex of the peptide of any one of [1] to [3] and an HLA antigen on their cell surface;

[0527] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on its cell surface; and

[0528] (e) CTLs targeting the peptide of any one of [1] to [3].

[0529]

[20] An antibody that binds to the peptide of any one of [1] to [3].

[0530]

[21] A method for screening a peptide having CTL inducibility, comprising the following steps:

[0531] (a) generating a candidate sequence consisting of an amino acid sequence, wherein one, two or several amino acid residues are substituted, deleted, inserted and / or added to an original amino acid sequence consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61;

[0532] (b) selecting, from the candidate sequences generated in (a), candidate sequences other than FOXM1 that do not have significant homology (sequence identity) to any known human gene product;

[0533] (c) contacting the APC with a peptide consisting of the candidate sequence selected in (b);

[0534] (d) contacting the APC of (c) with CD8-positive T cells; and

[0535] (e) Selecting peptides having equal or higher CTL inducibility than that of a peptide consisting of the original amino acid sequence.

[0536]

[22] Use of at least one component selected from the following groups (a) to (e) in the preparation of a composition for inducing an immune response against cancer:

[0537] (a) one or more types of peptides according to any one of [1] to [3];

[0538] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0539] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface;

[0540] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface; and

[0541] (e) CTLs targeting the peptide of any one of [1] to [3].

[0542]

[23] Use of at least one component selected from the following groups (a) to (e) in the preparation of a pharmaceutical composition for treating and / or preventing cancer and / or preventing its postoperative recurrence:

[0543] (a) one or more types of peptides according to any one of [1] to [3];

[0544] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0545] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface;

[0546] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface; and

[0547] (e) CTLs targeting the peptide of any one of [1] to [3].

[0548]

[24] Use of at least one component selected from the following groups (a) to (e) for inducing an immune response against cancer:

[0549] (a) one or more types of peptides according to any one of [1] to [3];

[0550] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0551] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface;

[0552] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface; and

[0553] (e) CTLs targeting the peptide of any one of [1] to [3].

[0554]

[25] Use of at least one component selected from the following groups (a) to (e) for treating and / or preventing cancer and / or preventing its recurrence after surgery:

[0555] (a) one or more types of peptides according to any one of [1] to [3];

[0556] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0557] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface;

[0558] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface; and

[0559] (e) CTLs targeting the peptide of any one of [1] to [3].

[0560]

[26] A method for inducing cytotoxic activity against cells expressing FOXM1, comprising the step of administering to a subject at least one component selected from the following groups (a) to (e):

[0561] (a) one or more types of peptides according to any one of [1] to [3];

[0562] (b) one or more types of polynucleotides encoding the peptide of any one of [1] to [3] in an expressible form;

[0563] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface;

[0564] (d) an exosome that presents a complex of the peptide of any one of [1] to [3] and an HLA antigen on the cell surface; and

[0565] (e) CTLs targeting the peptide of any one of [1] to [3].

[0566]

[27] A freeze-dried preparation comprising one or more types of the peptide of any one of [1] to [3].

[0567]

[28] A pharmaceutical composition prepared by a method comprising dissolving one or more types of the peptide of any one of [1] to [3] in a water-soluble carrier and performing filtration sterilization.

[0568]

[29] A filter-sterilized aqueous solution comprising one or more types of the peptide of any one of [1] to [3] and a water-soluble carrier.

[0569]

[30] An emulsion comprising one or more types of peptides according to any one of [1] to [3], a water-soluble carrier and an oil adjuvant.

[0570]

[31] A kit comprising a container containing the pharmaceutical composition of any one of [5] to

[11] and a container containing an adjuvant.

[0571]

[32] A kit comprising a container for storing a freeze-dried preparation comprising the peptide of any one of [1] to [3], a container for storing an adjuvant, and a container for storing the freeze-dried preparation in a dissolving solution.

[0572] The present invention is explained in detail herein with reference to its specific embodiments. However, it should be understood that the above explanation is actually illustrative and explanatory explanation, and is intended to explain the present invention and its preferred embodiments. Through routine experiments, those skilled in the art will easily recognize that various changes and modifications can be made therein without departing from the spirit and scope of the present invention. Therefore, the present invention is not limited to the above description, but is intended to be limited by the attached claims and their equivalents.

[0573] Hereinafter, the present invention is described in more detail with reference to examples. However, although the following materials, methods and examples can be used to help those of ordinary skill to carry out and use certain embodiments of the present invention, these are only intended to illustrate various aspects of the present invention, so by no means limit the scope of the present invention. As those of ordinary skill in the art will easily recognize that methods and materials similar or equivalent to those described herein can be used for implementation or testing of the present invention.

[0574] All prior art documents cited herein are incorporated into this specification by reference.

[0575] Example

[0576] [Example 1]

[0577] Materials and methods

[0578] Cell lines

[0579] C1R cells (HLA-A negative and HLA-B negative human B lymphoblastoid cell line), and COS7 cells (African green monkey kidney cell line) were purchased from ATCC.

[0580] Generation of target cells with stable HLA-A*33:03 expression

[0581] C1R stably expressing HLA-A*33:03 (C1R-A33) was used as a cell to stimulate CTL. The cDNA encoding HLA-A*33:03 was amplified by PCR and cloned into an expression vector. C1R cells introduced with the HLA-A*33:03 gene expression vector were cultured in a medium containing G418 (Invitrogen) for two weeks under drug selection. The G418-resistant C1R cell suspension was diluted, inoculated in a 96-well plate, and further selectively cultured in a medium containing G418 for 30 days. HLA-A*33:03 expression in C1R cells was confirmed by flow cytometry analysis.

[0582] Selection of FOXM1-derived peptides

[0583] FOXM1-derived 9- and 10-mer peptides predicted to bind to the HLA-A*33:03 molecule were identified using the binding prediction server "NetMHC pan2.8" (www.cbs.dtu.dk / services / NetMHCpan-2.8 / ) (Buus et al., Tissue Antigens. 2003, 62(5):378-84; Nielsen et al., Protein Sci. 2003, 12(5):1007-17; Bioinformatics. 2004, 20(9):1388-97).

[0584] Peptide synthesis

[0585] The peptide was synthesized by American Peptide Company (Sunnyvale, CA) according to standard solid phase synthesis and purified by reverse phase high performance liquid chromatography (HPLC). The quality of the peptide was ensured by HPLC and mass spectrometry (purity 90% or higher). The peptide was dissolved in dimethyl sulfoxide (final concentration: 20 mg / ml) and stored at -80°C.

[0586] In vitro CTL induction

[0587] Dendritic cells (DC) derived from monocytes are used as antigen presenting cells to induce specific cytotoxic T lymphocytes (CTL) responses for peptides presented on human leukocyte antigens (HLA). As reported in the literature, DC is generated in vitro (Nakahara S et al., Cancer Res 2003, 63 (14): 4112-8). Specifically, peripheral blood mononuclear cells (PBMC) collected from healthy volunteers with Ficoll-Paque plus solution (Pharmacia) are inoculated into plastic tissue culture dishes (Corning) so that the monocytes in PBMC adhere to the dish. This is cultured for 7 days in the presence of 1000IU / ml granulocyte macrophage colony stimulating factor (R&D System) and 1000IU / ml interleukin (IL)-4 (R&D System). AIM-V culture medium (Invitrogen) containing 5% inactivated AB type serum (ABS) is used as culture medium. DCs induced to differentiate from monocytes using cytokines were pulsed with 20 μg / ml of each synthesized peptide (37 degrees Celsius, 3 hours). Peptide pulses were performed in AIM-V medium containing 3 μg / ml beta2-microglobulin. DCs pulsed with these peptides were inactivated by X-ray irradiation (20 Gy) and mixed with autologous CD8-positive T cells obtained by using a CD8-positive isolation kit (Invitrogen) at a ratio of 1:20 (1.5x10 4 DC and 3x10 5CD8 positive T cells) were cultured in 48-well plates (Corning). Each well contained 0.5 ml of 5% ABS / AIM-V medium, and IL-7 (R&D System) was added thereto (final concentration: 10 ng / ml). Two days after the start of culture, IL-2 (Novartis) was added (final concentration: 20 IU / ml). On the 7th and 14th days of culture, CD8 positive T cells were further stimulated with peptide-pulsed DCs. DCs were prepared by the same method as described above when used. After 21 days (after 3 DC stimulations), human interferon (IFN)-gamma enzyme-linked immunospot (ELISPOT) assay (Tanaka H et al., Br J Cancer 2001, 84(1):94-9; Umano Y et al., Br J Cancer 2001, 84(8):1052-7; Uchida N et al., Clin Cancer Res 2004, 10(24):8577-86; Suda T et al., Cancer Sci 2006, 97(5):411-9; Watanabe T et al., Cancer Sci 2005, 96(8):498-506) was used to confirm IFN-gamma production in response to peptide-pulsed C1R-A33.

[0588] CTL Expansion Procedure

[0589] CTLs were expanded in culture using a method similar to that reported by Riddell et al. (Walter EA et al., N Engl J Med 1995, 333(16):1038-44; Riddell SR et al., Nat Med 1996, 2(2):216-23). ​​Two types of human B lymphoblastoid cell lines (5x10 6CTLs were cultured with anti-CD3 antibody (final concentration: 40 ng / ml) and IL-2 (final concentration: 120 IU / ml) one day after the start of culture. On days 5, 8 and 11, the culture medium was replaced with 5% ABS / AIM-V medium containing IL-2 (final concentration: 30 IU / ml) (Tanaka H et al., Br J Cancer 2001, 84(1):94-9; Umano Y et al., Br J Cancer 2001, 84(8):1052-7; Uchida N et al., Clin Cancer Res 2004, 10(24):8577-86; Suda T et al., Cancer Sci 2006, 97(5):411-9; Watanabe T et al., Cancer Sci 2005, 96(8):498-506).

[0590] Establishment of CTL clones

[0591] After induction of CTL in vitro, CTL were seeded at 1 cell / well or 10 cells / well in 96-well round-bottom microplates (NalgeNunc International). CTL were co-cultured with two types of human B lymphoblastoid cell lines treated with mitomycin C (1×10 4 cells / well). After 10 days, 50 microliters of 5% ABS / AIM-V containing 500 IU / ml IL-2 were added to the culture. On day 14 or later, CTLs showing peptide-specific IFN-gamma production in ELISPOT assays were expanded using the same method as described above (Uchida N et al., Clin Cancer Res 2004, 10(24):8577-86; Suda T et al., Cancer Sci 2006, 97(5):411-9; Watanabe T et al., Cancer Sci 2005, 96(8):498-506).

[0592] Confirmation of IFN-gamma production

[0593] In order to confirm the peptide-specific IFN-gamma production of CTLs induced by peptides, IFN-gamma ELISPOT assay and IFN-gamma ELISA were performed. C1R-A33 cells (1×10 4 IFN-gamma ELISPOT assay and IFN-gamma ELISA were performed according to the manufacturer's instructions.

[0594] Preparation of cells forced to express FOXM1 gene and HLA-A*33:03

[0595] cDNA encoding FOXM1 or HLA-A*33:03 gene was amplified by PCR. Each PCR amplification product was cloned into an expression vector. One or both of the FOXM1 gene expression vector and the HLA-A*33:03 gene expression vector were introduced into COS7 cells (which is an HLA-negative cell line) using Lipofectamine 2000 (Invitrogen). One day after the gene was introduced, COS7 cells were dissociated and harvested using Versene (Invitrogen) and used as target cells for confirming IFN-gamma production (5x10 4 cells / well).

[0596] result

[0597] Prediction of FOXM1-derived HLA-A*33:03 binding peptides

[0598] Tables 1a and 1b show the FOXM1-derived 9-mer peptides and 10-mer peptides predicted to bind to HLA-A*33:03 by "NetMHC pan2.8" in descending order of binding affinity. A total of 47 peptides with potential HLA-A*33:03 binding ability were used as epitope peptide candidates.

[0599] [Table 1a]

[0600] HLA-A*33:03 binding 9-mer peptide derived from FOXM1

[0601] Starting position Amino acid sequence Kd(nM) SEQ ID NO 180 SLSNIQWLR 13.57 1 308 WTIHPSANR 34.66 2 693 QVSGLAANR 75.29 3 190 MSSDGLGSR 132.48 4 515 MLVIQHRER 138.98 5 516 LVIQHRERR 156.26 6 146 AARDVNLPR 230.20 7 140 TLGPKPAAR 248.65 8 389 SLMSSELAR 262.06 9 246 QFAINSTER 277.30 10 289 LSLHDMFVR 439.26 11 228 SWQNSVSER 481.46 12 125 TQTSYDAK 519.42 13 489 DSSQSPTPR 646.55 14 270 FPYFKHIAK 766.69 15 216 QVKVEEPSR 835.12 16 502 YSGLRSPTR 855.62 17 321 DQVFKQQK 1343.52 18 393 SELARHSKR 1443.91 19 341 KTELPLGAR 1574.92 20 547 FSEGPSTSR 1679.70 21

[0602] The starting position indicates the number of amino acid residues from the N-terminus of FOXM1.

[0603] The dissociation constant [Kd (nM)] was derived from “NetMHC2.8”.

[0604] [Table 1b]

[0605] HLA-A*33:03 binding 10-mer peptide derived from FOXM1

[0606] Starting position Amino acid sequence Kd(nM) SEQ ID NO 514 EMLVIQHRER 26.71 22 139 ETLGPKPAAR 33.00 23 179 NSLSNIQWLR 34.89 24 288 NLSLHDMFVR 91.77 25 501 SYSGLRSPTR 109.92 26 515 MLVIQHRERR 114.82 27 307 FWTIHPSANR 131.76 28 398 HSKRVRIAPK 150.52 29 245 IQFAINSTER 172.14 30 269 HFPYFKHIAK 176.26 31 124 QTQTSYDAKR 190.55 32 595 STPSKSVLPR 240.63 33 394 ELARHSKRVR 278.30 34 247 FAINSTERKR 299.53 35 546 LFSEGPSTSR 332.38 36 392 SSELARHSKR 608.20 37 227 ASWQNSVSER 753.17 38 391 MSSELARHSK 875.46 39 326 QQKRPNPELR 880.19 40 607 ESWRLTPPAK 1179.88 41 265 WIEDHFPYFK 1214.75 42 497 RPKKSYSGLR 1332.87 43 471 EWPSPAPSFK 1418.64 44 4 SPRRPLILKR 1427.21 45 388 ASLMSSELAR 1586.13 46 651 SAPPLESPQR 1587.27 47

[0607] The starting position indicates the number of amino acid residues from the N-terminus of FOXM1.

[0608] The dissociation constant [Kd (nM)] was derived from “NetMHC2.8”.

[0609] Induction of CTL by predicted FOXM1-derived HLA-A*33:03-restricted peptide

[0610] FOXM1-derived peptide-specific CTLs were induced according to the protocol described in "Materials and Methods". Peptide-specific IFN-gamma production was confirmed by ELISPOT assay ( FIG. 1 ). Peptide-specific IFN-gamma production was observed in the following wells:

[0611] Well #6 using OXM1-A33-9-180 (SEQ ID NO: 1) (a)

[0612] Well #3 using FOXM1-A33-9-308 (SEQ ID NO: 2) (b)

[0613] Well #4 using FOXM1-A33-9-693 (SEQ ID NO: 3) (c)

[0614] Well #3 using FOXM1-A33-9-516 (SEQ ID NO: 6) (d)

[0615] Well #5 using FOXM1-A33-9-146 (SEQ ID NO: 7) (e)

[0616] Well #6 using FOXM1-A33-9-289 (SEQ ID NO: 11) (f)

[0617] Well #6 using FOXM1-A33-9-228 (SEQ ID NO: 12) (g)

[0618] Well #4 using FOXM1-A33-9-502 (SEQ ID NO: 17) (h)

[0619] Well #2 using FOXM1-A33-9-321 (SEQ ID NO: 18) (i)

[0620] Well #6 using FOXM1-A33-9-341 (SEQ ID NO: 20) (j)

[0621] Well #8 using FOXM1-A33-10-514 (SEQ ID NO: 22) (k)

[0622] Well #6 using FOXM1-A33-10-179 (SEQ ID NO: 24) (1)

[0623] Well #5 using FOXM1-A33-10-501 (SEQ ID NO: 26) (m)

[0624] Well #5 using FOXM1-A33-10-124 (SEQ ID NO: 32) (n)

[0625] Well #3 using FOXM1-A33-10-595 (SEQ ID NO: 33) (o)

[0626] Well #5 using FOXM1-A33-10-546 (SEQ ID NO: 36) (p)

[0627] Well #6 using FOXM1-A33-10-391 (SEQ ID NO: 39) (q)

[0628] Well #3 using FOXM1-A33-10-607 (SEQ ID NO: 41) (r)

[0629] Well #2(s) using FOXM1-A33-10-265 (SEQ ID NO:42)

[0630] Well #6 (t) and

[0631] Well #5 (u) using FOXM1-A33-10-388 (SEQ ID NO: 46).

[0632] Meanwhile, no specific IFN-gamma production was observed for other peptides shown in Tables 1a and 1b. For example, no specific IFN-gamma production was observed for FOXM1-A33-10-288 (SEQ ID NO: 25) (v). As a result, although all peptides had the potential to bind to HLA-A*33:03, 21 peptides were selected as peptides having CTL inducibility.

[0633] Establishment of CTL lines and clones specific for HLA-A*33:03-restricted FOXM1-derived peptides

[0634] CTL lines were established by proliferating cells in the following wells in IFN-gamma ELISPOT assay: in well #3 using FOXM1-A33-9-308 (SEQ ID NO: 2) (a), in well #5 using FOXM1-A33-9-146 (SEQ ID NO: 7) (b), in well #6 using FOXM1-A33-10-391 (SEQ ID NO: 39) (c), and in well #6 using FOXM1-A33-10-265 (SEQ ID NO: 42) (d). As a result of measuring IFN-gamma by ELISA, IFN-gamma production by CTL lines against the following target cells (C1R-A33) was observed ( Figure 2 ): target cells pulsed with well #3 having FOXM1-A33-9-308 (SEQ ID NO: 2) (a), target cells pulsed with well #5 having FOXM1-A33-9-146 (SEQ ID NO: 7) (b), target cells pulsed with well #6 having FOXM1-A33-10-391 (SEQ ID NO: 39) (c), or FOXM1-A33-10-265 (SEQID NO: 42). In addition, CTL clones were established by limiting dilution from the CTL lines as described in the “Materials and Methods” section above. As a result of measuring IFN-gamma by ELISA, CTL clones stimulated with FOXM1-A33-9-308 (SEQ ID NO: 2) (a) or FOXM1-A33-9-146 (SEQ ID NO: 7) (b) each showed peptide-specific IFN-gamma production ( Figure 3 ).

[0635] IFN-gamma production against target cells expressing FOXM1 and HLA-A*33:03

[0636] The IFN-gamma production of FOXM1-A33-9-308 (SEQ ID NO: 2)-specific CTL clones against target cells expressing FOXM1 and HLA-A*33:03 was studied. COS7 cells expressing both FOXM1 and HLA-A*33:03 were prepared as target cells. COS7 cells expressing either FOXM1 or HLA-A*33:03 were prepared as negative control cells. The FOXM1-A33-9-308 (SEQ ID NO: 2)-specific CTL clone showed IFN-gamma production against COS7 cells expressing both FOXM1 and HLA-A*33:03 ( Figure 4). On the other hand, no significant IFN-gamma production was observed against negative control cells. This clearly demonstrates that FOXM1-A33-9-308 (SEQ ID NO: 2) is a peptide produced by antigen processing and is presented on the cell surface with HLA-A*33:03 molecules and recognized by CTLs. This result suggests that FOXM1-A33-9-308 (SEQ ID NO: 2) can be useful as a cancer vaccine for patients whose cancer cells have enhanced FOXM1 expression.

[0637] Homology analysis of antigenic peptides

[0638] It has been confirmed that FOXM1-A33-9-180 (SEQ ID NO: 1), FOXM1-A33-9-308 (SEQ ID NO: 2), FOXM1-A33-9-693 (SEQ ID NO: 3), FOXM1-A33-9-516 (SEQ ID NO: 6), FOXM1-A33-9-146 (SEQ ID NO:7), FOXM1-A33-9-289 (SEQ ID NO:11), FOXM1-A33-9-228 (SEQ ID NO:12), FOXM1-A33-9-502 (SEQ ID NO:17), FOXM1-A33-9-321 (SEQ ID NO:18), FOXM1-A33-9-341 (SEQ IDNO:20), FOXM1-A33-10-514 (SEQ ID NO:22), FOXM1-A33-10-179 (SEQ ID NO:24), FOXM1-A33-10-501 (SEQ ID NO:26), FOXM1-A33-10-124 (SEQ ID NO:32), FOXM1-A33-10-595 (SEQID NO:33), FOXM1-A33-10-546 (SEQ ID NO:36), FOXM1-A33-10-391 (SEQ ID NO:39), FOXM1-A33-10-607 (SEQ ID NO:41), FOXM1-A33-10-265 (SEQ ID NO:42), FOXM1-A33-10-4 (SEQ ID NO:45) and FOXM1-A33-10-388 (SEQ ID NO:46) can induce CTLs that display peptide-specific IFN-gamma production.Therefore, in order to confirm FOXM1-A33-9-180 (SEQ ID NO: 1), FOXM1-A33-9-308 (SEQ ID NO: 2), FOXM1-A33-9-693 (SEQ ID NO: 3), FOXM1-A33-9-516 (SEQ ID NO: 6), FOXM1-A33-9-146 (SEQ ID NO: 7), FOXM1-A33-9-289 (SEQ ID NO: 11), FOXM1-A33-9-228 (SEQ ID NO: 12), FOXM1-A33-9-502 (SEQ ID NO: 17), FOXM1-A33-9-321 (SEQ ID NO: 18), FOXM1-A33-9-341 (SEQ ID NO: 20), FOXM1-A33-10-514 (SEQ ID NO: 21), FOXM1-A33-9-10-526 (SEQ ID NO: 22), FOXM1-A33-9-110 (SEQ ID NO: 23), FOXM1-A33-9-111 (SEQ ID NO: 24), FOXM1-A33-9-112 (SEQ ID NO: 25), FOXM1-A33-9-113 (SEQ ID NO: 26), FOXM1-A33-9-114 (SEQ ID NO: 27), FOXM1-A33-9-115 (SEQ ID NO: 28), FOXM1-A33-9-116 (SEQ ID NO: 29), FOXM1-A33-9-117 (SEQ ID NO: 30), FOXM1-A33-9-118 (SEQ ID NO: NO:22), FOXM1-A33-10-179 (SEQ ID NO:24), FOXM1-A33-10-501 (SEQ ID NO:26), FOXM1-A33-10-124 (SEQ ID NO:32), FOXM1-A33-10-595 (SEQ ID NO:33), FOXM1-A33-10-546 (SEQ ID NO:36), FOXM1-A33-10-391 (SEQ ID NO:39), FOXM1-A33-10-607 (SEQ ID NO:41), FOXM1-A33-10-265 (SEQ ID NO:42), FOXM1-A33-10-4 (SEQ ID NO:45) and FOXM1-A33-10-388 (SEQ ID NO:46) sequence was derived only from FOXM1, and homology analysis of the peptide sequence was performed using the BLAST algorithm (blast.ncbi.nlm.nih.gov / Blast.cgi).As a result, FOXM1-A33-9-180 (SEQ ID NO: 1), FOXM1-A33-9-308 (SEQ ID NO: 2), FOXM1-A33-9-693 (SEQ ID NO: 3), FOXM1-A33-9-516 (SEQ ID NO: 6), FOXM1-A33-9-146 (SEQ ID NO: 7), FOXM1-A33-9-289 (SEQ ID NO: 11), FOXM1-A33-9-228 (SEQ ID NO: 12), FOXM1-A33-9-502 (SEQ ID NO: 17), FOXM1-A33-9-321 (SEQ ID NO: 18), FOXM1-A33-9-341 (SEQ ID NO: 20), FOXM1-A33-10-514 (SEQ ID NO: 21), FOXM1-A33-9-10-526 (SEQ ID NO: 22), FOXM1-A33-9-110 (SEQ ID NO: 23), FOXM1-A33-9-111 (SEQ ID NO: 24), FOXM1-A33-9-112 (SEQ ID NO: 25), FOXM1-A33-9-113 (SEQ ID NO: 26), FOXM1-A33-9-114 (SEQ ID NO: 27), FOXM1-A33-9-115 (SEQ ID NO: 28), FOXM1-A33-9-116 (SEQ ID NO: 29), FOXM1-A33-9-1 NO:22), FOXM1-A33-10-179 (SEQ ID NO:24), FOXM1-A33-10-501 (SEQ ID NO:26), FOXM1-A33-10-124 (SEQ ID NO:32), FOXM1-A33-10-595 (SEQ ID NO:33), FOXM1-A33-10-546 (SEQ ID NO:36), FOXM1-A33-10-391 (SEQ ID NO:39), FOXM1-A33-10-607 (SEQ ID NO:41), FOXM1-A33-10-265 (SEQ ID NO:42), FOXM1-A33-10-4 (SEQ ID NO:45) and FOXM1-A33-10-388 (SEQ ID NO:46) sequences. Therefore, according to the present inventors' knowledge, these peptides are specific for FOXM1, making it almost impossible for these peptides to elicit undesirable immune responses against molecules other than FOXM1 (which are known to sensitize the human immune system). In conclusion, novel FOXM1-derived HLA-A*33:03-restricted epitope peptides were identified. It was demonstrated that FOXM1-derived epitope peptides are suitable for cancer immunotherapy.

[0639] [Example 2]

[0640] Materials and methods

[0641] Cell lines

[0642] C1R cells (HLA-A negative and HLA-B negative human B lymphoblastoid cell line), and COS7 cells (African green monkey kidney cell line) were purchased from ATCC.

[0643] Generation of target cells with stable HLA-A*01:01 expression

[0644] C1R stably expressing HLA-A*01:01 (C1R-A01) was used as a cell to stimulate CTL. The cDNA encoding HLA-A*01:01 was amplified by PCR and cloned into an expression vector. C1R cells introduced with the HLA-A*01:01 gene expression vector were cultured in a medium containing G418 (Invitrogen) for two weeks under drug selection. The G418-resistant C1R cell suspension was diluted, inoculated in a 96-well plate, and further selectively cultured in a medium containing G418 for 30 days. HLA-A*01:01 expression in C1R cells was confirmed by flow cytometry analysis.

[0645] Selection of FOXM1-derived peptides

[0646] FOXM1-derived 9- and 10-mer peptides predicted to bind to HLA-A*01:01 molecules were identified using the binding prediction server "NetMHC 3.4" (www.cbs.dtu.dk / services / NetMHC-3.4 / ) (Buus S et al., Tissue Antigens. 2003, 62(5):378-84; Nielsen M et al., Protein Sci. 2003, 12(5):1007-17; Nielsen M et al., Bioinformatics. 2004, 20(9):1388-97).

[0647] Peptide synthesis

[0648] The peptide was synthesized by American Peptide Company (Sunnyvale, CA) according to standard solid phase synthesis and purified by reverse phase high performance liquid chromatography (HPLC). The quality of the peptide was ensured by HPLC and mass spectrometry (purity 90% or higher). The peptide was dissolved in dimethyl sulfoxide (final concentration: 20 mg / ml) and stored at -80°C.

[0649] In vitro CTL induction

[0650] Dendritic cells (DC) derived from monocytes are used as antigen presenting cells to induce specific cytotoxic T lymphocytes (CTL) responses for peptides presented on human leukocyte antigens (HLA). As reported in the literature, DC is generated in vitro (Nakahara S et al., Cancer Res 2003, 63 (14): 4112-8). Specifically, peripheral blood mononuclear cells (PBMC) collected from healthy volunteers (HLA-A*01:01 positive) are inoculated into plastic tissue culture dishes (Corning) so that the monocytes in the PBMC adhere to the dish. This is cultured for 7 days in the presence of 1000IU / ml granulocyte macrophage colony stimulating factor (R&D System) and 1000IU / ml interleukin (IL)-4 (R&D System). AIM-V culture medium (Invitrogen) containing 5% inactivated AB type serum (ABS) is used as culture medium. DCs induced to differentiate from monocytes using cytokines were pulsed with 20 μg / ml of each synthesized peptide (37 degrees Celsius, 3 hours). Peptide pulses were performed in AIM-V medium containing 3 μg / ml beta 2-microglobulin. DCs pulsed with these peptides were inactivated by X-ray irradiation (20 Gy) and mixed with autologous CD8 positive T cells obtained by using a CD8 positive isolation kit (Invitrogen) at a ratio of 1:20 (1.5x10 4 DC and 3x10 5CD8 positive T cells) were cultured in 48-well plates (Corning). Each well contained 0.5 ml of 5% ABS / AIM-V medium, and IL-7 (R&D System) was added thereto (final concentration: 10 ng / ml). Two days after the start of culture, IL-2 (Novartis) was added (final concentration: 20 IU / ml). On the 7th and 14th days of culture, CD8 positive T cells were further stimulated with peptide-pulsed DCs. DCs were prepared by the same method as described above when used. After 21 days (after 3 DC stimulations), human interferon (IFN)-gamma enzyme-linked immunospot (ELISPOT) assay (Tanaka H et al., Br J Cancer 2001, 84(1):94-9; Umano Y et al., Br J Cancer 2001, 84(8):1052-7; Uchida N et al., Clin Cancer Res 2004, 10(24):8577-86; Suda T et al., Cancer Sci 2006, 97(5):411-9; Watanabe Tet al., Cancer Sci 2005, 96(8):498-506) was used to confirm IFN-gamma production in response to peptide-pulsed C1R-A01.

[0651] CTL Expansion Procedure

[0652] CTLs were expanded using a method similar to that disclosed by Riddell et al. (Walter EA et al., N Engl J Med 1995, 333(16):1038-44; Riddell SR et al., Nat Med 1996, 2(2):216-23). ​​Two types of human B lymphoblastoid cell lines (5x10 6CTLs were cultured with anti-CD3 antibody (final concentration 40 ng / ml) and 1% ABS / AIM-V medium (Tanaka H et al., Br J Cancer 2001, 84(1):94-9; Umano Y et al., Br J Cancer 2001, 84(8):1052-7; Uchida N et al., Clin Cancer Res 2004, 10(24):8577-86; Suda T et al., Cancer Sci 2006, 97(5):411-9; Watanabe T et al., Cancer Sci 2005, 96(8):498-506).

[0653] Establishment of CTL clones

[0654] After induction of CTL in vitro, CTL were seeded at 1 cell / well or 10 cells / well in 96-well round-bottom microplates (NalgeNunc International). CTL were co-cultured with two types of human B lymphoblastoid cell lines treated with mitomycin C (1×10 4 cells / well). After 10 days, 50 microliters of 5% ABS / AIM-V containing 500 IU / ml IL-2 were added to the culture. On day 14 or later, CTLs showing peptide-specific IFN-gamma production in ELISPOT assays were expanded using the same method as described above (Uchida N et al., Clin Cancer Res 2004, 10(24):8577-86; Suda T et al., Cancer Sci 2006, 97(5):411-9; Watanabe T et al., Cancer Sci 2005, 96(8):498-506).

[0655] Confirmation of IFN-gamma production

[0656] In order to confirm the peptide-specific IFN-gamma production by CTLs induced with peptides, IFN-gamma ELISPOT assay and IFN-gamma ELISA were performed. 4 IFN-gamma ELISPOT assay and IFN-gamma ELISA were performed according to the manufacturer's instructions.

[0657] Preparation of cells forced to express FOXM1 gene and HLA-A*01:01

[0658] cDNA encoding FOXM1 or HLA-A*01:01 gene was amplified by PCR. The PCR amplification products were each integrated into an expression vector. One or both of the FOXM1 gene expression vector and the HLA-A*01:01 gene expression vector were introduced into COS7 cells (which is an HLA-negative cell line) using Lipofectamine 2000 (Invitrogen). One day after the gene introduction, COS7 cells were dissociated and harvested using Versene (Invitrogen) and used as target cells for confirmation of IFN-gamma production (5x10 4 cells / well).

[0659] result

[0660] Prediction of FOXM1-derived HLA-A*01:01 binding peptides

[0661] Tables 2a and 2b show the FOXM1-derived 9-mer peptides and 10-mer peptides predicted to bind to HLA-A*01:01 by "NetMHC 3.4" in descending order of binding affinity. A total of 16 peptides with potential HLA-A*01:01 binding ability were used as epitope peptide candidates.

[0662] [Table 2a]

[0663] Predicted binding of FOXM1-derived 9-mer peptide to HLA-A*01:01

[0664] Starting position Amino acid sequence Kd(nM) SEQ ID NO 233 VSERPPYSY 73 48 539 CVDEPELLF 666 49 631 ASDPLPDPL 1807 50 703 LTEGLVLDT 6665 51 231 NSVSERPPY 6867 52 663 SSEPLDLIS 9729 53 392 SSELARHSK 18415 54 494 PTPRPKKSY 19761 55 341 KTELPLGAR 20899 20

[0665] The number of the starting position indicates the number of amino acids from the N-terminus of the FOXM1 protein to which the first amino acid of the peptide corresponds.

[0666] The dissociation constant [Kd (nM)] was calculated using “NetMHC3.4”.

[0667] [Table 2b]

[0668] FOXM1-derived 10-mer peptide predicted to bind HLA-A*01:01

[0669] Starting position Amino acid sequence Kd(nM) SEQ ID NO 566 SSDPASQLSY 15 56 263 YTWIEDHFPY 156 57 308 WTIHPSANRY 554 58 232 SVSERPPYSY 1482 59 663 SSEPlDLISV 10539 60 265 WIEDHFPYFK 18243 42 341 KTELPLGARR 26951 61

[0670] The number of the starting position indicates the number of amino acids from the N-terminus of the FOXM1 protein to which the first amino acid of the peptide corresponds.

[0671] The dissociation constant [Kd (nM)] was calculated using “NetMHC3.4”.

[0672] Induction of CTL by predicted FOXM1-derived HLA-A*01:01-restricted peptide

[0673] FOXM1-derived peptide-specific CTLs were induced according to the protocol described in "Materials and Methods". Peptide-specific IFN-gamma production was confirmed by ELISPOT assay ( FIG. 5 ). Peptide-specific IFN-gamma production was observed in the following wells:

[0674] In well #3 (a) using FOXM1-A01-9-233 (SEQ ID NO: 48),

[0675] In well #3 (b) using FOXM1-A01-9-539 (SEQ ID NO: 49),

[0676] Using well #3 with FOXM1-A01-9-631 (SEQ ID NO: 50) (c),

[0677] In well #2 (d) using FOXM1-A01-9-231 (SEQ ID NO: 52),

[0678] In well #2 (e) using FOXM1-A01-9-663 (SEQ ID NO: 53),

[0679] In well #5 (f) using FOXM1-A01-9-494 (SEQ ID NO: 55),

[0680] In well #2 (g) using FOXM1-A01-9-341 (SEQ ID NO: 20),

[0681] In well #1 using FOXM1-A01-10-566 (SEQ ID NO: 56) (h),

[0682] In well #2 (i) using FOXM1-A01-10-263 (SEQ ID NO: 57),

[0683] In well #4 (j) using FOXM1-A01-10-308 (SEQ ID NO: 58),

[0684] In well #6 (k) using FOXM1-A01-10-232 (SEQ ID NO: 59),

[0685] Well #6 (1) and

[0686] Well #6 (m) using FOXM1-A01-10-341 (SEQ ID NO: 61).

[0687] Meanwhile, no specific IFN-gamma production was observed for other peptides shown in Tables 1a and 1b. For example, no specific IFN-gamma production was observed for FOXM1-A01-10-265 (SEQ ID NO: 42) (n). As a result, although all peptides had the potential to bind to HLA-A*01:01, 13 peptides were selected as peptides having CTL inducibility.

[0688] Establishment of CTL lines and clones specific for HLA-A*01:01-restricted FOXM1-derived peptides

[0689] A CTL line was established by amplifying the cells in well 1#, which showed specific IFN-gamma production against FOXM1-A01-10-566 (SEQ ID NO: 56) in the IFN-gamma ELISPOT assay. As a result of measuring IFN-gamma by ELISA, IFN-gamma production of the CTL line against target cells (C1R-A01) impacted with FOXM1-A01-10-566 (SEQ ID NO: 56) was observed. In addition, CTL clones were established by the limiting dilution method. As a result of measuring IFN-gamma by ELISA, CTL clones stimulated with FOXM1-A01-9-233 (SEQ ID NO: 48) (a) or FOXM1-A01-10-566 (SEQ ID NO: 56) (b) each showed peptide-specific IFN-gamma production ( Figure 7 ).

[0690] IFN-gamma production against target cells expressing FOXM1 and HLA-A*01:01

[0691] The IFN-gamma production of FOXM1-A01-10-566 (SEQ ID NO: 56)-specific CTL clones against target cells expressing FOXM1 and HLA-A*01:01 was studied. COS7 cells expressing both FOXM1 and HLA-A*01:01 were prepared as target cells. COS7 cells expressing either FOXM1 or HLA-A*01:01 were prepared as negative control cells. The FOXM1-A01-10-566 (SEQ ID NO: 56)-specific CTL clone showed IFN-gamma production against COS7 cells expressing both FOXM1 and HLA-A*01:01 ( Figure 8 ). On the other hand, no significant IFN-gamma production was observed against negative control cells. This clearly demonstrates that FOXM1-A01-10-566 (SEQ ID NO: 56) is a peptide produced by antigen processing and is presented on the cell surface with HLA-A*01:01 molecules and recognized by CTLs. This result suggests that FOXM1-A01-10-566 (SEQ ID NO: 56) can be useful as a cancer vaccine for patients with enhanced FOXM1 expression in cancer cells.

[0692] Homology analysis of antigenic peptides

[0693] It has been confirmed that FOXM1-A01-9-233 (SEQ ID NO:48), FOXM1-A01-9-539 (SEQ ID NO:49), FOXM1-A01-9-631 (SEQ ID NO:50), FOXM1-A01-9-231 (SEQ ID NO:52), FOXM1-A01-9-663 (SEQ ID NO:53), FOXM1-A01-9-494 (SEQ ID NO:55), FOXM1-A01-9-341 (SEQ ID NO:20), FOXM1-A01-10-566 (SEQ ID NO:56), FOXM1-A01-10-263 (SEQ ID NO:57), FOXM1-A01-10-308 (SEQ ID NO:58), FOXM1-A01-10-232 (SEQ ID NO:59), FOXM1-A01-10-663 (SEQ ID NO:60) and FOXM1-A01-10-341 (SEQ ID NO:61) can induce CTLs showing peptide-specific IFN-gamma production. Therefore, in order to confirm FOXM1-A01-9-233 (SEQ ID NO: 48), FOXM1-A01-9-539 (SEQ ID NO: 49), FOXM1-A01-9-631 (SEQ ID NO: 50), FOXM1-A01-9-231 (SEQ ID NO: 52), FOXM1-A01-9-663 (SEQ ID NO: 53), FOXM1-A01-9-494 (SEQ ID NO: 55), FOXM1-A01-9-341 (SEQ ID NO: 20), FOXM1-A01-10-566 (SEQ ID NO: 56), FOXM1-A01-10-263 (SEQ ID NO: 57), FOXM1-A01-10-308 (SEQ ID NO: 58), FOXM1-A01-10-232 (SEQ ID NO: 59), FOXM1-A01-9-494 (SEQ ID NO: 50), FOXM1-A01-9-494 (SEQ ID NO: 51), FOXM1-A01-9-341 (SEQ ID NO: 20), FOXM1-A01-10-566 (SEQ ID NO: 52), FOXM1-A01-10-263 (SEQ ID NO: 53), FOXM1-A01-10-308 (SEQ ID NO: 54), FOXM1-A01-10-232 (SEQ ID NO: 55), FOXM1-A01-10-494 (SEQ ID NO: 56), FOXM1-A01-9-494 (SEQ ID NO: NO:59), FOXM1-A01-10-663 (SEQ ID NO:60) and FOXM1-A01-10-341 (SEQ ID NO:61) sequences were derived only from FOXM1, and homology analysis of peptide sequences was performed using the BLAST algorithm (blast.ncbi.nlm.nih.gov / Blast.cgi).As a result, FOXM1-A01-9-233 (SEQ ID NO: 48), FOXM1-A01-9-539 (SEQ ID NO: 49), FOXM1-A01-9-631 (SEQ ID NO: 50), FOXM1-A01-9-231 (SEQ ID NO: 52), FOXM1-A01-9-663 (SEQ ID NO: 53), FOXM1-A01-9-494 (SEQ ID NO: 55), FOXM1-A01-9-341 (SEQ ID NO: 20), FOXM1-A01-10-566 (SEQ ID NO: 56), FOXM1-A01-10-263 (SEQ ID NO: 57), FOXM1-A01-10-308 (SEQ ID NO: 58), and FOXM1-A01-10-494 (SEQ ID NO: 59). NO:58), FOXM1-A01-10-232 (SEQ ID NO:59), FOXM1-A01-10-663 (SEQ ID NO:60) and FOXM1-A01-10-341 (SEQ ID NO:61). Therefore, according to the knowledge of the inventors, these peptides are specific for FOXM1, making it almost impossible for these peptides to trigger undesirable immune responses against molecules other than FOXM1 (which are known to sensitize the human immune system). In summary, novel FOXM1-derived HLA-A*01:01-restricted epitope peptides were identified. It was demonstrated that FOXM1-derived epitope peptides are suitable for cancer immunotherapy.

[0694] [Example 3]

[0695] Preparation of emulsion formulations

[0696] The peptide is dissolved in an injection solvent or sterile saline to 1.0mg / ml to 10.0mg / ml, and collected in a syringe. This is connected to a syringe equipped with an IFA equivalent to an injection solvent or sterile saline via a connector, and then mixed by alternately pushing the syringe plungers of two connected syringes. After mixing for a few minutes, the completion of the emulsion is evaluated by a drop test method. The drop test method can be performed by dropping a drop of a mixed sample on water. When the sample dripped on the water is not immediately diffused in the water, the emulsion is evaluated to be completed; and when the sample dripped on the water is immediately diffused in the water, the emulsion is evaluated to be incomplete. When the emulsion is evaluated to be incomplete, further mixing is performed to complete the emulsion. The completed emulsion can be administered to cancer patients by subcutaneous injection. The cancer patient to be administered can be selected from patients affected by the following cancers: acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc.

[0697] Preparation of freeze-dried formulations

[0698] The peptide is dissolved in an injection solvent to make it 1.0mg / ml to 10.0mg / ml, and filtered and sterilized. It is loaded into a sterilized vial and semi-enclosed with a sterile rubber stopper. After the vial is freeze-dried, it is completely enclosed and sutured with an aluminum cap to produce a freeze-dried preparation. When used, an injection solvent or sterile saline is injected into the vial to redissolve the freeze-dried powder. The redissolved solution in the vial is collected using a syringe, and the syringe is connected to a syringe filled with the same amount of IFA as the amount of the collected redissolved solution by a connector. The redissolved solution and IFA are mixed by alternately pushing the syringe plungers of the two connected syringes. After mixing for a few minutes, the completion of the emulsion is evaluated by a drop test method. The completed emulsion can be administered to cancer patients by subcutaneous injection. The cancer patient to be administered can be selected from patients affected by the following cancers: acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, testicular tumors, etc.

[0699] Industrial Applicability

[0700] The present invention provides novel HLA-A33-restricted and HLA-A01-restricted epitope peptides derived from FOXM1, which induce strong and specific anti-tumor immune responses and thus have applicability to a wide range of cancer types. The peptides, compositions, APCs and CTLs of the present invention can be used as peptide vaccines against cancers expressing FOXM1, such as acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocarcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal cancer, small cell lung cancer (SCLC), soft tissue tumors, and testicular tumors.

[0701] Although the present invention is described in detail and with respect to its specific embodiments herein, it should be understood that the foregoing description is exemplary and explanatory in nature, and is intended to illustrate the present invention and its preferred embodiments. Through routine experiments, those skilled in the art will readily appreciate that various changes and modifications may be made to the present invention without departing from the spirit and scope of the present invention, the boundaries and scope of the present invention being defined by the appended claims.

[0702] The present invention includes

[0703] 1. A peptide of less than 15 amino acids having cytotoxic T cell (CTL) inducibility, comprising an amino acid sequence selected from the following group:

[0704] (a) an amino acid sequence selected from the group consisting of: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60, and 61; and

[0705] (b) an amino acid sequence in which 1, 2 or several amino acids are substituted, deleted, inserted and / or added in an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61.

[0706] 2. The peptide of item 1, which is selected from the following groups (i) to (ii):

[0707] (i) a peptide comprising an amino acid sequence comprising one or more substitutions selected from the following groups (a) to (c) introduced into an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45 and 46:

[0708] (a) replacing the first amino acid from the N-terminus with an amino acid selected from the group consisting of aspartic acid and glutamic acid;

[0709] (b) substituting the second amino acid from the N-terminus with an amino acid selected from the group consisting of phenylalanine, tyrosine, alanine, isoleucine, leucine and valine; and

[0710] (c) substituting the amino acid at the C-terminus with an amino acid selected from the group consisting of arginine and lysine;

[0711] (ii) a peptide comprising an amino acid sequence, wherein one or more substitutions selected from the following groups (a) to (c) are introduced into an amino acid sequence selected from the following group: SEQ ID NOs: 48, 49, 50, 52, 53, 55, 20, 56, 57, 58, 59, 60 and 61:

[0712] (a) substituting the second amino acid from the N-terminus with an amino acid selected from the group consisting of threonine and serine;

[0713] (b) substituting the third amino acid from the N-terminus with an amino acid selected from the group consisting of aspartic acid and glutamic acid; and

[0714] (c) Substitution of the C-terminal amino acid with tyrosine.

[0715] 3. The peptide of claim 1, which consists of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61.

[0716] 4. A polynucleotide encoding the peptide of any one of items 1 to 3.

[0717] 5. A composition comprising a pharmaceutically acceptable carrier and at least one component selected from the following groups (a) to (e):

[0718] (a) one or more types of peptides according to any one of items 1 to 3;

[0719] (b) one or more types of polynucleotides encoding the peptide of any one of items 1 to 3 in an expressible form;

[0720] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of Items 1 to 3 and an HLA antigen on its cell surface;

[0721] (d) an exosome that presents a complex of the peptide of any one of items 1 to 3 and an HLA antigen on its cell surface; and

[0722] (e) CTLs targeting the peptide of any one of items 1 to 3.

[0723] 6. The composition of item 5, which is a composition for inducing CTL, wherein the component is at least one component selected from the following groups (a) to (d):

[0724] (a) one or more types of peptides according to any one of items 1 to 3;

[0725] (b) one or more types of polynucleotides encoding the peptide of any one of items 1 to 3 in an expressible form;

[0726] (c) an antigen presenting cell (APC) which presents a complex of the peptide of any one of Items 1 to 3 and an HLA antigen on its cell surface; and

[0727] (d) An exosome which presents a complex of the peptide of any one of Items 1 to 3 and an HLA antigen on its cell surface.

[0728] 7. The composition of item 5, which is a pharmaceutical composition.

[0729] 8. The composition of claim 7, for use in one or more uses selected from the group consisting of: (i) cancer treatment, (ii) cancer prevention (prophylaxis), and (iii) prevention (prophylaxis) of cancer recurrence after surgery.

[0730] 9. The composition of claim 7, for use in inducing an immune response against cancer.

[0731] 10. The composition of claim 8 or 9, wherein the cancer is selected from the group consisting of acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocellular carcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, diffuse gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, small cell lung cancer (SCLC), soft tissue tumors, and testicular tumors.

[0732] 11. The composition of any one of items 5 to 10, formulated for administration to a subject who is positive for at least one HLA selected from the group consisting of HLA-A33 and HLA-A01.

[0733] 12. A method for inducing APCs with CTL inducibility, comprising a step selected from the group consisting of:

[0734] (a) contacting APC with the peptide of any one of items 1 to 3 in vitro, ex vivo or in vivo; and

[0735] (b) introducing a polynucleotide encoding the peptide of any one of items 1 to 3 into APC.

[0736] 13. A method for inducing CTL, comprising a step selected from the group consisting of:

[0737] (a) co-culturing CD8-positive T cells with APCs presenting a complex of an HLA antigen and a peptide according to any one of items 1 to 3 on its surface;

[0738] (b) co-culturing CD8-positive T cells with exosomes presenting a complex of an HLA antigen and a peptide according to any one of items 1 to 3 on its surface; and

[0739] (c) introducing a polynucleotide encoding each subunit of a T cell receptor (TCR) capable of binding to a peptide of any one of items 1 to 3 presented by an HLA antigen on the cell surface into a CD8-positive T cell.

[0740] 14. An APC which presents a complex of an HLA antigen and the peptide of any one of items 1 to 3 on its surface.

[0741] 15. The APC of item 14, which is induced by the method of item 12.

[0742] 16. CTL targeting the peptide of any one of items 1 to 3.

[0743] 17. The CTL of item 16, which is induced by the method of item 13.

[0744] 18. A method of inducing an immune response against cancer, comprising administering to a subject at least one component selected from the following groups (a) to (e):

[0745] (a) one or more types of peptides according to any one of items 1 to 3;

[0746] (b) one or more types of polynucleotides encoding the peptide of any one of items 1 to 3 in an expressible form;

[0747] (c) APCs which present a complex of the peptide of any one of Items 1 to 3 and an HLA antigen on their cell surface;

[0748] (d) an exosome that presents a complex of the peptide of any one of items 1 to 3 and an HLA antigen on its cell surface; and

[0749] (e) CTLs targeting the peptide of any one of items 1 to 3.

[0750] 19. A method for treating and / or preventing cancer, and / or preventing its post-operative recurrence, the method comprising administering to a subject at least one component selected from the following groups (a) to (e):

[0751] (a) one or more types of peptides according to any one of items 1 to 3;

[0752] (b) one or more types of polynucleotides encoding the peptide of any one of items 1 to 3 in an expressible form;

[0753] (c) APCs which present a complex of the peptide of any one of Items 1 to 3 and an HLA antigen on their cell surface;

[0754] (d) an exosome that presents a complex of the peptide of any one of items 1 to 3 and an HLA antigen on its cell surface; and

[0755] (e) CTLs targeting the peptide of any one of items 1 to 3.

[0756] 20. An antibody which binds to the peptide of any one of items 1 to 3.

[0757] 21. A method for screening a peptide having CTL inducibility, comprising the following steps:

[0758] (a) generating a candidate sequence consisting of an amino acid sequence, wherein one, two or several amino acid residues are substituted, deleted, inserted and / or added to an original amino acid sequence consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, 3, 6, 7, 11, 12, 17, 18, 20, 22, 24, 26, 32, 33, 36, 39, 41, 42, 45, 46, 48, 49, 50, 52, 53, 55, 56, 57, 58, 59, 60 and 61;

[0759] (b) selecting, from the candidate sequences generated in (a), candidate sequences that do not have significant homology (sequence identity) to any known human gene product other than FOXM1;

[0760] (c) contacting the APC with a peptide consisting of the candidate sequence selected in (b);

[0761] (d) contacting the APC in (c) with CD8-positive T cells; and

[0762] (e) Selecting peptides having equal or higher CTL inducibility than that of a peptide consisting of the original amino acid sequence.

[0763] 22. An emulsion comprising one or more types of the peptide of any one of items 1 to 3, a water-soluble carrier and an oil adjuvant.

[0764] 23. A kit comprising a container containing the composition of any one of items 5 to 11 and a container containing an adjuvant. Sequence Listing <110> Oncotherapy Sciences, Inc. <120> FOXM1-derived peptides and vaccines containing the same <130> ONC-A1604P <150> JP 2015-200221 <151> 2015-10-08 <160> 76 <170> PatentIn version 3.5 <210> 1 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 1 Ser Leu Ser Asn Ile Gln Trp Leu Arg 1 5 <210> 2 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 2 Trp Thr Ile His Pro Ser Ala Asn Arg 1 5 <210> 3 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 3 Gln Val Ser Gly Leu Ala Ala Asn Arg 1 5 <210> 4 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 4 Met Ser Ser Asp Gly Leu Gly Ser Arg 1 5 <210> 5 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 5 Met Leu Val Ile Gln His Arg Glu Arg 1 5 <210> 6 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 6 Leu Val Ile Gln His Arg Glu Arg Arg 1 5 <210> 7 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 7 Ala Ala Arg Asp Val Asn Leu Pro Arg 1 5 <210> 8 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 8 Thr Leu Gly Pro Lys Pro Ala Ala Arg 1 5 <210> 9 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 9 Ser Leu Met Ser Ser Glu Leu Ala Arg 1 5 <210> 10 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 10 Gln Phe Ala Ile Asn Ser Thr Glu Arg 1 5 <210> 11 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 11 Leu Ser Leu His Asp Met Phe Val Arg 1 5 <210> 12 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 12 Ser Trp Gln Asn Ser Val Ser Glu Arg 1 5 <210> 13 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 13 Thr Gln Thr Ser Tyr Asp Ala Lys Arg 1 5 <210> 14 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 14 Asp Ser Ser Gln Ser Pro Thr Pro Arg 1 5 <210> 15 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 15 Phe Pro Tyr Phe Lys His Ile Ala Lys 1 5 <210> 16 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 16 Gln Val Lys Val Glu Glu Pro Ser Arg 1 5 <210> 17 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 17 Tyr Ser Gly Leu Arg Ser Pro Thr Arg 1 5 <210> 18 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 18 Asp Gln Val Phe Lys Gln Gln Lys Arg 1 5 <210> 19 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 19 Ser Glu Leu Ala Arg His Ser Lys Arg 1 5 <210> 20 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 20 Lys Thr Glu Leu Pro Leu Gly Ala Arg 1 5 <210> twenty one <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> twenty one Phe Ser Glu Gly Pro Ser Thr Ser Arg 1 5 <210> twenty two <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> twenty two Glu Met Leu Val Ile Gln His Arg Glu Arg 1 5 10 <210> twenty three <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> twenty three Glu Thr Leu Gly Pro Lys Pro Ala Ala Arg 1 5 10 <210> twenty four <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> twenty four Asn Ser Leu Ser Asn Ile Gln Trp Leu Arg 1 5 10 <210> 25 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 25 Asn Leu Ser Leu His Asp Met Phe Val Arg 1 5 10 <210> 26 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 26 Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg 1 5 10 <210> 27 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 27 Met Leu Val Ile Gln His Arg Glu Arg Arg 1 5 10 <210> 28 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 28 Phe Trp Thr Ile His Pro Ser Ala Asn Arg 1 5 10 <210> 29 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 29 His Ser Lys Arg Val Arg Ile Ala Pro Lys 1 5 10 <210> 30 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 30 Ile Gln Phe Ala Ile Asn Ser Thr Glu Arg 1 5 10 <210> 31 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 31 His Phe Pro Tyr Phe Lys His Ile Ala Lys 1 5 10 <210> 32 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 32 Gln Thr Gln Thr Ser Tyr Asp Ala Lys Arg 1 5 10 <210> 33 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 33 Ser Thr Pro Ser Lys Ser Val Leu Pro Arg 1 5 10 <210> 34 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 34 Glu Leu Ala Arg His Ser Lys Arg Val Arg 1 5 10 <210> 35 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 35 Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg 1 5 10 <210> 36 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 36 Leu Phe Ser Glu Gly Pro Ser Thr Ser Arg 1 5 10 <210> 37 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 37 Ser Ser Glu Leu Ala Arg His Ser Lys Arg 1 5 10 <210> 38 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 38 Ala Ser Trp Gln Asn Ser Val Ser Glu Arg 1 5 10 <210> 39 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 39 Met Ser Ser Glu Leu Ala Arg His Ser Lys 1 5 10 <210> 40 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 40 Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg 1 5 10 <210> 41 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 41 Glu Ser Trp Arg Leu Thr Pro Pro Ala Lys 1 5 10 <210> 42 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 42 Trp Ile Glu Asp His Phe Pro Tyr Phe Lys 1 5 10 <210> 43 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 43 Arg Pro Lys Lys Ser Tyr Ser Gly Leu Arg 1 5 10 <210> 44 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 44 Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys 1 5 10 <210> 45 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 45 Ser Pro Arg Arg Pro Leu Ile Leu Lys Arg 1 5 10 <210> 46 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 46 Ala Ser Leu Met Ser Ser Glu Leu Ala Arg 1 5 10 <210> 47 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 47 Ser Ala Pro Pro Leu Glu Ser Pro Gln Arg 1 5 10 <210> 48 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 48 Val Ser Glu Arg Pro Pro Tyr Ser Tyr 1 5 <210> 49 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 49 Cys Val Asp Glu Pro Glu Leu Leu Phe 1 5 <210> 50 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 50 Ala Ser Asp Pro Leu Pro Asp Pro Leu 1 5 <210> 51 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 51 Leu Thr Glu Gly Leu Val Leu Asp Thr 1 5 <210> 52 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 52 Asn Ser Val Ser Glu Arg Pro Pro Tyr 1 5 <210> 53 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 53 Ser Ser Glu Pro Leu Asp Leu Ile Ser 1 5 <210> 54 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 54 Ser Ser Glu Leu Ala Arg His Ser Lys 1 5 <210> 55 <211> 9 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 55 Pro Thr Pro Arg Pro Lys Lys Ser Tyr 1 5 <210> 56 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 56 Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr 1 5 10 <210> 57 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 57 Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr 1 5 10 <210> 58 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 58 Trp Thr Ile His Pro Ser Ala Asn Arg Tyr 1 5 10 <210> 59 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 59 Ser Val Ser Glu Arg Pro Pro Tyr Ser Tyr 1 5 10 <210> 60 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 60 Ser Ser Glu Pro Leu Asp Leu Ile Ser Val 1 5 10 <210> 61 <211> 10 <212> PRT <213> Artificial sequence <220> <223> FOXM1-derived peptide <400> 61 Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg 1 5 10 <210> 62 <211> twenty two <212> DNA <213> Artificial sequence <220> <223> PCR primers for TCR analysis <400> 62 gtctaccagg cattcgcttc at 22 <210> 63 <211> twenty four <212> DNA <213> Artificial sequence <220> <223> PCR primers for TCR analysis <400> 63 tcagctggac cacagccgca gcgt 24 <210> 64 <211> twenty one <212> DNA <213> Artificial sequence <220> <223> PCR primers for TCR analysis <400> 64 tcagaaatcc tttctcttga c 21 <210> 65 <211> twenty four <212> DNA <213> Artificial sequence <220> <223> PCR primers for TCR analysis <400> 65 ctagcctctg gaatcctttc tctt 24 <210> 66 <211> 3506 <212> DNA <213> Homo sapiens <220> <221> CDS <222> (284)..(2530) <400> 66 tttcaaacag cggaacaaac tgaaagctcc ggtgccagac cccaccccg gcccggccc 60 gggaccccct cccctcccgg gatcccccgg ggttcccacc ccgcccgcac cgccggggac 120 ccggccggtc cggcgcgagc ccccgtccgg ggccctggct cggcccccag gttggaggag 180 cccggagccc gccttcggag ctacggccta acggcggcgg cgactgcagt ctggagggtc 240 cacacttgtg attctcaatg gagagtgaaa acgcagattc ata atg aaa act agc 295 Met Lys Thr Ser 1 ccc cgt cgg cca ctg att ctc aaa aga cgg agg ctg ccc ctt cct gtt 343 Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu Pro Leu Pro Val 5 10 15 20 caa aat gcc cca agt gaa aca tca gag gag gaa cct aag aga tcc cct 391 Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro Lys Arg Ser Pro 25 30 35 gcc caa cag gag tct aat caa gca gag gcc tcc aag gaa gtg gca gag 439 Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys Glu Val Ala Glu 40 45 50 tcc aac tct tgc aag ttt cca gct ggg atc aag att att aac cac ccc 487 Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile Ile Asn His Pro 55 60 65 acc atg ccc aac acg caa gta gtg gcc atc ccc aac aat gct aat att 535 Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn Asn Ala Asn Ile 70 75 80 cac agc atc atc aca gca ctg act gcc aag gga aaa gag agt ggc agt 583 His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys Glu Ser Gly Ser 85 90 95 100 agt ggg ccc aac aaa ttc atc ctc atc agc tgt ggg gga gcc cca act 631 Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly Gly Ala Pro Thr 105 110 115 cag cct cca gga ctc cgg cct caa acc caa acc agc tat gat gcc aaa 679 Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser Tyr Asp Ala Lys 120 125 130 agg aca gaa gtg acc ctg gag acc ttg gga cca aaa cct gca gct agg 727 Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys Pro Ala Ala Arg 135 140 145 gat gtg aat ctt cct aga cca cct gga gcc ctt tgc gag cag aaa cgg 775 Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys Glu Gln Lys Arg 150 155 160 gag acc tgt gca gat ggt gag gca gca ggc tgc act atc aac aat agc 823 Glu Thr Cys Ala Asp Gly Glu Ala Ala Gly Cys Thr Ile Asn Asn Ser 165 170 175 180 cta tcc aac atc cag tgg ctt cga aag atg agt tct gat gga ctg ggc 871 Leu Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser Asp Gly Leu Gly 185 190 195 tcc cgc agc atc aag caa gag atg gag gaa aag gag aat tgt cac ctg 919 Ser Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu Asn Cys His Leu 200 205 210 gag cag cga cag gtt aag gtt gag gag cct tcg aga cca tca gcg tcc 967 Glu Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg Pro Ser Ala Ser 215 220 225 tgg cag aac tct gtg tct gag cgg cca ccc tac tct tac atg gcc atg 1015 Trp Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser Tyr Met Ala Met 230 235 240 ata caa ttc gcc atc aac agc act gag agg aag cgc atg act ttg aaa 1063 Ile Gln Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg Met Thr Leu Lys 245 250 255 260 gac atc tat acg tgg att gag gac cac ttt ccc tac ttt aag cac att 1111 Asp Ile Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr Phe Lys His Ile 265 270 275 gcc aag cca ggc tgg aag aac tcc atc cgc cac aac ctt tcc ctg cac 1159 Ala Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn Leu Ser Leu His 280 285 290 gac atg ttt gtc cgg gag acg tct gcc aat ggc aag gtc tcc ttc tgg 1207 Asp Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys Val Ser Phe Trp 295 300 305 acc att cac ccc agt gcc aac cgc tac ttg aca ttg gac cag gtg ttt 1255 Thr Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu Asp Gln Val Phe 310 315 320 aag cag aaa cga ccg aat ca gag ctc cgc cgg aac atg acc atc 1303 Lys Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg Asn Met Thr Ile 325,330,335,340 aaa acc gaa ctc ccc ctg ggc gca cgg cgg aag atg aag ca cctg cta 1351 Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met Lys Pro Leu Leu 345 350 355 cca cgg gtc agc tca tac ctg gta cct atc cag ttc ccg gtg aac cag 1399 Pro Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe Pro Val Asn Gln 360 365 370 tca ctg gtg ttg cag ccc tcg gtg aag gtg ca ttg ccc ctg gcg gct 1447 Ser Val Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu Pro Leu Ala Ala 375 380 385 tcc ctc atg agc tca gag ctt gcc cgc cat agc aag cga gtc cgc att 1495 Ser Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys Arg Val Arg Ile 390,395,400 gcc ccc aag gtg ctg cta gct gag gag ggg ata gct cct ctt tct tct 1543 Ala Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala Pro Leu Ser Ser 405 410 415 420 gca gga cca ggg aaa gag gag aaa ctc ctg ttt gga gaa ggg ttt tct 1591 Ala Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly Glu Gly Phe Ser 425 430 435 cct ttg ctt cca gtt cag act atc aag gag gaa gaa atc cag cct ggg 1639 Pro Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu Ile Gln Pro Gly 440 445 450 gag gaa atg cca cac tta gcg aga ccc atc aaa gtg gag agc cct ccc 1687 Glu Glu Met Pro His Leu Ala Arg Pro Ile Lys Val Glu Ser Pro Pro 455 460 465 ttg gaa gag tgg ccc tcc ccg gcc cca tct ttc aaa gag gaa tca tct 1735 Leu Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys Glu Glu Ser Ser 470 475 480 cac tcc tgg gag gat tcg tcc caa tct ccc acc cca aga ccc aag aag 1783 His Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro Arg Pro Lys Lys 485 490 495 500 tcc tac agt ggg ctt agg tcc cca acc cgg tgt gtc tcg gaa atg ctt 1831 Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val Ser Glu Met Leu 505 510 515 gtg att caa cac agg gag agg agg gag agg agc cgg tct cgg agg aaa 1879 Val Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg Ser Arg Arg Lys 520 525 530 cag cat cta ctg cct ccc tgt gtg gat gag ccg gag ctg ctc ttc tca 1927 Gln His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu Leu Leu Phe Ser 535 540 545 gag ggg ccc agt act tcc cgc tgg gcc gca gag ctc ccg ttc cca gca 1975 Glu Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu Pro Phe Pro Ala 550 555 560 gac tcc tct gac cct gcc tcc cag ctc agc tac tcc cag gaa gtg gga 2023 Asp Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser Gln Glu Val Gly 565 570 575 580 gga cct ttt aag aca ccc att aag gaa acg ctg ccc atc tcc tcc acc 2071 Gly Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro Ile Ser Ser Thr 585 590 595 ccg agc aaa tct gtc ctc ccc aga acc cct gaa tcc tgg agg ctc acg 2119 Pro Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser Trp Arg Leu Thr 600 605 610 ccc cca gcc aaa gta ggg gga ctg gat ttc agc cca gta caa acc tcc 2167 Pro Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro Val Gln Thr Ser 615 620 625 cag ggt gcc tct gac ccc ttg cct gac ccc ctg ggg ctg atg gat ctc 2215 Gln Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly Leu Met Asp Leu 630 635 640 agc acc act ccc ttg caa agt gct ccc ccc ctt gaa tca ccg caa agg 2263 Ser Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu Ser Pro Gln Arg 645 650 655 660 ctc ctc agt tca gaa ccc tta gac ctc atc tcc gtc ccc ttt ggc aac 2311 Leu Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val Pro Phe Gly Asn 665 670 675 tct tct ccc tca gat ata gac gtc ccc aag cca ggc tcc ccg gag cca 2359 Ser Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly Ser Pro Glu Pro 680 685 690 cag gtt tct ggc ctt gca gcc aat cgt tct ctg aca gaa ggc ctg gtc 2407 Gln Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr Glu Gly Leu Val 695 700 705 ctg gac aca atg aat gac agc ctc agc aag atc ctg ctg gac atc agc 2455 Leu Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu Leu Asp Ile Ser 710 715 720 ttt cct ggc ctg gac gag gac cca ctg ggc cct gac aac atc aac tgg 2503 Phe Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp Asn Ile Asn Trp 725 730 735 740 tcc cag ttt att cct gag cta cag tag agccctgccc ttgcccctgt 2550 Ser Gln Phe Ile Pro Glu Leu Gln 745 gctcaagctg tccaccatcc cgggcactcc aaggctcagt gcaccccaag cctctgagtg aggacagcag gcagggactg ttctgctcct catagctccc tgctgcctga ttatgcaaaa gtagcagtca caccctagcc actgctggga ccttgtgttc cccaagagta tctgattcct ctgctgtccc tgccaggagc tgaagggtgg gacaacaaa ggcaatggtg aaaagagatt aggaaccccc cagcctgttt ccattctctg cccagcagtc tcttaccttc cctgatcttt 2850. gcagggtggt ccgtgtaat agtataaatt ctccaaatta tcctctaatt ataaatgtaa gcttatttcc ttagatcatt atccagagac tgccagaagg tgggtaggat gacctggggt 2970. ttcaattgac ttctgttcct tgcttttagt tttgatagaa gggaagacct gcagtgcacg gtttcttcca ggctgaggta cctggatctt gggttcttca ctgcagggac ccagacaagt ggatctgctt gccagagtcc tttttgcccc tccctgccac ctccccgtgt ttccaagtca 3150 gctttcctgc aagaagaat cctggttaaa aaagtctttt gtattgggtc aggagttgaa tttggggtgg gaggatggat gcaactgaag cagagtgtgg gtgcccagat gtgcgctatt 3270. agatgtttct ctgataatgt ccccaatcat accagggaga ctggcattga cgagaactca 3330 ggtggaggct tgagaaggcc gaaagggccc ctgacctgcc tggcttcctt agcttgcccc 3390 tcagctttgc aaagagccac cctaggcccc agctgaccgc atgggtgtga gccagcttga 3450 gaacactaac tactcaataa aagcgaaggt ggacatgaaa aaaaaaaaaa aaaaaa 3506 <210> 67 <211> 748 <212> PRT <213> Homo sapiens <400> 67 Met Lys Thr Ser Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu 1 5 10 15 Pro Leu Pro Val Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro 20 25 30 Lys Arg Ser Pro Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys 35 40 45 Glu Val Ala Glu Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile 50 55 60 Ile Asn His Pro Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn 65 70 75 80 Asn Ala Asn Ile His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys 85 90 95 Glu Ser Gly Ser Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly 100 105 110 Gly Ala Pro Thr Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser 115 120 125 Tyr Asp Ala Lys Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys 130 135 140 Pro Ala Ala Arg Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys 145 150 155 160 Glu Gln Lys Arg Glu Thr Cys Ala Asp Gly Glu Ala Ala Gly Cys Thr 165 170 175 Ile Asn Asn Ser Leu Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser 180 185 190 Asp Gly Leu Gly Ser Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu 195 200 205 Asn Cys His Leu Glu Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg 210 215 220 Pro Ser Ala Ser Trp Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser 225 230 235 240 Tyr Met Ala Met Ile Gln Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg 245 250 255 Met Thr Leu Lys Asp Ile Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr 260 265 270 Phe Lys His Ile Ala Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn 275 280 285 Leu Ser Leu His Asp Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys 290 295 300 Val Ser Phe Trp Thr Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu 305 310 315 320 Asp Gln Val Phe Lys Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg 325 330 335 Asn Met Thr Ile Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met 340 345 350 Lys Pro Leu Leu Pro Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe 355 360 365 Pro Val Asn Gln Ser Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu 370 375 380 Pro Leu Ala Ala Ser Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys 385 390 395 400 Arg Val Arg Ile Ala Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala 405 410 415 Pro Leu Ser Ser Ala Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly 420 425 430 Glu Gly Phe Ser Pro Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu 435 440 445 Ile Gln Pro Gly Glu Glu Met Pro His Leu Ala Arg Pro Ile Lys Val 450 455 460 Glu Ser Pro Pro Leu Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys 465 470 475 480 Glu Glu Ser Ser His Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro 485 490 495 Arg Pro Lys Lys Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val 500 505 510 Ser Glu Met Leu Val Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg 515 520 525 Ser Arg Arg Lys Gln His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu 530 535 540 Leu Leu Phe Ser Glu Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu 545 550 555 560 Pro Phe Pro Ala Asp Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser 565 570 575 Gln Glu Val Gly Gly Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro 580 585 590 Ile Ser Ser Thr Pro Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser 595 600 605 Trp Arg Leu Thr Pro Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro 610 615 620 Val Gln Thr Ser Gln Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly 625 630 635 640 Leu Met Asp Leu Ser Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu 645 650 655 Ser Pro Gln Arg Leu Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val 660 665 670 Pro Phe Gly Asn Ser Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly 675 680 685 Ser Pro Glu Pro Gln Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr 690 695 700 Glu Gly Leu Val Leu Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu 705 710 715 720 Leu Asp Ile Ser Phe Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp 725 730 735 Asn Ile Asn Trp Ser Gln Phe Ile Pro Glu Leu Gln 740 745 <210> 68 <211> 3506 <212> DNA <213> Homo sapiens <220> <221> CDS <222> (284)..(2530) <400> 68 tttcaaacag cggaacaaac tgaaagctcc ggtgccagac cccacccccg gccccggccc 60 gggaccccct cccctcccgg gatcccccgg ggttcccacc ccgcccgcac cgccggggac 120 ccggccggtc cggcgcgagc ccccgtccgg ggccctggct cggcccccag gttggaggag 180 cccggagccc gccttcggag ctacggccta acggcggcgg cgactgcagt ctggagggtc 240 cacacttgtg attctcaatg gagagtgaaa acgcagattc ata atg aaa act agc 295 Met Lys Thr Ser 1 ccc cgt cgg cca ctg att ctc aaa aga cgg agg ctg ccc ctt cct gtt 343 Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu Pro Leu Pro Val 5 10 15 20 caa aat gcc cca agt gaa aca tca gag gag gaa cct aag aga tcc cct 391 Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro Lys Arg Ser Pro 25 30 35 gcc caa cag gag tct aat caa gca gag gcc tcc aag gaa gtg gca gag 439 Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys Glu Val Ala Glu 40 45 50 tcc aac tct tgc aag ttt cca gct ggg atc aag att att aac cac ccc 487 Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile Ile Asn His Pro 55 60 65 acc atg ccc aac acg caa gta gtg gcc atc ccc aac aat gct aat att 535 Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn Asn Ala Asn Ile 70 75 80 cac agc atc atc aca gca ctg act gcc aag gga aaa gag agt ggc agt 583 His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys Glu Ser Gly Ser 85 90 95 100 agt ggg ccc aac aaa ttc atc ctc atc agc tgt ggg gga gcc cca act 631 Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly Gly Ala Pro Thr 105 110 115 cag cct cca gga ctc cgg cct caa acc caa acc agc tat gat gcc aaa 679 Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser Tyr Asp Ala Lys 120 125 130 agg aca gaa gtg acc ctg gag acc ttg gga cca aaa cct gca gct agg 727 Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys Pro Ala Ala Arg 135 140 145 gat gtg aat ctt cct aga cca cct gga gcc ctt tgc gag cag aaa cgg 775 Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys Glu Gln Lys Arg 150 155 160 gag acc tgt gat ggt gag gca gca ggc tgc act atc aac aat agc cta 823 Glu Thr Cys Asp Gly Glu Ala Ala Gly Cys Thr Ile Asn Asn Ser Leu 165 170 175 180 tcc aac atc cag tgg ctt cga aag atg agt tct gat gga ctg ggc tcc 871 Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser Asp Gly Leu Gly Ser 185 190 195 cgc agc atc aag caa gag atg gag gaa ag gag at tgt cac ctg gag 919 Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu Asn Cys His Leu Glu 200 205 210 cag cga cag gtt aag gtt gag gag cct tcg aga cca tca gcg tcc tgg 967 Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg Pro Ser Ala Ser Trp 215 220 225 cag aac tct gtg tct gag cgg cca ccc tac tct tac atg gcc atg ata 1015 Gln Asn Ser Val Ser Glu Arg Pro Tyr Ser Tyr Met Ala Met Ile 230 235 240 caa ttc gcc atc aac agc act gag agg aag cgc atg act ttg aaa gac 1063 Gln Phe Ala Ile Asn Served Thr Glu Arg Lys Arg Met Thr Leu Lys Asp 245 250 255 260 atc tat acg tgg att gag gac cac ttt ccc tac ttt aag cac att gcc 1111 They have Tyr Thr Trp They have Glu Asp they have Pro Tyr Phe Lys they have Ala 265 270 275 aag cca ggc tgg aag aac tcc atc cgc cac aac ctt tcc ctg cac gac 1159 Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn Leu Ser Leu His Asp 280 285 290 atg ttt gtc cgg gag acg tct gcc aat ggc aag gtc tcc ttc tgg acc 1207 Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys Val Ser Phe Trp Thr 295 300 305 att cac ccc agt gcc aac cgc tac ttg aca ttg gac cag gtg ttt aag 1255 Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu Asp Gln Val Phe Lys 310 315 320 cag cag cag aaa cga ccg aat cca gag ctc cgc cgg aac atg acc atc 1303 Gln Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg Asn Met Thr Ile 325 330 335 340 aaa acc gaa ctc ccc ctg ggc gca cgg cgg aag atg aag cca ctg cta 1351 Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met Lys Pro Leu Leu 345 350 355 cca cgg gtc agc tca tac ctg gta cct atc cag ttc ccg gtg aac cag 1399 Pro Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe Pro Val Asn Gln 360 365 370 tca ctg gtg ttg cag ccc tcg gtg aag gtg cca ttg ccc ctg gcg gct 1447 Ser Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu Pro Leu Ala Ala 375 380 385 tcc ctc atg agc tca gag ctt gcc cgc cat agc aag cga gtc cgc att 1495 Ser Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys Arg Val Arg Ile 390 395 400 gcc ccc aag gtg ctg cta gct gag gag ggg ata gct cct ctt tct tct 1543 Ala Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala Pro Leu Ser Ser 405 410 415 420 gca gga cca ggg aaa gag gag aaa ctc ctg ttt gga gaa ggg ttt tct 1591 Ala Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly Glu Gly Phe Ser 425 430 435 cct ttg ctt cca gtt cag act atc aag gag gaa gaa atc cag cct ggg 1639 Pro Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu Ile Gln Pro Gly 440 445 450 gag gaa atg cca cac tta gcg aga ccc atc aaa gtg gag agc cct ccc 1687 Glu Glu Met Pro His Leu Ala Arg Pro Ile Lys Val Glu Ser Pro Pro 455 460 465 ttg gaa gag tgg ccc tcc ccg gcc cca tct ttc aaa gag gaa tca tct 1735 Leu Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys Glu Glu Ser Ser 470 475 480 cac tcc tgg gag gat tcg tcc caa tct ccc acc cca aga ccc aag aag 1783 His Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro Arg Pro Lys Lys 485 490 495 500 tcc tac agt ggg ctt agg tcc cca acc cgg tgt gtc tcg gaa atg ctt 1831 Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val Ser Glu Met Leu 505 510 515 gtg att caa cac agg gag agg agg gag agg agc cgg tct cgg agg aaa 1879 Val Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg Ser Arg Arg Lys 520 525 530 cag cat cta ctg cct ccc tgt gtg gat gag ccg gag ctg ctc ttc tca 1927 Gln His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu Leu Leu Phe Ser 535 540 545 gag ggg ccc agt act tcc cgc tgg gcc gca gag ctc ccg ttc cca gca 1975 Glu Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu Pro Phe Pro Ala 550 555 560 gac tcc tct gac cct gcc tcc cag ctc agc tac tcc cag gaa gtg gga 2023 Asp Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser Gln Glu Val Gly 565 570 575 580 gga cct ttt aag aca ccc att aag gaa acg ctg ccc atc tcc tcc acc 2071 Gly Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro Ile Ser Ser Thr 585 590 595 ccg agc aaa tct gtc ctc ccc aga acc cct gaa tcc tgg agg ctc acg 2119 Pro Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser Trp Arg Leu Thr 600 605 610 ccc cca gcc aaa gta ggg gga ctg gat ttc agc cca gta caa acc tcc 2167 Pro Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro Val Gln Thr Ser 615 620 625 cag ggt gcc tct gac ccc ttg cct gac ccc ctg ggg ctg atg gat ctc 2215 Gln Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly Leu Met Asp Leu 630 635 640 agc acc act ccc ttg caa agt gct ccc ccc ctt gaa tca ccg caa agg 2263 Ser Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu Ser Pro Gln Arg 645 650 655 660 ctc ctc agt tca gaa ccc tta gac ctc atc tcc gtc ccc ttt ggc aac 2311 Leu Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val Pro Phe Gly Asn 665 670 675 tct tct ccc tca gat ata gac gtc ccc aag cca ggc tcc ccg gag cca 2359 Ser Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly Ser Pro Glu Pro 680 685 690 cag gtt tct ggc ctt gca gcc aat cgt tct ctg aca gaa ggc ctg gtc 2407 Gln Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr Glu Gly Leu Val 695 700 705 ctg gac aca atg aat gac agc ctc agc aag atc ctg ctg gac atc agc 2455 Leu Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu Leu Asp Ile Ser 710 715 720<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ttt cct ggc ctg gac gag gac cca ctg ggc cct gac aac atc aac tgg 2503<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> Phe Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp Asn Ile Asn Trp<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> 725 730 735 740<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> tcc cag ttt att cct gag cta cag tag agccctgccc ttgcccctgt 2550<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> Ser Gln Phe Ile Pro Glu Leu Gln<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> 745<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gctcaagctg tccaccatcc cgggcactcc aaggctcagt gcaccccaag cctctgagtg 2610<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> aggacagcag gcagggactg ttctgctcct catagctccc tgctgcctga ttatgcaaaa 2670<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gtagcagtca caccctagcc actgctggga ccttgtgttc cccaagagta tctgattcct 2730<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ctgctgtccc tgccaggagc tgaagggtgg gaacaacaaa ggcaatggtg aaaagagatt 2790<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> aggaaccccc cagcctgttt ccattctctg cccagcagtc tcttaccttc cctgatcttt 2850<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gcagggtggt ccgtgtaaat agtataaatt ctccaaatta tcctctaatt ataaatgtaa 2910<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gcttatttcc ttagatcatt atccagagac tgccagaagg tgggtaggat gacctggggt 2970<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ttcaattgac ttctgttcct tgcttttagt tttgatagaa gggaagacct gcagtgcacg 3030<h2 style=";text-align:left;direction:ltr"> gtttcttcca ggctgaggta cctggatctt gggttcttca ctgcagggac ccagacaagt 3090 ggatctgctt gccagagtcc tttttgcccc tccctgccac ctccccgtgt ttccaagtca 3150 gctttcctgc aagaagaaat cctggttaaa aaagtctttt gtattgggtc aggagttgaa 3210 tttggggtgg gaggatggat gcaactgaag cagagtgtgg gtgcccagat gtgcgctatt 3270 agatgtttct ctgataatgt ccccaatcat accagggaga ctggcattga cgagaactca 3330 ggtggaggct tgagaaggcc gaaagggccc ctgacctgcc tggcttcctt agcttgcccc 3390 tcagctttgc aaagagccac cctaggcccc agctgaccgc atgggtgtga gccagcttga 3450 gaacactaac tactcaataa aagcgaaggt ggacatgaaa aaaaaaaaaa aaaaaa 3506 <210> 69 <211> 748 <212> PRT <213> Homo sapiens <400> 69 Met Lys Thr Ser Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu 1 5 10 15 Pro Leu Pro Val Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro 20 25 30 Lys Arg Ser Pro Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys 35 40 45 Glu Val Ala Glu Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile 50 55 60 Ile Asn His Pro Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn 65 70 75 80 Asn Ala Asn Ile His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys 85 90 95 Glu Ser Gly Ser Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly 100 105 110 Gly Ala Pro Thr Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser 115 120 125 Tyr Asp Ala Lys Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys 130 135 140 Pro Ala Ala Arg Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys 145 150 155 160 Glu Gln Lys Arg Glu Thr Cys Asp Gly Glu Ala Ala Gly Cys Thr Ile 165 170 175 Asn Asn Ser Leu Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser Asp 180 185 190 Gly Leu Gly Ser Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu Asn 195 200 205 Cys His Leu Glu Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg Pro 210 215 220 Ser Ala Ser Trp Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser Tyr 225 230 235 240 Met Ala Met Ile Gln Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg Met 245 250 255 Thr Leu Lys Asp Ile Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr Phe 260 265 270 Lys His Ile Ala Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn Leu 275 280 285 Ser Leu His Asp Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys Val 290 295 300 Ser Phe Trp Thr Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu Asp 305 310 315 320 Gln Val Phe Lys Gln Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg 325 330 335 Asn Met Thr Ile Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met 340 345 350 Lys Pro Leu Leu Pro Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe 355 360 365 Pro Val Asn Gln Ser Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu 370 375 380 Pro Leu Ala Ala Ser Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys 385 390 395 400 Arg Val Arg Ile Ala Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala 405 410 415 Pro Leu Ser Ser Ala Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly 420 425 430 Glu Gly Phe Ser Pro Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu 435 440 445 Ile Gln Pro Gly Glu Glu Met Pro His Leu Ala Arg Pro Ile Lys Val 450 455 460 Glu Ser Pro Pro Leu Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys 465 470 475 480 Glu Glu Ser Ser His Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro 485 490 495 Arg Pro Lys Lys Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val 500 505 510 Ser Glu Met Leu Val Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg 515 520 525 Ser Arg Arg Lys Gln His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu 530 535 540 Leu Leu Phe Ser Glu Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu 545 550 555 560 Pro Phe Pro Ala Asp Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser 565 570 575 Gln Glu Val Gly Gly Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro 580 585 590 Ile Ser Ser Thr Pro Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser 595 600 605 Trp Arg Leu Thr Pro Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro 610 615 620 Val Gln Thr Ser Gln Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly 625 630 635 640 Leu Met Asp Leu Ser Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu 645 650 655 Ser Pro Gln Arg Leu Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val 660 665 670 Pro Phe Gly Asn Ser Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly 675 680 685 Ser Pro Glu Pro Gln Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr 690 695 700 Glu Gly Leu Val Leu Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu 705 710 715 720 Leu Asp Ile Ser Phe Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp 725 730 735 Asn Ile Asn Trp Ser Gln Phe Ile Pro Glu Leu Gln 740 745 <210> 70 <211> 3503 <212> DNA <213> Homo sapiens <220> <221> CDS <222> (284)..(2527) <400> 70 tttcaaacag cggaacaaac tgaaagctcc ggtgccagac cccacccccg gccccggccc 60 gggaccccct cccctcccgg gatcccccgg ggttcccacc ccgcccgcac cgccggggac 120 ccggccggtc cggcgcgagc ccccgtccgg ggccctggct cggcccccag gttggaggag 180 cccggagccc gccttcggag ctacggccta acggcggcgg cgactgcagt ctggaggttc 240 cacacttgtg attctcaatg gagagtgaaa acgcagatc ata atg aaa act agc 295 Met Lys Thr Ser 1 ccc cgt cgg cca ctg att ctc aaa aga cgg agg ctg ccc ctt cct gtt 343 Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Leu Pro Leu Pro Val 5 10 15 20 caa aat gcc cca agt gaa aca tca gag gag gaa cct aag aga tcc cct 391 Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro Lys Arg Ser Pro 25 30 35 gcc caa cag gag tct aat caa gca gag gcc tcc aag gaa gtg gca gag 439 Only Gln Gln Glu Ser Asn Gln Ala Glu Only Ser Lys Glu Val Ala Glu 40 45 50 tcc aac tct tgc aag ttt cca gct ggg atc aag att aac cac ccc 487 Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile Ile Asn His Pro 55 60 65 acc atg ccc aac acg caa gta gtg gcc atc ccc aac aat gct aat att 535 Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn Asn Ala Asn Ile 70 75 80 cac agc atc atc aca gca ctg act gcc aag gga aaa gag agt ggc agt 583 His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys Glu Ser Gly Ser 85 90 95 100 agt ggg ccc aac aaa ttc atc ctc atc agc tgt ggg gga gcc cca act 631 Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly Gly Ala Pro Thr 105 110 115 cag cct cca gga ctc cgg cct caa acc caa acc agc tat gat gcc aaa 679 Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser Tyr Asp Ala Lys 120 125 130 agg aca gaa gtg acc ctg gag acc ttg gga cca aaa cct gca gct agg 727 Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys Pro Ala Ala Arg 135 140 145 gat gtg aat ctt cct aga cca cct gga gcc ctt tgc gag cag aaa cgg 775 Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys Glu Gln Lys Arg 150 155 160 gag acc tgt gat ggt gag gca gca ggc tgc act atc aac aat agc cta 823 Glu Thr Cys Asp Gly Glu Ala Ala Gly Cys Thr Ile Asn Asn Ser Leu 165 170 175 180 tcc aac atc cag tgg ctt cga aag atg agt tct gat gga ctg ggc tcc 871 Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser Asp Gly Leu Gly Ser 185 190 195 cgc agc atc aag caa gag atg gag gaa aag gag aat tgt cac ctg gag 919 Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu Asn Cys His Leu Glu 200 205 210 cag cga cag gtt aag gtt gag gag cct tcg aga cca tca gcg tcc tgg 967 Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg Pro Ser Ala Ser Trp 215 220 225 cag aac tct gtg tct gag cgg cca ccc tac tct tac atg gcc atg ata 1015 Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser Tyr Met Ala Met Ile 230 235 240 caa ttc gcc atc aac agc act gag agg aag cgc atg act ttg aaa gac 1063 Gln Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg Met Thr Leu Lys Asp 245 250 255 260 atc tat acg tgg att gag gac cac ttt ccc tac ttt aag cac att gcc 1111 Ile Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr Phe Lys His Ile Ala 265 270 275 aag cca ggc tgg aag aac tcc atc cgc cac aac ctt tcc ctg cac gac 1159 Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn Leu Ser Leu His Asp 280 285 290 atg ttt gtc cgg gag acg tct gcc aat ggc aag gtc tcc ttc tgg acc 1207 Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys Val Ser Phe Trp Thr 295 300 305 att cac ccc agt gcc aac cgc tac ttg aca ttg gac cag gtg ttt aag 1255 Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu Asp Gln Val Phe Lys 310 315 320 cag cag aaa cga ccg aat cca gag ctc cgc cgg aac atg acc atc aaa 1303 Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg Asn Met Thr Ile Lys 325 330 335 340 acc gaa ctc ccc ctg ggc gca cgg cgg aag atg aag cca ctg cta cca 1351 Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met Lys Pro Leu Leu Pro 345 350 355 cgg gtc agc tca tac ctg gta cct atc cag ttc ccg gtg aac cag tca 1399 Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe Pro Val Asn Gln Ser 360 365 370 ctg gtg ttg cag ccc tcg gtg aag gtg cca ttg ccc ctg gcg gct tcc 1447 Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu Pro Leu Ala Ala Ser 375 380 385 ctc atg agc tca gag ctt gcc cgc cat agc aag cga gtc cgc att gcc 1495 Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys Arg Val Arg Ile Ala 390 395 400 ccc aag gtg ctg cta gct gag gag ggg ata gct cct ctt tct tct gca 1543 Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala Pro Leu Ser Ser Ala 405 410 415 420 gga cca ggg aaa gag gag aaa ctc ctg ttt gga gaa ggg ttt tct cct 1591 Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly Glu Gly Phe Ser Pro 425 430 435 ttg ctt cca gtt cag act atc aag gag gaa gaa atc cag cct ggg gag 1639 Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu Ile Gln Pro Gly Glu 440 445 450 gaa atg cca cac tta gcg aga ccc atc aaa gtg gag agc cct ccc ttg 1687 Glu Met Pro His Leu Ala Arg Pro Ile Lys Val Glu Ser Pro Pro Leu 455 460 465 gaa gag tgg ccc tcc ccg gcc cca tct ttc aaa gag gaa tca tct cac 1735 Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys Glu Glu Ser Ser His 470 475 480 tcc tgg gag gat tcg tcc caa tct ccc acc cca aga ccc aag aag tcc 1783 Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro Arg Pro Lys Lys Ser 485 490 495 500 tac agt ggg ctt agg tcc cca acc cgg tgt gtc tcg gaa atg ctt gtg 1831 Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val Ser Glu Met Leu Val 505 510 515 att caa cac agg gag agg agg gag agg agc cgg tct cgg agg aaa cag 1879 Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg Ser Arg Arg Lys Gln 520 525 530 cat cta ctg cct ccc tgt gtg gat gag ccg gag ctg ctc ttc tca gag 1927 His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu Leu Leu Phe Ser Glu 535 540 545 ggg ccc agt act tcc cgc tgg gcc gca gag ctc ccg ttc cca gca gac 1975 Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu Pro Phe Pro Ala Asp 550 555 560 tcc tct gac cct gcc tcc cag ctc agc tac tcc cag gaa gtg gga gga 2023 Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser Gln Glu Val Gly Gly 565 570 575 580 cct ttt aag aca ccc att aag gaa acg ctg ccc atc tcc tcc acc ccg 2071 Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro Ile Ser Ser Thr Pro 585 590 595 agc aaa tct gtc ctc ccc aga acc cct gaa tcc tgg agg ctc acg ccc 2119 Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser Trp Arg Leu Thr Pro 600 605 610 cca gcc aaa gta ggg gga ctg gat ttc agc cca gta caa acc tcc cag 2167 Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro Val Gln Thr Ser Gln 615 620 625 ggt gcc tct gac ccc ttg cct gac ccc ctg ggg ctg atg gat ctc agc 2215 Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly Leu Met Asp Leu Ser 630 635 640 acc act ccc ttg caa agt gct ccc ccc ctt gaa tca ccg caa agg ctc 2263 Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu Ser Pro Gln Arg Leu 645 650 655 660 ctc agt tca gaa ccc tta gac ctc atc tcc gtc ccc ttt ggc aac tct 2311 Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val Pro Phe Gly Asn Ser 665 670 675 tct ccc tca gat ata gac gtc ccc aag cca ggc tcc ccg gag cca cag 2359 Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly Ser Pro Glu Pro Gln 680 685 690 gtt tct ggc ctt gca gcc aat cgt tct ctg aca gaa ggc ctg gtc ctg 2407 Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr Glu Gly Leu Val Leu 695 700 705 gac aca atg aat gac agc ctc agc aag atc ctg ctg gac atc agc ttt 2455 Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu Leu Asp Ile Ser Phe 710 715 720 cct ggc ctg gac gag gac cca ctg ggc cct gac aac atc aac tgg tcc 2503 Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp Asn Ile Asn Trp Ser 725 730 735 740 cag ttt att cct gag cta cag tag agccctgccc ttgcccctgt gctcaagctg 2557 Gln Phe Ile Pro Glu Leu Gln 745 tccaccatcc cgggcactcc aaggctcagt gcaccccaag cctctgagtg aggacagcag 2617 gcagggactg ttctgctcct catagctccc tgctgcctga ttatgcaaaa gtagcagtca 2677 caccctagcc actgctggga ccttgtgttc cccaagagta tctgattcct ctgctgtccc 2737 tgccaggagc tgaagggtgg gaacaacaaa ggcaatggtg aaaagagatt aggaaccccc 2797 cagcctgttt ccattcttg cccagcagtc tcttaccttc cctgatcttt gcagggtggt 2857 ccgtgtaaat agtataaatt ctccaaatta tcctctaatt ataaatgtaa gcttatttcc 2917 ttagatcatt atccagagac tgccagaagg tgggtaggat gacctggggt ttcaattgac 2977 ttctgttcct tgcttttagt tttgatagaa gggaagacct gcagtgcacg gttctcttcca 3037 ggctgaggta cctggatctt gggttcttca ctgcagggac ccagacaagt ggatctgctt 3097 gccagagtcc ttttgcccc tccctgccac ctccccgtgt ttccaagtca gctttcctgc 3157 aagaagaaat cctggttaaa aaagtcttt gtattgggtc aggagttgaa tttggggtgg 3217 gaggatggat gcaactgaag cagagtgtgg gtgcccagat gtgcgctatt agatgtttct 3277 ctgataatgt ccccaatcat accagggaga ctggcattga cgagaactca ggtggaggct 3337 tgagaaggcc gaaagggccc ctgacctgcc tggcttcctt agcttgcccc tcagctttgc 3397 aaagagccac cctaggcccc agctgaccgc atgggtgtga gccagcttga gaacactaac 3457 tactcaataa aagcgaaggt ggacatgaaa aaaaaaaaaa aaaaaa 3503 <210> 71 <211> 747 <212> PRT <213> Homo sapiens <400> 71 Met Lys Thr Ser Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu 1 5 10 15 Pro Leu Pro Val Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro 20 25 30 Lys Arg Ser Pro Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys 35 40 45 Glu Val Ala Glu Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile 50 55 60 Ile Asn His Pro Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn 65 70 75 80 Asn Ala Asn Ile His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys 85 90 95 Glu Ser Gly Ser Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly 100 105 110 Gly Ala Pro Thr Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser 115 120 125 Tyr Asp Ala Lys Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys 130 135 140 Pro Ala Ala Arg Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys 145 150 155 160 Glu Gln Lys Arg Glu Thr Cys Asp Gly Glu Ala Ala Gly Cys Thr Ile 165 170 175 Asn Asn Ser Leu Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser Asp 180 185 190 Gly Leu Gly Ser Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu Asn 195 200 205 Cys His Leu Glu Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg Pro 210 215 220 Ser Ala Ser Trp Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser Tyr 225 230 235 240 Met Ala Met Ile Gln Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg Met 245 250 255 Thr Leu Lys Asp Ile Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr Phe 260 265 270 Lys His Ile Ala Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn Leu 275 280 285 Ser Leu His Asp Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys Val 290 295 300 Ser Phe Trp Thr Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu Asp 305 310 315 320 Gln Val Phe Lys Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg Asn 325 330 335 Met Thr Ile Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met Lys 340 345 350 Pro Leu Leu Pro Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe Pro 355 360 365 Val Asn Gln Ser Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu Pro 370 375 380 Leu Ala Ala Ser Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys Arg 385 390 395 400 Val Arg Ile Ala Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala Pro 405 410 415 Leu Ser Ser Ala Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly Glu 420 425 430 Gly Phe Ser Pro Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu Ile 435 440 445 Gln Pro Gly Glu Glu Met Pro His Leu Ala Arg Pro Ile Lys Val Glu 450 455 460 Ser Pro Pro Leu Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys Glu 465 470 475 480 Glu Ser Ser His Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro Arg 485 490 495 Pro Lys Lys Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val Ser 500 505 510 Glu Met Leu Val Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg Ser 515 520 525 Arg Arg Lys Gln His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu Leu 530 535 540 Leu Phe Ser Glu Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu Pro 545 550 555 560 Phe Pro Ala Asp Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser Gln 565 570 575 Glu Val Gly Gly Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro Ile 580 585 590 Ser Ser Thr Pro Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser Trp 595 600 605 Arg Leu Thr Pro Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro Val 610 615 620 Gln Thr Ser Gln Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly Leu 625 630 635 640 Met Asp Leu Ser Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu Ser 645 650 655 Pro Gln Arg Leu Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val Pro 660 665 670 Phe Gly Asn Ser Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly Ser 675 680 685 Pro Glu Pro Gln Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr Glu 690 695 700 Gly Leu Val Leu Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu Leu 705 710 715 720 Asp Ile Ser Phe Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp Asn 725 730 735 Ile Asn Trp Ser Gln Phe Ile Pro Glu Leu Gln 740 745 <210> 72 <211> 3551 <212> DNA <213> Homo sapiens <220> <221> CDS <222> (284)..(2575) <400> 72 tttcaaacag cggaacaaac tgaaagctcc ggtgccagac cccaccccg gcccggccc 60 gggaccccct cccctcccgg gatcccccgg ggttcccacc ccgcccgcac cgccggggac 120 ccggccggtc cggcgcgagc ccccgtccgg ggccctggct cggcccccag gttggaggag 180 cccggagccc gccttcggag ctacggccta acggcggcgg cgactgcagt ctggagggtc 240 cacacttgtg attctcaatg gagagtgaaa acgcagattc ata atg aaa act agc 295 Met Lys Thr Ser 1 ccc cgt cgg cca ctg att ctc aaa aga cgg agg ctg ccc ctt cct gtt 343 Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu Pro Leu Pro Val 5 10 15 20 caa aat gcc cca agt gaa aca tca gag gag gaa cct aag aga tcc cct 391 Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro Lys Arg Ser Pro 25 30 35 gcc caa cag gag tct aat caa gca gag gcc tcc aag gaa gtg gca gag 439 Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys Glu Val Ala Glu 40 45 50 tcc aac tct tgc aag ttt cca gct ggg atc aag att att aac cac ccc 487 Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile Ile Asn His Pro 55 60 65 acc atg ccc aac acg caa gta gtg gcc atc ccc aac aat gct aat att 535 Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn Asn Ala Asn Ile 70 75 80 cac agc atc atc aca gca ctg act gcc aag gga aaa gag agt ggc agt 583 His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys Glu Ser Gly Ser 85 90 95 100 agt ggg ccc aac aaa ttc atc ctc atc agc tgt ggg gga gcc cca act 631 Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly Gly Ala Pro Thr 105 110 115 cag cct cca gga ctc cgg cct caa acc caa acc agc tat gat gcc aaa 679 Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser Tyr Asp Ala Lys 120 125 130 agg aca gaa gtg acc ctg gag acc ttg gga cca aaa cct gca gct agg 727 Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys Pro Ala Ala Arg 135 140 145 gat gtg aat ctt cct aga cca cct gga gcc ctt tgc gag cag aaa cgg 775 Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys Glu Gln Lys Arg 150 155 160 gag acc tgt gca gat ggt gag gca gca ggc tgc act atc aac aat agc 823 Glu Thr Cys Ala Asp Gly Glu Ala Ala Gly Cys Thr Ile Asn Asn Ser 165 170 175 180 cta tcc aac atc cag tgg ctt cga aag atg agt tct gat gga ctg ggc 871 Leu Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser Asp Gly Leu Gly 185 190 195 tcc cgc agc atc aag caa gag atg gag gaa aag gag aat tgt cac ctg 919 Ser Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu Asn Cys His Leu 200 205 210 gag cag cga cag gtt aag gtt gag gag cct tcg aga cca tca gcg tcc 967 Glu Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg Pro Ser Ala Ser 215 220 225 tgg cag aac tct gtg tct gag cgg ccca ccc tac tct tac atg gcc atg 1015 Trp Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser Tyr Met Ala Met 230 235 240 ata caa ttc gcc atc aac agc act gag agg agg cgc atg act ttg aaa 1063 Gln Phe Ala Is Asn Served Thr Glu Arg Lys Arg Met Thr Leu Lys 245 250 255 260 gac atc tat acg tgg att gag gac cac ttt ccc tac ttt aag cac att 1111 Asp Ile Tyr Thr Trp Ile Glu Asp His Pro Tyr Phe Lys His Ile 265 270 275 gcc aag cca ggc tgg aag aac tc atc cgc cac aac ctt tcc ctg cac 1159 Only Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn Leu Ser Leu His 280 285 290 gac atg ttt gtc cgg gag acg tct gcc aat ggc aag gtc tcc ttc tgg 1207 Asp Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys Val Ser Phe Trp 295 300 305 acc att cac ccc agt gcc aac cgc tac ttg aca ttg gac cag gtg ttt 1255 Thr Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu Asp Gln Val Phe 310 315 320 aag cca ctg gac cca ggg tct cca caa ttg ccc gag cac ttg gaa tca 1303 Lys Pro Leu Asp Pro Gly Ser Pro Gln Leu Pro Glu His Leu Glu Ser 325 330 335 340 cag cag aaa cga ccg aat cca gag ctc cgc cgg aac atg acc atc aaa 1351 Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg Asn Met Thr Ile Lys 345 350 355 acc gaa ctc ccc ctg ggc gca cgg cgg aag atg aag cca ctg cta cca 1399 Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met Lys Pro Leu Leu Pro 360 365 370 cgg gtc agc tca tac ctg gta cct atc cag ttc ccg gtg aac cag tca 1447 Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe Pro Val Asn Gln Ser 375 380 385 ctg gtg ttg cag ccc tcg gtg aag gtg cca ttg ccc ctg gcg gct tcc 1495 Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu Pro Leu Ala Ala Ser 390 395 400 ctc atg agc tca gag ctt gcc cgc cat agc aag cga gtc cgc att gcc 1543 Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys Arg Val Arg Ile Ala 405 410 415 420 ccc aag gtg ctg cta gct gag gag ggg ata gct cct ctt tct tct gca 1591 Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala Pro Leu Ser Ser Ala 425 430 435 gga cca ggg aaa gag gag aaa ctc ctg ttt gga gaa ggg ttt tct cct 1639 Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly Glu Gly Phe Ser Pro 440 445 450 ttg ctt cca gtt cag act atc aag gag gaa gaa atc cag cct ggg gag 1687 Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu Ile Gln Pro Gly Glu 455 460 465 gaa atg cca cac tta gcg aga ccc atc aaa gtg gag agc cct ccc ttg 1735 Glu Met Pro His Leu Ala Arg Pro Ile Lys Val Glu Ser Pro Pro Leu 470 475 480 gaa gag tgg ccc tcc ccg gcc cca tct ttc aaa gag gaa tca tct cac 1783 Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys Glu Glu Ser Ser His 485 490 495 500 tcc tgg gag gat tcg tcc caa tct ccc acc cca aga ccc aag aag tcc 1831 Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro Arg Pro Lys Lys Ser 505 510 515 tac agt ggg ctt agg tcc cca acc cgg tgt gtc tcg gaa atg ctt gtg 1879 Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val Ser Glu Met Leu Val 520 525 530 att caa cac agg gag agg agg gag agg agc cgg tct cgg agg aaa cag 1927 Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg Ser Arg Arg Lys Gln 535 540 545 cat cta ctg cct ccc tgt gtg gat gag ccg gag ctg ctc ttc tca gag 1975 His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu Leu Leu Phe Ser Glu 550 555 560 ggg ccc agt act tcc cgc tgg gcc gca gag ctc ccg ttc cca gca gac 2023 Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu Pro Phe Pro Ala Asp 565 570 575 580 tcc tct gac cct gcc tcc cag ctc agc tac tcc cag gaa gtg gga gga 2071 Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser Gln Glu Val Gly Gly 585 590 595 cct ttt aag aca ccc att aag gaa acg ctg ccc atc tcc tcc acc ccg 2119 Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro Ile Ser Ser Thr Pro 600 605 610 agc aaa tct gtc ctc ccc aga acc cct gaa tcc tgg agg ctc acg ccc 2167 Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser Trp Arg Leu Thr Pro 615 620 625 cca gcc aaa gta ggg gga ctg gat ttc agc cca gta caa acc tcc cag 2215 Pro Ala Lys Val Gly Gly Leu Asp Phe Ser Pro Val Gln Thr Ser Gln 630 635 640 ggt gcc tct gac ccc ttg cct gac ccc ctg ggg ctg atg gat ctc agc 2263 Gly Ala Ser Asp Pro Leu Pro Asp Pro Leu Gly Leu Met Asp Leu Ser 645 650 655 660 acc act ccc ttg caa agt gct ccc ccc ctt gaa tca ccg caa agg ctc 2311 Thr Thr Pro Leu Gln Ser Ala Pro Pro Leu Glu Ser Pro Gln Arg Leu 665 670 675 ctc agt tca gaa ccc tta gac ctc atc tcc gtc ccc ttt ggc aac tct 2359 Leu Ser Ser Glu Pro Leu Asp Leu Ile Ser Val Pro Phe Gly Asn Ser 680 685 690 tct ccc tca gat ata gac gtc ccc aag cca ggc tcc ccg gag cca cag 2407 Ser Pro Ser Asp Ile Asp Val Pro Lys Pro Gly Ser Pro Glu Pro Gln 695 700 705 gtt tct ggc ctt gca gcc aat cgt tct ctg aca gaa ggc ctg gtc ctg 2455 Val Ser Gly Leu Ala Ala Asn Arg Ser Leu Thr Glu Gly Leu Val Leu 710 715 720 gac aca atg aat gac agc ctc agc aag atc ctg ctg gac atc agc ttt 2503 Asp Thr Met Asn Asp Ser Leu Ser Lys Ile Leu Leu Asp Ile Ser Phe 725 730 735 740 cct ggc ctg gac gag gac cca ctg ggc cct gac aac atc aac tgg tcc 2551 <h2 style=";text-align:left;direction:ltr">Pro Gly Leu Asp Glu Asp Pro Leu Gly Pro Asp Asn Ile Asn Trp Ser<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> 745 750 755<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> cag ttt att cct gag cta cag tag agccctgccc ttgcccctgt gctcaagctg 2605<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> Gln Phe Ile Pro Glu Leu Gln<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> 760<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> tccaccatcc cgggcactcc aaggctcagt gcaccccaag cctctgagtg aggacagcag 2665<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gcagggactg ttctgctcct catagctccc tgctgcctga ttatgcaaaa gtagcagtca 2725<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> caccctagcc actgctggga ccttgtgttc cccaagagta tctgattcct ctgctgtccc 2785<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> tgccaggagc tgaagggtgg gaacaacaaa ggcaatggtg aaaagagatt aggaaccccc 2845<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> cagcctgttt ccattctctg cccagcagtc tcttaccttc cctgatcttt gcagggtggt 2905<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ccgtgtaaat agtataaatt ctccaaatta tcctctaatt ataaatgtaa gcttatttcc 2965<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ttagatcatt atccagagac tgccagaagg tgggtaggat gacctggggt ttcaattgac 3025<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ttctgttcct tgcttttagt tttgatagaa gggaagacct gcagtgcacg gtttcttcca 3085<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ggctgaggta cctggatctt gggttcttca ctgcagggac ccagacaagt ggatctgctt 3145<h2 style=";text-align:left;direction:ltr"> gccagagtcc tttttgcccc tccctgccac ctccccgtgt ttccaagtca gctttcctgc 3205 aagaagaaat cctggttaaa aaagtctttt gtattgggtc aggagttgaa tttggggtgg 3265 gaggatggat gcaactgaag cagagtgtgg gtgcccagat gtgcgctatt agatgtttct 3325 ctgataatgt ccccaatcat accagggaga ctggcattga cgagaactca ggtggaggct 3385 tgagaaggcc gaaagggccc ctgacctgcc tggcttcctt agcttgcccc tcagctttgc 3445 aaagagccac cctaggcccc agctgaccgc atgggtgtga gccagcttga gaacactaac 3505 tactcaataa aagcgaaggt ggacatgaaa aaaaaaaaaa aaaaaa 3551 <210> 73 <211> 763 <212> PRT <213> Homo sapiens <400> 73 Met Lys Thr Ser Pro Arg Arg Pro Leu Ile Leu Lys Arg Arg Arg Leu 1 5 10 15 Pro Leu Pro Val Gln Asn Ala Pro Ser Glu Thr Ser Glu Glu Glu Pro 20 25 30 Lys Arg Ser Pro Ala Gln Gln Glu Ser Asn Gln Ala Glu Ala Ser Lys 35 40 45 Glu Val Ala Glu Ser Asn Ser Cys Lys Phe Pro Ala Gly Ile Lys Ile 50 55 60 Ile Asn His Pro Thr Met Pro Asn Thr Gln Val Val Ala Ile Pro Asn 65 70 75 80 Asn Ala Asn Ile His Ser Ile Ile Thr Ala Leu Thr Ala Lys Gly Lys 85 90 95 Glu Ser Gly Ser Ser Gly Pro Asn Lys Phe Ile Leu Ile Ser Cys Gly 100 105 110 Gly Ala Pro Thr Gln Pro Pro Gly Leu Arg Pro Gln Thr Gln Thr Ser 115 120 125 Tyr Asp Ala Lys Arg Thr Glu Val Thr Leu Glu Thr Leu Gly Pro Lys 130 135 140 Pro Ala Ala Arg Asp Val Asn Leu Pro Arg Pro Pro Gly Ala Leu Cys 145 150 155 160 Glu Gln Lys Arg Glu Thr Cys Ala Asp Gly Glu Ala Ala Gly Cys Thr 165 170 175 Ile Asn Asn Ser Leu Ser Asn Ile Gln Trp Leu Arg Lys Met Ser Ser 180 185 190 Asp Gly Leu Gly Ser Arg Ser Ile Lys Gln Glu Met Glu Glu Lys Glu 195 200 205 Asn Cys His Leu Glu Gln Arg Gln Val Lys Val Glu Glu Pro Ser Arg 210 215 220 Pro Ser Ala Ser Trp Gln Asn Ser Val Ser Glu Arg Pro Pro Tyr Ser 225 230 235 240 Tyr Met Ala Met Ile Gln Phe Ala Ile Asn Ser Thr Glu Arg Lys Arg 245 250 255 Met Thr Leu Lys Asp Ile Tyr Thr Trp Ile Glu Asp His Phe Pro Tyr 260 265 270 Phe Lys His Ile Ala Lys Pro Gly Trp Lys Asn Ser Ile Arg His Asn 275 280 285 Leu Ser Leu His Asp Met Phe Val Arg Glu Thr Ser Ala Asn Gly Lys 290 295 300 Val Ser Phe Trp Thr Ile His Pro Ser Ala Asn Arg Tyr Leu Thr Leu 305 310 315 320 Asp Gln Val Phe Lys Pro Leu Asp Pro Gly Ser Pro Gln Leu Pro Glu 325 330 335 His Leu Glu Ser Gln Gln Lys Arg Pro Asn Pro Glu Leu Arg Arg Asn 340 345 350 Met Thr Ile Lys Thr Glu Leu Pro Leu Gly Ala Arg Arg Lys Met Lys 355 360 365 Pro Leu Leu Pro Arg Val Ser Ser Tyr Leu Val Pro Ile Gln Phe Pro 370 375 380 Val Asn Gln Ser Leu Val Leu Gln Pro Ser Val Lys Val Pro Leu Pro 385 390 395 400 Leu Ala Ala Ser Leu Met Ser Ser Glu Leu Ala Arg His Ser Lys Arg 405 410 415 Val Arg Ile Ala Pro Lys Val Leu Leu Ala Glu Glu Gly Ile Ala Pro 420 425 430 Leu Ser Ser Ala Gly Pro Gly Lys Glu Glu Lys Leu Leu Phe Gly Glu 435 440 445 Gly Phe Ser Pro Leu Leu Pro Val Gln Thr Ile Lys Glu Glu Glu Ile 450 455 460 Gln Pro Gly Glu Glu Met Pro His Leu Ala Arg Pro Ile Lys Val Glu 465 470 475 480 Ser Pro Pro Leu Glu Glu Trp Pro Ser Pro Ala Pro Ser Phe Lys Glu 485 490 495 Glu Ser Ser His Ser Trp Glu Asp Ser Ser Gln Ser Pro Thr Pro Arg 500 505 510 Pro Lys Lys Ser Tyr Ser Gly Leu Arg Ser Pro Thr Arg Cys Val Ser 515 520 525 Glu Met Leu Val Ile Gln His Arg Glu Arg Arg Glu Arg Ser Arg Ser 530 535 540 Arg Arg Lys Gln His Leu Leu Pro Pro Cys Val Asp Glu Pro Glu Leu 545 550 555 560 Leu Phe Ser Glu Gly Pro Ser Thr Ser Arg Trp Ala Ala Glu Leu Pro 565 570 575 Phe Pro Ala Asp Ser Ser Asp Pro Ala Ser Gln Leu Ser Tyr Ser Gln 580 585 590 Glu Val Gly Gly Pro Phe Lys Thr Pro Ile Lys Glu Thr Leu Pro Ile 595 600 605 Ser Ser Thr Pro Ser Lys Ser Val Leu Pro Arg Thr Pro Glu Ser Trp 610 615 620 Arg Leu Thr ...

Claims

1. A peptide consisting of the amino acid sequence of SEQ ID NO:

56.

2. A polynucleotide encoding the peptide of claim 1.

3. A composition comprising a pharmaceutically acceptable carrier and at least one component selected from the following groups (a) to (e): (a) the peptide of claim 1; (b) a polynucleotide encoding the peptide of claim 1 in an expressible form; (c) an antigen presenting cell (APC) which presents a complex of the peptide of claim 1 and HLA-A01 on its cell surface; (d) an exosome presenting a complex of the peptide of claim 1 and HLA-A01 on its cell surface; and (e) CTLs which target the peptide of claim 1.

4. The composition of claim 3, which is a composition for inducing CTL, wherein the component is at least one component selected from the following groups (a) to (d): (a) the peptide of claim 1; (b) a polynucleotide encoding the peptide of claim 1 in an expressible form; (c) an antigen presenting cell (APC) which presents a complex of the peptide of claim 1 and HLA-A01 on its cell surface; and (d) An exosome presenting a complex of the peptide of claim 1 and HLA-A01 on its cell surface.

5. The composition of claim 3, which is a pharmaceutical composition.

6. The composition of claim 5, for use in one or more uses selected from the group consisting of: (i) cancer treatment, (ii) cancer prevention (prophylaxis) and (iii) prevention of cancer recurrence after surgery (prophylaxis).

7. The composition of claim 5, for use in inducing an immune response against cancer.

8. The composition of claim 6, wherein the cancer is selected from the group consisting of acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocellular carcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, small cell lung cancer (SCLC), soft tissue tumors, and testicular tumors.

9. The composition of any one of claims 3 to 8, formulated for administration to a subject who is at least HLA-A01 positive.

10. A method for inducing APCs having CTL inducibility, comprising a step selected from the group consisting of: (a) contacting APCs with the peptide of claim 1 in vitro or ex vivo; and (b) introducing a polynucleotide encoding the peptide of claim 1 into APCs in vitro or ex vivo.

11. An in vitro method for inducing CTLs, comprising a step selected from the group consisting of: (a) co-culturing CD8-positive T cells with APCs presenting a complex of an HLA antigen and the peptide of claim 1 on its surface; (b) co-culturing CD8-positive T cells with exosomes presenting a complex of an HLA antigen and the peptide of claim 1 on its surface; and (c) introducing a polynucleotide encoding each subunit of a T cell receptor (TCR) capable of binding to the peptide of claim 1 presented by an HLA antigen on the cell surface into a CD8-positive T cell.

12. APC which presents a complex of an HLA antigen and the peptide of claim 1 on its surface. The APC according to claim 12 , which is induced by the method according to claim 10 .

14. CTL which targets the peptide of claim 1.

15. The CTL of claim 14, which is induced by an in vitro method comprising a step selected from the group consisting of: (a) co-culturing CD8-positive T cells with APCs presenting a complex of an HLA antigen and the peptide of claim 1 on its surface; (b) co-culturing CD8-positive T cells with exosomes presenting a complex of an HLA antigen and the peptide of claim 1 on its surface; and (c) introducing a polynucleotide encoding each subunit of a T cell receptor (TCR) capable of binding to the peptide of claim 1 presented by an HLA antigen on the cell surface into a CD8-positive T cell.

16. An emulsion comprising the peptide of claim 1, a water-soluble carrier and an oil adjuvant.

17. A kit comprising a container containing the composition of any one of claims 3 to 9 and a container containing an adjuvant.

18. Use of an ingredient for preparing a pharmaceutical composition for inducing an immune response against a cancer expressing FOXM1, wherein the ingredient is selected from the group consisting of (a)-(e): (a) the peptide of claim 1; (b) a polynucleotide encoding the peptide of claim 1 in an expressible form; (c) APCs which present a complex of the peptide of claim 1 and HLA-A01 on their cell surface; (d) an exosome presenting a complex of the peptide of claim 1 and HLA-A01 on its cell surface; and (e) CTLs targeting the peptide of claim 1, wherein the cancer is selected from the group consisting of acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocellular carcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, small cell lung cancer (SCLC), soft tissue tumors and testicular tumors.

19. Use of a component for preparing a pharmaceutical composition for treating and / or preventing cancer expressing FOXM1, and / or preventing its postoperative recurrence, wherein the component is selected from the group consisting of the following (a)-(e): (a) the peptide of claim 1; (b) a polynucleotide encoding the peptide of claim 1 in an expressible form; (c) APCs which present a complex of the peptide of claim 1 and HLA-A01 on their cell surface; (d) an exosome presenting a complex of the peptide of claim 1 and HLA-A01 on its cell surface; and (e) CTLs targeting the peptide of claim 1, wherein the cancer is selected from the group consisting of acute myeloid leukemia (AML), bladder cancer, breast cancer, cervical cancer, cholangiocellular carcinoma, chronic myeloid leukemia (CML), colon cancer, esophageal cancer, gastric cancer, liver cancer, non-small cell lung cancer (NSCLC), lymphoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, small cell lung cancer (SCLC), soft tissue tumors and testicular tumors.

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