Method for activating T cells for cancer treatment

Abstract

The present invention relates to a cancer-specific tumor antigen neoepitope represented by any one of SEQ ID NOs: 1 to 184, an antigen-presenting cell loaded with the neoepitope, and a method for activating T cells for cancer treatment using the antigen-presenting cell. An antigen-presenting cell, that is, a dendritic cell, loaded with a cancer-specific tumor antigen epitope provided in the present invention enables rapid and effective induction of differentiation and proliferation of cancer antigen-specific T cells, preferably memory T cells, and the memory T cells thus activated can treat a cancerous or neoplastic condition or prevent recurrence, progression, or metastasis of cancer while avoiding the defense mechanism of cancer cells.

Claims

1. A method for preventing or treating cancer, comprising a step of administering to a target individual an antigen-presenting cell loaded with an Epstein-Barr virus (EBV)-positive cancer-specific tumor antigen neoepitope consisting of any one of SEQ ID NOs: 1, 5, 63, 99 and 136.

2. The method according to claim 1, wherein the antigen-presenting cell is a dendritic cell, a B cell, or a macrophage.

3. The method according to claim 1, wherein the antigen-presenting cell promotes proliferation or differentiation of T cells.

4. The method according to claim 1, wherein the Epstein-Barr virus (EBV)-positive cancer-specific tumor antigen neoepitope exhibits binding affinity with at least one of HLA-A, HLA-B, HLA-C, HLA-E, HLA-F, HLA-G, 2-microglobulin, HLA-DPA1, HLA-DPB1, HLA-DQA1, HLA-DQB1, HLA-DRA1, HLA-DRB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DM, HLA-DOA, and HLA-DOB loci.

5. The method according to claim 1, wherein the Epstein-Barr virus (EBV)-positive cancer-specific tumor antigen neoepitope exhibits binding affinity with at least one of HLA-A*2402, HLA-A*A0201, HLA-A*3303, HLA-A*1101, HLA-A*0206, HLA-A*3101, HLA-B*5101, HLA-B*4403, HLA-B*5401, HLA-B*5801, and HLA-B*3501.

6. The method according to claim 1, wherein the cancer is EBV-positive gastric cancer, EBV-positive cervical cancer, EBV-positive Burkitt's lymphoma, EBV-positive T cell lymphoma, EBV-positive breast cancer, EBV-positive leiomyosarcoma, EBV-positive smooth muscle tumor, EBV-positive Hodgkin lymphoma, EBV-positive nasopharyngeal cancer, or EBV-positive post-transplant lymphoproliferative disorder (PTLD).

Description

BRIEF DESCRIPTION OF DRAWINGS

(1) FIG. 1 illustrates results obtained by performing ELISPOT on respective EBNA-1-derived neoepitopes to identify degree of T cell activation caused by such neoepitopes in SNU719 cell line according to an embodiment of the present invention.

(2) FIG. 2 illustrates results obtained by performing ELISPOT on respective EBNA-1-derived neoepitopes to identify degree of T cell activation caused by such neoepitopes in EBV-infected MKN74 cell line according to an embodiment of the present invention.

(3) FIG. 3 illustrates results obtained by performing ELISPOT on respective LMP-2A-derived neoepitopes to identify degree of T cell activation caused by such neoepitopes in SNU719 cell line according to an embodiment of the present invention.

(4) FIG. 4 illustrates results obtained by performing ELISPOT on respective LMP-2A-derived neoepitopes to identify degree of T cell activation caused by such neoepitopes in EBV-infected MKN74 cell line according to an embodiment of the present invention.

(5) FIG. 5 illustrates results obtained by performing Cr51 release assay on each of three neoepitopes, which have been selected from EBNA-1-derived neoepitopes, in SNU-719 cell line, depending on HLA type, to identify targeted cancer cell-killing activity of EBV-positive gastric cancer cell antigen, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(6) FIG. 6 illustrates results obtained by performing Cr51 release assay on each of three neoepitopes, which have been selected from EBNA-1-derived neoepitopes, in EBV-infected MKN74 cell line, depending on HLA type, to identify targeted cancer cell-killing activity of EBV-positive gastric cancer cell antigen, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(7) FIG. 7 illustrates results obtained by performing Cr51 release assay on each of three neoepitopes, which have been selected from LMP-2A-derived neoepitopes, in SNU-719 cell line, depending on HLA type, to identify targeted cancer cell-killing activity of EBV-positive gastric cancer cell antigen, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(8) FIG. 8 illustrates results obtained by performing Cr51 release assay on each of three neoepitopes, which have been selected from LMP-2A-derived neoepitopes, in EBV-infected MKN74 cell line, depending on HLA type, to identify targeted cancer cell-killing activity of EBV-positive gastric cancer cell antigen, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(9) FIG. 9 illustrates results obtained by performing Cr51 release assay on each of three selected EBNA-1-derived neoepitopes in EBV-negative MKN74 cell line, to identify cancer cell specificity of such neoepitopes among EBV-positive gastric cancer cell antigens, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(10) FIG. 10 illustrates results obtained by performing Cr51 release assay on each of three selected EBNA-1-derived neoepitopes in EBV-negative MKN74 cell line, to identify cancer cell specificity of such neoepitopes among EBV-positive gastric cancer cell antigens, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(11) FIG. 11 illustrates results obtained by performing Cr51 release assay on each of three selected LMP-2A-derived neoepitopes in EBV-negative MKN74 cell line, to identify cancer cell specificity of such neoepitopes among EBV-positive gastric cancer cell antigens, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(12) FIG. 12 illustrates results obtained by performing Cr51 release assay on each of three selected LMP-2A-derived neoepitopes in EBV-negative MKN74 cell line, to identify cancer cell specificity of such neoepitopes among EBV-positive gastric cancer cell antigens, caused by human-derived memory T cells produced according to an embodiment of the present invention.

(13) FIG. 13 illustrates experimental results on effects of neoepitope-specific cytotoxic T cells in BALB/c nude mouse xenograft model (SNU-719), to verify an in vivo anticancer effect of EBNA-1-derived neoepitopes according to an embodiment of the present invention.

(14) FIG. 14 illustrates experimental results on effects of neoepitope-specific cytotoxic T cells in BALB/c nude mouse xenograft model (EBV-infected MKN74), to verify an in vivo anticancer effect of EBNA-1-derived neoepitopes according to an embodiment of the present invention.

(15) FIG. 15 illustrates experimental results on effects of neoepitope-specific cytotoxic T cells in BALB/c nude mouse xenograft model (SNU-719), to verify an in vivo anticancer effect of LMP-2A-derived neoepitopes according to an embodiment of the present invention.

(16) FIG. 16 illustrates experimental results on effects of neoepitope-specific cytotoxic T cells in BALB/c nude mouse xenograft model (EBV-infected MKN74), to verify an in vivo anticancer effect of LMP-2A-derived neoepitopes according to an embodiment of the present invention.

DETAILED DESCRIPTION OF INVENTION

(17) According to an embodiment of the present invention, there is provided an Epstein-Barr virus (EBV)-positive cancer-specific tumor antigen neoepitope, represented by any one of SEQ ID NOs: 1 to 184.

(18) According to another embodiment of the present invention, there is provided an antigen-presenting cell (APC) loaded with a cancer-specific tumor antigen neoepitope provided in the present invention.

(19) According to yet another embodiment of the present invention, there is provided a T cell activated by an antigen-presenting cell provided in the present invention.

(20) According to still yet another embodiment of the present invention, there is provided an anticancer vaccine or a pharmaceutical composition for preventing or treating cancer, comprising, as an active ingredient, an antigen-presenting cell loaded with a cancer-specific tumor antigen epitope provided in the present invention; and/or an activated T cell.

(21) Hereinafter, the present invention will be described in more detail by way of examples. These examples are only for describing the present invention in more detail, and it will be apparent to those skilled in the art that according to the gist of the present invention, the scope of the present invention is not limited by these examples.

EXAMPLES

Example 1 Production Method of Autologous Memory T Cells Specific to EBV-Positive Gastric Cancer Cells and Clinical Application Thereof

(22) 1. Selection of EBV-Positive Gastric Cancer Cell Antigen Neoepitopes

(23) Algorithms for predicting the most important sequence with accumulation of genetic mutations in gastric cancer cells and for predicting epitopes of this sequence which bind to HLA of T cells were developed using bioinformatics and proteomics. As algorithms for predicting neoepitopes, NetMHC and NetCTLpan were used. Here, peptide sequences were identified which are expected to have high binding affinity with various HLA types including HLA-A*1101, HLA-A*0206, HLA-A*3101, HLA-B*5101, HLA-B*4403, HLA-B*5401, HLA-B*5801, and HLA-B*3501, as well as HLA-A*2402, HLA-A*A0201, and HLA-A*3303 which are HLA types that Koreans express the most. To this end, existing EBV+gastric cancer cell lines were HLA-typed to investigate the HLA type of each cell line, and neoepitopes having high binding affinity with each HLA type were predicted and identified through NetMHC program for the representative proteins, LMP-2A and EBNA-1 proteins, which are present only in EBV+gastric cancer cells due to EBV virus and cause malignancies. The results are shown in Tables 1 and 2, respectively.

(24) TABLE-US-00001 TABLE1 NeoepitopesforLMP-2Aprotein HLA- HLA- HLA- HLA- HLA- HLA- #ID SEQ(9mer) A*24:02 A*02:01 A*33:03 A*11:01 A*02:06 A*31:01 LMP2A_SNU- MGS 27371.8 11991.5 19548.9 21044.7 2968.1 20300.8 719_1 LEM VPM (SEQ ID NO: 1) LMP2A_SNU- SMN 9889.5 9.9 6933.4 4394.9 14.9 1649.6 719_119 PVCL PV (SEQ ID NO: 2) LMP2A_SNU- NPV 40322.3 27033.9 31415.9 38783.6 19110.2 38909.2 719_121 CLPV IV (SEQ ID NO: 3) LMP2A_SNU- CLPV 13941.3 10007.3 11545.2 4017.4 5530.2 10510.1 719_124 IVAP Y (SEQ ID NO: 4) LMP2A_SNU- LPVI 29593.2 19280.3 23285.5 37445.1 8770.7 30098.9 719_125 VAP YL (SEQ ID NO: 5) LMP2A_SNU- VIVA 3787.8 21911.8 24575.2 10829.4 13529.2 16840.6 719_127 PYLF W (SEQ ID NO: 6) LMP2A_SNU- IVAP 2732.2 10.1 5448.5 8930.6 6.7 2015.8 719_128 YLF WL (SEQ ID NO: 7) LMP2A_SNU- VAP 6485.2 2405.6 10912 20624.5 159.1 10250.1 719_129 YLF WLA (SEQ ID NO: 8) LMP2A_SNU- APY 39372.1 14186.6 22554 23424.5 5249.7 20785.8 719_130 LFW LAA (SEQ ID NO: 9) LMP2A_SNU- PYLF 45.8 20451.8 20179 37721.6 12845 12993.1 719_131 WLA AI (SEQ ID NO: 10) LMP2A_SNU- YLF 36946.9 22.5 13714.8 22720.8 67.5 15292.2 719_132 WLA AIA (SEQ ID NO: 11) LMP2A_SNU- LAAI 11127 28133.2 32080.5 26643.1 12921.5 32823.8 719_136 AAS CF (SEQ ID NO: 12) LMP2A_SNU- AIAA 30319.9 200.5 24584.2 6731.1 71.3 12568.3 719_138 SCFT A (SEQ ID NO: 13) LMP2A_SNU- AAS 25117.4 224.6 17740.8 12369.6 32.8 11110 719_140 CFTA SV (SEQ ID NO: 14) LMP2A_SNU- FTAS 19464.9 88.6 13762.4 20136.1 14.6 19810.8 719_144 VST VV (SEQ ID NO: 15) LMP2A_SNU- ASVS 37052.1 5560.8 32852.6 8850.6 424.7 18831.4 719_146 TVV TA (SEQ ID NO: 16) LMP2A_SNU- TATG 28638.1 14742.6 31449.6 28040.2 4055.3 34638 719_153 LALS L (SEQ ID NO: 17) LMP2A_SNU- GLA 18374.7 83 33237 18495.4 254.8 19863.2 719_156 LSLL LL (SEQ ID NO: 18) LMP2A_SNU- LALS 35190.6 817.2 21235.6 14830.1 83.6 15856.8 719_157 LLLL A (SEQ ID NO: 19) LMP2A_SNU- ALSL 40341.5 157.1 31542.6 14530.5 190 17991.5 719_158 LLLA A (SEQ ID NO: 20) LMP2A_SNU- LSLL 25009.2 295.1 14717.7 21696.2 30.5 9125.3 719_159 LLA AV (SEQ ID NO: 21) LMP2A_SNU- SLLL 38650.3 362.5 30858 23970.9 591.4 22057.6 719_160 LAA VA (SEQ ID NO: 22) LMP2A_SNU- LLLA 39044.6 402.5 31171.1 21818.6 315.9 23555.9 719_162 AVAS S (SEQ ID NO: 23) LMP2A_SNU- LLA 26232.6 11237.2 17769.4 1693.5 8474.3 15035 719_163 AVAS SY (SEQ ID NO: 24) LMP2A_SNU- AAV 38779.4 7069.2 29746.7 10683.7 397.4 19865.3 719_165 ASSY AA (SEQ ID NO: 25) LMP2A_SNU- AVAS 39351.2 1556.8 21477.3 6403.6 124.8 14250.3 719_166 SYA AA (SEQ ID NO: 26) LMP2A_SNU- ASSY 40984.7 33783 367.6 173.6 25710.5 76.6 719_168 AAA QR (SEQ ID NO: 27) LMP2A_SNU- SSYA 36886.5 31558.3 5931.4 14.5 20818 930.3 719_169 AAQ RK (SEQ ID NO: 28) LMP2A_SNU- AQR 23509.8 882.2 24447.6 21770.7 70 5273.4 719_174 KLLT PV (SEQ ID NO: 29) LMP2A_SNU- KLLT 11776.9 93.5 28522.5 24228.3 193.5 7097.1 719_177 PVT VL (SEQ ID NO: 30) LMP2A_SNU- LLTP 37137.6 147.1 23494.6 22140.1 257.4 20809.9 719_178 VTV LT (SEQ ID NO: 31) LMP2A_SNU- TPVT 37734.2 18144.2 24195.8 32539.5 5162.9 31765.6 719_180 VLTA V (SEQ ID NO: 32) LMP2A_SNU- TVLT 1299.4 12607.8 21331.9 12630.8 2596 17811.8 719_183 AVV TF (SEQ ID NO: 33) LMP2A_SNU- VLTA 1880 1724.7 18858.8 13899.4 1269.2 11444.8 719_184 VVT FF (SEQ ID NO: 34) LMP2A_SNU- LTAV 28599.4 522 4795.7 3078.1 92.6 3678 719_185 VTFF A (SEQ ID NO: 35) LMP2A_SNU- TAV 7615.8 3348.3 12820.8 15877.6 134.9 13801.6 719_186 VTFF AI (SEQ ID NO: 36) LMP2A_SNU- TFFA 209.6 24870.2 17421.4 26303.1 22323.9 16909.8 719_190 ICLT W (SEQ ID NO: 37) LMP2A_SNU- FFAI 31407.7 22080.5 67.9 9575.9 20572.3 140.7 719_191 CLT WR (SEQ ID NO: 38) LMP2A_SNU- FAIC 2489.3 33.1 6603.3 20325.8 8.7 11143.4 719_192 LTW RI (SEQ ID NO: 39) LMP2A_SNU- SLLF 33552.8 28.8 22637.9 7154.3 26.2 12754.2 719_207 ALL AA (SEQ ID NO: 40) LMP2A_SNU- LLFA 39864.6 17.1 15143.5 19287.8 22.5 11438.6 719_208 LLA AA (SEQ ID NO: 41) LMP2A_SNU- ALL 30890.7 238.2 33730 31528.9 425.5 22061.4 719_211 AAA GGL (SEQ ID NO: 42) LMP2A_SNU- AAA 27565 3517.1 32696.6 24960 137.2 27509.3 719_214 GGL QGI (SEQ ID NO: 43) LMP2A_SNU- GLQ 19757.3 26.7 18903.5 20338.6 89.9 6778.5 719_218 GIYV LV (SEQ ID NO: 44) LMP2A_SNU- LQGI 21973.1 8022.8 37703.6 30905.4 2345.8 29019 719_219 YVL VM (SEQ ID NO: 45) LMP2A_SNU- GIYV 36065.3 151.8 21377.9 13972.3 448.8 8123.9 719_221 LVM LV (SEQ ID NO: 46) LMP2A_SNU- YVL 17490.5 293.6 15156 27530.7 153.6 11887.4 719_223 VML VLL (SEQ ID NO: 47) LMP2A_SNU- VML 35525.8 444.6 24613.5 18421.9 674.5 10636.3 719_226 VLLI LA (SEQ ID NO: 48) LMP2A_SNU- MLV 37081.4 19470 23627.1 6144.4 16629.6 20171.2 719_227 LLIL AY (SEQ ID NO: 49) LMP2A_SNU- LVLL 40296.1 15526.5 27.5 405.9 10119.1 20.7 719_228 ILAY R (SEQ ID NO: 50) LMP2A_SNU- VLLI 33407.1 15983.6 238.7 964.6 16825.3 563 719_229 LAY RR (SEQ ID NO: 51) LMP2A_SNU- LLIL 37733.4 194672 353 3782.7 15867.3 159.2 719_230 AYR RR (SEQ ID NO: 52) LMP2A_SNU- LILA 10191.2 25457.8 25429.9 27362.6 19540.7 17519.7 719_231 YRR RW (SEQ ID NO: 53) LMP2A_SNU- ILAY 39845.2 28729.1 71.9 7319.8 30463.6 29.4 719_232 RRR WR (SEQ ID NO: 54) LMP2A_SNU- LAY 24998.6 20425.9 7.8 424.1 14678.8 5.2 719_233 RRR WRR (SEQ ID NO: 55) LMP2A_SNU- RLTV 30154.3 7330.9 345477 24745.1 5573.4 16855.2 719_241 CGGI M (SEQ ID NO: 56) LMP2A_SNU- LTVC 9930 15172.7 21640.9 14515.6 3824 20535.8 719_242 GGI MF (SEQ ID NO: 57) LMP2A_SNU- TVC 17100.9 384.8 9819.9 8431.6 128.5 9625.5 719_243 GGI MFL (SEQ ID NO: 58) LMP2A_SNU- GGI 29382.6 2824.6 25826.7 29467 437.1 15686.2 719_246 MFL ACV (SEQ ID NO: 59) LMP2A_SNU- IMFL 13468.2 33.7 7969.8 17351.1 93.2 2903 719_248 ACV LV (SEQ ID NO: 60) LMP2A_SNU- FLAC 19384.4 10.5 12112.4 24245.6 16.8 10846 719_250 VLV LI (SEQ ID NO: 61) LMP2A_SNU- VLV 31115.8 63.7 27382.5 29178.3 81.1 22651.6 719_254 LIVD AV (SEQ ID NO: 62) LMP2A_SNU- LIVD 23873.3 143.2 25164.7 21696.4 59.9 25241.6 719_257 AVL QL (SEQ ID NO: 63) LMP2A_SNU- AVL 10808.3 395.5 23133.1 13646.3 117.1 9438.6 719_261 QLSP LL (SEQ ID NO: 64) LMP2A_SNU- LQLS 32042.4 469.3 29672.4 14340.6 29.8 15058.9 719_263 PLLG A (SEQ ID NO: 65) LMP2A_SNU- QLSP 33550.6 49.7 21128.2 26797.4 43.7 16755 719_264 LLG AV (SEQ ID NO: 66) LMP2A_SNU- SPLL 30117.5 17918.9 30290.4 33256.4 7730 31488.4 719_266 GAV TV (SEQ ID NO: 67) LMP2A_SNU- VTV 15006.2 5526.8 18969.7 15331.5 589.6 13347.5 719_272 VSM TLL (SEQ ID NO: 68) LMP2A_SNU- TVV 6049.7 652.7 13530.9 7958.2 177.7 15315.7 719_273 SMT LLL (SEQ ID NO: 69) LMP2A_SNU- VVS 8120.1 978.1 21960 10661 428 13853.2 719_274 MTL LLL (SEQ ID NO: 70) LMP2A_SNU- VSM 22765.4 920 16803.5 2487.7 79.4 5629.8 719_275 TLLL LA (SEQ ID NO: 71) LMP2A_SNU- SMT 2867.4 5041.4 25983.4 15082.4 3004.4 16672.7 719_276 LLLL AF (SEQ ID NO: 72) LMP2A_SNU- MTL 19035.5 27.4 1099.5 4770.9 8.4 582 719_277 LLLA FV (SEQ ID NO: 73) LMP2A_SNU- TLLL 12676.6 164.2 20545 25952.5 587.5 13178.3 719_278 LAF VL (SEQ ID NO: 74) LMP2A_SNU- LLLA 24803 40 23132.4 26986.3 202.3 10568 719_280 FVL WL (SEQ ID NO: 75) LMP2A_SNU- LLAF 33242 300.2 13010.6 11757 412.4 9140.2 719_281 VLW LS (SEQ ID NO: 76) LMP2A_SNU- TLG 7510.8 95.9 31160 23852.4 183.4 22528.4 719_296 AAL LTL (SEQ ID NO: 77) LMP2A_SNU- AAL 39908.7 1586.9 28364.3 16855.7 86.9 16521.3 719_299 LTLA AA (SEQ ID NO: 78) LMP2A_SNU- ALLT 23261.1 46.8 26860.1 27729.8 155.2 13582.5 719_300 LAA AL (SEQ ID NO: 79) LMP2A_SNU- LTLA 11578.4 949 20215.5 12875.3 129.5 13315.3 719_302 AAL AL (SEQ ID NO: 80) LMP2A_SNU- TLA 6696.7 21.8 13358.6 16222.7 28.2 11363.2 719_303 AAL ALL (SEQ ID NO: 81) LMP2A_SNU- LAA 36062.2 2985.8 25515.9 15003.8 414 25095.9 719_304 ALA LLA (SEQ ID NO: 82) LMP2A_SNU- AAL 16882.4 797.7 23744.5 21892.9 49.1 10793.2 719_306 ALL ASL (SEQ ID NO: 83) LMP2A_SNU- ALA 14588.2 191 30081.3 19870.7 541.2 18083.4 719_307 LLAS LI (SEQ ID NO: 84) LMP2A_SNU- LALL 17259.7 7925.7 33454.2 2993.87 1744.5 29244 719_308 ASLI L (SEQ ID NO: 85) LMP2A_SNU- YPSA 41679.6 39870.2 37645.8 39333.8 32297.5 42032 719-31 SGSY G (SEQ ID NO: 86) LMP2A_SNU- LLAS 41604.9 74.7 29015.5 23428.3 99.3 24439.7 719_310 LILG T (SEQ ID NO: 87) LMP2A_SNU- SLIL 12080.2 70.6 25306.7 15799.3 170.6 16385.8 719_313 GTL NL (SEQ ID NO: 88) LMP2A_SNU- LILG 41776.7 1274.6 27409.7 27342.2 220.3 24829.1 719_314 TLNL T (SEQ ID NO: 89) LMP2A_SNU- GTL 5555.5 13923.8 23808.1 6308.3 2682.7 9656.2 719_317 NLTT MF (SEQ ID NO: 90) LMP2A_SNU- TLNL 11744.1 70.1 13411.5 9317.2 331.4 9132.5 719_318 TTM FL (SEQ ID NO: 91) LMP2A_SNU- NLTT 13538.2 261.2 12244.1 16237.1 454.7 15909.4 719_320 MFL LM (SEQ ID NO: 92) LMP2A_SNU- TTM 2651.6 248.1 1781.8 2219 48.5 1513.6 719_322 FLL MLL (SEQ ID NO: 93) LMP2A_SNU- TMF 3988.3 16232.3 24882.1 18622.5 13501.9 17977.3 719_323 LLM LLW (SEQ ID NO: 94) LMP2A_SNU- FLL 8083.8 8.9 12745.7 33192.8 13.9 7882.1 719_325 MLL WTL (SEQ ID NO: 95) LMP2A_SNU- LLM 13488.2 26.7 7799.5 17571.3 52.8 3052 719_326 LLW TLV (SEQ ID NO: 96) LMP2A_SNU- LML 15787.8 31.7 15849.6 19663.7 119.1 6864 719_327 LWT LVV (SEQ ID NO: 97) LMP2A_SNU- MLL 16912 24.7 13754 24171.7 52.5 7678.7 719_328 WTL VVL (SEQ ID NO: 98) LMP2A_SNU- LLW 22368.6 24.7 11762 26130.3 141.6 4070.5 719_329 TLV VLL (SEQ ID NO: 99) LMP2A_SNU- CPLT 37670.6 21444.6 26652.3 29420.8 10764.3 29121.2 719_345 KILL A (SEQ ID NO: 100) LMP2A_SNU- ILLA 5233.8 3865.6 6694.9 236 2901.5 1264.9 719_350 RLFL Y (SEQ ID NO: 101) LMP2A_SNU- LLA 31672.9 34.9 11717.7 4604.7 45.8 6011.2 719_351 RLFL YA (SEQ ID NO: 102) LMP2A_SNU- RLFL 6856.4 20.3 14775.5 9547.6 92.6 2096.5 719_354 YAL AL (SEQ ID NO: 103) LMP2A_SNU- FLYA 11428.5 6 10051.6 14849.5 19.6 7883.2 719_356 LALL L (SEQ ID NO: 104) LMP2A_SNU- LYAL 101 12176 20911.6 31716.4 6586.9 16497.4 719_357 ALLL L (SEQ ID NO: 105) LMP2A_SNU- YAL 31846.1 159.2 16022.1 14850.3 20.8 15655.3 719_358 ALLL LA (SEQ ID NO: 106) LMP2A_SNU- LALL 40372 1722.3 23602.9 27980.2 127.2 20356.9 719_360 LLAS A (SEQ ID NO: 107) LMP2A_SNU- ALLL 25296.6 167 29618.5 30183.4 455.9 16547.6 719_361 LAS AL (SEQ ID NO: 108) LMP2A_SNU- LLLL 7740.7 55.9 20129.3 23602.9 116.9 13531.6 719_362 ASA LI (SEQ ID NO: 109) LMP2A_SNU- LLLA 37708.1 263.9 35188.7 16754.1 400.9 26159.2 719_363 SALI A (SEQ ID NO: 110) LMP2A_SNU- ALIA 25800.8 493.1 36320.6 28980.7 560.1 24630.6 719_368 GGSI L (SEQ ID NO: 111) LMP2A_SNU- GSIL 40801.5 35501.6 6602.4 16.2 26128.9 791.4 719_373 QTN FK (SEQ ID NO: 112) LMP2A_SNU- KSLS 5507.7 2533.5 21307.9 12976.3 221.3 3775.2 719_381 STEF I (SEQ ID NO: 113) LMP2A_SNU- SSTE 14988.3 2681.8 19581.3 13178 219.9 11100.4 719_384 FIPN L (SEQ ID NO: 114) LMP2A_SNU- FIPN 4505.5 72.9 10675 27973.3 19.8 14050.6 719_388 LFC ML (SEQ ID NO: 115) LMP2A_SNU- IPNL 16916.4 10587.1 20005.8 27848 6583.9 20799.9 719_389 FCM LL (SEQ ID NO: 116) LMP2A_SNU- NLFC 14595.1 20.1 7777.2 14671.3 74.9 7191.9 719_391 MLL LI (SEQ ID NO: 117) LMP2A_SNU- MLL 22235.9 20.4 9301.5 26251 43.9 6024.6 719_395 LIVA GI (SEQ ID NO: 118) LMP2A_SNU- LLIV 6873.4 7266.6 29271.6 21315.3 4210.1 22774.5 719_397 AGIL F (SEQ ID NO: 119) LMP2A_SNU- LIVA 23654.3 75.6 18461.8 15122.1 47.5 14693.4 719_398 GILFI (SEQ ID NO: 120) LMP2A_SNU- IVAG 6760.8 65.7 10025.3 14721.2 34.3 6870.5 719_399 ILFIL (SEQ ID NO: 121) LMP2A_SNU- FILAI 5075.9 3363.4 19730 23123.9 1176.5 21302.8 719_405 LTE W (SEQ ID NO: 122) LMP2A_SNU- LTE 45205.3 34231.1 404.4 1968.7 28770.8 787.8 719_410 WGS GNR (SEQ ID NO: 123) LMP2A_SNU- RTY 13674.3 2452.5 20810.3 7194.6 359.6 2242.3 719_418 GPVF MC (SEQ ID NO: 124) LMP2A_SNU- TYG 43.1 20573.4 17372.1 28767 11909.3 10962.4 719_419 PVF MCL (SEQ ID NO: 125) LMP2A_SNU- FMC 36749.1 418.4 26159.7 27774.5 543.7 23960.5 719_424 LGG LLT (SEQ ID NO: 126) LMP2A_SNU- MCL 25037.6 3987 27163.8 30133.1 848.4 26216.1 719_425 GGL LTM (SEQ ID NO: 127) LMP2A_SNU- CLG 29360.1 39.8 26562.2 29588.7 70 18962.1 719_426 GLLT MV (SEQ ID NO: 128) LMP2A_SNU- GLLT 42669.3 50.4 33575.3 27874.2 101.9 19703.9 719_429 MVA GA (SEQ ID NO: 129) LMP2A_SNU- LLT 37594.5 307.4 27810.6 35011.8 519 26238 719_430 MVA GAV (SEQ ID NO: 130) LMP2A_SNU- LTM 2451.6 22223.6 21501.8 15181.9 8332.6 17938.6 719_431 VAG AVW (SEQ ID NO: 131) LMP2A_SNU- TMV 14694.7 81.7 20692.6 26012.6 145.8 18008.3 719_432 AGA VWL (SEQ ID NO: 132) LMP2A_SNU- MVA 21800.9 136.2 5179.1 7819.4 24.4 7866.6 719_433 GAV WLT (SEQ ID NO: 133) LMP2A_SNU- VAG 8593.3 2606.6 25834.3 23105.9 411.4 17830.3 719_434 AVW LTV (SEQ ID NO: 134) LMP2A_SNU- WLT 11823.3 106 26126.6 35724.7 248.7 26604.7 719_439 VMT NTL (SEQ ID NO: 135) LMP2A_SNU- LTV 17661.7 3493.6 16154 19986.5 509.9 14004.8 719_440 MTN TLL (SEQ ID NO: 136) LMP2A_SNU- VMT 31556.9 156.8 28830.3 13858.3 115.4 18258.8 719_442 NTLL SA (SEQ ID NO: 137) LMP2A_SNU- MTN 7290.9 20237.4 12898.6 10001.4 8034.8 12238.2 719_443 TLLS AW (SEQ ID NO: 138) LMP2A_SNU- TLLS 30273 396.9 21252 15570.9 352.7 14095.9 719_446 AWIL T (SEQ ID NO: 139) LMP2A_SNU- LLSA 32764.6 32.4 24319.4 13902.6 93.4 16585.4 719_447 WILT A (SEQ ID NO: 140) LMP2A_SNU- SAWI 4573.1 11740.2 13701 17240.5 2700.5 8892.1 719_449 LTAG F (SEQ ID NO: 141) LMP2A_SNU- WILT 5519.8 57.6 10388.7 17443.7 33.4 8331.9 719_451 AGF LI (SEQ ID NO: 142) LMP2A_SNU- ILTA 769.6 2963.6 29225.1 16621.5 1785.3 21390.8 719_452 GFLI F (SEQ ID NO: 143) LMP2A_SNU- LTAG 16982.4 120 6508.1 8045.4 33.2 4517.1 719_453 FLIF L (SEQ ID NO: 144) LMP2A_SNU- FLIF 42365.3 42.7 13437.3 25165.8 52 16269.2 719_457 LIGF A (SEQ ID NO: 145) LMP2A_SNU- IFLIG 43.6 17773.3 13570.3 28036.3 10285.2 10021.4 719-459 FALF (SEQ ID NO: 146) LMP2A_SNU- FLIG 36214.7 277.8 25993.6 27631.3 461.1 25977.5 719_460 FALF G (SEQ ID NO: 147) LMP2A_SNU- LIGF 24962.4 20.3 11149.9 18296.8 16.6 6672.9 719_461 ALF GV (SEQ ID NO: 148) LMP2A_SNU- GFAL 35328.8 33229.1 582.4 8108.8 32269.6 210 719_463 FGVI R (SEQ ID NO: 149) LMP2A_SNU- LFG 28785.7 34993.2 78.8 12351.4 33974.7 38.3 719_466 VIRC CR (SEQ ID NO: 150) LMP2A_SNU- VPM 44669 36655.4 35264.6 36414.6 26600.4 39954.4 719_7 GAG PPS (SEQ ID NO: 151) HLA- HLA- HLA- HLA- HLA- HLA- #ID B*51:01 B*15:01 B*44:03 B*54:01 B*58:01 B*35:01 LMP2A_SNU- 19068 1393.9 31463.2 20902.1 2627.5 164.2 719_1 LMP2A_SNU- 18869.2 443.8 11789.2 12880.3 9730 9998.1 719_119 LMP2A_SNU- 1717.4 34671.7 29524.1 241.1 30432.6 868.3 719_121 LMP2A_SNU- 32273.1 161.2 17839.2 29638.7 17168.1 1118.9 719_124 LMP2A_SNU- 456.5 22503.1 30741.7 337.4 11162.4 39 719_125 LMP2A_SNU- 23369.9 7172.4 14029.8 36881.8 81 14816.8 719_127 LMP2A_SNU- 14093.9 7650 29346.4 18362.2 817.2 9877.3 719_128 LMP2A_SNU- 21984.5 20546.3 34583.3 6597 10554.1 13623 719_129 LMP2A_SNU- 9931.9 20808.5 14633.9 25.3 33807.9 1458.6 719_130 LMP2A_SNU- 32690.2 33027.6 35911.5 39974.3 31558.3 38018.7 719_131 LMP2A_SNU- 26173.6 3914.4 28938.1 5371.4 30695.2 11886.3 719_132 LMP2A_SNU- 7996.6 63.4 14661.8 18015.7 159.8 23.8 719_136 LMP2A_SNU- 37957.8 6013.5 31247.4 14492.5 17396.7 21943.8 719_138 LMP2A_SNU- 9860.7 3306.5 10812.2 5602.1 3334.9 7527.4 719_140 LMP2A_SNU- 5884.4 1074.2 32054.2 1077.5 1464.4 1864.9 719_144 LMP2A_SNU- 32015 3547.8 26008.7 9458.8 3367.2 18947.7 719_146 LMP2A_SNU- 12835.5 9454.7 26436.6 21854.8 1695.6 404.4 719_153 LMP2A_SNU- 38515.9 4023.7 31753.5 41085.5 15854.4 36429.6 719_156 LMP2A_SNU- 9880.7 9979.6 25269 909.2 2060 6184.2 719_157 LMP2A_SNU- 41561.2 6626.9 26400.6 22040.9 30091.4 31931.3 719_158 LMP2A_SNU- 7137.4 4766.5 30342.5 3013.5 2491.7 14007.7 719_159 LMP2A_SNU- 35597.7 6048.7 26564.8 16328.5 32493.8 27442.7 719_160 LMP2A_SNU- 34547.8 3633.5 37258.4 16083.4 27538.7 18507 719_162 LMP2A_SNU- 23097.6 9.1 9592.6 23151.9 2410.6 43 719_163 LMP2A_SNU- 30204.3 3134.7 19736.8 2929 8128.6 2163.8 719_165 LMP2A_SNU- 36692.7 1626.1 28170.4 6365.9 23419.2 10828.7 719_166 LMP2A_SNU- 45002.3 24905.2 34733.7 39245.8 16532.8 34429.5 719_168 LMP2A_SNU- 35890.1 17324.8 25977.8 33151.5 7653 29593.5 719_169 LMP2A_SNU- 29998.4 223.9 14359.9 20829.2 28749.6 32884.3 719_174 LMP2A_SNU- 26627.8 787.1 31556.2 34200.4 6615.5 25782.9 719_177 LMP2A_SNU- 35201.7 13541.2 40609.9 19821.1 20510.8 21753.5 719_178 LMP2A_SNU- 1073 27088.7 32545.2 36.9 30156.9 322.4 719_180 LMP2A_SNU- 17799.8 242.9 18087.5 26212.1 1633.7 303.3 719_183 LMP2A_SNU- 28882.1 119.1 16654.5 35748.6 1881.7 4251.7 719_184 LMP2A_SNU- 26352.3 14096.2 34361.4 2279.9 2239.6 11683.7 719_185 LMP2A_SNU- 1811.7 4697.9 20385.3 1210.6 1509.2 1494.9 719_186 LMP2A_SNU- 23353.2 17916.9 7418.5 35167.8 1245.1 14681.3 719_190 LMP2A_SNU- 46034.1 39399.8 40223.4 35558.1 43345.9 36240.1 719_191 LMP2A_SNU- 701.1 7610.2 11525.1 679.6 323.5 1097.9 719_192 LMP2A_SNU- 36465.9 3779.6 28678.8 11933.9 27135 21409.4 719_207 LMP2A_SNU- 25677.4 1228.6 30349.7 2809 28533 12044.2 719_208 LMP2A_SNU- 35326.5 2099.8 24292.3 40247.3 22863.4 30329.1 719_211 LMP2A_SNU- 18595.5 8783.3 20531.6 26576 4561.5 20074.7 719_214 LMP2A_SNU- 39230.9 15981.5 30612.2 35452.4 29224.7 42802.1 719_218 LMP2A_SNU- 28065.4 128 14998.1 33933.9 18243.6 8771 719_219 LMP2A_SNU- 35575.8 16510.1 32974.8 28738.1 24709.3 42830.8 719_221 LMP2A_SNU- 25310.5 10525.9 35050.4 21845.1 9208.3 13460.3 719_223 LMP2A_SNU- 38789 14580.5 34150.1 26232 28571.3 36984.1 719_226 LMP2A_SNU- 34382.6 171.6 11119.5 26821.8 11758.2 366.9 719_227 LMP2A_SNU- 42621.4 33896.1 40627.1 26214.4 30730.7 34800.3 719_228 LMP2A_SNU- 45282.7 36046.9 36738 40449.4 37005.7 40947.5 719_229 LMP2A_SNU- 45312.6 23309 39296.3 40381.7 37662.9 38250.9 719_230 LMP2A_SNU- 21706.8 9545.3 17745.4 34120.6 136 18865.5 719_231 LMP2A_SNU- 46423.7 30236.3 42353.4 42222 40589.7 40637.6 719_232 LMP2A_SNU- 33940.2 28971 35978.8 22147.5 14714.4 25705.5 719_233 LMP2A_SNU- 34894.5 216.9 31598.6 33239.9 10678.4 12768 719_241 LMP2A_SNU- 20266.8 151.3 23587 21495.2 512.1 692.3 719_242 LMP2A_SNU- 28314.3 5527.2 35112.3 30373.4 9552.1 13224.3 719_243 LMP2A_SNU- 32710.4 17841.9 32488.9 27472.7 21090.2 38076.7 719_246 LMP2A_SNU- 19631.8 4985.6 28744 20012.3 20098.4 27284.3 719_248 LMP2A_SNU- 23637.6 7804.8 28164.6 13581.5 18267.5 19826.9 719_250 LMP2A_SNU- 25904.3 6865.1 31934.1 18772.4 26216.4 27905.3 719_254 LMP2A_SNU- 17736.2 1148.6 32569.5 22865.1 6475.4 3550.7 719_257 LMP2A_SNU- 20883.6 4239.7 18649.1 30555 4025.1 11797.6 719_261 LMP2A_SNU- 28135.4 2373.8 17703.8 14681.8 21815.1 27746.3 719_263 LMP2A_SNU- 26585.2 3605.7 30283.5 19801.6 25397.5 24460.1 719_264 LMP2A_SNU- 1526.9 29720.3 28761.4 355 27198.2 892.6 719_266 LMP2A_SNU- 21080.7 3762.7 36890.5 30202.3 452.5 18717.1 719_272 LMP2A_SNU- 15908.7 1878.8 25562.4 21519.7 2484.5 3325.5 719_273 LMP2A_SNU- 19204.7 2901.8 32228.4 28795 1653.4 11146 719_274 LMP2A_SNU- 15701.6 6979.6 21951.2 3615.2 826.9 18865.5 719_275 LMP2A_SNU- 33160.8 60 8048.1 35866 7268.3 1565.1 719_276 LMP2A_SNU- 11617.1 11509.4 27039.5 2957.3 2041.4 20396.6 719_277 LMP2A_SNU- 31870.9 9836 27846.8 32047.2 21002.6 19043.3 719_278 LMP2A_SNU- 32574.4 13364.1 29749.2 34396.7 14753.9 32068.7 719_280 LMP2A_SNU- 42903.1 22877.5 34893 25882.1 23769.5 32067.4 719_281 LMP2A_SNU- 30173.3 1555.3 33269.7 37369 12634.3 20706.8 719_296 LMP2A_SNU- 23912.1 4561.1 23387.8 2285.5 13858.1 8526 719_299 LMP2A_SNU- 30515.3 753.6 16063.9 32101.7 21011.9 14789.4 719_300 LMP2A_SNU- 8078.8 561.8 27998.1 10984.6 188.7 641.6 719_302 LMP2A_SNU- 26107.4 612.3 25912.1 30321.5 10647 12400.7 719_303 LMP2A_SNU- 9761.3 5008 22831.3 743 798.5 1496.1 719_304 LMP2A_SNU- 13608.2 1354.4 14455.4 17031.9 928.5 3403.9 719_306 LMP2A_SNU- 25036 3146.9 16547.8 32603.7 12979.9 35350.6 719_307 LMP2A_SNU- 3865.7 3163.4 29938.1 12036 436.8 600.4 719_308 LMP2A_SNU- 13480.7 25105.7 38370.8 653.1 25946.6 290.6 719-31 LMP2A_SNU- 38239.8 10950.6 35332.6 27090.7 26687.8 31919.2 719_310 LMP2A_SNU- 29842.7 677.1 24228.8 35236.8 15685.2 15131.9 719_313 LMP2A_SNU- 36325.3 20529.8 43873.4 17673.7 30129.5 28730.9 719_314 LMP2A_SNU- 32438.7 337.5 19058.3 36479.3 182.7 7933.2 719_317 LMP2A_SNU- 26916.6 1882.1 25585.3 30786 12223.1 10896.2 719_318 LMP2A_SNU- 27240.3 2283.2 19381.3 29707.8 15477.7 3864.1 719_320 LMP2A_SNU- 18366.4 3871.8 21160.2 13386.7 625.4 11838.4 719_322 LMP2A_SNU- 28013.9 7739 5484.2 39287 377.9 19971.6 719_323 LMP2A_SNU- 29633 9838.9 29389.6 24180.3 18410.8 15012.5 719_325 LMP2A_SNU- 22552.3 6274.9 22518.5 11468.4 21324.5 27980.5 719_326 LMP2A_SNU- 15437.3 2864 29283.3 15663.8 15936 17141 719_327 LMP2A_SNU- 20823.4 1237.6 27968.7 15626.1 11511.6 5123.8 719_328 LMP2A_SNU- 26938.7 6287.6 30004.3 24819.7 16723.5 22476.8 719_329 LMP2A_SNU- 3795.9 33986.8 32242.7 40.5 27618.4 4556.3 719_345 LMP2A_SNU- 28103.1 675.1 15314.7 31828.5 3987.8 3393.9 719_350 LMP2A_SNU- 32842.3 4290.9 32551.5 9100.6 20431 25441.8 719_351 LMP2A_SNU- 22423.6 352.6 17945 23931.5 8333.2 11218.5 719_354 LMP2A_SNU- 14452.3 933.8 22566.5 14066.6 8976 7114.4 719_356 LMP2A_SNU- 27210.3 16018.8 33860.9 34946.7 16177.4 23396.4 719_357 LMP2A_SNU- 11348.3 9221.5 22015.4 558.3 2741.6 2807.8 719_358 LMP2A_SNU- 12404.7 7468.9 31337.1 726.9 8043.6 5539.5 719_360 LMP2A_SNU- 30580.1 982.2 18787.5 32238.5 24767.1 17945.6 719_361 LMP2A_SNU- 10169.7 2337.2 27104.5 20273.6 10813.2 22288.7 719_362 LMP2A_SNU- 28071.2 3395.5 34432.5 11384 20770 18889.4 719_363 LMP2A_SNU- 32742.3 102.5 21901.8 34824 20077.8 14272.5 719_368 LMP2A_SNU- 43973.3 24934.3 39868.5 40321.4 14126.7 38027.7 719_373 LMP2A_SNU- 18540.5 8780.4 27824.2 27091 48.4 29173.2 719_381 LMP2A_SNU- 18233.9 11500.3 25381.8 32592.1 521 12130.1 719_384 LMP2A_SNU- 18845.9 3217.7 37974.7 17265.7 27948.8 8245.1 719_388 LMP2A_SNU- 808.3 22119.5 23895.3 1101.4 14068.7 298.1 719_389 LMP2A_SNU- 17987 10340.4 16613.5 17723.3 17321.2 28299.3 719_391 LMP2A_SNU- 16888.9 5920 19812.8 13208.7 14228.6 26095.6 719_395 LMP2A_SNU- 28410.7 114.5 19338.1 30961 7162.9 4153.9 719_397 LMP2A_SNU- 15242 4774.9 34571.4 17750.6 7809.7 20384.2 719_398 LMP2A_SNU- 20780.6 3961.8 33101 18325.3 1341.3 6022.3 719_399 LMP2A_SNU- 14795.5 4255.1 11400.4 18955.3 74 1361.9 719_405 LMP2A_SNU- 43444.1 32402.9 44760 36137.2 34919.5 29665 719_410 LMP2A_SNU- 33440.4 22294 35295.9 20698.7 1940.2 38397.7 719_418 LMP2A_SNU- 31268.4 22881.4 30885.4 36459.2 27367.3 23665 719_419 LMP2A_SNU- 38175.3 6987.3 39059 27342.8 32303.8 26047.9 719_424 LMP2A_SNU- 15294.2 3697.3 26509.3 18716.3 1800.8 227.3 719_425 LMP2A_SNU- 33113.9 18368.6 35968.2 31684.9 29085.3 41790.7 719_426 LMP2A_SNU- 41913.5 12690.2 31166.7 32767.4 34065.6 41039.7 719_429 LMP2A_SNU- 18858.1 2762.7 34930.4 14473.4 28951.9 17382.8 719_430 LMP2A_SNU- 5820 331.9 4419.9 14170.2 3.5 575.5 719_431 LMP2A_SNU- 30304.4 1675.5 20684.8 36242.1 12071.7 10316.5 719_432 LMP2A_SNU- 26470.9 14570.8 28749 4146.1 5216 5224.5 719_433 LMP2A_SNU- 5520.8 11126.7 28783.8 8098 946.5 14748.6 719_434 LMP2A_SNU- 21999.9 1453.3 31394.8 22179.2 13818.5 2582.2 719_439 LMP2A_SNU- 14075.1 1202 34902.1 18094.4 339 6061.1 719_440 LMP2A_SNU- 30682.5 1108.2 32059.4 17305.7 16675.9 18845.3 719_442 LMP2A_SNU- 12564 770.3 3529.2 16440 4.4 383.6 719_443 LMP2A_SNU- 41198.1 22402.5 35064.1 28311.3 32570.2 33655.3 719_446 LMP2A_SNU- 29325.5 3688.2 31123.2 12418.8 16408.9 19392.4 719_447 LMP2A_SNU- 10008.4 210.4 4177.6 15491.4 309.7 149.2 719_449 LMP2A_SNU- 9748.3 10058.9 24364.2 8275.5 7233.1 10872.3 719_451 LMP2A_SNU- 21963.6 41.1 20551.4 32421.1 3330.1 1209.6 719_452 LMP2A_SNU- 24710.9 6108.3 33208.9 19951.9 1093.2 12051.5 719_453 LMP2A_SNU- 37931.1 9963.8 38572.6 13793.5 39126.6 29860.5 719_457 LMP2A_SNU- 27325.6 5467.1 19946.5 32621 14748.5 7339.5 719-459 LMP2A_SNU- 36790.5 11054.1 32861.9 25757.5 25267.6 26034.4 719_460 LMP2A_SNU- 30790.6 17419.9 38901.6 20134.8 18600.8 33095.9 719_461 LMP2A_SNU- 48129.4 37637.2 36865.8 42648.6 44000.4 40388.2 719_463 LMP2A_SNU- 46970.9 39068.3 42482.3 39408.7 43194.2 37968.9 719_466 LMP2A_SNU- 17537.7 28276.4 40505.1 191.3 38119.6 337.5 719_7

(25) TABLE-US-00002 TABLE2 NeoepitopesforEBNA-1protein SEQ HLA- HLA- HLA- HLA- HLA- HLA- #ID (9mer) A*24:02 A*02:01 A*33:03 A*11:01 A*02:06 A*31:01 EBNA- RGR 45316.5 39680 4373 18740.2 34826.3 289.3 1_SNU- GGS 719_277 GGR (SEQ ID NO: 152) EBNA- RGR 45316.5 39680 4373 18740.2 34826.3 289.3 1_SNU- GGS 719_285 GGR (SEQ ID NO: 153) EBNA- RGR 45316.5 39680 4373 18740.2 34826.3 289.3 1_SNU- GGS 719_293 GGR (SEQ ID NO: 154) EBNA- RGR 46032.1 42644 3895.5 29673.7 38658.7 229.8 1_SNU- GRER 719_302 AR (SEQ ID NO: 155) EBNA- RAR 45029.1 39602.4 1219 10439.7 31841.9 86.1 1_SNU- GGSR 719_308 ER (SEQ ID NO: 156) EBNA- GSRE 47184.8 41384.4 1249.1 15361.7 37002.5 175.9 1_SNU- RAR 719_312 GR (SEQ ID NO: 157) EBNA- RAR 44722.2 39536 884.7 17784.2 33601.8 65.1 1_SNU- GRG 719_316 RGR (SEQ ID NO: 158) EBNA- RGR 44053.2 42854.4 3516.3 22983.4 38461.4 136.3 1_SNU- GRG 719_320 EKR (SEQ ID NO: 159) EBNA- RGR 45689.7 42293.4 4657.9 28956.6 38106 233.2 1_SNU- GEK 719_322 RPR (SEQ ID NO: 160) EBNA- SSSS 42342.4 36614.5 425.2 125.9 28232.3 177.9 1_SNU- GSPP 719_336 R (SEQ ID NO: 161) EBNA- HPV 31231.2 36176.7 33692.8 36983.6 22741.8 41710.8 1_SNU- GDA 719_355 DYF (SEQ ID NO: 162) EBNA- KGG 42456.6 37922.1 4856.2 10120 33694.3 124.7 1_SNU- WFG 719_409 KHR (SEQ ID NO: 163) EBNA- RGR 45006.2 42700.3 3668.5 27677.6 39018.4 188.3 1_SNU- GRG 719_41 RGR (SEQ ID NO: 164) EBNA- EGLR 42050.2 37488.5 187.6 7955.5 29022.1 987.3 1_SNU- VLLA 719_431 R (SEQ ID NO: 165) EBNA- RVLL 19512.6 2581.5 11516.2 16095.2 353.6 1075 1_SNU- ARSH 719_434 V (SEQ ID NO: 166) EBNA- LLAR 36376.1 18739.8 116.8 577.6 17709.7 53 1_SNU- SHVE 719_436 R (SEQ ID NO: 167) EBNA- RGR 45332.2 41211.9 3488.8 26198.8 37068.6 236.3 1_SNU- GRG 719_45 GGR (SEQ ID NO: 168) EBNA- GVF 44626.5 27517.3 9143.4 81.9 20473.5 1986.2 1_SNU- VYG 719_454 GSK (SEQ ID NO: 169) EBNA- KTSL 34794.6 26595.5 446.4 53.6 20319.7 18.8 1_SNU- YNL 719_462 RR (SEQ ID NO: 170) EBNA- IALA 39511.7 32880.4 527.2 2884.4 26713.5 229.8 1_SNU- VPQC 719_472 R (SEQ ID NO: 171) EBNA- ITPLS 1491.1 25301 24504 16803.1 10821.1 17909.2 1_SNU- RLPF 719_481 (SEQ ID No: 172) EBNA- RESI 13257.4 17730.8 23507 22586.8 4054.7 14022.8 1_SNU- VCYF 719_503 M (SEQ ID NO: 173) EBNA- ESIV 17345.4 5738.8 5835.7 18953.5 465.9 9913.9 1_SNU- CYF 719_504 MV (SEQ ID NO: 174) EBNA- SIVC 2026.1 6030.4 15283.4 5288.3 1655.6 10406.6 1_SNU- YFM 719_505 VF (SEQ ID NO: 175) EBNA- IVCY 26724.5 521.2 7210.1 10733.8 480.8 3174.7 1_SNU- FMV 719_506 FL (SEQ ID NO: 176) EBNA- FMV 10141.7 16.1 10488.4 28823.4 26.1 10274.1 1_SNU- FLQT 719_510 HI (SEQ ID NO: 177) EBNA- MVF 773.1 6982.1 6844.4 6047.9 2152.6 8273.3 1_SNU- LQTH 719_511 IF (SEQ ID NO: 178) EBNA- FLQT 34284.9 428.3 14139.7 18590.5 402.8 12823.6 1_SNU- HIFA 719_513 E (SEQ ID NO: 179) EBNA- LQTH 17941 61.6 16078.5 25417.3 9.8 12729.3 1_SNU- IFAE 719_514 V (SEQ ID NO: 180) EBNA- AIKD 39676.1 22195.5 2666.4 29.4 14992.6 322.1 1_SNU- LVM 719_526 TK (SEQ ID NO: 181) EBNA- NIKV 15027.2 26519.7 16872.7 27649.5 17243.3 20453.6 1_SNU- TVCS 719_540 F (SEQ ID NO: 182) EBNA- TVCS 40262.6 2096.8 22454.7 24077.5 380.9 24941.6 1_SNU- FDD 719_544 GV (SEQ ID NO: 183) EBNA- FPPM 43694.8 29057.6 36446.2 42153.1 18949.4 43126.1 1_SNU- VEG 719_558 AA (SEQ ID NO: 184) HLA- HLA- HLA- HLA- HLA- HLA- #ID B*51:01 B*15:01 B*44:03 B*54:01 B*58:01 B*35:01 EBNA- 47765.8 29769.2 45986.8 44280.7 39982.1 43547.6 1_SNU- 719_277 EBNA- 47765.8 29769.2 45986.8 44280.7 39982.1 43547.6 1_SNU- 719_285 EBNA- 47765.8 29769.2 45986.8 44280.7 39982.1 43547.6 1_SNU- 719_293 EBNA- 47725.5 32620.6 45707.5 43198 39720.8 41805.7 1_SNU- 719_302 EBNA- 45468.3 19127.1 42510.4 37732.6 27999.9 30362.9 1_SNU- 719_308 EBNA- 47577.5 32963.7 43767.7 45223 37929.5 45253.8 1_SNU- 719_312 EBNA- 45327.3 23182.2 42450.6 38545.9 32356.3 39243.2 1_SNU- 719_316 EBNA- 47659.9 36580.1 45330.7 42924.9 37426.4 44343.5 1_SNU- 719_320 EBNA- 48265.6 33970.3 46416.2 44423.2 39930.2 43241.9 1_SNU- 719_322 EBNA- 43513.7 23101.1 39692.5 36562.7 27088.4 22741.3 1_SNU- 719_336 EBNA- 12468.9 13575.1 25926.7 10752.9 9492.1 13.2 1_SNU- 719_355 EBNA- 48473.3 35691.1 44147.2 45552.5 34497.4 44034.7 1_SNU- 719_409 EBNA- 47327.5 33030.8 45445.2 44144.4 39900.9 44790.1 1_SNU- 719_41 EBNA- 43250.4 39180.9 40538 35094.5 36902.5 29032.2 1_SNU- 719_431 EBNA- 22726.3 5093.3 32701.6 14056.7 7788.7 35120.3 1_SNU- 719_434 EBNA- 45126.7 22087.2 40498.5 35766.4 32575.8 26860.1 1_SNU- 719_436 EBNA- 47258.9 32002.9 45779.3 43911.9 41157.1 44732.4 1_SNU- 719_45 EBNA- 41747.3 17980.8 36473 31707.9 30633.8 30968.7 1_SNU- 719_454 EBNA- 45634.8 29342.9 38073.8 40031.5 13160.8 39576.3 1_SNU- 719_462 EBNA- 37853.6 31599.6 37944.3 32340.2 17579.5 24121.8 1_SNU- 719_472 EBNA- 26114.5 438.1 35685.7 33692.8 3771.2 2754 1_SNU- 719_481 EBNA- 33839.3 5262.5 440 26568.8 11335.6 16703 1_SNU- 719_503 EBNA- 20573.9 28021.1 28717.3 9180.2 7673.5 23247.5 1_SNU- 719_504 EBNA- 24843.9 52.8 11265 23057.2 8440 813.6 1_SNU- 719_505 EBNA- 34592.3 12094.1 39419 32020.9 11647.3 22473.2 1_SNU- 719_506 EBNA- 10650.8 402 22139.2 16201.6 5493.7 11412.7 1_SNU- 719_510 EBNA- 4430.3 28.8 10520 7239.1 217.3 20.3 1_SNU- 719_511 EBNA- 39689 12298.9 41389.8 23759.7 38950.5 15521.1 1_SNU- 719_513 EBNA- 17980.6 1123 14996.1 11684.8 17726.8 17732.5 1_SNU- 719_514 EBNA- 44286.5 16408 35567.3 36968.8 30457 32160.8 1_SNU- 719_526 EBNA- 29037.2 154.9 24316.2 34414.2 16844.2 1653.5 1_SNU- 719_540 EBNA- 34793.9 28022.7 35086.1 24905.8 21011.9 27572.1 1_SNU- 719_544 EBNA- 7576.1 30308.4 45375.9 48.7 39527.1 418.3 1_SNU- 719_558

(26) As shown in Tables 1 and 2 above, through in silico prediction, neoepitopes were identified which have high binding affinity for HLA-A*2402, HLA-A*A0201, HLA-A*3303, HLA-A*1101, HLA-A*0206, HLA-A*3101, HLA-B*5101, HLA-B*4403, HLA-B*5401, HLA-B*5801, or HLA-B*3501, and their binding affinity with respective HLA's was predicted by IC.sub.50 (nM) values. Sequences present in actual human gastric cancer cells and recognizable by T cells in the body were selected based on their binding affinity with the HLA's. As shown in Table 1, regarding the neoepitopes for LMP-2A, in a case of HLA-A*2402, TYGPVFMCL (IC.sub.50=43.1 nM), LLWTLVVLL (IC.sub.50=22368.6 nM), LTEWGSGNR (IC.sub.50=45205.3 nM) were selected in order of high to low binding affinity therefor, and in a case of HLA-A*3101, LAYRRRWRR (IC.sub.50=5.2 nM), LTVMTNTLL (IC.sub.50=14004.8 nM), YPSASGSYG (IC.sub.50=42032 nM) were selected. In addition, as shown in Table 2, regarding the neoepitopes for EBNA-1, in a case of HLA-A*2402, MVFLQTHIF (IC.sub.50=773.1 nM), AIKDLVMTK (IC.sub.50=39676.1 nM), GSRERARGR (IC.sub.50=47184.8 nM) were selected in order of high to low binding affinity therefor, and in a case of HLA-A*3101, KTSLYNLRR (IC.sub.50=18.8 nM), RGRGGSGGR (IC.sub.50=289.3 nM), FPPMVEGAA (IC.sub.50=43126.1 nM) were selected. The neoepitopes selected as above were synthesized into peptides for the following experiments.

(27) 2. ELISPOT Results for T Cells Activated by Dendritic Cells Loaded with Selected Neoepitope

(28) PBMCs extracted from healthy human blood were separated into monocytes and leukocytes through flow cytometry, and the monocytes were cultured for 2 days in a culture supplemented with cytokines GM-CSF and IL-4 to differentiate into dendritic cells. In addition, the leukocytes were cultured with anti-CD3/CD28 antibody for 3 days, and then cultured in a culture supplemented with cytokine IL-2. The neoepitope peptide selected as above was transferred to the monocyte-differentiated dendritic cells using electroporation. Subsequently, culture was performed for 2 days to identify that the neoepitope has been expressed on the surface of the dendritic cells. Then, the dendritic cells were co-cultured with the leukocytes, which were cultured in a culture supplemented with anti-CD3/CD28 antibody, at a ratio of 1:20 (dendritic cells:leukocytes). In a case of the co-culture, the culture was mixed with a cytokine cocktail containing both cytokine IL-4 that increases the antigen-presenting function of dendritic cells, and cytokines IL-2 and IL-7 that function to help conversion of T cells into memory cells, and culture was performed. After 16 hours, expression levels of IFN- in the T cells thus activated were measured with ELISPOT, and the results are illustrated in FIGS. 1 to 4. Here, for a negative control peptide used in the experiments, the amino acid sequence 9-mer (sequence: GGSRERARG) was employed which is of any EBNA-1 or LMP-2A protein that has not been extracted through NetMHC in EBNA-1 and LMP-2A.

(29) As a result, it was found that the T-cells cultured with dendritic cells loaded with a neoepitope peptide of the present invention secrete much more IFN- than the control regardless of binding affinity of the peptide for HLA.

(30) From these results, it was found that in the present invention, cytotoxic T lymphocytes (CTLs) can be activated by the dendritic cells loaded with each of neoepitopes, which have been selected by HLA-A types of respective cell lines in Tables 1 and 2 above, and the thus activated T cells have antigen specificity which enables recognition of the neoepitope that is a neoantigen.

(31) 3. Cancer Cell Lysis Effect of T Cells Activated by Dendritic Cells Loaded with Selected Neoepitope

(32) In order to select memory T cells to which an antigen had been presented through the dendritic cells after co-culture for 72 hours as in item no. 2 above, a magnetic-activated cell sorter (MACS) capable of extracting T cells secreting cytokine IFN- was used to extract EBV antigen-specific memory T cells. The extracted memory T cells were cultured in a culture supplemented with cytokines IL-2, IL-7, and IL-15 to maintain their memory function and increase the number of cells, in which the culture was performed until the memory T cells reach the number of cells that can be injected into mice.

(33) Such activated T cells were co-cultured with SNU-719 cell line, that is, EBV-positive gastric cancer cells, and with EBV-infected MKN74 cell line, to identify lysis of the cancer cells through Cr51 release assay. The results are illustrated in FIGS. 5 to 8. Also, such activated T cells were co-cultured with MKN-74 cell line, that is, EBV-negative gastric cancer cells, to identify lysis of the cancer cells through Cr51 release assay. The results are illustrated in FIGS. 9 to 12. Here, for a negative control peptide used in the experiments, the amino acid sequence 9-mer (sequence: GGSRERARG) was employed which is of any EBNA-1 or LMP-2A protein that has not been extracted through NetMHC in EBNA-1 and LMP-2A.

(34) As a result, it was found that the T cells activated by the neoepitope according to the present invention exhibit excellent lytic ability against EBV-positive gastric cancer cells while exhibiting insignificant lytic ability against EBV-negative gastric cancer cells. In other words, it can be seen that the T cells activated by the dendritic cells loaded with the EBV cancer-specific antigen neoepitope according to the present invention specifically lyse EBV-positive cancer cells.

(35) In addition, the T cells activated by the neoepitope according to the present invention tended to show an increase in the lytic ability, in a mixed-concentration dependent manner, especially in a case of being co-cultured with EBV-positive gastric cancer cells; and it was found that in a case where, among the neoepitopes for respective HLA types, in particular, a neoepitope with higher binding affinity with HLA is used, the activated T cells exhibit a more increased lytic ability against EBV-positive gastric cancer cell line.

(36) 4. Identification of In Vivo Anticancer Effect of T Cells Activated by Dendritic Cells Loaded with Selected Neoepitope

(37) The EBV-positive gastric cancer cell line, SNU-719 (HLA type: HLA-A*2402), or EBV-infected MKN74 (HLA type: HLA-A*3101), in an amount of 110.sup.7 cells, was mixed with Matrigel, and the mixture was transplanted into the flank of BALB/c nude mice, to produce a xenograft model. After 4 weeks had elapsed, in a case where cancer tissue is observed, the T cells activated in item no. 2 above, in an amount of 110.sup.7 cells, were intratumorally injected once a week, and then the size of cancer tissue was measured. The results are illustrated in FIGS. 13 to 16. Here, for a negative control peptide used in the experiments, the amino acid sequence 9-mer (sequence: GGSRERARG) was employed which is of any EBNA-1 or LMP-2A protein that has not been extracted through NetMHC in EBNA-1 and LMP-2A.

(38) As a result, it was found that in a case where activated T cells are subjected to treatment with the neoepitope according to the present invention, the size of EBV-positive gastric cancer is remarkably decreased as compared with the control; and it was found that in a case where, among the neoepitopes for respective HLA types, in particular, a neoepitope with higher binding affinity with HLA is used, a higher rate of decrease in size of gastric cancer is exhibited.

(39) As described above, it was found that all neoepitopes selected by the HLA-A type of each cell line exhibit an excellent effect in enhancing activity of T cells, and that as binding affinity obtained by bioinformatics increases (that is, IC.sub.50 value decreases), activity of the T cells is enhanced and activity thereof for targeted cancer cell killing is increased. From these results, it is easily predictable that dendritic cells together with the neoepitopes shown in Tables 1 and 2 can be used to enhance activity of T cells, and this also results in an excellent targeted cancer therapeutic effect.

(40) Although specific parts of the present invention have been described in detail as above, it is obvious to those skilled in the art that such a specific description is merely a preferred embodiment, and the scope of the present invention is not limited thereto. Therefore, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

INDUSTRIAL APPLICABILITY

(41) The present invention relates to a cancer-specific tumor antigen neoepitope, an antigen-presenting cell loaded with the neoepitope, and a method for activating T cells for cancer treatment using the antigen-presenting cell.