Induction of specific cellular immunity to EBV-positive T- and NK-lymphomas

诱导针对 EBV 阳性 T 和 NK 淋巴瘤的特异性细胞免疫

• 批准号: 12670802
• 负责人: KUZUSHIMA Kiyotaka
• 金额: $ 1.73万
• 依托单位: Aichi Cancer Center
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670802/
• 关键词: Epstein-Barr virus; CTL; HLA-A24; MHC-peptide tetramer; lymphoma; cytotoxic T lymphocyte; tetrameric MHN-peptide complex; HLA-A^*2402; LMP2; トランスレーショナルリサーチ; LMP1; TAT

EBV is associated with several malignant diseases, including a subset of lymphomas, nasopharyngeal carcinomas, Burkitt's lymphomas and immunoblastic lymphomas seen in immunocompromised hosts. We established an efficient approach for determination of CTL epitopes in the context of HLA A^*2402 molecules through multiple screenings, consisting of a computer-assisted algorithm, an in vitro MHC stabilization assay and an enzyme-linked immuno-spot (ELISPOT) assay. HLA A24 is the most frequently encountered HLA class I allele and the genotype is almost exclusively A^*2402. We first searched for candidate,peptides with the HLA A24-binding motif by a computer-assisted algorithm among EBV proteins reported to be targeted by specific CD8^+ T cells. We selected and synthesized 42 peptides after analyzing amino acid sequences of the 5 lytic and 6 latent cycle proteins. All of them functionally stabilized HLA A^*2402 molecules which had been expressed on the peptide transporter-deficient cell line T2. Next is screening of the peptides by ELISPOT assay for their recognition by bulk EBV-specific CD8^+ T cells, established from PBMCs of EBV-immune donors positive for HLA A24.-typing. We have identified 2 possible major lytic cycle EBV-specific CTL epitopes from the amino acid sequences of BRLF1 and BMLF1 and 1 from LMP2 presented by HLA A^*2402 molecules. The newly identified peptides should prove useful for detection and/or study of EBV-specific CD8^+ T cell responses among populations positive for HLA A^*2402. We produced HLA A^*2402tetramers-incorporating the 3 peptides and 2 known epitope peptide, one derived from EBNA3A (RYSIFFDYM), and the other derived from EBNA3B (TYSAGIVQI). All the 5 tetramers successfully stained various proportions of EBV-specific CD8+ T cells. The newly identified peptides should prove useful for detection and/or study of EBV-specific CD8^+ T cell responses among populations positive for HLAA^*2402.

EBV与几种恶性疾病有关，包括淋巴瘤的一部分，鼻咽癌，伯基特的淋巴瘤和免疫细胞淋巴瘤在免疫功能低下的宿主中可见。我们通过多个筛选建立了一种有效的方法，用于在HLA A^*2402分子的背景下确定CTL表位，包括计算机辅助算法，一种体外MHC稳定化测定法和酶 - 链接的Immuno Spot（ELISPOT）测定法。 HLA A24是最常见的HLA I类等位基因，基因型几乎完全是A^*2402。我们首先搜索了候选者，通过计算机辅助算法在EBV蛋白中具有HLA A24结合基序的肽，据报道是由特定CD8^+ T细胞靶向的。分析了5个裂解和6个潜在循环蛋白的氨基酸序列后，我们选择并合成了42个肽。所有这些都在功能上稳定的HLA A^*2402分子在肽转运蛋白缺陷型细胞系T2上表达。接下来是通过ELISPOT测定法对肽进行筛查，以通过EBV特异性CD8^+ T细胞识别，这些细胞是由EBV-immune供体的PBMC确定的，其hla a24.-typ.-typing。我们已经从BrlF1和BMLF1的氨基酸序列和1个来自HLA A^*2402分子提出的LMP2的氨基酸序列中确定了2个可能的主要裂解周期EBV特异性CTL表位。新确定的肽应被证明可用于检测和/或研究HLA A^*2402阳性的人群中EBV特异性CD8^+ T细胞反应。我们生产了HLA A^*2402TETRAMER，其中包括3种肽和2种已知的表位肽，一个肽来自EBNA3A（Rysiffdym），另一个源自EBNA3B（Tysagivqi）。所有5个四聚体成功染色了EBV特异性CD8+ T细胞的各种比例。新鉴定的肽应被证明可用于检测和/或研究HLAA^*2402阳性的人群中EBV特异性CD8^+ T细胞反应。

项目成果

葛島 清隆: "EBV感染症におけるウイルス特異的CD8+T細胞"臨床免疫. 34. 231-238 (2000)

Kiyotaka Katsurashima：“EBV 感染中的病毒特异性 CD8+ T 细胞”《临床免疫学》34. 231-238 (2000)。

Kuzushima K: "Longitudinal dynamics of EBV-specific CTL during the posttransplant lymphoproliferative disorder"Journal of Infections Disease. 182. 937-940 (2000)

Kuzushima K：“移植后淋巴增殖性疾病期间 EBV 特异性 CTL 的纵向动态”感染疾病杂志。

Hoshino Y: "Antigen-driven expansion and contraction of CD8^+ activated T cell in primary EBV infection"J.Immunol. 163・10. 5735-5740 (1999)

Hoshino Y：“原发性 EBV 感染中抗原驱动的 CD8^+ 激活 T 细胞的扩张和收缩”J.Immunol。163・10（1999）。

Yokoyama N, Hirata M, Ohtsuka K, Nishiyama Y, Fujii K, Fujita M, Kuzushima K, Kiyono T, and Tsurumi T: "Co-expression of human Chaperone hsp 70 and Hsdj or hsp 40 co-factor increase solubility of overexpressed Targwt proteins in insect cells"Biochim, Biop

Yokoyama N、Hirata M、Ohtsuka K、Nishiyama Y、Fujii K、Fujita M、Kuzushima K、Kiyono T 和 Tsurumi T：“人类伴侣 hsp 70 和 Hsdj 或 hsp 40 辅因子的共表达增加了过表达 Targwt 的溶解度

葛島清隆: "EBV関連lymphoproliferative disease:診断と治療の最近の動向."化学療法の領域. 15・9. 49-53 (1999)

Kiyotaka Katsurashima：“EBV 相关淋巴增殖性疾病：化疗的最新趋势”15・9（1999）。