Development of novel immunogene therapy for hematological malignancies

本研究旨在开发治疗血液系统恶性肿瘤的新型免疫疗法。1)建立了一个新的WT1衍生肽特异性CD8^+CTL细胞系，命名为NIM-1。NIM-1可裂解HL A-A24阳性的白血病细胞，但不能裂解HL A-A24阴性的白血病细胞或正常细胞。2)WT1特异性的、HL A-A24限制性的CTL克隆(命名为TAK-1)对携带HL A-A24的肺癌细胞株有杀伤作用，但不能裂解缺乏这种HL A的细胞。将TAK-1过继转移到移植有人类白细胞抗原A24阳性肺癌细胞系的裸鼠体内，可抑制癌细胞生长，延长生存期。这些发现有力地表明WT1是一种通用的肿瘤相关抗原，WT1靶向免疫治疗为肺癌和白血病提供了一种潜在的有效治疗选择。3)未成熟的树突状细胞(DC)装载了带有t(6；9)或t…的白血病细胞更多(9；22)，然后与DEK-CAN融合肽特异性或BCR ABL融合肽特异性CD4^+T淋巴细胞克隆共培养。DEK-CAN和BCR-ABLCD4+T淋巴细胞克隆分别与HLADR相合但不相合的树突状细胞(DC)共同培养时产生干扰素-γ(γ)。这些数据表明，急性髓系白血病相关融合蛋白DEK-CAN和慢性髓系白血病相关融合蛋白BCR-ABL都是由DC加工并呈递给融合多肽特异性的CD4^+T淋巴细胞。4)为了阐明Perform在人类CD4^+CTL介导的抗原特异性细胞毒中的作用，从一名遗传性穿孔素缺乏症患者中建立了抗原特异性的人CD4^+T淋巴细胞克隆，并对其细胞毒活性进行了研究。结果表明，穿孔素阴性的CD4^+CTL对Fas敏感的靶细胞具有杀伤作用，而穿孔素在人CD4^+和CD8^+CTL介导的抗原特异性杀伤中起重要作用。较少

This study was performed to develop the novel immunotherapy for hematological malignancies. The data obtained from the series of experiments are as follows.1) A novel WT1-derived peptide-specific CD8^+ CTL line, designated NIM-1 was established. NIM-1 lysed HLA-A24-positive leukaemia cells, but not HLA-A24-negative leukaemia cells or normal cells.2) A WT1-specific, HLA-A24-restricted CTL clone (designated TAK-1) exhibited cytotoxicity against lung cancer cell lines bearing HLA-A24 but did not lyse cells lacking this HLA. Adoptive transfer of TAK-1 into nude mice that had been engrafted with an HLA-A24-positive lung cancer cell line resulted in inhibition of the cancer cell growth and prolonged survival. These findings strongly suggest that WT1 is a universal tumor-associated antigen and that WT1 -targeting immunotherapy offers a potentially effective treatment option for lung cancer as well as leukemia.3) Immature dendritic cells (DCs) were loaded with leukemia cells with t(6 ; 9) or t … More (9 ; 22) and then cocultured with the dek-can fusion peptide-specific or the bcr abl fusion peptide-specific CD4^+ T-lymphocyte clone. The dek-can peptide-specific and bcr-abl peptide-specific CD4^+ T-lymphocyte clones produced interferon-γ (IFN-γ) when they were cocultured with HLA-DR-matched but not with mismatched DCs which had been loaded with apoptotic as well as necrotic leukemia cells with t(6 ; 9) and t(9 ; 22), respectively. These data indicate that the acute myelogenous leukemia-associated fusion protein, dek-can and chronic myelogenous leukemia-associated fusion protein, bcr-abl, are both processed and presented by DCs to the fusion peptide-specific CD4^+ T lymphocytes.4) In order to clarify the roles of perform in antigen-specific cytotoxicity mediated by human CD4^+ CTLs, antigen-specific human CD4^+ T-lymphocyte clones were established from a patient with hereditary perforin deficiency and their cytotoxic activities were investigated. The data demonstrated that perforin-negative CD4^+ CTLs can exert cytotoxicity against Fas-sensitive target cells ; however, perforin plays essential roles in antigen-specific cytotoxicity mediated by human CD4^+ as well as CD8^+ CTLs. Less