Development of novel immunogene therapy for hamatopietic malignancies

我们进行了一系列的实验，得到了以下数据：1)骨髓瘤细胞和淋巴瘤细胞WT1表达均呈弱阳性。然而，只有骨髓瘤细胞似乎对由wt1特异性细胞毒性T淋巴细胞（ctl）介导的穿孔依赖性细胞毒性敏感。这些数据表明，膜对穿孔素的敏感性是决定靶细胞对ctl介导的细胞毒性敏感性的重要因素，WT1是多发性骨髓瘤细胞免疫治疗的理想靶抗原。2)来自hla - a24限制性wt1特异性CTL克隆的T细胞受体（TCR）基因被转导到CD4+和CD8+外周血T淋巴细胞中。因此，TCR基因转导的T淋巴细胞可以发挥hla - a24限制性和WT1特异性反应性。因此，TCR基因修饰hla i类限制性Th1和Tc1细胞是应用于人类癌症过继性免疫治疗的有力策略。3)将人造血细胞移植到新生NOD/SCID/IL2rgamma（null）小鼠体内。在这些小鼠体内重建了人类免疫系统。因此，NOD/SCID/IL2R γ（零）新生儿系统可能是研究人类血淋巴系统的重要实验模型。4)我们确定了wt1衍生的辅助表位，该表位可被HLA ii类限制性CD4+ ctl识别。wt1特异性CD4+ T淋巴细胞能以HLA ii类限制的方式直接识别白血病细胞。5)建立cml66特异性和aurora - a特异性CTL系。由于这些ctl以hla限制的方式裂解白血病细胞，CML66和Aurora-A似乎是人类ctl识别的新型白血病相关抗原。6)采用WT1肽和hTERT肽接种癌肽。未发现不良事件，部分患者有明显的抗癌作用。

We performed the series of experiments and obtained the following data. 1) Myeloma cells and lymphoma cells were both weakly positive for WT1 expression. However, only myeloma cells appeared to be sensitive to perforin-dependent cytotoxicity mediated by WT1-specific cytotoxic T lymphocytes (CTLs). These data suggest that susceptibility of membranes to perforin is an important factor determining the sensitivity of target cells to CTL-mediated cytotoxicity and that WT1 is an ideal target antigen for cellular immunotherapy of multiple myeloma..2) T-cell receptor (TCR) genes derived from HLA-A24-restricted WT1-specific CTL clone were transduced into CD4+ and CD8+ peripheral blood T lymphocytes. Consequently, TCR gene-transduced T lymphocytes could exert HLA-A24-restricted and WT1 specific reactivity. Thus, TCR gene-modified HLA-class I-restricted Th1 and Tc1 cells are a powerful strategy for the application to adoptive immunotherapy of human cancer. 3) Human hematopietic cells were transferred into new born NOD/SCID/IL2rgamma(null) mice. Human immune system was reconstituted in these mice. Thus, the NOD/SCID/IL2R gamma(null) newborn system might be an important experimental model to study the human hemato-lymphoid system. 4) We identified WT1-derived helper epitope which is recognized by HLA class II-restricted CD4+ CTLs. WT1-specific CD4+ T lymphocytes could directly recognized leukemia cells in an HLA class II-restricted manner. 5) We established CML66-specific and Aurora-A-specific CTL lines. Since these CTLs lysed leukemia cells in HLA-restricted manner, CML66 and Aurora-A appeared to be novel leukemia-associated antigens which are recognized by human CTLs. 6) Cancer peptide vaccination using WT1 peptide and hTERT peptide has been performed. No adverse event was detected, and apparent anti-cancer effect was observed in some patients.