Immunotherapy of leukemias by targeting hematopoietic lineage-specific minor histocompatibility antigens

次要组织相容性抗原(MHAgs)特异性细胞毒性T淋巴细胞(CTL)的组织表达仅限于造血细胞，可用于异基因造血细胞移植(HCT)后复发白血病的免疫治疗。我们采集了研究期间接受HCT的26名患者的外周血样本，并试图产生受人类白细胞抗原-A24(日本最常见的人类白细胞抗原)限制的CTL。为了方便这一过程，我们设计了一种新的方法，通过逆转录病毒和基于干扰素-γ释放的检测来筛选感兴趣的人类白细胞抗原(HLA)所限制的CTL系中mHAgs特异性CTL的存在。我们成功地获得了4个不同的人类白细胞抗原-A24限制性CTL克隆。利用遗传标记分型结合连锁分析的CEPH大家系成员LCL，将编码mHAgs的基因(S)定位在染色体15q24.5上。在该区域内的39个基因中，我们利用各种在线数据库(DBSNP、SAGE、UCSC Human Genome Browser)和软件(BIMAS)确定了mHAgs的编码基因。该基因BCL2A1有三个单核苷酸多态，其中两个编码由两个CTL克隆检测到的两个mHAgs表位。BCL2A1仅在造血细胞中表达，因此被认为是白血病免疫治疗的良好候选者。我们已经成功地利用多肽冲击的抗原提呈细胞诱导了针对BCL2A1 mhag的CTL系。我们计划使用针对BCL2A1 mHAgs的体外扩增CTL克隆，对异基因HCT后复发白血病患者进行过继免疫治疗的I期临床研究。

Cytotoxic T lymphocytes (CTL) specific for minor histocompatibility antigens (mHAgs) whose tissue expression is limited to hematopoietic cells are useful for immunotherapy of relapsed leukemia following allogeneic hematopoietic cell transplantation (HCT). We have obtained peripheral blood samples from 26 patients receiving HCT during the research period and attempted to generate CTL restricted by HLA-A24, the most common HLA in Japanese. To facilitate the procedure, we devised a novel method to screen for the presence of mHAgs-specific CTL restricted by HLA of interest in CTL lines by using HLA transfected B lymphoblastoid cell lines retrovirally and interferon-gamma release-based assays. We successfully generated 4 distinct HLA-A24-restricted CTL clones. By using LCL from members of large CEPH families whose genetic markers had been typed in combination with linkage analysis, gene(s) encoding mHAgs detectable 2 distinct CTL restricted by HLA-A24 and B44 were located at chromosome 15q24.5. Of 39 genes within the region, we determined the gene encoding the mHAgs using various online databases (dbSNP, SAGE, UCSC Human Genome Browser) and softwares (BIMAS). The gene, BCL2A1, possesses three single nucleotide polymorphisms, two of which turned out to encode two mHAgs epitopes detected by the 2 CTL clones used. BCL2A1 is expressed in only hematopoietic cells, thus is thought to be a good candidate for immunotherapy of leukemia. We have successfully induced CTL lines specific for the BCL2A1 mHAg using peptide-pulsed antigen presenting cells. We plan on conducting a phase I clinical study of adoptive immunotherapy for patients with relapsed leukemia following allogeneic HCT using ex vivo expanded CTL clones specific for the BCL2A1 mHAgs.