Virus Specific CTL following T cell depleted SCT

描述(由申请人提供)：异基因干细胞移植(SCT)的T细胞耗竭(TCD)显著降低移植物抗宿主病(GVHD)的风险，但导致感染并发症的风险增加，其中最严重的两种是EB病毒(EBV)引起的淋巴增生性疾病和巨细胞病毒(CMV)感染。以前的研究表明，细胞毒性T淋巴细胞可以被培养并注入到SCT受者体内，从而导致抗原特异性免疫重建，但这可能是不现实的，因为需要用单独的抗原提呈细胞来扩增CTL。初步研究表明，逆转录病毒转导B淋巴母细胞系(BLCL)表达CMV pp65是可行的，表达pp65的BLCL可扩增识别CMVpp65和EBV抗原的CTL。在这项建议中，我们将比较预防性CMV/EBV特异性CTL在接受同种异体TCD SCT的受者中的临床、免疫学和病毒学效果，并与随机选择不接受CTL的TCD SCT患者进行对照。移植后每隔一段时间将比较两组患者的CD4和CD8抗原特异性免疫重建(CTL前体频率)以及CMV和EBV DNA水平(实时定量聚合酶链式反应)。我们将使用编码pp65缺失突变的重组牛痘疫苗来确定哪些CMVpp65表位在干细胞捐赠者中具有免疫优势，然后使用脉冲到APC上的合成肽来确定确切的表位。CTL供者的优势表位将在输血后与CTL接受者中发现的表位进行比较，以确定供者识别的免疫优势表位是否保留在接受者身上，如果不是，哪些表位在移植后最相关。CTL受者和对照组在CMV再激活过程中，效应细胞的类型和CD4和CD8 CMV特异性T细胞的动态变化也是有趣的。这些研究将确定预防性抗原特异性CTL对病毒再激活和免疫重建的影响，并将为将来将这一抗原提呈和T细胞扩增系统扩展到其他病原体和肿瘤抗原奠定基础。

DESCRIPTION (provided by applicant): T cell depletion (TCD) of allogeneic stem cell transplants (SCT) dramatically reduces the risk of graft vs host disease (GVHD) but results in a heightened risk for infectious complications, two of the most serious being Epstein Barr virus (EBV) induced lymphoproliferative disease and cytomegalovirus (CMV) infection. Previous studies have shown that cytotoxic T lymphocytes can be cultured and infused into SCT recipients resulting in antigen specific immune reconstitution, but this can be impractical due to the need of expanding CTL with separate antigen presenting cells. Preliminary studies have demonstrated the feasibility of retroviral transduction of B lymphoblastoid cell lines (BLCL) to achieve expression of CMV pp65, and stimulation with BLCL expressing pp65 results in expansion of CTL recognizing both CMVpp65 and EBV antigens. In this proposal, we will compare the clinical, immunologic, and virologic effects of prophylactic CMV/EBV specific CTL in recipients of allogeneic, TCD SCT with a control group of TCD SCT patients randomized to not receive CTL. CD4 and CD8 antigen specific immune reconstitution (CTL precursor frequencies) and CMV and EBV DNA levels (by real time PCR) will be compared in these groups at intervals post-transplant. We will determine which CMVpp65 epitopes are immunodominant in stem cell donors using recombinant vaccinia encoding pp65 deletion mutations, and then the exact epitopes will be determined using synthetic peptides pulsed onto APC. Dominant epitopes of the CTL donors will be compared with those found in CTL recipients post-infusion, to determine whether immunodominant epitopes recognized by the donor are retained in the recipient, and if not, which epitopes are most relevant post-transplant. Also of interest are the types of effector cells and the dynamic changes in CD4 and CD8 CMV specific T cells during CMV reactivation in recipients of CTL and the control group. These studies will determine the impact on viral reactivation and immune reconstitution of prophylactic antigen-specific CTL and will serve as a groundwork for extending this system of antigen presentation and T cell expansion to other pathogens as well as tumor antigens in the future.