Project 5: Enhancement of the Anti-Tumor Activity and Targeted Applications of Third Party Donor-Derived EBV-specific T-cells

用抗原特异性T细胞或经工程改造以表达嵌合抗原的T细胞的连续免疫疗法 受体（CAR）已成为治疗药物难治性病毒感染的潜在治愈方法 和同种异体造血细胞移植（HCT）后B系恶性肿瘤的复发。我们的节目 最近提供的证据表明，EB病毒特异性的，适当的HLA限制性T细胞从HLA部分 匹配的第三方供者也可以诱导70%的allo-HCT受者的持久完全或部分缓解 61%的实体器官移植（SOT）受者对Rituxan耐药，SOT患者接受化疗 难治性EBV+淋巴瘤。这与我们的实验一致，即同种异体反应性T细胞在 自体EBV+ BLCL的重复体外致敏过程，第三方EBVCTL的过继转移 与同种异体移植物功能受损或GVHD无关。引人注目的是，细胞因子释放 也没有观察到综合征。 在这个项目中，我们提出了战略，以扩大和加强银行第三方捐助者的应用， EBV特异性CTL，目前可以提供有效的和立即可用的，“现成的” 用于alloHCT和SOT患者的过继治疗，用于其他EBV相关恶性肿瘤患者，包括 血液疾病如EBV+霍奇金斯病、NK T细胞淋巴瘤、HLH和伯基特淋巴瘤， 以及EBV阴性B细胞谱系白血病和淋巴瘤，其指示了alloHCT。项目 将在体外和临床前模型中测试三种假设：1）表观遗传修饰剂如5-氮杂胞苷， 地西他滨和HDAC抑制剂伏立诺他和罗米地辛已经被发现诱导潜伏性感染， EBV+恶性肿瘤表达EBV的裂解周期基因，从而使它们对更昔洛韦敏感， 用于低毒性的重复短时间暴露，以诱导潜伏期I和II型恶性肿瘤，如Burkitt 淋巴瘤、NK T细胞淋巴瘤、EBV+霍奇金斯病和鼻咽癌中表达潜伏期 3和早期裂解性EBV蛋白如BZLF-1，从而使它们对EBV特异性、HLA限制性 用主要含有T细胞的自体EBV转化的BLCL致敏的第三方CTL 2）有限的第三方CD 19 CAR+ EBVCTL库可以提供即时和特异性的免疫应答， 对大多数移植后复发的CD 19 + B系ALL或DLBCL患者的有效过继治疗 没有GVHD。此外，它们的抗肿瘤活性和持久性可以通过汽车进一步增强 指导IL-12的表达。3)CD 19 CAR+ EBVCTL和CD 19 CAR+ EBVCTL的抗肿瘤活性 分泌IL-12的EBVCTL可以通过以下方式进一步增加：a）用低剂量的表观遗传修饰的预输注治疗， 修饰剂，从而改变活化和抑制配体的肿瘤表达，从而增强它们的敏感性 和/或B）共施用检查点抑制剂以增强CD 19 CAR+、EBVCTL的能力， CD 19 CAR+ EBVCTL接合并裂解B谱系ALL和DLBCL。

Adoptive immunotherapy with antigen-specific T cells or T cells engineered to express a chimeric antigen receptor (CAR) has emerged as a potentially curative approach to the treatment of drug refractory viral infections and relapses of B lineage malignancies following allogeneic hematopoietic cell transplants (HCT). Our program has recently provided evidence that EBV-specific, appropriately HLA restricted T cells from HLA partially matched third party donors can also induce durable complete or partial remissions in 70% of allo-HCT recipients and 61% of solid organ transplant (SOT) recipients with Rituxan resistant and, in SOT patients, chemotherapy refractory EBV+ lymphomas. Consistent with our demonstration that alloreactive T-cells are depleted in the course of repeated in vitro sensitizations with autologous EBV+ BLCL, adoptive transfer of 3rd party EBVCTL has not been associated with either impairment of allograft function or GVHD. Strikingly, cytokine release syndrome has also not been observed. In this project, we propose strategies to extend and enhance the application of banked third party donorderived EBV-specific CTLs that currently can provide effective and immediately accessible, “off the shelf” adoptive therapies for alloHCT and SOT patients, to patients with other EBV-associated malignancies including hematologic diseases such as EBV+ Hodgkins disease, NK T cell lymphomas, HLH and Burkitt lymphomas as well as EBV negative B cell lineage leukemias and lymphomas for which an alloHCT is indicated. The project will test in vitro and in preclinical models, three hypotheses: 1) Epigenetic modifiers such as 5-azacytidine, decitabine and the HDAC inhibitors, vorinostat and romidepsin which have been found to induce latently infected EBV+ malignancies to express lytic cycle genes of EBV so as to render them susceptible to ganciclovir can be used in repeated short exposures of low toxicity to induce latency I and II malignancies, such as Burkitt lymphomas, NK T cell lymphomas, EBV+ Hodgkins disease and nasopharyngeal carcinoma, to express latency 3 and early lytic EBV proteins such as BZLF-1, thereby rendering them susceptible to EBV-specific, HLArestricted 3rd party CTLs sensitized with autologous EBV transformed BLCLs that predominantly contain T cells specific for these EBV antigens; 2) A limited bank of 3rd party CD19 CAR+ EBVCTL can provide immediate and effective adoptive therapy for most patients relapsing with CD19+ B lineage ALL or DLBCL post transplant without GVHD. Furthermore, their anti-tumor activity and persistence can be further augmented by CARS directing the expression of IL-12. 3) The anti-tumor activity of the CD19 CAR+ EBVCTLs and CD19 CAR+ EBVCTLs secreting IL-12 can be further increased by a) pre-infusion treatment with low doses of epigenetic modifiers so as to alter tumor expression of activating and inhibitory ligands, and thereby enhance their sensitivity to the CD19 CAR+, EBVCTLs, and/or b) co-administration of a checkpoint inhibitory to enhance the capacity of the CD19 CAR+ EBVCTLs to engage and lyse B lineage ALL and DLBCL.