Project 2: Regulatory T Cells Generated by Gene Transfer to Prevent GVHD

项目2：项目总结 异基因造血干细胞移植（allo-HCT）的一个主要并发症是急性移植物抗宿主 疾病（aGvHD）。aGvHD是一种免疫介导的疾病，其在很大程度上有助于移植- 相关的发病率和死亡率。aGvHD的总体发病率在35%至50%之间，这取决于 供体-受体组织相容性差异，并携带约50%的死亡率。T调节细胞 （Tlc），一种促进和维持免疫耐受性的表型多样性淋巴细胞亚群， 作为aGvHD的细胞疗法已被广泛研究。临床前试验表明， 存在产生IL-10的1型调节性T细胞（Tr 1）或叉头盒P3（FOXP 3）调节性T细胞 （F0 XP 3 + TcB）与allo-HCT后aGvHD调节和增强的移植耐受性相关。T型 allo 10细胞是含有用宿主来源的致耐受性树突状细胞致敏的供体来源的Tr 1细胞的细胞产物 细胞（DC-10）以诱导免疫耐受。我们已获得FDA批准（IND# 17292），用于1期临床试验， 在接受错配相关或不相关治疗的儿童和年轻成人患者中， HCT用于血液系统恶性肿瘤，2例患者已成功治疗。的主要终点 1期试验是1）在我们的GMP设施中生产T-allo 10产品的可行性和2） T-allo 10产品的施用。目的1是完成本试验，包括免疫学评价 的T-allo 10产品和纵向评估的HCT患者的持续性的输注 使用TCR克隆型分析和免疫重建的Tr 1细胞。含有Tr 1的T-allo 10细胞提供 巨大的希望，但也含有T效应细胞的混合细胞产物，这可能会减少它们的免疫功能。 有效性尽管多克隆FOXP 3 + TGF 3在增强免疫后耐受性方面显示出一定的功效， 移植时，输注的细胞在体内存活有限，并且分离纯的FOXP 3 + T细胞是困难的。 因此，我们通过慢病毒载体（LV）基因开发了“第二代”Tr 1细胞和FOXP 3 + TcB 转移到CD 4 + T效应细胞中。LV介导的FOXP 3基因转移到CD 4 + T细胞中， “FOXP 3 + Treg样”（CD 4FOXP 3）细胞。类似地，可以通过LV-IL-10表达获得“Tr 1样”（CD 4 IL-10）细胞 在人CD 4 + T效应细胞中。在初步实验中，我们已经证明，第三方多克隆 在免疫缺陷小鼠的人源化模型中，CD 4 IL-10和CD 4FOXP 3 T细胞可以抑制GvHD。目标2 我们将比较自体多克隆和同种异体抗原特异性CD 4 IL-10和CD 4FOXP 3对免疫缺陷的预防作用。 aGvHD，它们对造血干细胞（HSC）植入、免疫重建、耐受和GvL的影响 临床前研究中的活性。这些研究将为开发未来的疗法提供临床前数据， 将改善接受allo-HCT的患者的结局。

Project 2: Project Summary A major complication of allogeneic hematopoietic stem cell transplantation (allo-HCT) is acute graft-versus-host disease (aGvHD). aGvHD is an immunologically mediated disease, that contributes substantially to transplant- related morbidity and mortality. The overall incidence of aGvHD is between 35% and 50% depending upon donor-recipient histocompatibility differences and carries approximately a 50% mortality rate. T regulatory cells (Tregs), a phenotypically diverse subset of lymphocytes that promote and maintain immunological tolerance, have been extensively researched as a cellular therapy for aGvHD. Preclinical trials have demonstrated that the presence of IL-10-producing T regulatory type 1 (Tr1) cells or the forkhead box P3 (FOXP3) T regulatory cells (FOXP3+Tregs) correlates with aGvHD modulation and enhanced transplantation tolerance after allo-HCT. T- allo10 cells are a cell product containing donor-derived Tr1 cells primed with host-derived tolerogenic dendritic cells (DC-10) to induce immune tolerance. We have FDA approval (IND# 17292 ) for a Phase 1 clinical trial of escalating doses of T-allo10 cells in pediatric and young adult patients receiving mismatched related or unrelated HCT for hematologic malignancies, and 2 patients have been successfully treated. The primary endpoints of the Phase 1 trial are 1) the feasibility of producing the T-allo10 product in our GMP facility and 2) the safety of the administration of the T-allo10 product. Aim 1 is the completion of this trial including the immunological evaluation of the T-allo10 product and the longitudinal assessment of the HCT patients for the persistence of the infused Tr1 cells using TCR clonotypic analyses and the immune reconstitution. Tr1 containing T-allo10 cells offer enormous promise, but are a mixed cell product also containing T effector cells, that potentially reduce their effectiveness. Although polyclonal FOXP3+Tregs have shown some efficacy in enhancing tolerance post- transplant, the infused cells have limited in vivo survival, and the isolation of pure FOXP3+Tregs is difficult. Therefore, we have developed “second-generation” Tr1 cells and FOXP3+Tregs by lentiviral-vector (LV) gene transfer into CD4+ T effector cells. LV-mediated gene transfer of FOXP3 into CD4+ T cells creates “FOXP3+Treg-like” (CD4FOXP3) cells. Similarly, “Tr1-like” (CD4IL-10) cells can be obtained via LV-IL-10 expression in human CD4+ T effector cells. In preliminary experiments, we have demonstrated that 3rd party polyclonal CD4IL-10 and CD4FOXP3 T cells can suppress GvHD in a humanized model of immune deficient mice. In Aims 2 and 3 we will compare autologous polyclonal and alloAg-specific CD4IL-10 and CD4FOXP3 for their prevention of aGvHD, their impact on hematopoietic stem cell (HSC) engraftment, immune reconstitution, tolerance and GvL activity in preclinical studies. These studies will provide preclinical data toward developing future therapies, that will improve the outcomes of patients receiving allo-HCT.