异基因靶向CD19嵌合抗原受体修饰的T细胞联合造血干细胞输注的免疫学演变及抗白血病机制研究

Relapsed and refractory B-cell acute lymphoblastic leukemia (r/r B-ALL) has poor prognosis. Although autologous chimeric antigen receptor-modified T cells (CART) improved remission rate, problems such as relapse and culture failure still exist. Whether non-gene-edited allogeneic CART (allo-CART) induce severe graft-versus-host disease (GVHD) or experience survival difficulty is uncertain. Previously we successfully used chemotherapy combined with mobilized HLA-mismatched donor stem cell infusion (microtransplantation) to treat elderly acute leukemia. Although abundant allogeneic T cells was infused, the risk of severe GVHD did not increase. Based on this, we utilized non-gene-edited allo-CART combining microtransplantation to treat r/r B-ALL. Remission rate was significantly improved, and long-term existence of CART and donor chimerism was detected. However, the immunological evolution pattern and anti-leukemic mechanism are unknown. We aim to utilize the humanized CD19+ leukemia mouse model, and co-infuse with donor-derived CART and hematopoietic stem cells, to elucidate immunological evolution pattern and impact on leukemic cells of allo-CART in the context of varied donor chimeric status. We also carry out clinical research to explore the influence of allo-CART survival and expansion, chimeric status and cytokine changes on minimal residual leukemia, patient survival and relapse. We believe our research will bring an alternative strategy for r/r B-ALL.

复发难治急性B淋巴细胞白血病（B-ALL）预后差，自体嵌合抗原受体修饰的T细胞（CART）仍存在复发、培养失败等问题，未经基因编辑的异基因CART是否诱发移植物抗宿主病（GVHD）以及体内存活仍不明确。化疗联合HLA不全相合造血干细胞输注（微移植）明显改善了老年白血病疗效，输注大量异基因T细胞并未增加严重GVHD；在此基础上我们利用异基因CART联合微移植治疗复发难治B-ALL，明显提高了缓解率，并检测到CART及供体不同比例嵌合长期存在，但其免疫学演变过程及抗白血病机制仍不明确。我们拟采用人源化CD19+白血病小鼠，同时输注供者来源CART及造血干细胞，以阐明异基因CART在不同比例供者嵌合状态下的免疫学演变规律及对白血病细胞的影响；另一方面，通过临床研究探讨异基因CART存活及扩增、嵌合状态持续时间、细胞因子变化对白血病残留、生存及复发的影响，为复发难治B-ALL开辟新的治疗手段。

复发难治急性B淋巴细胞白血病（acute B lymphoblastic leukemia, B-ALL）预后差，靶向CD19的自体嵌合抗原受体修饰的T（chimeric antigen receptor T, CART）细胞治疗显著改善了B-ALL患者预后，但仍存在复发等问题，异基因CART作为未来发展的趋势仍存在体内存活时间短及可能诱发移植物抗宿主病（graft-versus-host disease, GVHD）等问题，而为克服这些缺点而进行的额外基因编辑过程繁琐。我们既往应用化疗联合HLA不全相合造血干细胞输注（微移植）明显改善了老年白血病疗效，且未增加GVHD风险。基于以上前期研究，本项目探索了同一供者来源的G-CSF动员的单个核细胞联合CART细胞输注对CD19阳性B-ALL的疗效及相关机制。动物实验表明，同一供者来源高剂量单个核细胞输注增强了异基因CART细胞的体内扩增及对CD19阳性白血病细胞的杀伤能力。在长期存活的受鼠体内持续检测到供体嵌合成分存在，CD19阳性白血病细胞维持持续低水平，临床及组织病理均未出现GVHD表现，受鼠生存期明显延长。I期临床研究入组的9例复发难治ALL患者中获得了78%（7/9）的形态学缓解率，89%（8/9）的整体反应率，56%（5/9）的患者达到完全或高比例供体植入，未出现早期死亡，最长存活时间37个月。本项目初步证实了供者动员后高剂量单个核细胞输注对同一供者来源CART细胞体内扩增并发挥抗白血病效应的增强作用及安全性，为开展进一步临床研究及应用奠定基础。