CD44/CD49d介导骨髓微环境庇护和MIRL抵抗穿孔素/颗粒酶杀伤途径在CAR-T细胞治疗后急性淋巴细胞白血病CD19阳性复发中的作用和机制

难治复发急性淋巴细胞白血病(ALL)预后差、生存期短。靶向CD19的CAR-T(CART19)疗法是最具应用前景的靶向治疗，对CD19阳性ALL诱导缓解率约90%，但复发是治疗失败的主要原因。前期利用单细胞转录组测序首次发现治疗前高表达CD44/CD49d的亚克隆及治疗后CART19压力选择下的克隆演变是CD19阳性复发的两大原因。本项目首次围绕CART19治疗前后白血病亚克隆、骨髓微环境、克隆演变及穿孔素/颗粒酶途径等关键因素，拟应用单细胞、人源化小鼠、活体成像等手段揭示高表达CD44/CD49d亚克隆定植于骨髓龛，通过龛物理屏障、异常PI3K/AKT激活和骨髓基质细胞负向调控等逃逸CART19杀伤成为缓解期微小残留病并在CART19压力选择下上调MIRL表达以抵抗CART19的穿孔素/颗粒酶途径致复发的可能机制，为阐明CD19阳性复发的分子机理提供实验依据，为发现新的干预靶点奠定基础。

Patients with refractory/relapsed acute lymphoblastic leukemia (R/R ALL) are associated with very poor prognosis. Chimeric antigen receptor modified T cells targeting CD19 have potent anti-leukemia activities with a very high complete remission (CR) rate in R/R ALL (around 90%). Relapse, however, still remains to be the major problem. In our previous study for the first time we found that the sub-clone with high expression level of CD44/CD49d before CART19 therapy and clonal evolution after CART19 therapy were the 2 main causes for relapse by using single cell transcriptome sequencing. Based on these, our hypothesis is: the leukemia subclone with high expression of CD44/CD49d resides in the bone marrow niche, and escapes from CART19 cytotoxicity because of niche-protected physical barrier, hypoxia-activated PI3K/AKT pathway, and negative immune regulation from bone marrow stromal cells. Then the sub-clone will be the minimal residual leukemia cells during complete remission. Under the selection pressure of CART19 therapy, these minimal residual leukemia cells up-regulate MIRL expression which has shown to resist peforin/granzyme-mediated cell lysis. Our study tries to elucidate the above mechanisms of CD19 positive relapse after CART19 therapy which will provide expreriment evidences for CD19 positive relapse and lay the foundation for finding novel therapeutic targets.