LncRNA-ST3763/STAT3在急性髓系白血病发生发展中的作用机制研究

Leukemia stem cells (LSC) are the origins of relapse occurring in leukemia patients after remission, and effective removal of LSC is a big challenge in leukemia therapy. Most studies supported that LSC might be originated from dysregulation of the key regulation genes during the differentiation of hemopoietic stem/progenitor cells and STAT3 hyperactivation is the key factor involving in the transition of stem/progenitor cells to LSC. Although the role of STAT3 hyperactivation in the pathogenesis of acute myeloid leukemia (AML) has been widely studied, the exact mechanism by how STAT3 hyperactivation is regulated on multiple levels including transcriptional, post-transcriptional and epigenetic remains unclear. Considering the role of long chain non-coding RNA in the regulation of gene expression from several aspects, through gene chip analysis, we found that the long noncoding RNA-ST3763 directly and positively regulated the expression of STAT3 in the AML primary cell and cell line. Meanwhile, higher expression of ST3763 was observed in AML cells. Moreover, competitively inhibitiing the expression of ST3763 decreased the expression of STAT3, inhibited the proliferation of AML cell lines and promoted cell apoptosis. In this project, we propose to elucidate the multi-interface regulation mechanism of ST3763 on the expression and hyperactivation of STAT3 from epigenetic, transcriptional and post-transcriptional level using in vitro and in vivo model, aiming to explicit the role of ST3763 in the maintenance of LSC self-renewal as well as in the pathogenesis of AML. If finished on schedule, this study may enhance our understanding on the pathogenesis of AML and characteristics of LSC.

白血病干细胞（LSC）是白血病患者缓解后复发的根源，有效清除LSC是白血病治疗的难点。多数研究支持LSC起源于造血干/祖细胞关键调控基因的失调，而STAT3过激活是干/祖细胞向LSC转化的关键因素之一，虽然其在AML发生中的机制研究已较多，但STAT3本身在转录、转录后和表观遗传等水平被直接调控并过激活的机制尚不清楚。结合长链非编码RNA（LncRNA）可多层次调控基因表达的新视角，通过基因芯片，我们发现LncRNA-ST3763在AML原代细胞和细胞株中直接参与STAT3的正向表达调控，且ST3763高表达；若竞争抑制其表达，则STAT3表达下调，抑制AML细胞增殖，促进凋亡。为此，本课题拟采用离体和在体模型，从表观遗传学、转录和转录后水平探索ST3763多层次调控STAT3表达及过激活机制，明确其在LSC干性维持和AML发生发展中的作用。如期完成，有望加深对AML发病和和LSC的认识。

白血病干细胞（LSC）是患者缓解后复发的根源。STAT3过激活是LSC干性维持和AML发生的关键因素，但其过激活的机制尚不清楚。本研究结合LncRNA研究的新进展，探索了LncRNA调控STAT3过激活的机制。结果如下：①在AML细胞株和原代细胞中ST3763高表达，STAT3高表达，过表达ST3763导致STAT3高表达和过激活参与AML发生发展。②过表达ST3763促进细胞的增殖、抑制其凋亡，使其发生G1-S期的转变，提高对化疗药物的耐药性和克隆形成率促进LSC的干性维持；而竞争抑制其表达，则结果相反。③过表达ST3763促进AML原代细胞增殖、抑制对化疗药物的敏感性、细胞凋亡和分化、使细胞从G1向S期转化；而竞争抑制其表达，则结果相反，提示ST3763调控STAT3的活性调控AML细胞的生物学行为。④Chip分析提示ST3763在表观遗传学水平调控STAT3的甲基化；启动子区域敲除实验提示其招募结合蛋白调控STAT3的转录，不影响转录后调控。⑤ST3763参与LSC干性维持的调控，序贯移植实验提示：过表达ST3763提高LSC的含量、抑制其分化、使其处于静息状态；同时竞争实验显示：过表达ST3763提高LSC的数目、比例和干性维持能力；而竞争抑制其表达，则结果相反。⑥ST3763促进AML的发生发展，序贯移植实验提示：过表达ST3763缩短小鼠生存期、降低生存率、促进AML细胞的骨髓浸润；而竞争抑制其表达延长小鼠生存期、提高其生存率、减少AML细胞的骨髓浸润；STAT3敲除后，过表达ST3763不影响小鼠的生存率和生存期。本研究基本阐明了ST3763在多层次直接调控STAT3表达和过激活在LSC干性维持和AML发生发展中的作用机制，为AML的治疗提供了新的潜在靶点。意义：基本阐明了ST3763通过多层次调控STAT3表达和活性在LSC干性维持和AML发生发展中的作用机制，为其治疗提供了新的潜在靶点。