构建骨髓增生异常综合征/继发性白血病小鼠动物模型及其造血干细胞克隆演变机制研究

Myelodysplastic syndromes (MDS) are heterogeneous disorders in which the hematopoietic stem cells (HSCs) in the bone marrow are defective, resulting in insufficient normal blood cells and high risk progressing to secondary acute myeloid leukemia (sAML). More and more gene mutations are identified in MDS due to the Next Generation Sequencing (NGS). Those mutations involved in signaling pathways proteins (e.g. CBL, FLT3, JAK2, RAS), transcription factors (e.g. CEBPA, ETV6, RUNX1), epigenetic regulators (e.g. ASXL1, DNMT3A, EZH2, IDH1, IDH2,SUZ12, TET2, UTX), tumor suppressors (e.g. TP53), and components of the spliceosome (e.g. SF3B1, SRSF2). We built a robust MDS mouse model with MLL-PTD and RUNX1 mutations, which accurately mimic human MDS with pancytopenia, tri-lineage dysplasia and excess blasts in the bone marrow. However, it didn't develop AML. Little is known about the molecular mechanisms underlying MDS-associated clonal expansion, ineffective hematopoiesis, and sAML transformation, which significantly restricts the development of new therapies and pre-clinical models. The goals of this project are to build robust and faithful mouse MDS/sAML models combined with RAS mutations (NRAS G12D, KRAS G12D and Nf-/-)and to identify key targets that are critical for clonal evolution and pathogenesis of MDS/AML, and therapeutic intervention.

骨髓增生异常综合征是起源于造血干细胞恶性克隆性疾患，以骨髓无效造血，外周血细胞减少以及高危向急性髓系白血病进展为主要特点。随着二代测序技术的进步，发现了更多累及MDS的基因突变，主要涉及转录因子、表观遗传学调控因子、剪切体复合物、信号转导途径和抑癌基因。我们前期以高危MDS患者高频突变RUNX1和MLL-PTD为基础，构建了MDS小鼠模型。该模型具有全血细胞减少，形态发育异常，原始细胞轻度增多等表现，但是并没有发生AML转化。提示表观遗传学联合转录因子异常不足以促进MDS克隆演变，尚需要获得其他突变使MDS克隆具有增殖能力。因此我们提出本课题，在MDS小鼠模型基础上导入RAS信号途径基因突变（NRAS,KRAS和Nf-/-）,构建MDS/sAML模型，通过RNAseq、基因表达谱分析，ChIPseq等分析，明确MDS克隆演变的分子机制及途径，为MDS/sAML靶向治疗提供重要依据。

骨髓增生异常综合征（MDS）起源于造血干细胞，是一种最常见的血液系统恶性肿瘤性疾患。该病主要发生于老年人群，随着我国人口老龄化的加剧，患病人数急剧增多。国际流行病学数据表明，80岁以上人群中，MDS的发病率可达1/5000，是严重危害人民健康和生活质量的恶性疾病。已知MDS中存在40-60种MDS相关的基因突变，涉及到染色质修饰酶、RNA剪切、粘连蛋白复合体、代谢酶等基因，与造血干细胞自我更新、生存、分化相关。由这些基因突变导致的分子学改变差异大，如何致使MDS发生，目前机制尚不清楚我们通过MDS患者CD34+细胞基因表达谱分析发现，患者低氧诱导因子1A(HIF1A)信号通路存在激活，并且在患者骨髓病理中证实HIF1A蛋白表达明显增多。。我们分别检测了MDS相关基因突变小鼠（包括Dnmt3aΔ/Δ 小鼠, Tet2Δ/Δ 小鼠, Runx1 Δ/Δ 小鼠、Asxl1Δ/Δ 小鼠、以及 MllPTD/WT mice）骨髓c-kit+细胞HIF1A表达，结果发现HIF1A表达明显增多，提示提示HIF1α通路的激活是MDS的共同表现。通过构建血液系统可诱导表达HIF1A的TPM小鼠，这些小鼠出现典型MDS疾病表型，进一步证实HIF1A在MDS发生中的关键作用，而通过应用HIF1A抑制剂，可以明显改善MDS小鼠症状，延长生存期。以HIF1A为靶点有可能成为临床治疗MDS的新策略。