PDGFRB重排协同IKZF1突变在Ph-like ALL发生及耐药中作用机制的研究

Ph-like (BCR-ABL1-like) ALL is one of the newly discovered subtypes of acute lymphoblastic leukemia (ALL), with the characteristics of BCR-ABL1 negative, but exhibiting a gene expression profile similar to that of BCR-ABL1-positive ALL. Recent researches found that tyrosine kinase inhibitor (TKI) can also have effect on Ph-like ALL with specific kinase-activating alterations, including the rearrangements involving PDGFRB. To explore the pathogenesis and mechanism of TKI resistance in a 5-year-old Ph-like patient who experienced chemo-resistance and twice TKI resistance, we presented whole genome sequencing and mRNA sequencing of the samples at 4 time points: the diagnosis, the chemoUnCR (chemo-resistance), the first relapse after Imatinib (the first-generation TKI) and the second relapse after dasatinib (the second-generation TKI). We found a novel partner of PDGFRB fusion (AGGF1-PDGFRB) and IKZF1 deletions during the whole course of diseases; moreover, two somatic point mutations in IKZF1 and PDGFRB was found respectively only in the two samples at relapse (TKI resistance). This study was aimed to elucidate synergistic effects of PDGFRB translocation and IKZF1 mutation in the detailed pathogenesis and mechanisms of drug resistance in Ph-like ALL by constructing AGGF1-PDGFRB transducted Ba/F3 cell lines and mouse models wt/wo IKZF1 mutations, with a view to find potential targeting therapeutic ways in preclinical practice.

Ph-like ALL是近年来新发现的ALL亚型，其特点为具有与BCR-ABL ALL相似的基因表达谱。研究表明激酶异常激活是Ph-like ALL发生的原因之一，酪氨酸激酶抑制剂（Tyrosine kinase inhibitor，TKI）可用于治疗Ph-like ALL。通过对Ph-like ALL患儿初诊、常规化疗不缓解、TKI耐药等不同时间点的白血病细胞进行转录组测序和全基因组测序，发现在所有时间点均存在融合基因AGGF1-PDGFRB和IKZF1缺失突变，且在TKI处理后出现了IKZF1和PDGFRB的点突变。因此，本项目拟在细胞系及小鼠模型中探索融合基因AGGF1-PDGFRB协同IKZF缺失突变诱发Ph-like ALL的机制，阐明PDGFRB与IKZF1点突变与TKI耐药的关系，揭示B-ALL发生以及TKI耐药的机制，为临床治疗Ph-like ALL患儿提供靶向治疗新方案。

Ph-like ALL是近年来新发现的ALL亚型，其特点为具有与BCR-ABL ALL相似的基因表达谱。研究表明激酶异常激活是Ph-like ALL发生的原因之一，酪氨酸激酶抑制剂（Tyrosine kinase inhibitor，TKI）可用于治疗Ph-like ALL。通过对Ph-like ALL患儿初诊、常规化疗不缓解、TKI耐药等不同时间点的白血病细胞进行转录组测序和全基因组测序，发现在所有时间点均存在融合基因AGGF1-PDGFRB和IKZF1缺失突变，且在TKI处理后出现了IKZF1和PDGFRB的点突变，其中PDGFRB C843G点突变对ABL TKIs的所有系列均耐药，包括伊马替尼、达沙替尼、尼罗替尼和波纳替尼，但是，PDGFRB突变型白血病细胞对多靶点激酶抑制剂CHZ868高度敏感，提示对ABL-TKIs耐药的一些患者有潜在的治疗选择。.随后，我们构建了携带IKZF1缺失突变（IK-6亚型）的Ba/F3细胞，同时将发生率较高的几种Ph-like相关遗传学异常（EBF1-PDGFRB、NRAS G12D、KRAS G12V、P2RY8-CRLF2和JAK2 R683G）在IK-6 Ba/F3细胞中共表达，结果显示单纯IK-6基因的缺失改变不能直接引起Ba/F3细胞发生恶变，但在伴随其他遗传学异常或经受外界应激刺激后，往往能加快其恶变的速度。尤其在发生率最高的两种Ph like-ALL相关的遗传学异常-P2RY8-CRLF2和JAK2 R683G中，当协同出现后其加速恶变的效果尤其显著；电离辐射IKZF1缺失突变的Ba/F3细胞使之恶变，进行全基因组RNA-seq分析，结果表明IK-6缺失突变引起的IKAROS蛋白功能丧失造成了多种基因和信号通路的异常激活，这些途径是导致ALL发生和发展的潜在原因；我们采用多种ALL常用化疗药及300余种小分子靶向药的体外药敏试验来寻找针对IK-6缺失突变肿瘤细胞的有效治疗药物，结果显示携带IK-6缺失突变后，恶变的Ba/F3细胞对化疗药物和几乎所有的靶向药物敏感性均会降低，该研究为探索针对IK-6缺失ALL患者的敏感药物，提高其预后水平提供理论支持，为临床治疗Ph-like ALL患儿提供靶向治疗新方案。