Modeling chromosome 7 loss in Myelodysplasia-iPSCs

描述（由申请人提供）：骨髓增生异常综合征（MDS）是一种克隆性血液学疾病，其特征是造血功能低下，并倾向于进展为骨髓（BM）衰竭或急性白血病。尽管它们的发病率相对较高，但对其发病机制知之甚少，其治疗主要是姑息性的。这主要是由于缺乏良好的动物模型和原代MDS细胞的体外培养的挑战。7号染色体的全部或部分缺失[del(7/7q)]是MDS中一种复发性的细胞遗传学异常，与不良预后相关，强烈提示一个或多个关键基因存在于染色体7q中，而这些基因目前尚不清楚。随着人类多能干细胞（hPSC）研究的最新突破——直接重编程和新的基因工程技术——在等基因环境下通过精确破坏基因组元素进入同源基因组和细胞环境的反向遗传学——迄今为止对人类基因组来说是不可想象的——现在是一个现实的前景。我们的目标是利用我们和其他人开发的尖端重编程和基因工程技术，建立一种新的基于hpsc的模型来研究骨髓异常增生的细胞，分子和遗传发病机制。在初步研究中，我们从患者骨髓细胞中获得了染色体7q缺失的MDS-iPSC细胞系，发现它们概括了潜在的疾病相关表型：细胞增殖和造血分化受损。在我们提出的研究中，我们计划通过重编程和染色体工程获得额外的del(7q)-以及等基因核型正常的iPSCs，并表征它们在未分化状态和造血分化后的表型。为了确定在MDS发病机制中起作用的基因，我们将筛选位于染色体7q上的候选基因，以挽救我们的del(7q)- hPSCs的增殖。这些研究使用强大的新技术，将为骨髓发育不良的研究提供新的有价值的资源，对受影响的细胞过程和分子途径产生见解，并有可能确定MDS、骨髓衰竭和白血病前期发病机制的关键基因。！

DESCRIPTION (provided by applicant): Myelodysplastic syndromes (MDS) are clonal hematologic disorders characterized by ineffective hematopoiesis and a propensity for progression to bone marrow (BM) failure or acute leukemia. Despite their relatively high incidence, very little is known about their pathogenesis and their treatment remains mainly palliative. This is largely due to the unavailability of good animal models and the challenges of the ex vivo culture of primary MDS cells. Loss of the entire or part of chromosome 7 [del(7/7q)] is a recurrent cytogenetic abnormality in MDS, associated with unfavorable prognosis, strongly suggesting that one or more critical genes - that so far remain elusive - reside in chromosome 7q. With recent breakthroughs in human pluripotent stem cell (hPSC) research - direct reprogramming and new genetic engineering technologies - reverse genetics in an isogenic setting by precise disruption of genomic elements into their cognate genomic and cellular context - hitherto unthinkable for the human genome - are now a realistic prospect. Our goal is to harness cutting-edge reprogramming and genetic engineering technologies that we and others have developed to establish a novel hPSC-based model to study the cellular, molecular and genetic pathogenesis of myelodysplasia. In preliminary studies, we have derived MDS-iPSC lines with chromosome 7q deletions from patient BM cells and found that they recapitulate potential disease- associated phenotypes: impaired cell proliferation and hematopoietic differentiation. In the proposed study we plan to derive additional del(7q)- as well as isogenic karyotypically normal iPSCs, through reprogramming and chromosome engineering, and characterize their phenotype in the undifferentiated state and following hematopoietic differentiation. To identify genes with a role in MDS pathogenesis, we will perform a screen of candidate genes residing in chromosome 7q for rescue of proliferation in our del(7q)- hPSCs. These studies, using powerful new technologies, will provide a novel valuable resource for the study of myelodysplasia, generate insights into the cellular processes and molecular pathways affected and potentially identify critical genes in the pathogenesis of MDS, bone marrow failure and preleukemia, in general. !