Identifying Mechanisms of Glucocorticoid Resistance in Relapsed Pediatric T-ALL

While great strides have been made in the improvement of outcome for newly diagnosed pediatric acute lymphoblastic leukemia (ALL) patients, prognosis for relapsed leukemia patients remains poor. The synthetic glucocorticoid (GC) dexamethasone is part of the standard treatment for pediatric ALL and patient response to glucocorticoid treatment has proved to be a reliable prognostic indicator. Identifying the biological pathways responsible for glucocorticoid resistance may reveal novel therapeutic targets to prevent and treat relapsed ALL. Although genomic analyses of relapsed patients and matched diagnosis-relapse patient pairs have begun to define the genomic landscape of relapsed disease, discerning "driver from passenger" genetic lesions remains challenging. To identify glucocorticoid resistance genes in an unbiased, high-throughput manner, we conducted a genome wide, survival based, shRNA screen in dexamethasone sensitive murine T-ALL cells. Our preliminary data identify several hundred genes capable of mediating GC resistance, including several known GC resistance genes Nr3c1, Rcan1, Btg1 and Mllt10, thereby validating our experimental approach. Candidate genes identified in the screen including EP300 (p300), GATA3 and IKZF1 are known leukemia suppressors in pediatric ALL and the EP300 paralog CREBBP and IKAROS have been linked to GC resistance, indicating that suppressor genes involved in human leukemia and GC resistance are identified in our mouse screen. Consistently, we found the expression of several screen hits significantly decreased and/or mutated in relapse patient samples. Novel dexamethasone resistance genes identified in the screen interfere with GC-induced transcription (Stat3, Ikzf1), promote pluripotency (Esrrb, Sox2) or stimulate cAMP signaling (Adcy3, Gnas, Creb1). Silencing of these genes in multiple mouse T-ALL cell lines has no detectable effects on leukemic growth/survival in vitro, but confers resistance to dexamethasone treatment in vitro and in vivo. Moreover, we show that silencing of some candidate dexamethasone resistance genes accelerates leukemogenesis in vivo, demonstrating that leukemia suppressor genes were identified. Effect(s) of silencing or inhibiting these novel dexamethasone resistance genes/pathways in human T-ALL cell lines, primary patient samples and xenografts will be discussed. We predict that targeting these dexamethasone resistance pathways may re-sensitize relapse pediatric T-ALL cells to dexamethasone and/or contribute to more effective patient stratification to prevent relapse and induction failure.