Date of Award
Doctor of Philosophy (PhD)
Cancer and Developmental Biology
Gerard P. Zambetti, Ph.D. (for Richard T. Williams, M.D., Ph.D.)
Suzanne J. Baker, Ph.D. R. Kiplin Guy, Ph.D. Sima Jeha, Ph.D. Rajendra S. Raghow, Ph.D.
Oncogenic signaling by the Philadelphia chromosome-encoded BCR-ABL fusion kinase initiates and drives both Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL) and chronic myelogenous leukemia (CML). Food and Drug Administration (FDA)- approved BCR-ABL-specific kinase inhibitors (BCR-ABL–KIs) imatinib, dasatinib and nilotinib induce prolonged remissions in CML but poor leukemia-reduction and relapse-control in Ph+ ALL. The relative primary BCR-ABL–KI-resistance in Ph+ ALL patients carrying predominantly BCR-ABLWT disease cannot be attributed to drug-resistant BCR-ABL mutations (BCR-ABLMUTANTS), and remains poorly understood.
We established a cell-based platform to evaluate the modulation of anti-Ph+ ALL activity of drugs by both tumor-extrinsic cytokines normally present in the leukemia microenvironment and tumor-intrinsic vector-mediated alteration of candidate genes. We identified that BCR-ABLWT-driven Ph+ ALL cells are rendered significantly resistant against all FDAapproved BCR-ABL–KIs by 1) Several host-cytokines, but most dominantly by Interleukin7 (IL7), and 2) Cell-intrinsic functional-loss of IKAROS. Utilizing IL7-deficient recipient mice, we demonstrated that physiological levels of IL7 significantly attenuate the survival benefit derived from dasatinib monotherapy against Ph+ ALL. Follow-up mechanistic studies using cell signaling and gene expression comparisons indicated that IL7 imparts chemo-refractoriness by initiating IL7-pSTAT5-c-MYC signaling. Interestingly, IKAROS haploinsufficiency, which has been previously associated with poor clinical prognosis in patients with Ph+ ALL, was recently demonstrated to directly de-repress c-MYC expression. This suggested that both cell-extrinsic IL7 and cell-intrinsic IKAROS loss converge on c-MYC, which may act as node of leukemia BCR-ABL–KI-refractoriness. We confirmed that vector-mediated modifications imitating IL7 induction of STAT5 activity, functional-loss of IKAROS, and c-MYC over-expression, all selectively enrich Ph+ ALL cells during prolonged exposures to imatinib and dasatinib, thus revealing new chemo-refractory phenotypes.
Contemporary medicine advocates co-treatment with agents of complementary mechanism of drug-action to tackle drug resistance. To identify combination agents that could overcome the dasatinib-resistant phenotypes of Ph+ ALL, we screened a library of 3200 agents including known anti-infective and chemotherapy drugs. We discovered that a well known antimalarial drug dihydroartemisinin (DHA) killed host-IL7-protected BCR-ABLWT, c-MYC-overexpressing BCR-ABLWT and BCR-ABLMUTANT Ph+ ALL cells in vitro. In vivo, DHA displayed weak activity as a single agent but its addition synergistically augmented the leukemia reduction by dasatinib, relative to either of the two drugs alone. Remarkably, DHA and dasatinib combination regimen eliminated host-protected dasatinib-refractory persistent leukemia and improved long-term survival from 0 to >90% in a murine model that faithfully captures the BCR-ABL–KI drug-refractoriness of human Ph+ ALL.
This study: 1) Uncovers novel mechanisms of clinical drug-resistance against BCR-ABL–KIs, 2) Identifies increased levels of IL7, pSTAT5 and c-MYC protein, and IKAROS haploinsufficiency as potential biomarkers of BCR-ABL-targeted drug-resistance, 3) Strongly supports clinical exploration of the BCR-ABL–KI and DHA combinations for treating patients with Ph+ ALL, and 4) Establishes a paradigm for investigating frequently overlooked host-tumor-drug interactions.
Singh, Harpreet , "Discovery of Dihydroartemisinin and Dasatinib Drug Combination to Cure Pooroutcome BCR-ABL+ Acute Lymphoblastic Leukemia" (2012). Theses and Dissertations (ETD). Paper 249. http://dx.doi.org/10.21007/etd.cghs.2012.0292.