SFK-inhibitor enhancement of ATRA-mediated differentiation of APL

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PROJECT SUMMARY/ABSTRACT Retinoic acid signaling plays a key role in normal myeloid bone marrow cell development. Aberrant signaling leads to myeloid maturation arrest and continued cell proliferation. This is exemplified in Acute Promyelocytic Leukemia (APL), in which a mutated fusion protein of the receptor for retinoic acid (RAR) inhibits retinoic acid- dependent transcription. In APL, pharmacologic doses of all-trans retinoic acid (ATRA) overcome the inhibition of RAR-dependent transcription, leading to terminal differentiation of the blasts. The introduction of ATRA has transformed the treatment of APL, yet up to 30% of elderly and “high-risk” patients ultimately relapse and die of this disease. Current combination drug treatments are toxic. We propose a novel strategy to enhance the effectiveness of ATRA in APL. We have shown that Src family kinases (SFKs) are constitutively activated in APL and negatively regulate gene expression mediated by RAR. This suggests a model in which activated SFKs silence the endogenous RAR to aid in transcriptional repression of RAR-target genes. We predict that SFK inhibitors will enhance the activity of ATRA to activate RAR and increase differentiation in APL. Indeed, we found that inhibition of SFKs using the pan-SFK inhibitors PP1 and PP2 resulted in enhancement of ATRA- induced differentiation. Dasatinib, an FDA-approved inhibitor of SFKs, acted in a similar fashion to the PP1 and PP2 inhibitors, and dramatically enhanced ATRA-induced myeloid differentiation. Based on these findings, we hypothesize that SFK-inhibitors will enhance the activity of ATRA in therapies for APL. To test this hypothesis, we will first validate our observations in a panel of cryopreserved banked cells from patients with both standard and high-risk APL. We will test the combination of dasatinib and ATRA in an APL mouse model, assessing time to develop APL, time to relapse, and degree of decrease in tumor burden. We will also test the effects of ATRA and dasatinib in an ATRA-resistant mouse model. ATRA alone does not eradicate the APL leukemia initiating cell (LIC) that is responsible for relapse: we will assess the effects of the combination of ATRA plus SFK-inhibitors on the LIC. We propose that the combination of ATRA plus dasatinib will prove to be more efficacious and result in longer-term remissions. We will also determine the mechanism by which SFKs modulate RAR activity, which will identify other targetable pathways. These studies aim to develop a novel combination of drug classes to transform treatment of APL with less toxicity than current regimens. Both dasatinb and ATRA are FDA-approved, orally-administered agents, properties that will allow ready translation of our findings into the clinic.