Hyperactive CREB subpopulations increase during therapy in paediatric B lineage acute lymphoblastic leukaemia

Abstract

Persistence of residual disease in acute lymphoblastic leukaemia during the initial stages of chemotherapy is associated with inferior survival. To better understand clonal evolution and mechanisms of chemo-resistance, we used multi-parameter mass cytometry to functionally characterise pediatric B-ALL cells at disease presentation and those persisting during induction therapy at single cell resolution. Analysis of presentation ALL (n=42) showed the most abundant phosphosignals were pCREB, pH2AX and pHH3 and we identified JAKSTAT and RAS pathway activation in 5 from 6 patients with JAK or RAS genetic aberrations. The clonal composition of ALL was heterogeneous and dynamic during treatment but all viable cell clusters showed pCREB activation. Levels of pCREB in ALL cells were increased or maintained during therapy and high dimensional analysis revealed a subpopulation of ALL cells at presentation that were positive for pCREB/pHH3/pS6 which increased during treatment in some patients, implicating this signalling node in conferring a survival advantage to multi-agent induction therapy. The small molecule CREB inhibitor, 666-15, was shown to reduce CREB transcriptional activity and induce apoptosis in ALL PDX cells of varying cytogenetic subtypes in vitro, both in the presence and absence of stromal support. Together, these data suggest that the cAMP signalling pathway may provide an opportunity for MRD-directed therapy for many patients at high risk of relapse.