Transcriptional remodeling shapes therapeutic vulnerability to necroptosis in acute lymphoblastic leukemia.

Insufficient eradication of cancer cells and survival of drug tolerant clones are major relapse driving forces. Underlying molecular mechanisms comprise activated pro-survival and anti-apoptotic signaling leading to insufficient apoptosis and drug resistance. The identification of programmed cell death pathways alternative to apoptosis opens up for possibilities to antagonize apoptosis escape routes. We have earlier shown that acute lymphoblastic leukemia (ALL) harbours a distinct propensity to undergo cell death by RIPK1-dependent necroptosis, activated by small molecule second mitochondria-derived activators of caspase (SMAC) mimetics. Despite demonstrated safety and tolerability of SMAC mimetics in clinical trials, their efficacy as single agent appears still limited, highlighting the need for combinatorial treatments. Here, we investigate so far unexplored regulatory mechanisms of necroptosis and identify targets for interference to augment the necroptotic anti-leukemia response. Ex vivo drug response profiling in a model of the bone marrow microenvironment reveals powerful synergy of necroptosis induction with histone deacetylase inhibition. Subsequent transcriptome analysis and functional in vivo CRISPR screening identify gene regulatory circuitries through the master transcription regulators SP1, p300 and HDAC2 to drive necroptosis. While deletion of SP1 or p300 confers resistance to necroptosis, loss of HDAC2 sensitizes to RIPK1-dependent cell death by SMAC mimetics. Consequently, our data inform strong in vivo anti-leukemic activity of combinatorial necroptosis induction and HDAC inhibition in patient-derived human leukemia models. Thus, transcriptional dependency of necroptosis activation is a key regulatory mechanism that identifies novel targets for interference, pointing out a strategy to exploit alternative non-apoptotic cell death pathways to eradicate resistant disease.