Efficacy studies using genetic mouse models demonstrate that autophagy is strikingly important in the survival and growth of RAS-driven cancers. Specifically, inhibition of the RAF→MEK→ERK signaling pathway downstream of KRAS was recently shown to result in tumors dependent on autophagy for survival in vivo. Sole inhibition of the MEK pathway leads to the activation of the LKB1→AMPK→ULK1/ATG13 pathway that drives cells' survival under nutrient stress. The UNC-51-like kinase-1 (ULK1) is a central upstream regulator of the autophagy pathway – in effect an on/off switch – and selective molecular probes that function as ULK1-specific inhibitors would be an ideal strategy to abrogate autophagy and improve RAS-driven tumors sensitivity to approved therapeutics. However, there are currently no FDA-approved modulators that target ULK kinases. Here we report the discovery and characterization of potent cell-active ULK1 inhibitors with superior DMPK properties based on a 7-azaindole scaffold. Specifically, using tools of structure-based drug design, we have developed a series of inhibitors with excellent biochemical activity against ULK1 (IC₅₀< 100 nM). The target engagement of these analogs was confirmed using ULK1 NanoBRET intracellular kinase assay. Finally, the combinations of our compounds with known KRAS→RAF→MEK→ERK pathway inhibitors showed promising synergetic effects in vitro using PATC66 (KRAS^G12D) and H2030 (KRAS^G12C) cell lines. Importantly, no cellular toxicity was observed in cell lines not driven by RAS. This study reveals a new chemotype for ULK1 inhibition and proposes the therapeutic potential of where blocking autophagy in combination with MEK/ERK therapies will augment the efficacy of current chemotherapies across a broad spectrum of RAS-driven malignances.

Sean Chin Chan

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