The Wnt Pathway Inhibitor RXC004 Blocks Tumor Growth and Reverses Immune Evasion in Wnt Ligand–dependent Cancer Models

Wnt signaling is implicated in the etiology of gastrointestinal tract cancers. Targeting Wnt signaling is challenging due to on-target toxicity concerns and lack of druggable pathway components. We describe the discovery and characterization of RXC004, a potent and selective inhibitor of the membrane-bound o-acyl transferase Porcupine, essential for Wnt ligand secretion. Absorption, distribution, metabolism, and excretion and safety pharmacology studies were conducted with RXC004 in vitro, and pharmacokinetic exposure assessed in vivo. RXC004 effects on proliferation and tumor metabolism were explored in genetically defined colorectal and pancreatic cancer models in vitro and in vivo. RXC004 effects on immune evasion were assessed in B16F10 immune “cold” and CT26 immune “hot” murine syngeneic models, and in human cell cocultures. RXC004 showed a promising pharmacokinetic profile, inhibited Wnt ligand palmitoylation, secretion, and pathway activation, and demonstrated potent antiproliferative effects in Wnt ligand–dependent (RNF43-mutant or RSPO3-fusion) colorectal and pancreatic cell lines. Reduced tumor growth and increased cancer cell differentiation were observed in SNU-1411 (RSPO3-fusion), AsPC1 and HPAF-II (both RNF43-mutant) xenograft models, with a therapeutic window versus Wnt homeostatic functions. Additional effects of RXC004 on tumor cell metabolism were confirmed in vitro and in vivo by glucose uptake and (18)fluorodeoxyglucose-PET, respectively. RXC004 stimulated host tumor immunity; reducing resident myeloid-derived suppressor cells within B16F10 tumors and synergizing with anti-programmed cell death protein-1 (PD-1) to increase CD8(+)/regulatory T cell ratios within CT26 tumors. Moreover, RXC004 reversed the immunosuppressive effects of HPAF-II cells cocultured with human peripheral blood mononuclear cells, confirming the multiple anticancer mechanisms of this compound, which has progressed into phase II clinical trials. SIGNIFICANCE: Wnt pathway dysregulation drives many gastrointestinal cancers; however, there are no approved therapies that target the pathway. RXC004 has demonstrated the potential to block both tumor growth and tumor immune evasion in a genetically defined, clinically actionable subpopulation of Wnt ligand–dependent gastrointestinal cancers. The clinical utility of RXC004, and other Porcupine inhibitors, in such Wnt ligand–dependent cancers is currently being assessed in patient trials.