PIP5K1C Phosphoinositide Kinase Deficiency Distinguishes PIKFYVE-Dependent Cancer Cells from Non-Malignant Cells

Although PIKFYVE phosphoinositide kinase inhibitors can selectively eliminate PIKFYVE-dependent human cancer cells <i>in vitro</i> and <i>in vivo</i>, the basis for this selectivity has remained elusive. Here we show that the sensitivity of cells to the PIKFYVE inhibitor WX8 is not linked to PIKFYVE expression, macroautophagic/autophagic flux, the BRAF^V600E mutation, or ambiguous inhibitor specificity. PIKYVE dependence results from a deficiency in the PIP5K1C phosphoinositide kinase, an enzyme required for conversion of phosphatidylinositol-4-phosphate (PtdIns4P) into phosphatidylinositol-4,5-bisphosphate (PtdIns[4,5]P₂/PIP2), a phosphoinositide associated with lysosome homeostasis, endosome trafficking, and autophagy. PtdIns(4,5)P₂ is produced via two independent pathways. One requires PIP5K1C; the other requires PIKFYVE and PIP4K2C to convert PtdIns3P into PtdIns(4,5)P₂. In PIKFYVE-dependent cells, low concentrations of WX8 specifically inhibit PIKFYVE <i>in situ</i>, thereby increasing the level of its substrate PtdIns3P while suppressing PtdIns(4,5)P₂ synthesis and inhibiting lysosome function and cell proliferation. At higher concentrations, WX8 inhibits both PIKFYVE and PIP4K2C <i>in situ</i>, which amplifies these effects to further disrupt autophagy and induce cell death. WX8 did not alter PtdIns4P levels. Consequently, inhibition of PIP5K1C in WX8-resistant cells transformed them into sensitive cells, and overexpression of PIP5K1C in WX8-sensitive cells increased their resistance to WX8. This discovery suggests that PIKFYVE-dependent cancers could be identified clinically by low levels of PIP5K1C and treated with PIKFYVE inhibitors.