The cellular cytotoxicity of APY0201, a PIKfyve inhibitor, against multiple myeloma was initially identified in an unbiased in vitro chemical library screen. The activity of APY0201 was confirmed in all 25 cell lines tested and in 40% of 100 ex vivo patient-derived primary samples, with increased activity in primary samples harboring trisomies and lacking t(11;14). The broad anti-multiple myeloma activity of PIKfyve inhibitors was further demonstrated in confirmatory screens and showed the superior potency of APY0201 when compared to the PIKfyve inhibitors YM201636 and apilimod, with a mid-point half maximal effective concentration (EC50) at nanomolar concentrations in, respectively, 65%, 40%, and 5% of the tested cell lines. Upregulation of genes in the lysosomal pathway and increased cellular vacuolization were observed in vitro following APY0201 treatment, although these cellular effects did not correlate well with responsiveness. We confirm that PIKfyve inhibition is associated with activation of the transcription factor EB, a master regulator of lysosomal biogenesis and autophagy. Furthermore, we established an assay measuring autophagy as a predictive marker of APY0201 sensitivity. Overall, these findings indicate promising activity of PIKfyve inhibitors secondary to disruption of autophagy in multiple myeloma and suggest a strategy to enrich for likely responders.
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