Discovery Of A New PIKfyve Inhibitor

Phosphotidylinositol kinses and phosphatases phosphorylate ordephosphorylate phosphatidylinositol lipids,which are crucial for most eukaryotes and have diverse cellularfunctions.PI(3,5)P2 serves as a signal regulator in cellular homeostasis and adaptation,inhibition of PI(3,5)P2will lead to aberrant endosomal/lysosomal functions.PIKfyve is the sole lipid kinase that synthesizes PI(3,5)P2 from phosphatidylinositol-3-phosphate(PI3P),and can regulate exosomal release,endosomal pathways,and tumor cell invasion and migration.Several kinds of PIKfyve inhibitors have been reported to show inhibition effect in different cell lines,such as YM-201636,MF4,apilimoid and APY0201.However,there are some unavoidable defections among these inhibitors,including bad selectivityand efficacy.It’s urgent to find new inhibitors with high effiency and specificity.In this thesis,we have identified a small molecule inhibitor of PIKfyve,named HW-05-131-01,byphenotypic screening and subsequent KINOMEscan profiling.We found that HW-05-131-01 inhibited the function of PIKfyve in a time-and dose-dependent manner and led to endocellular vacuolization in breast cancer cell MDA-MB-231.Moreover,lack of nutrition affectsvacuolization upon HW-05-131-01 treatment,replenish of L-Glutamine and L-Threonine maintainsvacuolization,while other condition,such as glucose-free or hypoxemia has no effect.Long-time treatment of HW-05-131-01 can decrease cell survival rate.Finally,we found that HW-05-131-01 cause disfunction in endosomal trafficking pathwayand block the lysosomal degradation.In sunmmary,our results revealed that our new small molecular inhibitor of PIKfyve,HW-05-131-01,would lead to cell death and disrupt endosomal trafficking pathway.These data suggest that HW-05-131-01 could serve as a tool to pharmacologically investigate the biological function of PIKfyve.