Discovery of Small-Molecule Autophagy Modulators in Mammalian Systems
CitationKuo, Szu-Yu. 2016. Discovery of Small-Molecule Autophagy Modulators in Mammalian Systems. Doctoral dissertation, Harvard University, Graduate School of Arts & Sciences.
AbstractAutophagy is an evolutionarily conserved catabolic process in eukaryotes, which involves the formation of double-membrane vesicles that enclose cellular components and fuse with lysosomes. Autophagy is critical to the maintenance of cellular homeostasis by degrading cytosolic materials and redistributing biological building blocks to essential metabolic pathways. Furthermore, autophagy selectively clears cytosolic components, including protein aggregates, damaged organelles and invasive pathogens, to alleviate cellular stresses caused by these components. The importance of autophagy has been underscored by relationships between compromised autophagy and the pathogenesis of various human diseases. These observations have motivated us to explore whether autophagy enhancement can introduce beneficial effects in the context of these diseases.
Here I describe our efforts to study effects of autophagy enhancement in cellular models of human diseases using small-molecule probes. We identified 998 probes from screening 59,541 small molecules prepared by stereoselective, diversity oriented chemical synthesis. We further prioritized five probes based on their activities, chemical structures and toxicities to mammalian cells, and tested them in cellular models of various human diseases. One probe, BRD5631, elicited beneficial effects, which supports our hypothesis that enhancing autophagy is a promising strategy to rescue disease phenotypes related to defective autophagy. We also discovered BRD1240, a potent probe that suppressed the maturation of autophagy by disrupting lysosomal functions. An unbiased approach using bioactivity profiling suggested that BRD1240 might share the same MoA of a known V-ATPase inhibitor, bafilomycin A1, which was confirmed in vitro.
In parallel, I explored the potential to use small-molecule probes to directly modulate a variant of autophagic core protein ATG16L1 (T300A), which is associated with increased risk for Crohn’s disease due to susceptibility to caspase-3 cleavage. I developed an assay for screening small molecules that protect ATG16L1 T300A from caspase-3 cleavage and identified small molecules that elicited desirable effects. I also present plans to further study their mechanisms and effects of these small molecules on Crohn’s disease phenotypes.
Together, these findings provide more understanding of autophagy enhancement in the context of diseases. Moreover, small-molecule probes we discover can serve as valuable tools for studying autophagy in the context of cellular homeostasis and disease.
Citable link to this pagehttp://nrs.harvard.edu/urn-3:HUL.InstRepos:33840641
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