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Targeted and Phenotypic Approaches to Discover Autophagy Modulators for Therapeutic Development

thesis
posted on 01.12.2021, 00:00 authored by Maryna Salkovski
Macroautophagy (hereafter referred to as autophagy) is a conserved, eukaryotic, catabolic pathway that maintains cellular homeostasis by engulfing cytosolic contents into double membrane vesicles termed autophagosomes, which fuse with lysosomes to form autolysosomes where the contents are degraded and recycled. To assess the role of autophagy in different diseases, including cancer and Niemann-Pick Type C (NPC) disease, autophagy modulators were identified in high throughput using both targeted and phenotypic approaches. A compound-multiplexed fluorescence polarization assay was developed to discover small molecules that inhibit the ATG5-ATG16L1 protein-protein interaction (PPI) in early autophagy. Inhibition of this PPI would specifically inhibit autophagosomes formation, selectively inhibit autophagic flux, and potentially prevent cancer cell survival through nutrient deprivation and inhibition of the cellular stress response. A small-molecule inhibitor was identified, and future work will focus on its optimization for solubility, selectivity, and potency. A high-content phenotypic screen was also performed to identify small molecules that modulate autophagy and clear unesterified cholesterol in I1061T NPC1 fibroblasts. NPC is an autosomal recessive, fatal, neurodegenerative disease without FDA approved therapy that results from mutations in cholesterol transport proteins (NPC1 or NPC2), which causes impaired intracellular lipid trafficking and manifests as progressive cognitive decline, seizures, and eventually death in early adulthood. Two novel autophagy modulators were discovered that elicit opposing effects on the autophagy pathway, but display promising activity in NPC-relevant assays. Current work is focused on elucidation of the mechanisms of action of these two molecules and their optimization for in vivo studies.

History

Advisor

Aldrich, Leslie N

Chair

Aldrich, Leslie N

Department

Chemistry

Degree Grantor

University of Illinois at Chicago

Degree Level

Doctoral

Degree name

PhD, Doctor of Philosophy

Committee Member

Cologna, Stephanie M Fung, Leslie W Hu, Ying S Moore, Terry W

Submitted date

December 2021

Thesis type

application/pdf

Language

en