A growing body of evidence suggests that autophagy inhibition enhances the effectiveness of chemotherapy, especially in difficult-to-treat cancers. Existing autophagy inhibitors primarily target the lysosome and as a result are non-specific. More specific autophagy inhibitors have not yet been developed. The Atg8 family of proteins, including microtubule-associated protein 1A/1B light chain 3B protein, LC3B, and GABA<sub>A</sub> receptor associated protein, GABARAP, are adapter proteins that mediate key protein-protein interactions at several points in autophagy pathways. In this work, we used known peptide ligands as starting points to develop improved LC3B and GABARAP inhibitors. We developed new in vitro binding assays and used them to define structure-activity relationships. We quantified how peptide termini, individual charged residues, and hydrophobic interactions contributed to binding and selectivity for LC3B and GABARAP. Based on these data, we used artificial amino acids and diversity-oriented stapling to improve affinity, cytosolic penetration, and resistance to biological degradation while maintaining or improving LC3B or GABARAP affinity and selectivity. We found slight modifications to amino acids and staples that drastically alter LC3B binding, GABARAP binding, and relative selectivity between these Atg8 family members. These peptides include the highest-affinity LC3B-selective ligands reported to date and provide novel information on the structural determinants of selectivity for Atg8 proteins. Our designs will be useful tools for understanding the role of individual Atg8 proteins in autophagy and in cancer.

Stapled peptide inhibitors of Atg8 proteins

ACS Spring Meeting 2021

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.