Abstract 6873: Discovery and characterization of novel ULK1/2 inhibitors and their ability to degrade ATG13, inhibit autophagy, and treat lung cancer

Abstract Lung cancer is the leading cause of cancer-related mortality across all populations. Autophagy (ATG) is a cell survival process by which dysfunctional cellular components are removed and recycled to maintain homeostasis through a lysosomal dependent degradation mechanism. ATG upregulation is implicated as a bad actor in several diseases, particularly K-Ras driven non-small cell lung cancer (NSCLC), where it provides nutrients to lung cancer cells in the nutrient poor and hypoxic tumor microenvironment. ATG also contributes to chemotherapeutic resistance, making it a viable pathway for therapeutic intervention. However, the only clinically available drugs targeting ATG are chloroquine and hydroxychloroquine, which are not ATG-specific and have multiple adverse side effects. ATG is an intricate, multistep process beginning with the Unc-51-like autophagy-activating kinases 1 and 2 (ULK1/2) initiation complex. The supramolecular assembly of this complex relies on the adaptor protein ATG13, which binds ULK1/2. Knockout of ATG13 significantly reduces the viability of lung cancer cell lines, indicating that ATG13 is a viable therapeutic target for lung cancer. Furthermore, our group has characterized other autophagy inhibiting compounds that also robustly degrade ATG13 in cells and tissues, leading to cancer cell death. We have developed and optimized a high throughput screening (HTS) assay whereby ATG13 is HiBit tagged in A549 NSCLC cells (ATG13 HiBit assay) to allow identification of compounds that induce ATG13 protein loss, and therefore inhibit ATG. We have also developed a testing funnel that employs multiple assays to characterize compounds that degrade ATG13 independent of cellular toxicity (cell viability assay), confirm their activity against our protein of interest (In Cell Western blot ATG13 assay), measure their ability to inhibit autophagy (GFP-LC3-RFP-LC3∆G ATG flux assay) and eliminate compounds that cause assay interference (HiBit counterscreen). Using these assays we have identified a novel lead compound which degrade ATG13 and inhibits autophagy resulting in cell death in NSCLC. Citation Format: Patrick Martin Hagan, Allison Limpert, Douglas Sheffler, Huiyu Ren, Nicholas Cosford. Discovery and characterization of novel ULK1/2 inhibitors and their ability to degrade ATG13, inhibit autophagy, and treat lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6873.