Abstract 3239: Targeting the GAS41 YEATS domain in non-small cell lung cancer with novel small molecules
Authors: Winkler Alyssa , Listunov Dymytrii , Linhares Brian , Ray Joshua , Weaver Sidney , Zolov Sergey , Keshamouni Venkat , Grembecka Jolanta , Cierpicki Tomaszc 
Published: March 2024

Abstract
Background: Glioma amplified sequence 41 (GAS41) belongs to the Yaf9, ENL, AF9, Taf14, and Sas5 (YEATS) domain family of epigenetic reader proteins. This domain preferentially recognizes acetylated lysine residues on chromatin to facilitate gene expression by recruiting chromatin remodeling and transcription activating complexes. The YEATS4 gene encoding for GAS41 protein is frequently amplified in NSCLC patient samples relative to normal lung tissue, and GAS41 protein is overexpressed in NSCLC cells. In knockdown experiments using NSCLC xenograft mouse models, GAS41 has been shown to be essential for NSCLC tumor growth. In mutagenesis studies, the YEATS domain residues responsible for acetylation reader activity were found to be essential for NSCLC cellular proliferation and tumor progression. Thus, the YEATS domain of GAS41 represents an attractive target for the development of small-molecule inhibitors as potential therapeutics for NSCLC.
Results: We developed the first-in-class small-molecule inhibitors for the GAS41 YEATS domain. Using an NMR-based fragment screen, we identified a hit with low millimolar binding affinity. Structure-based drug design followed by extensive medicinal chemistry optimization yielded the potent and selective inhibitor DLG-369 with sub-micromolar inhibition of the GAS41 YEATS domain interaction with acylated histones. Using the NanoBRET platform, we showed that DLG-369 selectively disrupts the association of GAS41 YEATS with chromatin in mammalian cells. Furthermore, DLG-369 inhibits proliferation of NSCLC cell lines without affecting normal lung fibroblasts. In transcriptomic experiments, DLG-369 treatment regulates expression of multiple genes in NSCLC cells, including upregulation of the critical tumor suppressor genes CDKN1A and CYR61 and downregulation of chemokine genes CXCL5 and CXCR4 associated with NSCLC progression.
Conclusions: We developed DLG-369, a potent inhibitor of the GAS41 YEATS domain with activity in NSCLC cellular models. DLG-369 potently binds to the YEATS domain acetylation reader pocket and disrupts GAS41 recognition of acetylation in cells. DLG-369 potently reduces proliferation and drives differential expression of key genes associated with NSCLC. DLG-369 can serve as a chemical probe to further interrogate the biological role of the GAS41 YEATS domain in NSCLC and other cancers for potential therapeutic implications.
Citation Format: Alyssa Winkler, Dymytrii Listunov, Brian Linhares, Joshua Ray, Sidney Weaver, Sergey Zolov, Venkat Keshamouni, Jolanta Grembecka, Tomaszc Cierpicki. Targeting the GAS41 YEATS domain in non-small cell lung cancer with novel small molecules [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3239.