UC Berkeley

Cancer is a complex disease with high incidence and mortality rates. Conventional therapies, such as radiation or chemotherapy, often come with harmful side effects and have limited effectiveness against aggressive forms of cancer. Newer approaches, including immunotherapy and targeted therapy, have shown promise in improving therapeutic outcomes and sparing normal healthy cells from damage. While advanced sequencing technologies and research have uncovered cancer-driving molecular targets, specifically modulating those targets have remained challenging. Many proteins lack robust biochemical characterization or obvious binding pockets that traditional drugs target, rendering them “undruggable”. Chemoproteomics and fragment-based covalent ligand libraries have emerged as innovative tools for addressing these challenges, offering a more efficient and rational path to drug discovery. Furthermore, covalent drugs provide pharmacological advantages such as enhanced potency, selectivity, and prolonged duration of action.

Dysregulated oxidative stress plays a major role in cancer pathogenesis, offering a unique vulnerability for intervention. Targeting glutamate-cysteine ligase (GCL) to inhibit the synthesis of the major antioxidant glutathione (GSH) may be a promising therapeutic strategy for certain cancer types that are particularly sensitive to oxidative stress. In this dissertation, I use fragment library screening and chemoproteomic approaches for identifying new covalent inhibitors against GCL. We have discovered an acrylamide-containing cysteine-reactive ligand, EN25, that covalently targets cysteine C114 on GCLM, the modifier subunit of GCL. We show that EN25 allosterically reduces GCL activity, lowers cellular GSH levels, and impairs cell viability in ARID1A-deficient cancer cells that are particularly sensitive to glutathione depletion. We also performed a small-scale SAR study and identified additional covalent ligands with moderately enhanced binding affinity in vitro. Our studies reveal a new ligandable site within GCLM that can be targeted to inhibit the GSH synthesis in vulnerable cancer cell types.