Abstract 5437: Chemoproteomics-enabled discovery of covalent inhibitors targeted at AGPAT4: Unravelling tumor lineage plasticity to overcome drug resistance in hepatocellular carcinoma

Abstract The liver is a unique organ that is responsible for many metabolic functions. During hepatocellular carcinoma (HCC) development, these metabolic machineries are extensively reprogrammed to support the insatiable nutrient requirements of HCC. Tumor lineage plasticity, a recognized hallmark of cancer, is a phenomenon in which tumor cells co-opt developmental pathways to attain cellular plasticity, enabling them to evade targeted therapeutic interventions. Cancer cells can reprogram their metabolic pathways to match their increased metabolic needs for cancer cell survival under adverse conditions. Identifying novel metabolic targets related to stemness can offer promising strategies for targeting cancer stemness roots and hence to kill and control their growth. Through pathway enrichment analysis of genes that linked metabolism and stemness, we identified an aberrant glycerophospholipid metabolism signature, with AGPAT4 ranking as the top-hit. AGPAT4 upregulation in HCC is strongly correlated with aggressive clinical features, including survival, metastasis, and stemness signatures. AGPAT4 expression peaked during early liver progenitor development, decreased during hepatocyte maturation and progressively increased from well-differentiated to poorly differentiated HCCs. Enrichment of AGPAT4 in HCC is mediated by promoter binding of SOX9 to drive AGPAT4 transcriptional activity. AGPAT4 inhibition can mitigate tumor initiation, self-renewal, metastasis, and sorafenib resistance. Mechanistic studies revealed an AGPAT4-mediated phosphatidic acid production axis that promotes HCC through the regulation of mTOR signaling. Inhibition of Agpat4 by AAV8 shRNA reduced tumorigenicity and stemness, and sensitized HCC tumors to sorafenib. AGPAT4 overexpression can predict sorafenib response in clinical settings. Through chemoproteomics screening of a cysteine-reacting compound library using activity-based protein profiling, a cysteine-reacting compound with high binding affinity and selectivity towards AGPAT4 was identified and found to work synergistically with sorafenib to suppress HCC, as demonstrated in HCC patient-derived tumor xenograft (PDTX) models. Toxicity analysis through histological examination of organs, body weight measurements, and biochemical tests revealed minimal toxicity associated with the covalent inhibitor. In conclusion, AGPAT4 is a novel metabolic driver of oncogenic stemness, dedifferentiation, and metastasis in HCC. AGPAT4-induced tumor lineage plasticity may represent an Achilles heel for HCC treatment, and inhibition of AGPAT4 may widen the therapeutic window for sorafenib treatment in the clinic. Citation Format: Kai-Yu Ng, Tin-Yan Koo, Tsz-Lok Fong, Ya Gao, Tin-Lok Wong, Yuan Gao, Jing-Ping Yun, Xin-Yuan Guan, Ming Liu, Clive YS Chung, Stephanie Ma. Chemoproteomics-enabled discovery of covalent inhibitors targeted at AGPAT4: Unravelling tumor lineage plasticity to overcome drug resistance in hepatocellular carcinoma [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 5437.