Abstract 1519: Therapeutic targeting of phosphoglycerate mutase 1 disrupts cellular redox balance and promotes DNA damage in head and neck cancer

Head and neck squamous cell carcinomas (HNSCCs) are extremely aggressive cancers that exhibit notable metabolic reprogramming and resistance to therapy. Phosphoglycerate mutase 1 (PGAM1), a glycolytic enzyme, has emerged as an important regulator of cancer cell metabolism and is highly expressed in HNSCCs compared to normal (untransformed) tissues. PGAM1 catalyzes the conversion of 3-phosphoglycerate (3PG) into 2-phosphoglycerate (2PG) which is a highly important step in glycolysis due to the importance of 2PG in serine biosynthesis. In contrast, PGAM1 inhibition results in decreased 2PG production and 3PG accumulation which suppresses both glycolysis and serine metabolism. More importantly, the accumulation of 3PG downregulates the pentose phosphate pathway (PPP) via deceased glucose-6-phosphate dehydrogenase (G6PD) which is necessary for optimal NADP+/NADPH production and cellular redox balance. Given the critical function of PGAM1, this enzyme may be promising as a potential target for treating HNSCC. Here we test if PGAM1 inhibition affects cell viability, NADP+/NADPH ratios, and DNA integrity in HNSCC cells in vitro. Gene expression levels of PGAM1 were compared between control and HNSCC patient groups, and survival analysis based on PGAM1 expression was conducted utilizing data from the TCGA database. PGAM1 was shown to be highly overexpressed in HNSCCs relative to healthy/control patients (P<0.001), with its expression linked to poorer survival (HR = 1.43). In addition, the PGAM1 expression level was evaluated based on human papillomavirus (HPV) subtype and found that PGAM1 is expressed significantly higher in HPV-negative patients compared to HPV-positive. The PGAM1 inhibitor - HKB99, was used to detect changes in cell viability via MTT assays in FaDu, Cal-27 and SQ20B HPV-negative HNSCC cell lines. HKB99 over a range of 0.5 - 64 µM for 48 hours decreased cell viability in a dose-dependent manner and IC50 concentrations were established which were 4 µM, 3.4 µM and 6.4 µM for FaDu, Cal-27 and SQ20B respectively. Treatment of the 3 cell lines with HKB99 at respective IC50 doses did not affect PGAM1 mRNA expression level using PCR analyses. However, protein expression of PGAM1 using western blot was decreased in treated cell lines compared to untreated cells. Moreover, HKB99 at respective IC50 doses increased levels of γ-H2AX, a marker for DNA double-strand breaks and significantly increased NADP+/NADPH ratios. Lastly, the antioxidant N-acetylcysteine (NAC) partially rescued the cytotoxic effect of HKB99 in all 3 cell lines using MTT and clonogenic assays. These findings suggest that inhibition of PGAM1 with HKB99 in HNSCC cells increases DNA damage, disrupts redox homeostasis, and triggers cell death. Taken together, our findings indicate that PGAM1 may represent a promising therapeutic target and may potentially sensitize HNSCCs to DNA-damaging therapies. Citation Format: Md Roman Mogal, Krishna Awasthi, Ishrat Nourin Khan, Andrean Simons-Burnett. Therapeutic targeting of phosphoglycerate mutase 1 disrupts cellular redox balance and promotes DNA damage in head and neck 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 1519.