G0/G1 Switch 2 Regulates Radiation Sensitivity of Human Head and Neck Cancer through a G1-lipid Checkpoint

Head and neck squamous cell carcinoma (HNSCC) resistance to radiotherapy has prompted a need to develop adaptive radiation therapy protocols to improve patient outcomes. This study investigates the hypothesis that lipid metabolism regulates cell cycle phase-specific radiation sensitivity of HNSCC cells. Previous studies have shown that HNSCC tumors with a higher proportion of G0/G1 phase cells (low proliferative index, LPI) are more resistant to radiation compared to HNSCC tumors with a higher proportion of S/G2 phase cells (high proliferative index, HPI). RNA-seq and bioinformatics identified lipid metabolism as the major intrinsic pathway that differs between HPI and LPI HNSCC cultures. mRNA and protein levels of G0/G1 Switch 2 gene (G0S2), regulator of quiescence and lipid metabolism, were upregulated in LPI compared to HPI HNSCC cultures. G0S2 negatively regulates adipose triglyceride lipase (ATGL), resulting in less lipolytic activity. siG0S2 treatment of LPI cultures recruited cells into the proliferative cycle and exacerbated radiation sensitivity. To override G0S2 action, we incubated LPI cultures with the fatty acid palmitate and examined cellular metabolic stress markers. Compared to controls, LPI cultures treated with palmitate showed increased reactive oxygen species levels, lipid peroxidation and oxygen consumption rate coupled with increased mitochondrial fission. Furthermore, using the fluorescent based cell cycle real-time imaging system, we showed that palmitate treatment sustained cell proliferation (higher S/G2) compared to controls (higher G1). Palmitate treatment resulted in significant sensitization to radiation treatment and enhanced the efficacy of poly (ADP-ribose) polymerase (PARP) inhibitors. In summary, we demonstrate that G0S2-dependent lipid metabolism regulates cell cycle phase-specific radiation sensitivity of HNSCC cells and identify G0S2 and free fatty acids as novel targets for radiation therapy.