The influence of hyaluronan-CD44 interaction on topoisomerase II activity and etoposide cytotoxicity in head and neck cancer

Objective: To investigate the downstream molecular targets of hyaluronan (HA)-CD44 and phospholipase C (PLC)-mediated calcium ion (Ca²⁺) signaling in head and neck squamous cell carcinoma (HNSCC). Hyaluronan is a ligand for the CD44 receptor, which interacts with multiple signaling pathways to influence cellular behavior. We recently determined that HA-CD44 interaction promotes PLC-mediated Ca²⁺ signaling and cisplatin resistance in HNSCC. Design: Proliferation of HNSCC tumor cells and topoisomerase (Topo) II enzymatic activity, including DNA-cleavable complex formation and DNA decatenation, were analyzed in the presence or absence of HA, the Topo II poison etoposide (VP-16), and various inhibitors of PLC and Ca²⁺-calmodulin kinase II (CaMKII) signaling. Results: Treatment with HA promoted Topo II phosphorylation, suggesting that HA can modulate Topo II activity. Topoisomerase II-mediated DNA cleavable complex formation was increased by VP-16, and this increase was significantly enhanced by noncytotoxic doses of the PLC inhibitor U73122 and the CaMKII inhibitor KN-62, implicating PLC and CaMKII in Topo II regulation. However, the drug- and inhibitor-mediated increase in DNA cleavable complex formation was reduced with HA pretreatment. Inhibitors of PLC and CaMKII also enhanced VP-16 inhibition of Topo II-mediated DNA decatenation. Treatment with HA reduced VP-16 cytotoxic activity. On the other hand, U73122 and KN-62 enhanced VP-16 cytotoxic activity and reduced the ability of HA to promote VP-16 resistance. Conclusion: Our results suggest that HA, PLC, and CaMKII are upstream regulators of Topo II-mediated DNA metabolism in HNSCC and that this signaling pathway could be a promising target for the development of novel therapies against HNSCC.