Cell viability was determined by MTT assay, cell cycle status by propidium iodide staining, and apoptosis by Annexin-V staining and immunoblotting. Autophagy was assessed by immunofluorescence and immunoblotting.
All four HNSCC cell lines were highly sensitive to single-agent obatoclax with IC50鈥檚 ranging from 46 to 177 nM. Obatoclax induced apoptosis in all four HNSCC cell lines as evidenced by increases in sub-G1 DNA content, Annexin-V staining, and PARP cleavage. In addition, obatoclax induced autophagy in all 4 cell lines, and the addition of the autophagy inhibitor chloroquine enhanced obatoclax cytotoxicity.
Our findings demonstrate potent monotherapeutic activity of obatoclax against HNSCC cells, and enhancement of this activity in the presence of chloroquine. This preclinical study suggests that obatoclax might have therapeutic value in the treatment of HNSCC, either alone or in combination with inhibitors of autophagy.