Genes Involved in the Enhancement of Mild Hyperthermia Sensitivity by Chloroquine in Human Oral Squamous Cell Carcinoma HSC-3 Cells

Abstract: Chloroquine (CQ), an autophagy inhibitor, enhances thermosensitivity in cancer cells; however, the molecular mechanisms underlying this effect remain unclear. To investigate these mechanisms, we conducted a comprehensive gene expression analysis in human oral squamous cell carcinoma HSC-3 cells treated with both CQ and mild hyperthermia (MHT). GeneChip® analysis revealed that numerous differentially expressed probe sets following treatment with MHT (42 ℃ for 90 min) and its combination with CQ (30 μM). These probe sets were categorized into four clusters using K-means clustering: Up-I and Up-II (upregulated), and Down-I and Down-II (downregulated). Based on the Ingenuity® Pathways Knowledge Base, three gene networks － GN-I, GN-II, and GN-III － were identified from the upregulated clusters. GN-I, GN-II, and GN-III consisted of 8, 7, and 7 genes, respectively, and were associated with the biological functions of activation of autophagy, prevention of cell death, and induction of cell death. Real-time quantitative PCR confirmed that expression levels of HSPA5, BAG3, HSPA1A, DNAJA1, and HSPH1 － all belonging to GN-I or GN-II － were increased by MHT alone but significantly suppressed when combine with CQ. In contrast, the expression levels of FOS, EGR1, and JUN － members of GN-III － were upregulated by MHT and further enhanced by CQ. These findings provide new insights into the mechanisms through which CQ enhances thermosensitivity in cancer cells.