Stimulation of the toll-like receptor 3 promotes metabolic reprogramming in head and neck carcinoma cells

Abstract

In this study, a possible link between the innate immune recognition receptor TLR3 and metabolic reprogramming in Head and Neck carcinoma (HNC) cells was investigated. The effects of TLR3 stimulation/knock-down were assessed under several culture conditions in 4 HNC cell-lines by cell growth assays, targeted metabolomics, and glycolysis assays based on time-resolved analysis of proton release (Seahorse analyzer). The stimulation of TLR3 by its synthetic agonist Poly(A:U) resulted in a faster growth of HNC cells under low foetal calf serum conditions. Targeted analysis of glucose metabolism pathways demonstrated a tendency towards a shift from tricarboxylic acid cycle (Krebs cycle) to glycolysis and anabolic reactions in cells treated with Poly(A:U). Glycolysis assays confirmed that TLR3 stimulation enhanced the capacity of malignant cells to switch from oxidative phosphorylation to extra-mitochondrial glycolysis. We found evidence that HIF-1α is involved in this process: addition of the TLR3 agonist resulted in a higher cell concentration of the HIF-1α protein, even in normoxia, whereas knocking-down TLR3 resulted in a lower concentration, even in hypoxia. Finally, we assessed TLR3 expression by immunohistochemistry in a series of 7 HNSCC specimens and found that TLR3 was detected at higher levels in tumors displaying a hypoxic staining pattern. Overall, our results demonstrate that TLR3 stimulation induces the Warburg effect in HNC cells in vitro, and suggest that TLR3 may play a role in tumor adaptation to hypoxia.