Papillary Thyroid Cancer-Driving Oncogene BRAFV600E Promotes Aberrant Toll-Like Receptor 4 Overexpression

PEYRET V, ; NAZAR M; MARTÍN M; FUZIWARA CS; FERNANDEZ EA; MONTESINOS MM; PELLIZAS CG; KIMURA ET; NICOLA JP; MASINI - REPISO AM

Rio de Janeiro

Congreso; XVI Latin American Thyroid Congress (LATS); 2017

Latin American Thyroid Society (LATS)

Introduction: Emerging evidence suggests that hyperactivity of Toll like receptors (TLRs) signaling promotes tumor survival signals, thus favoring tumor progression. Recently, aberrant TLR4 overexpression was evidenced in papillary thyroid carcinomas (PTC)Objective: To study the mechanisms underlying TLR4 overexpression in PTC harboring the BRAFV600E mutation.Methods: TLR4 expression was studied in thyroid tissue derived from human PTCs and transgenic mice expressing BRAFV600E in thyrocytes (Tg-BRAFV600E mice) (IHC, RT/qPCR). BRAFV600E-positive PTC cell line BCPAP and PCCl3 cells expressing BRAFV600E in response to doxycycline (PC/BRAFV600E) were used to study BRAFV600E-driven TLR4 expression (western blot, RT/qPCR, siRNA silencing, luciferase assay). The Cancer Genome Atlas (TCGA) database was used to perform combined analysis.Results: We evidenced TLR4 overexpressionin PTCs compared to normal thyroid tissues. Moreover, match-samples of primary PTCs and its lymph node metastasis showed a significant upregulation of TLR4 levels in the metastatic tissues. In agreement, TLR4 expression was increased in the thyroid tissue of Tg-BRAFV600E mice compared to littermate controls. Furthermore, we demonstrated functional TLR4 expression in PTC cells models which evidenced an increased NF-κB transcriptional activity in response to the exogenous TLR4-agonist lipopolysaccharide.TCGA data analysis revealed that patients with BRAFV600E-positive tumors and high TLR4 expression have shorter disease-free survival. Consistently with transcriptomic data showing correlation between TLR4 expression and ERK activation score, conditional BRAFV600E expression in PC/BRAFV600E cells upregulates TLR4 protein levels. Moreover, chemical blockage of MAPK/ERK signaling abrogated BRAFV600E-induced TLR4 expression. Deletion analysis of TLR4 promoter revealed a critical MAPK/ERK-sensitive ETS binding-site involved in BRAFV600E responsiveness. Furthermore, we evidenced that the ETS binding factor ETS1 is critical for BRAFV600E-driven MAPK/ERK signaling-mediated TLR4 gene expression in PTCs.Conclusions: Increased TLR4 expression in PTCs would be a functional consequence of deregulated MAPK/ERK/ETS1 signaling as a result of thyroid tumors-drivers oncogenes such as BRAFV600E. Considering the oncogenic ability of aberrant NF-κB signaling activation in the promotion of thyroid tumor growth, our results suggest a pro-oncogenic potential of TLR4 downstream signaling in thyroid tumorigenesis.