BRAF V600E mutation has a significant association with tumor recurrences in patients with papillary thyroid cancer (PTC), reducing survival rate from 98% to 40%. Patients treated with BRAF V600E inhibitors, such as vemurafenib, acquire resistance to the drug over time (6 months) and tumor cells metastasize to distant organs, leading to patient death. Within the group of patients with BRAF V600E tumors, there are no reliable biomarkers to predict local or distant recurrences, especially when tumors acquire clonal resistance due to the effects of targeted therapies. PTC patients with the BRAF V600E and RAS mutations show an elevated ETV5 (Ets-transcript variant 5) expression. ETV5, a transcription factor, is known to promote epithelial to mesenchymal transition (EMT) in endometrial carcinomas and ovarian cancer by up-regulating ZEB1, a repressor of E-Cadherin, causing cell detachment and invasion. ETV5 also induces the expression of matrix metalloproteinase 2 (MMP-2) in human endometrial carcinomas, causing invasion.
We hypothesize that ETV5 drives cell proliferation and EMT in advanced papillary thyroid cancer. We used the cell line KTC1 (BRAF V600E mutant thyroid cancer cell line) as a model. ETV5 and BRAF V600E were knocked down using Silencer Select siRNA. Alternatively, cells were treated with standard pharmacological inhibitors (i.e. PI3Ki, AKTi, vemurafenib, ERKi, TGFBR1i). ETV5 expression was quantified using quantitative PCR and western blots. Proliferation/growth assays were performed for 5 days and the cell numbers were counted using the IN Cell 6000 confocal system and image analysis software post-transfection/treatment. Knockdown of ETV5 and/or BRAF decreased KTC1 cell proliferation. Cell growth was reduced upon treatment with vemurafenib, or PI3K-AKT and ERK inhibitors, but not with TGFBR1 inhibitors. In comparison to DMSO treatment (control), ETV5 levels were downregulated upon ERK inhibitor treatment, but were increased when treated with PI3K inhibitors. Shortterm vemurafenib treatment (48h) showed a reduction in ETV5, N-Cadherin, and CXCR4 expression, suggesting its role in EMT. In conclusion, ETV5 acts downstream of the RAS-MAPK (ERK1/2) pathway, is crucial for KTC1 cell proliferation, and may play a role in EMT in PTC.
Nguyet-Minh Hoang, ’16
Ho Chi Minh City, Vietnam
Biochemistry & Molecular Biology
Sponsor: Barbara Christie-Pope