Abstract
BRAF(V600E) mutation exerts an essential oncogenic function in many tumors, including papillary thyroid carcinoma (PTC). Although BRAF(V600E) inhibitors are available, lack of response has been frequently observed. To study the mechanism underlying intrinsic resistance to the mutant BRAF(V600E) selective inhibitor vemurafenib, we established short-term primary cell cultures of human metastatic/recurrent BRAF(V600E)-PTC, intrathyroidal BRAF(V600E)-PTC, and normal thyroid (NT). We also generated an early intervention model of human BRAF(V600E)-PTC orthotopic mouse. We find that metastatic BRAF(V600E)-PTC cells elicit paracrine-signaling which trigger migration of pericytes, blood endothelial cells and lymphatic endothelial cells as compared to BRAF(WT)-PTC cells, and show a higher rate of invasion. We further show that vemurafenib therapy significantly suppresses these aberrant functions in non-metastatic BRAF(V600E)-PTC cells but lesser in metastatic BRAF(V600E)-PTC cells as compared to vehicle treatment. These results concur with similar folds of down-regulation of tumor microenvironment-associated pro-metastatic molecules, with no effects in BRAF(WT)-PTC and NT cells. Our early intervention preclinical trial shows that vemurafenib delays tumor growth in the orthotopic BRAF(WT/V600E)-PTC mice. Importantly, we identify high copy number gain of MCL1 (chromosome 1q) and loss of CDKN2A (P16, chromosome 9p) in metastatic BRAF(V600E)-PTC cells which are associated with resistance to vemurafenib treatment. Critically, we demonstrate that combined vemurafenib therapy with BCL2/MCL1 inhibitor increases metastatic BRAF(V600E)-PTC cell death and ameliorates response to vemurafenib treatment as compared to single agent treatment. In conclusion, short-term PTC and NT cultures offer a predictive model for evaluating therapeutic response in patients with PTC. Our PTC pre-clinical model suggests that combined targeted therapy might be an important therapeutic strategy for metastatic and refractory BRAF(V600E)-positive PTC.
Highlights
The BRAFV600E mutation is the most common genetic alteration in papillary thyroid carcinoma (PTC) and may be associated with progression of PTC to anaplastic thyroid carcinoma (ATC) [1, 2], one of the most lethal human cancers [3, 4]
We have developed the first pre-clinical model of patient-derived PTC with BRAFWT/V600E (Figure 1A) using BRAFWT/V600E inhibitors
We established 7 short-term primary cell cultures in vitro of human PTC, including 2 from non angioinvasive and non-metastatic PTC, 2 from angioinvasive PTC with no clinical evidence of neck lymph node (LN) metastasis, 2 from angioinvasive PTC with positive clinical evidence of neck LN metastasis, and, 1 short-term cell culture from a PTC mediastinal LN metastasis
Summary
The BRAFV600E mutation is the most common genetic alteration in papillary thyroid carcinoma (PTC) and may be associated with progression of PTC to anaplastic thyroid carcinoma (ATC) [1, 2], one of the most lethal human cancers [3, 4]. We describe the development of the first patient-derived BRAFWT/V600E-PTC in vitro and in vivo early intervention pre-clinical model with some similar disease molecular characteristics that are recapitulated. This model offers interpretative insight into the concurrent vemurafenib human clinical trials in an independent cohort of patients with metastatic BRAFV600EPTC, may serve to provide rapid clinical translation of our findings. Our PTC pre-clinical model suggests that combination of anti-BRAFWT/V600E therapy (e.g. vemurafenib) with inhibitors of pro-survival molecules (i.e. pan-BCL2/MCL1 inhibitors) ameliorates intrinsic resistance to metastatic BRAFV600E-PTC cell death
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