Publications by Year: 2017

PURPOSE: Papillary thyroid carcinoma (PTC) is the most frequent endocrine tumor. BRAFV600E represents the PTC hallmark and is targeted with selective inhibitors (e.g. vemurafenib). Although there have been promising results in clinical trials using these inhibitors, most patients develop resistance and progress. Tumor clonal diversity is proposed as one mechanism underlying drug resistance. Here we have investigated mechanisms of primary and secondary resistance to vemurafenib in BRAFWT/V600E-positive PTC patient-derived cells with P16-/- (CDKN2A-/-).

EXPERIMENTAL DESIGN: Following treatment with vemurafenib, we expanded a sub-population of cells with primary resistance and characterized them genetically and cytogenetically. We have used exome sequencing, metaphase chromosome analysis, FISH and oligonucleotide SNP-microarray assays to assess clonal evolution of vemurafenib-resistant cells. Furthermore, we have validated our findings by networks and pathways analyses using PTC clinical samples.

RESULTS: Vemurafenib-resistant cells grow similarly to naïve cells but are refractory to apoptosis upon treatment with vemurafenib, and accumulate in G2-M phase. We find that vemurafenib-resistant cells show amplification of chromosome 5 and de novo mutations in the RBM (RNA-binding motifs) genes family (i.e. RBMX, RBM10). RBMX knockdown in naïve-cells contributes to tetraploidization, including expansion of clones with chromosome 5 aberrations (e.g. isochromosome 5p). RBMX elicits gene regulatory networks with chromosome 5q cancer-associated genes and pathways for G2-M and DNA damage-response checkpoint regulation in BRAFWT/V600E-PTC. Importantly, combined therapy with vemurafenib plus palbociclib (inhibitor of CDK4/6, mimicking P16 functions) synergistically induces stronger apoptosis than single agents in resistant-cells and in anaplastic thyroid tumor cells harboring the heterozygous BRAFWT/V600E mutation.

CONCLUSIONS: Critically, our findings suggest for the first time that targeting BRAFWT/V600E and CDK4/6 represents a novel therapeutic strategy to treat vemurafenib-resistant or vemurafenib-naïve radioiodine-refractory BRAFWT/V600E-PTC. This combined therapy could prevent selection and expansion of aggressive PTC cell sub-clones with intrinsic resistance, targeting tumor cells either with primary or secondary resistance to BRAFV600E inhibitor.

INTRODUCTION: The follicular variant of papillary thyroid cancer (FVPTC) can be noninvasive or invasive. The invasive form of FVPTC commonly harbors BRAF mutations whereas RAS mutations are more often associated with noninvasive FVPTC and a favorable clinical outcome.

CASE REPORT: A 47-year-old man presented with a metastasis to his right iliac bone as the initial manifestation of a 1.6cm invasive FVPTC. After total thyroidectomy, the patient underwent additional treatment, including thyroid hormone suppressive treatment to non-detectable TSH levels, repeated courses of radioiodine treatment, external beam radiation, and treatment with the tyrosine kinase inhibitor sorafenib. Despite these therapeutic efforts, the disease progressed with growth of the iliac mass and additional metastatic spread to cervical and lumbar vertebrae causing increasing pain and disability. The patient succumbed to the disease four years after presentation. Retrospective next-generation sequencing of the primary tumor using a pan-cancer targeted mutation and gene fusion panel revealed NRAS Q61K mutation and no other oncogenic alterations.

DISCUSSION: The study challenges the concept that thyroid neoplasms with isolated RAS mutations are often associated with favorable clinical behavior and may be candidates for conservative management.

CONCLUSION: An isolated RAS mutation in invasive FVPTC may be associated with an aggressive clinical behavior.