Significance of Allelic Percentage of BRAF c.1799T > A (V600E) Mutation in Papillary Thyroid Carcinoma

AbstractBackground Somatic BRAF mutation is frequently observed in papillary thyroid carcinoma (PTC). Recent evidence suggests that PTCs are heterogeneous tumors containing a subclonal or oligoclonal occurrence of BRAF mutation. Conflicting results have been reported concerning the prognostic significance of the mutant allele frequency. Our present aim was to investigate the association between the percentage of BRAF c.1799T > A (p.Val600Glu) alleles and clinicopathological parameters in PTC.Methods Genomic DNA was extracted from fresh-frozen specimens obtained from 50 PTC patients undergoing total thyroidectomy. The BRAF mutation status was determined by Sanger sequencing. The percentage of mutant BRAF alleles was quantified by mass spectrometric genotyping, pyrosequencing, and competitive allele-specific TaqMan PCR (castPCR).ResultsPositive rate of BRAF mutation was 72 % by Sanger sequencing, 82 % by mass spectrometric genotying, and 84 % by pyrosequencing or castPCR. The average percentage of mutant BRAF alleles was 22.5, 31, and 30.7 %, respectively. There was a good correlation among three quantification methods (Spearman’s rho = 0.87–0.97; p < 0.0001). The mutant allele frequency was significantly correlated with tumor size (rho = 0.47–0.52; p < 0.01) and extrathyroidal invasion. The frequency showed no difference in pathological lymph node metastasis.ConclusionsThe percentage of mutant BRAF alleles is positively associated with tumor burden and extrathyroidal invasion in PTC. Relatively good correlations exist among mass spectrometric genotyping, pyrosequencing, and castPCR in quantification of mutant BRAF allele frequency.

[1]  Q. Duh,et al.  Higher rate of BRAF mutation in papillary thyroid cancer over time , 2011, Cancer.

[2]  M. Schott.  Association of High Iodine Intake with the T1799A BRAF Mutation in Papillary Thyroid Cancer , 2010 .

[3]  Ed Y. Tom,et al.  Does addition of BRAF V600E mutation testing modify sensitivity or specificity of the Afirma Gene Expression Classifier in cytologically indeterminate thyroid nodules? , 2013, The Journal of clinical endocrinology and metabolism.

[4]  M. Cirillo,et al.  A high percentage of BRAFV600E alleles in papillary thyroid carcinoma predicts a poorer outcome. , 2012, The Journal of clinical endocrinology and metabolism.

[5]  Y. Youn,et al.  The association of the BRAFV600E mutation with prognostic factors and poor clinical outcome in papillary thyroid cancer , 2012, Cancer.

[6]  Y. Kim,et al.  BRAFV600E Mutation Analysis in Papillary Thyroid Carcinomas by Peptide Nucleic Acid Clamp Real-time PCR , 2013, Annals of Surgical Oncology.

[7]  S. Gabriel,et al.  High-throughput oncogene mutation profiling in human cancer , 2007, Nature Genetics.

[8]  J. Bishop,et al.  BRAF Mutation in Papillary Thyroid Cancer and Its Value in Tailoring Initial Treatment: A Systematic Review and Meta-Analysis , 2012, Medicine.

[9]  M. Pfaffl,et al.  A new mathematical model for relative quantification in real-time RT-PCR. , 2001, Nucleic acids research.

[10]  M. Zeiger,et al.  BRAF V600E mutation and its association with clinicopathological features of papillary thyroid cancer: a meta-analysis. , 2012, The Journal of clinical endocrinology and metabolism.

[11]  M. Santoro,et al.  The heterogeneous distribution of BRAF mutation supports the independent clonal origin of distinct tumor foci in multifocal papillary thyroid carcinoma. , 2007, The Journal of clinical endocrinology and metabolism.

[12]  V. Marotta,et al.  The primary occurrence of BRAF(V600E) is a rare clonal event in papillary thyroid carcinoma. , 2012, The Journal of clinical endocrinology and metabolism.

[13]  M. Zeiger,et al.  BRAF V600E Mutation Independently Predicts Central Compartment Lymph Node Metastasis in Patients with Papillary Thyroid Cancer , 2012, Annals of Surgical Oncology.

[14]  C. Sander,et al.  V600EBRAF is associated with disabled feedback inhibition of RAF–MEK signaling and elevated transcriptional output of the pathway , 2009, Proceedings of the National Academy of Sciences.

[15]  Lei Zhang,et al.  Targeted expression of BRAFV600E in thyroid cells of transgenic mice results in papillary thyroid cancers that undergo dedifferentiation. , 2005, Cancer research.

[16]  M. Xing,et al.  BRAFV600E mutation and papillary thyroid cancer: chicken or egg? , 2012, The Journal of clinical endocrinology and metabolism.

[17]  Somak Roy,et al.  Targeted next-generation sequencing panel (ThyroSeq) for detection of mutations in thyroid cancer. , 2013, The Journal of clinical endocrinology and metabolism.

[18]  J. Romijn,et al.  Survival and death causes in differentiated thyroid carcinoma. , 2006, The Journal of clinical endocrinology and metabolism.

[19]  S. Y. Park,et al.  Analysis of differential BRAF(V600E) mutational status in multifocal papillary thyroid carcinoma: evidence of independent clonal origin in distinct tumor foci. , 2006, Cancer.

[20]  M. Xing,et al.  BRAF mutation in papillary thyroid cancer: pathogenic role, molecular bases, and clinical implications. , 2007, Endocrine reviews.

[21]  H. Karim-Kos,et al.  Rising incidence, no change in survival and decreasing mortality from thyroid cancer in The Netherlands since 1989. , 2013, Endocrine-related cancer.

[22]  M. Belvin,et al.  RAF inhibitors prime wild-type RAF to activate the MAPK pathway and enhance growth , 2010, Nature.

[23]  Laura E. MacConaill,et al.  Existing and emerging technologies for tumor genomic profiling. , 2013, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[24]  B. Delahunt,et al.  Most multifocal papillary thyroid carcinomas acquire genetic and morphotype diversity through subclonal evolution following the intra‐glandular spread of the initial neoplastic clone , 2008, The Journal of pathology.

[25]  Carlos Caldas,et al.  The implications of clonal genome evolution for cancer medicine. , 2013, The New England journal of medicine.

[26]  K. Polyak,et al.  Tumor heterogeneity: causes and consequences. , 2010, Biochimica et biophysica acta.

[27]  Jong Min Lee,et al.  Factors influencing the detection of the BRAF T1799A mutation in papillary thyroid carcinoma. , 2011, Oncology reports.

[28]  J. Nortier,et al.  New treatment modalities in advanced thyroid cancer. , 2012, Annals of oncology : official journal of the European Society for Medical Oncology.

[29]  L. Kristensen,et al.  Evaluation of BRAF mutation testing methodologies in formalin-fixed, paraffin-embedded cutaneous melanomas. , 2013, The Journal of molecular diagnostics : JMD.

[30]  Guojun Wu,et al.  High prevalence and possible de novo formation of BRAF mutation in metastasized papillary thyroid cancer in lymph nodes. , 2005, The Journal of clinical endocrinology and metabolism.

[31]  M. McMahon,et al.  Mutationally activated BRAF(V600E) elicits papillary thyroid cancer in the adult mouse. , 2011, Cancer research.

[32]  D. Barford,et al.  Mechanism of Activation of the RAF-ERK Signaling Pathway by Oncogenic Mutations of B-RAF , 2004, Cell.

[33]  Joanna A Gibson,et al.  BRAFV600E mutation in papillary thyroid microcarcinoma: a genotype–phenotype correlation , 2013, Modern Pathology.

[34]  P. Fu,et al.  Analysis of the rising incidence of thyroid cancer using the Surveillance, Epidemiology and End Results national cancer data registry. , 2010, Surgery.

[35]  S. Piana,et al.  Allele percentage of the BRAF V600E mutation in papillary thyroid carcinomas and corresponding lymph node metastases: no evidence for a role in tumor progression. , 2013, The Journal of clinical endocrinology and metabolism.

[36]  Chien-Liang Liu,et al.  Expression and Biologic Significance of Adiponectin Receptors in Papillary Thyroid Carcinoma , 2013, Cell Biochemistry and Biophysics.