<p><strong>BACKGROUND: </strong>Genetic alterations in multiple cell signaling pathways are involved in the molecular pathogenesis of thyroid cancer. Oncogene mutation testing and gene-expression profiling are routinely used for the preoperative risk management of adult thyroid nodules. In this study, we evaluated the potential value of miRNA biomarkers for the classification of pediatric thyroid lesions.</p>

<p><strong>PROCEDURE: </strong>Double-blind case-control study with 113 resected pediatric lesions: 66 malignant and 47 benign. Quantitative and qualitative molecular data generated with a 10-miRNA expression panel (ThyraMIR) and a next-generation sequencing oncogene panel (ThyGeNEXT) were compared with clinicopathological parameters.</p>

<p><strong>RESULTS: </strong>miRNAs were differentially expressed in benign versus malignant tumors with distinct expression patterns in different histopathology categories. The 10-miRNA classifier identified 39 (59%) malignant lesions with 100% specificity. A positive classifier score was associated with lymph node metastasis, extrathyroidal extension and intrathyroidal spread. Genetic alterations associated with increased risk for malignancy were detected in 35 (53%) malignant cases, 20 positive for point mutations in BRAF, HRAS, KRAS, NRAS, PIK3CA, or TERT and 15 positive for gene rearrangements involving ALK, NTRK3, PPARG, or RET. The 10-miRNA classifier correctly identified 11 mutation-negative malignant cases. The performance of the combined molecular test was 70% sensitivity and 96% specificity with an area under the curve of 0.924.</p>

<p><strong>CONCLUSIONS: </strong>These data suggest that the regulatory miRNA pathways underlying thyroid tumorigenesis are similar in adults and children. miRNA expression can identify malignant lesions with high specificity, augment the diagnostic yield of mutation testing, and improve the molecular classification of pediatric thyroid nodules.</p>

<p><strong>BACKGROUND: </strong>Thyroid nodules are less common in pediatric patients (≤18 years) than in adults. The Bethesda System for Reporting Thyroid Cytopathology allows for individual risk stratification, but a significant number of nodules are indeterminate. Incorporating gene mutation panels and gene expression classifiers may aid in pre-operative diagnosis. The overall aim of this study was to assess the prevalence of oncogene alterations in a representative pediatric population and across a broad-spectrum of thyroid tumor diagnoses.</p>

<p><strong>METHODS: </strong>Retrospective cross-sectional evaluation of 115 archived samples, including: 47 benign [29 follicular adenoma (FA), 11 diffuse hyperplasia, 4 thyroiditis, and 3 multinodular goiter], 6 follicular thyroid cancer (FTC), 24 follicular variant of papillary thyroid cancer (fvPTC), 27 classic variant of PTC (cPTC), 8 diffuse sclerosing variant of PTC (dsvPTC) and 3 other PTC. Molecular testing was performed by multiplex qualitative PCR followed by bead array cytometry. Oncogene results were analyzed for association with age, gender, histology, lymph node metastasis and intrathyroidal spread.</p>

<p><strong>RESULTS: </strong>A mutation in one of the 17 molecular markers evaluated was found in: 2/6 (33%) FTC, 8/24 (33%) fvPTC, 17/27 (63%) cPTC, and 4/8 (50%) dsvPTC. Mutations in RAS or PAX8-PPARG were exclusive to FTC and fvPTC, BRAF was the most common mutation in cPTC (12/17; 71%) and RET-PTC was the only mutation associated with dsvPTC. Overall, a mutation was found in 32/68 (47%) malignant specimens with a single FA positive for PAX8-PPARG. The relative distribution of gene alterations in pediatric lesions was similar to adults. Presence of BRAF mutation in pediatric cPTC did not predict a more invasive phenotype.</p>

<p><strong>CONCLUSIONS: </strong>. 32/33 nodules with genetic alterations were malignant. Mutations in RAS were most frequently associated with FTC, RET-PTC rearrangements with dsvPTC and invasive fvPTC, and BRAF with cPTC. These results suggest a clinical role for mutational analysis of pediatric nodules to guide the surgical approach.</p>