Exp Clin Endocrinol Diabetes 2013; 121 - P12
DOI: 10.1055/s-0033-1336704

BRAF, NRAS, HRAS, KRAS, PAX8/PPARG, RET/PTC mutation screening in routine air dried Fine Needle Aspiration (FNA) smears from 310 patients with nodular thyroid disease

M Eszlinger 1, A Krogdahl 2, S Münz 1, C Rehfeld 1, E Jensen 2, C Ferraz 1, E Bösenberg 1, N Drieschner 3, M Scholz 4, L Hegedüs 5, R Paschke 1
  • 1Universität Leipzig, Klinik und Poliklinik für Endokrinologie und Nephrologie, Leipzig, Germany
  • 2Odense University Hospital, Department of Pathology, Odense, Denmark
  • 3Universität Bremen, Zentrum für Humangenetik, Bremen, Germany
  • 4Universität Leipzig, Institute for Medical Informatics, Statistics and Epidemiology (IMISE), Leipzig, Germany
  • 5Odense University Hospital, Department of Endocrinology and Metabolism, Odense, Denmark

Fine needle aspiration (FNA) is the most sensitive method to select suspicious thyroid nodules for surgery. However, this method has inherent limitations, e.g. “indeterminate” samples. As rearrangements (PAX8/PPARG, RET/PTC) and point mutations (BRAF, NRAS, HRAS, KRAS) have been detected in follicular carcinomas (FTC) and papillary carcinomas (PTC), their detection in FNA smears could improve the diagnosis. However, rearrangements have up to date only been detected in fresh FNA material and the number of FTCs was rather low in previous studies.

RNA and DNA was extracted from 310 routine air-dried FNA smears (164 indeterminate, 57 malignant, 89 non-neoplastic) and corresponding formalin-fixed paraffin-embedded tissue (FFPE) samples (156 follicular adenomas, 32 FTCs, 9 follicular variant PTCs, 44 PTCs, 69 goiters). PAX8/PPARG and RET/PTC1 and 3 rearrangements were detected by qPCR, while BRAF and RAS point mutations were detected by high resolution melting (HRM)-PCR and by pyrosequencing.

On average, 8% and 3.9% of routine FNA samples did not allow analysis of a point mutation or rearrangement, respectively. For the 164 indeterminate samples, NRAS mutations could be detected in 12 FNA samples. HRAS mutations were detected in 3 FNA samples. No KRAS mutation was detected in the FNA samples. PAX8/PPARG was detected in 6 FNA samples, while RET/PTC was not detected in any indeterminate sample. Molecular FNA screening increased the sensitivity from 67% (cytology alone) to 75% (cytology + molecular FNA screening) in the total set. In the indeterminate set with 19 FTCs the sensitivity of detecting carcinomas and mutation positive adenomas was 48% and specificity was 99%.

In summary, molecular screening for point mutations and rearrangements is feasible in routine air dried FNA smears. Analyzing this panel of mutations, especially in indeterminate routine air-dried FNA smears, may reduce the number of diagnostic thyroid surgeries.