The development of rapid and accurate screening test for RET hotspot somatic and germline mutations in MEN2 syndromes

•The development of high resolution melt analysis method for RET mutation detection•Novel germline RET proto-oncogene variant c.1915G > A (p.Ala639Thr)•A novel, high-throughput, accurate and cost-effective approach with faster turnaround time

Medullary thyroid carcinoma (MTC) is a rare endocrine malignancy with distinctive features separating it from other thyroid cancers. Cancer may be sporadic or occur as a consequence of the hereditary syndrome called multiple endocrine neoplasia type 2 (MEN2) with three distinct phenotypes in MEN2A, MEN2B and FMTC. Each variant of MEN2 results from different RET gene mutations, with a good genotype–phenotype correlation. The goal of the study was to develop a fast and accurate screening method for a reliable detection of hot-spot RET germline and sporadic tumor mutations. From a cohort of 191 patients with MTC and their relatives, 38 tested positive and 31 tested negative for a germline or somatic tumor RET mutation were selected. A positive HRM mutation pattern was detected in all mutation-positive patients and altogether the method was able to clearly differentiate between twenty different genotypes. A novel germline variant p.Ala639Thr was detected in MTC patient, which was determined to be likely benign. Analytical specificity was determined to be 98.6% and a sensitivity threshold was determined to be 30%. The fast and accurate HRM method reduces the turnaround time providing fast and important information, especially when targeted anti-tyrosine kinase therapy on tumor samples is considered. Overall, we developed a high-throughput, accurate and cost-effective approach for the detection of RET germline and sporadic tumor mutations.