A mutation panel comprising BRAF^V600E, NRAS^Q61R, and NRAS^Q61H replicated retrospective histopathological examination findings in differentiating benign goitre from malignant papillary thyroid cancer in a cohort of Malaysian patients

Thyroid malignancy status is usually confirmed through histopathological examination (HPE) following thyroidectomy. In Malaysia, the application of molecular markers in pre-operative diagnosis of thyroid cancer remains unexplored. In this study, BRAF and NRAS gene mutation panel was assessed, and the results were compared with retrospective HPE findings. Malaysian patients with benign goitre (BTG: n=33) and papillary thyroid cancer (PTC: n=25; PTCa: n=20, PTCb: n=5) were recruited at Universiti Malaya Medical Centre from September 2019 to December 2022. PCR-direct DNA sequencing of BRAF(V600), NRAS(G12), NRAS(G13), and NRAS(Q61) was conducted on DNA extracted from the patients' thyroid tissue specimens following thyroidectomy and HPE. BRAFV600E and NRAS(Q61R) mutations showed absolute PTC-specificity with PTC-sensitivity of 32% and 28%, respectively. NRAS(Q61H) demonstrated lower PTC-specificity (94%) but higher PTC-sensitivity (72%) compared to the BRAF(V600E) and NRAS(Q61R) mutations. Although the NRASG12 and NRAS(G13) variants were absent in this study, a novel NRASV14D mutation was detected in a PTCa patient. Unlike PTCb, coexistence of BRAFV600E and NRAS(Q61) variants was commonly observed among the PTCa patients. Notably, all PTCb patients had NRAS(Q61H) mutation with one patient carried both the NRAS(Q61H) and BRAF(V600E) mutations. Association analysis revealed potential link between gender, BRAF(V600E) mutation and lymph node metastasis. In conclusion, mutation panel comprising BRAF(V600E), NRAS(Q61R), and NRAS(Q61H) did not discriminate the two PTC subtypes but replicated the retrospective HPE findings in differentiating BTG from PTC. The application of this mutation panel in pre-operative diagnosis of thyroid nodules requires further validation in a larger sample size, preferably incorporating fineneedle aspirate biopsies.