Endocrine Abstracts

Introduction: Molecular testing is being increasingly used to detect malignancy in indeterminate thyroid nodules (Bethesda 3 & 4) which pose diagnostic dilemma. Long non-coding RNAs are crucial for metastasis, angiogenesis, and tumour growth of various cancers. Clinical utility of plasma lncRNAs as non-invasive diagnostic markers for thyroid cancer especially in indeterminate nodule remains unexplored.

Methodology: We selected lncRNAs relevant to thyroid neoplasm from LncRNA Disease 2.0 database. The lncRNAs thus selected were PVT1, HOTAIR, NEAT1, MALAT1, BANCR, HOTTIP, GAS5, H19, and GAS8-AS1. We compared the expression (using quantitative real-time PCR (qRT-PCR)) of these lncRNAs in biopsy proven differentiated thyroid cancer patients versus benign thyroid nodule subjects and healthy controls as part of first phase of study (discovery cohort). We then proceeded to determine cut-off of change of level of expression (ΔCT) for these lncRNAs (by performing a Receiver operating-characteristic (ROC) curve. These cut-offs was then applied in a validation cohort to test the ability of the same to help differentiate benign from differentiated thyroid malignancy (indeterminate nodules). We also determined the expression of these lncRNAs in FNAC materials and tissue samples for cross validation. Finally we rechecked the expression in plasma 2 weeks after surgery.

Results: We compared the plasma expression of the nine circulating lncRNAs in patients with thyroid cancer (n=25), benign thyroid nodules (n=40), and healthy controls (n=39) in the discovery cohort and the expression of PVT1, MALAT1, and BANCR were found to be significantly different between benign and malignant nodules (P<0.001). Cut off for ΔCT (obtained by ROC) for PVT1, MALAT1 and BANCR were 4.64, 6.28 and 5.94 respectively. We applied these cut-offs in a validation cohort (N=158, indeterminate nodules) and PVT1, MALAT1 and BANCR was able to differentiate malignant from benign nodule with 96.20%, 87.34% and 84.81% accuracy. PVT1 and BANCR were up-regulated in all differentiated thyroid cancer but MALAT1 was only up-regulated in PTC, but not in FV-PTC and FTC. Cross-validation of our data in FNAC material and histopathology sample reconfirmed our findings ( PVT1, MALAT1 and BANCR were up-regulated similar to that in plasma (similar quantum of increased expression)). Post-operatively plasma samples were tested and it was found that the over expression of PVT1, MALAT1 and BANCR normalized to levels similar to benign nodules.

Conclusion: Plasma PVT1, MALAT1, and BANCR may be useful molecular non-invasive diagnostic markers to help differentiate benign from malignant thyroid nodules, including in those presenting with indeterminate thyroid nodules.