Multicentre validation of a microRNA-based assay for diagnosing indeterminate thyroid nodules utilising fine needle aspirate smears

Abstract

AimsThe distinction between benign and malignant thyroid nodules has important therapeutic implications. Our objective was to develop an assay that could classify indeterminate thyroid nodules as benign or suspicious, using routinely prepared fine needle aspirate (FNA) cytology smears.MethodsA training set of 375 FNA smears was used to develop the microRNA-based assay, which was validated using a blinded, multicentre, retrospective cohort of 201 smears. Final diagnosis of the validation samples was determined based on corresponding surgical specimens, reviewed by the contributing institute pathologist and two independent pathologists. Validation samples were from adult patients (≥18 years) with nodule size >0.5 cm, and a final diagnosis confirmed by at least one of the two blinded, independent pathologists. The developed assay, RosettaGX Reveal, differentiates benign from malignant thyroid nodules, using quantitative RT-PCR.ResultsTest performance on the 189 samples that passed quality control: negative predictive value: 91% (95% CI 84% to 96%); sensitivity: 85% (CI 74% to 93%); specificity: 72% (CI 63% to 79%). Performance for cases in which all three reviewing pathologists were in agreement regarding the final diagnosis (n=150): negative predictive value: 99% (CI 94% to 100%); sensitivity: 98% (CI 87% to 100%); specificity: 78% (CI 69% to 85%).ConclusionsA novel assay utilising microRNA expression in cytology smears was developed. The assay distinguishes benign from malignant thyroid nodules using a single FNA stained smear, and does not require fresh tissue or special collection and shipment conditions. This assay offers a valuable tool for the preoperative classification of thyroid samples with indeterminate cytology.

Ancillary molecular testing of indeterminate thyroid nodules

“…These markers have been tested both individually and in panels even though only some of them (BRAF V600E mutation and RET/PCT rearrangements) show high specificity and likelihood of a malignant diagnosis in mutated thyroid lesions. Molecular thyroid tests that are commercially available in the United States are: 1) ThyroSeq (University of Pittsburgh Medical Center/Cytopath Biopsy Lab, Pittsburgh, PA), 2) Afirma Gene Expression Classifier (GEC; Veracyte, South San Francisco, CA), 3) RosettaGX Reveal (Rosetta Genomics, Philadelphia, PA), and 4) ThyGenX and ThyraMIR (both from Interpace Diagnostics, Parsippany, NJ) . Whereas the Thyroseq and ThyGenX tests have high positive predictive value (PPV) and negative predictive value (NPV) wherein high PPV helps as a “rule‐in malignancy” test, the Afirma GEC with its high NPV helps as a “rule‐out malignancy” test mostly for indeterminate thyroid lesions .…”

“…To select the set of miRNAs for classification, several screening stages were performed. In the first stage, 53 histological samples, 73 cell blocks of FNAC and 156 stained FNAC were profiled on Agilent custom‐designed miRNA microarrays containing over 2000 miRNA probes . Next, after a selection of 96 miRNAs that showed differential expression in benign and malignant tumors, a final set of 24 miRNAs was adopted.…”

“…Next, after a selection of 96 miRNAs that showed differential expression in benign and malignant tumors, a final set of 24 miRNAs was adopted. Lithwick‐Yanai et al highlighted the high level of reproducibility and concordance of yields (93% to 96%) in different laboratories and the advantage of using an existing diagnostic slide for molecular testing. Furthermore, the assay can be run on FNA slides with as little as 1% of thyroid cells present or from which only 5 ng of RNA is extracted.…”

Evaluation of Expression of 4 Mirnas in Cytological Samples as an Additional Method of Cervical Cancer Diagnosis

“…Очевид-но, практическое значение могла бы иметь разработка метода анализа микроРНК в материале цитологиче-ских мазков из шейки матки после их первичного ана-лиза, что позволило бы адаптировать этот метод к схе-мам традиционной морфологической диагностики. Этот подход активно внедряется для диагностики уз-ловых заболеваний щитовидной железы [14]. Недавно мы показали целесообразность его применения в слу-чае анализа материала тонкоигольной биопсии обра-зований молочной железы [15].…”

Utilization of direct smears of thyroid fine‐needle aspirates for ancillary molecular testing: A comparison of two proprietary testing platforms

“…Thyroid cancer is the most common endocrine malignancy, and its incidence has steadily increased over the past few decades. It may present as a nodule palpated on physical examination or as an incidental finding on imaging . As with any mass or nodule detected, there is a fundamental level of uncertainty regarding its diagnosis and prognosis.…”