AIMS: The 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. METHODS: A 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. RESULTS: Test 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%). CONCLUSIONS: A 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.

Cooper University Hospital Cooper Medical School of Rowan University at Camden New Jersey USA

Geha Mental Health Center Petach Tikva Israel

H Lee Moffitt Cancer Center and Research Institute Tampa Florida USA

Institute of Molecular and Translational Medicine Faculty of Medicine and Dentistry Palacky University Olomouc Czech Republic

National Centre of Clinical and Morphological Diagnostics St Petersburg Russia

Pathology Institute Meir Medical Center Kfar Saba Israel

Pathology Institute Rabin Medical Center Petach Tikva Israel

Pathology Institute Tel Aviv Sourasky Medical Center Tel Aviv Israel

Rosetta Genomics Inc Philadelphia Pennsylvania USA

Rosetta Genomics Ltd Rehovot Israel

Temple University Hospital Philadelphia Pennsylvania USA

The Johns Hopkins University School of Medicine Baltimore Maryland USA

The Sackler Faculty of Medicine Tel Aviv University Tel Aviv Israel

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Molecular markers in well-differentiated thyroid cancer