Assessing Bias and Limitations of Clinical Validation Studies of Molecular Diagnostic Tests for Indeterminate Thyroid Nodules: Systematic Review and Meta-Analysis

Background: Molecular tests for thyroid nodules with indeterminate fine needle aspiration results are increasingly used in clinical practice; however, true diagnostic summaries of these tests are unknown. A systematic review and meta-analysis were completed to (1) evaluate the accuracy of commercially available molecular tests for malignancy in indeterminate thyroid nodules and (2) quantify biases and limitations in studies that validate those tests.Summary: PubMed, EMBASE, and Web of Science were systematically searched through July 2021. English language articles that reported original clinical validation attempts of molecular tests for indeterminate thyroid nodules were included if they reported counts of true-negative, true-positive, false-negative, and false-positive results. We performed screening and full-text review, followed by assessment of eight common biases and limitations, extraction of diagnostic and histopathological information, and meta-analysis of clinical validity using a bivariate linear mixed-effects model. Forty-nine studies were included. Meta-analysis of Afirma Gene expression classifiers (GEC; n = 38 studies) revealed a sensitivity of 0.92 (confidence interval: 0.90-0.94), specificity of 0.26 (0.20-0.32), negative likelihood ratio (LR-) of 0.32 (0.23-0.44), positive LR+ of 1.24 (1.15-1.35), and area under the curve (AUC) of 0.83 (0.74-0.89). Afirma Genomic Sequencing Classifier (GSC; n = 10) had a sensitivity of 0.94 (0.89-0.96), specificity of 0.38 (0.27-0.50), LR- of 0.18 (0.10-0.30), LR+ of 1.52 (1.28-1.87), and AUC of 0.91 (0.62-0.92). ThyroSeq v1 and v2 (n = 10) had a sensitivity of 0.86 (0.82-0.90), specificity of 0.74 (0.59-0.85), LR- of 0.19 (0.13-0.26), LR+ of 3.52 (2.08-5.92), and AUC of 0.86 (0.81-0.90). ThyroSeq v3 (n = 6) had a sensitivity of 0.92 (0.86-0.95), specificity of 0.41 (0.18-0.69), LR- of 0.24 (0.09-0.62), LR+ of 1.67 (1.09-2.98), and AUC of 0.90 (0.63-0.92). Fourteen percent of studies conducted a blinded histopathologic review of excised thyroid nodules, and 8% made the decision to go to surgery blind to molecular test results.Conclusions: Meta-analyses reveal a high diagnostic accuracy of molecular tests for thyroid nodule assessment of malignancy risk; however, these studies are subject to several limitations. Limitations and their potential clinical impacts must be addressed and, when feasible, adjusted for using valid statistical methodologies.