A Thyroid Genetic Classifier Correctly Predicts Benign Nodules with Indeterminate Cytology: Two Independent, Multicenter, Prospective Validation Trials

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Background: Although most thyroid nodules with indeterminate cytology are benign, in most of the world surgery remains as the most frequent diagnostic management. We have previously reported a 10-gene thyroid genetic classifier which accurately predicts benign thyroid nodules. The assay is a prototype diagnostic kit suitable for reference laboratory testing and could potentially avoid unnecessary diagnostic surgery in patients with indeterminate thyroid cytology. Methods: Classifier performance was tested in two independent, ethnically diverse, prospective, multicenter trials (TGCT-1/Chile and TGCT-2/USA). A total of 4061 fine-needle aspirations were collected from 15 institutions of which 897 (22%) were called indeterminate. The clinical site was blind to the classifier score and the clinical laboratory blind to pathology report. A matched surgical pathology and valid classifier score was available for 270 samples. Results: Cohorts showed significant differences including; i) clinical site patient source (academic, 43% and 97% for TGCT-1 and 2, respectively), ii) ethnic diversity, with greater proportion of Hispanic (40% vs 3%) for TGCT-1 and greater proportion of African-American (11% vs 0%) and Asian (10% vs 1%) population for TGCT-2, and iii) tumor size, (mean of 1.7 cm and 2.5 cm for TGCT-1 and -2, respectively). Overall, there were no differences in the histopathological profile between cohorts. Forty-one of 155 and 45 of 115 nodules were malignant (cancer prevalence of 26% and 39% for TGCT 1 and 2, respectively). The classifier predicted 37 of 41 and 41 of 45 malignant nodules, yielding a sensitivity of 90% (95% confidence interval CI 77-97%) and 91% (95% CI 79-98%) for TGCTs 1 and 2, respectively. One hundred and one of 114 and 61 of 70 nodules were correctly predicted as benign yielding a specificity of 89% (95% CI 82-94%) and 87% (95% CI 77-94%), respectively. The negative predictive values for TGCT-1 and TGCT-2 were 96% and 94%, respectively, whereas the positive predictive values were 74% and 82%, respectively. The overall accuracy for both cohorts was 89%. Conclusions: Clinical validation of the classifier demonstrates equivalent performance in two independent and ethnically diverse cohorts, accurately predicting benign thyroid nodules that can undergo surveillance as an alternative to diagnostic surgery.

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... To solve this diagnostic problem of cytologically indeterminate thyroid nodules, molecular diagnostic tools have been utilized for routine thyroid nodule diagnostics primarily in the United States for presurgical risk stratification, with varying levels of success. ThyroSeq v3, Afirma GSC, ThyGeNEXT + ThyraMIR, and ThyroidPrint are each tests that have published multicenter validation studies showing high sensitivity and specificity, providing rule-in and rule-out molecular diagnosis for indeterminate thyroid FNAs (28,29,30,31,32). However, even these sophisticated molecular tests have drawbacks such as dependency on quality metrics of local diagnostic pathways, insufficient long-term follow-up data, limited or conflicting independent validation, high cost, and the need for additional dedicated FNA passes (33,34,35,36,37). ...
... A recent randomized controlled (albeit unblinded) trial found no significant difference in test performance between ThyroSeq v3 and Afirma GSC (44). ThyGeNEXT + ThyraMIR uses a mutation panel and a miRNA expression classifier, and ThyroidPrint uses an RT-qPCR mRNA classifier, both tests have reported similar high accuracy but lacked sufficient independent validation (30,31,32). ...
Objective: Genetic testing is increasingly used to diagnose or rule out thyroid cancer in indeterminate fine-needle aspirations. This review evaluates the usefulness of these methods with considerations of advantages and limitations. Design: Given the diagnostic problem associated with the increasing incidental detection of indeterminate thyroid nodules in the context of thyroid cancer overtreatment, we consider the conditions and respective necessary settings for the role of genetic testing to improve presurgical malignancy risk stratification. Methods: We review diagnostic pathway requirements and commercially available molecular tests with their respective advantages and disadvantages and discuss the prerequisites required for local application and implementation including quality assurance for local ultrasound and cytopathology practices. Results: Recent improvements in available molecular diagnostic tests have brought high sensitivity and specificity in initial validation studies, but whether these promising results translate to other clinical settings depends on the quality of the local thyroid nodule diagnostic pathway. Conclusions: Genetic testing can meaningfully improve presurgical malignancy risk assessment, but more work is needed to implement and use genetic testing effectively in local settings.
... On average, 15%-25% of the nodules with indeterminate cytology that are operated on are malignant. About 60% of them are PTC, of which up to 10% require totalization (28)(29)(30). Between 10%-15% are FTC; information on indication to totalization is scarce in this setting (27,28). In our series, of the 64 nodules with indeterminate cytology, 56 (87.5%) had a low risk of recurrence (minimally invasive FTC or angio-invasion <4 foci), so they did not require radioiodine ablation and, consequently, can be managed with lobectomy (18). ...
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Objective: Follicular thyroid carcinoma (FTC) is less frequent but has a worse prognosis than papillary carcinoma. The available evidence on pre-operative characteristics of FTC is controversial. Our objective was to characterize the clinical, ultrasound and histopathological presentation of FTC patients treated Chile. Subjects and methods: Retrospective analysis of 97 patients treated for FTC in 6 large centers in Chile. We analyzed their ultrasonographic features and classified the nodules according to ATA risk of malignancy and TI-RADS score, as well as the cytological findings according to the Bethesda system. We described their clinical and histopathological findings at diagnosis and classified their risk of recurrence and mortality according to ATA 2015 recurrence risk category and the eighth edition of the AJCC/UICC staging system, respectively. Results: Median age was 48 years and 73.2% were females. The median diameter was 38.8 mm; only 9.5% of them were microtumors. According to ATA risk of malignancy, 86% of the nodules were low or intermediate suspicious, while 78% were category 3 or 4A nodules according to the TI-RADS. Regarding the Bethesda system, 65.9% had indeterminate cytology (20.6% category III and 45.3% category IV). At histological examination, most were minimally-invasive and angio-invasive tumors with less than 4 foci (54.7% and 28.4% respectively). More than 90% of FTC were unifocal and there was no lymphovascular or extrathyroidal invasion or lymph node involvement. Four patients (4.1%) had distant metastases at diagnosis. Most patients (95%) had stage I or II disease according to the AJCC/UICC staging system, while the risk of recurrence was low at 51.5% when using the ATA risk of recurrence scale. Conclusion: At diagnosis, most FTCs were nodules of low or intermediate suspicion at ultrasound, nearly two thirds had indeterminate cytology according to the Bethesda system, and nearly 50% of them were of low risk of recurrence.
... The 2015 American Thyroid Association (ATA) guidelines [1] recommend surgical excision (usually a thyroid lobectomy) for Bethesda IV nodules, ☆ This article was written by members and invitees of the International Head and Neck Scientific Group ( although a total thyroidectomy could also be recommended depending on several characteristics, such as size, ultrasound pattern, bilateral disease, patient preference and genetic molecular testing [4]. ...
Thyroid nodules are a very common clinical condition. The 2015 American Thyroid Association (ATA) guidelines recommend surgical excision for Bethesda IV nodules. The use of intraoperative frozen section (FS) has been recommended as a strategy to tailor the extent of the initial surgery. We critically evaluated the literature that discusses the utility and cost-effectiveness of FS to make an intraoperative decision in patients with thyroid nodules classified as follicular neoplasm. FS should not be recommended as a routine intraoperative test to assess for malignancy in thyroid follicular patterned lesions due to its low performance; the high number of deferred results; the inability to adequately assess histologically defining features; the improvements in risk stratification guiding total thyroidectomy; and the low cost-effectiveness of FS.
... This type of testing takes a different approach from that of ThyroSeq v3 and Veracyte Afirma which require the original institution to submit the sample for molecular testing to centralized laboratories. Recent independent studies of international cohorts reported NPVs of 96% and 94%, which would be acceptable as "rule out" tests (36). However, additional independent studies for the 10gene panel (thyroid genetic classifier) with detailed performance characteristics and cost comparisons to the other molecular panels would be of interest. ...
The incidence of thyroid cancer is rising for a variety of reasons. At the same time, the nomenclature revision of non-invasive encapsulated follicular-variant PTC to noninvasive follicular neoplasm with papillary-like nuclear features (NIFTP) has modified the incidence of thyroid cancer. Given that thyroid neoplasia is a molecular event, it is important for the thyroid physician to evaluate each patient systematically. Most thyroid cancers are sporadic; however, some are familial and may be associated with syndromes with genetic implications. Advances in radiologic imaging have made ultrasonography a near equivalent of gross examination. The American College of Radiology Thyroid Imaging, Reporting and Data System (ACR TI-RADS) classifies nodules from TR1 to TR5 and is valuable in determining which patients should be guided toward fine-needle aspiration (FNA) sampling. While FNA procedures and processing may be varied, the key elements are cytologic diagnosis and collection of samples for potential molecular testing. The Bethesda System for Reporting Thyroid Cytology (BSRTC) is commonly used and categorizes each FNA specimen into one of six diagnoses. The indeterminate diagnoses with risk of malignancy (ROM) ranging from 10-75% comprise approximately 30% of thyroid FNA cases and for these, molecular testing is beneficial. In North America, the two most common molecular platforms are Veracyte Afirma GSC and ThyroSeq v3. Both panels cover an extensive array of genomic alterations associated with thyroid neoplasia and a negative result from either test effectively refines the ROM of an Atypia of Undetermined Significance/Follicular Lesion of Undetermined Significance (AUS/FLUS) or Follicular Neoplasm/Suspicious for a Follicular Neoplasm (FN/SFN) diagnosis to 3-4%. Given that the refined ROMs are comparable to that of a Benign BSRTC diagnosis, these patients are recommended for observation of their nodules. However, differences exist in the implication of Afirma GSC-Suspicious and ThyroSeq v3-Positive molecular results with regard to the probability of cancer. For either test, the molecular test result should be integrated with other clinical parameters to determine if surgery is indicated and, if so, the extent of surgery.
Objective: Advances in our understanding of the molecular biology of thyroid tumors is being rapidly translated into their clinical management. This review summarizes current use of molecular testing in thyroid tumors, focusing on their usefulness as diagnostic and prognostic tools to guide treatment with consideration of present limitations. Design: Considerations about molecular testing applications for the diagnosis and treatment of thyroid tumors are divided into four sections/roles: 1) evaluating cytologically indeterminate thyroid nodules; 2) guiding extent of surgery in indeterminate thyroid nodules; 3) completing histological characterization of thyroid tumors; and 4) identifying actionable mutations in advanced progressive thyroid cancers. Results: Genomic testing can improve the pre-surgical malignancy risk assessment in indeterminate thyroid nodules. However, a prior in-depth analysis of institutional quality and outcomes of sonographical, cytological and histological characterization of thyroid tumors is necessary. Presently, it remains uncertain whether knowing the molecular profile of a cytologically indeterminate thyroid nodule might be advantageous to modify the extent of initial surgery. Molecular characterization of thyroid tumors can be a valuable adjunct to morphological diagnosis in some challenging cases, such as in low-risk follicular cell-derived neoplasms, or rare tumors. Finally, as selective kinase inhibitors are available, molecular testing in locally advanced/metastatic progressive thyroid cancers should also be integrated into the institutional clinical management pathway to improve outcomes and limit toxicity. Conclusions: Molecular testing needs to be implemented into the local evidence based clinical management thyroid nodule/cancer pathways to improve its diagnostic and prognostic value and to optimize cost-effectiveness. This article is protected by copyright. All rights reserved.
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Importance Approximately 20% of fine-needle aspirations (FNA) of thyroid nodules have indeterminate cytology, most frequently Bethesda category III or IV. Diagnostic surgeries can be avoided for these patients if the nodules are reliably diagnosed as benign without surgery. Objective To determine the diagnostic accuracy of a multigene classifier (GC) test (ThyroSeq v3) for cytologically indeterminate thyroid nodules. Design, Setting, and Participants Prospective, blinded cohort study conducted at 10 medical centers, with 782 patients with 1013 nodules enrolled. Eligibility criteria were met in 256 patients with 286 nodules; central pathology review was performed on 274 nodules. Interventions A total of 286 FNA samples from thyroid nodules underwent molecular analysis using the multigene GC (ThyroSeq v3). Main Outcomes and Measures The primary outcome was diagnostic accuracy of the test for thyroid nodules with Bethesda III and IV cytology. The secondary outcome was prediction of cancer by specific genetic alterations in Bethesda III to V nodules. Results Of the 286 cytologically indeterminate nodules, 206 (72%) were benign, 69 (24%) malignant, and 11 (4%) noninvasive follicular thyroid neoplasms with papillary-like nuclei (NIFTP). A total of 257 (90%) nodules (154 Bethesda III, 93 Bethesda IV, and 10 Bethesda V) had informative GC analysis, with 61% classified as negative and 39% as positive. In Bethesda III and IV nodules combined, the test demonstrated a 94% (95% CI, 86%-98%) sensitivity and 82% (95% CI, 75%-87%) specificity. With a cancer/NIFTP prevalence of 28%, the negative predictive value (NPV) was 97% (95% CI, 93%-99%) and the positive predictive value (PPV) was 66% (95% CI, 56%-75%). The observed 3% false-negative rate was similar to that of benign cytology, and the missed cancers were all low-risk tumors. Among nodules testing positive, specific groups of genetic alterations had cancer probabilities varying from 59% to 100%. Conclusions and Relevance In this prospective, blinded, multicenter study, the multigene GC test demonstrated a high sensitivity/NPV and reasonably high specificity/PPV, which may obviate diagnostic surgery in up to 61% of patients with Bethesda III to IV indeterminate nodules, and up to 82% of all benign nodules with indeterminate cytology. Information on specific genetic alterations obtained from FNA may help inform individualized treatment of patients with a positive test result.
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Thyroid cancer is the most frequent endocrine malignancy, and its incidence is increasing. A current limitation of cytological evaluation of thyroid nodules is that 20-25% are reported as indeterminate. Therefore, an important challenge for clinicians is to determine whether an indeterminate nodule is malignant, and should undergo surgery, or benign, and should be recommended to follow-up. The emergence of precision medicine has offered a valuable solution for this problem, with four tests currently available for the molecular diagnosis of indeterminate nodules. However, efforts to critically analyze the quality of the accumulated evidence are scarce. This review and meta-analysis is aimed to contribute better knowledge about the four available molecular tests, their technical characteristics, clinical performance, and ultimately to help clinician to make better decisions to provide the best care options as possible. For this purpose, we address three critical topics: i) the proper theoretical accuracy, considering the intended clinical use of the test (rule-in versus rule-out) and the impact on clinical decisions; ii) the quality of the evidence reported for each test; iii) and how accurate and effective have the tests proved to be after their clinical use. Together with the upcoming evidence, this work provides significant and useful information for health-care system decision-makers to consider the use of molecular testing as a public health need, avoiding unnecessary surgical risks and costs.
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Background: In most of the world, diagnostic surgery remains as the most frequent approach for indeterminate thyroid cytology. Although several molecular tests are available for central-lab testing in the US, there are no available kits for local laboratory testing. The aim of this study was to develop a prototype in-vitro diagnostic (IVD) gene classifier for diagnosis of indeterminate thyroid cytology. Methods: In a first stage, the expression of 18 genes was determined by qPCR in a broad histopathological spectrum of fresh tissue biopsies (114). Expression data was used to train several classifiers by supervised machine learning approaches. Classifiers were tested in an independent set of 139 samples. In a second stage, the best classifier was chosen as a model to develop a multiplexed-qPCR IVD prototype assay, which was tested in a prospective multicenter cohort of fine needle aspiration (FNA) biopsies. Results: In tissue biopsies, the best classifier, using only 10 genes, reached an optimal and consistent performance in the 9-fold cross-validated testing set (sensitivity 93% and specificity 81%). In the multicenter cohort of FNA samples, the 10-gene signature, built into a multiplexed-qPCR IVD prototype showed an area under the curve of 0.96, a positive predictive value (PPV) of 87% and a negative predictive value (NPV) of 98%. By Bayes theorem, the IVD prototype is expected to achieve a PPV of 78%-90% and a NPV of 96%-99% in patients with a cancer prevalence range of 20%-40%. Conclusions: We report a new multiplexed-qPCR IVD prototype that accurately classifies thyroid nodules and may provide a future solution suitable for expanded local reference laboratory testing.
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Importance: Although growing evidence points to highly indolent behavior of encapsulated follicular variant of papillary thyroid carcinoma (EFVPTC), most patients with EFVPTC are treated as having conventional thyroid cancer. Objective: To evaluate clinical outcomes, refine diagnostic criteria, and develop a nomenclature that appropriately reflects the biological and clinical characteristics of EFVPTC. Design, setting, and participants: International, multidisciplinary, retrospective study of patients with thyroid nodules diagnosed as EFVPTC, including 109 patients with noninvasive EFVPTC observed for 10 to 26 years and 101 patients with invasive EFVPTC observed for 1 to 18 years. Review of digitized histologic slides collected at 13 sites in 5 countries by 24 thyroid pathologists from 7 countries. A series of teleconferences and a face-to-face conference were used to establish consensus diagnostic criteria and develop new nomenclature. Main outcomes and measures: Frequency of adverse outcomes, including death from disease, distant or locoregional metastases, and structural or biochemical recurrence, in patients with noninvasive and invasive EFVPTC diagnosed on the basis of a set of reproducible histopathologic criteria. Results: Consensus diagnostic criteria for EFVPTC were developed by 24 thyroid pathologists. All of the 109 patients with noninvasive EFVPTC (67 treated with only lobectomy, none received radioactive iodine ablation) were alive with no evidence of disease at final follow-up (median [range], 13 [10-26] years). An adverse event was seen in 12 of 101 (12%) of the cases of invasive EFVPTC, including 5 patients developing distant metastases, 2 of whom died of disease. Based on the outcome information for noninvasive EFVPTC, the name "noninvasive follicular thyroid neoplasm with papillary-like nuclear features" (NIFTP) was adopted. A simplified diagnostic nuclear scoring scheme was developed and validated, yielding a sensitivity of 98.6% (95% CI, 96.3%-99.4%), specificity of 90.1% (95% CI, 86.0%-93.1%), and overall classification accuracy of 94.3% (95% CI, 92.1%-96.0%) for NIFTP. Conclusions and relevance: Thyroid tumors currently diagnosed as noninvasive EFVPTC have a very low risk of adverse outcome and should be termed NIFTP. This reclassification will affect a large population of patients worldwide and result in a significant reduction in psychological and clinical consequences associated with the diagnosis of cancer.
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Molecular testing for oncogenic mutations or gene expression in fine-needle aspirations (FNAs) from thyroid nodules with indeterminate cytology identifies a subset of benign or malignant lesions with high predictive value. Evaluate a novel diagnostic algorithm combining mutation detection and miRNA expression to improve the diagnostic yield of molecular cytology. Surgical specimens and preoperative FNAs (n=638) were tested for 17 validated gene alterations using the miRInform Thyroid test and with a 10-miRNA gene expression classifier generating positive (malignant) or negative (benign) results. Cross-sectional sampling of thyroid nodules with AUS/FLUS or FN/SFN cytology (n=109) was conducted at 12 endocrinology centers across the United States. Qualitative molecular results were compared to surgical histopathology to determine diagnostic performance and model clinical impact. Mutations were detected in 69% of nodules with malignant outcome. Among mutation-negative specimens, miRNA testing correctly identified 64% of malignant cases and 98% of benign cases. The diagnostic sensitivity and specificity of the combined algorithm was 89% (95% confidence intervals (CI): 73-97%) and 85% (95% CI: 75-92%), respectively. At 32% cancer prevalence, 61% of the molecular results were benign with a negative predictive value of 94% (95% CI: 85-98%). Independently of variations in cancer prevalence, the test increased the yield of true benign results by 65% relative to mRNA-based gene expression classification and decreased the rate of avoidable diagnostic surgeries by 69%. Multi-platform testing for DNA, mRNA and miRNA can accurately classify benign and malignant thyroid nodules, increase the diagnostic yield of molecular cytology, and further improve the preoperative risk-based management of benign nodules with AUS/FLUS or FN/SFN cytology.
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Background EndoPredict (EP) is a clinically validated multianalyte gene expression test to predict distant metastasis in ER-positive, HER2-negative breast cancer treated with endocrine therapy alone. The test is based on the combined analysis of 12 genes in formalin-fixed, paraffin-embedded (FFPE) tissue by reverse transcription-quantitative real-time PCR (RT-qPCR). Recently, it was shown that EP is feasible for reliable decentralized assessment of gene expression. The aim of this study was the analytical validation of the performance characteristics of the assay and its verification in a molecular-pathological routine laboratory. Methods Gene expression values to calculate the EP score were assayed by one-step RT-qPCR using RNA from FFPE tumor tissue. Limit of blank, limit of detection, linear range, and PCR efficiency were assessed for each of the 12 PCR assays using serial samples dilutions. Different breast cancer samples were used to evaluate RNA input range, precision and inter-laboratory variability. Results PCR assays were linear up to Cq values between 35.1 and 37.2. Amplification efficiencies ranged from 75% to 101%. The RNA input range without considerable change of the EP score was between 0.16 and 18.5 ng/μl. Analysis of precision (variation of day, day time, instrument, operator, reagent lots) resulted in a total noise (standard deviation) of 0.16 EP score units on a scale from 0 to 15. The major part of the total noise (SD 0.14) was caused by the replicate-to-replicate noise of the PCR assays (repeatability) and was not associated with different operating conditions (reproducibility). Performance characteristics established in the manufacturer’s laboratory were verified in a routine molecular pathology laboratory. Comparison of 10 tumor samples analyzed in two different laboratories showed a Pearson coefficient of 0.995 and a mean deviation of 0.15 score units. Conclusions The EP test showed reproducible performance characteristics with good precision and negligible laboratory-to-laboratory variation. This study provides further evidence that the EP test is suitable for decentralized testing in specialized molecular pathological laboratories instead of a reference laboratory. This is a unique feature and a technical advance in comparison with existing RNA-based prognostic multigene expression tests.
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Gene expression profiles provide important information about the biology of breast tumors and can be used to develop prognostic tests. However, the implementation of quantitative RNA-based testing in routine molecular pathology has not been accomplished, so far. The EndoPredict assay has recently been described as a quantitative RT-PCR-based multigene expression test to identify a subgroup of hormone-receptor-positive tumors that have an excellent prognosis with endocrine therapy only. To transfer this test from bench to bedside, it is essential to evaluate the test-performance in a multicenter setting in different molecular pathology laboratories. In this study, we have evaluated the EndoPredict (EP) assay in seven different molecular pathology laboratories in Germany, Austria, and Switzerland. A set of ten formalin-fixed paraffin-embedded tumors was tested in the different labs, and the variance and accuracy of the EndoPredict assays were determined using predefined reference values. Extraction of a sufficient amount of RNA and generation of a valid EP score was possible for all 70 study samples (100%). The EP scores measured by the individual participants showed an excellent correlation with the reference values, respectively, as reflected by Pearson correlation coefficients ranging from 0.987 to 0.999. The Pearson correlation coefficient of all values compared to the reference value was 0.994. All laboratories determined EP scores for all samples differing not more than 1.0 score units from the pre-defined references. All samples were assigned to the correct EP risk group, resulting in a sensitivity and specificity of 100%, a concordance of 100%, and a kappa of 1.0. Taken together, the EndoPredict test could be successfully implemented in all seven participating laboratories and is feasible for reliable decentralized assessment of gene expression in luminal breast cancer.
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Thyroid fine needle aspiration (FNA) is a safe, cost effective, and relatively accurate method for guiding the initial management of a thyroid nodule. The popularity of thyroid FNA is reflected in the fact that over 350,000 thyroid FNAs are performed each year in the USA. As we move into the next decade, several issues pertaining to thyroid FNA are being addressed including: how to better apply thyroid FNA as a differential test for follicular-patterned thyroid tumors, how to manage the atypical thyroid FNA, and how to use thyroid FNA in the evaluation of poorly differentiated thyroid carcinomas.
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The overall interobserver reproducibility of thyroid fine-needle aspiration (FNA) has not been comprehensively assessed. A blinded 6-rater interobserver reproducibility study was conducted of 200 thyroid FNA cases using the UK System, which is similar to The Bethesda System for Reporting Thyroid Cytology: Thy1, nondiagnostic; Thy2, nonneoplastic; Thy3a, atypia, probably benign; Thy3f, follicular lesion; Thy4, suspicious of malignancy; and Thy5, malignant. There was good interobserver agreement for the Thy1 (κ = 0.69) and Thy5 (κ = 0.61), moderate agreement for Thy2 (κ = 0.55) and Thy3f (κ = 0.51), and poor agreement for Thy3a (κ = 0.11) and Thy4 (κ = 0.17) categories. Combining categories implying surgical management (Thy3f, Thy4, and Thy5) achieved good agreement (κ = 0.72), as did combining categories implying medical management (Thy1, Thy2, and Thy3a; κ = 0.72). The UK thyroid FNA terminology is a reproducible and clinically relevant system for thyroid FNA reporting. This study demonstrates that international efforts to harmonize and refine thyroid cytology classification systems can improve consistency in the clinical management of thyroid nodules.
Importance Use of next-generation sequencing of RNA and machine learning algorithms can classify the risk of malignancy in cytologically indeterminate thyroid nodules to limit unnecessary diagnostic surgery. Objective To measure the performance of a genomic sequencing classifier for cytologically indeterminate thyroid nodules. Design, Setting, and Participants A blinded validation study was conducted on a set of cytologically indeterminate thyroid nodules collected by fine-needle aspiration biopsy between June 2009 and December 2010 from 49 academic and community centers in the United States. All patients underwent surgery without genomic information and were assigned a histopathology diagnosis by an expert panel blinded to all genomic information. There were 210 potentially eligible thyroid biopsy samples with Bethesda III or IV indeterminate cytopathology that constituted a cohort previously used to validate the gene expression classifier. Of these, 191 samples (91.0%) had adequate residual RNA for validation of the genomic sequencing classifier. Algorithm development and independent validation occurred between August 2016 and May 2017. Exposures Thyroid nodule surgical histopathology diagnosis by an expert panel blinded to all genomic data. Main Outcomes and Measures The primary end point was measurement of genomic sequencing classifier sensitivity, specificity, and negative and positive predictive values in biopsies from Bethesda III and IV nodules. The secondary end point was measurement of classifier performance in biopsies from Bethesda II, V, and VI nodules. Results Of the 183 included patients, 142 (77.6%) were women, and the mean (range) age was 51.7 (22.0-85.0) years. The genomic sequencing classifier had a sensitivity of 91% (95% CI, 79-98) and a specificity of 68% (95% CI, 60-76). At 24% cancer prevalence, the negative predictive value was 96% (95% CI, 90-99) and the positive predictive value was 47% (95% CI, 36-58). Conclusions and Relevance The genomic sequencing classifier demonstrates high sensitivity and accuracy for identifying benign nodules. Its 36% increase in specificity compared with the gene expression classifier potentially increases the number of patients with benign nodules who can safely avoid unnecessary diagnostic surgery.
Thyroid nodules affect nearly two-thirds of the world population. Fine-needle biopsy with cytologic evaluation remains the diagnostic test of choice to distinguish benign from malignant thyroid nodules yet fails to discriminate as benign or malignant in up to one-third of cases. This review discusses the limitation of current cytopathologic evaluation, the development of thyroid molecular testing, and the strengths and limitations of commercially available tests. Initial cytomolecular testing sought to identify specific gene mutations associated with thyroid cancer. Although the presence of a mutation was strongly associated with cancer, the likelihood of identifying a mutation was low; therefore, the test had low sensitivity. Subsequent tests developed have sought to improve the accuracy of cytomolecular testing for thyroid fine-needle aspirations, both to reassure patients and providers when malignancy may be absent and to confirm the malignancy when present. The development of cytomolecular testing for thyroid nodules has informed and improved current understanding of thyroid nodule formation and progression. When used appropriately and with clear understanding of the advantages and disadvantages, cytomolecular testing has the potential to improve patient care in the setting of indeterminate thyroid nodules by helping to guide both the need for and the extent of thyroid surgery. Cancer 2017.
Background: Thyroid nodules are a common clinical problem, and differentiated thyroid cancer is becoming increasingly prevalent. Since the American Thyroid Association's (ATA's) guidelines for the management of these disorders were revised in 2009, significant scientific advances have occurred in the field. The aim of these guidelines is to inform clinicians, patients, researchers, and health policy makers on published evidence relating to the diagnosis and management of thyroid nodules and differentiated thyroid cancer. Methods: The specific clinical questions addressed in these guidelines were based on prior versions of the guidelines, stakeholder input, and input of task force members. Task force panel members were educated on knowledge synthesis methods, including electronic database searching, review and selection of relevant citations, and critical appraisal of selected studies. Published English language articles on adults were eligible for inclusion. The American College of Physicians Guideline Grading System was used for critical appraisal of evidence and grading strength of recommendations for therapeutic interventions. We developed a similarly formatted system to appraise the quality of such studies and resultant recommendations. The guideline panel had complete editorial independence from the ATA. Competing interests of guideline task force members were regularly updated, managed, and communicated to the ATA and task force members. Results: The revised guidelines for the management of thyroid nodules include recommendations regarding initial evaluation, clinical and ultrasound criteria for fine-needle aspiration biopsy, interpretation of fine-needle aspiration biopsy results, use of molecular markers, and management of benign thyroid nodules. Recommendations regarding the initial management of thyroid cancer include those relating to screening for thyroid cancer, staging and risk assessment, surgical management, radioiodine remnant ablation and therapy, and thyrotropin suppression therapy using levothyroxine. Recommendations related to long-term management of differentiated thyroid cancer include those related to surveillance for recurrent disease using imaging and serum thyroglobulin, thyroid hormone therapy, management of recurrent and metastatic disease, consideration for clinical trials and targeted therapy, as well as directions for future research. Conclusions: We have developed evidence-based recommendations to inform clinical decision-making in the management of thyroid nodules and differentiated thyroid cancer. They represent, in our opinion, contemporary optimal care for patients with these disorders.
Background: Fine-needle aspiration (FNA) cytology is a common approach to evaluating thyroid nodules, although 20% to 30% of FNAs have indeterminate cytology, which hampers the appropriate management of these patients. Follicular (or oncocytic) neoplasm/suspicious for a follicular (or oncocytic) neoplasm (FN/SFN) is a common indeterminate diagnosis with a cancer risk of approximately 15% to 30%. In this study, the authors tested whether the most complete next-generation sequencing (NGS) panel of genetic markers could significantly improve cancer diagnosis in these nodules. Methods: The evaluation of 143 consecutive FNA samples with a cytologic diagnosis of FN/SFN from patients with known surgical outcomes included 91 retrospective samples and 52 prospective samples. Analyses were performed on a proprietary sequencer using the targeted ThyroSeq v2 NGS panel, which simultaneously tests for point mutations in 13 genes and for 42 types of gene fusions that occur in thyroid cancer. The expression of 8 genes was used to assess the cellular composition of FNA samples. Results: In the entire cohort, histologic analysis revealed 104 benign nodules and 39 malignant nodules. The most common point mutations involved the neuroblastoma RAS viral oncogene homolog (NRAS), followed by the Kirsten rat sarcoma viral oncogene homolog (KRAS), the telomerase reverse transcriptase (TERT) gene, and the thyroid-stimulating hormone receptor (TSHR) gene. The identified fusions involved the thyroid adenoma associated (THADA) gene; the peroxisome proliferator-activated receptor γ (PPARG) gene; and the neurotrophic tyrosine kinase, receptor, type 3 (NTRK3) gene. Performance characteristics were similar in the retrospective and prospective groups. Among all FN/SFN nodules, preoperative ThyroSeq v2 performed with 90% sensitivity (95% confidence interval [CI], 80%-99%), 93% specificity (95% CI, 88%-98%), a positive predictive value of 83% (95% CI, 72%-95%), a negative predictive value of 96% (95% CI, 92%-100%), and 92% accuracy (95% CI, 88%-97%). Conclusions: The current results indicate that comprehensive genotyping of thyroid nodules using a broad NGS panel provides a highly accurate diagnosis for nodules with FN/SFN cytology and should facilitate the optimal management of these patients.
To provide population-based estimates of trends in thyroid nodule fine-needle aspirations (FNA) and operative volumes, we used multiple claims databases to quantify rates of these procedures and their association with the increasing incidence of thyroid cancer in the United States. Private and public insurance claims databases were used to estimate procedure volumes from 2006 to 2011. Rates of FNA and thyroid operations related to thyroid nodules were defined by CPT4 codes associated with International Classification of Diseases, Ninth Revision Clinical Modification codes for nontoxic uni- or multinodular goiter and thyroid neoplasms. Use of thyroid FNA more than doubled during the 5-year study period (16% annual growth). The number of thyroid operations performed for thyroid nodules increased by 31%. Total thyroidectomies increased by 12% per year, whereas lobectomies increased only 1% per year. In 2011, total thyroidectomies accounted for more than half (56%) of the operations for thyroid neoplasms in the United States. Thyroid operations became increasingly (62%) outpatient procedures. Thyroid FNA and operative procedures have increased rapidly in the United States, with an associated increase in the incidence of thyroid cancer. The more substantial increase in number of total versus partial thyroid resections suggests that patients undergoing thyroid operation are perceived to have a greater risk of cancer as determined by preoperative assessments, but this trend could also increase detection of incidental microcarcinomas.
Chinese translation Clinical management of thyroid neoplasms is based on light microscopic diagnosis, but its accuracy and precision are poorly defined. To assess inter- and intraobserver variability of preoperative cytopathologic and postoperative histopathologic thyroid diagnoses. Samples were collected in a prospective, multicenter trial validating a gene expression classifier between June 2009 and December 2010. 14 academic and 35 community clinical sites. 653 patients with 776 surgically resected thyroid nodules of 1 cm or greater. Intraobserver concordance among 2 or more central histopathologists who independently read histopathology slides was calculated. Interobserver concordance between the diagnoses made by the central histopathologists and those made by local pathologists were calculated. Intra- and interobserver concordance for cytopathology was similarly calculated by comparing diagnoses made by local pathologists with those made by a central panel of 3 cytopathologists. Concordance on the histopathologic distinction between benign and malignant diagnoses was 91% comparing local with central histopathologists and 90% comparing 2 central histopathologists. Using the 6-category Bethesda System, 64.0% of diagnoses made by local and central cytopathologists and 74.7% of intraobserver diagnoses were concordant. Central cytopathologists made fewer indeterminate diagnoses than local pathologists (41.2% vs. 55.0%). Many local pathologists did not use the Bethesda System, so their reports were translated to allow comparison. The study required histopathology, and the study population and specimens did not encompass all newly evaluated patients with a thyroid nodule. Substantial inter- and intraobserver variability exists in the cytopathologic and histopathologic evaluation of thyroid nodules, confirming an inherent limitation of visual microscopic diagnosis. Veracyte.
Objectives Next-generation sequencing (NGS) allows for high-throughput sequencing analysis of large regions of the human genome. We explored the use of targeted NGS for simultaneous testing for multiple mutations in thyroid cancer. Design A custom panel (ThyroSeq) was designed to target 12 cancer genes with 284 mutational hot spots. Sequencing was performed to analyze DNA from 228 thyroid neoplastic and nonneoplastic samples including 105 frozen, 72 formalin-fixed, and 51 fine-needle aspiration samples representing all major types of thyroid cancer. Results Only 5–10 ng of input DNA was sufficient for successful analysis of 99.6% of samples. The analytical accuracy for mutation detection was 100% with the sensitivity of 3%–5% of mutant allele. ThyroSeq DNA assay identified mutations in 19 of 27 of classic papillary thyroid carcinomas (PTCs) (70%), 25 of 30 follicular variant PTCs (83%), 14 of 18 conventional (78%) and 7 of 18 oncocytic follicular carcinomas (39%), 3 of 10 poorly differentiated carcinomas (30%), 20 of 27 anaplastic (ATCs) (74%), and 11 of 15 medullary thyroid carcinomas (73%). In contrast, 5 of 83 benign nodules (6%) were positive for mutations. Most tumors had a single mutation, whereas several ATCs and PTCs demonstrated two or three mutations. The most common mutations detected were BRAF and RAS followed by PIK3CA, TP53, TSHR, PTEN, GNAS, CTNNB1, and RET. The BRAF mutant allele frequency was 18%–48% in PTCs and was lower in ATCs. Conclusions The ThyroSeq NGS panel allows simultaneous testing for multiple mutations with high accuracy and sensitivity, requires a small amount of DNA and can be performed in a variety of thyroid tissue and fine-needle aspiration samples, and provides quantitative assessment of mutant alleles. Using this approach, the point mutations were detected in 30%–83% of specific types of thyroid cancer and in only 6% of benign thyroid nodules and were shown to be present in the majority of cells within the cancer nodule.
To address terminology and other issues related to thyroid fine-needle aspiration (FNA), the National Cancer Institute (NCI) hosted The NCI Thyroid FNA State of the Science Conference. The conclusions regarding terminology and morphologic criteria from the NCI meeting led to the Bethesda Thyroid Atlas Project and form the framework for the Bethesda System for Reporting Thyroid Cytopathology. Participants of the Atlas Project were selected from among the committee members of the NCI FNA State of the Science Conference and other participants at the live conference. The terminology framework was based on a literature search of English language publications dating back to 1995 using PubMed as the search engine; online forum discussions ( ); and formal interdisciplinary discussions held on October 22 and 23, 2007, in Bethesda, MD. For clarity of communication, the Bethesda System for Reporting Thyroid Cytopathology recommends that each report begin with one of the six general diagnostic categories. Each of the categories has an implied cancer risk that links it to an appropriate clinical management guideline. The project participants hope that the adoption of this framework will facilitate communication among cytopathologists, endocrinologists, surgeons, and radiologists; facilitate cytologic-histologic correlation for thyroid diseases; facilitate research into the understanding of thyroid diseases; and allow easy and reliable sharing of data from different laboratories for national and international collaborative studies.
Thyroid nodules are common and are commonly benign. The reported prevalence of nodular thyroid disease depends on the population studied and the methods used to detect nodules. Nodule incidence increases with age, and is increased in women, in people with iodine deficiency, and after radiation exposure. Numerous studies suggest a prevalence of 2-6% with palpation, 19-35% with ultrasound, and 8-65% in autopsy data. With widespread use of sensitive imaging in clinical practice, incidental thyroid nodules are being discovered with increasing frequency. Ultrasonography is the most accurate and cost-effective method for evaluating and observing thyroid nodules. Current ultrasonography machines are relatively inexpensive, sensitive, and easy to operate. Most endocrinologists are now using ultrasound examination in the initial evaluation of a patient with known or suspected thyroid nodule. The management of thyroid incidentalomas is a matter of controversy.
To study the degree of interobserver variability in the interpretation of fine needle aspiration (FNA) biopsies of the thyroid, specifically in the categorization of follicular lesions (FLs), and to examine the accuracy of FNA diagnosis of FLs with surgical follow-up. Fifty cases were chosen with surgical follow-up and a cytologic diagnosis of either FL (21) or follicular neoplasms (29). Representative slides were selected for each case and circulated to 4 pathologists for review. Interobserver variability was assessed using pairwise K statistics. Accuracy of the cytologic diagnoses in predicting a nonneoplastic or neoplastic outcome was determined by measuring sensitivity and specificity. Likelihood ratios and receiver operator characteristic curves were calculated for each reviewer. Interobserver agreement between the 4 pathologists was fair to substantial (K scores, 0.199-0.617). The accuracy of the 4 pathologists' cytologic diagnoses in predicting the surgical outcome was 77-90% for follicular neoplasms and 53-74% for nonneoplastic diagnoses. FLs present diagnostic difficulties as to cytologic categorization. A wide range of interobserver agreement was found in this study of 4 pathologists from the same institution. Some pathologists make greater use of intermediate categories, such as FL, favor nonneoplastic, or FL, favor neoplastic, whereas others show more definitive categorization into benign and neoplastic groups.
Understanding patterns of gene-expression variation within and among human populations will provide important insights into the molecular basis of phenotypic diversity and the interpretation of patterns of expression variation in disease. However, little is known about how gene-expression variation is apportioned within and among human populations. Here, we characterize patterns of natural gene-expression variation in 16 individuals of European and African ancestry. We find extensive variation in gene-expression levels and estimate that approximately 83% of genes are differentially expressed among individuals and that approximately 17% of genes are differentially expressed among populations. By decomposing total gene-expression variation into within- versus among-population components, we find that most expression variation is due to variation among individuals rather than among populations, which parallels observations of extant patterns of human genetic variation. Finally, we performed allele-specific quantitative polymerase chain reaction to demonstrate that cis-regulatory variation in the lymphocyte adaptor protein (SH2B adapter protein 3) contributes to differential expression between European and African samples. These results provide the first insight into how human population structure manifests itself in gene-expression levels and will help guide the search for regulatory quantitative trait loci.
Performance of a genomic sequencing classifier for the preoperative diagnosis of cytologically surgery indeterminate thyroid nodules
  • K N Patel
  • T E Angell
  • J Babiarz
  • N M Barth
  • T Blevins
  • Q-Y Duh
  • R A Ghossein
  • R M Harrell
  • J Huang
  • G C Kennedy
  • S Y Kim
  • R T Kloos
  • V A Livolsi
  • G W Randolph
  • P M Sadow
  • M H Shanik
  • J A Sosa
  • S T Taweek
  • P S Walsh
  • D Whitney
  • M W Yeh
  • P W Ladenson
Patel KN, Angell TE, Babiarz J, Barth NM, Blevins T, Duh Q-Y, Ghossein RA,Harrell RM, Huang J, Kennedy GC,Kim SY,Kloos RT, LiVolsi VA,Randolph GW, Sadow PM,Shanik MH, Sosa JA, Taweek ST,Walsh PS,Whitney D, Yeh MW,Ladenson PW 2018 Performance of a genomic sequencing classifier for the preoperative diagnosis of cytologically surgery indeterminate thyroid nodules. JAMA Surg 153:817-824.
Nomenclature revision for encapsulated follicular variant of papillary thyroid carcinoma: a paradigm shift to reduce overtreatment of indolent tumors
  • Y E Nikiforov
  • R R Seethala
  • G Tallini
  • Z W Baloch
  • F Basolo
  • L D Thompson
  • J A Barletta
  • B M Wenig
  • Al Ghuzlan
  • A Kakudo
  • K Giordano
  • T J Alves
  • V A Khanafshar
  • E Asa
  • S L El-Naggar
  • A K Gooding
  • W E Hodak
  • S P Lloyd
  • R V Maytal
  • G Mete
  • O Nikiforova
  • M N Nose
  • V Papotti
  • M Poller
  • D N Sadow
  • P M Tischler
  • A S Tuttle
  • R M Wall
  • K B Livolsi
  • V A Randolph
  • G W Ghossein
Nikiforov YE, Seethala RR, Tallini G, Baloch ZW, Basolo F, Thompson LD, Barletta JA, Wenig BM, Al Ghuzlan A, Kakudo K, Giordano TJ, Alves VA, Khanafshar E, Asa SL, El-Naggar AK, Gooding WE, Hodak SP, Lloyd RV, Maytal G, Mete O, Nikiforova MN, Nose V, Papotti M, Poller DN, Sadow PM, Tischler AS, Tuttle RM, Wall KB, LiVolsi VA, Randolph GW, Ghossein RA 2016 Nomenclature revision for encapsulated follicular variant of papillary thyroid carcinoma: a paradigm shift to reduce overtreatment of indolent tumors. JAMA Oncol2:1023-1029.