Preoperative Evaluation of Thyroid Cancer: A Review of Current Best Practices.

OBJECTIVE: The incidence of thyroid cancer has significantly increased in recent decades. Although most thyroid cancers are small and carry an excellent prognosis, a subset of patients present with advanced thyroid cancer, which is associated with increased rates of morbidity and mortality. The management of thyroid cancer requires a thoughtful individualized approach to optimize oncologic outcomes and minimize morbidity associated with treatment. Because endocrinologists usually play a key role in the initial diagnosis and evaluation of thyroid cancers, a thorough understanding of the critical components of the preoperative evaluation facilitates the development of a timely and comprehensive management plan. The following review outlines considerations in the preoperative evaluation of patients with thyroid cancer. METHODS: A clinical review based on current literature was generated by a multidisciplinary author panel. RESULTS: A review of considerations in the preoperative evaluation of thyroid cancer is provided. The topic areas include initial clinical evaluation, imaging modalities, cytologic evaluation, and the evolving role of mutational testing. Special considerations in the management of advanced thyroid cancer are discussed. CONCLUSION: Thorough and thoughtful preoperative evaluation is critical for formulating an appropriate treatment strategy in the management of thyroid cancer.

Thyroid Carcinoma, Version 2.2022, NCCN Clinical Practice Guidelines in Oncology.

Differentiated thyroid carcinomas is associated with an excellent prognosis. The treatment of choice for differentiated thyroid carcinoma is surgery, followed by radioactive iodine ablation (iodine-131) in select patients and thyroxine therapy in most patients. Surgery is also the main treatment for medullary thyroid carcinoma, and kinase inhibitors may be appropriate for select patients with recurrent or persistent disease that is not resectable. Anaplastic thyroid carcinoma is almost uniformly lethal, and iodine-131 imaging and radioactive iodine cannot be used. When systemic therapy is indicated, targeted therapy options are preferred. This article describes NCCN recommendations regarding management of medullary thyroid carcinoma and anaplastic thyroid carcinoma, and surgical management of differentiated thyroid carcinoma (papillary, follicular, Hürthle cell carcinoma).

NCCN Guidelines Insights: Thyroid Carcinoma, Version 2.2018.

The NCCN Guidelines for Thyroid Carcinoma provide recommendations for the management of different types of thyroid carcinoma, including papillary, follicular, Hürthle cell, medullary, and anaplastic carcinomas. These NCCN Guidelines Insights summarize the panel discussion behind recent updates to the guidelines, including the expanding role of molecular testing for differentiated thyroid carcinoma, implications of the new pathologic diagnosis of noninvasive follicular thyroid neoplasm with papillary-like nuclear features, and the addition of a new targeted therapy option for BRAF V600E-mutated anaplastic thyroid carcinoma.

Evaluation and Management of Indeterminate Thyroid Nodules: The Revolution of Risk Stratification Beyond Cytological Diagnosis.

In accordance with National Guidelines, we currently follow a linear approach to the diagnosis of thyroid nodules, with management decision based primarily on a cytological diagnosis following fine-needle aspiration biopsy. However, 25% of these biopsies render an indeterminate cytology, leaving uncertainty regarding appropriate management. Individualizing the risk of malignancy of these nodules could improve their management significantly. We summarize the current evidence on the relevance of clinical information, radiological features, cytological features, and molecular markers tests results and describe how these can be integrated to personalize the management of thyroid nodules with indeterminate cytology. Several factors can be used to stratify the risk of malignancy in thyroid nodules with indeterminate cytology. Male gender, large tumors (>4 cm), suspicious sonographic patterns, and the presence of nuclear atypia on the cytology are all associated with an increased cancer prevalence. The added value of current molecular markers in the risk stratification process needs further study because their performance seems compromised in some clinical settings and remains to be validated in others. Risk stratification is possible in thyroid nodules with indeterminate cytology using data that are often underused by current guidelines. Future guidelines should integrate these factors and personalize the recommended diagnostic and therapeutic approaches accordingly.

Molecular Assays in Cytopathology for Thyroid Cancer.

BACKGROUND: Despite lack of adequate, validated, independently performed clinical studies, several molecular tests are commercially available on the market and are being used on indeterminate thyroid nodules to guide patient-care decisions. METHODS: We summarize the current evidence on the role and limitations of molecular tests used in combination with thyroid cytopathology to refine the presurgical diagnosis of thyroid nodules. RESULTS: The clinical performance of molecular tests depends on the pretest risk of malignancy within the specific cytological group being assessed. This risk is variable and should be assessed at each institution to optimize the selection of the molecular test and the interpretation of its results. Next-generation sequencing has increased the sensitivity of oncogene panels while maintaining high specificity. Tests assessing the gene expression pattern have shown promising results, with high sensitivity but low specificity. The impacts of molecular markers on clinical practice remains in flux and their effect on health care costs remains poorly understood. CONCLUSIONS: Further large, independent, confirmatory, clinical validation studies and real-world, cost-effectiveness studies are necessary before the widespread adoption of these tests can be endorsed as standard of care.

A qualitative evaluation of the validity of published health utilities and generic health utility measures for capturing health-related quality of life (HRQL) impact of differentiated thyroid cancer (DTC) at different treatment phases.

PURPOSE: This study explored the impact of differentiated thyroid cancer (DTC) on health-related quality of life (HRQL) at different treatment phases and evaluated the validity of published DTC utilities and generic health utility measures (EQ-5D and SF-6D) for economic evaluation of treatments for radio-iodine (RAI) refractory DTC. METHODS: Focus groups and interviews were conducted with DTC patients grouped by treatment phase. Qualitative thematic analysis was conducted on interview/focus group transcripts. A thematic coding framework was developed to compare experiences between treatment phases and inform development of a conceptual model. Model concepts were mapped to EQ-5D and SF-6D domains/items. RESULTS: Eight focus groups and 11 individual interviews were conducted with 52 DTC patients. Fifty symptoms and HRQL concepts were identified. The impact of DTC and DTC treatment on emotional and cognitive functioning was reported across the treatment phases. The impact on daily activities, mobility, and energy levels was greatest for patients with recurring/persistent or RAI-refractory DTC. Of the 50 concepts, 25 and 27 mapped directly onto domains/items in the EQ-5D and SF-6D, respectively. The SF-6D covered a broader range of DTC impact on emotional/physical problems and daily/social activities than did the EQ-5D. CONCLUSIONS: The conceptual model summarizes the wide-ranging impact of DTC and its treatment on patients' HRQL, particularly for those with recurring/persistent or RAI-refractory DTC. Findings suggest that published DTC utilities lack validity for RAI-refractory DTC and that the SF-6D may be more sensitive to HRQL impact of DTC than the EQ-5D.

Clinical consequences of a change in anti-thyroglobulin antibody assays during the follow-up of patients with differentiated thyroid cancer.

OBJECTIVE: Thyroglobulin (Tg) quantitation by immunometric assays is compromised by anti-thyroglobulin antibodies (TgAbs), potentially resulting in falsely low Tg concentrations. TgAb screening is recommended when measuring Tg, but current TgAb immunoassays do not detect all possible TgAbs in circulation. We assessed the impact of a change in TgAb assay on apparent disease status of patients with differentiated thyroid carcinoma (DTC). METHODS: Patients seen at the Mayo Clinic, Rochester, Minnesota, for follow-up of DTC, who had been tested using 2 different TgAb assays (Beckman Access and Roche Elecsys) were identified. Electronic medical records were reviewed to evaluate any impact the change in TgAb assay had on clinical disease status assessment and follow-up. RESULTS: A total of 1,457 patients were tested using both assays. A change in TgAb status was found in 124 (8.5%) patients; a total of 117 patients who were TgAb-negative on the Beckman assay became TgAb-positive with the Roche assay. Additional testing was performed in 5 of these patients. Seven patients previously considered TgAb-positive were now TgAb-negative. In all 7 cases, physicians documented that they considered these patients now to be truly TgAb-negative and free of disease. CONCLUSION: Discrepancies in TgAb status are seen when using different TgAb assays. Relying on Tg and TgAb measurements to determine disease status may lead to underestimation of residual cancer. A multimodal (clinical, biochemical, and radiologic) approach to follow up on patients with DTC should be continued, pending the development of Tg quantitation methods that are highly sensitive and not affected by TgAb interference.

RET splice site variants in medullary thyroid carcinoma.

Introduction: Medullary thyroid carcinoma (MTC) is an aggressive cancer that is often caused by driver mutations in RET. Splice site variants (SSV) reflect changes in mRNA processing, which may alter protein function. RET SSVs have been described in thyroid tumors in general but have not been extensively studied in MTC. Methods: The prevalence of RET SSVs was evaluated in 3,624 cases with next generation sequence reports, including 25 MTCs. Fisher exact analysis was performed to compare RET SSV frequency in cancers with/without a diagnosis of MTC. Results: All 25 MTCs had at least one of the two most common RET SSVs versus 0.3% of 3,599 cancers with other diagnoses (p < 0.00001). The 11 cancers with non-MTC diagnoses that had the common RET SSVs were 4 neuroendocrine cancers, 4 non-small cell lung carcinomas, 2 non-MTC thyroid cancers, and 1 melanoma. All 25 MTCs analyzed had at least one of the two most common RET SSVs, including 4 with no identified mutational driver. Discussion: The identification of RET SSVs in all MTCs, but rarely in other cancer types, demonstrates that these RET SSVs distinguish MTCs from other cancer types. Future studies are needed to investigate whether these RET SSVs play a pathogenic role in MTC.

Characterization of the Thyroid Cancer Genomic Landscape by Plasma-Based Circulating Tumor DNA Next-Generation Sequencing.

Background: The limited availability of targeted therapies in thyroid cancer (TC) has challenged conventional treatment algorithms and has established urgency for the identification of targetable genomic abnormalities. In addition to widely adopted tissue-based next-generation sequencing (NGS), plasma-based circulating tumor DNA (ctDNA) NGS is rapidly emerging as a genomic biomarker detection method and is steadily gaining utility across solid tumors. To date, plasma-based genomic alterations in TC have not been determined. Herein, we profile potential actionable mutations detected through ctDNA in patients with TC subtypes. Methods: A retrospective data analysis of the Guardant Health, Inc. database was performed using the commercially available Guardant360® plasma-NGS test on TC samples from adult patients collected between 2016 and 2021. The landscape of genomic alterations and blood tumor mutation burden (bTMB) were analyzed in patients with different types of TC: anaplastic TC (ATC), papillary TC (PTC), follicular TC (FTC), oncolytic carcinoma of the thyroid (OCA), poorly differentiated TC (PDTC), medullary TC (MTC), and TC not otherwise specified (TC NOS). Results: Of the 1094 patients included most of the patients n = 876 had TC NOS, and 20% had a specific diagnosis (92 ATC, 62 PTC, 14 FTC, 16 OCA, 2 PDTC, and 32 MTC patients). The median age was 65 (range 10-98) and 47.3% were male. 78.3% of patients had one or more genomic alteration detected by ctDNA NGS. TP53 (46.9%) was the most common mutation detected among all TC. BRAFV600E was detected in 27.2% of ATC, 35.7% of PTC, and in none of FTC. RAS was detected in 18.5% of ATC, 11.9% of PTC, and 62.5% of FTC. RET, ALK, and NTRK fusions were seen in 1.1%, 0.5%, and 0.2% of all TC, respectively. RET mutations were detected in 66.7% of MTC. bTMB analysis was performed on 159 patients. The mean bTMB was higher in ATC compared with other types of TC (p = 0.0011, 0.0557, and <0.0001, respectively). Conclusions: Plasma-based comprehensive NGS is a promising NGS method in TC; however, future validation of the clinical utility by analysis of paired tumor and plasma samples is needed.

Molecular Alterations and Comprehensive Clinical Management of Oncocytic Thyroid Carcinoma: A Review and Multidisciplinary 2023 Update.

Importance: Oncocytic (Hürthle cell) thyroid carcinoma is a follicular cell-derived neoplasm that accounts for approximately 5% of all thyroid cancers. Until recently, it was categorized as a follicular thyroid carcinoma, and its management was standardized with that of other differentiated thyroid carcinomas. In 2022, given an improved understanding of the unique molecular profile and clinical behavior of oncocytic thyroid carcinoma, the World Health Organization reclassified oncocytic thyroid carcinoma as distinct from follicular thyroid carcinoma. The International Thyroid Oncology Group and the American Head and Neck Society then collaborated to review the existing evidence on oncocytic thyroid carcinoma, from diagnosis through clinical management and follow-up surveillance. Observations: Given that oncocytic thyroid carcinoma was previously classified as a subtype of follicular thyroid carcinoma, it was clinically studied in that context. However, due to its low prevalence and previous classification schema, there are few studies that have specifically evaluated oncocytic thyroid carcinoma. Recent data indicate that oncocytic thyroid carcinoma is a distinct class of malignant thyroid tumor with a group of distinct genetic alterations and clinicopathologic features. Oncocytic thyroid carcinoma displays higher rates of somatic gene variants and genomic chromosomal loss of heterozygosity than do other thyroid cancers, and it harbors unique mitochondrial DNA variations. Clinically, oncocytic thyroid carcinoma is more likely to have locoregional (lymph node) metastases than is follicular thyroid carcinoma-with which it was formerly classified-and it develops distant metastases more frequently than papillary thyroid carcinoma. In addition, oncocytic thyroid carcinoma rarely absorbs radioiodine. Conclusions and Relevance: The findings of this review suggest that the distinct clinical presentation of oncocytic thyroid carcinoma, including its metastatic behavior and its reduced avidity to radioiodine therapy, warrants a tailored disease management approach. The reclassification of oncocytic thyroid carcinoma by the World Health Organization is an important milestone toward developing a specific and comprehensive clinical management for oncocytic thyroid carcinoma that considers its distinct characteristics.

Molecular Profiling of 50 734 Bethesda III-VI Thyroid Nodules by ThyroSeq v3: Implications for Personalized Management.

CONTEXT: Comprehensive genomic analysis of thyroid nodules for multiple classes of molecular alterations detected in a large series of fine needle aspiration (FNA) samples has not been reported. OBJECTIVE: To determine the prevalence of clinically relevant molecular alterations in Bethesda categories III-VI (BCIII-VI) thyroid nodules. METHODS: This retrospective analysis of FNA samples, tested by ThyroSeq v3 using Genomic Classifier and Cancer Risk Classifier at UPMC Molecular and Genomic Pathology laboratory, analyzed the prevalence of diagnostic, prognostic, and targetable genetic alterations in a total of 50 734 BCIII-VI nodules from 48 225 patients. RESULTS: Among 50 734 informative FNA samples, 65.3% were test-negative, 33.9% positive, 0.2% positive for medullary carcinoma, and 0.6% positive for parathyroid. The benign call rate in BCIII-IV nodules was 68%. Among test-positive samples, 73.3% had mutations, 11.3% gene fusions, and 10.8% isolated copy number alterations. Comparing BCIII-IV nodules with BCV-VI nodules revealed a shift from predominantly RAS-like alterations to BRAF V600E-like alterations and fusions involving receptor tyrosine kinases (RTK). Using ThyroSeq Cancer Risk Classifier, a high-risk profile, which typically included TERT or TP53 mutations, was found in 6% of samples, more frequently BCV-VI. RNA-Seq confirmed ThyroSeq detection of novel RTK fusions in 98.9% of cases. CONCLUSION: In this series, 68% of BCIII-IV nodules were classified as negative by ThyroSeq, potentially preventing diagnostic surgery in this subset of patients. Specific genetic alterations were detected in most BCV-VI nodules, with a higher prevalence of BRAF and TERT mutations and targetable gene fusions compared to BCIII-IV nodules, offering prognostic and therapeutic information for patient management.

Radiofrequency ablation and related ultrasound-guided ablation technologies for treatment of benign and malignant thyroid disease: An international multidisciplinary consensus statement of the American Head and Neck Society Endocrine Surgery Section with the Asia Pacific Society of Thyroid Surgery, Associazione Medici Endocrinologi, British Association of Endocrine and Thyroid Surgeons, European Thyroid Association, Italian Society of Endocrine Surgery Units, Korean Society of Thyroid Radiology, Latin American Thyroid Society, and Thyroid Nodules Therapies Association.

BACKGROUND: The use of ultrasound-guided ablation procedures to treat both benign and malignant thyroid conditions is gaining increasing interest. This document has been developed as an international interdisciplinary evidence-based statement with a primary focus on radiofrequency ablation and is intended to serve as a manual for best practice application of ablation technologies. METHODS: A comprehensive literature review was conducted to guide statement development and generation of best practice recommendations. Modified Delphi method was applied to assess whether statements met consensus among the entire author panel. RESULTS: A review of the current state of ultrasound-guided ablation procedures for the treatment of benign and malignant thyroid conditions is presented. Eighteen best practice recommendations in topic areas of preprocedural evaluation, technique, postprocedural management, efficacy, potential complications, and implementation are provided. CONCLUSIONS: As ultrasound-guided ablation procedures are increasingly utilized in benign and malignant thyroid disease, evidence-based and thoughtful application of best practices is warranted.

Efficacy of pazopanib in progressive, radioiodine-refractory, metastatic differentiated thyroid cancers: results of a phase 2 consortium study.

BACKGROUND: Chemotherapy has historically proven ineffective in advanced differentiated thyroid cancers, but the realisation that various tyrosine kinases are activated in the disease suggested a potential therapeutic role for tyrosine-kinase inhibitors. We investigated the safety and efficacy of pazopanib. METHODS: This phase 2 trial was done from Feb 22, 2008, to Jan 31, 2009, in patients with metastatic, rapidly progressive, radioiodine-refractory differentiated thyroid cancers. Each patient received 800 mg continuous pazopanib daily in 4-week cycles until disease progression, drug intolerance, or both occurred. Up to two previous therapies were allowed, and measurable disease with radiographic progression in the 6-month period before enrolment was a requirement for inclusion. The primary endpoint was any tumour response, according to the Response Evaluation Criteria in Solid Tumors 1.0. This study is registered with ClinicalTrials.gov, number NCT00625846. FINDINGS: 39 patients were enrolled. One patient had received no previous radioiodine therapy and another withdrew consent before treatment. Clinical outcomes could, therefore, be assessed in 37 patients (19 [51%] men, median age 63 years). The study is closed to accrual of new patients, but several enrolled patients are still being treated. Patients received a median of 12 cycles (range 1 to >23, total >383). Confirmed partial responses were recorded in 18 patients (response rate 49%, 95% CI 35-68), with likelihood of response lasting longer than 1 year calculated to be 66%. Maximum concentration of pazopanib in plasma during cycle one was significantly correlated with radiographic response (r=-0·40, p=0·021). 16 (43%) patients required dose reductions owing to adverse events, the most frequent of which (any grade) were fatigue (29 patients), skin and hair hypopigmentation (28), diarrhoea (27), and nausea (27). Two patients who died during treatment had pre-existing contributory disorders. INTERPRETATION: Pazopanib seems to represent a promising therapeutic option for patients with advanced differentiated thyroid cancers. The correlation of the patient's response and pazopanib concentration during the first cycle might indicate that treatment can be individualised to achieve optimum outcomes. Assessment of pazopanib in an expanded cohort of patients with differentiated thyroid cancer, as well as in cohorts of patients with medullary and anaplastic thyroid cancers, is presently being done. FUNDING: National Cancer Institute, supported in part by NCI CA15083 and CM62205.

Bilateral testicular adrenal rest tumors in a patient with nonclassical congenital adrenal hyperplasia.

INTRODUCTION: Solid testis tumors in post-pubertal males usually represent germ cell malignancies; however, other uncommon or rare histologies must be considered. CASE PRESENTATION: We present a case of an 18-year-old male undergoing attempted bilateral partial orchiectomies for suspected germ cell tumors. Tumor pathology, laboratory results, radiographic studies, and post-surgical elevated adrenocorticotropic hormone levels supported the diagnosis of testicular adrenal rest tumors secondary to previously undiagnosed nonclassical congenital adrenal hyperplasia. CONCLUSION: Testicular adrenal rest tumors are rare in patients with nonclassical congenital adrenal hyperplasia and may be accompanied by adrenal insufficiency and hypogonadism, which can be treated with glucocorticoid therapy and testosterone replacement. Differential diagnosis of tumors is challenging but necessary for proper symptom-based management.

Outcomes of repeat fineneedle aspiration biopsy for AUS/FLUS thyroid nodules.

OBJECTIVE: To determine the proportion of aspirates reclassified into each Bethesda category and to assess the rates of malignancy in each of them on repeat fine-needle aspiration biopsy (RFNA) following an AUS/FLUS diagnosis. DESIGN: Systematic review and meta-analysis. METHODS: On February 2019, Pubmed/MEDLINE, EMBASE, WoS, and the Cochrane Library were searched for articles published from January 1, 2007. All studies published in English describing RFNA outcomes in AUS/FLUS nodules were included. PRISMA and MOOSE guidelines were followed. Five investigators independently assessed the eligibility of the studies. Two investigators extracted summary data and assessed the risk of bias. Data were pooled using a random-effects model. The rate of malignancy was calculated on resected nodules only (upper limit of true value); and considering all unresected nodules were benign (lower limit of true value). The protocol was registered in PROSPERO (CRD42019123114). RESULTS: Of 2937 retrieved studies, 27 were eligible. The meta-analysis was conducted on summary data of 3932 AUS/FLUS thyroid nodules with RFNA. RFNA cytology would reclassify into categories I through VI of Bethesda: 4% (3%, 5%), 48% (43%, 54%), 26% (20%, 32%), 4% (3%, 6%), 5% (3%, 6%), and 2% (1%, 2%) of AUS/FLUS nodules. Malignancy rates of resected nodules were 24% (9%, 38%), 4% (1%, 7%), 40% (28%, 52%), 37% (27%, 47%), 79% (69%, 90%), and 99% (95%, 100%) for categories I through VI of Bethesda. There was high heterogeneity in these data. CONCLUSIONS: RFNA reclassified two-thirds of the AUS/FLUS specimens into a more definitive cytological category, with a benign call rate of nearly 50% and a negative predictive value greater than 96%.

Integrating Somatic and Germline Next-Generation Sequencing Into Routine Clinical Oncology Practice.

Next-generation sequencing (NGS) is rapidly expanding into routine oncology practice. Genetic variations in both the cancer and inherited genomes are informative for hereditary cancer risk, prognosis, and treatment strategies. Herein, we focus on the clinical perspective of integrating NGS results into patient care to assist with therapeutic decision making. Five key considerations are addressed for operationalization of NGS testing and application of results to patient care as follows: (1) NGS test ordering and workflow design; (2) result reporting, curation, and storage; (3) clinical consultation services that provide test interpretations and identify opportunities for molecularly guided therapy; (4) presentation of genetic information within the electronic health record; and (5) education of providers and patients. Several of these key considerations center on informatics tools that support NGS test ordering and referencing back to the results for therapeutic purposes. Clinical decision support tools embedded within the electronic health record can assist with NGS test utilization and identifying opportunities for targeted therapy including clinical trial eligibility. Challenges for project and change management in operationalizing NGS-supported, evidence-based patient care in the context of current information technology systems with appropriate clinical data standards are discussed, and solutions for overcoming barriers are provided.

Evaluation of the PAX8/PPARG translocation in follicular thyroid cancer with a 4-color reverse-transcription PCR assay and automated high-resolution fragment analysis.

BACKGROUND: Molecular testing of thyroid malignancies, in combination with cytologic and histologic examination, is becoming increasingly attractive as a tool for refining traditional morphologic diagnosis. The molecular changes associated with follicular thyroid carcinoma (FTC) are point mutations in RAS oncogenes or the presence of PAX8/PPARG (paired box 8/peroxisome proliferator-activated receptor gamma) rearrangement. METHODS: We developed and validated a clinical assay for the detection of PAX8/PPARG rearrangements that uses a 4-color reverse-transcription PCR (RT-PCR) assay and high-resolution fragment analysis. RESULTS: The RT-PCR assay is applicable for detecting the various described fusion transcripts of PAX8/PPARG in formalin-fixed, paraffin-embedded thyroid tissue and in fine-needle aspirate biopsy washes from thyroid nodules. The analytical sensitivity of the assay is 1 abnormal cell in a background of 100-10 000 translocation-negative cells. A comparison of the RT-PCR assay with dual-fusion fluorescence in situ hybridization showed an overall concordance of 95%. With this assay, we obtained a prevalence for the PAX8/PPARG rearrangement in FTC of 62% (13 of 21 cases), compared with a 5% prevalence (3 of 55) for other follicular cell-derived neoplasms. CONCLUSIONS: The introduction of this assay into clinical practice could provide useful information for the diagnosis and possibly for the prognosis and treatment of thyroid cancer in the future.

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

Background: Although most thyroid nodules with indeterminate cytology are benign, in most of the world, surgery remains as the most frequent diagnostic approach. 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 the 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 a greater proportion of the Hispanic population (40% vs. 3%) for TGCT-1 and a greater proportion of African American (11% vs. 0%) and Asian (10% vs. 1%) populations for TGCT-2; and (iii) tumor size (mean of 1.7 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 TGCT-1 and -2, respectively. One hundred 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.