Gene Expression Classifier Testing Research Articles

Trials in the Key of G: Building Level 1 Evidence on the Real-world Effectiveness of Prostate Biomarkers

A number of genomic classifiers are available to aid in shared decision-making for men with localized prostate cancer; however, there is no high-level evidence assessing their clinical utility. The two randomized controlled trials in this report prospectively evaluate the use of gene expression classifier testing at the time of cancer diagnosis and after surgical treatment.

Treatment patterns and outcomes in prostate cancer patients tested with Decipher in SEER.

e17006 Background: In 2021, national data between the Decipher 22-gene prognostic gene expression classifier (GC) for men with prostate cancer and the Surveillance, Epidemiology, and End Results (SEER) cancer registries were linked. The purpose of the work is to report on the linkage by characterizing national GC usage and its association with treatment decisions for men with prostate cancer. Methods: Patients in the SEER registries with primary prostate cancer diagnosis from 2010 to 2018 were included and linked to data from GC testing conducted between 2014 to 2020 (Veracyte, San Diego, CA). GC scores (range 0-1) and GC risk groups (low, intermediate, and high) were used for continuous and categorical analyses. Multivariable logistic regression was used to quantify the association between GC and active surveillance and watchful waiting (AS/WW) use and adverse pathology at radical prostatectomy (RP). Adverse pathology was defined as pathological grade group ≥3, pathological stage ≥pT3b, or lymph node invasion. Results: A total of 575,363 patients were eligible for analysis, of which 10,528 patients underwent GC testing (5,015 GC biopsy test, and 5,513 GC RP test). The median age was 67 for both tested and untested, but more white patients underwent testing (82% vs 76%, p < 0.001). For GC biopsy tested patients, AS/WW was highest for those with GC low risk results (41%) as compared to those with intermediate (32%) or high (17%) GC risk (p < 0.001). RP rates were lower in the tested compared to untested (25% vs. 36%, p < 0.001), and among the tested patients, RP use increased by GC risk group (19% of low, 25% of intermediate, and 34% of high GC risk, p < 0.001). A similar trend by GC risk group in management for radiation therapy was observed (13% of low, 19% of intermediate, and 29% of high GC risk, p < 0.001). In a multivariable logistic regression adjusted for age, race, NCCN risk group, and year of diagnosis, GC tested patients were more likely to undergo AS/WW compared to untested (OR 2.9 [95% CI, 2.8-3.1], p < 0.001). Within the subset of patients classified as NCCN low/favorable intermediate risk at biopsy and who were subsequently treated with RP (n = 594), GC high risk (> 0.6) was associated with more than 3 times the odds of harboring adverse pathology (OR 3.2 [95% CI 1.6-6.4], p < 0.001). Conclusions: Using the first ever linked SEER-Decipher data, we demonstrate that population-based treatment patterns are independently associated with GC test results. Patients with lower GC scores are independently more likely to undergo AS/WW; those with higher scores are more likely to have adverse pathology at time of RP.

Molecular Testing Has Limited Utility in the Surgical Evaluation of Bethesda III Thyroid Nodules

The Bethesda System for Reporting Thyroid Cytopathology has 6 diagnostic categories, each with an implied cancer risk of malignancy (ROM). Bethesda III, defined as atypia or follicular lesions of undetermined significance (AUS/FLUS) on fine needle aspiration (FNA), has an indeterminate ROM. This study investigates the utility of Afirma Gene Expression Classifier (GEC) and Thyroid Sequencing (ThyroSeq) molecular testing to predict malignancy in AUS/FLUS thyroid nodules. A retrospective review of prospectively collected data of 1457 patients with index thyroid nodules who underwent FNA and thyroidectomy at a single academic institution was performed. Use of GEC or ThyroSeq for AUS/FLUS thyroid nodules was examined. GEC testing was reported benign or suspicious for malignancy whereas ThyroSeq testing was reported on a spectrum of low, intermediate or high ROM. Descriptive statistics were utilized to compare the ROM among AUS/FLUS thyroid nodules. Of 1457 patients with FNA thyroid cytology, 359 (25%) corresponded to AUS/FLUS results. There were 132 (37%) patients with GEC testing and 88 (24%) had ThyroSeq testing. ROM without GEC or ThyroSeq testing was 49%, whereas ROM with suspicious GEC was 55%. ROM with positive ThyroSeq was 73%. Among ThyroSeq patients, 43 had intermediate-risk mutations with 60% malignancy, and 23 had high-risk mutations with 96% malignancy (P < 0.01). Surgical patients with AUS/FLUS thyroid nodules have a high ROM. High-risk ThyroSeq testing may have some utility in predicting malignancy, but GEC and intermediate-risk TGC results have limited value. Surgeons should carefully consider the utility of molecular tests to determine surgical resection.

Combined molecular and histologic end points inform cancer risk estimates for thyroid nodules classified as atypia of undetermined significance.

Thyroid nodules classified as atypia of uncertain significance (AUS) on fine-needle aspiration cytology are heterogeneous. Prior studies reported a higher risk of noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP)/cancer among AUS nodules that had cytologic (AUS-C) versus architectural (AUS-A) atypia; however, such studies were generally confined to resected cohorts, introducing bias into risk calculations. The authors hypothesized that combined histologic and molecular end points would permit clinically meaningful calculations of NIFTP/malignancy risk among AUS nodules. The study consisted of 279 thyroid nodules classified as AUS on initial fine-needle aspiration and tested by the Afirma Gene Expression Classifier (GEC) between June 2013 and October 2017. Results of GEC testing and histopathologic diagnoses were stratified by AUS classifiers. The AUS-A category was further subclassified as 1) hypocellular microfollicular or 2) cellular with mixed but predominantly microfollicular architecture. NIFTP/cancer risk was calculated for each subgroup, with the inclusion of unresected nodules that had benign GEC results as low-risk end points comparable to histologically benign nodules. When only histologic end points were considered, there was no difference in NIFTP/cancer risk (25% vs 23%; P = .82). By using molecular and histologic end points, AUS cases with cytologic atypia trended toward higher NIFTP/cancer risk than AUS-A cases (14% vs 6%; P = .06). Furthermore, AUS-A cases showed a trend toward lower NIFTP/cancer risk for hypocellular microfollicular aspirates (3%) compared with cellular samples that had mixed/predominantly microfollicular architecture (13%; P = .18). The inclusion of unresected benign GEC nodules in risk-of-malignancy calculations provides more accurate results, which may be helpful for informing patient management as well as quality improvement in the cytopathology laboratory.

Repeat Fine Needle Aspiration Cytology Refines the Selection of Thyroid Nodules for Afirma Gene Expression Classifier Testing.

Background: Molecular testing (MT) refines risk stratification for thyroid nodules that are indeterminate for cancer by fine needle aspiration (FNA) cytology. Criteria for selecting nodules for MT vary and remain largely untested, raising questions about the best strategy for maximizing the usefulness of MT while minimizing the harms of overtesting. We used a unique data set to examine the effects of repeat FNA cytology-based criteria for MT on management decisions and nodule outcomes. Methods: This was a study of adults (age 25-90 years; 281 women and 72 men) with cytologically indeterminate (Bethesda III/IV) thyroid nodules who underwent repeat FNA biopsy and Afirma Gene Expression Classifier (GEC) testing (N = 363 nodules from 353 patients) between June 2013 and October 2017 at a single institution, with follow-up data collected until December 2019. Subgroup analysis was performed based on classification of repeat FNA cytology. Outcomes of GEC testing, clinical/sonographic surveillance of unresected nodules, and histopathologic diagnoses of thyroidectomies were compared between three testing approaches: (i) Reflex (MT sent on the basis of the initial Bethesda III/IV FNA), (ii) SemiRestrictive (MT sent if repeat FNA is Bethesda I-IV), and (iii) Restrictive (MT sent only if repeat FNA is Bethesda III/IV) testing approaches. Results: Restricting MT to nodules that remain Bethesda III/IV on repeat FNA would have missed 4 low-risk cancers and 3 noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTP) (collectively 2% of the test population) but would have avoided diagnostic surgery for 42 benign nodules (12% of the test population). The Restrictive testing strategy was more specific (delta 0.126 confidence interval [CI 0.093 to 0.159] and 0.129 [CI 0.097 to 0.161], respectively) but less sensitive (delta -0.339 [CI -0.424 to -0.253] and -0.340 [CI -0.425 to -0.255], respectively) than the Reflex and SemiRestrictive approaches for detecting NIFTP or cancer. Conclusions: Repeat FNA cytology can guide the selection of cytologically indeterminate thyroid nodules that warrant MT. The Restrictive model of performing Afirma GEC only on nodules with two separate biopsies showing Bethesda III/IV cytology would reduce the rate of diagnostic surgery for histologically benign nodules while missing only rare low-risk tumors. Given the low but nontrivial risks of thyroidectomy, the higher specificity of the Restrictive testing approach disproportionately outweighs the potential harms.

Prognostic capabilities and clinical utility of cell cycle progression testing, prostate imaging reporting and data system, version 2, and clinicopathologic data in management of localized prostate cancer

ObjectivesTo compare the prognostic capabilities and clinical utility of the cell cycle progression (CCP) gene expression classifier test, multiparametric magnetic resonance imaging (mpMRI) with Prostate Imaging Reporting and Data System (PI-RADS) scoring, and clinicopathologic data in select prostate cancer (PCa) medical management scenarios. Patients and methodsRetrospective, observational analysis of patients (N = 222) ascertained sequentially from a single urology practice from January 2015 to June 2018. Men were included if they had localized PCa, a CCP score, and an mpMRI PI-RADS v2 score. Cohort 1 (n = 156): men with newly diagnosed PCa, with or without a previous negative biopsy. Cohort 2 (n = 66): men who initiated active surveillance (AS) without CCP testing, but who received the test during AS. CCP was combined with the UCSF Cancer of the Prostate Risk Assessment (CAPRA) score to produce a clinical cell-cycle risk (CCR) score, which was reported in the context of a validated AS threshold. Spearman's rank correlation test was used to evaluate correlations between variables. Generalized linear models were used to predict binary Gleason score category and medical management selection (AS or curative therapy). Likelihood-ratio tests were used to determine predictor significance in both univariable and multivariable models. ResultsIn the combined cohorts, modest but significant correlations were observed between PI-RADS score and CCP (rs = 0.22, P = 8.1 × 10−4), CAPRA (rs= 0.36, P = 4.8 × 10−8), or CCR (rs = 0.37, P = 2.0 × 10−8), suggesting that much of the prognostic information captured by these measures is independent. When accounting for CAPRA and PI-RADS score, CCP was a significant predictor of higher-grade tumor after radical prostatectomy, with the resected tumor approximately 4 times more likely to harbor Gleason ≥4+3 per 1-unit increase in CCP in Cohort 1 (Odds Ratio [OR], 4.10 [95% confidence interval [CI], 1.46, 14.12], P = 0.006) and in the combined cohorts (OR, 3.72 [95% CI, 1.39, 11.88], P = 0.008). On multivariable analysis, PI-RADS score was not a significant predictor of post-radical prostatectomy Gleason score. Both CCP and CCR were significant and independent predictors of AS versus curative therapy in Cohort 1 on multivariable analysis, with each 1-unit increase in score corresponding to an approximately 2-fold greater likelihood of selecting curative therapy (CCP OR, 2.08 [95% CI, 1.16, 3.94], P = 0.014) (CCR OR, 2.33 [95% CI, 1.48, 3.87], P = 1.5 × 10−4). CCR at or below the AS threshold significantly reduced the probability of selecting curative therapy over AS (OR, 0.28 [95% CI, 0.13, 0.57], P = 4.4 × 10−4), further validating the clinical utility of the AS threshold. ConclusionCCP was a better predictor of both tumor grade and subsequent patient management than was PI-RADS. Even in the context of targeted biopsy, molecular information remains essential to ensure precise risk assessment for men with newly diagnosed PCa.

Afirma Testing in Thyroid Nodules with Indeterminate Cytology: A single Medical Institute Experience

Abstract Introduction/Objective About 10 to 30% of thyroid fine needle aspiration (FNA) nodules have indeterminate cytology, including Bethesda III, IV, and V. Afirma Gene Expression Classifier (GEC) measuring the expression of 167 mRNA was designed to classify the indeterminate thyroid nodules into benign and suspicious categories. This study aimed to evaluate the clinical performance of the Afirma GEC testing in these indeterminate thyroid lesions. Methods Medical records of patients with indeterminate thyroid FNA results and corresponding Afirma GEC results from November 2012 to December 2019 were retrieved. Subsequent surgical follow-up results were obtained. GEC results were compared to the histologic diagnoses. Results 1. There were 77 cases with indeterminate thyroid FNA results. Cytology diagnosis included 67 Bethesda III and 10 Bethesda IV. Afirma GEC results were benign for 41 cases (53%), suspicious for 32 cases (42%), and non- diagnostic for 4 cases (5%). Twenty seven of 32 cases (84%) with suspicious GEC results had surgical follow-up which revealed 12 malignant and 15 benign histologic diagnoses. Only 11 of 41 cases (27%) with benign GEC results had surgery which showed 8 benign and 3 malignant diagnoses. Based on the data, the sensitivity of this test is 80% and specificity is 65%. Positive predictive value (PPV) is 44% and negative predictive value (NPV) is 73%. The false positive cases include 5 Hurthle cell adenoma, 4 nodular hyperplasia, 3 follicular adenoma, 2 Noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFPT), and 1 lymphocytic thyroiditis. The false negative cases include 2 minimally invasive follicular carcinoma (from same patient) and 1 papillary thyroid carcinoma. Conclusion 1. We demonstrated in this study that a little more than half of the cases with indeterminate thyroid cytology had negative Afirma GEC results. 2.Afirma test has relatively low specificity (65%) and PPV (44%). The sensitivity (80%) and NPV (73%) is relatively higher but is lower than the values in most literature. This could be due to the fact that majority of the Afirma GEC negative cases in this study did not have surgical follow-up and the sample size is small. 3.Afirma GEC test is a relatively good “rule-out” molecular test for indeterminate thyroid nodules but is not a reliable “rule-in” test due to the low specificity and PPV.

Utility of Afirma Gene Expression Classifier for Evaluation of Indeterminate Thyroid Nodules and Correlation with Ultrasound Risk Assessment: Single Institutional Experience

Objective: We assessed our experience with Afirma gene expression classifier (GEC) combined with sono-graphic risk assessment, using both the American Thyroid Association (ATA) and the Thyroid Imaging Reporting and Data System (TI-RADS) in evaluating indeterminate thyroid nodules. Methods: We identified 98 patients with 101 nodules who had a second fine needle aspiration biopsy (FNA) between January 1, 2014, and September 30, 2017, and sent to Veracyte for cytopathology and subsequent Afirma GEC testing. A second FNA biopsy was performed if the initial cytopathology was either Bethesda III or IV (n = 94) or nondiagnostic (n = 7). We correlated cytopathology, histopathology, and Afirma GEC results with sonographic risk assessment using both the ATA system and TI-RADS. Results: The mean age of the cohort was 57.4 ± 12.3 years; 84% women and 60% white. Repeat FNA was benign in 51 of 101 nodules, and of the remaining 50 nodules, 18 (36%) were GEC-benign and 32 (64%) GEC-suspicious. Eighteen of the 32 GEC-suspicious nodules underwent surgery with the following results: 7 benign (39%), 1 follicular thyroid carcinoma (6%), 6 follicular variant of papillary thyroid cancer (33%), and 4 noninvasive follicular tumor with papillary-like nuclear features (22%). The malignancy rate among the surgical cohort was 39% (without noninvasive follicular tumor with papillary-like nuclear features [NIFTP]) and 61% (with NIFTP) and about 50% and 20% of this group scored in the high suspicion category by ATA and TR5 by TI-RADS, respectively. Conclusion: Afirma GEC was useful in avoiding surgery in one-third of indeterminate nodules and performed similarly to ATA and TI-RADS. However, the use of echogenicity in scoring may underestimate the risk of malignancy in patients with indeterminate nodules. Abbreviations: ATA = American Thyroid Association; AUS = Atypia of Undetermined Significance; FLUS = Follicular Lesion of Undetermined Significance; FN = follicular neoplasm; FNA = fine needle aspiration; FTC = follicular thyroid cancer; FVPTC = follicular variant of papillary thyroid cancer; GEC = Gene Expression Classifier; ND = nondiagnostic; NIFTP = noninvasive follicular tumor with papillary-like nuclear features; TI-RADS = Thyroid Imaging Reporting and Data System; TR = TI-RADS.

Long-Term Outcomes of Thyroid Nodule AFIRMA GEC Testing and Literature Review: An Institutional Experience.

To assess outcomes of thyroid nodules analyzed with the AFIRMA gene expression classifier (GEC) and to perform a comprehensive literature review. Retrospective analysis of patients with thyroid nodules who underwent AFIRMA GEC testing at our institution. A tertiary care academic institution. We collected clinical outcomes for 416 thyroid nodules that were analyzed with AFIRMA GEC between 2011 and 2015, including long-term follow-up through 2019. We performed a comprehensive literature review. The resection rate for nodules with "suspicious" GEC results was 85% with a positive predictive value of 37%. The resection rate for nodules with "benign" GEC results was 24% with a negative predictive value of 90%. The prevalence of thyroid malignancy in patients with thyroid nodules with indeterminate cytology at our institution during this timeframe was 41%, thus lowering our negative predictive value. Mean follow-up duration for unresected nodules was 27.8 months. Our resection rates for nodules with "benign" GEC were among the highest reported in the literature. Molecular marker testing of thyroid nodules with indeterminate cytology can aid in the surgical decision making by obviating the need for diagnostic surgery and/or guiding extent of resection. Patients with other indications for surgery may not benefit from such costly testing.

Chronic Lymphocytic Thyroiditis May Lower Accuracy of AUS/FLUS Cytopathology in Surgical Patients

BackgroundChronic lymphocytic thyroiditis (CLT) increases cytologic atypia on fine-needle aspiration of thyroid nodules, and its effect on rate of malignancy in atypia of undetermined significance (AUS)/follicular lesions of undetermined significance (FLUS) thyroid nodules remains unclear. This study evaluates the effect of concomitant CLT on malignancy rates of AUS/FLUS thyroid nodules in surgical patients. MethodsRetrospective review of 1061 patients who underwent thyroidectomy for a dominant thyroid nodule from a single institution was performed. Fine-needle aspiration was classified according to the Bethesda System for Reporting Thyroid Cytopathology. Patients with AUS/FLUS cytopathology were classified into two cohorts: AUS/FLUS with CLT and AUS/FLUS without CLT. Final pathology was reviewed, and the cohorts were further stratified into benign and malignant subgroups. When applicable, patients with gene expression classifier (GEC) testing were reviewed and the positive predictive value (PPV) was calculated. ResultsOf the entire surgical series, 293 (28%) patients had AUS/FLUS cytopathology with a rate of malignancy of 56% (163/293) on final pathology. Seventy-three (25%) patients had AUS/FLUS with CLT, of which 44% (n = 32) were malignant by final pathology. The remaining 75% (n = 220) had AUS/FLUS without CLT, 60% (n = 131) of which were malignant. GEC testing was performed in 36 of the AUS/FLUS with CLT patients, where of the 33 suspicious results, 17 were malignant on final pathology, yielding a PPV of 52%. ConclusionsThe rate of malignancy for AUS/FLUS thyroid nodules is lower with coexisting CLT, and similar to previous studies, the PPV of GEC testing is approximately 50%. Cytologic atypia due to CLT may increase more AUS/FLUS results in thyroid nodules, which may lead to overestimation of malignancy rates in this patient population.

Hurthle cell predominance impacts results of Afirma gene expression classifier and ThyroSeq molecular panel performance in indeterminate thyroid nodules

Molecular tests such as the Afirma gene expression classifier (GEC) and mutational panels (such as ThyroSeq) have been introduced to help risk stratify cytologically indeterminate thyroid nodules with the aim to reduce the number of unnecessary thyroidectomies. Some reports have suggested that samples with Hurthle cell predominance have higher false-positive rates on GEC testing, but data are limited. We reviewed thyroid nodules with indeterminate (Bethesda III/IV) cytology at our institution. Patient demographics, cytologic and histologic diagnoses (where available), and molecular test results were collected. GEC was performed on 202 nodules, and ThyroSeq was performed on 81 nodules. In the GEC cohort, 66% of nodules with Hurthle cell predominance yielded "suspicious" result vs 46% of nodules without Hurthle cell predominance, with risk of malignancy (ROM) for surgically resected nodules of 16% and 33%, respectively. In ThyroSeq cohort, 8% of nodules with Hurthle cell predominance yielded a high-risk mutation vs 19% of nodules without Hurthle cell predominance, with ROM of 50% and 33%, respectively. For ThyroSeq molecular panel, while it did not appear that there was an increase in rate of high-risk mutations detected in the samples with Hurthle cell predominance, small numbers limit the generalizability of these results. For the GEC cohort, indeterminate thyroid nodules with predominance of Hurthle cells showed an increased rate of "suspicious" results compared to samples without Hurthle cell predominance. The ROM for GEC "suspicious" nodules with Hurthle cell predominance on surgical resection was lower in our study. Repeat FNA may be of use in patients with these types of nodules. In the context of a Hurthle cell predominant lesion, positive results on molecular testing may not carry a high rate of malignancy.

MON-569 Performance of American Thyroid Association Sonographic Risk and Gene Expression Classifier in the Diagnosis of Thyroid Nodules with Indeterminate Fine Needle Aspiration Cytology

Introduction: The 2015 American Thyroid Association (ATA) guidelines recommend decision making for nodules with indeterminate cytology to include consideration of sonographic features prior to performing molecular testing. However, only a few studies have evaluated how ATA sonographic risk categories predict risk of malignancy along with gene expression classifier (GEC). It has been suggested that in ATA high suspicion nodules, GEC may not add much value. Objective: Evaluate, among indeterminate nodules, how ATA sonographic risk categories perform in comparison to GEC and if the combination of these tools may increase the predictability of malignancy. Methods: A retrospective single center study included Bethesda III thyroid nodules that had undergone evaluation by GEC between 2012-2015. Each nodule was reviewed by 3 radiologists and classified using the 2015 ATA risk categories based on sonographic features: a) high suspicion (HS), b) intermediate suspicion (IS), c) low suspicion (LS), and d) very low suspicion (VLS) of malignancy or e) benign. Nodules were determined to be benign or malignant based on surgical pathology or minimum 2 year follow up if available. Positive predictive values (PPV) and negative predictive values (NPV) were calculated. Overall PPV and NPV for ATA classification were calculated using HS and IS nodules as a positive test and LS and VLS nodules as a negative test. Results: 97 nodules with Bethesda III cytology that had undergone GEC testing were included. Of these, 72 were deemed benign or malignant (17 of 25 nodules that lost follow up had benign GEC). One nodule with suspicious GEC and malignant pathology had no agreement between the readers for ATA sonographic risk category. Prevalence of malignancy was 29%. Overall PPV for malignancy of GEC was calculated at 40% and NPV at 92% (95% CI 0.75-0.98). The overall PPV of ATA classification was 39% and NPV was 82%. The ATA HS nodules (n=31) had 65% suspicious GEC, a PPV of 43% alone, and a combined PPV and NPV with GEC, of 59% and 100%, respectively. The IS nodules (n=21) had 57% suspicious GEC, a PPV of 29%, and combined PPV and NPV with GEC of 44% and 100%. The LS nodules (n=37) had 49% suspicious GEC, a PPV of 18%, and combined PPV and NPV with GEC of 19% and 83% (95% CI 0.55-0.95). The VLS nodules (n=7) had 57% suspicious GEC, a PPV of 17%, and combined PPV and NPV with GEC of 25% and 100%. Conclusion: Among thyroid nodules with Bethesda III cytology that underwent GEC, overall PPV based on ATA classification is comparable to the PPV of GEC, but NPV was higher with GEC. The HS category had the highest rate of suspicious GEC. Within each ATA risk category, the PPV increased moving from VLS to HS categories. Combining ATA sonographic risk with GEC resulted in improved predictive values within each category. Overall, the NPV across all ATA categories was very high and suggests that it is a reliable rule-out test.

Molecular testing for indeterminate thyroid nodules: Performance of the Afirma gene expression classifier and ThyroSeq panel.

The ThyroSeq mutational panel and Afirma gene expression classifier (GEC) are used to risk stratify cytologically indeterminate thyroid nodules. In the current study, the authors evaluated the performance of these tests within the context of ultrasonographic features and with the incorporation of the noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) nomenclature. The authors reviewed nodules using ThyroSeq or Afirma GEC testing. For nodules that were surgically resected, both tests were studied within the context of ultrasound findings, comparing performance stratified by the 2015 American Thyroid Association guideline (ATA 2015) sonographic patterns and assessing the positive predictive value (PPV) of these tests both including and excluding NIFTP in the malignant category. A total of 304 cases were identified, 119 of which were resected. All cases that met the criteria for NIFTP on excision demonstrated either high-risk mutations on ThyroSeq or a "suspicious" result on Afirma GEC. When NIFTP cases were shifted from the malignant to nonmalignant category, the PPV of "positive" tests for both ThyroSeq and Afirma GEC decreased from 42.9% to 14.3% (an absolute decrease of 28.6%) and 30.1% to 25.3% (an absolute decrease of 4.8%), respectively. No cases of malignancy were found in the ATA 2015 "very low suspicion" group, even with a "suspicious" Afirma GEC result. Both the ThyroSeq and Afirma GEC tests demonstrated decreases in the PPV when NIFTP was considered nonmalignant. In the era of NIFTP, a "positive" test result for either the Afirma GEC or ThyroSeq should be interpreted in light of clinical factors and should not exclude conservative (ie, lobectomy) surgical management. ATA 2015 "very low suspicion" nodules, even with "suspicious" Afirma GEC results, were not found to demonstrate malignancy in this series. Cancer Cytopathol 2018. © 2018 American Cancer Society.

Shear wave elastography and Afirma\u2122 gene expression classifier in thyroid nodules with indeterminate cytology: a comparison study

PurposeTo compare shear wave elastography (SWE) and Afirma™ gene expression classifier (GEC) for diagnosis of malignancy in thyroid nodules (TNs) with Bethesda Classification (BC) III or IV indeterminate cytology.MethodsThis preliminary single-center prospective study was approved by the Institutional Review Board. We evaluated 151 consented patients with 151 indeterminate TNs (123 BC III, 28 BC IV) on fine-needle aspiration biopsy (FNAB). B-mode ultrasound, vascularity, and SWE were performed prior to FNAB. TN stiffness was measured as shear wave velocity (SWV) in meters per second (m/s). The stiffest area of the TN was selected for SWV measurement. GEC testing was performed with a second FNAB. Surgery was recommended for GEC-suspicious TNs, or GEC-benign TNs with two or more worrisome B-mode US features.ResultsSurgical pathology confirmed 31 malignant TNs. Among the GEC-suspicious group, 28 of 59 TNs were malignant. The SWV value of ≥3.59 m/s was the best cut-off for malignancy risk based on the receiver operating curve (ROC). Twenty-six malignant TNs had SWV ≥ 3.59 m/s. The sensitivity and specificity for SWV ≥ 3.59 m/s were 83.9 and 79.2%, respectively. Positive predictive value (PPV) was 51.0% and negative predictive value (NPV) was 95.0%. For the GEC-suspicious group, sensitivity, specificity, PPV, and NPV were 90.3, 74.2, 47.5, and 96.7%, respectively. In multivariate analysis, SWV and GEC-suspicious were significant predictors of malignancy, but B-mode features and vascularity were not.ConclusionThis preliminary study indicates that SWE and GEC are independent predictors of malignancy in TNs with BC III or IV.

Pre-operative features of non-invasive follicular thyroid neoplasms with papillary-like nuclear features: An analysis of their cytological, Gene Expression Classifier and sonographic findings.

To investigate the corresponding cytological diagnoses, Gene Expression Classifier (GEC) results and ultrasound features of thyroid nodules diagnosed as non-invasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTP), as well as any coexisting pathology. We performed a retrospective review of thyroid nodules histologically diagnosed as NIFTP at our institution between 1st April 2016 and 1st April 2017. The following data points were collected: demographics, nodule size, ultrasound features, cytological diagnosis, GEC results, origin of sample (in-house vs outside hospital) and any additional pathology identified in the resection specimen. The case cohort included 87 nodules diagnosed as NIFTP (size range: 1-7cm, mean: 2.5cm) from 82 patients (age range: 22-82, mean age: 50.4, M:F-1:4.1). Corresponding FNA results were available for 72 nodules (82.8%) and were categorised as follows: benign (n=5, 6.9%), atypia of unknown significance/follicular lesion of undetermined significance (n=29, 40.3%), follicular neoplasm/suspicious for follicular neoplasm/follicular neoplasm with oncocytic features (n=27, 37.5%), suspicious for papillary thyroid carcinoma (n=6, 8.3%) and malignant (n=5, 6.9%). GEC results were available for 32 (44.4%) nodules, with the majority of cases classified as suspicious (81.3%). On ultrasound, most of the nodules were predominantly solid (81.8%), vascular (93.8%), non-calcified (86.5%), and either hypoechoic (44.9%) or isoechoic (38.8%). In addition to NIFTP and other benign findings in the background thyroid, 75 separate malignant tumours were identified in 38 (46.3%) patients, many of which were papillary thyroid microcarcinomas (86.5%) with lymph node metastases present in two cases. The majority of thyroid nodules histologically diagnosed as NIFTP have indeterminate cytology (77.8%) and are classified as suspicious (81.3%) by GEC testing. Taken together, these findings can guide clinicians toward a more conservative therapeutic approach.

Cost-effectiveness of Gene Expression Classifier Testing of Indeterminate Thyroid Nodules Utilizing a Real Cohort Comparator.

Objective To predict the cost-effectiveness of implementing routine gene expression classifier testing for thyroid nodules with indeterminate fine-needle aspiration cytology, by utilizing a real cohort of patients as a comparator. Study Design Cost-effectiveness analysis of a retrospective cohort compared with a simulated cohort. Setting Tertiary academic medical center. Subjects and Methods We reviewed the records of all patients who underwent ultrasound-guided fine-needle aspiration from 2010 to 2014 at a tertiary academic medical center. All patients with Bethesda class III or IV cytopathology had the details of their management catalogued over the subsequent 2 years of care. These patients were assigned to the standard-of-care arm of the study. We compared the third-party payer costs of care and the proportion of patients who underwent surgery with a simulated cohort who underwent gene expression classifier testing after an initial indeterminate fine-needle aspiration (molecular test arm). Results The cost of managing 1 nodule over 2 years was $2399 higher (range, $397-$4399) for the molecular test group than the standard of care group. The molecular test group had a 13.1% decrease (base parameters; range, 0.73%-45.09%) in the number of patients undergoing surgery. Conclusion When applied to a real cohort of patients, routine gene expression classifier is predicted to reduce the number of patients undergoing surgery but will increase cost of care. Cost-effectiveness is heavily dependent on prevalence of malignancy and gene expression classifier specificity.

The impact of noninvasive follicular thyroid neoplasm with papillary-like nuclear features on the performance of the Afirma gene expression classifier.

A recent revision in thyroid tumor nomenclature has resulted in a change from a malignant diagnosis (noninvasive follicular variant of papillary thyroid carcinoma) to one that is nonmalignant (noninvasive follicular thyroid neoplasm with papillary-like nuclear features [NIFTP]). The objective of the current study was to evaluate the impact of this change on the performance of the Afirma gene expression classifier (GEC). The authors retrospectively analyzed consecutive thyroid fine-needle aspiration specimens with indeterminate diagnoses on which GEC was performed. Surgical pathology material was reviewed with the reclassification of nodules into NIFTP. GEC testing was performed on 384 fine-needle aspiration specimens diagnosed as atypia of undetermined significance (AUS) (304 cases) and suspicious for a follicular neoplasm (SFN) (80 cases) and yielded a suspicious result in 152 of the AUS cases (50%) and 50 of the SFN cases (63%). Thyroidectomy was performed on 177 patients. After reclassifying NIFTP, the positive predictive value of GEC decreased from 42% (95% confidence interval [95% CI], 39%-45%) to 24% (95% CI, 22%-26%) in the AUS group and from 23% (95% CI, 19%-27%) to 13% (95% CI, 9%-18%) in the SFN group. Total thyroidectomy was performed more frequently than a partial thyroidectomy in patients with AUS with a suspicious GEC result compared with pre-GEC controls (68% vs 49%; P = .037). Reclassification of NIFTP significantly decreases the positive predictive value of GEC in indeterminate thyroid nodules. Nevertheless, the majority of patients with indeterminate thyroid nodules with a suspicious GEC result in the study institution have undergone total thyroidectomy. This finding raises concerns over reliance on a suspicious GEC result by clinicians to justify total thyroidectomy. Cancer Cytopathol 2017;125:683-91. © 2017 American Cancer Society.

The first Canadian experience with the Afirma\xae gene expression classifier test

BackgroundThyroid nodules are common and often benign, although prove to be malignant upon surgical pathology in 5–15% of cases. When assessed with ultrasound-guided fine-needle aspiration (USFNA), 15–30% of the nodules yield an indeterminate result. The Afirma® gene expression classifier (AGEC) was developed to improve management of indeterminate thyroid nodules (ITNs) by classifying them as “benign” or “suspicious.” Objectives were (1) to assess the performance of the AGEC in two Canadian academic medical centres (2), to search for inter-institutional variation and (3) to compare AGEC performance in Canadian versus American institutions.MethodsWe undertook a retrospective cohort study of patients with indeterminate cytopathology (Bethesda Class III or IV) as per USFNA who underwent AGEC testing. We reviewed patient demographics, cytopathological results, AGEC data and, if the patient underwent surgery, results from their final pathology.ResultsIn total, we included 172 patients with Bethesda Class III or IV thyroid nodules underwent AGEC testing, 109 in Montreal, Quebec and 63 in St. John’s, Newfoundland, in this study. Among the nodules sent for testing, 55% (60/109) in Montreal and 46% (29/63) in St. John’s returned as “benign.” None of these patients underwent surgery. On the other hand, 45% (49/109) nodules in Montreal and 54% (34/63) in St. John’s were found to be “suspicious,” for a total of 83 specimens. Seventy seven of these patients underwent surgery. Both in Montreal and St. John’s, the final pathology yielded malignant thyroid disease in approximately 50% of the specimens categorized as “suspicious.” Since 2013, no patient diagnosed with a benign nodule as per AGEC testing was found to harbor a malignant thyroid nodule on follow-up.ConclusionsMolecular analysis is increasingly used in the management of indeterminate thyroid nodules. This study highlights the experience of two Canadian centres with AGEC testing. We found inter-institutional variability in the rate of nodules returning as “benign,” however we found similar rates of confirmed malignancy in nodules returning as “suspicious.” According the literature, results for AGEC testing in two Canadian institutions align with results reported in American centres.

Qualifiers of atypia in the cytologic diagnosis of thyroid nodules are associated with different Afirma gene expression classifier results and clinical outcomes.

BACKGROUNDThyroid nodules with atypia of undetermined significance (AUS) on fine‐needle aspiration (FNA) have a low risk of malignancy that appears to vary based on specific features described in the AUS diagnosis. The Afirma gene expression classifier (GEC) is a molecular test designed to improve preoperative risk stratification of thyroid nodules, but its performance for different patterns of AUS has not been defined. The objective of this study was to assess GEC results and clinical outcomes in AUS nodules with architectural atypia (AUS‐A), cytologic atypia (AUS‐C) or both (AUS‐C/A).METHODSThis was a retrospective review of all thyroid nodules with AUS cytopathology that underwent GEC testing at the authors' institution over a period of >4 years.RESULTSIn 227 nodules that had AUS cytology results and Afirma GEC testing, the rate of benign GEC results was higher in AUS‐A nodules (70 of 107; 65%) than in AUS‐C/A nodules (25 of 65; 38%; P = .0008), and AUS‐C nodules exhibited an intermediate rate of benign results (27 of 55 nodules; 59%). The risk of cancer among patients who had GEC‐suspicious nodules, 86% of whom underwent resection, was 19% (6 of 25) for AUS‐A nodules compared with 57% (21 of 37) for AUS‐C/A nodules (P = .003) and 45% (10 of 22) for AUS‐C nodules (P = .07). In nodules that had an indeterminate repeat cytology result, no difference was observed in the rate of benign GEC results or in the malignancy rate compared with nodules that had a single cytology result.CONCLUSIONSThe performance characteristics of Afirma GEC testing vary, depending on qualifiers of cytologic atypia. Recognition of these differences may enable clinicians to provide improved counseling and treatment recommendations to patients. Cancer Cytopathol 2017;125:313–322. © 2017 American Cancer Society.

Utility of Ultrasound Versus Gene Expression Classifier in Thyroid Nodules with Atypia of Undetermined Significance

Thyroid nodules with fine-needle aspiration (FNA) cytology categorized as atypia of undetermined significance (AUS) often undergo additional diagnostic analysis with the Afirma Gene Expression Classifier (GEC), which classifies these as either high probability of being benign (GEC-B) or suspicious for malignancy (GEC-S). Our goal was to assess the clinical validity and utility of GEC in the evaluation of AUS cytology and evaluate the performance of ultrasonography (USG) for predicting malignancy in this subset. We conducted a study with a retrospective cohort of patients from January 2012 to January 2014 who had FNA of thyroid nodules >1 cm in size with AUS cytology. Cleveland Clinic Florida has an overall prevalence of AUS of 5%. A total of 119 cases with nodules >1 cm in size were reported as AUS. Forty-eight (40.3%) had a GEC performed after the first FNA (AUS-1), and 27 of these were GEC-S. Of those 27, 21 went for surgery and 14 (66.6%) had thyroid cancer on histopathology. The remaining 71 with AUS-1 were sent for a second FNA: 19 nodules were benign and did not undergo further evaluation, while the remaining 52 were reported as AUS for the second consecutive time (AUS-2). AUS-2 samples were sent for GEC. Of these 52 AUS-2, 38 (73.1%) were reported as GEC-S. Thirty-five went for surgery and 32 (91.4%) had confirmed malignancy on histopathology. Positive predictive value (PPV) was 91.4% for AUS-2 and 66.6% for AUS-1. Moreover, AUS-2 nodules that were hypoechoic and solid on USG showed a PPV of 92% for malignancy. In our practice, the diagnostic accuracy to predict malignancy with GEC for AUS-1 nodules was poor (PPV, 66.6%). The PPV of GEC testing was markedly higher at 91.4% performed after two consecutive AUS cytologies. AUS-2 nodules that were solid and hypoechoic on USG also had a high probability to be malignant (PPV, 92%). We recommend repeat FNA on AUS-1 nodules rather than proceeding directly to GEC testing. Also, we suggest that among AUS-2 nodules, surgery can be recommended when USG shows solid and hypoechoic features with GEC testing reserved for the remainder. AUS = atypia of undetermined significance FNA = fine-needle aspiration GEC = gene expression classifier GEC-B = GEC-benign GEC-S = GEC-suspicious for malignancy NPV = negative predictive value PPV = positive predictive value USG = ultrasonography.