Context The Afirma® GSC aids in risk stratifying indeterminate thyroid nodule cytology (ITN). The 2018 GSC validation study (VS) reported a sensitivity (SN) of 91%, specificity (SP) of 68%, positive predictive value (PPV) of 47%, and negative predictive value (NPV) of 96%. Since then, 13 independent real world (RW) post-validation studies have been published. Objective This study’s objective is to compare the RW GSC performance to the VS metrics. Methods Rules and assumptions applying to this analysis include: 1. At least one patient with molecular benign results must have surgery for that study to be included in SN, SP and NPV analyses. 2. Molecular benign results without surgical histology are considered true negatives (TN) (as are the molecular benign results with benign surgical histology) 3. Unoperated patients with suspicious results are either excluded from the analysis (observed PPV (oPPV) and observed SP (oSP)) or assumed as histology negatives (false positives - conservative PPV (cPPV) and conservative SP (cSP)) 4. NIFTP is considered malignant. Results In RW studies, the GSC demonstrates a SN, oSP, oPPV and NPV of 97%, 88%, 65%, 99% respectively, and conservative RW performance showed cSP of 80% and cPPV of 49%, all significantly higher than the VS save for SN and cPPV. There was also a higher benign call rate (BCR) of 67% in RW studies compared to 54% in the VS (p < 0.05). Conclusion RW data for the Afirma GSC demonstrates significantly better oSP and oPPV performance compared to the VS, indicating an increased yield of cancers for resected GSC suspicious nodules. The higher BCR likely increases the overall rate of clinical observation in lieu of surgery.

Background Analysis of cytologically indeterminate thyroid nodules with Afirma Gene Expression Classifier (GEC) and Genomic Sequencing Classifier (GSC) can reduce surgical rate and increase malignancy rate of surgically resected indeterminate nodules. Methods Retrospective cohort analysis of all adults with cytologically indeterminate thyroid nodules from January 2013 through December 2019. We compared surgical and malignancy rates of those without molecular testing to those with GEC or GSC, analyzed test performance between GEC and GSC, and identified variables associated with molecular testing. Results 468 indeterminate thyroid nodules were included. No molecular testing was performed in 273, 71 had GEC, and 124 had GSC testing. Surgical rate was 68% in the group without molecular testing, 59% in GEC, and 40% in GSC. Malignancy rate was 20% with no molecular testing, 22% in GEC, and 39% in GSC (p = 0.022). GEC benign call rate (BCR) was 46%, sensitivity 100%, specificity 61% and PPV 28%. GSC BCR was 60%, sensitivity 94%, specificity 76%, and PPV 41%. Those with no molecular testing had larger nodule size, pre-operative growth of nodules, and constrictive symptoms, and those who underwent surgery in the no molecular testing group had higher BMI, constrictive symptoms, higher TIRADS and Bethesda classification. Type of provider was also associated with the decision to undergo surgery. Conclusion Implementation of GEC showed no effect on surgical or malignancy rate, but GSC resulted in significantly lower surgical and higher malignancy rates. This study provides insight into the factors that affect the real- world use of these molecular markers preoperatively in indeterminate thyroid nodules.

The Afirma GSC aids in the clinical decision making for patients with indeterminate thyroid nodule cytology (ITN). The 2018 GSC validation study was a prospective, multi-center study, conducted on a patient cohort with ITN. All patients underwent surgery without known genomic information and were assigned a histopathology diagnosis by an expert panel blinded to all genomic information. The results showed a sensitivity (SN) of 91%, specificity (SP) of 68%, positive predictive value (PPV) of 47%, and negative predictive value (NPV) of 96% at a cancer prevalence of 24%. Since then, 13 independent GSC post-validation studies have been published. This study's objective is to compare the real world (RW) GSC performance to the validation study metrics. Rules and assumptions applying to this analysis include: 1. At least one patient with molecular benign results must have surgery for that study to be included in SN, SP and NPV analysis and in these studies, molecular benign results without surgical histology are considered true negatives (TN) (as are the molecular benign results with benign surgical histology) 2. Patients with suspicious results that do not have surgery are either excluded from the analysis (generating an observed PPV (oPPV) and observed SP (oSP)) or assumed as histology negatives (false positives - generating a conservative PPV (cPPV) and conservative SP (cSP)) 3. NIFTP is considered malignant. Rule #1 excluded two studies from SN, SP and NPV analysis. Data from all studies were pooled using a random-effects model. All analyses were done using R package meta (version 4.18-2). In the RW, the GSC demonstrates a SN, oSP, oPPV and NPV of 97%, 88%, 65%, 99% respectively, and conservative RW performance of cSP at 80% and cPPV at 49%. A statistically significant improvement is observed for oSP, cSP, oPPV, and NPV (p<0. 05) relative to the validation study with no statistical difference in SN. An overrepresentation of Hurthle subtypes in the validation study (20% Hürthle carcinoma (HCC) of malignant histology and 11% Hürthle adenoma of benign histology) relative to RW estimates of HCC prevalence (<5%) may partly explain improved RW GSC performance (as well as other histology subtype differences). Additionally, there may be an enrichment of malignancy in the operated cohort based upon the selection of patients with GSC-S nodules that have surgery (higher clinical risk or more worrisome ultrasound features compared to GSC-S nodules without surgical follow-up). The high benign call rate, predicted by the higher cSP and oSP, likely increases the overall rate of clinical observation in lieu of surgery. The high oPPV indicates an increased yield of cancers for resected GSC-S lesions relative to the validation study (65% vs 47%). In summary, RW GSC data indicates significantly better performance on several metrics as compared to the validation study, most notably on cSP, oSP and oPPV. Presentation: Saturday, June 11, 2022 1:00 p.m. - 3:00 p.m.