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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) postvalidation 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 1 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 molecular benign results with benign surgical histology). (3) Unoperated patients with suspicious results are either excluded from analysis (observed PPV [oPPV] and observed SP [oSP]) or assumed histology negatives (false positives; conservative PPV [cPPV] and conservative SP [cSP]) 4. Noninvasive follicular thyroid neoplasm with papillary-like nuclear features 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 except 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 than 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: Cytopathological evaluation of thyroid fine-needle aspiration biopsy (FNAB) specimens can fail to raise preoperative suspicion of medullary thyroid carcinoma (MTC). The Afirma RNA-sequencing MTC classifier identifies MTC among FNA samples that are cytologically indeterminate, suspicious, or malignant (Bethesda categories III-VI). In this study we report the development and clinical performance of this MTC classifier. Methods: Algorithm training was performed with a set of 483 FNAB specimens (21 MTC and 462 non-MTC). A support vector machine classifier was developed using 108 differentially expressed genes, which includes the 5 genes in the prior Afirma microarray-based MTC cassette. Results: The final MTC classifier was blindly tested on 211 preoperative FNAB specimens with subsequent surgical pathology, including 21 MTC and 190 non-MTC specimens from benign and malignant thyroid nodules independent from those used in training. The classifier had 100% sensitivity (21/21 MTC FNAB specimens correctly called positive; 95% confidence interval [CI] = 83.9-100%) and 100% specificity (190/190 non-MTC FNAs correctly called negative; CI = 98.1-100%). All positive samples had pathological confirmation of MTC, while all negative samples were negative for MTC on surgical pathology. Conclusions: The RNA-sequencing MTC classifier accurately identified MTC from preoperative thyroid nodule FNAB specimens in an independent validation cohort. This identification may facilitate an MTC-specific preoperative evaluation and resulting treatment.

Despite its wide-ranging benefits, whole-transcriptome or RNA exome profiling is challenging to implement in a clinical diagnostic setting. The Unified Assay is a comprehensive workflow wherein exome-enriched RNA-sequencing (RNA-Seq) assays are performed on clinical samples and analyzed by a series of advanced machine learning-based classifiers. Gene expression signatures and rare and/or novel genomic events, including fusions, mitochondrial variants, and loss of heterozygosity were assessed using RNA-Seq data generated from 120,313 clinical samples across three clinical indications (thyroid cancer, lung cancer, and interstitial lung disease). Since its implementation, the data derived from the Unified Assay have allowed significantly more patients to avoid unnecessary diagnostic surgery and have played an important role in guiding follow-up decisions regarding treatment. Collectively, data from the Unified Assay show the utility of RNA-Seq and RNA expression signatures in the clinical laboratory, and their importance to the future of precision medicine.

Context Broad genomic analyses among thyroid histologies have been described from relatively small cohorts. Objective Investigate the molecular findings across a large, real-world cohort of thyroid fine-needle aspiration (FNA) samples. Design Retrospective analysis of RNA sequencing data files. Setting Clinical Laboratory Improvement Amendments laboratory performing Afirma Genomic Sequencing Classifier (GSC) and Xpression Atlas (XA) testing. Participants A total of 50 644 consecutive Bethesda III-VI nodules. Intervention None. Main Outcome Measures Molecular test results. Results Of 48 952 Bethesda III/IV FNAs studied, 66% were benign by Afirma GSC. The prevalence of BRAF V600E was 2% among all Bethesda III/IV FNAs and 76% among Bethesda VI FNAs. Fusions involving NTRK, RET, BRAF, and ALK were most prevalent in Bethesda V (10%), and 130 different gene partners were identified. Among small consecutive Bethesda III/IV sample cohorts with one of these fusions and available surgical pathology excision data, the positive predictive value of an NTRK or RET fusion for carcinoma or noninvasive follicular thyroid neoplasm with papillary-like nuclear features was >95%, whereas for BRAF and ALK fusions it was 81% and 67%, respectively. At least 1 genomic alteration was identified by the expanded Afirma XA panel in 70% of medullary thyroid carcinoma classifier–positive FNAs, 44% of Bethesda III or IV Afirma GSC suspicious FNAs, 64% of Bethesda V FNAs, and 87% of Bethesda VI FNAs. Conclusions This large study demonstrates that almost one-half of Bethesda III/IV Afirma GSC suspicious and most Bethesda V/VI nodules had at least 1 genomic variant or fusion identified, which may optimize personalized treatment decisions.

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 Presentation: Saturday, June 11, 2022 1:00 p.m. - 3:00 p.m.

Item does not contain fulltext Background: The American Thyroid Association (ATA), the European Association of Nuclear Medicine, the European Thyroid Association, and the Society of Nuclear Medicine and Molecular Imaging have established an intersocietal working group to address the current controversies and evolving concepts in thyroid cancer management and therapy. The working group annually identifies topics that may significantly impact clinical practice and publishes expert opinion articles reflecting intersocietal collaboration, consensus, and suggestions for further research to address these important management issues. Summary: In 2019, the intersocietal working group identified the following topics for review and interdisciplinary discussion: (i) perioperative risk stratification, (ii) the role of diagnostic radioactive iodine (RAI) imaging in initial staging, and (iii) indicators of response to RAI therapy. Conclusions: The intersocietal working group agreed that (i) initial patient management decisions should be guided by perioperative risk stratification that should include the eighth edition American Joint Committee on Cancer staging system to predict disease specific mortality, the modified 2009 ATA risk stratification system to estimate structural disease recurrence, with judicious incorporation of molecular theranostics to further refine management recommendations; (ii) diagnostic RAI scanning in ATA intermediate risk patients should be utilized selectively rather than being considered mandatory or not necessary for all patients in this category; and (iii) a consistent semiquantitative reporting system should be used for response evaluations after RAI therapy until a reproducible and clinically practical quantitative system is validated.

Background Preoperative diagnosis of medullary thyroid carcinoma (MTC) is often difficult, given the poor sensitivity of fine-needle aspiration (FNA) cytology for MTC. This study investigates this issue and presents recommendations for improving preoperative diagnostic paradigms in MTC cases. Design/method Histopathologically confirmed MTC patients with preoperative cytologic assessment of index nodules were enrolled. FNA diagnosis, final pathology, and surgery details were collected. Results Out of 71 patients, 49 (69%) were diagnosed by FNA as either definitive MTC (35, 49%) or suspected MTC (14, 20%) and 22 (31%) patients had no indication of MTC on FNA. Conclusion In a tertiary-care setting, one-third of subjects had an FNA interpretation that did not suggest the possibility of MTC. The limitations of preoperative diagnosis are especially problematic for MTC as they can cause delayed or incomplete treatment. Additional testing is proposed to improve preoperative diagnosis and surgical care of MTC patients.

Background: Gene panels are routinely used to assess predisposition to hereditary cancers by simultaneously testing multiple susceptibility genes and/or variants. More recently, genetic panels have been implemented as part of solid tumor malignancy testing assessing somatic alterations. One example is targeted variant panels for thyroid nodules that are not conclusively malignant or benign upon fine-needle aspiration (FNA). We systematically reviewed published studies from 2009 to 2018 that contained genetic data from preoperative FNA specimens on cytologically indeterminate thyroid nodules (ITNs) that subsequently underwent surgical resection. Pooled prevalence estimates per gene and variant, along with their respective positive predictive values (PPVs) for malignancy, were calculated. Summary: Our systematic search identified 540 studies that were supplemented by 18 studies from bibliographies or personal files. Sixty-one studies met all inclusion criteria and included >4600 ITNs. Overall, 26% of nodules contained at least 1 variant or fusion. However, half of them did not include details on the specific gene, variant, and/or complete fusion pair reported for inclusion toward PPV calculations. The PPVs of genomic alterations reported at least 10 times were limited to BRAFV600E (98%, 95% confidence interval [CI 96–99%]), PAX8/PPARG (55% [CI 34–78%]), HRASQ61R (45% [CI 22–72%]), BRAFK601E (42% [CI 19–68%]), and NRASQ61R (38% [CI 23–55%]). Excluding BRAFV600E, the pooled PPV for all other specified variants and fusions was 47%. Multiple variants within the same nodule were identified in ∼1% of ITN and carried a cumulative PPV of 77%. Conclusions: The chance that a genomic alteration predicts malignancy depends on the individual variant or fusion detected. Only five alterations were reported at least 10 times; BRAFV600E had a PPV of 98%, while the remaining four had individual PPVs ranging from 38% to 55%. The small sample size of most variants and fusion pairs found among ITNs, however, limits confidence in their individual PPV point estimates. Better specific reporting of genomic alterations with cytological category, histological subtype, and cancer staging would facilitate better understanding of cancer prediction, and the independent contribution of the genomic profile to prognosis.

The Afirma Genomic Sequencing Classifier (GSC) is a rule‐out test for malignancy/noninvasive follicular thyroid neoplasms with papillary‐like nuclear features among patients with Bethesda category III/IV nodules, whereas the complimentary Xpression Atlas provides genomic insights from a curated panel of 511 genes among GSC suspicious and Bethesda category V/VI nodules. Together, they facilitate personalized treatment decisions based on genomic insights derived from the transcriptome of the biopsied target and extend the diagnostic and therapeutic reach of cytopathologists and fine‐needle aspiration biopsy sample collection.

BACKGROUND. Current recommendations for persistent or recurrent locoregional papillary thyroid cancer (PTC) include consideration of surgical resection versus active surveillance. The purpose of this study is to determine long-term outcomes after surgical resection of recurrent or persistent metastatic PTC in cervical lymph nodes after failure of initial surgery and radioactive iodine therapy using newer validated clinical outcomes measures. METHODS. Outcomes of 70 patients who underwent cervical lymphadenectomy (n = 110) from 1999 to 2013 for recurrent or persistent locoregional PTC metastases were reviewed. Measures included biochemical remission (BCR) based on Tg levels, American Thyroid Association classifications for response to treatment [biochemical incomplete response (BIR), structural incomplete response (SIR), indeterminate response (IR), and excellent response (ER)], need for reoperation, surgical complications, disease progression, and death. RESULTS. The median follow-up was 13.1 years, with only two additional reoperations since 2010, one of which had no metastasis on pathology with the other developing anaplastic thyroid cancer in background PTC. ER was achieved in 31 (44%) patients, all of whom remained in ER at time of last follow-up (median 14.1 years). There were no structural recurrences in patients with persistent BIR or IR after reoperation. Patients with SIR had stable disease, except for one who died due to anaplastic thyroid cancer. CONCLUSIONS. Patients who achieved ER after reoperation had no need for further treatment. Patients with persistent detectable Tg levels after reoperation rarely developed structural recurrence. ATA outcomes can be safely used to guide treatment decisions over a decade after reoperation for PTC.

Our understanding of the genomics of benign and malignant thyroid nodules has advanced rapidly, and so have the applications that utilize this information for clinical decision-making. Not long ago, it was “cutting-edge” medicine to test for a single genomic variant. Now, everyday practitioners have access to gene panels and genomic classifiers that utilize hundreds and thousands of genes. Here, we discuss current cytological nomenclature of thyroid aspirates, review cellular genomics, discuss current genomic tests for cytologically indeterminate nodules, consider surgical decision-making, explore the potential role of genomics among suspicious and malignant cytology categories, and consider future directions.

To determine the association between pathologic features and molecular classes (BRAF-like, RAS-like, and non-BRAF-like non-RAS-like [NBNR]).Retrospective review of a merged database containing 676 patients, 84% (571/676) were assigned to a molecular class from publicly accessible sequenced data of thyroid neoplasms.The merged cohort included 571 neoplasms: 353 (62%) BRAF-like, 172 (30%) RAS-like, and 46 (8.1%) NBNR. Lymph node metastasis (any N1 disease) was present in 166/337 (49%) of BRAF-like, 23/164 (14%) of RAS-like, and 0/46 (0%) of NBNR and are significantly different (P.001). Gross extra-thyroidal extension was observed in 27 patients, including 24/331 (7%) of BRAF-like, 2/160 (1%) of RAS-like, and 1/46 (2%) of NBNR (P = .01). N1B lymph node metastases or T4 disease was present in 74/333 (22%) of BRAF-like, 10/160 (6%) of RAS-like, and 1/46 (2%) of NBNR (P.0001). Distant metastasis was present in 4/151 (2.6%) of BRAF-like, 2/50 (4%) of RAS-like and 0/46 for NBNR (P = .627). Angioinvasion was present in 0/81 (0%) of BRAF-like, 3/53 (6%) of RAS-like, and 3/46 (7%) of NBNR (P = .08); and multifocality was present in 27/81 (33%) of BRAF-like, 9/53 (17%) of RAS-like, and 1/46 (2%) for NBNR (P = .0001).Pathological features of metastasis, gross extra-thyroidal extension, and multifocality were more prevalent in BRAF-like samples compared to RAS-like and NBNR. A trend towards increased frequency of angioinvasion in RAS-like and NBNR cancers compared to BRAF-like samples was observed. Further studies are needed to evaluate if preoperative knowledge of molecular mutations in thyroid tumors aids in decision-making regarding extent of surgery.

Introduction: Receptor tyrosine kinases (RTKs) initiate signaling cascades, including growth and differentiation. Activation can occur through chromosomal rearrangements that lead to gene fusions. RTK fusions are potential targets for small molecule inhibitors to treat advanced cancers. The original Afirma Xpression Atlas (XA) reported 761 selected variants and 130 fusion pairs in Bethesda III/IV Afirma Genomic Sequencing Classifier (GSC) suspicious or Bethesda V/VI nodules. The landscape of additional potentially actionable gene fusions has not been explored in treatment-naïve patients. Methods: Anonymized RNA-seq data from >37,000 Bethesda III-VI samples were examined with STAR-fusion to determine gene/gene fusions. All samples were examined for NTRK1, NTRK3, RET, ALK, and BRAF fusions, regardless of fusion partner. Fusions were evaluated for being in-frame, with an intact kinase domain at the 3’ end of the fusion pair. Fusion pairs not currently reported by XA and not reported in thyroid TCGA fusion data are denoted “additional”. All fusion pairs were searched for in the literature and public fusion databases. Results: Examining the Veracyte clinical database revealed 7 additional NTRK1/3 fusions, with 3 NTRK fusions observed more than once - SQSTM1/NTRK3, VIM/NTRK3, and EML4/NTRK3. One of the 7 NTRK fusions had not been previously reported. Eight additional ALK fusions were identified, with 4 observed more than once- ITSN2/ALK, PPP1R21/ALK, PDE8B/ALK, NPAT/ALK. Five of these 8 ALK fusions had not been previously described. Seventeen additional RET fusions were identified, with 5 observed recurrently - KIAA1217/RET, AFAP1L2/RET, ACBD5/RET, SQSTM1/RET, and TFG/RET. Six of the 17 RET fusions had not been previously reported. Seventy-two additional BRAF fusions were identified, and 58 of them have not been previously reported. Eight of the 72 BRAF fusions were observed more than once. Examining >50,000 Afirma samples, NTRK1, NTRK3, RET, ALK, or BRAF fusions were not identified among the Afirma GSC Benign, and were present in 3.2% of 16,594 Bethesda III/IV Afirma GSC Suspicious samples, and 8.0% of 1,692 Bethesda V/VI samples. Correlation with surgical histology is unknown. Conclusions: By examining a large cohort of patients with an unbiased, whole-transcriptome RNA-seq assay, we identified potentially actionable kinase fusions in thyroid nodules beyond those described in TCGA. All fusions described here are either novel and not previously reported, rarely reported in one or two case studies, or not described in thyroid cancers. Additional NTRK, ALK, RET and BRAF fusions were found, all of which may be targeted with specific kinase inhibitors currently available. Future studies may determine genotype-phenotype correlations regarding the natural history of these neoplasms. Because of the potential clinical implications of these genomic markers for patient management, all 104 fusions described here are now included among the 235 gene pairs reported by the expanded Afirma XA.

Nuclear medicine is an essential tool in the delivery of high-quality medical care, going beyond simple anatomic imaging to the use of physiologic processes for both imaging and therapy. One of the first uses of nuclear medicine techniques occurred in the 1930s to demonstrate the physiology of

RAS mutations are common in the available mutational analysis of cytologically indeterminate (Cyto-I) thyroid nodules. However, their reported positive predictive value (PPV) for cancer is widely variable. The reason for this variability is unknown, and it causes clinical management uncertainty. A systematic review was performed, evaluating the PPV for cancer in RAS mutation positive Cyto-I nodules, and variables that might affect residual heterogeneity across the different studies were considered.PubMed was searched through February 22, 2017, including studies that evaluated at least one type of RAS mutation in Cyto-I nodules, including any (or all) of the Bethesda III/IV/V categories or their equivalents and where the histological diagnosis was available. The PPV residual heterogeneity was investigated after accounting for Bethesda classification, blindedness of the histopathologist to the RAS mutational status, Bethesda category-specific cancer prevalence for each study, and which RAS genes and codons were tested. This was studied using five meta-regression models fit to different sets of Bethesda classification categories: Bethesda III, IV, or V (III/IV/V); Bethesda III or IV (III/IV); Bethesda III only; Bethesda IV only; and Bethesda V only.Of 1831 studies, 23 were eligible for data inclusion. Wide ranges of PPV were found at 0-100%, 28-100%, and 0-100% in Bethesda III, IV, and V, respectively. Residual heterogeneity remained moderately high for PPV after accounting for the above moderators for Bethesda III/IV/V (21 studies; IThe PPV of RAS mutations in Bethesda III and IV categories is quite heterogeneous across different studies, creating low confidence in the accuracy of a single estimate of PPV. Clinicians must appreciate this wide variability when managing a RAS-mutated Cyto-I nodule. Future studies should seek to resolve this unexplained variability.

Background Glucagon‐like peptide‐1 receptor agonists, such as dulaglutide, exenatide and liraglutide, are approved to treat Type 2 diabetes mellitus. Although these drugs provide substantial glycaemic control, studies in rodents have prompted concerns about the development of medullary thyroid carcinoma. These data are reflected in the US package insert, with boxed warnings and product labelling noting the occurrence of these tumours after clinically relevant exposures in rodents, and contraindicating glucagon‐like peptide‐1 receptor agonist use in people with a personal or family history of medullary thyroid carcinoma, or in people with multiple endocrine neoplasia type 2. However, there are substantial differences between rodent and human responses to glucagon‐like peptide‐1 receptor agonists. This report presents the case of a woman with pre‐existing medullary thyroid carcinoma who exhibited no significant changes in serum calcitonin levels despite treatment with dulaglutide 2.0 mg for 6 months in the Assessment of Weekly AdministRation of LY2189265 [dulaglutide] in Diabetes‐5 clinical study (NCT00734474). Case report Elevated serum calcitonin was noted in a 56‐year‐old woman with Type 2 diabetes mellitus at the 6‐month discontinuation visit in a study of long‐term dulaglutide therapy. Retroactive assessment of serum collected before study treatment yielded an elevated calcitonin level. At 3 months post‐study, calcitonin level remained elevated; ultrasonography revealed multiple bilateral thyroid nodules. Eventually, medullary thyroid carcinoma was diagnosed; the woman was heterozygous positive for a germline RET proto‐oncogene mutation. Conclusion The tumour was not considered stimulated by dulaglutide therapy because calcitonin remained stable throughout., What's new? A woman with an unrecognized pre‐existing medullary thyroid carcinoma (MTC) who received a glucagon‐like peptide‐1 receptor agonist (GLP‐1RA) in a clinical trial provides a possibly unique case in the GLP‐1RA drug development literature.A lack of both serum calcitonin stimulation and functional glucagon‐like peptide‐1 receptor in this MTC was observed.

6083 Background: Receptor tyrosine kinase (RTK) fusions may be targeted by small molecule inhibitors to treat various advanced tumors, including thyroid cancer. Clinical trials have studied selective inhibitors of ALK, BRAF, NTRK and RET, leading to several FDA-approved therapies. The Afirma Genomic Sequencing Classifier (GSC) classifies cytologically indeterminate thyroid nodules as molecularly benign or suspicious. The Xpression Atlas reports 905 genomic variants and 235 fusion pairs on GSC Suspicious, Suspicious for Malignancy (SFM), and Malignant FNA samples at the time of diagnosis. Here we report the prevalence of these fusion genes in real-world clinical practice. Methods: We analyzed anonymized data from 50,644 consecutive Bethesda III-VI nodule FNA samples submitted to the Veracyte CLIA laboratory for molecular testing using whole transcriptome RNA sequencing (RNA-Seq). Gene pairs are listed alphabetically. Results: 32,080 Bethesda III/IV nodules were classified as GSC Benign and 278 were Parathyroid Classifier positive. No ALK, BRAF, NTRK1/3, or RET fusions were identified among these samples. Among 16,594 Bethesda III/IV GSC Suspicious FNAs, 3% (n = 529) were positive for ALK, BRAF, NTRK1/3 or RET fusions. Among the 1,692 Bethesda V/VI FNAs, the proportion of positive nodules was 8% (n = 135). Among these combined cohorts of Bethesda III/IV GSC Suspicious and Bethesda V/VI, the most common gene fusions observed for each of the 5 studied RTK genes was: ETV6/NTRK3 (n = 164, 72% of NTRK3 fusions), CCDC6/RET (n = 104, 55% of RET), BRAF/SND1 (n = 32, 20% of BRAF), ALK/STRN (n = 20, 37% of ALK), and NTRK1/TPM3 (n = 14, 50% of NTRK1). BRAF showed the highest diversity of fusions, with 80 gene partners. Different gene partners with RET, ALK, NTRK1, and NTRK3 numbered 25, 11, 9, and 5 , respectively . Conclusions: Whole-transcriptome RNA-seq on small sample thyroid FNA specimens can identify clinically relevant ALK, BRAF, NTRK, and RET fusions across Bethesda categories. The prevalence ranges from 3% in Bethesda III/IV Afirma GSC Suspicious specimens to 8% among Bethesda V/VI specimens. Future studies need to determine if detection of precision medicine candidates by pre-operative FNA can optimize initial treatment, predict response to treatment, and prioritize selective targeted therapy should systemic treatment be needed.[Table: see text]

Thyroid fine-needle aspiration biopsy results are cytologically indeterminate in 15-30% of cases. When these nodules undergo diagnostic surgery, approximately three-quarters are histologically benign. These unnecessary surgeries diminish quality of life, generate complications, and increase healthcare costs. The Afirma gene expression classifier (GEC) is validated to pre-operatively identify cytologically indeterminate nodules likely to be truly benign so that surgery can be avoided. Its performance is supported by robust multicenter prospective and blinded clinical validation studies, and supported by extensive independent clinical utility publications which show a marked reduction in surgery among patients with benign Afirma GEC results. To rule-out cancer and avoid unnecessary diagnostic surgery, Afirma's quality and depth of validation stand alone. The accuracy of a benign result is the negative predictive value (NPV). Afirma achieves an NPV ≥94% among cytologically indeterminate nodules (Bethesda III or IV). Thirteen clinical utility studies describing 1468 GEC benign patients demonstrate that few Afirma GEC benign nodules undergo surgery, including after 3 years of follow-up. With a specificity of 52%, over half of the truly benign nodules with indeterminate cytology receive a benign GEC result. High test sensitivity is critical to safely rule out cancer. The Afirma GEC's 90% sensitivity means that regardless of the pre-test risk of malignancy, 90% of all malignant nodules are GEC suspicious. The Afirma GEC has transformed patient care. Where the majority of cytologically indeterminate patients were once operated to determine if the nodule was benign or malignant, now nearly half of these surgeries can be avoided.Belirsiz sitoloji tiroid ince iğne biyopsi örneklerinin %15-30’unda bulunur. Bu nodüller tanı amaçlı ameliyat edildiğinde yaklaşık dörtte üçünün histolojik olarak benign olduğu saptanmıştır. Gereksiz ameliyatlar yaşam kalitesini azaltır, komplikasyonlara neden olur ve sağlık giderlerini artırır. Afirma gen ekspresyonu sınıflandırıcı [Gene Expression Classifier (GEC)] operasyon öncesi benign olma olasılığı olan belirsiz sitolojili nodülleri tespit ederek cerrahi girişim gerekliliğini önleyebilir. Testin etkinliği güvenilir çok merkezli prospektif kör klinik doğrulama çalışmalarıyla, benign Afirma GEC sonuçları ile bu hasta grubunda cerrahinin belirgin bir azalma gösterdiği ise geniş, bağımsız klinik yarar yayınları ile desteklenmiştir. Kanseri ekarte etmek ve gereksiz tanısal cerrahiyi önlemek amacıyla Afirma, kalitesi ve doğrulama oranı ile alanında tektir. Benign bir sonucun doğruluğu testin negatif prediktif değerini (NPD) gösterir. Afirma’nın NPD’yi belirsiz sitolojili nodüller için (Bethesda III veya IV) ≥%94’e ulaşmaktadır. GEC benign 1,468 hastayı içeren on üç klinik yarar çalışması, sadece birkaç Afirma GEC benign nodülde ameliyat ihtiyacı olduğunu 3 yıllık takip süresi ile göstermektedir. Testin %52 özgüllüğü ile, gerçekten benign olan belirsiz sitolojili nodüllerin yarısından fazlasında benign GEC sonucu alınmaktadır. Kanseri güvenle ekarte etmek için yüksek test duyarlılığı kritik öneme sahiptir. Afirma GEC’nin %90 duyarlılığı, test incesi malignite riski ne olursa olsun, tüm malign nodüllerin %90’ının GEC şüpheli olduğu anlamına gelir. Afirma GEC hasta yaklaşımını değiştirmiştir. Belirsiz sitolojili hastaların çoğunluğu nodülün iyi veya kötü huylu olup olmadığını belirlemek için ameliyat edilirken, günümüzde bu ameliyatların neredeyse yarısı önlenebilir.

Context: Sialadenitis and xerostomia are major adverse effects of 131I therapy in thyroid cancer patients. The risk factors for these adverse effects, other than administered activity of 131I, have not been investigated. Objective: The aim of this study is to identify risk factors for 131I-induced salivary gland damage among follicular cell-derived thyroid cancer patients. Design: We enrolled 216 thyroid cancer patients who visited The Ohio State University Wexner Medical Center between April 2013 and April 2014. Symptoms of xerostomia and sialadenitis were identified via questionnaire and medical record search. To validate the findings in a large cohort, we retrospectively searched for ICD-9/10 codes for sialadenitis, xerostomia, and autoimmune disease associated with Sjogren's syndrome (AID-SS) in our existing database (n = 1507). Demographic and clinical information was extracted from medical records. Multivariate analyses were performed to identify independent predictors for salivary gland damage. Res...

The aim of this study was to demonstrate the analytical validity of an RNA classifier for medullary thyroid carcinoma (MTC).Fresh-frozen tissue specimens were obtained from commercial sources, and MTC diagnoses were confirmed by histopathology review. De-identified patient fine-needle aspiration biopsies (FNABs) and whole blood from normal donors were obtained. Total RNA was extracted, amplified, and hybridized to custom microarrays for gene expression analysis. Gene expression data were normalized and classified via a machine learning algorithm. Positive control materials were produced from MTC tissues and tested across multiple experiments and laboratories. Twenty-seven MTC tissue specimens were used to evaluate the sensitivity of the MTC classifier. Gene expression data from tissues and FNABs were used to model classifier response to mixtures of MTC samples with normal thyroid tissue, a benign thyroid nodule, a Hürthle cell adenoma, and whole blood. Select mixture conditions were confirmed in vitro. Assay tolerance to RNA input variation (5-25 ng) and genomic DNA contamination (30% by mass) was evaluated. The intra- and inter-run reproducibility and inter-laboratory accuracy of MTC classifier results were characterized.The MTC classifier sensitivity of 96.3% [confidence interval 81.0-99.9%] was determined retrospectively using 27 MTC confirmed tissue specimens. One false-negative result in a necrotic tissue implicated sample necrosis in reduced classifier sensitivity. Dilution modeling of MTC samples with normal or benign tissues showed consistent detection of MTC down to 20% sample proportions, with in vitro confirmation of 20% analytical sensitivity. Classifier tolerance to RNA input variation (5-25 ng), genomic DNA contamination (30% by mass), and an interfering substance (blood) was demonstrated with 100% accurate classifier results under all tested conditions. The maximum observed run-to-run score difference for a single FNAB sample was ∼1 unit compared with the average score difference between 38 MTC and non-MTC FNABs of ∼32 units. MTC classifier results for 20 tissues processed from total RNA in two different laboratories showed 100% concordance.The MTC classifier, offered as part of the routine molecular testing of cytology-indeterminate thyroid nodules, demonstrates robust analytical sensitivity, specificity, accuracy, and reproducibility.

Introduction: The Afirma® Xpression Atlas (XA) detects gene variants and fusions in thyroid nodule FNA samples from a curated panel of 511 genes using whole-transcriptome RNA-sequencing. Its intended use is among cytologically indeterminate nodules that are Afirma GSC suspicious, Bethesda V/VI nodules, or known thyroid metastases. Here we report its analytical and clinical validation. Methods: DNA and RNA were purified from the same sample across 943 blinded FNAs and compared by multiple methodologies, including whole-transcriptome RNA-seq, targeted RNA-seq, and targeted DNA-seq. An additional 695 blinded FNAs were used to define performance for fusions between whole-transcriptome RNA-seq and targeted RNA-seq. We quantified the reproducibility of the whole-transcriptome RNA-seq assay across laboratories and reagent lots. Finally, variants and fusions were compared to histopathology results. Results: Of variants detected in DNA at 5 or 20% variant allele frequency, 74 and 88% were also detected by XA, respectively. XA variant detection was 89% when compared to an alternative RNA-based detection method. Low levels of expression of the DNA allele carrying the variant, compared with the wild-type allele, was found in some variants not detected by XA. 82% of gene fusions detected in a targeted RNA fusion assay were detected by XA. Conversely, nearly all variants or fusions detected by XA were confirmed by an alternative method. Analytical validation studies demonstrated high intra-plate reproducibility (89%-94%), inter-plate reproducibility (86-91%), and inter-lab accuracy (90%). Multiple variants and fusions previously described across the spectrum of thyroid cancers were identified by XA, including some with approved or investigational targeted therapies. Among 190 Bethesda III/IV nodules, the sensitivity of XA as a standalone test was 49%. Conclusion: When the Afirma Genomic Sequencing Classifier (GSC) is used first among Bethesda III/IV nodules as a rule-out test, XA supplements genomic insight among those that are GSC suspicious. Our data clinically and analytically validate XA for use among GSC suspicious, or Bethesda V/VI nodules. Genomic information provided by XA may inform clinical decision-making with precision medicine insights across a broad range of FNA sample types encountered in the care of patients with thyroid nodules and thyroid cancer.

Background Identification of Hürthle cell cancers by non-operative fine-needle aspiration biopsy (FNAB) of thyroid nodules is challenging. Resultingly, non-cancerous Hürthle lesions were conventionally distinguished from Hürthle cell cancers by histopathological examination of tissue following surgical resection. Reliance on histopathological evaluation requires patients to undergo surgery to obtain a diagnosis despite most being non-cancerous. It is highly desirable to avoid surgery and to provide accurate classification of benignity versus malignancy from FNAB preoperatively. In our first-generation algorithm, Gene Expression Classifier (GEC), we achieved this goal by using machine learning (ML) on gene expression features. The classifier is sensitive, but not specific due in part to the presence of non-neoplastic benign Hürthle cells in many FNAB. Results We sought to overcome this low-specificity limitation by expanding the feature set for ML using next-generation whole transcriptome RNA sequencing and called the improved algorithm the Genomic Sequencing Classifier (GSC). The Hürthle identification leverages mitochondrial expression and we developed novel feature extraction mechanisms to measure chromosomal and genomic level loss-of-heterozygosity (LOH) for the algorithm. Additionally, we developed a multi-layered system of cascading classifiers to sequentially triage Hürthle cell-containing FNAB, including: 1. presence of Hürthle cells, 2. presence of neoplastic Hürthle cells, and 3. presence of benign Hürthle cells. The final Hürthle cell Index utilizes 1048 nuclear and mitochondrial genes; and Hürthle cell Neoplasm Index leverages LOH features as well as 2041 genes. Both indices are Support Vector Machine (SVM) based. The third classifier, the GSC Benign/Suspicious classifier, utilizes 1115 core genes and is an ensemble classifier incorporating 12 individual models. Conclusions The accurate algorithmic depiction of this complex biological system among Hürthle subtypes results in a dramatic improvement of classification performance; specificity among Hürthle cell neoplasms increases from 11.8% with the GEC to 58.8% with the GSC, while maintaining the same sensitivity of 89%. Electronic supplementary material The online version of this article (10.1186/s12918-019-0693-z) contains supplementary material, which is available to authorized users.

Background The Afirma Genomic Sequencing Classifier (GSC) uses RNA sequencing to assess FNA specimens from cytologically indeterminate thyroid nodules, which are also tested for specific molecular aberrations associated with thyroid cancer via a suite of highly accurate malignancy classifiers. This suite can also be applied independently to Bethesda V/VI nodules. The Afirma Xpression Atlas (XA) is an optional add-on test to GSC and the malignancy classifiers that reports nucleotide variants and fusions across 511 cancer-associated genes. Medullary thyroid cancer (MTC) is a rare subtype of thyroid cancer that can appear in every Bethesda category. Herein, we report the prevalence and genomic landscape of MTC classifier positive (MTC+) FNA samples in a CLIA certified clinical laboratory. Methods All Afirma GSC and malignancy classifier tests run between July 2017 and November 2018 were deidentified and examined for MTC+ cases. Afirma XA was run on all MTC cases and all variants and fusions were tabulated. Results Examination of 22,130 FNAs revealed 77 MTC cases. Among consecutive GSC tests, 28 were Bethesda III (0.16% out of 17,245) and 27 were Bethesda IV (0.65% out of 4,182). Provider-ordered testing was done on an additional 14 and 8 MTC cases from Bethesda V and VI nodules, respectively. Examining all MTC+ samples revealed that 55.8% harbored a RET variant (+/- others), 9.1% contained a KRAS variant (+/- others), 6.5% had an HRAS variant, 2.6% possessed fusions, and 26.0% included no variant/fusion. Conclusions In indeterminate FNA samples, the Afirma GSC can help to clarify the risk of MTC. In our cohort, the Afirma XA identified a variant or fusion in 74.0% of MTC+ FNAs. Limitations of this study include the lack of knowledge regarding germline RET status (which should be checked in all patients with an identified RET variant or confirmed MTC) and final pathology on MTC+ samples. Future studies may investigate how the preoperative identification of a known MTC driver mutation in a biopsy sample can inform the pre-operative evaluation, the surgical plan, and the potential role of targeted therapy. Unless otherwise noted, all abstracts presented at ENDO are embargoed until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. Abstracts presented at a news conference are embargoed until the date and time of the news conference. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.

© 2018 American Medical Association. All rights reserved. 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.

Along with breast and endometrial cancers, thyroid cancer is a major component cancer in Cowden syndrome (CS). Germline variants in SDHB/C/D (SDHx) genes account for subsets of CS/CS-like cases, conferring a higher risk of breast and thyroid cancers over those with only germline PTEN mutations. To investigate whether SDHx alterations at both germline and somatic levels occur in apparently sporadic breast cancer and differentiated thyroid cancer (DTC), we analyzed SDHx genes in the following four groups: i) 48 individuals with sporadic invasive breast adenocarcinoma for germline mutation; ii) 48 (expanded to 241) DTC for germline mutation; iii) 37 pairs DTC tumor-normal tissues for germline and somatic mutation and mRNA expression levels; and iv) data from 476 patients in the Cancer Genome Atlas thyroid carcinoma dataset for validation. No germline SDHx variant was found in a pilot series of 48 breast cancer cases. As germline SDHx variants were found in our pilot of 48 thyroid cancer cases, we expanded to three series of DTC comprising a total 754 cases, and found 48 (6%) with germline SDHx variants (PSDHC. Both papillary and follicular thyroid tumors showed consistent loss of SDHC/D gene expression (PSDHx mutations/variants occur in sporadic DTC but are very rare in sporadic breast cancer, and overall loss of SDHx gene expression is a signature of DTC.

Background: Treatment options for patients with advanced or metastatic medullary thyroid cancer (MTC) have, in recent years, expanded with the approval of two tyrosine kinase inhibitors (TKIs): vandetanib and cabozantinib. Other agents, including TKIs, are under clinical investigation for MTC. Although patients treated with TKIs are at risk of developing dermatologic adverse events (AE), these untoward events may be mitigated through AE-driven algorithms. Summary: AE-driven algorithms combine effective nonpharmaceutical and pharmaceutical treatment modalities implemented by a multidisciplinary effort that incorporates nursing interventions, patient education, and referrals to pain-management specialists, podiatrists, and dermatologists, as appropriate. Effective AE prevention and management reduce the need for dose interruptions and modifications, allowing patients the opportunity to derive the maximal benefit from TKI therapy, while maintaining quality of life. Conclusions: Optimal use of targeted therapies in the treatment of MTC depends on careful patient selection, interdisciplinary communication, and patient education and encouragement to enhance compliance and safety, optimize consistent dosing, and maximize the use of effective therapies.

Sialadenitis and xerostomia are major adverse effects ofThe aim of this study is to identify risk factors forWe enrolled 216 thyroid cancer patients who visited The Ohio State University Wexner Medical Center between April 2013 and April 2014. Symptoms of xerostomia and sialadenitis were identified via questionnaire and medical record search. To validate the findings in a large cohort, we retrospectively searched for ICD-9/10 codes for sialadenitis, xerostomia, and autoimmune disease associated with Sjögren's syndrome (AID-SS) in our existing database (n = 1507). Demographic and clinical information was extracted from medical records. Multivariate analyses were performed to identify independent predictors for salivary gland damage.Risk factors for

6087 Background: Systemic therapies targeting specific genomic alterations in advanced MTC are available or under investigation. The Afirma Genomic Sequencing Classifier (GSC) uses RNA sequencing to assess FNA specimens from cytologically indeterminate thyroid nodules, which are also tested for specific molecular aberrations associated with thyroid cancer via a suite of highly accurate malignancy classifiers. This suite can be applied independently to Bethesda V/VI nodules. The Afirma Xpression Atlas (XA) is an additional test that can be combined with Afirma GSC to report nucleotide variants and fusions across 511 cancer-associated genes. Here we report the prevalence and genomic landscape of MTC classifier positive (MTC+) FNA samples. Methods: All Afirma GSC and malignancy classifier tests run in the Veracyte Clinical Laboratory between July 2017 and January 2019 were deidentified and examined for MTC+ cases. Afirma XA was run on all such cases, and all variants and fusions were tabulated. Results: Examination of 29,895 FNAs revealed 90 MTC cases. Of 22,793 Bethesda III cases, 32 (0.14%) were MTC+. Of 5,491 Bethesda IV cases, 33 (0.60%) were MTC+. Provider-ordered testing was done on an additional 16 and 9 MTC cases from Bethesda V and VI nodules, respectively. 58% of all MTC+ samples harbored a RET variant (+/- others), 9% contained a KRAS variant (+/- others), 6% included an HRAS variant, 1% had a BRAF fusion, 1% demonstrated other fusions, and 26% held no variant/fusion. Conclusions: In our cohort, Afirma XA identified a variant or fusion in 74% of MTC+ FNAs. Currently approved or investigational therapies exist for cancers with RET, BRAF and HRAS alterations, suggesting that 64% of our series might be eligible for treatment based on genomic information from FNA. In advanced MTC, noninvasive FNA sample collection at the time of diagnosis may ultimately impact on targeted therapy selection, with the option to repeat FNA testing should the disease progress. Future studies may investigate how finding a genomic alteration by FNA can inform the management of MTC and, in the case of progressive disease, improve our understanding of the mechanisms of disease progression and drug resistance.

The purpose of this study was to determine the frequency of BRAF mutation in cytologically indeterminate thyroid nodules and to investigate whether adding the BRAF test improves diagnostic accuracy of the Afirma Gene Expression Classifier (GEC).BRAF V600E mutational status was determined for DNA extracted from cytologically benign (n = 40), indeterminate (n = 208), and malignant (n = 48) fine-needle aspiration specimens previously categorized by GEC as molecularly Benign or Suspicious. Analytical performance of the BRAF assay was assessed to establish reproducibility and limits of detection. Molecular testing results were correlated with blinded expert histopathological diagnoses.The BRAF assay detected mutations reproducibly to 2.5% mutant allele frequency. The prevalence of BRAF mutations in cytologically benign specimens was 2 of 40 (5.0%, 95% confidence interval [CI], 0-16) and in cytologically malignant specimens was 36 of 48 (75.0%, 95% CI, 60-86). In the cytologically indeterminate category, 10.1% of specimens were BRAF+: 2 of 95 were subcategorized as atypia of undetermined significance or follicular lesion of undetermined significance (2.1%, 95% CI, 0-7); 1 of 70 as follicular neoplasm or suspicious for follicular neoplasm (1.4%, 95% CI, 0-9); and 18 of 43 as suspicious for malignancy (41.9%, 95% CI, 27-58). All BRAF+ specimens were classified as Suspicious by the GEC.BRAF mutations are uncommon in nodules with atypia of undetermined significance or follicular lesion of undetermined significance or follicular neoplasm or suspicious for follicular neoplasm cytology. Most cytologically indeterminate nodules that proved to be malignant were also BRAF-, and all nodules that were false-negative by GEC were also BRAF-. Similarly, all BRAF+ specimens were also GEC Suspicious. Neither GEC test sensitivity nor specificity was improved by addition of BRAF mutation testing.

Objectives: The gene expression classifier (GEC, Afirma) reclassifies as molecularly benign approximately one half of thyroid nodule fine-needle aspiration (FNA) biopsies with an initial indeterminate cytopathology diagnosis, facilitating clinical monitoring in lieu of diagnostic thyroid surgery. This study evaluated the long-term management patterns and thyroid surgery rates of GEC benign patients compared to a control group of cytopathology benign patients and also described the costs of thyroid surgery. Methods: This retrospective cohort study used laboratory test results linked to payer medical claims data. Patients who underwent FNA biopsy between 1 January 2011 and 31 July 2013 were selected. GEC benign patients were matched 1:3 to cytopathology benign patients on biopsy year, gender, nodule size and age. Outcomes measured included thyroid-related follow-up clinic visits, ultrasound examinations and surgeries. Results: Out of 2059 patients, matched groups consisting of 201 GEC benign patients and 603 cytopathology benign patients were evaluated over an average follow-up of 20 months. The proportions of GEC benign and cytopathology benign patients that underwent thyroid surgery (11.4% versus 10.1%, p = 0.594), and received a follow-up ultrasound exam (60.2% versus 61.7%, p = 0.706), respectively, were not significantly different. Average thyroid-related medical cost for the surgical episode and during 6 months following surgery were $10,432.00 (SD $8301) and $10,939.00 (SD $9656) respectively. Limitations: The study cohort included only patients whose diagnostic laboratory test results and administrative claims data were uniquely identifiable and could be linked on multiple identifiers; the rigorous matching and patient selection algorithms utilized improved the robustness and internal validity of the study. Claims were used as a proxy for clinical utilization without chart review confirmation. Conclusion: Patients with GEC and cytopathology benign diagnoses were managed similarly. The majority of patients in both groups did not undergo surgery and were managed with usual care, consisting of clinical follow-up and ultrasound exams.

Background Thyroid carcinomas are known to harbor oncogenic driver mutations and advances in sequencing technology now allow the detection of these in fine needle aspiration biopsies (FNA). Recent work by The Cancer Genome Atlas (TCGA) Research Network has expanded the number of genetic alterations detected in papillary thyroid carcinomas (PTC). We sought to investigate the prevalence of these and other genetic alterations in diverse subtypes of thyroid nodules beyond PTC, including a variety of samples with benign histopathology. This is the first clinical evaluation of a large panel of TCGA-reported genomic alterations in thyroid FNAs. Results In FNAs, genetic alterations were detected in 19/44 malignant samples (43 % sensitivity) and in 7/44 histopathology benign samples (84 % specificity). Overall, after adding a cohort of tissue samples, 38/76 (50 %) of histopathology malignant samples were found to harbor a genetic alteration, while 15/75 (20 %) of benign samples were also mutated. The most frequently mutated malignant subtypes were medullary thyroid carcinoma (9/12, 75 %) and PTC (14/30, 47 %). Additionally, follicular adenoma, a benign subtype of thyroid neoplasm, was also found to harbor mutations (12/29, 41 %). Frequently mutated genes in malignant samples included BRAF (20/76, 26 %) and RAS (9/76, 12 %). Of the TSHR variants detected, (6/7, 86 %) were in benign nodules. In a direct comparison of the same FNA also tested by an RNA-based gene expression classifier (GEC), the sensitivity of genetic alterations alone was 42 %, compared to the 91 % sensitivity achieved by the GEC. The specificity based only on genetic alterations was 84 %, compared to 77 % specificity with the GEC. Conclusions While the genomic landscape of all thyroid neoplasm subtypes will inevitably be elucidated, caution should be used in the early adoption of published mutations as the sole predictor of malignancy in thyroid. The largest set of such mutations known to date detects only a portion of thyroid carcinomas in preoperative FNAs in our cohort and thus is not sufficient to rule out cancer. Due to the finding that variants are also found in benign nodules, testing only GEC suspicious nodules may be helpful in avoiding false positives and altering the extent of treatment when selected mutations are found. Electronic supplementary material The online version of this article (doi:10.1186/s12859-015-0849-9) contains supplementary material, which is available to authorized users.

Item does not contain fulltext BACKGROUND: Current surgical standard of care in sporadic medullary thyroid carcinoma (sMTC) consists of a minimum of total thyroidectomy with central neck dissection. Some have suggested thyroid lobectomy with isthmusectomy and central neck dissection for patients with sMTC, given their lower frequency of bilateral disease, although this topic has not been thoroughly studied. This study assessed the prevalence of multifocality in sMTC via a large international multi-institutional retrospective review to quantify this prevalence, including the impact of geography, to assess more accurately the risks associated with alternative surgical approaches. METHODS: A retrospective chart review of sMTC patients from 11 institutions over 29 years (1983-2011) was undertaken. Data regarding focality, extent of disease, RET germline analysis plus family and clinical history for multiple endocrine neoplasia type 2 (MEN2), and demographic data were collected and analyzed. RESULTS: Patients from four continents and seven countries were included in the sample. Data for 313 patients with documented sMTC were collected. Of these, 81.2% were confirmed with negative RET germline testing, while the remaining 18.8% demonstrated a negative family history and no manifestations of MEN2 syndromes other than MTC. Bilateral disease was identified in 17/306 (5.6%) patients, while multifocal disease was noted in 50/312 (16.0%) sMTC patients. When only accounting for germline negative patients, these rates were not significantly different (5.6% and 17%, respectively). Among them, when disease was unifocal in the ipsilateral lobe and isthmus, bilateral disease was present in 6/212 (2.8%) cases. When disease was multifocal in the ipsilateral lobe or isthmus, then bilateral disease was present in 8/37 (21.6%) cases (p < 0.001). No geographic differences in focality were identified. CONCLUSIONS: The 5.6% prevalence of bilateral foci in sMTC suggests that total thyroidectomy should remain the standard of care for initial surgery, as less complete thyroid surgery may fail to address fully the primary site of disease. Whether ipsilateral tumor focality should be an independent factor determining the need for completion thyroidectomy when sMTC is diagnosed after hemithyroidectomy remains to be determined.

BACKGROUND Approximately 15 to 30% of thyroid nodules evaluated by means of fine-needle aspiration are not clearly benign or malignant. Patients with cytologically indeterminate nodules are often referred for diagnostic surgery, though most of these nodules prove to be benign. A novel diagnostic test that measures the expression of 167 genes has shown promise in improving preoperative risk assessment. METHODS We performed a 19-month, prospective, multicenter validation study involving 49 clinical sites, 3789 patients, and 4812 fine-needle aspirates from thyroid nodules 1 cm or larger that required evaluation. We obtained 577 cytologically indeterminate aspirates, 413 of which had corresponding histopathological specimens from excised lesions. Results of a central, blinded histopathological review served as the reference standard. After inclusion criteria were met, a gene-expression classifier was used to test 265 indeterminate nodules in this analysis, and its performance was assessed. RESULTS Of the 265 indeterminate nodules, 85 were malignant. The gene-expression classifier correctly identified 78 of the 85 nodules as suspicious (92% sensitivity; 95% confidence interval [CI], 84 to 97), with a specificity of 52% (95% CI, 44 to 59). The negative predictive values for “atypia (or follicular lesion) of undetermined clinical significance,” “follicular neoplasm or lesion suspicious for follicular neoplasm,” or “suspicious cytologic findings” were 95%, 94%, and 85%, respectively. Analysis of 7 aspirates with false negative results revealed that 6 had a paucity of thyroid follicular cells, suggesting insufficient sampling of the nodule. CONCLUSIONS These data suggest consideration of a more conservative approach for most patients with thyroid nodules that are cytologically indeterminate on fine-needle aspiration and benign according to gene-expression classifier results. (Funded by Veracyte.)

Micro-single-photon emission computed tomography (SPECT) provides a noninvasive way to evaluate the effects of genetic and/or pharmacological modulation on sodium-iodide symporter (NIS)-mediated radionuclide accumulation in mouse thyroid and salivary glands. However, parameters affecting image acquisition and analysis of mouse thyroids and salivary glands have not been thoroughly investigated. In this study, we investigated the effects of region-of-interest (ROI) selection, collimation, scan time, and imaging orbit on image acquisition and quantification of thyroidal and salivary radionuclide accumulation in mice.The effects of data window minima and maxima on thyroidal and salivary ROI selection using a visual boundary method were examined in SPECT images acquired from mice injected with (123)I NaI. The effects of collimation, scan time, and imaging orbit on counting linearity and signal intensity were investigated using phantoms filled with various activities of (123)I NaI or Tc-99m pertechnetate. Spatial resolution of target organs in whole-animal images was compared between circular orbit with parallel-hole collimation and spiral orbit with five-pinhole collimation. Lastly, the inter-experimental variability of the same mouse scanned multiple times was compared with the intra-experimental variability among different mice scanned at the same time.Thyroid ROI was separated from salivary glands by empirically increasing the data window maxima. Counting linearity within the range of 0.5-14.2 μCi was validated by phantom imaging using single- or multiple-pinhole collimators with circular or spiral imaging orbit. Scanning time could be shortened to 15 minutes per mouse without compromising counting linearity despite proportionally decreased signal intensity. Whole-animal imaging using a spiral orbit with five-pinhole collimators achieved a high spatial resolution and counting linearity. Finally, the extent of inter-experimental variability of NIS-mediated radionuclide accumulation in the thyroid and salivary glands by SPECT imaging in the same mouse was less than the magnitude of variability among the littermates.The impacts of multiple variables and experimental designs on micro-SPECT imaging and quantification of radionuclide accumulation in mouse thyroid and salivary glands can be minimized. This platform will serve as an invaluable tool to screen for pharmacologic reagents that differentially modulate thyroidal and salivary radioiodine accumulation in preclinical mouse models.

The role of prophylactic central lymph node dissection in papillary thyroid cancer (PTC) is controversial in patients who have no pre- or intraoperative evidence of nodal metastasis (clinically N0; cN0). The controversy relates to its unproven role in reducing recurrence rates while possibly increasing morbidity (permanent hypoparathyroidism and unintentional recurrent laryngeal nerve injury).We examined the design and feasibility of a multi-institutional prospective randomized controlled trial of prophylactic central lymph node dissection in cN0 PTC. Assuming a 7-year study with 4 years of enrollment, 5 years of average follow-up, a recurrence rate of 10% after 7 years, a 25% relative reduction in the rate of the primary endpoint (newly identified structural disease; i.e., persistent, recurrent, or distant metastatic disease) with central lymph node dissection and an annual dropout rate of 3%, a total of 5840 patients would have to be included in the study to achieve at least 80% statistical power. Similarly, given the low rates of morbidity, several thousands of patients would need to be included to identify a significant difference in rates of permanent hypoparathyroidism and unintentional recurrent laryngeal nerve injury.Given the low rates of both newly identified structural disease and morbidity after surgery for cN0 PTC, prohibitively large sample sizes would be required for sufficient statistical power to demonstrate significant differences in outcomes. Thus, a prospective randomized controlled trial of prophylactic central lymph node dissection in cN0 PTC is not readily feasible.

Context: The family risk ratio for papillary thyroid carcinoma (PTC) is among the highest of all cancers. Collectively, familial cases (fPTC) and sporadic cases (sPTC) are not known to show molecular differences. However, one study reported that telomeres were markedly shorter and the telomerase reverse transcriptase (TERT) gene was amplified and up-regulated in germline DNA from patients with fPTC compared with sPTC. Objective: The aim of this study was to evaluate telomere length and TERT gene amplification and expression in blood samples of fPTC and sPTC patients in a genetically distinct population from the previous study. Design: In 42 fPTC and 65 sPTC patients, quantitative real-time PCR was employed to measure the relative telomere length (RTL) and TERT gene copy number and RNA level. To validate the results using alternative methods, we further studied a subset of the original cohort consisting of randomly chosen fPTC (n = 10) and sPTC (n = 14) patients and controls (n = 21) by assessing both telomere length by flow fluorescent in situ hybridization and TERT gene expression by quantitative real-time PCR. Results: RTL and TERT gene copy number did not differ between fPTC and sPTC (P = 0.957 and P = 0.998, respectively). The mean RTL and TERT gene expression were not significantly different among the groups of the validation series (P = 0.169 and P = 0.718, respectively). Conclusion: Our data show no difference between familial and sporadic PTC with respect to telomere length, TERT copy number, or expression in our cohort. Further investigations in additional cohorts of patients are desirable.

The prevalence of occult medullary thyroid carcinoma (MTC) in the general population is unknown but may be important when considering strategies to diagnose clinically relevant MTC in nodular goiter or other populations.Our objective was to determine the prevalence of occult MTC in a series of autopsies.We conducted a systematic review of autopsy series from 1970 to present using a PubMed search.The patients came from 21 countries, ages ranged from 6-95 yr, both genders were represented, and none had clinical evidence of thyroid disease before autopsy.Three series were excluded based on tumor size less than 500 μm, non-English language, or insufficient information.Prevalence of occult MTC was calculated.An average prevalence of 0.14 and 7.6% for occult MTC and papillary thyroid carcinoma, respectively, was found among 7897 autopsies from 24 published series. Greater than 75% of patients with MTC were more than 60 yr old, and male to female ratio was comparable. Tumor size was virtually all subcentimeter, and there was no lymph node spread, extrathyroidal extension, or distant metastases reported.A small number of people in the general population, who do not have known thyroid disease, have occult MTC and die of other causes. This finding of untreated occult MTC without morbidity or mortality should be considered in population prevalence studies, when strategies to detect thyroid neoplasia are considered (e.g. serum calcitonin or ultrasound), and included in cost-effectiveness models of routine serum calcitonin screening for nodular thyroid disease.

Selective iodide uptake and prolonged iodine retention in the thyroid is the basis for targeted radioiodine therapy for thyroid cancer patients; however, salivary gland dysfunction is the most frequent nonthyroidal complications. In this study, we have used noninvasive single photon emission computed tomography functional imaging to quantify the temporal dynamics of thyroidal and salivary radioiodine accumulation in mice. At 60 min post radionuclide injection, radionuclide accumulation in the salivary gland was generally higher than that in thyroid due to much larger volume of the salivary gland. However, radionuclide accumulation per anatomic unit in the salivary gland was lower than that in thyroid and was comparable among mice of different age and gender. Differently, radionuclide accumulation per anatomic unit in thyroid varied greatly among mice. The extent of thyroidal radioiodine accumulation stimulated by a single dose of exogenous bovine TSH (bTSH) in triiodothyronine (T₃)-supplemented mice was much less than that in mice received neither bTSH nor T₃ (nontreated mice), suggesting that the duration of elevated serum TSH level is important to maximize thyroidal radioiodine accumulation. Furthermore, the extent and duration of radioiodine accumulation stimulated by bTSH was less in the thyroids of the thyroid-targeted RET/PTC1 (thyroglobulin (Tg)-PTC1) mice bearing thyroid tumors compared with the thyroids in wild-type (WT) mice. Finally, the effect of 17-allyamino-17-demothoxygeldanamycin on increasing thyroidal, but not salivary, radioiodine accumulation was validated in both WT mice and Tg-PTC1 preclinical thyroid cancer mouse model.