Abstract 5639: Diagnosing indeterminate thyroid nodules: combining next generation sequencing and gene expression analysis
Authors: Weiss Vivian L. , Daber Robert , Moreau Michael , Gaskell Alisa , Ely Kim , Coogan Alice , Stricker Thomas 
Published: July 2017

Abstract
Thyroid nodules are common in the United States and are currently diagnosed using fine needle aspiration (FNA) biopsy. FNA evaluation provides a definitive diagnosis in the majority of cases, but about 20-30% of nodules are deemed indeterminate. Targeted next generation sequencing (NGS) and gene expression analysis have been used as ancillary tests to provide additional information about the malignancy risk for indeterminate thyroid lesions. Here we develop a 31 gene NGS panel, as well as a 14 gene expression panel, and test their performance on thyroid lesions with known diagnosis. Formalin-fixed paraffin embedded tissue from 23 resected thyroid lesions (1 non-neoplastic, 3 follicular adenomas (FA), 1 medullary carcinoma (MC), 7 follicular carcinomas (FC), 7 papillary carcinomas (PC), 4 follicular variant of papillary carcinomas (FVPC)) was evaluated. Total nucleic acid extraction, NGS (31 genes including: NRAS, HRAS, KRAS, CTNNB1, PIK3CA, BRAF, RET, PTEN, AKT1, TP53, GNAS, ALK, DDR2, EGFR, ERBB2, FGFR1/2/3, GNA11, GNAQ, IDH1/2, KIT, MAP2K1, MET, MTOR, NOTCH1, PDGFRA, EPHA2, ESR1, RAC1, and ROS1) through Bio Reference Laboratories, and gene expression analysis (14 genes: KRT7, TTF-1, TG, PGK-1, CK-20, PTH, CALCA, RAP2A, PLAB, HMGA2, FN-1, GAL-3, CK19, and NIS) were performed. This assay detected mutations in 14/19 malignant lesions including 4/7 FCs (NRAS, TP53), 4/4 FVPC (BRAF, NRAS, HRAS, PTEN), 5/7 PCs (BRAF, NRAS), and 1/1 MC (RET). No mutations were detected in the non-neoplastic thyroid and 1/3 FAs contained a KRAS mutation. All three mutation-negative FCs were oncocytic variants and showed upregulation of ≥2 cancer-associated genes including FN-1, GAL-3, HMGA2, PLAB, and RAP2a. The two mutation-negative PCs showed upregulation of 4 cancer-associated genes including FN-1, CK-19, GAL-3, HMGA2, and PLAB. This upregulation was not seen in non-neoplastic thyroid and was seen in only 1 of the FAs (FN-1, GAL-3, and PLAB). This small study demonstrates that a combination of NGS and gene expression increases the assay sensitivity (100% versus 74% sensitivity) and negative predictive value (100% versus &lt;45% for NGS or gene expression alone). The positive predictive value of the combined assay was 90% versus 93% for either assay alone. The addition of gene fusions will likely further improve the assay’s diagnostic capacity. In conclusion, the addition of a gene expression assay to this thyroid NGS assay allowed for identification of mutation-negative thyroid malignancies. The improved detection of malignant thyroid lesions will allow for better risk assessment for indeterminate nodules as well as improved treatment algorithms.
Citation Format: Vivian L. Weiss, Robert Daber, Michael Moreau, Alisa Gaskell, Kim Ely, Alice Coogan, Thomas Stricker. Diagnosing indeterminate thyroid nodules: combining next generation sequencing and gene expression analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5639. doi:10.1158/1538-7445.AM2017-5639