Molecular Determinants of Thyroid Nodules with Indeterminate Cytology andRASMutations

Background:RASgene family mutations are the most prevalent in thyroid nodules with indeterminate cytology and are present in a wide spectrum of histological diagnoses. We evaluated differentially expressed genes and signaling pathways across the histological/clinical spectrum ofRAS-mutant nodules to determine key molecular determinants associated with a high risk of malignancy. Methods:Sixty-one thyroid nodules withRASmutations were identified. Based on the histological diagnosis and biological behavior, the nodules were grouped into five categories indicating their degree of malignancy: non-neoplastic appearance, benign neoplasm, indeterminate malignant potential, low-risk cancer, or high-risk cancer. Gene expression profiles of these nodules were determined using the NanoString PanCancer Pathways and IO 360 Panels, and Angiopoietin-2 level was determined by immunohistochemical staining. Results:The analysis of differentially expressed genes using the five categories as supervising parameters unearthed a significant correlation between the degree of malignancy and genes involved in cell cycle and apoptosis (BAX,CCNE2,CDKN2A,CDKN2B,CHEK1,E2F1,GSK3B,NFKB1, andPRKAR2A),PI3Kpathway (CCNE2,CSF3,GSKB3,NFKB1,PPP2R2C, andSGK2), and stromal factors (ANGPT2andDLL4). The expression of Angiopoietin-2 by immunohistochemistry also showed the same trend of increasing expression from non-neoplastic appearance to high-risk cancer (p < 0.0001). Conclusions:The gene expression analysis ofRAS-mutant thyroid nodules suggests increasing upregulation of key oncogenic pathways depending on their degree of malignancy and supports the concept of a stepwise progression. The utility ofANGPT2expression as a potential diagnostic biomarker warrants further evaluation.