Risk Stratification Study of Indeterminate Thyroid Nodules with a next-generation Sequencing Assay with Residual ThinPrep® Material

Abstract

Objective: The management of indeterminate thyroid nodules is challenging. Molecular testing has emerged as a promising method for stratifying this gray area of fine-needle aspiration (FNA) cytology. Next-generation sequencing (NGS) can be used to test a large variety of genetic changes with very small amounts of nucleic acids obtained from FNA samples. Methods: Thyroid FNA assays were classified according to the Bethesda System for Reporting Thyroid Cytopathology after routine ThinPrep® slide preparation. Indeterminate nodules with surgical outcomes were assayed with an 18-gene NGS panel with the residual ThinPrep® material, including nodules categorized as atypia of undetermined significance (AUS)/follicular lesions of undetermined significance (FLUS) or follicular neoplasm (FN)/suspicious for a follicular neoplasm (SFN). We evaluated the diagnostic efficacy of the 18-gene panel for thyroid malignancies and potential malignancies and compared it with a well-accepted examination, ThyroSeq v2 testing. Results: A total of 36 indeterminate nodules were assayed, seven were categorized as AUS/FLUS and 29 as FN/SFN. All of them had adequate DNA for the NGS procedure. When noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) was considered malignant, the risk of malignancy was 71.4% for AUS/FLUS nodules, and 69.0%for FN/SFN nodules. The 18-gene panel showed 72.0% sensitivity, 72.7% specificity, 85.7% positive predictive value (PPV), and 53.3% negative predictive value (NPV) in identifying malignancies and potential malignancies in the indeterminate nodules. Compared with a multicenter report from ThyroSeq v2 testing, 18-gene panel showed a lower NPV (p=0.005), but a higher PPV (p=0.02). Conclusions: NGS assays are feasible on residual ThinPrep® material, with the advantage of not requiring additional FNA procedure. The 18-gene panel testing can be used as a 'rule in' test for surgical management based on indeterminate nodules and showed a lower NPV but a higher PPV compared to ThyroSeq v2 testing.

Diagnostic performance of next-generation sequencing and genetic profiling in thyroid nodules from a single center in China

“…In the literature, Nikiforov et al also reported that only one of three TSHR mutation-positive nodules was identified as cancer after surgery and two NRAS -mutated samples were benign on histology ( 30 ). Furthermore, RAS mutations are considered to be not specifically associated with malignant (75–88%) or potentially malignant outcomes ( 7 , 27 , 31 , 32 , 33 ) and are highly predictive of predominantly low-risk follicular-pattern carcinomas ( 34 ), which is further supported by our results of a significantly lower frequency of lymph node metastasis in RAS -mutated malignancies. Additional gene mutations including SPOP , EZH1 , and ZNF148 are also suggested to be benign markers ( 32 ).…”

“…However, approximately 20–30% of FNA samples reveal an indeterminate state (AUS/FLUS and FT/SFT), with only a 10–40% risk of malignancy ( 6 ). In recent years, several molecular tests were developed to improve the accuracy of preoperative diagnosis of cytologically indeterminate thyroid nodules and minimize false-positive and false-negative cytology ( 7 , 12 , 27 , 28 ). The multigene NGS assay in the current study, targeting point mutations and gene fusions in thyroid cancer, showed superior diagnostic values.…”

Performance of repeat cytology with reflex ThyroSeq genomic classifier for indeterminant thyroid cytology

“…We recognize that the optimal strategy for facilitating the performance of ancillary tests can vary, depending on many factors unique to each institution: the laboratories, the type of cytology preparations they make (eg, direct smears, liquid‐based preparations, cell blocks), and their patient populations. Recent studies have demonstrated the ability perform ThyroSeq on cytology smear slides 22 and residual ThinPrep material, 23 and these are potential alternative avenues that could help mitigate the some of the costs of collecting separate aliquots for molecular testing, particularly for very busy practices in which the resources needed for routine molecular collection are perceived as prohibitive. Another rational for using rebiopsy as part of the diagnostic algorithm for an AUS/FLUS finding is that rebiopsy may have the potential to obtain additional diagnostic material that is not present or detectable in the initial sampling.…”

Thyroid Cancer Screening Using Tumor-Associated DN T Cells as Immunogenomic Markers

“…Approximately 600,000 fine-needle aspirations (FNAs) are performed annually in the United States, and almost a fifth of those (i.e., 120,000 nodules) carry features consistent with atypia of undetermined significance and are suspicious for follicular neoplasms or for thyroid malignancy (3)(4)(5). These nodules are commonly subjected to thyroid surgery in order to establish a definitive diagnosis (6)(7)(8). These strategies lead to extravagant costs in the care of thyroid cancer that could reach $3.5 billion annually by 2030 (2).…”