No abstract

Bioinformatics pipelines are an integral component of next-generation sequencing (NGS). Processing raw sequence data to detect genomic alterations has significant impact on disease management and patient care. Because of the lack of published guidance, there is currently a high degree of variability in how members of the global molecular genetics and pathology community establish and validate bioinformatics pipelines. Improperly developed, validated, and/or monitored pipelines may generate inaccurate results that may have negative consequences for patient care. To address this unmet need, the Association of Molecular Pathology, with organizational representation from the College of American Pathologists and the American Medical Informatics Association, has developed a set of 17 best practice consensus recommendations for the validation of clinical NGS bioinformatics pipelines. Recommendations include practical guidance for laboratories regarding NGS bioinformatics pipeline design, development, and operation, with additional emphasis on the role of a properly trained and qualified molecular professional to achieve optimal NGS testing quality.

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The EIF1AX gene mutations have been recently found in papillary thyroid carcinoma (PTC) and anaplastic thyroid carcinoma (ATC). The prevalence of these mutations in other types of thyroid cancers and benign nodules is unknown. In this study, we analyzed the occurence of EIF1AX mutations in exons 2, 5 and 6 of the gene in a series of 266 thyroid tumors and hyperplastic nodules by either Sanger or next-generation sequencing (ThyroSeq v.2). In addition, 647 thyroid FNA samples with indeterminate cytology were analyzed. Using surgically removed samples, EIF1AX mutations were detected in 3/86 (2.3%) PTC, 1/4 (25%) ATC, 0/53 follicular carcinomas, 0/12 medullary carcinomas, 2/27 (7.4%) follicular adenomas and 1/80 (1.3%) hyperplastic nodules. Among 5 mutation-positive FNA samples with surgical follow-up, one nodule was PTC and others were benign follicular adenomas or hyperplastic nodules. Overall, among 33 mutations identified, A113_splice mutation at the intron 5/exon 6 splice site of EIF1AX was the most common. All 4 carcinomas harbored A113_splice mutation and three of them had one or more coexisting mutations, typically RAS. All PTC carrying EIF1AX mutation were encapsulated follicular variants. In summary, this study shows that EIF1AX mutations occur not only in thyroid carcinomas, but also in benign nodules. The most common mutation hotspot is the A113_splice, followed by a cluster of mutations in exon 2. When found in thyroid FNA samples, EIF1AX mutations confer ~20% risk of cancer; the risk is likely to be higher in nodules carrying a A113_splice mutation and when EIF1AX co-exists with RAS mutations.

The BRAF(K601E) mutation is the second most common BRAF mutation found in thyroid nodules. Unlike BRAF(V600E), the most common mutation, K601E is strongly associated with follicular-patterned cancer, particularly with the encapsulated follicular variant of PTC, and may also be found in follicular thyroid carcinomas. Overall, BRAF(K601E) mutant tumors show better clinical outcomes than BRAF(V600E) positive tumors, and preoperative BRAF(K601E) analysis may provide important prognostic information for use in clinical management.