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AMP, NSGC Issue New Recommendations for Confirming Germline Variants From NGS Tests

NEW YORK – The Association for Molecular Pathology and the National Society of Genetic Counselors have jointly issued new recommendations for confirming germline variants detected by next-generation sequencing.

Released in the Journal of Molecular Diagnostics on Wednesday, the guidelines reflect conclusions of a working group convened by AMP to assess current evidence and standardize orthogonal confirmation practices to help limit the reporting of false positive results. The authors include representatives from several commercial laboratories including Ambry Genetics, Invitae, and Veritas Genetics/LetsGetChecked.

"While NGS has quickly transformed the field of clinical molecular genetic testing, orthogonal confirmation practices for germline variants may vary between laboratories," Kristy Crooks, laboratory director at the University of Colorado Anschutz Medical Campus and chair of the working group, said in a statement.

The new report comprises eight recommendations, informed by a comprehensive review of the literature, empirical data, current laboratory practice surveys, feedback from open public comment, and professional experience.

According to the authors, the guide applies only to germline DNA sequencing. "Other clinical DNA tests are not in scope, including tumor sequencing and liquid biopsy tests, both intended to detect somatic mutations, as well as noninvasive prenatal screening, intended to detect fetal variants in maternal blood," the group wrote.

Among other recommendations, AMP and NSGC advise clinical laboratories to establish a written policy regarding orthogonal confirmation of NGS results, and to make sure that policy is overseen and approved by a qualified and appropriately certified medical professional with training and experience in clinical NGS. They should also be prepared to provide this written policy, as well as information about their criteria and methods, upon request.

When delivering clinical test reports, labs should routinely summarize their orthogonal confirmation policy and clearly indicate if exceptions have been made.

The recommendations also state that laboratories’ confirmatory methods, including sequencing platforms and associated bioinformatics, should be validated and maintained under appropriate regulatory oversight. Confirmation of germline findings should be orthogonal, and in cases of discrepancy, further investigation should be performed, rather than defaulting to either the original or confirmatory assay as correct.

While labs can forego confirmatory testing for variant calls that meet certain technical criteria that have been rigorously demonstrated to ensure a high positive predictive value from NGS alone, they should make sure to confirm all other germline variants with "significant clinical implications."

Susan Hsiao, associate professor of pathology at Columbia University's Vagelos College of Physicians and Surgeons and chair of the 2023 AMP clinical practice committee, said that the association will continue to reassess and modify these guidelines as methodologies and bioinformatics underlying NGS-based variant detection evolve.