BACKGROUND Genetic testing frequently identifies variants of uncertain significance (VUSs). Providers, however, are often ill-prepared or too time-constrained to manage these findings, and insurers are concerned about impacts on clinical care and cost. Here we compared the contribution of panel-based and genomic (exome and genome) testing to the generation of inconclusive results due to VUSs. METHODS Rates of inconclusive results due to VUS and diagnostic yield were determined from over 1.5 million sequencing test results from 19 clinical laboratories in North America from 2020 - 2021. RESULTS We found a lower rate of inconclusive test results due to VUSs from genomic tests (22.5%) compared to multi-gene panel tests (32.6%; p<0.001) and a higher diagnostic yield (17.5% vs 10.3%; p<0.001). For panel tests, the rate of inconclusive results correlated with panel size. The use of trios improved yield (19.5% vs 15.2%; p<0.001) and reduced inconclusive rates (18.9% vs 27.6%; p<0.001). The use of genome sequencing compared to exome improved yield (25.1% vs 16.6%; p<0.001) without increasing the rate of inconclusive results (22.2% vs 22.6%). CONCLUSION Genomic sequencing demonstrated reduced uncertainty and higher molecular diagnostic yield compared to panel testing. This difference is best explained by obligatory reporting of all VUSs in panel-based testing compared to genomic testing where correlation with phenotype is used to constrain variant reporting. These results may inform future genetic testing practices and heighten appreciation for the professional skills involved in genomic test interpretation.

All authors are employed by clinical laboratories offering genetic testing services, as indicated by their affiliations. Additional conflicts include: Swaroop Aradhya, Elaine Chen, Kathryn E Hatchell, and Dianalee McKnight - Stockholders of Invitae; Christina DiVincenzo, Izabela D Karbassi - Stockholders of Quest Diagnostics; Kyle Retterer - Stockholder of Sema4 and Opko Health; Kyle W Davis, Nir Neerman, and Christine Stanley - Stockholders of Variantyx

HR's participation in this study was funded in part by the National Human Genome Research Institute under award U24HG006834.

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NOT HUMAN SUBJECT RESEARCH DETERMINATION Date: September 21, 2022 Title of Project: Comparing the rate of uncertainty generated from panels versus genomic sequencing tests Project Lead Name: Heidi Rehm The above referenced project does not meet the criteria for human subject research as defined by Mass General Brigham Human Research Office policies and Health and Human Services regulations set forth in 45 CFR 46. Based on the information you provided this activity is not human subjects research because does not involve human subjects. The project does not require IRB approval. This NHSR activity is not applicable for Clinicaltrials.gov registration. Please retain a copy of this letter in your project file. Please feel free to contact our office directly (partnersirb@partners.org) with any questions related to this determination. Sincerely, Ben McGill, MS, CIP Expedited Specialist II, Human Research Affairs bmcgill1@partners.org

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