Evaluating novel in silico tools for accurate pathogenicity classification in epilepsy-associated genetic missense variants.

Objective: Determining the pathogenicity of missense variants in clinical genetic tests for individuals with epilepsy is crucial for guiding personalized treatment. However, achieving a definitive pathogenic classification remains challenging, with most missense variants still classified as variants of uncertain significance (VUS) and with the availability of many computational tools which may provide conflicting predictions. Here, we aim to evaluate the performance of state-of-the-art computational tools in pathogenicity prediction of missense variants in epilepsy-associated genes. This will assist in selecting the most appropriate tool and critically assess their use in clinical setting.

Methods: We assessed the performance of nine in silico pathogenicity prediction tools for missense variants in epilepsy-associated genes on three carefully curated data sets. The first two data sets comprise missense variants in epilepsy associated genes that have been uploaded to ClinVar in the last year and were, therefore, not part of the training set of any of the nine considered tools. These two data sets are based on two different lists of epilepsy-associated genes and comprise ~700 and ~ 250 missense variants, respectively. The third data set includes ~400 missense variants within epilepsy-associated genes for which the functional effects have been determined experimentally and are therefore used here to infer pathogenicity. These three data sets represent the best available approximation to blind and independent test sets.

Results: Among the nine assessed tools, AlphaMissense (area under the curve [AUC]: .93, .88, and .95) and REVEL (AUC: .93, .88, and .93) showed the best classification performance, also outperforming other tools in the number of classified variants.

Significance: We show which recently developed prediction tools achieve higher performance in epilepsy-associated genes and should be integrated, therefore, into the American College of Medical Genetics and Genomics/Association of Molecular Pathology (AGMC/AMP) variant classification process. Periodic reevaluation of genetic test results with newly developed or updated tools should be incorporated into standard clinical practice to improve diagnostic yield and better inform precision medicine.