An evaluation of pipelines for DNA variant detection can guide a reanalysis protocol to increase the diagnostic ratio of genetic diseases

Raquel Romero,Irene Perea-Romero,Pablo Mínguez,Berta Almoguera,María José Trujillo-Tiebas,Carmen Ayuso,Alejandra Damián,Gonzalo Núñez-Moreno,Inmaculada Martín-Mérida,Rosa Riveiro-Álvarez,Cristina Rodilla,Ionut-Florin Iancu,Marta Cortón,Almudena Ávila-Fernández,Marta Del Pozo-Valero,Lorena De La Fuente

doi:10.1038/s41525-021-00278-6

Abstract

Clinical exome (CE) sequencing has become a first-tier diagnostic test for hereditary diseases; however, its diagnostic rate is around 30–50%. In this study, we aimed to increase the diagnostic yield of CE using a custom reanalysis algorithm. Sequencing data were available for three cohorts using two commercial protocols applied as part of the diagnostic process. Using these cohorts, we compared the performance of general and clinically relevant variant calling and the efficacy of an in-house bioinformatic protocol (FJD-pipeline) in detecting causal variants as compared to commercial protocols. On the whole, the FJD-pipeline detected 99.74% of the causal variants identified by the commercial protocol in previously solved cases. In the unsolved cases, FJD-pipeline detects more INDELs and non-exonic variants, and is able to increase the diagnostic yield in 2.5% and 3.2% in the re-analysis of 78 cancer and 62 cardiovascular cases. These results were considered to design a reanalysis, filtering and prioritization algorithm that was tested by reassessing 68 inconclusive cases of monoallelic autosomal recessive retinal dystrophies increasing the diagnosis by 4.4%. In conclusion, a guided NGS reanalysis of unsolved cases increases the diagnostic yield in genetic disorders, making it a useful diagnostic tool in medical genetics.

Highlights

The clinical and genetic heterogeneity of many genetic disorders can hinder the determination of their molecular causes, complicating diagnosis for affected families[1]
Two other targeted sequencing panels: TruSight Cancer (TSCa), and Hereditary Cancer Solution by Sophia Genetics (HCS) were applied to hereditary cancers, and the Nextera Rapid Capture (NRC) panel was used for cardiopathies
Variant detection was performed with different commercial bioinformatics software programs depending on the sequencing panel: (1) the Illumina-pipeline and (2) the Sophia-pipeline (Sophia Genetics pipeline applied to Clinical Exome Solution (CES) and HCS)

Summary

Introduction

The clinical and genetic heterogeneity of many genetic disorders can hinder the determination of their molecular causes, complicating diagnosis for affected families[1]. CE sequencing has been reported to be a cost-effective first-tier molecular test[6–8] Despite these advances, the diagnostic yield of rare hereditary diseases using CE remains around 30–50% 5,9,10. There are several causes underlying this relatively low diagnostic yield, such as limitations of analytical methods (variant calling and annotation);[11,12] the genetic and phenotypic diversity of some genetic disorders;[13] knowledge gaps in gene-disease and variantdisease associations; and a lack of structured databases of these associations[14]. New bioinformatics developments and new disease related knowledge are needed in order to obtain a conclusive diagnosis for the majority of the unsolved cases. In this sense, automatic reanalysis might be a helpful and cost-effective tool to complement the process of diagnosis

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Conclusion