Successful application of genome sequencing in a diagnostic setting: 1007 index cases from a clinically heterogeneous cohort

Abstract

Despite clear technical superiority of genome sequencing (GS) over other diagnostic methods such as exome sequencing (ES), few studies are available regarding the advantages of its clinical application. We analyzed 1007 consecutive index cases for whom GS was performed in a diagnostic setting over a 2-year period. We reported pathogenic and likely pathogenic (P/LP) variants that explain the patients' phenotype in 212 of the 1007 cases (21.1%). In 245 additional cases (24.3%), a variant of unknown significance (VUS) related to the phenotype was reported. We especially investigated patients which had had ES with no genetic diagnosis (n = 358). For this group, GS diagnostic yield was 14.5% (52 patients with P/LP out of 358). GS should be especially indicated for ES-negative cases since up to 29.6% of them could benefit from GS testing (14.5% with P/LP, n = 52 and 15.1% with VUS, n = 54). Genetic diagnoses in most of the ES-negative/GS-positive cases were determined by technical superiority of GS, i.e., access to noncoding regions and more uniform coverage. Importantly, we reported 79 noncoding variants, of which, 41 variants were classified as P/LP. Interpretation of noncoding variants remains challenging, and in many cases, complementary methods based on direct enzyme assessment, biomarker testing and RNA analysis are needed for variant classification and diagnosis. We present the largest cohort of patients with GS performed in a clinical setting to date. The results of this study should direct the decision for GS as standard second-line, or even first-line stand-alone test.

Combining exome/genome sequencing with data repository analysis reveals novel gene–disease associations for a wide range of genetic disorders

“…When carrying out GS, genomic DNA was fragmented by sonication, and Illumina adapters were ligated to generated fragments for subsequent sequencing on the HiSeqX platform (Illumina) to yield an average coverage depth of at least 30×. Data analysis, including base calling, de-multiplexing, alignment to the hg19 human reference genome (Genome Reference Consortium GRCh37), and variant calling, was performed using the HiSeq Analysis Software v2.0 pipelines (Illumina, Inc., San Diego, CA), as previously described 8 (Supplementary information ).…”

An X‐linked syndrome with severe neurodevelopmental delay, hydrocephalus, and early lethality caused by a missense variation in the OTUD5 gene

“…GS was performed as described previously 7 . X‐inactivation analysis was performed by the Rare & Inherited Disease Genomic Laboratory in London, U.K., using blood samples from four females (V‐2, V‐13, IV‐2, IV‐14, Figure 1).…”

The importance of genetic testing for dystonia patients and translational research

“…Of note, WGS is even more comprehensive than WES and may detect disease-causing variants in deep intronic or relevant regulatory regions of dystonia genes that would be overlooked by sequencing exons alone. Indeed, the merits of using WGS even as a first-tier stand-alone genetic test are being increasingly recognized (Lionel et al 2018;Kumar et al 2019;Bertoli-Avella et al 2021). The conventional practice includes sequential testing that resorts to WGS only in patients who remain without a diagnosis after several other variant detection strategies have been exhausted [with reported medians of three tests and over 5000 US$ costs per patient (Lionel et al 2018)].…”

“…The conventional practice includes sequential testing that resorts to WGS only in patients who remain without a diagnosis after several other variant detection strategies have been exhausted [with reported medians of three tests and over 5000 US$ costs per patient (Lionel et al 2018)]. Clearly, this approach leads to a significant diagnostic delay that may average 5 years and in extreme cases reach 50 years (Bertoli-Avella et al 2021).…”