Human Mutation

Scott, Stuart A.; Wang, Kai; Spinner, Nancy B.

doi: 10.1002/humu.24485pmid: N/A

doi: 10.1002/humu.24233pmid: N/A

Scott, Stuart A.; Wang, Kai; Spinner, Nancy B.

doi: 10.1002/humu.24474pmid: 36116036

This special issue of Human Mutation focuses on Innovations in Genomic Diagnostics. The increasing interest in genomic medicine, and the growing possibilities for treatment and management of genetic disease, make complete and accurate diagnosis mission critical. This issue describes leading‐edge technologies with emerging utility for genomic diagnostics. Genomic testing has dramatically evolved as a result of advances in technology, data analytics, and the continuing pace of disease gene discovery. Since 2011, clinical laboratories have increasingly employed next‐generation sequencing‐based tests in addition to historical techniques to identify a spectrum of germline and somatic variants implicated in human disease. However, common testing platforms have known limitations, including failure to detect disease‐causing variants in certain regions, inability to identify all variant types, variant phasing, measuring epigenetic changes, and ongoing challenges with variant interpretation. Innovative solutions are emerging, including increasingly rapid genome sequencing, long‐read sequencing, clinical RNA sequencing, epigenomic profiling, facial phenotyping, and an array of computational tools for variant identification and interpretation.

Sanford Kobayashi, Erica F.; Dimmock, David P.

doi: 10.1002/humu.24422pmid: 35723630

The rapid pace of advancement in genomic sequencing technology has recently reached a new milestone, with a record‐setting time to molecular diagnosis of a mere 8 h. The catalyst behind this achievement is the accumulation of evidence indicating that quicker results more often make an impact on patient care and lead to healthcare cost savings. Herein, we review the diagnostic and clinical utility of rapid whole genome and rapid whole exome sequencing, the associated reduction in healthcare costs, and the relationship between these outcome measures and time‐to‐diagnosis.

Jezkova, Jana; Shaw, Sophie; Taverner, Nicola V.; Williams, Hywel J.

doi: 10.1002/humu.24466pmid: 36086948

The advancements made in next‐generation sequencing (NGS) technology over the past two decades have transformed our understanding of genetic variation in humans and had a profound impact on our ability to diagnose patients with rare genetic diseases. In this review, we discuss the recently developed application of rapid NGS techniques, used to diagnose pediatric patients with suspected rare diseases who are critically ill. We highlight the challenges associated with performing such clinical diagnostics tests in terms of the laboratory infrastructure, bioinformatic analysis pipelines, and the ethical considerations that need to be addressed. We end by looking at what future developments in this field may look like and how they can be used to augment the genetic data to further improve the diagnostic rates for these high‐priority patients.

Hou, Ying‐Chen C.; Neidich, Julie A.; Duncavage, Eric J.; Spencer, David H.; Schroeder, Molly C.

doi: 10.1002/humu.24381pmid: 35471774

Characterizing the genomic landscape of cancers is a routine part of clinical care that began with the discovery of the Philadelphia chromosome and has since coevolved with genomic technologies. Genomic analysis of tumors at the nucleotide level using DNA sequencing has revolutionized the understanding of cancer biology and identified new molecular drivers of disease that have led to therapeutic advances and improved patient outcomes. However, the application of next‐generation sequencing in the clinical laboratory has generally been limited until very recently to targeted analysis of selected genes. Recent technological innovations and reductions in sequencing costs are now able to deliver the long‐promised goal of tumor whole‐genome sequencing as a practical clinical assay.

Conlin, Laura K.; Aref‐Eshghi, Erfan; McEldrew, Deborah A.; Luo, Minjie; Rajagopalan, Ramakrishnan

doi: 10.1002/humu.24465pmid: 36086952

Long‐read sequencing (LRS) has been around for more than a decade, but widespread adoption of the technology has been slow due to the perceived high error rates and high sequencing cost. This is changing due to the recent advancements to produce highly accurate sequences and the reducing costs. LRS promises significant improvement over short read sequencing in four major areas: (1) better detection of structural variation (2) better resolution of highly repetitive or nonunique regions (3) accurate long‐range haplotype phasing and (4) the detection of base modifications natively from the sequencing data. Several successful applications of LRS have demonstrated its ability to resolve molecular diagnoses where short‐read sequencing fails to identify a cause. However, the argument for increased diagnostic yield from LRS remains to be validated. Larger cohort studies may be required to establish the realistic boundaries of LRS's clinical utility and analytical validity, as well as the development of standards for clinical applications. We discuss the limitations of the current standard of care, and contrast with the applications and advantages of two major LRS platforms, PacBio and Oxford Nanopore, for molecular diagnostics of constitutional disorders, and present a critical argument about the potential of LRS in diagnostic settings.

Holt, Giles S.; Batty, Lois E.; Alobaidi, Bilal K. S.; Smith, Hannah E.; Oud, Manon S.; Ramos, Liliana; Xavier, Miguel J.; Veltman, Joris A.

doi: 10.1002/humu.24450pmid: 36047340

De novo mutations (DNMs) play an important role in severe genetic disorders that reduce fitness. To better understand their role in disease, it is important to determine the parent‐of‐origin and timing of mutational events that give rise to these mutations, especially in sex‐specific developmental disorders such as male infertility. However, currently available short‐read sequencing approaches are not ideally suited for phasing, as this requires long continuous DNA strands that span both the DNM and one or more informative single‐nucleotide polymorphisms. To overcome these challenges, we optimized and implemented a multiplexed long‐read sequencing approach using Oxford Nanopore technologies MinION platform. We focused on improving target amplification, integrating long‐read sequenced data with high‐quality short‐read sequence data, and developing an anchored phasing computational method. This approach handled the inherent phasing challenges of long‐range target amplification and the normal accumulation of sequencing error associated with long‐read sequencing. In total, 77 of 109 DNMs (71%) were successfully phased and parent‐of‐origin identified. The majority of phased DNMs were prezygotic (90%), the accuracy of which is highlighted by an average mutant allele frequency of 49.6% and standard error of 0.84%. This study demonstrates the benefits of employing an integrated short‐read and long‐read sequencing approach for large‐scale DNM phasing.

Scott, Erick R.; Yang, Yao; Botton, Mariana R.; Seki, Yoshinori; Hoshitsuki, Keito; Harting, John; Baybayan, Primo; Cody, Neal; Nicoletti, Paola; Moriyama, Takaya; Chakraborty, Shreyasee; Yang, Jun J.; Edelmann, Lisa; Schadt, Eric E.; Korlach, Jonas;

Eisfeldt, Jesper; Rezayee, Fatemah; Pettersson, Maria; Lagerstedt, Kristina; Malmgren, Helena; Falk, Anna; Grigelioniene, Giedre; Lindstrand, Anna

doi: 10.1002/humu.24440pmid: 35842787

Prader–Willi syndrome (PWS; MIM# 176270) is a neurodevelopmental disorder caused by the loss of expression of paternally imprinted genes within the PWS region located on 15q11.2. It is usually caused by either maternal uniparental disomy of chromosome 15 (UPD15) or 15q11.2 recurrent deletion(s). Here, we report a healthy carrier of a balanced X;15 translocation and her two daughters, both with the karyotype 45,X,der(X)t(X;15)(p22;q11.2),−15. Both daughters display symptoms consistent with haploinsufficiency of the SHOX gene and PWS. We explored the architecture of the derivative chromosomes and investigated effects on gene expression in patient‐derived neural cells. First, a multiplex ligation‐dependent probe amplification methylation assay was used to determine the methylation status of the PWS‐region revealing maternal UPD15 in daughter 2, explaining her clinical symptoms. Next, short read whole genome sequencing and 10X genomics linked read sequencing was used to pinpoint the exact breakpoints of the translocation. Finally, we performed transcriptome sequencing on neuroepithelial stem cells from the mother and from daughter 1 and observed biallelic expression of genes in the PWS region (including SNRPN) in daughter 1. In summary, our multi‐omics analysis highlights two different PWS mechanisms in one family and provide an example of how structural variation can affect imprinting through long‐range interactions.