Expediting rare disease diagnosis: a call to bridge the gap between clinical and functional genomics

Hartin, Samantha N.; Means, John C.; Alaimo, Joseph T.; Younger, Scott T.

doi:10.1186/s10020-020-00244-5

Cited by 18 publications

(14 citation statements)

References 39 publications

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“…Aberrant splicing is a common outcome in the presence of exonic or intronic variants that might hamper the intricate network of interactions defining an exon in a specific gene context. Therefore, the evaluation of the functional, and potentially pathological, role of nucleotide changes remains one of the major challenges in the modern genomic era (Taneri et al 2012 ; Hartin et al 2020 ). This aspect has also to be taken into account during the pre-clinical evaluation of innovative therapeutic approaches in animal models of human diseases.…”

Section: Discussionmentioning

confidence: 99%

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

Sacchetto

Peretto²,

Baralle³

et al. 2021

Mol Med

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Background Aberrant splicing is a common outcome in the presence of exonic or intronic variants that might hamper the intricate network of interactions defining an exon in a specific gene context. Therefore, the evaluation of the functional, and potentially pathological, role of nucleotide changes remains one of the major challenges in the modern genomic era. This aspect has also to be taken into account during the pre-clinical evaluation of innovative therapeutic approaches in animal models of human diseases. This is of particular relevance when developing therapeutics acting on splicing, an intriguing and expanding research area for several disorders. Here, we addressed species-specific splicing mechanisms triggered by the OTC c.386G>A mutation, relatively frequent in humans, leading to Ornithine TransCarbamylase Deficiency (OTCD) in patients and spfash mice, and its differential susceptibility to RNA therapeutics based on engineered U1snRNA. Methods Creation and co-expression of engineered U1snRNAs with human and mouse minigenes, either wild-type or harbouring different nucleotide changes, in human (HepG2) and mouse (Hepa1-6) hepatoma cells followed by analysis of splicing pattern. RNA pulldown studies to evaluate binding of specific splicing factors. Results Comparative nucleotide analysis suggested a role for the intronic +10-11 nucleotides, and pull-down assays showed that they confer preferential binding to the TIA1 splicing factor in the mouse context, where TIA1 overexpression further increases correct splicing. Consistently, the splicing profile of the human minigene with mouse +10-11 nucleotides overlapped that of mouse minigene, and restored responsiveness to TIA1 overexpression and to compensatory U1snRNA. Swapping the human +10-11 nucleotides into the mouse context had opposite effects. Moreover, the interplay between the authentic and the adjacent cryptic 5′ss in the human OTC dictates pathogenic mechanisms of several OTCD-causing 5′ss mutations, and only the c.386+5G>A change, abrogating the cryptic 5′ss, was rescuable by engineered U1snRNA. Conclusions Subtle intronic variations explain species-specific OTC splicing patterns driven by the c.386G>A mutation, and the responsiveness to engineered U1snRNAs, which suggests careful elucidation of molecular mechanisms before proposing translation of tailored therapeutics from animal models to humans.

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Section: Discussionmentioning

confidence: 99%

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

Sacchetto

Peretto²,

Baralle³

et al. 2021

Mol Med

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“…Such screening approaches include germline mutagenesis, CRISPR/CAS9, plasmid-based reporter assays, RNA interference, chemical screens [ 249 ], and multiplexed assays of variant effect [ 250 , 251 ]. Advancements in CRISPR/Cas9 technology make it a robust tool to profile cellular phenotypes resulting from each of the thousand genetic perturbations in a high-throughput manner [ 252 ]. The underlying principle behind CRISPR screens [ 253 , 254 ] is to introduce thousands of variants in a large cell population but only one gene is perturbed per cell.…”

Section: Functional Studiesmentioning

confidence: 99%

A guide for the diagnosis of rare and undiagnosed disease: beyond the exome

2022

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Rare diseases affect 30 million people in the USA and more than 300–400 million worldwide, often causing chronic illness, disability, and premature death. Traditional diagnostic techniques rely heavily on heuristic approaches, coupling clinical experience from prior rare disease presentations with the medical literature. A large number of rare disease patients remain undiagnosed for years and many even die without an accurate diagnosis. In recent years, gene panels, microarrays, and exome sequencing have helped to identify the molecular cause of such rare and undiagnosed diseases. These technologies have allowed diagnoses for a sizable proportion (25–35%) of undiagnosed patients, often with actionable findings. However, a large proportion of these patients remain undiagnosed. In this review, we focus on technologies that can be adopted if exome sequencing is unrevealing. We discuss the benefits of sequencing the whole genome and the additional benefit that may be offered by long-read technology, pan-genome reference, transcriptomics, metabolomics, proteomics, and methyl profiling. We highlight computational methods to help identify regionally distant patients with similar phenotypes or similar genetic mutations. Finally, we describe approaches to automate and accelerate genomic analysis. The strategies discussed here are intended to serve as a guide for clinicians and researchers in the next steps when encountering patients with non-diagnostic exomes.

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“…6 It is estimated that approximately 400 million people live with a RD globally, and on average, it can take up to 5 years for people to receive an accurate diagnosis. 7,8 Though more than 7000 RDs have been identified, only 10% of these conditions have US Food and Drug Administration (FDA) approved treatments. 9 These barriers, along with an overall paucity of medical knowledge, research, and literature, leave millions of people living with RDs with inadequate resources to manage their care.…”

Section: Defining Disparitiesmentioning

confidence: 99%

What Rare Disease Patient Advocacy Groups Are Doing to Mitigate the Effects of Disparities

Drell

Silva

Lee

2022

Advances in Pulmonary Hypertension

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Compared with chronic diseases affecting larger populations, rare disease (RD) patients experience great inequities in diagnosis, care, and research. Within RDs, health disparities compound these inequities, as marginalized communities experience additional barriers in accessing clinical care and are often underrepresented from participation in research and clinical trials. For almost 40 years, the National Organization for Rare Disorders (NORD), a RD umbrella organization with over 300 nonprofit organizational members, has led efforts to understand and address inequities for the RD community through innovative research, programming, and collaboration with patients, caregivers, practitioners, and external stakeholders. The beginning of the COVID-19 pandemic in 2020 brought to light longstanding disparities and discrimination for marginalized communities as well as pivotal racial justice movements. These events spurred many RD nonprofit organizations’ interest in increasing outreach and engagement with minoritized communities within RDs and diversifying their organization internally. Building on the increased interest in diversity, equity, and inclusion (DEI), NORD has focused on collecting case studies from within NORD and its member organizations to capture current efforts to improve DEI within the RD ecosystem. One way clinicians can work to mitigate the effects of disparities is to collaborate with RD patient organizations; this article provides a means by which clinicians and researchers can understand some of the challenges RD nonprofit organizations face in bridging disparities and learn about solutions to supporting marginalized patients within their communities. Clinicians are encouraged to join NORD in our policy efforts advocating to ensure patient access to health care providers practicing in a different state vis-à-vis telehealth.

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Expediting rare disease diagnosis: a call to bridge the gap between clinical and functional genomics

Cited by 18 publications

References 39 publications

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

A guide for the diagnosis of rare and undiagnosed disease: beyond the exome

What Rare Disease Patient Advocacy Groups Are Doing to Mitigate the Effects of Disparities

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