Exome Sequencing

Exome sequencing may be used as a first-line test when a genetic condition is suspected but the proband’s (patient's) clinical features are not suggestive of a single disorder. The entire exome, including all known nuclear genes (approximately 1-2% of the human genome), is analyzed to identify the causative variant(s). Submission of comparator samples from the proband’s parents, and, when relevant, other similarly affected family members, is recommended to aid in results interpretation.

The American College of Medical Genetics and Genomics (ACMG) strongly recommends exome or genome sequencing as a first- or second-tier test for probands with congenital anomalies or developmental delay/intellectual disability. ^, 

The American Academy of Pediatrics supports exome or genome sequencing as a first-tier test for children with congenital anomalies or global developmental delay/intellectual disability. 

Test Overview

• Approximately 85% of pathogenic variants can be detected by exome sequencing.
  • The function of approximately 4,500 genes is currently known.
• Results of exome sequencing may or may not:
  • Identify the etiology of the proband's clinical presentation
  • Determine prognosis
  • Predict the severity of the proband's condition
  • Guide medical management, including disease surveillance
  • Provide information about recurrence risk

Test Submission Requirements

A completed Exome Sequencing Intake Form is required for the proband.

Parental/familial comparator specimens: Submission of parental specimens is recommended to identify de novo variants and the chromosomal phase of variants, and to optimally interpret proband results. Specimens from additional informative relatives (e.g., family members suspected to have the same condition as the proband) may also be submitted as comparators to aid interpretation. Submit comparator specimens within 7 days of the proband's specimen.

Medical records: Medical records detailing the proband’s clinical findings, relevant previous testing/imaging results, and family history are required for optimal interpretation of the proband’s results. The ability to identify causative variant(s) for the proband’s presentation is influenced by the quality of the clinical information provided.

Informed consent: Healthcare provider attestation of informed consent is required. Reporting of secondary findings (SF) is available for the proband and familial comparators, if desired.

Test Description

Methodology

• Targeted capture of all (or selected) coding exons and exon-intron junctions of the targeted genes is performed, followed by massively parallel sequencing (MPS).
• Sanger sequencing is performed as necessary to confirm reported variants.
• Human genome build 19 (Hg 19) is used for data analysis.

Clinical Sensitivity

Clinical sensitivity varies based on clinical testing indication, family history, previous clinical evaluations, and availability of parental comparator specimens.

• A diagnosis is determined in approximately 20-40% of individuals; higher diagnostic rates are reported when parental specimens are submitted as exome sequencing comparators. ^, , 
• Mode of inheritance, reduced penetrance, and genetic heterogeneity can reduce clinical sensitivity.

Reporting

Primary Findings

• Variants that are known or suspected to be causative for the proband’s provided clinical presentation are reported.
  • Inheritance of primary findings is reported when this information is available using familial comparator specimen data.
  • Familial comparator individuals are not issued clinical reports for inherited primary findings identified in the proband.
• Candidate variants that are not known to be causative for the proband’s phenotype may be reported (e.g., de novo variants in a gene of uncertain significance, trans-heterozygous variants of uncertain significance in an autosomal recessive gene, heterozygous variants in an autosomal recessive gene that has clinical overlap with the proband’s phenotype).

Secondary Findings

SF refer to medically actionable disease-associated variants that are not associated with clinical findings or phenotypes provided upon submission for phenotype-driven analysis from exome testing. The ACMG recommends reporting of certain variants involving genes from a specified, curated gene list for consented individuals undergoing exome sequencing. 

The current list utilized in analysis can be found in the ACMG Secondary Findings Gene List supplemental resource.

Medically actionable variants involving genes outside of the ACMG SF list, which do not overlap with submitted clinical findings and phenotypes (non-ACMG SF), may also be reported as SF.

• The Exome Sequencing Intake Form provides the option for each individual sequenced (proband and familial comparators) to opt out of receiving SF.
  • SF will be reported for any individual(s) sequenced unless “opt out” is selected on the exome intake form.
  • SF reported in a proband will generally include inheritance information unless one or both parents do not elect to receive SF.
  • ACMG SF will be reported for familial comparators who elect to receive this information regardless of whether the SF was also identified in the proband.
• Heterozygous, single variants involving autosomal recessive ACMG SF genes are generally not reported.

Interpretation and Reanalysis

• Disclosure of biological relationships among tested family members is imperative for accurate analysis and interpretation. Misattributed parentage may impact results interpretation.
• Variant classification and results interpretation for exome sequencing is phenotype-driven and based on clinical information provided at the time of testing. As phenotype information may change over time, reanalysis is available.
• Data reanalysis is available for probands whose testing was originally performed at ARUP Laboratories after April 1, 2015.
  • Order Exome Reanalysis (Originally Tested at ARUP - No Specimen Required) (ARUP test code 3001457).
  • The first request for exome data reanalysis is available at no cost; additional charges will apply for subsequent reanalysis requests.
  • Reanalysis is recommended for undiagnosed individuals 12-18 months after completion of initial testing or upon encounter of new significant clinical findings.

Storage and Data Sharing

• Raw exome sequencing data will be stored for a minimum of 5 years in compliance with ARUP’s data retention policy.
• Raw exome sequencing data may be requested by the ordering healthcare provider and/or institution that submitted testing to ARUP.
• Original or derivative samples from patient testing may be used for internal test validation, research, and/or training purposes. All samples are deidentified before use outside of clinical testing. Individuals may request that their sample be discarded after testing by contacting ARUP Laboratories at 800-242-2787 ext. 3301.
• ARUP participates in responsible genetic data sharing through submission of deidentified variant-level information to publicly accessible databases such as ClinVar.
  • Individuals may request to opt out of data sharing by calling ARUP Laboratories at 800-242-2787 ext. 3301.
  • Depending on timing of requests, variant records will be either omitted or removed from external databases.
  • Please note that separate opt-out requests are generally required for each genetic test performed at ARUP.
• Probands have the opportunity to participate in proband registries and research. For more information, refer to the ARUP Genetics web page. 

Analytic Sensitivity

The analytic sensitivity of this test is approximately 98% for single nucleotide variants (SNVs) and greater than 93% for insertions/duplications/deletions from 1-10 base pairs in size. Deletions/duplications greater than 10 base pairs may be detected, but the analytic sensitivity may be reduced.

Limitations

• A negative result does not exclude a genetic cause for the proband’s condition.
• The human exome cannot be completely analyzed.
  • Some genes have not been identified.
  • Some genes cannot be sequenced or interpreted due to technical limitations.
• The following will not be evaluated:
  • Variants outside the coding regions and intron-exon boundaries
  • Large deletions/duplications
  • Mitochondrial DNA (mtDNA)
• The following may not be detected:
  • Deletions/duplications/insertions of any size by MPS
  • Some variants due to the presence of pseudogenes, repetitive, or homologous regions
  • Low-level somatic variants
  • Chromosomal phase of identified variants
  • Pathogenic ACMG SF variants that cannot be detected by routine exome analysis
• Results interpretation may be impacted if the tested individual has had an allogeneic stem cell transplant.
• Diagnostic errors can occur due to rare sequence variations.
• Results interpretation may be impacted by the absence of parental data, whether through nonsubmission of parental comparator samples, technical failure, or misattributed parentage.