Test Information: Exome sequencing is used to identify an underlying genetic cause for an individual’s clinical presentation. It is indicated for individuals with a genetically heterogeneous disease, individuals with a long list of differential diagnoses, individuals with an atypical presentation of a genetic disorder, and individuals who have exhausted other currently available genetic testing options. This test targets the exons, which are the protein coding regions of the genome.
This test is a phenotype-driven analysis. Parental samples are used to create filters aimed at identifying de novo variants and to determine the inheritance of identified variants. Variants included on the report are limited to those identified in genes related to the reported phenotype of the individual and classified as either likely pathogenic or pathogenic.
Secondary findings are likely pathogenic and pathogenic variants identified in one or more of the genes included in the American College of Medical Genetics (ACMG) list of genes that are suggested as returnable results for individuals undergoing whole exome/genome sequencing (PMID: 27854360). Secondary findings are only reported if the patient opts in to receive them. If parents elect to receive secondary findings, they will receive a separate report with their results.
Incidental findings are genetic changes that are of medical importance that are not directly relevant to the indication for testing that are incidentally detected during testing. Incidental findings will be reported if they have significant healthcare implications for the patient.
Methodology: This test has been designed to assess variants occurring within protein coding regions and splice junctions of RefSeq genes. Nucleic acid from the submitted specimen is enriched for coding and adjacent noncoding (splice) regions of the genes using the IDT xGen Exome (Integrated DNA Technologies, Coralville, IA). The library products are sequenced with 2 by 150 bp reads using either the NextSeq or NovaSeq sequencing instruments (Illumina, San Diego, CA). After alignment to the reference genome (GRCh37/hg19), off target, low quality and duplicate reads are removed from the analysis. The targeted regions are assessed for average depth of coverage and other data quality thresholds. Variants are called using several different variant calling algorithms, including ones developed by Invitae.
This test detects single nucleotide substitutions (SNVs) and small insertions, deletions, and indels located in the targeted regions. Sequence alterations are described according to the Human Genome Variation Society (HGVS) nomenclature guidelines and are classified according to the standards and guidelines for the interpretation and reporting of sequencing variants by joint ACMG/AMP guidelines (PMID: 25741868). Variants are filtered based on the clinical information provided by the ordering clinician. Parental samples are used to create filters aimed at identifying de novo variants and to determine the inheritance of identified variants. Only variants in genes associated with diseases that overlap with the phenotype information provided that are classified as likely pathogenic or pathogenic are included on the report. Variants of uncertain significance are reported at the discretion of the laboratory. Clinically significant variants included on the report are confirmed by an appropriate orthogonal method.
Secondary findings are only reported if elected by the patient. Under certain circumstances, such as a single pathogenic variant identified in a gene with autosomal recessive inheritance, a likely pathogenic or pathogenic variant in one of these genes may not be reported. The secondary findings gene list is subject to change over time as it is updated by the ACMG. The current gene list for secondary findings in the following 78 genes ( PMID: 35802134): ACTA2, ACTC1, ACVRL1, APC, APOB, ATP7B, BAG3, BMPR1A, BRCA1, BRCA2, BTD, CACNA1S, CASQ2, COL3A1, DES, DSC2, DSG2, DSP, ENG, FBN1, FLNC, GAA, GLA, HFE (p.Cys282Tyr homozygotes only), HNF1A, KCNH2, KCNQ1, LDLR, LMNA, MAX, MEN1, MLH1, MSH2, MSH6, MUTYH, MYBPC3, MYH11, MYH7, MYL2, MYL3, NF2, OTC, PALB2, PCSK9, PKP2, PMS2, PRKAG2, PTEN, RB1, RBM20, RET, RPE65, RYR1, RYR2, SCN5A, SDHAF2, SDHB, SDHC, SDHD, SMAD3, SMAD4, STK11, TNNC1, TGFBR1, TGFBR2, TMEM127, TMEM43, TNNI3, TNNT2, TP53, TPM1, TRDN, TSC1, TSC2, TTN, TTR, VHL, WT1 .
Incidental findings are distinct from secondary findings which are included when elected by the patient. All incidental findings are reported at the discretion of the geneticist signing out the report and/or the laboratory director. In general, the following types of incidental findings will NOT be reported: those associated with adult-onset disorders for which there are no interventions available, those associated with adult-onset disorders when the patient is a child, and carrier status variants in genes unrelated to the reported phenotype.
The test was developed, and its performance characteristics were determined by Invitae. The report and methods used to generate the report, have not been cleared or approved by the U.S. Food and Drug Administration (FDA). The FDA has determined that such clearance or approval is not necessary. This test is used for both clinical and clinical research purposes. Pursuant to the requirements of federal regulations listed in the Clinical Laboratory Improvement Amendments of 1988 (CLIA), this laboratory has established and verified this test's accuracy and precision. However, the possibility of a false positive or false negative result incurred during any phase of the testing cannot be completely excluded.
Limitations: A negative or inconclusive result does not eliminate the possibility of a genetic basis for disease in this individual and does not guarantee present or future health. Some types of genomic aberrations are not be detectable by the technologies used to perform this test. This test is not intended to detect the following types of genetic variants: deletions/insertions of >15 bp, gross rearrangements, variants in promotor regions and other non-coding regions, copy number variations, variations occurring within repetitive sequences and repeat expansion mutations, variants in regions with high homology or in genes with pseudogenes, mitochondrial genome mutations, epigenetic effects, mosaic or somatic events, and other complex aberrations. In addition, not all regions of the exome are fully covered by this analysis and variants in regions of low coverage may not be detected.
The interpretation of this test is based on the assumption that the clinical information and family relationships provided to the laboratory are accurate. Not all variants identified have been analyzed. In addition, the interpretation of these results is based on currently available scientific knowledge. Not all disease-associated genes have been identified and the clinical significance of variation in many genes is not well understood. Variant interpretation may change over time if more information becomes available. This test is not intended to examine variants associated with non-Mendelian patterns of inheritance. It is recommended that genetic test results be periodically reinterpreted, particularly if new symptoms arise. Healthcare providers can contact the laboratory to determine if there have been any changes to the interpretation of a variant or test result.
Although rare, the accuracy of results may also be impacted due to sample mix-up, DNA contamination, poor DNA quality, or the presence of pre-malignant or malignant cells in the sample, as well as in the setting of bone marrow transplantation or a recent blood transfusion.
Laboratory Information: Invitae is a College of American Pathologists (CAP)-accredited and Clinical Laboratory Improvement Amendments (CLIA)-certified clinical diagnostic laboratory (CAP#: 8289311; CLIA #31D2142534). This test was performed at Invitae located at 485F US Route 1 South, Suite 110, Iselin, NJ 08830.