PGxome® - Whole Exome Sequencing

Test Requisition Form

PGxome® Prenatal Exome Test

Name Test Code Description CPT Code(s) Price
Family - Trio 14013 WES of patient + 2 additional family members 81415, 81416x2, 81265 $3,590

If report is needed for any additional family members, add $490 per family member.

Family - Duo 14012 WES of patient + 1 additional family member 81415, 81416, 81265 $3,360

If report is needed for any additional family members, add $490 per family member.

Patient Only 14010 WES of patient 81415, 81265 $2,860

Sequencing cost to additional family members beyond trio: $390 (no report); additional CPT Code 81416

We require at least one parental specimen be sent as part of prenatal testing for quality assurance purposes. A maternal specimen is strongly encouraged to be sent for maternal cell contamination (MCC) testing of the fetal specimen. We do not charge extra for MCC studies for any fetal testing, but the CPT Code 81265 will be included on invoices and insurance claims as appropriate.

PGxome - Prenatal is ideal for:

  • Ongoing pregnancies with ultrasound findings consistent with a serious genetic condition.
  • In the case of fetal demise or pregnancy termination, PGxome Diagnostic can be ordered.

For Ordering via paper requisition: Prenatal PGxome Test Requisition Form & Provider Statement

What is PGxome Prenatal?

PGxome is PreventionGenetics' whole exome sequencing (WES) test. The PGxome assesses almost all genes from the human genome including coding regions and adjacent introns. This test is an appropriate choice for health care providers who are looking for an urgent genetic diagnosis. This is important as more than 50% of patients with genetic diseases are not given a specific diagnosis even after repeat clinical examinations and tests (Shashi et al. 2014). The standard clinical practice often involves examinations for specific phenotypes, imaging, biochemical testing for inborn errors of metabolism, genomic tests such as karyotyping or microarrays, and single gene or panel tests (Iglesias et al. 2014). However, patients remain without a genetic diagnosis, and patients and health care providers are caught in a long term diagnosis search, known as a diagnostic odyssey. This can lead to failures in identifying potential treatments and unknown recurrence and prognosis risks (Yang et al. 2013).

Reported diagnostic rates from commercial and academic laboratories have found that WES assays have a ~20-40% positive diagnostic rate, with higher rates being reported from trio analysis (i.e. proband and parents) compared to singleton analysis (Atwal et al. 2014; Iglesias et al. 2014; Farwell et al. 2015). Notably, ~5-7% of individuals who have WES have had dual diagnoses (i.e. two non-overlapping clinical presentations) (Yang et al. 2014; Farwell et al. 2015; Posey et al. 2016). The inclusion of copy number variant (CNV) calling should increase diagnostic rates. One study reported that 30% of genetics diagnoses have only been recently resolved due to new literature reports, highlighting the fast pace of gene-disease discovery, and the need of genetic testing laboratories to be current of the medical literature (Yang et al. 2014). The use of a whole exome sequencing test may aid in altering clinical management, predict recurrence and prognosis risks, reduce costs of additional testing, and may offer advantages over traditional molecular tests in certain patients (Valencia et al. 2015).

PGxome - Prenatal is ideal for:

  • Ongoing pregnancies with ultrasound findings consistent with a serious genetic condition.
  • In the case of fetal demise or pregnancy termination, PGxome Diagnostic can be ordered.

For Ordering via paper requisition: Prenatal PGxome Test Requisition Form & Provider Statement

TURN AROUND TIME (TAT)

PGxome Prenatal has an expedited TAT of 13 calendar days on average.

We recommend that providers choose expedited shipping to decrease the time samples spend in transit to PreventionGenetics.

Inclusion of detailed clinical notes/completion of the clinical data checklist and a pedigree are required. The ability to select variants that may be involved with the fetal health problem directly correlates with the quality of clinical information provided.

ORDERING / SPECIMENS

Our PGxome Prenatal offers the traditional options of Patient Only testing or Family testing (e.g., Duo, Trio, etc.). For the highest diagnostic rate, Family - Trio testing is recommended.

Specimen Requirements and Shipping Details

 

TEST METHODS

For the PGxome we use Next Generation Sequencing (NGS) technologies to cover the coding regions of targeted genes plus ~10 bases of non-coding DNA flanking each exon. As required, genomic DNA is extracted from patient specimens. Patient DNA corresponding to these regions is captured using hybridization probes. Captured DNA is sequenced on the NovaSeq 6000 using 2x150 bp paired-end reads (Illumina, San Diego, CA, USA). The following quality control metrics are generally achieved: >97% of target bases are covered at >20x, and mean coverage of target bases >100x. Data analysis and interpretation is performed by the internally developed Infinity pipeline. Variant calls are made by the GATK Haplotype caller and annotated using in house software and Jannovar. Common benign, likely benign, and low quality variants are filtered from analysis.

Copy number variants (CNVs) are also detected from NGS data. We utilize a CNV calling algorithm that compares mean read depth and distribution for each target in the test sample against multiple matched controls. Neighboring target read depth and distribution and zygosity of any variants within each target region are used to reinforce CNV calls. All reported CNVs are confirmed using another technology such as aCGH, MLPA, or PCR.

REPORTING

Reports will consist of up to four different sections:

Primary Findings (related to the indication for testing)

  • Variants in genes known to be associated with phenotype
  • Variants in genes possibly associated with phenotype

Pathogenic, likely pathogenic and uncertain variants are reported.

Secondary Findings (if opted in on the requisition form)

  • Guideline Recommended Genes: The American College of Medical Genetics and Genomics recommends all labs performing WES or WGS report pathogenic variants in specific  genes that cause certain, mostly dominantly inherited disorders (Version 3.1, Miller et al. 2022. PubMed ID: 35802134). These disorders are treatable and/or preventable. Included on this list are some cancer predisposition conditions, heart conditions associated with sudden death, and conditions that could result in severe health consequences if surgery is performed with certain anesthetics. Only pathogenic and likely pathogenic variants are reported.
  • Childhood Onset Disorders: The American College of Medical Genetics and Genomics recommends all labs performing prenatal WES report pathogenic or likely pathogenic variants detected in genes unrelated to the fetal clinical features, but known to cause moderate to severe childhood onset disorders (Monaghan et al. 2020. PubMed ID: 31911674). Many of these disorders, especially those associated with nonsyndromic intellectual disability/neurodevelopmental disorders and metabolic conditions, are not detectable with fetal imaging. If this option is selected, we will report pathogenic and likely pathogenic variants in genes that would result in a Mendelian (single gene) childhood onset disorder (i.e., one variant in a dominant gene or X-linked gene or two variants in a recessive gene). Only pathogenic and likely pathogenic variants are reported.

A preliminary report prior to confirmation may be issued in cases with a clear positive finding.

All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories (pathogenic, likely pathogenic, variant of uncertain significance, likely benign and benign) per ACMG guidelines (Richards et al. 2015). Benign and likely benign variants are not reported.  Sequencing data is available to the ordering physician upon request.

Nomenclature for sequence variants comes from Human Genome Variation Society (HGVS) (http://www.hgvs.org).

LIMITATIONS AND OTHER TEST NOTES

Interpretation of the test results is limited by the information that is currently available. Better interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder are accumulated.

Sequencing: When sequencing does not reveal any heterozygous differences from the reference sequence, we cannot be certain that we were able to detect both patient alleles.

For technical reasons, the PGxome test is not 100% sensitive. Some exons cannot be efficiently captured, and some genes cannot be accurately sequenced because of the presence of multiple copies in the genome. Therefore, a small fraction of sequence variants relevant to the patient's health will not be detected.

We sequence coding exons for most given transcripts, plus ~10 bp of flanking non-coding DNA for each exon. Unless specifically indicated, test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions, uncharacterized alternative exons, chromosomal rearrangements, repeat expansions, epigenetic effects, and mitochondrial genome variants.

In most cases, we are unable to determine the phase of sequence variants. In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.

The ability to detect low-level mosaicism of variants is limited.

Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during amplification.

Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes if taken from whole blood). Test reports contain no information about the DNA sequence in other cell-types.

We cannot be certain that the reference sequences are correct.

Copy Number Variant Analysis: The PGxome test detects most deletions and duplications including intragenic CNVs and large cytogenetic events; however aberrations in a small percentage of regions may not be accurately detected due to sequence paralogy (e.g., pseudogenes, segmental duplications), sequence properties, deletion/duplication size (e.g., 1-3 exons vs. 4 or more exons), and inadequate coverage. In general, sensitivity for single, double, or triple exon CNVs is ~70% and for CNVs of 4 exons or larger is >95% but may vary from gene-to-gene based on exon size, depth of coverage, and characteristics of the region.

Balanced translocations or inversions are only rarely detected.

Certain types of sex chromosome aneuploidy may not be detected.

In nearly all cases, our ability to determine the exact copy number change within a targeted region is limited.

Our ability to detect CNVs due to somatic mosaicism is limited.

The sensitivity of this test is dependent on DNA quality.

CONTACTS

Genetic Counselors: GC Team - support@preventiongenetics.com

Geneticist: Diane Allingham-Hawkins, PhD, FCCMG, FACMG - diane.allingham-hawkins@preventiongenetics.com

REFERENCES

Atwal P.S. et al. 2014. Genetics in Medicine : Official Journal of the American College of Medical Genetics. 16: 717-9. PubMed ID: 24525916

Caudle et al. 2016. Genetics in Medicine. PubMed ID: 27441996

Farwell K.D. et al. 2015. Genetics in Medicine : Official Journal of the American College of Medical Genetics. 17: 578-86. PubMed ID: 25356970

Iglesias A. et al. 2014. Genetics in Medicine : Official Journal of the American College of Medical Genetics. 16: 922-31. PubMed ID: 24901346

Kalia S.S. et al. 2016. Genetics in Medicine: Official Journal of the American College of Medical Genetics. Advance online publication. doi:10.1038/gim.2016.190. PubMed ID: 27854360

Miller D. et al. 2022. Genetics in Medicine : Official Journal of the American College of Medical Genetics. PubMed ID: 35802134

Posey et al. 2016. Genetics in Medicine : Official Journal of the American College of Medical Genetics. 18: 678-85. PubMed ID: 26633545

Richards S et al. 2015. Genetics in Medicine : Official Journal of the American College of Medical Genetics. 17: 405-24. PubMed ID: 25741868

Shashi V. et al. 2014. Genetics in Medicine : Official Journal of the American College of Medical Genetics. 16: 176-82. PubMed ID: 23928913

Valencia C.A. et al. 2015. Frontiers in Pediatrics. 3: 67. PubMed ID: 26284228

Yang Y. et al. 2013. The New England Journal of Medicine. 369: 1502-11. PubMed ID: 24088041

Yang Y. et al. 2014. JAMA. 312: 1870-9. PubMed ID: 25326635