Considerations on Implementing the ACMG IF Recommendations: The Laboratory Perspective

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2 Considerations on Implementing the ACMG IF Recommendations: The Laboratory Perspective Elaine Lyon, PhD Medical Director, Molecular Genetics, Associate Professor of Pathology University of Utah School of Medicine President, Association for Molecular Pathology

3 Disclosures The presenter has the following financial relationship(s) with commercial interest(s): Complete Genomics: Advisory board

4 The Clinical Exome/Genome Genes associated with disease (symptom-guided analysis) OMIM, HGMD Incidental Findings Medically significant genes Actionable ACMG recommendations 56 genes associated with increased risk for cancer, cardiovascular problems, genes influencing response to anesthesia Expand the gene list? What is actionable? Include pharmacogenetics? Report pathogenic or expected pathogenic variants Expected vs Likely

5 Technical Limitations Regions of no or low coverage Regulatory, Exon 1 Repeat regions Deletions/duplications Not filled in by Sanger sequencing Pseudogenes PMS2, SDHD, PTENP, PKP2P1

6 Consent Patient autonomy Standard principle in genetic testing Models of consent Opt out: perform unless request not to perform Opt in: perform only when requested ACMG, expanded gene list, PGx Choose specific genes? Elements Indications for testing/ordering considerations Limitations Risks General Case specific Manage consent process and choices

7 Consent Form

8 Important Points in Informed Consent Regarding Incidental Findings Check here if you do NOT want variants analyzed and reported for the ACMG genes listed above. Exome sequencing may fail to detect mutations in some of the ACMG recommended genes, which means a mutation may be present that is not detected. We will only report disease-causing mutations that can be identified using the routine exome analysis. If you have symptoms or a family history of one of the conditions tested in the ACMG list of genes, we recommend you order additional testing specifically for that condition, as coverage of the associated gene(s) may not be complete using exome sequencing. Each family member whose exome is sequenced will receive a separate report, either listing the ACMG mutations detected, or stating no pathogenic mutations were identified.

9 Analysis/Assessment Incidental findings not always incidental Depends of filtering Filter parameters may filter out variants with higher frequency Specific searches Set list of incidental genes Set filter parameters Set list of curated known pathogenic variants? If frequency is higher than filter parameters Variant assessment Similar to symptom guided analysis Differences for pharmacogenomics Confirm findings by Sanger sequencing?

10 Filters Population frequencies Filtering Set MAF exclusion criteria Set exclude criteria for internal data Exclude types of variants? Intergenic, noncoding RNA, UTR, synonymous Set Quality and Depth parameters Genes: Set for specific genes (i.e. incidental genes only)

11 PGx from Exome/Genome Limitations/Challenges Not disease associated Requires exogenous challenge Common variants Can t filter by population frequency Known variants outside of gene or intron/exon boundaries. Example: CYP2D6 involved in metabolism of 25% of drugs pseudogene Copy number variants (large deletions/duplications) Structural variants: translocations, inversions, gene conversion Nomenclature Translate * alleles, haplotypes, rs numbers to standard nomenclature Haplotype Ex: 100C>T = *10 Ex: 100C>T G>A = *4 Interpretation Uncharacterized variants

12 Interrogate Known PGX Genes CYPs: CYP1A1,CYP1A2,CYP2B6,CYP2C19,CYP2C8,CYP2C9,CYP2D6, CYP2E1, CYP3A4, CYP3A5,CYP4F2 SULTs: SULT1A1,SULT4A1 UGTs: UGT1A,GT1A3,UGT1A4,UGT1A6,UGT1A9,UGT2B15,UGT2B7 NATs: NAT1,NAT2 Others: APOE, BCHE, CHRNA4, CHRNB2, COMT, CPS1, CYB5R3, DPYD,DRD2,DRD4, G6PD,,HPRT1,HTR1A,HTR2A,IL28B,ITPA,MC1R,MT-CO1, MT, CO2, OPRM1, OTC, PNPLA3, SLC6A4, SLCO1B1,TPMT, TYMS, VKORC1

13 Reporting How to present complex and detailed explanation of variant/gene in an understandable manner Gene: PKP2: encodes plakophilin-2, which forms part of the link between the cytoplasmic tail of cadherins and the intermediate filament cytoskeleton. Heterozygous PKP2 mutations have been shown to cause arrhythmogenic right ventricular dysplasia /cardiomyopathy. Mutations in PKP2 are inherited in an autosomal dominant manner with reduced penetrance Nucleotide Change: c.336+1g>a not previously reported in the literature, but it is expected to be pathogenic because it disrupts a canonical splice site Recommendation: evaluation by a cardiologist for possible symptoms of this condition. Parental testing for PKP2 c.336+1g>a is recommended as they may also be at risk.

14 Conclusions Considerations for implementing incidental findings: Determine gene list ACMG, Expanded gene list, pharmacogenomics Determine technical ability/limitations Pseudogenes, repeat regions, regulatory etc Decide informed consent model Always performed, opt-out, opt-in Analysis/Assessment Setting filtering parameters Reporting Detailed and complex information

15 Acknowledgement ARUP Laboratories and ARUP Institute for Clinical & Experimental Pathology Rong Mao, MD Pinar Bayrak-Toydemir, MD, PhD Gwen McMillin, PhD Roberta Melis, PhD Genevieve Pont-Kingdon, PhD Shale Dames, MS (PhD candidate) Whitney Donahue, PhD Brendan O Fallon, PhD Jacob Durtschi, MS Marc Singleton (PhD candidate) Karl Voelkerding, MD Rebecca Margraff, PhD Emily Coonrod, PhD Cindy Meadows Tyler Wayman Jennifer Stocks Maria Erali, MS Christina Goleman AMP Clinical Practice Committee chair: Madhuri Hegde, PhD