2015 TriCon - Clinical Grade Annotations - Public Data Resources for Interpreting Genomic Variants
Download as PPTX, PDF3 likes1,554 views
This document discusses the advancements in genomic variant interpretation and the increasing importance of public data resources for high-quality variant calls. It covers various tools and workflows used in clinical genomics, including annotations, data sharing, and variant interpretation, while emphasizing the role of genomic sequencing in diagnostics. Key public initiatives such as ClinVar and databases like HGMD are highlighted for their contributions to understanding genomic variations and their clinical significance.
2015 TriCon - Clinical Grade Annotations - Public Data Resources for Interpreting Genomic Variants
- 1. Clinical Grade Annotations: Public Data Resources for Interpreting Genomic Variants February 19, 2105 Gabe Rudy @gabeinformatics VP Product Management and Engineering Golden Helix
- 2. My Background Golden Helix - Founded in 1998 - Genetic association software - Analytic services - Thousands of users worldwide - Over 800 customer citations in journals Products I Build with My Team - SNP & Variation Suite (SVS) - SNP, CNV, NGS tertiary analysis - Import and deal with all flavors of upstream data - VarSeq - Annotate and filter variants in gene panels, exomes and genomes for clinical labs and researchers. - GenomeBrowse (Free!) - Visualization of everything with genomic coordinates. All standardized file formats.
- 3. Agenda Getting High Quality Variant Calls Data Sharing and the Maturing of Public Resources 2 3 4 Clinical Grade Candidate Variant Identification How I Met My Exomes1 NGS Clinical Utopia: Are We There Yet?5
- 4. Exome Sequencing in Consumer Genomics Exomes done as part of Pilot Program 80x coverage Raw data with no interpretation Erin JIA Gabe (me) Ethan
- 5. Research or clinical grade? Total Reads 140M Unique Align 87% Mean Target 105x % Target at 2x 97% % Target at 10x 94% % Target at 20x 89% % Target at 30x 83%
- 6. Agenda Getting High Quality Variant Calls Data Sharing and the Maturing of Public Resources 2 3 4 Clinical Grade Candidate Variant Identification How I Met My Exomes1 NGS Clinical Utopia: Are We There Yet?5
- 12. GRCh38 – Here Now, but no hurry A better human reference - Revised Cambridge Reference Sequence (rCRS) MT - Has centromere models - ~2000 incorrect alleles fixed - ~100 assembly gaps updated NCBI Annotations 106 on 38 - dbSNP 141, ClinVar, RefSeqGene - Ensembl 76 on both No Poplulation Catalogs - Some being ported (by Ensembl, dbSNP) GRCh37 GRCh38 Ts/Tv 2.06558 2.10171 snps snps mnps mnps indels indels complex complex 270000 280000 290000 300000 310000 320000 330000 340000 GRCh37 GRCh38 My Exome 331,824 319,442
- 13. Blog Post
- 14. Agenda Getting High Quality Variant Calls Data Sharing and the Maturing of Public Resources 2 3 4 Clinical Grade Candidate Variant Identification How I Met My Exomes1 NGS Clinical Utopia: Are We There Yet?5
- 15. Baylor Workflow - Clinical Exomes Paper Disease gene related Medically actionable deleterious variants Deleterious variants in ACMG gene list Deleterious variants VUS in dominant gene or homozygous in recessive gene Deleterious variant in gene with no known disease
- 16. Annotate, Then Filter and Interpret
- 17. Data Sources to Replicate Workflow 1000 Genomes (Phase 1) “ESP” (NHLBI 6500 Exomes v2) HGMD (Public vs Professional) Variant’s Protein Coding Effect RNA Splicing Effect (dbscSNV) - −3 to +8 at the 5’, −12 to +2 at the 3’ Genes Lists: - Single-Gene Disorder (OMIM with Inheritance) - Medically Actionable (114 genes NHLBI study) - Dominant Inheritance (MedGen) - ACMG Carrier Panel (ACMG Incidental Findings guidelines)
- 18. My Exome Analyzed Start: 235,689 847 234,842 224,914 9,928 9,069 807 859 40 242 13 59 565 0 624 624 255 20 20 20 0 0 598 644
- 19. • Pathogenic OTC Variant • What if I got this through BabySeq?
- 20. Agenda Getting High Quality Variant Calls Data Sharing and the Maturing of Public Resources 2 3 4 Clinical Grade Candidate Variant Identification How I Met My Exomes1 NGS Clinical Utopia: Are We There Yet?5
- 21. Annotating against Transcripts RefSeqGenes – Versioned on RNA sequence - Annotated against human reference by “Annotation Releases” - Last on GRCh37 was 105 (2013-08-20) – GRCh38 release 106 (2014-01-17) - 84,950 transcripts, most are “predicted” (XM_” and non-coding) - Standard in US for reporting variation (NM_016335.4:c.123C>T etc) - UCSC grabs RNA from RefSeq directly and maps to their genome references “continuously” Ensembl – Versioned on Alignment - GENCODE: Well curated subset of high-quality, validated transcripts - V75 last version of GRCh37, 2014-06-27 - Many specific bio-types, but protein_coding usued for annotation - Has mappings to RefSeq IDs, but
- 22. Reference Sequence Versus Gene Sequence EMG1 on GRCh37 “Gap” of the mRNA coding sequence versus reference seq: Handled differently by 3 different “gene alignments”
- 23. Reference Sequence Versus Gene Sequence EMG1 on GRCh38 Reference sequence patched, no gap Alignments agree
- 24. RefSeq Accession Not Sufficient for Var-Tx Interaction RefSeq defines transcripts as mRNA sequence NCBI “Annotation Releases” (like v105) provides alignments using “Splign” UCSC pulls RefSeq mRNA and aligns themselves using “BLAT” They can choose equally valid but different alignments for the same assession This alignment of NM_052814.3 places the exon at dramatically different loci. Will result in different annotations of any variant overlapping these exons
- 25. COSMIC Does not provide data in easy to use form for NGS Just announced change in licensing affective in March - Access to the COSMIC website will stay free for all users. - The new licensing strategy will charge for-profit organisations to download COSMIC datasets. - Download by academic and non- profit organisations will remain free 2015 Roadmap: - GRCh38 - More curation - Visualization improvements
- 26. ClinVar Submitters: - OMIM: Johns Hopkins - Samuels - Lab for Molecular Medicine - Invitae - Emory Genetics Lab Star rating system - 0-4 stars – level of review ClinVar is designed to provide a freely accessible, public archive of reports of the relationships among human variations and phenotypes, with supporting evidence.
- 27. ClinVitae: ClinVar and Friends by Invitae Sources: - ClinVar (62,913) - Emory (13,365) - ARUP (2,850) - Carver Mut (199) - K Cunningham (581) 79,907 V, 9,189 G - 32,523 Pathogenic - 38,796 Likely Pathogenic Provided in HGVS - 59,878 after mapping to genomic space
- 28. BRCA: The back door to Myriad’s database 1995 – Patent issued to Myriad Genetics June 2013 – Patents invalidated by ruling Lab setting up Dx has a lot of catch up “Free the Data” and other ways in which Mryiad’s data is in ClinVar, etc. Sharing Clinical Reports Project
- 29. BRCA: In my wife
- 30. HGMD Data mines academic papers for reported functional variants Also takes submissions, corrections reviewed by team First available in 1996 - Originally 10k variants - 105k in Public (2014) - 148k in “Pro” (2014)
- 31. Left-Align Delta F508 to Make it Match
- 32. Left-Align Annotations Using a Smith- Waterman algorithm to left- align variants from public databases show non-obvious differences NGS alignment and variant calling always left-aligned Left-align your database so they can be annotated
- 33. Changes in Monthly Updates • 36 variants went missing from December to Jan release • Some where Pathogenic
- 34. ClinVar’s VCF File • ClinVar current relies on their dbSNP identifier mappings to “build” VCF files • There are ~14,000 small variants in their database without dbSNP identifiers, and thus missing from the VCF • ~5K Pathogenic • Often these variants are in newer dbSNP builds, and the ClinVar mappings are just not updated. • This variant was in ClinVar, with genomic coordinates, but no RSID: - HGVS(c.): NM_002894.2:c.298C>T - Chromosome:Start:Stop: 18:20548818:20548818 - (Recently RSID was added)
- 35. dbSNP 141 Had Allele Errors I reported the issue 7/22/2014 Confirmed, 8/12 generated better VCF and placed in “test” folder Found more issues Replaced official VCF in 02/09/2014 We waited until fixed to publish official support
- 36. Agenda Getting High Quality Variant Calls Data Sharing and the Maturing of Public Resources 2 3 4 Clinical Grade Candidate Variant Identification How I Met My Exomes1 NGS Clinical Utopia: Are We There Yet?5
- 37. asdf
- 39. NM_002626.4:c.1877G>C in PFKL NP_002617.3:p.Arg626Pro missense mutation Predicted damaging by 4/5 functional predictions VEST3: 0.948, GERP++: 4.59 ExAC and 1kG have a G>A, but G>C is novel Variants in region are extremely rare (G>C ExAC 4 of 122,364 alleles) – 0.003% No ClinVar variants for gene OMIM entry has no known disease association PubMed search shows few recent articles: Most recent 1998 paper showed - phosphofructokinase (PFKL) overexpressed in Down syndrome (DS) - Transgenic PFKL mice had an abnormal glucose metabolism with reduced clearance rate from blood and enhanced metabolic rate in brain.
- 40. d
- 41. d 35 LoF Variants, None Homozygous
- 42. Training Most variants are rare or novel - Training to interpret these is extensive MD/Pathology background is insufficient Need a PhD in molecular genetics There’s only 500 board certified Clinical Molecular Geneticists since started Let’s share in the learning process Baylor Exome Sign-Out
- 43. Phenotypeing and Matchmaking Portals Diagnosis often requires finding another family to confirm a novel gene to phenotype association Finding a second family: - Social media - PhenoDB - PhenomeCentral.org - Orphanet – Resources on over 6000 rare diseases and orphan drugs. - European centric: GEN2PHEN (G2P) Matt Might found a second family with NGLY1 deficiency through a blog post that went viral.
- 44. N-Glycanase Deficiency http://www.ngly1.org/ Matthew Might and Matt Wilsey. The shifting model in clinical diagnostics: how next-generation sequencing and families are altering the way rare diseases are discovered, studied, and treated. Genetics in Medicine. March 2014.
- 45. Thank you Heidi Rehm – Chief Laboratory Director at Laboratory for Molecular Medicine, PCPGM Joel Parker – Cancer Genetics, UNC Chapel Hill Gerry Higgins – VP, Pharmacogenomic Science, Assure Rx Health Frank Schacherer – Chief Technical Officer, BIOBASE Reece Hart – Computational Biologist, Invitae (now 23andMe) Greta Linse Peterson – Director of Product Management and Quality, Golden Helix
- 46. Questions?