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MESA Family

MESA Family. Publications & Presentations Kent Taylor Monday, 18 September 2006. P&P History. Candidate Genes selected Phenotypes identified Concerns: normal MESA P&P process would not be efficient for Candidate Gene proposals

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MESA Family

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  1. MESA Family Publications & Presentations Kent Taylor Monday, 18 September 2006

  2. P&P History • Candidate Genes selected • Phenotypes identified • Concerns: • normal MESA P&P process would not be efficient for Candidate Gene proposals • Possible for early investigators to scoop important genetic papers without appropriate oversight of the process • Formal process needed • Two stage process developed: • Initial basic proposal (list genes and phenotypes only) • Online Investigator survey • Follow-up e-mail with additional questions • Formal MESA P&P proposal • Following MESA Policies and Procedures • Using MESA Internal Website • Potential to overwhelm P&P Committee prompted proposal to create a second P&P Genetics Review Committee (to be discussed by MESA SC)

  3. “Prior” Gene Studies: MESA

  4. “Prior” Gene Studies: MESA

  5. Gene Studies: MESA Family • MESA Family grant originally proposed to genotype 6-8 candidate genes with ~80 SNPs in MESA, AND • Genome scan in MESA Family subjects • Rapid technological developments allow a project of greater scope for the same budget

  6. Expanded Candidate Gene Study • Conduct an association study of 2,880 MESA participants (parent study) • 720 randomly selected from each ethnic group of Caucasian-, African-, Mexican-, and Chinese-Americans • Include the well-phenotyped “MESA 1000” • Genotype 1536 SNPs in candidate genes proposed by MESA Investigators, TWICE

  7. Candidate Gene Selection • 267 candidate genes were initially proposed by MESA investigators, collated by DCC, and then ranked by MESA investigators • Further input by MESA laboratory and MESA Family Study Genetics Committee (294 genes) • Genes have continued to be added to list for the second 1536 genotyping run

  8. Selection of SNPs • SNPs proposed by MESA investigators • SNPs with feasibility of 80% or greater on Illumina platform • SNPs with minor allele frequency >0.05 • tagSNPs for Caucasian-American and African-American haplotype blocks as defined by Tagger or LDSelect for each candidate gene • List of SNPs sent to Illumina for evaluation • Candidate gene was re-analyzed for tagSNPs if necessary after feasibility scores were returned from Illumina • Ancestry informative SNPs added to panel (96)

  9. Data Overview • 1440/1536 SNPs were successfully genotyped in 3036 samples • 4 ethnic groups • MESA “1000” • 119 genes represented • 96 ethnic-specific markers • Only 6 DNA samples did not genotype • These same 3036 samples are available for Phase 2 genotyping

  10. Phase 1 Genotyping QC (1)Laboratory • Illumina report: • Data quality was very high • DNA success rate unprecedented (for such a large project) • Being pleased with the locus conversion rate (higher than predicted) • Excellent DNA quality (aided in achieving the high locus success rate) • Non-blinded & blinded sample pair genotyping concordance rates: >99.99%

  11. Phase 1 Genotyping QC (2)Analysis at Wake Forest Total Samples 3036 Sample Failures 6 Expected Samples to Delete (QC duplicate) 156 Actual Samples Deleted 183 Duplicate 168 Duplicate+Unresolved Gender Err 4 Triplicate 3 Triplicate+Unresolved Gender Err 1 Identical Twins 1 Unresolved Gender Error 6 Remaining Samples for Analysis: 2847

  12.  2847 Samples, 4 ethnic groups Total Number of Genotypes Expected: 2847 x 1440 4,099,680 Total Sporadic Missing Genotypes: 2,944 Total Genotypes: 4,096,736 And all MESA phenotypes

  13. Phase 1 Data Analyses • Identify key phenotypes for analysis • Transformations provided by Coordinating Center • Transfer of data from Coordinating Center to WFU • Initial data analyses using pre-specified models • Ethnic-specific {4 ethnic groups} phenotype = SNP + age + sex + center • Total MESA phenotype = SNP + age + sex + center/ethnic • Within each of the 5 sets of analyses for each SNP (and each phenotype), perform generalized test of association (2 df) and model SNP as dominant, additive and recessive

  14. Publication Process … which leads to the obvious problem of writing it all up: Status Report

  15. BACKGROUND • The MESA P&P committee had too much to do prior to these developments • The possible number of paper proposals is daunting • MESA P&P wanted additional genetic analytical expertise to augment effort

  16. Call for Proposals • Call was put out for MESA investigators to propose lead authors, senior authors, candidate genes, & phenotypes for projects of interest • These were collected by Coordinating Center • Committee met by phone in August to: • put groups interested in the same gene/phenotype combinations together • spread the proposals throughout MESA and over many investigators so that all groups had good proposals • narrow some proposals if too many phenotypes were requested or phenotypes too vague

  17. Result • Lead & senior authors for 41 proposals have been identified • More to come: • Phenotypes narrowed and made specific • Additional authors are identified in MESA groups

  18. Publications Process In order not to burden the existing MESA P&P: • Formation of separate Genetics P&P Committee to be proposed to the Steering Committee at this meeting • Collate authors, genes, & phenotypes • Help lead author to find other investigators interested in a particular gene/phenotype combination • Make gene/phenotype data available

  19. Lead Authors • Once approved, lead author will be asked to confirm acceptance of gene/phenotype assignment • Lead author will be given access to the genetic & phenotype data by the Coordinating Center upon acceptance • Lead develops paper proposal for MESA P&P Committee

  20. Phase 2 Genotyping • Additional SNPs genotyped using older technologies (being considered in the future) • ACE insertion/deletion, APOE • 1536 SNPs using Illumina run • DNA already at Illumina from Phase 1 • Some of same genes with additional SNPs • Additional genes proposed since Phase 1 was completed • Next 50 genes on the Phase 1 list • Additional AIMs

  21. Phase 2 Gene List

  22. Phase 1 Genotyping • Candidate genes and analyses • Focus of Mychaleckyj presentation (MESA Family Steering Committee meeting and MESA SC mtg) • High degree of performance • Few SNPs eliminated • High accuracy • Specific genes: focus of Bowden & Tsai presentations (MESA Family SC mtg) • Ancestry Informative Markers • 97 picked, 96 genotyped • Focus of Arnett presentation (MESA Family SC mtg)

  23. What to do about APOE ? Illumina Marker Panel #2: Results of Genotyping #SNPs Gene/Marker Set Picked Succeed Fail APOE 7 2 5 ADORA2A 14 10 4 3 SNP FAIL (6 Genes) 3 2 SNP FAIL (17 Genes) 2 1 SNP FAIL (35 Genes) 1 TOTAL FAIL 96 # SUCCESSFUL SNPS 1440 (93.75%)

  24. Addn Candidate Gene Slides

  25. Characteristics of Variation • Tests for Hardy-Weinberg Equilibrium • For each polymorphism (SNP) • Deviations from HWE • Genotyping errors • Deviations from underlying assumptions • No mutation, migration, selection • Small sample size (genetic drift) • True association with phenotype • Estimates of Linkage Disequilibrium • D’ and r2 • Examination of LD block structure

  26. Implications • Consideration of ethnic differences in disease risk • Exposures • Behaviors • Genetics • How to incorporate complexity and reduce heterogeneity • Translation of molecular features into clinical applications to benefit everyone

  27. MESA Family Study

  28. Genetic Plan • Recruitment of families • Wonderfully well .. Congrats to all! • Permits two general types of analyses • Phenotypic (including environmental covariates) • Familial Aggregation (heritability) • Bivariate (genetic/environmental correlations) • Genotypic • Candidate Gene (family-based association) • Genome-wide Linkage Scan (6K SNP panel) (Optional)

  29. Familial Aggregation • Partition variation into variance components based upon familial resemblance • Additive effects of genes (heritability, h2) • Proportion of variation due to additive effects • h2 = s2A/s2P, where s2A estimated from • Parent-offspring ~ 50% s2A • Siblings ~ 50% s2A and s2D and common environment • Higher h2, more of phenotypic variation due to ‘familial effects’; however, may not be due to major gene effects • Phenotypic data available on ~50% of MESA Family participants (primary analyses at CSMC/Minnesota)

  30. Familial Aggregation Results • Focus of Raffel presentation • MESA Family Steering Committee meeting and MESA Steering Committee meeting • Focus of Pankow presentation • MESA Family Steering Committee meeting

  31. Genetic Analyses • Uses families in the analysis of • Family-based association • Candidate genes from MESA • Testing in MESA Family • Linkage • Candidate genes from MESA • Genome-wide Linkage Scan • No longer MGS, scan to be performed within MESA • Other analytic methods • Genetic and epidemiologic data • Gene-environment interaction • Longitudinal genetic data analyses

  32. Operational Issues • Genome wide scan • Handling small genotyping projects (Tsai, Taylor, Bowden subcommittee proposal) • Handling collaborations when genes/ markers are unknown at onset

  33. Genome Wide Scan • Originally planned to utilize Mammalian Genotyping Service • Issue: no more Mammalian Genotyping Service • Plan: Supplemental funding request

  34. Supplemental Funding Request • Bids for genome screen solicited from deCode, Prevention Genetics, Illumina • Recommended 6000 SNP Illumina panel to NHLBI: • 47 parent visits/565 families • Increased SNP density for information content when parents are not available* *Hum Mol Genet 2004 13:1943-9

  35. Comparison of 400 Microsatellites with 4763 SNPs Hum Mol Genet 2004 13:1943-9

  36. Proposal on Handling Small Genotyping Projects

  37. Proposed Small-Scale (1 to 100 SNPs) Genotyping Policy • Sending DNA of MESA out to investigators is an inefficient use of the available resource for genotyping of only a few (1 to 100) SNPs: • Amount of DNA is finite and large amounts will be wasted • Too labor-intensive for a large number of requests • Therefore genotyping will be performed at one of the three MESA genotyping laboratories by Bowden, Tsai, or Taylor • Investigator(s) will be responsible for supply costs

  38. Proposed Small-Scale (1 to 100 SNPs) Genotyping Policy • Preferred method for conservation of DNA • Illumina technology in batch: • 6 Illumina runs use the same amount of DNA as one TaqMan MGB run (for 1 SNP) • Genotyping 1536 SNPs uses the same amount of DNA as 96 SNPs (but costs a lot more)

  39. Proposed Small-Scale (1 to 100 SNPs) Genotyping Policy • Investigator(s) will make request to MESA Genetics Committee • Genotyping Sub-Committee is chaired by Mike Tsai, with Don Bowden and Kent Taylor as members • Genotyping Sub-Committee will: • Decide best way to perform the genotyping (where & how) • Combine requests into fewer Illumina runs in order to save DNA resource • Work with MESA Family genotyping committee • If Illumina design not possible, then TaqMan MGB is available. Check feasibility at dbSNP • If these designs are not possible, then MESA family genetics committee will determine the best way to genotype the particular SNP. Emphasis will be placed on conservation of DNA

  40. Proposed Small-Scale (1 to 100 SNPs) Genotyping Policy • MESA Genotyping Sub-Committee will consider work load/ scheduling and decide where genotyping will be done • Supply costs to be born by the investigator (costs change constantly): • 96 snp run using Illumina: ~12 cents/genotype • TaqMan MGB: ~50-80 cents/genotype • Other snps depend on technology required (eg restriction enzyme, dye-primers, gel or polymer, etc.) • There will likely be DCC costs and there may be overhead and technician costs, depending on the project

  41. Handling Collaborations When Genes are Unknown

  42. Overview of CARE Study • To test a large number of candidate genes in NHLBI cohort studies • Goal: 1700 candidate genes (8-10 SNPs per gene); 15,000 markers on ~50,000 participants from eight NHLBI funded-studies • Comparatively smaller sample for genome-wide association

  43. Eight NHLBI CARE Cohorts • ARIC - Atherosclerosis Risk In Communities • CARDIA - Coronary Artery Risk Development in Young Adults • CHS - Cardiovascular Health Study • CSSCD - The Cooperative Study of Sickle Cell Disease • FHS - Framingham Heart Study • JHS - Jackson Heart Study • MESA - Multi-Ethnic Study of Atherosclerosis • SHHS - Sleep Heart Health Study

  44. Organization of CARE Study • RFP to establish a Center • coordination and utilization of genetic and phenotypic data • from well-characterized NHLBI cohorts • Center funded at the Broad Institute Center for Genotyping and Analysis

  45. CARE Center Goals 1. Receipt and management of DNA samples and phenotypic information to facilitate cross study analysis 2. Design genotyping experiments with software tools integrating sample management, SNP selection, and SNP genotyping platform 3. Support genotyping experiment execution in candidate genes and a set of genome-wide SNPs 4. Manage data and develop and apply statistical methods required to identify associations between genotypes and HLB phenotypes

  46. Larry Atwood, Boston, MAFHS Eric Boerwinkle, Houston, TXARIC Richard Fabsitz, Bethesda, MDNHLBI Myriam Fornage, Houston, TXCARDIA Stacey Gabriel, Cambridge, MABroad Joel Hirschhorn, Cambridge, MABroad Ronald Krauss, Oakland, CAHeart Abdullah Kutlar, August, GABlood Deborah Meyers, Winston-Salem, NCLung Emanual Mignot, Palo Alto, CASleep Dina Paltoo, Bethesda, MDNHLBI Susan Redline, Cleveland, OHSHHS Jerome Rotter, Los Angeles, CAMESA Jeanne Smith, Englewood, NJCSSCD Russell Tracy, Colchester, VTCHS James Wilson, Jackson, MSJHS CARE Steering Committee

  47. Mediawiki-based web pagehttp://www.broad.mit.edu/gen_analysis/care/index.php/Main_Page • Numerous conference calls • Steering Committee Meeting, July 25,2006 • Discussion Items • Define pilot project (phenotypes & SNPs) • Establish principles of data release • Discuss genotyping study design • Select phenotypes to be analyzed

  48. Susan Heckbert Craig Johnson Richard Kronmal Kiang Liu Joe Mychaleckyj James Pankow Wendy Post Bruce Psaty Stephen Rich Jerome Rotter Kent Taylor Russell Tracy Michael Tsai MESA CARE Working Group

  49. Data Release/IRB - James Wilson DNA Transfer/Genotyping - Larry Atwood Analysis/Study Design - Stephen Rich Candidate Gene/SNP Selection - Myriam Fornage Phenotypes - Bruce Psaty- Susan Heckbert Informatics - Joe Mychaleckyj Publications- to be determined CARE Subcommittees & Chair

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