Improving Reporting of Association Study Results Timothy R. Rebbeck, Ph.D. University of Pennsylvania School of Medicin - PowerPoint PPT Presentation

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Improving Reporting of Association Study Results Timothy R. Rebbeck, Ph.D. University of Pennsylvania School of Medicin

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  1. Improving Reporting of Association Study Results Timothy R. Rebbeck, Ph.D. University of Pennsylvania School of Medicine

  2. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?

  3. Example: CYP3A Locus(Chromosome 7q22) CYP3A5 CYP3A7 CYP3AP1 CYP3A4 CYP3A43 *1B (5’ UTR)

  4. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?

  5. CYP3A4 Substrate Heterogeneity R=Estrogens, Progesterone, Testosterone, Carcinogens, Drugs CYP3A4 R R-OH Phase II X=Sulfate, Glucuronate, Glutathione,Methyl R-OX

  6. G G G Exposure Disease Outcome G G Treatment Chemoprevention Where Are Genes Acting?

  7. Lupron Finasteride Pleiotropic Effects of CYP3A4 and Fatal Prostate Cancer CYP3A4 CYP3A4 Prostate Cancer Androgens PSA Failure CYP3A4 CYP3A4

  8. Linkage Disequilibrium in the CYP3A Gene Cluster CYP3A43 CYP3A4 CYP3AP1 CYP3A7 CYP3A5 3’ 5’ LD LD LD LD Chang 2000, Kuehl 2001, Wojnowski 2002, Zeigler-Johnson 2004

  9. Inference • How Many Tests Were Performed? • Correction for Multiple Comparisons • Bonferroni • Bonferroni-Holm, Westfall-Young • False Discovery Rate (Benjamini-Hochberg) • FPRP/FNRP (Wacholder) • Others?

  10. Novel Analytical Methods • Recursive Partitioning (CART; Breiman 1984, Foulkes 2005) • Random Forests (Pavolov 1997) • Combinatorial Partitioning (Nelson 2001) • Multifactor-Dimensionality Reduction (Ritchie 2001) • Permutation-Based Procedures (Trimming/Weighting; Hoh 2000) • Multivariate Adaptive Regression Spines (Friedman 1991) • Boosting (Schapire 1990) • Support Vector Machines (Vapnik 2000) • Neural Networks (Friedman & Tukey 1974, Friedman & Stuetzle 1981) • Bayesian Pathway Modeling (Conti 2003, Cortessis & Thomas 2004) • Clique-Finding (Mushlin 2006)

  11. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?

  12. Laboratory Error • Non-Differential Misclassification: Bias Toward the Null Hypothesis • Differential Misclassification (e.g., cryptic variants): Unpredictable Bias • Possible Type I or Type II Error and Lack of Replication Among Studies • Appropriate reporting of QC

  13. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?

  14. Racial Differences: CYP3A4*1B Example Zeigler-Johnson 2002, 2004

  15. CYP3A4*1B Frequency and Prostate Cancer Incidence by Race AfricanAmerican European American ChineseAmerican

  16. CYP3A Haplotypes Zeigler-Johnson 2004

  17. (When) Is Population Stratification a Problem? • Can be found if you look hard enough (Freedman 2004) • May be more pronounced in recently admixed populations (Ardlie 2003) • Adjustment or matching can be undertaken when race can be appropriately measured • Other approaches available when race cannot be measured: • Genomic Control (Pritchard 1999; Devlin 1999) • Structured Association (Pritchard 2000; Satten 2001) • Small biases likely in most situations (Wacholder 2000, Millikan 2000, Wang 2004, 2005, 2006) • Other forms of bias exist that may be more important (Palmer and Cardon 2003)

  18. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?

  19. Effect of CYP3A4*1B on “Function” Effect of CYP3A4*1BCompared with CYP3A4*1A 190%↑ in Testosterone oxidation 90%↑ in Luciferase Expression 40%↑ in Luciferase Expression 40%↑ in Nifedipine Oxidase Activity 110%↑in CYP3A4 Protein Expression No Change in Luciferase Expression 20%↑in Luciferase Expression 20%↑in Luciferase Expression 40%↑in Luciferase Expression 20-90%↑ in Luciferase Expression 20-117%↑in transcriptional activation* 75-147%↑ in transcriptional activation* *Upon Xenobiotic Exposure CYP3A4*1B ATG 5’ 3’ Westlind(Hepatocytes) Amirimani (MCF7) Amirimani (HepG2) Ando (Hepatocytes) Ando (Hepatocytes) Spurdle (HepG2) Spurdle (HepG2+E) Amirimani (MCF7) Amirimani (HepG2) Amirimani (Hepatocytes) Hamzeiy (HepG2) Hamzeiy (HuH7)

  20. Sources of “Functional” Information Evidence Example Experimental In vivo, In vitro assays Nucleotide Sequence Mutation Consequences Evolutionary Conservation Sequence Conservation (e.g., SIFT, CODDLE) Population Genetics Hardy-Weinberg, Linkage Disequilibrium Exposures Metabolism of relevant carcinogens Epidemiology Association consistency Structural Protein conformation (e.g., PolyPhen)

  21. Assessing SNP Functional Significance(Rebbeck, Wu, Spitz 2004)

  22. Assessing SNP Functional Significance(Rebbeck, Wu, Spitz 2004)

  23. Assessing SNP Functional Significance(Rebbeck, Wu, Spitz 2004)

  24. Infer SNP to be Functional?

  25. Infer SNP to be Non-Functional?

  26. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?

  27. Informative Irreproducibility:Is Replication Always Necessary or Expected? • When might “Irreproducibility” reflect meaningful biological phenomena? • Differences may reflect • Incorrect model (interaction vs. main effects) • Frequency (power?) differences • Context of association • Distinguish error from true differences

  28. Reporting Genetic Associations • Appropriate • Study Design? • Biological Model? • Statistical Analysis? • Laboratory QC? • Race/Ethnicity? • Biological Plausibility? • Replication?