1. A coalescent computational platform to predict strength of association for clinical samples Gabor T. Marth Department of Biology, Boston College marth@bc.edu MIT Bioinformatics Seminar April 25, 2005 Cambridge, Massachusetts, USA

2. Sequence variations cause inherited diseases allow tracking ancestral human history

3. Automated polymorphism discovery tools Marth et al. Nature Genetics 1999

4. Genome scale sequence variation resources Sachidanandam et al. Nature 2001 ~ 10 million EST WGS BAC genome reference

5. How to use markers to find disease? problem: genotyping cost precludes using millions of markers simultaneously for an association study genome-wide, dense SNP marker map depends on the patterns of allelic association in the human genome question: how to select from all available markers a subset that captures most mapping information (marker selection, marker prioritization)

6. Allelic association allelic association is the non- random assortment between alleles i.e. it measures how well knowledge of the allele state at one site permits prediction at another site marker site functional site by necessity, the strength of allelic association is measured between markers significant allelic association between a marker and a functional site permits localization (mapping) even without having the functional site in our collection there are pair-wise and multi-locus measures of association

7. Pair-wise: linkage disequilibrium (LD) LD measures the deviation from random assortment of the alleles at a pair of polymorphic sites D=f( ) – f( ) x f( ) other measures of LD are derived from D, by e.g. normalizing according to allele frequencies (r 2 )

8. strong association: most chromosomes carry one of a few common haplotypes – reduced haplotype diversity Multi-marker: haplotype diversity the most useful multi-marker measures of associations are related to haplotype diversity 2 n possible haplotypesn markers random assortment of alleles at different sites

9. The main determinants of allelic association recombination: breaks down allelic association by “randomizing” allele combinations demographic history of effective population size: bottlenecks increase allelic association by non-uniform re- sampling of allele combinations (haplotypes) bottleneck

10. Block-like patterns in the human genome Wall & Pritchard Nature Rev Gen 2003 Daly et al. Nature Genetics 2001

11. The promise for medical genetics CACTACCGA CACGACTAT TTGGCGTAT within blocks a small number of SNPs are sufficient to distinguish the few common haplotypes  significant marker reduction is possible if the block structure is a general feature of human variation structure, whole-genome association studies will be possible at a reduced genotyping cost this motivated the HapMap project Gibbs et al. Nature 2003

12. The HapMap initiative goal: to map out human allele and association structure of at the kilobase scale deliverables: a set of physical and informational reagents

13. HapMap physical reagents reference samples: 4 world populations, ~100 independent chromosomes from each markers: millions of SNPs from the US public variation database (dbSNP) genotypes: high-accuracy assays using various platforms; fast public data release

14. Informational: HapMap annotations (“HaploView”, Daly lab)

15. Focal questions about the HapMap CEPH European samples 1. Required marker densityYoruban samples 4. How general the answers are to these questions among different human populations 2. How to quantify the strength of allelic association in genome region 3. How to choose tagging SNPs

16. Across samples from a single population? (random 60-chromosome subsets of 120 CEPH chromosomes from 60 independent individuals)

17. Consequence for marker performance Markers selected based on the allele structure of the HapMap reference samples… … may not work well in another set of samples such as those used for a clinical study.

18. How to assess sample-to-sample variability? 1. Understanding intrinsic properties of a given genome region, e.g. estimating local recombination rate from the HapMap data 3. It would be a desirable alternative to generate such additional sets with computational means McVean et al. Science 2004 2. Experimentally genotype additional sets of samples, and compare association structure across consecutive sets directly

19. Towards a marker selection tool 2. generate computational samples for this genome region 3. test the performance of markers across consecutive sets of computational samples 1. select markers (tag SNPs) with standard methods

20. Modeling variations: the Coalescent past the Coalescent is a simulation technique produces possible genealogies backwards, towards MRCA generates mutations (neutral, non-recurrent) used to describe the statistical properties of DNA samples these statistical properties depend on model structure and model parameters

21. 1. marker density (MD): distribution of number of SNPs in pairs of sequences Two simple statistics “rare” “common” 2. allele frequency spectrum (AFS): distribution of SNPs according to allele frequency in a set of samples

22. The effects of demographic history past present stationaryexpansioncollapse MD (simulation) AFS (direct form) history bottleneck

23. Generating haplotypes with the Coalescent

24. Global haplotypes vs. local data relevance

25. Generating data-relevant haplotypes 1. Generate a pair of haplotype sets with Coalescent genealogies. This “models” that the two sets are “related” to each other by being drawn from a single population. 3. Use the second haplotype set induced by the same mutations as our computational samples. 2. Only accept the pair if the first set reproduces the observed haplotype structure of the HapMap reference samples. This enforces relevance to the observed genotype data in the specific region.

26. Generating computational samples Problem: The efficiency of generating data- relevant genealogies (and therefore additional sample sets) with standard Coalescent tools is very low even for modest sample size (N) and number of markers (M). Despite serious efforts with various approaches (e.g. importance sampling) efficient generation of such genealogies is an unsolved problem. N M We are developing a method to generate “approximative” M-marker haplotypes by composing consecutive, overlapping sets of data-relevant K-site haplotypes (for small K) Motivation from composite likelihood approaches to recombination rate estimation by Hudson, Clark, Wall, and others.

27. Approximating M-site haplotypes as composites of overlapping K-site haplotypes 1. generate K-site sets 2. build M-site composites M

28. Piecing together neighboring K-site sets 000 100 001 101 010 110 011 111 000 001 010 011 100 101 110 111 this should work to the degree to which the constraint at overlapping markers preserves long-range marker association

29. Building composite haplotypes A composite haplotype is built from a complete path through the (M-K+1) K-sites.

30. Initial results: 3-site composite haplotypes a typical 3-site composite 30 CEPH HapMap reference individuals (60 chr) Hinds et al. Science, 2005

31. 3-site composite vs. data

32. 3-site composites: the “best case” “short-range” “long-range” 1. generate K-site sets

33. Variability across sets The purpose of the composite haplotypes sets … … is to model sample variance across consecutive data sets. But the variability across the composite haplotype sets is compounded by the inherent loss of long-range association when 3-sites are used.

34. 4-site composite haplotypes 4-site composite

35. “Best-case” 4 site composites Composite of exact 4-site sub-haplotypes

36. Variability across 4-site composites

37. … is comparable to the variability across data sets.

38. Software engineering aspects: efficiency To do larger-scale testing we must first improve the efficiency of generating composite sets. Currently, we run fresh Coalescent runs at each K-site (several hours per region). Total # genotyped SNPs is ~ 1 million -> 1 million different K-sites to match. Any given Coalescent genealogy is likely to match one or more of these. Computational hap sets can be databased efficiently. 4 HapMap populations x 1 million K-sites x 1,000 comp sets x 50 bytes < 200 Gigabytes

39. Un-phased genotypes (AC)(CG)(AT)(CT) A G A C C C T T http://pga.gs.washington.edu/ the primary data represent diploid genotypes one has the choice to “reconstruct” the haplotypes with statistical methods as shown (e.g. the PHASE program); this may be inaccurate or one may account for all possible reconstructions when evaluating data-relevance; this is computationally very expensive

40. Conclusions 3-site composites are unlikely to work 4-site composites are very promising both the initial results and the expected payoff justify going ahead more thorough statistical analyses, performance evaluations, and algorithmic development work ahead

41. Acknowledgements Eric Tsung Aaron Quinlan Ike Unsal Eva Czabarka (Dept. Mathematics, William & Mary)