Genetic Epidemiology

Developments in genome‐wide association studies and the increasing availability of summary genetic association data have made application of Mendelian randomization relatively straightforward. However, obtaining reliable results from a Mendelian randomization investigation remains problematic, as the conventional inverse‐variance weighted method only gives consistent estimates if all of the genetic variants in the analysis are valid instrumental variables. We present a novel weighted median estimator for combining data on multiple genetic variants into a single causal estimate. This estimator is consistent even when up to 50% of the information comes from invalid instrumental variables. In a simulation analysis, it is shown to have better finite‐sample Type 1 error rates than the inverse‐variance weighted method, and is complementary to the recently proposed MR‐Egger (Mendelian randomization‐Egger) regression method. In analyses of the causal effects of low‐density lipoprotein cholesterol and high‐density lipoprotein cholesterol on coronary artery disease risk, the inverse‐variance weighted method suggests a causal effect of both lipid fractions, whereas the weighted median and MR‐Egger regression methods suggest a null effect of high‐density lipoprotein cholesterol that corresponds with the experimental evidence. Both median‐based and MR‐Egger regression methods should be considered as sensitivity analyses for Mendelian randomization investigations with multiple genetic variants.

Các nghiên cứu liên kết toàn bộ hệ gen (GWAS) có thể xác định các alen phổ biến có đóng góp vào sự nhạy cảm với các bệnh phức tạp. Mặc dù số lượng lớn SNPs được đánh giá trong mỗi nghiên cứu, tác động của phần lớn các SNP phổ biến phải được đánh giá gián tiếp bằng cách sử dụng các dấu hiệu đã được genotyped hoặc các haplotype của chúng làm đại diện. Chúng tôi đã triển khai một khung Markov Chain hiệu quả về mặt tính toán cho việc ước tính kiểu gen và haplotyping trong gói phần mềm MaCH miễn phí có sẵn. Phương pháp tiếp cận này mô tả các nhiễm sắc thể mẫu như những hình khảm của nhau và sử dụng dữ liệu kiểu gen hiện có và chuỗi shotgun để ước tính các kiểu gen và haplotype chưa quan sát, cùng với các thước đo hữu ích về chất lượng của những ước tính này. Phương pháp của chúng tôi đã được sử dụng rộng rãi để tạo điều kiện so sánh kết quả giữa các nghiên cứu cũng như phân tích tổng hợp GWAS. Tại đây, chúng tôi sử dụng các mô phỏng và kiểu gen thực nghiệm để đánh giá độ chính xác và tính hữu ích của nó, xem xét các lựa chọn bảng genotyping, cấu hình bảng tham chiếu và các thiết kế genotyping được thay bằng chuỗi shotgun. Điều quan trọng, chúng tôi cho thấy ước tính kiểu gen không chỉ tạo điều kiện cho phân tích giữa các nghiên cứu mà còn tăng công suất của các nghiên cứu liên kết di truyền. Chúng tôi cho rằng việc ước tính kiểu gen các biến thể phổ biến bằng cách sử dụng haplotypes HapMap làm tham chiếu là rất chính xác khi sử dụng dữ liệu SNP toàn bộ hệ gen hoặc số lượng nhỏ dữ liệu điển hình trong các nghiên cứu phác thảo chi tiết hơn. Hơn nữa, chúng tôi cho thấy phương pháp này có thể áp dụng trong nhiều quần thể khác nhau. Cuối cùng, chúng tôi minh họa làm thế nào phân tích liên kết các biến thể chưa quan sát sẽ được hưởng lợi từ những tiến bộ hiện tại như các bảng tham chiếu HapMap lớn hơn và công nghệ chuỗi shotgun toàn bộ hệ gen. Genet. Epidemiol. 34: 816-834, 2010. © 2010 Wiley‐Liss, Inc.

Association tests of multilocus haplotypes are of interest both in linkage disequilibrium mapping and in candidate gene studies. For case‐parent trios, I discuss the extension of existing multilocus methods to include ambiguous haplotypes in tests of models which distinguish between the cis and trans phase. A likelihood‐ratio test is proposed, using the expectation‐maximization (E‐M) algorithm to account for haplotype ambiguities. Assumptions about the population structure are required, but realistic situations, including population stratification, which violate the assumptions lead to conservative tests. I describe a permutation procedure for the null hypothesis of interest, which controls for violation of the assumptions. For general pedigrees, I describe extensions of the pedigree disequilibrium test to include uncertain haplotypes. The summary statistics are replaced by their expected values over prior distributions of haplotype frequencies. If prior distributions are not available, a valid test is possible by using the E‐M algorithm to estimate the null distribution of haplotype frequencies. Similar methods are available for quantitative traits. Exact permutation tests are difficult to construct in small samples, but an approximate procedure is appropriate in large samples, and can be used to account for dependencies between tests of multiple haplotypes and loci. Genet Epidemiol 25:115–121, 2003. © 2003 Wiley‐Liss, Inc.

Genomewide association studies are an exciting strategy in genetics, recently becoming feasible and harvesting many novel genes linked to multiple phenotypes. Determining the significance of results in the face of testing a genomewide set of multiple hypotheses, most of which are producing noisy, null‐distributed association signals, presents a challenge to the wide community of association researchers. Rather than each study engaging in independent evaluation of significance standards, we have undertaken the task of developing such standards for genomewide significance, based on data collected by the International Haplotype Map Consortium. We report an estimated testing burden of a million independent tests genomewide in Europeans, and twice that number in Africans. We further identify the sensitivity of the testing burden to the required significance level, with implications to staged design of association studies. Genet. Epidemiol. 2008. © 2008 Wiley‐Liss, Inc.

The genome of an admixed individual represents a mixture of alleles from different ancestries. In the United States, the two largest minority groups, African‐Americans and Hispanics, are both admixed. An understanding of the admixture proportion at an individual level (individual admixture, or IA) is valuable for both population geneticists and epidemiologists who conduct case‐control association studies in these groups. Here we present an extension of a previously described frequentist (maximum likelihood or ML) approach to estimate individual admixture that allows for uncertainty in ancestral allele frequencies. We compare this approach both to prior partial likelihood based methods as well as more recently described Bayesian MCMC methods. Our full ML method demonstrates increased robustness when compared to an existing partial ML approach. Simulations also suggest that this frequentist estimator achieves similar efficiency, measured by the mean squared error criterion, as Bayesian methods but requires just a fraction of the computational time to produce point estimates, allowing for extensive analysis (e.g., simulations) not possible by Bayesian methods. Our simulation results demonstrate that inclusion of ancestral populations or their surrogates in the analysis is required by any method of IA estimation to obtain reasonable results. Genet. Epidemiol. © 2005 Wiley‐Liss, Inc.

The number of Mendelian randomization (MR) analyses including large numbers of genetic variants is rapidly increasing. This is due to the proliferation of genome‐wide association studies, and the desire to obtain more precise estimates of causal effects. Since it is unlikely that all genetic variants will be valid instrumental variables, several robust methods have been proposed. We compare nine robust methods for MR based on summary data that can be implemented using standard statistical software. Methods were compared in three ways: by reviewing their theoretical properties, in an extensive simulation study, and in an empirical example. In the simulation study, the best method, judged by mean squared error was the contamination mixture method. This method had well‐controlled Type 1 error rates with up to 50% invalid instruments across a range of scenarios. Other methods performed well according to different metrics. Outlier‐robust methods had the narrowest confidence intervals in the empirical example. With isolated exceptions, all methods performed badly when over 50% of the variants were invalid instruments. Our recommendation for investigators is to perform a variety of robust methods that operate in different ways and rely on different assumptions for valid inferences to assess the reliability of MR analyses.

In the haplotype relative risk (HRR) statistic (Rubinstein et al.: Human Immunol 3:384 [abstract], 1981), a disease sample is constructed along with its own internal control by comparing those marker alleles passed from the parents to an affected child with the other parental marker alleles not transmitted. Based on the conditional parental genotype distribution given that they have an affected child, statistical properties of the HRR statistic are derived. It is shown that the HRR is different from 1 only when allelic association is present and the recombination fraction is different from 1/2. Transmitted and nontransmitted marker alleles are shown to be statistically independent only in the absence of either allelic association or recombination.

Mutations in the gene encoding interferon regulatory factor 6 (IRF6) underlie a common form of syndromic clefting known as Van der Woude syndrome. Lip pits and missing teeth are the only additional features distinguishing the syndrome from isolated clefts. Van der Woude syndrome, therefore, provides an excellent model for studying the isolated forms of clefting. From a population‐based case‐control study of facial clefts in Norway (1996–2001), we selected 377 cleft lip with or without cleft palate (CL/P), 196 cleft palate only (CPO), and 763 control infant‐parent triads for analysis. We genotyped six single nucleotide polymorphisms within the IRF6 locus and estimated the relative risks (RR) conferred on the child by alleles and haplotypes of the child and of the mother. On the whole, there were strong statistical associations with CL/P but not CPO in our data. In single‐marker analyses, mothers with a double‐dose of the ‘a’‐allele at rs4844880 had an increased risk of having a child with CL/P (RR=1.85, 95% confidence interval: 1.04–3.25; P=0.036). An RR of 0.38 (95% confidence interval: 0.16–0.92; P=0.031) was obtained when the child carried a single‐dose of the ‘a’‐allele at rs2235371 (the p.V274I polymorphism). The P‐value for the overall test was <0.001. In haplotype analyses, several of the fetal and maternal haplotype relative risks were statistically significant individually but were not strong enough to show up on the overall test (P=0.113). Taken together, these findings further support a role for IRF6 variants in clefting of the lip and provide specific risk estimates in a Norwegian population. Genet. Epidemiol. 2008. © 2008 Wiley‐Liss, Inc.

Searching for rare genetic variants associated with complex diseases can be facilitated by enriching for diseased carriers of rare variants by sampling cases from pedigrees enriched for disease, possibly with related or unrelated controls. This strategy, however, complicates analyses because of shared genetic ancestry, as well as linkage disequilibrium among genetic markers. To overcome these problems, we developed broad classes of “burden” statistics and kernel statistics, extending commonly used methods for unrelated case‐control data to allow for known pedigree relationships, for autosomes and the X chromosome. Furthermore, by replacing pedigree‐based genetic correlation matrices with estimates of genetic relationships based on large‐scale genomic data, our methods can be used to account for population‐structured data. By simulations, we show that the type I error rates of our developed methods are near the asymptotic nominal levels, allowing rapid computation of P‐values. Our simulations also show that a linear weighted kernel statistic is generally more powerful than a weighted “burden” statistic. Because the proposed statistics are rapid to compute, they can be readily used for large‐scale screening of the association of genomic sequence data with disease status.

We selected 262 case‐parent triads from a population‐based study of orofacial clefts in Norway, and examined variants of developmental genes TGFA, TGFB3, and MSX1 in the etiology of orofacial clefts. One hundred seventy‐four triads of cleft lip cases (CL±P) and 88 triads of cleft palate only cases (CPO) were analyzed. There was little evidence for an association of any of these genes with CL±P. The strongest association was a 1.7‐fold risk with two copies of the TGFB3‐CA variant (95% CI=0.9–3.0). Among CPO cases, there was a 3‐fold risk with two copies of the TGFA TaqI A2 allele, and no increase with one copy. Assuming this to be a recessive effect, we estimated a 3.2‐fold risk among babies homozygous for the variant (95% CI=1.1–9.2). Furthermore, there was strong evidence of gene‐gene interaction. While there was only a weak association of the MSX1‐CA variant with CPO, the risk was 9.7‐fold (95% CI=2.9–32) among children homozygous for both the MSX1‐CA A4 allele and the TGFA A2 allele. No association of CPO with the TGFA variant was seen among the other MSX1‐CA genotypes. In conclusion, no strong associations were found between CL±P and variants at these three genes. There was a possible recessive effect of the TGFA TaqI variant on the risk of CPO, with a 3‐fold risk among children homozygous for the variant. The effect of this TGFA genotype was even stronger among children homozygous for the MSX1‐CA A4 allele, raising the possibility of interaction between these two genes. Genet Epidemiol 24:230–239, 2003. © 2003 Wiley‐Liss, Inc.