Sequence variability and candidate gene analysis in complex disease: association of micro opioid receptor gene variation with substance dependence

Abstract

To analyze candidate genes and establish complex genotype–phenotype relationships against a background of high natural genome sequence variability, we have developed approaches to (i) compare candidate gene sequence information in multiple individuals; (ii) predict haplotypes from numerous variants; and (iii) classify haplotypes and identify specific sequence variants, or combinations of variants (pattern), associated with the phenotype. Using the human µ opioid receptor gene (OPRM1) as a model system, we have combined these approaches to test a potential role of OPRM1 in substance (heroin/cocaine) dependence. All known functionally relevant regions of this prime candidate gene were analyzed by multiplex sequence comparison in 250 cases and controls; 43 variants were identified and 52 different haplotypes predicted in the subgroup of 172 African-Americans. These haplotypes were classified by similarity clustering into two functionally related categories, one of which was significantly more frequent in substance-dependent individuals. Common to this category was a characteristic pattern of sequence variants [–1793T→A, –1699Tins, –1320A→G, –111C→T, +17C→T (A6V)], which was associated with substance dependence. This study provides an example of approaches that have been successfully applied to the establishment of complex genotype–phenotype relationships in the presence of abundant DNA sequence variation.