Barbara Truitt

Genetic evidence for role of carotenoids in age-related macular degeneration in the carotenoids in age-related eye disease study (CAREDS)

PURPOSE We tested variants in genes related to lutein and zeaxanthin status for association with age-related macular degeneration (AMD) in the Carotenoids in Age-Related Eye Disease Study (CAREDS). METHODS Of 2005 CAREDS participants, 1663 were graded for AMD from fundus photography and genotyped for 424 single nucleotide polymorphisms (SNPs) from 24 candidate genes for carotenoid status. Of 337 AMD cases 91% had early or intermediate AMD. The SNPs were tested individually for association with AMD using logistic regression. A carotenoid-related genetic risk model was built using backward selection and compared to existing AMD risk factors using the area under the receiver operating characteristic curve (AUC). RESULTS A total of 24 variants from five genes (BCMO1, BCO2, NPCL1L1, ABCG8, and FADS2) not previously related to AMD and four genes related to AMD in previous studies (SCARB1, ABCA1, APOE, and ALDH3A2) were associated independently with AMD, after adjusting for age and ancestry. Variants in all genes (not always the identical SNPs) were associated with lutein and zeaxanthin in serum and/or macula, in this or other samples, except for BCO2 and FADS2. A genetic risk score including nine variants significantly (P = 0.002) discriminated between AMD cases and controls beyond age, smoking, CFH Y402H, and ARMS2 A69S. The odds ratio (95% confidence interval) for AMD among women in the highest versus lowest quintile for the risk score was 3.1 (2.0-4.9). CONCLUSIONS Variants in genes related to lutein and zeaxanthin status were associated with AMD in CAREDS, adding to the body of evidence supporting a protective role of lutein and zeaxanthin in risk of AMD.

Discovery of Antivirulence Agents against Methicillin-Resistant Staphylococcus aureus

ABSTRACT Antivirulence agents inhibit the production of disease-causing virulence factors but are neither bacteriostatic nor bactericidal. Antivirulence agents against methicillin-resistant Staphylococcus aureus (MRSA) strain USA300, the most widespread community-associated MRSA strain in the United States, were discovered by virtual screening against the response regulator AgrA, which acts as a transcription factor for the expression of several of the most prominent S. aureus toxins and virulence factors involved in pathogenesis. Virtual screening was followed by similarity searches in the databases of commercial vendors. The small-molecule compounds discovered inhibit the production of the toxins alpha-hemolysin and phenol-soluble modulin α in a dose-dependent manner without inhibiting bacterial growth. These antivirulence agents are small-molecule biaryl compounds in which the aromatic rings either are fused or are separated by a short linker. One of these compounds is the FDA-approved nonsteroidal anti-inflammatory drug diflunisal. This represents a new use for an old drug. Antivirulence agents might be useful in prophylaxis and as adjuvants in antibiotic therapy for MRSA infections.

Genome-Wide Association and Trans-ethnic Meta-Analysis for Advanced Diabetic Kidney Disease: Family Investigation of Nephropathy and Diabetes (FIND).

Diabetic kidney disease (DKD) is the most common etiology of chronic kidney disease (CKD) in the industrialized world and accounts for much of the excess mortality in patients with diabetes mellitus. Approximately 45% of U.S. patients with incident end-stage kidney disease (ESKD) have DKD. Independent of glycemic control, DKD aggregates in families and has higher incidence rates in African, Mexican, and American Indian ancestral groups relative to European populations. The Family Investigation of Nephropathy and Diabetes (FIND) performed a genome-wide association study (GWAS) contrasting 6,197 unrelated individuals with advanced DKD with healthy and diabetic individuals lacking nephropathy of European American, African American, Mexican American, or American Indian ancestry. A large-scale replication and trans-ethnic meta-analysis included 7,539 additional European American, African American and American Indian DKD cases and non-nephropathy controls. Within ethnic group meta-analysis of discovery GWAS and replication set results identified genome-wide significant evidence for association between DKD and rs12523822 on chromosome 6q25.2 in American Indians (P = 5.74x10-9). The strongest signal of association in the trans-ethnic meta-analysis was with a SNP in strong linkage disequilibrium with rs12523822 (rs955333; P = 1.31x10-8), with directionally consistent results across ethnic groups. These 6q25.2 SNPs are located between the SCAF8 and CNKSR3 genes, a region with DKD relevant changes in gene expression and an eQTL with IPCEF1, a gene co-translated with CNKSR3. Several other SNPs demonstrated suggestive evidence of association with DKD, within and across populations. These data identify a novel DKD susceptibility locus with consistent directions of effect across diverse ancestral groups and provide insight into the genetic architecture of DKD.

Genetic determinants of macular pigments in women the carotenoids in age-related eye disease study

PURPOSE To investigate genetic determinants of macular pigment optical density in women from the Carotenoids in Age-Related Eye Disease Study (CAREDS), an ancillary study of the Womens Health Initiative Observational Study. METHODS 1585 of 2005 CAREDS participants had macular pigment optical density (MPOD) measured noninvasively using customized heterochromatic flicker photometry and blood samples genotyped for 440 single nucleotide polymorphisms (SNPs) in 26 candidate genes related to absorption, transport, binding, and cleavage of carotenoids directly, or via lipid transport. SNPs were individually tested for associations with MPOD using least-squares linear regression. RESULTS Twenty-one SNPs from 11 genes were associated with MPOD (P ≤ 0.05) after adjusting for dietary intake of lutein and zeaxanthin. This includes variants in or near genes related to zeaxanthin binding in the macula (GSTP1), carotenoid cleavage (BCMO1), cholesterol transport or uptake (SCARB1, ABCA1, ABCG5, and LIPC), long-chain omega-3 fatty acid status (ELOVL2, FADS1, and FADS2), and various maculopathies (ALDH3A2 and RPE65). The strongest association was for rs11645428 near BCMO1 (βA = 0.029, P = 2.2 × 10(-4)). Conditional modeling within genes and further adjustment for other predictors of MPOD, including waist circumference, diabetes, and dietary intake of fiber, resulted in 13 SNPs from 10 genes maintaining independent association with MPOD. Variation in these single gene polymorphisms accounted for 5% of the variability in MPOD (P = 3.5 × 10(-11)). CONCLUSIONS Our results support that MPOD is a multi-factorial phenotype associated with variation in genes related to carotenoid transport, uptake, and metabolism, independent of known dietary and health influences on MPOD.

Vitamin D Intake and Season Modify the Effects of the GC and CYP2R1 Genes on 25-Hydroxyvitamin D Concentrations

Vitamin D deficiency {defined by the blood concentration of 25-hydroxyvitamin D [25(OH)D]} has been associated with many adverse health outcomes. Genetic and nongenetic factors account for variation in 25(OH)D, but the role of interactions between these factors is unknown. To assess this, we examined 1204 women of European descent from the Carotenoids in Age-Related Eye Disease Study, an ancillary study of the Womens Health Initiative Observational Study. Twenty-nine single nucleotide polymorphisms (SNPs) in 4 genes, GC, CYP2R1, DHCR7, and CYP24A1, from recent meta-analyses of 25(OH)D genome-wide association studies were genotyped. Associations between these SNPs and 25(OH)D were tested using generalized linear regression under an additive genetic model adjusted for age, blood draw month, and ancestry. Results were stratified by season of blood draw and, separately, vitamin D intake for the 6 SNPs showing a significant association with 25(OH)D at the P < 0.01 level. Two nonsynonymous SNPs in GC and 4 SNPs in CYP2R1 were strongly associated with 25(OH)D in individuals whose blood was drawn in summer (P ≤ 0.002) but not winter months and, independently, in individuals with vitamin D intakes ≥400 (P ≤ 0.004) but not <400 IU/d (10 μg/d). This effect modification, if confirmed, has important implications for the design of genetic studies for all health outcomes and for public health recommendations and clinical practice guidelines regarding the achievement of adequate vitamin D status.

Differing Roles for TCF4 and COL8A2 in Central Corneal Thickness and Fuchs Endothelial Corneal Dystrophy

Fuchs endothelial corneal dystrophy (FECD) is the most common late-onset, vision-threatening corneal dystrophy in the United States, affecting about 4% of the population. Advanced FECD involves a thickening of the cornea from stromal edema and changes in Descemet membrane. To understand the relationship between FECD and central corneal thickness (CCT), we characterized common genetic variation in COL8A2 and TCF4, genes previously implicated in CCT and/or FECD. Other genes previously associated with FECD (PITX2, ZEB1, SLC4A11), and genes only known to affect CCT (COL5A1, FOXO1, AVGR8, ZNF469) were also interrogated. FECD probands, relatives and controls were recruited from 32 clinical sites; a total of 532 cases and 204 controls were genotyped and tested for association of FECD case/control status, a 7-step FECD severity scale and CCT, adjusting for age and sex. Association of FECD grade with TCF4 was highly significant (OR  = 6.01 at rs613872; p = 4.8×10−25), and remained significant when adjusted for changes in CCT (OR  = 4.84; p = 2.2×10−16). Association of CCT with TCF4 was also significant (p = 6.1×10−7), but was abolished with adjustment for FECD grade (p = 0.92). After adjusting for FECD grade, markers in other genes examined were modestly associated (p ∼ 0.001) with FECD and/or CCT. Thus, common variants in TCF4 appear to influence FECD directly, and CCT secondarily via FECD. Additionally, changes in corneal thickness due to the effect of other loci may modify disease severity, age-at-onset, or other biomechanical characteristics.

Exome array analysis identifies CAV1/CAV2 as a susceptibility locus for intraocular pressure

PURPOSE Intraocular pressure (IOP) is an important clinical parameter in the evaluation of ocular health. Elevated IOP is a major risk factor for primary open-angle glaucoma (POAG). The goal of this study was to identify rare and less common variants that influence IOP. METHODS We performed an exome array analysis in a subset of 1660 individuals from a population-based cohort, the Beaver Dam Eye Study. Associations with IOP were tested on 45,849 single nucleotide variants and 12,390 autosomal genes across the genome. RESULTS Intraocular pressure was suggestively associated with novel variants located in FAR2 at 12p11.22 (rs4931170, P = 1.2 × 10(-5)), in GGA3 at 17q25.1 (rs52809447, P = 6.7 × 10(-5)), and in PKDREJ at 22q13.31 (rs7291444, P = 7.4 × 10(-5)). Gene-based analysis found suggestive associations between IOP and the genes HAP1, MTBP, FREM3, and PHF12. We successfully replicated the associations with GAS7 (P = 7.4 × 10(-3)) for IOP, and also identified a previously reported POAG locus in the CAV1/CAV2 region to be associated with IOP (P = 3.3 × 10(-3)). This association was confirmed in a meta-analysis with three published genome-wide association studies (Pcombined = 4.0 × 10(-11)). CONCLUSIONS Our results suggest that novel genetic variants and genes with multiple, less common variants may play a role in the control of IOP. The implication of the caveolin genes, CAV1/CAV2, as a common genetic factor influencing both IOP variations and POAG may provide new insights of the underlying mechanism leading to glaucoma and glaucomatous visual field loss.

Rare and common variants in extracellular matrix gene Fibrillin 2 (FBN2) are associated with macular degeneration

Neurodegenerative diseases affecting the macula constitute a major cause of incurable vision loss and exhibit considerable clinical and genetic heterogeneity, from early-onset monogenic disease to multifactorial late-onset age-related macular degeneration (AMD). As part of our continued efforts to define genetic causes of macular degeneration, we performed whole exome sequencing in four individuals of a two-generation family with autosomal dominant maculopathy and identified a rare variant p.Glu1144Lys in Fibrillin 2 (FBN2), a glycoprotein of the elastin-rich extracellular matrix (ECM). Sanger sequencing validated the segregation of this variant in the complete pedigree, including two additional affected and one unaffected individual. Sequencing of 192 maculopathy patients revealed additional rare variants, predicted to disrupt FBN2 function. We then undertook additional studies to explore the relationship of FBN2 to macular disease. We show that FBN2 localizes to Bruchs membrane and its expression appears to be reduced in aging and AMD eyes, prompting us to examine its relationship with AMD. We detect suggestive association of a common FBN2 non-synonymous variant, rs154001 (p.Val965Ile) with AMD in 10 337 cases and 11 174 controls (OR = 1.10; P-value = 3.79 × 10(-5)). Thus, it appears that rare and common variants in a single gene--FBN2--can contribute to Mendelian and complex forms of macular degeneration. Our studies provide genetic evidence for a key role of elastin microfibers and Bruchs membrane in maintaining blood-retina homeostasis and establish the importance of studying orphan diseases for understanding more common clinical phenotypes.

Genome-wide association study identifies three novel loci in Fuchs endothelial corneal dystrophy

The structure of the cornea is vital to its transparency, and dystrophies that disrupt corneal organization are highly heritable. To understand the genetic aetiology of Fuchs endothelial corneal dystrophy (FECD), the most prevalent corneal disorder requiring transplantation, we conducted a genome-wide association study (GWAS) on 1,404 FECD cases and 2,564 controls of European ancestry, followed by replication and meta-analysis, for a total of 2,075 cases and 3,342 controls. We identify three novel loci meeting genome-wide significance (P<5 × 10−8): KANK4 rs79742895, LAMC1 rs3768617 and LINC00970/ATP1B1 rs1200114. We also observe an overwhelming effect of the established TCF4 locus. Interestingly, we detect differential sex-specific association at LAMC1, with greater risk in women, and TCF4, with greater risk in men. Combining GWAS results with biological evidence we expand the knowledge of common FECD loci from one to four, and provide a deeper understanding of the underlying pathogenic basis of FECD.

Polymorphisms in TICAM2 and IL1B are associated with TB

Human genetic susceptibility for tuberculosis (TB) has been demonstrated by several studies, but few have examined the multiple innate and adaptive immunity genes comprehensively, age-specific effects and/or resistance to Mycobacterium tuberculosis (Mtb) infection (resistors (RSTRs)). We hypothesized that RSTRs, defined by a persistently negative tuberculin skin test, may have different genetic influences than Mtb disease. We examined 29 candidate genes in pathways that mediate immune responses to Mtb in subjects in a household contact study in Kampala, Uganda. We genotyped 546 haplotype-tagging single-nucleotide polymorphisms (SNPs) in 835 individuals from 481 families; 28.7% had TB, 10.5% were RSTRs, and the remaining 60.8% had latent Mtb infection. Among our most significant findings were SNPs in TICAM2 (P=3.6 × 10−6) and IL1B (P=4.3 × 10−5) associated with TB. Multiple SNPs in IL4 and TOLLIP were associated with TB (P<0.05). Age–genotype interaction analysis revealed SNPs in IL18 and TLR6 that were suggestively associated with TB in children aged ⩽10 years (P=2.9 × 10−3). By contrast, RSTR was associated with SNPs in NOD2, SLC6A3 and TLR4 (nominal P<0.05); these genes were not associated with TB, suggesting distinct genetic influences. We report the first association between TICAM2 polymorphisms and TB and between IL18 and pediatric TB.