A Scan of Pleiotropic Immune Mediated Disease Genes Identifies Novel Determinants of Baseline FVIII Inhibitor Status in Hemophilia-A.

Hemophilia-A (HA) is caused by heterogeneous loss-of-function factor (F)VIII gene (F8)-mutations and deficiencies in plasma-FVIII-activity that impair intrinsic-pathway-mediated coagulation-amplification. The standard-of-care for severe-HA-patients is regular infusions of therapeutic-FVIII-proteins (tFVIIIs) but ~30% develop neutralizing-tFVIII-antibodies called "FVIII-inhibitors (FEIs)" and become refractory. We used the PATH study and ImmunoChip to scan immune-mediated-disease (IMD)-genes for novel and/or replicated genomic-sequence-variations associated with baseline-FEI-status while accounting for non-independence of data due to genetic-relatedness and F8-mutational-heterogeneity. The baseline-FEI-status of 450 North American PATH subjects-206 with black-African-ancestry and 244 with white-European-ancestry-was the dependent variable. The F8-mutation-data and a genetic-relatedness matrix were incorporated into a binary linear-mixed model of genetic association with baseline-FEI-status. We adopted a gene-centric-association-strategy to scan, as candidates, pleiotropic-IMD-genes implicated in the development of either ³2 autoimmune-/autoinflammatory-disorders (AADs) or ³1 AAD and FEIs. Baseline-FEI-status was significantly associated with SNPs assigned to NOS2A (rs117382854; p=3.2E-6) and B3GNT2 (rs10176009; p=5.1E-6), which have functions in anti-microbial-/-tumoral-immunity. Among IMD-genes implicated in FEI-risk previously, we identified strong associations with CTLA4 assigned SNPs (p=2.2E-5). The F8-mutation-effect underlies ~15% of the total heritability for baseline-FEI-status. Additive genetic heritability and SNPs in IMD-genes account for >50% of the patient-specific variability in baseline-FEI-status. Race is a significant determinant independent of F8-mutation-effects and non-F8-genetics.

Endogenous factor VIII synthesis from the intron 22-inverted F8 locus may modulate the immunogenicity of replacement therapy for hemophilia A.

Neutralizing antibodies (inhibitors) to replacement factor VIII (FVIII, either plasma derived or recombinant) impair the effective management of hemophilia A. Individuals with hemophilia A due to major deletions of the FVIII gene (F8) lack antigenically cross-reactive material in their plasma ("CRM-negative"), and the prevalence of inhibitors in these individuals may be as high as 90%. Conversely, individuals with hemophilia A caused by F8 missense mutations are CRM-positive, and their overall prevalence of inhibitors is <10% (ref. 2). Individuals with the F8 intron 22 inversion (found in ∼50% of individuals with severe hemophilia A) have been grouped with the former on the basis of their genetic defect and CRM-negative status. However, only ∼20% of these individuals develop inhibitors. Here we demonstrate that the levels of F8 mRNA and intracellular FVIII protein in B lymphoblastoid cells and liver biopsies from individuals with the intron 22 inversion are comparable to those in healthy controls. These results support the hypothesis that most individuals with the intron 22 inversion are tolerized to FVIII and thus do not develop inhibitors. Furthermore, we developed a new pharmacogenetic algorithm that permits the stratification of inhibitor risk for individuals and subpopulations by predicting the immunogenicity of replacement FVIII using, as input, the number of putative T cell epitopes in the infused protein and the competence of major histocompatibility complex class II molecules to present such epitopes. This algorithm showed statistically significant accuracy in predicting the presence of inhibitors in 25 unrelated individuals with the intron 22 inversion.

Impact of the definition utilized on the rate of contrast-induced nephropathy in percutaneous coronary intervention.

Several definitions have been used to assess rates of contrast-induced nephropathy (CIN) in patients undergoing percutaneous coronary intervention (PCI). Whether the definition influences observed rates of CIN is unclear. The Oxilan Registry was the first-ever prospective analysis of the efficacy and safety of ioxilan (low-osmolar and low-viscosity contrast medium), including rates of CIN assessed by multiple definitions, in PCI. From July 2006 to June 2007, consecutive patients undergoing PCI using ioxilan were enrolled. Serum creatinine (SCr) and estimated glomerular filtration rate (eGFR) were assessed at baseline and 3 to 5 days after PCI. CIN was defined as SCr increase >or=0.5 mg/dl, eGFR decrease >or=25%, SCr increase >or=25%, or the composite. Of 400 patients (age 62 +/- 11 years), 19% were women, 37% were diabetic, 22% were anemic, and 8% had a history of congestive heart failure. Baseline SCr was 1.12 +/- 0.3 mg/dl and 24% had an eGFR <60 ml/min. CIN rates were 3.3% (SCr increase >or=0.5 mg/dl), 7.6% (eGFR decrease >or=25%), 10.2% (SCr increase >or=25%), and 10.5% (composite). Hospitalization was prolonged in 3.4% of patients with CIN and none required dialysis. There were no deaths or severe allergic reactions. Non-ST-elevation myocardial infarction and repeat revascularization each occurred in 0.8%. In conclusion, in this unselected population undergoing PCI, CIN ranged in frequency from 3.3% to 10.5% depending on the definition used and was not associated with in-hospital mortality or substantial morbidity, such as dialysis. The wide variation in CIN and its lack of association with adverse outcomes underscore the need for a standardized, clinically relevant definition.

Inhibitors of factor VIII in black patients with hemophilia.

BACKGROUND: Black patients with hemophilia A (factor VIII deficiency) are twice as likely as white patients to produce inhibitors against factor VIII proteins given as replacement therapy. There are six wild-type factor VIII proteins, designated H1 through H6, but only two (H1 and H2) match the recombinant factor VIII products used clinically. H1 and H2 are found in all racial groups and are the only factor VIII proteins found in the white population to date. H3, H4, and H5 have been found only in blacks. We hypothesized that mismatched factor VIII transfusions contribute to the high incidence of inhibitors among black patients. METHODS: We sequenced the factor VIII gene (F8) in black patients with hemophilia A to identify causative mutations and the background haplotypes on which they reside. Results from previous Bethesda assays and information on the baseline severity of hemophilia, age at enrollment, and biologic relationships among study patients were obtained from review of the patients' medical charts. We used multivariable logistic regression to control for these potential confounders while testing for associations between F8 haplotype and the development of inhibitors. RESULTS: Of the 78 black patients with hemophilia enrolled, 24% had an H3 or H4 background haplotype. The prevalence of inhibitors was higher among patients with either of these haplotypes than among patients with haplotype H1 or H2 (odds ratio, 3.6; 95% confidence interval, 1.1 to 12.3; P=0.04), despite a similar spectrum of hemophilic mutations and degree of severity of illness in these two subgroups. CONCLUSIONS: These preliminary results suggest that mismatched factor VIII replacement therapy may be a risk factor for the development of anti-factor VIII alloantibodies.

A linkage analysis of cigarette and alcohol consumption in an unselected Mexican American population.

The use of alcohol and tobacco is highly prevalent. Studying the rate of consumption in a non-selected population could contribute to the elucidation of pathways involved in addiction or to the development of prevention programs. The San Antonio Family Heart Study has approximately 1,400 members with longitudinal data and did not select the proband with regard to exposure status. The goal of this study was to perform genome-wide linkage analysis of the rate of alcohol and cigarette consumption in a "normal" population. We used SOLAR to perform variance-components based analysis of the transformed maximal rate of consumption. Despite estimated heritabilities of 0.52 (P < 0.001) for cigarette and 0.39 (P < 0.001) for alcohol consumption, univariate linkage analyses produced only suggestive LOD scores, however the second suggestive linkage peak for the alcohol phenotype was present at 148 cM on chromosome 10, in the exact vicinity of the peak for the cigarette phenotype. In a bivariate analyses, the environmental correlation between alcohol and cigarette consumption was not significantly different from zero (rho(e) = -0.15, P = 0.18) and the overall genetic correlation was not different from zero (rho(g) = 0.16, P = 0.34). The results from the bivariate linkage analysis found a maximum LOD score of 3.82 (genome-wide P = 0.0054) at 151 cM on chromosome 10, at the location of the overlapping peaks from the univariate analyses.

A sequence variation scan of the coagulation factor VIII (FVIII) structural gene and associations with plasma FVIII activity levels.

Plasma factor VIII coagulant activity (FVIII:C) level is a highly heritable quantitative trait that is strongly correlated with thrombosis risk. Polymorphisms within only 1 gene, the ABO blood-group locus, have been unequivocally demonstrated to contribute to the broad population variability observed for this trait. Because less than 2.5% of the structural FVIII gene (F8) has been examined previously, we resequenced all known functional regions in 222 potentially distinct alleles from 137 unrelated nonhemophilic individuals representing 7 racial groups. Eighteen of the 47 variants identified, including 17 single-nucleotide polymorphisms (SNPs), were previously unknown. As the degree of linkage disequilibrium across F8 was weak overall, we used measured-genotype association analysis to evaluate the influence of each polymorphism on the FVIII:C levels in 398 subjects from 21 pedigrees known as the Genetic Analysis of Idiopathic Thrombophilia project (GAIT). Our results suggested that 92714C>G, a nonsynonymous SNP encoding the B-domain substitution D1241E, was significantly associated with FVIII:C level. After accounting for important covariates, including age and ABO genotype, the association persisted with each C-allele additively increasing the FVIII:C level by 14.3 IU dL(-1) (P = .016). Nevertheless, because the alleles of 56010G>A, a SNP within the 3' splice junction of intron 7, are strongly associated with 92714C>G in GAIT, additional studies are required to determine whether D1241E is itself a functional variant.

Dissection of heterogeneous phenotypes for quantitative trait mapping.

We discuss analyses of Genetic Analysis Workshop 14 data from the Collaborative Study on the Genetics of Alcoholism (COGA) as well as from a simulated complex disease, Kofendrerd personality disorder (KPD), with both genetic and phenotypic heterogeneity. Both data sets included numerous related phenotypes in addition to disease definitions. All analyses either chose from the given selection of phenotypes or defined new ones, including traits that may not have been related to alcoholism or KPD. Some contributors evaluated the genetic components of the trait. Many investigated genome-wide linkage and/or association, using microsatellites and/or single-nucleotide polymorphism (SNP) chip data. Here we will focus on methodological issues that the investigators faced. Their results depended on phenotype selection, whether continuous or discrete, the covariates included, and ethnicity of the study population. For SNP chip data, members of our group detected no difference in results for Affymetrix or Illumina chips, although higher marker density for association studies appeared to be advantageous. Overall, there were some observations that different chromosomal segments, i.e., physical locations on the p-arm, q-arm, or middle segment, might lead to possible differences in type I error rates. This finding and others highlight the importance of empirical determination of P-values to determine significance.

A comparison of discrete versus continuous environment in a variance components-based linkage analysis of the COGA data.

BACKGROUND: The information content of a continuous variable exceeds that of its categorical counterpart. The parameterization of a model may diminish the benefit of using a continuous variable. We explored the use of continuous versus discrete environment in variance components based analyses examining gene x environment interaction in the electrophysiological phenotypes from the Collaborative Study on the Genetics of Alcoholism. RESULTS: The parameterization using the continuous environment produced a greater number of significant gene x environment interactions and lower AICs (Akaike's information criterion). In these cases, the genetic variance increased with increasing cigarette pack-years, the continuous environment of interest. This did not, however, result in enhanced LOD scores when linkage analyses incorporated the gene x continuous environment interaction. CONCLUSION: Alternative parameterizations may better represent the functional relationship between the continuous environment and the genetic variance.

Administrative data as a tool for arthritis surveillance: estimating prevalence and utilization of services.

The public health burden of arthritis and related conditions is incompletely described by commonly used public health surveillance systems. We examined the potential of administrative data as a supplement. The administrative data sources we used underestimated the prevalence of arthritis and overestimated service utilization for persons with arthritis when data from only one year were used. The use of five year's data doubled the prevalence estimate and reduced the service utilization estimate by half. The demographics of the population covered by administrative data also influence the prevalence estimate. Administrative data may usefully supplement routine public health surveillance systems but must be used with caution.