Michael Mosteller

Genetic variations in HLA-B region and hypersensitivity reactions to abacavir.

Hypersensitivity to abacavir affects about 4% of patients who receive the drug for HIV-1 infection. We did a retrospective, case-control study to identify multiple markers in the vicinity of HLA-B associated with hypersensitivity reactions. HLA-B57 was present in 39 (46%) of 84 patients versus four (4%) of 113 controls (p<0 small middle dot0001). However, because of low numbers of women and other ethnic groups enrolled, these findings relate largely to white men. The lower sensitivity of HLA-B57 for predicting hypersensitivity to abacavir identified in this study compared with a previous report highlights that predictive values for markers will vary across populations. Clinical monitoring and management of hypersensitivity reactions among patients receiving abacavir must remain unchanged.

High Sensitivity of Human Leukocyte Antigen-B*5701 as a Marker for Immunologically Confirmed Abacavir Hypersensitivity in White and Black Patients

BACKGROUND Although the human leukocyte antigen (HLA)-B*5701 is highly associated with a hypersensitivity reaction (HSR) to abacavir (ABC), variable sensitivities have been reported when clinical data alone have been used to define an ABC HSR. This study evaluated the sensitivity of detection of the HLA-B*5701 allele as a marker of ABC HSRs in both white and black patients, using skin patch testing to supplement clinical diagnosis. METHODS White and black patients, identified through chart review, were classified as having received a diagnosis of an ABC HSR based on clinical findings only (a clinically suspected ABC HSR) or based on clinical findings and a positive skin patch test result (an immunologically confirmed [IC] ABC HSR). Control subjects were racially matched subjects who tolerated ABC for >/=12 weeks without experiencing an ABC HSR. Patients and control subjects were tested for the presence of HLA-B*5701. Sensitivity, specificity, and odds ratios for the detection of HLA-B*5701 as a marker for an ABC HSR were calculated for white and black participants. RESULTS Forty-two (32.3%) of 130 white patients and 5 (7.2%) of 69 black patients who met the criteria for clinically suspected HSRs had IC HSRs. All 42 white patients with IC HSRs were HLA-B*5701 positive (sensitivity, 100%; odds ratio, 1945; 95% confidence interval, 110-34,352). Among all white patients with clinically suspected HSRs, sensitivity was 44% (57 of 130 patients tested positive for HLA-B*5701); specificity among white control subjects was 96%. Five of 5 black patients with IC HSRs were HLA-B*5701 positive (sensitivity, 100%; odds ratio, 900; 95% confidence interval, 38-21,045). Among black patients with clinically suspected HSRs, the sensitivity was 14% (10 of 69 tested positive for HLA-B*5701); specificity among black control subjects was 99%. CONCLUSIONS Although IC ABC HSRs are uncommon in black persons, the 100% sensitivity of HLA-B*5701 as a marker for IC ABC HSRs in both US white and black patients suggests similar implications of the association between HLA-B*5701 positivity and risk of ABC HSRs in both races.

Genetic association studies to detect adverse drug reactions: abacavir hypersensitivity as an example

Abacavir hypersensitivity (ABC HSR) is a treatment-limiting adverse event associated with the use of the antiretroviral medicine, abacavir. The objective of the ABC HSR pharmacogenetics program was to identify clinically useful genetic risk factors to predict an individual patients risk for ABC HSR. The major histocompatibility complex allele, HLA-B*5701, was identified retrospectively and confirmed with independent sample sets. The clinical utility of prospective HLA-B*5701 screening was demonstrated in a blinded randomized clinical trial and in open-label cohorts. Screening has been incorporated into clinical practice and the ABC HSR pharmacogenetics program has been highlighted as a success by pharmacogenetics researchers. Important lessons from this pharmacogenetics program will be discussed in this paper.

GLCCI1 rs37973 does not influence treatment response to inhaled corticosteroids in white subjects with asthma

To The Editor: Inhaled corticosteroids (ICS) are the primary anti-inflammatory therapy for the control and management of asthma, but their effects are characterised by some inter-individual variability that may have a genetic basis1–4. Identification of genetic markers that predict ICS treatment response will facilitate individualised treatment for asthma patients in the future, particularly in those with more severe disease. An association between genetic variation in GLCCI1 and response to ICS therapy in non-Hispanic white asthma subjects was observed by Tantisira et al.5 Genome wide association analysis of 118 trios (one asthmatic child and two parents) from the NHLBI Childhood Asthma Management Program (CAMP) identified 13 single nucleotide polymorphisms (SNPs) with evidence for association with the level of ICS treatment response. These 13 SNPs which included the GLCCI1 promoter polymorphism rs37972, were genotyped and evaluated in four additional independent collections: adult studies SOCS (Salmeterol Or CorticosteroidS) and SLIC (SalmeteroL ± Inhaled CorticosteroidS) (n=264); a second adult study (n=385); LOCCS (Leukotriene modifier Or Corticosteroid or Corticosteroid-Salmeterol) (n=185) and CARE (Childhood Asthma Research and Education) (n=101). In three of the four replicate populations GLCCI1 rs37973, a functional promoter polymorphism5 in complete linkage disequilibrium with rs37972 in white populations, was associated (P<0.05) with change in FEV1 (forced expiratory volume in one second) after 4–8 weeks of ICS treatment. The combined P value measuring association between rs37973 and ICS response over the four collections (n=935) was 0.0007. Tantisira et al. concluded that this functional GLCCI1 polymorphism, rs37973, was associated with response to ICS in asthma patients. Using data from a recently completed genome wide association study of response to steroid therapy, we sought to confirm this observation in a homogeneous, well-characterised population of n=1,924 non-Hispanic white subjects by testing for association between rs37973 and measures of corticosteroid response in subjects treated with either fluticasone furoate (FF) or fluticasone propionate (FP) in seven GSK-sponsored clinical trials (NCT01165138; NCT01134042; NCT01086384; NCT01159912; NCT00603382; NCT00603278; NCT00603746). All seven studies were randomised, double blind, placebo controlled, parallel group multicentre studies in adolescent and adult subjects and employed change from baseline in FEV1 as their primary end-point, apart from HZA106837 which also investigated asthma exacerbations. HZA106837 (N=616) required each subject to have an asthma exacerbation within the 12 months prior to enrolment, whereas the other studies excluded subjects with previous asthma exacerbations. FEV1 was assessed at week 8 for all studies except HZA106837, which used week 12 data. Overall, except for FF and FP dose, baseline demographics and study characteristics were similar across all seven studies (Table 1). TABLE 1 Baseline, demographic and steroid response characteristics in seven GSK-sponsored clinical studies Germline DNA was extracted from peripheral blood collected from all 1,924 subjects all of whom provided consent for genetic analysis. Genotyping used either the KBiosciences Competitive Allele Specific PCR SNP genotype System (KASPar) (Hoddesdon, Herts, UK) or the Illumina Omni1-Quad panel (Expression Analysis, Durham, NC, USA). Analysis was undertaken in 1,916 subjects, including four with imputed data from rs37972. Eight subjects had missing covariate data. Genetic association between rs37973 and ICS response was evaluated, with ICS treatment response defined as change from baseline in trough FEV1 using the last observation carried forward (in any subject who did not complete the specific trial), to week 8 or week 12 of FF or FP treatment. Change in trough FEV1 was regressed against covariates identified in this asthma population: age, percent of predicted baseline FEV1, study, height, asthma duration and drug (FF versus FP). We also evaluated the influence of rs37973 on subject placement within the highest and lowest response quartiles. Subjects who fell into the lowest (n=479) and highest (n=479) response quartiles were identified. Logistic regression was used to fit a model with quartile of response as the dependent variable, and genotype and relevant covariates as the independent variables. The P value measuring heterogeneity among the seven studies was 0.98, allowing pooling of data at the subject level. The minor allele frequency of rs37973 was 0.44 and its genotype frequencies were consistent with Hardy-Weinberg equilibrium (P=0.48). Covariate-adjusted FEV1 change was regressed on rs37973 genotype (Figure 1). Rs37973 did not influence change from baseline in FEV1 in this sample of 1,916 non-Hispanic white subjects treated with either FF or FP (P=0.15). However, this regression analysis suggested a trend toward a slightly lower ICS response for each additional copy of the rs37973 G allele. This direction of effect was consistent with that observed by Tantisira et al. In addition, the percentage change from baseline FEV1 (unadjusted for covariates, results not shown) was 10.4±0.8% in AA homozygotes and 8.8±1.0% in GG homozygotes, while the overall mean response was 9.8±0.4%. FIGURE 1 GLCCI1 rs37973 genotype does not significantly influence steroid response in 1,916 asthma patients This genetic marker did not influence subject membership within response quartiles (P=0.08, Odds Ratio (OR) =1.39, 95% CI 0.96–2.00). The ORs in each clinical study ranged from 0.61 (HZA106827) (N=616) to 4.42 (FFA112059), which is one of two smallest clinical trials, N=143 (Table 1). Meta-analysis of the influence of rs37973 on subject placement within response quartile across all seven clinical studies, revealed similar results to the subject level pooled data, suggesting a non-significant trend towards lower ICS response in GLCCI1 rs37973 GG homozygotes, compared to AA homozygotes (P=0.11, OR 1.32, 95% CI 0.94–1.87). In order to closely mimic the analyses of Tantisira et al 5, change in trough FEV1 was also regressed against age, sex and height. All analyses in our sample were repeated using these covariates; there were no qualitative differences between the two sets of results. Asthma is currently estimated to affect ~315 million people worldwide6. A robust genetic predictor of ICS response in asthma patients would provide clinical value7 as inter-individual variability in ICS treatment response is commonly observed. Tantisira et al 5 reported an association (P=0.0007) in a pooled analysis between GLCCI1 rs37973 and ICS treatment response as measured over 4–8 weeks in 935 white non-Hispanic adults and children, and an OR of 2.36 in a subject level pooled analysis evaluating subject placement with response quartiles. In this larger sample set, n=1,916, drawn from seven clinical studies, we did not confirm GLCCI1 rs37973 as a predictor of ICS response. However, the discrepant outcomes might have been due to various factors: the GSK studies were clinical trials specifically designed with FEV1 change as the primary endpoint for all studies except one, whereas those of Tantisira were designed around a range of other primary endpoints, including lung growth, time to treatment failure and the percentage of asthma control days; paediatric study participants were only included in the initial Tantisira5 cohort; and the duration of ICS treatment at the time of FEV1 assessment varied between the two evaluations. Further genetic studies will be required to fully elucidate the potential role of GLCCI1 in ICS treatment response in asthma patients.

Distribution of HLA-B alleles in a Ugandan HIV-infected adult population: NORA pharmacogenetic substudy of DART.

Objectives  To determine the frequencies of HLA‐B alleles in Ugandan patients in the NORA substudy of the DART trial and to compare HLA‐B allele frequencies in those with and without clinically diagnosed hypersensitivity reaction (HSR).

Genetic variants in the epithelial sodium channel associate with oedema in type 2 diabetic patients receiving the peroxisome proliferator-activated receptor gamma agonist farglitazar

Peroxisome proliferator-activated receptor gamma (PPAR&ggr;) agonists are highly effective in the treatment of type 2 diabetes. In some patients, PPAR&ggr; ligands are associated with fluid retention/oedema, for which the mechanism is not fully understood. A pharmacogenetic study was undertaken to investigate effects of variations in 21 candidate genes related to epithelial sodium channel (ENaC) pathways on oedema. This study used DNA samples collected from type 2 diabetes phase III clinical trials of the PPAR&ggr; agonist farglitazar (administered alone or in combination with insulin or glyburide) and investigated oedema reported as an adverse event as phenotype. Initial case–control analysis of oedema identified candidate gene single nucleotide polymorphisms with significant associations. These included three polymorphisms in ENaC&bgr; subunit (SCNN1B) that showed significant associations (P<0.05) with the two combination treatments in discrete regions of the gene, but not farglitazar treatment alone. Sequencing of SCNN1B in 207 Caucasian participants receiving farglitazar plus insulin or glyburide combination therapies, identified additional polymorphisms that were also significantly associated with oedema (P<0.0005) and maintained the treatment-regional associations. Further covariate analysis accounting for clinical factors influencing oedema supported these observations. One of the SCNN1B polymorphisms, at position −405 of the 5′ flanking region (rs34241435), was predicted to modify transcriptional interactions and in a transfected COS cell luciferase reporter gene assay exhibited higher promoter activity. These exploratory studies provide clinical pharmacogenetic and functional genomic evidence to support a pivotal role for ENaC regulation in PPAR&ggr;-induced oedema and provide insight into mechanisms and possible management of this side effect.

No evidence of large genetic effects on steroid response in asthma patients

Background: Inhaled corticosteroids (ICSs) are considered the most effective anti‐inflammatory therapy for asthma control and management; however, there is substantial treatment response variability. Objective: We sought to identify genetic markers of ICS response by conducting the largest pharmacogenetic investigation to date in 2672 ICS‐treated patients with asthma. Methods: Genotyping and imputation was performed in fluticasone furoate (FF) or fluticasone propionate–treated patients with asthma from 3 phase IIB and 4 phase IIIA randomized, double‐blind, placebo‐controlled, parallel group, multicenter studies. The primary end point analyzed was change in trough FEV1 (&Dgr;FEV1) from baseline to 8 to 12 weeks of treatment. Results: More than 9.8 million common genetic variants (minor allele frequency ≥ 1%) were analyzed to test for association with &Dgr;FEV1. No genetic variant met the prespecified threshold for statistical significance. Conclusions: This study provides no evidence to confirm previously reported associations between candidate genetic variants and ICS response (&Dgr;FEV1) in patients with asthma. In addition, no variant satisfied the criterion for genome‐wide significance in our study. Common genetic variants are therefore unlikely to prove useful as predictive biomarkers of ICS response in patients with asthma.

Exploring the roles of UGT1A1 and UGT1A3 in oral clearance of GSK2190915, a 5-lipoxygenase-activating protein inhibitor.

Pharmacokinetic variability in drug exposure is a concern for all compounds in development including those for the treatment of asthma and other respiratory disorders. Substantial variability in the oral clearance of GSK2190915, a 5-lipoxygenase-activating protein inhibitor that attenuates the production of leukotriene B4 and cysteinyl leukotrienes, is largely unaccounted for by clinical variables. A study of 41 patients, 78% (32/41) of whom were non-Hispanic whites, with mild to moderate asthma identified an association of UGT1A1*28 and UGT1A3*2 with the oral clearance of GSK2190915 (P=3.8×10⁻⁴ and 1.2×10⁻⁵, respectively). However, in a subsequent replication study of 403 non-Hispanic white patients with asthma, we failed to observe a statistically significant association between oral clearance of GSK2190915 and either UGT1A1*28 or UGT1A3*2 (P>0.05). Therefore, genetic effects that could explain the systemic exposure level variability of GSK2190915 were not identified.

Genetic analysis of asthma exacerbations

BACKGROUND Identifying genetic markers of susceptibility to exacerbations may improve patient management, decrease morbidity, and lead to drug development. OBJECTIVES To assess whether genetic markers associated with severe asthma exacerbations in previous reports are associated with less severe events that do not require intensive care and intubation and to identify additional markers in candidate genes and throughout the genome. METHODS A total of 199 patients and 502 controls (individuals without an exacerbation) were identified from 4 clinical trials. We genotyped 51 markers from 17 genes previously reported to be associated with exacerbations; a whole genome scan was used to identify additional markers. Admixture analysis was conducted to characterize the presence of ancestral groups. The genetic marker effects were assessed by logistic regression for each study followed by a meta-analysis. RESULTS Several coding variants in the IL4R gene had a genetic effect across 3 studies, including rs1805011 in IL4R (P < .0006). In addition, 3 markers in the IFNB1 gene showed evidence of association (P < .002) but only in the study with African Americans. Because these markers did not meet the prespecified multiplicity-adjusted significance level of P = .0002, we were unable to confirm previously published results for less severe events. The whole genome scan identified genes related to mast cell mediator release. The admixture analysis suggests that ancestry was best characterized by the presence of 3 ancestral groups. CONCLUSION We were unable to confirm previously reported associations of genetic markers with asthma exacerbations. Although, in general, the patients studied had less severe asthma than patients in earlier reports, these results suggest involvement of similar pathways. TRIAL REGISTRATION clinicaltrials.gov Identifiers: NTC00452699, NCT00452348, NTC00102765, NCT00843193.

Distribution of HLA-B alleles in a Ugandan HIV-infected adult population: NORA pharmacogenetic substudy of DART: HLA-B*5701 and hypersensitivity in Africa

Objectives  To determine the frequencies of HLA‐B alleles in Ugandan patients in the NORA substudy of the DART trial and to compare HLA‐B allele frequencies in those with and without clinically diagnosed hypersensitivity reaction (HSR).