Genome-wide approaches to identify pharmacogenetic contributions to adverse drug reactions.

Adverse drug reactions (ADRs) have a major impact on patients, physicians, health care providers, regulatory agencies and pharmaceutical companies. Identifying the genetic contributions to ADR risk may lead to a better understanding of the underlying mechanisms, identification of patients at risk and a decrease in the number of events. Technological advances have made the routine monitoring and investigation of the genetic basis of ADRs during clinical trials possible. We demonstrate through simulation that genome-wide genotyping, coupled with the use of clinically matched or population controls, can yield sufficient statistical power to permit the identification of strong genetic predictors of ADR risk in a prospective manner with modest numbers of ADR cases. The results of a 500,000 single nucleotide polymorphism analysis of abacavir-associated hypersensitivity reaction suggest that the known HLA-B gene region could be identified with as few as 15 cases and 200 population controls in a sequential analysis.

Pharmacogenetics of hypersensitivity to abacavir: from PGx hypothesis to confirmation to clinical utility.

The hypersensitivity (HSR) to abacavir (ABC) pharmacogenetics (PGx) program represents the progression from an exploratory discovery to a validated biomarker. Within the program, two retrospective PGx studies were conducted to identify HIV-1 patients at increased risk for ABC HSR, a treatment-limiting and potentially life-threatening adverse event. A strong statistical association between the major histocompatibility complex allele, HLA-B*5701, and clinically diagnosed ABC HSR was identified but varied between racial populations. Subsequently, ABC skin patch testing was introduced as a research tool to supplement clinical case ascertainment. In a randomized, prospective study evaluating the clinical utility of HLA-B*5701 screening, avoidance of ABC in HLA-B*5701-positive patients significantly reduced clinically diagnosed ABC HSR and eliminated patch test-positive ABC HSR. Finally, a retrospective PGx study supports the generalizability of the association across races. Prospective HLA-B*5701 screening should greatly reduce the incidence of ABC HSR by identifying patients at high risk for ABC HSR before they are treated.

Use of pairwise marker combination and recursive partitioning in a pharmacogenetic genome-wide scan.

The objective of pharmacogenetic research is to identify a genetic marker, or a set of genetic markers, that can predict how a given person will respond to a given medicine. To search for such marker combinations that are predictive of adverse drug events, we have developed and applied two complementary methods to a pharmacogenetic study of the hypersensitivity reaction (HSR) associated with treatment with abacavir, a medicine that is used to treat HIV-infected patients. Our results show that both of these methods can be used to uncover potentially useful predictive marker combinations. The pairwise marker combination method yielded a collection of marker pairs that featured a spectrum of sensitivities and specificities. Recursive partitioning results led to the genetic delineation of multiple risk categories, including those with extremely high and extremely low risk of HSR. These methods can be readily applied in pharmacogenetic candidate gene studies as well as in genome-wide scans.

Maximizing the value of medicines by including pharmacogenetic research in drug development and surveillance.

Genetics provides significant opportunities to maximize the safety and efficacy of medicines. Over the next 3--5 years, it may be possible to develop tools that use selective information from patients' DNA to enable healthcare professionals to predict more accurately those patients at risk of serious adverse events to some medicines currently available. This is likely to be followed, over the next 5--10 years, by the application of the technology to predict more accurately if individual patients will obtain a therapeutic benefit from a particular medicine. The ability to accurately predict patient response will inevitably change the way medicines are developed, evaluated, and prescribed. Advances in single nucleotide polymorphism (SNP) map technology are likely to drive this innovation. Abbreviated SNP profiles will provide the means to define medicine response tests, thereby allowing clinicians to select the medicine to which the patient is likely to gain the greatest benefit and least risk. This will help to maximize efficacy and reduce the incidence of drug-related adverse events. It may be possible to identify SNP profiles during larger Phase II clinical trials which predict efficacy, and use these to form the basis of Phase III entry criteria. As a result, Phase III trials may be streamlined for many medicines making them smaller, more efficient, and more focused. In addition it may be possible to incorporate pharmacogenetics into postmarketing surveillance strategies to provide a means to identify SNPs which predict uncommon serious adverse drug reactions, and so refine the initial medicine response test. The ability to develop drugs with a predictable response will allow clinicians to provide targeted treatment for patients, with greater confidence of safety and efficacy. Patients therefore will receive more efficacious, timely, and well-tolerated medicines. The challenge for those involved in drug development is to model and evaluate the application of pharmacogenetics so that steps can be taken to realize this potential.

Applications of pharmacogenetics to drug development: the Glaxo Wellcome experience.

The Genetics Directorate was established at Glaxo Wellcome (GW) in 1997. The goals of the Directorate are to identify susceptibility genes for common diseases with large unmet therapeutic need, apply genetic methods for the targeted development of medicines so that the right medicine is developed for the right patient, assist in translating gene discoveries into target selection, and represent genetics accurately internally within GW and externally (to laypersons, the medical community, the business community, government representatives, and regulatory agencies). As part of the goal of developing the right medication for the right patient, GW has added genetic research to its clinical drug studies in every major therapeutic area. GW worked closely with international ethicists experienced in the area of genetic research to develop the process and documents that are currently in use; these undergo frequent, rigorous review in light of the changing regulatory and legal environment and the needs of clinical investigators and patients. The addition of genetic research to clinical studies was accompanied by significant education efforts within GW and for study-site personnel, ethics committees, and regulatory authorities. Feedback from all those involved is an integral part of implementing GW's genetic research and is used to fine-tune the processes and protocol and consent document templates. A recently completed review of the approval rate from ethics committees in several countries has revealed trends in EC/IRB (Ethics Committees/Institutional Review Boards) questions and concerns about genetic research. This article will focus on the lessons learned from incorporating genetic research into clinical studies at over 1,500 international sites and will include summaries of the feedback from investigators, EC/IRBs, and regulatory authorities. It also will include a discussion of the potential applications of SNP (single nucleotide polymorphism) map technologies to pharmacogenetics by increasing the ability to correlate patients' genetic information with their response to medicines.

Cardiovascular reactivity in isometric exercise and mental arithmetic in children.

In children, we studied noninvasively the cardiovascular stress responses, including changes over time of systolic blood pressure (SBP), heart rate (HR), and stroke volume (SV) in isometric handgrip (IHG) and mental arithmetic. Specifically, we asked whether 1) these cardiovascular stress responses were different for the two stress conditions in children, 2) these responses differed in boys and girls, and 3) the anthropometric variables related to these stress responses. SV differed significantly between IHG and mental arithmetic over the entire stress period. This may reflect higher systemic vascular resistance during IHG. HR in boys was lower than in girls over the entire period of stress in both stress tests. This observation cannot be attributed to differences in conditioning, because this should not influence responses to isometric or mental stress. A larger left ventricular mass was related to higher SVs. A marked relationship was found between HR and SBP and between HR and SV. No relationship was found between SBP and SV.

Univariate genetic analysis of oxygen transport regulation in children: the Medical College of Virginia Twin Study.

We investigated the relative contributions of genetic, individual environmental, and shared environmental effects on 2,3-diphosphoglycerate (DPG) regulation in preadolescent children. In a population of 165 early pubescent boy and girl twin pairs (11.4 y old), of whom 63 were passive smokers, we asked: 1) Are there differences in the control of DPG levels between early pubertal boys and girls? 2) If present, are these differences influenced by exposure to passive cigarette smoke? Non-passive-smoking boys and girls had similar DPG levels. With exposure to passive smoke, DPG levels increased in boys (p = 0.02) but not in girls. Analysis of variance on DPG demonstrated a parental smoking effect (p = 0.008) and suggested an interactive effect between parental smoking and sex of the child (p = 0.08). Univariate genetic analyses suggested that genes operated at different magnitudes in boys (9%) and girls (39%) in explaining a significant portion of the variance in DPG. The magnitude of shared environmental influences was greater in boys (62%) than in girls (34%), whereas individual environmental effects were similar in boys (29%) and girls (26%). Early pubertal boys differ from girls in their regulation of DPG. Environmental stressors such as passive cigarette smoke may elicit different responses in males and females, even at an early age. The use of path analysis may provide important insights into the mechanisms and interactions of genetic and environmental effects that underly the childhood antecedents of atherosclerotic heart disease.

A genetic analysis of cardiovascular disease risk factor clustering in adult female twins.

Risk factors for cardiovascular disease have been shown to cluster in adult populations of men and women [Criqui et al., 1980]. In a population of adult female twin pairs (ages 18-85), body mass index, systolic and diastolic blood pressure, and high and low density lipoprotein levels were found to exhibit significant clustering. Path analysis was used to resolve the risk factor correlations into genetic, environmental, and age-related components.

Multivariate genetic analysis of blood pressure and body size. The Medical College of Virginia Twin Study.

BACKGROUND: In subjects of all ages, those who weigh the most often have the highest blood pressure. Thus, in epidemiological studies, weight is the most important correlate of blood pressure. Using the data from the Medical College of Virginia Twin Study, we asked these questions: 1) Do the same genetic paths that regulate body size also regulate systolic and diastolic blood pressure? 2) Are there distinct genetic pathways that regulate each of these variables? 3) Does environment play a major regulatory role? 4) Are the correlations among these variables mainly due to genetic or environmental effects? 5) Do genetic paths that regulate body size mediate the correlation between systolic blood pressure and diastolic blood pressure? METHODS AND RESULTS: We ascertained 253 Caucasian twin pairs living in the Commonwealth of Virginia. The average age was 11.2 +/- 0.2 years. We used multivariate path analyses to investigate the genetic relations among systolic blood pressure, diastolic blood pressure, and body size. We found that there was a highly significant genetic relation between systolic blood pressure and body size and between systolic and diastolic blood pressure. There are genetic paths that are shared within these two sets of variables, but in each case, the paths for each pair appear to be separate from one another. CONCLUSIONS: These analyses provide a method to partition correlation coefficients found in epidemiological studies into genetic and environmental components. The correlations found among these three variables are in large part due to these genetic relations. We found no genetic relation between diastolic blood pressure and body size.

Sex differences in the determinants of left ventricular mass in childhood. The Medical College of Virginia Twin Study.

BACKGROUND: Left ventricular (LV) hypertrophy is a predictor of cardiovascular events in adults and has been observed in children and adolescents with hypertension. We wanted to establish the determinants of LV mass in normotensive preadolescent children. Our objectives were 1) to produce a simplified and generalizable model of the clinical variables that determine normal cardiac growth during childhood and 2) to understand better why males have an increased LV mass relative to females, even as children. METHODS AND RESULTS: In a group of 243 eleven-year-old children, we analyzed anthropometric, hemodynamic, and echocardiographic data to define which variables were predictors of echocardiographically determined LV mass. Stepwise regression was used to predict LV mass overall, by sex, and by body size (body mass index). Overall, LV mass was directly related to weight, male sex, and systolic and diastolic blood pressure and inversely related to resting heart rate and skin-fold thicknesses. Systolic blood pressure was a determinant in boys but not in girls. Heart rate was a weak inverse correlate in both sexes. When the data were analyzed by body mass index quartile, weight was the sole predictor of LV mass in the largest children. CONCLUSIONS: We conclude that in normotensive preadolescent children, 1) weight, but not pondersity, is a strong predictor of LV mass; 2) body fat is negatively associated with LV mass; 3) boys have an increased LV mass relative to girls; and 4) boys and girls have similar anthropometric determinants and may have different hemodynamic determinants. Our data suggest that body size, and in particular lean body mass, explains much of the variability in cardiac growth seen in children. The influence of hemodynamic variables seems to be more limited. Our findings are of general interest because, although hypertensive heart disease is well described, the early developmental stages are not well understood.

Bivariate genetic analysis of left ventricular mass and weight in pubertal twins (the Medical College of Virginia twin study).

Left ventricular (LV) hypertrophy in adults is a recognized risk factor for the subsequent development of cardiovascular morbidity. To make informed preventive health decisions it is important to understand the interaction of genes and environment on LV mass. In both children and adults, weight is a strong correlate of LV mass. We hypothesized that genetic influences common to both of these variables could in part explain the strong relation between weight and LV mass in children. In a population of 341 twins (11 years old), these questions were asked: (1) How much of the total variance of LV mass is under genetic control? (2) After accounting for weight and weight adjusted for sexual maturity, how much of the remaining variance is genetic? (3) Of the total genetic variance, what proportion is specific for LV mass and what proportion is common to both weight and LV mass? (4) How much of the correlation between these 2 variables is explained by genes common to both LV mass and weight? Univariate genetic analyses documented that genes operating at different magnitudes in boys (63%) and girls (71%) explain a significant proportion of the variance of LV mass. After removing the effect of weight and sexual maturity by regression methods, genes remain an important influence. Bivariate genetic analyses confirmed that genes common to LV mass and weight significantly influence the covariation of these variables and that greater than 90% of the correlation of LV mass and weight is due to common genes.

Genetic and environmental contributions to cholesterol and its subfractions in 11-year-old twins. The Medical College of Virginia Twin Study.

We conducted a cross-sectional analysis of the genetic and environmental contributions to the variance of lipoprotein cholesterol and its subfractions in children during early adolescence. Univariate path analysis was used to determine the relative contributions of genes, individual environment, and family environment to these measures in 233 11-year-old Caucasian twin pairs. For high density lipoprotein, high density lipoprotein2, low density lipoprotein, very low density lipoprotein, and triglycerides, a model that incorporated genes and individual environmental variation but not common environment was sufficient to explain the variation. Different magnitudes of genetic effects were seen for total cholesterol in boys and girls. High density lipoprotein3 showed different magnitudes by sex for genetic and individual environmental effect. Intermediate density lipoprotein was the only cholesterol subfraction in which shared, or common, environment was found to make a statistically significant contribution to the variation.

Genetic analysis of anthropometric measures in 11-year-old twins: the Medical College of Virginia Twin Study.

We have conducted a cross-sectional analysis of the genetic and environmental contributions to the variance of anthropometric measurements in children during early adolescence. Univariate path analysis was used to estimate the relative contributions of genes, individual environment, and family environment to measures of childhood obesity in 259 11-y-old Caucasian twin pairs. Triceps, subcapular, and suprailiac skinfold thicknesses, as well as waist circumferences, ht, and wt were measured in a standardized protocol. In this sample, a parsimonious model that included only additive genetic effects and environmental factors unique to the individual provided an adequate explanation for the variation in ht, wt, quetelet index, and subscapular and triceps skinfolds. In this largely preadolescent population, different magnitudes of genetic effects were seen in males and females for waist circumference, biiliac diameter, and suprailiac skinfold.

Altered systolic and diastolic function in children after "successful" repair of coarctation of the aorta.

We investigated whether left ventricular (LV) structural or functional abnormalities persist in children on long-term follow-up after successful correction of coarctation of the aorta. Two-dimensional directed M-mode and Doppler echocardiographic examinations were performed in 11 such subjects and 22 age-matched control subjects. Digitized tracings were made from M-mode recordings of the LV and Doppler mitral valve inflow recordings to measure septal, posterior wall, and LV dimensions, LV mass, shortening fraction, peak shortening and lengthening velocities, diastolic filling time, peak E velocity, peak A velocity, and velocity time integrals. Despite group similarities in age, body size, and systolic blood pressure, greater fractional shortening (p = 0.0001), indexed peak shortening velocity (p less than 0.001), and greater LV mass index (p less than 0.05) were seen in the coarctation group in the face of lower LV wall stress (p = 0.0001). LV mass index correlated with the resting arm-leg gradient, which ranged from -4 to +10 mm Hg. The coarctation group had decreased early filling (p less than 0.006) with compensatory increased late diastolic filling (p less than 0.05). Diastolic filling abnormalities were prominent in the older coarctation subjects and were related to both systolic blood pressure (p less than 0.001) and LV mass index (p less than 0.01). Despite apparently successful repair of coarctation of the aorta, persistent alterations in both systolic and diastolic LV function and LV mass are present in children at long-term follow-up, which are related to the resting arm-leg gradient.(ABSTRACT TRUNCATED AT 250 WORDS)

Lipoprotein and oxygen transport alterations in passive smoking preadolescent children. The MCV Twin Study.

We investigated the cardiovascular effects of lifelong passive cigarette smoke exposure in preadolescent children and examined the following questions: 1) Is systemic oxygen transport altered? 2) Are coronary heart disease risk factors adversely affected? We recruited 216 families from the MCV Twin Study; 105 had at least one smoking parent. Serum thiocyanate and cotinine levels were used as measures of smoke exposure in the children and thiocyanate was proportional to the number of parental cigarettes smoked each day (p = 0.0001). Paternal smoking had no effect on these measures. Whole blood 2,3-diphosphoglycerate was higher in smoke-exposed than unexposed children (p less than 0.01) and was related to the thiocyanate level (p less than 0.02). High density lipoprotein (HDL) cholesterol was lower in passive smoking children (p less than 0.05); the HDL2 subfraction was reduced in passive smoking boys, while the HDL3 subfraction was reduced in passive smoking girls. Significant adverse alterations in systemic oxygen transport and lipoprotein profiles are already present in preadolescent children exposed to long-term passive cigarette smoke, primarily from maternal smoke. Children with long-term exposure to passive smoke may be at elevated risk for the development of premature coronary heart disease.

Univariate genetic analysis of blood pressure in children (the Medical College of Virginia Twin Study).

The relative contributions of genetic, individual environmental and shared environmental effects on resting blood pressure (BP) and heart rate (HR) were studied in prepubescent twins. The study population consisted of 251 caucasian 11-year-old twin pairs. Correlations were higher for all variables in monozygotic twins compared to dizygotic twins; this is consistent with a significant genetic effect. Path analysis revealed that the model of additive genetic and individual environmental effects fit systolic BP, diastolic BP and HR. In boys and girls, sex-specific genetic effects controlled systolic BP. The magnitudes of the sex-specific genetic effects on systolic BP were similar in both boys and girls and accounted for 66% of the variance. In boys, for diastolic BP, genetic effects accounted for 64% of the variance while in girls they accounted for 51%. These results provide no evidence for different genetic effects on HR in boys or girls. No shared environmental effects were detected. The large sample size and design, using different-sex dizygotic twins of the same age, establish that genes play an important role in the influence of resting BP and HR and that there are sex-specific genetic contributions in early pubertal children.

Aortic stiffness: a new Doppler echocardiographic measure predictive of systolic blood pressure in children.

Aortic stiffness, the maximal frequency shift in the descending aorta divided by the Doppler acceleration time, was studied in 384 eleven year old twin children. The extent to which this measurement provided a prediction of systolic blood pressure that was independent of body size, heart rate, cardiac contractility and left ventricular mass was investigated. Aortic stiffness, after adjustment for height and weight, correlated significantly with systolic blood pressure (r = 0.22, p less than 0.01), but not with diastolic blood pressure. The short- (r = 0.82) and longer- (r = 0.68) term reproducibility of aortic stiffness was high. This measure appears to be a more powerful predictor of systolic blood pressure than is left ventricular mass. Aortic stiffness is a highly reproducible Doppler variable that may explain in part the contribution of the aortic wall elastic properties to the level of systolic blood pressure in preadolescent children at rest.

Corneal pseudocyst following acute hydrops.

A 20-year-woman with bilateral keratoconus developed acute hydrops of her right cornea. The amount of corneal oedema progressively increased over eight weeks while the central cornea thinned. Histologically the cornea contained a stromal pseudocyst that was in continuity with the anterior chamber through breaks in Descemet's membrane. Pseudocyst formation is a rare complication of acute corneal hydrops that can simulate severe corneal ectasia.

Denominator definition by the utilization correction factor method.

Most clinical research in the United States has been carried out in atypical populations. This study, done in a network of primary care populations in Virginia, calculates the denominator by the utilization correction factor method and compares the demography with that of the population of the state. The demographic characteristics of the patients in the network are very similar to those of the underlying populations and on the same order of state-of-the-art sampling methods in current use.