Analytical validity of a genotyping assay for use with personalized antihypertensive and chronic kidney disease therapy.

Hypertension and chronic kidney disease are inextricably linked. Hypertension is a well-recognized contributor to chronic kidney disease progression and, in turn, renal disease potentiates hypertension. A generalized approach to drug selection and dosage has not proven effective in managing these conditions, in part, because patients with heterogeneous kidney disease and hypertension etiologies are frequently grouped according to functional or severity classifications. Genetic testing may serve as an important tool in the armamentarium of clinicians who embrace precision medicine. Increasing scientific evidence has supported the utilization of genomic information to select efficacious antihypertensive therapy and understand hereditary contributors to chronic kidney disease progression. Given the wide array of antihypertensive agents available and diversity of genetic renal disease predictors, a panel-based approach to genotyping may be an efficient and economic means of establishing an individualized blood pressure response profile for patients with various forms of chronic kidney disease and hypertension. In this manuscript, we discuss the validation process of a Clinical Laboratory Improvement Amendments-approved genetic test to relay information on 72 genetic variants associated with kidney disease progression and hypertension therapy. These genomic-based interventions, in addition to routine clinical data, may help inform physicians to provide personalized therapy.

Case report: Severe central nervous system manifestations associated with aberrant efavirenz metabolism in children: the role of CYP2B6 genetic variation.

BACKGROUND: Efavirenz, widely used as part of antiretroviral drug regimens in the treatment of paediatric human immunodeficiency virus infection, has central nervous system side effects. We describe four children presenting with serious, persistent central nervous system adverse events who were found to have elevated plasma efavirenz concentrations as a result of carrying CYP2B6 single nucleotide polymorphisms, known to play a role in the metabolism of EFV. None of the children had a CYP2B6 wildtype haplotype. We believe this is the first case of cerebellar dysfunction associated with efavirenz use to be described in children. CASE PRESENTATION: Four black African children, between the ages of 4 and 8 years presenting between 1 and 20 months post-efavirenz initiation, are described. Cerebellar dysfunction, generalised seizures and absence seizures were the range of presenting abnormalities. Plasma efavirenz levels ranged from 20-60 mg/L, 5-15 times the upper limit of the suggested reference range. All abnormal central nervous system manifestations abated after efavirenz discontinuation. CONCLUSION: Efavirenz toxicity should always be considered in human immunodeficiency virus-infected children with unexplained central nervous system abnormalities. Our findings further our understanding of the impact of genetic variants on antiretroviral pharmacokinetics in children across various ethnic groups. Screening for potential EFV-toxicity based on the CYP2B6 c.516 SNP alone, may not be adequate.

High predictive value of CYP2B6 SNPs for steady-state plasma efavirenz levels in South African HIV/AIDS patients.

INTRODUCTION: Efavirenz is primarily metabolized by CYP2B6, with a minor contribution from CYP1A2, CYP2A6, CYP3A4 and CYP3A5. Genetic variability in these genes contributes towards differences in plasma efavirenz concentration, which ultimately leads to either development of adverse drug events or emergence of virus resistance. However, the clinical utility or validity of introducing genotype-assisted dosing is not known. The aim of this study was therefore to evaluate the effects of 14 single-nucleotide polymorphisms (SNPs) in five drug-metabolizing enzyme genes on steady-state plasma efavirenz levels in South African HIV/AIDS patients as well as their clinical validity. METHODS: HIV/AIDS patients were recruited from Themba Lethu Clinic, at Helen Joseph Hospital, Johannesburg. Blood samples for plasma drug levels and DNA extraction were obtained from each participant. PCR/RFLP and SNaPshot genotyping were used for SNPs in CYP1A2, CYP2A6, CYP2B6, CYP3A4 and CYP3A5 among 464 Bantu-speaking South Africans. Plasma efavirenz concentrations were measured using LC/MS/MS. Genotypes and plasma efavirenz levels were used to calculate predictive values. Multivariate analysis was used to select the minimal set of SNPs with significant clinical validity. RESULTS: Qualitative and quantitative differences in allele frequencies were observed when comparing South Africans with African, Caucasian and Asian populations. CYP2B6 516T and 785G (*6) and CYP2B6 983C (*18) alleles were significantly associated with high plasma efavirenz levels. CYP2B6 A-G-A-C-C and A-T-G-T-C haplotypes (with respect to CYP2B6 136A>G; CYP2B6 516G>T; CYP2B6 785A>G; CYP2B6 983T>C; and CYP2B6 1459C>T) were associated with higher levels of efavirenz, whereas G-G-A-T-C and A-G-A-T-C haplotypes showed significantly lower levels of efavirenz. The CYP2B6*1/*6 genotype was significantly associated with an increased risk of loss to follow-up. The sensitivity, specificity and positive predictive values for the CYP2B6*6/*6 genotype in predicting efavirenz levels above 4 µg/ml were 46, 97 and 88%, respectively. However, these values improved to 49, 100 and 100%, respectively, when either the CYP1A2 -163A (*1F) allele or the NR1I3 8784C/C genotype was present. CONCLUSION: Screening for CYP2B6 516G>T SNP has a high specificity and positive predictive value for efavirenz levels above 4 µg/ml and could be used in deciding on efavirenz dosage among individuals homozygous for this variant, which could lead to better precision medication.

PXR and CAR single nucleotide polymorphisms influence plasma efavirenz levels in South African HIV/AIDS patients.

BACKGROUND: This study investigated variation in NR1I2 and NR1I3 and its effect on plasma efavirenz levels in HIV/AIDS patients. Variability in plasma drug levels has largely led research on identifying causative variants in drug metabolising enzyme (DME) genes, with little focus on the nuclear receptor genes NR1I2 and NR1I3, coding for PXR and CAR, respectively, that are involved in regulating DMEs. METHODS: 464 Bantu-speaking South Africans comprising of HIV/AIDS patients on efavirenz-based treatment (n=301) and 163 healthy subjects were genotyped for 6 SNPs in NR1I2 and NR1I3. 32 of the 301 patients had their DNA binding domains (DBDs) in NR1I2 and NR1I3 sequenced. RESULTS: Significantly decreased efavirenz plasma concentrations were observed in patients carrying the NR1I3 rs3003596C/C and T/C genotypes (P=0.015 and P=0.010, respectively). Sequencing resulted in the discovery of a further 13 SNPs, 3 of which are novel variants in the DBD of NR1I2. There were significant differences in the distribution of NR1I2 and NR1I3 SNPs between South Africans when compared to Caucasian, Asian and Yoruba population groups. CONCLUSION: For the realisation of personalised medicine, PXR and CAR genetic variation should be taken into consideration because of their involvement in the regulation of DMEs.

MicroRNA Mediated Changes in Drug Metabolism and Target Gene Expression by Efavirenz and Rifampicin In Vitro: Clinical Implications.

Efavirenz (EFV) and rifampicin (RMP) are widely prescribed in Africa for treatment of HIV/AIDS and tuberculosis epidemics. Exposure to medicines can alter drug metabolism, for example, through changes in expression of microRNAs. We report, in this study, novel observations on the ways in which EFV and RMP change microRNA expression signatures in vitro in HepaRG cells. Additionally, we discuss the clinical implications of changes in expression of drug-metabolizing enzyme genes, such as CYP3A4, CYP3A5, UGT1A1, CYP2B6, and NR1I3. Differentiated HepaRG cells were treated with EFV (6.4 µM) or RMP (24.4 µM) for 24 h. Treatment of HepaRG cells with EFV resulted in a significant increase in messenger RNA (mRNA) expression for CYP3A4 (12.51-fold, p = 0.002), CYP3A5 (2.10-fold, p = 0.019), and UGT1A1 (2.52-fold, p = 0.005), whereas NR1I3 expression decreased (0.41-fold, p = 0.02). On the other hand, treatment of HepaRG cells with RMP resulted in a significant increase in mRNA expression for CYP2B6 (6.68-fold, p = 0.007) and CYP3A4 (111.96-fold, p = 0.001), whereas NR1I3 expression decreased (0.46-fold, p = 0.033). These data point to several important clinical implications through changes in drug/drug interaction risks and achieving optimal therapeutics. All in all, this study shows that differential expression of microRNAs after treatment with EFV and RMP adds another layer of complexity that should be incorporated in pharmacogenomic algorithms to render drug response more predictable.

Analytical Validation of Variants to Aid in Genotype-Guided Therapy for Oncology.

The Clinical Laboratory Improvement Amendments of 1988 require that pharmacogenetic genotyping methods need to be established according to technical standards and laboratory practice guidelines before testing can be offered to patients. Testing methods for variants in ABCB1, CBR3, COMT, CYP3A7, C8ORF34, FCGR2A, FCGR3A, HAS3, NT5C2, NUDT15, SBF2, SEMA3C, SLC16A5, SLC28A3, SOD2, TLR4, and TPMT were validated in a Clinical Laboratory Improvement Amendments-accredited laboratory. Because no known reference materials were available, existing DNA samples were used for the analytical validation studies. Pharmacogenetic testing methods developed here were shown to be accurate and 100% analytically sensitive and specific. Other Clinical Laboratory Improvement Amendments-accredited laboratories interested in offering pharmacogenetic testing for these genetic variants, related to genotype-guided therapy for oncology, could use these publicly available samples as reference materials when developing and validating new genetic tests or refining current assays.

Clinical Pharmacogenetics Implementation Consortium (CPIC) Guideline for CYP2B6 and Efavirenz-Containing Antiretroviral Therapy.

The HIV type-1 nonnucleoside reverse transcriptase inhibitor, efavirenz, is widely used to treat HIV type-1 infection. Efavirenz is predominantly metabolized into inactive metabolites by cytochrome P450 (CYP)2B6, and patients with certain CYP2B6 genetic variants may be at increased risk for adverse effects, particularly central nervous system toxicity and treatment discontinuation. We summarize the evidence from the literature and provide therapeutic recommendations for efavirenz prescribing based on CYP2B6 genotypes.

Pharmacogenetics of Antiretroviral Drug Response and Pharmacokinetic Variations in Indigenous South African Populations.

Interindividual and interethnic differences in response to antiretroviral drugs (ARVs) are influenced by genetic variation. The few genomic studies conducted among African-Americans and African ethnic groups do not reflect the extensive genetic diversity within African populations. ARVs are widely used in Africa. Therefore, genomic characterization of African populations is required before genotype-guided dosing becomes possible. The aim of this study was to determine and report on the frequency of genetic variants in genes implicated in metabolism and transport of ARVs in South African populations. The study comprised 48 self-reported South African Colored (SAC) and 296 self-reported Black African (BA) individuals. Allele and genotype frequency distributions for 93 variants contributing to metabolism and transport of ARVs were compared between groups, and other global populations. Fifty-three variants had significant differences in allele and genotype frequencies when comparing SAC and BA groups. Thirteen of these have strong clinical annotations, affecting efavirenz and tenofovir pharmacokinetics. This study provides a summary of the genetic variation within genes implicated in metabolism and transport of ARVs in indigenous South African populations. The observed differences between indigenous population groups, and between these groups and global populations, demonstrate that data generated from specific African populations cannot be used to infer genetic diversity within other populations on the continent. These results highlight the need for comprehensive characterization of genetic variation within indigenous African populations, and the clinical utility of these variants in ARV dosing for global precision medicine. Population pharmacogenetics is a nascent field of global health and warrants further research and education.

Report of Confirmation of the rs7853758 and rs885004 Haplotype in SLC28A3.

AIMS: To validate a laboratory-developed test for the nucleoside transporter, SLC28A3, which has been associated with an increased risk of anthracycline-induced cardiomyopathy. METHODS: We used Taqman® allele discrimination to test for two variants of the SLC28A3 gene: rs7853758 (c.1381C>T) and rs885004 (c.862-360C>T). RESULTS: During the validation process, we noted that several DNA samples obtained from the Coriell Cell Repository (Camden, NJ) were positive for both the c.1381 C > T and c.862-360C>T variants and another variant allele for either c.1381 C > T or c.862-360C>T (e.g., c.1381C>T homozygous/c.862-360C>T heterozygous, c.1381C>T homozygous/c.862-360C>T homozygous). We used de-identified DNA samples from trios of family members (mother, father, and child) to establish that the c.1381 C > T and c.862-360C>T variant alleles could be inherited in cis on the same chromosome. CONCLUSIONS: Samples containing three variant alleles suggest that the c.1381 C > T and c.862-360C>T are in cis on the chromosome in some individuals and may have implications when calculating anthracycline-induced cardiomyopathy risk. In this study, we confirm a novel haplotype of SLC28A3 using familial studies.

Variants in the CYP2B6 3'UTR Alter In Vitro and In Vivo CYP2B6 Activity: Potential Role of MicroRNAs.

CYP2B6*6 and CYP2B6*18 are the most clinically important variants causing reduced CYP2B6 protein expression and activity. However, these variants do not account for all variability in CYP2B6 activity. Emerging evidence has shown that genetic variants in the 3'UTR may explain variable drug response by altering microRNA regulation. Five 3'UTR variants were associated with significantly altered efavirenz AUC0-48 (8-OH-EFV/EFV) ratios in healthy human volunteers. The rs70950385 (AG>CA) variant, predicted to create a microRNA binding site for miR-1275, was associated with a 33% decreased CYP2B6 activity among normal metabolizers (AG/AG vs. CA/CA (P < 0.05)). In vitro luciferase assays were used to confirm that the CA on the variant allele created a microRNA binding site causing an 11.3% decrease in activity compared to the AG allele when treated with miR-1275 (P = 0.0035). Our results show that a 3'UTR variant contributes to variability in CYP2B6 activity.

A Global Health Diagnostic for Personalized Medicine in Resource-Constrained World Settings: A Simple PCR-RFLP Method for Genotyping CYP2B6 g.15582C>T and Science and Policy Relevance for Optimal Use of Antiretroviral Drug Efavirenz.

The use of pharmacogenomics (PGx) knowledge in treatment of individual patients is becoming a common phenomenon in the developed world. However, poorly resourced countries have thus far been constrained for three main reasons. First, the cost of whole genome sequencing is still considerably high in comparison to other (non-genomics) diagnostics in the developing world where both science and social dynamics create a dynamic and fragile healthcare ecosystem. Second, studies correlating genomic differences with drug pharmacokinetics and pharmacodynamics have not been consistent, and more importantly, often not indexed to impact on societal end-points, beyond clinical practice. Third, ethics regulatory frames over PGx testing require improvements based on nested accountability systems and in ways that address the user community needs. Thus, CYP2B6 is a crucial enzyme in the metabolism of antiretroviral drugs, efavirenz and nevirapine. More than 40 genetic variants have been reported, but only a few contribute to differences in plasma EFV and NVP concentrations. The most widely reported CYP2B6 variants affecting plasma drug levels include c.516G>T, c.983T>C, and to a lesser extent, g.15582C>T, which should be considered in future PGx tests. While the first two variants are easily characterized, the g.15582C>T detection has been performed primarily by sequencing, which is costly, labor intensive, and requires access to barely available expertise in the developing world. We report here on a simple, practical PCR-RFLP method with vast potentials for use in resource-constrained world regions to detect the g.15582C>T variation among South African and Cameroonian persons. The effects of CYP2B6 g.15582C>T on plasma EFV concentration were further evaluated among HIV/AIDS patients. We report no differences in the frequency of the g.15582T variant between the South African (0.08) and Cameroonian (0.06) groups, which are significantly lower than reported in Asians (0.39) and Caucasians (0.31). The g.15582C/T and T/T genotypes were associated with significantly reduced EFV levels (p=0.006). This article additionally presents the policy relevance of the PGX global health diagnostics and therefore, collectively makes an original interdisciplinary contribution to the field of integrative biology and personalized medicine in developing world. Such studies are, in fact, broadly important because resource-constrained regions exist not only in developing world but also in major geographical parts of the G20 nations and the developed countries.

An Expanded Analysis of Pharmacogenetics Determinants of Efavirenz Response that Includes 3'-UTR Single Nucleotide Polymorphisms among Black South African HIV/AIDS Patients.

INTRODUCTION: Efavirenz (EFV) is a non-nucleoside reverse transcriptase inhibitor prescribed as part of first-line highly active antiretroviral therapy (HAART) in South Africa. Despite administration of fixed doses of EFV, inter-individual variability in plasma concentrations has been reported. Poor treatment outcomes such as development of adverse drug reactions or treatment failure have been linked to EFV plasma concentrations outside the therapeutic range (1-4 µg/mL) in some studies. The drug metabolizing enzyme (DME), CYP2B6, is primarily responsible for EFV metabolism with minor contributions by CYP1A2, CYP2A6, CYP3A4, CYP3A5, and UGT2B7. DME coding genes are also regulated by microRNAs through targeting the 3'-untranslated region. Expanded analysis of 30 single nucleotide polymorphisms (SNPs), including those in the 3'-UTR, was performed to identify pharmacogenetics determinants of EFV plasma concentrations in addition to CYP2B6 c.516G>T and c.983T>C SNPs. METHODS: SNPs in CYP1A2, CYP2B6, UGT2B7, and NR1I2 (PXR) were selected for genotyping among 222 Bantu-speaking South African HIV-infected patients receiving EFV-containing HAART. This study is a continuation of earlier pharmacogenetics studies emphasizing the role of genetic variation in the 3'-UTR of genes which products are either pharmacokinetic or pharmacodynamic targets of EFV. RESULTS: Despite evaluating thirty SNPs, CYP2B6 c.516G>T and c.983T>C SNPs remain the most prominent predictors of EFV plasma concentration. CONCLUSION: We have shown that CYP2B6 c.516G>T and c.983T>C SNPs are the most important predictors of EFV plasma concentration after taking into account all other SNPs, including genetic variation in the 3'-UTR, and variables affecting EFV metabolism.

Effects of CYP2B6 and CYP1A2 Genetic Variation on Nevirapine Plasma Concentration and Pharmacodynamics as Measured by CD4 Cell Count in Zimbabwean HIV-Infected Patients.

The extremely high prevalence of HIV/AIDS in sub-Saharan Africa and limitations of current antiretroviral medicines demand new tools to optimize therapy such as pharmacogenomics for person-to-person variations. African populations exhibit greater genetic diversity than other world populations, thus making it difficult to extrapolate findings from one population to another. Nevirapine, an antiretroviral medicine, displays large plasma concentration variability which adversely impacts therapeutic virological response. This study, therefore, aimed to identify sources of variability in nevirapine pharmacokinetics and pharmacodynamics, focusing on genetic variation in CYP2B6 and CYP1A2. Using a cross-sectional study design, 118 HIV-infected adult Zimbabwean patients on nevirapine-containing highly active antiretroviral therapy (HAART) were characterized for three key functional single nucleotide polymorphisms (SNPs), CYP2B6 c.516G>T (rs3745274), CYP2B6 c.983T>C (rs28399499), and CYP1A2 g.-163C>A (rs762551). We investigated whether genotypes at these loci were associated with nevirapine plasma concentration, a therapeutic biomarker, and CD4 cell count, a biomarker of disease progression. CYP2B6 and CYP1A2 were chosen as the candidate genes based on reports in literature, as well as their prominence in the metabolism of efavirenz, a drug in the same class with nevirapine. Nevirapine plasma concentration was determined using LC-MS/MS. The mean nevirapine concentration for CYP2B6 c.516T/T genotype differed significantly from that of 516G/G (p < 0.001) and 516G/T (p < 0.01) genotypes, respectively. There were also significant differences in mean nevirapine concentration between CYP2B6 c.983T > C genotypes (p = 0.04). Importantly, the CYP1A2 g.-163C>A SNP was significantly associated with the pharmacodynamics endpoint, the CD4 cell count (p = 0.012). Variant allele frequencies for the three SNPs observed in this Zimbabwean group were similar to other African population groups but different to observations among Caucasian and Asian populations. We conclude that CYP2B6 c.516G>T and CYP2B6 c.983T>C could be important sources of nevirapine pharmacokinetic variability that could be considered for dosage optimization, while CYP1A2 g.-163C>A seems to be associated with HIV disease progression. These inter- and intra-population pharmacokinetic and pharmacodynamics differences suggest that a single prescribed dosage may not be appropriate for the treatment of disease. Further research into a personalized nevirapine regimen is required.

Genetic variation in the 3'-UTR of CYP1A2, CYP2B6, CYP2D6, CYP3A4, NR1I2, and UGT2B7: potential effects on regulation by microRNA and pharmacogenomics relevance.

INTRODUCTION: Pharmacogenomics research has concentrated on variation in genes coding for drug metabolizing enzymes, transporters and nuclear receptors. However, variation affecting microRNA could also play a role in drug response. This project set out to investigate potential microRNA target sites in 11 genes and the extent of variation in the 3'-UTR of six selected genes; CYP1A2, CYP2B6, CYP2D6, CYP3A4, NR1I2, and UGT2B7. METHODS: Fifteen microRNA target prediction algorithms were used to identify microRNAs predicted to regulate 11 genes. The 3'-UTR of the 6 genes which topped the list of potential microRNA targets was sequenced in 30 black South Africans. In addition, genetic variants within these genes were investigated for interference with mRNA-microRNA interactions. Potential effects of observed variants were determined using in silico prediction tools. RESULTS: The 11 genes coding for DMEs, transporters and nuclear receptors were predicted to be targets of microRNAs with CYP2B6, NR1I2 (PXR), CYP3A4, and CYP1A2, interacting with the most microRNAs. The majority of identified genetic variants were predicted to interfere with microRNA regulation. For example, the variant, rs1054190C in NR1I2 was predicted to result in the presence of a binding site for the microRNA miR-1250-5p, while the variant rs1054191G was predicted to result in the absence of a recognition site for miR-371b-3p, miR-4258 and miR-4707-3p. Fifteen of the seventeen, novel variants occurred within microRNA target sequences. CONCLUSION: The 3'-UTR harbors variation that is likely to influence regulation of specific genes by microRNA. In silico prediction followed by functional validation could aid in decoding the contribution of variation in the 3'-UTR, to some unexplained heritability that affects drug response. Understanding the specific role of each microRNA may lead to identification of markers for targeted therapy and therefore improve personalized drug treatment.

The role of genetic polymorphisms in cytochrome P450 and effects of tuberculosis co-treatment on the predictive value of CYP2B6 SNPs and on efavirenz plasma levels in adult HIV patients.

Efavirenz (EFV) exhibits interindividual pharmacokinetic variability caused by differences in cytochrome P450 (CYP) expression. Most tuberculosis (TB) drugs interact with the CYP metabolizing enzymes, while the clinical validity of genotyping in predicting EFV plasma levels in Rwandan subjects is not known. We investigated in patients co-infected with human immunodeficiency virus (HIV) and TB recruited in Rwanda the effects of 10 SNPs in five drug-metabolizing enzymes on EFV plasma levels and treatment response when patients are treated with EFV-containing therapy alone (n=28) and when combined with rifampicin-based TB treatment (n=62), and the validity of genotyping for CYP2B6 single nucleotide polymorphisms in predicting supra-therapeutic EFV levels. There was a significant difference between CYP1A2 -739T/G and T/T genotypes when patients were treated with EFV-containing therapy combined with rifampicin-based TB treatment, but not when EFV-containing therapy was alone. CYP2B6 516T/T genotype was associated with high EFV levels compared to other CYP2B6 516G>T genotypes in the presence and in the absence of rifampicin-based TB treatment. Predictive factors of EFV plasma levels in the presence of rifampicin-based TB treatment were CYP2A6 1093G>A, CYP2B6 516G>T, and CYP2B6 983T>C accounting for 27%, 43%, and 29% of the total variance in EFV levels, respectively. There was a high positive predictive value (PPV) (100%) for CYP2B6 516T/T and 983T/T genotypes in predicting EFV plasma levels above the therapeutic range, but this PPV decreased in the presence of rifampicin-based TB treatment. Rifampicin-based TB treatment was also shown to affect EFV plasma levels significantly, but did not affect the significant reduction of HIV-RNA copies. These results indicate that genotyping for CYP2B6 SNPs could be used as a tool in predicting supra-therapeutic EFV plasma levels, which could minimize adverse drug events.

Cytochrome P450 pharmacogenetics in African populations: implications for public health.

INTRODUCTION: Africa harbors a disproportionate burden of disease when taking into account the triple challenge caused by HIV/AIDS, tuberculosis (TB) and malaria, against a backdrop of an increasing burden of noncommunicable diseases. More than 80% of therapeutic drugs used in the management of these diseases/conditions are metabolized by CYP enzymes that exhibit genetic polymorphisms. AREAS COVERED: There is variability in the expression and activities of CYPs resulting in interindividual differences in the response to standard doses of therapeutic drugs, due to genetic polymorphisms, which exhibit both quantitative and qualitative differences between racial and between ethnic groups. The review aims to evaluate the implications of the genetic variation in CYPs on the public health of Africans. The CYPs reviewed here metabolize most of the commonly used therapeutic drugs and include CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 3A4 and 3A5. Allele frequencies are compared between African ethnic groups and among populations of African, Asian and European origin. Data are obtained from our own studies and literature. EXPERT OPINION: The variability in the pattern of genetic variation between populations translates into differences in drug response. Understanding CYP variability improves rational drug use and has public health significance.

9ß Polymorphism of the glucocorticoid receptor gene appears to have limited impact in patients with Addison's disease.

BACKGROUND: Addison's disease (AD) has been associated with an increased risk of cardiovascular disease. Glucocorticoid receptor polymorphisms that alter glucocorticoid sensitivity may influence metabolic and cardiovascular risk factors in patients with AD. The 9ß polymorphism of the glucocorticoid receptor gene is associated with relative glucocorticoid resistance and has been reported to increase the risk of myocardial infarction in the elderly. We explored the impact of this polymorphism in patients with AD. MATERIALS AND METHODS: 147 patients with AD and 147 age, gender and ethnicity matched healthy controls were recruited. Blood was taken in a non-fasted state for plasma lipid determination, measurement of cardiovascular risk factors and DNA extraction. RESULTS: Genotype data for the 9ß polymorphism was available for 139 patients and 146 controls. AD patients had a more atherogenic lipid profile characterized by an increase in the prevalence of small dense LDL (p = 0.003), increased triglycerides (p = 0.002), reduced HDLC (p<0.001) an elevated highly sensitive C-reactive protein (p = 0.01), compared with controls. The 9ß polymorphism (at least one G allele) was found in 28% of patients and controls respectively. After adjusting for age, gender, ethnicity, BMI and hydrocortisone dose per metre square of body surface area in patients, there were no significant metabolic associations with this polymorphism and hydrocortisone doses were not higher in patients with the polymorphism. CONCLUSIONS: This study did not identify any associations between the 9ß polymorphism and cardiovascular risk factors or hydrocortisone dose and determination of this polymorphism is therefore unlikely to be of clinical benefit in the management of patients with AD.

How does mother-to-child transmission of HIV differ among African populations? Lessons from MBL2 genetic variation in Zimbabweans.

Mannose binding lectin (MBL) is a pathogen pattern recognition protein involved in antimicrobial activities. Variation in MBL2 gene has been extensively implicated in differential outcomes of infectious diseases in studies conducted outside Africa, but virtually very little is known on the role of this candidate gene in the African continent. We investigated human genetic variations in MBL2 in a Zimbabwean pediatric population and their putative associations with HIV infection in perinatally exposed children. One hundred and four children aged 7 to 9 years comprising 68 perinatally exposed to HIV (32 who were born infected and 36 who were uninfected) and 36 unexposed controls were recruited. DNA samples were genotyped for MBL2 polymorphisms using PCR-RFLP and sequencing. HIV infected children had markedly variable and significantly lower mean height (p=0.03) and weight (p=0.005) when compared to the uninfected children. Using all samples, frequencies for MBL2 genetic variants for the Zimbabwean population were calculated. Twelve single nucleotide polymorphisms were observed and minor alleles occurred with the following frequencies: -550C>G (G: 0.02), -435G>A (A: 0.08), -428A>C (C: 0.39), -394A>G (A: 0.39), -328AGAGAA ins/del (AGAGAA ins: 0.44), -245G>A (A: 0.05), -221C>G (C: 0.12), -111A>T (T: 0.10), -70C>T (C: 0.46), +4C>T (C: 0.45), novel -595G>A (A: 0.02), and 170G>A (0.24). We found that the MBL2 +4T variant displayed a trend for association with reduced risk of HIV transmission from mother-to-child but the remaining vast majority of the genetic markers did not show a significant association. We conclude (1) the MBL2 gene is highly polymorphic in the Zimbabwean population, and (2) MBL2 genetic variation does not appear to play a major role in influencing the risk of mother-to-child HIV transmission in our study sample. These observations contest the hitherto significant role of this candidate gene for HIV transmission from mother-to-child in non-African populations and thus, further speak to the limits of extrapolating genomic association studies directly to the African populations from studies conducted elsewhere. It is hoped that more OMICS research in a diverse set of African countries can shed further light on the putative role (or the lack thereof ) of this candidate gene in HIV transmission in the continent, a major global health burden in Africa.

ABCB1 4036A>G and 1236C>T Polymorphisms Affect Plasma Efavirenz Levels in South African HIV/AIDS Patients.

The ABCB1 gene encodes P-glycoprotein, an ATP-dependent drug efflux pump, which is responsible for drug transport across extra- and intra-cellular membranes. The variability in the expression of ABCB1 may contribute to variable plasma efavirenz concentration which results in variability in the levels of suppression of the human immunodeficiency syndrome virus (HIV). The aim of the study was to evaluate the role of polymorphisms in ABCB1 gene on plasma efavirenz levels and treatment response in the form of change in viral load and CD-4 cell count in HIV/AIDS patients receiving efavirenz-containing highly active antiretroviral treatment regimens. Two hundred and eighty-two HIV-infected patients were recruited from Themba Lethu Clinic in Johannesburg and plasma efavirenz drug concentration levels were measured using LC-MS/MS. SNaPshot was used to genotype five known ABCB1 single nucleotide polymorphisms (SNPs). Genotype-phenotype correlations were computed. The ABCB1 4036A/G and 4036G/G genotypes were significantly associated with low plasma efavirenz concentrations (P = 0.0236), while the ABCB1 1236C/T and 1236T/T genotypes were associated with high efavirenz concentrations (P = 0.0282). A haplotype ABCB1 T-G-T-A is reported that is associated with significantly increased plasma efavirenz levels. This is the first report on 61A>G, 2677G>T/A, and 4036A>G SNPs in the South African population. ABCB1 plays a role in determining the plasma concentrations of efavirenz and should be taken into account in future design of assays for genotype-based dosing of efavirenz-containing regimens.