<i>APOL1</i> risk alleles among individuals with CKD in Northern Tanzania: A pilot study

<div><p>Introduction</p><p>In sub-Saharan Africa, approximately 100 million people have CKD, yet genetic risk factors are not well-understood. Despite the potential importance of understanding <i>APOL1</i> risk allele status among individuals with CKD, little genetic research has been conducted. Therefore, we conducted a pilot study evaluating the feasibility of and willingness to participate in genetic research on kidney disease, and we estimated <i>APOL1</i> risk allele frequencies among individuals with CKD.</p><p>Methods</p><p>In 2014, we conducted a community-based field study evaluating CKD epidemiology in northern Tanzania. We assessed for CKD using urine albumin and serum creatinine to estimate GFR. We invited participants with CKD to enroll in an additional genetic study. We obtained dried-blood spots on filter cards, from which we extracted DNA using sterile punch biopsies. We genotyped for two single nucleotide polymorphisms (SNPs) defining the <i>APOL1</i> G1 risk allele and an insertion/deletion polymorphism defining the G2 risk allele. Genotyping was performed in duplicate.</p><p>Results</p><p>We enrolled 481 participant, 57 (12%) of whom had CKD. Among these, enrollment for genotyping was high (n = 48; 84%). We extracted a median of 19.4 ng of DNA from each dried-blood spot sample, and we genotyped the two <i>APOL1</i> G1 SNPs and the <i>APOL1</i> G2 polymorphism. Genotyping quality was high, with all duplicated samples showing perfect concordance. The frequency of <i>APOL1</i> risk variants ranged from 7.0% to 11.0%, which was similar to previously-reported frequencies from the general population of northern Tanzania (p>0.2).</p><p>Discussion</p><p>In individuals with CKD from northern Tanzania, we demonstrated feasibility of genotyping <i>APOL1</i> risk alleles. We successfully genotyped three risk variants from DNA extracted from filter cards, and we demonstrated a high enrollment for participation. In this population, more extensive genetic studies of kidney disease may be well-received and will be feasible.</p></div

Frequency of <i>APOL1</i> G1 and <i>APOL1</i> G2 risk variants for CKD-AFRiKA study sample and other populations of interest.

<p>Frequency of <i>APOL1</i> G1 and <i>APOL1</i> G2 risk variants for CKD-AFRiKA study sample and other populations of interest.</p

Reported frequencies of <i>APOL1</i> G1 and G2 risk alleles for several Bantu populations across sub-Saharan Africa.

<p>Reported frequencies of <i>APOL1</i> G1 and G2 risk alleles for several Bantu populations across sub-Saharan Africa.</p

Characteristics of participants with CKD, by genotyping status; n = 57.

<p>Characteristics of participants with CKD, by genotyping status; n = 57.</p

Feasibility of a bilateral 4000–6000 Hz notch as a phenotype for genetic association analysis

<div><p></p><p><i>Objective:</i> Noise-induced hearing loss (NIHL) is a worldwide health problem and a growing concern among young people. Although some people appear to be more susceptible to NIHL, genetic association studies lack a specific phenotype. We tested the feasibility of a bilateral 4000–6000 Hz audiometric notch as a phenotype for identifying genetic contributions to hearing loss in young adults. <i>Design:</i> A case-control-control study was conducted to examine selected SNPs in 52 genes previously associated with hearing loss and/or expressed in the cochlea. A notch was defined as a minimum of a 15-dB drop at 4000–6000 Hz from the previous best threshold with a 5-dB ‘recovery’ at 8000 Hz. <i>Study sample:</i> Participants were 252 individuals of European descent taken from a population of 640 young adults who are students of classical music. Participants were grouped as No-notch (NN), Unilateral Notch (UN), or Bilateral Notch (BN). <i>Results:</i> The strongest evidence of a genetic association with the 4000–6000 Hz notch was a nonsynonymous SNP variant in the ESRR– gene (rs61742642:C> T, P386S). Carriers of the minor allele accounted for 26% of all bilateral losses. <i>Conclusion:</i> This study indicates that the 4000–6000 Hz bilateral notch is a feasible phenotype for identifying genetic susceptibility to hearing loss.</p></div

Case-Only Survival Analysis Reveals Unique Effects of Genotype, Sex, and Coronary Disease Severity on Survivorship

<div><p>Survival bias may unduly impact genetic association with complex diseases; gene-specific survival effects may further complicate such investigations. Coronary artery disease (CAD) is a complex phenotype for which little is understood about gene-specific survival effects; yet, such information can offer insight into refining genetic associations, improving replications, and can provide candidate genes for both mortality risk and improved survivorship in CAD. Building on our previous work, the purpose of this current study was to: evaluate <i>LSAMP</i> SNP-specific hazards for all-cause mortality post-catheterization in a larger cohort of our CAD cases; and, perform additional replication in an independent dataset. We examined two <i>LSAMP</i> SNPs—rs1462845 and rs6788787—using CAD case-only Cox proportional hazards regression for additive genetic effects, censored on time-to-all-cause mortality or last follow-up among Caucasian subjects from the Catheterization Genetics Study (CATHGEN; <i>n</i> = 2,224) and the Intermountain Heart Collaborative Study (IMHC; <i>n</i> = 3,008). Only after controlling for age, sex, body mass index, histories of smoking, type 2 diabetes, hyperlipidemia and hypertension (<i>HR</i> = 1.11, 95%<i>CI</i> = 1.01–1.22, <i>p</i> = 0.032), rs1462845 conferred significantly increased hazards of all-cause mortality among CAD cases. Even after controlling for multiple covariates, but in only the primary cohort, rs6788787 conferred significantly improved survival (<i>HR</i> = 0.80, 95% <i>CI</i> = 0.69–0.92, <i>p</i> = 0.002). Post-hoc analyses further stratifying by sex and disease severity revealed replicated effects for rs1462845: even after adjusting for aforementioned covariates and coronary interventional procedures, males with severe burden of CAD had significantly amplified hazards of death with the minor variant of rs1462845 in both cohorts (<i>HR</i> = 1.29, <i>95% CI</i> = 1.08–1.55, <i>p</i> = 0.00456; replication <i>HR</i> = 1.25, <i>95% CI</i> = 1.05–1.49, <i>p</i> = 0.013). Kaplan-Meier curves revealed unique cohort-specific genotype effects on survival. Additional analyses demonstrated that the homozygous risk genotype (‘A/A’) fully explained the increased hazard in both cohorts. None of the post-hoc analyses in control subjects were significant for any model. This suggests that genetic effects of rs1462845 on survival are unique to CAD presence. This represents formal, replicated evidence of genetic contribution of rs1462845 to increased risk for all-cause mortality; the contribution is unique to CAD case status and specific to males with severe burden of CAD.</p></div

Hazards of Death in Caucasian Control Subjects.

<p>Hazards of Death in Caucasian Control Subjects.</p

Kaplan–Meier survival curves for IMHC males with severe CAD by genotype for <i>LSAMP</i> SNP rs6788787.

<p><i>X</i>-axis displays the number of days from catheterization to death (all-cause mortality). <i>Y</i>-axis displays the Kaplan–Meier survival probability by genotype. A is the minor allele; GG, wild-type genotype (reference; black curve); GA, heterozygous genotype; and AA, risk homozygous genotype (red curve). This model did not demonstrate significant genotype effects on survival probability (<i>HR</i> = 1.04, <i>95% CI</i> = 0.83–1.29, <i>p</i> = 0.752, additive genetic model).</p

Kaplan–Meier survival curves for IMHC males with severe CAD by genotype for <i>LSAMP</i> SNP rs6788787.

<p><i>X</i>-axis displays the number of days from catheterization to death (all-cause mortality). <i>Y</i>-axis displays the Kaplan–Meier survival probability by genotype. A is the minor allele; GG, wild-type genotype (reference; black curve); GA, heterozygous genotype; and AA, risk homozygous genotype (red curve). No significant differences were found in any statistical modeling (data not shown).</p

Kaplan–Meier survival curves for CATHGEN males with severe CAD by genotype for <i>LSAMP</i> SNP rs6788787.

<p><i>X</i>-axis displays the number of days from catheterization to death (all-cause mortality). <i>Y</i>-axis displays the Kaplan–Meier survival probability by genotype. A is the minor allele; GG, wild-type genotype (reference; black curve); GA, heterozygous genotype; and AA, risk homozygous genotype (red curve). Only after controlling for covariates was this model significant for reduced hazards of death by genotype (<i>HR</i> = 0.71, <i>95% CI</i> = 0.54–0.93, <i>p</i> = 0.0148, additive genetic model).</p