Hypercholesterolemia Is Associated with the Apolipoprotein C-III (<em>APOC3</em>) Genotype in Children Receiving HAART: An Eight-Year Retrospective Study

<div><p>Polymorphisms in apolipoprotein genes have shown to be predictors of plasma lipid levels in adult cohorts receiving highly active antiretroviral therapy (HAART). Our objective was to confirm the association between the <em>APOC3</em> genotype and plasma lipid levels in an HIV-1-infected pediatric cohort exposed to HAART. A total of 130 HIV-1-infected children/adolescents that attended a reference center in Argentina were selected for an 8-year longitudinal study with retrospective data collection. Longitudinal measurements of plasma triglycerides, total cholesterol, HDL-C and LDL-C were analyzed under linear or generalized linear mixed models. The contribution of the <em>APOC3</em> genotype at sites −482, −455 and 3238 to plasma lipid levels prediction was tested after adjusting for potential confounders. Four major <em>APOC3</em> haplotypes were observed for sites −482/−455/3238, with estimated frequencies of 0.60 (C/T/C), 0.14 (T/C/C), 0.11 (C/C/C), and 0.11 (T/C/G). The <em>APOC3</em> genotype showed a significant effect only for the prediction of total cholesterol levels (p<0.0001). However, the magnitude of the differences observed was dependent on the drug combination (p = 0.0007) and the drug exposure duration at the time of the plasma lipid measurement (p = 0.0002). A lower risk of hypercholesterolemia was predicted for double and triple heterozygous individuals, mainly at the first few months after the initiation of Ritonavir-boosted protease inhibitor-based regimens. We report for the first time a significant contribution of the genotype to total cholesterol levels in a pediatric cohort under HAART. The genetic determination of <em>APOC3</em> might have an impact on a large portion of HIV-1-infected children at the time of choosing the treatment regimens or on the counter-measures against the adverse effects of drugs.</p> </div

Predicted risk of dyslipidemia according to different treatment options after one year of exposure.

<p>Odds ratio (OR) estimates for individuals with no <i>APOC3</i> variants (“000” haplotype) were adjusted by the variables listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039678#pone.0039678.s004" target="_blank">Table S1</a> under Generalized Liner Mixed-effects Model (GLMM), and after backward elimination algorithm. Dots depict punctual estimations for the mean effect of the exposure to each drug while lines depict the 95% confidence intervals.</p

Prediction of lipids levels under linear mixed models (LMM).

<p>The contribution of each factor was evaluated with Wald test on 127 individuals with full haplotype characterization; p-values are depicted. Significant p-values (p<0.003125, after Bonferroni correction) are indicated in bold numbers. Correlation sign is depicted between parentheses for p-values below 0.05. NA: variable excluded by stepwise backward elimination. ¹Alternative 1 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039678#pone.0039678.s001" target="_blank">Figure S1</a>). ²Alternative 2 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039678#pone.0039678.s001" target="_blank">Figure S1</a>). ³Alternative 3 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039678#pone.0039678.s001" target="_blank">Figure S1</a>).</p

Frequency of haplotype pairs (loci−482, 455 and 3238 on the −<i>APOC3</i> gene, respectively) as observed in 127 HIV-1-infected pediatric patients.

<p>Inset on top right shows haplotype frequency estimation by an Expectation-Maximization (EM) algorithm.</p

Generalized Linear Mixed-effects Model (GLMM) projections of hypercholesterolemia risk for patients carrying the indicated haplotype pairs.

<p>GLMM projections for a ARV-experienced male (treatment regimen starting after 62 months on HAART, the observed mean time on HAART at new regimen initiation) under a RTV-boosted PI regimen without D4T. Upper and lower panels depict contrasts to WT before exposure or at the same exposure time. Haplotype notation indicates the gene dose at each locus. Dots represent punctual contrast projections. Dots with labels indicate haplotype pairs statistically significantly different from WT at the same time (only lower panel). Odds ratios for common haplotype pairs “020”, “120” and “121” could not be estimated due to power issues. The contribution of all the factors included in GLMM were included in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039678#pone.0039678.s007" target="_blank">Table S4</a>.</p

Hierarchical test for differences in absolute plasma levels (LMM)¹.

1<p>Bonferroni corrected significance level was α* = 0.003125.</p>2<p>Test for the contribution of <i>APOC3</i> genotypes taking into account interactions with specific treatment scheme (inclusion of PIs boosted with RTV and/or D4T) and time of exposure (alternative 3 vs. null, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039678#pone.0039678.s001" target="_blank">Figure S1</a>).</p>3<p>Test for the contribution of the interaction between <i>APOC3</i> genotypes and time of exposure (alternative 3 vs. alternative 2).</p>4<p>Test for the contribution of the interaction between <i>APOC3</i> genotypes and treatment scheme (alternative 2 vs. alternative 1).</p>5<p>Test for the contribution of <i>APOC3</i> genotypes without interaction (alternative 1 vs. null).</p

Characteristics of the population studied.

1<p>96.2% of patients were born in Buenos Aires, 2.3% in other provinces, and 1.5% in other countries.</p>2<p>Study endpoint: December 31^st, 2008.</p>3<p>n = 1484.</p>4<p>n = 1574.</p>5<p>≥12 hours fasting status was presumed for 97.5% of blood samples.</p

Prediction of mean total cholesterol (TC) plasma levels variations for individuals carrying <i>APOC3</i> minor alleles.

<p>Linear Mixed-effects Model (LMM) projections for a ARV-experienced male under his first HAART drug regimen. Basal levels for total cholesterolemia were estimated subtracting the effect of adjusted treatment options (RTV, NFV, NNRTIs and D4T) and the effect of minor alleles on other loci. Thick lines depict punctual estimates, whereas dotted lines depict the 95% confidence intervals. *WT =  projections for individuals without any minor alleles.</p

Treatment changes and actions after viral rebound.

<p>Treatment changes and actions after viral rebound.</p

Virology, immunology and secondary endpoints over all follow-up.

<p>Virology, immunology and secondary endpoints over all follow-up.</p