The goal of this project is to identify genetic and epigenetic factors contributing to human infectious diseases. HIV, EBV, and the hepatitis C virus (HCV) and hepatitis B virus (HBV) contribute to AIDS-associated malignancies, nasopharyngeal carcinoma (NPC) and hepatocellular carcinoma (HCC), respectively. Little is understood about the interplay between chronic viral infection and host genetic variation leading to pathogenic outcomes in persons infected with these viruses. Genome wide association studies (GWAS) have revealed that HLA class I alleles are most strongly associated with HBV clearance, HIV control of viral load, and with NPC. However, HLA explains only a small fraction of the variance for these infections and cancers. In addition, with advancing age of the HIV-infected population, diseases commonly associated with aging and affecting all organ systems are appearing at younger ages. We are embarking on genetic association studies to identify correlates of eye disease in the Longitudinal Study of Ocular Complications with AIDS (LSOCA) and of HIV-associated non-AIDS serious diseases in longitudinal natural and treated cohorts. Our laboratory is moving to integrate epigenetic signatures, gene expression, and results of GWAS and pathway analysis in a systems-based approach to further define genes and pathways important in the pathophysiology and carcinogenesis of infectious diseases. Accomplishments 1. Pneumocystis pneumonia (PCP) is a major AIDS-defining condition and a common complication of immunosuppression. Chromosome 3p21-22 harbors two clusters of chemokine receptor genes, several of which serve as major or minor coreceptors of HIV-1. We resequenced 7 chemokine receptors in 144 individuals representing extreme phenotypes for HIV progression and infection;six codon-changing SNPs were discovered. We identified a non-conservative amino acid change in CCRL2 (Y167F) associated with more rapid progression to AIDS defining conditions, and specifically to the AIDS-defining condition PCP. CCRL2 is involved in lung dendritic cell trafficking and may affect PCP by inducing inflammation. This is the first genetic association with a major AIDS defining condition and brings renewed attention to the chemokine-chemokine receptor nexus in HIV pathogenesis. Highlighting the role of inflammation may lead to better therapeutic options for persons presenting with PCP infection. The results of this study were published in PLoS Genetics. 2. GWAS is an agnostic, unbiased method for detecting genetic factors associated with disease and traits. Because of the large number of tests, rigorous criteria for genome wide significance (by convention, p<5 x 10-8) and independent replication are critical to avoid false positive associations. Since the number of persons with longitudinal clinical and laboratory data for infectious diseases is generally limited, the discovery of genetic factors associated with infectious disease phenotypes is problematic. To overcome this barrier, we have joined forces with the International HIV Consortium to contribute genotype-phenotypes from over 1000 HIV-infected participants with longitudinal follow-up for a meta-analysis. We are also part of the analytical team for viral load and CD4 T cell trajectories for this international collaboration. We are currently in the process of rigorous quality control (QC) for the genotype calls for three previous CCR GWAS for HBV and HIV clinical outcomes and for NPC. Using the ISHAPE software, we have determined the haplotype structure of participants enrolled in the NPC, HIV, and HBV studies with GWAS data. The next step will be to impute SNPs not represented on the Affymetric chip using data from the International HapMap Program and the 1000 Genome Project representing the major continental populations. With the enriched SNPs, we will reanalyze the data for association with multiple phenotypes using the 5 natural history cohorts and the LSOCA cohort, including HIV-associated non-AIDS and AIDS clinical outcomes. As mentioned above, a hindrance to the identification of host variants influencing HIV acquisition and pathogenesis is a lack of power to identify small effect size host variants. 3) The lifetime risk for African Americans for HIV-associated nephropathy is approximately 10% prior to widespread use of effective antiretroviral therapies. Two APOL1 variant alleles, one comprising two codon-changing SNPs and the other a 6-base pair deletion removing two amino acids are found in 35% of African Americans;these variants are also extremely frequent in west African populations, presumably driven by survival advantage in APOL1 mutation carriers, which confers resistance to African sleeping sickness. Carriers of two APOL1 risk alleles (approximately 13-15% of African Americans) are at a much higher risk of several forms of non-diabetic kidney disease. We showed that the lifetime incidence for HIVAN among HIV-infected African Americans carrying two APOL1 variants is 50%. The attributable risk (AR) (68%) and explained fraction (EF) (37%) for APOL1 variants with HIVAN are quantitatively similar to the role of smoking in non-small-cell lung cancer risk (AR, 90% for males;EF, 12%). This study, published in the Journal of American Society of Nephrology, suggests that screening for APOL1 risk variants may have a role in personalized medicine for persons with African ancestry and HIV infection. 4. To determine if APOL1 variants drive distinctive pathological characteristics, we genotyped 60 patients with biopsy-proven HIVAN. In this group, 37 had two risk alleles, 18 were heterozygous, and 5 had neither risk variant. There were no differences in the pathological findings of HIVAN and the number of APOL1 risk alleles. Further, the progression to end-stage kidney disease or death did not differ by the number of risk alleles. Median renal survival was 9.3 months in patients with zero or one risk allele compared to 11.7 months in patients with two APOL1 risk alleles. Our study suggests that although the majority of African-American patients with HIVAN have two APOL1 risk alleles other as yet unknown factors, including other genetic risk variants or environmental or viral factors, may influence the development of this disorder in those with zero or one APOL1 risk allele. This study was published in Kidney International. 5. APOL1 variants are associated with reduced estimated glomerular filtration in treated HIV-infected persons. The epidemiology of HIV kidney disease has changed over time. In the pre-cART era, HIVAN was prominent, but in the post-cART era, HIVAN has waned but kidney disease still remains a major complication due in part to drug toxicities, immune activation, and chronic inflammatory state. We hypothesized that APOL1/ MYH9 risk alleles could be associated with decreased kidney function in HIV-infected subjects with African ancestry. HIV-infected persons with African ancestry from the SMART cohort were genotyped for APOL1 (G1 and G2 variants) and MYH9 (4 intronic SNPs composing the E1 haplotype) risk variants. Kidney function using estimates of glomerular filtration rates (eGFR), was evaluated at baseline and during a 60 month follow-up using linear regression and linear mixed effects regression, respectively. APOL1 risk alleles were strongly associated with decreased eGFR at baseline (P=3x10-8);MYH9 renal risk alleles were independently associated with eGFR;APOL1 and MYH9 risk alleles remained persistently associated with lower eGFR during follow-up. The results of this study were presented at the CROI conference and are being formalized for publication.
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