Influence of HLA class I haplotypes on HIV-1 seroconversion and disease progression in Pumwani sex worker cohort.

We examined the effect of HLA class I haplotypes on HIV-1 seroconversion and disease progression in the Pumwani sex worker cohort. This study included 595 HIV-1 positive patients and 176 HIV negative individuals. HLA-A, -B, and -C were typed to 4-digit resolution using sequence-based typing method. HLA class I haplotype frequencies were estimated using PyPop 32-0.6.0. The influence of haplotypes on time to seroconversion and CD4+ T cell decline to <200 cells/mm3 were analyzed by Kaplan-Meier analysis using SPSS 13.0. Before corrections for multiple comparisons, three 2-loci haplotypes were significantly associated with faster seroconversion, including A*23∶01-C*02∶02 (p = 0.014, log rank(LR) = 6.06, false-discovery rate (FDR) = 0.056), B*42∶01-C*17∶01 (p = 0.01, LR = 6.60, FDR = 0.08) and B*07∶02-C*07∶02 (p = 0.013, LR = 6.14, FDR = 0.069). Two A*74∶01 containing haplotypes, A*74∶01-B*15∶03 (p = 0.047, LR = 3.942, FDR = 0.068) and A*74∶01-B*15∶03-C*02∶02 (p = 0.045, LR = 4.01, FDR = 0.072) and B*14∶02-C*08∶02 (p = 0.021, LR = 5.36, FDR = 0.056) were associated with slower disease progression. Five haplotypes, including A*30∶02-B*45∶01 (p = 0.0008, LR = 11.183, FDR = 0.013), A*30∶02-C*16∶01 (p = 0.015, LR = 5.97, FDR = 0.048), B*53∶01-C*04∶01 (p = 0.010, LR = 6.61, FDR = 0.08), B*15∶10-C*03∶04 (p = 0.031, LR = 4.65, FDR = 0.062), and B*58∶01-C*03∶02 (p = 0.037, LR = 4.35, FDR = 0.066) were associated with faster progression to AIDS. After FDR corrections, only the associations of A*30∶02-B*45∶01 and A*30∶02-C*16∶01 with faster disease progression remained significant. Cox regression and deconstructed Kaplan-Meier survival analysis showed that the associations of haplotypes of A*23∶01-C*02∶02, B*07∶02-C*07∶02, A*74∶01-B*15∶03, A*74∶01-B*15∶03-C*02∶02, B*14∶02-C*08∶02 and B*58∶01-C*03∶02 with differential seroconversion or disease progression are due to the dominant effect of a single allele within the haplotypes. The true haplotype effect was observed with A*30∶02-B*45∶01, A*30∶02-C*16∶02, B*53∶01-C*04∶01 B*15∶10-C*03∶04, and B*42∶01-C*17∶01. In these cases, the presence of both alleles accelerated the disease progression or seroconversion than any of the single allele within the haplotypes. Our study showed that the true effects of HLA class I haplotypes on HIV seroconversion and disease progression exist and the associations of HLA class I haplotype can also be due to the dominant effect of a single allele within the haplotype.

Immunogenicity of sequences around HIV-1 protease cleavage sites: potential targets and population coverage analysis for a HIV vaccine targeting protease cleavage sites.

Developing an effective preventative vaccine against HIV-1 has proved to be a great challenge. The classical and proven vaccine approach has failed so far or produced a modest effect, new approaches are needed. In this study we evaluated the immunogenicity of the sequences around the protease cleavage sites (PCS) and the population coverage of a vaccine targeting HIV-1 PCS. The sequence conservation was evaluated by comparing entropy score of sequences around PCS with Gag and Pol. The immunogenicity of sequences around the 12 PCS (+10/-10 amino acids) was analyzed by identifying epitopes of HLA class I alleles in PCS region using four approaches: (1) identification of previously reported HLA class I allele epitopes around PCS region; (2) screening and validating epitopes of 8 HLA class I alleles common to most world populations using iTopia Epitope Discovery system and IFN-γ ELISpot assays; (3) screening of 151 patients of Pumwani cohort for PBMC IFN-γ ELISPOT responses to the subtype A and D consensus around PCS region; and (4) prediction of HLA alleles with epitopes around the PCS using NetMHCpan. Population coverage was calculated using the web-based analysis tool of the Immune Epitope Database based on HLA class I genotype frequencies from dbMHC database. The results showed that many HLA class I alleles have multiple epitopes in the 12 PCS regions, indicating sequence immunogenicity around PCS. Multiple epitopes of many HLA class I alleles common to >95% world populations have been identified around the 12 PCS region. Targeting these sites is a feasible vaccine approach.

Clade-specific evolution mediated by HLA-B*57/5801 in human immunodeficiency virus type 1 clade A1 p24.

HLA-B*57-mediated selection pressure leads to a typical escape pathway in human immunodeficiency virus type 1 (HIV-1) CD8 epitopes such as TW10. Whether this T242N pathway is shared by all clades remains unknown. We therefore assessed the nature of HLA-B*57 selection in a large, observational Kenyan cohort where clades A1 and D predominate. While T242N was ubiquitous in clade D HLA-B*57(+) subjects, this mutation was rare (15%) in clade A1. Instead, P243T and I247L were selected by clade A1-infected HLA-B*57 subjects but not by HLA-B*5801(+) subjects. Our data suggest that clade A1 consensus proline at Gag residue 243 might represent an inherent block to T242N escape in clade A1. We confirmed immunologically that P243T and I247L likely represent escape mutations. HLA-B*57 evolution also differed between clades in the KF11 and IW9 epitopes. A better understanding of clade-specific evolution is important for the development of HIV vaccines in regions with multiple clades.

Multiple T-cell epitopes overlap positively-selected residues in the p1 spacer protein of HIV-1 gag.

OBJECTIVES: The p1 region of HIV-1 gag contains the frameshift stem-loop, gag-pol transframe and a protease cleavage site that are crucial for viral assembly, replication and infectivity. Identifying and characterizing CD8+ epitopes that are under host immune selection in this region will help in designing effective vaccines for HIV-1. DESIGN: An approach combining bioinformatical analysis and interferon gamma enzyme-linked immunosorbent spot (ELISPOT) assays is used to identify and characterize the epitopes. Potential human leukocyte antigen (HLA)-restricted epitopes were identified by correlating the positively-selected mutations with host HLA alleles. METHODS: ELISPOT analysis with overlapping peptides was used to confirm and characterize the epitopes. RESULTS: Four positively-selected residues were significantly associated with HLA class I alleles, including HLA B*1302 (K4R, P = 0.0008 and I5L, P = 0.0108), A*7401 (S9N, P = 0.0002) and A*30 genotypes (P7S, P = 0.009), suggesting epitopes restricted by these alleles are present in this region. ELISPOT analysis with patient peripheral blood mononuclear cells (PBMCs) identified seven novel epitopes restricted by the 3 alleles. Two types of epitopes were observed in this region based on the ELISPOT responses, Type I: the positively-selected variation does not affect CD8+ T-cell responses; and Type II: the CD8+ T-cell responses are determined by the epitope variants. CONCLUSION: We identified and characterized seven novel CD8+ epitopes in the p1 spacer protein region. Classifying the effects of positively-selected variants on CD8+ T-cell responses will help in designing effective vaccines for HIV-1.

An integrative bioinformatic approach for studying escape mutations in human immunodeficiency virus type 1 gag in the Pumwani Sex Worker Cohort.

Human immunodeficiency virus type 1 (HIV-1) is able to evade the host cytotoxic T-lymphocyte (CTL) response through a variety of escape avenues. Epitopes that are presented to CTLs are first processed in the presenting cell in several steps, including proteasomal cleavage, transport to the endoplasmic reticulum, binding by the HLA molecule, and finally presentation to the T-cell receptor. An understanding of the potential of the virus to escape CTL responses can aid in designing an effective vaccine. To investigate such a potential, we analyzed HIV-1 gag from 468 HIV-1-positive Kenyan women by using several bioinformatic approaches that allowed the identification of positively selected amino acids in the HIV-1 gag region and study of the effects that these mutations could have on the various stages of antigen processing. Correlations between positively selected residues and mean CD4 counts also allowed study of the effect of mutation on HIV disease progression. A number of mutations that could create or destroy proteasomal cleavage sites or reduce binding affinity of the transport antigen processing protein, effectively hindering epitope presentation, were identified. Many mutations correlated with the presence of specific HLA alleles and with lower or higher CD4 counts. For instance, the mutation V190I in subtype A1-infected individuals is associated with HLA-B*5802 (P = 4.73 x 10(-4)), a rapid-progression allele according to other studies, and also to a decreased mean CD4 count (P = 0.019). Thus, V190I is a possible HLA escape mutant. This method classifies many positively selected mutations across the entire gag region according to their potential for immune escape and their effect on disease progression.

Slow continuous intravenous plasmapheresis (SCIP): clinical applications and hemostability of extracorporeal ultrafiltration.

An intravenous plasmapheresis catheter which excludes >99.4% of platelets from external ultrafiltration circuits is currently undergoing safety and efficacy trials for fluid removal from NYHA class II-IV congestive heart failure patients resistant to diuretic drug therapy. In animals, the SCIP catheter allowed a four fold increase in ultrafiltration efficiency without hemolysis, hemoinstability or external cartridge changes in 72 hours of treatment. Further, systemic anticoagulation was not required. These techniques might be envisioned for treatment of fluid overload in heart failure, surgery or trauma and may have applications in therapeutic apheresis, venous thrombosis, liver disease or autologous tissue engineering.