B Cell Depletion in HIV-1 Subtype A Infected Ugandan Adults: Relationship to CD4 T Cell Count, Viral Load and Humoral Immune Responses

To better understand the nature of B cell dysfunctions in subjects infected with HIV-1 subtype A, a rural cohort of 50 treatment-naïve Ugandan patients chronically infected with HIV-1 subtype A was studied, and the relationship between B cell depletion and HIV disease was assessed. B cell absolute counts were found to be significantly lower in HIV-1+ patients, when compared to community matched negative controls (p<0.0001). HIV-1-infected patients displayed variable functional and binding antibody titers that showed no correlation with viral load or CD4+ T cell count. However, B cell absolute counts were found to correlate inversely with neutralizing antibody (NAb) titers against subtype A (p = 0.05) and subtype CRF02_AG (p = 0.02) viruses. A positive correlation was observed between subtype A gp120 binding antibody titers and NAb breadth (p = 0.02) and mean titer against the 10 viruses (p = 0.0002). In addition, HIV-1 subtype A sera showed preferential neutralization of the 5 subtype A or CRF02_AG pseudoviruses, as compared with 5 pseudoviruses from subtypes B, C or D (p<0.001). These data demonstrate that in patients with chronic HIV-1 subtype A infection, significant B cell depletion can be observed, the degree of which does not appear to be associated with a decrease in functional antibodies. These findings also highlight the potential importance of subtype in the specificity of cross-clade neutralization in HIV-1 infection

Patients infected with HIV-1 subtype A show greater frequency and magnitude of neutralization against subype A and CRF02_AG viruses.

<p>Sera from 50 HIV-1 subtype A-infected patients were tested in the TZM-bl neutralization assay against 1–3 pseudoviruses of each subtype indicated. The frequency of neutralization (% of titers >10) of viruses in a given subtype (black bars), as well as the geometric mean titer (ID₅₀, white bars) against each viral subtype, is shown. For the geometric mean titers, the error bars represent the 95% confidence intervals.</p

The absolute B lymphocyte count in HIV-1 subtype A infected patients is significantly lower than in HIV-1 negative controls.

<p>Samples of whole blood from HIV-1 infected and uninfected individuals were analysed using flow cytometry and the MultiTEST™ IMK Kit (BD). Lymphocyte subpopulations were quantified and samples from 261 HIV-1 negative individuals (white box) were compared with HIV positives. The absolute counts of CD19+ B lymphocytes in 50 HIV-1 subtype A infected patients (grey box) and in 192 patients with all HIV-1 subtypes combined (hatched box) were significantly reduced in comparison to uninfected participants. (p<0.0001, Mann-Whitney test). The whiskers represent the range from the 10th–90th percentile.</p

B lymphocyte absolute counts in HIV-1 subtype A infected Ugandans correlate inversely with neutralizing antibody titers against different HIV-1 subtypes.

<p>Neutralizing antibody responses in sera of 50 HIV-1 subtype A infected patients were determined against a panel of 10 pseudoviruses in the TZM-bl Assay. The mean ID₅₀ using each serum sample against all 10 pseudoviruses or against all pseudoviruses within a subtype was calculated. B lymphocyte numbers showed a significant inverse correlation with neutralizing antibody titers against the mean ID₅₀ of all 10 pseudoviruses (A) and with the mean ID₅₀ for subtype CRF02_AG (B); the same trend was observed for subtype A viruses (C). B lymphocyte numbers also showed a trend towards inverse correlation with breadth (number of panel viruses neutralized) (D).</p

Absolute CD4+ T lymphocyte counts correlate with B lymphocyte absolute counts, but not with HIV-1 viral Load.

<p>The absolute counts of B lymphocytes correlated with the absolute number of CD4+ T lymphocytes (A). The number of plasma HIV-1 RNA copies/ml showed a significant inverse correlation with the CD4+ T lymphocyte percentage (C) and a trend towards correlation with CD4 absolute number (B).</p

Breadth of neutralization in HIV-1 subtype A infected patient sera.

<p>Shown is the breadth of neutralization for each serum sample against a panel of 10 pseudoviruses from five different subtypes (Purple columns = subtype CRF02_AG, yellow = subtype A, green = subtype B, blue = subtype C and orange = subtype D). Red boxes indicate ID₅₀s>10 (positive neutralization). HIV-1 subtype A infected subjects showed more frequent neutralization against subtypes CRF02_AG and A, as compared to non-A subtypes (p<0.001, Fisher's exact test).</p

Binding antibodies against HIV-1 gp120 correlate with mean NAb Titer and breadth of neutralization.

<p>Subtype A gp120 binding antibody titers correlate with the mean NAb titers against all 10 pseudoviruses (A) and the mean ID₅₀s of subtype CRF02_AG (B) and subtype A (C) pseudoviruses. A significant correlation was also seen between the gp120 binding antibody titers and neutralization breadth, as measured by the number of viruses neutralized (D).</p

International seroepidemiology of adenovirus serotypes 5, 26, 35, and 48 in pediatric and adult populations

Recombinant adenovirus serotype 5 (rAd5) vaccine vectors for HIV-1 and other pathogens have been shown to be limited by high titers of Ad5 neutralizing antibodies (NAbs) in the developing world. Alternative serotype rAd vectors have therefore been constructed. Here we report Ad5, Ad26, Ad35, and Ad48 NAb titers in 4381 individuals from North America, South America, sub-Saharan Africa, and Southeast Asia. As expected, Ad5 NAb titers were both frequent and high magnitude in sub-Saharan Africa and Southeast Asia. In contrast, Ad35 NAb titers proved infrequent and low in all regions studied, and Ad48 NAbs were rare in all regions except East Africa. Ad26 NAbs were moderately common in adults in sub-Saharan Africa and Southeast Asia, but Ad26 NAb titers proved markedly lower than Ad5 NAb titers in all regions, and these relatively low Ad26 NAb titers did not detectably suppress the immunogenicity of 4×10(10)vp of a rAd26-Gag/Pol/Env/Nef vaccine in rhesus monkeys. These data inform the clinical development of alternative serotype rAd vaccine vectors in the developing worl