Tiered Categorization of a Diverse Panel of HIV-1 Env Pseudoviruses for Assessment of Neutralizing Antibodies

Abstract

The restricted neutralization breadth of vaccine-elicited antibodies is a major limitation of current human immunodeficiency virus-1 (HIV-1) candidate vaccines. In order to permit the efficient identification of vaccines with enhanced capacity for eliciting cross-reactive neutralizing antibodies (NAbs) and to assess the overall breadth and potency of vaccine-elicited NAb reactivity, we assembled a panel of 109 molecularly cloned HIV-1 Env pseudoviruses representing a broad range of genetic and geographic diversity. Viral isolates from all major circulating genetic subtypes were included, as were viruses derived shortly after transmission and during the early and chronic stages of infection. We assembled a panel of genetically diverse HIV-1-positive (HIV-1 ؉ ) plasma pools to assess the neutralization sensitivities of the entire virus panel. When the viruses were rank ordered according to the average sensitivity to neutralization by the HIV-1 ؉ plasmas, a continuum of average sensitivity was observed. Clustering analysis of the patterns of sensitivity defined four subgroups of viruses: those having very high (tier 1A), above-average (tier 1B), moderate (tier 2), or low (tier 3) sensitivity to antibody-mediated neutralization. We also investigated potential associations between characteristics of the viral isolates (clade, stage of infection, and source of virus) and sensitivity to NAb. In particular, higher levels of NAb activity were observed when the virus and plasma pool were matched in clade. These data provide the first systematic assessment of the overall neutralization sensitivities of a genetically and geographically diverse panel of circulating HIV-1 strains. These reference viruses can facilitate the systematic characterization of NAb responses elicited by candidate vaccine immunogens.

Antibody‐virus co‐evolution in HIV infection: paths for HIV vaccine development

“…Very few bnAbs were known prior to this time (e.g., b12, 2G12, 2F5, 4E10), and the breadth and potency of these early bnAbs pale in comparison to the newer bnAbs (12). High throughput assay technologies with molecularly cloned Env-pseudotyped viruses (67–69), combined with a heightened awareness of the importance of the “Tier 2” neutralization phenotype of most circulating strains (70, 71) were major contributors. New reporter gene assays in either TZM-bl or U87.CD4.CCR5.CXCR4 became available that are more rapid, sensitive and less costly than other assays, and are amenable to high standards of optimization and validation (72).…”

Identification and specificity of broadly neutralizing antibodies against HIV

“…The ability to analyze mAb and serum activity against large panels of viruses was demonstrated20 and subsequently used to evaluate large numbers of HIV‐infected donors in the International AIDS Vaccine Initiative (IAVI) Protocol G and C studies to identify those with exceptionally potent and broad sera,8 map the specificities underlying these responses,7, 21 and then isolate bnAbs from these individuals 22, 23, 24, 25, 26, 27, 28. Independently, the standardization of the TZM‐bl neutralization assay and the definition of neutralization sensitivity tiers29, 30, 31 allowed much more rigorous serum analysis.…”

Generation of a neutralization-resistant CCR5 tropic simian/human immunodeficiency virus (SHIV-MK38) molecular clone, a derivative of SHIV-89.6

“…The stock viruses were prepared with 293T cells by transfection or with C8166-CCR5 cells by infection Seaman et al, 2010). The titration was performed with TZM-bl cells for in vitro assays and with C8166-CCR5 cells for the in vivo inoculation experiment.…”

“…Based on these results, most previously used SHIVs were thought to be neutralization sensitive because they contained the env gene of laboratory strains obtained by passaging through T-cell lines. Another report showed that the neutralization phenotype against neutralizing antibodies can be categorized as: tier 1 (neutralization sensitive), tier 2 (neutralization resistant) and tier 3 (extremely neutralization resistant) (Seaman et al, 2010). The majority of HIV-1 clinical isolates are categorized as tier 2, whilst HIV-1 laboratory strains are categorized as tier 1.…”