HIV-2 is a somewhat understudied virus with a lower pathogenicity and transmission rate than HIV-1. Gulsen Özkaya Sahin and Marianne Jansson from Lund University in Sweden demonstrated that the breadth and potency of neutralizing activity in plasma differ between HIV-1 and HIV-2 infected individuals. Plasma from HIV-2 patients was shown to neutralize HIV-2 isolates at a high frequency, whereas plasma from HIV-1 patients had much lower intratype neutralizing activity against HIV-1 isolates. The intratype neutralizing activity correlated with viral load in patients infected with HIV-1 but not with HIV-2. Hence the neutralizing activity in HIV-2 infection does not seem to require high levels of antigen stimulation, whereas this is more important in HIV-1 infection. Jansson concluded that the viral structure of HIV-2 may promote a more potent humoral immune response and/or HIV-2 itself may be more neutralization sensitive than HIV-1.
Stefania Dispinseri (San Raffaele Scientific Institute) presented data from paediatric infections, correlating disease progression with neutralizing responses. HIV-1 infected children with slow disease progression developed an increasing breadth of neutralizing response over time, whereas fast-progressing children did not.
A similar study, conducted by Katharina Raue, a student from the German Primate Center, analysed the immune responses of infected rhesus monkeys, grouped into controllers or progressors. Animals controlling the infection demonstrated higher immune response than progressors, both humoral and cellular, and the response was directed against Gag proteins and not Envelope.
Not all patients develop broadly neutralizing antibodies; in fact, only 10-30% of HIV-1 infected patients possess antibodies with a strong cross-reactive neutralizing activity and less than 1% have broadly neutralizing antibodies. Zelda Euler from the University of Amsterdam performed a genome-wide association study, to learn more about which genetic markers are involved in the development of antibodies with broadly neutralizing activity. The study included 335 HIV-infected men who have sex with men from the Amsterdam Cohort with known neutralizing activity of their sera. Single nucleotide polymorphisms (SNP), in the major histocompatibility complex gene region, close to MICA and HCP5 genes on chromosome 6, were found to associate with broadly neutralizing activity. HCP5 encodes HLA complex protein 5, and has previously been associated with viral load and disease progression in HIV infected individuals [28, 29]. None of the identified SNPs which were associated with broadly neutralizing activity were associated with the clinical course of disease progression. In addition, the association between SNPs or broadly neutralizing activity of antibodies was not related to viral load or CD4+ T-cell count at viral load set-point. Although this finding is remarkable, its application to our understanding of the factors modulating the mounting of an effective immune response is not yet clear.
Marie Borggren, from the University of Lund, compared how CCR5 and CxCR4-using HIV-1 evolve during end-stage disease and how the immune responses correlate with the evolution. The Env proteins of end-stage viruses were often more positively charged and had fewer glycans than virus from the chronic stage of disease. End-stage R5 viruses were shown to use the trans-infection route from DC to T-cells less efficiently, which might be due to loss of a specific glycan in gp120.
David Reinhart (Polymun and University of Natural Resources and Life Sciences, Vienna) discussed the switching of the isotypes of anti-HIV-1 IgG antibodies 3D6 and 4B3 recognizing the ectodomain of gp41 to an IgA isotype and their expression in CHO cells in serum-free medium as dimeric IgA. The purification scheme mainly relies on IgA affinity chromatography, using VHH ligands. Unlike previously published protocols, the purification schedule can be used for any IgA irrespective of alpha heavy chains, light chains and their subtypes . These new reagents will allow the study of the influence of antibody isotype on their function with respect to antiviral activity against HIV. Studies are pending to investigate the potential role of IgA during viral exposure at the mucosa.
Enas Sheik-Khalil (University of Lund) described the development of an image-based high-throughput as well as high-content HIV neutralization assay. The assay is based on a plaque reduction assay along with a specialized image analysis tool tailored to analyse neutralization. An additional parameter, mean plaque area, enables the detection of virus fusogenicity to characterize the duration in vitro of neutralization. A decreased plaque size may indicate avidity of neutralizing antibodies. The assay demonstrated equal or higher sensitivity compared to conventional neutralization assays.
Eliciting new type of antibodies and defining novel epitopes will be critical for future vaccine development. Mark Hassell et al. (NIBSC) have produced novel C clade specific antibodies by priming mice with plasmid DNA encoding either CN54 or ZM96 gp140 genes, followed by a CN54 trimeric gp140 boost, all of subtype C. Applying the trimeric immunogens, 18 HIV-specific antibodies were successfully generated. However, no novel epitopes were defined. Although all isolated antibodies bound CN54, none were able to recognise and neutralise epitopes that are specific for the trimeric envelope structure.
Another approach to elicit novel antibodies, presented by Pierpaolo Racchiolli et al. (University of Verona), established new anti-HIV-1 antibodies by immunizing mice with CHO cells expressing gp120/gp41 and CHO cells expressing CD4 and CCR5 to generate fusion intermediate structures of the HIV envelope. One hybridoma clone, 10 F12, which showed the highest HIV-1 neutralization activity and lowest cell reactivity, was further investigated and stably produced in CHO cells as a chimeric antibody. Hybridoma and chimeric antibodies both neutralized several HIV viruses.
Other roles for antibodies
HIV infection can be inhibited by both neutralizing and non-neutralizing antibodies via the Fc receptor. Infection of myeloid dendritic cells, mDC, has been shown to be inhibited using both types of antibody. In contrast, Alexandre Lederle, a student from University of Strasbourg, demonstrated that infection of plasmacytoid dendritic cells, pDC, can be inhibited only by neutralizing antibodies, not by non-neutralizing antibodies, despite the expression of Fc receptors on the surface of pDC.
As discussed previously, transmission of HIV-1 via mucosal tissues during sexual intercourse globally represents the main route of infection. Immature dendritic cells reside in the genital mucosal tissues and are believed to be among the first cells to be in contact with HIV. An infected DC may act as a “Trojan horse” for HIV and further transmit viruses to primary CD4+ T lymphocytes. Antibodies that inhibit such viral transfer could prevent HIV infection. Bin Su from the University of Strasbourg addressed this hypothesis and demonstrated that neutralizing antibodies could efficiently inhibit the transfer of several R5 HIV-1 strains from primary DCs to autologous CD4+ T lymphocytes. Furthermore, he showed that neutralizing as well as non-neutralizing antibodies could decrease HIV-1 replication in DCs. Induction of both types of antibodies directly at mucosal sites may thus be beneficial when designing a future vaccine.
Besides neutralization, HIV-specific antibodies could thus have an impact on protection against HIV-1 transmission in several ways. Such HIV-specific antibodies could target not only free virions, but also infected cells, as well as the HIV transfer from infected cells to other cells. In order to understand the contribution of antibodies induced against these targets and especially the role of Fc-mediated protection, Marina Biedma from the University of Strasbourg investigated several inhibitory functions of anti-HIV IgGs. It was found that non-neutralizing inhibitory antibodies (NNIAbs) could mediate inhibitory activities such as ADCC or phagocytosis. Indeed, it was demonstrated that NNIAbs such as 4B3 and 246D were highly efficient in capturing free virions when compared to neutralizing antibody 2G12, 4E10 or 2F5. Furthermore, NNIAbs 4B3 and 246D revealed Fc-mediated inhibitory activities on viral replication in macrophages and DCs similar to some neutralizing antibody. When applied locally at the vaginal site of macaques, these NNIAbs were able to significantly decrease the peak viral load following vaginal experimental challenge with SHIV SF162P3. These results suggest that NNIAbs may be important in controlling viral infections.
In addition to broadly neutralizing antibodies, other targets for antibodies with alternative antiviral function are being explored. Donato Zipeto from the University of Verona described the identification of one such new target by unravelling the association between HLA-C and gp120. Using bimolecular fluorescence complementation analysis, it was shown that HLA-C competes for binding between β2m and gp120. Furthermore, association of gp120 with HLA-C happens early in the endoplasmic reticulum and continues during passage through the Golgi apparatus and early endosomes, until presentation at the cell membrane. It was found that HLA-C specifically incorporates into HIV-1 envelopes and increases viral infectivity by increasing viral envelope fusion with R5 and X4 HIV strains. A compound or antibody that could disturb this association might decrease viral infectivity. Furthermore, antibody to HLA-C displayed on the virusor virus-infected cell surface might participate in ADCC.