Replicating adenovirus HIV/SIV recombinant priming alone or in combination with a gp140 protein boost results in significant control of viremia following a SHIV89.6P challenge in Mamu-A*01 negative rhesus macaques.

Previously, replicating adenovirus type 5 host range (Ad5hr)-HIV/SIV recombinant priming in combination with SIV envelope boosting, resulted in significant, durable protection in 39% of rhesus macaques after SIVmac251 challenge. Both Env-specific antibody mediating ADCC, and cellular immunity correlated with protection. Here we evaluate the relative immunogenicities of novel HIV proteins and their contribution to protection in a SHIV89.6P model. All groups were primed with Ad-HIVenv89.6P, SIVgag239, and SIVnef239 recombinants. One group was not boosted, one received HIV89.6Pgp140DeltaCFI protein, and one a novel HIV-1 poly-peptide "peptomer". The HIV89.6Pgp140DeltaCFI protein in adjuvant strongly boosted Env-specific antibody and memory T cell responses in blood and tissue, resulting in significant reductions in acute and set point viremia. Macaques not boosted, showed a significant reduction in set point viremia, a full 32 weeks after the last Ad priming immunization. The HIV peptomer-boosted group showed a trend toward chronic viremia reduction, but was not protected.

Evaluation of passively transferred, nonneutralizing antibody-dependent cellular cytotoxicity-mediating IgG in protection of neonatal rhesus macaques against oral SIVmac251 challenge.

Previously, Ab-dependent cellular cytotoxicity (ADCC) was significantly correlated with reduced acute viremia upon intrarectal SIVmac251 challenge of immunized rhesus macaques. To directly assess ADCC protective efficacy, six neonatal macaques were infused s.c. with immune IgG (220 mg/kg) purified from the immunized animals and positive for ADCC and Ab-dependent cell-mediated viral inhibition (ADCVI) activities. Six neonates received control IgG. The neonates were challenged twice orally with 10(5) 50% inhibiting tissue culture-infective dose of SIVmac251 2 days post-IgG infusion. At challenge, plasma of neonates that received immune IgG did not neutralize SIVmac251 but had geometric mean ADCC titers of 48,130 and 232,850 against SIVmac251 -infected and gp120-coated targets, respectively. Peak ADCVI activity varied from 62 to 81%. ADCC activity declined with the 2-wk IgG half-life but was boosted at wk 4, together with de novo ADCC-mediating Abs in controls, by postchallenge viremia. ADCVI activity was similarly induced. No protection, assessed by viral burdens, CD4 counts, and time to euthanasia was observed. Possible factors contributing to the discrepancy between the previous correlation and lack of protection here include: the high oral challenge dose compared with the 400-fold lower intrarectal dose; the challenge route with regard to viral dissemination and distribution of infused IgG; insufficient NK effector activity and/or poor functionality in newborns; insufficient immune IgG; and the possibility that the previous correlation of ADCC with protection was augmented by cellular immune responses also present at challenge. Future studies should explore additional challenge routes in juvenile macaques using higher amounts of potent IgG preparations.

A simplified method for the rapid fluorometric assessment of antibody-dependent cell-mediated cytotoxicity.

We demonstrate that the FATAL cytolysis assay can be adapted into a rapid and fluorometric antibody-dependent cellular cytotoxicity assay (RFADCC). The RFADCC relies on double-staining target cells with a membrane dye (PKH-26) and a viability dye (CFSE) prior to the addition of antibody and effector cells. We used the RFADCC to assess dose-dependent and envelope-specific anti-human immunodeficiency virus (HIV) ADCC responses mediated by monoclonal antibody-2G12 and human sera. Using the assay, we also detected early anti-simian immunodeficiency virus (SIV) ADCC responses in rhesus macaques infected with pathogenic SIV(mac251). Importantly, the RFADCC was further useful in monitoring anti-HIV and anti-SIV ADCC responses elicited by immunizing chimpanzees and rhesus macaques with replicating adenovirus-based AIDS vaccine candidates. In comparison to the standard chromium release assay, the RFADCC provides a higher cell killing readout and is advantageous in allowing use of viably frozen as well as fresh effector cells, thus facilitating assay standardization. The RFADCC is therefore a simple, reliable, and highly sensitive method that can be applied to assess the ADCC activity of monoclonal antibodies as well as ADCC responses elicited by HIV or SIV infection or by AIDS vaccine candidates.

Enhanced cellular immunity to SIV Gag following co-administration of adenoviruses encoding wild-type or mutant HIV Tat and SIV Gag.

Among candidate antigens for human immunodeficiency virus (HIV) prophylactic vaccines, the regulatory protein Tat is a critical early target, but has a potential for immune suppression. Adenovirus (Ad) recombinants encoding wild-type HIV Tat (Tat-wt) and a transdominant negative mutant HIV Tat (Tat22) were constructed and administered to mice separately or together with Ad-SIVgag. Immunogenicity and effects on immune responses to the co-administered Gag immunogen were evaluated. Wild-type and mutant Tat recombinants elicited similar Tat-specific cellular and humoral immune responses. Co-administration of either Tat immunogen with Ad-SIVgag induced modest but significant enhancement of Gag-specific interferon-gamma secreting T cells and lymphoproliferative responses. Neither the Ad-recombinant encoding Tat-wt nor Tat22 suppressed induction of anti-Tat or anti-Gag antibodies. Based on the immune responses observed in mice, both recombinants appear to be suitable vaccine candidates. Their contribution to protective efficacy remains to be determined in a non-human primate model.

Vaccine-elicited antibodies mediate antibody-dependent cellular cytotoxicity correlated with significantly reduced acute viremia in rhesus macaques challenged with SIVmac251.

Effector cells armed with Abs can eliminate virus-infected target cells by Ab-dependent cellular cytotoxicity (ADCC), an immune mechanism that has been largely overlooked in HIV vaccine development. Here, we show that a prime/boost AIDS vaccine approach elicits potent ADCC activity correlating with protection against SIV in rhesus macaques (Macacca mulatta). Priming with replicating adenovirus type 5 host range mutant-SIV recombinants, followed by boosting with SIV gp120, elicited Abs with ADCC activity against SIV(mac251)-infected cells. In vitro ADCC activity correlated with in vivo reduced acute viremia after a mucosal challenge with pathogenic SIV. Our findings expose ADCC activity as an immune correlate that may be relevant in the rational design of an efficacious vaccine against HIV.