Gerald K. Chege

Broad, high-magnitude and multifunctional CD4 + and CD8 + T-cell responses elicited by a DNA and modified vaccinia Ankara vaccine containing human immunodeficiency virus type 1 subtype C genes in baboons

Candidate human immunodeficiency virus (HIV) vaccine regimens based on DNA boosted with recombinant modified vaccinia Ankara (MVA) have been in development for some time, and there is evidence for improved immunogenicity of newly developed constructs. This study describes immune responses to candidate DNA and MVA vaccines expressing multiple genes (gag, RT, tat, nef and env) from HIV-1 subtype C in chacma baboons (Papio ursinus). The vaccine regimen induced (i) strong T-cell responses, with a median of 4103 spot forming units per 10(6) peripheral blood mononuclear cells by gamma interferon (IFN-gamma) ELISPOT, (ii) broad T-cell responses targeting all five vaccine-expressed genes, with a median of 12 peptides targeted per animal and without any single protein dominating the response, (iii) balanced CD4(+) and CD8(+) responses, which produced both IFN-gamma and interleukin (IL)-2, including IL-2-only responses not detected by the ELISPOT assay, (iv) vaccine memory, which persisted 1 year after immunization and could be boosted further, despite strong anti-vector responses, and (v) mucosal T-cell responses in iliac and mesenteric lymph nodes in two animals tested. The majority of peptide responses mapped contained epitopes previously identified in human HIV infection, and two high-avidity HIV epitope responses were confirmed, indicating the utility of the baboon model for immunogenicity testing. Together, our data show that a combination of DNA and MVA immunization induced robust, durable, multifunctional CD4(+) and CD8(+) responses in baboons targeting multiple HIV epitopes that may home to mucosal sites. These candidate vaccines, which are immunogenic in this pre-clinical model, represent an alternative to adenoviral-based vaccines and have been approved for clinical trials.

Recombinant Mycobacterium bovis BCG as an HIV Vaccine Vector

HIV-1 has resulted in a devastating AIDS pandemic. An effective HIV/AIDS vaccine that can be used to either, prevent HIV infection, control infection or prevent progression of the disease to AIDS is needed. In this review we discuss the use of Mycobacterium bovis BCG, the tuberculosis vaccine, as a vaccine vector for an HIV vaccine. Numerous features make BCG an attractive vehicle to deliver HIV antigens. It has a good safety profile, elicits long-lasting cellular immune responses and in addition manufacturing costs are affordable, a necessary consideration for developing countries. In this review we discuss the numerous factors that influence generation of a genetically stable recombinant BCG vaccine for HIV.

A prime-boost immunisation regimen using recombinant BCG and Pr55gag virus-like particle vaccines based on HIV type 1 subtype C successfully elicits Gag-specific responses in baboons

Mycobacterium bovis BCG is considered an attractive live bacterial vaccine vector. In this study, we investigated the immune response of baboons to a primary vaccination with recombinant BCG (rBCG) constructs expressing the gag gene from a South African HIV-1 subtype C isolate, and a boost with HIV-1 subtype C Pr55(gag) virus-like particles (Gag VLPs). Using an interferon enzyme-linked immunospot assay, we show that although these rBCG induced only a weak or an undetectable HIV-1 Gag-specific response on their own, they efficiently primed for a Gag VLP boost, which strengthened and broadened the immune responses. These responses were predominantly CD8+ T cell-mediated and recognised similar epitopes as those targeted by humans with early HIV-1 subtype C infection. In addition, a Gag-specific humoral response was elicited. These data support the development of HIV-1 vaccines based on rBCG and Pr55(gag) VLPs.

HIV-1 subtype C Pr55gag virus-like particle vaccine efficiently boosts baboons primed with a matched DNA vaccine

A DNA vaccine expressing human immunodeficiency virus type 1 (HIV-1) southern African subtype C Gag (pTHGag) and a recombinant baculovirus Pr55gag virus-like particle prepared using a subtype C Pr55gag protein (Gag VLP) was tested in a prime-boost inoculation regimen in Chacma baboons. The response of five baboons to Gag peptides in a gamma interferon (IFN-gamma) enzyme-linked immunospot (ELISPOT) assay after three pTHGag immunizations ranged from 100 to 515 spot-forming units (s.f.u.) per 10(6) peripheral blood mononuclear cells (PBMCs), whilst the response of two baboons to the Gag VLP vaccine ranged from 415 to 465 s.f.u. per 10(6) PBMCs. An increase in the Gag-specific response to a range of 775-3583 s.f.u. per 10(6) PBMCs was achieved by boosting with Gag VLPs the five baboons that were primed with pTHGag. No improvement in Gag responses was achieved in this prime-boost inoculation regimen by increasing the number of pTHGag inoculations to six. IFN-gamma responses were mapped to several peptides, some of which have been reported to be targeted by PBMCs from HIV-1 subtype C-infected individuals. Gag VLPs, given as a single-modality regimen, induced a predominantly CD8+ T-cell IFN-gamma response and interleukin-2 was a major cytokine within a mix of predominantly Th1 cytokines produced by a DNA-VLP prime-boost modality. The prime-boost inoculation regimen induced high serum p24 antibody titres in all baboons, which were several fold above that induced by the individual vaccines. Overall, this study demonstrated that these DNA prime/VLP boost vaccine regimens are highly immunogenic in baboons, inducing high-magnitude and broad multifunctional responses, providing support for the development of these products for clinical trials.

Robust Immunity to an Auxotrophic Mycobacterium bovis BCG-VLP Prime-Boost HIV Vaccine Candidate in a Nonhuman Primate Model

ABSTRACT We previously reported that a recombinant pantothenate auxotroph of Mycobacterium bovis BCG expressing human immunodeficiency virus type 1 (HIV-1) subtype C Gag (rBCGpan-Gag) efficiently primes the mouse immune system for a boost with a recombinant modified vaccinia virus Ankara (rMVA) vaccine. In this study, we further evaluated the immunogenicity of rBCGpan-Gag in a nonhuman primate model. Two groups of chacma baboons were primed or mock primed twice with either rBCGpan-Gag or a control BCG. Both groups were boosted with HIV-1 Pr55gag virus-like particles (Gag VLPs). The magnitude and breadth of HIV-specific cellular responses were measured using a gamma interferon (IFN-γ) enzyme-linked immunosorbent spot (ELISPOT) assay, and the cytokine profiles and memory phenotypes of T cells were evaluated by polychromatic flow cytometry. Gag-specific responses were detected in all animals after the second inoculation with rBCGpan-Gag. Boosting with Gag VLPs significantly increased the magnitude and breadth of the responses in the baboons that were primed with rBCGpan-Gag. These responses targeted an average of 12 Gag peptides per animal, compared to an average of 3 peptides per animal for the mock-primed controls. Robust responses of Gag-specific polyfunctional T cells capable of simultaneously producing IFN-γ, tumor necrosis alpha (TNF-α), and interleukin-2 (IL-2) were detected in the rBCGpan-Gag-primed animals. Gag-specific memory T cells were skewed toward a central memory phenotype in both CD4+ and CD8+ T cell populations. These data show that the rBCGpan-Gag prime and Gag VLP boost vaccine regimen is highly immunogenic, inducing a broad and polyfunctional central memory T cell response. This report further indicates the feasibility of developing a BCG-based HIV vaccine that is safe for childhood HIV immunization.

Stimulation of rotavirus IgA, IgG and neutralising antibodies in baboon milk by parenteral vaccination

Abstract A rhesus rotavirus vaccine adjuvanted with ISCOMs was injected intramuscularly to 5 pregnant baboons, with repeated doses 1–2 and 14 weeks after delivery. Maternal blood and milk samples and blood samples from their babies were collected at 2-weekly intervals until 26 weeks after parturition. Samples were assayed for rotavirus antibodies by ELISAs and neutralisation tests. Vaccination produced statistically significant increases in maternal serum IgG and neutralising antibodies, and in milk IgA, IgG, and neutralising antibodies. Control baboon mothers sampled from 12 weeks after delivery had lower serum and milk antibody titres, but responded to vaccination at 16 weeks by producing a similar antibody profile in serum and milk to those previously vaccinated. Because of the endemic nature of human rotaviral infections, similar maternal vaccinations have potential as a means of increasing milk antibodies to a level at which they may be protective to infants.

Detection of natural infection with Mycobacterium intracellulare in healthy wild-caught Chacma baboons (Papio ursinus) by ESAT-6 and CFP-10 IFN-γ ELISPOT tests following a tuberculosis outbreak

BackgroundBoth tuberculous and non-tuberculous mycobacteria can cause infection in nonhuman primates (NHP), indicating the existence of potential zoonotic transmission between these animals and visitors to zoos or animal handlers in primate facilities. Screening of mycobacterial infections in NHP is traditionally done by tuberculin skin test (TST), which is unable to distinguish between pathogenic and non-pathogenic mycobacterial infections. In this study, we investigated the use of ESAT-6 and CFP-10 for detection of mycobacterial infections in a wild-caught baboon colony after one baboon died of tuberculosis (TB).MethodsPeripheral blood lymphocytes for interferon-gamma enzyme-linked immunospot assay (IFN-γ ELISPOT) assay were obtained from TST positive baboons and those in contact with tuberculous baboons before being euthanased, autopsied and lung tissues taken for histology and mycobacterial culture.ResultsBoth ESAT-6 and CFP-10 IFN-γ ELISPOT assays were able to detect early M. tuberculosis but also M. intracellulare infection. Although this indicates potential cross-reactivity with M. intracellulare antigens, the method was able to distinguish M. bovis BCG vaccination from M. tuberculosis infection. This assay performed better than the TST, which failed to detect one M. tuberculosis and two early M. intracellulare infections.ConclusionThese results suggest that the IFN-γ ELISPOT assay could improve the detection of M tuberculosis infections when screening NHP. There is some doubt, however, concerning specificity, as the assay scored positive three animals infected with M. intracellulare.

The novel capripoxvirus vector lumpy skin disease virus efficiently boosts modified vaccinia Ankara human immunodeficiency virus responses in rhesus macaques.

Poxvirus vectors represent promising human immunodeficiency virus (HIV) vaccine candidates and were a component of the only successful HIV vaccine efficacy trial to date. We tested the immunogenicity of a novel recombinant capripoxvirus vector, lumpy skin disease virus (LSDV), in combination with modified vaccinia Ankara (MVA), both expressing genes from HIV-1. Here, we demonstrated that the combination regimen was immunogenic in rhesus macaques, inducing high-magnitude, broad and balanced CD4(+) and CD8(+) T-cell responses, and transient activation of the immune response. These studies support further development of LSDV as a vaccine vector.

HIV-1 Subtype C Mosaic Gag Expressed by BCG and MVA Elicits Persistent Effector T Cell Responses in a Prime-Boost Regimen in Mice.

Over 90% of HIV/AIDS positive individuals in sub-Saharan Africa are infected with highly heterogeneous HIV-1 subtype C (HIV-1C) viruses. One of the best ways to reduce the burden of this disease is the development of an affordable and effective prophylactic vaccine. Mosaic immunogens are computationally designed to overcome the hurdle of HIV diversity by maximizing the expression of potential T cell epitopes. Mycobacterium bovis BCG ΔpanCD auxotroph and modified vaccinia Ankara (MVA) vaccines expressing HIV-1C mosaic Gag (GagM) were tested in a prime-boost regimen to demonstrate immunogenicity in a mouse study. The BCG-GagM vaccine was stable and persisted 11.5 weeks post vaccination in BALB/c mice. Priming with BCG-GagM and boosting with MVA-GagM elicited higher Gag-specific IFN-γ ELISPOT responses than the BCG-GagM only and MVA-GagM only homologous vaccination regimens. The heterologous vaccination also generated a more balanced and persistent CD4+ and CD8+ T cell Gag-specific IFN-γ ELISPOT response with a predominant effector memory phenotype. A Th1 bias was induced by the vaccines as determined by the predominant secretion of IFN-γ, TNF-α, and IL-2. This study shows that a low dose of MVA (104 pfu) can effectively boost a BCG prime expressing the same mosaic immunogen, generating strong, cellular immune responses against Gag in mice. Our data warrants further evaluation in non-human primates. A low dose vaccine would be an advantage in the resource limited countries of sub-Saharan Africa and India (where the predominating virus is HIV-1 subtype C).

DNA-MVA-protein vaccination of rhesus macaques induces HIV-specific immunity in mucosal-associated lymph nodes and functional antibodies

Successful future HIV vaccines are expected to generate an effective cellular and humoral response against the virus in both the peripheral blood and mucosal compartments. We previously reported the development of DNA-C and MVA-C vaccines based on HIV-1 subtype C and demonstrated their immunogenicity when given in a DNA prime-MVA boost combination in a nonhuman primate model. In the current study, rhesus macaques previously vaccinated with a DNA-C and MVA-C vaccine regimen were re-vaccinated 3.5 years later with MVA-C followed by a protein vaccine based on HIV-1 subtype C envelope formulated with MF59 adjuvant (gp140Env/MF59), and finally a concurrent boost with both vaccines. A single MVA-C re-vaccination elicited T cell responses in all animals similar to previous peak responses, with 4/7 demonstrating responses >1000 SFU/106 PBMC. In contrast to an Env/MF59-only vaccine, concurrent boosting with MVA-C and Env/MF59 induced HIV-specific cellular responses in multiple mucosal associated lymph nodes in 6/7 animals, with high magnitude responses in some animals. Both vaccine regimens induced high titer Env-specific antibodies with ADCC activity, as well as neutralization of Tier 1 viruses and modest Tier 2 neutralization. These data demonstrate the feasibility of inducing HIV-specific immunity in the blood and mucosal sites of viral entry by means of DNA and poxvirus-vectored vaccines, in combination with a HIV envelope-based protein vaccine.