Associate Professor
Godwin Nchinda
Project Title
Pre-clinical evaluation of dendritic cell targeted consensus B, C, CRFO2_AG and MOSAIC HIV gag protein vaccines in PBMC from chronically infected patients in Central Africa
EDCTP Project
TA.2010.40200.016
EDCTP Program
EDCTP1
EDCTP Project Call
Senior Fellowship (SF)
Project Objectives
To examine if a DC targeted consensus B HIV gag p24 protein vaccine could recall in vitro pre-existing gag specific T cells in PBMCs of subjects chronically infected with unrelated HIV-1 strains prevalent in Africa To compare in terms of magnitude, breadth, and depth T cell responses recalled in vitro in PBMCs of subjects chronically infected with HIV-1 in central Africa by 4 four different DC targeted HIV gag p24 protein vaccines based on CRFO2_AG, C, B and MOSAIC HIV gag sequences, which are designed to address the problems associated with HIV-1 diversity.
Study Design
Laboratory pre-clinical evaluation of dendritic cell antigens and HIV gag protein vaccines
Host Organisation
| Department | Institution | Country |
|---|---|---|
| Vaccinology/Biobanking Lab | CIRCB | Cameroon |
| Immunology Laboratory | CIRCB | Cameroon |
| Microbiology and Immunology Laboratroy | CIRCB | Cameroon |
Results & Outcomes
Current Organisation
The chantal Biya international reference center for HIV
Current Job Title
Deputy Director General
Biography
Publications
Viral vaccine vectors have emerged as an attractive strategy for the development of a human immunodeficiency
virus (HIV) vaccine. Recombinant Newcastle disease virus (rNDV) stands out as a vaccine vector since
it has a proven safety profile in humans, it is a potent inducer of both alpha interferon (IFN-) and IFN-)
production, and it is a potent inducer of dendritic cell (DC) maturation. Our group has previously generated
an rNDV vector expressing a codon-optimized HIV Gag protein and demonstrated its ability to induce a
Gag-specific CD8 T cell response in mice. In this report we demonstrate that the Gag-specific immune
response can be further enhanced by the targeting of the rNDV-encoded HIV Gag antigen to DCs. Targeting
of the HIV Gag antigen was achieved by the addition of a single-chain Fv (scFv) antibody specific for the
DC-restricted antigen uptake receptor DEC205 such that the DEC205 scFv-Gag molecule was encoded for
expression as a fusion protein. The vaccination of mice with rNDV coding for the DC-targeted Gag antigen
induced an enhanced Gag-specific CD8 T cell response and enhanced numbers of CD4 T cells and CD8 T
cells in the spleen relative to vaccination with rNDV coding for a nontargeted Gag antigen. Importantly, mice
vaccinated with the DEC205-targeted vaccine were better protected from challenge with a recombinant vaccinia
virus expressing the HIV Gag protein. Here we demonstrate that the targeting of the HIV Gag antigen to DCs
via the DEC205 receptor enhances the ability of an rNDV vector to induce a potent antigen-specific immune
response.
BACKGROUND:
Targeting antigens encoded by DNA vaccines to dendritic cells (DCs) in the presence of adjuvants enhances their immunogenicity and efficacy in mice.
METHODOLOGY/PRINCIPAL FINDINGS:
To explore the immunogenicity of this approach in non-human primates, we generated a single chain antibody to the antigen uptake receptor DEC-205 expressed on rhesus macaque DCs. DNA vaccines encoding this single chain antibody fused to the SIV capsid protein were delivered to six monkeys each by either intramuscular electroporation or conventional intramuscular injection co-injected or not with poly ICLC, a stabilized poly I: C analogue, as adjuvant. Antibodies to capsid were induced by the DC-targeting and non-targeting control DNA delivered by electroporation while conventional DNA immunization at a 10-fold higher dose of DNA failed to induce detectable humoral immune responses. Substantial cellular immune responses were also observed after DNA electroporation of both DNAs, but stronger responses were induced by the non-targeting vaccine. Conventional immunization with the DC-targeting DNA at a 10-fold higher dose did not give rise to substantial cellular immune responses, neither when co-injected with poly ICLC.
CONCLUSIONS/SIGNIFICANCE:
The study confirms the potent immunogenicity of DNA vaccines delivered by electroporation. Targeting the DNA via a single chain antibody to DEC-205 expressed by DCs, however, does not improve the immunogenicity of the antigens in non-human primates.
Abstract
Background:
In sub-Saharan Africa, intense perennial Plasmodium species transmission coincides with areas of high prevalence of the human immunodeficiency virus type 1 (HIV) infection. This implies that antiretroviral naïve HIV-infected people living within these regions are repeatedly exposed to Plasmodium species infection and consequently malaria. Natural killer (NK) cells are known to contribute to malaria immunity through the production of IFN-γafter exposure to Plasmodium falciparum-infected erythrocytes (infected red blood cells [iRBC]). However, in antiretroviral naïve HIV-1 infection, these functions could be impaired. In this study we assess the ability of NK cells from antiretroviral naïve HIV-1-infected people to respond to iRBC.
Method: Magnetically sorted NK cells from antiretroviral naïve HIV-1-infected people were tested for their ability to respond to iRBC following in vitro coculture. NK cell IFN-γ production after coculture was measured through multiparametric flow cytometry analysis.
Results:
Our data show a significant reduction ( p = 0.03) in IFN-γ production by NK
cells from antiretroviral naïve HIV-1-infected people after coculture with iRBCs. This was in
contrast to the NK cell response from healthy controls, which demonstrated elevated IFN-γ
production. NK cell IFN-γ production from untreated HIV-1-infected participants correlated
inversely with the viral load ( r = –0.5, p = 0.02) and positively with total helper CD4 + T-cell
count ( r = 0.4, p = 0.04). Thus, antiretroviral naïve HIV-1 infection can dampen NK cell-mediated
immunity to P. falciparum infection in malaria-intense regions. This could in effect escalate
morbidity and mortality in people chronically infected with HIV-1.
Dengue fever is regarded as the most important arboviral disease worldwide. This study was designed to investigate the prevalence of denque virus seropositivity among children with febrile illness at the Nnamdi Azikiwe University Teaching Hospital, Nnewi, Anambra State, Nigeria. A total of 96 subjects were recruited for the study. The demographic data and dietary pattern of the subjects was obtained using well structured questionnaire. Dengue Virus IgM was analysed using ELISA techniques. The results showed a prevalence of 77.1% Dengue Virus seropositivity among children with febrile illness in the study population with a greater prevalence found among the males (54.1%) than female (45.9%) subjects. The dengue virus seropositive participants had significantly greater IgM levels compared with the seronegative participants (2.02 ± 0.76 vs 0.84 ± 0.28; p < 0.001). Children who had dengue virus had significantly greater WBC (p = 0.017), esinophil (p<0.001), but lower RBC (<0.001), Hb (p = 0.001) and basophil (p = 0.001) compared with dengue virus negative children. The result showed a strong positive association between anemia and dengue viral status (χ2, 6.31; p = 0.012) with dengue virus seropositive partcipants at greater risk (OR, 3.52; p = 0.015) of developing anemia compared to those who had no dengue virus. More so, the incidence of anemia was higher in those who had malaria and dengue virus co-infection (86.8%) compared with those who had no malaria and were Dengue virus negative (7.5%) and those who presented with malaria but had no Dengue virus (5.7%). Furthermore, the result indicates a significant association (p = 0.005) between dengue virus seropositivity and malaria co-infection with anemia. Our report has revealed that dengue virus may is a major cause of fever among the study population. This calls for urgent attention and large scale research to confirm the circulating strains of the dengue virus.
Abstract
INTRODUCTION:
Recombinant Newcastle Disease virus (rNDV) vectored vaccines are safe mucosal applicable vaccines with intrinsic immune-modulatory properties for the induction of efficient immunity. Like all viral vectored vaccines repeated inoculation via mucosal routes invariably results to immunity against viral vaccine vectors. To obviate immunity against viral vaccine vectors and improve the ability of rNDV vectored vaccines in inducing T cell immunity in murine air way we have directed dendritic cell targeted HIV-1 gag protein (DEC-Gag) vaccine; for the induction of helper CD4+ T cells to a Recombinant Newcastle disease virus expressing codon optimized HIV-1 Gag P55 (rNDV-L-Gag) vaccine.
METHODS:
We do so through successive administration of anti-DEC205-gagP24 protein plus polyICLC (DEC-Gag) vaccine and rNDV-L-Gag. First strong gag specific helper CD4+ T cells are induced in mice by selected targeting of anti-DEC205-gagP24 protein vaccine to dendritic cells (DC) in situ together with polyICLC as adjuvant. This targeting helped T cell immunity develop to a subsequent rNDV-L-Gag vaccine and improved both systemic and mucosal gag specific immunity.
RESULTS:
This sequential DEC-Gag vaccine prime followed by an rNDV-L-gag boost results to improved viral vectored immunization in murine airway, including mobilization of protective CD8+ T cells to a pathogenic virus infection site.
CONCLUSION:
Thus, complementary prime boost vaccination, in which prime and boost favor distinct types of T cell immunity, improves viral vectored immunization, including mobilization of protective CD8+ T cells to a pathogenic virus infection site such as the murine airway.
Summary
Regulatory T (Treg) cells play a key role in dampening excessive immune
activation. However, antiretroviral therapy (ART) -naive HIV-1 infection
maintains the immune system in a sustained state of activation that could
alter both Treg cell surface markers and functions. As Treg cell surface markers
are directly linked to their functions the overall objective of this study
was to determine how ART-naive HIV infection affects the phenotypic properties
of Treg cells. Our data showed that in ART-naive HIV-1 infection,
Treg cells are dominated by effector (CD45RA+ CD27
CCR7
CD62L
)
and effector memory (CD45RA
CD27
CCR7
CD62L
) cells. In contrast
Treg cells from HIV-negative individuals were mainly naive
(CD45RA+ CD27+ CCR7+ CD62L+) and central memory (CD45RA
CD27+ CCR7+ CD62L+) cells. Whereas effector and effector memory Treg
cells showed enhanced expression of CD39 (P < 005), CD73 (P < 0001),
HLA-DR and CD38 (P < 0001); naive and central memory Treg cells
showed a significant reduction in the expression of these markers. Overall
Treg cell frequencies within total CD4+ T cells correlated positively with
plasmatic HIV-1 viral load. As increased viral load is associated with augmented
CD4+ T-cell destruction; this could suggest a resistance of peripheral
Treg cells to HIV-1 destruction. Hence the modulation of Treg cell phenotype
and frequencies could be considered in designing immunotherapeutic
strategies targeting immune system restoration during HIV-1 infection.
Recent outbreaks of Ebola virus disease and Zika virus disease highlight the need for disseminating accurate predictions of emerging zoonotic viruses to national governments for disease surveillance and response. Although there are published maps for many emerging zoonotic viruses, it is unknown if there is agreement among different models or if they are concordant with national expert opinion. Therefore, we reviewed existing predictions for five high priority emerging zoonotic viruses with national experts in Cameroon to investigate these issues and determine how to make predictions more useful for national policymakers. Predictive maps relied primarily on environmental parameters and species distribution models. Rift Valley fever virus and Crimean-Congo hemorrhagic fever virus predictions differed from national expert opinion, potentially because of local livestock movements. Our findings reveal that involving national experts could elicit additional data to improve predictions of emerging pathogens as well as help repackage predictions for policymakers.
Abstract
Although human immunodeficiency type 1 (HIV-1) infection induces strong antibody responses to the viral envelope glycoprotein (Env) only a few of these antibodies possess the capacity to neutralize a broad range of strains. The induction of such antibodies represents an important goal in the development of a preventive vaccine against the infection. Among the broadly neutralizing monoclonal antibodies discovered so far, three (2F5, Z13 and 4E10) target the short and hidden membrane proximal external region (MPER) of the gp41 transmembrane protein. Antibody responses to MPER are rarely observed in HIV-infected individuals or after immunization with Env immunogens. To initiate antibody responses to MPER in its membrane-embedded native conformation, we generated expression plasmids encoding the membrane-anchored ectodomain of gp41 with N-terminal deletions of various sizes. Following transfection of these plasmids, the MPER domains are displayed on the cell surface and incorporated into HIV virus like particles (VLP). Transfected cells displaying MPER mutants bound as efficiently to both 2F5 and 4E10 as cells transfected with a plasmid encoding full-length Env. Mice immunized with VLPs containing the MPER mutants produced MPER-specific antibodies, the levels of which could be increased by the trimerization of the displayed proteins as well as by a DNA prime-VLP boost immunization strategy. Although 2F5 competed for binding to MPER with antibodies in sera of some of the immunized mice, neutralizing activity could not be detected. Whether this is due to inefficient binding of the induced antibodies to MPER in the context of wild type Env or whether the overall MPER-specific antibody response induced by the MPER display mutants is too low to reveal neutralizing activity, remains to be determined.
Abstract Introduction: The membrane proximal external region (MPER) of HIV-1 envelope glycoprotein-41 (gp41) is targeted by several broadly neutralizing antibodies whose conserved linear epitopes are promising targets for vaccine design. However, a formidable challenge has remained the difficulty to design and deliver MPER based immunogens for the efficient induction of such broadly neutralizing HIV-1 specific antibodies (bnAb). This is mainly because the linear bnAb MPER epitopes are poorly accessible to the immune system. The overall objective of this study therefore was the development of a novel RNA Qβ phage display system not only for monitoring anti-MPER specific antibody responses but equally as potential immunogens in future HIV-1 vaccine designs. Method: To overcome the challenge of effective presentation of MPER to the immune system we have selectively engineered the surface of the RNA coliphage Qβ to expose all MPER bnAb epitopes. Briefly, DNA representing a 50 amino acids consensus region within the HIV-1 gp41 MPER was fused in frame with the minor coat protein A1of the Qβ phage. Three variant MPER expression cassettes were obtained with the MPER cDNA in frame with the minor coat protein A1 gene, including pQβMPER, pQβMPERHis and pQβMPERN. The expression cassettes were used for the production of QβMPER recombinant phages after transformation of E. coli HB101 strain. Antigencity of the phages was assessed with plasma from long standing anti-retroviral naïve HIV-1 infected people from the CIRCB AFRODEC cohort while immunogenicity studies were done in female Balb/c mice. Results: The initial titers of all recombinant phages including QβMPER, QβMPERHis and QβMPERN were 104 plaque forming units/ml (pfu/ml). This was significantly lower (P<0.001) relative to the 108 pfu/ml of wild type phage, but was scaled up to 1014pfu/ml. The fusion of MPER and Qβ genes was confirmed by RT-PCR followed by gel electrophoresis and sequencing. Specific recognition of some reported bnAb epitopes within MPER were confirmed in ELISA using the three recombinant QβMPER phages together with an MPER restrictive peptide as antigens and the bnAb 4E10, Z13e1, 2F5 and 10E8 as antibodies. Next the prevalence of MPER-specific antibodies was determined in plasma from long standing antiretroviral naïve HIV-1 infected participants of the CIRCB AFRODEC cohort. The greater majority (84%) of participants’ plasma showed MPER peptide specific reactivity with anti-MPER specific IgG antibody titers ranging from 200 to 409600 comparative to background IgG antibody titer with the Qβ phage alone as antigen or plasma from seronegative participants. In immunogenicity studies in Balb/c mice the recombinant phages QβMPERN and QβMPERHis induced significantly high Anti-MPER-specific IgG antibody responses (P<0.04) in at least 60 % of mice following three inoculations of each recombinant phage. Conclusion: Thus, these novel recombinantQβMPER phages can be used to monitor MPER-specific immune responses in HIV-1 exposed or infected people. In addition the recombinant QβMPER phages could be used as immunogens either alone as demonstrated here in mice or in combination with other strategies for the induction of MPER specific immunity against HIV-1
Abstract
Background.
Severe adverse reactions have been observed in individuals with Loa loainfection treated with either diethylcarbamazine (DEC), the drug of choice for loiasis, or ivermectin (IVM), which is used in mass drug administration programs for control of onchocerciasis and lymphatic filariasis in Africa. In this study, posttreatment clinical and immunologic reactions were compared following single-dose therapy with DEC or IVM to assess whether these reactions have the same underlying pathophysiology.
Methods.
Twelve patients with loiasis and microfilarial counts <2000 mf/mL were randomized to receive single-dose DEC (8 mg/kg) or IVM (200 µg/kg). Clinical and laboratory assessments were performed at 4, 8, 24, 48, and 72 hours and 5, 7, 9, and 14 days posttreatment.
Results.
Posttreatment adverse events were similar following DEC or IVM, but peaked earlier in subjects who received DEC, consistent with a trend toward more rapid and complete microfilarial clearance in the DEC group. After a transient rise (post-IVM) or fall (post-DEC) in the first 24 hours posttreatment, the eosinophil count rose significantly in both groups, peaking at day 5 in the DEC group and day 9 in the IVM group. Serum interleukin 5 levels and eosinophil activation, as assessed by surface expression of CD69 and serum levels of eosinophil granule proteins, were increased posttreatment in both groups.
Conclusions.
Despite differences in eosinophil and lymphocyte counts during the first 24 hours posttreatment, the overall pattern of hematologic and immunologic changes suggest that posttreatment reactions following DEC and IVM share a common pathophysiology.
Background
Dengue is a major public health problem worldwide. Assessment of adaptive immunity is important to understanding immunopathology and to define correlates of protection against dengue virus (DENV). To enable global assessment of CD4+ T cell responses, we mapped HLA-DRB1-restricted DENV-specific CD4+ T cell epitopes in individuals previously exposed to DENV in the general population of the dengue-endemic region of Managua, Nicaragua.
Methods
HLA class II epitopes in the population of Managua were identified by an in vitro IFNγ ELISPOT assay. CD4+ T cells purified by magnetic bead negative selection were stimulated with HLA-matched epitope pools in the presence of autologous antigen-presenting cells, followed by pool deconvolution to identify specific epitopes. The epitopes identified in this study were combined with those previously identified in the DENV endemic region of Sri Lanka, to generate a “megapool” (MP) consisting of 180 peptides specifically designed to achieve balanced HLA and DENV serotype coverage. The DENV CD4MP180 was validated by intracellular cytokine staining assays.
Results
We detected responses directed against a total of 431 epitopes, representing all 4 DENV serotypes, restricted by 15 different HLA-DRB1 alleles. The responses were associated with a similar pattern of protein immunodominance, overall higher magnitude of responses, as compared to what was observed previously in the Sri Lanka region. Based on these epitope mapping studies, we designed a DENV CD4 MP180 with higher and more consistent coverage, which allowed the detection of CD4+ T cell DENV responses ex vivo in various cohorts of DENV exposed donors worldwide, including donors from Nicaragua, Brazil, Singapore, Sri Lanka, and U.S. domestic flavivirus-naïve subjects immunized with Tetravalent Dengue Live-Attenuated Vaccine (TV005). This broad reactivity reflects that the 21 HLA-DRB1 alleles analyzed in this and previous studies account for more than 80% of alleles present with a phenotypic frequency ≥5% worldwide, corresponding to 92% phenotypic coverage of the general population (i.e., 92% of individuals express at least one of these alleles).
Conclusion
The DENV CD4 MP180 can be utilized to measure ex vivo responses to DENV irrespective of geographical location.
Abstract
INTRODUCTION:
Targeting antigens to dendritic cells (DCs) in vivo via a DC-restricted endocytic receptor, DEC205, has been validated to enhance immunity in several vaccine platforms. Particularly atttractive is selected delivery of proteins to DCs in vivo because it enables proteins to be more immunogenic and provides a cheaper and effective way for repeated immunizations.
METHODS:
In this study, we tested the efficacy of a single chain antibody to DEC205 (scDEC) to deliver protein antigens selectively to DCs in vivo and to induce protective immunity.
RESULTS:
In comparison to soluble Ovalbumin (OVA) antigen, when recombinant scDEC:OVA protein was injected subcutaneously (s.c.) into mice, the OVA protein was selectively presented by DCs to both TCR transgenic CD8+ and CD4+ T cells approximately 500 and 100 times more efficient than soluble OVA, respectively, and could persist for seven days following s.c. injection of the scDEC205:OVA. Similarly selective targeting of HIV Gag P24 to DCs in vivo using scDEC-Gag protein plus polyICLC vaccine resulted in strong, long lasting, polyfuntional CD4+ T cells in mice which were protective against airway challenge by a recombinant vaccinia-gag virus.
CONCLUSION:
Thus targeting protein antigens to DCs using scDEC can be used either alone or in combination with other strategies for effective immunization.
Replication-defective adenoviral vectors have emerged as promising vaccine candidates for diseases relying on strong CD8 + T-cell responses for protection. In this study, we modified a non-replicative adenoviral vector to selectively deliver, in situ, an encoded ovalbumin (OVA) model antigen to dendritic cells (DCs). Efficient uptake and presentation of OVA was achieved through fusion of the antigen to a single-chain antibody directed against DEC205, an endocytic receptor expressed on DCs. The immunogenicity of the vaccine was thereby enhanced as demonstrated by elevated antibody levels and increased T-cell responses after low-dose vaccination with 10 7 viral particles compared with a non-targeted control. Nevertheless, after immunization with higher doses of the targeted vaccine, the capacity of vaccine-induced CD8 + T cells to produce the cytokine IL-2 was diminished and the CD8 + T-cell response was dominated by an effector memory phenotype (CD62L – /CD127 + ) in contrast to the effector phenotype (CD62L – /CD127 – ) observed after non-targeted antigen delivery. Interestingly, the protective capacity of the non-targeted vaccine was superior to that of the targeted vaccine in an antigen-specific vaccinia virus infection as well as in a tumor challenge model. In the latter, the low dose of the DC-targeted vaccine also conferred partial protection from tumor growth, demonstrating dose-dependent effects of the DC-targeting on the quality of the vaccine-induced immune response. Significant differences could be observed in regard to the antibody pattern, the functional and phenotypic T-cell repertoire, and to the protective capacity.