Safety and immunogenicity of an FP9-vectored candidate tuberculosis vaccine (FP85A), alone and with candidate vaccine MVA85A in BCG-vaccinated healthy adults: a phase I clinical trial.

The safety and immunogenicity of a new candidate tuberculosis (TB) vaccine, FP85A was evaluated alone and in heterologous prime-boost regimes with another candidate TB vaccine, MVA85A. This was an open label, non-controlled, non-randomized Phase I clinical trial. Healthy previously BCG-vaccinated adult subjects were enrolled sequentially into three groups and vaccinated with FP85A alone, or both FP85A and MVA85A, with a four week interval between vaccinations. Passive and active data on adverse events were collected. Immunogenicity was evaluated by Enzyme Linked Immunospot (ELISpot), flow cytometry and Enzyme Linked Immunosorbent assay (ELISA). Most adverse events were mild and there were no vaccine-related serious adverse events. FP85A vaccination did not enhance antigen 85A-specific cellular immunity. When MVA85A vaccination was preceded by FP85A vaccination, cellular immune responses were lower compared with when MVA85A vaccination was the first immunisation. MVA85A vaccination, but not FP85A vaccination, induced anti-MVA IgG antibodies. Both MVA85A and FP85A vaccinations induced anti-FP9 IgG antibodies. In conclusion, FP85A vaccination was well tolerated but did not induce antigen-specific cellular immune responses. We hypothesize that FP85A induced anti-FP9 IgG antibodies with cross-reactivity for MVA85A, which may have mediated inhibition of the immune response to subsequent MVA85A. ClinicalTrials.gov identification number: NCT00653770

A Phase I study evaluating the safety and immunogenicity of MVA85A, a candidate TB vaccine, in HIV-infected adults

Objectives Control of the tuberculosis (TB) epidemic is a global health priority and one that is likely to be achieved only through vaccination. The critical overlap with the HIV epidemic requires any effective TB vaccine regimen to be safe in individuals who are infected with HIV. The objectives of this clinical trial were to evaluate the safety and immunogenicity of a leading candidate TB vaccine, MVA85A, in healthy, HIV-infected adults. Design This was an open-label Phase I trial, performed in 20 healthy HIV-infected, antiretroviral-naïve subjects. Two different doses of MVA85A were each evaluated as a single immunisation in 10 subjects, with 24 weeks of follow-up. The safety of MVA85A was assessed by clinical and laboratory markers, including regular CD4 counts and HIV RNA load measurements. Vaccine immunogenicity was assessed by ex vivo interferon γ (IFN-γ) ELISpot assays and flow-cytometric analysis. Results MVA85A was safe in subjects with HIV infection, with an adverse-event profile comparable with historical data from previous trials in HIV-uninfected subjects. There were no clinically significant vaccine-related changes in CD4 count or HIV RNA load in any subjects, and no evidence from qPCR analyses to indicate that MVA85A vaccination leads to widespread preferential infection of vaccine-induced CD4 T cell populations. Both doses of MVA85A induced an antigen-specific IFN-γ response that was durable for 24 weeks, although of a lesser magnitude compared with historical data from HIV-uninfected subjects. The functional quality of the vaccine-induced T cell response in HIV-infected subjects was remarkably comparable with that observed in healthy HIV-uninfected controls, but less durable. Conclusion MVA85A is safe and immunogenic in healthy adults infected with HIV. Further safety and efficacy evaluation of this candidate vaccine in TB- and HIV-endemic areas is merited

Preclinical Development of an In Vivo BCG Challenge Model for Testing Candidate TB Vaccine Efficacy

There is an urgent need for an immunological correlate of protection against tuberculosis (TB) with which to evaluate candidate TB vaccines in clinical trials. Development of a human challenge model of Mycobacterium tuberculosis (M.tb) could facilitate the detection of such correlate(s). Here we propose a novel in vivo Bacille Calmette-Guérin (BCG) challenge model using BCG immunization as a surrogate for M.tb infection. Culture and quantitative PCR methods have been developed to quantify BCG in the skin, using the mouse ear as a surrogate for human skin. Candidate TB vaccines have been evaluated for their ability to protect against a BCG skin challenge, using this model, and the results indicate that protection against a BCG skin challenge is predictive of BCG vaccine efficacy against aerosol M.tb challenge. Translation of these findings to a human BCG challenge model could enable more rapid assessment and down selection of candidate TB vaccines and ultimately the identification of an immune correlate of protection

Evaluation of the Immunological Mechanisms Induced by Mycobacteria and the Potential Effect this may have on Immunity Induced by Tuberculosis Vaccines

The efficacy of Bacille-Calmette Guerin (BCG) vaccination in protection against pulmonary tuberculosis (TB) is highly variable between populations. One possible explanation is increased exposure of certain populations to non-tuberculous mycobacteria (NTM). Given the variable efficacy of BCG an improved vaccine against TB is required. The novel TB vaccine MVA85A has shown promising results, however, the immunogenicity of the vaccine is reduced when it is administered in the Expanded Programme on Immunisation (EPI) schedule. This thesis aims to explore: (A) the effect of exposure to NTM on the level of protection afforded by BCG vaccination against Mycobacterium tuberculosis (M. tb) and (B) the immunological mechanisms behind EPI interference with MVA85A. The effect of M. avium (MA) exposure via systemic and oral routes on the efficacy of BCG was tested using M. tb aerosol infection in a mouse model. The adaptive immune response was profiled in BCG vaccinated mice with and without exposure to MA pre- and post- M. tb infection. The results showed BCG efficacy could be enhanced by exposure to dead MA by a systemic route; T helper 1 and T helper 17 responses were associated with increased protection. In contrast, BCG efficacy may have been reduced by exposure to live MA by the oral route; T helper 2 and regulatory T cells were associated with reduced protection. To answer the second aim MVA85A was co-administered to mice with aluminium adjuvants or aluminium-containing vaccines to replicate the effect of co-administration in the EPI schedule; the adaptive immune response was profiled. T helper 2 and regulatory T cell responses induced by aluminium-containing vaccines were associated with a reduction in the immunogenicity of MVA85A.</p

Effect of single dose vaccines (subunits MVA85A and Ad85A, and BCG) on an <i>id</i> BCG challenge.

<p>(<b>a</b>) BALB/c mice were immunized <i>id</i> with 1×10⁶ pfu MVA85A or 2×10⁹ vp Ad85A. Control mice (Naïve) received no immunization. Four weeks later all mice were challenged <i>id</i> with 1×10⁵ CFU BCG, contralaterally to the site of vaccination. Ears and LNs were harvested 4 weeks after BCG challenge and processed for CFU quantification. (*<i>P</i><0.05, <i>n</i> = 10 except naïves, <i>n</i> = 5). (<b>b</b>) Corresponding intracellular cytokine staining (ICS) of the local draining LNs. Red bars represent the proportion of IFN-γ-secreting CD4⁺ T cells in response to 85A, blue bars represent the same for CD8⁺ T cells (M, <i>n</i> = 4; Naïve, <i>n</i> = 3; Ad, <i>n</i> = 4. *<i>P</i><0.05). (<b>c</b>) Effect of BCG vaccine compared to subunit MVA85A on an <i>id</i> BCG challenge. BALB/c mice were immunized <i>id</i> with either 1×10⁶ pfu MVA85A or 2.2×10⁴ cfu BCG. “Naïve” and antibiotic-treated (I+R) mice received no immunization. Four weeks later all mice were challenged with 6×10³ CFU BCG, except the BCG control group who received no challenge. In the I+R group, challenge was followed by 4 weeks treatment with isoniazid and rifampicin. Ears and LNs were harvested 4 weeks after BCG challenge and processed for CFU quantification. Log₁₀ BCG CFU individual data points for each mouse are shown. Bars represent the median per group. (<b>c</b>) Ears (**<i>P</i><0.01, “non-significant, ND”, <i>n</i> = 10); (<b>d</b>) LNs (**<i>P</i><0.01, “non-significant, ND”, <i>n</i> = 10).</p

Splenic IFN-γ responses to PPD up to 12 weeks post BCG immunization.

<p>In (<b>a</b>) high dose group (7000 CFU <i>id</i>); (<b>b</b>) mid dose group (60 CFU <i>id</i>) and (<b>c</b>) low dose group (1 CFU <i>id</i>). Datasets include individual data points for each mouse; the bars represent the median value per group. Results expressed as SFC/million splenocytes.</p

Timescale showing BCG persistence in the ears and LNs of <i>id</i>-injected mice up to 12 weeks post immunization.

<p>Log BCG CFU in the ears (estimated by culture) are shown for (<b>a</b>) the high dose group (7000 CFU <i>id</i>); and (<b>b</b>) the low dose group (60 CFU <i>id</i>). Log BCG CFU in the auricular LNs are shown in (<b>c</b>) the high dose group (7000 CFU <i>id</i>); and (<b>d</b>) the mid dose group (60 CFU <i>id</i>). Datasets include individual data points for each mouse; the bars represent the median per group in (<b>a</b>) and (<b>b</b>), and a line connects the means for each group in (<b>c</b>) and (<b>d</b>). * indicates <i>P</i><0.05. Both ears and LNs were homogenized and plated onto 7H11 Middlebrook agar. Log BCG CFU in the ears estimated by culture (<b>e</b>) and BCG genome copies/mHPRT copies estimated by PCR (<b>f</b>) are shown in a timescale for the high dose group, up to 12 weeks post BCG immunization. Quantitative PCR was performed with BCG-specific primers. Individual data points are shown for each mouse with a line connecting the means for each group.</p

Effect of BCG immunization on a 16-week <i>id</i> BCG challenge.

<p>Timeline is shown in (<b>a</b>). BALB/c mice were immunized <i>id</i> with 1×10⁴ CFU BCG. Naïve mice received no immunization. 16 weeks later all mice were challenged with 10³ CFU BCG, except the (BCG+(I+R)(no challenge)) group. This group and the BCG+(I+R) group received 6 weeks of isoniazid and rifampicin (starting 8 weeks post initial BCG immunization). In the BCG+(I+R) group, there was a 2 week wash-out period between cessation of antibiotics and subsequent BCG challenge. (<b>b</b>) shows the splenic IFN-γ ELISpot responses to PPD and TB10.3, in naïve, BCG-vaccinated, and BCG+(I+R)(no challenge) animals, 4 weeks post BCG challenge. Whiskers represent minimum to maximum values, boxes the interquartile range, and the bars the median values for each group (*<i>P</i><0.05, **<i>P</i><0.01, “non-significant”, ND. <i>n</i> = 10, except for BCG+(I+R); <i>n</i> = 2). Ears and LNs were harvested 4 weeks after BCG challenge and processed for CFU quantification. Log₁₀ BCG CFU of challenge are shown, for groups 1 (Naïve), 2 (BCG) and 3 (BCG+(I+R)). For group 4 (BCG+(I+R) no challenge), the ear and LN CFU correspond to the CFU remaining from the priming BCG immunization (zero in all animals). Individual data points for each mouse are shown. Bars represent the median per group. (<b>c</b>) Ears (*<i>P</i><0.05,**<i>P</i><0.01, <i>n</i> = 10); (<b>d</b>) LNs (**<i>P</i><0.01, <i>n</i> = 10).</p

Final sequences of BCG-specific primers, C3/5 and ET1/3, showing base pair differences from the original published sequences underlined.

<p>Final sequences of BCG-specific primers, C3/5 and ET1/3, showing base pair differences from the original published sequences underlined.</p