The status of tuberculosis vaccine development.

Tuberculosis represents the leading global cause of death from an infectious agent. Controlling the tuberculosis epidemic thus represents an urgent global public health priority. Epidemiological modelling suggests that, although drug treatments for tuberculosis continue to improve, WHO timelines to control the spread of the disease require a new vaccine capable of preventing tuberculosis, particularly in adolescents and adults. The spread of strains resistant to multiple drugs adds additional urgency to the vaccine development effort yet attempts to develop new vaccines with wider applicability and better, longer-lasting efficacy than BCG-the only tuberculosis vaccine licensed for use globally-have proven challenging. Results from clinical efficacy trials, particularly a completed, phase 2b trial for preventing tuberculosis disease in people infected with Mycobacterium tuberculosis using the adjuvanted protein subunit vaccine M72/AS01E give hope. We review the current status of tuberculosis vaccine candidates and outline the diversified vaccine development that are underway.

Toward Tuberculosis Vaccine Development: Recommendations for Nonhuman Primate Study Design.

Clinical trials of novel tuberculosis (TB) vaccines are expensive, while global resources for TB vaccine development are limited. Therefore, there is a need for robust and predictive preclinical data to support advancement of candidate vaccines into clinical trials. Here, we provide a rationale for using the nonhuman primate as an essential component of these efforts, as well as guidance to the TB community for standardizing experimental design and aligning endpoints to facilitate development of new TB vaccines.

A temperature sensitive Mycobacterium paragordonae induces enhanced protective immune responses against mycobacterial infections in the mouse model.

Recently, we introduced a temperature sensitive Mycobacterium spp., Mycobacterium paragordonae (Mpg). Here, we checked its potential as a candidate for live vaccination against Mycobacterium tuberculosis and Mycobacterium abscessus. Intravenous infections of mice with Mpg led to lower colony forming units (CFUs) compared to infection with BCG, suggesting its usefulness as a live vaccine. The analyses of immune responses indicated that the highly protective immunity elicited by Mpg was dependent on effective dendritic maturation, shift of cytokine patterns and antibody production toward a Th1 phenotype, and enhanced cytotoxic T cell response. Compared to BCG, Mpg showed a more effective protective immune response in the vaccinated mice against challenges with 2 different mycobacterial strains, M. tuberculosis H37Ra or M. abscessus Asan 50594. Our data suggest that a temperature sensitive Mpg may be a potentially powerful candidate vaccine strain to induce enhanced protective immune responses against M. tuberculosis and M. abscessus.

MTBVAC from discovery to clinical trials in tuberculosis-endemic countries.

INTRODUCTION: BCG remains the only vaccine against tuberculosis (TB) in use today and despite its impressive global coverage, the nature of BCG protection against the pulmonary forms of TB remains subject to ongoing debate. Because of the limitations of BCG, novel TB vaccine candidates have been developed and several have reached the clinical pipeline. One of these candidates is MTBVAC, the first and only TB vaccine in the clinical pipeline to date based on live-attenuated Mycobacterium tuberculosis that has successfully entered clinical evaluation, a historic milestone in human vaccinology. Areas covered: This review describes development of MTBVAC from discovery to clinical development in high burden TB-endemic countries. The preclinical experiments where MTBVAC has shown to confer improved safety and efficacy over BCG are presented and the clinical development plans for MTBVAC are revealed. The search of all supportive literature in this manuscript was carried out via Pubmed. Expert commentary: Small experimental medicine trials in humans and preclinical efficacy studies with a strong immunological component mimicking clinical trial design are considered essential by the scientific community to help identify reliable vaccine-specific correlates of protection in order to support and accelerate community-wide efficacy trials of new TB vaccines.

Protective efficacy of a lipid antigen vaccine in a guinea pig model of tuberculosis.

The bacillus Calmette Guérin (BCG) vaccine, the only licensed vaccine against TB, displays partial and variable efficacy, thus making the exploitation of novel vaccination strategies a major priority. Most of the current vaccines in pre-clinical or clinical development are based on the induction of T cells recognizing protein antigens. However, a large number of T cells specific for mycobacterial lipids are induced during infection, suggesting that lipid-based vaccines might represent an important component of novel sub-unit vaccines. Here, we investigated whether immunization with defined mycobacterial lipid antigens induces protection in guinea pigs challenged with M. tuberculosis. Two purified mycobacterial lipid antigens, the diacylated sulfoglycolipids (Ac2SGL) and the phosphatidyl-myo-inositol dimannosides (PIM2) were formulated in biophysically characterized liposomes made of dimethyl-dioctadecyl-ammonium (DDA) and synthetic trehalose 6,6'-dibehenate (TDB). In three protection trials, a reduction of bacterial load in the spleen of inoculated animals was consistently observed compared to the unvaccinated group. Moreover, a reduction in the number of lesions and severity of pathology was detected in the lungs and spleen of the lipid vaccine group compared to unvaccinated controls. As the degree of protection achieved is similar to that observed using protein antigens in the same guinea pig model, these promising results pave the way to future investigations of lipid antigens as subunit vaccines.

Immunogencity of antigens from Mycobacterium tuberculosis self-assembled as particulate vaccines.

Traditional approaches to vaccine development have failed to identify better vaccines to replace or supplement BCG for the control of tuberculosis (TB). Subunit vaccines offer a safer and more reproducible alternative for the prevention of diseases. In this study, the immunogenicity of bacterially derived polyester beads displaying three different Rv antigens of Mycobacterium tuberculosis was evaluated. Polyester beads displaying the antigens Rv1626, Rv2032, Rv1789, respectively, were produced in an endotoxin-free Escherichia coli strain. Beads were formulated with the adjuvant DDA and subcutaneously administered to C57BL/6 mice. Cytokine responses were evaluated by CBA and antibody responses by ELISA. Specificity of the IgG response was assessed by immunoblotting cell lysates of the vaccine production strains using sera from the vaccinated mice. Mice vaccinated with beads displaying Rv1626 had significantly greater IgG1 responses compared to mice vaccinated with Rv1789 beads and greater IgG2 responses than the group vaccinated with Rv2032 beads (p<0.05). Immunoblotting of antisera from these mice indicated the antibody responses were Rv1626 antigen-specific and there was no detectable immune response to the polyester component of the vaccine. Overall, this study suggested that selected TB antigens derived from reverse vaccinology approaches can be displayed on polyester beads to produce antigen-specific immune responses potentially relevant to the prevention of TB.

Experimental Infection Models of Tuberculosis in Domestic Livestock.

In this article we present experimental Mycobacterium bovis infection models in domestic livestock species and how these models were applied to vaccine development, biomarker discovery, and the definition of specific antigens for the differential diagnosis of infected and vaccinated animals. In particular, we highlight synergies between human and bovine tuberculosis (TB) research approaches and data and propose that the application of bovine TB models could make a valuable contribution to human TB vaccine research and that close alignment of both research programs in a one health philosophy will lead to mutual and substantial benefits.

A novel multivalent tuberculosis vaccine confers protection in a mouse model of tuberculosis.

Mycobacterium tuberculosis infects one third of the world's population. Due to variable efficacy of the Bacille Calmette Guerin (BCG) vaccine, development of novel TB vaccines remains a priority. Here, we demonstrate the protective efficacy of a novel multivalent DNA vaccine, which contains 15 synthetic antigens targeting the Mtb ESX secretion system.

Why don't we have an effective tuberculosis vaccine yet?

Mycobacterium tuberculosis (M.tb) has co-evolved with humans for thousands of years, to cause tuberculosis (TB). The success of M.tb as a pathogen is in part because of the ways in which M.tb evades and exploits different cell subsets, to persist and cause disease. M.tb expresses numerous molecules to prevent its recognition and destruction by immune cells. The only licensed vaccine against TB, Bacillle Calmette-Guerin (BCG), is effective at preventing disseminated disease in infants but confers highly variable efficacy against pulmonary TB in adults, particularly in the developing world. A greater understanding of the reasons for this variability, together with a better understanding of the early, innate, and non-antigen specific mechanisms of protection would facilitate the design and development of more effective vaccines.

Complement component 3: a new paradigm in tuberculosis vaccine.

Vaccines are critical for the control of tuberculosis (TB) affecting humans and animals worldwide. First-generation vaccines protect from active TB but new vaccines are required to protect against pulmonary disease and infection. Recent advances in post-genomics technologies have allowed the characterization of host-pathogen interactions to discover new protective antigens and mechanisms to develop more effective vaccines against TB. Studies in the wild boar model resulted in the identification of complement component 3 (C3) as a natural correlate of protection against TB. Oral immunization with heat-inactivated mycobacteria protected wild boar against TB and showed that C3 plays a central role in protection. These results point at C3 as a target to develop novel vaccine formulations for more effective protection against TB in humans and animals.

Evolutionary landscape of the Mycobacterium tuberculosis complex from the viewpoint of PhoPR: implications for virulence regulation and application to vaccine development.

Different members of the Mycobacterium genus have evolved to cause tuberculosis in diverse human populations and in a variety of animal species. Our cumulative knowledge of mycobacterial genomes indicates that mutations in the PhoPR two-component virulence system were acquired not only during the natural evolution of mycobacterial species but also during in vitro subculture, which has given rise to the attenuated reference strain H37Ra or to different daughter strains of Mycobacterium bovis BCG. PhoPR is a well-known regulator of pathogenic phenotypes, including secretion of the virulence factor ESAT-6, biosynthesis of acyltrehalose-based lipids, and modulation of antigen export, in members of the Mycobacterium tuberculosis complex (MTBC). Evolutionarily conserved polymorphisms in PhoPR from Mycobacterium africanum, M. bovis, or M. tuberculosis H37Ra result in loss of functional phenotypes. Interestingly, some members of the MTBC have acquired compensatory mutations to counteract these polymorphisms and, probably, to maintain their pathogenic potential. Some of these compensatory mutations include the insertion of the IS6110 element upstream from phoPR in a particular M. bovis strain that is able to transmit between humans or polymorphisms in M. africanum and M. bovis that affect the regulatory region of the espACD operon, allowing PhoPR-independent ESAT-6 secretion. This review highlights the increasing knowledge of the significance of PhoPR in the evolution of the MTBC and its potential application in the construction of new attenuated vaccines based on phoPR inactivation. In this context, the live attenuated vaccine MTBVAC, based on a phoP fadD26 deletion mutant of M. tuberculosis, is the first vaccine of this kind to successfully enter into clinical development, representing a historic milestone in the field of human vaccinology.

Current efforts and future prospects in the development of live mycobacteria as vaccines.

The development of more effective vaccines against Mycobacterium tuberculosis (Mtb) remains a major goal in the effort to reduce the enormous global burden of disease caused by this pathogen. Whole-cell vaccines based on live mycobacteria with attenuated virulence represent an appealing approach, providing broad antigen exposure and intrinsic adjuvant properties to prime durable immune responses. However, designing vaccine strains with an optimal balance between attenuation and immunogenicity has proven to be extremely challenging. Recent basic and clinical research efforts have broadened our understanding of Mtb pathogenesis and created numerous new vaccine candidates that have been designed to overcome different aspects of immune evasion by Mtb. In this review, we provide an overview of the current efforts to create improved vaccines against tuberculosis based on modifications of live attenuated mycobacteria. In addition, we discuss the use of such vaccine strains as vectors for stimulating protective immunity against other infectious diseases and cancers.

Tuberculosis vaccine development: Shifting focus amid increasing development challenges.

A new tuberculosis vaccine is needed to replace or enhance BCG, which induces variable protection against Mycobacterium tuberculosis pulmonary infections in adults. Development of new TB vaccine candidates is severely hampered by the lack of a correlate of immunity, unproven animal models, and limited funding opportunities. One candidate, MVA85A, recently failed to meet its efficacy endpoint goals despite promising early-phase trial data. As a result, some in the field believe we should now shift our focus away from product development and toward a research-oriented approach. Here, we outline our suggestions for this research-oriented strategy including diversification of the candidate pipeline, expanding measurements of immunity, improving pre-clinical animal models, and investing in combination pre-clinical/experimental medicine studies. As with any evolution, this change in strategy comes at a cost but may also represent an opportunity for advancing the field.

Protective capacity of proteoliposomes from Mycobacterium bovis BCG in a mouse model of tuberculosis.

Tuberculosis (TB) is one of the most important causes of mortality and morbidity due to infectious diseases. BCG, the vaccine in use, is not fully protective against TB. In a previous study, we have shown that proteoliposomes (outer membrane extracts), obtained from BCG (PLBCG) were able to induce humoral immune responses against Mycobacterium tuberculosis (Mtb) antigens. With the objective to evaluate the protective capability of PLBCG alone or as a booster with BCG, a murine model of progressive pulmonary TB was used. Animals immunized with PLBCG adjuvanted with alum (PLBCG-Al) showed similar protection to that conferred by BCG. The group immunized with PLBCG-Al as a booster to BCG gave superior protection than BCG as evidenced by a reduction of bacterial load in lungs 2 months after infection with Mtb. Animals immunized with BCG, PLBCG-Al and this formulation as a booster of BCG, showed a significant decrease of tissue damage (percentage of pneumonic area/lung) compared with non-immunized animals. These results demonstrate that immunization with PLBCG-Al alone or as a booster to BCG induce appropriate protection against challenge with Mtb in mice and support the future evaluation of PLBCG as a promising vaccine candidate against Mtb.

A PE_PGRS33 protein of Mycobacterium tuberculosis: an ideal target for future tuberculosis vaccine design.

It is known that cellular immune response is relevant to fight against tuberculosis (TB); hence, identification of mycobacterial antigens that induce a protective immune cellular response is of great interest, especially for the development of effective TB vaccines. Genomic data have an impact on the identification of potential antigens as new vaccine targets. In this review, we summarize the current knowledge about the advances in new TB vaccine designs as well as the features reported for the pro-glu_polymorphic GC-rich sequence (PE_PGRS33) protein, considering this molecule as a prototype of the PE_PGRS family to better understand the biological function of this protein family that could be considered an ideal target for future vaccine design.

Development of a new DNA vaccine based on mycobacterial ESAT-6 antigen delivered by recombinant invasive Lactococcus lactis FnBPA+.

The use of the food-grade bacterium Lactococcus lactis as a vehicle for the oral delivery of DNA vaccine plasmids constitutes a promising strategy for vaccination. The delivery of DNA plasmids into eukaryotic cells is of critical importance for subsequent DNA expression and effectiveness of the vaccine. In this context, the use of the recombinant invasive L. lactis FnBPA+ (fibronectin-binding protein A) strain for the oral delivery of the eukaryotic expression vector vaccination using lactic acid bacteria (pValac), coding for the 6-kDa early secreted antigenic target (ESAT-6) gene of Mycobacterium tuberculosis, could represent a new DNA vaccine strategy against tuberculosis. To this end, the ESAT-6 sequence was cloned into the pValac vector; the L. lactis fibronectin-binding protein A (FnBPA)+ (pValac:ESAT-6) strain was obtained, and its immunological profile was checked in BALB/c mice. This strain was able to significantly increase interferon gamma (IFN-γ) production in spleen cells, showing a systemic T helper 1 (Th1) cell response. The mice also showed a significant increase in specific secretory immunoglobulin A (sIgA) production in colon tissue and fecal extracts. Thus, this is the first time that L. lactis has been used to deliver a plasmid DNA harboring a gene that encodes an antigen against tuberculosis through mucous membranes.

Assessment of phagosomes infected with Mycobacterium tuberculosis as a vaccine candidate against tuberculosis.

The present study describes a novel and simple vaccination strategy that involve culturing of M. tuberculosis in the macrophage cells. Isolation of phagosome from macrophage (cell line J774) infected with M. tuberculosis (H37) and M. bovis (BCG) at early and late phase of infection was done ensuing the identification and characterization of these phagosome. In vitro study of apoptosis induced by phagosome infected with (H37) and (BCG) was performed. The vaccine candidate with H1137 MOI- 1:10 at 3 h, MOI- 1:20 at 1, 1.5, 2.5 and 3 h and BCG MOI- 1:20 at 3.5 h showed percentage apoptosis as 38.64, 39.93, 34.66, 22.56,34.59 and 37.81% respectively. The results designates that macrophages provide cellular niche during infection and illustrate considerable immunogenic property. Novel antigens expressed or secreted by H37 in infected macrophages can provide evidence to be a successful vaccine candidate as it endures enhanced immune response than BCG.

In search of a new paradigm for protective immunity to TB.

Clinical trials of vaccines against Mycobacterium tuberculosis are well under way and results are starting to come in. Some of these results are not so encouraging, as exemplified by the latest Aeras-422 and MVA85A trials. Other than empirically determining whether a vaccine reduces the number of cases of active tuberculosis, which is a daunting prospect given the chronic nature of the disease, we have no way of assessing vaccine efficacy. Therefore, investigators seek to identify biomarkers that predict vaccine efficacy. Historically, focus has been on the production of interferon-γ by CD4(+) T cells, but this has not been a useful correlate of vaccine-induced protection. In this Opinion article, we discuss recent advances in our understanding of the immune control of M. tuberculosis and how this knowledge could be used for vaccine design and evaluation.

Comparative genomics of the Mycobacterium signaling architecture and implications for a novel live attenuated Tuberculosis vaccine.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (M.tb), remains a major threat to global public health. A new TB vaccine affording superior immune protection to M. bovis Bacillus Calmette-Guérin (BCG) is imperative. The advantage of a live attenuated vaccine is that it can mimic the bona fide pathogen, elicit immune responses similar to natural infection, and potentially provide more protection than other vaccines. BCG, the only vaccine and a live attenuated vaccine that is the result of cumulative mutations by serial passage of M. bovis, has provided clues for the construction of novel improved vaccines. A strategy is put forward for identifying a new live attenuated TB vaccine generated by cumulative mutation based on M.tb. Given the important role of the M.tb signaling network consisting of a two-component system, eukaryotic-like Ser/Thr protein kinase system and sigma factor system based on comparisons among M.tb H37Rv, M. bovis, and BCG, we have put a premium on this signaling circuit as the starting point for the generation of an attenuated TB vaccine.