**6.3. Vaccines**

The induction of specific protective antibody responses by vaccination, either alone or as an addition to the stimulation of cell mediated immunity could be a novel strategy for the development of new generation of prophylactic and therapeutic vaccines against TB.

The prevailing past dogma that discounted the role of antibodies in host protection against TB has resulted in a limited study of B cell immunodominant epitopes as targets for protective immunity [87].

#### *6.3.1. Polysaccharide conjugate vaccines*

Polysaccharide conjugate vaccines are considered to elicit specific protective antibody responses against a variety of pathogens [88]. However, the polysaccharide conjugate vaccine against *Salmonella typhi* [89] demonstrates the feasibility of this kind of vaccines for the prevention of infectious diseases caused by intracellular pathogens. In the case of *M. tuberculosis*, several authors reported the use of polysaccharide conjugated vaccine candi‐ dates [61, 90, 91, 92].

All these vaccine candidates induced the production of specific IgG [61, 90, 91, 92] and some of them conferred variable levels of protection [61, 91] which validate this strategy as one of the potential avenues for the development of new generation of vaccines against tuberculosis

#### *6.3.2. Identifying other B-cell immunodominant epitopes*

With the development of bioinformatics tools for bacterial genome analysis, it has been possible to predict *in silico* microbial regions that trigger immune responses relevant for protection and vaccine development. A candidate experimental vaccine based on proteolipo‐ somes from *M. smegmatis* is currently in development [93].

In one study, bibliographic search was used to identify highly expressed proteins in active, latent and reactivation phases of TB [94]. The subcellular localization of the selected proteins was defined according to the report on the identification and localization of 1044 *M. tubercu‐ losis* proteins using two-dimensional, capillary high-performance liquid chromatography coupled with mass spectrometry (2DLC/MS) method [95] and using prediction algorithms. Taking into consideration the cell fractions potentially included in the proteoliposome, from the previously identified proteins, the ones located in the cell membrane and cell wall, as well as those which are secreted and homologous to those of *M. smegmatis* were selected.

The regions of the selected proteins containing promiscuous B and T cell epitopes were determined [94]. Thus the *M. smegmatis* proteoliposomes were predicted to contain multiple B and T epitopes which are potentially cross reactive with those of *M. tuberculosis*. It is important to note that there could be conformational B epitopes and additional epitopes related with lipids and carbohydrates included in the proteoliposomes that could reinforce the humoral cross reactivity.

Considering the results of the *in silico* analysis, proteoliposomes of *M. smegmatis* were obtained and their immunogenicity was studied in mice [93]. In addition to cellular immune effectors recognizing antigens from *M. tuberculosis*, cross reactive humoral immune responses of several IgG subclasses corresponding with a combined Th1 and Th2 pattern against antigenic components of *M. tuberculosis* were elicited. These findings were in concordance with the *in silico* predictions [93, 94]. It is interesting to note that differences in the pattern of humoral recognition of lipidic components was dependent on the characteristics of the adjuvant used, which could have relevance for the development of vaccines which includes lipidic compo‐ nents [93]. Currently studies are underway to evaluate the protective capacity of *M. smegma‐ tis* proteoliposomes in challenge models with *M. tuberculosis* in mice.

Bioinformatics tools for prediction of T and B epitopes were also employed for the design of multiepitopic constructions, which were used to obtain recombinant BCG strains. Based on this prediction, B cell epitopes from ESAT-6, CFP-10, Ag87B and MTP40 proteins were selected and combined with T cell epitopes of the 87B protein and fused to Mtb8.4 protein [96].

A significant IgG antibody response against specific B cell epitopes of ESAT-6 and CFP-10 was obtained in mice immunized with the recombinant strain. After studying the specific response of spleen cells by lymphoproliferation assay and detection of intracellular cytokines in CD4 + and CD8 + subpopulations, the recognition of T epitopes was also observed. The response showed a Th1 pattern after immunization with this recombinant strain (Mohamud, R, et al. manuscript in preparation). In another series of experiments, recombinant BCG strains expressing several combinations of multiepitopic constructions were used to immunize BALB/ c mice subcutaneously and challenged intratracheally with the *M. tuberculosis* H37Rv strain. Recombinant BCG strains expressing T epitopes from 87BAg fused to Mtb8.4 protein and BCG expressing a HSP62 T cell epitope plus different combinations of B cell epitopes from 87BAg, Mce1A, L7/L12, 16 kDa, HBHA, ESAT6, CFP10 and MTP40 and combinations of B cell epitopes alone produced significant reductions in lung CFU compared to BCG (Norazmi MN, et al. manuscript in preparation).

#### *6.3.3. Diagnosis*

**6.2. Prophylactic use**

66 Tuberculosis - Current Issues in Diagnosis and Management

**6.3. Vaccines**

immunity [87].

dates [61, 90, 91, 92].

*6.3.1. Polysaccharide conjugate vaccines*

*6.3.2. Identifying other B-cell immunodominant epitopes*

somes from *M. smegmatis* is currently in development [93].

Prophylactic use of antibodies could be applied in recent contacts of TB patients, with special attention to risk groups [84]. In this regard, successful prophylactic use of antibodies in exposed individuals has been shown in the case of several other pathogens such as varicella, tetanus,

The induction of specific protective antibody responses by vaccination, either alone or as an addition to the stimulation of cell mediated immunity could be a novel strategy for the

The prevailing past dogma that discounted the role of antibodies in host protection against TB has resulted in a limited study of B cell immunodominant epitopes as targets for protective

Polysaccharide conjugate vaccines are considered to elicit specific protective antibody responses against a variety of pathogens [88]. However, the polysaccharide conjugate vaccine against *Salmonella typhi* [89] demonstrates the feasibility of this kind of vaccines for the prevention of infectious diseases caused by intracellular pathogens. In the case of *M. tuberculosis*, several authors reported the use of polysaccharide conjugated vaccine candi‐

All these vaccine candidates induced the production of specific IgG [61, 90, 91, 92] and some of them conferred variable levels of protection [61, 91] which validate this strategy as one of the potential avenues for the development of new generation of vaccines against tuberculosis

With the development of bioinformatics tools for bacterial genome analysis, it has been possible to predict *in silico* microbial regions that trigger immune responses relevant for protection and vaccine development. A candidate experimental vaccine based on proteolipo‐

In one study, bibliographic search was used to identify highly expressed proteins in active, latent and reactivation phases of TB [94]. The subcellular localization of the selected proteins was defined according to the report on the identification and localization of 1044 *M. tubercu‐ losis* proteins using two-dimensional, capillary high-performance liquid chromatography coupled with mass spectrometry (2DLC/MS) method [95] and using prediction algorithms. Taking into consideration the cell fractions potentially included in the proteoliposome, from the previously identified proteins, the ones located in the cell membrane and cell wall, as well

The regions of the selected proteins containing promiscuous B and T cell epitopes were determined [94]. Thus the *M. smegmatis* proteoliposomes were predicted to contain multiple

as those which are secreted and homologous to those of *M. smegmatis* were selected.

development of new generation of prophylactic and therapeutic vaccines against TB.

Respiratory Synsicial Virus (RSV), rabies and Hepatitis B [85, 86]

Although no serological assays are currently recommended for diagnosis of TB [97], largely due to the possibility of false results and thus incorrect treatments, for many other pathogens, serological diagnostic tests has been of great value, particularly in poor countries. In some cases, antibody responses can constitute useful correlates of protection [98]. In the specific case of TB, several studies of the antibody response have been reported [99]. There is a substantial amount of variability in antibody response to TB [100]. This variability has been attributed to several factors. Some of these factors are associated with the pathogen (strain variation, microenvironment and growth state of bacteria) and others are related to the host, primarily previous exposure to related antigens and host genetics [99].

However, it is important to consider that only a small fraction of the genomic regions of *M. tuberculosis* encoding proteins has been explored. Currently, novel immunoassay platforms are being used to dissect the entire proteome of *M. tuberculosis*, including reacting protein microarrays with sera from TB patients and controls [101,102]. These studies could lead to the discovery of new antigens that may constitute suitable diagnostic markers and tools for the identification of protection correlates.

2 School of Health Sciences and Institute for Research in Molecular Medicine, Universiti

The Role of Antibodies in the Defense Against Tuberculosis

http://dx.doi.org/10.5772/53950

69

3 Experimental Pathology Section, National Institute of Medical Sciences and Nutrition,

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