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Journal of Biomedicine and Biotechnology
Volume 2010 (2010), Article ID 832341, 8 pages
http://dx.doi.org/10.1155/2010/832341
Methodology Report

Proposing Low-Similarity Peptide Vaccines against Mycobacterium tuberculosis

Department of Biochemistry and Molecular Biology “Ernesto Quagliariello”, University of Bari, Bari 70126, Italy

Received 23 September 2009; Revised 2 December 2009; Accepted 24 March 2010

Academic Editor: Yongqun Oliver He

Copyright © 2010 Guglielmo Lucchese et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Using the currently available proteome databases and based on the concept that a rare sequence is a potential epitope, epitopic sequences derived from Mycobacterium tuberculosis were examined for similarity score to the proteins of the host in which the epitopes were defined. We found that: (i) most of the bacterial linear determinants had peptide fragment(s) that were rarely found in the host proteins and (ii) the relationship between low similarity and epitope definition appears potentially applicable to T-cell determinants. The data confirmed the hypothesis that low-sequence similarity shapes or determines the epitope definition at the molecular level and provides a potential tool for designing new approaches to prevent, diagnose, and treat tuberculosis and other infectious diseases.

1. Introduction

Mycobacterium tuberculosis is the causative agent of tuberculosis (TB) and is a major infectious pathogen. A resurgence of tuberculosis has marked the beginning of the third millennium, with about 8 million new cases and 1.8 million deaths annually worldwide attributed to M. tuberculosis [1]. More recently, the World Health Organization’s Global Tuberculosis Control Report 2009 estimated 9.27 million cases of TB in 2007 [2], an increase from 9.24 million cases in 2006, 8.3 million cases in 2000, and 6.6 million cases in 1990. These numbers underscore the need to develop and characterize rational and effective TB therapies and diagnostic reagents. Currently, the M. bovis bacillus Calmette-Guérin (BCG) vaccine is used against TB. BCG has excellent immuno-potentiating properties, but it is not uniformly effective [3], and efforts to develop a more effective M. bovis-derived vaccine against TB have not had satisfactory results [4, 5]. Moreover, the emergence of multiple-drug-resistant TB [6] and the increasing prevalence of mycobacterial disease in people with acquired immunodeficiency syndrome [7] add urgency to the need for an effective vaccine against TB.

Recent studies have provided convincing evidence that antibodies can protect against mycobacterial infections [8]. Passive immunization using mono- and polyclonal antibodies and humoral immune responses against specific mycobacterial antigens in experimental animals have shown that antibodies may play an anti-TB role [911]. The epitopic characterization of mycobacterial antigens is performed antigen-by-antigen, searching for specific peptide features and epitope-paratope interaction domains for each mycobacterial protein antigen [1214]. In essence, the common molecular basis and the fundamental rules underlying the immune function of specific peptide units remain unexplained [15, 16]. Little is known about the structure-function relationship(s) of protein sequences in immunology. This lack of knowledge may underlie the confusion surrounding neutralizing versus nonneutralizing antibodies, harmful vaccine side effects, and imprecise diagnostic tools.

Within this framework, we propose that rare peptide sequences, that is, peptide motifs rarely found in host proteins, elicit an immune response. Unique protein sequences are more likely to evoke an immune response than are highly repeated motifs, which would be immunogenically silenced by tolerance. We have previously validated the relationship between sequence rarity and peptide immunoreactivity in cancer, autoimmunity, and infectious disease models. We reported several examples of specific targeting of peptide regions with no or limited sequence similarity to the host proteome, including mono- and polyclonal antibodies raised against EC Her-2/Neu oncoprotein [17], desmoglein-3 [18], melan-a/MART-1 [19], high-molecular-weight melanoma-associated-antigen [20, 21], tyrosinase [22, 23], prostate-specific-antigen [24], HPV16 E7 [25], HCV [26], and influenza A [27] proteins. In addition, a review of the epitope mapping literature revealed that most peptide epitopes obey the low-similarity rule [28], further supporting our hypothesis. Taken together, our data [1727] and findings from other research [28] suggest that unique peptide sequences are more likely to be immunogenic than are highly repeated peptide sequences, which may be immuno-tolerated.

In the present study, we investigated the low-similarity hypothesis in M. tuberculosis antigen epitopes with the aim of defining immunogenic peptide sequences that may be effective for anti-TB immunotherapy. We studied validated M. tuberculosis epitopes currently cataloged in the Immune Epitope Database and Analysis Resources (IEDB) (http://www.immuneepitope.org/) [29, 30]. We report that most of the mycobacterial epitopes involved in the immune response are characterized by a low level of similarity to the host proteome.

2. Methods

The IEDB contains hundreds of mycobacterial epitopes derived from various strains and with different characteristics [30]. Specifically, the IEDB contains B- and T-cell mycobacterial epitopes that are (a) linear or conformational; (b) derived from M. tuberculosis, M. bovis, M. Avium, and M. leprae; (c) of different lengths (up to 36 aa) [31]; and (d) have sequences that were negative, positive, or produced mixed negative/positive results in immunoassays. The present report focused on B-cell linear epitopes derived from M. tuberculosis that were positive in immunoassays.

These mycobacterial epitopes were analyzed for similarity to the host proteins with the aim of identifying identical amino acid groupings that were common sequences [32]. The amino acid groupings were linear pentapeptides. Pentapeptides were used because the canonical literature indicates that five to six amino acids are a sufficient minimal determinant for an epitope-paratope interaction, and thus a pentapeptide can act as an immune unit and play a crucial role in cell immunoreactivity and antigen-antibody recognition [33, 34].

Fifty-six M. tuberculosis epitopes cataloged as immuno-positive at IEDB resource were dissected into pentamer motifs offset by one residue: that is, WDEDG, DEDGE, EDGEK, DGEKR, and so forth. Then, each 5-mer was used as a probe to scan the entire host proteome searching for identical matches using the PIR protein database and PIR perfect peptide match program (http://pir.georgetown.edu/pirwww/) [35]. The number of matches of each mycobacterial pentamer to the host proteome indicates the similarity score that can range from no matches to hundreds of matches. Following previous experimental data, a pentapeptide fragment that has about five (or fewer) perfect matches to the host proteome was considered to be a low-similarity sequence, that is, a rare fragment [1728].

3. Results

3.1. Low-Similarity Sequences and B-Cell Epitopes

We examined the linear M. tuberculosis-derived B-cell epitopes cataloged in the IEDB [30], using pentapeptide probes as described above and report the similarity scores (Table 1).

tab1
Table 1: Similarity analysis of M. tuberculosis B-cell epitopes to the host proteome.

As shown in Table 1, most of the mycobacterial B-cell epitopes contain pentapeptides that are rare or absent in the host proteins. Furthermore, preliminary screening revealed that a few B-cell epitopes were also T-cell epitopes (Table 1, footnote 5) [36, 4952].

The relationship between low-similarity sequences and mycobacterial epitopes was further confirmed by extending the similarity analysis to the COOH domain of the mycobacterial DNAK protein. The DNAK chaperon protein is a member of the highly conserved heat shock protein family and is a protein antigen involved in the immune response to mycobacteria [53, 54]. In particular, the DNAK carboxy-terminus domain is highly immunogenic [41, 55] and hosts several of the epitopes described in the IEDB database (Table 1) [41]. Thus, the COOH domain of the mycobacterial DNAK protein was suitable for verifying the similarity-immunogenicity link. In brief, the primary sequence of the DNAK COOH-terminus domain (aa 500–609) was dissected into pentamers that were probed against the human proteome by the computer-assisted similarity analysis described in the Methods section. Then, the epitopic sequences were allocated along the mycobacterial-versus-human similarity profile.

Figure 1 shows the similarity profile of the DNAK COOH-terminus protein to the human proteome and the localization of the linear epitopes along the antigenic sequence. It can be seen that the carboxy terminal sequence of the mycobacterial chaperon protein has pentapeptides occurring frequently in the human proteome as well as unique fragments. Five pentamers (i.e., TWRIGY, WRIGY, VTGHW, TGHWR, and GHWRC) were unique to the DNAK COOH-terminus domain and these accounted for 5% of the total DNAK pentamers. The remaining 100 pentapeptides were repeated at different extent throughout the human proteome and occurred a total of 2468 times. Furthermore, and most importantly, Figure 1 shows that the peptide epitopes recognized by the human sera, as demonstrated by Elsaghier et al. [41], were among the mycobacterial peptide fragments that had a low similarity to the human proteome. That is, the unique pentapeptide signatures of the DNAK carboxy-terminus domain coincided with the epitopic motifs involved in immune recognition, indicating a direct relationship between low similarity and the humoural antibody response. Conversely, M. tuberculosis peptides not responsive to the human sera had a high number of pentamer matches to the human proteome (Figure 1, black arrows numbered from 6 to 15).

832341.fig.001
Figure 1: Similarity profile of the mycobacterial DNAK carboxy-terminus domain versus the human proteome at the pentapeptide level. The columns indicate the number of DNAK pentapeptide occurrences in the human proteome. DNAK corresponds to hsp70 protein, UniProt accession: P0A5B9; length: 609 aa. The numbered arrows indicate the location of mycobacterial peptide sequences immunoassayed with human sera [41]. Black arrows refer to the immuno-negative peptide sequences corresponding to IEDB ID: (6) 1212577 (aa FVKEQREA); (7) 1212579 (aa VKEQREAE); (8) 1212581 (aa KEQREAEG); (9) 1212582 (aa EQREAEGG); (10) 1212585 (aa QREAEGGS); (11) 1212586 (aa REAEGGSK); (12) 1212589 (aa EAEGGSKV); (13) 1212590 (aa AEGGSKVP); (14) 1212593 (aa EGGSKVPE); (15) 1212595 (aa GGSKVPED). Red arrows refer to the positive epitopic sequences corresponding to IEDB ID: (1) 1052559; (2) 1052560; (3) 1052558; (4) 1052561; (5) 1052562; (16) 1052583; (17) 1052584; (18) 1052589; (19) 1052591; (20) 1052593; (21) 1052594; (22) 1052595; (23) 1052596; (24) 1052597; (25) 1052598; (26) 24658; (27) 1052627; (28) 1052628; (29) 1052629; (30) 1052630; (31) 1052631; (32) 1052632; (33) 1052633; (34) 1052549; (35) 1052550. See Table 1 for amino acid sequence details. IEDB ID Reference Dataset of Mycobacterial Immune Epitopes: http://iedb.zendesk.com/entries/18171-reference- datasets-of-mycobacterial-immune-epitopes.
3.2. Low-Similarity Sequences and T-Cell Epitopes

The low-similarity M. tuberculosis B-cell epitopes hosted T-cell epitopes [36, 4952] in several cases (Table 1). These data suggest that low-similarity influences the T- as well as the B-cell immune response, a finding that might be used in defining T-cell epitopes. Characterization of T-cell epitopes is complicated by different sets of rules governing the immune response. The problems include, but are not limited to, epitope hierarchy, immunoprevalence and immunodominance [56], variable epitopic length, the epitope major histocompatibility complex (MHC) binding potential and the difficulty of correctly predicting MHC binding activities in the majority of the test-set peptides [57], and the degenerate T-cell receptor recognition [22].

In this intricate context, the low-similarity hypothesis may provide insight for the structural characterization of immunogenic T-cell epitopic sequences. We know that the optimal peptide length for MHC binding is between nine and fifteen amino acids, and the central 5-6 residues contribute the majority of the specific contacts [59]. Thus, as previously proposed [19], the structural features that define an immunogenic T-cell epitope may be characterized by the MHC binding terminal residues flanking a central low-similarity core formed by 5-6 residues. This model is illustrated in Figure 2 using an immunodominant mycobacterial T-cell epitope validated and described by Ivanyi et al. [58]. The immunodominant p61–80/PT19 mycobacterial epitope (VTGSVVCTTAAGNVNIAIGG) has a critical core formed by five residues (AGNVN, aa 71–75): a single amino acid substitution with N73A impairs T-cell immunogenicity of the target p61–80/PT19 epitope. Our similarity analysis reveals that this substitution causes a shift in the proteomic similarity level of the five core residues (aa 71–75) that changes from one match (aa sequence AGNVN) to eight matches (aa sequence AGAVN) and “hides” the 5-mer core from the immune system.

fig2
Figure 2: The immunogenic impact of a single amino acid substitution in the five core residues (AGNVN, aa 71–75) of the immunodominant p61–80/PT19 mycobacterial epitope (VTGSVVCTTAAGNVNIAIGG). The substitution of N73A impairs T immunogenicity for the target epitope and changes the proteomic similarity level of the five core residues (aa 71–75 underlined) from one match (aa sequence AGNVN) to eight matches (aa sequence AGAVN). (a) The immunogenic MHC-peptide complex containing the central low-similarity pentapeptide AGNVN (in white); (b) The nonimmunogenic MHC-peptide complex containing the higher similarity pentapeptide AGAVN (in gray). Immunoreactivity and mutagenesis experiments are described in [58]. The figures are modified versions of the original drawings from the website http://www.iayork.com/ by kind authorization of Professor Ian York, University of Michigan.

The model illustrated in Figure 2 is supported by reports that the HFMPT pentapeptide is a minimal antigenic determinant for MHC class I-restricted T lymphocytes [60], and the KYVKQ pentapeptide is a minimal antigenic determinant for CD4(+) T-cell clones [61]. Moreover, the proteomic similarity values for HFMPT and KYVKQ were no matches and one match, respectively.

4. Discussion

In this study, we explored the IEDB resources to define and characterize immunogenic mycobacterial epitopes that could be used for effective anti-TB immunotherapy. We observed that the immunological information carried by the M. tuberculosis antigens was localized in rare peptide fragments, confirming the relationship between low-similarity and immunogenicity. Sequences containing pentapeptide fragment(s) with low similarity to the host proteome appear to be those involved in the humoral antibody recognition of the mycobacterial antigen and may also influence T-cell immunoreactivity too.

Our findings provide a method for investigating the immune potential of the mycobacterial proteome, and thus, may help elucidate M. tuberculosis immunobiology, one of the primary challenges in current tuberculosis research. This is an important issue, particularly because the epitopic characterization of mycobacterial antigens advances slowly, antigen-by-antigen [1214].

Moreover, mycobacterial antigenic motifs rarely found in host proteins are more likely to serve as potential immunogenic antigens. Use of short peptide modules rather than full-length mycobacterial antigens in vaccines may increase specificity and efficacy. It has been understood since the 1980s that chemically synthesized small peptides can induce antibodies that react with intact proteins [62]. Furthermore, peptide-based vaccines have been successfully used against several pathologies and infectious diseases. For example, tumour antigen-derived peptides evoked potent antitumour immunity in the murine melanoma M-3 [63]; cytotoxic CD4+ and CD8+ T lymphocytes generated by mutant p21-ras (12Val) peptide vaccination selectively killed autologous tumour cells carrying this mutation [64]; Her-2/neu peptide (aa 657–665) is an immunogenic epitope of Her-2/neu oncoprotein with potent antitumour properties [65]; synthetic peptides identified as antigenic sites on the S1 subunit of pertussis toxin induced especially high antibody titres against native pertussis toxin in mice [66]; a linear peptide containing minimal T- and B-cell epitopes of Plasmodium falciparum circumsporozoite protein provided protection against a transgenic sporozoite challenge [67]. These findings indicate that the use of exact immunogenic mycobacterial peptide sequences may provide the most effective active and passive immunotherapeutic anti-TB approaches. In addition, a major obstacle to antibody-based anti-TB therapy is the risk of adverse effects that range from Lupus vulgaris to granulomatous hepatitis [6872], possibly caused by cross-reactivity [73]. The use of low-similarity peptides may allow the development of effective vaccines without adverse side-effects [74]. In conclusion, the findings of the present study may provide guidance in the analysis, identification, and utilization of the B- and T-cell response in vaccination against mycobacterial and other infectious diseases.

Acknowledgments

This study was made possible by the free access to the Immune Epitope Database at the National Health Institutes, Bethesda, MD. We thank Professor Ian York, University of Michigan, for allowing us to use his figures. We are indebted to the Referees for precious comments and suggestions. Of course, we remain responsible for any remaining errors.

References

  1. E. L. Corbett, C. J. Watt, N. Walker, et al., “The growing burden of tuberculosis: global trends and interactions with the HIV epidemic,” Archives of Internal Medicine, vol. 163, no. 9, pp. 1009–1021, 2003. View at Publisher · View at Google Scholar · View at Scopus
  2. The World Health Organization Report, Global Tuberculosis Control, WHO, Geneva, Switzerland, 2009.
  3. P. E. M. Fine, “BCG: the challenge continues,” Scandinavian Journal of Infectious Diseases, vol. 33, no. 4, pp. 243–245, 2001. View at Publisher · View at Google Scholar · View at Scopus
  4. K. Huygen, “DNA vaccines against mycobacterial diseases,” Future Microbiology, vol. 1, pp. 63–73, 2006. View at Publisher · View at Google Scholar · View at Scopus
  5. A. Tanghe, J.-P. Dangy, G. Pluschke, and K. Huygen, “Improved protective efficacy of a species-specific DNA vaccine encoding mycolyl-transferase Ag85A from Mycobacterium ulcerans by homologous protein boosting,” PLoS Neglected Tropical Diseases, vol. 2, no. 3, article e199, 2008. View at Publisher · View at Google Scholar · View at Scopus
  6. D. B. Meya and K. P. McAdam, “The TB pandemic: an old problem seeking new solutions,” Journal of Internal Medicine, vol. 261, no. 4, pp. 309–329, 2007. View at Publisher · View at Google Scholar · View at Scopus
  7. V. Nissapatorn, I. Kuppusamy, F. P. Josephine, I. Jamaiah, M. Rohela, and A. Khairul Anuar, “Tuberculosis: a resurgent disease in immunosuppressed patients,” The Southeast Asian Journal of Tropical Medicine and Public Health, vol. 37, supplement 3, pp. 153–160, 2006.
  8. F. Abebe and G. Bjune, “The protective role of antibody responses during Mycobacterium tuberculosis infection,” Clinical and Experimental Immunology, vol. 157, no. 2, pp. 235–243, 2009. View at Publisher · View at Google Scholar · View at Scopus
  9. Y. Xu, B. Zhu, Q. Wang, et al., “Recombinant BCG coexpressing Ag85B, ESAT-6 and mouse-IFN-γ confers effective protection against Mycobacterium tuberculosis in C57BL/6 mice,” FEMS Immunology and Medical Microbiology, vol. 51, no. 3, pp. 480–487, 2007. View at Publisher · View at Google Scholar · View at Scopus
  10. Y. Q. Qie, J. L. Wang, B. D. Zhu, et al., “Evaluation of a new recombinant BCG which contains mycobacterial antigen ag85B-mpt64190-198-mtb8.4 in C57/BL6 mice,” Scandinavian Journal of Immunology, vol. 67, no. 2, pp. 133–139, 2008. View at Publisher · View at Google Scholar · View at Scopus
  11. S. Chang-Hong, W. Xiao-Wu, Z. Hai, Z. Ting-Fen, W. Li-Mei, and X. Zhi-Kai, “Immune responses and protective efficacy of the gene vaccine expressing Ag85B and ESAT6 fusion protein from Mycobacterium tuberculosis,” DNA and Cell Biology, vol. 27, no. 4, pp. 199–207, 2008. View at Publisher · View at Google Scholar · View at Scopus
  12. R. Hussain, H. M. Dockrell, and T. J. Chiang, “Dominant recognition of a cross-reactive B-cell epitope in Mycobacterium leprae 10 K antigen by immunoglobulin G1 antibodies across the disease spectrum in leprosy,” Immunology, vol. 96, no. 4, pp. 620–627, 1999. View at Publisher · View at Google Scholar · View at Scopus
  13. K. K. Singh, N. Sharma, D. Vargas, et al., “Peptides of a novel Mycobacterium tuberculosis-specific cell wall protein for immunodiagnosis of tuberculosis,” Journal of Infectious Diseases, vol. 200, no. 4, pp. 571–581, 2009. View at Publisher · View at Google Scholar · View at Scopus
  14. L. Baassi, K. Sadki, F. Seghrouchni, et al., “Evaluation of a multi-antigen test based on B-cell epitope peptides for the serodiagnosis of pulmonary tuberculosis,” International Journal of Tuberculosis and Lung Disease, vol. 13, no. 7, pp. 848–854, 2009. View at Scopus
  15. U. Gowthaman and J. N. Agrewala, “In silico tools for predicting peptides binding to HLA-class II molecules: more confusion than conclusion,” Journal of Proteome Research, vol. 7, no. 1, pp. 154–163, 2008. View at Publisher · View at Google Scholar · View at Scopus
  16. M. J. Blythe and D. R. Flower, “Benchmarking B cell epitope prediction: underperformance of existing methods,” Protein Science, vol. 14, no. 1, pp. 246–248, 2005. View at Publisher · View at Google Scholar · View at Scopus
  17. A. Mittelman, A. Lucchese, A. A. Sinha, and D. Kanduc, “Monoclonal and polyclonal humoral immune response to EC HER-2/neu peptides with low similarity to the host's proteome,” International Journal of Cancer, vol. 98, no. 5, pp. 741–747, 2002. View at Publisher · View at Google Scholar · View at Scopus
  18. A. Lucchese, A. Mittelman, M.-S. Lin, D. Kanduc, and A. A. Sinha, “Epitope definition by proteomic similarity analysis: identification of the linear determinant of the anti-Dsg3 MAb 5H10,” Journal of Translational Medicine, vol. 2, article 43, 2004. View at Publisher · View at Google Scholar · View at Scopus
  19. R. Tiwari, J. Geliebter, A. Lucchese, A. Mittelman, and D. Kanduc, “Computational peptide dissection of Melan-a/MART-1 oncoprotein antigenicity,” Peptides, vol. 25, no. 11, pp. 1865–1871, 2004. View at Publisher · View at Google Scholar · View at Scopus
  20. A. Mittelman, R. Tiwari, G. Lucchese, J. Willers, R. Dummer, and D. Kanduc, “Identification of monoclonal anti-HMW-MAA antibody linear peptide epitope by proteomic database mining,” Journal of Investigative Dermatology, vol. 123, no. 4, pp. 670–675, 2004. View at Publisher · View at Google Scholar · View at Scopus
  21. R. Dummer, A. Mittelman, F. P. Fanizzi, G. Lucchese, J. Willers, and D. Kanduc, “Non-self-discrimination as a driving concept in the identification of an immunodominant HMW-MAA epitopic peptide sequence by autoantibodies from melanoma cancer patients,” International Journal of Cancer, vol. 111, no. 5, pp. 720–726, 2004. View at Publisher · View at Google Scholar · View at Scopus
  22. A. Lucchese, J. Willers, A. Mittelman, D. Kanduc, and R. Dummer, “Proteomic scan for tyrosinase peptide antigenic pattern in vitiligo and melanoma: role of sequence similarity and HLA-DR1 affinity,” Journal of Immunology, vol. 175, no. 10, pp. 7009–7020, 2005. View at Scopus
  23. J. Willers, A. Lucchese, A. Mittelman, R. Dummer, and D. Kanduc, “Definition of anti-tyrosinase MAb T311 linear determinant by proteome-based similarity analysis,” Experimental Dermatology, vol. 14, no. 7, pp. 543–550, 2005. View at Publisher · View at Google Scholar · View at Scopus
  24. A. Stufano and D. Kanduc, “Proteome-based epitopic peptide scanning along PSA,” Experimental and Molecular Pathology, vol. 86, no. 1, pp. 36–40, 2009. View at Publisher · View at Google Scholar · View at Scopus
  25. D. Kanduc, A. Lucchese, and A. Mittelman, “Individuation of monoclonal anti-HPV16 E7 antibody linear peptide epitope by computational biology,” Peptides, vol. 22, no. 12, pp. 1981–1985, 2001. View at Scopus
  26. D. Kanduc, L. Tessitore, G. Lucchese, A. Kusalik, E. Farber, and F. M. Marincola, “Sequence uniqueness and sequence variability as modulating factors of human anti-HCV humoral immune response,” Cancer Immunology, Immunotherapy, vol. 57, no. 8, pp. 1215–1223, 2008. View at Publisher · View at Google Scholar · View at Scopus
  27. G. Lucchese, A. Stufano, and D. Kanduc, “Proteome-guided search for influenza A B-cell epitopes,” FEMS Immunology and Medical Microbiology, vol. 57, no. 1, pp. 88–92, 2009. View at Publisher · View at Google Scholar · View at Scopus
  28. D. Kanduc, ““Self-nonself” peptides in the design of vaccines,” Current Pharmaceutical Design, vol. 15, no. 28, pp. 3283–3289, 2009. View at Publisher · View at Google Scholar · View at Scopus
  29. B. Peters, J. Sidney, P. Bourne, et al., “The immune epitope database and analysis resource: from vision to blueprint,” PLoS Biology, vol. 3, no. 3, article e91, 2005.
  30. M. J. Blythe, Q. Zhang, K. Vaughan, et al., “An analysis of the epitope knowledge related to Mycobacteria,” Immunome Research, vol. 3, no. 1, article 10, 2007. View at Publisher · View at Google Scholar · View at Scopus
  31. P. Chakhaiyar, Y. Nagalakshmi, B. Aruna, K. J. R. Murthy, V. M. Katoch, and S. E. Hasnain, “Regions of high antigenicity within the hypothetical PPE major polymorphic tandem repeat open-reading frame, Rv2608, show a differential humoral response and a low T cell response in various categories of patients with tuberculosis,” Journal of Infectious Diseases, vol. 190, no. 7, pp. 1237–1244, 2004. View at Publisher · View at Google Scholar · View at Scopus
  32. A. C. W. May, “Percent sequence identity: the need to be explicit,” Structure, vol. 12, no. 5, pp. 737–738, 2004. View at Publisher · View at Google Scholar · View at Scopus
  33. M. B. A. Oldstone, “Molecular mimicry and immune-mediated diseases,” FASEB Journal, vol. 12, no. 13, pp. 1255–1265, 1998. View at Scopus
  34. G. Lucchese, A. Stufano, B. Trost, A. Kusalik, and D. Kanduc, “Peptidology: short amino acid modules in cell biology and immunology,” Amino Acids, vol. 33, no. 4, pp. 703–707, 2007. View at Publisher · View at Google Scholar · View at Scopus
  35. C. W. Wu, L.-S. Yeh, H. Huang, et al., “The protein information resource,” Nucleic Acids Research, vol. 31, no. 1, pp. 345–347, 2003. View at Publisher · View at Google Scholar · View at Scopus
  36. R. Hussain, F. Shahid, S. Zafar, M. Dojki, and H. M. Dockrell, “Immune profiling of leprosy and tuberculosis patients to 15-mer peptides of Mycobacterium leprae and M. tuberculosis GroES in a BCG vaccinated area: implications for development of vaccine and diagnostic reagents,” Immunology, vol. 111, no. 4, pp. 462–471, 2004. View at Publisher · View at Google Scholar · View at Scopus
  37. B. Chua-Intra, S. Peerapakorn, N. Davey, et al., “T-cell recognition of mycobacterial GroES peptides in Thai leprosy patients and contacts,” Infection and Immunity, vol. 66, no. 10, pp. 4903–4909, 1998. View at Scopus
  38. B. Chua-Intra, J. Ivanyi, A. Hills, J. Thole, C. Moreno, and H. M. Vordermeier, “Predominant recognition of species-specific determinants of the GroES homologues from Mycobacterium leprae and M. tuberculosis,” Immunology, vol. 93, no. 1, pp. 64–72, 1998. View at Publisher · View at Google Scholar · View at Scopus
  39. B. Chua-Intra, R. J. Wilkinson, and J. Ivanyi, “Selective T-cell recognition of the N-terminal peptide of GroES in tuberculosis,” Infection and Immunity, vol. 70, no. 3, pp. 1645–1647, 2002. View at Publisher · View at Google Scholar · View at Scopus
  40. M. Harboe, A. S. Malin, H. S. Dockrell, et al., “B-cell epitopes and quantification of the ESAT-6 protein of Mycobacterium tuberculosis,” Infection and Immunity, vol. 66, no. 2, pp. 717–723, 1998. View at Scopus
  41. A. Elsaghier, R. Lathigra, and J. Ivanyi, “Localisation of linear epitopes at the carboxy-terminal end of the mycobacterial 71 kDa heat shock protein,” Molecular Immunology, vol. 29, no. 9, pp. 1153–1156, 1992. View at Publisher · View at Google Scholar · View at Scopus
  42. J. S. Spencer, H. J. Kim, A. M. Marques, et al., “Comparative analysis of B- and T-cell epitopes of Mycobacterium leprae and Mycobacterium tuberculosis culture filtrate protein 10,” Infection and Immunity, vol. 72, no. 6, pp. 3161–3170, 2004. View at Publisher · View at Google Scholar · View at Scopus
  43. P. W. Roche, C. G. Feng, and W. J. Britton, “Human T-cell epitopes on the Mycobacterium tuberculosis secreted protein MPT64,” Scandinavian Journal of Immunology, vol. 43, no. 6, pp. 662–670, 1996. View at Scopus
  44. D. P. Harris, H.-M. Vordermeier, A. Arya, K. Bogdan, C. Moreno, and J. Ivanyi, “Immunogenicity of peptides for B cells is not impaired by overlapping T-cell epitope topology,” Immunology, vol. 88, no. 3, pp. 348–354, 1996. View at Scopus
  45. K. R. Ashbridge, B. T. Bäckström, H.-X. Liu, et al., “Mapping of T helper cell epitopes by using peptides spanning the 19-kDa protein of Mycobacterium tuberculosis: evidence for unique and shared epitopes in the stimulation of antibody and delayed-type hypersensitivity responses,” Journal of Immunology, vol. 148, no. 7, pp. 2248–2255, 1992. View at Scopus
  46. J. C. Falla, C. A. Parra, M. Mendoza, et al., “Identification of B- and T-cell epitopes within the MTP40 protein of Mycobacterium tuberculosis and their correlation with the disease course,” Infection and Immunity, vol. 59, no. 7, pp. 2265–2273, 1991. View at Scopus
  47. K. A. L. De Smet, H. M. Vordermeier, and J. Ivanyi, “A versatile system for the production of recombinant chimeric peptides,” Journal of Immunological Methods, vol. 177, no. 1-2, pp. 243–250, 1994. View at Publisher · View at Google Scholar · View at Scopus
  48. H.-M. Vordermeier, D. P. Harris, C. Moreno, M. Singh, and J. Ivanyi, “The nature of the immunogen determines the specificity of antibodies and T cells to selected peptides of the 38 kDa mycobacterial antigen,” International Immunology, vol. 7, no. 4, pp. 559–566, 1995.
  49. Y. López-Vidal, S. P. de León-Rosales, M. Castañón-Arreola, M. S. Rangel-Frausto, E. Meléndez-Herrada, and E. Sada-Díaz, “Response of IFN-γ and IgG to ESAT-6 and 38 kDa recombinant proteins and their peptides from Mycobacterium tuberculosis in tuberculosis patients and asymptomatic household contacts may indicate possible early-stage infection in the latter,” Archives of Medical Research, vol. 35, no. 4, pp. 308–317, 2004. View at Publisher · View at Google Scholar · View at Scopus
  50. H. Shams, P. Klucar, S. E. Weis, et al., “Characterization of a Mycobacterium tuberculosis peptide that is recognized by human CD4+ and CD8+ T cells in the context of multiple HLA alleles,” Journal of Immunology, vol. 173, no. 3, pp. 1966–1977, 2004. View at Scopus
  51. A. S. Mustafa, F. A. Shaban, R. Al-Attiyah, et al., “Human Th1 cell lines recognize the Mycobacterium tuberculosis ESAT-6 antigen and its peptides in association with frequently expressed HLA class II molecules,” Scandinavian Journal of Immunology, vol. 57, no. 2, pp. 125–134, 2003. View at Publisher · View at Google Scholar · View at Scopus
  52. D. P. Harris, H. M. Vordermeier, G. Friscia, et al., “Genetically permissive recognition of adjacent epitopes from the 19-kDa antigen of Mycobacterium tuberculosis by human and murine T cells,” Journal of Immunology, vol. 150, no. 11, pp. 5041–5050, 1993. View at Scopus
  53. D. B. Young, “The immune response to mycobacterial heat shock proteins,” Autoimmunity, vol. 7, no. 4, pp. 237–244, 1990. View at Scopus
  54. T. M. Shinnick, “Heat shock proteins as antigens of bacterial and parasitic pathogens,” Current Topics in Microbiology and Immunology, vol. 167, pp. 145–160, 1991.
  55. D. B. Young and T. R. Garbe, “Heat shock proteins and antigens of Mycobacterium tuberculosis,” Infection and Immunity, vol. 59, no. 9, pp. 3086–3093, 1991. View at Scopus
  56. C. Oseroff, B. Peters, V. Pasquetto, et al., “Dissociation between epitope hierarchy and immunoprevalence in CD8 responses to vaccinia virus western reserve,” Journal of Immunology, vol. 180, no. 11, pp. 7193–7202, 2008. View at Scopus
  57. D. A. Winkler and F. R. Burden, “Nonlinear predictive modeling of MHC class II-peptide binding using Bayesian neural networks,” Methods in Molecular Biology, vol. 409, pp. 365–377, 2007. View at Scopus
  58. D. P. Harris, M. Hill, H.-M. Vordermeier, et al., “Mutagenesis of an immunodominant Y cell epitope can affect recognition of different T and B determinants within the same antigen,” Molecular Immunology, vol. 34, no. 4, pp. 315–322, 1997. View at Publisher · View at Google Scholar · View at Scopus
  59. J. B. Rothbard and M. L. Gefter, “Interactions between immunogenic peptides and MHC proteins,” Annual Review of Immunology, vol. 9, pp. 527–565, 1991.
  60. M. J. Reddehase, J. B. Rothbard, and U. H. Koszinowski, “A pentapeptide as minimal antigenic determinant for MHC class I-restricted T lymphocytes,” Nature, vol. 337, no. 6208, pp. 651–653, 1989. View at Scopus
  61. B. Hemmer, T. Kondo, B. Gran, et al., “Minimal peptide length requirements for CD4+ T cell clones—implications for molecular mimicry and T cell survival,” International Immunology, vol. 12, no. 3, pp. 375–383, 2000. View at Scopus
  62. H. L. Niman, R. A. Houghten, L. E. Walker, et al., “Generation of protein-reactive antibodies by short peptides is an event of high frequency: implications for the structural basis of immune recognition,” Proceedings of the National Academy of Sciences of the United States of America, vol. 80, no. 16, pp. 4949–4953, 1983. View at Scopus
  63. W. Schmidt, M. Buschle, W. Zauner, et al., “Cell-free tumor antigen peptide-based cancer vaccines,” Proceedings of the National Academy of Sciences of the United States of America, vol. 94, no. 7, pp. 3262–3267, 1997. View at Publisher · View at Google Scholar · View at Scopus
  64. M. K. Gjertsen, J. Bjorheim, I. Saeterdal, J. Myklebust, and G. Gaudernack, “Cytotoxic CD4+ and CD8+ T lymphocytes, generated by mutant p21-ras (12Val) peptide vaccination of a patient, recognize 12Val-dependent nested epitopes present within the vaccine peptide and kill autologous tumour cells carrying this mutation,” International Journal of Cancer, vol. 72, no. 5, pp. 784–790, 1997. View at Scopus
  65. A. D. Gritzapis, A. Fridman, S. A. Perez, et al., “HER-2/neu (657-665) represents an immunogenic epitope of HER-2/neu oncoprotein with potent antitumor properties,” Vaccine, vol. 28, no. 1, pp. 162–170, 2009. View at Publisher · View at Google Scholar · View at Scopus
  66. P. Askelöf, K. Rodmalm, G. Wrangsell, et al., “Protective immunogenicity of two synthetic peptides selected from the amino acid sequence of Bordetella pertussis toxin subunit S1,” Proceedings of the National Academy of Sciences of the United States of America, vol. 87, no. 4, pp. 1347–1351, 1990. View at Scopus
  67. J. M. Calvo-Calle, G. A. Oliveira, C. O. Watta, J. Soverow, C. Parra-Lopez, and E. H. Nardin, “A linear peptide containing minimal T- and B-cell epitopes of Plasmodium falciparum circumsporozoite protein elicits protection against transgenic sporozoite challenge,” Infection and Immunity, vol. 74, no. 12, pp. 6929–6939, 2006. View at Publisher · View at Google Scholar · View at Scopus
  68. K. Farsinejad, M. Daneshpazhooh, H. Sairafi, M. Barzegar, and M. Mortazavizadeh, “Lupus vulgaris at the site of BCG vaccination: report of three cases,” Clinical and Experimental Dermatology, vol. 34, no. 5, pp. e167–e169, 2009. View at Publisher · View at Google Scholar · View at Scopus
  69. M. Enserink, “Public health: in the HIV era, an old TB vaccine causes new problems,” Science, vol. 318, no. 5853, p. 1059, 2007. View at Publisher · View at Google Scholar · View at Scopus
  70. E. Attia, “BCG vaccine-induced lupus vulgaris,” European Journal of Dermatology, vol. 17, no. 6, pp. 547–548, 2007. View at Scopus
  71. A. Hristea, A. Neacsu, D. A. Ion, A. Streinu-Cercel, and F. Stăniceanu, “BCG-related granulomatous hepatitis,” Pneumologia, vol. 56, no. 1, pp. 32–34, 2007. View at Scopus
  72. A. Mandavilli, “When the vaccine causes disease,” Nature Medicine, vol. 13, no. 3, p. 274, 2007. View at Scopus
  73. A. Spratt, T. Key, and A. J. Vivian, “Chronic anterior uveitis following bacille Calmette-Guérin vaccination: molecular mimicry in action?” Journal of Pediatric Ophthalmology and Strabismus, vol. 45, no. 4, pp. 252–253, 2008. View at Publisher · View at Google Scholar · View at Scopus
  74. D. Kanduc, “Epitopic peptides with low similarity to the host proteome: towards biological therapies without side effects,” Expert Opinion on Biological Therapy, vol. 9, no. 1, pp. 45–53, 2009. View at Publisher · View at Google Scholar · View at Scopus