Identification and selection of immunodominant B and T cell epitopes for dengue multi-epitope-based vaccine


Dengue virus (DENV) comprises four serotypes (DENV1–4) which cause 390 million global infections with 500,000 hospitalizations and 25,000 fatalities annually. Currently, the only FDA approved DENV vaccine is the chimeric live-attenuated vaccine, Dengvaxia®, which is based on the yellow fever virus (YFV) genome that carries the prM and E genes of the respective DENV 1, 2, 3, and 4 serotypes. However, it has lower efficacies against serotypes DENV1 (51%) and DENV2 (34%) when compared with DENV3 (75%) and DENV4 (77%). The absence of T cell epitopes from non-structural (NS) and capsid (C) proteins of the yellow fever vaccine strain might have prevented Dengvaxia® to elicit robust cellular immune responses, as CD8+ T cell epitopes are mainly localized in the NS3 and NS5 regions. Multi-epitope-based peptide vaccines carrying CD4+, CD8+ T cell and B cell epitopes represent a novel approach to generate specific immune responses. Therefore, assessing and selecting epitopes that can induce robust B and T cell responses is a prerequisite for constructing an efficient multi-epitope peptide vaccine. Potent B and T cell epitopes can be identified by utilizing immunoinformatic analysis, but the immunogenicity of the epitopes have to be experimentally validated. In this review, we presented T cell epitopes that have been predicted by bioinformatic approaches as well as recent experimental validations of CD4+ and CD8+ T cell epitopes by ex-vivo stimulation of PBMCs with specific peptides. Immunoproteomic analysis could be utilized to uncover HLA-specific epitopes presented by DENV-infected cells. Based on various approaches, immunodominant epitopes capable of inducing strong immune responses could be selected and incorporated to form a universally applicable multi-epitope-based peptide dengue vaccine.

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  1. 1.

    Ferreira-de-Lima VH, Lima-Camara TN (2018) Natural vertical transmission of dengue virus in Aedes aegypti and Aedes albopictus: a systematic review. Paras Vectors 11(1):77.

    Article  Google Scholar 

  2. 2.

    Martina BE, Koraka P, Osterhaus AD (2009) Dengue virus pathogenesis: an integrated view. Clin Microbiol Rev 22(4):564–581.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Murray NE, Quam MB, Wilder-Smith A (2013) Epidemiology of dengue: past, present and future prospects. Clin Epidemiol 5:299–309.

    Article  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Deng S-Q, Yang X, Wei Y, Chen J-T, Wang X-J, Peng H-J (2020) A review on dengue vaccine development. Vaccines (Basel) 8(1):63.

    CAS  Article  Google Scholar 

  5. 5.

    Guy B, Guirakhoo F, Barban V, Higgs S, Monath TP, Lang J (2010) Preclinical and clinical development of YFV 17D-based chimeric vaccines against dengue, West Nile and Japanese encephalitis viruses. Vaccine 28(3):632–649.

    CAS  Article  PubMed  Google Scholar 

  6. 6.

    Rosa BR, Cunha A, Medronho RA (2019) Efficacy, immunogenicity and safety of a recombinant tetravalent dengue vaccine (CYD-TDV) in children aged 2–17 years: systematic review and meta-analysis. BMJ Open 9(3):e019368.

    Article  PubMed  PubMed Central  Google Scholar 

  7. 7.

    Brewoo JN, Kinney RM, Powell TD, Arguello JJ, Silengo SJ, Partidos CD, Huang CYH, Stinchcomb DT, Osorio JE (2012) Immunogenicity and efficacy of chimeric dengue vaccine (DENVax) formulations in interferon-deficient AG129 mice. Vaccine 30(8):1513–1520.

    CAS  Article  PubMed  Google Scholar 

  8. 8.

    Durbin AP, Kirkpatrick BD, Pierce KK, Elwood D, Larsson CJ, Lindow JC, Tibery C, Sabundayo BP, Shaffer D, Talaat KR, Hynes NA, Wanionek K, Carmolli MP, Luke CJ, Murphy BR, Subbarao K, Whitehead SS (2013) A single dose of any of four different live attenuated tetravalent dengue vaccines is safe and immunogenic in flavivirus-naive adults: a randomized, double-blind clinical trial. J Infect Dis 207(6):957–965.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Biswal S, Reynales H, Saez-Llorens X, Lopez P, Borja-Tabora C, Kosalaraksa P, Sirivichayakul C, Watanaveeradej V, Rivera L, Espinoza F, Fernando L, Dietze R, Luz K, Venâncio da Cunha R, Jimeno J, López-Medina E, Borkowski A, Brose M, Rauscher M, LeFevre I, Bizjajeva S, Bravo L, Wallace D (2019) Efficacy of a tetravalent dengue vaccine in healthy children and adolescents. N Engl J Med 381(21):2009–2019.

    CAS  Article  PubMed  Google Scholar 

  10. 10.

    Kirkpatrick BD, Durbin AP, Pierce KK, Carmolli MP, Tibery CM, Grier PL, Hynes N, Diehl SA, Elwood D, Jarvis AP, Sabundayo BP, Lyon CE, Larsson CJ, Jo M, Lovchik JM, Luke CJ, Walsh MC, Fraser EA, Subbarao K, Whitehead SS (2015) Robust and balanced immune responses to all 4 dengue virus serotypes following administration of a single dose of a live attenuated tetravalent dengue vaccine to healthy, flavivirus-naive adults. J Infect Dis 212(5):702–710.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  11. 11.

    Rivino L (2016) T cell immunity to dengue virus and implications for vaccine design. Exp Rev Vacc 15(4):443–453.

    CAS  Article  Google Scholar 

  12. 12.

    Sant AJ, McMichael A (2012) Revealing the role of CD4+ T cells in viral immunity. J Exp Med 209(8):1391–1395.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Tian Y, Grifoni A, Sette A, Weiskopf D (2019) Human T cell response to dengue virus infection. Front Immunol. 10:2125.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Kurane I, Meager A, Ennis FA (1989) Dengue virus-specific human T cell clones. Serotype crossreactive proliferation, interferon gamma production, and cytotoxic activity. J Exp Med 170(3):763–775.

    CAS  Article  PubMed  Google Scholar 

  15. 15.

    Mathew A, Kurane I, Rothman AL, Zeng LL, Brinton MA, Ennis FA (1996) Dominant recognition by human CD8+ cytotoxic T lymphocytes of dengue virus nonstructural proteins NS3 and NS12a. J Clin Invest. 98(7):1684–1691.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Gagnon SJ, Ennis FA, Rothman AL (1999) Bystander target cell lysis and cytokine production by dengue virus-specific human CD4+ cytotoxic T-lymphocyte clones. J Virol 73(5):3623

    CAS  Article  Google Scholar 

  17. 17.

    Hatch S, Endy TP, Thomas S, Mathew A, Potts J, Pazoles P, Libraty DH, Gibbons R, Rothman AL (2011) Intracellular cytokine production by dengue virus–specific T cells correlates with subclinical secondary infection. J Infect Dis 203(9):1282–1291.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  18. 18.

    de Matos AM, Carvalho KI, Rosa DS, Villas-Boas LS, da Silva WC, Rodrigues CL, Oliveira OM, Levi JE, Araújo ES, Pannuti CS, Luna EJ, Kallas EG (2015) CD8+ T lymphocyte expansion, proliferation and activation in dengue fever. PLoS Negl Trop Dis 9(2):e0003520.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Alves RPdS, Pereira LR, Fabris DLN, Salvador FS, Santos RA, Zanotto PMdA, Romano CM, Amorim JH, Ferreira LCdS (2016) Production of a recombinant dengue virus 2 NS5 protein and potential use as a vaccine antigen. Clin Vaccine Immunol 23(6):460.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Tian Y, Babor M, Lane J, Seumois G, Liang S, Goonawardhana NDS, De Silva AD, Phillips EJ, Mallal SA, da Silva AR, Grifoni A, Vijayanand P, Weiskopf D, Peters B, Sette A (2019) Dengue-specific CD8+ T cell subsets display specialized transcriptomic and TCR profiles. J Clin Invest 130(4):1727–1741.

    Article  Google Scholar 

  21. 21.

    Yauch LE, Zellweger RM, Kotturi MF, Qutubuddin A, Sidney J, Peters B, Prestwood TR, Sette A, Shresta S (2009) A protective role for dengue virus-specific CD8 T cells. J Immunol 182(8):4865.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Yauch LE, Prestwood TR, May MM, Morar MM, Zellweger RM, Peters B, Sette A, Shresta S (2010) CD4 T cells are not required for the induction of dengue virus-specific CD8 T cell or antibody responses but contribute to protection after vaccination. J Immunol 185(9):5405.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  23. 23.

    Zompi S, Santich BH, Beatty PR, Harris E (2012) Protection from secondary dengue virus infection in a mouse model reveals the role of serotype cross-reactive B and T cells. J Immunol 188(1):404.

    CAS  Article  PubMed  Google Scholar 

  24. 24.

    Pinto PBA, Assis ML, Vallochi AL, Pacheco AR, Lima LM, Quaresma KRL, Pereira BAS, Costa SM, Alves AMB (2019) T cell responses induced by DNA vaccines based on the DENV2 E and NS1 proteins in mice: importance in protection and immunodominant epitope Identification. Front Immunol 10:1522.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Weiskopf D, Yauch LE, Angelo MA, John DV, Greenbaum JA, Sidney J, Kolla RV, De Silva AD, de Silva AM, Grey H, Peters B, Shresta S, Sette A (2011) Insights into HLA-restricted T cell responses in a novel mouse model of dengue virus infection point toward new implications for vaccine design. J Immunol. 187(8):4268–4279.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  26. 26.

    Zellweger RM, Tang WW, Eddy WE, King K, Sanchez MC, Shresta S (2015) CD8 T cells can mediate short-term protection against heterotypic dengue virus reinfection in mice. J Virol 89(12):6494.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  27. 27.

    Li W, Joshi MD, Singhania S, Ramsey KH, Murthy AK (2014) Peptide vaccine: progress and challenges. Vaccines (Basel) 2(3):515–536.

    CAS  Article  Google Scholar 

  28. 28.

    Khan AM, Heiny AT, Lee KX, Srinivasan KN, Tan TW, August JT, Brusic V (2006) Large-scale analysis of antigenic diversity of T-cell epitopes in dengue virus. BMC Bioinform 7(Suppl 5):S4.

    Article  Google Scholar 

  29. 29.

    Jin X, Newman MJ, De-Rosa S, Cooper C, Thomas E, Keefer M, Fuchs J, Blattner W, Livingston BD, McKinney DM, Noonan E, Decamp A, Defawe OD, Wecker M (2009) A novel HIV T helper epitope-based vaccine elicits cytokine-secreting HIV-specific CD4+ T cells in a Phase I clinical trial in HIV-uninfected adults. Vaccine 27(50):7080–7086.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Muderspach L, Wilczynski S, Roman L, Bade L, Felix J, Small LA, Kast WM, Fascio G, Marty V, Weber J (2000) A phase I trial of a human papillomavirus (HPV) peptide vaccine for women with high-grade cervical and vulvar intraepithelial neoplasia who are HPV 16 positive. Clin Cancer Res 6(9):3406–3416

    CAS  PubMed  Google Scholar 

  31. 31.

    Dong X-N, Wei K, Liu Z-Q, Chen Y-H (2002) Candidate peptide vaccine induced protection against classical swine fever virus. Vaccine 21(3):167–173.

    CAS  Article  PubMed  Google Scholar 

  32. 32.

    Hiremath J, Kang K-i, Xia M, Elaish M, Binjawadagi B, Ouyang K, Dhakal S, Arcos J, Torrelles JB, Jiang X, Lee CW, Renukaradhya GJ (2016) Entrapment of H1N1 influenza virus derived conserved peptides in PLGA nanoparticles enhances T cell response and vaccine efficacy in pigs. PLoS ONE 11(4):e0151922–e0151922.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  33. 33.

    Purcell AW, McCluskey J, Rossjohn J (2007) More than one reason to rethink the use of peptides in vaccine design. Nat Rev Drug Discovery 6(5):404–414.

    CAS  Article  PubMed  Google Scholar 

  34. 34.

    Robinson J, Halliwell JA, Hayhurst JD, Flicek P, Parham P, Marsh SG (2015) The IPD and IMGT/HLA database: allele variant databases. Nucleic Acids Res 43(Database issue):D423–431. doi:

  35. 35.

    Roche PA, Furuta K (2015) The ins and outs of MHC class II-mediated antigen processing and presentation. Nat Rev Immunol 15(4):203–216.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Crotzer VL, Blum JS (2009) Autophagy and its role in MHC-mediated antigen presentation. J Immunol. 182(6):3335–3341.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Weiskopf D, Angelo MA, de Azeredo EL, Sidney J, Greenbaum JA, Fernando AN, Broadwater A, Kolla RV, De Silva AD, de Silva AM, Mattia KA, Doranz BJ, Grey HM, Shresta S, Peters B, Sette A (2013) Comprehensive analysis of dengue virus-specific responses supports an HLA-linked protective role for CD8+ T cells. Proc Natl Acad Sci USA 110(22):E2046–E2053.

    Article  PubMed  Google Scholar 

  38. 38.

    Ali M, Pandey RK, Khatoon N, Narula A, Mishra A, Prajapati VK (2017) Exploring dengue genome to construct a multi-epitope based subunit vaccine by utilizing immunoinformatics approach to battle against dengue infection. Sci Rep 7(1):9232.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Shi J, Sun J, Wu M, Hu N, Li J, Li Y, Wang H, Hu Y (2015) Inferring protective CD8+ T-cell epitopes for NS5 protein of four serotypes of dengue virus chinese isolates based on HLA-A, -B and -C allelic distribution: implications for epitope-based universal vaccine design. PLoS ONE 10(9):e0138729.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Chong LC, Khan AM (2019) Identification of highly conserved, serotype-specific dengue virus sequences: implications for vaccine design. BMC Genomics 20(Suppl 9):921–921.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Appanna R, Huat TL, See LL, Tan PL, Vadivelu J, Devi S (2007) Cross-reactive T-cell responses to the nonstructural regions of dengue viruses among dengue fever and dengue hemorrhagic fever patients in Malaysia. Clin Vacc Immunol 14(8):969–977.

    CAS  Article  Google Scholar 

  42. 42.

    Khan AM, Miotto O, Nascimento EJ, Srinivasan KN, Heiny AT, Zhang GL, Marques ET, Tan TW, Brusic V, Salmon J, August JT (2008) Conservation and variability of dengue virus proteins: implications for vaccine design. PLoS Negl Trop Dis 2(8):e272.

  43. 43.

    Bui HH, Sidney J, Peters B, Sathiamurthy M, Sinichi A, Purton KA, Mothé BR, Chisari FV, Watkins DI, Sette A (2005) Automated generation and evaluation of specific MHC binding predictive tools: ARB matrix applications. Immunogenetics 57(5):304–314.

    CAS  Article  PubMed  Google Scholar 

  44. 44.

    Bian H, Hammer J (2004) Discovery of promiscuous HLA-II-restricted T cell epitopes with TEPITOPE. Methods 34(4):468–475.

    CAS  Article  PubMed  Google Scholar 

  45. 45.

    Nascimento EJM, Mailliard RB, Khan AM, Sidney J, Sette A, Guzman N, Paulaitis M, Melo ABd, Cordeiro MT, Gil LVG, Lemonnier F, Rinaldo C, August JT, Marques ETA Jr (2013) Identification of conserved and HLA promiscuous DENV3 T-cell epitopes. PLoS Negl Trop Dis 7(10):e2497.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  46. 46.

    Rivino L, Kumaran EA, Jovanovic V, Nadua K, Teo EW, Pang SW, Teo GH, Gan VC, Lye DC, Leo YS, Hanson BJ, Smith KG, Bertoletti A, Kemeny DM, MacAry PA (2013) Differential targeting of viral components by CD4+ versus CD8+ T lymphocytes in dengue virus infection. J Virol 87(5):2693–2706.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Li S, Peng L, Zhao W, Zhong H, Zhang F, Yan Z, Cao H (2011) Synthetic peptides containing B- and T-cell epitope of dengue virus-2 E domain III provoked B- and T-cell responses. Vaccine 29(20):3695–3702.

    CAS  Article  PubMed  Google Scholar 

  48. 48.

    Rocha RP, Livonesi MC, Fumagalli MJ, Rodrigues NF, da Costa LC, Dos Santos MC, de Oliveira Rocha ES, Kroon EG, Malaquias LC, Coelho LF (2014) Evaluation of tetravalent and conserved synthetic peptides vaccines derived from Dengue virus Envelope domain I and II. Virus Res 188:122–127.

    CAS  Article  PubMed  Google Scholar 

  49. 49.

    Piazza P, Campbell D, Marques E, Hildebrand WH, Buchli R, Mailliard R, Rinaldo CR (2014) Dengue virus-infected human dendritic cells reveal hierarchies of naturally expressed novel NS3 CD8 T cell epitopes. Clin Exp Immunol 177(3):696–702.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  50. 50.

    Weiskopf D, Cerpas C, Angelo MA, Bangs DJ, Sidney J, Paul S, Peters B, Sanches FP, Silvera CGT, Costa PR, Kallas EG, Gresh L, de Silva AD, Balmaseda A, Harris E, Sette A (2015) Human CD8+ T-cell responses against the 4 dengue virus serotypes are associated with distinct patterns of protein targets. J Infect Dis 212(11):1743–1751.

    Article  PubMed  PubMed Central  Google Scholar 

  51. 51.

    Hertz T, Beatty PR, MacMillen Z, Killingbeck SS, Wang C, Harris E (2017) Antibody epitopes identified in critical regions of dengue virus nonstructural 1 protein in mouse vaccination and natural human infections. Journal of immunology (Baltimore, Md : 1950) 198 (10):4025–4035. doi:

  52. 52.

    Hunt DF, Michel H, Dickinson TA, Shabanowitz J, Cox AL, Sakaguchi K, Appella E, Grey HM, Sette A (1992) Peptides presented to the immune system by the murine class II major histocompatibility complex molecule I-Ad. Science 256(5065):1817.

    CAS  Article  PubMed  Google Scholar 

  53. 53.

    Cox AL, Skipper J, Chen Y, Henderson RA, Darrow TL, Shabanowitz J, Engelhard VH, Hunt DF, Slingluff CL (1994) Identification of a peptide recognized by five melanoma-specific human cytotoxic T cell lines. Science 264(5159):716.

    CAS  Article  PubMed  Google Scholar 

  54. 54.

    Testa JS, Shetty V, Hafner J, Nickens Z, Kamal S, Sinnathamby G, Philip R (2012) MHC class I-presented T cell epitopes identified by immunoproteomics analysis are targets for a cross reactive influenza-specific T cell response. PLoS ONE 7(11):e48484.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  55. 55.

    Testa JS, Shetty V, Sinnathamby G, Nickens Z, Hafner J, Kamal S, Zhang X, Jett M, Philip R (2012) Conserved MHC class I-presented dengue virus epitopes identified by immunoproteomics analysis are targets for cross-serotype reactive T-cell response. J Infect Dis 205(4):647–655.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  56. 56.

    Bassani-Sternberg M, Barnea E, Beer I, Avivi I, Katz T, Admon A (2010) Soluble plasma HLA peptidome as a potential source for cancer biomarkers. Proc Natl Acad Sci 107(44):18769.

    Article  PubMed  Google Scholar 

  57. 57.

    Ternette N, Yang H, Partridge T, Llano A, Cedeño S, Fischer R, Charles P, Dudek N, Mothe B, Crespo M, Fischer W, Korber B, Nielsen M, Borrow P, Purcell A, Brander C, Dorrell L, Kessler B, Hanke T (2015) Defining the HLA class I-associated viral antigen repertoire from HIV-1-infected human cells. Eur J Immunol.

    Article  PubMed  PubMed Central  Google Scholar 

  58. 58.

    Comber JD, Karabudak A, Huang X, Piazza PA, Marques ET, Philip R (2014) Dengue virus specific dual HLA binding T cell epitopes induce CD8+ T cell responses in seropositive individuals. Human Vacc Immunother 10(12):3531–3543.

    Article  Google Scholar 

  59. 59.

    Purcell AW, Ramarathinam SH, Ternette N (2019) Mass spectrometry–based identification of MHC-bound peptides for immunopeptidomics. Nat Protoc 14(6):1687–1707.

    CAS  Article  PubMed  Google Scholar 

  60. 60.

    Shetty V, Nickens Z, Testa J, Hafner J, Sinnathamby G, Philip R (2012) Quantitative immunoproteomics analysis reveals novel MHC class I presented peptides in cisplatin-resistant ovarian cancer cells. J Proteom 75(11):3270–3290.

    CAS  Article  Google Scholar 

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This study was funded by the Sunway University Internal Grant 2020 (GRTIN-RSF-SST-CVVR-01-2020) and Sunway University Research Centre Grant 2020 (STR-RCTR-CVVR-01-2020) to the Centre for Virus and Vaccine Research (CVVR).

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Lim, H.X., Lim, J. & Poh, C.L. Identification and selection of immunodominant B and T cell epitopes for dengue multi-epitope-based vaccine. Med Microbiol Immunol 210, 1–11 (2021).

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  • CD4+ T cell
  • CD8+ T cell
  • B cell
  • Epitope
  • Dengue
  • Vaccine