Identification and evaluation of protective activity of a T cell epitope targeting nucleoprotein of H5N1 influenza virus

Abstract

The outbreaks of human influenza caused by highly pathogenic avian influenza

H5N1 virus have attracted a lot of attention and public concern. Effective and

universal vaccines may be the best means for prevention and control of the influenza.

Taking into account that viral clearance and recovery from influenza A virus infection

have been demonstrated to be correlated to specific cytoxic T lymphocyte (CTL)

instead of neutralizing antibodies, it is important to develop effective vaccines which

are capable of inducing not only neutralizing antibody but also CTL responses.

Furthermore, T cell epitopes are usually more conserved than neutralizing epitopes.

However, rare information concerning human T cell epitopes specific to H5N1 virus

has been reported so far.

This study was designed to test our hypothesis that novel and potent human CTL and

Th epitopes specific to NP protein of H5N1 virus may be identified in vaccinated

and/or infected HLA-A2/DR1 transgenic mice (SURE/L1), while protective epitopes

may be further defined from the identified T cell epitopes in the mice challenged with

lethal dose of the virus. We used SURE/L1 mouse model because it contains HLA-A2

(*0201) and -DR1 (*0701), both are the second highest frequency of HLA class I and

II in Chinese. Since the NP gene is relatively conserved among different clades or

strains of H5N1 virus, we selected viral protein NP as the target. Furthermore, we

screened the T cell epitopes in splenocytes not only from vaccinated mice but also

from survived mice infected with gradually increased dose of H5N1 virus, because

the T cell epitopes identified in both vaccinated and infected mice or in infected mice

alone might have higher potential to be protective epitopes.

In this study, a novel HLA-DR1 (class II) restricted T cell epitope NP368-382, NPII-7,

was identified in both vaccinated and infected mice. Two doses of NPII-7 peptide

boosting in the mice induced very strong Th1 and CTL responses but no NP specific

antibody responses. The vaccination of additional 2 doses of NPII-7 also provided

partial protection against lethal challenge of H5N1 virus in the mice, whereas NP

DNA vaccination alone did not show any protective effect. The protective effect may

be attributed to the strong Th1 and CTL responses induced by the NPII-7 vaccination,

because both NP DNA and NPII-7 vaccinations could not induce neutralizing

antibody response.

Notably, a HLA class II restricted peptide, NPII-7, may induce not only Th1

responses but also more strong HLA class I restricted T cell (CTL) responses. It may

probably due to that the HLA-DR1 restricted T cell epitope

(NENMEAMDSNTLELR) contained the full sequence of a reported HLA-A2

restricted CTL epitope (AMDSNTLEL), named NP-17 in this study. Although it

needs to be further defined whether this novel epitope is really a HLA-DR1 restricted

T cell epitope, or it shares the activity of HLA-A2 restricted T cell epitope, or it is

just an alternate HLA-A2 restricted T cell epitope, this study has identified a novel T

cell epitope and proved that it is a protective T cell epitope.