T cell responses to the human papillomavirus type 16 E7 protein in mice of different haplotypes

Abstract

The response of murine T cells to the E7 molecule of human papillomavirus type 16 (HPV-16) was studied using eight different mouse strains of six distinct H-2 haplotypes. HPV-16 E7 protein was prepared as a fusion protein with glutathione-S-transferase, purified by affinity chromatography and used for immunization. Cells from the lymph nodes were cultured with whole fusion protein, glutathione-S-transferase or HPV-16 E7 protein synthetic peptides. All the mouse strains tested, with the exception of BALB/c, recognized the E7 molecule, as evidenced by a proliferative response to at least two of the peptides. The profile of responses to peptides varied between and within a strain, but five distinct immunodominant regions could be identified. These regions were defined on the basis of a reaction to one or more peptides in a given part of the E7 molecule by at least four strains. The five regions were encompassed by amino acid residues 1 to 9, 17 to 32, 42 to 59, 62 to 77 and 87 to 98. The findings suggest that in an outbred population, such as man, the E7 molecule of HPV-16 would be recognized by a large proportion of the population. However, the poor response of two mouse strains [B10.RIII (71NS) and BALB/c] could also have a corollary in man.Certain human papillomavirus (HPV) types have been implicated in the aetiology of benign and malignant lesions of the ano-genital region. We are interested in identifying immune responses to one of the more important HPV types, namely HPV-16. Little is known about the immunobiology of HPV-16 because the virus cannot easily be grown in vitro or in vivo, and few virions can be isolated from genital lesions. To circumvent these problems, both early and late regions of the HPV-16 genome have been expressed as bacterial fusion proteins (Jenison et al., 1989;Jochmus-Kudielka et al., 1989).

Cell-mediated immune responses to E7 peptides of human papillomavirus (HPV) type 16 are dependent on the HPV type infecting the cervix whereas serological reactivity is not type-specific

“…The 'public' T cell epitope DRAHYNI (48-54) described by Tindle et al (1991) was included within one of the peptides (105) frequently recognized by our patients in an HPV-16 group-specific manner. The HPV-16 E7 T cell epitopes T1 (2(~29), T2 (45-54), T3 (6(~79) and T4 (85-94) identified in studies of mice immunized with E7 fusion proteins by Comerford et al (1991) and peptides 17-32, 42-59, 62-77 and 87-98 observed by Shepherd et al (1992) were similar to those observed in this study of human 'naturally induced' LP responses to HPV-16 E7. HPV-16 type-specific delayed hypersensitivity reactions mediated by CD4 + T cells have recently been reported in mice engrafted with syngeneic keratinocytes transfected with HPV-16 E7 DNA (McLean et al, 1993), supporting the type-specific CMI reactivity reported here for humans with natural HPV infection.…”

Proliferative T cell responses to the human papillomavirus type 16 E7 protein in women with cervical dysplasia and cervical carcinoma and in healthy individuals

“…The HPV-16 GST-E7 fusion protein was induced in Escherichia coli transformed with the plasmid pGEX-2T.HPV-16E7 (Smith & Johnson, 1988;Comerford, 1991) and a crude extract of insoluble inclusion bodies prepared according to the method of Marston (i986). The protein was then isolated essentially as previously described (Shepherd et aL, 1992). Briefly, the inclusion bodies were solubilized in 8 M-urea then diluted in nine volumes of phosphate buffer (50 mMKHaPO 4, 100 mM-EDTA, 1 M-NaC1 pH 10'7).…”

“…After thorough washing with PBS plus 1% Triton X-I00, GST-E7 was eluted by stepwise addition of elution buffer (50 mM-Tris-HC1, 5 mM reduced glutathione in PBS pH 7"5). Fractions were analysed for the presence of the protein by standard Western blotting procedures as previously described (Shepherd et al, 1992).…”

“…A number of studies of both human and murine responses have concentrated on the identification of cytotoxic T cell epitopes within this protein (Kast et al, 1993(Kast et al, , 1994Sadovnikova et al, 1994) and immunization of mice with peptides derived from the E6 and E7 proteins has been demonstrated to protect against challenge with HPV-16-transformed tumour cells (Feltkamp et al, 1993). Studies of proliferative T cell responses to HPV-16 E7 in mice 58[ 0001-3867 © 1996 SGM have identified epitopes spanning almost the entire molecule (Shepherd et aL, 1992;Comerford eta]., 1991; Tindle e~ a]., 199i). A 'promiscuous' T helper cell epitope between amino acid residues 48-54 (DRAHYNI) was identified in mice which was capable of providing help for the production of several HPV-I6 ET-specific antibodies simultaneously (Tindle et al,I991).…”

Proliferative T cell responses to human papillomavirus type 16 L1 peptides in patients with cervical dysplasia

“…Most have used peptides or fusion proteins produced by bacterial expression systems as antigens and have focused on the immune response to HPV-16. Using this type of approach in mice, T cell epitopes in the HPV-16 L1 and E7 proteins have been described (Davies et al, 1990;Tindle et al, 1991;Comerford et al, 1991;Shepherd et at., 1992). In humans, epitopes in the HPV-16 E6, E7 and L1 proteins have been described (Strang et al, 1990;Altmann et al, 1992).…”