title: A mutant B7-1/Ig fusion protein that selectively binds to CTLA-4 ameliorates anti-tumor DNA vaccination and counters regulatory T cell activity.
abstract: We have shown that a plasmid encoding a B7-1/Ig fusion protein enhanced DNA vaccination against human carcinoembryonic antigen (CEA) more effectively than the plasmid encoding membrane-bound B7-1. However, it was not known if B7-1/Ig acted only by binding CD28 (amplifying a stimulatory signal) or by blocking CTLA-4 on T cells (removing inhibitory signals). Here, we aimed to determine this using a plasmid encoding mutant B7-1/Ig (B7-1wa/Ig), which binds only to CTLA-4 but not to CD28. Our results showed that both the B7-1/Ig and B7-1wa/Ig plasmids, when co-administered with a CEA plasmid, enhanced tumor rejection and the in vitro anti-CEA response. Therefore, B7-1wa/Ig ameliorates DNA vaccination, presumably by binding to CTLA-4. This could result from a number of non-exclusive mechanisms, such as a reduced threshold for T-cell activation, or blockade of CTLA-4/B7-mediated tolerogenic signals in DCs or T cells. We found that, in vitro, a significant fraction of CD3/CD28-activated T cells (in the absence of DCs) expressed CTLA-4 and B7-1. Primed T cells of CTLA-4(+)B7-1(+/-) phenotype acted as regulatory T cells by inhibiting IFNgamma production by re-stimulated CTLA-4(-)B7-1(-) cells, and this was reversed by antibodies against IL-10 or TGF-beta1. Both B7-1wa/Ig and CTLA-4/Ig, which bind to CTLA-4 and B7-1/B7-2 respectively, enhanced IFNgamma production, but not the proliferation or IL-4 release in mixed T-cell populations containing these two cell types. In contrast, CTLA-4(-)B7-1(-) T cells produced IFNgamma which was not affected by B7-1wa/Ig or CTLA-4/Ig. These results suggest that blocking of CTLA-4/B7-1 binding in T cell/T cell interactions blocks negative regulatory signals. This might be the mechanism, at least in part, of the enhancement of anti-tumor immunity by the B7-1wa/Ig and B7-1/Ig plasmids.
authors.affiliation: Department of Laboratory Medicine and Pathobiology, St. Michael's Hospital and University of Toronto, 30 Bond Street, Toronto, Ontario, Canada M5B 1W8. chakrabartir@smh.toronto.on.ca
authors.forename: Rabindranath
authors.initials: R
authors.lastname: Chakrabarti
authors.forename: Zheng F
authors.initials: ZF
authors.lastname: Zhou
authors.forename: Yigang
authors.initials: Y
authors.lastname: Chang
authors.forename: Gérald J
authors.initials: GJ
authors.lastname: Prud'homme
mesh_terms: D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000276:immunology; D018122:B7-1 Antigen / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D018106:CD28 Antigens / Q000276:immunology; D060908:CTLA-4 Antigen; D019496:Cancer Vaccines / Q000008:administration & dosage / Q000276:immunology; D004357:Drug Synergism; D018796:Immunoconjugates / Q000494:pharmacology*; D007136:Immunoglobulins / Q000235:genetics / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D051379:Mice; D008817:Mice, Mutant Strains / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology; D019444:Vaccines, DNA / Q000008:administration & dosage* / Q000276:immunology
chemical_list: D015703:Antigens, CD; D000943:Antigens, Differentiation; D018122:B7-1 Antigen; D018106:CD28 Antigens; D060908:CTLA-4 Antigen; C556706:CTLA4 protein, human; D019496:Cancer Vaccines; C556707:Ctla4 protein, mouse; D018796:Immunoconjugates; D007136:Immunoglobulins; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA