title: Antibody-peptide-MHC fusion conjugates target non-cognate T cells to kill tumour cells.
abstract: Attempts to generate robust anti-tumour cytotoxic T lymphocyte (CTL) responses using immunotherapy are frequently thwarted by exhaustion and anergy of CTL recruited to tumour. One strategy to overcome this is to retarget a population of virus-specific CTL to kill tumour cells. Here, we describe a proof-of-principle study using a bispecific conjugate designed to retarget ovalbumin (OVA)-specific CTL to kill tumour cells via CD20. A single-chain trimer (SCT) consisting of MHCI H-2K(b)/SIINFEKL peptide/beta 2 microglobulin/BirA was expressed in bacteria, refolded and chemically conjugated to one (1:1; F2) or two (2:1; F3) anti-hCD20 Fab' fragments. In vitro, the [SCT × Fab'] (F2 and F3) redirected SIINFEKL-specific OT-I CTL to kill CD20(+) target cells, and in the presence of CD20(+) target cells to provide crosslinking, they were also able to induce proliferation of OT-I cells. In vivo, activated OT-I CTL could be retargeted to kill [SCT × Fab']-coated B cells from hCD20 transgenic (hCD20 Tg) mice and also EL4 and B16 mouse tumour cells expressing human CD20 (hCD20). Importantly, in a hCD20 Tg mouse model, [SCT × Fab'] administered systemically were able to retarget activated OT-I cells to deplete normal B cells, and their performance matched that of a bispecific antibody (BsAb) comprising anti-CD3 and anti-CD20. [SCT × Fab'] were also active therapeutically in an EL4 tumour model. Furthermore, measurement of serum cytokine levels suggests that [SCT × Fab'] are associated with a lower level of inflammatory cytokine release than the BsAb and so may be advantageous clinically in terms of reduced toxicity.
authors.affiliation: Antibody and Vaccine Group MP88, Cancer Sciences Unit, Faculty of Medicine, General Hospital, University of Southampton, Tremona Road, Southampton SO16 6YD, UK.
authors.forename: Ben C
authors.initials: BC
authors.lastname: King
authors.forename: Angela D
authors.initials: AD
authors.lastname: Hamblin
authors.forename: Philip M
authors.initials: PM
authors.lastname: Savage
authors.forename: Leon R
authors.initials: LR
authors.lastname: Douglas
authors.forename: Ted H
authors.initials: TH
authors.lastname: Hansen
authors.forename: Ruth R
authors.initials: RR
authors.lastname: French
authors.forename: Peter W M
authors.initials: PW
authors.lastname: Johnson
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Glennie
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000235:genetics / Q000276:immunology*; D018951:Antigens, CD20 / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism; D045744:Cell Line, Tumor; D003602:Cytotoxicity, Immunologic; D004195:Disease Models, Animal; D023061:Gene Order; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000276:immunology*; D006801:Humans; D018796:Immunoconjugates / Q000008:administration & dosage / Q000276:immunology*; D008213:Lymphocyte Activation / Q000276:immunology; D008212:Lymphocyte Depletion; D051379:Mice; D009369:Neoplasms / Q000188:drug therapy / Q000276:immunology* / Q000401:mortality; D010047:Ovalbumin / Q000276:immunology; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology
chemical_list: D018033:Antibodies, Bispecific; D018951:Antigens, CD20; D015395:Histocompatibility Antigens Class I; D018796:Immunoconjugates; D010455:Peptides; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D010047:Ovalbumin