title: Clearance mechanism of a mannosylated antibody-enzyme fusion protein used in experimental cancer therapy.
abstract: MFECP1 is a mannosylated antibody-enzyme fusion protein used in antibody-directed enzyme prodrug therapy (ADEPT). The antibody selectively targets tumor cells and the targeted enzyme converts a prodrug into a toxic drug. MFECP1 is obtained from expression in the yeast Pichia pastoris and produced to clinical grade. The P. pastoris-derived mannosylation of the fusion protein aids rapid normal tissue clearance required for successful ADEPT. The work presented provides evidence that MFECP1 is cleared by the endocytic and phagocytic mannose receptor (MR), which is known to bind to mannose-terminating glycans. MR-transfected fibroblast cells internalize MFECP1 as revealed by flow cytometry and confocal microscopy. Immunofluorescence microscopy shows that in vivo clearance in mice occurs predominantly by MR on liver sinusoidal endothelial cells, although MR is also expressed on adjacent Kupffer cells. In the spleen, MFECP1 is taken up by MR-expressing macrophages residing in the red pulp and not by dendritic cells which are found in the marginal zone and white pulp. Clearance can be inhibited in vivo by the MR inhibitor mannan as shown by increased enzyme activities in blood. The work improves understanding of interactions of MFECP1 with normal tissue, shows that glycosylation can be exploited in the design of recombinant anticancer therapeutics and opens the ways for optimizing pharmacokinetics of mannosylated recombinant therapeutics.
authors.affiliation: Cancer Research UK Targeting and Imaging Group, Department of Oncology, Royal Free & University College Medical School, London, UK.
authors.forename: Heide
authors.initials: H
authors.lastname: Kogelberg
authors.forename: Berend
authors.initials: B
authors.lastname: Tolner
authors.forename: Surinder K
authors.initials: SK
authors.lastname: Sharma
authors.forename: Mark W
authors.initials: MW
authors.lastname: Lowdell
authors.forename: Uzma
authors.initials: U
authors.lastname: Qureshi
authors.forename: Mathew
authors.initials: M
authors.lastname: Robson
authors.forename: Tim
authors.initials: T
authors.lastname: Hillyer
authors.forename: R Barbara
authors.initials: RB
authors.lastname: Pedley
authors.forename: Wouter
authors.initials: W
authors.lastname: Vervecken
authors.forename: Roland
authors.initials: R
authors.lastname: Contreras
authors.forename: Richard H J
authors.initials: RH
authors.lastname: Begent
authors.forename: Kerry A
authors.initials: KA
authors.lastname: Chester
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000493:pharmacokinetics / Q000627:therapeutic use; D002272:Carcinoembryonic Antigen / Q000276:immunology; D002478:Cells, Cultured; D018796:Immunoconjugates / Q000737:chemistry / Q000493:pharmacokinetics* / Q000627:therapeutic use*; D007167:Immunotherapy / Q000379:methods; D037181:Lectins, C-Type / Q000378:metabolism; D008099:Liver / Q000737:chemistry; D008351:Mannans / Q000494:pharmacology; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins / Q000378:metabolism; D008657:Metabolic Clearance Rate / Q000187:drug effects; D051379:Mice; D008807:Mice, Inbred BALB C; D009374:Neoplasms, Experimental / Q000628:therapy*; D011134:Polysaccharides / Q000032:analysis; D011355:Prodrugs / Q000138:chemical synthesis / Q000737:chemistry / Q000493:pharmacokinetics / Q000627:therapeutic use; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000493:pharmacokinetics* / Q000627:therapeutic use*; D013154:Spleen / Q000737:chemistry; D014018:Tissue Distribution; D014162:Transfection
chemical_list: D000970:Antineoplastic Agents; D002272:Carcinoembryonic Antigen; D018796:Immunoconjugates; D037181:Lectins, C-Type; D008351:Mannans; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D011134:Polysaccharides; D011355:Prodrugs; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins