title: Granzyme M as a novel effector molecule for human cytolytic fusion proteins: CD64-specific cytotoxicity of Gm-H22(scFv) against leukemic cells.
abstract: Immunotoxins are promising targeted therapeutic agents comprising an antibody-based ligand that specifically binds to diseased cells, and a pro-apoptotic protein. Toxic components from bacteria or plants can trigger a neutralizing immune response, so that human effector molecules are more suitable. In this context, the protease granzyme B has been successfully tested in cytotoxicity assays against different cancer cells in vitro and in vivo. Our aim here was to introduce granzyme M as an alternative and novel component of human cytolytic fusion proteins. We fused it to the humanized single-chain antibody fragment (scFv) H22 which specifically binds to CD64, an FcγRI receptor overexpressed on activated myeloid cells and leukemic cells. We show that the humanized cytolytic fusion protein Gm-H22(scFv) specifically targets the acute myeloid leukemia cell line HL60 in vitro and is cytotoxic with an IC50 between 1.2 and 6.4 nM. These findings were confirmed ex vivo using leukemic primary cells from patients, which were killed by granzyme M despite the presence of the granzyme B inhibitor serpin B9. In conclusion, granzyme M is a promising new cell-death inducing component for hCFPs because it specifically and efficiently kills target cells when fused to a targeting component.
authors: Department of Experimental Medicine and Immunotherapy, RWTH Aachen, Institute for Applied Medical Engineering, Aachen, Germany; Department of Pharmaceutical Product Development, Fraunhofer Institute for Molecular Biology and Applied Ecology, Aachen, Germany.
authors: Sonja
authors: S
authors: Schiffer
authors: Soriba
authors: S
authors: Letzian
authors: Edgar
authors: E
authors: Jost
authors: Radoslav
authors: R
authors: Mladenov
authors: Dmitrij
authors: D
authors: Hristodorov
authors: Michael
authors: M
authors: Huhn
authors: Rainer
authors: R
authors: Fischer
authors: Stefan
authors: S
authors: Barth
authors: Theo
authors: T
authors: Thepen
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000276:immunology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000276:immunology; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D053804:Granzymes / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D057809:HEK293 Cells; D018922:HL-60 Cells; D006801:Humans; D000922:Immunotoxins / Q000276:immunology* / Q000494:pharmacology; D020014:K562 Cells; D007938:Leukemia / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D017452:Receptors, IgG / Q000276:immunology / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000276:immunology* / Q000378:metabolism / Q000494:pharmacology; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D000922:Immunotoxins; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D053804:Granzymes
keywords: Immunotherapy; Immunotoxin; Serine protease; Serpin B9