Targeted macrophage cytotoxicity using a nonreplicative live vector expressing a tumor-specific single-chain variable region fragment

Antigen-specific recognition and subsequent destruction of tumor cells is t he goal of vaccine-based immunotherapy of cancer. Often, however, tumor ant igen-specific cytotoxic T lymphocytes (CTLs) are either not available or in a state of anergy. In addition, MHCI expression on tumor cells is often do wnregulated. Either or both of these situations can allow tumor growth to p roceed unchecked by CTL control. We have shown previously that tumor antige n-specific monoclonal antibodies can be expressed in vaccinia virus and tha t activated macrophages infected with this virus acquire the ability to kil l tumor cells expressing that antigen. Here we show that a membrane-anchore d form of the scFv portion of the MUC1 tumor antigen-specific monoclonal an tibody, SM3, can be expressed on activated macrophages with the highly atte nuated poxvirus, modified vaccinia Ankara (MVA), as a gene transfer vector. Cells infected with the MVA-scFv construct were shown to express the membr ane-bound scFv by Western blot and FACS analysis. That cells expressing the membrane-anchored scFv specifically bind antigen was shown by FACS and by BIAcore analysis. GM-CSF-activated macrophages were infected with the const ruct and shown to recognize specifically MUC1-expressing tumor cells as mea sured by IL-12 release. Furthermore, activated macrophages expressing the m embrane-bound scFv specifically lyse target cells expressing the MUC1 antig en but not cells that do not express MUC1.