DEVELOPMENT OF LYMPHOID-SPECIFIC IMMUNOTOXINS

THE GOAL OF THE PROPOSED RESEARCH IS TO DEVELOP AN EXTENSIVEPANEL OF RICIN A CHAIN IMMUNOTOXINS WHICH ARE CYTOTOXIC AND SPECIFIC FOR SUBSETS OF HUMAN LYMPHOCYTES AND OTHER CELL TYPES OF IMMUNOLOGICAL IMPORTANCE SUCH AS GRANULOCYTES, MONOCYTES, AND ANTIGEN PRESENTING CELLS. THE SPECIFIC AIM OF THE RESEARCH IS TO PREPARE RICIN A CHAIN IMMUNOTOXINS WITH A PANEL OF SEVENTEEN MURINE ANTI-HUMAN MONOCLONAL ANTIBODIES, AND TO TEST THE CYTOTOXICITY OF THESE IMMUNOTOXINS ON HUMAN CELL LINES WHICH EXPRESS THE TARGET ANTIGENS. THE FIRST STEP WILL BE TO SCREEN THE MONOCLONAL ANTIBODIES FOR THEIR ABILITY TO MEDIATE CYTOTOXICITY USING AN INDIRECT IMMUNOTOXIN CYTOTOXICITY ASSAY IN WHICH A SECONDARY IMMUNOTOXIN, PREPARED WITH FAB FRAGMENTS OF ANTI-MOUSE IGG ANTIBODY AND RICIN A CHAIN, IS USED TO TREAT TARGET CELLS COATED WITH THE PRIMARY MONOCLONAL ANTIBODY. IMMUNOTOXINS WILL BE PREPARED WITH THE MONOCLONAL ANTIBODIES WHICH ARE CYTOTOXIC IN THE INDIRECT ASSAY BY CONJUGATION WITH RICIN A CHAIN USING THE HETEROBIFUNCTIONAL CROSSLINKER, N-SUCCINIMIDYL 3-(2-PYRDYLDITHIO) PROPIONATE. THESE IMMUNOTOXINS WILL BE TESTED IN DIRECT CYTOTOXICITY ASSAYS. BOTH DIRECT AND INDIRECT CYTOTOXICITY ASSAYS WILL BE PERFORMED WITH IN VITRO HUMAN CELL LINES. THE COMMERCIALAVAILABILITY OF THESE REAGENTS WILL PROVIDE BOTH BASIC AND CLINICAL RESEARCHERS WITH A SAFE, SIMPLE, AND MORE EFFECTIVEMETHOD THAN IS CURRENTLY AVAILABLE FOR ELIMINATING SUBPOPULATIONS OF CELLS IN VITRO. THE LONG TERM OBJECTIVE IS TO DEVELOP, AND MAKE COMMERCIALLY AVAILABLE, IMMUNOTOXINSWHICH CAN BE UTILIZED FOR SPECIFIC IN VIVO THERAPY OF HUMAN MALIGNANCY, AUTOIMMUNITY, ACQUIRED IMMUNE DEFICIENCY SYNDROME (AIDS) AND OTHER DISEASES.