Basic StudiesCytotoxicity of Antiosteosarcoma Recombinant Immunotoxins Composed of TP-3 Fv Fragments and a Truncated Pseudomonas Exotoxin AOnda, Masanori*; Olafsen, Tove*; Tsutsumi, Yasuo*; Bruland, Øyvind S.†; Pastan, Ira* Author Information *Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, U.S.A.; and †Department of Oncology, The Norwegian Radium Hospital, Oslo, Norway. Received September 7, 2000; accepted November 9, 2000. Address correspondence and reprint requests to Dr. Ira Pastan, Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Building 37, Room 4E16, 37Convent Drive MSC 4255, Bethesda, MD 20892-4255, USA. Journal of Immunotherapy 24(2):p 144-150, March 2001. Buy Abstract Regrowth of drug-resistant tumor cells is responsible for approximately half of an unselected osteosarcoma population still dying of the disease despite aggressive combination therapy. Two monoclonal antibodies, TP-1 (immunoglobulin 2a) and TP-3 (immunoglobulin 2b) are available, which specifically recognize an antigen on osteosarcoma cells. In this work, we have fused the variable (V) genes of TP-3 to a truncated fragment of Pseudomonas exotoxin A, referred to as PE38. Two immunotoxins were made that differed in the Fv portion: TP-3(scFv)-PE38, which contains a peptide linker, and TP-3(dsFv)-PE38, which contains a disulfide bond for stabilization of the association between the V domains. Recombinant TP-3 immunotoxins were expressed in Escherichia coli and purified from inclusion bodies. We describe the design and expression of these immunotoxins, and their properties with regard to antigen binding, stability, and cytotoxicity. Toxicity studies were done in mice. We found that the immunotoxins exhibited very similar in vitro properties, whereas in vivo TP-3(dsFv)-PE38 was much better tolerated than TP-3(scFv)-PE38. © 2001 Lippincott Williams & Wilkins, Inc.