Tumor selective monoclonal antibodies chemically linked to toxin polypeptides (IMT) inhibit the growth of respective cancer cells. While these conjugates are very efficient in eliminating target tumor cells in vitro, their application in vivo is limited due to i) variable cytotoxicity, ii) escape of a minor population of cells from IMT treatment, iii) instability of the conjugate in circulation and iv) reduced tumor localization. The last two issues are critical for the systemic application of IMT but could be circumvent in treatment of tumors confined to restricted anatomical space like the peritoneum (e.g. ovarian epithelial carcinoma). Many IMTs have been prepared in the recent past that are selective against breast and ovarian cancer cells. Some of these conjugates made from recombinant ricin A-chain are inactive inspite of efficient internalization. The chemical linkage between antibody and the toxin moiety also plays a critical role in determining the cytotoxicity. Therefore, in the proposed study, attempts will be made to understand the reasons for reduced immunotoxin activity and develop methods to overcome the lack of efficacy. Earlier studies have shown that the presence of lysosomotropic reagents increase the cytotoxicity of antibody- toxin conjugates. However, this approach could not be used in vivo due to faster clearance of the potentiating compound. Thus, conjugation of lysosomotropic reagent like monensin to macromolecules could increase its biological half-life and in turn could improve the efficacy of IMT in vivo. Monensin would be directed to tumor cells via a second antibody (carrier) recognizing a distinct antigen. Alternatively, monensin would be encapsulated in immunoliposomes for the specific delivery to tumor cells. Improved tumor cell elimination could also be achieved by targeting toxin molecules via dual epitopes present on the antigen or via two distinct antigens present on the cancer cells. As toxin polypeptides differ in their mode and site of action, it may be possible to improve the IMT activity by linking two toxin moieties to antibodies. These reagents will be tested for their ability to enhance tumor cell elimination in vitro and in vivo in a nude mouse xenograft tumor model system. Like in chemotherapy, a small fraction of tumor cells do escape IMT treatment. Investigations will be carried out to understand the mechanism of resistance and based on the above work, methods to inhibit the growth of these tumor cell variants will be studied.
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