Scientific Journal of

  Abstract

  Background and Objectives

  Immunotoxins are comprised of cell targeting and cell killing moieties. Immunotoxins have been proposed as new strategy for cancer treatment. In this study, catalytic domain of Shiga-toxin (A1) was fused to human Granulocyte Macrophage-Colony Stimulating Factor (h GM-CSF) . The fused gene was already expressed in E.coli and the expressed protein was analysed for its cytotoxic activity on human cancer cell lines expressing GM-CSF receptor (GM-CSFR). Moreover, the impact of GM-CSF receptor on inhibition of cytotoxic effect of Shiga-toxin-GM-CSF recombinant hybrid in cell lines expressing GM-CSF receptor (GM-CSFR) was evaluated.

 Materials and Methods

  Cell lines were grown in RPMI-1640 medium supplemented with 10-20% heat inactivated fetal bovine serum (Gibco-BRL). Cytotoxic activity was checked on the cell lines and was measured by trypan blue staininig and MTT assay. GM-CSF receptor was blocked by anti-GM-CSF antibody.

  Results

  Cytotoxicity studies revealed that the chimeric protein induced cytotoxic effect on different cell lines. This effect was found to be specific due to the presence of killing moiety (A1) that exerts its effect through specific cell targeting domain i.e. GM-CSF, by binding to its receptor present on those cell lines. K562 and LS174T were the most sensitive .

  Conclusions

  Our results revealed that the hybrid protein is toxic to the cancer cell lines bearing GM-CSF receptor this effect could be a potential factor for further consideration of hybrid protein as a therapeutic agent.

  Key words: Shiga-toxin , Granulocyte macrophage - colony stimulating factor ( GM–CSF ) , Cytotoxicity, Cancer cells