C-Terminal Amidation of Chlorotoxin Does Not Affect Tumour Cell Proliferation and Has No Effect on Toxin Cytotoxicity

Abstract

Chlorotoxin (Cltx), a cell-penetrating peptide from the venom of the scorpion Leiurus quinquestriatus has selective activity on a range of neuroectodermal tumours and related cell lines. Although the precise mechanism of Cltx action is not clear, Cltx has been shown to be a useful tumour imaging agent and radiolabelled Cltx has undergone clinical trials for targeted glioma radiotherapy. The native peptide (and synthetic material, TM601) is amidated at its C-terminus (Cltx-CONH2). A recombinant form produced in E. coli is not processed and has a free C-terminal carboxyl group (Cltx-COOH). The amidated form appears to have no effects on proliferation and cytotoxicity, whereas studies on the free carboxyl form are contradictory. Cltx-CONH2 and Cltx-COOH have been used interchangeably over many years and this has resulted in confusion in the literature; the significance of any difference is extremely important one with respect to the use of the toxin in a clinical diagnostic or therapeutic setting. This study was carried out to investigate, in parallel, the effects of Cltx-CONH2 and Cltx-COOH on cell proliferation and cytotoxicity on a variety of representative tumour cell lines (U87MG, MCF-7, PC3 and A549) using cell Titer Glo and cell Tox Green assays, respectively. Neither Cltx-CONH2 nor Cltx-COOH affected cell proliferation or cytotoxicity, indicating that terminal arginine amidation does not play an important role in cytotoxic or cytostatic properties of Cltx.