Abstract
BackgroundChlorotoxin (TM601), a scorpion venom- derived 36-AA peptide, is an experimental drug against recurrent glioma with tumor specificity but unknown route of intracellular distribution. The aim of this study was to evaluate the route of entry and cellular localization of TM601 in glioma cells.ResultsWe have found that in human gliomas, lung carcinoma and normal vascular endothelial cells, TM601 localizes near trans-Golgi while in normal human dermal fibroblasts (NHDF) and astrocytes it is dispersed in the cytoplasm. The uptake of TM601 by U373 glioma cells is rapid, concentration and time dependent, not affected by inhibitors such as filipin (caveolae-dependent endocytosis) and amiloride (non-selective macropinocytosis), but significantly affected by chlorpromazine (clathrin-dependent intracellular transport of coated pits) resulting in intracellular build-up of the drug and clathrin near the Golgi. In contrast, TM601 uptake by NHDF cells was significantly affected by amiloride indicating that macropinocytosis is the dominant uptake route of TM601 in these cells.ConclusionsIn conclusion, we found a distinct cellular localization pattern and uptake of TM601 by glioma cells differing from that found in normal cells. Further insight into the cellular processing of TM601 should assist in the development of effective anti-glioma therapeutic modalities.
Highlights
Chlorotoxin (TM601), a scorpion venom- derived 36-AA peptide, is an experimental drug against recurrent glioma with tumor specificity but unknown route of intracellular distribution
We found that TM601 uptake by glioma cells resembles that of normal umbilical vascular endothelial cells, but is different from the one seen in normal astrocytes and fibroblasts
Clathrin-mediated endocytosis involves the concentration of high-affinity trans-membrane receptors and receptor- bound ligands into “coated pits” and adaptor protein-2 (AP-2) mediated clathrin binding to coated pits, followed by their maturation, fission and formation of clathrin-coated vesicles (CCVs)
Summary
Chlorotoxin (TM601), a scorpion venom- derived 36-AA peptide, is an experimental drug against recurrent glioma with tumor specificity but unknown route of intracellular distribution. TM601 is a pure, chemically synthesized chlorotoxin of 36 amino acids which was first purified from the venom of the scorpion Leiurus quinquestriatus. It was first found to be an inhibitor of small conductance chloride channels [2]. Both inhibition of invasion and inhibition of metalloproteinase-2 (MMP-2) activity have been previously observed in glioma cells treated with chlorotoxin [3,4]. A similar in vitro finding was reported for human umbilical vein endothelial cells (HUVEC) treated with TM601 [5]. TM601 is not cytostatic or cytotoxic to tumor or vascular endothelial cells in vitro
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