Mycalamide A: Comparison between Caenorhabditis elegans and cultured mammalian cells. Searching for targeted therapeutics

DAVIES CAROLINA; TEESDALE-SPITTLE PAUL

Queenstown, New Zealand

Congreso; 14th Queenstown Molecular Biology Meeting; 2004

Queenstown Molecular Biology Society and New Zealand Society for Biochemistry and Molecular Biology

Mycalamide A is produced by the marine sponge Mycale sp and has cytotoxic and anti-viral properties, but its molecular targets, metabolism and pharmacokinetics remain unknown. In order to improve our understanding of these factors, cultured mammalian cells (HepG2 and HL60) and a multicellular organism, the fiee-living nematode C. elegans, were incubated with mycalamide. An ECSo (measuring population susceptibility to the toxin following a 48 hour incubation) was determined. The ECSo for the nematodes was 23.7 +/- 12.3 nM, whereas for HepG2 cells it was 3.4 +/- 2.2 nM and for HL60 was 2.4 +/- 1.3 nM. By fluorescence microscopy, following propidium iodide staining, the observed toxicity in C. elegans appeared in the pharynx and mouth cells, where P-gp is highly expressed but not MDR pumps. In contrast, there was little apparent toxicity in the gastrointestinal region of the nematode where MDR pumps are expressed. Incubation with verapamil, a Pgp pump inhibitor, and probenecid (PB), an inhibitor of MDR pumps, did not alter the staining pattern following mycalamide treatment of nematodes. This indicated that efflux pumps are not responsible for the regiospecific toxicity. Mycalamide is a known protein synthesis inhibitor and its targets must be ubiquitously expressed, however strong sequestration by pharyngeal cells, the first that become exposed to the toxin, could prevent its distribution throughout the nematode. In contrast, ivermectin (a nematocide) was widely distributed. In parallel experiments using verapamil and probenecid in combination with mycalamide in HL60 and HepG2 cells, it was shown that inhibition of P-gp and MDR pumps also has no influence on mycalamide cytotoxicity in mammalian cells.