PO-420 Mechanisms of action of anti-proliferative lichen compound protolichesterinic acid

Abstract

Introduction Protolichesterinic acid (PA), an aliphatic α-methylene γ-lactone isolated from the lichen Iceland moss ( Cetraria islandica ) has an anti-proliferative effect against a variety human cancer cells. In multiple myeloma cells PA induces apoptosis. The anti-proliferative effect of PA is not mediated by the known inhibitory action of PA on 5- and 12-lipoxygenases and effects on growth factor signalling are secondary. PA inhibits DNA replication and earlier studies indicated direct effects of PA on HIV reverse transcriptase (RT) and DNA polymerase. RT inhibitors are known to affect mitochondria. Material and methods Molecular modelling software (Glide, Schrodinger, LLC) was used to test for the ability of PA to bind onto DNA polymerases α, β, γ and θ. The crystal structures of polymerases were acquired from Protein Data Bank (PDB ID: 4QCL, 5TB8, 4ZTZ and 5A9F, respectively) and PA was docked into each structure. The effects of PA on mitochondria were assessed morphologically by electron microscopy and metabolically by measuring extracellular glucose and lactate concentrations in cell medium after 6 and 24 hour (ABL90 FLEX blood gas analyzer) in the pancreatic cancer cell line Aspc-1, the breast cancer cell line T47D, normal skin fibroblast and MCF-10 non-tumorigenic breast epithelial cell line. Results and discussions PA docked to all four types of DNA polymerases, creating multiple hydrogen bonds directly with the enzymes/polymerases and with the metal ion (Mg 2+ ) cofactor. Hydrogen bonds to the metal ions bridging the interaction to the enzyme appeared important and might be necessary/important for the activity of protolichesterinic acid and explain its mechanism of action. Morphological changes in mitochondria were observed following exposure to PA in both cancer cell lines, but these were difficult to interpret. Aspc-1 pancreatic cancer showed a significant increase in lactate production after 24 hour exposure to PA. This was not observed in the breast cell lines, T47D and MCF-10. Aspc-1 is less sensitive to the anti-proliferative effects of PA than the breast cell lines. Further metabolomics studies are in progress. Conclusion Docking study of crystal structures of DNA polymerases indicates that PA could affect activity of several types of DNA polymerase, but this remains to be tested experimentally. PA may affect mitochondrial structure but increased lactate production is associated with relative resistance to its anti-proliferative effects.