Planta Med 2014; 80 - P1L85
DOI: 10.1055/s-0034-1394742

iTRAQ-based proteomic analysis of ellagitannins-enriched fraction from Fragaria vesca leaves on HepG2 cells

J Liberal 1, 2, 3, G Costa 1, 2, 3, C Marques 4, H Girão 4, R Vitorino 5, R Domingues 5, P Domingues 5, MT Cruz 1, 2, MT Batista 1, 2, 3
  • 1Center for Neuroscience and Cell Biology, University of Coimbra, Portugal
  • 2Faculty of Pharmacy, University of Coimbra, Coimbra, Portugal
  • 3Center for Pharmaceutical Studies, University of Coimbra, Portugal
  • 4Centre of Ophthalmology and Vision Sciences, Institute for Biomedical Imaging and Life Sciences, University of Coimbra, Portugal
  • 5Department of Chemistry, Mass Spectrometry Center, QOPNA, University of Aveiro, Portugal

The hepatocellular carcinoma is a primary malignancy of the liver with a very poor prognosis. The inefficacy and the side effects of the conventional therapies highlight the importance of discovering new compounds with anti-tumoral effects. We previously demonstrated that an ethanol crude extract from Fragaria vesca leaves modulated autophagy and ubiquitin-proteasome system (UPS), two proteolytic pathways that are therapeutic targets in the treatment of cancer. Herein, this extract was fractionated and the ellagitannins-enriched fraction (EEF) was selected to perform the succeeding experiments. To study the cytotoxic mechanism of EEF, an iTRAQ (isobaric tag for relative and absolute quantitation) based proteomics approach was used. Several proteins (296) were identified, among which 133 proteins were significantly modulated after cells treatment with EEF for 24 hours. These results were validated, by western blot, for the most modulated proteins, namely fatty acid synthase and insulin-like growth factor-binding protein 1 that are involved in cell death and metabolic pathways, respectively. Furthermore, and since the proteome analysis also showed that several proteasome subunits were downregulated, we hypothesized that a decrease in proteasome activity would lead to an accumulation of polyubiquitinated proteins. Accordingly, we found an increase of ubiquitin conjugates, suggesting that EEF decreases the UPS protein degradation. Overall, these results suggest that ellagitannins present in this bioactive fraction, obtained from Fragaria vesca leaves, could be a promising source of anti-cancer agents.

Acknowledgements: FCT and POFC/FEDER for financial support. Research supported by FCT PhD fellowship SFRH/BD/72918/2010, projects PTDC/SAU-FCF/105429/2008, PEst-OE/SAU/UI0177/2011, QOPNA research unit (project PEst-C/QUI/UI0062/2013; FCOMP-01 – 0124-FEDER-037296), and RNEM (REDE/1504/REM/2005 that concerns the Portuguese National Mass Spectrometry Network).

Keywords: Fragaria vesca; Ellagitannins; Ubiquitin-proteasome system; iTRAQ; Proteomics