Date Published: August 14, 2019
Publisher: Public Library of Science
Author(s): Limor Goren, George Zhang, Susmita Kaushik, Paul A. S. Breslin, Yi-Chieh Nancy Du, David A. Foster, Yi-Hsien Hsieh.
(-)-Oleocanthal (oleocanthal) is a phenolic compound found in varying concentrations in extra virgin olive oil oleocanthal has been shown to be active physiologically, benefiting several diseased states by conferring anti-inflammatory and neuroprotective benefits. Recently, we and other groups have demonstrated its specific and selective toxicity toward cancer cells; however, the mechanism leading to cancer cell death is still disputed. The current study demonstrates that oleocanthal, as well as naturally oleocanthal-rich extra virgin olive oils, induced damage to cancer cells’ lysosomes leading to cellular toxicity in vitro and in vivo. Lysosomal membrane permeabilization following oleocanthal treatment in various cell lines was assayed via three complementary methods. Additionally, we found oleocanthal treatment reduced tumor burden and extended lifespan of mice engineered to develop pancreatic neuroendocrine tumors. Finally, following-up on numerous correlative studies demonstrating consumption of olive oil reduces cancer incidence and morbidity, we observed that extra virgin olive oils naturally rich in oleocanthal sharply reduced cancer cell viability and induced lysosomal membrane permeabilization while oleocanthal-poor oils did not. Our results are especially encouraging since tumor cells often have larger and more numerous lysosomes, making them especially vulnerable to lysosomotropic agents such as oleocanthal.
Olive oil has been consumed by humans for millennia and is frequently associated with health-related properties. The Mediterranean diet and olive oil consumption in particular are correlated with lower cancer incidence and mortality [1–3]. A meta analyses of nineteen observational studies performed between 1990 and 2011 that included approximately 35,000 individuals found that olive oil intake is inversely related to cancer prevalence . More recently, a randomized trial found that women who adhered to a Mediterranean diet supplemented with extra virgin olive oil (EVOO) had 62% less invasive breast cancer incidence than a control group that was advised to restrict dietary fats . These studies, however, did not distinguish between the protective effects of EVOO’s triglycerides and its phenolic components. Furthermore, to the best of our knowledge, no controlled study tested the effect of high phenolic EVOO on cancer.
Although several groups have demonstrated oleocanthal’s ability to inhibit key proteins that promote cell growth and survival [12, 14–17, 39], a unifying mechanism for the specific and irreversible cellular death-inducing properties of oleocanthal has not been established. In this report, we observed that a transient exposure of cancer cells to oleocanthal for one hour resulted in the loss of cell viability after 24 hours. Although a classic apoptotic mechanism has been proposed [16, 20, 21], in our hands the rapid cell death caused by oleocanthal was necrotic. Specifically, viable cells were not observed to display phosphatidylserine on the outer membrane leaflet as evidenced by staining with AV, a well-established phase in the apoptotic cascade. Furthermore, using three different and complementary methods, we demonstrated that oleocanthal-treated cells undergo LMP. The latest, most robust method to assess LMP is the galectin translocation assay . We observed that oleocanthal treated MCF7 breast cancer cells showed robust galectin-3 translocation to lysosomes, similar to that observed with the established LMP inducer LLOMe. In a biochemical assay that checks the leakage of lysosomal enzymes into the cytosol, we observed a pronounced leakage of both cathepsin D and cathepsin B to the cytosol in PC3 prostate and MDA-MB-231 breast cancer cells. The translocation of cathepsins of two different sizes suggests that the lysosomal membrane undergoes permeabilization. Agents that are known to cause LMP with only minimal cathepsin release, such as LLMOe  enable cells to survive the initial LMP and repair their lysosomal membrane. Other agents that cause the release of cathepsin D (a small hydrolase) but not the release of cathepsin B (a larger hydrolase) are often associated with apoptosis . We, therefore, conclude that the degree of lysosomal damage in the case of oleocanthal is massive and leads to rapid necrosis in the affected cancer cells with less and survivable damage to normal cells.