Identifying Mitochondrial Targets for Pancreatic Cancer Employing Unconventional Combinations of Middle Eastern Phytoextracts with Raloxifene

Abstract

PCa has a high mortality, substantial chemoresistance, and a poor 5‐year survival rate of 6 to 8%. It is the fourth leading cause of deaths due to cancer in the United States, killing over 53,000 Americans each year, and 250,000 people worldwide. Little is known about the regulation of mitophagy in PCa by drugs and dietary components. There is also a gap in knowledge concerning molecular interactions between drugs and food and the K‐ras pathway in PCa. This is important because mutations in the human homologue of the Kirsten Rat Sarcoma (KRAS) viral oncogene occur in 90% of PCa patients and drive the early development of PCa. Our previous work showed that enhanced mitophagy, which prevents the accumulation of damaged mitochondria in PCa cells was alleviated by repurposing the FDA‐approved selective estrogen receptor modulator raloxifene to treat PCa cells. Here, we used a combination of Middle Eastern dietary components (MEDC) and raloxifene to mitigate mitochondrial renewal and identify novel mitochondrial targets for PCa. We screened crude aqueous extracts from six spices and plants used in Middle Eastern and Indian cuisines, including artichoke (Cynara scolymus), curry leaf (Murraya koenigii), dill (Anethum graveolens), noni (Morinda citrifolia), olive (Olea europaea), rose (Rosa damascena), and sumac (Rhus coriaria) for cytotoxicity in the PCa cell lines, MIA PaCa‐2 (similar KRAS gene mutation as majority of patients) and PANC‐1 (less common KRAS gene mutation). Specifically, we treated PCa cells in vitro with exponentially increasing doses of aqueous extracts at 300 ng/mL, 3 μg/mL, 30 μg/mL, and 300 μg/mL for 48 hours followed by an MTT assays. Aqueous extracts of sumac and artichoke (AESA) induced maximum cytotoxicity, which was exacerbated by raloxifene (AESA‐R). To address whether cytotoxicity was limited to PCa cells, we used NIH‐3T3 cells as a non‐cancer control, showing a sparing effect. We also tested 1:1 combinations of AESA and found evidence of activity. To identify specific compounds, using a fractional approach based on preparative HPLC, 100 mg of the residue was chromatographed onto a silica gel column and gradient eluted using dichloromethane/methanol (gradient, 0, 100%, v/v). Fractions and eluates were monitored by thin layer chromatography using precoated silica gel 60 F254, 0.25 mm aluminum backed plates. Cytotoxic fractions were reconstituted in deuterated solvents and analyzed by NMR spectroscopy to identify pentagalloyl glucose as the bioactive component in sumac. To elucidate the mechanism for the cytotoxicity, we showed that AESA and AESA‐R activated caspase‐3 and 7, induced mitochondrial ROS, and reduced mitochondrial membrane potential in PCa cells, starting within 20 minutes and sustained for 48 hours following treatment. Western blot showed changes in the expression of proapoptotic (bim) and the translocation of antiapoptotic proteins (Bcl2), sustained over 48 hours. Our data demonstrate for the first time that specific food‐drug combinations alter mitochondrial biology and response including the regulation of mitophagy in PCa cells.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.