Pancreatic cancer is the most common cancer among men and women; the fourth leading cause of cancer death in the United States and the fifth leading cause of cancer death worldwide. This disease has a poor prognosis with a 5-year overall survival rate of less than 20%. Multiple mechanisms have been postulated for the development of benign and malignant pancreatic diseases. However, the nature and origin of the precursor cells for pancreatic cancer have not yet been delineated. Based on several molecular mechanism(s) proposed for pancreatic cancer, the phosphoinositide 3-kinase (PI3K)/Akt pathway is found to be constitutively activated and the mammalian target of rapamycin (mTOR) kinase is reported to be an important mediator for its signaling. The phosphoinositide 3-kinase-AKT-mammalian target of rapamycin (PI3K-AKT-mTOR) pathway is a frequently hyper activated pathway in cancer and is important for tumor cell growth and survival. The development of targeted therapies against mTOR pathway led to the approval of drugs including everolimus and temsirolimus, for the treatment pancreatic and other cancer types. However, the effectiveness and response among high risk patients still remains unclear. Epidemiologic and laboratory studies suggest that plant-derived bioactive food components reverse or prevent the development and progression of early-stage disease before it becomes aggressive and malignant. Lately, naturally occurring non-toxic dietary compounds are increasingly used as a novel strategy for the prevention of more aggressive cancers. Previous reports from our laboratory suggest that prolonged exposure of cancer cells to natural agents may effectively modulate mTOR signaling and promote anti-proliferative effects. In the present study, we evaluated the effectiveness of celastrol in human (AsPC-1) and mouse (Pan-02) pancreatic cancer cells. Celastrol is a plant extract isolated from the root extract of Tripterygium Wilfordi (Thunder of God vine -TGV) and Celastrus Regelii, is also known as tripterine, a multiple bioactive natural product. Recent studies suggest that celastrol may exhibit antioxidant, anti-inflammatory, anticancer, and insecticidal activities. However, a significant lack of information on the bioavailability and the effect on specific molecular targets of celastrol is unknown and is therefore limiting its use for future clinical trials. our preliminary findings on the oral administration of celastrol (10mg/KgBW) and TGV separately to C57BL/6 black mice showed no toxicity or animal death (n=24). Subsequent analysis of serum samples showed bioavailability of celastrol with a retention time between 12 to 15 minutes. Human pancreatic cancer cells AsPC-1, and mouse pancreatic cancer cell Pan-02, treated with celastrol showed a significant decrease in cell proliferation (>4 fold) after 24h with a dose of 20ug/ml (confirmed by MTS assay). In order to determine whether the cell growth inhibition is associated with an effect on cell cycle, we conducted flow cytometry analysis for DNA content. Pan-02 cells treated with celastrol at an optimum dose of 10ug/ml showed cell cycle arrest at the G1 phase. Further analysis of the flow cytometry data showed a pre-G1 peak of apoptosis (>10%). Based on the findings on celastrol inducing cell cycle arrest and apoptosis, we extended our studies to determine the effect on the downstream targets of PI3K-AKT-mTOR pathways. RT-PCR analysis for AKT using specific primers with RNA isolated from pancreatic cancer cells showed a decrease in the mRNA (>2 fold). Further studies are in progress to confirm a specific dose dependent effect celastrol and TGV on the downstream targets of the PI3K-AKT-mTOR and cell cycle regulatory proteins including cyclin D1 in pancreatic cancer cells. The future goal will be to test the efficacy of celastrol and TGV in preclinical animal models of pancreatic cancer. Overall, our findings suggest the potential molecular targets of celastrol in pancreatic cancer cells.

Citation Format: Di Tian, Bhagavathi Narayanan, Amitabha Mazumder, George Miller, Narayanan K. Narayanan. Tripterine induced cell cycle arrest and apoptosis is associated with modulation of PI3K-AKT-mTOR pathway in human pancreatic cancer cell. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr B27.