Supercritical Extract Of Azadirachta Indica In Chemoprevention Of Colon Cancer

The leaves bark, and seed of Azadirachta Indica (Neem tree) have long been used in Ayurvedic medicine for health benefits. We have investigated a Supercritical CO2 Neem Extract (SCNE) in human colon cancer cell lines for chemopreventive potential. A SCNE of leaf was procured from Nisarga Ltd, Sartara, India and subjected to HPLC and LC-MS to develop of signature of triterpenoids with potential cancer preventive activity. One compound, nimbolide, suggested to account for anti-proliferative activity against cancer cells was found to be present in the extract. The effects of the SCNE were determined on HT29 and HCT116 human colon cancer cell lines. The proliferative activity of HT29 and HCT116 cells were studied after 48 and 72 hours treatment with SCNE and Nimbolide at various concentrations. We also evaluated the effects of SCNE and nimbolide on cell cycle parameters, and examined relevant proteins associated with cancer cell proliferation. Cell cycle analysis was performed by flow cytometry and confirmed a pre-apoptotic peak in SCNE treated HCT116 cells. The modulation of apoptotic markers (intrinsic pathway: Bax, Bcl-2, and caspase-3, 9), proliferation pathway (cyclin D1) were studied at the protein and mRNA level. Treatment with SCNE and Nimbolide in a range from 8- 18 µg/ml resulted in dose -dependent growth inhibition in 48 and 72 hours. At these concentrations SCNE caused changes in cyclin D1. The induction of apoptosis by SCNE in HT29 and HCT116 was shown by caspase activation, cleavage of PARP, increased levels of pro-apoptotic protein Bax, and reduced levels of the anti-apoptotic protein Bcl-2. The effect of SCNE on colon cancer cell migration will be reported by scratch wound healing assay. Overall, our data suggested that SCNE can suppress the growth of human colorectal cancer through induction of apoptosis. Current experiments are now focused on NF-kB and associated proteins as targets of SCNE inhibition. Supported by grant P30CAO54174. Citation Format: Roxanne Khoogar, Keya Mukhophadyay, Jay Morris, Girish Soman, Michael Wargovich. Supercritical extract of Azadirachta indica in chemoprevention of colon cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3210. doi:10.1158/1538-7445.AM2014-3210