Isolation, Structure Elucidation, And Mechanistic Studies Of Natural Compounds Against Gastric Cancer From Traditional Chinese Medicinal Herbs

Throughout the history of civilization, the humans have relied on natural products as aprimary source of medicine. Natural products have traditionally provided a rich source ofdrugs for many diseases including cancer and plants are an important source of novel naturalproducts. Only a fraction from biosphere diversity has been tested for biological activity andnovel cancer therapeutics still remains to be discovered. Cancer is a leading cause of humandeath worldwide ensuing in a paramount body of research to curtail it. However gastric canceris found to be the second most common cause of mortality over the globe. Prevailingtreatment options had limited therapeutic success in gastric cancer, particularly at an advancedstage; owing to its undiagonistic behaviour at an early stage so it’s becomes resistant totherapy. Hence, the current therapy for gastric cancer is not satisfactory. New and bettertherapeutic options are immediately required to develop a more effective therapy for gastriccancer. Secondly, chemotherapy, cancer treatment, has a major limitation that in spite of itsefficacy usage of different chemical agents produces some untoward side effects at highereffective doses. The present study was designed to find the novel natural anticancer agents fortreatment of gastric cancer through screening of Chinese herb extracts and TCM (traditionalChinese medicine) compounds library and to investigate their possible molecular mechanism.Moreover, we focused to test the hypothesis that combined use of low doses ofchemotherapeutic agents with different mode of actions, rather than the administration of asingle agent at a higher dose, not only to obtain increased efficacy and minimize side effectsbut also this approach may prove helpful in reducing drug resistance and may prolong thesurvival after tumor challenge. For these purposes, the crude ethanolic extracts of300speciesof herbal plants, traditionally used in China for the treatment of a variety of diseases, and fourhundred TCM compounds were screened to evaluate the cytotoxic activity on human gastricadenocarcinoma SGC-7901cells and splenocytes (normal cells). Extracts, which exhibitedcytotoxicity at100μM, were considered active.33of these raw ethanolic extractsdemonstrated growth inhibitory activity on SGC-7901cells. Interestingly, of the33activeextracts on cancer cells, nine showed less toxicity against the normal spleen cells. Of the400natural compounds,18significantly inhibited proliferation of SGC-7901cells. These resultsindicate the potential use of traditional Chinese medicinal herbs as antineoplastic agents andsuggest that further studies evaluating their mechanism(s) of action and the isolation of activeanti-tumor compounds are warranted. We have successfully isolated three compounds withanti-proliferative effect on SGC-7901cells. The chemical structures of these compounds were determined as Costunolide, Xanthoxyletin, Kuraridin and Nor-kurarinone by using MS,1H-NMR and13C-NMR. Understanding the interplay of different cancer-related signalingpathways is important for the development of efficacious multi-targeted anticancer drugs.This dissertation reports mechanisms of these TCM compounds or their derivatives(Costunolide, Xanthoxyletin, Kuraridin and Nor-kurarinone, Evodiamine, Magnolol,Tubeimoside-1, Isoalantolactone, and Dracorhodin perchlorate) that induced cell death inhuman gastric adenocarcinoma SGC-7901cells. DNA staining assays (Hoechst, PI), andannexin V-FITC flow cytometric analysis showed that these compounds induced apoptosis inSGC-7901cells. To investigate the mechanism underlying these compounds activities, weused western blotting to identify proteins whose expressions are altered in tumor cells. Thesecompounds decreased the expression of anti-apoptotic mitochondrial protein Bcl-2andincreased the expression of pro-apoptotic protein, Bax. These compounds also induced thedissipation of mitochondrial membrane potential (ΔΨm), activation and cleavage of caspase-3and its substrates. Furthermore, it was found that these compounds arrested the cell cycle atdifferent phases; Magnolol, Xanthoxyletin at S-phase, mixture of Kuraridin and Nor-kurarinone, Costunolide, Evodiamine, Tubeimoside-1, and Isoalantolactone at G2/M phaseand Dracorhodin perchlorate arrested the cell cycle at G1/S phase. We also found thatMagnolol, Isoalantolactone, and Dracorhodin perchlorate treatment results in selectivedownregulation of a specific catalytic subunit of the phosphoinositide3-kinase (PI3K) family,p85. Further exploration of this observation demonstrated that the mechanism of action ofthese compounds involves the inhibition of the pro-survival Akt pathway that lies downstreamof PI3K. In addition, Dracorhodin perchlorate modulated the p53and NF-κB pathways aswell. These results reveal the functional interplay among the PI3K-Akt, p53, and NF-κBpathways that are frequently deregulated in cancer and suggest that their simultaneoustargeting by Dracorhodin perchlorate could result in efficacious and selective killing oftransformed cells. Together with all the above-mentioned results indicate that all thesecompounds are promising gastric cancer therapeutic agents. Traditional medicines not onlyprovide an important notion for finding novel drugs, also may supportive to reallocate drugdiscovery paradigm from ‘finding new-entity drugs’ to ‘combining existing agents and mighteven the combinations between such agents. As mentioned above, Evodiamine significantlyinduces apoptosis in gastric cancer cells. Tumor necrosis factor-related apoptosis-inducingligand (TRAIL) selectively induces apoptosis and kills cancer cells with little or no adverseeffects on normal cells. However, gastric cancer cells are less sensitive than other cancer cellsto TRAIL-induced apoptosis. Effects of Evodiamine on TNF-α-and TRAIL-inducedapoptosis in human gastric cancer cells were remained unexplored. In the present study, we found that Evodiamine enhanced TNF-α-and TRAIL-induced apoptosis in human gastriccancer cells. Here, we demonstrate the first evidence that this compound effectively enhancesTNF-α-and TRAIL-mediated cytotoxicity by suppressing pro-survival proteins includingXIAP and survivin. Together with all the above-mentioned results indicate that all these TCMcompounds are promising therapeutic agents for treatment of gastric cancer. These anticanceragents underline the potential utility as a new cancer treatment modality and are thought to berepresentative candidates for in vivo studies of monotherapies or combined anti-tumortherapies.