Effects And Mechanisms Of Active Compounds With Acanthopanax Trifoliatus In Combination On Cancer Cells

Acanthopanax trifoliatus(L.)Merr.,a shrub belonging to the Araliaceae family.It has been utilized as a folk vegetable and medicinal plant to take advantage of its anti-oxidant,anti-inflammatory,and anti-cancer effects.Our research group previously found that impressic acid(E12-1)and acankoreanogein(E13-1)isolated from A.trifoliatus(L.)Merr.exhibited potent cytotoxic effects against cancer cells.The plant also contained chlorogenic acid(CA)and ursolic acid(UA).Cancer is a malignant disease threating human health seriously and it is a common aspiration of mankind to conquer cancer.Currently,the mainly employed treatment of cancer is chemotherapy which often causes serious side-effect and poor tolerance.As a result,combination therapy emerges as an alternative for chemotherapy to fight against cancer.The combination of first-line medicine and different drugs can not only reduce the side effects and toxicity,but also increase the synergy effects.In order to isolated the compound E12-1 and E13-1 effectively,the separation condition is:speed 60mL·min-1,time 2 h,EtOAc/petroleum ether 15%,18%and 20%.The column chromatography over silica gel eluted with petroleum ether-EtOAc,80:1,to get E12-1 and petroleum ether-EtOAc,60:1,to get E13-1.In this study,we investigated the potential synergistic effects of compounds from A.trifoliatus(L.)Merr.in combination with docetaxel(DOC)or gemcitabine(GEM)for the VCap prostate cancer or PANC-1 pancreatic cancer cells and the underlying mechanisms of their action.The proliferation,viability,and apoptosis of cancer cells were evaluated by the MTT assay,trypan blue exclusion assay,and morphological assessments to investigate the effects and action mechanisms in cancer cells.In the research,cell migration was examined under a light microscope.Besides,NF-κB activity was assessed by the luciferase reporter assay and Western blot was used to determine the levels of protein.The result showed that:1)The combination of E12-1,E13-1,CA,UA and DOC for the VCaP cells were evaluated by MTT assays,the IC50 of E12-1 E13-1 and DOC were 80.16μM,61.17μM,20.67μM,12.5μM and 15.71nM,respectively.Treatment with combined drugs decreased cell viability by 27-36%compared to the single drug.The treatment with the combination also strongly promoted the apoptosis of VCaP cells compared with treatment with the compounds.The percent of apoptotic cells increased to 36.58%,38.64%,40.58%and 38.78%,respectively.Especially,the combination of E12-1 and DOC showed best apotosis effect.Besides,closure of the wound area with the combination significantly decelerated.Inhibition of cell migration by combination treatment with E13-1 and DOC was decreased by 26.48%,which showed that the best decrease in relative wound closure was significantly related to cell proliferation and migration.2)The combination of E12-1,E13-1,CA,UA and GEM for the PANC-1 cells were evaluated by MTT assays,the IC50 of E12-1 E13-1 and GEM were 66.13μM,98.38μM,21.52μM,13.67 p.M and 2.78 μM,respectively.The inhibition of cell viability was increased by the treatment with combination.Treatment with combination was decreased cell viability by 25-40%.Treatment with combination caused more visible morphological changes compared with single drug.The combination of E12-1 and DOC showed best inhibited relative wound closure to 26.48%.3)The combination of E12-1,E13-1,CA and UA interact with each other in VCap cells.The rate of cell viability were 50,35%,65,28%,63,67%,61,84%and 57.93%。The combination of E12-1 and E13-1 showed best inhibited cell viability activity.The percentage of apoptosis with combination were 42.89%,30.04%,33.18%,32.82%and 35.97%,respectively.Compared to the single drug,treatment with combination caused more visible morphological changes.Treatment with combination with E12-1 and E13-1,the NF-κB activity was decreased by about 20%,showed the best inhibited NF-κB activity.4)The combination of E12-1,E13-1,CA,UA and each other for the PANC-1 cells.Treatment with combination with E12-1 and E13-1 showed best apotosis effect compared with single drug increased by 25%and compared with combination for other compounds increased by 7-10%.The combination of E12-1 and E13-1 showed best inhibited relative wound closure to 32-35%.Western blots results showed that the combinations of compounds had stronger stimulatory effects on p-JNK expression and caused decreases on Survivin,P-Erk1/2,Bcl-2,Akt,NF-κB and P-Stat 3 levels in VCap cells and PANC-1 cells.The present study demonstrated that the antitumor activity for the combination of E12-1,E13-1,CA and UA for the first time in VCap prostate cancer or PANC-1 pancreatic cancer cells.The conclusion showed that the combination group had more potent inhibitory effect on the growth and a more potent effect on the apotosis induction of cancer cells than ether agent alone,showing a synergistic effect.It will provide a scientific basic for the later research about compound from A.trifoliatus(L.)Merr.and first line drugs.