Effect Of Novel Aza-chalcone Derivatives On The Growth And Sensitization Of Cervical Cells

Background and Objective Cervical cancer is one of leading causes of mortality in women currently. Persistent infection with high risk Human Papillomaviruses(HPVs) is the main factor.Although the incidence and mortality have been gradually decreasing with earlier detection and advanced medical treatment,the prognosis of patients with cervical cancer remains dismal in the presence of advanced metastatic disease.Due to the minimal success of cytotoxic therapies for cervical cancer.At present,cisplatin combined with paclitaxel is the main treatment for advanced cervical cancer,which can extend survival time for 3 months.Once this combined treatment fails,there are no second- or third-line drugs available.When themotherapeutic drugs with different effects have been combined,drug fast and intolerable side effects still can appear sometimes. Therefore, it is necessary to develop new bioactive compounds which can improve the effect of chemotherapy on cancer cells. Flavonoids compounds is kind of polyphenols secondary metabolites existing in nature.Most of nature plant contains abundant flavonoids with various complex structures.It can be divided into several types including flavone,flavonol, flavanone,flavanone alcohol, alcohol, isoflavones, isoflavones alcohol,aurone, flavanoid and flavanol and so on,with mutiple pharmacological activity such as anti-oxidative, anti-aging,anti-virus, anti-cardiovascular disease, anti-cancer,analgesia,regulating tumor drug resistance and improving the immune regulating funtion.Flavonoids can inhibit many malignant tumor growth obviously in vivo and vitro,which treament mechanism is becoming more and more attention.Natural flavonoids were varied,exists in the form of combination,which lead the extraction process is relatively complex.In recent years,there have been new synthsis of flavonoids. In our study,we choose novel aza-chalcone derivatives alone or combined with nedaplatin to observe the effect of inhibiting the growth of cervical cancer Hela and Siha,and the sensitized effect of nedaplatin in vitro.Our study comfired that the novel synthesis of flavonoids has better activity against cervical cancer cells and sensitized effect.Materials and Methods We synthesized seven novel aza-chalcone derivatives. These compounds were structural confirmed and their activitiy was measured. MTT assay was used to detect the effect anti-proliferative of seven compounds on Hela cells.According to the value of IC50,we choose one compound with the lowest IC50 for the next step of the experiment,and observed their inhibitory effect on Hela,Siha and Ha Ca T cells,and the chemosensitive effect when combined with nedaplatin.By scratch test,we observed the migration effect.The cell morphological changes were observed and photographed under inverted microscope after treatment for 24 hours with different concentration of the compound.Flow cytometry was performed to test the cycles and apoptosis of the cells. Westernblot to detect the impact of the compound on the expression level of p53 in cervical cancer cells.Results 1.The inhibition effect of novel compounds on Hela cell: Seven compounds have different degree inhibition effect, and was dose dependent(the range of IC50 values from 14.133?M to 173.236?M).The strengthen of inhibition is number 10 g, 10 f, 10 b, 10 a, 10 d,10e,10 c,in that order. 2.The inhibition effect of No. 10 g on Hela,Siha and Ha Ca T cells:No.10 g dose-dependently inhibited the growth of He La and Siha cells.The effect on Hela(IC50=14.133?M) is more obvious than on Siha(IC50=18.281?M).However,mild cytotoxicity was observed on Ha Ca T(IC50=243.751?M) at the high concentration ofthe compound, p<0.05. 3.The chemosensitive effect of No.10 g when combined with nedaplatin: Hela and Siha cells were treated with combined nedaplatin of different concentration and 2?M of No.10 g compound for 48 hours. The combined inhibitory effect at low concentration is more obvious than nedaplatin alone.Moreover,the chemosensitivity effect on Hela(IC50 NDP:2.878ng/L;NDP+10g:0.937 ng/L) is better than on Siha(IC50 NDP:11.204 ng/L; NDP+10g:5.927 ng/L), p<0.05. 4.The cell migration effect of No.10g:After treatment with No.10 g compound of 15?M for 24 hours, the inhibitory effect of migration on Hela >Siha> Ha Ca T. 5.The cell morphology change of No.10g:The Hela and Siha cell morphology under inverted phase contrast microscope became floating,death with the increasing concentration of No.410 g compound after treatment for 24 hours.However, no obvious change in Ha Ca T cells. 6.The change of cell cycle and apoptosis of No.10g:After 24 hours treatment of 15?M No.10 g,flow cytometry found that Hela and Siha cells were arrested significantly into G0/G1 phase,and the opoptosis rates on He La,Siha and Ha Ca T((12.483±0.49)%?(1.819±0.78)%?(0.785±0.32)%)were higher than in control groups((0.882±0.81)%?(0.820±0.21)%?(0.084±0.41)%),all p<0.05. 7.Influence on the expression of P53 of No.10g: Westernblot showed that the relative expression level of p53(0.71±0.095?1.02±1.134)were higher after treament with No.10 g than the control groups(0.53±0.071?0.69±0.109), all p<0.01.Conclusions 1.All of the seven novel aza-chalcone derivatives can inhibit the growth of He La cells. 2.The inhibitory effect of No.10 g compound on Hela cells with HPV 18 positive is higher than on Siha cells with HPV 16 positive,however, the cytotoxic effect on Ha Ca T cells with HPV negative is mild. 3.No.10 g compound sensitized cervical cancer cells on nedaplatin at low concentration. 4. Aza-chalcone derivatives improve the apotosis and inhibit the growth of cervical cancer cells by up-regulating the expression level of wild-type p53.