| BackgroundAt present,The pathogenesis of tumor research mainly focus on tumor related gene cloning and functional analysis,cell signal transduction pathways and cell cycle regulation for the three fields.Signaling pathways are closely related on occurrence and development of cancer.Janus Kinase/Signal and activators of transcription(JAK/STAT)is one cell signal transduction pathways.JAK/STAT pathway activation affect cell proliferation,differentiation,apoptosis and advanced mitotic cycle,promote occurrence and development on the tumor.In addition, Study had confirmed JAK/STAT pathway activation can raise Bcl-2 protein expression, Enhance cell antiapoptotic ability.thus,to obstruct the access to therapy tumor.Currently,More research of AG490 is JAK/STAT signaling pathways blockers,AG490 by blocking its of signal transduction process,Can significantly inhibit the growth of tumor cells,Induce tumor cells apoptosis, And participate in the body's immune response, adjust inflammation, etc.It has become a research focus now.So far,Docetaxel is the first and only a proven to hormone independence prostate cancer have definite survival benefit of chemotherapy.Docetaxel is a kind of half synthesis of derivatives on paclitaxel,compared with paclitaxel has more strong microtubules protein affinity and longer intracellular retention time (is 2 to 12 times of paclitaxel).So the destructive power of tumor cells is more strong.It is the main mechanism antitumor by promote cell microtubules polymerization,Stop microtubules normal physiological depolymerization, the cell cycle is blocked in the G2/M period,Affecting cell mitosis, Causing cancer death, Producing antitumor effect. Nowadays discovery,it can inhibit Bcl-2 and Bcl-XL gene expression effect.Bcl-2 is related to occurrence,development and prognosis of prostate cancer closely,It has become a research hotspot in recent years.Currently,people do not know pathogenesis of prostate cancer. so scholars from home and abroad are researching pathogenesis of prostate cancer from cytology. Prostate cancer has three cell strains (LNCaP, DU- 145 and PC-3). LNCaP is androgen dependent prostate cancer. DU- 145 and PC-3 are prostate androgen independent prostate cancer. In fact, from confirmed diagnosis, most of prostate cancer patients have already had local progress and metastatic prostate cancer. There by they lose the chance of radical cure, and may only choose conservative treatment.The clinical prostate cancer is changing from the androgen dependent prostate cancer to the androgen-independent prostate cancer. The later is not sensitive to hormonal treatment, The treatment effect is not ideal. The above situation indicates,The focal point problem is to defer the mechanism of transformation from the androgen dependent prostate cancer to the androgen-independent prostate cancer and to treat the androgen independent prostate cancer.Objectives:To investigate the anti-proliferation effects and mechanism of decotaxel with AG490 on hormone-refractory Prostate cancer cell lines DU-145 in vitro,which may provide some clues to clinical application.Methods:1.We cultured the human prostate carcinoma cell line DU-145 cell ,To observe cell growth and photographed.2.This study was divided into two Parts.One was single agent experiment and the other was agents combination experiment.There were three groups for single ageniexperimeni:docetaxel,AG490 and negative control group. Cell morphology, flow cytometer,MTT method was used to observe the effect of all groups on Prostate cancer cell lines DU-145 invitro. The growth inhibition ratio and half inhibiting concentration(IC50) of AG490 were measured.3.The concentrations of decotaxel and AG490 was selected for combination experimental use,which inhibition rate on DU-145 cell was from 10%-40% insingle used.MTT method was used to investigate the growth inhibition.4.The cell cycle distribution were analyzed by flow cytometry,Treated with 20umol/L,40umol/L and 80umol/L AG490,40umol/L AG490 combined with 100nmol/L docetaxel for 48 hours.5.The expression of Bcl-2 protein was determined by immunocytochemistry in different groups.6.Using the statistical software SPSS16.0 make statistical analysis.Results:1. Observing cell morphology confirm prostate cancer cell is the truth from The Chinese Academy of Sciences cell bank.2. The DU-145 cells became round,small,cell shrinkage,membrane blebbing and so on,after exposed to AG490.3. In single agent experiment,with the inereasing concent ration and prolonging of treatment time,AG490 and docetaxel can significantly inhibited the proliferation of DU-145 cells in a dose and time-dependent manner.MTT colorimetric method was performed to evaluate the potential cytostatic effect of combining AG490 with docetaxol.It showed that low concentration of docetaxel had the enhancing synergistic effect combined with AG490 (P<0.05).4. The DU-145 cells were blocked at G1-Phase,and the S-Phase fraction (SPF) of the cells were decreased by AG490 treatment.The cells were blocked at G2/M-Phase by docetaxel treatment.When treated with both of them,G1-Phase and S-Phase cell fraction were between the values of single agent. 5. Immunocytochemistry experimental result display,In contrast to control group,the Bcl-2 protein expression level obvious down-regulated in AG490 group,AG490 could significantly decrease the level of Bcl-2 Proteins in DU-145 cells when combined with docetaxel (P<0.05).Conclusions:1. Janus Kinase inhibitor AG490 inhibited the growth of the prostate carcinoma cells by concentration and time dependent,the mechanism of which relates to its cell cycle arrest and apoptosis induction on prostate carcinoma cells.2. Janus Kinase inhibitor AG490 down-regulates the expression Bcl-2 protein via inhibiting the JAK-STAT3 signaling pathway,further inhibits the growth of prostate cancer cells and promotes its apoptosis.3.We confirmed that AG490 combined with low concentration docetaxel had a synergistic effect on inhibiting the proliferation of DU-145 cells.Their effects of inhibiting the proliferation of DU-145 cells seem to due to down-regulation of the expression of Bcl-2 protein collectively.The research first time showed that AG490 combined with docetaxel had a synergistic effect on inhibiting the proliferation of DU-145 cells,Which provided a new theoretical basement for reasonable clinical treatment of hormone independent prostate cancer. |
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