Experimental Studies On Therapeutic Effect And Mechanism Of Brucea Javanica Emulsion Associated With Mitoxantrone For Hormone Refractory Prostate Cancer(HRPC)

It is recognized by urologists in the world that the treatment of hormone refractory prostate cancer (HRPC) is very difficult. So far, the research in-depth on the therapeutic effect and mechanism of Brucea javanica emulsion associated with Mitoxantrone for HRPC have not been reported, and we will try to explore it and find the experimental evidence on clinical use of Brucea javanica emulsion associated with Mitoxantrone for patients with HRPC.Part OneExperimental study on the effect of Brucea javanica emulsion associated with mitoxantrone for PC3 cell: an in vitro investigationObjectiveTo explore the therapeutic effect and mechanism of Brucea javanicaemulsion associated with mitoxantrone for HRPC by investigation in vitro.MethodsAccording to the use of drug, the experiment groups were randomly divided into four groups: Control group(no drug), A group (Brucea javanica emulsion only), B group (Mitoxantrone only), and C group (Brucea javanica emulsion plus Mitoxantrone). To analyze the proliferation, apoptosis and gene expression of PC3 cell among four groups by the technique of MTT, RT-PCR, Western-Blot and FCM.ResultsIt was showed by MTT examination that the suppression effect of Brucea javanica emulsion and Mitoxantrone on PC3 cell proliferation were all dependent on drug concentration and time. After being exposed to Brucea javanica emulsion(500 μ g/ml) plus low dosage Mitoxantrone(2.5 μ, g/ml) for 24, 48h, 72h, the suppression rates on PC3 cells were respectively 45.6%, 69.9%, 87.3%, being significantly superior to that in A group and B group, P<0.05.After being exposed to Brucea javanica emulsion (500 μ g/ml, A group), low dosage Mitoxantrone(2.5 μ g/ml, B group) ,and Brucea javanica emulsion(500 μ g/ml) plus low dosage Mitoxantrone(2.5 μ g/ml, C group) for 48h, (1) it was showed by RT-PCR that the mRNA ofBcl-2, Bax, MMP9, MRP and PSA were all expressed in PC3 cells. In contrast to Control group, A group and B group, The mRNA expression of Bcl-2, MMP9, MRP and PSA in C group were all significantly lower than that in Control group, A group and B group(/)<0.05), while the mRNA expression of Bax was significantly superior to that in Control group, A group and B group(P<0.05);(2) it was showed by Western Blot that the protein expression of Bcl-2 and Bax in C group were respectively lower and higher than that in Control group, A group and B group, with a significant difference(/)<0.05);(3)it was showed by FCM that the cell ratio of G0/G1 stage was 70.6% in C group, being significantly higher than that in Control group, A group and B group(P<0.05), whereas the the cell ratio of S stage and G2/M stage in C group were respectively 13.6%, 15.8%, being lower.Conclusion1. The inhibitory effect on PC3 cell growth in C group (Brucea javanica emulsion plus low dosage Mitoxantrone) is stronger than that in A group (Brucea javanica emulsion only) and B group (Mitoxantrone only).2. Brucea javanica emulsion associated with low dosage Mitoxantrone can induce cell apoptosis of HRPC by obviously down-regulating the expression of Bcl-2 and up-regulating theexpression of Bax in PC3 cells.3. Brucea javanica emulsion associated with low dosage Mitoxantrone might debase cell aggressiveness and multidrug resistance of HRPC by obviously down-regulating the expression of MMP-9 and MRP in PC3 cells.4. Brucea javanica emulsion associated with low dosage Mitoxantrone can suppress cell DNA synthesis and block cell proliferation of HRPC by obviously elevating the ratio of GO/Gl stage and debasing the ratio of S stage in PC3 cells.Part twoInhibitory effect of Brucea javanica emulsion associated withMitoxantrone on HRPC xenotransplanted tumor model ofnude mice : an in vivo investigationObjectiveTo analyze the inhibitory effect of Brucea javanica emulsion associated with Mitoxantrone on Xenotransplanted tumor growth of HRPC by an investigation in vivo, and to provide the scientific evidence for further clinical treatmentMethodsXenotransplanted tumor models of HRPC were established by injecting PC3 cell in the dorsal skin of 24 male nude mice, and then begin drug experiment for 3 weeks. 24 mice were randomly divided into four groups: Control group(no drug), A group (Brucea javanica emulsion, 200mg/kg/d), B group (Mitoxantrone, lmg/kg/d), and C group (Brucea javanica emulsion 200 mg/kg/d plus Mitoxantrone lmg/kg/d), 6 mice for every group. To observe and analyze the tumor weight, tumor size, and tissue pathology of tumor, liver, kidney on mice among four groups.ResultsAfter executing nude mice in the end of experiment, we found following results:1. The inhibition rate on tumor growth in C group was 76.2%, significantly higher than that in A group and B group(P<0.05), and the average tumor weight in Control group, A group, B group, C group was respectively 436. lmg, 184.3mg, 197.2mg, 103.9mg.2. The tumor size in C group was significantly smaller than that in Control group, A group and B group(P<0.05), and the average tumor size in Control group, A group, B group, C group was respectively 359.2 mm3,158.6 mm3,175.7 mm3,93.8mm3.3. By microscopy observing, the obviously abnormal pathologysigns of liver and kidney were not found in Control group, A group, B group, C group, and the pathology signs of tumor necrocytosis in C group was significantly obvious.Conclusion1. Brucea javanica emulsion associated with low dosage Mitoxantrone have good anti-tumor effect on Xenotransplanted tumor models of HRPC.2. Brucea javanica emulsion associated with low dosage Mitoxantrone can bring little toxic damage on liver and kidney of nude mice, the associated treatment for HRPC will be safe and feasible in a certain extent.