The Efforts Research Of Combined GM-CSF And MTDH Gene Vaccine To Inhibit The Prostate Cancer’s Growth And Metastasis

Background:Tumor metastasis and tumor resistance to chemotherapeutic drugs is one of theleading cause of death for cancer patients. The advanced prostate cancer often occursorgan metastasis, such as lymph nodes, bone and lung. And it is prone resistant tochemotherapy and androgen and becomes the hormone-refractory prostate cancer(Hormone-refractory prostate cancer, HRPC). In recent years, the study found thatMTDH had the function of oncogene, which regulates the cell’s transformation,proliferation, invasion and metastasis, promotes angiogenesis and reduces thesensitivity of tumor cells to chemotherapeutic drugs; the express of the MTDH inprostate cancer and cultured prostate cancer cells significantly increased. In this paper,basing on the preliminary study, we have further research of the MTDH gene vaccineto inhibit the growth and metastasis of prostate cancer.Objective:To evaluate the effect of intramuscular MTDH gene vaccine in mice withprostate cancer RM-1xenograft tumor, and to preliminary study the mechanism of theGM-CSF, which is one of immune adjuvant, effecting on the MTDH gene vaccine.Methods:Preventive model group: We used the method of quadriceps injection to injectthe plasmid of MTDH into the immune male C57BL/6mice,2times a week, a totalof three times, and when1day,3days and5days after each gene immunization, wesubcutaneously injected GM-CSF100μL (1μg/100μL).7days after the last geneimmunization, we caught five mice from each group, then test their spleen CD4+andCD8+T cell subsets by the flow cytometry and detect cytotoxic T lymphocytes (CTL)response by using CytoTox96Non-Radioactive Cytotoxicity Assay Kit. Bloodsamples from their angular veins were taken to assay the serum IFN-γ and IL-4level by ELISA.2weeks after the last gene immunization, the remaining mice weresubcutaneously inoculated with RM-1cells(105/100μL).1week after the inoculationof tumor cells, we observed the growth sitution of tumors, measured the size of thetumors via the vernier caliper estimated the survival time of these tumor-bearing miceand drew the tumor’s growth curve chart.21days after the inoculation of tumor cells,we executed all of the mice, weighed the weight of these tumors,calculated theirinhibition rate, and then evaluated the anti-tumor effect of the DNA vaccine in mice.We used the TUNEL labeling method (Terminal–deoxynucle-oitidyl TransferaseMediated Nick End Labeling) to detect the apoptosis of tumor cell,used the routineHE staining to observe the tumor tissue structure.Results:MTDH gene highly expressed in the RM-1cells, but low in the major organs andtissues of the mice. Comparing with the control group of Pub and PBS group, MTDHDNA vaccine could significantly stimulate the CD8+T cells（p＜0.01）, but the CD4+T cells’ difference was not obvious（p＞0.05）; Comparing with the alone MTDHvaccine-preventable group, the prevention group with GM-CSF adjuvant’s CD4+Tand CD8+T cells increased significantly different(p＜0.01).Anti-experimental results showed that the kill rate of RM-1target cell wasdifferent under different effector-target ratio. When the effector-target ratio was100:1,the kill rate of RM-1target cell was highest, which come to12.99%.Comparing to thecontrol group, the killing rate of the prevention group and adjuvanted group hadsignificant difference（p＜0.05）; The prevention model group’s tumor growth curveshowed that comparing with the control group, tumors growth of MTDH preventiongroup was significantly inhibited (p<0.05).GM-CSF adjuvant had the effect ofenhancing the MTDH’s antitumor (p <0.05). Analysis of the survival period, thesurvival period of the MTDH prevention group and the combined GM-CSF group wasno significant difference (p>0.05),but them were significantly longer than the controlgroup. We could detect that the mice serum cell factor IFN-γ in the MTDHprevention group and joint GM-CSF group were significantly increased. Found noevidence of autoimmune inflammation and tissue damage in the main organs and tissue in mice after vaccination. The Tunel showed the tumor cell apoptosis ofadjuvant GM-CSF group and MTDH prevent group were significantly higher than theother two control groups, and the adjuvant GM-CSF could significantly promoteMTDH vaccine against RM-1tumor cell apoptosis.Conclusions:Using the MTDH gene vaccine to immune the male C57BL/6mice, couldgenerate protective cellular immune responses and humoral immune response in mice,and then inhibits the growth of transplanted tumor. And the joint with GM-CSFadjuvant has stronger antineoplastic effect, but combining with GM-CSF has nosignificant effect on the survival time of tumor-bearing mice.