The Research For Anti-tumor Activity Of Axitinib To Glioblastoma And Glioma Stem Cell

Based on the NIH grant program, we evaluated the new oral selective pan-VEGFR (VEGFR-1, VEGFR-2, VEGFR-3), PDGFR-alpha, PDGFR-beta, c-kit and tyrosine kinase inhibitor axitinib (AG-013736) against gliomablastoma and glioma caner stem cells.Here, we explored the anti-tumor activity of axitinib to glioblastoma U87 cell, glioblastoma cancer stem cell GBM4EF, GBM8EF, GBM18EF, BT74 derived from patients and prostate cancer PC-3 cancer cells (positive control) as well as anti-antigenosis to MS-1 cells with tube formation. Moreover, We developed a novel gene engeering virus oHSV47△-IL 12 which could infect and kill the gliomablastoma cell and glioma caner stem cells. Pointedly, Axitinib with oHSV47△-IL 12 combined would improve the effect for anti-tumor growth, primally have the synergy treatment whatever from morphous or amount.After the positive vitro data, we set up a series of xenograft and orthotopic models. Compared various drug administrations, we replaced the PEG400 with CMC as the solution of Axitinib, changed gavages into ip,25 mg/kg, qd.As a result, it has shown that Axitinib could decrease the volume of PC-3 tumor significantly and was generally safe toxity in PC-3 subcutaneous model. It also demonstrated the tumors at Axitinib group much smaller than mock group significantly and the time of ulcer happening also delayed for the malignant U87 subcutaneous model. Thus, we went on setting up the U87 intracranial model. For survival experiment, the median survival was 25 days at Axitinib group and 20 days at mock group. For safe experiment, the mean weights between two groups hasn’t any significant difference.For pathological experiment, H.E staining showed that all the mice had been implanted tumor successfully. The density of tumor cells was higher at mock group than Axitinib group. Meanwhile, the CD34 immunohistochemical staining demonstrated the endothelial cells in mock group were more than Axitinib group singinificantly. Axitinib downregulated VEGFR phosphorylation resulting in significantly decreased microvessel density and tumor vessels in all xenografts as measured. So far, these results indicated that Axitinib have anti-tumor growth effect to glioblastoma U87 cell, glioblastoma cancer stem cell GBM4EF, GBM8EF, GBM18EF, BT74 and prostate cancer PC-3 cancer cells in vitro. In vivo, Axitinib can markedly prolong the survival than mock group and it’s safe in toxity primarily. Immunohisto-chemical staining revealed that it inhibited the formation and growth of microvessel in tumor.PartⅠ:The killing effect of oHSV47△-IL 12 to glioblastoma and glioma cancer stem cellsObjective:To set up the glioma cancer stem cells and to create the novel oHSV47△-IL 12, then explore it’s killing effect to glioblastoma and glioma cancer stem cells.Methods:We abstracted, verified and cultured the glioma cancer stem cells from glioblastoma from patients and gene-engeneer IL12 by vector into the oHSV47△; then, we tried oHSV47△-IL 12 infected GBM stem cell experiments.Results:The cells can self-renew, differentiate, and the CD133、Nestin were also positve. oHSV47△-IL 12 was named after IL12 gene-engineering into oHSV47△.Conclusion:The glioma stem cell lines GBM4EF、GBM8EF、GBM18EF、BT74 have been cultured, which have stem potency; oHSV47△-IL 12 has been engineered and could infect and kill the glioblastoma and glioma stem cells GBM4EF、GBM8EF、GBM18EF、 BT74.PartⅡ:The anti-tumor activity of Axitinib to glioblastoma and glioma cancer stem cellsObjective:To explore whether Axitinib alone or combined with oHSV47A-IL 12 has antitumor activity to glioblastoma, prostate caner, especially for glioblastoma cancer stem cells.Methods:We used cell survival counters, MTS assay for cell survival; migration observation, tube formation test for endothelial cell proliferation. Moreover, We explored the synergy treatment of combination of Axitinib and oHSV47△-IL 12.Results:Axitinib reduced proliferation of tumor cells and cancer stem cells in a dose-dependent manner. These angiogenic agents were more effective in combination than when used alone in vitro. And Axitinib inhibited the tube forming of endothelial cells.Conclusion:Axitinib has anti-tumor activity and anti-angenosesis activity in vitro. Axitinib and oHSV47△-IL 12 have somewhat synergetic treatment.PartⅢ:The treatment of Axitinib to glioblastoma xenograft and orthotopic modelsObjective:Presently, we investigated the effect of the vascular endothelial growth factor receptor tyrosine kinase inhibitor axitinib (AG-013736) on xenograft and orthotopic models.Methods:Prostate cancer PC-3 xenografts were used to evaluate the effect of axitinib treatment on tumor vascular morphology, tumor volume, microvessel density, by measurement and examination. Then glioblastoma U87 xenograft was used for testing the anti-tumor growth activity in vivo. Following, intracranial model were set up to assess the survival and toxicity of Axitinib as well as anti-tumor effect for the malignant brain tumor.Results:Axitinib monotherapy induced prohibit growth stasis in PC-3 tumors xenograft model (positive control). A substantial decrease in tumor volume was observed (P<0.001). Moreover, Antitumor activity was significantly enhanced in both xenograft (P<0.001) and orthotopic models(P<0.01) of U87 tumors treated using an optimized schedule of administration (ip,25 mg/kg, qd), demonstrated promising anti-tumor activity with a favorable toxicity profile at the same time.Conclusion:PC-3 subcutaneous model showed the Axitinib could decrease the volume of PC-3 tumor significantly and was generally safe. For U87’s xenograft and orthotopic models, Axitinib can markedly prolong the survival than mock group and primarily it’s safe in toxity. Immunohistochemical staining revealed that it inhibit the formation and growth of microvessel in tumor.