A basic trial of gene therapy against peritoneal dissemination of ovarian cancer by calponin h1 gene transfer

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Calponin h1(CNh1), one of the actin-binding proteins, stabilizes the filaments of alpha-smooth muscle actin(α-SMA) and modulates cellular biological phenotypes. We have reported that the ovarian cancer cells probably make favorable environment for fheir intravasation through fhe down-regulation of CNh1 and α-SMA in the cells forming blood vessel walls. In this study, we investigated (1)the ovarian cancer-derived effects on peritoneal mesothelial cells and fibroblasfs, (2)the changes occurredin CNh1 gene-transfected cells, and (3)the efficacy of CNh1 gene therapy against the peritoneal dissemination of ovarian cancer.By adding the conditioned culture medium of human ovarian cancer cells, the reduction of both CNh1 andα-SMA expressions occurred in peritoneal cells and fibroblasts resulting in cellular retraction and suppression of actin stress fiber-formation. Especially in the monolayer of cultured peritoneal cells, intercellular dissociation was observed. The concentration of platelet- … More derived growth factor(PDGF) in an intratumoral fluid was significantly higher in ovarian cancers compared with that in benign ovarian tumors. Moreover, the ovarian cancer-derived inhibitory effect on α-SMA expression, which observed in cultured fibroblasts, was partially avoided by adding anti-PDGF neutralizing antibody into the conditioned medium of ovarian cancer cells. These observations opened up a possibility that ovarian cancer-derived secretor factors including PDGF made favorable environment for cancer invasion via down-regulated CNh1 and α-SMA in peritoneal cells and fibroblasts.Adenoviral vector-mediated CNh1 gene transfections into peritoneal cell lines and ovarian cancer cell lines induced the formation of longer and thicker actin steress fibers in each cell line, and the localization of fibers coincided with that of externally transducted CNh1 expression. Simultaneously, a monolayer of the infected peritoneal cells gained stability against the intercellular dissociation induced by the conditioned medium of ovarian cancer cells, and inhibited cancer cell invasion confirmed by cancer cell penetration through the peritoneal cell monolayer. On the other hand, CNh1-transfected ovarian cancer cells showed retarded growth property and invasiveness, the latter of which accompanied the impaired cell motility. When both the peritoneal cells and ovarian cancer cells were transfected at the same time, an additive inhibitory effect against the cancer cell invasion through the peritoneal cell monolayer was confirmed. By in vivo treatment experiments using nude mice, intraperitoneally injected CNh1-adenovirus successfully avoided ovarian cancer-induced change in peritoneal cell surface, and significantly prolonged the survival time of mice inoculated intraperitoneally with ovarian cancer cells.Consequently, CNh1 gene therapy against the peritoneal dissemination ot ovarian cancer is expected to be effective through the inhibitory effect in infected peritoneal cell layers against cancer invasion and the direct anti-tumor effect against cancer invasion and growth. The present therapy may be considered as a novel gene therapy based on the concept that the same gene plays bifunctional therapeutic roles against cancer : one is to repress cancer cells and the other is to reinforce host defense mechanism. Less