The molecular mechanism of protein kinase C inhibitors on anti-metastasis

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We investigated the mechanism of protein kinase C(PKC) inhibitors on anti-metastasis. PKC412 (4'-N-benzoyl staurosporine), a conventional PKC(alpha, beta and gamma) inhibitor, reduced the ability of mouse malignant melanoma(B16-BL6) cells to form lung colonies in mice and reduced invasion of the extracellular matrix in vitro when pie-incubated with the cells for 1 hour. Further, PKC412(200mg/kg/day for 4 weeks, p.o.) significantly prolonged survival time in a spontaneous metastatic mouse model, produced by subcutaneous inoculation of B16-BL6 cells(1×10^6 cells) into the right footpad of C57BL/6Cr mice Mowed by surgical amputation of the primary tumor 2 weeks after tumor inoculation. To elucidate mechanisms of anti-invasive action for PKC412, we measured cell motility, matrix metalloproteinase(MMP) activity secreted from cells and expression of integrin beta 1 protein in cells. As a result, PKC412 decreased the expression of integrin beta 1 protein of B16-BL6 cells in a dose-dependent manner. However, PKC412 could not inhibit cell motility and MMP activity of the melanoma cells. These results suggest that PKC412 inhibits the process of invasion in the metastatic pathway of melanoma cells by attenuation of integrin beta 1 expression. Finally, we examined metastasis-related PKC isoforms using rottlerin, a specific PKC delta inhibitor. Since rottlerin could not block the hematogenic lung metastasis in mice using B16-BL6 cells, conventional PKC isoforms were considered to induce tumor metastasis, while the delta isoform did not.