Modulation of pancreatic tumor potential by overexpression of protein kinase C ß1.

OBJECTIVE: This study aimed to investigate whether the overexpression of protein kinase C ß1 (PKCß1) is able to modulate the malignant phenotype displayed by the human ductal pancreatic carcinoma cell line PANC1. METHODS: PKCß1 overexpression was achieved using a stable transfection approach. PANC1-PKCß1 and control cells were analyzed both in vitro and in vivo. RESULTS: PANC1-PKCß1 cells displayed a lower growth capacity associated with the down-regulation of the MEK/ERK pathway and cyclin expression. Furthermore, PKCß1 overexpression was associated with an enhancement of cell adhesion to fibronectin and with reduced migratory and invasive phenotypes. In agreement with these results, PANC1-PKCß1 cells showed an impaired ability to secrete proteolytic enzymes. We also found that PKCß1 overexpressing cells were more resistant to cell death induced by serum deprivation, an event associated with G0/G1 arrest and the modulation of PI3K/Akt and NF-κB pathways. Most notably, the overexpression of PKCß1 completely abolished the ability of PANC1 cells to induce tumors in nude mice. CONCLUSIONS: Our results established an important role for PKCß1 in PANC1 cells suggesting it would act as a suppressor of tumorigenic behavior in pancreatic cancer.

PKC Delta (PKCdelta) promotes tumoral progression of human ductal pancreatic cancer.

OBJECTIVE: Our objective was to study the role of protein kinase C delta (PKCdelta) in the progression of human pancreatic carcinoma. METHODS: Protein kinase C delta expression in human ductal carcinoma (n = 22) was studied by immunohistochemistry. We analyzed the effect of PKCdelta overexpression on in vivo and in vitro properties of human ductal carcinoma cell line PANC1. RESULTS: Human ductal carcinomas showed PKCdelta overexpression compared with normal counterparts. In addition, in vitro PKCdelta-PANC1 cells showed increased anchorage-independent growth and higher resistance to serum starvation and to treatment with cytotoxic drugs. Using pharmacological inhibitors, we determined that phosphatidylinositol-3-kinase and extracellular receptor kinase pathways were involved in the proliferation of PKCdelta-PANC1. Interestingly, PKCdelta-PANC1 cells showed a less in vitro invasive ability and an impairment in their ability to migrate and to secrete the proteolytic enzyme matrix metalloproteinase-2. In vivo experiments indicated that PKCdelta-PANC1 cells were more tumorigenic, as they developed tumors with a significantly lower latency and a higher growth rate with respect to the tumors generated with control cells. Besides, only PKCdelta-PANC1 cells developed lung metastasis. CONCLUSION: Our results showed that the overexpression of PKCdelta in PANC1 cells induced a more malignant phenotype in vivo, probably through the modulation of cell proliferation and survival, involving phosphatidylinositol-3-kinase and extracellular receptor kinase signaling pathways.

Opposite effects of protein kinase C beta1 (PKCbeta1) and PKCepsilon in the metastatic potential of a breast cancer murine model.

In this paper we investigated whether protein kinase C (PKC) beta1 and PKCepsilon, members of the classical and novel PKC family, respectively, induce phenotypic alterations that could be associated with tumor progression and metastatic dissemination in a murine model of breast cancer. Stable overexpression of PKCbeta1 in LM3 cells altered their ability to proliferate, adhere, and survive, and impaired their tumorigenicity and metastatic capacity. Moreover, PKCbeta1 induced the re-expression of fibronectin, an extracellular matrix glycoprotein which loss has been associated with the acquisition of a transformed phenotype in different cell models, and exerted an important inhibition on proteases production, effects that probably impact on LM3 invasiveness and dissemination. Conversely, PKCepsilon overexpression enhanced LM3 survival, anchorage-independent growth, and caused a significant increase in spontaneous lung metastasis. Our results suggest PKCbeta1 functions as an inhibitory protein for tumor growth and metastasis dissemination whereas PKCepsilon drives metastatic dissemination without affecting primary tumor growth.

Protein kinase C delta inhibits the production of proteolytic enzymes in murine mammary cells.

In previous studies we have determined that protein kinase C (PKC) delta, a widely expressed member of the novel PKC serine-threonine kinases, induces in vitro changes associated with the acquisition of a malignant phenotype in NMuMG murine mammary cells. In this study we show that PKCdelta overexpression significantly decreases urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-9 (MMP-9) production, two proteases associated with migratory and invasive capacities. This effect is markedly enhanced by treatment with phorbol 12-myristate 13-acetate (PMA). On the other hand, depletion of PKCdelta using RNAi led to a marked increase in both uPA and MMP-9 secretion, suggesting a physiological role for PKCdelta in controlling protease secretion. The MEK-1 inhibitor PD98059 reverted the characteristic pattern of proteases secretion and phospho-ERK1/2 up-regulation observed in PKCdelta overexpressors, suggesting that the PKCdelta effect is mediated by the MEK/ERK pathway. Our results suggest a dual role for PKCdelta in murine mammary cell cancer progression. While this kinase clearly promotes mitogenesis and favors malignant transformation, it also down-modulates the secretion of proteases probably limiting metastatic dissemination.

Protein kinase C delta enhances proliferation and survival of murine mammary cells.

Protein kinase C (PKC) delta, a member of the novel family of PKC serine-threonine kinases, has been implicated in negative regulation of proliferation and apoptosis in a large number of cell types, including breast cancer cell lines, and postulated as a tumor suppressor gene. In this study we show that in murine NMuMG mammary cells PKCdelta promotes a mitogenic response. Overexpression of PKCdelta in NMuMG cells leads to a significant increase in [3H]-tymidine incorporation and cell proliferation, as well as enhanced extracellular signal-regulated kinase (ERK)-mitogen-activated protein kinase (MAPK) activation. Activation of PKCdelta with a phorbol ester leads to elevated cyclin D1 expression and an hyperphosphorylated Rb state. Surprisingly, ectopic expression of PKCdelta conferred anchorage-independent growth capacity to NMuMG cells. PKCdelta overexpressors showed enhanced resistance to apoptotic stimuli, such as serum deprivation or doxorubicin treatment, an effect that correlates with hyperactivation of the Akt survival pathway. Our results provide evidence for a role of PKCdelta as a positive modulator of proliferative and survival signals in immortalized mammary cells. The fact that PKCdelta exerts differential responses depending on the cell context not only highlights the necessity to carefully understand the signaling events controlled by this PKC in each cell type but also suggests that we should be cautious in considering this kinase a target for cancer therapy.

Atypical protein kinase C-zeta modulates clonogenicity, motility, and secretion of proteolytic enzymes in murine mammary cells.

In this paper, we investigated whether protein kinase C-zeta (PKC zeta), a member of the atypical PKC family, induces phenotypic alterations associated with malignant transformation and tumor progression in mammary cells. The stable overexpression of PKC zeta in immortalized mammary epithelial cells (NMuMG), activates the mitogenic extracellular signal-regulated kinase (ERK) pathway, enhanced clonal cell growth and exerts profound effects on proteases secretion. The effect on proteases expression seems to be specific for urokinase-type plasminogen activator and metalloproteinase-9 (MMP-9) because no modulation in MMP-2 and MMP-3 production could be detected. In addition, our experiments demonstrated that PKC zeta overexpression markedly altered the adhesive, spreading, and migratory abilities of NMuMG cells. The overexpression of this enzyme was not sufficient to confer an anchorage-independent growth capacity. An extensive mutational analysis of PKC zeta revealed that the effects observed in NMuMG cells were strictly dependent on the kinase (catalytic) domain of the enzyme. Taken together, these results suggest that in mammary cells PKC zeta modulates several of the critical events involved in tumor development and dissemination through the activation of mitogen activated protein kinase (MAPK)/ERK pathway.