Blocking of glioma proliferation in vitro and in vivo and potentiating the effects of BCNU and cisplatin: UCN-01, a selective protein kinase C inhibitor

1996 ◽  
Vol 84 (6) ◽  
pp. 1024-1032 ◽  
Author(s):  
Ian F. Pollack ◽  
Stephanie Kawecki ◽  
John S. Lazo
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Lines ◽  
Glioma Cell ◽  
Significant Activity ◽  
Inhibition Of Proliferation ◽  
Kinase C ◽  
Glioma Cell Lines

✓ Seven-hydroxystaurosporine (UCN-01) is a derivative of the nonselective protein kinase inhibitor staurosporine that exhibits significant selectivity for protein kinase C (PKC) in comparison to a variety of other intracellular kinases and appears to be well tolerated in vivo at concentrations sufficient to achieve effective inhibition of PKC. Because recent studies have indicated that the proliferation of malignant gliomas may result from activation of PKC-mediated pathways and, conversely, may be inhibited by blocking PKC, the authors examined the efficacy of this agent as an inhibitor of proliferation in three established and three low-passage malignant glioma cell lines in vitro. A striking inhibition of proliferation was produced by UCN-01 in each of the cell lines, with a median effective concentration of 20 to 100 nM, which correlated with the median in vitro PKC inhibitory concentration of 20 to 60 nM for this agent in the U-87 and SG-388 glioma cell lines. Inhibition-recovery studies of clonogenic activity indicated that UCN-01 had both cytostatic and cytotoxic effects on the treated cells. Proliferation resumed after short-term (6- and 24-hour) exposures to this agent; in contrast, with longer exposures, recovery of proliferative activity was severely compromised. In addition, UCN-01 enhanced the inhibition of glioma cell proliferation achieved with conventional chemotherapeutic agents, exhibiting synergistic effects with cisplatin and additive effects with 1,3-bis(2-chloroethyl)-1-nitrosourea. In vivo studies in which UCN-01 was administered by continuous intraperitoneal infusion in subcutaneous and intracranial intraparenchymal nude rat models demonstrated significant activity against U-87 glioma xenografts at dose levels that were well tolerated. It is concluded that UCN-01 is an effective agent for the inhibition of glioma proliferation in vitro and in vivo and has potential for clinical applicability in the treatment of human gliomas.

scholarly journals Semi-Synthetic Ingenol Derivative from Euphorbia tirucalli Inhibits Protein Kinase C Isotypes and Promotes Autophagy and S-phase Arrest on Glioma Cell Lines

Molecules ◽  
2019 ◽  
Vol 24 (23) ◽  
pp. 4265 ◽  
Author(s):  
Silva ◽  
Rosa ◽  
Tansini ◽  
Martinho ◽  
Tanuri ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Lines ◽  
Glioma Cell ◽  
Glioma Cells ◽  
S Phase ◽  
Kinase C ◽  
Phase Arrest ◽  
S Phase Arrest ◽  
Glioma Cell Lines

The identification of signaling pathways that are involved in gliomagenesis is crucial for targeted therapy design. In this study we assessed the biological and therapeutic effect of ingenol-3-dodecanoate (IngC) on glioma. IngC exhibited dose-time-dependent cytotoxic effects on large panel of glioma cell lines (adult, pediatric cancer cells, and primary cultures), as well as, effectively reduced colonies formation. Nevertheless, it was not been able to attenuate cell migration, invasion, and promote apoptotic effects when administered alone. IngC exposure promoted S-phase arrest associated with p21CIP/WAF1 overexpression and regulated a broad range of signaling effectors related to survival and cell cycle regulation. Moreover, IngC led glioma cells to autophagy by LC3B-II accumulation and exhibited increased cytotoxic sensitivity when combined to a specific autophagic inhibitor, bafilomycin A1. In comparison with temozolomide, IngC showed a mean increase of 106-fold in efficacy, with no synergistic effect when they were both combined. When compared with a known compound of the same class, namely ingenol-3-angelate (I3A, Picato®), IngC showed a mean 9.46-fold higher efficacy. Furthermore, IngC acted as a potent inhibitor of protein kinase C (PKC) activity, an emerging therapeutic target in glioma cells, showing differential actions against various PKC isotypes. These findings identify IngC as a promising lead compound for the development of new cancer therapy and they may guide the search for additional PKC inhibitors.

scholarly journals Protein kinase C inhibitors induce apoptosis in human malignant glioma cell lines

FEBS Letters ◽  
1994 ◽  
Vol 345 (1) ◽  
pp. 43-46 ◽  
Author(s):  
William T. Couldwell ◽  
David R. Hinton ◽  
Shikun He ◽  
Thomas C. Chen ◽  
Ibrahim Sebat ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Malignant Glioma ◽  
Cell Lines ◽  
Glioma Cell ◽  
Protein Kinase C Inhibitors ◽  
Kinase C ◽  
Glioma Cell Lines ◽  
Human Malignant Glioma

scholarly journals Protein Kinase C Signaling Mediates a Program of Cell Cycle Withdrawal in the Intestinal Epithelium

2000 ◽  
Vol 151 (4) ◽  
pp. 763-778 ◽  
Author(s):  
Mark R. Frey ◽  
Jennifer A. Clark ◽  
Olga Leontieva ◽  
Joshua M. Uronis ◽  
Adrian R. Black ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Intestinal Epithelium ◽  
Molecular Mechanisms ◽  
Model Systems ◽  
Intestinal Epithelial ◽  
Kinase C

Members of the protein kinase C (PKC) family of signal transduction molecules have been widely implicated in regulation of cell growth and differentiation, although the underlying molecular mechanisms involved remain poorly defined. Using combined in vitro and in vivo intestinal epithelial model systems, we demonstrate that PKC signaling can trigger a coordinated program of molecular events leading to cell cycle withdrawal into G0. PKC activation in the IEC-18 intestinal crypt cell line resulted in rapid downregulation of D-type cyclins and differential induction of p21waf1/cip1 and p27kip1, thus targeting all of the major G1/S cyclin-dependent kinase complexes. These events were associated with coordinated alterations in expression and phosphorylation of the pocket proteins p107, pRb, and p130 that drive cells to exit the cell cycle into G0 as indicated by concomitant downregulation of the DNA licensing factor cdc6. Manipulation of PKC isozyme levels in IEC-18 cells demonstrated that PKCα alone can trigger hallmark events of cell cycle withdrawal in intestinal epithelial cells. Notably, analysis of the developmental control of cell cycle regulatory molecules along the crypt–villus axis revealed that PKCα activation is appropriately positioned within intestinal crypts to trigger this program of cell cycle exit–specific events in situ. Together, these data point to PKCα as a key regulator of cell cycle withdrawal in the intestinal epithelium.

Mitogen stimulation of Na+-H+ exchange: differential involvement of protein kinase C

1987 ◽  
Vol 253 (2) ◽  
pp. C219-C229 ◽  
Author(s):  
L. L. Muldoon ◽  
G. A. Jamieson ◽  
A. C. Kao ◽  
H. C. Palfrey ◽  
M. L. Villereal
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Types ◽  
Intact Cells ◽  
Kinase C ◽  
Protein Kinase C Activity ◽  
Human Fibroblast Strains ◽  
Stimulation Of

The mitogen-induced activation of Na+-H+ exchange was investigated in two cultured human fibroblast strains (HSWP and WI-38 cells) that, based on previous studies, differed in their response to the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) (L. M. Vincentini and M. L. Villereal, Proc. Natl. Acad. Sci. USA 82: 8053-8056, 1985). The role of protein kinase C in the activation of Na+-H+ exchange was investigated by comparing the effects of TPA on Na+ influx, in vitro phosphorylation, and in vivo phosphorylation in both cell types. Although both cell types have significant quantities of protein kinase C activity that can be activated by TPA in intact cells, the addition of TPA to intact cells stimulates Na+ influx in WI-38 cells but not in HSWP cells, indicating that in HSWP cells the stimulation of protein kinase C is not sufficient to activate the Na+-H+ exchanger. Cells were then depleted of protein kinase C activity by chronic treatment with high doses of TPA. Both HSWP and WI-38 cells were rendered protein kinase C deficient by this treatment as determined by in vitro and in vivo phosphorylation studies. Protein kinase C-deficient HSWP cells lose the ability for TPA to inhibit the serum-induced activation of Na+-H+ exchange, but there is no reduction in the stimulation of Na+ influx by serum, bradykinin, vasopressin, melittin, or vanadate, indicating that protein kinase C activity is not necessary for the mitogen-induced activation of Na+-H+ exchange in HSWP cells by agents known to stimulate phosphatidylinositol turnover (G. A. Jamieson and M. Villereal. Arch. Biochem. Biophys. 252: 478-486, 1987). In contrast, depletion of protein kinase C activity in WI-38 cells significantly reduces both the TPA- and the serum-induced activation of the Na+-H+ exchange system, suggesting that protein kinase C activity is necessary for at least a portion of the mitogen-induced activation of the Na+-H+ exchanger in WI-38 cells. These results indicate that the mechanisms for regulating Na+-H+ exchange can differ dramatically between different types of fibroblasts.

14-3-3 Isotypes facilitate coupling of protein kinase C-ζ to Raf-1: negative regulation by 14-3-3 phosphorylation

2000 ◽  
Vol 345 (2) ◽  
pp. 297-306 ◽  
Author(s):  
Paulus C. J. VAN DER HOEVEN ◽  
José C. M. VAN DER WAL ◽  
Paula RUURS ◽  
Marc C. M. VAN DIJK ◽  
Wim J. VAN BLITTERSWIJK
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Complex Formation ◽  
Dependent Manner ◽  
Cos Cells ◽  
Kinase C ◽  
Pkc Ζ ◽  
Co Precipitation

14-3-3 Proteins may function as adapters or scaffold in signal-transduction pathways. We found previously that protein kinase C-ζ (PKC-ζ) can phosphorylate and activate Raf-1 in a signalling complex [van Dijk, Hilkmann and van Blitterswijk (1997) Biochem. J. 325, 303-307]. We report now that PKC-ζ-Raf-1 interaction is mediated by 14-3-3 proteins in vitro and in vivo. Co-immunoprecipitation experiments in COS cells revealed that complex formation between PKC-ζ and Raf-1 is mediated strongly by the 14-3-3β and -θ isotypes, but not by 14-3-3ζ. Far-Western blotting revealed that 14-3-3 binds PKC-ζ directly at its regulatory domain, where a S186A mutation in a putative 14-3-3-binding domain strongly reduced the binding and the complex formation with 14-3-3β and Raf-1. Treatment of PKC-ζ with lambda protein phosphatase also reduced its binding to 14-3-3β in vitro. Preincubation of an immobilized Raf-1 construct with 14-3-3β facilitated PKC-ζ binding. Together, the results suggest that 14-3-3 binds both PKC-ζ (at phospho-Ser-186) and Raf-1 in a ternary complex. Complex formation was much stronger with a kinase-inactive PKC-ζ mutant than with wild-type PKC-ζ, supporting the idea that kinase activity leads to complex dissociation. 14-3-3β and -θ were substrates for PKC-ζ, whereas 14-3-3ζ was not. Phosphorylation of 14-3-3β by PKC-ζ negatively regulated their physical association. 14-3-3β with its putative PKC-ζ phosphorylation sites mutated enhanced co-precipitation between PKC-ζ and Raf-1, suggesting that phosphorylation of 14-3-3 by PKC-ζ weakens the complex in vivo. We conclude that 14-3-3 facilitates coupling of PKC-ζ to Raf-1 in an isotype-specific and phosphorylation-dependent manner. We suggest that 14-3-3 is a transient mediator of Raf-1 phosphorylation and activation by PKC-ζ.

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