Photo-Induced Inactivation of Protein Kinase Cα by Dequalinium Inhibits Motility of Murine Melanoma Cells
- 1Department of Chemistry and Biochemistry, Queens College-City University of New York, Flushing, New York (R.M.S., S.A.R.), 2Richard Dimbleby Department of Cancer Research, Imperial Cancer Research Fund Laboratory, St. Thomas' Hospital, London, United Kingdom (M.S., J.F.M.), and 3Institute of Medical Chemistry and Biochemistry, University of Innsbruck, Innsbruck, Austria (F.U.)
Abstract
Dequalinium (DECA) is a potent antitumor agent and inhibitor of protein kinase C (PKC). Previously it was shown that PKCα activity in vitro could be irreversibly inhibited when treated with DECA at low micromolar concentrations and irradiated with 366 nm of light. This approach was used to probe the role of intracellular PKC activity in the motility of metastatic murine melanoma B16 F10 cells and as a target for DECA analogs with increasing PKC inhibitory potencies. Pretreatment of a monolayer of B16 F10 cells with 250 nM of a DECA analog in the presence of UV irradiation for 5 min resulted in 1) complete inhibition of cell motility for up to 4 h in a time-lapse motility assay and 40 to 60% inhibition of cell migration in a Boyden chamber, and 2) inhibition by 40 to 60% of intracellular phosphatidylserine/Ca2+-dependent PKC catalytic activity, signifying inactivation of a conventional PKC isoform. Because PKCα is the only conventional PKC isoform detected in B16 F10 cells, a stably transfected clone expressing a kinase-defective mutant of PKCα was developed that exhibited a substantial loss of adhesion and motility and was refractory to further inhibition by DECA. These findings identify PKCα catalytic activity both as a mechanistic component of cell motility and adhesion and as a critical intracellular target of DECA. These studies further suggest that the combined use of UV with nanomolar concentrations of DECA offers an effective chemotherapeutic approach to inhibit metastatic behavior of melanoma cells.
Footnotes
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Send reprint requests to: Susan A. Rotenberg, Ph.D., Department of Chemistry and Biochemistry, Queens College-City University of New York, 65-30 Kissena Boulevard, Flushing, NY 11367-1597. E-mail: Susan_Rotenberg{at}qc.edu
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↵1 Src kinase does not udnergo inhibition in vitro by concentrations of C10-DECA up to 1 mM (S. A. Rotenberg, unpublished results).
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This work was supported by grants from the National Institutes of Health (CA60618), the Elsa U. Pardee Foundation, the Professional Staff Congress of the City University of New York Research Foundation, and the Austrian Fond SFB, F208, Biological communication systems. Aspects of this work appeared in the Proceedings of the American Association for Cancer Research, Vol. 88 (abstract 1551) and Vol. 90 (abstract 3702).
- Abbreviations:
- PKC
- protein kinase C
- DECA
- 1,1′-decamethylene-bis-4-aminoquinaldinium di-iodide
- PS
- phosphatidylserine
- TPA
- 12-O-tetradecanoylphorbol-13-acetate
- DMSO
- dimethyl sulfoxide
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- Received October 29, 1999.
- Accepted June 2, 2000.
- The American Society for Pharmacology and Experimental Therapeutics
