Effect of verapamil on cell cycle transit and c-myc gene expression in normal and malignant murine cells

Abstract

Summary Verapamil, the prototype calcium channel blocker, reversibly inhibits cell proliferation in many normal and tumour cell lines (Schmidt et al., Cancer Res., 48, 3617, 1988). We The calcium channel blockers have generated much interest in cancer research since the demonstration that at low concentration (5-10Mm) they augment the cytotoxicity of many standard anti-cancer agents in a variety of tumour cell types (Tsuruo et al., 1983a,b;Yalowich & Ross, 1984Robinson et al., 1985;Ince et al., 1986;Merry et al., 1986). Verapamil, the prototype calcium channel blocker, enhances the cytotoxic effects of both vincristine and adriamycin in vitro as well as in vivo in cells previously resistant to these drugs (Tsuruo et al., 1981(Tsuruo et al., , 1983a. Although the precise mechanism of this increased cytotoxic effect is not completely understood, the calcium channel blockers are thought to act by blocking efflux of the chemotherapeutic agents from the cell (Tsuruo et al., 1982).At higher concentrations (10-100yM), verapamil by itself reversibly inhibits cell growth in several human cell lines (Schmidt et al., 1988). Protein synthesis, DNA synthesis and RNA synthesis are all inhibited within minutes of addition of OO M verapamil to the cells; removal of the drug by simple washing of the cells results in a rapid resumption of cell growth (Schmidt et al., 1988). These reversible antiproliferative effects of verapamil make it an ideal compound to study cell cycle related events.Cell growth is controlled by a cascade of events that ultimately leads to DNA synthesis. Briefly, cell proliferation begins when growth factors interact with the cell membrane, sending a signal via inositol phospholipids to increase cytoplasmic ionised calcium by one pathway and to increase cytoplasmic pH by another pathway (Berridge, 1984). However, stimulation of protein kinase C by phorbol esters (e.g. TPA) directly causes cytoplasmic alkalinisation and in at least some cells this stimulation can by-pass the calcium pathway (Rozengurt & Mendoza, 1985). Furthermore, the involvement of c-onc genes in cell proliferation has been extensively documented (Kahn & Graf, 1986).One of the first genes linked to cell growth was the protooncogene c-myc. C-myc gene expression is known to be linked tightly to cell proliferation, increasing [10][11][12][13][14][15][16][17][18][19][20] al., 1983). The cmyc gene, which is expressed in both malignant and normal cells, encodes a protein that is functionally involved in DNA synthesis (Studzinski et al., 1986). This protein is believed to directly regulate the rate at which cells divide (Cole, 1986 Fidler & Kripke (1977). After injection of 50,000 to 100,000 cells into the tail veins of C57BL/6 mice, the B16 FIO cells form many more pulmonary metastases than the B16 Fl cells within 2-3 weeks. All experiments were performed at least twice.Radioisotope incorporation Actively growing cells (in the exponential phase) were always used when assessing isotope incorporation, Methyl-3H-thymidine was added to cell cultu...

Thrombospondin-1 Signaling through CD47 Inhibits Self-renewal by Regulating c-Myc and Other Stem Cell Transcription Factors

“…This, combined with previous evidence that c-Myc expression is dysregulated in B16 cells53, suggested that CD47 signaling might not regulate c-Myc in these cells. Consistent with this hypothesis, transiently over-expressing CD47 in B16 melanoma cells did not inhibit their growth (Fig.…”

Targeting specificity of dendritic cells on breast cancer stem cells: in vitro and in vivo evaluations

“…Verapamil, an anticancer drug, has been used as a drug target in many different types of cancer. In 1988, Huber et al 38 showed an antiproliferative effect of verapamil on growth rates of certain human brain tumor lines. Growth rates were inhibited by 10%–100% with 10–100 μM verapamil.…”

xCT expression reduces the early cell cycle requirement for calcium signaling

“…The calcium influx phase of this response is inhibited both by inorganic calcium channel blockers and therapeutically important dihydropyridines. Although poorly understood, these inhibitors as wells as benzodiazepine and phenylalkamine-type calcium channel antagonists all arrest cell cycle progression in G1 [12][13][14]. Although the targets of the calcium channel antagonists in non-excitable cells are unknown, their use has been proposed as an approach to the treatment of proliferative diseases such as atherosclerosis and tumorigenesis [15][16][17].…”