The Ca²⁺Channel Antagonists Mibefradil and Pimozide Inhibit Cell Growth via Different Cytotoxic Mechanisms

Abstract

We show that mitogenic cells expressing T-type Ca 2ϩ channels (T-channels) are more sensitive to the antiproliferative effects of the drugs pimozide and mibefradil than cells without significant T-channel expression. The growth of Y79 and WERI-Rb1 retinoblastoma cells, as well as MCF7 breast cancer epithelial cells, all of which express T-channel current and mRNA for T-channel subunits, is inhibited by pimozide and mibefradil with IC 50 values between 0.6 and 1.5 M. Proliferation of glioma C6 cells, which show little T-channel expression, is less sensitive to these drugs (IC 50 ϭ 8 and 5 M for pimozide and mibefradil, respectively). Neither drug seems to alter cell cycle or the expression of cyclins. Although this strong correlation between T-channel expression and growth inhibition exists, the following results suggest that the drugs inhibit cell growth via different cytotoxic pathways: 1) pimozide and mibefradil have additive effects on T-channel current inhibition, whereas the antiproliferative activity of the drugs together is synergistic; 2) an increase in the number of apoptotic Y79 and MCF7 cells and a decrease in the mRNA for the antiapoptotic gene Bcl-2 is detected only in pimozide-treated cells, whereas in mibefradiltreated cells, the toxicity is primarily necrotic; and 3) growth inhibition by mibefradil is reduced in Y79 cells transfected with T-channel antisense and in differentiated Y79 cells (which have decreased T-channel expression), but growth inhibition by pimozide is affected to a lesser extent. These results suggest that pimozide and mibefradil inhibit cell proliferation via different cytotoxic pathways and that in the case of pimozide, it is unlikely that this effect is mediated solely by T-channel inhibition.

Melatonin affects voltage-dependent calcium and potassium currents in MCF-7 cell line cultured either in growth or differentiation medium

“…In relation to I Ca , our findings demonstrate that melatonin treatment was able to reduce both T-and L-type Ca 2 þ current amplitude and to provoke an alteration of the current voltage dependence along with a decrease of intracellular Ca 2 þ concentration. Since Ca 2 þ seems to be a crucial regulator of the cell cycle necessary for proper proliferation (Bertolesi et al, 2002;Gray et al, 2004;Naziroğlu et al, 2012;Wang et al, 2000), the associated reduction of free [Ca 2 þ ] i observed in our experiments supports the clue that melatonin may contribute to the decline of proliferation also disturbing internal Ca 2 þ homeostasis. The G-protein-coupled melatonin MT1 receptors are expressed in MCF-7 cells (Girgert et al, 2009;Hill et al, 2011b;Jablonska et al, 2013;Jawed et al, 2007;Rich et al, 1999) and, via the inhibition of adenylate cyclase activity (Jablonska et al, 2013;Jawed et al, 2007), their activation leads to the decrease of adenosine 3 0 , 5 0 -cyclic monophosphate (cAMP) synthesis.…”

“…Since several studies demonstrated that human mammary cancer cell lines express voltage-dependent Ca 2 þ channels (Bertolesi et al, 2002;Ohkubo and Yamazaki, 2012), we intended to evaluate the effects of melatonin on voltage-dependent Ca 2 þ currents in MCF-7 cells cultured in growth medium. In control conditions we could observe a low voltage-activated inward transient current starting from À 60 mV up to À 40 mV.…”

“…In particular, by electrophysiological technique, we evaluated the action of melatonin on resting membrane potential and on voltage-dependent ionic channels known to play a role in proliferation (Baglioni et al, 2012;Bertolesi et al, 2002;Currò, 2014;Enomoto et al, 1986;Gamper et al, 2002;Girgert et al, 2009;Grant et al, 2009;Gray et al, 2004;Haren et al, 2010;Kunzelmann, 2005).…”

T-Type Ca2+ Channel Inhibition Induces p53-Dependent Cell Growth Arrest and Apoptosis through Activation of p38-MAPK in Colon Cancer Cells

“…Voltage gated Ca 2þ channels facilitate transient Ca 2þ influx from the environment into the cytoplasm and appear mostly in excitable tissues, but also are unusually expressed in cancer cells. Low-voltage activated Ca 2þ channels, termed T-type Ca 2þ channels, recently gained attention in cancer therapy, because their inhibition decreased proliferation of glioblastoma cells (3,4), breast adenocarcinoma cells (5,6), melanoma cells (7), and esophageal carcinoma cells (8). In addition, an antagonist selective for T-type Ca 2þ channels, mibefradil, has been proposed recently as a sensitizing agent with activity in vivo in combination with chemo-(9) or radiotherapy (10).…”

T-type voltage-activated calcium channel Cav3.1, but not Cav3.2, is involved in the inhibition of proliferation and apoptosis in MCF-7 human breast cancer cells

“…In previous studies, T-type channel blockers such as mibefradil and pimozide, which cannot distinguish between Ca v 3.1 and Ca v 3.2 channels, were used to investigate the regulation of cancer cell proliferation (13)(14)(15)(16)(17). Since there is a possibility that these isoforms exhibit different functions in tumors, if both isoforms are inhibited by such drugs to the same extent, the results should be interpreted with caution.…”