Inhibition of T‐type Ca²⁺ channels by endostatin attenuates human glioblastoma cell proliferation and migration

Abstract

BACKGROUND AND PURPOSEEndostatin (ES) is a c-terminal proteolytic fragment of collagen XVIII with promising antitumour properties in several tumour models, including human glioblastoma. We hypothesized that this peptide could interact with plasma membrane ion channels and modulate their functions. EXPERIMENTAL APPROACHUsing cell proliferation and migration assays, patch clamp and Western blot analysis, we studied the effects of ES on the proliferation and migration of human glioblastoma U87 cells, mediated by T-type Ca 2+ channels. KEY RESULTSExtracellular application of ES reversibly inhibited T-type Ca 2+ channel currents (T-currents) in U87 cells, whereas L-type Ca 2+ currents were not affected. This inhibitory effect was associated with a hyperpolarizing shift in the voltage-dependence of inactivation but was independent of G-protein and protein tyrosine kinase-mediated pathways. All three a1 subunits of T-type Ca 2+ channels (CaV3), a1G (CaV3.1), a1H (CaV3.2) and a1I (CaV3.3), were endogenously expressed in U87 cells. Using transfected HEK293 or CHO cells, we showed that only CaV3.1 and CaV3.2, but not CaV3.3 or CaV1.2 (L-type), channel currents were significantly inhibited. More interestingly, ES inhibited the proliferation and migration of U87 cells in a dose-dependent manner. Pretreatment of the cells with the specific T-type Ca 2+ channel blocker mibefradil occluded these inhibitory effects of ES. CONCLUSION AND IMPLICATIONSThis study provides the first evidence that the antitumour effects of ES on glioblastoma cells is through direct inhibition of T-type Ca 2+ channels and gives new insights into the future development of a new class of antiglioblastoma agents that target the proliferation and migration of these cells. LINKED ARTICLEThis article is commented on by Santoni et al., pp. 1244Santoni et al., pp. -1246

The Physiology, Pathology, and Pharmacology of Voltage-Gated Calcium Channels and Their Future Therapeutic Potential

“…They appear to be a contributor to abnormal growth of ventricular cells (Martínez et al, 1999) and may dispose hypertrophic tissue to arrhythmias (Nuss and Houser, 1993). Finally, there is accumulating evidence that T-type calcium channels may also participate in the growth of certain cancers (Ohkubo and Yamazaki, 2012;Rim et al, 2012;Zhang et al, 2012;Das et al, 2013;Gackière et al, 2013;Dziegielewska et al, 2014).…”

Remodelling of Ca2+transport in cancer: how it contributes to cancer hallmarks?

“…The following Ca 2þ permeable channels have been implicated in the enhanced migration of various types of cancer cells (figure 3): TRPC1 [64], TRPM7 [65 -71], TRPM8 [43,72,73], TRPV1 [74], TRPV2 [75], TRPV6 [40], STIM1 and ORAI1 SOC constituents [13,[76][77][78][79], some types of VGCCs [35,80]. Owing to the presence of mechanical stimulusdependent and Mg-ATP-dependent modes of activation TRPM7 was especially implicated in providing spatially restricted Ca 2þ entry in response to local membrane stretch in front of migrating cells [66,69] as well as promoting m-calpain-mediated disassembly of peripheral adhesions under decreased intracellular Mg-ATP levels [65] typically found in hypoxic tumour conditions.…”

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

“…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). Aberrant expression of T-type Ca 2þ channels in cancer cells is thought to promote cell survival, proliferation, and motility (3,11); however, the molecular mechanisms for these effects are poorly understood.…”

“…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).…”