Differential role of cathepsins B and L in autophagy-associated cell death induced by arsenic trioxide in U87 human glioblastoma cells

Abstract

Arsenic trioxide (arsenite) was the first chemotherapeutic drug to be described and is now being rediscovered in cancer treatment, including glioblastoma multiforme. Arsenite toxicity triggers autophagy in cancer cells, although final stages of the process involve executive caspases, suggesting an interplay between autophagic and apoptotic pathways that awaits to be explained at a molecular level. We evaluated the contribution of the lysosomal cathepsins (Cat) L and B, which are upregulated in glioblastomas, in the mechanism of arsenite toxicity in human glioblastoma cells. Arsenite treatment induced autophagosome formation and permeabilization of mitochondria, followed by caspase 3/7-mediated apoptosis. The autophagy inhibitor 3-methyladenine protected from arsenite toxicity, whereas bafilomycin A1 did not. Furthermore, arsenite significantly decreased CatB levels and selectively inhibited its cellular and recombinant protein activity, while not affecting CatL. However, downregulation of CatL greatly enhanced apoptosis by arsenite. Our results show that arsenite toxicity involves a complex interplay between autophagy and apoptosis in human glioblastoma cells and is associated with inhibition of CatB, and that this toxicity is highly exacerbated by simultaneous CatL inhibition. The latter points to a synergy that could be used in clinical treatment to lower the therapeutic dose, thus avoiding the toxic side effects of arsenite in glioblastoma management.

Synergistic Effects of Arsenic Trioxide and Silibinin on Apoptosis and Invasion in Human Glioblastoma U87MG Cell Line

“…In our study, even though the mRNA level of cathepsin B was decreased dramatically by ATO and in the combination treatments; however, we did not observe any significant differences in the activity of intracellular cathepsin B between the treated cells and untreated cells (data not shown). These results confirm the recently reported study that ATO (2 lM) did [32]. Several studies have reported the roles of extracellular cathepsin B in tumor invasion [33,34]; however, there are also other reports that introduce cathepsin B as an inducer of autophagic cell death, apoptosis or as a molecular link between autophagy and apoptosis [30,31,[34][35][36].…”

Inhibition of cathepsin L lowers the apoptotic threshold of glioblastoma cells by up-regulating p53 and transcription of caspases 3 and 7

“…The results are similar to those achieved with anti-sense transfection [10]. Cathepsin L was anti-apoptotic regardless of whether the induction of apoptosis triggered the intrinsic or extrinsic pathway or even an indirect pathway via autophagy upon treatment with arsenic trioxide [21].…”

“…CatL inhibition strongly increases the sensitivity of GBM cells to the induction of cell death by arsenic trioxide [21], which is already in a clinical trial to treat malignant glioma (www.clinicaltrials.gov). Drug targets topoisomerase IIa, estrogen receptor, Bcr-Abl, and histone deacetylase have also been shown to accumulate after CatL inhibition, and cells with less CatL activity were consequently less resistant to their corresponding chemotherapeutics [34].…”

Retinoblastoma protein regulates the crosstalk between autophagy and apoptosis, and favors glioblastoma resistance to etoposide

“…Katayama et al 12 showed that the pharmacological inhibition of the autophagic process, in its final steps, increased the apoptotic rate induced by the compound. This same cross talk between autophagy and apoptosis has also been shown for other chemotherapy agents, including temozolomide, arsenic trioxide, resveratrol, camptothecin, cisplatin and 5-fluoracil, in many types of tumor cells 12, 13, 14, 15, 16 But still, the molecular mechanisms regulating the interplay between these two events are poorly understood.…”