Dysfunctional autophagy induced by the pro-apoptotic natural compound climacostol in tumour cells

Abstract

Autophagy occurs at a basal level in all eukaryotic cells and may support cell survival or activate death pathways. Due to its pathophysiologic significance, the autophagic machinery is a promising target for the development of multiple approaches for anti-neoplastic agents. We have recently described the cytotoxic and pro-apoptotic mechanisms, targeting the tumour suppressor p53, of climacostol, a natural product of the ciliated protozoan Climacostomum virens. We report here on how climacostol regulates autophagy and the involvement of p53-dependent mechanisms. Using both in vitro and in vivo techniques, we show that climacostol potently and selectively impairs autophagy in multiple tumour cells that are committed to die by apoptosis. In particular, in B16-F10 mouse melanomas climacostol exerts a marked and sustained accumulation of autophagosomes as the result of dysfunctional autophagic degradation. We also provide mechanistic insights showing that climacostol affects autophagosome turnover via p53-AMPK axis, although the mTOR pathway unrelated to p53 levels plays a role. In particular, climacostol activated p53 inducing the upregulation of p53 protein levels in the nuclei through effects on p53 stability at translational level, as for instance the phosphorylation at Ser15 site. Noteworthy, AMPKα activation was the major responsible of climacostol-induced autophagy disruption in the absence of a key role regulating cell death, thus indicating that climacostol effects on autophagy and apoptosis are two separate events, which may act independently on life/death decisions of the cell. Since the activation of p53 system is at the molecular crossroad regulating both the anti-autophagic action of climacostol and its role in the apoptosis induction, it might be important to explore the dual targeting of autophagy and apoptosis with agents acting on p53 for the selective killing of tumours. These findings also suggest the efficacy of ciliate bioactive molecules to identify novel lead compounds in drug discovery and development