Clarithromycin inhibits autophagy in colorectal cancer by regulating the hERG1 potassium channel interaction with PI3K

Giulia Petroni,Matteo Stefanini,Giacomo Bagni,Andrea Becchetti,Tiziano Lottini,Annarosa Arcangeli,Goran Kragol,Claudia Duranti,Jessica Iorio

doi:10.1038/s41419-020-2349-8

Giulia Petroni, Matteo Stefanini + Show 7 more

Open Access

https://doi.org/10.1038/s41419-020-2349-8

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Abstract

We have studied how the macrolide antibiotic Clarithromycin (Cla) regulates autophagy, which sustains cell survival and resistance to chemotherapy in cancer. We found Cla to inhibit the growth of human colorectal cancer (CRC) cells, by modulating the autophagic flux and triggering apoptosis. The accumulation of cytosolic autophagosomes accompanied by the modulation of autophagic markers LC3-II and p62/SQSTM1, points to autophagy exhaustion. Because Cla is known to bind human Ether-à-go-go Related Gene 1 (hERG1) K+ channels, we studied if its effects depended on hERG1 and its conformational states. By availing of hERG1 mutants with different gating properties, we found that fluorescently labelled Cla preferentially bound to the closed channels. Furthermore, by sequestering the channel in the closed conformation, Cla inhibited the formation of a macromolecular complex between hERG1 and the p85 subunit of PI3K. This strongly reduced Akt phosphorylation, and stimulated the p53-dependent cell apoptosis, as witnessed by late caspase activation. Finally, Cla enhanced the cytotoxic effect of 5-fluorouracil (5-FU), the main chemotherapeutic agent in CRC, in vitro and in a xenograft CRC model. We conclude that Cla affects the autophagic flux by impairing the signaling pathway linking hERG1 and PI3K. Combining Cla with 5-FU might be a novel therapeutic option in CRC.

Highlights

Autophagy is a homeostatic and evolutionarily conserved process characterized by cellular self-digestion and the removal of defective organelles and proteins, aimed to maintain cellular biosynthesis during nutrient deprivation or metabolic stress[1]
The only identified Cla targets in mammalian cell membranes are the K+ channel encoded by the ether-à-go-go related gene 135,36, and the solute carrier organic anion transporters SLCOB1 and SLCOB337. human Ether-à-go-go Related Gene 1 (hERG1) is often aberrantly expressed in cancers[38,39,40,41], including CRC42, and we have previously shown that hERG1 operates in a peculiar way in cancer cells, mainly modulating the intracellular signaling triggered by cell adhesion[43]
We investigated how the interplay between hERG1 and the PI3K/Akt pathway is regulated by Cla, and how this mechanism regulates the autophagic effects of the macrolide

Summary

Introduction

Autophagy is a homeostatic and evolutionarily conserved process characterized by cellular self-digestion and the removal of defective organelles and proteins, aimed to maintain cellular biosynthesis during nutrient deprivation or metabolic stress[1]. In certain circumstances, autophagy constitutes a major mechanism for cell killing, or can activate other death pathways[4]. Such dual role of autophagy is clearly observed in colorectal cancer (CRC), one of the most aggressive cancer types. Many genes and proteins involved in the autophagic process are implicated in CRC tumor progression[5,6,7,8,9]. The role of autophagy in progression and survival of CRC patients is still controversial[7,10,11,12,13,14]

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