Newly identified properties of known pharmaceuticals and myxobacterial small molecules revealed by screening for autophagy modulators.

Abstract

Autophagy is a cellular degradation and recycling process important for maintaining cellular health and function. It is constitutively active at a low level in eukaryotic cells and can be induced by conditions of cellular stress, such as nutrient starvation. Moreover, autophagy plays an important role in diverse processes such as immunobiology, pathogen infection, ageing, and neurodegenerative and other diseases. Using a high-content fluorescence assay for microtubule-associated protein 1 light chain 3 beta (LC3B), a major player in the autophagic pathway, we screened a library of commercial drugs and natural products for activators and inhibitors of LC3B-positive vesicle accumulation. Positive hits for known autophagy modulators included anisomycin, amphotericin B, carbonyl cyanide m-chlorophenylhydrazone (CCCP) and cytochalasin D. Importantly, we identified several new autophagy modulators, such as aciclovir and myxobacterial vioprolides. Anisomycin, aciclovir and vioprolides promoted intracellular growth of Staphylococcus aureus, a bacterium that is known to be a target of autophagy. In contrast, anisomycin strongly inhibited influenza A virus and SARS-CoV-2 replication. Subsequently, we investigated the influence of these autophagy modulators in a cellular disease model of neuronal vacuolation and spinocerebellar degeneration (NVSD), which is associated with cysteine protease ATG4D mutations. We provide evidence that anisomycin and famciclovir, an aciclovir analogue, can normalise the elevated amount of LC3-positive vesicles in mutant fibroblasts, highlighting their potential for the treatment of NVSD. Thus, the screening method enabled the identification of autophagy-modulating compounds with therapeutic potential.