Development of a p62 biodegrader for autophagy targeted degradation

Harnessing autophagy for targeted degradation is a promising extension to proteasome-based targeted protein degradation because of the capacity and versatility of lysosomes to degrade large and complex cargo, thus broadening the scope of therapeutic targets. While small-molecule degraders recruiting the autophagy machinery to targets are starting to emerge, it remains unclear which component of the autophagy lysosomal pathway is most efficacious to induce selective target degradation. Here, we describe two orthogonal induced-proximity strategies to identify and prioritize autophagy effectors that are sufficient to degrade organelles as well as soluble targets. We show that induced proximity of different effectors such as autophagy cargo receptors, ATG8-like proteins or the kinases ULK1 and TBK1 are sufficient to trigger mitophagy. In contrast, self-oligomerizing autophagy cargo receptors outperform ATG8- like effectors and autophagy-related kinases in clearing a soluble cytosolic protein. We further explore the importance of avidity for targeted degradation via autophagy and reveal that the PB1 domain of p62 fused to a LIR peptide is a minimal degron to induce the degradation of mitochondria as well as cytosolic proteins. By developing a novel and highly selective intrabody against the autophagy cargo receptor p62 into a heterobifunctional degrader, we demonstrate that recruitment of endogenous p62 is sufficient to clear mitochondria. This biodegrader, however, is unable to induce degradation of soluble cytosolic proteins due to its inhibitory effect on p62 self-oligomerization. Our study highlights the importance of avidity and suggests that autophagy cargo receptors are attractive entry points for the development of heterobifunctional degraders for complex targets such as organelles or protein aggregates.