Development of a p62 biodegrader for autophagy targeted degradation

Harnessing autophagy for targeted degradation of intracellular cargo is a promising extension to proteasome-based targeted protein degradation because of the capacity and versatility of lysosomes, broadening the scope of therapeutic targets. First autophagy-based degraders and glues have been identified; however, it remains unclear which component of the autophagy lysosomal pathway is most efficacious to induce the selective degradation of targets when brought into proximity. Here, we describe a platform to systematically evaluate induced proximity of autophagy candidates with mitochondria using a nanobody-based and small molecule heterobifunctional dimerizer approach. 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 degradation of mitochondria. In contrast, self-oligomerizing autophagy cargo receptors outperform ATG8-like effectors and autophagy kinases in clearing a soluble cytosolic protein. By developing an intrabody against the autophagy cargo receptor p62, one of the strongest effectors for autophagy targeted degradation, we demonstrate that recruitment of endogenous p62 with a heterobifunctional biodegrader is sufficient to clear mitochondria. This biodegrader, however, is unable to induce degradation of soluble cytosolic proteins due to its inhibitory effect on the self-oligomerization of p62. Our study highlights the importance of avidity for autophagy targeted degradation and suggests that autophagy cargo receptors are attractive entry points for the development of heterobifunctional degraders.