Expanding the horizons of targeted protein degradation: A non-small molecule perspective

Abstract

Targeted protein degradation (TPD) represented by proteolysis targeting chimeras (PROTACs) marks a significant stride in drug discovery. A plethora of innovative technologies inspired by PROTAC have not only revolutionized the landscape of TPD but have the potential to unlock functionalities beyond degradation. Non-small-molecule-based approaches play an irreplaceable role in this field. A wide variety of agents spanning a broad chemical spectrum, including peptides, nucleic acids, antibodies, and even vaccines, which not only prove instrumental in overcoming the constraints of conventional small molecule entities but also provided rapidly renewing paradigms. Herein we summarize the burgeoning non-small molecule technological platforms inspired by PROTACs, including three major trajectories, to provide insights for the design strategies based on novel paradigms.

Keywords: Autophagy; Endocytosis; Post-translational modification; Proteolysis targeting chimera; Proximity-inducing modality; Targeted protein degradation.

Graphical abstract

Figure 1

Constructional considerations and mechanisms of antibody-based PROTACs. (A) Schematic representation of DAC construction. (B) Details of DAC mechanism-of-action. (C) Ligase-based PROTAB-mediated membrane POI degradation.

Figure 2

Structure of DACs and their corresponding small molecule PROTACs. The POI ligands are marked in magenta, while the E3 ligands are highlighted in blue. DAR, drug antibody ratio.

Figure 3

Structures and mechanisms of TF-PROTACs and RNA-PROTACs. (A) Schematic diagram of TRAFTAC and its degradation mechanism. Heterobifunctional TRAFTAC consists of an RNA moiety recruits the dCas9-HT7 engineered fusion protein, while the dsDNA sequence binds to the transcription factor of interest (TOI). In the presence of HaloPROTAC, TRAFTAC recruits the E3 ligase complex to the vicinity of the TOI, leading to its degradation by mediating proximity-induced ubiquitination. (B) Diagram illustrating the design strategies and chemical structures of representative TF-PROTACs and RNA-PROTAC.

Figure 4

Schematic representation of the mechanism-of-action and representative structures of aptamer-conjugated and aptamer-constructed PROTACs. (A) Degradation of POI or nucleolin triggered by aptamer-conjugated or aptamer-constructed PROTACs. (B) Representative structures of aptamer-conjugated and aptamer-constructed PROTACs.

Figure 5

Chemical structures and design rationale of G4-PROTACs, TeloTAC and Bivalent nucleic PROTAC.

Figure 6

Schematic illustration of the mechanism of attenuation and critical genome of PROTAC virus. (A) Schematic illustration of the PROTAC virus production system. PROTAC viruses replicate efficiently in TEVp-expressing stable cells, which cleaves the PTD. Whereas in conventional cells, deficient protein synthesis and attenuation of replication are triggered by proteasome-mediated viral protein destabilization, leading to live attenuated PROTAC viruses. Ub, ubiquitin; M1, matrix protein; PTD, proteasome-targeting domain. (B) Diagrammatic representation of the PTD genome.

Figure 7

Summary of Endocytosis-based lysosomal degraders. The construction of LYTAC, BAC, DENTAC and KineTAC is summarized in the left column. LYTACs and BACs tame CI-M6PR and ASGPR as the lysosome-targeting receptor (LTRs) to achieve the degradation of membrane proteins and extracellular proteins. KineTAC acquires lysosomal targeting properties by binding CXCLR7 and CXCL12, with the ability to degrade membrane targets. DENTAC utilizes SR as a novel kind of LTRs to promote the lysosomal degradation of membrane POI.

Figure 8

Schematic diagram of the design rationale of AUTAC, ATTEC, AUTOTAC and CMA-based degrader. AUTAC, ATTEC and AUTOTAC promote the degradation of POI via the formation of POI-specific autophagosome. CMA-based degrader mimics KFERQ motifs thereby capturing HSP70, facilitating POI degradation via the chaperone-mediate process.

Figure 9

Chemical structures and construction considerations of autophagy-based degraders.

Figure 10

Summary of non-small bifunctional chimeras with the functions beyond TPD. (A) Chemical structure and design rationale of TF-DUBTAC. (B) RIPR promotes the dephosphorylation of targeted proteins by simultaneous binding with POI and CD45. (C) Design rationale and amino acid sequence of DEPTAC. (D) Bidirectional regulation of O-GlcNAcylation generated by nanobody‒enzyme conjugates. (E) Targeted O-GlcNAcylation based on modular dual-specificity (DS) RNA aptamer. (F) HER2 antibody‒sialidase conjugates induce catalytic degradation of sialoglycans in a tumor-specific manner.

Figure 11

Graphical summary of the mechanism of bifunctional non-small molecule entities to induce degradation and other proximity-mediated effects. (A) The mechanism of the non-small molecule PROTACs. POIs are ubiquitin-tagged via recruitment of the E3 ligase complex and subsequently degraded via the 26S protease. (B) Lysosomal degraders utilizing endocytosis or autophagy processes. (C) Macromolecular entities for triggering proximity-mediated effects to regulate multiple post-translational modifications (PTMs).