Prey for the Proteasome: Targeted Protein Degradation-A Medicinal Chemist's Perspective

Abstract

Targeted protein degradation (TPD), the ability to control a proteins fate by triggering its degradation in a highly selective and effective manner, has created tremendous excitement in chemical biology and drug discovery within the past decades. The TPD field is spearheaded by small molecule induced protein degradation with molecular glues and proteolysis targeting chimeras (PROTACs) paving the way to expand the druggable space and to create a new paradigm in drug discovery. However, besides the therapeutic angle of TPD a plethora of novel techniques to modulate and control protein levels have been developed. This enables chemical biologists to better understand protein function and to discover and verify new therapeutic targets. This Review gives a comprehensive overview of chemical biology techniques inducing TPD. It explains the strengths and weaknesses of these methods in the context of drug discovery and discusses their future potential from a medicinal chemist's perspective.

Keywords: PROTACs; chemical biology; medicinal chemistry; molecular glue; targeted protein degradation.

Figure 1

Native degradation pathways: a) Substrate ubiquitination occurs via a cascade of activation and transfer reactions mediated by the E1, E2 and E3 ubiquitin enzymes. b) Polyubiquitinated proteins are recognized and degraded by the proteasome. Organelles and protein aggregates are removed via autophagy, whereas a double membrane structure encapsulates the substrate and forms the autophagosome. Upon fusion with a lysosome the autophagosome becomes an autolysosome initiating the degradation process. c) Schematic representation of different Cullin RING ubiquitin ligases (CRLs). CRLs consist of a core protein Cullin (CUL), which is regulated by neddylation, and the RING protein ligase RBX1, interacting with the E2 conjugating enzyme. Substrate specificity is achieved via adaptors and substrate receptors such as Cereblon (CRBN) or the von Hippel‐Lindau protein (VHL) as well as the E2 ligases. Ub=Ubiquitin.

Figure 2

Summary of degron mediated degradation. a) Temperature sensitive degrons (tsD), b) Auxin‐inducible degron (AID), c) Destabilization Domain (DD) Degrons, d) Ligand‐induced degron (LID), e) Small molecule‐assisted shutoff (SMASh), f) Light‐activated degrons (LAD). POI=protein of interest; Deg=degron.

Figure 3

Schematic representation of the HaloTag, dTag and Nanobody approach. a) The HaloTag ligand covalently binds to the HaloTag and induces POI degradation via VHL. b) The dTag degrader brings the FKBP^F36V‐tagged POI in close proximity to CRBN which subsequently ubiquitinates the fusion protein. c) The GFP nanobody fused to VHL recognizes the GFP‐POI fusion protein and mediates its removal via the UPS. POI=protein of interest; GFP=green fluorescent protein; Ub=ubiquitin.

Figure 4

Schematic representation of hydrophobic tagging (HyT), molecular glue and PROTAC mode of action. a) The hydrophobic moiety of the HyT partially unfolds the POI resulting in chaperone recognition and subsequent proteasomal degradation. b) Upon binding of the molecular glue to the E3 receptor protein it reshapes the receptor's surface inducing the recognition of neo‐substrates. c) In contrast to molecular glues the PROTAC binds to both, the E3 receptor as well as the POI, bringing the POI in spatial proximity to the E3 and thus enabling its ubiquitination. POI=protein of interest; Ub=ubiquitin.

Figure 5

Structures of protein degrader. a) Structures of the BCL6 degrader BI‐3802 and its close analog BI‐3812 a BCL‐6 inhibitor. b) Structures of the GSPT1 degrading molecular glue and the mouse double minute 2 (MDM2) degrading PROTAC.

Figure 6

Structures of typical E3 ligase binders. a) The VHL binding peptidomimetics are based on the essential hydroxyproline. The linker for PROTAC synthesis can be either attached at the N‐terminus or side‐on (e.g. at the phenyl ring) to the peptidomimetic VHL binder. b) CRBN ligands are typically based on the IMiD scaffolds of thalidomide, pomalidomide and lenalidomide. c) IAP binders comprise peptidomimetics bearing a terminal N‐methylated alanine which is essential for binding. d) For MDM2 PROTACs idasanutlin is a commonly used MDM2 ligand.

Figure 7

Representative PROTAC screening tree.

Figure 8

Schematic overview of PHOTACs, LYTACs and AUTACs. a) PHOTACs can be switched between an inactive and active conformation upon irradiation with different wavelength. b) Photo caged PROTACs are activated after removal of the photocleavable protecting group with light. c) LYTACs are degrading extracellular proteins by inducing endocytosis via the mannose‐6‐phosphate receptor (M6PR) and subsequent lysosomal degradation. d) Compound 1 induces the degradation of MetAP2 (POI) by means of autophagy. e) Schematic representation of AUTAC mode of action. POI=protein of interest.

Figure 9

Schematic representation of TRIM‐Away and bioPROTACs. a) TRIM21 selectively recognizes the Fc‐region of an antibody which results in proteasomal degradation of the POI, the antibody as well as TRIM21. b) bioPROTACs are fused to the E3 receptor proteins and bind to the POI via a peptide or protein recognition domain. Subsequently, the POI gets ubiquitinated and degraded by the proteasome. POI=protein of interest; Ub=Ubiquitin.