PROteolysis TArgeting Chimeras (PROTACs) - Past, present and future

Abstract

The majority of currently used therapeutics are small molecule-based and utilize occupancy-driven pharmacology as the mode of action (MOA), in which the protein function is modulated via temporary inhibition. New modalities that operate using alternative MOAs are essential for tapping into the "undruggable" proteome. The PROteolysis Targeting Chimera (PROTAC) technology provides an attractive new approach that utilizes an event-driven MOA. Small molecule-based heterobifunctional PROTACs modulate protein target levels by hijacking the ubiquitin-proteasome system to induce degradation of the target. Here, we address important milestones in the development of the PROTAC technology, as well as emphasize key findings from this previous year and highlight future directions of this promising drug discovery modality.

Figure 1.

Schematic illustration of: a) Occupancy driven pharmacology – a small molecule-based drug, often an inhibitor, modulates protein function employing a non-catalytic MOA. b) Event-driven pharmacology (using PROTAC MOA as an example) – protein function is modulated by induced degradation. The PROTACs initiates a degradation cascade with POI ubiquitination followed by subsequent 26S proteasomal degradation of the POI. c) Schematic illustration of a PROTAC, POI ligand (royal blue) and an E3 ligand (dark blue), linker (black) and examples of PROTACs.

Figure 2.

Time table describing the evolution of PROTACs (2001-2016): above time arrow (left): first time different protein or protein families were targeted using the PROTAC technology. Box: Example of targets that have been explored with the PROTAC technology [–41]. Below time arrow (right): important milestones for the PROTAC technology development.

Figure 3.

a) PROTAC-mediated ternary complex formation and Hook effect as a function of PROTAC concentration. PROTAC compound; POI ligand (royal blue), E3 ligand (dark blue) connected by a linker (black), POI (light blue), E3 (green). b) Top: Positive cooperativity is obtained when stabilizing PPIs are obtained between POI and E3. Bottom: Negative cooperativity is observed when charge repulsion and/or steric clashes is observed between POI and E3.

Figure 4.

a) PROTAC compound; POI ligand (royal blue), E3 ligand (dark blue) connected by a linker (black). PROTAC platform with iterative design, synthesis, test and analyze cycle. b) Schematic illustration of the CRISPR NanoBiT-BET protein system [86]. This system enables PROTAC characterization in live-cells using endogenous protein levels. c) Protein tagging systems can be exploited for target selection and/or validation. The POI is fused to e.g. HT7 [61] or FKBP[87] and PROTAC mediated ternary complex formation results in polyubiquitination of the POI followed by proteasomal degradation. d) System for assessing E3 utility in the PROTAC technology [88]. Schematic illustration of the E3 reporter substrate system. E3 is fused to HT7, chloroalkane PROTAC covalently binds to HT7, and the recruitment of the reporter substrate (FKBP-GFP fusion protein) results in ternary complex, polyubiquitination and subsequent proteasomal degradation of the reporter substrate.