SCF ubiquitin ligase-targeted therapies

Abstract

The clinical successes of proteasome inhibitors for the treatment of cancer have highlighted the therapeutic potential of targeting this protein degradation system. However, proteasome inhibitors prevent the degradation of numerous proteins, which may cause adverse effects. Increased specificity could be achieved by inhibiting the components of the ubiquitin-proteasome system that target specific subsets of proteins for degradation. F-box proteins are the substrate-targeting subunits of SKP1-CUL1-F-box protein (SCF) ubiquitin ligase complexes. Through the degradation of a plethora of diverse substrates, SCF ubiquitin ligases control a multitude of processes at the cellular and organismal levels, and their dysregulation is implicated in many pathologies. SCF ubiquitin ligases are characterized by their high specificity for substrates, and these ligases therefore represent promising drug targets. However, the potential for therapeutic manipulation of SCF complexes remains an underdeveloped area. This Review explores and discusses potential strategies to target SCF-mediated biological processes to treat human diseases.

Figure 1. Ubiquitin-mediated degradation

Ubiquitin is attached to substrates by the consecutive activities of three enzymes. An E1 enzyme activates ubiquitin in an ATP-dependent reaction. An E2 enzyme subsequently transfers the activated ubiquitin to the substrate that is specifically bound to the E3 substrate selection factor. Polyubiquitylated substrates are targeted to the proteasome, a multisubunit protease, to undergo degradation. Although protein degradation is irreversible, the ubiquitylation signal can be attenuated through the action of de-ubiquitylating enzymes (DUBs). Multiple enzymatic steps within the ubiquitylation process are potentially druggable. As the selectivity factors, E3 ubiquitin ligases represent the most specific point of intervention. In contrast, proteasome inhibitors block the degradation of a large number of substrates.

Figure 2. The modular structure of SCF ubiquitin ligases and points of potential therapeutic intervention

CUL1 acts as a scaffold that brings the E2 enzyme in proximity to the substrate. It binds to SKP1 at its N-terminus, and the E2 enzyme via RBX1 at its C-terminus. The F-box protein functions as the interchangeable component (69 F-box proteins in humans) that interacts with SKP1 via its F-box domain and with its cognate substrate through its specific substrate recognition domain. CUL1 is activated by covalent conjugation with Nedd8. This modification induces a conformational change in CUL1 that facilitates the transfer of ubiquitin from the E2 to the substrate and is required for CRL function. MLN4924, a small molecule inhibitor of the Nedd8-activating enzyme, shows therapeutic potential and has progressed to phase I/II clinical trials. Because it inhibits the activity of all CRLs, it affects the degradation of a large number substrates. Strategies to inhibit SCF ligase function with more selectivity include blocking SCF complex assembly, blocking the interaction between substrate and F-box protein, and inhibiting E2 enzyme binding and/or function.

Figure 3. Strategies to manipulate SCF ubiquitin ligase activity

A) Unperturbed substrate recognition by an F-box protein. B) PROTACs can act to restore the function of a deleted or mutated F-box protein. They are hetero-bivalent molecules that simultaneously bind to the substrate and the SCF ligase, thereby bringing the ligase into proximity with the substrate. C) Small molecules can restore the function of mutated F-box proteins by acting as molecular glue, providing stable interaction surfaces. D) Small molecules can also function to re-purpose substrate receptors, targeting them to other substrates. E) Inhibition of SCF ligases can be accomplished by competitive inhibition of substrate binding or G) SCF complex formation. F) Allosteric inhibitors can bind to the F-box protein at a region remote from the degron site, distorting the substrate recognition site.