Chemical remodeling of a cellular chaperone to target the active state of mutant KRAS.

The discovery of small-molecule inhibitors requires suitable binding pockets on protein surfaces. Proteins that lack this feature are considered undruggable and require innovative strategies for therapeutic targeting. KRAS is the most frequently activated oncogene in cancer, and the active state of mutant KRAS is such a recalcitrant target. We designed a natural product-inspired small molecule that remodels the surface of cyclophilin A (CYPA) to create a neomorphic interface with high affinity and selectivity for the active state of KRASG12C (in which glycine-12 is mutated to cysteine). The resulting CYPA:drug:KRASG12C tricomplex inactivated oncogenic signaling and led to tumor regressions in multiple human cancer models. This inhibitory strategy can be used to target additional KRAS mutants and other undruggable cancer drivers. Tricomplex inhibitors that selectively target active KRASG12C or multiple RAS mutants are in clinical trials now (NCT05462717 and NCT05379985).

Adaptor-Specific Antibody Fragment Inhibitors for the Intracellular Modulation of p97 (VCP) Protein-Protein Interactions.

Protein-protein interactions (PPIs) form complex networks to drive cellular signaling and cellular functions. Precise modulation of a target PPI helps explain the role of the PPI in cellular events and possesses therapeutic potential. For example, valosin-containing protein (VCP/p97) is a hub protein that interacts with more than 30 adaptor proteins involved in various cellular functions. However, the role of each p97 PPI during the relevant cellular event is underexplored. The development of small-molecule PPI modulators remains challenging due to a lack of grooves and pockets in the relatively large PPI interface and the fact that a common binding groove in p97 binds to multiple adaptors. Here, we report an antibody fragment-based modulator for the PPI between p97 and its adaptor protein NSFL1C (p47). We engineered these antibody modulators by phage display against the p97-interacting domain of p47 and minimizing binding to other p97 adaptors. The selected antibody fragment modulators specifically disrupt the intracellular p97/p47 interaction. The potential of this antibody platform to develop PPI inhibitors in therapeutic applications was demonstrated through the inhibition of Golgi reassembly, which requires the p97/p47 interaction. This study presents a unique approach to modulate specific intracellular PPIs using engineered antibody fragments, demonstrating a method to dissect the function of a PPI within a convoluted PPI network.

Modulating protein-protein interaction networks in protein homeostasis.

Protein-protein interactions (PPIs) occur in complex networks. These networks are highly dependent on cellular context and can be extensively altered in disease states such as cancer and viral infection. In recent years, there has been significant progress in developing inhibitors that target individual PPIs either orthosterically (at the interface) or allosterically. These molecules can now be used as tools to dissect PPI networks. Here, we review recent examples that highlight the use of small molecules and engineered proteins to probe PPIs within the complex networks that regulate protein homeostasis. Researchers have discovered multiple mechanisms to modulate PPIs involved in host/viral interactions, deubiquitinases, the ATPase p97/VCP, and HSP70 chaperones. However, few studies have evaluated the effect of such modulators on the target's network or have compared the biological implications of different modulation strategies. Such studies will have an important impact on next generation therapeutics.

Quantifying the chameleonic properties of macrocycles and other high-molecular-weight drugs.

Key to the pharmaceutical utility of certain macrocyclic drugs is a 'chameleonic' ability to change their conformation to expose polar groups in aqueous solution, but bury them when traversing lipid membranes. Based on analysis of the structures of 20 macrocyclic compounds that are approved oral drugs, we propose that good solubility requires a topological polar surface area (TPSA, in Å(2)) of ≥0.2×molecular weight (MW). Meanwhile, good passive membrane permeability requires a molecular (i.e., 3D) PSA in nonpolar environments of ≤140Å(2). We show that one or other of these limits is almost invariably violated for compounds with MW>600Da, suggesting that some degree of chameleonic behavior is required for most high MW oral drugs.