Small molecule stabilization of the KSR inactive state antagonizes oncogenic Ras signalling

Abstract

Deregulation of the Ras–mitogen activated protein kinase (MAPK) pathway is an early event in many different cancers and a key driver of resistance to targeted therapies1. Sustained signalling through this pathway is caused most often by mutations in K-Ras, which biochemically favours the stabilization of active RAF signalling complexes2. Kinase suppressor of Ras (KSR) is a MAPK scaffold3–5 that is subject to allosteric regulation through dimerization with RAF6,7. Direct targeting of KSR could have important therapeutic implications for cancer; however, testing this hypothesis has been difficult owing to a lack of small-molecule antagonists of KSR function. Guided by KSR mutations that selectively suppress oncogenic, but not wild-type, Ras signalling, we developed a class of compounds that stabilize a previously unrecognized inactive state of KSR. These compounds, exemplified by APS-2-79, modulate KSR-dependent MAPK signalling by antagonizing RAF heterodimerization as well as the conformational changes required for phosphorylation and activation of KSR-bound MEK (mitogen-activated protein kinase kinase). Furthermore, APS-2-79 increased the potency of several MEK inhibitors specifically within Ras-mutant cell lines by antagonizing release of negative feedback signalling, demonstrating the potential of targeting KSR to improve the efficacy of current MAPK inhibitors. These results reveal conformational switching in KSR as a druggable regulator of oncogenic Ras, and further suggest co-targeting of enzymatic and scaffolding activities within Ras–MAPK signalling complexes as a therapeutic strategy for overcoming Ras-driven cancers.

Advances of small molecule targeting of kinases

“…A small molecule ligand of the JH2 pseudokinase domain of Tyk2, a member of the Janus tyrosine kinase family, was able to lock in a conformation that stabilizes an autoinhibitory interaction with the JH1 catalytic domain, thereby blocking Tyk2-mediated signaling important in autoimmunity [52]. A new class of pseudokinase antagonists was developed that stabilize a previously unrecognized inactive state of the scaffold protein KSR (kinase suppressor of Ras) [53]. In combination with MEK inhibitors, these compounds inhibited growth of Ras mutant cell lines, providing a new therapeutic strategy against Ras-driven cancers.…”

The evolving world of pseudoenzymes: proteins, prejudice and zombies

“…Indeed, based on the direction of pharma and some of the patent literature, it is certain that scientists are attempting to target pseudoenzymes, both knowingly and (thought provokingly) unknowingly. The likely conservation of structural fold, and a reliance on protein–protein interactions for outputs, mean that pseudoenzymes might be targeted therapeutically in essentially the same ways as other enzymes (see, for example, [26–28]). Indeed, in the case of pseudokinases, it was found that promiscuous inhibitors that target the ATP-binding cleft of conventional protein kinases could also bind the equivalent site in pseudokinases despite most of the examined pseudokinases exhibiting no detectable ATP-binding capacity [10].…”

Arresting kinase suppressor of Ras in an inactive state

“…These studies includes the nucleotide exchange blockage [6, 7], Ras association with son of sevenless homologue (SOS) [8–10], and Ras–Raf interaction [11, 12]. In the past, some approaches were used to target Ras via the KSR due to its pseudokinase standing and non-catalytic function, but now it is a more favorable targeted pathway for designing drugs [13, 14]. The RAS–RAF–KSR–MEK1 pathway proteins work in a cascade.…”

“…Recently, Dhawan et al [14] published a paper entitled “Small molecule stabilization of the KSR inactive state antagonizes oncogenic Ras signalling” in Nature journal. They targeted the RAS signaling pathway by interfering in KSR, using small molecules as inhibitors that arrest the KSR–MEK1 in an inactive conformational state.…”

“…They targeted the RAS signaling pathway by interfering in KSR, using small molecules as inhibitors that arrest the KSR–MEK1 in an inactive conformational state. Dhawan et al [14] started their work on the hypothesis that if the KSR–MEK1 interface is disrupted through small molecules that can bind within the adenosine triphosphate (ATP)-binding pocket, these small molecules may disrupt the signaling pathway. They expected that if they used an inhibitor that take the structure of KSR into a similar state in complex with MEK1 and ATP as in the recent crystal structure form, it will not be possible for KSR to regulate RAF and MEK proteins.…”