Approach for targeting Ras with small molecules that activate SOS-mediated nucleotide exchange

Abstract

Aberrant activation of the small GTPase Ras by oncogenic mutation or constitutively active upstream receptor tyrosine kinases results in the deregulation of cellular signals governing growth and survival in ∼30% of all human cancers. However, the discovery of potent inhibitors of Ras has been difficult to achieve. Here, we report the identification of small molecules that bind to a unique pocket on the Ras:Son of Sevenless (SOS):Ras complex, increase the rate of SOS-catalyzed nucleotide exchange in vitro, and modulate Ras signaling pathways in cells. X-ray crystallography of Ras:SOS:Ras in complex with these molecules reveals that the compounds bind in a hydrophobic pocket in the CDC25 domain of SOS adjacent to the Switch II region of Ras. The structureactivity relationships exhibited by these compounds can be rationalized on the basis of multiple X-ray cocrystal structures. Mutational analyses confirmed the functional relevance of this binding site and showed it to be essential for compound activity. These molecules increase Ras-GTP levels and disrupt MAPK and PI3K signaling in cells at low micromolar concentrations. These small molecules represent tools to study the acute activation of Ras and highlight a pocket on SOS that may be exploited to modulate Ras signaling.

Small Molecule Binding Sites on the Ras:SOS Complex Can Be Exploited for Inhibition of Ras Activation

“…Binding site A corresponds to the site recently reported by Burns, et al 11 The three X-ray crystal structures they describe (4NYI, 4NYJ, 4NYM) show minimal side chain movements in site A compared with the unbound structure of SOS, and correspond most closely with our structures of 1 and 2 in complex with HRas:SOS (Figs. 2a & 2b).…”

“…10 Compounds have recently been reported by Burns et al which bind to the Ras:SOS complex and increase the rate of SOS-mediated nucleotide exchange. 11 This is the reverse of the desired effect for an oncology therapy but it is notable that a small molecule can affect this pathway.…”

In situ selectivity profiling and crystal structure of SML-8-73-1, an active site inhibitor of oncogenic K-Ras G12C

“…In other words, there is no requirement to achieve one specific inactive conformation; there may be many inactive conformations that can be achieved with a range of different inhibitor scaffolds targeting the GN-binding pocket and surrounding regions that will likely have the same inhibitory effect on Ras signaling. We therefore speculate that although binding of SML to K-Ras G12C places the protein in a conformation that is known to be inactive, a number of other approaches may also achieve the goal of impairing Ras signaling (41).…”

Direct Modulation of Small GTPase Activity and Function

“…However, the potential of this indole scaffold to serve as as tarting point for fragment growing appears to be limited, as compound 19 was also reported to be an activator of SOS-catalyzed nucleotide exchange (EC 50 = 14 mm). [93] Compound 19 binds to ah ydrophobic pocket formed by the CDC25 domain of SOS and increases cellular Ras-GTP levels in ac oncentration-dependent manner,w hereas no effect on the Ras-specific GEF RasGRF-1 was observed. Further investigation of 19 will be required, as it shows ab iphasic response and decreases AKT and ERK activation downstream of Ras through an unknown mode of action, thereby underlining the complexity of the Ras signaling network.…”