Validation of a small molecule inhibitor of PDE6D-RAS interaction with favorable anti-leukemic effects

Abstract

RAS mutations prevalent in high-risk leukemia have been linked to relapse and chemotherapy resistance. Efforts to directly target RAS proteins have been largely unsuccessful. However, since RAS-mediated transformation is dependent on signaling through the RAS-related C3 botulinum toxin substrate (RAC) small GTPase, we hypothesized that targeting RAC may be an effective therapeutic approach in RAS mutated tumors. Here we describe multiple small molecules capable of inhibiting RAC activation in acute lymphoblastic leukemia cell lines. One of these, DW0254, also demonstrates promising anti-leukemic activity in RAS-mutated cells. Using chemical proteomics and biophysical methods, we identified the hydrophobic pocket of phosphodiester 6 subunit delta (PDE6D), a known RAS chaperone, as a target for this compound. Inhibition of RAS localization to the plasma membrane upon DW0254 treatment is associated with RAC inhibition through a phosphatidylinositol-3-kinase/AKT-dependent mechanism. Our findings provide new insights into the importance of PDE6D-mediated transport for RAS-dependent RAC activation and leukemic cell survival.

An Improved PDE6D Inhibitor Combines with Sildenafil To Inhibit KRAS Mutant Cancer Cell Growth

“…44 While some PDE6Di were shown to dislodge K-Ras more or less from the plasma membrane within 60−90 min, 7,15,16,26 only in some cases was evidence for a moderate effect on Ras signaling provided. 16,24,26 Nevertheless, all of these PDE6Di demonstrated cell killing activity in KRAS mutant pancreatic or colorectal cancer cells; however, these are assays that cannot detect off-target activities.…”

“…For instance, farnesyl-transferase inhibitors that block the enzyme-mediating Ras farnesylation are now applied with some success in HRAS mutant head and neck cancers . While some PDE6Di were shown to dislodge K-Ras more or less from the plasma membrane within 60–90 min, ,,, only in some cases was evidence for a moderate effect on Ras signaling provided. ,, Nevertheless, all of these PDE6Di demonstrated cell killing activity in KRAS mutant pancreatic or colorectal cancer cells; however, these are assays that cannot detect off-target activities.…”

“…25 Another PDE6Di emerged from a Rac-inhibitor screen, which led to the oxadiazole DW0254 as a submicromolar active compound (K D = 436 ± 6 nM). 26 This compound inhibited downstream signaling of Ras above 20 μM, and in vivo activity was observed with pretreatment of the transplanted T-cell cancer cell line or subcutaneous implantation of an osmotic pump due to limited solubility.…”

Selected Approaches to Disrupting Protein–Protein Interactions within the MAPK/RAS Pathway

“…Recently, in silico techniques have also been used to identify novel inhibitors of the PDEδ:RAS PPI. In 2022, Williams and coworkers conducted a virtual screening for novel RAC inhibitors followed by profiling these inhibitors for their ability to inhibit growth of leukemic cell lines [ 108 ]. Hit optimization efforts led to the discovery of DW0441 and DW0254 ( 31 and 32 , respectively, Figure 9 E), which were found to have IC 50 s in the submicromolar range when screening the viability of T-cell acute lymphoblastic leukemia cells.…”