Discovery of novel diaryl urea-oxindole hybrids as BRAF kinase inhibitors targeting BRAF and KRAS mutant cancers

In the current study, a novel series of diaryl urea incorporating oxindole moiety was rationally designed as type II BRaf inhibitors targeting BRaf and KRAS mutant cancers. Molecular hybridization between the diaryl urea scaffold which binds to the inactive conformation of protein kinases on one side and the oxindole core which exhibit adenine mimic properties to be settled in the hinge region on the other side was performed. Studying the antiproliferative activity of the synthesized candidates 9a-t on NCI Cancer cell lines showed that they exhibit potent and broad spectrum of antiproliferative activity on the tested Cancer cell lines with compounds 9c, 9p, 9q, 9s, and 9t demonstrating potent GI₅₀ reaching 0.01 µM. Noteworthy, compound 9s demonstrated a potent GI₅₀ on cell lines expressing mutant KRAS and those express BRaf_V600E with GI₅₀ ranges of 1.79 and 7.94 µM and 1.68 to 2.0 µM, respectively. Further analysis on A375 and Mel501 cell lines expressing BRaf_V600E revealed that compound 9s has a potent growth inhibitory activity with IC₅₀ of 0.7 and 1.5 µM, respectively, in reference to sorafenib (IC₅₀ = 8.7 and 0.3 µM, respectively). Additionally, nearly all the target candidates did not show any cytotoxic effect on the normal fibroblast cell line BJ-1 with compound 9s showing IC₅₀ of 20.2 µM in reference to sorafenib (IC₅₀ = 6.1 µM). Further cellular assays on A375 cell line, revealed the ability of compound 9s to halt the cell cycle progression at the G2 phase besides its ability to induce Apoptosis. In parallel, all the synthesized candidates 9a-t were biochemically evaluated for their inhibitory activity on BRaf_WT and compounds 9b, 9c, and 9n revealed a sub-micromolar IC₅₀ of 0.11, 0.84 and 0.80 µM, respectively. Further investigation of selected compounds on BRaf_V600E showed that compounds 9c, 9n, 9s, and 9t exhibit a sub-micromolar IC₅₀ range of 0.17 to 0.89 µM. Noteworthy, the examined candidates demonstrated a higher selectively towards BRaf_V600E over BRaf_WT highlighting their promising optimization for treating BRaf_V600E expressing cancers. Molecular docking and molecular dynamics simulations in the inactive DFG-out kinase domain of BRaf_WT/_V600E protein kinases confirmed the planned design strategy.

Anticancer; BRAF; Diaryl Urea − Oxindole.