Discovery of potent and selective Cdc25 phosphatase inhibitors via rapid assembly and in situ screening of Quinonoid-focused libraries

Abstract

Cell division cycle 25 (Cdc25) phosphatase is an attractive target for drug discovery. The rapid assembly and in situ screening of focused combinatorial fragment libraries using efficient modular reactions is a highly robust strategy for discovering bioactive molecules. In this study, we have utilized miniaturized synthesis to generate several quinonoid-focused libraries, by standard CuAAC reaction and HBTU-based amide coupling chemistry. Then the enzyme inhibition screening afforded some potent and selective Cdc25s inhibitors. Compound M5N36 (Cdc25A: IC₅₀ = 0.15 ± 0.05 μM; Cdc25B: IC₅₀ = 0.19 ± 0.06 μM; Cdc25C: IC₅₀ = 0.06 ± 0.04 μM) exhibited higher inhibitory activity than the initial lead NSC663284 (Cdc25A: IC₅₀ = 0.27 ± 0.02 μM; Cdc25B: IC₅₀ = 0.42 ± 0.01 μM; Cdc25C: IC₅₀ = 0.23 ± 0.01 μM). Moreover, M5N36 displayed about three-fold more potent against Cdc25C than Cdc25A and B, indicating that M5N36 could act as a relatively selective Cdc25C inhibitor. Cell viability evaluation, western blotting and molecular simulations provided a mechanistic understanding of the activity of M5N36. It showed promising anti-growth activity against the MDA-MB-231 cell line and desirable predicted physicochemical properties. Overall, M5N36 was proven to be a promising novel Cdc25C inhibitor.

Keywords: Cancer; Cdc25 phosphatase; Click chemistry; Miniaturized synthesis; Molecular docking.