Discovery and biological evaluation of N5-substituted 6,7-dioxo-6,7-dihydropteridine derivatives as potent Bruton's tyrosine kinase inhibitors

Abstract

Bruton's tyrosine kinase (BTK) plays a critical role in B cell receptor (BCR)-mediated signaling pathways responsible for the development and function of B cells, which makes it an attractive target for the treatment of many types of B-cell malignancies. Herein, a series of N5-substituted 6,7-dioxo-6,7-dihydropteridine-based, irreversible BTK inhibitors were reported with IC₅₀ values ranging from 1.9 to 236.6 nM in the enzymatic inhibition assay. Compounds 6 and 7 significantly inhibited the proliferation of Ramos cells which overexpress the BTK enzyme, as well as the autophosphorylation of BTK at Tyr223 and the activation of its downstream signaling molecule PLCγ2. Overall, this series of compounds could provide a promising starting point for further development of potent BTK inhibitors for B-cell malignancy treatment.

Fig. 1. Representative structures of reported BTK inhibitors.

Fig. 2. (A) Putative binding pose of compound 1 (purple) against BTK (purple). BTK is shown in a cartoon representation with the inhibitor in a stick representation. Key residues are shown as sticks and the hydrogen bonds are highlighted as black dashed lines. (B) Representation of bad (yellow) and ugly (red) contacts between the N5-substituted methyl group and residues Ala428 and Thr474. Hydrogen atoms involved are displayed for explanation purposes. (C) Putative binding pose of compound 3 (green) against BTK. (D) Superposition of the BTK structure in complex with compound 5 (magenta) and EGFR (blue) in complex with 5 (blue). (E) Putative binding pose of compound 6 (orange) against BTK. (F) Alignment of putative binding poses of compounds 1 and 6.

Fig. 3. Phosphorylation blocking effects of compounds 1, 6 and 7 for BTK Tyr223 and the downstream molecule PLCγ2. Cells were starved for 2 h and treated with indicated concentration of compounds 1, 6, 7 or ibrutinib for 1.5 h and then stimulated by anti-IgM (2.5 μg mL^–1) for 15 min.

Fig. 4. Kinase selectivities of compounds 5, 6 and 7 against 25 kinases. Measurements were conducted at a concentration of 100 nM and the values were determined from the results of two independent tests.