Linking Phenotype to Kinase: Identification of a Novel Benzoxaborole Hinge-Binding Motif for Kinase Inhibition and Development of High-Potency Rho Kinase Inhibitors

Abstract

Benzoxaboroles are a novel class of drug-like compounds that have been rich sources of novel inhibitors for various enzymes and of new drugs. While examining benzoxaborole activity in phenotypic screens, our attention was attracted by the (aminomethylphenoxy)benzoxaborole family, which potently inhibited Toll-like receptor-stimulated cytokine secretion from leukocytes. After considering their structure-activity relationships and the central role of kinases in leukocyte biology, we performed a kinome-wide screen to investigate the members of the (aminomethylphenoxy)benzoxaborole family. This technique identified Rho-activated kinase (ROCK) as a target. We showed competitive behavior, with respect to ATP, and then determined the ROCK2-drug cocrystal structure. The drug occupies the ATP site in which the oxaborole moiety provides hydrogen bond donors and acceptors to the hinge, and the aminomethyl group interacts with the magnesium/ATP-interacting aspartic acid common to protein kinases. The series exhibits excellent selectivity against most of the kinome, with greater than 15-fold selectivity against the next best member of the AGC protein kinase subfamily. Medicinal chemistry efforts with structure-based design resulted in a compound with a K i of 170 nM. Cellular studies revealed strong enzyme inhibition rank correlation with suppression of intracellular phosphorylation of a ROCK substrate. The biochemical potencies of these compounds also translated to functional activity, causing smooth muscle relaxation in rat aorta and guinea pig trachea. The series exhibited oral availability and one member reduced rat blood pressure, consistent with ROCK's role in smooth muscle contraction. Thus, the benzoxaborole moiety represents a novel hinge-binding kinase scaffold that may have potential for therapeutic use.

RhoA/Rho-Kinase in the Cardiovascular System

“…Currently, many pharmaceutical companies and manufacturers have strong interests in the RhoA/Rho-kinase signaling and the development of its inhibitors. 3,112,125,237 Among them, Akama et al 238 performed a kinome-wide screen to investigate the members of the benzoxaborole family and identified Rho-kinase as a target. They observed a competitive behavior, with respect to ATP, and determined the ROCK2-drug cocrystal structure.…”

“…They observed a competitive behavior, with respect to ATP, and determined the ROCK2-drug cocrystal structure. 238 On the basis of the role of Rho-kinase in disease processes, we found that the target and therapeutic applications for Rho-kinase inhibitors are mainly in the field of cardiovascular diseases. However, our recent study demonstrated a crucial role for Rho-kinase in cardiac development, 13 which may warn against the use of Rho-kinase inhibitors during pregnancy as in the case of inhibitors of the renin-angiotensin system.…”

2015 ATVB Plenary Lecture

“…They showed competitive behavior, with respect to ATP, and determined the ROCK2-drug cocrystal structure. 180 They exhibited oral availability and 1 member reduced rat blood pressure, consistent with ROCK's role in smooth muscle contraction. Thus, the benzoxaborole moiety may possess a novel hinge-binding kinase scaffold that may have potential for therapeutic use.…”

“…179 Among them, Akama et al 180 performed a kinome-wide screen to investigate the members of the benzoxaborole family and identified ROCK as a target. They showed competitive behavior, with respect to ATP, and determined the ROCK2-drug cocrystal structure.…”

“…Thus, the benzoxaborole moiety may possess a novel hinge-binding kinase scaffold that may have potential for therapeutic use. 179,180 On the basis of the role of ROCK in disease processes that include smooth muscle contraction, fibrosis, and inflammation, the target and therapeutic applications for ROCK inhibitors are mainly in the field of cardiovascular diseases, such as VSA, cardiac hypertrophy, and PH. Indeed, the role of the ROCK pathway has been emerging and the indications of ROCK inhibitors have been expanding especially in cardiovascular medicine (Figure 4).…”