Discovery and characterization of an iminocoumarin scaffold as an inhibitor of MEKK2 (MAP3K2)

Syed Ahmad; Valentine R St Hilaire; Srinivasa R Dandepally; Gary L Johnson; Alfred L Williams; John E Scott

doi:10.1016/j.bbrc.2018.01.027

Discovery and characterization of an iminocoumarin scaffold as an inhibitor of MEKK2 (MAP3K2)

Biochem Biophys Res Commun. 2018 Jan 29;496(1):205-211. doi: 10.1016/j.bbrc.2018.01.027. Epub 2018 Jan 5.

Authors

Syed Ahmad¹, Valentine R St Hilaire¹, Srinivasa R Dandepally¹, Gary L Johnson², Alfred L Williams¹, John E Scott³

Affiliations

¹ Department of Pharmaceutical Sciences, Biomanufacturing Research Institute and Technology Enterprise (BRITE), North Carolina Central University, Durham, NC, USA.
² Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, NC, USA; Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC, USA.
³ Department of Pharmaceutical Sciences, Biomanufacturing Research Institute and Technology Enterprise (BRITE), North Carolina Central University, Durham, NC, USA. Electronic address: jscott@nccu.edu.

Abstract

The kinase MEKK2 (MAP3K2) activates the MEK5/ERK5 cell signaling pathway and may play an important role in tumor growth and metastasis. Thus, MEKK2 may represent a novel kinase target for cancer. In order to identify inhibitors of MEKK2, we screened a library of compounds using a high throughput MEKK2 intrinsic ATPase enzyme assay. We identified two hits with validated structures and confirmed activity in the primary assay (IC₅₀ values = 322 nM and 7.7 μM) and two orthogonal MEKK2 biochemical assays. Compound 1, the more potent hit, was the subject of further investigation. Limited structure-activity relationship (SAR) studies were performed on this iminocoumarin hit which resulted in ≥20-fold more potent analogs (e.g. 8 and 16 nM IC₅₀). Two analogs had improved selectivity in a 50-member kinase profiling panel compared to the hit. These studies suggested that substitutions around the phenoxy ring of this scaffold can impart improved potency and selectivity for MEKK2. Analog Compound 1s (16 nM IC₅₀) was further verified by external testing to inhibit MEKK2 and MEKK3 with similar potencies. Compound 1s displayed activity in cell-based assays in which it inhibited ERK5 pathway activation in cells and inhibited cell migration in a scratch assay. Thus, we have identified a scaffold that has promising potential to be developed into a highly selective and potent inhibitor of MEKK2. Information from these SAR studies provides specific guidance for the future design of MEKK2 inhibitor probes.

Keywords: ADP; ATP; DMSO; ELISA; HTS; Inhibitor; Kinase; MAP3K2; MAPK; MEKK2; MS; SAR; SD; TR-FRET; adenosine diphosphate; adenosine triphosphate; dimethyl sulfoxide; enzyme-linked immunosorbent assay; high throughput screening; mass spectrometry; mitogen-activated protein kinase; standard deviation; structure-activity relationship; time-resolved fluorescence resonance energy transfer.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Cells, Cultured
Coumarins / administration & dosage
Coumarins / chemistry*
Coumarins / metabolism*
Drug Delivery Systems / methods
Drug Discovery
Drug Evaluation, Preclinical / methods
Humans
MAP Kinase Kinase Kinase 2 / antagonists & inhibitors*
MAP Kinase Kinase Kinase 2 / metabolism*
Protein Interaction Mapping / methods*
Protein Kinase Inhibitors / administration & dosage
Protein Kinase Inhibitors / chemistry*
Protein Kinase Inhibitors / metabolism*

Substances

Coumarins
Protein Kinase Inhibitors
MAP Kinase Kinase Kinase 2

Grants and funding

U54 CA156735/CA/NCI NIH HHS/United States