Identification of Activators of ERK5 Transcriptional Activity by High-Throughput Screening and the Role of Endothelial ERK5 in Vasoprotective Effects Induced by Statins and Antimalarial Agents

Abstract

Because extracellular signal-regulated kinase 5 (ERK5) inhibits endothelial inflammation and dysfunction, activating ERK5 might be a novel approach to protecting vascular endothelial cells (ECs) against various pathological conditions of the blood vessel. We have identified small molecules that protect ECs via ERK5 activation and determined their contribution to preventing cardiac allograft rejection. Using high throughput screening (HTS), we identified certain statins and anti-malarial agents including chloroquine (CQ), hydroxychloroquine (HCQ), and quinacrine (QC) as strong ERK5 “activators”. Pitavastatin enhanced ERK5 transcriptional activity and Kruppel-like factor-2 (KLF2) expression in cultured human and bovine ECs, but these effects were abolished by the depletion of ERK5. CQ and HCQ up-regulated ERK5 kinase activity and inhibited VCAM-1 expression in an ERK5-dependent but MEK5- and KLF2/4-independent manner. Leukocyte rolling and vascular reactivity were used to evaluate endothelial function in vivo, and we found that EC-specific ERK5 knockout (ERK5-EKO) mice exhibited increased leukocyte rolling and impaired vascular reactivity, which could not be corrected by pitavastatin. The role of endothelial ERK5 in acute cardiac allograft rejection was also examined by heterotopic grafting of the heart obtained from either wild type (WT) or ERK5-EKO mice into allomismatched recipient mice. A robust increase in both inflammatory gene expression and CD45-positive cell infiltration into the graft was observed. These tissue rejection responses were inhibited by pitavastatin in WT but not ERK5-EKO hearts. Our study has identified statins and anti-malarial drugs as strong ERK5 activators and shown that ERK5 activation is preventive of endothelial inflammation and dysfunction and acute allograft rejection.

Dipeptidyl Peptidase‐4 Inhibitor Anagliptin Prevents Intracranial Aneurysm Growth by Suppressing Macrophage Infiltration and Activation

“…Krüppel‐like transcription factors KLF2 and KLF4 are downstream targets for statin‐mediated ERK5 activation14; however, we detected no significant increase in KLF2 or KLF4 expression in RAW264.7 cells treated with anagliptin (data not shown), suggesting that the downstream effects of anagliptin‐mediated ERK5 activation in macrophages may be different from those of statins. Previous reports have indicated that ERK5 activation reduces cellular inflammatory responses by inhibiting the NF‐κB pathway 14, 32.…”

“…Statins robustly activate ERK5 in endothelial cells, preventing endothelial dysfunction 14. In macrophages, ERK5 activation by statins enhances efferocytosis and plays a key role in atherosclerotic plaque formation in hyperlipidemic mice 15.…”

The Vitamin E Analog Gamma-Tocotrienol (GT3) and Statins Synergistically Up-Regulate Endothelial Thrombomodulin (TM)

“…Sen-Banerjee et al show that statins enhance KLF2 promoter activity in human endothelial cells via monocyte enhancer factor binding sites and thus induce KLF2 expression [41]. This statin-mediated KLF2 up-regulation in endothelial cells depends on enhancement of ERK5 kinase activity [44]. Others show that statins can induce KLF2 expression in endothelial cells by regulating the apelin/APJ signaling pathway [45].…”

Endothelial Cell Dysfunction and the Pathobiology of Atherosclerosis

“…Activation of MEK5 and/or ERK5 per se leads to endothelial vasoprotection in both a KLF2-dependent and KLF2-independent manner, suggesting that additional targets of these kinases may be involved in mediating the beneficial effects of flow 181, 182 . One such target is the transcription factor Kruppel-like factor 4 (KLF4).…”