The quinic acid derivative KZ-41 prevents glucose-induced caspase-3 activation in retinal endothelial cells through an IGF-1 receptor dependent mechanism.

Hui He,Jena J Steinle,Jayaprakash Pagadala,Jerome Baudry,Rebecca L Weir,Jordan J Toutounchian,Duane D Miller,Charles R Yates,Rajesh Mohanraj

doi:10.1371/journal.pone.0180808

Abstract

Retinal microaneurysms, an early disease manifestation of diabetic retinopathy, are associated with retinal endothelial cell (REC) death and macular edema. We previously demonstrated that a quinic acid (QA) analog, KZ-41, promoted REC survival by blunting stress-induced p38 MAPK activation. Herein, we sought to expand our understanding of the pro-survival signal transduction pathways actuated by KZ-41. Using human RECs exposed to high glucose (25 mM, 72 hours), we demonstrated that KZ-41 blocks caspase-3 activation by triggering phosphorylation of the PI3K regulatory subunit (p85; Tyr458) and its downstream target Akt (Ser473). Akt signal transduction was accompanied by autophosphorylation of the receptor tyrosine kinase, insulin growth factor-1 receptor (IGF-1R). IGF-1R knockdown using either the tyrosine kinase inhibitor AG1024 or silencing RNA abolished KZ-41’s pro-survival effect. Under high glucose stress, caspase-3 activation correlated with elevated ERK1/2 phosphorylation and decreased insulin receptor substrate-1 (IRS-1) levels. KZ-41 decreased ERK1/2 phosphorylation and reversed the glucose-dependent reduction in IRS-1. To gain insight into the mechanistic basis for IGF-1R activation by KZ-41, we used molecular modeling and docking simulations to explore a possible protein:ligand interaction between the IGF-1R kinase domain and KZ-41. Computational investigations suggest two possible KZ-41 binding sites within the kinase domain: a region with high homology to the insulin receptor contains one potential allosteric binding site, and another potential site on the other side of the kinase domain, near the hinge domain. These data, together with previous proof-of-concept efficacy studies demonstrating KZ-41 mitigates pathologic retinal neovascularization in the murine oxygen-induced retinopathy model, suggests that QA derivatives may offer therapeutic benefit in ischemic retinopathies.

Highlights

Diabetic retinopathy (DR), the most frequently occurring microvascular complication of diabetes, is a leading cause of vision loss
We have discovered a new class of orally bioavailable quinic acid (QA) analogs [13], which counteract p38 MAPK-dependent pro-apoptotic signaling in human retinal endothelial cell (REC) exposed to genotoxic stress including radiation and melphalan [14, 15]
To determine the effect of KZ-41 on glucose-induced apoptosis, cultured REC cells were exposed to HG, NG, or M and either KZ-41 or normal saline

Summary

Introduction

Diabetic retinopathy (DR), the most frequently occurring microvascular complication of diabetes, is a leading cause of vision loss. An early disease manifestation, are associated with retinal endothelial cell (REC) death, capillary dropout, and macular edema [1]. The resultant ischemia triggers hypoxia-induced factor-1 (HIF-1) driven VEGF, eNOS, and ET-1 expression, which are biomarkers of retinal neovascularization (RNV) [2]. Prolonged high glucose exposure inactivates Akt-dependent pro-survival signaling leading to reduced endothelial cell viability [3]. Overexpression of constitutively active Akt mutants rescues endothelial cells from glucose-induced apoptosis [4]. Insulin-dependent IRS-1/2 recruitment of p85, a subunit of phosphatidylinositide 3-kinase (PI3K), and Akt activation are significantly reduced in isolated microvessels from obese rats compared to lean controls. As evidenced by a reduction in IRS-1-dependent Akt activation, is evident in RECs exposed to high glucose [6]

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