Administration of Menadione, Vitamin K3, Ameliorates Off-Target Effects on Corneal Epithelial Wound Healing Due to Receptor Tyrosine Kinase Inhibition
Administration of Menadione, Vitamin K3, Ameliorates Off-Target Effects on Corneal Epithelial Wound Healing Due to Receptor Tyrosine Kinase Inhibition
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DOI：
10.1167/iovs.16-19952
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发表时间：
2016-11-01
影响因子：
4.4
通讯作者：
Ceresa, Brian P.
中科院分区：
文献类型：
--
作者：
Rush, Jamie S.;Bingaman, David P.;Ceresa, Brian P.
目的.抗血管生成受体酪氨酸激酶抑制剂（RTKi），3-[（4-溴-2，6-二氟苯基）甲氧基]-5-[4-（1-吡咯烷基）丁基]氨基]羰基]氨基]-4-异噻唑甲酰胺盐酸盐，靶向VEGFR 2（半数最大抑制浓度[IC 50] = 11 nM）;然而，在较高浓度下发生表皮生长因子受体（EGFR）的脱靶抑制。(IC50= 5.8 μ M）。本研究旨在确定局部RTKi治疗对EGF介导的角膜上皮伤口愈合的影响，并开发新的策略以最大限度地减少脱靶EGFR抑制。使用转化的人细胞系（hTCEpi细胞）测量响应于EGF的体外角膜上皮伤口愈合。使用鼠模型评估体内角膜伤口愈合。在这些补充测定中，在不同RTKi浓度的存在下测量伤口愈合。免疫印迹分析用于检查EGFR和VEGFR 2磷酸化和EGFR降解的动力学。Alcohol Blue测定测量VEGFR 2介导的细胞生物学。受体酪氨酸激酶抑制剂暴露在体外和体内引起EGFR介导的角膜上皮伤口愈合的剂量依赖性抑制。纳摩尔浓度的甲萘醌，维生素K3类似物，当与RTKi共同给药时，减缓了EGFR降解，并改善了体外和体内对上皮伤口愈合的抑制作用。甲萘醌不改变RTKi对VEGFR 2磷酸化的IC 50或其对VEGF诱导的视网膜内皮细胞增殖的抑制。抗血管生成RTKi对角膜上皮表现出脱靶效应，可以通过甲萘醌最小化，而不会不利地影响其靶向VEGFR 2阻断。这些数据表明，甲萘醌具有作为局部补充剂用于患有角膜上皮内稳态紊乱的个体的潜力，特别是作为激酶抑制剂的不良副作用。
PURPOSE. The antiangiogenic receptor tyrosine kinase inhibitor (RTKi), 3-[(4-bromo-2,6-difluorophenyl) methoxy]-5-[[[[4-(1-pyrrolidinyl) butyl] amino] carbonyl] amino]-4-isothiazolecarboxamide hydrochloride, targets VEGFR2 (half maximal inhibitory concentration [IC50] = 11 nM); however, off-target inhibition of epidermal growth factor receptor (EGFR) occurs at higher concentrations. (IC50 = 5.8 mu M). This study was designed to determine the effect of topical RTKi treatment on EGF-mediated corneal epithelial wound healing and to develop new strategies to minimize off-target EGFR inhibition.METHODS. In vitro corneal epithelial wound healing was measured in response to EGF using a transformed human cell line (hTCEpi cells). In vivo corneal wound healing was assessed using a murine model. In these complementary assays, wound healing was measured in the presence of varying RTKi concentrations. Immunoblot analysis was used to examine EGFR and VEGFR2 phosphorylation and the kinetics of EGFR degradation. An Alamar Blue assay measured VEGFR2-mediated cell biology.RESULTS. Receptor tyrosine kinase inhibitor exposure caused dose-dependent inhibition of EGFR-mediated corneal epithelial wound healing in vitro and in vivo. Nanomolar concentrations of menadione, a vitamin K3 analog, when coadministered with the RTKi, slowed EGFR degradation and ameliorated the inhibitory effects on epithelial wound healing both in vitro and in vivo. Menadione did not alter the RTKi's IC50 against VEGFR2 phosphorylation or its inhibition of VEGF-induced retinal endothelial cell proliferation.CONCLUSIONS. An antiangiogenic RTKi exhibited off-target effects on the corneal epithelium that can be minimized by menadione without deleteriously affecting its on-target VEGFR2 blockade. These data indicate that menadione has potential as a topical supplement for individuals suffering from perturbations in corneal epithelial homeostasis, especially as an untoward side effect of kinase inhibitors.