Effects of receptor tyrosine kinase inhibitors on VEGF165 a- and VEGF165 b-stimulated gene transcription in HEK-293 cells expressing human VEGFR2.
Effects of receptor tyrosine kinase inhibitors on VEGF165 a- and VEGF165 b-stimulated gene transcription in HEK-293 cells expressing human VEGFR2.
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DOI：
10.1111/bph.13116
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发表时间：
2015-06
影响因子：
7.3
通讯作者：
Woolard J
中科院分区：
文献类型：
--
作者：
Carter JJ;Wheal AJ;Hill SJ;Woolard J
针对 VEGF 受体 2 (VEGFR2) 的受体酪氨酸激酶抑制剂 (RTKI) 因其减少血管生成的能力而被证明是一种有吸引力的癌症治疗方法。在这里，我们通过研究活 HEK-293 细胞中活化 T 细胞核因子 (NFAT) 报告基因的活性，对 RTKI 和两种 VEGF 剪接变体 VEGF165a 和 VEGF165b 与 VEGFR2 的相互作用进行了定量分析。使用表达人 VEGFR2 和受 NFAT 响应元件调控的萤火虫荧光素酶报告基因的 HEK-293 细胞来定量分析 RTKI 对 VEGF165a 和 VEGF165b 刺激的荧光素酶基因表达的影响。 VEGF165a 在活细胞中产生 NFAT 荧光素酶报告基因的浓度依赖性激活，该激活被四种不同的 RTKI（西地尼布、帕唑帕尼、索拉非尼和凡德他尼）以非竞争性方式抑制。从该分析中获得的每个 RTKI 的效力与使用纯化的 VEGFR2 激酶结构域的结合研究中获得的效力相似。与 VEGF165a 相比，VEGF165b 是 NFAT 荧光素酶反应的低效激动剂。使用激动操作模型对浓度-反应数据进行分析表明，两种 VEGF165 亚型对 VEGFR2 具有相似的亲和力。对完整活细胞中 VEGF165 亚型和 RTKI 与 VEGFR2 相互作用的定量药理学分析，为 VEGF165a 和 VEGF165b 通过该酪氨酸激酶受体激活钙调磷酸酶-NFAT 信号通路的相对亲和力和功效提供了重要见解。
Receptor tyrosine kinase inhibitors (RTKIs) targeted at VEGF receptor 2 (VEGFR2) have proved to be attractive approaches to cancer therapy based on their ability to reduce angiogenesis. Here we have undertaken a quantitative analysis of the interaction of RTKIs and two VEGF splice variants, VEGF165a and VEGF165b, with VEGFR2 by studying nuclear factor of activated T-cells (NFAT) reporter gene activity in live HEK-293 cells. HEK-293 cells expressing the human VEGFR2 and a firefly luciferase reporter gene regulated by an NFAT response element were used for quantitative analysis of the effect of RTKIs on VEGF165a- and VEGF165b-stimulated luciferase gene expression. VEGF165a produced a concentration-dependent activation of the NFAT-luciferase reporter gene in living cells that was inhibited in a non-competitive fashion by four different RTKIs (cediranib, pazopanib, sorafenib and vandetanib). The potency obtained for each RTKI from this analysis was similar to those obtained in binding studies using purified VEGFR2 kinase domains. VEGF165b was a lower-efficacy agonist of the NFAT-luciferase response when compared with VEGF165a. Analysis of the concentration–response data using the operational model of agonism indicated that both VEGF165 isoforms had similar affinity for VEGFR2. Quantitative pharmacological analysis of the interaction of VEGF165 isoforms and RTKIs with VEGFR2 in intact living cells has provided important insights into the relative affinity and efficacy of VEGF165a and VEGF165b for activation of the calcineurin- NFAT signalling pathway by this tyrosine kinase receptor.
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影响因子：
7.3
作者：
Alexander SP;Benson HE;Faccenda E;Pawson AJ;Sharman JL;Spedding M;Peters JA;Harmar AJ;CGTP Collaborators
通讯作者：
CGTP Collaborators
影响因子：
3.3
作者：
Fearnley GW;Odell AF;Latham AM;Mughal NA;Bruns AF;Burgoyne NJ;Homer-Vanniasinkam S;Zachary IC;Hollstein MC;Wheatcroft SB;Ponnambalam S
通讯作者：
Ponnambalam S
影响因子：
11.4
作者：
Macián, F;García-Rodríguez, C;Rao, AJN
通讯作者：
Rao, AJN
影响因子：
14.9
作者：
Voon, DC;Subrata, LS;Daly, JM
通讯作者：
Daly, JM
DOI：
10.1073/pnas.0914318107
发表时间：
2010-02-09
影响因子：
11.1
作者：
Leppanen, Veli-Matti;Prota, Andrea E.;Alitalo, Kari
通讯作者：
Alitalo, Kari