Endothelial Cell Biology Unit, Institute of Molecular and Cellular Biology, University of Leeds, Leeds, UK.
Br J Pharmacol. 2012 Jan;165(1):245-59. doi: 10.1111/j.1476-5381.2011.01545.x.
The potent pro-angiogenic growth factors VEGF-A and basic fibroblast growth factor (bFGF) exert their effects by binding VEGF receptor 2 and FGF receptor tyrosine kinases, respectively. Indolinones (e.g. SU5416 and Sutent) and anilinophthalazines (e.g. PTK787) are potent small molecule inhibitors of VEGFR2 and other tyrosine kinases, but their effects on VEGF-A- and bFGF-stimulated endothelial responses are unclear. Here we assess the ability of these compounds to inhibit pro-angiogenic responses through perturbation of receptor activity and endothelial function(s).
We used in silico modelling, in vitro tyrosine kinase assays, biochemistry and microscopy to evaluate the effects of small molecules on receptor tyrosine kinase activation and intracellular signalling. Primary human endothelial cells were used to assess intracellular signalling, cell migration, proliferation and tubulogenesis.
We predicted that the anilinophthalazine PTK787 binds the tyrosine kinase activation loop whereas indolinones are predicted to bind within the hinge region of the split kinase domain. Sutent is a potent inhibitor of both VEGFR2 and FGFR1 tyrosine kinase activity in vitro. The compounds inhibit both ligand-dependent and -independent VEGFR2 trafficking events, are not selective for endothelial cell responses and inhibit both VEGF-A- and bFGF-mediated migration, wound healing and tubulogenesis at low concentrations. CONCLUSIONS AND IMPLICATIONS; We propose that these compounds have novel properties including inhibition of bFGF-mediated endothelial responses and perturbation of VEGFR2 trafficking. Differential inhibitor binding to receptor tyrosine kinases translates into more potent inhibition of bFGF- and VEGF-A-mediated intracellular signalling, cell migration and tubulogenesis. Indolinones and anilinophthalazines thus belong to a class of multi-kinase inhibitors that show clinical efficacy in disease therapy.
血管内皮生长因子 A（VEGF-A）和碱性成纤维细胞生长因子（bFGF）这两种强效的促血管生成生长因子，分别通过与血管内皮生长因子受体 2（VEGFR2）和 FGF 受体酪氨酸激酶结合来发挥作用。吲唑酮（如 SU5416 和 Sutent）和苯胺基萘并嗪（如 PTK787）是 VEGFR2 和其他酪氨酸激酶的强效小分子抑制剂，但它们对 VEGF-A 和 bFGF 刺激的内皮反应的影响尚不清楚。在这里，我们评估了这些化合物通过干扰受体活性和内皮功能来抑制促血管生成反应的能力。
我们使用了计算机模拟、体外酪氨酸激酶测定、生物化学和显微镜技术来评估小分子对受体酪氨酸激酶激活和细胞内信号转导的影响。我们使用原代人内皮细胞来评估细胞内信号转导、细胞迁移、增殖和小管形成。
我们预测苯胺基萘并嗪 PTK787 结合于酪氨酸激酶的激活环，而吲唑酮则预测结合于分裂激酶结构域的铰链区。Sutent 是体外 VEGFR2 和 FGFR1 酪氨酸激酶活性的强效抑制剂。这些化合物抑制配体依赖性和非依赖性 VEGFR2 转运事件，对内皮细胞反应没有选择性，并在低浓度下抑制 VEGF-A 和 bFGF 介导的迁移、伤口愈合和小管形成。
我们提出，这些化合物具有新的特性，包括抑制 bFGF 介导的内皮反应和干扰 VEGFR2 转运。受体酪氨酸激酶的不同抑制剂结合导致 bFGF 和 VEGF-A 介导的细胞内信号转导、细胞迁移和小管形成的抑制作用更强。因此，吲唑酮和苯胺基萘并嗪属于一类多激酶抑制剂，在疾病治疗中具有临床疗效。
