Focal Adhesion Kinase (FAK) tyrosine 397E mutation restores the vascular leakage defect in endothelium-specific FAK-kinase dead mice.
Focal Adhesion Kinase (FAK) tyrosine 397E mutation restores the vascular leakage defect in endothelium-specific FAK-kinase dead mice.
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DOI：
10.1002/path.4911
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发表时间：
2017-07
期刊：
影响因子：
--
通讯作者：
Hodivala-Dilke K
中科院分区：
文献类型：
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作者：
Alexopoulou AN;Lees DM;Bodrug N;Lechertier T;Fernandez I;D'Amico G;Dukinfield M;Batista S;Tavora B;Serrels B;Hodivala-Dilke K
粘着斑激酶（FAK）抑制剂已被开发为潜在的抗癌药物，并正在进行临床试验。在体外激活FAK激酶结构域触发Y397的自磷酸化，Src激活，以及随后的其他FAK酪氨酸残基的磷酸化。然而，FAK Y397突变如何影响体内肿瘤血管生成中FAK激酶死亡（KD）表型尚不清楚。我们开发了三种Pdgfb‐iCreert‐驱动的内皮细胞（EC）特异性、他莫昔芬诱导型纯合突变小鼠系：FAK野生型（WT）、FAK KD和FAK双突变体（DM），即具有puerodine磷酸模拟物Y397 E突变的KD。这些ECCre+;FAKWT / WT、ECCre+;FAKKD / KD和ECCre+;FAKDM / DM小鼠皮下注射同基因B16 F0黑素瘤细胞。ECCre+;FAKWT / WT和ECCre-;FAKWT / WT对照小鼠之间的肿瘤生长和肿瘤血管功能没有变化。相反，与Cre −同窝小鼠相比，ECCre+;FAKKD / KD和ECCre+;FAKDM / DM小鼠的肿瘤生长和血管密度降低。尽管在任一基因型中灌注血管或周细胞覆盖率的百分比没有变化，但在ECCre+;FAKKD / KD和ECCre+;FAKDM / DM小鼠中肿瘤缺氧升高。此外，虽然ECCre+; FAKKD / KD小鼠显示血管渗漏减少，ECCre+;FAKDM / DM和ECCre−;FAKDM / DM小鼠显示渗漏无差异。从机制上讲，与Cre +;FAKWT / WT细胞相比，Cre+;FAKKD / KD EC中纤连蛋白刺激的Y397自磷酸化减少，已知Src靶点FAK-Y 577、FAK-Y861、FAK-Y 925、桩蛋白-Y118、p130 Cas-Y 410的磷酸化没有变化。Cre+;FAKDM / DM EC显示Src靶磷酸化水平降低，表明Y397 E取代实际上破坏了Src活化。在Cre+FAKDM / DM EC中，Cre+;FAKKD / KD EC中VE-钙粘蛋白-pY 658水平降低，这与ECCre+;FAKDM / DM小鼠中血管渗漏的拯救相对应。我们发现EC特异性FAK激酶活性是肿瘤生长、血管生成和血管通透性所必需的。ECCre+;FAKDM/DM小鼠恢复了在体内ECCre+;FAKKD/KD小鼠中观察到的KD依赖性肿瘤血管渗漏。这项研究为体内FAK信号转导开辟了新的领域。版权所有© 2017作者.病理学杂志由John Wiley & Sons Ltd代表大不列颠和爱尔兰病理学会出版。
Focal adhesion kinase (FAK) inhibitors have been developed as potential anticancer agents and are undergoing clinical trials. In vitro activation of the FAK kinase domain triggers autophosphorylation of Y397, Src activation, and subsequent phosphorylation of other FAK tyrosine residues. However, how FAK Y397 mutations affect FAK kinase‐dead (KD) phenotypes in tumour angiogenesis in vivo is unknown. We developed three Pdgfb‐iCreert‐driven endothelial cell (EC)‐specific, tamoxifen‐inducible homozygous mutant mouse lines: FAK wild‐type (WT), FAK KD, and FAK double mutant (DM), i.e. KD with a putatively phosphomimetic Y397E mutation. These ECCre+;FAKWT / WT, ECCre+;FAKKD / KD and ECCre+;FAKDM / DM mice were injected subcutaneously with syngeneic B16F0 melanoma cells. Tumour growth and tumour blood vessel functions were unchanged between ECCre+;FAKWT / WT and ECCre−;FAKWT / WT control mice. In contrast, tumour growth and vessel density were decreased in ECCre+;FAKKD / KD and ECCre+;FAKDM / DM mice, as compared with Cre − littermates. Despite no change in the percentage of perfused vessels or pericyte coverage in either genotype, tumour hypoxia was elevated in ECCre+;FAKKD / KD and ECCre+;FAKDM / DM mice. Furthermore, although ECCre+;FAKKD / KD mice showed reduced blood vessel leakage, ECCre+;FAKDM / DM and ECCre−;FAKDM / DM mice showed no difference in leakage. Mechanistically, fibronectin‐stimulated Y397 autophosphorylation was reduced in Cre+;FAKKD / KD ECs as compared with Cre+;FAKWT / WT cells, with no change in phosphorylation of the known Src targets FAK‐Y577, FAK‐Y861, FAK‐Y925, paxillin‐Y118, p130Cas‐Y410. Cre+;FAKDM / DM ECs showed decreased Src target phosphorylation levels, suggesting that the Y397E substitution actually disrupted Src activation. Reduced VE‐cadherin‐pY658 levels in Cre+;FAKKD / KD ECs were rescued in Cre+FAKDM / DM ECs, corresponding with the rescue in vessel leakage in the ECCre+;FAKDM / DM mice. We show that EC‐specific FAK kinase activity is required for tumour growth, angiogenesis, and vascular permeability. The ECCre+;FAKDM/DM mice restored the KD‐dependent tumour vascular leakage observed in ECCre+;FAKKD/KD mice in vivo. This study opens new fields in in vivo FAK signalling. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
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影响因子：
11.1
作者：
Maschler, Sabine;Gebeshuber, Christoph A.;Wiedemann, Eva-Maria;Alacakaptan, Memetcan;Schreiber, Martin;Custic, Ivana;Beug, Hartmut
通讯作者：
Beug, Hartmut
影响因子：
8.7
作者：
Balconi, G;Spagnuolo, R;Dejana, E
通讯作者：
Dejana, E
影响因子：
4.8
作者：
Lebrun, P;Baron, V;Van Obberghen, E
通讯作者：
Van Obberghen, E
影响因子：
9.7
作者：
Luo, Ming;Guan, Jun-Lin
通讯作者：
Guan, Jun-Lin
DOI：
10.1158/1078-0432.ccr-14-2021
发表时间：
2016-08-01
期刊：
Clinical cancer research : an official journal of the American Association for Cancer Research
影响因子：
--
作者：
Roy-Luzarraga M;Hodivala-Dilke K
通讯作者：
Hodivala-Dilke K