Stylianou D C, Auf der Maur A, Kodack D P, Henke R T, Hohn S, Toretsky J A, Riegel A T, Wellstein A
Lombardi Cancer Center, Georgetown University, Washington, DC 20057, USA.
Oncogene. 2009 Sep 17;28(37):3296-306. doi: 10.1038/onc.2009.184. Epub 2009 Jul 27.
The tyrosine kinase receptor anaplastic lymphoma kinase (ALK) and its ligand, the growth factor pleiotrophin (PTN), are highly expressed during the development of the nervous system and have been implicated in the malignant progression of different tumor types. Here, we describe human single-chain variable fragment (scFv) antibodies that target the ligand-binding domain (LBD) in ALK and show the effect in vitro and in vivo. The ALK LBD was used as a bait in a yeast two-hybdrid system to select human scFv from a library with randomized complementarity-determining region 3 domains. Surface plasmon resonance showed high-affinity binding of the selected scFv. The anti-ALK scFv competed for binding of PTN to ALK in intact cells and inhibited PTN-dependent signal transduction through endogenous ALK. Invasion of an intact endothelial cell monolayer by U87MG human glioblastoma cells was inhibited by the anti-ALK scFv. In addition, the growth of established tumor xenografts in mice was reversed after the induction of the conditional expression of the anti-ALK scFv. In archival malignant brain tumors expression levels of ALK and PTN were found elevated and appear correlated with poor patient survival. This suggests a rate-limiting function of the PTN/ALK interaction that may be exploited therapeutically.
酪氨酸激酶受体间变性淋巴瘤激酶（ALK）及其配体生长因子多效蛋白（PTN）在神经系统发育过程中高度表达，并与不同肿瘤类型的恶性进展有关。在此，我们描述了靶向ALK中配体结合域（LBD）的人单链可变片段（scFv）抗体，并展示了其在体外和体内的作用效果。ALK LBD在酵母双杂交系统中用作诱饵，从具有随机互补决定区3结构域的文库中筛选人scFv。表面等离子体共振显示所选scFv具有高亲和力结合。抗ALK scFv在完整细胞中竞争PTN与ALK的结合，并抑制通过内源性ALK的PTN依赖性信号转导。抗ALK scFv抑制了U87MG人胶质母细胞瘤细胞对完整内皮细胞单层的侵袭。此外，在诱导抗ALK scFv的条件表达后，小鼠体内已建立的肿瘤异种移植物的生长得到逆转。在存档的恶性脑肿瘤中，发现ALK和PTN的表达水平升高，且似乎与患者的不良生存相关。这表明PTN/ALK相互作用的限速功能可能在治疗中得到利用。
