Kardinal C, Konkol B, Lin H, Eulitz M, Schmidt E K, Estrov Z, Talpaz M, Arlinghaus R B, Feller S M
Laboratory of Molecular Oncology, Institut für Medizinishe Strahleukunde und Zellforschung, Universität Würzburg, Germany.
Blood. 2001 Sep 15;98(6):1773-81. doi: 10.1182/blood.v98.6.1773.
Chronic myelogenous leukemia (CML) is commonly characterized by the presence of the p210(Bcr-Abl) oncoprotein. Many downstream effectors of Bcr-Abl have been described, including activation of the Grb2-SoS-Ras-MAP kinase (Erk) pathway. The precise contributions of these signal-transduction proteins in CML blast cells in human patients are not yet well defined. To gain further insight into the importance of Grb2 for CML, peptides that disrupt Grb2-SoS complexes were tested. These high-affinity Grb2-binding peptides (HAGBPs) can autonomously shuttle into cells and function by binding to the N-terminal SH3 domain of Grb2. The HAGBPs were analyzed for their effects on Bcr-Abl-expressing cell lines and freshly isolated CML blast cells from patients. They induced a dramatic decrease in the proliferation of CML cell lines. This was not observed with point-mutated control peptides with abolished Grb2SH3(N) binding. As expected, Grb2-SoS complexes were greatly diminished in the HAGBP-treated cells, and MAP kinase activity was significantly reduced as determined by an activation-specific phospho-MAPK antibody. Furthermore, cell fractions that are enriched for blast cells from CML patients with active disease were also incubated with the Grb2 blocker peptides. The HAGBPs led to a significant proliferation reduction of these cells in the majority of the isolates, but not in all patients' cells. These results show that, in addition to the direct targeting of Bcr-Abl, selective inhibition of Grb2 protein complexes may be a therapeutic option for a significant number of CML patients.
慢性粒细胞白血病（CML）通常以p210（Bcr-Abl）癌蛋白的存在为特征。已经描述了许多Bcr-Abl的下游效应器，包括Grb2-SoS-Ras-MAP激酶（Erk）途径的激活。这些信号转导蛋白在人类患者的CML原始细胞中的精确作用尚未明确界定。为了进一步深入了解Grb2对CML的重要性，测试了破坏Grb2-SoS复合物的肽。这些高亲和力的Grb2结合肽（HAGBPs）可以自主穿梭进入细胞，并通过与Grb2的N端SH3结构域结合发挥作用。分析了HAGBPs对表达Bcr-Abl的细胞系和从患者新鲜分离的CML原始细胞的影响。它们导致CML细胞系的增殖显著降低。对于Grb2SH3（N）结合被消除的点突变对照肽，未观察到这种情况。正如预期的那样，在HAGBP处理的细胞中，Grb2-SoS复合物大大减少，并且通过激活特异性磷酸化-MAPK抗体测定，MAP激酶活性显著降低。此外，来自患有活动性疾病的CML患者的富含原始细胞的细胞组分也与Grb2阻断肽一起孵育。在大多数分离物中，HAGBPs导致这些细胞的增殖显著降低，但并非在所有患者的细胞中都如此。这些结果表明，除了直接靶向Bcr-Abl之外，选择性抑制Grb2蛋白复合物可能是许多CML患者的一种治疗选择。
