Brain Tumor Center, Departments of Neuro-Oncology.
Genomic Medicine, The University of Texas MD Anderson Cancer Center, Houston, Texas.
Neuro Oncol. 2018 Jan 10;20(1):78-91. doi: 10.1093/neuonc/nox128.
Oncogenic activation of phosphatidylinositol-3 kinase (PI3K) signaling plays a pivotal role in the development of glioblastoma (GBM). However, pharmacological inhibition of PI3K has so far not been therapeutically successful due to adaptive resistance through a rapid rewiring of cancer cell signaling. Here we identified that WEE1 is activated after transient exposure to PI3K inhibition and confers resistance to PI3K inhibition in GBM.
Patient-derived glioma-initiating cells and established GBM cells were treated with PI3K inhibitor or WEE1 inhibitor alone or in combination, and cell proliferation was evaluated by CellTiter-Blue assay. Cell apoptosis was analyzed by TUNEL, annexin V staining, and blotting of cleaved caspase-3 and cleaved poly(ADP-ribose) polymerase. Both subcutaneous xenograft and orthotropic xenograft studies were conducted to evaluate the effects of the combination on tumorigenesis; the tumor growth was monitored by bioluminescence imaging, and tumor tissue was analyzed by immunohistochemistry to validate signaling changes.
PI3K inhibition activates WEE1 kinase, which in turn phosphorylates cell division control protein 2 homolog (Cdc2) at Tyr15 and inhibits Cdc2 activity, leading to G2/M arrest in a p53-independent manner. WEE1 inhibition abrogated the G2/M arrest and propelled cells to prematurely enter into mitosis and consequent cell death through mitotic catastrophe and apoptosis. Additionally, combination treatment significantly suppressed tumor growth in a subcutaneous model but not in an intracranial model due to limited blood-brain barrier penetration.
Our findings highlight WEE1 as an adaptive resistant gene activated after PI3K inhibition, and inhibition of WEE1 potentiated the effectiveness of PI3K targeted inhibition, suggesting that a combinational inhibition of WEE1 and PI3K might allow successful targeted therapy in GBM.
磷脂酰肌醇-3 激酶（PI3K）信号的致癌激活在胶质母细胞瘤（GBM）的发展中起着关键作用。然而，由于癌细胞信号的快速重布线导致适应性耐药，PI3K 的药理抑制迄今为止在治疗上并未取得成功。在这里，我们发现 WEE1 在短暂暴露于 PI3K 抑制后被激活，并在 GBM 中赋予对 PI3K 抑制的抗性。
用 PI3K 抑制剂或 WEE1 抑制剂单独或联合处理患者来源的神经胶质瘤起始细胞和建立的 GBM 细胞，并通过 CellTiter-Blue 测定评估细胞增殖。通过 TUNEL、膜联蛋白 V 染色和裂解的 caspase-3 和裂解的多聚（ADP-核糖）聚合酶的印迹分析来分析细胞凋亡。进行皮下异种移植和原位异种移植研究以评估组合对肿瘤发生的影响；通过生物发光成像监测肿瘤生长，并通过免疫组织化学分析验证信号变化。
PI3K 抑制激活 WEE1 激酶，其反过来使细胞分裂控制蛋白 2 同源物（Cdc2）在 Tyr15 处磷酸化并抑制 Cdc2 活性，导致 p53 非依赖性的 G2/M 期阻滞。WEE1 抑制消除了 G2/M 期阻滞，并通过有丝分裂灾难和细胞凋亡促使细胞过早进入有丝分裂和随后的细胞死亡。此外，由于血脑屏障渗透有限，联合治疗在皮下模型中显著抑制肿瘤生长，但在颅内模型中没有抑制。
我们的研究结果强调 WEE1 是 PI3K 抑制后激活的适应性耐药基因，WEE1 抑制增强了 PI3K 靶向抑制的有效性，表明 WEE1 和 PI3K 的联合抑制可能允许在 GBM 中成功进行靶向治疗。
