Mukherjee Bipasha, McEllin Brian, Camacho Cristel V, Tomimatsu Nozomi, Sirasanagandala Shyam, Nannepaga Suraj, Hatanpaa Kimmo J, Mickey Bruce, Madden Christopher, Maher Elizabeth, Boothman David A, Furnari Frank, Cavenee Webster K, Bachoo Robert M, Burma Sandeep
Department of Radiation Oncology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390, USA.
Cancer Res. 2009 May 15;69(10):4252-9. doi: 10.1158/0008-5472.CAN-08-4853. Epub 2009 May 12.
Glioblastoma multiforme (GBM) is the most lethal of brain tumors and is highly resistant to ionizing radiation (IR) and chemotherapy. Here, we report on a molecular mechanism by which a key glioma-specific mutation, epidermal growth factor receptor variant III (EGFRvIII), confers radiation resistance. Using Ink4a/Arf-deficient primary mouse astrocytes, primary astrocytes immortalized by p53/Rb suppression, as well as human U87 glioma cells, we show that EGFRvIII expression enhances clonogenic survival following IR. This enhanced radioresistance is due to accelerated repair of DNA double-strand breaks (DSB), the most lethal lesion inflicted by IR. The EGFR inhibitor gefitinib (Iressa) and the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 attenuate the rate of DSB repair. Importantly, expression of constitutively active, myristylated Akt-1 accelerates repair, implicating the PI3K/Akt-1 pathway in radioresistance. Most notably, EGFRvIII-expressing U87 glioma cells show elevated activation of a key DSB repair enzyme, DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Enhanced radioresistance is abrogated by the DNA-PKcs-specific inhibitor NU7026, and EGFRvIII fails to confer radioresistance in DNA-PKcs-deficient cells. In vivo, orthotopic U87-EGFRvIII-derived tumors display faster rates of DSB repair following whole-brain radiotherapy compared with U87-derived tumors. Consequently, EGFRvIII-expressing tumors are radioresistant and continue to grow following whole-brain radiotherapy with little effect on overall survival. These in vitro and in vivo data support our hypothesis that EGFRvIII expression promotes DNA-PKcs activation and DSB repair, perhaps as a consequence of hyperactivated PI3K/Akt-1 signaling. Taken together, our results raise the possibility that EGFR and/or DNA-PKcs inhibition concurrent with radiation may be an effective therapeutic strategy for radiosensitizing high-grade gliomas.
多形性胶质母细胞瘤（GBM）是最致命的脑肿瘤，对电离辐射（IR）和化疗具有高度抗性。在此，我们报告一种分子机制，即关键的胶质瘤特异性突变——表皮生长因子受体变体III（EGFRvIII）赋予辐射抗性。利用Ink4a/Arf基因缺失的原代小鼠星形胶质细胞、通过p53/Rb抑制永生化的原代星形胶质细胞以及人U87胶质瘤细胞，我们发现EGFRvIII的表达增强了IR后的克隆形成存活率。这种增强的放射抗性归因于DNA双链断裂（DSB）修复加速，DSB是IR造成的最致命损伤。EGFR抑制剂吉非替尼（易瑞沙）和磷脂酰肌醇3激酶（PI3K）抑制剂LY294002可减弱DSB修复速率。重要的是，组成型活性的肉豆蔻酰化Akt-1的表达加速了修复，这表明PI3K/Akt-1途径与放射抗性有关。最值得注意的是，表达EGFRvIII的U87胶质瘤细胞显示关键的DSB修复酶——DNA依赖性蛋白激酶催化亚基（DNA-PKcs）的活化增强。DNA-PKcs特异性抑制剂NU7026消除了增强的放射抗性，并且EGFRvIII在DNA-PKcs缺陷细胞中无法赋予放射抗性。在体内，与U87来源的肿瘤相比，原位U87-EGFRvIII来源的肿瘤在全脑放疗后显示出更快的DSB修复速率。因此，表达EGFRvIII的肿瘤具有放射抗性，在全脑放疗后继续生长，对总生存期几乎没有影响。这些体外和体内数据支持我们的假设，即EGFRvIII的表达促进DNA-PKcs活化和DSB修复，这可能是PI3K/Akt-1信号过度激活的结果。综上所述，我们的结果提出了一种可能性，即EGFR和/或DNA-PKcs抑制与放疗同时进行可能是使高级别胶质瘤增敏的有效治疗策略。
