Beltzig Lea, Schwarzenbach Christian, Leukel Petra, Frauenknecht Katrin B M, Sommer Clemens, Tancredi Alessandro, Hegi Monika E, Christmann Markus, Kaina Bernd
Institute of Toxicology, University Medical Center, 55131 Mainz, Germany.
Institute of Neuropathology, University Medical Center, 55131 Mainz, Germany.
Cancers (Basel). 2022 Apr 29;14(9):2233. doi: 10.3390/cancers14092233.
First-line drug in the treatment of glioblastoma, the most severe brain cancer, is temozolomide (TMZ), a DNA-methylating agent that induces the critical damage O6-methylguanine (O6MeG). This lesion is cytotoxic through the generation of mismatch repair-mediated DNA double-strand breaks (DSBs), which trigger apoptotic pathways. Previously, we showed that O6MeG also induces cellular senescence (CSEN). Here, we show that TMZ-induced CSEN is a late response which has similar kinetics to apoptosis, but at a fourfold higher level. CSEN cells show a high amount of DSBs, which are located outside of telomeres, a high level of ROS and oxidized DNA damage (8-oxo-guanine), and sustained activation of the DNA damage response and histone methylation. Despite the presence of DSBs, CSEN cells are capable of repairing radiation-induced DSBs. Glioblastoma cells that acquired resistance to TMZ became simultaneously resistant to TMZ-induced CSEN. Using a Tet-On glioblastoma cell system, we show that upregulation of MGMT immediately after TMZ completely abrogated apoptosis and CSEN, while induction of MGMT long-term (>72 h) after TMZ did not reduce apoptosis and CSEN. Furthermore, upregulation of MGMT in the senescent cell population had no impact on the survival of senescent cells, indicating that O6MeG is required for induction, but not for maintenance of the senescent state. We further show that, in recurrent GBM specimens, a significantly higher level of DSBs and CSEN-associated histone H3K27me3 was observed than in the corresponding primary tumors. Overall, the data indicate that CSEN is a key node induced in GBM following chemotherapy.
治疗最严重的脑癌——胶质母细胞瘤的一线药物是替莫唑胺（TMZ），它是一种DNA甲基化剂，可诱导产生关键损伤O6-甲基鸟嘌呤（O6MeG）。这种损伤通过错配修复介导的DNA双链断裂（DSB）产生细胞毒性，从而触发凋亡途径。此前，我们发现O6MeG也会诱导细胞衰老（CSEN）。在此，我们表明TMZ诱导的CSEN是一种晚期反应，其动力学与凋亡相似，但水平高出四倍。CSEN细胞显示出大量位于端粒之外的DSB、高水平的活性氧（ROS）和氧化DNA损伤（8-氧代鸟嘌呤），以及DNA损伤反应和组蛋白甲基化的持续激活。尽管存在DSB，但CSEN细胞能够修复辐射诱导的DSB。对TMZ产生耐药性的胶质母细胞瘤细胞同时对TMZ诱导的CSEN产生耐药性。使用Tet-On胶质母细胞瘤细胞系统，我们发现TMZ处理后立即上调甲基鸟嘌呤-DNA甲基转移酶（MGMT）可完全消除凋亡和CSEN，而TMZ处理后长期（>72小时）诱导MGMT则不会降低凋亡和CSEN。此外，在衰老细胞群体中上调MGMT对衰老细胞的存活没有影响，这表明O6MeG是诱导衰老所必需的，但不是维持衰老状态所必需的。我们进一步表明，在复发性胶质母细胞瘤标本中，观察到的DSB和与CSEN相关的组蛋白H3K27me3水平明显高于相应的原发性肿瘤。总体而言，数据表明CSEN是化疗后胶质母细胞瘤中诱导产生的关键节点。
