MMS induced expression changes (Mouse)

Despite the high toxicity, alkylating agents are still at the forefront of several clinical protocols used to treat cancers. In this study, we investigated the mechanisms underlying alkylation damage responses, aiming to identify novel strategies to augment alkylating therapy efficacy. In this pursuit, we compared gene expression profiles of evolutionary distant cell types (D. melanogaster Kc167 cells, mouse embryonic fibroblasts and human cancer cells) in response to the alkylating agent methyl-methanesulfonate (MMS). We found that many responses to alkylation damage are conserved across species independent on their tumor/normal phenotypes. Key amongst these observations was the protective role of NRF2-induced GSH production primarily regulating GSH pools essential for MMS detoxification but also controlling activation of unfolded protein response (UPR) needed for mounting survival responses across species. An interesting finding emerged from a non-conserved mammalian-specific induction of mitogen activated protein kinase (MAPK)-dependent inflammatory responses following alkylation, which was not directly related to cell survival but stimulated the production of a pro-inflammatory, invasive and angiogenic secretome in cancer cells. Appropriate blocking of this inflammatory component blocked the invasive phenotype and angiogenesis in vitro and facilitated a controlled tumor killing by alkylation in vivo through inhibition of alkylation-induced angiogenic response, and induction of tumor healing. Gene expression of four biological replicates of primary C57BL6/J mouse embryonic fibroblasts. Cell were plated and medium was exchanged 24 hours later for media either without (Ctrl) or with MMS (MMS at 40 µg/mL) treatment. RNA was harvested either 0, 1, 8, 24 or 72 hours following initiation of MMS exposure.

尽管烷化剂具有较高毒性，但其仍是多种临床抗癌治疗方案的核心用药。本研究聚焦烷化剂损伤应答的潜在机制，旨在探索可提升烷化治疗疗效的全新策略。为此，我们对比了三类进化亲缘关系较远的细胞类型——黑腹果蝇（D. melanogaster）Kc167细胞、小鼠胚胎成纤维细胞以及人类癌细胞——在烷化剂甲磺酸甲酯（MMS）处理下的基因表达谱。研究发现，诸多烷化损伤应答通路在不同物种间具有保守性，且与细胞的肿瘤/正常表型无关。其中核心发现为：核因子E2相关因子2（NRF2）介导的谷胱甘肽（GSH）合成发挥了关键保护作用——其不仅调控用于MMS解毒的GSH池稳态，还可控制未折叠蛋白反应（UPR）的激活，而该反应是跨物种启动细胞存活应答所必需的。另有一项有趣的非保守发现：烷化处理后仅在哺乳动物中特异性诱导丝裂原活化蛋白激酶（MAPK）依赖的炎症应答，该通路并不直接参与细胞存活调控，却可促进癌细胞分泌促炎、侵袭性及血管生成相关的分泌组。适度阻断该炎症通路，可在体外抑制癌细胞的侵袭表型与血管生成；在体内则可通过抑制烷化诱导的血管生成应答、促进肿瘤愈合，实现对烷化治疗杀伤肿瘤效果的精准调控。本数据集包含原代C57BL/6J小鼠胚胎成纤维细胞的4次生物学重复的基因表达数据。实验流程为：细胞接种培养24小时后更换培养基，分别设置对照组（未添加MMS）与处理组（添加40 µg/mL的MMS）；分别在MMS处理启动后的0、1、8、24及72小时收集细胞RNA。