SOD2 在肺腫瘤進展的角色

已知 Manganese superoxide dismutase (SOD2) 能抑制 p53 之轉錄活性，但有研究指出 SOD2 亦會被 p53 所調控。因此 SOD2 在不同癌症扮演不同之角色，但 SOD2 和 p53 之交互作用在肺癌進展之角色至今仍不清楚。本研究在肺癌細胞觀察到 SOD2 與 NKX2-1 的表現呈負相關性，但卻與肺癌細胞之 p53 基因突變與否並無相關性。進一步發現 NKX2-1 可做為 p53 之轉錄因子，直接調控 p53 之轉錄作用。又發現 SOD2 會降低 NKX2-1 之表現而抑制 p53 之轉錄作用，因此解除 p53 抑制 Sp1 結合至 IKKβ 啟動子之能力，進而活化 IKKβ 的轉錄作用。另外，本研究以Immunoprecipitation, GST-pull down 和 EMSA 實驗技術證實 NKX2-1會與 Sp1 蛋白結合，使得 Sp1 無法結合至 IKKβ 的啟動子，而抑制 IKKβ 之轉錄作用，因此 SOD2 會促進 IKKβ 之轉錄作用，而活化 NF-kB 訊號路徑。SOD2 活化之 NF-kB 訊號路徑能促進肺癌細胞在軟瓊酯培養皿之生長 (soft agar growth)、細胞侵襲 (invasion) 和裸鼠異殖腫瘤形成 (xenograft tumor formation) 之能力。另外，本研究探討 SOD2 增加 FOXM1 和 MMP2 之表現，是否會促進肺腺癌進展? Luciferase gene reporter assay和 immunoprecipitation 分析之結果顯示 SOD2 會經由抑制 p53 之表現而促進 E2F1 游離出 Rb-E2F1 complex，以及解除 p53 抑制 SP1 結合至 FOXM1 之啟動子之能力。 SOD2 會經由促進FOXM1 和 MMP2 之表現，而促進肺腺細胞之轉移、侵襲和軟瓊酯培養皿生長之能力。在肺腺癌患者，NKX2-1 mRNA 之表現與 SOD2 蛋白以及 IKKβ mRNA之表現均呈逆相關性。SOD2 和 IKKβ mRNA 之表現都能獨立用於評估肺腺癌患者之overall survival (OS) 以及 relapse free survival (RFS)。因此本研究發現SOD2 經由活化 IKKβ/NF-kB 和 FOXM1/MMP2 路徑而促進肺腺癌腫瘤進展，因此 SOD2 之表現可做為肺腺癌患者之預後因子。

MnSOD (SOD2) has been shown to cause dysfunction of p53 transcriptional activity, while conversely, SOD2 expression is further regulated by p53 to modulate lung tumorigenesis. However, the underlying mechanism remains elusive. In the present study, SOD2 expression in a panel of lung cancer cells was negatively correlated with NKX2-1, but not related with p53 status. We demonstrated that NKX2-1 may act a direct transcription factor to upregulate p53 transcription. Mechanistic studies indicated a decrease in NKX2-1 caused by SOD2 activated IKKβ transcription by de-repression of the binding activity of SP1 to the IKKβ promoter due to suppression of p53 function. In addition, immunoprecipitation, GST-pull down, and EMSA assays demonstrated a direct interaction between NKX2-1 and Sp1, which blocked Sp1-mediated IKKβ transcription. SOD2-mediated NF-κB activation via elevation of IKKβ transcription promoted anchorage-independent soft-agar growth, invasion capability, and xenograft tumor formation. We further investigated the possibility that FOXM1 and MMP2 elevated by SOD2 could be responsible for migration, soft-agar growth and invasion capability in lung adenocarcinoma cells. Western blotting showed that FOXM1 and MMP2 expression was dependent on SOD2 expression. Luciferase gene reporter and chromatin immunoprecipitation assays indicated that SOD2 overexpression in lung cancer cells promoted E2F1 and Sp1 binding to FOXM1 promoter and activated reporter activity due to p53 function suppressed by SOD2 overexpression. SOD2 enhanced the potential for cell migration, invasion, and soft agar growth via the FOXM1-MMP-2 axis. In lung adenocarcinoma patients, the expression of NKX2-1 mRNA was negatively associated with SOD2 immunostaining and with IKKβ mRNA expression levels in lung tumors. SOD2 immunostaining and IKKβ mRNA levels could independently predict overall survival and relapse-free survival in lung adenocarcinoma patients. We therefore demonstrated that SOD2 activates IKKβ/NF-kB and FOXM1-MMP2 axis, which results in lung adenocarcinoma progression and poorer patient outcomes.