Upregulation of FoxM1 by MnSOD Overexpression Contributes to Cancer Stem-Like Cell Characteristics in the Lung Cancer H460 Cell Line

Abstract

Manganese superoxide dismutase promotes migration and invasion in lung cancer cells via upregulation of the transcription factor forkhead box M1. Here, we assessed whether upregulation of forkhead box M1 by manganese superoxide dismutase overexpression mediates the acquisition of cancer stem-like cell characteristics in non-small cell lung cancer H460 cells. The second-generation spheroids from H460 cells were used as lung cancer stem-like cells. The levels of manganese superoxide dismutase, forkhead box M1, stemness markers (CD133, CD44, and ALDH1), and transcription factors (Bmi1, Nanog, and Sox2) were analyzed by Western blot. Sphere formation in vitro and carcinogenicity of lung cancer stem-like cells were evaluated by spheroid formation assay and limited dilution xenograft assays. Knockdown or overexpression of manganese superoxide dismutase or/and forkhead box M1 by transduction with short hairpin RNA(shRNA) or complementary DNA were performed for mechanistic studies. We showed that manganese superoxide dismutase and forkhead box M1 amounts as well as the expression levels of stemness markers and transcription factors sphere formation in vitro, and carcinogenicity of lung cancer stem-like cells were higher than in monolayer cells. Lung cancer stem-like cells transduced with manganese superoxide dismutase shRNA or FoxM1 shRNA exhibited decreased sphere formation and lower amounts of stemness markers and transcription factors. Overexpression of manganese superoxide dismutase or FoxM1 in H460 cells resulted in elevated sphere formation rates and protein levels of stemness markers and transcription factors. Meanwhile, manganese superoxide dismutase knockdown or overexpression accordingly altered forkhead box M1 levels. However, forkhead box M1 knockdown or overexpression had no effect on manganese superoxide dismutase levels but inhibited or promoted lung cancer stem-like cell functions. Interestingly, forkhead box M1 overexpression alleviated the inhibitory effects of manganese superoxide dismutase knockdown in lung cancer stem-like cells. In a panel of non-small cell lung cancer cells, including H441, H1299, and H358 cells, compared to the respective monolayer counterparts, the expression levels of manganese superoxide dismutase and forkhead box M1 were elevated in the corresponding spheroids. These findings revealed the role of forkhead box M1 upregulation by manganese superoxide dismutase overexpression in maintaining lung cancer stem-like cell properties. Therefore, inhibition of forkhead box M1 upregulation by manganese superoxide dismutase overexpression may represent an effective therapeutic strategy for non-small cell lung cancer.

Genistein inhibits lung cancer cell stem‑like characteristics by modulating MnSOD and FoxM1 expression

“…MnSOD increases lung tumor invasion by modulating FoxM1 expression (10). It was also recently reported that MnSOD and FoxM1 expression levels were increased in H460-derived LCSLCs (17), and isovitexin reduced hepatic carcinoma stem-like cell carcinogenicity and stemness by modulating MnSOD and FoxM1 expression levels (34). The present study indicated that genistein inhibited LCSLC CSLC characteristics via modulation of MnSOD and FoxM1 expression.…”

“…FoxM1 knockdown did not affect MnSOD expression in lung cancer cells, but upregulated FoxM1 via upregulation of E2F transcription factor 1 and Sp1 transcription factor (10). Similar results were obtained in a study using H460 cells (17). The present study investigated the potential of MnSOD and FoxM1 as drug targets of genistein in LCSLCs.…”

“…The majority of the studies have suggested that MnSOD overexpression suppresses the malignant phenotype of melanoma (14), and pancreatic (15) and colorectal carcinoma (16). In addition, a previous study provided mechanistic evidence demonstrating that the LCSLC properties of the NSCLC H460 cell line were enhanced by Forkhead box protein M1 (FoxM1) activation, which occurred via MnSOD overexpression (17).…”

<p>Modulation of MnSOD and FoxM1 Is Involved in Invasion and EMT Suppression by Isovitexin in Hepatocellular Carcinoma Cells</p>

“… 15 Our laboratory and others demonstrated that elevated FoxM1 by manganese superoxide dismutase (MnSOD) promotes migration and invasion. 16 , 17 Whether and how FoxM1 upregulated by MnSOD induces the migratory and invasive capabilities and EMT phenotype in HCC stem-like cells (HCSLCs), thereby stimulating tumor progression, remains unknown.…”

“… 17 Our recent study demonstrated that MnSOD is overexpressed in lung CSLCs from H460 cells, and confers carcinogenesis and lung CSLC properties through activation of the FoxM1 transcriptional factor. 16 Because FoxM1 contributes to migration, invasion and EMT in various cancers, 7 , 8 , 14 we initially aimed to evaluate whether FoxM1 upregulation by MnSOD overexpression leads to migration, invasion and EMT in HCSLCs.…”