全反式维甲酸对结肠癌细胞XIAP相关因子1表达的影响

目的了解全反式维甲酸对结肠癌lovo细胞XIAP相关因子1(XAF1)表达及细胞增殖的影响。方法以不同浓度的ATRA刺激lovo细胞,RT-PCR检测XAF1的mR-NA水平变化,Westernblot分析XAF1蛋白水平的表达;MTT法检测ATRA对lovo细胞生长抑制率。构建含有XAF1启动子序列荧火虫荧光素酶报告质粒,分析ATRA作用对XAF1启动子活性的影响。结果经ATRA作用,lovo细胞mRNA和蛋白的表达水平上调,细胞生长受到抑制。上述效应均具有药物浓度依赖性。报告基因分析结果显示,ATRA使含有XAF1启动子序列荧火虫荧光素酶报告质粒的虫荧光素酶的活性增加。结论ATRA通过促进启动子转录活性上调XAF1的mRNA和蛋白表达水平,并抑制结肠癌细胞的生长。     

Restoration of XAF1 expression induces apoptosis and inhibits tumor growth in gastric cancer

XAF1 (XIAP‐associated factor 1) is a novel XIAP binding protein that can antagonize XIAP and sensitize cells to other cell death triggers. Our previous results have shown that aberrant hypermethylation of the CpG sites in XAF1 promoter is strongly associated with lower expression of XAF1 in gastric cancers. In our study, we investigated the effect of restoration of XAF1 expression on growth of gastric cancers. We found that the restoration of XAF1 expression suppressed anchorage‐dependent and ‐independent growth and increased sensitivity to TRAIL and drug‐induced apoptosis. Stable cell clones expressing XAF1 exhibited delayed tumor initiation in nude mice. Restoration of XAF1 expression mediated by adenovirus vector greatly increased apoptosis in gastric cancer cell lines in a time‐ and dose‐dependent manner and sensitized cancer cells to TRAIL and drugs‐induced apoptosis. Adeno‐XAF1 transduction induced cell cycle G2/M arrest and upregulated the expression of p21 and downregulated the expression of cyclin B1 and cdc2. Notably, adeno‐XAF1 treatment significantly inhibited tumor growth, strongly enhanced the antitumor activity of TRAIL in a gastric cancer xenograft model in vivo, and significantly prolonged the survival time of animals bearing tumor xenografts. Complete eradication of established tumors was achieved on combined treatment with adeno‐XAF1 and TRAIL. Our results document that the restoration of XAF1 inhibits gastric tumorigenesis and tumor growth and that XAF1 is a promising candidate for cancer gene therapy. © 2009 UICC     

Tumor suppressor XIAP‐Associated factor 1 (XAF1) cooperates with tumor necrosis factor‐related apoptosis‐inducing ligand to suppress colon cancer growth and trigger tumor regression

BACKGROUND:

XIAP‐associated factor 1 (XAF1) antagonizes the anticaspase activity of XIAP (X‐linked inhibitor of apoptosis) and functions as a tumor suppressor in colon cancer. The tumor necrosis factor‐related apoptosis‐inducing ligand (TRAIL) is known as a potential anticancer agent. In this study, the synergistic effect of XAF1 and TRAIL on colon cancer growth was investigated.

METHODS:

Adeno‐XAF1 virus was generated and purified. Cell apoptosis was detected by flow‐cytometry and terminal deoxynucleotidyl transferase‐mediated dUTP nick‐end labeling assay. Protein expression of the different genes was determined by Western blot analysis. Tumorigenesis and tumor growth were assessed in subcutaneous nude mouse xenograft experiments.

RESULTS:

Stable overexpression of XAF1‐sensitized colon cancer cells to TRAIL‐induced apoptosis significantly increased the activity of caspase 3, 7, 8, and 9; released cytochrome c; and down‐regulated XIAP, survivin, and c‐IAP‐2. The restoration of XAF1 expression mediated by adenovirus (adeno‐XAF1) directly induced apoptosis, and synergized TRAIL‐induced apoptosis in colon cancer cells. Ex vivo transduction of adeno‐XAF1 suppressed colon cancer formation in vivo. Furthermore, adeno‐XAF1 treatment of mice significantly inhibited tumor growth, strongly enhanced TRAIL‐induced apoptosis and antitumor activity in colon cancer xenograft models in vivo, and markedly prolonged the survival. Notably, the combined treatment with adeno‐XAF1 and TRAIL completely eradicated the established tumors without detectable toxicity in normal tissue.

CONCLUSIONS:

The combined restoration of XAF1 expression and TRAIL treatment may be a potent strategy for colon cancer therapy. Cancer 2010. © 2010 American Cancer Society.     

The XAF1 tumor suppressor induces autophagic cell death via upregulation of Beclin-1 and inhibition of Akt pathway

Autophagy is designated as type II programmed cell death and may confer a tumor-suppressive function. Our previous studies have shown that XIAP-associated factor 1 (XAF1) induced apoptosis and inhibited tumor growth in gastric cancer cells. In this study, we investigated the effect of XAF1 on the induction of autophagy in gastric cancer cells. We found that adenovirus vector-mediated XAF1 (adeno-XAF1) expression markedly induced autophagy, upregulated the level of Beclin-1 and inhibited phospho-Akt and phospho-p70S6K in gastric cancer cells. The downregulation of Beclin 1 or 3-methyladenine treatment suppressed adeno-XAF1-induced autophagy, but significantly enhanced adeno-XAF1-induced apoptosis. A pan-caspase inhibitor prevented adeno-XAF1-induced apoptosis, but significantly increased adeno-XAF1-induced autophagy. Furthermore, adeno-XAF1 induced autophagy in xenograft tumor and inhibited tumor growth. Our results document that adeno-XAF1 induces autophagy through upregulation of Beclin 1 expression and inhibition of Akt/p70S6K pathway, and reveal a new mechanism of XAF1 tumor suppression.