5925

RB1

Ubiquitinlike with plant homeodomain (PHD) and RINGfinger domain 1 (UHRF1) maintains methylation patterns following DNA replication and is expressed at high levels in various types of human cancer. UHRF1 has been identified as a novel oncogene involved in the pathogenesis of hepatocellular carcinoma. Previous studies have demonstrated that inhibition of the expression of UHRF1 suppresses the proliferation of cancer cells. However, the role of UHRF1 in human osteosarcoma has not been investigated. The present study examined the expression levels of UHRF1 and retinoblastoma 1 (Rb1) in human osteosarcoma cell lines by western blot analysis. Stable overexpression of UHRF1 or knockdown of Rb1 was achieved by lentiviral transfection. Subsequently, a Cell Counting Kit-8 assay and a cell invasion assay were performed to detect the biological functions of UHRF1 in vitro. The results of the present study demonstrated that UHRF1 promoted the proliferation of human osteosarcoma cells. The present study also reported that UHRF1 was able to enhance the invasion of osteosarcoma cells in a retinoblastoma 1 (Rb1)dependent manner. UHRF1 promoted invasion in Rb1positive osteosarcoma cells, but not in Saos2 cells with homozygous loss of Rb1. Similarly, knockdown of Rb1 in Rb1positive osteosarcoma cells enhanced levels of invasion and eliminated the regulation of invasion by UHRF1. UHRF1 was found to inhibit the mRNA and protein expression levels of Rb1. Furthermore, deletion of Rb1 was found to suppress the expression of Ecadherin and promote epithelialtomesenchymal transition (EMT). In addition, the overexpression of UHRF1 inhibited the expression of Ecadherin and promoted EMT via the suppression of Rb1. These data demonstrated that UHRF1 promotes osteosarcoma cell invasion by downregulating the expression of Ecadherin and increasing EMT in an Rb1dependent manner.

BACKGROUND: Aberrant microRNA expression has been implicated in metastasis of cancers. MiR-661 accelerates proliferation and invasion of breast cancer and ovarian cancer, while impedes that of glioma. Its role in non small cell lung cancer (NSCLC) and underlying mechanism are worthy elucidation. METHODS: expression of miR-661 was measured with real-time PCR in both NSCLC tissues and cell lines. The effects of miR-661 on migration, invasion and metastasis capacity of NSCLC were evaluated using wound healing, transwell assay and animal models. Dual reporter luciferase assay and complementary experiments were performed to validate RB1 as a direct target of miR-661 for participation in the progression of NSCLC. RESULTS: MiR-661 was upregulated in NSCLC tissues as compared to paired adjacent tissues and associated with shorter overall survival. Furthermore, miR-661 promoted proliferation, migration and metastasis of NSCLC. Then, we identified RB1 as a direct target of miR-661 through which miR-661 affected EMT process and metastasis of NSCLC. RB1 interacted with E2F1 and both could mediate EMT process in NSCLC. CONCLUSION: MiR-661 promotes metastasis of NSCLC through RB/E2F1 signaling and EMT events, thus may serves as a negative prognostic factor and possible target for treatment of NSCLC patient.

Prediction of lung cancer metastasis relies on post-resection assessment of tumor histology, which is a severe limitation since only a minority of lung cancer patients are diagnosed with resectable disease. Therefore, characterization of metastasis-predicting biomarkers in pre-resection small biopsy specimens is urgently needed. Here we report a biomarker consisting of the phosphorylation of the retinoblastoma protein (Rb) on serine 249 combined with elevated p39 expression. This biomarker correlates with epithelial-to-mesenchymal transition traits in non-small cell lung carcinoma (NSCLC) cells. Immunohistochemistry staining of NSCLC tumor microarrays showed that strong phospho-Rb S249 staining positively correlated with tumor grade specifically in the squamous cell carcinoma (SCC) subtype. Strong immunoreactivity for p39 positively correlated with tumor stage, lymph node invasion, and distant metastases, also in SCC. Linear regression analyses showed that the combined scoring for phospho-Rb S249, p39 and E-cadherin in SCC is even more accurate at predicting tumor staging, relative to each score individually. We propose that combined immunohistochemistry staining of NSCLC samples for Rb phosphorylation on S249, p39, and E-cadherin protein expression could aid in the assessment of tumor staging and metastatic potential when tested in small primary tumor biopsies. The intense staining for phospho-Rb S249 that we observed in high grade SCC could also aid in the precise sub-classification of poorly differentiated SCCs.