OBJECTIVE: To assess the value of m⁷G-lncRNAs in predicting the prognosis and microenvironment of colorectal cancer (CRC). METHODS: We screened m⁷G-lncRNAs from TCGA to construct an m⁷G-lncRNAs risk model using multivariate Cox analysis, which was validated using ROC and C-index curves. Calibration and nomogram were used to predict the prognosis of CRC patients. Point-bar charts and K-M survival curves were used to assess the correlation of risk scores with the patients' clinical staging and prognosis. CIBERSORT and ESTIMATE were used to explore the association between the tumor microenvironment and immune cell infiltration in patients in high and low risk groups and the correlation of risk scores with microsatellite instability, stem cell index and immune checkpoint expression. A protein-protein interaction network was constructed, and the key targets regulated by m⁷G-lncRNAs were identified and validated in paired samples of CRC and adjacent tissues by immunoblotting. RESULTS: We identified a total of 1722 m⁷G-lncRNAs from TCGA database, from which 12 lncRNAs were screened to construct the risk model. The AUCs of the risk model for predicting survival outcomes at 1, 3 and 5 years were 0.727, 0.747 and 0.794, respectively. The AUC of the nomogram for predicting prognosis was 0.794, and the predicted results were consistent with actual survival outcomes of the patients. The patients in the high-risk group showed more advanced tumor stages and a greater likelihood of high microsatellite instability than those in the low-risk group (P < 0.05). The tumor stemness index was negatively correlated with the risk score (r=-0.19; P=7.3e-05). Patients in the high-risk group had higher stromal cell scores (P=0.0028) and higher total scores (P=0.007) with lowered expressions of activated mast cells (r=-0.11; P=0.045) and resting CD4+ T cells (r=-0.14; P=0.01) and increased expressions of most immune checkpoints (P < 0.05). ATXN2 (P= 0.006) and G3BP1 (P=0.007) were identified as the key targets regulated by m⁷G-lncRNAs, and their expressions were both higher in CRC than in adjacent tissues. CONCLUSION The risk model based on 12 m⁷G-lncRNAs has important prognostic value for CRC and can reflect the microenvironment and the efficacy of immunotherapy in the patients.
Biomarkers, Tumor/metabolism* ; Colonic Neoplasms ; DNA Helicases/metabolism* ; Gene Expression Regulation, Neoplastic ; Humans ; Microsatellite Instability ; Poly-ADP-Ribose Binding Proteins/metabolism* ; Prognosis ; RNA Helicases/metabolism* ; RNA Recognition Motif Proteins/metabolism* ; RNA, Long Noncoding/metabolism* ; Tumor Microenvironment

OBJECTIVETo investigate the effect of small interfering RNA（siRNA）for MSI-2 on the growth, apoptosis and NUMB expression of THP-1 cells.

METHODSThree siRNA for MSI-2 gene was designed and transfected into THP- 1 cells. The cell inhibition, colony formation and apoptosis were determined. The protein expression of NUMB, caspase- 3 and PARP were detected by Western blotting.

RESULTSAfter MSI- 2 expression of THP- 1 cells was down- regulated for 24 hours, cell inhibition of siRNA MSI-2 group was（47.89±7.64）%, obviously higher than that of negative control group（P=0.005）. After 9 days, cell colony count of siRNA MSI-2 group was 7.50±1.53, also lower than that of negative control group（35.75±7.46, P<0.001）. In addition, apoptotic rates of siRNA MSI- 2 group at 24 hours ［（15.22±1.52）%］and 48 hours［（33.83±3.96）%］were significantly higher than those of negative control group（P=0.008 and P=0.001, respectively）. Accordingly, activations of caspase-3 and PARP and increased NUMB were observed in siRNA MSI- 2 group.

CONCLUSIONsiRNA for MSI- 2 gene could increase the expressions of NUMB to inhibit the proliferation and induce apoptosis of THP-1 cells.

Apoptosis ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Down-Regulation ; Humans ; Membrane Proteins ; genetics ; metabolism ; Nerve Tissue Proteins ; genetics ; metabolism ; Poly (ADP-Ribose) Polymerase-1 ; Poly(ADP-ribose) Polymerases ; metabolism ; RNA Interference ; RNA, Small Interfering ; RNA-Binding Proteins ; genetics ; metabolism ; Transfection

OBJECTIVETo study the role of poly (ADP-ribose) polymerase-l (PARP-1) in formaldehyde-induced DNA damage response in human bronchial epithelial (HBE) cells and to investigate the mechanism of formaldehyde carcinogenicity.

METHODSThe protein levels were measured by Western blot. The interaction between different proteins was determined by co-immunoprecipitation assay. The chemical inhibitor was used to confirm the relationship between PARP-1 and DNA damage repair.

RESULTSAfter being exposed to different concentrations of formaldehyde for 4 h, HBE cells showed no significant changes in cell viability. Cell viability was significantly reduced after 24-h exposure to 80 and 160 µmol/L formaldehyde (P < 0.05). The 10 µmol/L formaldehyde resulted in significant increases in the protein levels of PARP-1 and XRCC-1. However, 80 µmol/L formaldehyde led to a significant decrease in the protein level of PARP-1 of 124 KD molecular weight but a significant increase in the protein level of PARP-1 of 89 KD molecular weight; there was no significant change in the protein level of XRCC-1. The co-immunoprecipitation assay showed that 10 µmol/L formaldehyde induced increased binding between PARP-1 and XRCC-1, but 80 µmol/L formaldehyde led to no significant change in binding between PARP-1 and XRCC-1. Here, we confirmed the role of 10 µmol/L formaldehyde in strand breaks by comet assay which showed an increase in the tail DNA content of HBE cells after 4-h formaldehyde exposure. No significant difference was observed in tail DNA content between treated HBE cells and control cells at 2 h after formaldehyde was removed. Moreover, compared with control, inhibition of PARP-1 induced a significant increase in tail DNA content, and a significant difference was observed in tail DNA content between inhibited HBE cells and control cells at 2 h after formaldehyde was removed. Inhibition of PARP-1 significantly reduced DNA repair capacity.

CONCLUSIONPARP-1 mediated the repair of DNA damage induced by low-concentration formaldehyde through recruiting XRCC-1 protein, and may be involved in the regulation of cell apoptosis induced by high-concentration formaldehyde.

Apoptosis ; drug effects ; Cells, Cultured ; DNA Damage ; drug effects ; DNA Repair ; drug effects ; DNA-Binding Proteins ; metabolism ; Epithelial Cells ; drug effects ; metabolism ; pathology ; Formaldehyde ; toxicity ; Humans ; Poly (ADP-Ribose) Polymerase-1 ; Poly(ADP-ribose) Polymerases ; metabolism ; X-ray Repair Cross Complementing Protein 1

OBJECTIVE: To explore the effects of 5-Aza-2'-deoxycitydine(5-Aza-dC) and trichostatin A (TSA) on the expression and methylation of CHFR in human laryngeal carcinoma cell line. METHOD: The mRNA expression and promoter hypermethylation and were detected by Realtime fluro-genetic quantitative PCR and methylation specific PCR in Hep-2 cell line, which were cultured in vitro and then treated with different concentrations of 5-Aza-dC and TSA. RESULT: Compared with the control team, 5-Aza-dC alone reactivated expression of the CHFR in Hep-2 cell line (1.75 +/- 0.21). TSA had no effect on gene expression (1.05 +/- 0.13). The combined treatment with 5-Aza-dC and TSA increased gene expression (2.15 +/- 0.18). The cell lines showed a characteristic DNA methylation status. 5-Aza-dC and combined 5-Aza-dC and TSA resulted in demethylation of CHFR. In contrast, TSA alone did not affect the DNA methylation status of CHFR. CONCLUSION Hypermethylation of CHFR gene promoter is a common event in the occurrence and development of laryngeal carcinoma. The promoter aberrant methylation of CHFR is a main cause for down-expression of CHFR. After either treatment with 5-Aza-dC alone or in combination with TSA, the expression of CHFR is up-regulated duo to the reversal methylation. It can be a new idea to the therapy of laryngeal carcinoma.
Azacitidine ; analogs & derivatives ; pharmacology ; Cell Cycle Proteins ; genetics ; metabolism ; DNA Methylation ; drug effects ; Decitabine ; Gene Expression ; drug effects ; Hep G2 Cells ; Humans ; Hydroxamic Acids ; pharmacology ; Laryngeal Neoplasms ; metabolism ; Methylation ; drug effects ; Neoplasm Proteins ; genetics ; metabolism ; Poly-ADP-Ribose Binding Proteins ; Promoter Regions, Genetic ; Ubiquitin-Protein Ligases

OBJECTIVEThe aim of this study was to assess the TOP2A RNA expression and the relationship of TOP2A protein expression with metastasis-free interval in breast cancer patients.

METHODSTOP2A expression was analyzed prior to surgery in 86 patients. The level of TOP2A gene amplification was analyzed by fluorescence in situ hybridization (FISH), its RNA expression level with RT-PCR, and their correlation with TOP2A protein expression was assessed by immunohistochemistry (IHC). The correlation between RNA expression level and metastasis-free interval in breast cancer patients was also analyzed.

RESULTSAberrations (amplification or deletion) of TOP2A copy number was observed in 25.6% (22/86) of the cases. TOP2A protein expression was detected in 66.3% (57/86) of the samples. There was a significant correlation between the TOP2A RNA expression and protein expression (P < 0.001). TOP2A gene expression was significantly associated with the metastasis-free interval in the breast cancer patients (P = 0.001). There was no significant correlation between TOP2A gene amplification and TOP2A protein expression (P = 0.211).

CONCLUSIONSTOP2A RNA level is an objective and reliable prognostic indicator in breast cancer.

Antigens, Neoplasm ; genetics ; metabolism ; Breast Neoplasms ; drug therapy ; genetics ; metabolism ; surgery ; Carcinoma, Ductal, Breast ; drug therapy ; genetics ; metabolism ; surgery ; Carcinoma, Intraductal, Noninfiltrating ; drug therapy ; genetics ; metabolism ; surgery ; Carcinoma, Lobular ; drug therapy ; genetics ; metabolism ; surgery ; Chemotherapy, Adjuvant ; DNA Topoisomerases, Type II ; genetics ; metabolism ; DNA-Binding Proteins ; genetics ; metabolism ; Disease-Free Survival ; Female ; Gene Amplification ; Gene Deletion ; Gene Expression Regulation, Neoplastic ; Humans ; Middle Aged ; Neoadjuvant Therapy ; Poly-ADP-Ribose Binding Proteins ; RNA ; metabolism ; Remission Induction

G3BP (Ras-GTPase-activating protein SH3 domain binding protein), a protein which binds to RasGAP SH3 domain, belongs to RNA-binding protein family, implicating in the downstream of Ras signaling. G3BP harbors the activities of endoribonuclease and DNA helicase, and can induce stress granules formation. G3BP plays a general role in the signal pathways of cell proliferation, differentiation, apoptosis and RNA metabolism. It has been shown to be over-expressed in a number of human malignancies and has a close relationship with tumor invasion and metastasis. Given that it has been implicated in several pathways that are known to be involved in cancer biology, G3BP may provide a new target for cancer therapy.
Animals ; Carrier Proteins ; genetics ; metabolism ; DNA Helicases ; Drug Delivery Systems ; GTPase-Activating Proteins ; therapeutic use ; Humans ; Molecular Sequence Data ; Neoplasms ; drug therapy ; metabolism ; pathology ; Peptide Fragments ; therapeutic use ; Phosphorylation ; Poly-ADP-Ribose Binding Proteins ; RNA Helicases ; RNA Recognition Motif Proteins ; Signal Transduction ; ras GTPase-Activating Proteins ; metabolism ; src Homology Domains ; genetics

OBJECTIVETo detect the correlation between the expression of topoisomerase 2 alpha (TOP2A) and the biological behaviors of human colorectal carcinoma.

METHODSImmunohistochemistry and real-time RT-PCR were used to detect the expression of TOP2A in colorectal carcinomas and normal mucosa.

RESULTSThe protein and mRNA expressions of TOP2A in the metastatic lymph nodes were significantly higher than those in matched primary lesions and normal tissues (P<0.05). No significant difference was found in TOP2A expressions between normal mucosa and colorectal carcinomas. The protein and mRNA expressions of TOP2A were significantly correlated to the lymph node metastasis and invasion depth (P<0.05), but not to the differentiation of the tumor (P>0.05).

CONCLUSIONTOP2A plays an important role in the invasion and metastasis of the colorectal carcinomas, and may serve as a valuable indicator for the diagnosis, treatment and the prognostic evaluation of the malignancy.

Antigens, Neoplasm ; metabolism ; Colorectal Neoplasms ; metabolism ; pathology ; DNA Topoisomerases, Type II ; metabolism ; DNA-Binding Proteins ; metabolism ; Female ; Humans ; Lymphatic Metastasis ; Male ; Poly-ADP-Ribose Binding Proteins

OBJECTIVETo investigate the expressions and significances of Ras-GTPase-activating protein SH 3 domain binding protein(G3BP) and osteopontin (OPN) proteins in esophageal squamous carcinoma (ESC).

METHODSThe expressions of G3BP and OPN proteins in 80 cases of ESC were detected by immunohistochemistry. The relationships between the 2 protein expression and tumor size, differentiation degree, TNM stage, lymph node metastasis and prognosis of ESC were also explored.

RESULTS(1) The positive expression rate of G3BP in ESC was 71.3%, and the rate in lymphoid metastatic group was significantly higher than that in non lymphoid metastatic group (Z=-2.283, P=0.022), but no relations were found between G3BP expression and diameter of tumor, differentiation and TNM grade (P>0.05). The G3BP positive expression group had shorter survival time than G3BP negative expression group (P=0.000). (2) The positive expression rate of OPN in ESC was 100%, and the degree of OPN expression was correlated with the differentiation (chi(2)=10.766, P=0.005) and lymphoid metastasis (Z=-2.289, P=0.022), but no relationship was found between the diameter of tumor and TNM grade (P>0.05). The expression of OPN were significantly related to survivals in a negative time-dependent manner in ESC patients (P=0.000). (3) The expression of G3BP protein correlated positively with the degree of OPN expression in ESC tissue (r(s)=0.376, P=0.001).

CONCLUSIONSThe expressions of G3BP and OPN proteins have a close relationship with lymphoid metastasis and survival in ESC patients. G3BP and OPN proteins can be considered as predictors of prognosis in ESC patients.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Carrier Proteins ; metabolism ; DNA Helicases ; Esophageal Neoplasms ; metabolism ; pathology ; Female ; Follow-Up Studies ; Humans ; Immunohistochemistry ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Osteopontin ; metabolism ; Poly-ADP-Ribose Binding Proteins ; Prognosis ; RNA Helicases ; RNA Recognition Motif Proteins

OBJECTIVETo investigate the changes of topoisomerase IIalpha (TOP2A) and HER2/neu genes in pancreatic ductal adenocarcinomas of Chinese patients, and to determine their roles during carcinogenesis and tumor progression.

METHODSExpressions of TOP2A and HER2/neu proteins were detected by using immunohistochemistry, while gene amplifications of TOP2A and HER2/neu were assessed by using multi-color fluorescence in situ hybridization (FISH). All the samples were of paraffin embedded and 10% formalin fixed tissue, including 26 cases of pancreatic ductal adenocarcinomas with adjacent non-neoplastic pancreatic tissues, 10 cases of chronic panreatitis, and 10 cases of normal pancreas. The correlation between TOP2A and HER2/neu gene status was analyzed.

RESULTSBy immunohistochemistry, the nuclear positive index of TOP2A in pancreatic ductal adenocarcinomas varied from 0.5% to 70%, and the positive rate of HER2/neu in pancreatic ductal adenocarcinomas was 46.2% (12/26). By FISH, 9/10 TOP2A amplified adenocarcinomas showed TOP2A and HER2/neu gene coamplification, while one case with HER2/neu gene amplification adenocarcinoma showed no TOP2A amplification. No expression of TOP2A, HER2/neu proteins and no amplification of TOP2A and HER2/neu gene were detected in adjacent non-neoplastic pancreatic tissues, chronic pancreatitis tissues and normal pancreas. No relationship was found between protein expression and gene amplification of TOP2A and HER2/neu (P > 0.05). TOP2A gene amplification was significantly correlated with HER2/neu gene amplification (P < 0.01).

CONCLUSIONSProtein expression of TOP2A and HER2/neu are not associated with the gene amplification. There is a significant correlation between TOP2A amplification and HER2/neu gene amplification. Co-amplification of TOP2A and HER2/neu may play an important role in the carcinogenesis and progression of pancreatic carcinoma. Evaluation of the status of TOP2A and HER2/neu may be helpful to achieve target therapy of pancreatic carcinoma.

Adenocarcinoma ; genetics ; metabolism ; pathology ; secondary ; Adult ; Aged ; Antigens, Neoplasm ; genetics ; metabolism ; Carcinoma, Pancreatic Ductal ; genetics ; metabolism ; pathology ; secondary ; DNA Topoisomerases, Type II ; genetics ; metabolism ; DNA-Binding Proteins ; genetics ; metabolism ; Female ; Gene Amplification ; Gene Expression Regulation, Neoplastic ; Genes, erbB-2 ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Liver Neoplasms ; metabolism ; secondary ; Lymphatic Metastasis ; Male ; Middle Aged ; Pancreatic Neoplasms ; genetics ; metabolism ; pathology ; Poly-ADP-Ribose Binding Proteins ; Receptor, ErbB-2 ; genetics ; metabolism

This study was purposed to investigate the significance of mitosis checkpoint gene chfr expression in acute leukemia (AL). 2 ml of bone marrow were extracted from each of 46 AL patients and 10 normal donors as control and their mononuclear cells were isolated. Then, their chfr expression was detected by using RT-PCR and immunohistochemistry. Normal control blood samples were also analyzed. The results showed that in 15 out of 28 cases of acute non-lymphocytic leukemia and 13 out of 18 cases of acute lymphocytic leukemia expression of chfr gene mRNA and protein significantly decreased as compared with control. The cytogenetic analysis of patients with a decreased Chfr expression revealed abnormal chromosome. In conclusion, Chfr gene is a leukemia-related gene and may play an important role in leukemia pathogenesis.
Bone Marrow Cells ; metabolism ; Cell Cycle Proteins ; biosynthesis ; genetics ; Humans ; Leukemia, Myeloid, Acute ; genetics ; metabolism ; Mitosis ; Neoplasm Proteins ; biosynthesis ; genetics ; Poly-ADP-Ribose Binding Proteins ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; metabolism ; Ubiquitin-Protein Ligases