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Background and Objective: The prognosis of advanced squamous cell carcinoma of the larynx is poor Prognostic factors of this disease vary in different studies. This study was to analyze the most important factors affecting the prognosis of the patients with advanced (stage III and IV) squamous cell carcinoma (SCC) of the larynx., Methods: Clinical data of 221 patients with advanced SCC of the larynx were retrospectively analyzed. Survival analysis was performed by the life table method; comparison among/between groups was performed using the log-rank test; and multivariate analysis was carried out using the Cox proportional hazard model., Results: The two- and five-year overall survival rates of the 221 patients were 76.9% and 51.1%; while the 2-and 5-year disease free survival rates were 60.0% and 43.0%. Patients in stage III had better prognosis than those in stage IV. Post-operative radiotherapy improved the survival rate in patients with positive surgical margins. There was no difference in the survival rate between patients underwent partial laryngectomy and those underwent total laryngectomy. Multivariate analyses indicated that age, anatomic type, post-surgical stage, surgical margin and radiotherapy influenced the disease free survival (p<0.05), whereas, age, post-surgical stage and surgical margin affected the overall survival (p<0.05)., Conclusions: The prognosis of patients with advanced SCC of the larynx receiving surgery is poor. Age, post-surgical stage and surgical margin are the most important factors affecting the overall survival.

Background & Objective: At present, the molecular mechanisms of development and progression of bladder cancer are still poorly understood. This study was to explore the expression and significance of Bmi-1 in bladder cancer, and analyze its correlations to clinicopathologic features and prognosis of bladder cancer., Methods: The expression of Bmi-1 protein in 137 specimens of bladder cancer and 30 specimens of normal bladder tissues was detected by SP immunohistochemistry, and its correlations to clinicopathologic features and prognosis of the patients were analyzed., Results: The positive rate of Bmi-1 protein was significantly higher in bladder cancer than in normal bladder tissues (54.3% vs. 16.7%, P<0.05). The positive rate of Bmi-1 protein was 20.6% in grade G1 bladder cancer, 54.3% in grade G2 bladder cancer, and 78.8% in grade G3 bladder cancer, with significant difference (P<0.05). It was significantly lower in superficial bladder cancer than in invasive bladder cancer (32.5% vs. 81.5%,P<0.05). The expression intensity of Bmi-1 was related to tumor size, classification, TNM stage and prognosis (P<0.05), but not to tumor number and tumor recurrence (P>0.05). The patients were followed up for 44-86 months, with a median of 66 months. The 5-year survival rate was significantly higher in superficial bladder cancer patients than in invasive bladder cancer patients (89.6% vs. 39.2%, P<0.05), and higher in Bmi-1-negative patients than in Bmi-1-positive patients (78.5% vs. 50.8%,P<0.05)., Conclusions: Bmi-1 protein is highly expressed in bladder cancer. Detecting Bmi-1 protein is helpful for diagnosis and prognostic evaluation of bladder cancer.

Background & Objective: DNA double-strand break (DSB) is the main mechanism of tumor cell death after irradiation. Homologous recombination (HR) and DNA nonhomologous end-joining (NHEJ) are two important ways to repair DSB. The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), an essential protein of NHEJ, plays a major role during DSB. This study was to investigate the expression of DNA-PKcs in nasopharyngeal carcinoma (NPC), and analyze its correlation to the clinicopathologic features and prognosis of NPC., Methods: The expression of DNA-PKcs protein in 223 specimens of NPC tissues was detected by immunohistochemistry. The correlation of DNA-PKcs expression to clinicopathologic features and prognosis of NPC were analyzed., Results: The overexpression rate of DNA-PKcs in 223 NPC specimens was 36.8%. The expression of DNA-PKcs had no significant correlations to gender, age, pathological type and N staging of NPC (P>0.05), but had remarkable correlations to TNM, T and M staging (P<0.05). The 5-year overall survival rate was significantly lower in the patients with overexpression of DNA-PKcs than in those with low expression of DNA-PKcs (54.6% vs. 79.4%, P<0.05). T, N, M staging and the expression of DNA-PKcs were independent predictors for the overall survival of NPC (P<0.05)., Conclusions: DNA-PKcs is positively expressed in the majority of NPC tissues. The expression level of DNA-PKcs is an important factor affecting the prognosis of NPC, which could be used as a prognostic predictor for NPC.

Background & Objective: Epidermal growth factor receptor (EGFR) is overexpressed in many tumors, and is correlated to poor prognosis. However, the prognostic value of EGFR expression in nasopharyngeal carcinoma (NPC) is controversial. This study was to investigate the expression of EGFR and its phosphorylated form (pEGFR) in NPC, and explore their correlations to the prognosis., Methods: NPC samples were obtained from 110 NPC patients treated at Cancer Center of Sun Yat-sen University from Jan. 1999 to Dec. 1999. The expression of EGFR and pEGFR in 110 specimens of NPC and 20 specimens of normal nasopharyngeal tissues were detected by immunohistochemistry. The correlations of EGFR and pEGFR expression to clinical characteristics and prognosis of NPC were analyzed by univariate and multivariate analyses., Results: The positive rates of EGFR and pEGFR were significantly higher in NPC than in normal tissues (100% vs. 10.0%, 60.0% vs. 15.0%, both P<0.001). High expression rate of EGFR was significantly higher in stage T3-4 tumors than in stage T1-2 tumors (63.6% vs. 43.2%, P=0.034), but it had no relationship with other clinical parameters. The 5-year metastasis-free survival rate was significantly lower in the patients with high pEGFR expression than in those with low pEGFR expression (72.2% vs. 91.0%, P=0.012). However, multivariate analysis revealed that pEGFR expression was not an independent prognostic factor for metastasis-free survival of NPC patients., Conclusion: Phosphorylation of EGFR is related with metastasis-free survival of NPC patients, suggesting an important role of activation of EGFR in the dissemination of NPC.

Objective: To analyze the global gene expression profile of primary cultured nasopharyngeal carcinoma (NPC) cells using cDNA microarray techniques to screen new candidate genes related to the occurrence and progression of NPC., Methods: A NPC cell line C666 and primary cultured NPC cells from biopsy specimens in 5 cases were analyzed with microarray techniques in comparison with 3 normal nasopharyngeal epithelial (NPE) biopsy specimens. Several differentially expressed genes identified from the microarray results were verified by fluorescence real-time PCR (FQ-PCR) and immunohistochemistry (IHC)., Results: Primary cultured cells of both NPC and NPE were verified by cytokeratin IHC, EBER1 in situ hybridization and EBV-DNA real-time PCR. Compared with NPE cells, a total of 493 genes in at least 4/6 of the samples were identified to be differentially expressed in the primary cultured NPC cells, including 264 up-regulated and 229 down-regulated ones. Several differentially expressed genes according to the microarray results were confirmed by real-time PCR and IHC., Conclusion: cDNA microarray technique provides an effective and accurate means for global gene expression profiling of primary cultured NPC cells to screen the differentially expressed genes, which may serve as an important basis for studying the mechanism, classification and diagnosis of NPC at the molecular level.

Objective: To investigate the association of B cell-specific MLV integration site-1 (Bmi-1) mRNA expression level with the differentiation, metastasis and prognosis of gastric carcinoma., Methods: Tissue specimens were obtained from 42 patients undergoing surgery for gastric carcinomas. Reverse transcriptional PCR (RT-PCR) was performed for amplification of Bmi-1 mRNA from the 42 tumor tissues and matched adjacent normal tissues, and the differential Bmi-1 mRNA expression was analyzed for its association with the clinical manifestations of the patients., Results: Bmi-1 mRNA expression was detected in all the gastric carcinoma tissues and normal tissues adjacent to the carcinoma using fluorescence method, and in 29 cases, Bmi-1 mRNA expression was significantly higher in the tumor tissues than in the adjacent tissues. Expression of Bmi-1 mRNA was highly correlated with tumor size, lymph node metastasis and T classification (P<0.05), but not with the patients' gender, age, or tumor differentiation (P>0.05). The survival rate was much lower in patients positive for Bmi-1 mRNA expression than in those without Bmi-1 mRNA expression., Conclusions: Bmi-1 mRNA expression is correlated to differentiation and metastasis of gastric carcinoma and may facilitate its prognostic evaluation. Bmi-1 may serve as a marker for assessing the progression of gastric carcinoma and provide assistance for clinical therapeutic decisions.

Background & Objective: Bmi-1, a putative oncogene, is a member of the polycomb group genes family. It is widely expressed in many kinds of tumors. This study was to investigate the expression and significance of Bmi-1 in breast cancer., Methods: The expression of Bmi-1 protein in 58 specimens of breast cancer was detected by immunohistochemistry. The correlation of Bmi-1 expression to clinicopathologic features of breast cancer was analyzed., Results: The intensive positive rate of Bmi-1 in breast cancer was 82.8%. The intensive expression of Bmi-1 was positively correlated to clinical stage and lymph node metastasis (P < 0.05), but not to tumor size, estrogen receptor (ER), progesterone receptor (PR), c-erbB-2 and vascular endothelial growth factor (VEGF) expression (P>0.05)., Conclusions: The intensive expression of Bmi-1 in breast cancer is related to tumor progression. Bmi-1 may serve as a new molecular marker of metastasis of breast cancer.

Objective: To synthesize a novel gadolinium-loaded nanoparticle as a molecular imaging contrast agent for magnetic resonance imaging., Methods: Gadolinium ion was incorporated within a silica nanoparticle. The size of this nanosized particle was determined by using transmission electron microscope. The spin-echo (SE) images of nine nanoparticle dilutions in vitro were obtained from a 1.5 T clinical scanner, compared with gadolinium diethylene triaminepenta acetate (Gd-DTPA). In vivo distribution of nanoparticle in Balb/c nude mice and Balb/c nude mice with nasopharyngeal carcinoma (NPC) xenografted CNE-2 tumors was studied at MR imaging, 30 sec, 5 min, 30 min, 1 h, 2 h, 4 h, and 24 h after intravenous administration., Results: The gadolinium-loaded nanoparticle was short rod-shaped, and approximately 30 to 40 nm in diameter. The value of longitudinal relaxativity (r(1)) of gadolinium nanoparticle was much higher than that of Gd-DTPA. Thirty minutes after injection the gadolinium nanoparticle, the signal intensities of liver, kidney and xenografted tumor increased from 226 +/- 10, 283 +/- 7 and 195 +/- 5 to 352 +/- 12, 328 +/- 10 and 245 +/- 7, respectively. The dynamic MRI scanning showed that gadolinium nanoparticles were mainly distributed in liver after intravenous administration. Strong enhancement was also detected in CNE-2 xenografted tumors., Conclusion: A new gadolinium-loaded nanoparticle with high relaxativity was synthesized successfully, and might serve as a carrier for magnetic resonance molecular imaging.

Objective: To investigate the effect of membrane type-1 matrix metalloproteinase (MTI-MMP) on the invasive potential of breast cancer cell and analyze its mechanisms., Methods: After treatment of breast cancer MDA-MB-453 cell line with concanavalin A ( ConA, 20 microg/ml) for 24 h, MT1-MMP protein was detected in cancer cells by Western analysis and immunocytochemistry. MDA-MB-453 cells were cultured with exogenous latent proMMP-2 and MMP-2 activity was analyzed by gelatin zymography. The invasive potential of the tumor cells was measured with a membrane invasion culture system. Cancer cells of the cell line were divided into four groups: the control group treated by neither reagent, group ConA was only treated by ConA, group MMP-2 was treated only by MMP-2, and group ConA + MMP-2 was treated by both ConA and MMP-2. RESULTS The expression of MTI-MMP protein could be detected in groups ConA and ConA + MMP-2, but nothing was detected in control and group MMP-2. There was only 72 000 precursor form of MMP-2 in group MMP-2 and there were both 72 000 precursor form and 64 000 active enzyme form of MMP-2 in group ConA + MMP-2, but there was no forms of MMP-2 in the other two groups detected by gelatin zymography. The largest amount of cells penetrated through Matrigel was observed in group ConA + MMP-2 than in the other three groups., Conclusion: MTI-MMP can remarkably promote the invasive potential of breast cancer cells mainly through its ability of activating latent proMMP-2 to degrade

Background & Objective: Epstein-Barr virus (EBV) in nasopharyngeal carcinoma (NPC) cells expresses Epstein-Barr nuclear antigen 1 (EBNA1), latent membrane protein 1 (LMP1), and LMP2. LMP2 is an ideal target for immunotherapy because LMP2 mRNA is detected in 100% nasopharyngeal carcinoma cells and LMP2 protein shows stronger immunogenity than the rest 2 viral proteins. This study was to analyze the sequence of CTL epitopes in the transmembrane region of LMP2 to optimize LMP2-targeted immunotherapy., Methods: Genomic DNA was extracted from 20 biopsies of NPC and 3 biopsies of normal nasopharynx from Cantonese. The transmembrane region of LMP2 gene was amplified with hemi-nest polymerase chain reaction (PCR), and then sequenced directly., Results: As compared with prototype B95.8 cells, the transmembrane region of LMP2 gene, derived from Cantonese NPC and normal nasopharynx tissues, had 14 base pair substitutions, resulting in 6 amino acid substitutions. Among these substitutions, 3 changed amino acids were located in 4 HLA-restricted CTL epitopes (SSC, TYG, CLG, and VMS). Among these polymorphisms, the VMS variation was first identified. The sequence changes of the LMP2 derived from NPC was the same as those of the LMP2 from normal nasopharynx, indicating that those variations were due to geographic-associated polymorphisms rather than NPC-associated mutations., Conclusion: Polymorphisms of LMP2 exist in EBV derived from Cantonese, resulting in 4 CTL epitope variations, which implicates that the effect of LMP2 polymorphisms should be considered when LMP2-targeted vaccine is designed for immunotherapy.

Objective: To investigate the patterns of expression of key cell cycle proteins targeting on the human papillomavirus (HPV) sites E6 and E7 in cervical carcinoma., Methods: Semiquantitative immunohistochemistry was used to determine the expression of the cell cycle regulatory factor p53, retinoblastoma gene product pRb, cyclin-dependent kinase inhibitors p21(CIP1/WAF1) (p21), p16(INK4A) (p21), and p27(KIP1) (p21), and cyclin D1 targeting on the HPV sites E6 and E7 in 73 HPV 16-positive specimens of cervical squamous cell carcinoma, 35 from Australia patients and 38 age, lymph node status, and grade of tumor-matched Chinese patients., Results: There were no significant differences in age, lymph node status, and grade of tumor between the Chinese and Australian groups, however, the number of the patients in advanced stage (>Stage IIa) was greater in the Chinese group than in the Australian group (19:7). The positive rate of p53 in the Australian group was 3%, significantly lower than that in the Chinese group (23%, P = 0.028). The upregulation rate of pRb, p21, and p27 in the Australian group were 56%, 63% and 34% respectively, all significantly higher than those in the Chinese group (89%, 97%, and 89% respectively, P = 0.01, P < 0.001, and P < 0.001). There were no significant differences in the upregulation rate of p16 and cyclin D1 expression rate between the Australian group and Chinese group (97% versus 100%, and 3% versus 12%, both P > 0.05). In the combined data of both groups, the positive rate of p53 was 13%; and the upregulation rates of pRb, p16, p21, p27, and cyclin D1 were 74%, 99%, 81%, 63% and 7% respectively., Conclusion: Cervical carcinoma overexpresses pRb, p16, p21, and p27. Tumors from Chinese patients are significantly more likely to be positive in p53 and to have upregulated pRb, p21, and p27 than their Australian counterparts. The molecular pathways to human papillomavirus-induced cervical cancer may be influenced by ethnicity and further investigation is necessary.

Background & Objective: Since three-dimensional culture simulates in vivo microenvironment better than planar culture, the biological character of cells in three-dimensional culture model are more similar with that of living tissues. This study was to establish three-dimensional culture models that represent different stages of nasopharyngeal caicinogenesis, and provide information to elucidate the related molecular events., Methods: Non-tumor nasopharyngeal biopsy specimens were used to culture for normal nasopharyngeal epithelial cells. Early and late passages of normal nasopharyngeal epithelial cell line NPNE2 from nasopharyngeal biopsy specimens, immortalized nasopharyngeal epithelial cell line NP69SV40T, nasopharyngeal carcinoma (NPC) cell line SUNE-1 and its high metastatic subclone 5-8F were cultured in Matrigel to establish three-dimensional models., Results: The cells grew out of non-tumor nasopharyngeal biopsy specimens displayed the morphologic characteristics as epithelia, and could proliferate in vitro for 8-10 passages before senescence. Immunocytochemical staining of cytokeratin revealed that the cells were of epithelial origin. All of the cells, except late passages of NPNE2 cells, could proliferate in the three-dimensional culture system. NPNE2 and NP69SV40T cells mainly developed reticular structures in morphology, and formed few clones with clear and smooth edges as well as tight intercellular junctions. SUNE-1 and 5-8F cells formed clones with irregular morphology, unclear edge, and loose intercellular junctions. In addition, the clones formed by 5-8F cells also developed a lot of pseudopodia, but developed no reticular structure. Late passages of NPNE2 cells formed no clone and reticular structure in the three-dimensional culture., Conclusions: Normal nasopharyngeal epithelial cells can be successfully cultured in vitro from naspharyngeal biopsy specimens. The three-dimensional culture models, established with normal nasopharyngeal epithelial cells, immortalized nasopharyngeal epithelial cells, and NPC cells, may represent the different stages of nasopharyngeal carcinogenesis.