OBJECTIVETo investigate the relationship between HPV-DNA status and p16 protein expression in oropharyngeal squamous cell carcinoma (OSCC) and their clinical significance.

METHODSSixty-six patients with oropharyngeal squamous cell carcinomas treated in the Cancer Hospital of Chinese Academy of Medical Sciences from Jan. 1999 to Dec. 2009 were included in this study. Their formalin-fixed and paraffin-embedded tumor tissue blocks met the eligibility criteria and were used in this study. A "sandwich" technique was used to prepare paraffin sections for HPV-DNA analysis. HPV-DNA was detected using the SPF10 LiPA25 version 1 assay. The expression of p16 protein was detected by immunohistochemistry. The survival rates of patients with different HPV-DNA and p16 protein status were analyzed.

RESULTSHPV-DNA was detected in 11 (16.7%) of all specimens. Expression of p16 protein was detected in 9 of the 11 patients with HPV-positive tumors, and in 12 patients of 55 HPV-negative tumors. The expression of p16 protein was highly correlated with the presence of HPV-DNA (P < 0.001). The tumors were classified into three groups based on the p16 protein expression and HPV-DNA status: group A (9 patients): HPV(+) and p16 protein(+); group B (14 patients): HPV-DNA(+)/p16 protein(-) or HPV-DNA(-)/p16 protein(+); and group C (43 patients): HPV-DNA(-)/p16 protein(-). The 3-year OS rates of these 3 groups were 100%, 77.8% and 42.0% (P = 0.001), and their DSS rates were 100%, 77.8% and 46.4%, respectively(P = 0.004).

CONCLUSIONSIn oropharyngeal squamous cell carcinomas, p16 protein expression is highly correlated with the presence of HPV-DNA, and might be a surrogate marker for HPV-positive OSCC. Combination of p16 protein and HPV-DNA status detection may help to more accurately stratify oropharyngeal carcinomas and predict their prognosis.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Squamous Cell ; genetics ; metabolism ; virology ; Cyclin-Dependent Kinase Inhibitor p16 ; metabolism ; DNA, Viral ; isolation & purification ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Neoplasm Metastasis ; Neoplasm Recurrence, Local ; Oropharyngeal Neoplasms ; genetics ; metabolism ; virology ; Papillomaviridae ; Papillomavirus Infections ; genetics ; metabolism ; Survival Rate

Adenocarcinoma ; genetics ; pathology ; virology ; Carcinoma in Situ ; genetics ; pathology ; virology ; Cervical Intraepithelial Neoplasia ; genetics ; pathology ; virology ; Cervix Uteri ; pathology ; virology ; Cytological Techniques ; DNA, Viral ; analysis ; Female ; Follow-Up Studies ; Humans ; Neoplasm Grading ; Papillomaviridae ; genetics ; isolation & purification ; Papillomavirus Infections ; diagnosis ; Uterine Cervical Neoplasms ; genetics ; pathology ; virology

PURPOSE: Circulating free DNA (cfDNA) in plasma is promising to be a surrogate for tumor tissue DNA. However, not all epidermal growth factor receptor (EGFR) mutations in tumor tissue DNA has been detected in matched cfDNA, at least partly due to inefficient cfDNA extraction method. The purpose of this study was to establish an efficient plasma cfDNA extraction protocol. MATERIALS AND METHODS: The yield of plasma cfDNA extracted by our modified phenol-chloroform (MPC) method from non-small-cell lung cancer (NSCLC) patients was compared with that by QIAamp MinElute Virus Spin kit (Qiagen kit) as control, using the Wilcoxon rank-sum test. TaqMan quantitative polymerase chain reaction (qPCR) assays were used to quantify the plasma cfDNA extracted. Both Mutant-enriched PCR (ME-PCR) coupled sequencing and DxS EGFR mutation test kit were used to evaluate the impact of extraction method on EGFR mutation analysis. RESULTS: MPC method extracted more plasma cfDNA than Qiagen kit method (p=0.011). The proportion of longer fragment (> or =202 bp) in cfDNA extracted by MPC method was significantly higher than by Qiagen kit method (p=0.002). In the sequencing maps of ME-PCR products, a higher mutant peak was observed on plasma cfDNA extracted by MPC method than by Qiagen kit method. In DxS EGFR mutation test kit results, plasma cfDNA extracted by MPC method contained more tumor-origin DNA than by Qiagen kit method. CONCLUSION: An improved plasma cfDNA extraction method of MPC is provided, which will be beneficial for EGFR mutation analysis for patients with NSCLC.
Base Sequence ; Carcinoma, Non-Small-Cell Lung/*genetics ; Chloroform ; DNA Mutational Analysis/*methods ; DNA, Neoplasm/*blood/*isolation & purification ; Genetic Testing/methods ; Humans ; Lung Neoplasms/*genetics ; Molecular Sequence Data ; Phenol ; Polymerase Chain Reaction/methods ; Receptor, Epidermal Growth Factor/*genetics

OBJECTIVETo evaluate the expression of BRAF V600E mutation in 240 Chinese patients with thyroid lesions.

METHODSTwo hundred and forty Chinese patients with thyroid lesions, including 129 papillary thyroid carcinomas (PTC), 12 follicular carcinomas, 4 medullary carcinomas, 30 adenomas, 30 nodular goiters, and 35 papillary hyperplasia. DNA was extracted from thyroid biopsy and paraffin embedded thyroid tissues, and the expression of BRAF V600E mutation was detected by polymerase chain reaction and DNA sequencing assays.

RESULTSThe presence of BRAF V600E mutation was found in 61 of the total group of 240 cases (25.4%). It was only detected in PTC (47.3%), and not detected in other types of malignant and benign thyroid lesions. There was a statistically significant difference between the expression of BRAF V600E mutation in classic type PTC (49.6%) and in follicular type PTC (12.5%,P < 0.05), but statistical data did not show any correlation between BRAF V600E mutation and clinicopathologic parameters in PTC (P > 0.05).

CONCLUSIONSBRAF V600E mutation has a significant correlation with PTC and the detection of BRAF V600E mutation may be used as an important prognostic marker of PTC. Our new method of DNA extraction from paraffin embedded tissues is efficient and inexpensive.

Adenocarcinoma, Follicular ; genetics ; metabolism ; Adenoma ; genetics ; metabolism ; Adult ; Biomarkers, Tumor ; genetics ; Carcinoma, Papillary ; genetics ; metabolism ; Codon ; DNA Mutational Analysis ; DNA, Neoplasm ; genetics ; isolation & purification ; Female ; Goiter, Nodular ; genetics ; metabolism ; Humans ; Male ; Middle Aged ; Point Mutation ; Proto-Oncogene Proteins B-raf ; genetics ; metabolism ; Thyroid Neoplasms ; genetics ; metabolism

Carcinoma, Non-Small-Cell Lung ; genetics ; metabolism ; pathology ; DNA Mutational Analysis ; DNA, Neoplasm ; genetics ; isolation & purification ; Genes, erbB-1 ; Humans ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Mutation ; Paraffin Embedding ; methods ; Polymerase Chain Reaction ; methods ; Real-Time Polymerase Chain Reaction ; methods ; Receptor, Epidermal Growth Factor ; genetics

Transitional-CpG methylation between unmethylated promoters and nearby methylated retroelements plays a role in the establishment of tissue-specific transcription. This study examined whether chromosomal losses reducing the active genes in cancers can change transitional-CpG methylation and the transcription activity in a cancer-type-dependent manner. The transitional-CpG sites at the CpG-island margins of nine genes and the non-island-CpG sites round the transcription start sites of six genes lacking CpG islands were examined by methylation-specific polymerase chain reaction (PCR) analysis. The number of active genes in normal and cancerous tissues of the stomach, colon, breast, and nasopharynx were analyzed using the public data in silico. The CpG-island margins and non-island CpG sites tended to be hypermethylated and hypomethylated in all cancer types, respectively. The CpG-island margins were hypermethylated and a low number of genes were active in the normal stomach compared with other normal tissues. In gastric cancers, the CpG-island margins and non-island-CpG sites were hypomethylated in association with high-level chromosomal losses, and the number of active genes increased. Colon, breast, and nasopharyngeal cancers showed no significant association between the chromosomal losses and methylation changes. These findings suggest that chromosomal losses in gastric cancers are associated with the hypomethylation of the gene-control regions and the increased number of active genes.
Alu Elements/genetics ; *Chromosome Deletion ; CpG Islands/*genetics ; *DNA Methylation ; DNA, Neoplasm/chemistry/isolation & purification ; Gene Expression Profiling ; *Genes, Neoplasm ; Humans ; Long Interspersed Nucleotide Elements/genetics ; Polymerase Chain Reaction ; *Promoter Regions, Genetic ; Stomach Neoplasms/*genetics

OBJECTIVETo explore the effect of Rg3 on inhibiting and inducing apoptosis of bladder cancer cells.

METHODThe bladder cancer cell line EJ was treated with Rg3 of various concentrations. Cell proliferation was measured by MTT assay. Morphological changes of cells were observed by fluorescent staining of Hoechst 33258. Cell cycle and apoptosis rate were analyzed by flow cytometry (FCM). The expression of caspase-3 in cells was detected by immunocytochemistry. DNA ladder was showed by agarose gel electrophoresis.

RESULTRg3 inhibited proliferation of EJ cells in a manner of concentration-dependent relationship, IC50 of Rg3 in 48 h treatment was 125.5 mg x L(-1) to EJ cells. When treated with 150 mg x L(-1) of Rg3 for 24 h and 48 h, the cells showed apoptotic morphological characteristics including the condensed chromatin, the nuclear fragmentation, the apoptotic body and bright fluorescent granules as well as a higher caspase-3 expression. FCM assay indicated that Rg3 regulated cell cycle and induced apoptosis of EJ cells. When treated for 24 h and 48 h with 75 mg x L(-1) of Rg3 as well as for 48 h with 150 mg x L(-1) of Rg3, the percentages of cells in S phase and G2/M phase were increased, whereas the percentage of cells in G0-G1 was decreased. The apoptosis rates were increased from (1.05 +/- 0.17)% in control group cells to (8.41 +/- 0.98)%, (18.57 +/- 2.20)% and (33.98 +/- 1.64)%, respectively. Remarkable DNA ladders were revealed. The effects showed a manner in dose and time dependent of Rg3.

CONCLUSIONThe results suggest that ginsenoside Rg3 exerts an inhibiting effect on proliferation of EJ cells by inducing apoptosis.

Antineoplastic Agents, Phytogenic ; administration & dosage ; pharmacology ; Apoptosis ; drug effects ; Carcinoma, Transitional Cell ; genetics ; metabolism ; pathology ; Caspase 3 ; metabolism ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; DNA, Neoplasm ; genetics ; Dose-Response Relationship, Drug ; Electrophoresis, Agar Gel ; Flow Cytometry ; Ginsenosides ; administration & dosage ; isolation & purification ; pharmacology ; Humans ; Immunohistochemistry ; Inhibitory Concentration 50 ; Panax ; chemistry ; Plants, Medicinal ; chemistry ; Urinary Bladder Neoplasms ; genetics ; metabolism ; pathology

The article submitted the new progress of the study in antitumor mechanism of bererine (Ber). Reports indicated that Ber suppressed growth of tumor cells through impacting tumor cells growth cycle, inhibiting synthesises of DNA and protein, and reducing the activity of topoisomerase. Ber improved tumor cells apoptosis through several ways such as regulating apoptotic gene expression, inducing the decline of transmembrane potential on mitochondria. And Ber still could inhibit tumor metastasis through suppressing the formation of tumor angiogenesis, blocking signal transduction pathway, antagonizing extralumen, et al. In addition, Ber could induce tumor cells to differentiate to antagonize tumor.
Animals ; Antineoplastic Agents, Phytogenic ; isolation & purification ; pharmacology ; Berberine ; isolation & purification ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; DNA, Neoplasm ; biosynthesis ; Humans ; Neoplasm Proteins ; biosynthesis ; Neoplasms ; genetics ; metabolism ; pathology ; Plants, Medicinal ; chemistry

OBJECTIVETo investigate the association between p53 Arg72Pro polymorphism and cervical carcinomas HPV-associated cervical carcinoma in Uigur and Han women.

METHODSThe distribution and frequencies of p53 Arg72Pro genotypes were determined by PCR-RFLP in 152 cases of cervical carcinoma in ethnic Uigur women with 110 cases of normal control and 120 cases of cervical carcinoma in Han women with 122 cases of normal control.

RESULTSThe omni-constituent ratio of p53 genotype was statistically different between cervical carcinoma and normal control groups in the Uigur (chi(2) = 7.196, P < 0.05) group. The proportion of Arg/Arg was higher in cervical carcinomas than that in control. The omni-constituent ratio of p53 genotype was statistically different between cervical carcinoma and normal control groups in Han (chi(2) = 8.231, P < 0.025). The proportion of Pro/Pro was higher in cervical carcinoma than that in normal control. The omni-constituent ratio was statistically different between HPV 16 positive and negative groups of cervical carcinoma in the Uigur group (chi(2) = 7.177, P < 0.05). The proportion of Arg/Arg was higher in HPV 16 positive group than that in HPV 16 negative group.

CONCLUSIONSp53 Arg72Pro polymorphism may be associated with the development of cervical carcinoma in Uigur and Han women in Xinjiang. p53 Arg/Arg genotype may be a genetically susceptible factor to HPV-associated cervical carcinoma in Uigur. p53 Pro/Pro genotype may be a genetically susceptible factor to cervical carcinoma in Han. There may be different susceptibilities to cervical cancer between Uigur and Han women in Xinjiang.

Adult ; Aged ; Alleles ; Base Sequence ; China ; ethnology ; Codon ; DNA, Neoplasm ; genetics ; Exons ; Female ; Gene Frequency ; Genes, p53 ; Genetic Predisposition to Disease ; ethnology ; Human papillomavirus 16 ; isolation & purification ; Humans ; Middle Aged ; Molecular Sequence Data ; Papillomavirus Infections ; ethnology ; Polymorphism, Genetic ; Tumor Suppressor Protein p53 ; genetics ; Uterine Cervical Neoplasms ; ethnology ; genetics ; virology ; Young Adult

OBJECTIVETo investigate the clinicopathological significance of p16(INK4A) expression and DNA ploidy status in HPV-negative uterine cervical cancers and their precursors.

METHODSHPV-negative cervical lesions, including 20 cases of cervicitis, 20 cases of cervical intraepithelial neoplasm (CIN), 3 cases of cervical glandular intraepithelial neoplasm (CGIN), 38 cases of invasive squamous cell carcinoma (SCCs) and 15 cases of invasive adenocarcinoma were selected and subject to screening for HPV infection by PCR method. The p16(INK4A) protein expression and DNA ploidy status were studied by immunohistochemistry and flow cytometry respectively.

RESULTSSpecific expression of p16(INK4A) was seen in both the nucleus and cytoplasm of the dysplastic and malignant cells of CIN, CGIN, cervical SCC and adenocarcinoma. In contrast, no expression was present in normal and inflammatory squamous or glandular epithelium. DNA aneuploidy was significantly more frequent in invasive SCCs and adenocarcinomas than in CIN (P < 0.01). Aneuploid was also more frequent in the lymph node positive group than lymph node negative group, although no statistic significance was found. Among the 8 cases of p16(INK4A) negative SCCs, two showed DNA aneuploidy.

CONCLUSIONSImmunohistochemical detection for p16(INK4A) can be an early diagnostic marker for HPV-negative cervical SCC and adenocarcinoma. DNA ploidy analysis may further assist the diagnosis of cervical malignancies.

Adenocarcinoma ; genetics ; metabolism ; pathology ; Aneuploidy ; Carcinoma, Squamous Cell ; genetics ; metabolism ; pathology ; Cervical Intraepithelial Neoplasia ; genetics ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p16 ; metabolism ; DNA, Neoplasm ; genetics ; Female ; Flow Cytometry ; Humans ; Immunohistochemistry ; Papillomaviridae ; genetics ; isolation & purification ; Polymerase Chain Reaction ; Uterine Cervical Neoplasms ; genetics ; metabolism ; pathology ; Uterine Cervicitis ; genetics ; metabolism ; pathology