BACKGROUND: The p16 protein is a cyclin-dependent kinase inhibitor (CDKI) that inhibits cell cycle progression from phase G1 to phase S in the cell cycle. Many p16 gene mutations have been noted in many cancer-cell lines and in some primary cancers. These mutated genes caused abnormal or aberrant expression of the p16 protein, which might have contributed to the malignant progression of the cells by deranging the cell cycle. This study was to examine the abnormal or aberrant expression of the p16 protein in breast cancer tissue by using p16 protein specific immunohistochemical staining. METHODS: p16-protein-specific immunohistochemical staining was performed on 31 breast-cancer tissue samples. Twenty-four cases among the 31 tissue staining slides simultaneously showed a normal breast-tissue portion on the same staining slide. Microscopic photographs of both the breast-cancer and the normal- tissue portion were taken at the same magnification to compare the statistically analyzed fraction of red or brown colored p16 stained nuclei. RESULTS: In the breast cancer tissue, 7 (22.6%) showed totally negative, with less than 5% of the nuclei staining. The completely negative cases were not related to the stage of the disease (p=0.096) or to the histopathologic grade (p=0.20). The staining ratios of the breast-cancer tissue and the normal tissue were 26.2 ( +/- 18.7)% and 72.4 ( +/- 18.8)%, respectively. In the breast-cancer tissue, the ratio of expression of the p16 protein was significantly lower than in the normal tissue (p=0.001). CONCLUSIONS: In the carcinogenesis of some breast cancers, low expression of the p16 protein may play an important role in the unlimited proliferation of tumor cell due to a loss of the cell-cycle-regulating role of the p16 protein.
Breast Neoplasms* ; Breast* ; Carcinogenesis ; Cell Cycle ; Cyclin-Dependent Kinase Inhibitor p16* ; Genes, p16 ; Humans* ; Phosphotransferases

OBJECTIVE: To overcome the limitations of the single gene transfer, the authors present the results of wild-type p16 and p53 combined genes transfer in vitro to the U251MG and U373MG cell lines using cationic liposome as a vector. METHODS: To compare the therapeutic effect of the combined p16 and p53 genes transfer with the single p16 and p53 gene transfer, full length of wild-type human p16 and p53 gene, and combined p16-p53 genes were transferred in vitro to the U251MG and U373MG cell lines using cationic liposome as a vector. As the U251MG and U373MG cell lines are devoid of p16 and p53 genes, the therapeutic effect of the three groups of gene transfer could be evaluated by the growth suppression or percentage of the viable cells. Reverse transcription polymerase chain reaction(RT-PCR), flow cytometry, and electron microscopy(EM) were used for evaluation of the growth suppression or apoptosis of the tumor cells. RESULTS: p16 gene, p53 gene and the combined p16-p53 genes were effectively transferred to the cell lines using cationic liposome as a vector resulting in dramatic decrease of the viable tumor cells in comparison to the control group(p=0.004). The cytotoxic effect of the gene transfer in the U251MG cell line was the most significant in the combined p16-p53 group. However, in the U373MG cell line p53 single gene transfer group showed more significant effect than the combined gene transfer group. Apoptosis was confirmed by EM in the combined p16-p53 genes group. The G1 phase arrest effect, confirmed by the flow cytometry was more prevalent in the p16 gene transfer group than the other groups. CONCLUSION: Cationic liposome-mediated transfer of combined p16-p53 genes to the human glioblastoma cell lines is proven effective. However, the therapeutic effect of the combined p16-p53 genes transfer was not consistently superior to the single p16 or p53 gene transfer.
Apoptosis ; Cell Line* ; Flow Cytometry ; G1 Phase ; Genes, p16 ; Genes, p53* ; Glioblastoma* ; Humans* ; Liposomes ; Reverse Transcription

BACKGROUND: Loss of heterozygosity (LOH) and mutation of the p16 tumor suppressor gene have been detected in non-Hodgkin's lymphomas (NHLs). Recently, hypermethylation of the p16 gene has been reported. The role of p16 gene alterations in the genesis of NHLs and their high-grade transformations require explanation. METHODS: LOH of D9S171 and IFNA microsatellite markers, DNA hypermethylation, and mutation of exon 1 and 2A were assessed in 43 cases of NHLs. The genetic abnormalities were compared with the protein expression by immunohistochemistry, and they were evaluated according to the histologic subtypes, grades and immunophenotypes. RESULTS: DNA hypermethylation was the most common p16 gene abnormality and was found in 30 of 39 cases (76.9%). Eight cases (18.6%) showed LOH in one or both microsatellite markers, and five cases (11.6%) showed mutations in exon 1 or 2A. Loss of protein expression was seen in 17 cases (39.5%) and was associated with mutation and LOH. Loss of protein was more frequent in high-grade lymphomas than in low-grade lymphomas. CONCLUSION: These results suggest that the functional loss of the p16 gene contributes to the development of NHLs, especially to the development of high-grade lymphomas.
DNA ; Exons ; Genes, p16* ; Genes, Tumor Suppressor ; Immunohistochemistry ; Loss of Heterozygosity ; Lymphoma ; Lymphoma, Non-Hodgkin* ; Microsatellite Repeats

OBJECTIVE: Hypermethylation of p16, a tumor suppressor gene, has been frequently detected in a variety of cancer cells and is known to represent the level of p16 transcription. In human meningiomas, genetic alterations of p16 have shown to be infrequent. The purpose of this study is to investigate the role of p16 associated with the progression of meningiomas. METHODS: Sixty-eight meningiomas(randomly sampled 29 benign, 16 atypical and 23 malignant formalin-fixed, paraffin-embedded tissues) were analyzed. We examined the molecular mechanism of inactivation of p16 in these benign, atypical and malignant meningiomas by detecting the methylation status of p16 using methylation-specific polymerase chain reaction. RESULTS: One out of 29(3.4%) revealed hypermethylation of p16 in benign meningiomas. Atypical and malignant meningiomas showed hypermethylation of p16 in 2 out of 16 cases(12.5%) and in 5 out of 23 cases(21.7%), respectively. Immunohistochemical analysis of methylation-positive tumors demonstrated that tumor cells had reduced immunoreactivity compared to normal lymphocytes. CONCLUSIONS: Our results suggest that inactivation of p16 gene plays a role in the pathogenesis of meningioma and hypermethylation is one of the processes for gene inactivation.
CpG Islands* ; Gene Silencing ; Genes, p16 ; Genes, Tumor Suppressor ; Humans ; Lymphocytes ; Meningioma* ; Methylation* ; Polymerase Chain Reaction

The tumor suppressor gene p53 and p16, both of which play an important role in inhibition of tumorigenesis, are homozygously deleted in human myeloid leukemia cell line K562. To explore the inhibition of K562 cell proliferation by wild type p16 and p53 genes, both p16 and p53 genes were co-transfected into K562 cells mediated by liposome. The expression of the two genes was measured by immunocytochemical method, the cell cycle was analysed by flow cytometry, and the number of recovered viable cells was assessed after transfection. After co-transfection, the p53 and p16 positive cells were 23% and 28%, respectively. The results showed that co-transfection of p16 and p53 genes significantly inhibits cell proliferation comparing with transfection either by p16 gene or by p53 gene (P < 0.05). Expression of p16 and p53 proteins increased the cell number in G(1) phase but decreased the cell number in S phase. It is concluded that co-transfection of p16 and p53 genes has a stronger growth-inhibitory effect on K562 cell growth than that of transfection only by p16 gene or by p53 gene, may be a pathway for gene therapy in leukemia.
Cell Division ; Genes, p16 ; physiology ; Genes, p53 ; physiology ; Humans ; K562 Cells ; Plasmids ; Transfection

Animals ; Genes, BRCA2 ; Genes, erbB-1 ; Genes, erbB-2 ; Genes, p16 ; Genes, p53 ; Genes, ras ; Genetic Predisposition to Disease ; genetics ; Humans ; Pancreatic Neoplasms ; classification ; genetics ; pathology

PURPOSE: Alterations of the p15(INK4B) and p16(INK4A) gene which are separated by 25 kb on chromosome 9p21 have been reported in various tumor derived cell lines and primary tumors, but the role of these genes in cervical cancer is unknown. MATERIAL AND METHOD: To determine the frequency of deletions and point mutations of these genes in human cervical cancer, we examined 57 primary tumors and matched normal tissues, and 3 cervical cancer derived cell lines. All the tumor tissues and cell lines were human papil- INK48 lomavirus (HPV)-positive. Deletions or point mutations of exon 2 of the pl5 gene and exons 1, 2, and 3 of the p16(INK4A) gene were examined by polymerase chain reaction (PCR) and direct sequencing, respectively. RESULT: Our data indicate no evidence for intragenic homozygous deletion or point mutation in the cervical cancer or cervical cancer derived cell lines. INK48 INK4A CONCLUSION: Deletions or point mutations in the p15 or p16 gene may not be required for the development of HPV-positive cervical cancer or for establishment of cervical cancer cell lines.
Cell Line* ; Cyclin-Dependent Kinase Inhibitor p16 ; Exons ; Genes, p16 ; Humans ; Point Mutation ; Polymerase Chain Reaction ; Uterine Cervical Neoplasms

OBJECTIVETo explore the correlation between homozygous deletions and mutation of p16 gene and the carcinogenesis and progression of squamous cell carcinoma of buccal mucosa.

METHODSThirty buccal cancers, 10 leukoplakias and 8 buccal mucosas were involved. DNA was extracted from the tissues. PCR was used to analyses homozygous deletion of p16 gene. PCR-SSCP-DNA sequencing was performed to detect the point mutation of p16 gene. Immunohistochemical techniques were used to detect the expression of P16 protein.

RESULTSGene deletions and point mutations were not found in leukoplakia and normal buccal mucosa. Gene deletions were found in 7 samples out of 30 cases of squamous cell carcinoma of buccal mucosa (23.3%), while point mutations were found in 5 samples out of 30 cases of squamous cell carcinoma of buccal mucosa (16.7%). Sequencing analysis showed that 5 cases point mutations were missense mutations, occurred on exon 2. Three cases occurred in the same point, codon 99 (GAT --> AAT). The result of immunohistochemical stains showed that 11 out of 12 cases gene inactivation did not expressed P16 protein.

CONCLUSIONHomozygous deletion and point mutation of p16 were the main pattern of gene inactivation in squamous cell carcinoma of buccal mucosa. There was a closely correlation between p16 gene inactivation and the carcinogenesis of squamous cell carcinoma of buccal mucosa.

BACKGROUND: Two tumor suppressor genes, p53 and p16, which have different roles in controlling the cell cycle and inducing apoptosis, are frequently inactivated during carcinogenesis including lung cancer. Single tumor suppressor gene therapies using tither with p53 or p16 have been studied extensively. However, there is a paucity of reports regarding a combined gene therapy using these two genes. METHODS: The combined effect of p53 and p16 gene transfer by the adenoviral vector on the growth of lung cancer cell lines and its interactive mechanism was investigated. RESULTS: An isobologram showed that the co-transduction of p53 and p16 exhibited a synergistic growth inhibitory effect on NCI H358 and an additive effect on NCI H23. Cell cycle analysis demonstrated the induction of a synergistic G1/S arrest by a combined p53 and p16 transfer. This synergistic interaction was again confirmed in a soft agar clonogenic assay. CONCLUSION: These observations suggest the potential of a p53 and p16 combination gene therapy as another potent strategy in cancer gene therapy.
Adenoviridae ; Agar ; Apoptosis ; Carcinogenesis ; Cell Cycle ; Cell Line* ; Genes, Neoplasm ; Genes, p16 ; Genes, Tumor Suppressor ; Genetic Therapy ; Lung Neoplasms* ; Lung*

Pancreatic cancer is a disease with poor prognosis mainly due to low resection rates and late diagnosis. To increase resectability and improve survival rates, a better understanding of pancreatic cancer pathogenesis and more effective screening techniques are required. New methods, such as genetic and molecular alterations, may suggest novel approaches for pancreatic cancer diagnosis and treatment. We immunohistochemically investigated 44 formalin-fixed, paraffin-embedded specimens of pancreatic ductal adenocarcinoma using monoclonal anti-p16 antibodies and monoclonal anti-p53 antibodies. The expressions of p16 and p53 proteins were compared using the Chi-square test with SPSS. Disease-free survival was analyzed using the Kaplan-Meier method, verified by the Log- Rank test. Loss of p16 expression was noted in 20 (45.5%) cases and aberrant p53 protein expression was detected in 14 (31.8%) cases. Loss of p16 expression was associated with a higher incidence of lymph node metastasis (p=0.040) and a more advanced stage (p=0.015), although there was no significant correlation between p16 expression and survival. Aberrant p53 protein expression correlated with histologic grade (p= 0.038). Disease-free survival rate was significantly lower in the aberrant p53 protein positive group compared to the negative group (p=0.029). From our results, we suggest that p53 is not a prognostic factor; however, p16 and p53 genes do play important roles in the progression of pancreatic ductal adenocarcinoma.
Adult ; Aged ; Female ; Genes, p16 ; Genes, p53 ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Neoplasm Staging ; Pancreatic Neoplasms/*chemistry/genetics/mortality/pathology ; Protein p16/*analysis ; Protein p53/*analysis ; Sex Characteristics