Background: Development of squamous cell cancer of head and neck (SCCHN) is associated with human papillomavirus (HPV) infection, which in turn is closely related with expression of $p16^{INK4A}$. Loss of $p16^{INK4A}$ expression by deletion, mutation, or hypermethylation is common in SCCHN. We here evaluated $p16^{INK4A}$ as a prognostic marker of treatment response and survival in our SCCHN patients with laryngeal, hypopharyngeal or nasopharyngeal cancers. Materials and Methods: 131 patients diagnosed with SCCHN between January 2,2006 and July 17, 2010 were examined for $p16^{INK4A}$. The median age was 60 years (15-82 years). Fifty one patients were stage I-II and 80 were stage III-IV. Immunohistochemical expression of $p16^{INK4A}$ was analyzed in pretreatment paraffin-embedded tumor blocks. The influence of $p16^{INK4A}$ status on disease-free survival, and overall survival after treatment was evaluated. Results: $p16^{INK4A}$ positivity was found in 58 patients (44%). Tumor-positivity for$ p16^{INK4A}$ was correlated with improved disease free survival (70.1 months vs 59 months) and improved overall survival (2, 3 and 5-year values; 77% vs 72%, 70% vs 63% and, 63% vs 55%; respectively). On multivariate analysis, stage was determined as independent prognostic factor for disease-free survival. Conclusions: Stage was the major prognostic factor on treatment response and survival in our patients. $p16^{INK4A}$ status predicts better outcome in laryngeal, hypopharyngeal or nasopharyngeal cancer cases treated with surgery plus adjuvant radiochemotherapy as well as with definitive radiation therapy and/or chemotherapy.
Journal of the Korean Association of Oral and Maxillofacial Surgeons
/
v.38
no.2
/
pp.78-84
/
2012
Objectives: Odontogenic keratocysts (OKCs) have a tendency to recur and possess an aggressive nature. the aim of the present study was to evaluate cytokeratin (CK) expression patterns as a method for the differentiation between dentigerous cysts (DCs) and OKCs, as their histomorphologic appearance are often indistinguishable. Materials and Methods: Formalin-fixed, paraffin-embedded tissue sections of 43 OKCs and 38 DCs were immunohistochemically analyzed with i-solution in a quantitative manner in order to evaluate the immunoreactivity of CK 10, 16 and 17. Results: CK 10 expression was evident in 79.1% of OKCs but found in only 18.4% of DCs (P<0.05), and CK 10 expression was observed to occur more frequently in OKCs (mean 25.45%) than in DCs (2.19%) (P<0.05). The expression of CK 16 was evident in 79.1% of OKCs but found in only 7.9% of the DCs (P<0.05) and CK 16 expression was observed to occur more frequently in OKCs (mean 4.33%) than in the DCs (0.61%) (P<0.05). The expression of CK 17 was evident in 88.4% of OKCs but seen in only 15.7% of the DCs (P<0.05) and CK 17 expression was observed to occur more frequently in OKCs (mean 31.11%) than in the DCs (2.37%) (P<0.05). Conclusion: The immunohistochemical detection of CK 10, 16 and 17 can be utilized as a valuable biomarker for use in distinguishing between OKCs and DCs, which have clinically significant differential diagnoses.
Journal of the Korean Association of Oral and Maxillofacial Surgeons
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v.38
no.2
/
pp.101-109
/
2012
Objectives: The inactivation of the tumor suppressor gene $p16^{INK4a}$ plays an important role in the development of malignant tumors, including oral squamous cell carcinoma. The p16 gene is involved in the p16/cyclin-dependent kinase/retinoblastoma (Rb) gene pathway of cell cycle control. The p16 protein is considered a negative regulator of this pathway. The p16 gene encodes an inhibitor of cyclin-dependent kinases 4 and 6 which regulate the phosphorylation of the retinoblastoma gene and G1 to S phase transition in the cell cycle. However, the p16 gene can lose its functionality through point mutations, loss of heterozygosity or methylation of its promoter region. Materials and Methods: In this study, the authors analyzed the correlation between various clinicopathological findings- patient age, gender and smoking, disease recurrence, tumor size, stage, and differentiation- and p16 protein expression or p16 promoter hypermethylation in 59 cases of head and neck squamous cell carcinoma. Results: The results revealed p16 protein expression and p16 promoter hypermethylation in 28 cases (47.5%) and 21 cases (35.6%), respectively, of head and neck squamous cell carcinoma. However, neither p16 protein expression nor p16 promoter hypermethylation had any statistical influence on clinicopathological findings or survival rate. Conclusion: This data, and a review of the literature, suggest that p16 promoter hypermethylation cannot yet be used as an independent prognostic factor influencing carcinogenesis, but must be considered as an important factor along with other genetic alterations affecting the pRb pathway.
Background: Cancer of the gallbladder is a relatively rare neoplasm with a poor prognosis. The exact mechanisms of its genesis are not known and very little information is available on molecular events leading to labeling this as an orphan cancer. Materials and Methods: In this prospective case control study we evaluated the expression of p16, pRb and cyclin D1 by immunohistochemistry to study the G1-S cell-cycle check point and its possible role in gallbladder carcinogenesis. A total of 25 patients with gallbladder carcinoma (group I), 25 with cholelithiasis (group II) and 10 normal controls. were enrolled Results: Cyclin D1 expression was seen in 10 (40%) patients each with carcinoma and cholelithiasis while only in 2 (20%) of the normal gallbladders but differences were not statistically significant (p value=0.488). p16 was expressed in 12% patients of carcinoma of the gallbladder and 28% of cholelithiasis, however this difference was not statistically significant (p value=0.095). Retinoblastoma protein was found to be expressed in 50% of normal gallbladders and 6 (24%) of carcinoma and 8 (32%) of gallstones. The present study failed to demonstrate any conclusive role of cyclin D1/RB/ p16 pathway in carcinoma of the gallbladder. Conclusions: The positive relation observed between tumor metastasis and cyclinD1 expression and p16 with nodal metastasis suggested that higher cyclin D1/p16 expression may act as a predictive biomarker for aggressive behavior of gallbladder malignancies.
Kim, Mi-Suk;Kwon, Hee-Chung;Kang, Hee-Seog;Park, In-Chul;Rhee, Chang-Hun;Kim, Chang-Min;Lee, Choon-Taek;Hong, Seok-Il;Lee, Seung-Hoon
Journal of Korean Neurosurgical Society
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v.29
no.4
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pp.471-476
/
2000
Objective : p16/INK4a, a kind of tumor suppressor genes, encodes a specific inhibitor of the cyclin D-dependent kinases CDK4 and CDK6. This prevents the association of CDK4 with cyclin D1, and subsequently inhibits phosphorylation of retinoblastoma tumor suppressor protein(pRb), thus preventing exit from the G1 phase. According to previous reports, over 50% of glioma tissue and 80% of glioma cell lines have been demonstrated inactivation of p16/INK4a gene. The purpose of this study was to determine whether recombinant adenovirus-p16 virus is a suitable candidate for gene replacement therapy in cases of glioma. Methods : Three human glioma cell lines(U251MG, U87MG and U373MG) that express mutant p16 protein were used. Replication-deficient adenovirus was utilized as an expression vector to transfer exogenous p16 cDNA into the cells ; control cells were infected with the Ad-${\beta}$-gal expressing ${\beta}$-galactosidase. To monitor gene transfer and the expression of exogenous genes, we used Western Blotting analysis. Flow cytometry studies of cellular DNA content were performed to determine the cell cycle phenotype of the glioma cells before and after treatment. Results : We showed here that restoration of p16/INK4a expression in p16 negative U87MG, U251MG and partially deleted U373MG by Ad-CMV-p16 induced growth suppression in vitro. Flow cytometric study revealed that Ad-CMV-p16 infected U87MG cells were arrested during the G0-G1 phase of the cell cycle. Expression of p16 transferred by Ad-CMV-p16 in glioma cells was highly efficient and maintained for more than seven days. Conclusions : Our results suggest that Ad-CMV-p16 gene therapy strategy is potentially useful and warrants further clinical investigation for the treatment of gliomas.
Background: Invasive ductal (IDC) and lobular (ILC) carcinomas are the common histological types of breast carcinoma which are difficult to distinguish when poorly differentiated. Discoidin domain receptor (DDR1) and Drosophila dishevelled protein (DVL1) were recently suggested to differentiate IDC from ILC. Objectives: To assess the expression of DDR1 and DVL1 and their association with histological type, grading and hormonal status of IDC and ILC. Materials and Methods: This cross sectional study was conducted on IDC and ILC breast tumours. Tumours were immunohistochemically stained for (DDR1) and (DVL1) as well as estrogen receptor (ER), progesterone receptor (PR) and C-erbB2 receptor. Demographic data including age and ethnicity were obtained from patient records. Results: A total of 51 cases (30 IDCs and 21 ILCs) were assessed. DDR1 and DVL1 expression was not significantly associated with histological type (p=0.57 and p=0.66 respectively). There was no association between DDR1 and DVL1 expression and tumour grade (p=0.32 and p=1.00 respectively), ER (p=0.62 and 0.50 respectively), PR (p=0.38 and p=0.63 respectively) and C-erbB2 expression (p=0.19 and p=0.33 respectively) in IDC. There was no association between DDR1 and DVL1 expression and tumour grade (p=0.52 and p=0.33 respectively), ER (p=0.06 and p=0.76 respectively), PR (p=0.61 and p=0.43 respectively) and C-erbB2 expression (p=0.58 and p=0.76 respectively) in ILC. Conclusions: This study revealed that DDR1 and DVL1 are present in both IDC and ILC regardless of the tumour differentiation. More studies are needed to assess the potential of these two proteins in distinguishing IDC from ILC in breast tumours.
Objectives: We aimed to identify the dose-dependent inhibitory effects of Maekmundongcheongpye-eum and Liriopis Tuber on the mRNA expression of IL-6, IL-16, GM-CSF involved in the asthma model. Methods: In the study BEAS-2B cell lines, human epithelial cells were used. These cells were stimulated with tumor necrosis factor $(TNF)-{\alpha}$ for artificial inflammatory expression. ${\beta}-actin$ messenger RNA (mRNA) was used by internal standard. After 24 hours of Maekmundongcheongpye-eum, Liriopis Tuber-treatment, total cellular RNAs were collected, treating RNAzol directly on the alive cells. Then the transcriptional activities of IL-6, 16, GM-CSF were measured by RT-PCR with electrophoresis. Results: In the Maekmundongcheongpye-eum study, the mRNA expression of IL-6 showed 48% transcriptional inhibitory effect compared to the control group in the $100{\;}{\mu}l/ml$ category (P<0.001). In the IL-16, there was 53% and 57% transcriptional inhibitory effect compared to the control group in the $20{\;}{\mu}l/ml$ and $100{\;}{\mu}l/ml$ categories (P<0.001). In the GM-CSF, there was no inhibitory effect. In the Liriopis Tuber study, the mRNA expression of IL-6 showed 43% transcriptional inhibitory effect compared to the control group in the $100{\;}{\mu}l/ml$ category (p<0.005). In the IL-16, 34% and 26% of transcriptional inhibitory effect was shown compared to the control group in the $20{\;}{\mu}l/ml$ and $100{\;}{\mu}l/ml$ categories, respectively (P<0.05). In the GM-CSF, there was no inhibitory effect. Conclusions: This study shows that Maekmundongcheongpye-eum and Liriopis Tuber have dose-dependent inhibitory effects on the mRNA expression of IL-6 and IL-16 in BEAS-2B cell lines, human epithelial cells. Advanced studies are required to investigate the mechanisms of inhibition by herbal medicine in the asthma model.
Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3) is an important serine/threonine protein kinase and a member of the MAPK family. MEKK3 can effectively activate the MEK/ERK signaling pathway and promote an autocrine growth loop critical for tumor genesis, cell proliferation, terminal differentiation, apoptosis and survival. To explore the relationship between MEKK3 and cell apoptosis, clinicopathology and prognosis, we characterize the expression of MEKK3, pERK and FoxP3 in the renal clear cell carcinoma (RCCC). Protein expression was detected by tissue microarray and immunochemistry in 46 cases of RCCC and 28 control cases. Expression levels of CD3+,CD3+CD4+,CD3+CD8+,CD4+CD25+, CD4+CD25+ FoxP3+ were assessed by flow cytometry and analyzed for their association with pathological factors, correlation and prognosis in RCCC. Expression of MEKK3, pERK and FoxP3 was significantly up-regulated in RCCC as compared to control levels (p<0.01), associated with pathological grade (p<0.05)and clinical stage (p<0.05). CD4+CD25+ Foxp3+ Treg cells were also significantly increased in RCCC patients (p<0.05). Cox multivariate regression analysis showed that MEKK3, pERK expression and patholigical stage were independent prognostic factors in patients with RCCC (p<0.05). MEKK3 can be used as an important marker of early diagnosis and prognostic evaluation in RCCC. It may be associated with imbalance of anti-tumor immunity and overexpression of pERK. Expression of MEKK3 and pERK are significantly increased in RCCC, with protein expression and clinical stage acting as independent prognostic factors.
Kim, Seon-Young;Lee, Chang-Hun;Kim, Kyung-Joo;Kim, Yeon-Soo
Journal of Microbiology and Biotechnology
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v.11
no.2
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pp.234-241
/
2001
The C-terminal 393 bp region of the human interleukin-16 (IL-16) gene was cloned and expressed in E. coli along with mammalian cell lines. Recombinant IL-16 expressed from E. coli was 22 kDa on SDS-PAGE and showed 260% of chemoattractant activity at a concentration of $0.1\;{\mu}g/ml$. HeLa, COS, and Neuro-2a cells were transduced by recombinant retrovirus vector pLNC/IL-16/IRES/TK and the intracellular and secreted amounts of IL-16 produced by HeLa/IL-16/TK, COS/IL-16/TK, and Neuro-2a/IL-16/TK cells were determined by enzyme-linked immunosorbent assay (ELISA). HeLa/IL-16/TK $(1{\times}10^5)$ and COS/IL-16/TK $(1{\times}10^5)$ cells secreted 36.1 and 13.3 ng of IL-16 for 48 h, respectively. Forty-nine ng and 86.4 ng of IL-16 remained in the cell lysates of HeLa/IL-16/TK and COS/IL-16/TK. Intracellular and secreted amounts of IL-16 from Neuro-2a/IL-16/TK $(5{\times}10^5)$ cells during 24 h cultivation were 50 ng and 3.3 ng, respectively. Also, HeLa and COS cells wee stably transfected with mammalian expression vector pCRIII/IL-16. Both culture media and cell lysates prepared from HeLa/IL-16 cells and COS/IL-16 cells showed chemoattractant activity ranging from 190% to 460% as compared to the control experiment. Expression of the herpes simplex virus thymidine kinase (HSV0tk) gene in pLNC/IL-16/ IRES/TK bicistronic retroviral expression vector was verified by performing a genciclovir (GCV) sensitivity assay. Finally, IL-16 repressed Tat-transactivated human immunodeficiency virus type 1 long terminal repeat (HIV-1 LTR) promoter activity.
Purpose: We investigated the impacts of the methylation states of the P16 and the hMLH1 genes on pathogenesis and genetic expression of stomach cancer and their relationships with Helicobater pylori infection, and with other clinico-pathologic factors. Material and Methods: In our study, to detect protein expression and methylation status of the P16 and the hMLH1 genes in 100 advanced gastric adenocarcinomas, used immunohistochemical staining and methylation-specific PCR (MSP) and direct automatic genetic sequencing analysis. Results: Methylation of the P16 gene was observed in 19 out of 100 cases (19%) and in the 18 of those cases (94.7%) loss of protein expression was seen. We were sble to show that loss of P16 gene expression was related to methylation of the P16 gene (kappa coefficient=0.317, p=0.0011). Methylation of the hMLH1 gene was observed in 27 cases (27%), and in 24 cases of those 27 cases (88.8%), loss of protein expression was seen, which suggested that loss of protein expression in the hMLH1 gene is related to methylation of hMLH1 gene (kappa coefficient=0.675, P<0.0001). Also methylation of the hMLH1 gene was related to age, size of the mass, and lauren's classification. Conclusion: We found that methylation of DNA plays an important role in inactivation of the P16 and the hMLH1 genes. The methylation of the hMLH1 genes is significantly related to age, size of the mass, and lauren's classification.
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