The effect of elemene on lung adenocarcinoma A549 cell radiosensitivity and elucidation of its mechanism

OBJECTIVE: To investigate the effect of elemene on the radiosensitivity of A549 cells and its possible molecular mechanism. METHODS: Apoptosis of A549 cells was detected by flow cytometry and fluorescence microscopy. The effect of double-strand break (DSB) damage repair in A549 cells was evaluated using the neutral comet assay. Protein expression levels were detected using western blotting, and the correlation between protein levels was analyzed. RESULTS: Elemene exhibited a radiosensitizing effect on A549 cells. The level of apoptosis induced by elemene combined with radiation was significantly greater (p<0.01) than that elicited by either radiation or elemene alone. Following radiation and subsequent repair for 24 h, the tail intensity of A549 cells treated with a combination of elemene and radiation was greater than that of cells treated with either elemene or radiation alone (p<0.01). This result indicates that elemene inhibits cellular DSB repair. Both elemene combined with radiation and radiation alone decreased the protein expression of DNA-PKcs and Bcl-2 compared to elemene alone (p<0.01), while p53 protein expression was increased (p<0.01). A negative correlation was observed between DNA-PKcs and p53 expression (r=−0.569, p=0.040), while a positive correlation was found between DNA-PKcs and Bcl-2 expression (r=0.755, p=0.012). CONCLUSIONS: Elemene exhibits a radiosensitizing effect on A549 cells, and its underlying molecular mechanism of action may be related to the downregulation of DNA-PKcs gene expression. .

Inhibitors of DNA-dependent protein kinase promote p53-independent apoptosis induced by 1, 4-benzoquinone in HL60 cells / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To investigate the impact of NU7026 and Wortmannin, inhibitors of DNA-dependent protein kinase (DNA-PK), in HL60 cells apoptosis induced by 1, 4-benzoquinone (1, 4-BQ).</p><p><b>METHODS</b>HL60 cells were divided into three groups according to the exposures: the poisoned groups which were treated with 0, 5, 10, 25 and 50 µmol/L 1, 4-BQ for 24 h, respectively, the NU7026 groups which were preincubated with 10 µmol/L NU7026 for 1h prior to the 24h treatment of 0, 5, 10, 25 and 50 µmol/L 1, 4-BQ and the Wortmannin groups which were preincubated with 25 µmol/L Wortmannin for 1h prior to the 24 h treatment of 0, 5, 10, 25 and 50 µmol/L 1, 4-BQ. Then we detected the apoptosis via flowcytometry Annexin V-FITC/PI and the DNA Ladder, respectively. We also tested the expressions of Bax mRNA with Real-Time PCR in HL60 cells which were exposed to 10 µmol/L NU7026 for 24 h, 25 µmol/L Wortmannin 24 h, 10 µmol/L 1, 4-BQ 24 h, 10 µmol/L NU7026 1h+10 µmol/L 1, 4-BQ 24 h and 25 µmol/L Wortmannin 1 h+10 µmol/L 1, 4-BQ 24 h, as well as null (control). We also examed the expressions of p53 in HL60 cells with Western blot.</p><p><b>RESULTS</b>Annexin V-FITC/PI apoptosis tests suggested that apoptosis rates of NU7026+10 µmol/L 1, 4-BQ group and Wortmannin +10 µmol/L 1, 4-BQ were 17.6±1.19% and 15.2±1.22%, respectively. Both of results were higher than that of 10 µmol/L 1, 4-BQ group (6.3±1.04%); Apoptosis of NU7026+25 µmol/L 1, 4-BQ group was 46.2±3.55%, and Wortmannin +25 µmol/L 1, 4-BQ group 26.9±2.62%. Both of results were also higher than that of 25 µmol/L 1, 4-BQ group (14.1±1.54%); Apoptosis of NU7026+50 µmol/L 1, 4-BQ group (61.8±1.78%) was higher than that of 50 µmol/L 1, 4-BQ group (35.9±4.51%). The above results were all statistically significant (P < 0.05).</p><p><b>RESULTS</b>of DNA-Ladder were basically consistent with those of Annexin V-FITC/PI apoptosis tests. In addition, both NU7026 and Wortmannin pretreatment elicited the higher expression of Bax mRNA in HL60 treated by 1, 4-benzoquinone with statistically significance (P < 0.05). However, p53 protein was not detected in HL60 cells as the western blot indicated.</p><p><b>CONCLUSION</b>Inhibitors of DNA-PK, NU7026 and Wortmannin, promote p53-independent apoptosis induced by 1, 4-benzoquinone in HL60 cells.</p>

El Estado, la espera y la dominación política en los sectores populares: entrevista al sociólogo Javier Auyero / The State, waiting and political domination among the poor: interview with the sociologist Javier Auyero

La extensa obra de Javier Auyero sobre los sectores populares en América Latina inquieta por su complejidad sociológica y política. Alejada de los lugares comunes sobre cómo viven, sufren y se relacionan los habitantes de los márgenes de nuestras ciudades, su programa de veinte años de investigación aborda las consecuencias del neoliberalismo en la marginalidad urbana. Por la publicación de su último libro, Pacientes del Estado (2013), Salud Colectiva lo invita a reflexionar sobre las conexiones, no siempre observadas, entre la espera y la dominación política en oficinas estatales, escuelas y hospitales. Su estrategia etnográfica le permite ingresar sin prejuicios a un universo social atravesado por posicionamientos sociales polarizantes. En los encuentros cotidianos de los pobres con diversas formas de poder estatal, afirma, se reproducen prácticas -no todas ellas igualmente conscientes y planificadas- que imparten educación política y culminan convirtiendo a quienes deberían ser ciudadanos con derechos en pacientes del Estado.

The extensive work of Javier Auyero regarding the poor in Latin America is disturbing in its sociological and political complexity. Instead of falling into the commonplace explorations of how inhabitants at the margins of our cities live, suffer and relate, his twenty years of research have focused on the consequences of neoliberalism in urban marginality. In light of the publication of his last book Patients of the State (2013), Salud Colectiva invited Auyero to reflect on the connections, not always observed, between waiting and political domination in government offices, schools and hospitals. His ethnographic strategy allows him to enter without prejudices into a social universe marked by polarizing political positions. He affirms that in the everyday encounters of poor people with the diverse forms of state power, practices are reproduced - not all of which are equally conscious and planned - that impart a political education and end up turning those who should be citizens into patients of the State.

Expression and significance of DNA-dependent protein kinase in human laryngeal squamous cell carcinoma / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To study the expression of DNA-dependent protein kinase (DNA-PK) in human laryngeal squamous cell carcinoma (LSCC) and normal laryngeal mucosa (NLM), and to analysize the relationship between the expression and the clinicopathologic parameters of LSCC.@*METHOD@#Immunohistochemical technique (Envision) was used to detect the expression of DNA-PK in 64 cases of LSCC and 15 cases of NLM. To investigate an investigation was conducted on the relationship between the expression and clinico-pathological features of LSCC.@*RESULT@#DNA-PK was lowly expressed in NLM and highly expressed in LSCC,the positive rate of DNA-PK expression was 26.67% (4/15), 78.13% (50/64), respectively, and there was significant different difference between the two groups (P 0.05).@*CONCLUSION@#DNA-PK may be involved in disease development of LSCC.

Inhibition of DNA-dependent protein kinase catalytic subunit by small molecule inhibitor NU7026 sensitizes human leukemic K562 cells to benzene metabolite-induced apoptosis / 华中科技大学学报（医学）（英德文版）

Benzene is an established leukotoxin and leukemogen in humans. We have previously reported that exposure of workers to benzene and to benzene metabolite hydroquinone in cultured cells induced DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to mediate the cellular response to DNA double strand break (DSB) caused by DNA-damaging metabolites. In this study, we used a new, small molecule, a selective inhibitor of DNA-PKcs, 2-(morpholin-4-yl)-benzo[h]chomen-4-one (NU7026), as a probe to analyze the molecular events and pathways in hydroquinone-induced DNA DSB repair and apoptosis. Inhibition of DNA-PKcs by NU7026 markedly potentiated the apoptotic and growth inhibitory effects of hydroquinone in proerythroid leukemic K562 cells in a dose-dependent manner. Treatment with NU7026 did not alter the production of reactive oxygen species and oxidative stress by hydroquinone but repressed the protein level of DNA-PKcs and blocked the induction of the kinase mRNA and protein expression by hydroquinone. Moreover, hydroquinone increased the phosphorylation of Akt to activate Akt, whereas co-treatment with NU7026 prevented the activation of Akt by hydroquinone. Lastly, hydroquinone and NU7026 exhibited synergistic effects on promoting apoptosis by increasing the protein levels of pro-apoptotic proteins Bax and caspase-3 but decreasing the protein expression of anti-apoptotic protein Bcl-2. Taken together, the findings reveal a central role of DNA-PKcs in hydroquinone-induced hematotoxicity in which it coordinates DNA DSB repair, cell cycle progression, and apoptosis to regulate the response to hydroquinone-induced DNA damage.

Association between Exudative Age-related Macular Degeneration and the G6721T Polymorphism of XRCC7 in Outdoor Subjects

PURPOSE: To investigate whether the G6721T polymorphism (rs.7003908) of the non-homologous end-joining DNA repair XRCC7 gene contributes to the development of exudative age-related macular degeneration (ARMD). METHODS: The present case-control study consisted of 111 patients with exudative ARMD and 112 sex frequency-matched healthy controls that were randomly selected from unrelated volunteers in the same clinic. Genotypes were determined by the Restriction Fragment Length Polymorphism (PCR-RFLP) based method. Logistic regression was used to calculate odds ratios (ORs) and 95% confidence intervals (CIs) for ARMD risk associated with polymorphism of XRCC7. In all analysis the GG genotype was considered to be the reference genotype. RESULTS: There was no significant association between genotypes of XRCC7 and susceptibility to ARMD. Considering the significant difference in age distribution between cases and controls, age was used as a covariate in further analysis. After ORs were adjusted for age, the same result was observed. In the next step we stratified our subjects into outdoor and indoor groups according to their job titles. The outdoor and indoor patients were occupationally exposed to sunlight and not exposed to sunlight, respectively. Our present study showed that among indoor subjects there was no association between XRCC7 polymorphism and susceptibility to ARMD. However, among outdoor subjects, the GT + TT genotypes compared to the GG genotype increased the risk of ARMD (OR, 3.13; 95% CI, 1.04-9.39; p = 0.042). CONCLUSIONS: Our study revealed that the T allele of the G6721T polymorphism of XRCC7 increased the risk of ARMD among outdoor subjects.

Nampt is involved in DNA double-strand break repair / 癌症

DNA double-strand break (DSB) is the most severe form of DNA damage, which is repaired mainly through high-fidelity homologous recombination (HR) or error-prone non-homologous end joining (NHEJ). Defects in the DNA damage response lead to genomic instability and ultimately predispose organs to cancer. Nicotinamide phosphoribosyltransferase (Nampt), which is involved in nicotinamide adenine dinucleotide metabolism, is overexpressed in a variety of tumors. In this report, we found that Nampt physically associated with CtIP and DNA-PKcs/Ku80, which are key factors in HR and NHEJ, respectively. Depletion of Nampt by small interfering RNA (siRNA) led to defective NHEJ-mediated DSB repair and enhanced HR-mediated repair. Furthermore, the inhibition of Nampt expression promoted proliferation of cancer cells and normal human fibroblasts and decreased β-galactosidase staining, indicating a delay in the onset of cellular senescence in normal human fibroblasts. Taken together, our results suggest that Nampt is a suppressor of HR-mediated DSB repair and an enhancer of NHEJ-mediated DSB repair, contributing to the acceleration of cellular senescence.

p53 and DNA-dependent protein kinase catalytic subunit independently function in regulating actin damage-induced tetraploid G1 arrest

We previously reported that the p53 tumor suppressor protein plays an essential role in the induction of tetraploid G1 arrest in response to perturbation of the actin cytoskeleton, termed actin damage. In this study, we investigated the role of p53, ataxia telangiectasia mutated protein (ATM), and catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) in tetraploid G1 arrest induced by actin damage. Treatment with actin-damaging agents including pectenotoxin-2 (PTX-2) increases phosphorylation of Ser-15 and Ser-37 residues of p53, but not Ser-20 residue. Knockdown of ATM and DNA-PKcs do not affect p53 phosphorylation induced by actin damage. However, while ATM knockdown does not affect tetraploid G1 arrest, knockdown of DNA-PKcs not only perturbs tetraploid G1 arrest, but also results in formation of polyploidy and induction of apoptosis. These results indicate that DNA-PKcs is essential for the maintenance of actin damage induced-tetraploid G1 arrest in a p53-independent manner. Furthermore, actin damage-induced p53 expression is not observed in cells synchronized at G1/S of the cell cycle, implying that p53 induction is due to actin damage-induced tetraploidy rather than perturbation of actin cytoskeleton. Therefore, these results suggest that p53 and DNA-PKcs independently function for tetraploid G1 arrest and preventing polyploidy formation.

Comparative analysis of two component signal transduction systems of the Lactobacillus acidophilus group

The Lactobacillus acidophilus group is a phylogenetically distinct group of closely related lactobacilli. Members of this group are considered to have probiotic properties and occupy different environmental niches. Bacteria generally sense and respond to environmental changes through two component systems (TCSs) which consist of a histidine protein kinase (HPK) and its cognate response regulator (RR). With the use of in silico techniques, the five completely sequenced L. acidophilus group genomes were scanned in order to predict TCSs. Five to nine putative TCSs encoding genes were detected in individual genomes of the L. acidophilus group. The L. acidophilus group HPKs and RRs were classified into subfamilies using the Grebe and Stock classification method. Putative TCSs were analyzed with respect to conserved domains to predict biological functions. Putative biological functions were predicted for the L. acidophilus group HPKs and RRs by comparing them with those of other microorganisms. Some of TCSs were putatively involved in a wide variety of functions which are related with probiotic ability, including tolerance to acid and bile, production of antimicrobial peptides, resistibility to the glycopeptide antibiotic vancomycin, and oxidative condition.

Roles of DNA dependent protein kinase in silica-induced cyclin E and CDK2 expressions and cell cycle changes in human embryo lung fibroblasts / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the roles of DNA dependent protein kinase (DNA-PK)in silica-induced cell cycle changes and expressions of CyclinE and CDK2 in human embryo lung fibroblasts (HELF).</p><p><b>METHODS</b>The expressions of Ku80 and DNA-PKcs proteins were inhibited by siRNA plasmids, respectively. Flow cytometry was used to detect the distributions of cell cycle and western blot assay was used to determine the expression levels of CyclinE and CDK2 after cells were exposed to 200 microg/ml silica for 0, 3, 6, 12, 24 h.</p><p><b>RESULTS</b>The proportion of G1 phases in negative control cells decreased from 83.53% +/- 2.24% to 69.11% +/- 3.12% after exposure to silica; the proportion of G1 phases in H-Ku80 and H-PKcs cells exposed to silica decreased from 85.16% +/- 3.73% to 59.92% +/- 3.31% and from 75.06% +/- 2.23% to 58.32% +/- 1.35%, respectively (P < 0.05). The exposure to silica resulted in the increasing protein expression levels of CyclinE and CDK2 in negative control cells, and the expression levels of CyclinE were obviously suppressed in H-Ku80 and H-PKcs as compared with control cells. However, the expression level of CDK2 protein did not change significantly.</p><p><b>CONCLUSION</b>DNA-PK might play a role in silica-induced alternations of cell cycle and regulate silica-induced overexpression of CyclinE in human embryo lung fibroblasts.</p>

Role of heterogeneous nuclear ribonucleoprotein A2/B1 protein in the pathogenesis of non-small cell lung cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the expression of heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) in non-small cell lung cancer (NSCLC), and the interaction between hnRNP A2/B1 protein and mRNA of DNA repair enzymes O(6)-methylguanine DNA-methyltransferase (MGMT), 8-oxoguanine DNA glycosylase (OGG1), redox factor 1(Ref-1), DNA-dependent protein kinase (including DNA-PKcs and ku).</p><p><b>METHODS</b>The expression and distribution of hnRNP A2/B1 were detected by immunohistochemistry and Western blot on 50 NSCLC samples from patients who underwent resection in Zhongshan Hospital. The hnRNP A2/B1 mRNA expression was tested by real-time PCR. Co-immunoprecipitation (co-IP) combined RT-PCR was used to investigate whether hnRNP A2/B1 could be bound with the mRNA of the above mentioned 5 DNA repair enzymes in human lung cancer cell line (HTB-182). Then immunohistochemistry and real-time PCR were used to detect the expression of MGMT in the same group of patients.</p><p><b>RESULTS</b>HnRNP A2/B1 protein and mRNA expressions were increased in the NSCLC tissues than that in the corresponding normal lung tissues. HnRNP A2/B1 was expressed predominantly in the nuclei of tumor cells. The positive rate and immunohistochemistry score of hnRNP A2/B1 in tumor tissue were significantly higher than that in normal tissue (P < 0.01). In stage III-IV NSCLC, hnRNP A2/B1 expression was higher than that in stage I-II. There was no significant differences of hnRNP A2/B1 expression among patients of different age, sex, histological type, and smoking history. The results of co-IP combined RT-PCR suggested that hnRNP A2/B1 is bound with MGMT mRNA, and MGMT expression is decreased in tumor tissue of NSCLC.</p><p><b>CONCLUSIONS</b>The results of this study show that hnRNP A2/B1 protein and mRNA are highly expressed in NSCLC, and hnRNP A2/B1 is bound with MGMT mRNA, which indicate that it might be one of the mechanisms of hnRNP A2/B1 participating in the pathogenesis of NSCLC.</p>

Expression of DNA-dependent protein kinase catalytic subunit in adult acute leukemia and its significance / 中国实验血液学杂志

This study was aimed to explore the expression of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) in adult acute leukemia and its correlation with clinical characteristics, karyotype and prognosis. Indirect immunofluorescent cytometry was used to detect the expression of DNA-PKcs in bone marrow mononuclear cells of 105 patients with acute leukemia before chemotherapy and 41 of them after 2 cycles of chemotherapy. Cytogenetic data were obtained from 26 of them by R band karyotypic analysis. The results showed that the expression of DNA-PKcs was correlated with higher WBC count level in peripheral blood (p < 0.05), but was not obviously associated with median age, gender, percentage of bone marrow blasts, clinical classification, median hemoglobin level and median platelet count (p > 0.05). The middle and strong positive expression of DNA-Pkcs in non-remission group was significantly higher than that in remission group (p < 0.05). The positive rate of DNA-PKcs in abnormal chromosome group was significantly higher than that in chromosome normal group (p < 0.05). It is concluded that the DNA-PKcs expression level is closely related with the increased WBC count, and the expression of DNA-PKcs is correlated also with karyotype and clinical prognosis in adult acute leukemia.

DNA-PKcs silencing inhibit the DNA repair induced by low dose radiation on human breast epithelial cells / 生物工程学报

DNA-PKcs, the catalytic subunit of DNA-dependent protein kinase (DNA-PK), plays an important role in the nonhomologous end-joining (NHEJ) pathway of DNA double-strand breaks (DSBs) repair. To investigate the effects of DNA-PKcs down-regulation on cell growth and sensitization to low dose radiation (LDR), we transfected DNA-PKcs siRNA into human mammary epithelia cells MCF10F, then, detected the proliferation curve of the cells and the expression of protiens in DNA repair pathways. The results showed that DNA-PKcs gene silencing, induced by the transfection of DNA-PKcs siRNA could suppress significantly the cell proliferation and inhibit the expression of the DNA repair proteins, such as Ku80, ATM and P53 after 50 cGy 137Cs gamma-irradiation.The results suggested that DNA-PKcs gene silencing could increase the sensitivity of human breast epithelial cells to the LDR, which might be relative with the decrease of the proteins.

Role of DNA-dependent protein kinase catalytic subunit in silica-induced DNA double-strand break repair in human embryo lung fibroblasts / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the role of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) in silica-induced DNA double-strand break repair in human embryo lung fibroblasts (HELF).</p><p><b>METHODS</b>Two stable transfectants, HELF transfected with DNA-PKcs siRNA (HELF-PKcs) and with negative control siRNA (HELF-NC), were established. HELF cells were treated with 0, 25, 50, 100, 200, 300 and 400 microg/ml silica for 12 h and with 200 microg/ml silica for different times (0, 1, 2, 6, 12 and 24 h). HELF-PKcs and HELF-NC were treated with 200 microg/ml silica for 0, 12 and 24 h. The expression levels of DNA-PKcs and phosphor-H2AX (H2AX) were determined by Western blot. DNA double strand breaks were measured by neutral comet assay.</p><p><b>RESULTS</b>After treatment with different doses of silica for 12 h, the levels of H2AX and the percentages of tail DNA increased in concentration-dependent manner. After treatment with 200 microg/ml silica for different times, the levels of H2AX increased in a time-dependent manner. The percentages of tail DNA increased significantly at 6 h, and reaching maximum at 12 h and then decreasing at 24 h. The expression level of DNA-PKcs was suppressed in HELF-PKcs. After treatment with silica at 12 h, the level of H2AX was lower in HELF-PKcs than in HELF-NC, and the percentages of tail DNA increased obviously in both HELF-PKcs and HELF-NC compared with non-treated cells, but no significant difference was found in the percentages of tail DNA between them. The percentages of tail DNA decreased markedly in silica-treated HELF-NC and was significantly lower than in HELF-PKcs at 24 h (P < 0.05).</p><p><b>CONCLUSION</b>Silica can induce DNA double strand breaks in human embryo lung fibroblasts. DNA-PKcs might play a major role in silica-induced DNA double strand break repair. Silica-induced histone H2AX phosphorylation was dependent on DNA-PKcs.</p>

DNA-dependent protein kinase activity and radiosensitivity of nasopharyngeal carcinoma cell lines CNE1/CNE2 / 生理学报

The present study investigated the relationship between DNA-dependent protein kinase (DNA-PK) and radiosensitivity of nasopharyngeal carcinoma (NPC) cell lines. The dose-survival relationship for NPC cell lines, CNE1 and CNE2, was analyzed using clonogenic formation assay, the activity of DNA-PK of the two cell lines was measured using the Signa TECT DNA-PK assay kit, and the localization and expression of Kus (a heterodimer) and DNA-PKcs protein in CNE1 and CNE2 before irradiation and 15 min, 1 h, 6 h, 12 h, 24 h after 4 Gy irradiation were analyzed by immunofluorescence, laser scanning confocal microscope (LSCM) and Western blot. The results showed that the surviving fraction of CNE1 was higher than that of CNE2 at each dose. The DNA-PK activity of CNE1 was also significantly higher than that of CNE2 before and after irradiation (P<0.05), while the expression of total Ku70/Ku80 in CNE1 and CNE2 had no significant difference. Increasing translocation of Ku70 and Ku80 from the cytoplasm to the nuclei in the two cell lines was observed with increase of irradiation time as detected by Western blot, and the immunofluorescence of the DNA-PK complex subunits showed greater nuclear translocation in CNE1 than CNE2 after irradiation. The results suggest that the relatively higher radio-resistance of CNE1 correlates with the higher activity of DNA-PK as compared to that of more radiosensitive CNE2 (or lower radio-resistance) before and after irradiation. Thus, DNA-PK activity may be a useful predictor of radiosensitivity of NPC.

Gene of DNA-dependent protein kinase catalylic subunit in chronic myeloid leukemia / 中国实验血液学杂志

This study was aimed to investigate the expression and regulation mechanism of DNA-dependent protein kinase catalylic subunit (DNA-PKcs) in chronic myeloid leukemia (CML) and its role in blast crisis of CML. Expression of DNA-PKcs mRNA was detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and DNA-PKcs protein by Western blot in 62 CML patients and K562, as compared to those of 23 normal individual controls. In 26 CML patients received allogeneic peripheral blood stem cell transplantation (allo-PBSCT) and 4 CML patients treated with imatinib, the expression of bcr-abl mRNA and DNA-PKcs protein was detected by RT-PCR and Western blot, respectively. After treatment with imatinib in mononuclear cell (MNC) of CML patients and K562 in vitro, expression of DNA-PKcs mRNA was detected by RT-PCR and DNA-PKcs protein level, tyrosine phosphorylation of bcr-abl fusion protein were detected by Western blot. The results showed that the expression of DNA-PKcs protein was significantly lower in CML and K562 than those in normal control (P<0.05). In 26 CML patients received allo-PBSCT and 4 CML patients treated with imatinib, the expression of DNA-PKcs protein was enhanced while the expression of bcr-abl mRNA decreased. After treatment of MNC of CML and K562 with imatinib in vitro, the expression of DNA-PKcs protein was enhanced while tyrosine phosphorylation of bcr-abl fusion protein decreased. It is concluded that the expression of DNA-PKcs protein is down-regulate by bcr-abl fusion gene, and the bcr-abl fusion gene down-regulate the expression of DNA-PKcs protein by post-transcriptional mechanism; the decrease of DNA-PKcs protein expression may be one of mechanisms underlying the acute transformation of CML.

Expression of GLUT-1, p63 and DNA-Pkcs in serous ovarian tumors and their significance / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the expression of GLUT1, p63 and DNA-Pkcs in serous ovarian tumors and their significance.</p><p><b>METHODS</b>GTUL1, p63 and DNA-Pkcs expression at protein level was detected by immunohistochemistry in patients with serous ovarian tumors. Chi-square analysis was used to assess if their expression is associated with clinicopathologic characteristics of the tumors.</p><p><b>RESULTS</b>Cells in the normal ovarian tissues were not stained with GTUL1 and p63 antiserum, but DNA-Pkcs was positively stained. The intensity of GTUT1 and p63 expression was stronger in malignant ovarian serous tumors compared with other subtypes (P < 0.01). There were significant differences of DNA-PKcs among normal ovaries (100.0%), benign (95.0%), borderline (90.0%) and malignant (60.0%) serious ovarian neoplasms (P < 0.01). The level of GLUT-1 expression was correlated with FIGO staging, intraperitoneal implantation, ascites and lymph node metastasis (P < 0.05). p63 expression was associated with clinicopathologic characteristics except ascites (P < 0.05). DNA-PKcs was only correlated with FIGO staging and lymph node metastasis (P < 0.05).</p><p><b>CONCLUSION</b>The results suggest that the abnormal expression of GTUT1, p63 and DNA-Pkcs may perhaps participate in serous ovarian tumor occurrence and development and may be considered as a marker reflecting tumor malignant behavior.</p>

Correlation of DNA-PK activity with anti-cancer drug-sensitivity in human gliomas / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the relationship between DNA-dependent protein kinase (DNA-PK) activity and anti-cancer drug sensitivity in human glioma tissues.</p><p><b>METHODS</b>Human glioma specimens were primarily cultured and its sensitivity to several anti-cancer drugs were evaluated by MTT assay. Nuclear protein was extracted from the glioma sample of the same patient and its DNA-PK activity was determined by a biotinylated DNA-PK assay with p53-derived peptide as a specific substrate.</p><p><b>RESULTS</b>DNA-PK activity varied widely among these glioma samples. Of all 36 samples, 16 showed higher DNA-PK activity (relative activity > or = 0.40) and 20 samples with lower DNA-PK activity (relative activity < 0.40). The gliomas sensitive to DDP and VCR as evaluated by inhibition rate (IR > or = 50%) under plasma peak concentration (PPC) showed lower DNA-PK activity than the resistant ones (IR < 50%) (t = -3.445, P < 0.01). Furthermore, the gliomas with higher DNA-PK activity showed lower inhibition rate (IR < 50%) than those with lower DNA-PK activity ones (t = -2.145, P < 0.05).</p><p><b>CONCLUSION</b>DNA-PK activity is significantly associated with anti-cancer drug sensitivity to DDP and VCR in human gliomas. DNA-PK activity could be used as a new biomarker for the chemotherapy sensitivity of human gliomas.</p>

Loss of DNA-dependent Protein Kinase Catalytic Subunit (DNA- PKcs) Expression in Gastric Cancers / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: DNA-PKcs is one of the DNA repair genes. It was recently found that hyperplasia and dysplasia of the intestinal mucosa and the production of aberrant crypt foci were developed in DNA-PKcs-null mice, and this suggests a suppressive role for DNA-PKcs in tumorigenesis. MATERIALS AND METHODS: To investigate the possible relationship between the clinico-pathologic characteristics and the survival of gastric cancer patients, the expression status of DNA-PKcs was determined in 279 consecutive gastric cancers. Immunohistochemical analysis was performed to evaluate the expression levels of DNA-PKcs protein by using the tissue array method. RESULTS: Out of 279 consecutive gastric cancers, 63 cases (22.6%) showed the loss of DNA-PKcs expression. The loss of DNA-PKcs expression was significantly associated with advanced cancer (p <0.001), lymphatic invasion (p=0.001), lymph node metastasis (p=0.009), and advanced pTNM stage (p=0.009). Univariate survival analysis revealed that patients with the loss of DNA-PKcs expression had significantly poorer survival than those patients with intact DNA-PKcs expression (p=0.004). Moreover, the loss of DNA-PKcs expression was identified to correlate with a lower survival in the subgroup of stage I gastric cancer patients (p=0.037). CONCLUSION: The loss of DNA-PKcs expression was found in 23% of human gastric cancers and this was identified to significantly correlate with poor patient survival, especially for stage I gastric cancer patients.

The Significance of Ku70 Expression in Patients with Nasopharyngeal Carcinoma / 대한방사선종양학회지

PURPOSE: The objective of this study was to determine whether the expressions of the two components of DNA-dependent protein kinase, Ku70 and DNA-PKcs, influence the response to radiotherapy (RT) and outcome of treatment of non-disseminated nasopharyngeal carcinoma (NPC) in patients who received definitive RT. MATERIASL AND METHODS: Sixty-six patients with NPC who were treated with radiotherapy alone or with concurrent chemotherapy between June 1995 and December 2001 were divided into groups based on the levels of immunoreactivity for Ku70 and DNA-PKcs in pretreatment biopsy specimens. The over-expression of Ku70 or DNA-PKcs groups included patients whose biopsy specimens showed at least 50% immunopositive tumor cells; patients in which less than 50% of the tumor cells in the biopsy tissues were immunopositive were placed in the low Ku70 and DNA-PKcs groups. The immunoreactivities for Ku70 and DNA-PKcs were retrospectively compared with the sensitivity of the tumor to radiation and the patterns of therapy failure. Univariate analyses were performed to determine the prognostic factors that influenced locoregional control of NPC. RESULTS: The five-year locoregional control rate was significantly higher in the low Ku70 group (Ku(-)) (85%) than in the high Ku70 group (Ku(+)) (42%) (p=0.0042). However, there were no differences in the metastases-free survival rates between the two groups (Ku70 (+), 82%; Ku70 (-), 78%; p=0.8672). Univariate analysis indicated that the over-expression of Ku70 surpassed other well-known predictive clinocopathologic parameters as an independent prognostic factor for locoregional control. Eighteen of 22 patients who had locoregional recurrences of the tumor displayed an over-expression of Ku70. No significant association was found between the level of DNA-PKcs expression and the clinical outcome. CONCLUSION: Our data suggest that the level of Ku70 expression can be used as a molecular marker to predict the response to RT and the locoregional control after RT and concurrent chemotherapy in patients with non-disseminated NPC.