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Various c-mesenchymal-to-epithelial transition (c-MET) inhibitors are effective in the treatment of non-small cell lung cancer; however, the inevitable drug resistance remains a challenge, limiting their clinical efficacy. Therefore, novel strategies targeting c-MET are urgently required. Herein, through rational structure optimization, we obtained novel exceptionally potent and orally active c-MET proteolysis targeting chimeras (PROTACs) namely D10 and D15 based on thalidomide and tepotinib. D10 and D15 inhibited cell growth with low nanomolar IC50 values and achieved picomolar DC50 values and >99% of maximum degradation (Dmax) in EBC-1 and Hs746T cells. Mechanistically, D10 and D15 dramatically induced cell apoptosis, G1 cell cycle arrest and inhibited cell migration and invasion. Notably, intraperitoneal administration of D10 and D15 significantly inhibited tumor growth in the EBC-1 xenograft model and oral administration of D15 induced approximately complete tumor suppression in the Hs746T xenograft model with well-tolerated dose-schedules. Furthermore, D10 and D15 exerted significant anti-tumor effect in cells with c-METY1230H and c-METD1228N mutations, which are resistant to tepotinib in clinic. These findings demonstrated that D10 and D15 could serve as candidates for the treatment of tumors with MET alterations.
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Pediatric solid tumors are mainly mother cell-derived tumors. It is more common in children under the age of 15 and is the main cause of non-accidental death in children. Pediatric solid tumors exist with varying epidemiology. The risk factors that have already been studied regarding to its onset are birth weight, birth order, age of parents at the time of pregnancy, parental smoking, assisted reproduction, abnormal birth, economic status, maternal education, and environmental factors. This paper reviews the current research status of epidemic factors in pediatric solid tumors.
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Objective To clinically validate the precision of diagnostic Sepsis-3 criteria, and to guide and generalize its clinical application.Methods A multicenter retrospective observational study was conducted. The patients admitted to intensive care unit (ICU) of 6 tertiary hospitals in Zhejiang Province from January to June 2015 were enrolled, and the patients satisfying the diagnostic criteria of Sepsis-2 and Sepsis-3 were screened. Population characteristics between the patients satisfying two editions were compared, and the diagnosis accuracy rate in different degree hospitals were investigated. According to the doctor's diagnosis, the patients who met the criteria of Sepsis-2 were divided into diagnosis group and non-diagnosis group, and the factors influencing the diagnosis of sepsis were analyzed by logistic regression. The patients meeting Sepsis-2 but no meeting Sepsis-3 were served as exclusion group, and those meeting Sepsis-2 and Sepsis-3 were served as enroll group, and the characteristics of patients between the two groups were compared. Receiver operating characteristic (ROC) curve was plotted to evaluate the predictive value of systemic inflammatory response syndrome (SIRS) score, sepsis-related quick sequential organ failure assessment (qSOFA) and sequential organ failure assessment (SOFA) on death, and whether the consistency of qSOFA and SOFA would affect the sensitivity of definition. The patients meeting Sepsis-2 were divided into non-survived group and survived group, and the factors associated with death were analyzed by logistic regression.Results Finally, 1423 patients were enrolled, 3 patients with age 0.05) except for acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ) score [19.10 (8.00) vs. 20.28 (8.00),P 0.05], longer length of ICU stay [days: 22.42 (22.00) vs. 15.13 (16.00),P < 0.01], and higher 28-day mortality [45.29% (149/329) vs. 14.71% (5/34),P < 0.01], indicating that the diagnostic efficiency of Sepsis-2 was low, the diagnostic specificity of Sepsis-3 was high, and the prognosis of Sepsis-3 patients was worse. It was shown by ROC curve analysis that the prognostic value of SIRS, qSOFA and SOFA to mortality was gradually increased [area under ROC curve (AUC) was 0.567, 0.597, 0.683, respectively], but the prognostic value were all low. Comparing patients meeting qSOFA and (or) SOFA in Sepsis-2, significant differences were found in APACHE Ⅱ score [17.55 (7.00) vs. 23.24 (8.00)] and 28-day mortality [38.75% (31/80) vs. 58.59% (75/128), bothP < 0.01]. The patients who just met the qSOFA or SOFA, their 28-day mortality was up to 38.75%, suggesting that qSOFA should not be ignored. Compared with survived group, the patients in survived group were older with higher APACHE Ⅱ score and shorter length of ICU stay (allP < 0.05). It was shown by logistic regression analysis that APACHE Ⅱ score (OR = 1.199,P = 0.000) and length of ICU stay (OR = 0.949,P = 0.000) were related with death.Conclusion Patients satisfied Sepsis-3 were easier to develop more organ failure, Sepsis-3 and higher death prediction than Sepsis-2 and higher diagnosis specificity, but data shows that there is extra room for improvement.
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Objective To construct the luciferase report vector carrying a disintegrin and metalloprotease 10(adam10) gene promoter,to screen its stable expression cell line and to analyze its activity.Methods The genome DNA of human neuroblastoma SH-SY5Y cells was extracted as the template.The adam10 gene promoter was amplified by PCR and was cloned into luciferase reporter vector pGL4.17.The adam10 gene promoter luciferase reporter vector pGL4.17-adam10 was constructed and transfected in to SH-SY5Y cells(pGL4.17 vector without promotoer as the negative control and pGL4.17 vector with CMV promoter as the positive control).Then the stable expression cell line was screened by G418 and its fluorescence activity was detect after treating with 1 tμmol/L retinoic acid(RA) for 4 d.Results About 438 bp adam10 gene promoter was successfully amplified by PCR.The pGL4.17-adam10 vector was correct by pCR and double enzyme digestion identification.The cell line stably expressing adam10 gene promoter was obtained after transfecting SH-SY5Y cells by this vector and screening by G418,which had stronger transcriptional activity by detection;1 μmol/L RA could induce high efficiency expression of adam10 gene promoter.Conclusion Human adam10 gene promoter luciferase vector is successfully constructed.adam10 gene promoter can be stably expressed in SH-SY5Y cells,which provides a basis for deeply studying adam10 gene expression regulation,polymorphism analysis and high-throughput drug screening.
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Objective To construct the luciferase report vector carrying a disintegrin and metalloprotease 10(adam10) gene promoter,to screen its stable expression cell line and to analyze its activity.Methods The genome DNA of human neuroblastoma SH-SY5Y cells was extracted as the template.The adam10 gene promoter was amplified by PCR and was cloned into luciferase reporter vector pGL4.17.The adam10 gene promoter luciferase reporter vector pGL4.17-adam10 was constructed and transfected in to SH-SY5Y cells(pGL4.17 vector without promotoer as the negative control and pGL4.17 vector with CMV promoter as the positive control).Then the stable expression cell line was screened by G418 and its fluorescence activity was detect after treating with 1 tμmol/L retinoic acid(RA) for 4 d.Results About 438 bp adam10 gene promoter was successfully amplified by PCR.The pGL4.17-adam10 vector was correct by pCR and double enzyme digestion identification.The cell line stably expressing adam10 gene promoter was obtained after transfecting SH-SY5Y cells by this vector and screening by G418,which had stronger transcriptional activity by detection;1 μmol/L RA could induce high efficiency expression of adam10 gene promoter.Conclusion Human adam10 gene promoter luciferase vector is successfully constructed.adam10 gene promoter can be stably expressed in SH-SY5Y cells,which provides a basis for deeply studying adam10 gene expression regulation,polymorphism analysis and high-throughput drug screening.
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<p><b>OBJECTIVE</b>To evaluate the gene therapeutic efficiency of apoptosis-inducing ligand (TRAIL) related to tumor necrosis factor on human colon cancer cell line HT29.</p><p><b>METHODS</b>Human colon cancer cell line HT29 was transfected with adenovirus-mediated TRAIL gene Ad/GT-TRAIL. The morphological changes, cell growth and apoptosis were measured by phase contrast microscope, MTT method and flow cytometry.</p><p><b>RESULTS</b>Obvious morphological changes in HT29 cells was induced by Ad/GT-TRAIL and Ad/PGK-GV16. The cell suppression percentage and the percentage of apoptotic cells were 54.3% and 11.1%, respectively. When used in combination with Ad/PGK-GV16, HT29 was suppressed to 82.7% and the percentage of apoptotic cells was 24.6%. This result showed significantly enhanced therapeutic efficiency on HT29 and thus inhibiting of its growth (P < 0.05).</p><p><b>CONCLUSION</b>Ad/GT-TRAIL is able to induce apoptosis of HT29 and inhibit its growth. Ad/GT-TRAIL shows significantly enhanced therapeutic efficiency for HT29 when used in combination with Ad/PGK-GV16.</p>
Subject(s)
Humans , Adenoviridae , Genetics , Apoptosis , Genetics , Physiology , Apoptosis Regulatory Proteins , Cell Division , Genetics , Physiology , Colonic Neoplasms , Genetics , Pathology , Therapeutics , Genetic Therapy , Methods , Genetic Vectors , Genetics , HT29 Cells , Membrane Glycoproteins , Genetics , Physiology , TNF-Related Apoptosis-Inducing Ligand , Transfection , Tumor Necrosis Factor-alpha , Genetics , PhysiologyABSTRACT
Objective To study the bystander effect of tumor necrosis factor related apoptosis inducing ligand(TRAIL) gene and it's mechanism. Methods Full length cDNA of human TRAIL was transfected into SMMC7721 with binary adenoviral vectors system. RT PCR was used to determine the expression of TRAIL gene. By MTT assay the effects of the TRAIL gene on the proliferation of SMMC7721 was studied; The apoptosis inducing ability of TRAIL gene on SMMC7721 was tested by fluorescence activated cell sorting (FACS). Bystander effect by testing the proliferation of the mixed cells of SMMC7721/TRAIL and SMMC7721 with different ratios, and the mechanism of bystander effect by testing the proliferation of SMMC7721 cultured with media of SMMC7721/TRAIL without cell components were also studied. Results TRAIL gene expressed by binary adenoviral vector system was able to inhibit proliferation(91.2%) and induce apoptosis(29.07%) of SMMC7721, significant difference between TRAIL gene and the other three controls were observed(PBS, LacZ, Bax)( P
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Objective To evaluate the expression and activity of GFP/TRAIL gene activated by the hTERT promoter on colon cancer cell line DLD-1. MethodsGFP/TRAIL gene activated by the hTERT promoter was transfected into DLD-1 with adenoviral vector,expression and apoptosis inducing ability of GFP/TRAIL protein was determined by fluorescence-activated cell sorting (FACS). [WT5”HZ]ResultsThe expression of GFP gene is 41.63% and 54.07% with either hTERT promoter or CMV promoter in DLD1 cells;GFP/TRAIL gene was able to inhibit cell growth(93.50%) and induce apoptosis(56.97%) of DLD-1 ,there was significant difference between Ad/hTERT-gTRAIL and the other two control groups (PBS and Ad/hTERT-LacZ)( P
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Objective The purpose of this study was to investigate the role of p38 mitogen-activated protein kinase ( p38 MAPK) in the lung injury induced by mechanical ventilation.Methods Fifteen healthy 80 day-old pigs weighing (22.5 ? 1.5)kg were randomly divided into three groups according to the tidal volume(VT) and PEEP of mechanical ventilation: group A (VT = 16ml?kg-1, PEEP = 0) ; group B (VT = 6 ml?kg-1, PEEP= 16cm H2O) and group C(VT = 16ml?kg-1, PEEP = 8cm H2O). The animals were mechanically ventilated for 3h, then sacrificed by exsanguination. Right lower lobe was immediately removed for identification of intercellular adhesion molecule-1 ( ICAM-1 ) expression using immunohistological technique, determination of phosphorylated p38 MAPK content using Western Blot and microscopic examination. Results There was significant histological changes in the lung tissue in group A and B, but no significant histological changes were found in group C. The expression of ICAM-1 was positive in the lung in group A and B but negative in group C. The level of phosphorylated p38 MAPK among the 3 groups. Conclusion Acute lung injury can be induced by mechanical ventilation with high tidal volume or low tidal volume plus high PEEP, p38 MAPK may mediate the inflammatory response-induced lung injury.
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Objective To investigate the expression of double minutes (DM) in bladder transitional cell carcinoma (TCC) and its significance. Methods 15 control individuals and 14 patients were examined. Whole blood samples from these patients were cultured before and 10-15 days after operations. The relationship between expression of DM and clinic-pathologic factors and prognosis was ana- lyzed by chi-square test. Results There is no DM in control individuals, but there are respectively 35 pairs and 31 pairs DM in preop- erative and optoperative patients with TCC (P 0. 05). Conclusions DM may be an independent prognosis indication of cancer gene amplication. DM always show treatment failure and rapid recurrence.
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AIM: To explore the influence of IFN-? on the role of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in inducing apoptosis of RKO cell line. METHODS: Survival fraction and apoptosis were measured by MTT method and flow cytometry (FACS). RESULTS: Survival fraction of IFN-? group, TRAIL and IFN-? 72 h+TRAIL group were 99.28%, 85.45%, 52.60%, respectively. The percentage of apoptotic cells of IFN-? group, TRAIL group, IFN-? 24 h+TRAIL group, IFN-? 48 h+TRAIL and IFN-? 72 h+TRAIL group were 1.51%, 2.38%, 4.97%, 13.30%, 21.00%, respectively. The percentage of apoptotic cells of IFN-? 24 h+TRAIL group was higher than the sum of IFN-? group and TRAIL group (P