CHK1 expression in Gastric Cancer is modulated by p53 and RB1/E2F1: implications in chemo/radiotherapy response.

Radiation has a limited but relevant role in the adjuvant therapy of gastric cancer (GC) patients. Since Chk1 plays a critical function in cellular response to genotoxic agents, we aimed to analyze the role of Chk1 in GC as a biomarker for radiotherapy resistance. We analyzed Chk1 expression in AGS and MKN45 human GC cell lines by RT-QPCR and WB and in a small cohort of human patient's samples. We demonstrated that Chk1 overexpression specifically increases resistance to radiation in GC cells. Accordingly, abrogation of Chk1 activity with UCN-01 and its expression with shChk1 increased sensitivity to bleomycin and radiation. Furthermore, when we assessed Chk1 expression in human samples, we found a correlation between nuclear Chk1 accumulation and a decrease in progression free survival. Moreover, using a luciferase assay we found that Chk1's expression is controlled by p53 and RB/E2F1 at the transcriptional level. Additionally, we present preliminary data suggesting a posttranscriptional regulation mechanism, involving miR-195 and miR-503, which are inversely correlated with expression of Chk1 in radioresistant cells. In conclusion, Chk1/microRNA axis is involved in resistance to radiation in GC, and suggests Chk1 as a potential tool for optimal stratification of patients susceptible to receive adjuvant radiotherapy after surgery.

Have winter fuel payments reduced excess winter mortality in England and Wales?

BACKGROUND: The historical series of excess winter mortality (EWM) in England and Wales presents a negative trend. Winter fuel payments (WFPs) are the most important benefits for people aged 65 or over directly related to Winter Mortality in the UK. METHODS: This study presents a time series analysis of the direct effect of WFPs on EWM in England and Wales. RESULTS: We find a significant structural break in trend and volatility in the EWM series in England and Wales in 1999-2000. After controlling for a number of covariates, an ARIMA-X model finds that WFPs can account for almost half of the reduction in EWM in England and Wales since 1999/2000. CONCLUSIONS: Almost half of the reduction in EWM since 1999/2000 is attributable to WFPs.

Mad2 and BubR1 modulates tumourigenesis and paclitaxel response in MKN45 gastric cancer cells.

Aneuploidy and chromosomal instability (CIN) are common features of gastric cancer (GC), but their contribution to carcinogenesis and antitumour therapy response is still poorly understood. Failures in the mitotic checkpoint induced by changes in expression levels of the spindle assembly checkpoint (SAC) proteins cause the missegregation of chromosomes in mitosis as well as aneuploidy. To evaluate the possible contribution of SAC to GC, we analyzed the expression levels of proteins of the mitotic checkpoint complex in a cohort of GC cell lines. We found that the central SAC proteins, Mad2 and BubR1, were the more prominently expressed members in disseminated GC cell lines. Silencing of Mad2 and BubR1 in MKN45 and ST2957 cells decreased their cell proliferation, migration and invasion abilities, indicating that Mad2 and BubR1 could contribute to cellular transformation and tumor progression in GC. We next evaluated whether silencing of SAC proteins could affect the response to microtubule poisons. We discovered that paclitaxel treatment increased cell survival in MKN45 cells interfered for Mad2 or BubR1 expression. However, apoptosis (assessed by caspase-3 activation, PARP proteolysis and levels of antiapoptotic Bcl 2-family members), the DNA damage response (assessed by H2Ax phosphorylation) and exit from mitosis (assessed by Cyclin B degradation and Cdk1 regulation) were activated equally between cells, independently of Mad2 or BubR1-protein levels. In contrast, we observed that the silencing of Mad2 or BubR1 in MKN45 cells showed the induction of a senescence-like phenotype accompanied by cell enlargement, increased senescence-associated ß-galactosidase activity and increased IL-6 and IL-8 expression. In addition, the senescent phenotype is highly increased after treatment with PTX, indicating that senescence could prevent tumorigenesis in GC. In conclusion, the results presented here suggest that Mad2 and BubR1 could be used as prognostic markers of tumor progression and new pharmacological targets in the treatment for GC.

Targeting Chk2 improves gastric cancer chemotherapy by impairing DNA damage repair.

Our results demonstrate that the addition of cisplatin after paclitaxel-induced mitotic arrest was more effective than individual treatment on gastric adenocarcinoma cells (MKN45). However, the treatment did not induce benefits in cells derived from lymph node metastasis (ST2957). Time-lapse microscopy revealed that cell death was caused by mitotic catastrophe and apoptosis induction, as the use of the caspase inhibitor z-VAD-fmk decreased cell death. We propose that the molecular mechanism mediating this cell fate is a slippage suffered by these cells, given that our Western blot (WB) analysis revealed premature cyclin B degradation. This resulted in the cell exiting from mitosis without undergoing DNA damage repair, as demonstrated by the strong phosphorylation of H2AX. A comet assay indicated that DNA repair was impaired, and Western blotting showed that the Chk2 protein was degraded after sequential treatment (paclitaxel-cisplatin). Based on these results, the modulation of cell death during mitosis may be an effective strategy for gastric cancer therapy.

Reaction of phenyl radical with propylene as a possible source of indene and other polycyclic aromatic hydrocarbons: an ab initio/RRKM-ME study.

Ab initio G3(MP2,CC)//B3LYP/6-311G** calculations have been performed to investigate the potential energy surface (PES) and mechanism of the reaction of phenyl radical with propylene followed by kinetic RRKM-ME calculations of rate constants and product branching ratios at various temperatures and pressures. The reaction can proceed either by direct hydrogen abstraction producing benzene and three C(3)H(5) radicals [1-propenyl (CH(3)CHCH), 2-propenyl (CH(3)CCH(2)), and allyl (CH(2)CHCH(2))] or by addition of phenyl to the CH or CH(2) units of propylene followed by rearrangements on the C(9)H(11) PES producing nine different products after H or CH(3) losses. The H abstraction channels are found to be kinetically preferable at temperatures relevant to combustion and to contribute 55-75% to the total product yield in the 1000-2000 K temperature range, with the allyl radical being the major product (~45%). The relative contributions of phenyl addition channels are calculated to be ~35% at 1000 K, decreasing to ~15% at 2000 K, with styrene + CH(3) and 3-phenylpropene + H being the major products. Collisional stabilization of C(6)H(5) + C(3)H(6) addition complexes is computed to be significant only at temperatures up to 1000-1200 K, depending on the pressure, and maximizes at low temperatures of 300-700 K reaching up to 90% of the total product yield. At T > 1200 K collisional stabilization becomes negligible, whereas the dissociation products, styrene plus methyl and 3-phenylpropene + H, account for up to 45% of the total product yield. The production of bicyclic aromatic species including indane C(9)H(10) is found to be negligible at all studied conditions indicating that the phenyl addition to propylene cannot be a source of polycyclic aromatic hydrocarbons (PAH) on the C(9)H(11) PES. Alternatively, the formation of a PAH molecule, indene C(9)H(8), can be accomplished through secondary reactions after activation of a major product of the C(6)H(5) + C(3)H(6) addition reaction, 3-phenylpropene, by direct hydrogen abstraction by small radicals, such as H, OH, CH(3), etc. It is shown that at typical combustion temperatures 77-90% of C(9)H(9) radicals formed by H-abstraction from 3-phenylpropene undergo a closure of a cyclopentene ring via low barriers and then lose a hydrogen atom producing indene. This results in 7.0-14.5% yield of indene relative to the initial C(6)H(5) + C(3)H(6) reactants within the 1000-2000 K temperature range.

Crossed beam reaction of phenyl and D5-phenyl radicals with propene and deuterated counterparts--competing atomic hydrogen and methyl loss pathways.

We conducted the crossed molecular beams reactions of the phenyl and D5-phenyl radicals with propylene together with its partially deuterated reactants at collision energies of ~45 kJ mol(-1) under single collision conditions. The scattering dynamics were found to be indirect and were mainly dictated by an addition of the phenyl radical to the sterically accessible CH(2) unit of the propylene reactant. The resulting doublet radical isomerized to multiple C(9)H(11) intermediates, which were found to be long-lived, decomposing in competing methyl group loss and atomic hydrogen loss pathways with the methyl group loss leading to styrene (C(6)H(5)C(2)H(3)) and the atomic hydrogen loss forming C(9)H(10) isomers cis/trans 1-phenylpropene (CH(3)CHCHC(6)H(5)) and 3-phenylpropene (C(6)H(5)CH(2)C(2)H(3)). Fractions of the methyl versus hydrogen loss channels of 68 ± 16% : 32 ± 10% were derived experimentally, which agrees nicely with RRKM theory. As the collision energy rises to 200 kJmol(-1), the contribution of the methyl loss channel decreases sharply to typically 25%; the decreased importance of the methyl group loss channel was also demonstrated in previous crossed beam experiments conducted at elevated collision energies of 130-193 kJ mol(-1). The presented work highlights the interesting differences of the branching ratios with rising collision energies in the reaction dynamics of phenyl radicals with unsaturated hydrocarbons related to combustion processes. The facility of forming styrene, a common molecule found in combustion against the elusiveness of forming the cyclic indane molecule demonstrates the need to continue to explore the potential surfaces through the combinative single collision experiment and electronic structure calculations.