Cell cycle progression or translation control is not essential for vesicular stomatitis virus oncolysis of hepatocellular carcinoma.

The intrinsic oncolytic specificity of vesicular stomatitis virus (VSV) is currently being exploited to develop alternative therapeutic strategies for hepatocellular carcinoma (HCC). Identifying key regulators in diverse transduction pathways that define VSV oncolysis in cancer cells represents a fundamental prerequisite to engineering more effective oncolytic viral vectors and adjusting combination therapies. After having identified defects in the signalling cascade of type I interferon induction, responsible for attenuated antiviral responses in human HCC cell lines, we have now investigated the role of cell proliferation and translation initiation. Cell cycle progression and translation initiation factors eIF4E and eIF2Bepsilon have been recently identified as key regulators of VSV permissiveness in T-lymphocytes and immortalized mouse embryonic fibroblasts, respectively. Here, we show that in HCC, decrease of cell proliferation by cell cycle inhibitors or siRNA-mediated reduction of G(1) cyclin-dependent kinase activities (CDK4) or cyclin D1 protein expression, do not significantly alter viral growth. Additionally, we demonstrate that translation initiation factors eIF4E and eIF2Bepsilon are negligible in sustaining VSV replication in HCC. Taken together, these results indicate that cellular proliferation and the initiation phase of cellular protein synthesis are not essential for successful VSV oncolysis of HCC. Moreover, our observations indicate the importance of cell-type specificity for VSV oncolysis, an important aspect to be considered in virotherapy applications in the future

GENDER DIFFERENCES IN PHYSICAL ACTIVITY AMONG THE UNIVERSITY STUDENTS IN THE VISEGRAD (V4) COUNTRIES

Introduction. Sedentary way of life has become a global phenomenon in the past decades. Therefore, the number of people with excess weight has doubled in the past 30 years. Besides this fact, it has been justified that more than half of the population is overweight. Even young adults are affected by the problem. It is an important issue because 60% of the overweight young people keep their excess body weight in later adulthood increasing the risk of different diseases. Material and method. Our study aims at assessing the differences between the health status and the physical activity among young people (secondary school and university students) in the Visegrad (V4) countries. Our current research examines the differences in the physical activity among university students regarding their sexes (n=2237). SPSS 22.00 software was used for statistical analysis. Results. According to the results, we found significant differences (p0.05). In Slovakia, we found significant differences between sexes in total MET/week and walking activities (MET/week) (p<0.001), thus, female students were found to be more active than males caused by the higher rates of walking activities of women. Conclusion. The V4 countries are not in an advantageous situation concerning physical activity in the European framework because only 21-35% of the population does sports once a week. According to our results, university students show a more positive picture on physical activity than the adult population. However, there are some specific risk groups. 43.8% of female and 57.3% male students can be considered as persons with high physical activity. Our findings may play a major role in the development of intervention programs targeting young people and in the concern of the differences between sexes. Furthermore, these results may call young people’s attention to health maintenance to preserve their fitness for getting better activity figures

Revisionsutskott - påverkas revisorns oberoende?

Bakgrund och problem: Revision har fått en ökad betydelse efter flera bolagsskandaler som har orsakat en stor brist på förtroende för bolagsstyrning och finansiell rapportering. Reaktioner till följd av dessa skandaler och krav från den internationella kapitalmarknaden har lett till implementeringen av Svensk Kod för Bolagsstyrning. Syftet med Koden är att bidra till förbättrad styrning av svenska bolag och därigenom öka förtroendet för svenskt näringsliv. Ett av kraven i Koden är att vissa noterade svenska företag skall införa revisionsutskott. Syftet med dessa utskott är att öka kommunikationen mellan revisorn och styrelsen samt att öka fokus på redovisnings- och revisionsfrågor. Inom utskottens ansvarsområde ligger även att fastställa riktlinjer för vilka andra tjänster än revision som bolaget får använda sina revisorer till. Syfte: Vi vill med denna uppsats undersöka om revisorns oberoende kommer att påverkas av revisionsutskotten och i så fall på vilket sätt. Avgränsningar: Respondenter av intresse för denna uppsats har varit revisorer på någon av de fyra stora revisionsbyråerna samt sakkunniga inom det behandlade ämnet. Kodens effekter för näringslivet i övrigt kommer inte att behandlas, utan endast de avsnitt som berör revisionsutskotten och revisorns oberoende. Oberoendet som utreds är i första hand oberoendet gentemot revisionsutskott och i andra hand oberoendet gentemot företagsledning och styrelse. Metod: Empirin för denna studie baseras på kvalitativ datainsamling genom intervjuer med revisorer och sakkunniga. Koden har varit en viktig källa för att förstå revisionsutskottens kontext och syfte. Tonvikten i denna uppsats ligger på den deskriptiva undersökningsansatsen även om vissa element kan sägas vara explorativa. Analys och slutsatser: Revisionsutskotten kommer främst att ha en positiv effekt på revisorns oberoende förutsatt att en praxis för utskotten upparbetas samt att ledamöterna i utskotten besitter den kompetens inom redovisning och revision som krävs. En annan slutsats är att ordet oberoende inte är lämpligt för att definiera förhållandet mellan revisorn och dennes uppdragsgivare då revisorn alltid kommer att vara beroende av lön eller motsvarande och då det är ett uppdragsförhållande. Förslag till fortsatt forskning: Det vore intressant att göra om uppsatsen när Koden varit igång ett tag. Vidare vore ett annat ämne att undersöka skillnaderna mellan revisorer och sakkunniga vad gäller synen på oberoende

Abstract 5302: Early prediction of response to therapy in malignant lymphoma using non-invasive [18F]FLT-PET imaging from animal to human model

Abstract Purpose: Preclinical and clinical studies have recently shown that the thymidine analogue [18F]fluorothymidine (FLT) accumulates in tumor tissue and enables non-invasive imaging of tumor proliferation, using positron emission tomography (PET). The aim of this study is to establish FLT-PET as an adequate and robust surrogate marker for very early response assessment to therapy in malignant Lymphoma. Methods: In animal models we assessed the thymidine metabolism in SCID mice bearing SUDHL-1 or Karpas 299 lymphoma prior to and early in the course of therapy with Hsp90 inhibitor NVP-AUY922 or mTOR inhibitor RAD001, using a small animal PET system. Tumor-to-background ratios (TBR) of FLT-PET were compared to that of FDG-PET. PET findings were correlated with histopathology and in vitro data comprising cellular tracer uptake, cell cycle related protein expression, cell cycle distribution and viability assessment. In human models, FLT-PET was performed additionally to routine staging procedures in 66 patients with malignant lymphoma undergoing R-CHOP chemotherapy. FLT-uptake was correlated with response and survival. Results: SUDHL-1 cells proved to be sensitive to both inhibitors whereas Karpas299 showed relative resistance to NVP-AUY922. Tumor volume in treated animals bearing SUDHL-1 lymphomas showed modest increase within the first week (median increase = + 25%, range -30% to + 80%, n=8) as opposed to a 3.8-fold increase in control mice. After 14 days a clear reduction of tumor mass was observed (median = – 25%, range -40% to + 30%, n=4). Median TBR of FLT-PET decreased significantly to 40% compared to baseline as early as 5d after therapy (range 32-67%, n=8, p=0,008). In contrast, TBR in FDG-PET did not show any clear tendency (median TBR 79%, range 36%-161%, n=8, p=0.73). According to our in vitro results, less effect was seen during treatment with AUY299 resulting in an increase of median TBR in FLT-PET to 154% (p=0,008, n=11) on day 2. In contrast, mice receiving RAD001 showed a stronger inhibition of tumor development and early FLT-PET imaging indicated a decrease of TBR to 78% (n=9, p=0.008). In human data, the initial SUVmesn in complete responders was significantly lower than in patients with progressive disease or partial response (p=0.049). Conclusion: FLT-PET has a high specificity for the detection of malignant lymphomas, both in animal xenotransplants model and in human disease. Compared with FDG-PET, FLT-PET is able to predict response to specific inhibitors treatment very early in the therapeutic course. Our human data indicate that high initial FLT uptake is a negative predictor of response to R-CHOP treatment in aggressive B-cell NHL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5302. doi:10.1158/1538-7445.AM2011-5302</jats:p

[18F]FLT Is Superior to [18F]FDG to Early Predict Response to Specific Inhibitors of NPM-ALK-Dependent Pathways In a Human ALCL Xenograft Model

Abstract Abstract 2849 Purpose: The thymidine analogue [18F]fluorothymidine (FLT) has been shown to reflect proliferation of high-grade lymphoma cells both in preclinical and clinical studies. In this preclinical in vitro and in vivo study we assessed early FLT-uptake as an adequate and robust surrogate marker for response to inhibitors of Nucleophosmin-anaplastic lymphoma kinase (NPM-ALK)-dependent pathways in an anaplastic large cell lymphoma (ALCL) xenotransplant model. Methods: In vitro investigations included viability assessment (MTT assay), cell cycle analysis using propidium iodide staining and western blotting to characterize response of the ALCL cell lines SUDHL-1 and Karpas299 to treatment with heat shock protein 90 (Hsp90) inhibitor NVP-AUY922, the Phosphoinositide 3-kinase (PI3K) inhibitor BGT226 or the mammalian target of rapamycin (mTOR) inhibitor RAD001. Thymidine metabolism in severe combined immunodeficient (SCID) mice bearing SUDHL-1 or Karpas 299 lymphoma xenotransplants was assessed non-invasively prior to and early in the course of therapy (48h to 7 days) by FLT and FDG positron emission tomography (FLT-PET and FDG-PET) using a dedicated small animal PET system. Tumor-to-background ratios (TBR) of FLT-PET were compared to that of PET using the standard radiotracer [18F]fluorodeoxyglucose (FDG). Reference for tumor response was local control of the tumor measured by shifting calliper and histopathological analysis of explanted lymphomas. Results: In vitro, SUDHL-1 cells were sensitive to all three inhibitors (IC50 AUY922= 50 nM; IC50 BGT266= 10 nM; IC50 RAD001= 1 nM). These cells showed a dose-dependent induction of cell-cycle arrest in G1-phase and reduction of S-Phase after 24 to 48 hours and - to a lesser extent - increase of apoptosis. Incubation of SUDHL-1 cells with NVP-AUY922 (50 nM) for 24 hours led to a 70% reduction of ALK level and a abrogation of Akt phosphorylation as determined by western blot analysis. Likewise, no phosphorylation of Akt was detectable after incubation with BGT266 (10 nM) already after 4 hours. RAD001 (0.1-1nM, 24h) completely inhibited phosphorylation of p70 S6K. In contrast, Karpas299 cells were only sensitive to RAD001-induced cell cycle arrest, but insensitive to NVP-AUY922 and BGT266. In vivo, we performed FLT- and FDG-PET scans to monitor inhibition of tumor growth in the course of therapy with NVP-AUY922. Tumor volume in treated animals bearing SUDHL-1 lymphomas showed modest increase within the first week (median increase= + 25%, range -30% to + 80%, n=8) as opposed to a 3.8-fold increase in untreated control animals. After 14 days a clear reduction of tumor mass could be observed (median= - 25%, range -40% to + 30%, n=4). Median TBR of FLT-PET decreased significantly to 40% compared to baseline as earlier as 5 days after initiation of therapy (range 32–67%, n=8, p=0,008). In contrast, the pattern of TBR in FDG-PET did not show any clear tendency (median TBR 79%, range 36%-161%, n=8, p=0,73). We then investigated the ability of FLT-PET to differentiate between sensitive and resistant lymphoma cells. Therefore, mice bearing Karpas299 lymphomas were treated with NVP-AUY922 (resistant in vitro) or RAD001 (sensitive in vitro). According to our in vitro results, no effect was seen during treatment with NVP-AUY299 as indicated by about 3-fold tumor growth on day 7 and increase of median TBR in FLT-PET to 162% (range 106–177%, p=0,008, n=8) on day 2. In contrast, mice receiving RAD001 showed a deceleration of tumor development with doubling of tumor volume within the first week (range -20% to + 320%, n=10) that remained fairly constant over the following weeks. FLT-PET imaging indicated a slight increase of TBR correctly reflecting tumor growth kinetics (median=126%, range 60–129%, no p-value). A larger cohort is currently investigated as well as histopathological analysis of explanted lymphomas. The updated data will be presented at the meeting. Conclusion: In contrast to FDG-PET, FLT-PET is able to predict response to specific inhibitors early in the course of the therapy using a anaplastic large cell lymphoma xenograft model and is able to distinguish between sensitive and resistant lymphoma cells. Disclosures: No relevant conflicts of interest to declare. </jats:sec

PHYSICAL ACTIVITY PATTERNS AMONG HIGH SCHOOL AND UNIVERSITY STUDENTS IN THE V4 COUNTRIES: A COMPARATIVE STUDY

Insufficient levels of physical activity are a significant contributor to chronic noncommunicable diseases such as diabetes, obesity, and cardiovascular disease. Recommendations from the World Health Organization include both general guidelines and activity recommendations specific to age, pregnancy, disease, and disability to help maintain good health and fitness. Assessing physical activity levels, as well as researching its correlates and determinants, is essential for understanding why people are physically active or inactive. This, in turn, contributes to evidencebased planning of public health interventions

Cell cycle inhibitors.

<p>A broader range of concentration was tested until the appearance of cytotoxic effects. HCC cells (HepG2 and Huh-7) and immortalized human hepatocytes (PH5CH8) were mock-treated (DMSO) or treated with increasing concentrations of cell-cycle inhibitors: Roscovitine; Ly294002 and aphidicolin. Cultures were infected with VSV-wt at an MOI of 0.1 and viral titers were determined 24 hr post-infection by TCID 50. Data represent the average of 3 independent experiments.</p

Cell cycle arrest by siRNA.

<p><b>A)</b> Cells were transfected without siRNA, with scramble siRNA (scramb) or with siRNA against cyclin D1 or cyclin-kinase (CDK4). Forty-eight hours post-transfection cells were infected with VSV-wt at an MOI of 0.1 for 24 hours. Results show the average of at least three independent experiments. <b>B)</b> Mock-infected lysates from PH5CH8 and Huh-7 cells of the experiment describe above are shown for the expression of cyclin D1 and CDK4. <b>C)</b> FACS analysis of cell cycle arrest in Huh-7 cells upon treatment with siRNA targeting CDK4 and cyclin D1 or control siRNA (SCR). Experiments were conducted at least three times and triplicate values of one experiment are shown as representative (p<0.001).</p