Chemoprevention with the metabolism modifying drugs dichloroacetate and metformin in the Li-Fraumeni Syndrome model, Trp53+/- mice

BACKGROUND: While genetic testing for familial cancer has excelled, the prevention options for those carrying high risk alleles have not. Altered bioenergetics is now acknowledged as a hallmark of cancer, and several very safe drugs are available that can target this phentoype. Dichloroacetate (DCA) inactivates pyruvate dehydrogenase kinase, resulting in activation of pyruvate dehydrogenase, reduced lactic acid production and increased mitochondrial activity. Metformin, a type 2 diabetes treatment which activates AMPK, thereby inhibiting mTOR, has unambiguously been demonstrated to reduce the risk of many cancer types in diabetics. We have tested these drugs as chemopreventive agents against the mammary tumours that occur in the BALB/c-Trp53+/- mouse spontaneous tumour model. MATERIALS and METHODS Breast cancer cell lines were examined for cell viability after DCA and/or metformin treatment in vitro (neutral red uptake assay). Four groups of female BALB/c-Trp53+/- mice were given distilled water (n=75), DCA (1.5 g/L in drinking water, ~180 mg/ kg/day, n=53), metformin (0.25 g/L in drinking water, ~30 mg/kg/day, n=61) or DCA +metformin (n=51) from 8 weeks of age, and monitored for tumour development over 78 weeks, and Kaplan-Meier survival analysis was performed. RESULTS In vitro, DCA (1-5 mM) and metformin (30-300 uM), alone or combined, significantly inhibited breast cancer cell growth. In vivo, the overall tumour-free survival curves for BALB/c-Trp53+/- mice were not significantly different between treatment groups, suggesting that metformin does not reduce cancer risk in non-diabetics. However, analysis of mammary tumours alone found that DCA reduced the number and increased their latency (28.0% vs 20.8% of mice with mean latency of 55.0 vs 63.8 weeks, untreated vs DCA respectively), whereas metformin had no effect (26.2% of mice, mean latency 54.7 weeks). DCA appeared to eliminate the early onset mammary tumours (latency <52 weeks, p=0.02), while not affecting the occurrence of longer latency tumours. In contrast, the two drug combination had worse outcomes for tumour development, (35.3% of mice, latency 48.8 weeks, p<0.02 compared to DCA alone). Preliminary western blotting results in MDA-MB-468 breast cancer cells found that DCA could block the activation of AMPK by metformin, indicating the potential for drug interactions.Supported by NHMRC Career Development Award, National Breast Cancer Foundation Novel Concept Award, and Cancer Australia

Nuclear export factor 3 regulates the localization of small nucleolar RNAs

Metabolic diseases, such as obesity and diabetes, are associated with excess levels of lipids, which can lead to organelle dysfunction, cell death and eventually organ dysfunction. This process, termed lipotoxicity, is still not completely understood. In a genetic screen used to identify genes critical for lipotoxicity, the Schaffer lab has identified small nucleolar RNAs (snoRNAs) within the ribosomal protein L13a (Rpl13a) locus that mediate the cellular response to lipotoxic and general metabolic stress. These snoRNAs are non-canonical in that they accumulate in the cytosol after metabolic stresses like lipotoxicity and oxidative stress, suggesting that cells have specific mechanisms for regulating snoRNA distribution within the cell. To begin elucidating the underlying mechanism of snoRNA transport from the nucleus to the cytoplasm during metabolic stress, we modeled oxidative stress by treating cells with the chemotherapeutic drug doxorubicin (DOX). DOX is a potent inducer of superoxide through the activation of NADPH oxidase splice isoform 4D (NOX4D) and results in the cytosolic accumulation of Rpl13a snoRNAs. NOX inhibitors and genetic knockdown of NOX4D led to a decrease in the accumulation of Rpl13a snoRNAs in the cytosol after DOX treatment. Furthermore, RNA-sequencing studies demonstrated that snoRNAs as a class accumulated in the cytosol after DOX treatment, while simultaneous treatment with NOX inhibitors blunted the increase in cytosolic levels of snoRNAs. Together, these data indicate that snoRNAs as a class are present in the cytoplasm, where their levels are dynamically regulated by NOX4D during oxidative stress. The finding that loss-of-function of NXF3, a member of the nuclear export family of RNA transporters, protects cells from lipotoxic cell death suggested that this protein may function as a regulator of snoRNA distribution. In transient transfection assays, NXF3 knockdown increased abundance of cytosolic Rpl13a snoRNAs, whereas NXF3 overexpression led to decreased cytosolic levels. These observations suggested that snoRNAs traffic constitutively between the nucleus and cytoplasm, and NXF3 functions to efficiently concentrate snoRNAs in the nucleus. Consistent with a role for NXF3 as a snoRNA transporter, we found the Rpl13a snoRNAs co-immunoprecipitated with NXF3. Treatment of cells with forskolin caused a rapid decrease in cytosolic snoRNAs and increased both nuclear localization of NXF3 and association of NXF3 with Rpl13a snoRNAs. These effects of forskolin were abrogated by NXF3 knockdown. Our findings identify a novel trafficking pathway for snoRNAs that cycle between the nucleus and the cytosol. This pathway is regulated by oxidative stress and levels of the RNA transporter NXF3

A Mathematical Theory of the Topological Vertex

We have developed a mathematical theory of the topological vertex--a theory that was original proposed by M. Aganagic, A. Klemm, M. Marino, and C. Vafa in hep-th/0305132 on effectively computing Gromov-Witten invariants of smooth toric Calabi-Yau threefolds derived from duality between open string theory of smooth Calabi-Yau threefolds and Chern-Simons theory on three manifolds.Comment: 66 pages, 10 figures; notation simplified, references adde

Synthesising Resource-Advantage and Hofstede Cultural Dimensions: Customer Orientation, Knowledge Creation and Firm Performance

There is a dearth of studies on synthesising the resource-advantage theory and Hofstedecultural dimensions that explore the influence of customer orientation and knowledgecreation on the firm performance for small hotels. This paper reviews extant literaturethat provides insights on the influence of customer orientation and knowledge creationon firm performance. In-depth interviews with three hotel owners and three hotelmanagers were performed in this study. The results disclose that forging memorableand positive customer experiences and developing informed knowledge databases arebusiness practices that sustain the firm performance. From the theoretical aspect, it is apreliminary step to blend the culturally relevant customer orientation and knowledgecreation elements into forming a firm performance framework designed for small hotels

Preventing Firearm Suicide Among White Men Who Own Firearms in Greater Minnesota: Findings from Interviews with Firearm Owners and National Messaging Experts

Wilder Research conducted interviews with national messaging experts and white, male, firearm owners in greater Minnesota. Respondents were asked for their suggestions for trusted messengers who could share communications about firearm suicide prevention, suggestions for framing messaging and the types of content that should be shared, safe storage practices, and how they and other firearm owners would respond to a mental health crisis and the barriers that prevent people from intervening in a crisis

Hydrodynamics of impinging liquid jets used in cleaning

Impinging liquid jets are widely used in industrial cleaning-in-place (CIP) systems to remove residual product or soil layers from the internal surfaces of process equipment such as tanks and vessels. The optimisation of these cleaning operations is often done empirically as a large number of parameters are needed to define the problem. Three aspects of jet behaviour were investigated as a step towards enabling the systematic design and optimisation of CIP systems. The effect of jet length and wall curvature were explored. The flow patterns generated by the impingement of a coherent, turbulent, horizontal water jet on a flat, vertical target were characterised as a benchmark and compared with existing models that predict the shape of the radial flow zone (RFZ). As the jet length increased, some liquid was lost to splatter through jet breakup into droplets and rebound of liquid droplets off the target. The shape of the RFZ agreed with existing models once the fraction of liquid lost to splatter was accounted for. Tests on horizontal and vertical cylinders with curvatures in the range 6.9 to 20 m⁻¹ showed that wall curvature did not have a significant effect on the shape of the RFZ unless the liquid film wrapped around the inside of the cylinder, observed at high jet flow rates. There was no appreciable effect of wall curvature on cleaning behaviour. Soaking a water-soluble soil prior to cleaning increased its cleaning rate. In many CIP systems, the liquid jet impinges at an oblique angle to the target surface and an understanding of the liquid flow distribution created by inclined jets is needed to predict the cleaning behaviour. Tests with inclined jets were carried out to establish the shape of the hydraulic jump formed and their cleaning behaviour. Three flow distribution models were developed and compared with the experimental data, providing an insight into the liquid flow distribution, but further work remains to enable the liquid flow distribution to be predicted a priori. The cleaning of viscous soil layers (petroleum jelly, tomato ketchup and two toothpastes) from vertical walls by an intermittent water jet was investigated by using a moving interrupter plate to periodically disrupt the impingement of a continuous water jet. The use of intermittent jets was found to provide no improvement in the cleaning of these soil layers in the absence of soaking.PhD Studentship, Newnham College, Cambridg