A preclinical evaluation of the PI3K alpha/delta dominant inhibitor BAY 80-6946 in HER2-positive breast cancer models with acquired resistance to the HER2-targeted therapies trastuzumab and lapatinib.

The PI3K pathway is a key mechanism of trastuzumab resistance, but early attempts to indirectly target this pathway with mTOR inhibitors have had limited success. We present the results of a preclinical study of the selective alpha/delta isoform dominant PI3K inhibitor BAY 80-6946 tested alone and in combination with HER2-targeted therapies in HER2-positive cell lines, including models with acquired resistance to trastuzumab and/or lapatinib. A panel of HER2-positive breast cancer cells were profiled for their mutational status using Sequenom MassARRAY, PTEN status by Western blot, and anti-proliferative response to BAY 80-6946 alone and in combination with the HER2-targeted therapies trastuzumab, lapatinib and afatinib. Reverse phase protein array was used to determine the effect of BAY 80-6946 on expression and phosphorylation of 68 proteins including members of the PI3K and MAPK pathways. The Boyden chamber method was used to determine if BAY 80-6946 affected cellular invasion and migration. BAY 80-6946 has anti-proliferative and anti-invasive effects when used alone in our panel of cell lines (IC50s 3.9-29.4 nM). BAY 80-6946 inhibited PI3K signalling and was effective in cells regardless of their PI3K, P53 or PTEN status. The combination of HER2-targeted therapies and BAY 80-6946 inhibited growth more effectively than either therapy used alone (with clear synergism in many cases), and can restore sensitivity to trastuzumab and lapatinib in cells with acquired resistance to either trastuzumab and/or lapatinib. The addition of BAY 80-6946 to HER2-targeted therapy could represent an improved treatment strategy for patients with refractory metastatic HER2-positive breast cancer, and should be considered for clinical trial evaluation

Bi-directional gene set enrichment and canonical correlation analysis identify key diet-sensitive pathways and biomarkers of metabolic syndrome

peer-reviewedBackground Currently, a number of bioinformatics methods are available to generate appropriate lists of genes from a microarray experiment. While these lists represent an accurate primary analysis of the data, fewer options exist to contextualise those lists. The development and validation of such methods is crucial to the wider application of microarray technology in the clinical setting. Two key challenges in clinical bioinformatics involve appropriate statistical modelling of dynamic transcriptomic changes, and extraction of clinically relevant meaning from very large datasets. Results Here, we apply an approach to gene set enrichment analysis that allows for detection of bi-directional enrichment within a gene set. Furthermore, we apply canonical correlation analysis and Fisher's exact test, using plasma marker data with known clinical relevance to aid identification of the most important gene and pathway changes in our transcriptomic dataset. After a 28-day dietary intervention with high-CLA beef, a range of plasma markers indicated a marked improvement in the metabolic health of genetically obese mice. Tissue transcriptomic profiles indicated that the effects were most dramatic in liver (1270 genes significantly changed; p < 0.05), followed by muscle (601 genes) and adipose (16 genes). Results from modified GSEA showed that the high-CLA beef diet affected diverse biological processes across the three tissues, and that the majority of pathway changes reached significance only with the bi-directional test. Combining the liver tissue microarray results with plasma marker data revealed 110 CLA-sensitive genes showing strong canonical correlation with one or more plasma markers of metabolic health, and 9 significantly overrepresented pathways among this set; each of these pathways was also significantly changed by the high-CLA diet. Closer inspection of two of these pathways - selenoamino acid metabolism and steroid biosynthesis - illustrated clear diet-sensitive changes in constituent genes, as well as strong correlations between gene expression and plasma markers of metabolic syndrome independent of the dietary effect. Conclusion Bi-directional gene set enrichment analysis more accurately reflects dynamic regulatory behaviour in biochemical pathways, and as such highlighted biologically relevant changes that were not detected using a traditional approach. In such cases where transcriptomic response to treatment is exceptionally large, canonical correlation analysis in conjunction with Fisher's exact test highlights the subset of pathways showing strongest correlation with the clinical markers of interest. In this case, we have identified selenoamino acid metabolism and steroid biosynthesis as key pathways mediating the observed relationship between metabolic health and high-CLA beef. These results indicate that this type of analysis has the potential to generate novel transcriptome-based biomarkers of disease.Department of Agriculture and Food, Ireland - Food Institutional Research Measure (project no. 5254); IRCSET postgraduate scholarship scheme (MJM); Science Foundation Ireland Principal Investigator Programme (HMR) Programme

Transcriptomic Coordination in the Human Metabolic Network Reveals Links between n-3 Fat Intake, Adipose Tissue Gene Expression and Metabolic Health

Understanding the molecular link between diet and health is a key goal in nutritional systems biology. As an alternative to pathway analysis, we have developed a joint multivariate and network-based approach to analysis of a dataset of habitual dietary records, adipose tissue transcriptomics and comprehensive plasma marker profiles from human volunteers with the Metabolic Syndrome. With this approach we identified prominent co-expressed sub-networks in the global metabolic network, which showed correlated expression with habitual n-3 PUFA intake and urinary levels of the oxidative stress marker 8-iso-PGF2α. These sub-networks illustrated inherent cross-talk between distinct metabolic pathways, such as between triglyceride metabolism and production of lipid signalling molecules. In a parallel promoter analysis, we identified several adipogenic transcription factors as potential transcriptional regulators associated with habitual n-3 PUFA intake. Our results illustrate advantages of network-based analysis, and generate novel hypotheses on the transcriptomic link between habitual n-3 PUFA intake, adipose tissue function and oxidative stress

Inhibition of the IGF signalling pathway in MDA-MB-231 triple-negative breast cancer cells

Introduction: Triple-negative breast cancer (TNBC) is characterised by the absence of oestrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER-2) on malignant cells. Insulin-like growth factors (IGFs) stimulate cell proliferation and promote cell survival in TNBC via receptor phosphorylation and activation of adaptor proteins. The aim of this project is to characterise the expression and activation of the IGF signalling pathway in a TNBC cell line, namely MDA-MB-231. Methods: Expression of oestrogen, progesterone and growth hormone receptors and activation of the IGF signalling pathway in MDA-MB-231 cells was analysed by western blotting. The effect of stimulation with IGF1 or inhibition of epidermal growth factor receptor (EGFR)/IGF1R tyrosine kinase activity on proliferation was assessed using an MTS cell proliferation assay. Proliferation was expressed relative to untreated controls, and data was analysed by ANOVA with Tukey’s multiple comparison post hoc test. Results: MDA-MB-231 cells express EGFR and high levels of insulin-like growth factor binding protein 4 (IGFBP4). Moreover, MDA-MB-231 cells express type I IGF1 receptors and proteins in the IGF signalling cascade, namely Erk and Akt. The presence of phosphorylated forms of these proteins suggests activation of the IGF1R signal transduction pathway in MDA-MB-231 cells. Proliferation is increased by IGF1 (E3R), a recombinant IGF1 resistant to binding by IGFBPs. Inhibition of EGFR tyrosine kinase activity or IGF1R tyrosine kinase activity inhibits proliferation of MDA-MB-231 cells. Conclusion: These results suggest that the IGF1 signalling pathway is activated in MDA-MB-231 TNBC cells. Therefore, inhibition of the IGF1R and/or its downstream targets may be of benefit in the treatment of TNBC </p

Growing a Collaborative Online Community

Cycling Without Age (CWA) transitioned to its digital Community of Practice (COP), The Hood, from an outdated platform. An effective COP benefits from a well-defined governance structure and a flexible change management plan to encourage productive use of the platform throughout the community. Through communications with the platform's administrators and moderators, this project assisted CWA's transition by effectively introducing and planning for the sustainable operation of The Hood. This project also provided recommendations to support the COP's future growth

Design, synthesis and evaluation of novel 2,2-dimethyl-2,3-dihydroquinolin-4(1H)-one based chalcones as cytotoxic agents

We designed and synthesised a series of novel chalcones, incorporating the heterocyclic framework of 2,2-dimethyl-2,3-dihydro-4(1H)-quinolinone, which was prepared via Sonogashira coupling of a substituted orthoaniline under aqueous conditions using Pd catalysis followed by acid-mediated cyclisation. The compounds were screened against the NCI-N87 and DLD-1 cancer cell lines, with most compounds showing low micromolar cytotoxic activity

Design, synthesis and evaluation of novel 2,2-dimethyl-2,3-dihydroquinolin-4(1H)-one based chalcones as cytotoxic agents

We designed and synthesised a series of novel chalcones, incorporating the heterocyclic framework of 2,2-dimethyl-2,3-dihydro-4(1H)-quinolinone, which was prepared via Sonogashira coupling of a substituted orthoaniline under aqueous conditions using Pd catalysis followed by acid-mediated cyclisation. The compounds were screened against the NCI-N87 and DLD-1 cancer cell lines, with most compounds showing low micromolar cytotoxic activity. </p

Fusobacterium nucleatum: caution with interpreting historical patient sample cohort

Fusobacterium nucleatum (Fn) was first noted to be associated with colorectal cancer (CRC) in 2012. Since then there have been several publications in retrospective cohorts analyzing the relationship between colorectal tumor fusobacterial abundance, clinical and molecular characteristics and clinical outcomes. The majority of studies, including ‘Prognostic impact of Fusobacterium nucleatum depends on combined tumor location and microsatellite instability status in stage II/III colorectal cancers treated with adjuvant chemotherapy’ by Oh et al. published in Journal of Pathology amd Translational Medicine, utilize quantative polymerase chain reaction for the NusG gene in Fn relative to a control prostaglandin transporter gene (SLCO2A1). Many of these studies utilize formalin-fixed paraffin-embedded (FFPE) tissue that has been stored for up to two decades. Although this enables a long follow-up period for clinical outcomes, recent data from our lab shows that advanced age of sample significantly impairs the ability to detect Fn.</p