Inhibiting the glycerophosphodiesterase EDI3 in ER-HER2+ breast cancer cells resistant to HER2-targeted therapy reduces viability and tumour growth.

BACKGROUND: Intrinsic or acquired resistance to HER2-targeted therapy is often a problem when small molecule tyrosine kinase inhibitors or antibodies are used to treat patients with HER2 positive breast cancer. Therefore, the identification of new targets and therapies for this patient group is warranted. Activated choline metabolism, characterized by elevated levels of choline-containing compounds, has been previously reported in breast cancer. The glycerophosphodiesterase EDI3 (GPCPD1), which hydrolyses glycerophosphocholine to choline and glycerol-3-phosphate, directly influences choline and phospholipid metabolism, and has been linked to cancer-relevant phenotypes in vitro. While the importance of choline metabolism has been addressed in breast cancer, the role of EDI3 in this cancer type has not been explored. METHODS: EDI3 mRNA and protein expression in human breast cancer tissue were investigated using publicly-available Affymetrix gene expression microarray datasets (n = 540) and with immunohistochemistry on a tissue microarray (n = 265), respectively. A panel of breast cancer cell lines of different molecular subtypes were used to investigate expression and activity of EDI3 in vitro. To determine whether EDI3 expression is regulated by HER2 signalling, the effect of pharmacological inhibition and siRNA silencing of HER2, as well as the influence of inhibiting key components of signalling cascades downstream of HER2 were studied. Finally, the influence of silencing and pharmacologically inhibiting EDI3 on viability was investigated in vitro and on tumour growth in vivo. RESULTS: In the present study, we show that EDI3 expression is highest in ER-HER2 + human breast tumours, and both expression and activity were also highest in ER-HER2 + breast cancer cell lines. Silencing HER2 using siRNA, as well as inhibiting HER2 signalling with lapatinib decreased EDI3 expression. Pathways downstream of PI3K/Akt/mTOR and GSK3ß, and transcription factors, including HIF1α, CREB and STAT3 were identified as relevant in regulating EDI3 expression. Silencing EDI3 preferentially decreased cell viability in the ER-HER2 + cells. Furthermore, silencing or pharmacologically inhibiting EDI3 using dipyridamole in ER-HER2 + cells resistant to HER2-targeted therapy decreased cell viability in vitro and tumour growth in vivo. CONCLUSIONS: Our results indicate that EDI3 may be a potential novel therapeutic target in patients with HER2-targeted therapy-resistant ER-HER2 + breast cancer that should be further explored.

Spatio-Temporal Multiscale Analysis of Western Diet-Fed Mice Reveals a Translationally Relevant Sequence of Events during NAFLD Progression.

Mouse models of non-alcoholic fatty liver disease (NAFLD) are required to define therapeutic targets, but detailed time-resolved studies to establish a sequence of events are lacking. Here, we fed male C57Bl/6N mice a Western or standard diet over 48 weeks. Multiscale time-resolved characterization was performed using RNA-seq, histopathology, immunohistochemistry, intravital imaging, and blood chemistry; the results were compared to human disease. Acetaminophen toxicity and ammonia metabolism were additionally analyzed as functional readouts. We identified a sequence of eight key events: formation of lipid droplets; inflammatory foci; lipogranulomas; zonal reorganization; cell death and replacement proliferation; ductular reaction; fibrogenesis; and hepatocellular cancer. Functional changes included resistance to acetaminophen and altered nitrogen metabolism. The transcriptomic landscape was characterized by two large clusters of monotonously increasing or decreasing genes, and a smaller number of 'rest-and-jump genes' that initially remained unaltered but became differentially expressed only at week 12 or later. Approximately 30% of the genes altered in human NAFLD are also altered in the present mouse model and an increasing overlap with genes altered in human HCC occurred at weeks 30-48. In conclusion, the observed sequence of events recapitulates many features of human disease and offers a basis for the identification of therapeutic targets.

Soybean Meal Can Be Replaced by Faba Beans, Pumpkin Seed Cake, Spirulina or Be Completely Omitted in a Forage-Based Diet for Fattening Bulls to Achieve Comparable Performance, Carcass and Meat Quality.

The aim of the study was to investigate the complete substitution of imported soybean meal in beef cattle diets and the consequences on performance, meat, and adipose tissue quality. Thirty growing crossbred Limousin bulls, with an initial bodyweight of 164 ± 13 kg and 4.3 ± 0.3 months of age, were fed a grass/maize-silage based diet with little additional concentrate (0.5:0.3:0.2). Concentrates contained either soybean meal (positive control), faba beans, pumpkin seed cake, or spirulina (Arthrospira platensis), resulting in about 226 g crude protein (CP)/kg concentrate dry matter (DM) and 158 g CP/kg total diet DM. A grain-based concentrate providing just 135 g CP/kg concentrate DM and 139 g CP/total diet DM served as a negative control. Bulls of all groups had comparable average daily gains (1.43 ± 0.1 kg) and feed intakes (6.92 ± 0.37 kg). Carcass and meat quality did not differ among groups. The fatty acid profile of meat lipids was hardly affected. These results indicate that soybean meal can be replaced by any of the tested protein sources without impairing performance or meat quality. Importantly, bulls fed the negative control achieved a fattening and slaughter performance comparable to that of the protein-supplemented groups without affecting meat and adipose tissue quality. Thus, the present findings suggest that feeding crossbred bulls a grass/maize-silage based diet does not require additional protein supplementation.

Antioxidant and Inflammatory Gene Expression Profiles of Bovine Peripheral Blood Mononuclear Cells in Response to Arthrospira platensis before and after LPS Challenge.

Oxidative stress and inflammatory diseases are closely related processes that need to be controlled to ensure the desirable high performance of livestock. The microalga spirulina has shown antioxidant and anti-inflammatory properties in monogastric species. To investigate potential beneficial effects in ruminants, we replaced soybean meal (SOY) in the diets of dairy cows and fattening bulls by spirulina (SPI) and analyzed plasma concentrations of antioxidants (ß-carotene, α-tocopherol, polyphenols) and serum total antioxidant capacity. Following in vitro stimulation with lipopolysaccharide (LPS), peripheral blood mononuclear cells (PBMCs) were isolated for expression analysis of inflammation- and antioxidant-defense-related genes. Plasma ß-carotene concentration was higher in SPI, compared to SOY cows, but did not differ in bulls. Plasma total phenol concentration was significantly higher in SPI, compared to SOY bulls, but not in cows. Stimulation of bovine PBMCs with LPS increased the expression of most cytokines and some antioxidant enzymes. Gene expression of PBMCs derived from SPI animals, compared to SOY animals, hardly differed. Our results indicate that in ruminants, spirulina might not have potent antioxidant and anti-inflammatory properties. Future studies should evaluate the microbial degradation of spirulina and its bioactive compounds in the rumen to provide further data on potential beneficial health effects in ruminants.