cMET inhibitor crizotinib impairs angiogenesis and reduces tumor burden in the C3(1)-Tag model of basal-like breast cancer

Epidemiologic studies have associated obesity with increased risk of the aggressive basal-like breast cancer (BBC) subtype. Hepatocyte growth factor (HGF) signaling through its receptor, cMET, is elevated in obesity and is a pro-tumorigenic pathway strongly associated with BBC. We previously reported that high fat diet (HFD) elevated HGF, cMET, and phospho-cMET in normal mammary gland, with accelerated tumor development, compared to low fat diet (LFD)-fed lean controls in a murine model of BBC. We also showed that weight loss resulted in a significant reversal of HFD-induced effects on latency and elevation of HGF/cMET signaling in normal mammary and cMET in normal mammary and tumors. Here, we sought to inhibit BBC tumor progression in LFD- and HFD-fed C3(1)-Tag BBC mice using a small molecule cMET inhibitor, and began crizotinib treatment (50mg/kg body weight by oral gavage) upon identification of the first palpable tumor. We next investigated if administering crizotinib in a window prior to tumor development would inhibit or delay BBC tumorigenesis. Treatment: Crizotinib significantly reduced mean tumor burden by 27.96 and 37.29%, and mean tumor vascularity by 35.04 and 33.52%, in our LFD- and HFD-fed C3(1)-Tag BBC mice, respectively. Prevention: Crizotinib significantly accelerated primary tumor progression in both diet groups but had no effect on total tumor progression or total tumor burden. In sum, cMET inhibition by crizotinib limited tumor development and microvascular density in basal-like tumor-bearing mice but did not appear to be an effective preventive agent for BBC.Electronic supplementary materialThe online version of this article (doi:10.1186/s40064-016-1920-3) contains supplementary material, which is available to authorized users

Dysregulation of fatty acid synthesis and glycolysis in non-Hodgkin lymphoma

The metabolic differences between B-NHL and primary human B cells are poorly understood. Among human B-cell non-Hodgkin lymphomas (B-NHL), primary effusion lymphoma (PEL) is a unique subset that is linked to infection with Kaposi's sarcoma-associated herpesvirus (KSHV). We report that the metabolic profiles of primary B cells are significantly different from that of PEL. Compared with primary B cells, both aerobic glycolysis and fatty acid synthesis (FAS) are up-regulated in PEL and other types of nonviral B-NHL. We found that aerobic glycolysis and FAS occur in a PI3K-dependent manner and appear to be interdependent. PEL overexpress the fatty acid synthesizing enzyme, FASN, and both PEL and other B-NHL were much more sensitive to the FAS inhibitor, C75, than primary B cells. Our findings suggest that FASN may be a unique candidate for molecular targeted therapy against PEL and other B-NHL

Metabolic reprogramming through fatty acid transport protein 1 (FATP1) regulates macrophage inflammatory potential and adipose inflammation

OBJECTIVE: A novel approach to regulate obesity-associated adipose inflammation may be through metabolic reprogramming of macrophages (MΦs). Broadly speaking, MΦs dependent on glucose are pro-inflammatory, classically activated MΦs (CAM), which contribute to adipose inflammation and insulin resistance. In contrast, MΦs that primarily metabolize fatty acids are alternatively activated MΦs (AAM) and maintain tissue insulin sensitivity. In actuality, there is much flexibility and overlap in the CAM-AAM spectrum in vivo dependent upon various stimuli in the microenvironment. We hypothesized that specific lipid trafficking proteins, e.g. fatty acid transport protein 1 (FATP1), would direct MΦ fatty acid transport and metabolism to limit inflammation and contribute to the maintenance of adipose tissue homeostasis. METHODS: Bone marrow derived MΦs (BMDMs) from Fatp1 (-/-) and Fatp1 (+/+) mice were used to investigate FATP1-dependent substrate metabolism, bioenergetics, metabolomics, and inflammatory responses. We also generated C57BL/6J chimeric mice by bone marrow transplant specifically lacking hematopoetic FATP1 (Fatp1 (B-/-)) and controls Fatp1 (B+/+). Mice were challenged by high fat diet (HFD) or low fat diet (LFD) and analyses including MRI, glucose and insulin tolerance tests, flow cytometric, histologic, and protein quantification assays were conducted. Finally, an FATP1-overexpressing RAW 264.7 MΦ cell line (FATP1-OE) and empty vector control (FATP1-EV) were developed as a gain of function model to test effects on substrate metabolism, bioenergetics, metabolomics, and inflammatory responses. RESULTS: Fatp1 is downregulated with pro-inflammatory stimulation of MΦs. Fatp1 (-/-) BMDMs and FATP1-OE RAW 264.7 MΦs demonstrated that FATP1 reciprocally controled metabolic flexibility, i.e. lipid and glucose metabolism, which was associated with inflammatory response. Supporting our previous work demonstrating the positive relationship between glucose metabolism and inflammation, loss of FATP1 enhanced glucose metabolism and exaggerated the pro-inflammatory CAM phenotype. Fatp1 (B-/-) chimeras fed a HFD gained more epididymal white adipose mass, which was inflamed and oxidatively stressed, compared to HFD-fed Fatp1 (B+/+) controls. Adipose tissue macrophages displayed a CAM-like phenotype in the absence of Fatp1. Conversely, functional overexpression of FATP1 decreased many aspects of glucose metabolism and diminished CAM-stimulated inflammation in vitro. FATP1 displayed acyl-CoA synthetase activity for long chain fatty acids in MΦs and modulated lipid mediator metabolism in MΦs. CONCLUSION: Our findings provide evidence that FATP1 is a novel regulator of MΦ activation through control of substrate metabolism. Absence of FATP1 exacerbated pro-inflammatory activation in vitro and increased local and systemic components of the metabolic syndrome in HFD-fed Fatp1 (B-/-) mice. In contrast, gain of FATP1 activity in MΦs suggested that Fatp1-mediated activation of fatty acids, substrate switch to glucose, oxidative stress, and lipid mediator synthesis are potential mechanisms. We demonstrate for the first time that FATP1 provides a unique mechanism by which the inflammatory tone of adipose and systemic metabolism may be regulated

Impact of Tumor Microenvironment and Epithelial Phenotypes on Metabolism in Breast Cancer

Cancer cells have altered metabolism, with increased glucose uptake, glycolysis, and biomass production. This study performed genomic and metabolomic analyses to elucidate how tumor and stromal genomic characteristics influence tumor metabolism

Role of HGF in obesity-associated tumorigenesis: C3(1)-TAg mice as a model for human basal-like breast cancer

Obesity is associated with basal-like breast cancer (BBC), an aggressive breast cancer subtype. The objective of this study was to determine whether obesity promotes BBC onset in adulthood and to evaluate the role of stromal-epithelial interactions in obesity-associated tumorigenesis. We hypothesized that hepatocyte growth factor (HGF) plays a promoting role in BBC, which express the HGF receptor, c-Met. In C3(1)-Tag mice, a murine model of BBC, we demonstrated that obesity leads to a significant increase in HGF secretion and an associated decrease in tumor latency. By immunohistochemical analysis, normal mammary gland exhibited obesity-induced HGF, c-Met and phospho-c-Met, indicating that activation of the cascade was obesity-driven. HGF secretion was also increased from primary mammary fibroblasts isolated from normal mammary glands and tumors of obese mice compared to lean. These results demonstrate that obesity-induced elevation of HGF expression is a stable phenotype, maintained after several passages, and after removal of dietary stimulation. Conditioned media from primary tumor fibroblasts from obese mice drove tumor cell proliferation. In co-culture, neutralization of secreted HGF blunted tumor cell migration, further linking obesity-mediated HGF-dependent effects to in vitro measures of tumor aggressiveness. In sum, these results demonstrate that HGF/c-Met plays an important role in obesity-associated carcinogenesis. Understanding the effects of obesity on risk and progression is important given that epidemiologic studies imply a portion of BBC could be eliminated by reducing obesity

Metabolic Reprogramming of Macrophages: GLUCOSE TRANSPORTER 1 (GLUT1)-MEDIATED GLUCOSE METABOLISM DRIVES A PROINFLAMMATORY PHENOTYPE

Glucose is a critical component in the proinflammatory response of macrophages (MΦs). However, the contribution of glucose transporters (GLUTs) and the mechanisms regulating subsequent glucose metabolism in the inflammatory response are not well understood. Because MΦs contribute to obesity-induced inflammation, it is important to understand how substrate metabolism may alter inflammatory function. We report that GLUT1 (SLC2A1) is the primary rate-limiting glucose transporter on proinflammatory-polarized MΦs. Furthermore, in high fat diet-fed rodents, MΦs in crown-like structures and inflammatory loci in adipose and liver, respectively, stain positively for GLUT1. We hypothesized that metabolic reprogramming via increased glucose availability could modulate the MΦ inflammatory response. To increase glucose uptake, we stably overexpressed the GLUT1 transporter in RAW264.7 MΦs (GLUT1-OE MΦs). Cellular bioenergetics analysis, metabolomics, and radiotracer studies demonstrated that GLUT1 overexpression resulted in elevated glucose uptake and metabolism, increased pentose phosphate pathway intermediates, with a complimentary reduction in cellular oxygen consumption rates. Gene expression and proteome profiling analysis revealed that GLUT1-OE MΦs demonstrated a hyperinflammatory state characterized by elevated secretion of inflammatory mediators and that this effect could be blunted by pharmacologic inhibition of glycolysis. Finally, reactive oxygen species production and evidence of oxidative stress were significantly enhanced in GLUT1-OE MΦs; antioxidant treatment blunted the expression of inflammatory mediators such as PAI-1 (plasminogen activator inhibitor 1), suggesting that glucose-mediated oxidative stress was driving the proinflammatory response. Our results indicate that increased utilization of glucose induced a ROS-driven proinflammatory phenotype in MΦs, which may play an integral role in the promotion of obesity-associated insulin resistance

Metabolomic Profiling Reveals Mitochondrial-Derived Lipid Biomarkers That Drive Obesity-Associated Inflammation

Obesity has reached epidemic proportions worldwide. Several animal models of obesity exist, but studies are lacking that compare traditional lard-based high fat diets (HFD) to “Cafeteria diets" (CAF) consisting of nutrient poor human junk food. Our previous work demonstrated the rapid and severe obesogenic and inflammatory consequences of CAF compared to HFD including rapid weight gain, markers of Metabolic Syndrome, multi-tissue lipid accumulation, and dramatic inflammation. To identify potential mediators of CAF-induced obesity and Metabolic Syndrome, we used metabolomic analysis to profile serum, muscle, and white adipose from rats fed CAF, HFD, or standard control diets. Principle component analysis identified elevations in clusters of fatty acids and acylcarnitines. These increases in metabolites were associated with systemic mitochondrial dysfunction that paralleled weight gain, physiologic measures of Metabolic Syndrome, and tissue inflammation in CAF-fed rats. Spearman pairwise correlations between metabolites, physiologic, and histologic findings revealed strong correlations between elevated markers of inflammation in CAF-fed animals, measured as crown like structures in adipose, and specifically the pro-inflammatory saturated fatty acids and oxidation intermediates laurate and lauroyl carnitine. Treatment of bone marrow-derived macrophages with lauroyl carnitine polarized macrophages towards the M1 pro-inflammatory phenotype through downregulation of AMPK and secretion of pro-inflammatory cytokines. Results presented herein demonstrate that compared to a traditional HFD model, the CAF diet provides a robust model for diet-induced human obesity, which models Metabolic Syndrome-related mitochondrial dysfunction in serum, muscle, and adipose, along with pro-inflammatory metabolite alterations. These data also suggest that modifying the availability or metabolism of saturated fatty acids may limit the inflammation associated with obesity leading to Metabolic Syndrome

BCAT1 controls metabolic reprogramming in activated human macrophages and is associated with inflammatory diseases

This work was supported by the Medical Research Council (MR/M004716/1 and MR/N01121X/1 to J.B.) and by Kidney Research UK (RP9/2013 to J.B.). D.C. is supported by the Institute Pasteur, Fondazione Cenci Bolognetti. C.M. is supported by the British Hearth Foundation Fellowship (FS/12/3829640)

Longitudinal mechanical tension induces growth in the small bowel of juvenile rats

Introduction: The aim of our study was to apply longitudinal force to the small bowel to increase the length of intestine in juvenile rats. Methods: Fifty juvenile rats had double barrelled, blind loop ostomies created using an isolated segment of bowel. Our intestinal lengthening device was inserted into one of the loops and the second loop served as a control. Once the device was deployed, the experimental, control, and in situ segments of bowel were evaluated for length, weight, histology, and disaccharidase enzyme activity. Results: Mechanical tension increased intestinal length by 149%. The lengthened bowel also exhibited a greater total weight (218%), greater mucosal weight (122%), and increased protein mass (164%) compared with the control limb of bowel. Histologically, there was a markedly increased thickness of the muscularis propria in the lengthened bowel (200% increase compared with the control limb). Functionally, we found increased total disaccharidase activity in the lengthened bowel (between 47% and 350%, depending on the particular enzyme tested; p<0.01). Conclusion: Mechanical tension induces intestinal growth by increasing length, weight of the bowel and mucosa, and protein mass. Histological changes, such as increases in Paneth cells, suggest that increased proliferation and reorganisation of the mucosa and muscularis propria are a response to mechanical tension. Functionally, increased intestinal length corresponds with increased disaccharidase activity, thus implying potential increased absorptive capacity of the lengthened bowel