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Prostate cancer (PCa) is the most commonly diagnosed malignancy and the second leading cause of cancer-related death in American men. Androgen deprivation therapy (ADT) has become a standard treatment strategy for advanced PCa. Although a majority of patients initially respond to ADT well, most of them will eventually develop castration-resistant PCa (CRPC). Previous studies suggest that ADT-induced changes in the immune microenvironment (mE) in PCa might be responsible for the failures of various therapies. However, the role of the immune system in CRPC development remains unclear. To systematically understand the immunity leading to CRPC progression and predict the optimal treatment strategy in silico, we developed a 3D Hybrid Multi-scale Model (HMSM), consisting of an ODE system and an agent-based model (ABM), to manipulate the tumor growth in a defined immune system. Based on our analysis, we revealed that the key factors (e.g. WNT5A, TRAIL, CSF1, etc.) mediated the activation of PC-Treg and PC-TAM interaction pathways, which induced the immunosuppression during CRPC progression. Our HMSM model also provided an optimal therapeutic strategy for improving the outcomes of PCa treatment. [ABSTRACT FROM AUTHOR]

Objective: To explore the use of Contrast-enhanced Ultrasound (CEUS) in evaluating angiogenesis in a xenograft nasopharyngeal carcinoma (NPC) model in nude mice and the evolution of CEUS parameters according to the growth of NPC. Methods: Nude mice were divided into three groups according to experiments conducted at various times from tumor implantation (8 mice/group; group A: 4 weeks from implantation; group B:6 weeks from implantation; group C:8 weeks from implantation). CNE-2 cells were transplanted in 24 nude mice and CEUS evaluations of the tumors were performed at 4, 6 or 8 weeks from implantation. CEUS parametric perfusion images and pathological findings were recorded. R version 3.4.4 software was used to analyze the CEUS parameters and pathological findings. Results: One-way anova analysis indicated statistically significant differences among the three groups with the parameters of peak intensity (PI) (p<0.001), area wash in (AWI) (p<0.001), area wash out (AWO) (p<0.001) and tumor volumes (p<0.001).Pearson correlation coefficient analysis indicated that microvessel density (MVD) was correlated with tumor volume (r = 0.644, p = 0.001), PI (r = 0.904, p<0.0001), AWI (r = 0.547, p = 0.008) and AWO (r = 0.744, P<0.0001). Tumor volume was correlated with MVD (r = 0.644, p = 0.001), PI (r = 0.625, p = 0.002), AWI (r = 0.528, p = 0.012) and AWO (r = 0.784, p<0.001). The percentage of necrosis in histological sections was correlated with the percentage of CEUS unperfused area (r = 0.446,p = 0.038). Spearman rank correlation coefficient analysis indicated that vascular endothelial growth factor (VEGF) was correlated with PI (r = 0.462, P = 0.032). Welch t test indicated PI, AWI and AWO parameters were significantly lower than that of kidneys (p<0.001, p = 0.009, p = 0.005). Conclusions: The CEUS parameters PI, AWI and AWO indirectly reflect the MVD and the tumor volume in our model of subcutaneous transplanted NPC in nude mice, providing precious information on angiogenesis and tumor growth. VEGF may play a role in promoting angiogenesis of NPC. [ABSTRACT FROM AUTHOR]

Cancers exhibit spatially heterogeneous, unique vascular architectures across individual samples, cell-lines and patients. This inherently disorganised collection of leaky blood vessels contribute significantly to suboptimal treatment efficacy. Preclinical tools are urgently required which incorporate the inherent variability and heterogeneity of tumours to optimise and engineer anti-cancer therapies. In this study, we present a novel computational framework which incorporates whole, realistic tumours extracted ex vivo to efficiently simulate vascular blood flow and interstitial fluid transport in silico for validation against in vivo biomedical imaging. Our model couples Poiseuille and Darcy descriptions of vascular and interstitial flow, respectively, and incorporates spatially heterogeneous blood vessel lumen and interstitial permeabilities to generate accurate predictions of tumour fluid dynamics. Our platform enables highly-controlled experiments to be performed which provide insight into how tumour vascular heterogeneity contributes to tumour fluid transport. We detail the application of our framework to an orthotopic murine glioma (GL261) and a human colorectal carcinoma (LS147T), and perform sensitivity analysis to gain an understanding of the key biological mechanisms which determine tumour fluid transport. Finally we mimic vascular normalization by modifying parameters, such as vascular and interstitial permeabilities, and show that incorporating realistic vasculatures is key to modelling the contrasting fluid dynamic response between tumour samples. Contrary to literature, we show that reducing tumour interstitial fluid pressure is not essential to increase interstitial perfusion and that therapies should seek to develop an interstitial fluid pressure gradient. We also hypothesise that stabilising vessel diameters and permeabilities are not key responses following vascular normalization and that therapy may alter interstitial hydraulic conductivity. Consequently, we suggest that normalizing the interstitial microenvironment may provide a more effective means to increase interstitial perfusion within tumours. [ABSTRACT FROM AUTHOR]

Mesenchymal stem cells (MSCs) can influence the tumour microenvironment (TEM) and play a major role in tumourigenesis. Triple-negative [Ostrogen receptor (ER-), Progesterone receptor (PgR-), and HER2/neu receptor (HER2-)] breast cancer (TNBC) is an aggressive class of BC characterized by poor prognosis and lacks the benefit of routinely available targeted therapies. This study aims to investigate the effect of human placental chorionic villi derived MSCs (CVMSCs) on the behavior of TNBC in vitro. This was done by assaying different cancer hallmarks including proliferation, migration and angiogenesis. Cell proliferation rate of TNBC cell line (MDA-MB231) was monitored in real time using the xCELLigence system. Whereas, Boyden chamber migration assay was used to measure MDA-MB231 motility and invasiveness toward CVMSCs. Finally, a three-dimensional (3D) model using a co-culture system of CVMSCs with MDA-MB231 with or without the addition of human umbilical vein endothelial cells (HUVECs) was created to assess tumour angiogenesis in vitro. CVMSCs were able to significantly reduce the proliferative and migratory capacity of MDA-MB231 cells. Co-culturing of MDA-MB231 with CVMSCs, not only inhibited the tube formation ability of HUVECs but also reduced the expression of the BC characteristic cytokines; IL-10, IL-12, CXCL9 and CXCL10 of CVMSCs. These results support the hypothesis that CVMSCs can influence the behavior of TNBC cells and provides a basic for a potential therapeutic approach in a pre-clinical settings. The data from this study also highlight the complexity of the in vitro cancer angiogenesis model settings and regulations. [ABSTRACT FROM AUTHOR]

The delivery of blood-borne therapeutic agents to solid tumours depends on a broad range of biophysical factors. We present a novel multiscale, multiphysics, in-silico modelling framework that encompasses dynamic tumour growth, angiogenesis and drug delivery, and use this model to simulate the intravenous delivery of cytotoxic drugs. The model accounts for chemo-, hapto- and mechanotactic vessel sprouting, extracellular matrix remodelling, mechano-sensitive vascular remodelling and collapse, intra- and extravascular drug transport, and tumour regression as an effect of a cytotoxic cancer drug. The modelling framework is flexible, allowing the drug properties to be specified, which provides realistic predictions of in-vivo vascular development and structure at different tumour stages. The model also enables the effects of neoadjuvant vascular normalisation to be implicitly tested by decreasing vessel wall pore size. We use the model to test the interplay between time of treatment, drug affinity rate and the size of the vessels endothelium pores on the delivery and subsequent tumour regression and vessel remodelling. Model predictions confirm that small-molecule drug delivery is dominated by diffusive transport and further predict that the time of treatment is important for low affinity but not high affinity cytotoxic drugs, the size of the vessel wall pores plays an important role in the effect of low affinity but not high affinity drugs, that high affinity cytotoxic drugs remodel the tumour vasculature providing a large window for the normalisation of the vascular architecture, and that the combination of large pores and high affinity enhances cytotoxic drug delivery efficiency. These results have implications for treatment planning and methods to enhance drug delivery, and highlight the importance of in-silico modelling in investigating the optimisation of cancer therapy on a personalised setting. [ABSTRACT FROM AUTHOR]

Background: Angiogenesis is a key process for tumor progression and a target for treatment. However, the regulation of breast cancer angiogenesis and its relevance for clinical resistance to antiangiogenic drugs is still incompletely understood. Recent developments on the contribution of microRNA to tumor angiogenesis and on the oncogenic effects of miR-17-92, a miRNA cluster, point to their potential role on breast cancer angiogenesis. The aim of this work was to establish the contribution of miR-20a, a member of miR-17-92 cluster, to tumor angiogenesis in patients with invasive breast carcinoma. Methods: Tube-formation in vitro assays with conditioned medium from MCF7 and MDA-MB-231 breast cancer cell lines were performed after transfection with miR-20a and anti-miR20a. For clinical validation of the experimental findings, we performed a retrospective analysis of a series of consecutive breast cancer patients (n = 108) treated with neoadjuvant chemotherapy and with a full characterization of their vessel pattern and expression of angiogenic markers in pre-treatment biopsies. Expression of members of the cluster miR-17-92 and of angiogenic markers was determined by RT-qPCR after RNA purification from FFPE samples. Results: In vitro angiogenesis assays with endothelial cells and conditioned media from breast cancer cell lines showed that transfection with anti-miR20a in MDA-MB-231 significantly decreased mean mesh size and total mesh area, while transfection with miR-20a in MCF7 cells increased mean mesh size. MiR-20a angiogenic effects were abrogated by treatment with aflibercept, a VEGF trap. These results were supported by clinical data showing that mir-20a expression was higher in tumors with no estrogen receptor or with more extensive nodal involvement (cN2-3). A higher miR-20a expression was associated with higher mean vessel size (p = 0.015) and with an angiogenic pattern consisting in larger vessels, higher VEGFA expression and presence of glomeruloid microvascular proliferations (p<0.001). This association was independent of tumor subtype and VEGFA expression. Conclusions: Transfection of breast cancer cells with miR-20a induces vascular changes in endothelial tube-formation assays. Expression of miR-20a in breast invasive carcinomas is associated with a distinctive angiogenic pattern consisting in large vessels, anomalous glomeruloid microvascular proliferations and high VEGFA expression. Our results suggest a role for miR-20a in the regulation of breast cancer angiogenesis, and raise the possibility of its use as an angiogenic biomarker. [ABSTRACT FROM AUTHOR]

Tumour angiogenesis is an important hallmark of cancer and the study of its metabolic adaptations, downstream to any cellular change, can reveal attractive targets for inhibiting cancer growth. In the tumour microenvironment, endothelial cells (ECs) interact with heterogeneous tumour cell types that drive angiogenesis and metastasis. In this study we aim to characterize the metabolic alterations in ECs influenced by the presence of tumour cells with extreme metastatic abilities. Human umbilical vein endothelial cells (HUVECs) were subjected to different microenvironmental conditions, such as the presence of highly metastatic PC-3M and highly invasive PC-3S prostate cancer cell lines, in addition to the angiogenic activator vascular endothelial growth factor (VEGF), under normoxia. Untargeted high resolution liquid chromatography-mass spectrometry (LC-MS) based metabolomics revealed significant metabolite differences among the various conditions and a total of 25 significantly altered metabolites were identified including acetyl L-carnitine, NAD+, hypoxanthine, guanine and oleamide, with profile changes unique to each of the experimental conditions. Biochemical pathway analysis revealed the importance of fatty acid oxidation and nucleotide salvage pathways. These results provide a global metabolic preview that could help in selectively targeting the ECs aiding in either cancer cell invasion or metastasis in the heterogeneous tumour microenvironment. [ABSTRACT FROM AUTHOR]

Tumors exploit angiogenesis, the formation of new blood vessels from pre-existing vasculature, in order to obtain nutrients required for continued growth and proliferation. Targeting factors that regulate angiogenesis, including the potent promoter vascular endothelial growth factor (VEGF), is therefore an attractive strategy for inhibiting tumor growth. Computational modeling can be used to identify tumor-specific properties that influence the response to anti-angiogenic strategies. Here, we build on our previous systems biology model of VEGF transport and kinetics in tumor-bearing mice to include a tumor compartment whose volume depends on the “angiogenic signal” produced when VEGF binds to its receptors on tumor endothelial cells. We trained and validated the model using published in vivo measurements of xenograft tumor volume, producing a model that accurately predicts the tumor’s response to anti-angiogenic treatment. We applied the model to investigate how tumor growth kinetics influence the response to anti-angiogenic treatment targeting VEGF. Based on multivariate regression analysis, we found that certain intrinsic kinetic parameters that characterize the growth of tumors could successfully predict response to anti-VEGF treatment, the reduction in tumor volume. Lastly, we use the trained model to predict the response to anti-VEGF therapy for tumors expressing different levels of VEGF receptors. The model predicts that certain tumors are more sensitive to treatment than others, and the response to treatment shows a nonlinear dependence on the VEGF receptor expression. Overall, this model is a useful tool for predicting how tumors will respond to anti-VEGF treatment, and it complements pre-clinical in vivo mouse studies. [ABSTRACT FROM AUTHOR]

Lupeol and stigmasterol, major phytosterols in various herbal plants, possess anti-inflammatory activities and have been proposed as candidates for anti-cancer agents, but their molecular mechanisms are still unclear. Here, we investigated the effects of lupeol and stigmasterol on tumor and endothelial cells in vitro and their anti-cancer activities in vivo. Our results demonstrated that lupeol and stigmasterol suppressed cell viability, migration, and morphogenesis of human umbilical vein endothelial cells (HUVECs) but not cholangiocarcinoma (CCA) cells. Expression analyses showed that the treatment of both compounds significantly reduced the transcript level of tumor necrosis factor-α (TNF-α), and Western blot analyses further revealed a decrease in downstream effector levels of VEGFR-2 signaling, including phosphorylated forms of Src, Akt, PCL, and FAK, which were rescued by TNF-α treatment. In vivo, lupeol and stigmasterol disrupted tumor angiogenesis and reduced the growth of CCA tumor xenografts. Immunohistochemical analyses confirmed a decrease in CD31-positive vessel content and macrophage recruitment upon treatment. These findings indicate that lupeol and stigmasterol effectively target tumor endothelial cells and suppress CCA tumor growth by their anti-inflammatory activities and are attractive candidates for anti-cancer treatment of CCA tumors. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to investigate the link between the inhibitory effect of ginsenoside Rg3 on the ectopic endometrium growth and the VEGFR-2-mediated PI3K/Akt/mTOR signaling pathway, a mechanism known to inhibit angiogenesis and induce ectopic endometrial cell apoptosis. Materials and methods: A model of endometriosis was established by allotransplantation in rats. The rats were randomly divided into 5 groups: the ginsenoside Rg3 low-dose group (group A,5mg/kgBW/d of ginsenoside Rg3), the ginsenoside Rg3 high-dose group (group B, 10mg/kgBW/d of ginsenoside Rg3), the gestrinone group (group C, 0.5mg/kgBW/d of gestrinone), the control group (groupD, 10ml/kg BW/d of 0.5%CMC-Na) and the ovariectomized group (group E, 10ml/kgBW/d of 0.5%CMC-Na). Rats were executed after 21 days of continuous administration. The ectopic endometrium volume was measured and the inhibitory rate was calculated. The levels of serum estradiol (E2) and progesterone (P) were detected by Electro-Chemiluminescence Immunoassay (ECLI). The protein expressionof VEGF, VEGFR-2, p-Akt, and p-mTOR inthe ectopic endometrium wastested by immunohistochemistry(IHC) and Western Blotting. The mRNA expression levels of VEGF, VEGFR-2, Akt, and mTOR were tested by Real-Time Polymerase Chain Reaction (PCR). The apoptosis rate of the ectopic endometrial cells was detected by Terminal Deoxynucleotidyl Transferase-mediated Digoxigenin-dUTP Nick-End Labeling Assay(TUNEL). Main results: Tissue measurements revealed a dose-dependent inhibition effect of ginsenoside Rg3 on the growth of the ectopic endometrium in treated rats compared to controls. Immunohistochemical and Western Blotting assays confirmed that the expression of VEGF, p-Akt, and p-mTOR was down-regulated in ginsenoside Rg3 -treated lesions. Real-time PCR results also showed that the mRNA expression levels of VEGF, Akt, and mTOR in the ectopic endometrium were reduced. Conclusions: The present study demonstrates, for the first time, that ginsenoside Rg3 suppresses angiogenesis in developing endometrial lesions. The ginsenoside Rg3 inhibitory effect on the growth of the ectopic endometrium in EMs rats might occur through the blocking of the VEGFR-2-mediated PI3K/Akt/mTOR signaling pathway, thus halting angiogenesis and promoting the apoptosis of ectopic endometrial cells. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the therapeutic responses of lung cancer mice models with adenocarcinoma HCC827 (gefitinib sensitive) and HCC827R (gefitinib resistant) to the epidermal growth factor receptor-tyrosine kinase inhibitor erlotinib alone and in combination with the anti-angiogenesis agent bevacizumab using dynamic contrast enhanced (DCE) and diffusion-weighted MRI. In the HCC827 model, temporal changes in DCE-MRI derived parameters (Ktrans, kep, and iAUC90) and apparent diffusion coefficient (ADC) were significantly correlated with tumor size. Ktrans and iAUC90 significantly decreased at week 2 in the groups receiving erlotinib alone and in combination with bevacizumab, whereas kep decreased at week 1 and 2 in both treatment groups. In addition, there was a significant difference in iAUC90 between the treatment groups at week 1. Compared to the control group of HCC827, there was a significant reduction in microvessel density and increased tumor apoptosis in the two treatment group. ADC value increased in the erlotinib alone group at week 1 and week 2, and in the erlotinib combined with bevacizumab group at week 2. Enlarged areas of central tumor necrosis were associated with a higher ADC value. However, progressive enlargement of the tumors but no significant differences in DCE parameters or ADC were noted in the HCC827R model. These results showed that both erlotinib alone and in combination with bevacizumab could effectively inhibit tumor growth in the gefitinib-sensitive lung cancer mice model, and that this was associated with decreased vascular perfusion, increased ADC percentage, decreased microvessel density, and increased tumor apoptosis with a two-week treatment cycle. [ABSTRACT FROM AUTHOR]

In the course of searching for angiogenesis inhibitors from microorganisms, two cyclic peptides, PF1171A (1) and PF1171C (2) were isolated from the soil fungus Penicillium sp. FN070315. In the present study, we investigated the antiangiogenic efficacy and associated mechanisms of 1 and 2 in vitro using human umbilical vein endothelial cells (HUVECs). Compounds 1 and 2 inhibited the proliferation of HUVECs at concentrations not exhibiting cytotoxicity. Moreover, 1 and 2 significantly suppressed vascular endothelial growth factor (VEGF)-induced migration, invasion, proliferation and tube formation of HUVECs as well as neovascularization of the chorioallantoic membrane in developing chick embryos. We also identified an association between the antiangiogenic activity of 1 and 2 and the downregulation of both the phosphorylation of VEGF receptor 2 and the expression of hypoxia inducible factor-1α at the protein level. Taken together, these results further suggest that compounds 1 and 2 will be promising angiogenesis inhibitors. [ABSTRACT FROM AUTHOR]

The mechanisms that link inflammatory responses to cancer development remain a subject of intense investigation, emphasizing the need to better understand the cellular and molecular pathways that create a tumor promoting microenvironment. The myeloid differentiation primary response protein MyD88 acts as a main adaptor molecule for the signaling cascades initiated from Toll-like receptors (TLRs) and the interleukin 1 receptor (IL-1R). MyD88 has been shown to contribute to tumorigenesis in many inflammation-associated cancer models. In this study, we sought to better define the role of MyD88 in neoplastic cells using a murine melanoma model. Herein, we have demonstrated that MyD88 expression is required to maintain the angiogenic switch that supports B16 melanoma growth. By knocking down MyD88 we reduced TLR-mediated NF-κB activation with no evident effects over cell proliferation and survival. In addition, MyD88 downregulation was associated with a decrease of HIF1α levels and its target gene VEGF, in correlation with an impaired capability to induce capillary sprouting and tube formation of endothelial cells. Melanomas developed from cells lacking MyD88 showed an enhanced secretion of chemoattractant ligands such as CCL2, CXCL10 and CXCL1 and have an improved infiltration of macrophages to the tumor site. Our results imply that cell-autonomous signaling through MyD88 is required to sustain tumor growth and underscore its function as an important positive modulator of tumor angiogenesis. [ABSTRACT FROM AUTHOR]

We investigate the effects of intermittent hypoxia (IH), a characteristic feature of obstructive sleep apnea (OSA), on renal cancer progression in an animal and cell model. An in vivo mouse model (Balb/c, n = 50) of kidney cancer was used to assess the effect of IH on tumor growth, metastatic capacity, angiogenesis and tumor immune response. An in vitro model tested the effect of IH on RENCA cells, macrophages and endothelial cells. Tumor growth, metastatic capacity, circulating vascular endothelial growth factor (VEGF) and content of endothelial cells, tumor associated macrophages and their phenotype were assessed in the tumor. In vitro, VEGF cell expression was quantified.Although IH did not boost tumor growth, it significantly increased endothelial cells (p = 0.001) and circulating VEGF (p<0.001) in the in vivo model. Macrophages exposed to IH in vitro increased VEGF expression, whereas RENCA cells and endothelial cells did not. These findings are in keeping with previous clinical data suggesting that OSA has no effect on kidney cancer size and that the association observed between OSA and higher Fuhrman grade of renal cell carcinoma may be mediated though a proangiogenic process, with a key role of macrophages. [ABSTRACT FROM AUTHOR]

Increases in nuclear calcium concentration generate specific biological outcomes that differ from those resulting from increased cytoplasmic calcium. Nuclear calcium effects on tumor cell proliferation are widely appreciated; nevertheless, its involvement in other steps of tumor progression is not well understood. Therefore, we evaluated whether nuclear calcium is essential in other additional stages of tumor progression, including key steps associated with the formation of the primary tumor or with the metastatic cascade. We found that nuclear calcium buffering impaired 4T1 triple negative breast cancer growth not just by decreasing tumor cell proliferation, but also by enhancing tumor necrosis. Moreover, nuclear calcium regulates tumor angiogenesis through a mechanism that involves the upregulation of the anti-angiogenic C-X-C motif chemokine 10 (CXCL10-IP10). In addition, nuclear calcium buffering regulates breast tumor cell motility, culminating in less cell invasion, likely due to enhanced vinculin expression, a focal adhesion structural protein. Together, our results show that nuclear calcium is essential for triple breast cancer angiogenesis and cell migration and can be considered as a promising strategic target for triple negative breast cancer therapy. [ABSTRACT FROM AUTHOR]

Esophageal squamous cell carcinoma (ESCC) is the second common cancer in Henan province and is well-known for aggressiveness and dismal prognosis. Adjuvant therapies, chemotherapy, radiotherapy and endoscopic treatment have not improved survival rates in patients with late stage esophageal carcinoma. All-trans retinoic acid (ATRA) is the active ingredient of Vitamin A and affects a wide spectrum of biological processes including development, growth, neural function, immune function, reproduction, and vision. It is one of the most potent therapeutic agents used for treating cancers, especially lung adenocarcinomas. ATRA inhibits metastatic potential and angiogenesis in several tumor models. We investigated the effects of ATRA on the expression of angiopoietin 1 (Ang-1), angiopoietin 2 (Ang-2) and receptor Tie-2 in EC1 cells in vitro. We also assessed the growth and migration of EC1 cells in vitro. ATRA treatment caused 29.5% and 40.3% reduction of the growth of EC1 cells after 24 hours and 48 hours, relative to the control. ATRA plus fluorouracil treatment reduced the viability more strongly than either drug alone, indicating an additive effect. Moreover, ATRA decreased EC1 migration by 87%. Furthermore, ATRA treatment led to a marked decrease of the transcript levels of Ang-1, Ang-2, Tie-2, VEGF, and VEGF receptors, as assessed by real-time RT-PCR. Importantly, the protein levels of Ang-1, Ang-2 and Tie-2 were reduced by ATRA treatment. In vivo, we found ATRA treatment suppressed the tumor growth and improved the cachexia of mice. Importantly, ATRA treatment decreased the expression of CD31, Ang-1, Ang-2 and Tie-2 in subcutaneous tumors of EC1 cells. Collectively, our findings demonstrate that ATRA exhibits a dose- and temporal-dependent effect on the metastatic behavior, suppresses the angiopoietin-Tie2 pathway and inhibits angiogenesis and the progression of xenograft tumors of EC1 cells. [ABSTRACT FROM AUTHOR]

Secreted frizzled related protein 2 (SFRP2) is a tumor endothelial marker expressed in angiosarcoma. Previously, we showed ultrasound molecular imaging with SFRP2-targeted contrast increased average video pixel intensity (VI) of angiosarcoma vessels by 2.2 ± 0.6 VI versus streptavidin contrast. We hypothesized that redesigning our contrast agents would increase imaging performance. Improved molecular imaging reagents were created by combining NeutrAvidin™-functionalized microbubbles with biotinylated SFRP2 or IgY control antibodies. When angiosarcoma tumors in nude mice reached 8 mm, time-intensity, antibody loading, and microbubble dose experiments optimized molecular imaging. 10 minutes after injection, the control-subtracted time-intensity curve (TIC) for SFRP2-targeted contrast reached a maximum, after subtracting the contribution of free-flowing contrast. SFRP2 antibody-targeted VI was greater when contrast was formulated with 10-fold molar excess of maleimide-activated NeutrAvidin™ versus 3-fold (4.5 ± 0.18 vs. 0.32 ± 0.15, VI ± SEM, 5 x 106 dose, p < 0.001). Tumor vasculature returned greater average video pixel intensity using 5 x 107 versus 5 x 106 microbubbles (21.2 ± 2.5 vs. 4.5 ± 0.18, p = 0.0011). Specificity for tumor vasculature was confirmed by low VI for SFRP2-targeted, and control contrast in peri-tumoral vasculature (3.2 ± 0.52 vs. 1.6 ± 0.71, p = 0.92). After optimization, average video pixel intensity of tumor vasculature was 14.2 ± 3.0 VI units higher with SFRP2-targeted contrast versus IgY-targeted control (22.1 ± 2.5 vs. 7.9 ± 1.6, p < 0.001). After log decompression, 14.2 ΔVI was equal to ~70% higher signal, in arbitray acoustic units (AU), for SFRP2 versus IgY. This provided ~18- fold higher acoustic signal enhancement than provided previously by 2.2 ΔVI. Basing our targeted contrast on NeutrAvidin™-functionalized microbubbles, using IgY antibodies for our control contrast, and optimizing our imaging protocol significantly increased the SFRP2-specific signal returned from angiosarcoma vasculature, and may provide new opportunities for targeted molecular imaging. [ABSTRACT FROM AUTHOR]

Objective: The brain’s functional architecture of interconnected network-related oscillatory patterns in discrete cortical regions has been well established with functional magnetic resonance imaging (fMRI) studies or direct cortical electrophysiology from electrodes placed on the surface of the brain, or electrocorticography (ECoG). These resting state networks exhibit a robust functional architecture that persists through all stages of sleep and under anesthesia. While the stability of these networks provides a fundamental understanding of the organization of the brain, understanding how these regions can be perturbed is also critical in defining the brain’s ability to adapt while learning and recovering from injury. Methods: Patients undergoing an awake craniotomy for resection of a tumor were studied as a unique model of an evolving injury to help define how the cortical physiology and the associated networks were altered by the presence of an invasive brain tumor. Results: This study demonstrates that there is a distinct pattern of alteration of cortical physiology in the setting of a malignant glioma. These changes lead to a physiologic sequestration and progressive synaptic homogeneity suggesting that a de-learning phenomenon occurs within the tumoral tissue compared to its surroundings. Significance: These findings provide insight into how the brain accommodates a region of “defunctionalized” cortex. Additionally, these findings may have important implications for emerging techniques in brain mapping using endogenous cortical physiology. [ABSTRACT FROM AUTHOR]

Drug transport and its uptake by tumour cells are strongly dependent on tumour properties, which vary in different types of solid tumours. By simulating the key physical and biochemical processes, a numerical study has been carried out to investigate the transport of anti-cancer drugs in 3-D tumour models of different sizes. The therapeutic efficacy for each tumour is evaluated by using a pharmacodynamics model based on the predicted intracellular drug concentration. Simulation results demonstrate that interstitial fluid pressure and interstitial fluid loss vary non-linearly with tumour size. Transvascular drug exchange, driven by the concentration gradient of unbound drug between blood and interstitial fluid, is more efficient in small tumours, owing to the low spatial-mean interstitial fluid pressure and dense microvasculature. However, this has a detrimental effect on therapeutic efficacy over longer periods as a result of enhanced reverse diffusion of drug to the blood circulation after the cessation of drug infusion, causing more rapid loss of drug in small tumours. [ABSTRACT FROM AUTHOR]

Vascularisation is a key feature of cancer growth, invasion and metastasis. To better understand the governing biophysical processes and their relative importance, it is instructive to develop physiologically representative mathematical models with which to compare to experimental data. Previous studies have successfully applied this approach to test the effect of various biochemical factors on tumour growth and angiogenesis. However, these models do not account for the experimentally observed dependency of angiogenic network evolution on growth-induced solid stresses. This work introduces two novel features: the effects of hapto- and mechanotaxis on vessel sprouting, and mechano-sensitive dynamic vascular remodelling. The proposed three-dimensional, multiscale, in-silico model of dynamically coupled angiogenic tumour growth is specified to in-vivo and in-vitro data, chosen, where possible, to provide a physiologically consistent description. The model is then validated against in-vivo data from murine mammary carcinomas, with particular focus placed on identifying the influence of mechanical factors. Crucially, we find that it is necessary to include hapto- and mechanotaxis to recapitulate observed time-varying spatial distributions of angiogenic vasculature. [ABSTRACT FROM AUTHOR]

Objectives: To investigate contrast-enhanced ultrasound (CEUS) with VEGFR2-targeted microbubbles for monitoring therapy effects of regorafenib on experimental colon carcinomas in rats with correlation to dynamic contrast-enhanced MRI (DCE-MRI) and immunohistochemistry. Materials and Methods: Human colorectal adenocarcinoma xenografts (HT-29) were implanted subcutaneously in n = 21 (n = 11 therapy group; n = 10 control group) female athymic nude rats (Hsd: RH-Foxn1rnu). Animals were imaged at baseline and after a one-week daily treatment with regorafenib or a placebo (10 mg/kg bodyweight), using CEUS with VEGFR2-targeted microbubbles and DCE-MRI. In CEUS tumor perfusion was assessed during an early vascular phase (wash-in area under the curve = WiAUC) and VEGFR2-specific binding during a late molecular phase (signal intensity after 8 (SI8min) and 10 minutes (SI10min)), using a conventional 15L8 linear transducer (transmit frequency 7 MHz, dynamic range 80 dB, depth 25 mm). In DCE-MRI functional parameters plasma flow (PF) and plasma volume (PV) were quantified. For validation purposes, CEUS parameters were correlated with DCE-MRI parameters and immunohistochemical VEGFR2, CD31, Ki-67 and TUNEL stainings. Results: CEUS perfusion parameter WiAUC decreased significantly (116,989 ± 77,048 a.u. to 30,076 ± 27,095a.u.; p = 0.005) under therapy with no significant changes (133,932 ± 65,960 a.u. to 84,316 ± 74,144 a.u.; p = 0.093) in the control group. In the therapy group, the amount of bound microbubbles in the late phase was significantly lower in the therapy than in the control group on day 7 (SI8min: 283 ± 191 vs. 802 ± 460 a.u.; p = 0.006); SI10min: 226 ± 149 vs. 645 ± 461 a.u.; p = 0.009). PF and PV decreased significantly (PF: 147 ± 58 mL/100 mL/min to 71 ± 15 mL/100 mL/min; p = 0.003; PV: 13 ± 3% to 9 ± 4%; p = 0.040) in the therapy group. Immunohistochemistry revealed significantly fewer VEGFR2 (7.2 ± 1.8 vs. 17.8 ± 4.6; p < 0.001), CD31 (8.1 ± 3.0 vs. 20.8 ± 5.7; p < 0.001) and Ki-67 (318.7 ± 94.0 vs. 468.0 ± 133.8; p = 0.004) and significantly more TUNEL (672.7 ± 194.0 vs. 357.6 ± 192.0; p = 0.003) positive cells in the therapy group. CEUS parameters showed significant (p < 0.05) correlations to DCE-MRI parameters and immunohistochemistry. Conclusions: CEUS with VEGFR2-targeted microbubbles allowed for monitoring regorafenib functional and molecular therapy effects on experimental colorectal adenocarcinomas with a significant decline of CEUS and DCE-MRI perfusion parameters as well as a significant reduction of specifically bound microbubbles under therapy, consistent with a reduced expression of VEGFR2. [ABSTRACT FROM AUTHOR]

• Comparison and quantification of effects of MNP injection rates on MNP distribution pattern. • Comparison and quantification of effects tumor physiological aspects in terms of diffusion coefficient on MNP distribution pattern. • Temperature profile evaluation based on different MNP distribution profiles. Effects of injection rate and tumor physiology on the diffusion of magnetic nano-particles (MNPs) and temperature profile during magnetic hyperthermia are investigated in this work. The study considers three injection rates (2.5 μL/min, 10 μL/min, and 40 μL/min), and two MNP diffusion coefficients (10−9 m2/s and 10−11 m2/s). The simulation of this physics has been done on 3D tumor surrounded by healthy tissue. Transient MNP distribution in tissue is evaluated using Darcy's flow model and the MNP transport (convection-diffusion) equation. The temperature profile in the tumor model is computed by solving Penne's bioheat transfer equation (PBHTE). Results show tumors with high collagen content (with low MNP diffusivity) are more restrictive towards MNP transport than tumors having low collagen content. Thus, tumors with low MNP diffusivity need a higher injection rate to increase the homogeneity of MNP concentration as well as temperature profile during thermo-therapy. Results also show that, MNP fluid injected with a higher injection rate produces a more uniform MNP concentration up to greater depth than the lower injection rate. [ABSTRACT FROM AUTHOR]

Hypoxia in the tumor microenvironment is the leading factor in angiogenesis. Angiogenesis can be identified by dynamic contrast-enhanced breast MRI (DCE MRI). Here we investigate the relationship between perfusion parameters on DCE MRI and angiogenic and prognostic factors in patients with invasive ductal carcinoma (IDC). Perfusion parameters (Ktrans, kep and ve) of 81 IDC were obtained using histogram analysis. Twenty-fifth, 50th and 75th percentile values were calculated and were analyzed for association with microvessel density (MVD), vascular endothelial growth factor (VEGF) and conventional prognostic factors. Correlation between MVD and ve50 was positive (r = 0.33). Ktrans50 was higher in tumors larger than 2 cm than in tumors smaller than 2 cm. In multivariate analysis, Ktrans50 was affected by tumor size and MVD with 12.8% explanation. There was significant association between Ktrans50 and tumor size and MVD. Therefore we conclude that DCE MRI perfusion parameters are potential imaging biomarkers for prediction of tumor angiogenesis and aggressiveness. [ABSTRACT FROM AUTHOR]

Approximately 30% of tumor endothelial cells have over-duplicated (>2) centrosomes, which may contribute to abnormal vessel function and drug resistance. Elevated levels of vascular endothelial growth factor A induce excess centrosomes in endothelial cells, but how other features of the tumor environment affect centrosome over-duplication is not known. To test this, we treated endothelial cells with tumor-derived factors, hypoxia, or reduced p53, and assessed centrosome numbers. We found that hypoxia and elevated levels of bone morphogenetic protein 2, 6 and 7 induced excess centrosomes in endothelial cells through BMPR1A and likely via SMAD signaling. In contrast, inflammatory mediators IL-8 and lipopolysaccharide did not induce excess centrosomes. Finally, down-regulation in endothelial cells of p53, a critical regulator of DNA damage and proliferation, caused centrosome over-duplication. Our findings suggest that some tumor-derived factors and genetic changes in endothelial cells contribute to excess centrosomes in tumor endothelial cells. [ABSTRACT FROM AUTHOR]

Both normal wound healing and tumor angiogenesis are mitigated by the sequential, carefully orchestrated release of growth stimulators and inhibitors. These regulators are released from platelet clots formed at the sites of activated endothelium in a temporally and spatially controlled manner, and the order of their release depends on their affinity to glycosaminoglycans (GAG) such as heparan sulfate (HS) within the extracellular matrix, and platelet open canallicular system. The formation of vessel sprouts, triggered by angiogenesis regulating factors with lowest affinities for heparan sulfate (e.g. VEGF), is followed by vessel-stabilizing PDGF-B or bFGF with medium affinity for HS, and by inhibitors such as PF-4 and TSP-1 with the highest affinities for HS. The invasive wound-like edge of growing tumors has an overabundance of angiogenesis stimulators, and we propose that their abundance out-competes angiogenesis inhibitors, effectively preventing inhibition of angiogenesis and vessel maturation. We evaluate this hypothesis using an experimentally motivated agent-based model, and propose a general theoretical framework for understanding mechanistic similarities and differences between the processes of normal wound healing and pathological angiogenesis from the point of view of competitive inhibition. [ABSTRACT FROM AUTHOR]

Purpose: To develop a general model that utilises a stochastic method to generate a vessel tree based on experimental data, and an associated irregular, macroscopic tumour. These will be used to evaluate two different methods for computing oxygen distribution. Methods: A vessel tree structure, and an associated tumour of 127 cm3, were generated, using a stochastic method and Bresenham’s line algorithm to develop trees on two different scales and fusing them together. The vessel dimensions were adjusted through convolution and thresholding and each vessel voxel was assigned an oxygen value. Diffusion and consumption were modelled using a Green’s function approach together with Michaelis-Menten kinetics. The computations were performed using a combined tree method (CTM) and an individual tree method (ITM). Five tumour sub-sections were compared, to evaluate the methods. Results: The oxygen distributions of the same tissue samples, using different methods of computation, were considerably less similar (root mean square deviation, RMSD≈0.02) than the distributions of different samples using CTM (0.001< RMSD<0.01). The deviations of ITM from CTM increase with lower oxygen values, resulting in ITM severely underestimating the level of hypoxia in the tumour. Kolmogorov Smirnov (KS) tests showed that millimetre-scale samples may not represent the whole. Conclusions: The stochastic model managed to capture the heterogeneous nature of hypoxic fractions and, even though the simplified computation did not considerably alter the oxygen distribution, it leads to an evident underestimation of tumour hypoxia, and thereby radioresistance. For a trustworthy computation of tumour oxygenation, the interaction between adjacent microvessel trees must not be neglected, why evaluation should be made using high resolution and the CTM, applied to the entire tumour. [ABSTRACT FROM AUTHOR]

The processes governing tumor growth and angiogenesis are codependent. To study the relationship between them, we proposed a coupled hybrid continuum-discrete model. In this model, tumor cells, their microenvironment (extracellular matrixes, matrix-degrading enzymes, and tumor angiogenic factors), and their network of blood vessels, described by a series of discrete points, were considered. The results of numerical simulation reveal the process of tumor growth and the change in microenvironment from avascular to vascular stage, indicating that the network of blood vessels develops gradually as the tumor grows. Our findings also reveal that a tumor is divided into three regions: necrotic, semi-necrotic, and well-vascularized. The results agree well with the previous relevant studies and physiological facts, and this model represents a platform for further investigations of tumor therapy. [ABSTRACT FROM AUTHOR]

Our previous studies revealed molecular alterations of tumor vessels, varying from immature to mature alterations, resulting from Abraxane, and demonstrated that the integrin-specific PET tracer 18F-FPPRGD2 can be used to noninvasively monitor such changes. However, changes in the tumor vasculature at functional levels such as perfusion and permeability are also important for monitoring Abraxane treatment outcomes in patients with cancer. The purpose of this study is to further investigate the vascular response during Abraxane therapy and the effectiveness of its synergistic interaction with cisplatin using Dynamic contrast enhanced-magnetic resonance imaging (DCE-MRI). Thirty MDA-MB-435 tumor mice were randomized into three groups: PBS control (C group), Abraxane only (A group), and sequential treatment with Abraxane followed by cisplatin (A-P group). Tumor volume was monitored based on caliper measurements. A DCE-MRI protocol was performed at baseline and day 3. The Ktrans, Kep and Ve were calculated and compared with CD31, α-SMA, and Ki67 histology data. Sequential treatment with Abraxane followed by cisplatin produced a significantly greater inhibition of tumor growth during the three weeks of the observation period. Decreases in Ktrans and Kep for the A and A-P groups were observed on day 3. Immunohistological staining suggested vascular remodeling during the Abraxane therapy. The changes in Ktrans and Kep values were correlated with alterations in the permeability of the tumor vasculature induced by the Abraxane treatment. In conclusion, Abraxane-mediated permeability variations in tumor vasculature can be quantitatively visualized by DCE-MRI, making this a useful method for studying the effects of early cancer treatment, especially the early vascular response. Vascular remodeling by Abraxane improves the efficiency of cisplatin delivery and thus results in a favorable treatment outcome. [ABSTRACT FROM AUTHOR]

Cancer-associated fibroblasts (CAFs), the most common constituent of the tumor stoma, are known to promote tumor initiation, progression and metastasis. However, the mechanism of how cancer cells transform normal fibroblasts (NFs) into CAFs is largely unknown. In this study, we determined the contribution of miRNAs in the transformation of NFs into CAFs. We found that miR-1 and miR-206 were down-regulated, whereas miR-31 was up-regulated in lung CAFs when compared with matched NFs. Importantly, modifying the expression of these three deregulated miRNAs induced a functional conversion of NFs into CAFs and vice versa. When the miRNA-reprogrammed NFs and CAFs were co-cultured with lung cancer cells (LCCs), a similar pattern of cytokine expression profiling were observed between two groups. Using a combination of cytokine expression profiling and miRNAs algorithms, we identified VEGFA/CCL2 and FOXO3a as direct targets of miR-1, miR-206 and miR-31, respectively. Importantly, systemic delivery of anti-VEGFA/CCL2 or pre-miR-1, pre-miR-206 and anti-miR-31 significantly inhibited tumor angiogenesis, TAMs accumulation, tumor growth and lung metastasis. Our results show that miRNAs-mediated FOXO3a/VEGF/CCL2 signaling plays a prominent role in LCCs-mediated NFs into CAFs, which may have clinical implications for providing novel biomarker(s) and potential therapeutic target(s) of lung cancer in the future. [ABSTRACT FROM AUTHOR]

Heparan sulphate (HS), a ubiquitously expressed glycosaminoglycan (GAG), regulates multiple cellular functions by mediating interactions between numerous growth factors and their cell surface cognate receptors. However, the structural specificity of HS in these interactions remains largely undefined. Here, we used completely synthetic, structurally defined, alternating N-sulfated glucosamine (NS) and 2-O-sulfated iduronate (IS) residues to generate dodecasaccharides ([NSIS]6) that contained no, one or six glucosamine 6-O-sulfates (6S). The aim was to address how 6S contributes to the potential of defined HS dodecasaccharides to inhibit the angiogenic growth factors FGF2 and VEGF165, in vitro and in vivo. We show that the addition of a single 6S at the non-reducing end of [NSIS]6, i.e. [NSIS6S]-[NSIS]5, significantly augments the inhibition of FGF2-dependent endothelial cell proliferation, migration and sprouting in vitro when compared to the non-6S variant. In contrast, the fully 6-O-sulfated dodecasaccharide, [NSIS6S]6, is not a potent inhibitor of FGF2. Addition of a single 6S did not significantly improve inhibitory properties of [NSIS]6 when tested against VEGF165-dependent endothelial cell functions.In vivo, [NSIS6S]-[NSIS]5 blocked FGF2-dependent blood vessel formation without affecting tumor growth. Reduction of non-FGF2-dependent ovarian tumor growth occurred when [NSIS6S]-[NSIS]5 was combined with cisplatin. The degree of inhibition by [NSIS6S]-[NSIS]5 in combination with cisplatin in vivo equated with that induced by bevacizumab and sunitinib when administered with cisplatin. Evaluation of post-treatment vasculature revealed that [NSIS6S]-[NSIS]5 treatment had the greatest impact on tumor blood vessel size and lumen formation. Our data for the first time demonstrate that synthetic, structurally defined oligosaccharides have potential to be developed as active anti-angiogenic agents that sensitize tumors to chemotherapeutic agents. [ABSTRACT FROM AUTHOR]

Endothelium-dependent angiogenesis is thought to be a crucial step in cancer progression. We previously reported that metastasis-associated in colon cancer-1 (MACC1) contributed to the vasculogenic mimicry in gastric cancer (GC), but it remains unknown whether MACC1 promotes endothelium-dependent angiogenesis of GC and whether TWIST1 is involved in this process. In the present study, we detected MACC1 expression and microvessel density (MVD) by immunohistochemistry in 159 patients with stage I-III GC, and investigated the role of TWIST1 and vascular endothelial growth factor A (VEGF-A) in MACC1-induced endothelium-dependent angiogenesis using nude mice with GC xenografts, and human umbilical vein endothelial cells (HUVECs) that were co-cultured with conditioned media from overexpression and interference MACC1 GC cells. We found that MACC1 expression was positively correlated with an increased MVD and tumor recurrence in GC patients. In GC xenograft models, MACC1 elevated MVD and upregulated the expression of VEGF-A as well as accelerated tumor growth. In addition, MACC1 obviously increased the expression of TWIST1 and induced tube-like formation of HUVECs, whereas attenuation of TWIST1 suppressed the protein expression of VEGF-A and repealed the effect of MACC1 on tube formation. Our findings shed light on the function of MACC1 in endothelium-dependent angiogenesis of GC and suggest potential prognostic and therapeutic value. [ABSTRACT FROM AUTHOR]

The Angiopoietin-2 (Ang2, Angpt2) growth factor is a context-dependent antagonist/agonist ligand of the endothelial Tie2 receptor tyrosine kinase and known to promote tumour angiogenesis and metastasis. Angiopoietin antagonists have been tested in clinical cancer trials in combination with VEGF-based anti-angiogenic therapy, including sunitinib, which is widely used as a first-line therapy for metastatic renal cell carcinoma (mRCC). However, little is known about Ang2 protein expression in human tumours and the correlation of tumour Ang2 expression with tumour vascularization, tumour cell proliferation and response to anti-angiogenic therapies. Here, we evaluated, using immunohistochemistry, the expression of Ang2, CD31 and the cell proliferation marker Ki-67 in the primary kidney cancer from 136 mRCC patients, who received first-line sunitinib after nephrectomy. Ang2 protein expression was restrained to RCC tumour vessels, and correlated with tumour vascularization and response to sunitinib. High pre-therapeutic Ang2 expression, and more strongly, combined high expression of both Ang2 and CD31, were associated with a high clinical benefit rate (CBR). Low cancer Ki-67 expression, but not Ang2 or CD31 expression, was associated with favourable progression-free (PFS) and overall survival (OS) as compared to patients with high Ki-67 expression (PFS 6.5 vs. 10.6 months, P = 0.009; OS, 15.7 vs. 28.5 months, P = 0.015). In summary, in this study to investigate endothelial Ang2 in mRCC patients treated with first-line sunitinib, high cancer Ang2 expression was associated with the CBR, but not PFS or OS, whereas low Ki-67 expression was significantly associated with long PFS and OS. [ABSTRACT FROM AUTHOR]

Background: Metronomic chemotherapy has shown promising activity against solid tumors and is believed to act in an antiangiogenic manner. The current study describes and quantifies the therapeutic efficacy, and mode of activity, of metronomic gemcitabine and a dedicated antiangiogenic agent (DC101) in patient-derived xenografts of pancreatic cancer. Methods: Two primary human pancreatic cancer xenograft lines were dosed metronomically with gemcitabine or DC101 weekly. Changes in tumor growth, vascular function, and metabolism over time were measured with magnetic resonance imaging, positron emission tomography, and immunofluorescence microscopy to determine the anti-tumor effects of the respective treatments. Results: Tumors treated with metronomic gemcitabine were 10-fold smaller than those in the control and DC101 groups. Metronomic gemcitabine, but not DC101, reduced the tumors' avidity for glucose, proliferation, and apoptosis. Metronomic gemcitabine-treated tumors had higher perfusion rates and uniformly distributed blood flow within the tumor, whereas perfusion rates in DC101-treated tumors were lower and confined to the periphery. DC101 treatment reduced the tumor's vascular density, but did not change their function. In contrast, metronomic gemcitabine increased vessel density, improved tumor perfusion transiently, and decreased hypoxia. Conclusion: The aggregate data suggest that metronomic gemcitabine treatment affects both tumor vasculature and tumor cells continuously, and the overall effect is to significantly slow tumor growth. The observed increase in tumor perfusion induced by metronomic gemcitabine may be used as a therapeutic window for the administration of a second drug or radiation therapy. Non-invasive imaging could be used to detect early changes in tumor physiology before reductions in tumor volume were evident. [ABSTRACT FROM AUTHOR]

CD73 (ecto-5'-nucleotidase), a cell surface enzyme hydrolyzing AMP to adenosine, was lately demonstrated to play a direct role in tumor progression including regulation of tumor vascularization. It was also shown to stimulate tumor macrophage infiltration. Interstitial adenosine, accumulating in solid tumors due to CD73 enzymatic activity, is recognized as a main mediator regulating the production of pro- and anti-angiogenic factors, but the engagement of specific adenosine receptors in tumor progression in vivo is still poorly researched. We have analyzed the role of high affinity adenosine receptors A1, A2A, and A3 in B16F10 melanoma progression using specific agonists (CCPA, CGS-21680 and IB-MECA, respectively). We limited endogenous extracellular adenosine background using CD73 knockout mice treated with CD73 chemical inhibitor, AOPCP (adenosine α,β-methylene 5’-diphosphate). Activation of any adenosine receptor significantly inhibited B16F10 melanoma growth but only at its early stage. At 14th day of growth, the decrease in tumor neovascularization and MAPK pathway activation induced by CD73 depletion was reversed by all agonists. Activation of A1AR primarily increased angiogenic activation measured by expression of VEGF-R2 on tumor blood vessels. However, mainly A3AR activation increased both the microvessel density and expression of pro-angiogenic factors. All agonists induced significant increase in macrophage tumor infiltration, with IB-MECA being most effective. This effect was accompanied by substantial changes in cytokines regulating macrophage polarization between pro-inflammatory and pro-angiogenic phenotype. Our results demonstrate an evidence that each of the analyzed receptors has a specific role in the stimulation of tumor angiogenesis and confirm significantly more multifaceted role of adenosine in its regulation than was already observed. They also reveal previously unexplored consequences to extracellular adenosine signaling depletion in recently proposed anti-CD73 cancer therapy. [ABSTRACT FROM AUTHOR]

Although zinc finger E-box binding homeobox 1 (ZEB1) has been identified as a key factor in the regulation of breast cancer differentiation and metastasis, its potential role in modulating tumor angiogenesis has not been fully examined. Here, we present the novel finding that conditioned medium derived from ZEB1-expressing MDA-MB-231 cells significantly increased the capillary tube formation of human umbilical vein endothelial cells (HUVECs), whereas ZEB1 knockdown by RNA interference had the opposite effect. ZEB1 caused marked upregulation of the expression of vascular endothelial growth factor A (VEGFA) at both mRNA and protein levels. Pre-incubation of HUVECs with anti-VEGFA neutralized antibody attenuated ZEB1-mediated tube formation of HUVECs. In breast cancer tissues, expression of ZEB1 was positively correlated with those of VEGFA and CD31. At the molecular level, ZEB1 activated VEGFA transcription by increasing SP1 recruitment to its promoter, which was mediated via the activation of PI3K and p38 pathways. Using a nude mouse xenograft model, we demonstrated that elevated expression of ZEB1 promotes in vivo tumorigenesis and angiogenesis in breast cancer. Collectively, we found that ZEB1-expressing breast cancer cells increase VEGFA production and thus stimulate tumor growth and angiogenesis via a paracrine mechanism. [ABSTRACT FROM AUTHOR]

Objective: To prepare arginine-glycine-aspartate (RGD)-targeted ultrasound contrast microbubbles (MBs) and explore the feasibility of their use in assessing dynamic changes in αvβ3 integrin expression in a murine model of tumor angiogenesis. Methods: RGD peptides were conjugated to the surfaces of microbubbles via biotin-avidin linkage. Microbubbles bearing RADfK peptides were prepared as controls. The RGD-MBs were characterized using an Accusizer 780 and optical microscopy. The binding specificity of the RGD-MBs for ανβ3-expressing endothelial cells (bEnd.3) was demonstrated in vitro by a competitive inhibition experiment. In an in vivo study, mice bearing tumors of three different stages were intravenously injected with RGD-MBs and subjected to targeted, contrast-enhanced, high-frequency ultrasound. Subsequently, tumors were harvested and sectioned for immunofluorescence analysis of ανβ3 expression. Results: The mean size of the RGD-MBs was 2.36 ± 1.7 μm. The RGD-MBs showed significantly higher adhesion levels to bEnd.3 cells compared to control MBs (P < 0.01). There was rarely binding of RGD-MBs to αvβ3-negative MCF-7 cells. Adhesion of the RGD-MBs to the bEnd.3 cells was significantly inhibited following treatment with anti-alpha(v) antibodies. The quantitative acoustic video intensity for high-frequency, contrast-enhanced ultrasound imaging of subcutaneous human laryngeal carcinoma (Hep-2) tumor xenografts was significantly higher in small tumors (19.89 ± 2.49) than in medium tumors (11.25 ± 2.23) and large tumors (3.38 ± 0.67) (P < 0.01). Conclusions: RGD-MBs enable noninvasive in vivo visualization of changes in tumor angiogenesis during tumor growth in subcutaneous cancer xenografts. [ABSTRACT FROM AUTHOR]

Glioblastoma multiforme is the most aggressive type of glioma and the most common malignant primary intra-axial brain tumor. In an effort to predict the evolution of the disease and optimize therapeutical decisions, several models have been proposed for simulating the growth pattern of glioma. One of the latest models incorporates cell proliferation and invasion, angiogenic net rates, oxygen consumption, and vasculature. These factors, particularly oxygenation levels, are considered fundamental factors of tumor heterogeneity and compartmentalization. This paper focuses on the initialization of the cancer cell populations and vasculature based on imaging examinations of the patient and presents a feasibility study on vasculature prediction over time. To this end, pharmacokinetic parameters derived from dynamic contrast-enhanced magnetic resonance imaging using Toft's model are used in order tofeed the model. Ktrans is used as a metric of the density of endothelial cells (vasculature); at the same time, it also helps to discriminate distinct image areas of interest, under a set of assumptions. Feasibility results of applying the model to a real clinical case are presented, including a study on the effect of certain parameters on the pattern of the simulated tumor. [ABSTRACT FROM AUTHOR]

The tumour microenvironment is complex and composed of many different constituents, including matricellular proteins such as connective tissue growth factor (CCN2), and is characterized by gradients in oxygen levels. In various cancers, hypoxia and CCN2 promote stem and progenitor cell properties, and regulate the proliferation, migration and phenotype of cancer cells. Our study was aimed at investigating the effects of hypoxia and CCN2 on chordoma cells, using the human U-CH1 cell line. We demonstrate that under basal conditions, U-CH1 cells express multiple CCN family members including CCN1, CCN2, CCN3 and CCN5. Culture of U-CH1 cells in either hypoxia or in the presence of recombinant CCN2 peptide promoted progenitor cell-like characteristics specific to the notochordal tissue of origin. Specifically, hypoxia induced the most robust increase in progenitor-like characteristics in U-CH1 cells, including increased expression of the notochord-associated markers T, CD24, FOXA1, ACAN and CA12, increased cell growth and tumour-sphere formation, and a decrease in the percentage of vacuolated cells present in the heterogeneous population. Interestingly, the effects of recombinant CCN2 peptide on U-CH1 cells were more pronounced under normoxia than hypoxia, promoting increased expression of CCN1, CCN2, CCN3 and CCN5, the notochord-associated markers SOX5, SOX6, T, CD24, and FOXA1 as well as increased tumour-sphere formation. Overall, this study highlights the importance of multiple factors within the tumour microenvironment and how hypoxia and CCN2 may regulate human chordoma cell behaviour. [ABSTRACT FROM AUTHOR]

Despite continuous improvements in therapeutic protocols, cancer-related mortality is still one of the main problems facing public health. The main cause of treatment failure is multi-drug resistance (MDR: simultaneous insensitivity to different anti-cancer agents), the underlying molecular and biological mechanisms of which include the activity of ATP binding cassette (ABC) proteins and drug compartmentalisation in cell organelles. We investigated the expression of the main ABC proteins and the role of cytoplasmic vacuoles in the MDR of six hepatocellular carcinoma (HCC) cell lines, and confirmed the accumulation of the yellow anti-cancer drug sunitinib in giant (four lines) and small cytoplasmic vacuoles of lysosomal origin (two lines). ABC expression analyses showed that the main ABC protein harboured by all of the cell lines was PGP, whose expression was not limited to the cell membrane but was also found on lysosomes. MTT assays showed that the cell lines with giant lysosomes were more resistant to sorafenib treatment than those with small lysosomes (p<0.01), and that verapamil incubation can revert this resistance, especially if it is administered after drug pre-incubation. The findings of this study demonstrate the involvement of PGP-positive lysosomes in drug sequestration and MDR in HCC cell lines. The possibility of modulating this mechanism using PGP inhibitors could lead to the development of new targeted strategies to enhance HCC treatment. [ABSTRACT FROM AUTHOR]

Objectives: To investigate the role of pericytes in constructing the malformed microvessels (MVs) and participating microvascular architecture heterogeneity of glioma. Methods: Forty human glioma tissue samples (WHO grade II-IV) were included in present study. Observation of blood vessel patterns, quantitative analysis of endothelial cells (ECs)- and pericyte-labeled MVs and comparison between malignant grades based on single- or double-immunohistochemical staining. The MV number density (MVND), microvascular pericyte number density (MPND), and microvascular pericyte area density (MPAD) were calculated. The expression of PDGFβ was also scored after immunostaining. Results: In grade II glioma, most of tumor MVs were the thin-wall CD34+ vessels with near normal morphology. In addition to thin-wall CD34+ MVs, more thick-wall MVs were found in grade III glioma, which often showed α-SMA positive. Most of MVs in grade IV glioma were in the form of plexus, curled cell cords and glomeruloid microvascular proliferation while the α-SMA+ cells were the main components. The MVs usually showed disordered arrangement, loose connection and active cell proliferation as shown by Ki67 and α-SMA coexpression. With the increase of glioma grades, the α-SMA+ MVND, CD34+ MVND and MPND were significantly augmented although the increase of CD34+ MVND but not MPAD was statistically insignificant between grade III and IV. It was interesting that some vessel-like structures only consist of α-SMA+ cells, assuming the guiding role of pericytes in angiogenesis. The expression level of PDGFβ was upregulated and directly correlated with the MPND in different glioma grades. Conclusion: Hyperplasia of pericytes was one of the significant characteristics of malignant glioma and locally proliferated pericytes were the main constituent of MVs in high grade glioma. The pathological characteristics of pericytes could be used as indexes of malignant grades of glioma. [ABSTRACT FROM AUTHOR]

The kynurenine pathway (KP) is the principal route of L-tryptophan (TRP) catabolism leading to the production of kynurenine (KYN), the neuroprotectants, kynurenic acid (KYNA) and picolinic acid (PIC), the excitotoxin, quinolinic acid (QUIN) and the essential pyridine nucleotide, nicotinamide adenine dinucleotide (NAD+). The enzymes indoleamine 2,3-dioxygenase-1 (IDO-1), indoleamine 2,3-dioxygenase-2 (IDO-2) and tryptophan 2,3-dioxygenase (TDO-2) initiate the first step of the KP. IDO-1 and TDO-2 induction in tumors are crucial mechanisms implicated to play pivotal roles in suppressing anti-tumor immunity. Here, we report the first comprehensive characterisation of the KP in 1) cultured human glioma cells and 2) plasma from patients with glioblastoma (GBM). Our data revealed that interferon-gamma (IFN-γ) stimulation significantly potentiated the expression of the KP enzymes, IDO-1 IDO-2, kynureninase (KYNU), kynurenine hydroxylase (KMO) and significantly down-regulated 2-amino-3-carboxymuconate semialdehyde decarboxylase (ACMSD) and kynurenine aminotransferase-I (KAT-I) expression in cultured human glioma cells. This significantly increased KP activity but significantly lowered the KYNA/KYN neuroprotective ratio in human cultured glioma cells. KP activation (KYN/TRP) was significantly higher, whereas the concentrations of the neuroreactive KP metabolites TRP, KYNA, QUIN and PIC and the KYNA/KYN ratio were significantly lower in GBM patient plasma (n = 18) compared to controls. These results provide further evidence for the involvement of the KP in glioma pathophysiology and highlight a potential role of KP products as novel and highly attractive therapeutic targets to evaluate for the treatment of brain tumors, aimed at restoring anti-tumor immunity and reducing the capacity for malignant cells to produce NAD+, which is necessary for energy production and DNA repair. [ABSTRACT FROM AUTHOR]

Recently, the correlation between the efficacy of platinum-based chemotherapy and ERCC1 expression in patients with SCLC has attracted wide-spread attention, and a lot of investigations have been conducted, whereas conflicting results were presented. Therefore, we performed the present meta-analysis of eligible studies to derive a more precise evaluation of the association between ERCC1 expression and the clinical outcome in SCLC patients receiving platinum-based chemotherapy. A literature search for relevant studies was conducted in the electronic databases of PubMed, EMBASE and Web of Science. The inclusive criteria were SCLC patients treated by platinum-based chemotherapy, and evaluated the relationship between ERCC1 expression and the clinical outcomes [including overall response rate (ORR), overall survival (OS) or progression-free survival (PFS)]. Odds ratio (OR) or hazard ratio (HR) with 95% confidence interval (CI) was calculated to assess the risk. A total of nine studies including 1129 patients were included in final analysis. Our analysis indicated that positive/high ERCC1 expression was associated with unfavorable OS (HR = 1.18, 95%CI = 1.02–1.37) and PFS (HR = 1.46, 95%CI = 1.14–1.88). Subgroup analysis according to disease stage suggested the significant relationship was found in limited stage (LS-SCLC), but not in extensive stage (ES-SCLC). However, no significant association was found between ERCC1 expression and ORR. Our analysis suggested ERCC1 expression may be a prognostic factor in SCLC patients receiving platinum-based chemotherapy, especially for LS-SCLC. [ABSTRACT FROM AUTHOR]

Galectin-3 (gal-3) is a β-galactoside binding protein related to many tumoral aspects, e.g. angiogenesis, cell growth and motility and resistance to cell death. Evidence has shown its upregulation upon hypoxia, a common feature in solid tumors such as glioblastoma multiformes (GBM). This tumor presents a unique feature described as pseudopalisading cells, which accumulate large amounts of gal-3. Tumor cells far from hypoxic/nutrient deprived areas express little, if any gal-3. Here, we have shown that the hybrid glioma cell line, NG97ht, recapitulates GBM growth forming gal-3 positive pseudopalisades even when cells are grafted subcutaneously in nude mice. In vitro experiments were performed exposing these cells to conditions mimicking tumor areas that display oxygen and nutrient deprivation. Results indicated that gal-3 transcription under hypoxic conditions requires previous protein synthesis and is triggered in a HIF-1α and NF-κB dependent manner. In addition, a significant proportion of cells die only when exposed simultaneously to hypoxia and nutrient deprivation and demonstrate ROS induction. Inhibition of gal-3 expression using siRNA led to protein knockdown followed by a 1.7–2.2 fold increase in cell death. Similar results were also found in a human GBM cell line, T98G. In vivo, U87MG gal-3 knockdown cells inoculated subcutaneously in nude mice demonstrated decreased tumor growth and increased time for tumor engraftment. These results indicate that gal-3 protected cells from cell death under hypoxia and nutrient deprivation in vitro and that gal-3 is a key factor in tumor growth and engraftment in hypoxic and nutrient-deprived microenvironments. Overexpression of gal-3, thus, is part of an adaptive program leading to tumor cell survival under these stressing conditions. [ABSTRACT FROM AUTHOR]