WNT/β-catenin signalling is crucial for intestinal homoeostasis. The intestinal epithelium and stroma are the major source of WNT ligands but their origin and role in intestinal stem cell (ISC) and ...epithelial repair remains unknown. Macrophages are a major constituent of the intestinal stroma. Here, we analyse the role of macrophage-derived WNT in intestinal repair in mice by inhibiting their release using a macrophage-restricted ablation of Porcupine, a gene essential for WNT synthesis. Such Porcn-depleted mice have normal intestinal morphology but are hypersensitive to radiation injury in the intestine compared with wild-type (WT) littermates. Porcn-null mice are rescued from radiation lethality by treatment with WT but not Porcn-null bone marrow macrophage-conditioned medium (CM). Depletion of extracellular vesicles (EV) from the macrophage CM removes WNT function and its ability to rescue ISCs from radiation lethality. Therefore macrophage-derived EV-packaged WNTs are essential for regenerative response of intestine against radiation.
Angiogenesis, the development of new capillary vessels, has a host of clinical manifestations. The identification of agents that increase or decrease angiogenesis is of great pharmaceutical interest. ...Classically, in vitro angiogenesis utilizes human umbilical vein endothelial cells (HUVEC) grown in matrigel. This valid and simple method has the drawbacks that each cell population is distinct and the constraint of obtaining primary source material. Herein we utilize the established EA.hy926 endothelial cell line as our model for in vitro angiogenesis and present a novel formula to quantify endothelial cell remodeling to identify pro- and anti-angiogenic agents. Furthermore, our technique details the procedures to identify and quantify compounds that have the capacity to generate pro- or anti-angiogenic factors when given to non-endothelial cells, which we define herein as angiogenic potential. In conclusion, we propose a novel formula that we are confident accurately reflects the degree of in vitro angiogenesis allowing the quantification of prospective angiogenic compounds.
The current process of medical terminology coding in health standards at the Hospital of Clinics of Paraguay is performed by physicians, normally interns or residents. They search the medical ...terminology code in impressed coding manuals or in internet by using their cell phones. This search process takes a lot time to be done during the medical consultation. This work proposes and evaluates a friendly medical terminology server thought web services, using Apache Lucene, as a search engine library, and the Metathesaurus of Unified System of Medical Language (UMLS), as an information source. The server is developed for Spanish speakers. Results show that physicians can find medical terminology code with the terminology server, using friendly or familiar terms, 18 times faster than with the current search process. The user satisfaction degree is ``Good” according to an adjective rating of the System Usability Scale (SUS). In addition, a comparison with a search engine of medical terminology called Metamorphosys shows that the implemented terminology server is quite competitive and it responses in similar average time
The majority of breast cancer specific deaths in women with estrogen receptor positive (ER+) tumors occur due to metastases that are resistant to therapy. There is a critical need for novel ...therapeutic approaches to achieve tumor regression and/or maintain therapy responsiveness in metastatic ER+ tumors. The objective of this study was to elucidate the role of metabolic pathways that undermine therapy efficacy in ER+ breast cancers. Our previous studies identified Exportin 1 (XPO1), a nuclear export protein, as an important player in endocrine resistance progression and showed that combining selinexor (SEL), an FDA-approved XPO1 antagonist, synergized with endocrine agents and provided sustained tumor regression. In the current study, using a combination of transcriptomics, metabolomics and metabolic flux experiments, we identified certain mitochondrial pathways to be upregulated during endocrine resistance. When endocrine resistant cells were treated with single agents in media conditions that mimic a nutrient deprived tumor microenvironment, their glutamine dependence for continuation of mitochondrial respiration increased. The effect of glutamine was dependent on conversion of the glutamine to glutamate, and generation of NAD+. PGC1α, a key regulator of metabolism, was the main driver of the rewired metabolic phenotype. Remodeling metabolic pathways to regenerate new vulnerabilities in endocrine resistant breast tumors is novel, and our findings reveal a critical role that ERα-XPO1 crosstalk plays in reducing cancer recurrences. Combining SEL with current therapies used in clinical management of ER+ metastatic breast cancer shows promise for treating and keeping these cancers responsive to therapies in already metastasized patients.
Progesterone and progestins have been demonstrated to enhance breast cancer cell migration, although the mechanisms are still not fully understood. The protease-activated receptors (PARs) are a ...family of membrane receptors that are activated by serine proteases in the blood coagulation cascade. PAR1 (F2R) has been reported to be involved in cancer cell migration and overexpressed in breast cancer. We herein demonstrate that PAR1 mRNA and protein are upregulated by progesterone treatment of the breast cancer cell lines ZR-75 and T47D. This regulation is dependent on the progesterone receptor (PR) but does not require PR phosphorylation at serine 294 or the PR proline-rich region mPRO. The increase in PAR1 mRNA was transient, being present at 3 h and returning to basal levels at 18 h. The addition of a PAR1-activating peptide (aPAR1) to cells treated with progesterone resulted in an increase in focal adhesion (FA) formation as measured by the cellular levels of phosphorylated FA kinase. The combined but not individual treatment of progesterone and aPAR1 also markedly increased stress fiber formation and the migratory capacity of breast cancer cells. In agreement with in vitro findings, data mining from the Oncomine platform revealed that PAR1 expression was significantly upregulated in PR-positive breast tumors. Our observation that PAR1 expression and signal transduction are modulated by progesterone provides new insight into how the progestin component in hormone therapies increases the risk of breast cancer in postmenopausal women.
Lysosomes degrade cellular proteins and organelles and regulate cell signaling by providing a surface for the formation of critical protein complexes, notably molecular target of rapamycin (mTOR) ...complex 1 (mTORC1). Striking differences in the lysosomes of cancer versus normal cells suggest that they could be targets for drug development. Although the lysomotropic drugs chloroquine (CQ) and hydroxychloroquine (HCQ) have been widely investigated, studies have focused on their ability to inhibit autophagy. We synthesized a novel compound, called EAD1, which is structurally related to CQ but is a 14-fold more potent inhibitor of cell proliferation. Here we find that EAD1 causes rapid relocation, membrane permeabilization (LMP), and deacidification of lysosomes, and it induces apoptosis and irreversibly blocks proliferation of human lung cancer H460, H520, H1299, HCC827, and H1703 cells. EAD1 causes dissociation of mTOR from lysosomes and increases mTOR's perinuclear versus cytoplasmic localization, changes previously shown to inactivate mTORC1. The effect on mTOR was not seen with HCQ, even at >10-fold greater concentrations. Phosphorylation of a downstream target of mTORC1, ribosomal protein S6, was inhibited by EAD1. Although EAD1 also inhibited autophagy, it retained full antiproliferative activity in autophagy-deficient H1650 lung cancer cells, which have a biallelic deletion of Atg7, and in H460 Atg7-knockout cells. As Atg7 is critical for the canonical autophagy pathway, it is likely that inhibition of autophagy is not how EAD1 inhibits cell proliferation. Further studies are needed to determine the relationship of LMP to mTORC1 disruption and their relative contributions to drug-induced cell death. These studies support the lysosome as an underexplored target for new drug development.
Angiogenesis, the growth of new blood vessels, involves specification of endothelial cells to tip cells and stalk cells, which is controlled by Notch signalling, whereas vascular endothelial growth ...factor receptor (VEGFR)-2 and VEGFR-3 have been implicated in angiogenic sprouting. Surprisingly, we found that endothelial deletion of Vegfr3, but not VEGFR-3-blocking antibodies, postnatally led to excessive angiogenic sprouting and branching, and decreased the level of Notch signalling, indicating that VEGFR-3 possesses passive and active signalling modalities. Furthermore, macrophages expressing the VEGFR-3 and VEGFR-2 ligand VEGF-C localized to vessel branch points, and Vegfc heterozygous mice exhibited inefficient angiogenesis characterized by decreased vascular branching. FoxC2 is a known regulator of Notch ligand and target gene expression, and Foxc2(+/-);Vegfr3(+/-) compound heterozygosity recapitulated homozygous loss of Vegfr3. These results indicate that macrophage-derived VEGF-C activates VEGFR-3 in tip cells to reinforce Notch signalling, which contributes to the phenotypic conversion of endothelial cells at fusion points of vessel sprouts.
Abstract
Angiogenesis is a critical component of the initiation, maintenance and progression of tumors, and has been the target of extensive drug development. In clinical practice there are several ...approved and investigational anti-angiogenesis drugs, the majority of which primarily target the VEGF pathway, however their long term effectiveness remains limited. In designing new drug regimens, it is important to consider other known angiogenic factors that likely contribute to the pro-angiogenic phenotype. We evaluated two such factors: thymidine phosphorylase (TP) (aka platelet-derived endothelial cell growth factor, PD-ECGF), an enzyme that can stimulate endothelial cell migration, and angiopoietin 2 (Ang2), a member of the Ang/Tie2 signaling pathway. To target TP, we used a novel, highly specific, orally available small molecule inhibitor, which we had previously designed, named AEAC. To target Ang2, we used an Ang2 neutralizing peptibody, L1-7N (provided by CTEP-NCI), which is a genetically engineered peptide-Fc fusion protein that specifically binds Ang2. In a mouse NCI-H460 NSCLC sc xenograft model, AEAC and L1-7N produced 33% and 67% reductions, respectively, in tumor growth, and the combination produced a small, but statistically significant further reduction, when compared to either agent used alone. Tumor growth inhibition was paralleled by a reduction in Ki-67 staining. L1-7N also produced a large infiltration of macrophages (F4/80+) into the tumors after treatment. We also tested the drug combination in a mouse genetic model of pancreatic neuroendocrine tumors (PNET). PNET are one the few human solid tumors that are clinically responsive to angiogenesis inhibitors used alone. These mice have a conditional knockout of the Men1 gene in the endocrine pancreatic cells (Pdx1-Cre;Men1 mice) mimicking a primary genetic alteration seen in human disease. Men1 KO PNET experiments were initiated when the mice were 10-11 months old, and drug treatment was daily (5 days per week) for 3 weeks. To date, we have not seen any antiangiogenic or antitumor effects of the drugs. Upon sacrifice, the mice were found to have extensive insulinomas, involving around 70% of the entire pancreas. CD31(+) and F4/80(+) cells in the pancreas showed that most of them concentrated inside the PNET, however there was no change after the treatments. Studies are currently underway to determine if higher doses and/or starting treatment at an earlier time would be more efficacious.
Citation Format: Evelyn Aranda, Ziqiang Yuan, Steven K. Libutti, Edward L. Schwartz. Antitumor activity of agents targeting the angiogenic factors angiopoietin-2 and thymidine phosphorylase in mouse models of lung and pancreatic neuroendocrine cancer abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1822. doi:10.1158/1538-7445.AM2017-1822
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