bluuebunny/crossref_metadata_2025 huggingface dataset:
Transcriptional Regulation of the Gene Cluster Encoding Allantoinase and Guanine Deaminase in Klebsiella pneumoniae
Authors: Guzmán Karla , Badia Josefa , Giménez Rosa , Aguilar Juan , Baldoma Laura
Published: February 2011

Purines can be used as the sole source of nitrogen by several strains of
K. pneumoniae
under aerobic conditions. The genes responsible for the assimilation of purine nitrogens are distributed in three separated clusters in the
K. pneumoniae
genome. Here, we characterize the cluster encompassing genes KPN_01787 to KPN_01791, which is involved in the conversion of allantoin into allantoate and in the deamination of guanine to xanthine. These genes are organized in three transcriptional units,
hpxSAB
,
hpxC
, and
guaD
. Gene
hpxS
encodes a regulatory protein of the GntR family that mediates regulation of this system by growth on allantoin. Proteins encoded by
hpxB
and
guaD
display allantoinase and guanine deaminase activity, respectively. In this cluster,
hpxSAB
is the most tightly regulated unit. This operon was activated by growth on allantoin as a nitrogen source; however, addition of allantoin to nitrogen excess cultures did not result in
hpxSAB
induction. Neither
guaD
nor
hpxC
was induced by allantoin. Expression of
guaD
is mainly regulated by nitrogen availability through the action of NtrC. Full induction of
hpxSAB
by allantoin requires both HpxS and NAC. HpxS may have a dual role, acting as a repressor in the absence of allantoin and as an activator in its presence. HpxS binds to tandem sites, S1 and S2, overlapping the −10 and −35 sequences of the
hpxSAB
promoter, respectively. The NAC binding site is located between S1 and S2 and partially overlaps S2. In the presence of allantoin, interplay between NAC and HpxS is proposed.
Republished paper: Managing HBV in patients with impaired immunity
Authors: Wursthorn Karsten , Wedemeyer Heiner , Manns Michael P
Published: February 2011

Chronic hepatitis B is one of the most common infectious diseases worldwide. In patients with an impaired immune system the prevalence of HBsAg is even higher and the course of hepatitis B infection is often aggravated. In HIV/HBV co-infected patients, liver related morbidity and mortality can be reduced by implementing highly active antiretroviral treatment (HAART) that contains substances active against HBV. Reactivation of HBV during chemotherapy may occur in HBsAg positive patients but can even occur in serologically recovered anti-HBc positive, HBsAg negative patients resulting in high mortality from liver disease. HBsAg positive patients irrespective of HBV DNA levels should receive preemptive treatment with HBV polymerase inhibitors which should be continued for 12 months after cessation of chemo- and or immunosuppressive therapy. The combination prophylaxis of passive immunisations with hepatitis B immunoglobulins (HBIG) and nucleos(t)ide analogues (NUC) is able to reduce HBV recurrence rates after transplantation to 0–10%. This review will summarise the current knowledge on pathogenesis, frequency and treatment options of HBV reactivations in patients with impaired immunity.
Cadmium Inhibition of Erythropoiesis in Goldfish,Carassius auratus
Authors: Houston A. H. , Keen J. E.
Published: February 2011

Goldfish (Carassius auratus) maintained on a highly nutritional diet under normoxic circumstances at 25 °C in soft water had mean half-recovery times of 22.5 h (hemoglobin) and 21.0 h (hematocrit) following anemia induced with phenylhydrazine HCl. Corresponding values for fish exposed under similar conditions to 1, 5, 15, and 25% of the softwater 240-h cadmium LC50 were 41.9, 53.1, 164.8, and 344.3 h (hemoglobin) and 44.7, 46.5, 348.9, and 263.2 h (hematocrit), respectively. We suggest that whereas cadmium has relatively little effect upon circulating erythrocytes, it seriously impedes red cell formation and hemoglobin synthesis at concentrations well below the acutely lethal level.
Impact of body position on central and peripheral hemodynamic contributions to movement-induced hyperemia: implications for rehabilitative medicine
Authors: Trinity Joel D. , McDaniel John , Venturelli Massimo , Fjeldstad Anette S. , Ives Stephen J. , Witman Melissa A. H. , Barrett-O'Keefe Zachary , Amann Markus , Wray D. Walter , Richardson Russell S.
Published: February 2011

This study used alterations in body position to identify differences in hemodynamic responses to passive exercise. Central and peripheral hemodynamics were noninvasively measured during 2 min of passive knee extension in 14 subjects, whereas perfusion pressure (PP) was directly measured in a subset of 6 subjects. Movement-induced increases in leg blood flow (LBF) and leg vascular conductance (LVC) were more than twofold greater in the upright compared with supine positions (LBF, supine: 462 ± 6, and upright: 1,084 ± 159 ml/min, P < 0.001; and LVC, supine: 5.3 ± 1.2, and upright: 11.8 ± 2.8 ml·min−1·mmHg−1, P < 0.002). The change in heart rate (HR) from baseline to peak was not different between positions (supine: 8 ± 1, and upright: 10 ± 1 beats/min, P = 0.22); however, the elevated HR was maintained for a longer duration when upright. Stroke volume contributed to the increase in cardiac output (CO) during the upright movement only. CO increased in both positions; however, the magnitude and duration of the CO response were greater in the upright position. Mean arterial pressure and PP were higher at baseline and throughout passive movement when upright. Thus exaggerated central hemodynamic responses characterized by an increase in stroke volume and a sustained HR response combined to yield a greater increase in CO during upright movement. This greater central response coupled with the increased PP and LVC explains the twofold greater and more sustained increase in movement-induced hyperemia in the upright compared with supine position and has clinical implications for rehabilitative medicine.
Human cord blood CD34+ progenitor cells acquire functional cardiac properties through a cell fusion process
Authors: Avitabile Daniele , Crespi Alessia , Brioschi Chiara , Parente Valeria , Toietta Gabriele , Devanna Paolo , Baruscotti Mirko , Truffa Silvia , Scavone Angela , Rusconi Francesca , Biondi Andrea , D'Alessandra Yuri , Vigna Elisa , DiFrancesco Dario , Pesce Maurizio , Capogrossi Maurizio C. , Barbuti Andrea
Published: February 2011

The efficacy of cardiac repair by stem cell administration relies on a successful functional integration of injected cells into the host myocardium. Safety concerns have been raised about the possibility that stem cells may induce foci of arrhythmia in the ischemic myocardium. In a previous work ( 36 ), we showed that human cord blood CD34+ cells, when cocultured on neonatal mouse cardiomyocytes, exhibit excitation-contraction coupling features similar to those of cardiomyocytes, even though no human genes were upregulated. The aims of the present work are to investigate whether human CD34+ cells, isolated after 1 wk of coculture with neonatal ventricular myocytes, possess molecular and functional properties of cardiomyocytes and to discriminate, using a reporter gene system, whether cardiac differentiation derives from a (trans)differentiation or a cell fusion process. Umbilical cord blood CD34+ cells were isolated by a magnetic cell sorting method, transduced with a lentiviral vector carrying the enhanced green fluorescent protein (EGFP) gene, and seeded onto primary cultures of spontaneously beating rat neonatal cardiomyocytes. Cocultured EGFP+/CD34+-derived cells were analyzed for their electrophysiological features at different time points. After 1 wk in coculture, EGFP+ cells, in contact with cardiomyocytes, were spontaneously contracting and had a maximum diastolic potential (MDP) of −53.1 mV, while those that remained isolated from the surrounding myocytes did not contract and had a depolarized resting potential of −11.4 mV. Cells were then resuspended and cultured at low density to identify EGFP+ progenitor cell derivatives. Under these conditions, we observed single EGFP+ beating cells that had acquired an hyperpolarization-activated current typical of neonatal cardiomyocytes (EGFP+ cells, −2.24 ± 0.89 pA/pF; myocytes, −1.99 ± 0.63 pA/pF, at −125 mV). To discriminate between cell autonomous differentiation and fusion, EGFP+/CD34+ cells were cocultured with cardiac myocytes infected with a red fluorescence protein-lentiviral vector; under these conditions we found that 100% of EGFP+ cells were also red fluorescent protein positive, suggesting cell fusion as the mechanism by which cardiac functional features are acquired.
Heterogeneous function of ryanodine receptors, but not IP3receptors, in hamster cremaster muscle feed arteries and arterioles
Authors: Westcott Erika B. , Jackson William F.
Published: February 2011

The roles played by ryanodine receptors (RyRs) and inositol 1,4,5-trisphosphate receptors (IP3Rs) in vascular smooth muscle in the microcirculation remain unclear. Therefore, the function of both RyRs and IP3Rs in Ca2+signals and myogenic tone in hamster cremaster muscle feed arteries and downstream arterioles were assessed using confocal imaging and pressure myography. Feed artery vascular smooth muscle displayed Ca2+sparks and Ca2+waves, which were inhibited by the RyR antagonists ryanodine (10 μM) or tetracaine (100 μM). Despite the inhibition of sparks and waves, ryanodine or tetracaine increased global intracellular Ca2+and constricted the arteries. The blockade of IP3Rs with xestospongin D (5 μM) or 2-aminoethoxydiphenyl borate (100 μM) or the inhibition of phospholipase C using U-73122 (10 μM) also attenuated Ca2+waves without affecting Ca2+sparks. Importantly, the IP3Rs and phospholipase C antagonists decreased global intracellular Ca2+and dilated the arteries. In contrast, cremaster arterioles displayed only Ca2+waves: Ca2+sparks were not observed, and neither ryanodine (10–50 μM) nor tetracaine (100 μM) affected either Ca2+signals or arteriolar tone despite the presence of functional RyRs as assessed by responses to the RyR agonist caffeine (10 mM). As in feed arteries, arteriolar Ca2+waves were attenuated by xestospongin D (5 μM), 2-aminoethoxydiphenyl borate (100 μM), and U-73122 (10 μM), accompanied by decreased global intracellular Ca2+and vasodilation. These findings highlight the contrasting roles played by RyRs and IP3Rs in Ca2+signals and myogenic tone in feed arteries and demonstrate important differences in the function of RyRs between feed arteries and downstream arterioles.
Phosphatidylinositol 3′-kinase signaling pathway is essential for Rac1-induced hypoxia-inducible factor-1α and vascular endothelial growth factor expression
Authors: Xue Yan , Li Nan-Lin , Yang Jing-Yue , Chen Yan , Yang Lu-Lu , Liu Wen-Chao
Published: February 2011

We have previously demonstrated the roles of RhoA, Rac1, and Cdc42 in hypoxia-driven angiogenesis. However, the role of oncogenes in hypoxia signaling is poorly understood. Given the importance of Rho proteins in the hypoxic response, we hypothesized that Rho family members could act as mediators of hypoxic signal transduction. We investigated the cross-talk between hypoxia and oncogene-driven signal transduction pathways and explored the role of Rac1 on hypoxia-induced hypoxia-inducible factor (HIF)-1α and VEGF expression. Since the phosphatidylinositol 3′-kinase (PI3K) pathway is involved in signal transduction of many oncogenes, we explored the role of PI3K on Rac1-mediated expression of HIF-1α and VEGF in hypoxia. We showed that LY-294002, a PI3K inhibitor, suppressed HIF-1α and VEGF induction under hypoxic conditions by up to 50%. Activation of Rac1 resulted in an upregulation of hypoxia-induced HIF-1α expression, which was blocked by LY-294002. These data suggested that Rac1 is an intermediate in the PI3K-mediated induction of HIF-1α. Interestingly, there was a significant downregulation of the tumor suppressor genes p53 and von Hippel-Lindau tumor suppressor (VHL) in cells expressing a constitutively active form of Rac1. Rac1-mediated inhibition of p53 and VHL could therefore be implicated in the upregulation of HIF-1α expression.
Rapamycin inhibits hydrogen peroxide-induced loss of vascular contractility
Authors: Gao Ge , Li Jing-Jing , Li Yuenan , Li Dong , Wang Yuebing , Wang Lifeng , Tang Xiang D. , Walsh Michael P. , Gui Yu , Zheng Xi-Long
Published: February 2011

Rapamycin, an inhibitor of the mammalian target of rapamycin (mTOR) pathway, has been shown to extend the life span of mice, and oxidative stress plays critical roles in vascular aging involving loss of compliance of arteries. We examined, therefore, whether rapamycin has protective effects on the inhibition of vascular contractility by hydrogen peroxide (H2O2). Prolonged (3 h) exposure to H2O2 induced complete loss of contraction of mouse aortic rings and mesenteric (resistance) arteries to either KCl or phenylephrine, which was attenuated by pretreatment with rapamycin. H2O2-induced loss of contractility was unaffected by treatment with actinomycin D or cycloheximide, inhibitors of gene transcription and protein synthesis, respectively. Western blot analysis showed that there was no increase in phosphorylation of S6 kinase 1 (S6K) or factor 4E binding protein 1 (4EBP1) in response to H2O2 treatment, suggesting involvement of the mTOR complex-2 (mTORC2) rather than mTORC1. H2O2 treatment inhibited phosphorylation of the 20-kDa regulatory light chains of myosin (LC20), which was partially blocked by rapamycin treatment. Interestingly, the calcineurin inhibitors cyclosporine A and FK506 were found to mimic the rapamycin effect, and rapamycin inhibited calcineurin activation induced by H2O2. We conclude that rapamycin inhibits H2O2-induced loss of vascular contractility, likely through an mTORC2-calcineurin pathway.
Myocardin is differentially required for the development of smooth muscle cells and cardiomyocytes
Authors: Hoofnagle Mark H. , Neppl Ronald L. , Berzin Erica L. , Teg Pipes G. C. , Olson Eric N. , Wamhoff Brian W. , Somlyo Avril V. , Owens Gary K.
Published: February 2011

Myocardin is a serum response factor (SRF) coactivator exclusively expressed in cardiomyocytes and smooth muscle cells (SMCs). However, there is highly controversial evidence as to whether myocardin is essential for normal differentiation of these cell types, and there are no data showing whether cardiac or SMC subtypes exhibit differential myocardin requirements during development. Results of the present studies showed the virtual absence of myocardin−/−visceral SMCs or ventricular myocytes in chimeric myocardin knockout (KO) mice generated by injection of myocardin−/−embryonic stem cells (ESCs) into wild-type (WT; i.e., myocardin+/+ESC) blastocysts. In contrast, myocardin−/−ESCs readily formed vascular SMC, albeit at a reduced frequency compared with WT ESCs. In addition, myocardin−/−ESCs competed equally with WT ESCs in forming atrial myocytes. The ultrastructural features of myocardin−/−vascular SMCs and cardiomyocytes were unchanged from their WT counterparts as determined using a unique X-ray microprobe transmission electron microscopic method developed by our laboratory. Myocardin−/−ESC-derived SMCs also showed normal contractile properties in an in vitro embryoid body SMC differentiation model, other than impaired thromboxane A2 responsiveness. Together, these results provide novel evidence that myocardin is essential for development of visceral SMCs and ventricular myocytes but is dispensable for development of atrial myocytes and vascular SMCs in the setting of chimeric KO mice. In addition, results suggest that as yet undefined defects in development and/or maturation of ventricular cardiomyocytes may have contributed to early embryonic lethality observed in conventional myocardin KO mice and that observed deficiencies in development of vascular SMC may have been secondary to these defects.
Infection-induced coronary dysfunction and systemic inflammation in piglets are dampened in hypercholesterolemic milieu
Authors: Birck Malene M. , Pesonen Erkki , Odermarsky Michal , Hansen Axel K. , Persson Kenneth , Frikke-Schmidt Henriette , Heegaard Peter M. H. , Liuba Petru
Published: February 2011

The synergism of infection with conventional cardiovascular risk factors in atherosclerosis is much debated. We hypothesized that coronary arterial injury correlates with infection recurrence and pathogen burden and is further aggravated by hypercholesterolemia. Forty-two Göttingen minipigs were assigned to repeated intratracheal inoculation of PBS, Chlamydia pneumoniae (Cpn), or both Cpn and influenza virus at 8, 11, and 14 wk of age. Animals were fed either standard or 2% cholesterol diet (chol-diet). At 19 wk of age coronary vasomotor responses to acetylcholine (ACh) and adenosine were assessed in vivo and blood and tissue samples were collected. Nonparametric tests were used to compare the groups. In cholesterol-fed animals, total cholesterol/HDL was significantly increased in infected animals compared with noninfected animals [3.13 (2.17–3.38) vs. 2.03 (1.53–2.41), respectively; P = 0.01]. C-reactive protein (CRP) rose in infected animals [10.60 (4.96–18.00) vs. 2.47 (1.44–3.01) μg/ml in noninfected; P < 0.01] without significant difference between the mono- and coinfected groups. Among coinfected animals, both CRP and haptoglobin were lower in those fed chol-diet than in those fed standard diet ( P < 0.05). The vasoconstricting response to ACh was most prominent in coinfected animals {769.3 (594–1,129) cm; P = 0.03 vs. noninfected [342 (309–455) cm] and P = 0.07 vs. monoinfected [415 (252.5–971.8) cm]}. Among monoinfected animals, similar to CRP, a trend for less vasoconstriction was observed in those fed chol-diet ( P = 0.08). Coinfection of piglets appears to be associated with more pronounced coronary muscarinic vasomotor dysfunction. In monoinfected animals, use of chol-diet seems to dampen both coronary dysfunction and systemic inflammation induced by infection.
A model of chronic inflammation and pulmonary emphysema after multiple ozone exposures in mice
Authors: Triantaphyllopoulos Kostas , Hussain Farhana , Pinart Mariona , Zhang Min , Li Feng , Adcock Ian , Kirkham Paul , Zhu Jie , Chung Kian Fan
Published: February 2011

Oxidative stress plays a role in the pathophysiology of emphysema through the activation of tissue proteases and apoptosis. We examined the effects of ozone exposure by exposing BALB/c mice to either a single 3-h exposure or multiple exposures over 3 or 6 wk, with two 3-h exposures per week. Compared with air-exposed mice, the increase in neutrophils in bronchoalveolar lavage fluid and lung inflammation index was greatest in mice exposed for 3 and 6 wk. Lung volumes were increased in 3- and 6-wk-exposed mice but not in single-exposed. Alveolar space and mean linear intercept were increased in 6- but not 3-wk-exposed mice. Caspase-3 and apoptosis protease activating factor-1 immunoreactivity was increased in the airway and alveolar epithelium and macrophages of 3- and 6-wk-exposed mice. Interleukin-13, keratinocyte chemoattractant, caspase-3, and IFN-γ mRNA were increased in the 6-wk-exposed group, but heme oxygenase-1 (HO-1) mRNA decreased. matrix metalloproteinase-12 (MMP-12) and caspase-3 protein expression increased in lungs of 6-wk-exposed mice. Collagen area increased and epithelial area decreased in airway wall at 3- and 6-wk exposure. Exposure of mice to ozone for 6 wk induced a chronic inflammatory process, with alveolar enlargement and damage linked to epithelial apoptosis and increased protease expression.
Analysis of the Oxidative Damage Repair GenesNUDT1,OGG1, andMUTYHin Patients from Mismatch Repair Proficient HNPCC Families (MSS-HNPCC)
Authors: Garre Pilar , Briceño Verónica , Xicola Rosa M. , Doyle Brian J. , de la Hoya Miguel , Sanz Julián , Llovet Patricia , Pescador Paula , Puente Javier , Díaz-Rubio Eduardo , Llor Xavier , Caldés Trinidad
Published: February 2011

Purpose: Several studies have described molecular differences between microsatellite stable hereditary nonpolyposis colorectal cancer (MSS-HNPCC) and microsatellite unstable Lynch syndrome tumors (MSI-HNPCC). These differences highlight the possibility that other instability forms could explain cancer susceptibility in this group of families.

The base excision repair (BER) pathway is the major DNA repair pathway for oxidative DNA damage. A defect in this pathway can result in DNA transversion mutations and a subsequent increased cancer risk. Mutations in MUTYH have been associated with increased colorectal cancer (CRC) risk while no association has been described for OGG1 or NUDT1.

Experimental Design: We performed mutational screening of the three genes involved in defense against oxidative DNA damage in a set of 42 MSS-HNPCC families.

Results: Eight rare variants and 5 frequent variants were found in MSS-HNPCC patients. All variants were previously described by other authors except variant c.285C>T in OGG1. Segregation studies were done and in silico programs were used to estimate the level of amino acid conservation, protein damage prediction, and possible splicing alterations. Variants OGG1 c.137G>A; MUTYH c.1187G>A were detected in Amsterdam I families and cosegregate with cancer. Analysis of OGG1 c.137G>A transcripts showed an inactivation of the splicing donor of exon 1.

Conclusions: Two rare variants (OGG1 c.137G>A; MUTYH c.1187G>A) and one common polymorphism (NUDT1 c.426C>T) were associated with CRC risk. We show that the BER pathway can play a significant role in a number of MSS-HNPCC colorectal cancers. More studies could be of interest in order to gain further understanding of yet unexplained CRC susceptibility cases. Clin Cancer Res; 17(7); 1701–12. ©2011 AACR.
The A/G Allele of Rs16906252 Predicts for MGMT Methylation and Is Selectively Silenced in Premalignant Lesions from Smokers and in Lung Adenocarcinomas
Authors: Leng Shuguang , Bernauer Amanda M. , Hong Chibo , Do Kieu C. , Yingling Christin M. , Flores Kristina G. , Tessema Mathewos , Tellez Carmen S. , Willink Randall P. , Burki Elizabeth A. , Picchi Maria A. , Stidley Christine A. , Prados Michael D. , Costello Joseph F. , Gilliland Frank D. , Crowell Richard E. , Belinsky Steven A.
Published: February 2011

Purpose: To address the association between sequence variants within the MGMT (O6-methylguanine-DNA methyltransferase) promoter–enhancer region and methylation of MGMT in premalignant lesions from smokers and lung adenocarcinomas, their biological effects on gene regulation, and targeting MGMT for therapy.

Experimental Design: Single nucleotide polymorphisms (SNP) identified through sequencing a 1.9 kb fragment 5′ of MGMT were examined in relation to MGMT methylation in 169 lung adenocarcinomas and 1,731 sputum samples from smokers. The effect of promoter haplotypes on MGMT expression was tested using a luciferase reporter assay and cDNA expression analysis along with allele-specific sequencing for methylation. The response of MGMT methylated lung cancer cell lines to the alkylating agent temozolomide (TMZ) was assessed.

Results: The A allele of rs16906252 and the haplotype containing this SNP were strongly associated with increased risk for MGMT methylation in adenocarcinomas (ORs ≥ 94). This association was observed to a lesser extent in sputum samples in both smoker cohorts. The A allele was selectively methylated in primary lung tumors and cell lines heterozygous for rs16906252. With the most common haplotype as the reference, a 20 to 41% reduction in promoter activity was seen for the haplotype carrying the A allele that correlated with lower MGMT expression. The sensitivity of lung cancer cell lines to TMZ was strongly correlated with levels of MGMT methylation and expression.

Conclusions: These studies provide strong evidence that the A allele of a MGMT promoter–enhancer SNP is a key determinant for MGMT methylation in lung carcinogenesis. Moreover, TMZ treatment may benefit a subset of lung cancer patients methylated for MGMT. Clin Cancer Res; 17(7); 2014–23. ©2011 AACR.
Nelfinavir Induces Liposarcoma Apoptosis through Inhibition of Regulated Intramembrane Proteolysis of SREBP-1 and ATF6
Authors: Guan Min , Fousek Kristen , Jiang Chunling , Guo Song , Synold Tim , Xi Bixin , Shih Chu-Chih , Chow Warren A.
Published: February 2011

Purpose: We previously reported that nelfinavir (NFV) induces G1 cell-cycle block and apoptosis selectively in liposarcoma cell lines due to increased SREBP-1 (sterol regulatory element binding protein-1) expression in the absence of increased transcription. We postulate that NFV interferes with regulated intramembrane proteolysis of SREBP-1 and ATF6 (activating transcription factor 6).

Experimental Design: Time-lapse, confocal microscopic studies show that NFV inhibits the nuclear translocation of full-length SREBP-1–EGFP and ATF6–EGFP fusion proteins. siRNA-mediated knockdown of site-1 protease (S1P) and/or site-2 protease (S2P) leads to inhibition of SREBP-1 intracellular trafficking to the nucleus and reduces liposarcoma cell proliferation. Treatment of LiSa-2 liposarcoma cells with 3,4-dichloroisocoumarin, a serine protease inhibitor of S1P, did not affect SREBP-1 processing. In contrast, 1,10-phenanthroline, an S2P-specific inhibitor, reproduces the molecular and biological phenotypes observed in NFV-treated cells, which implicates S2P as a target of NFV. In vivo evaluation of NFV in a murine liposarcoma xenograft model leads to inhibition of tumor growth without significant toxicity.

Results: NFV-induced upregulation of SREBP-1 and ATF6 results from inhibition of S2P, which together with S1P mediates regulated intramembrane proteolysis from their precursor to their transcriptionally active forms. The resulting endoplasmic reticulum (ER) stress and concurrent inhibition of the unfolded protein response induce caspase-mediated apoptosis.

Conclusions: These results provide new insight into the mechanism of NFV-mediated induction of ER stress and cell death in liposarcomas and are the first to report targeting S2P for cancer therapy. Clin Cancer Res; 17(7); 1796–806. ©2011 AACR.
A YKL-40–Neutralizing Antibody Blocks Tumor Angiogenesis and Progression: A Potential Therapeutic Agent in Cancers
Authors: Faibish Michael , Francescone Ralph , Bentley Brooke , Yan Wei , Shao Rong
Published: February 2011

Accumulating evidence has indicated that expression levels of YKL-40, a secreted glycoprotein, were elevated in multiple advanced human cancers. Recently, we have identified an angiogenic role of YKL-40 in cancer development. However, blockade of the function of YKL-40, which implicates therapeutic value, has not been explored yet. Our current study sought to establish a monoclonal anti–YKL-40 antibody as a neutralizing antibody for the purpose of blocking tumor angiogenesis and metastasis. A mouse monoclonal anti–YKL-40 antibody (mAY) exhibited specific binding with recombinant YKL-40 and with YKL-40 secreted from osteoblastoma cells MG-63 and brain tumor cells U87. In the functional analysis, we found that mAY inhibited tube formation of microvascular endothelial cells in Matrigel induced by conditioned medium of MG-63 and U87 cells, as well as recombinant YKL-40. mAY also abolished YKL-40–induced activation of the membrane receptor VEGF receptor 2 (Flk-1/KDR) and intracellular signaling mitogen-activated protein (MAP) kinase extracellular signal–regulated kinase (Erk) 1 and Erk 2. In addition, mAY enhanced cell death response of U87 line to γ-irradiation through decreased expression of pAKT and AKT and accordingly, abrogated angiogenesis induced by the conditioned medium of U87 cells in which YKL-40 levels were elevated by treatment with γ-irradiation. Furthermore, treatment of xenografted tumor mice with mAY restrained tumor growth, angiogenesis, and progression. Taken together, this study has shown the therapeutic use for the mAY in treatment of tumor angiogenesis and metastasis. Mol Cancer Ther; 10(5); 742–51. ©2011 AACR.
Abstract 1230: Nicotine induces expression of integrin-linked kinase in human lung carcinoma cells through A7 nAChR-mediated inhibition of AP-2A
Authors: Zhong XiaoRong , Zheng Ying , Sun XiaoJuan , Ritzenthaler Jeffrey D. , Roman Jesse , Han Shouwei
Published: February 2011

We previously demonstrated that nicotine stimulates non small cell lung carcinoma (NSCLC) cell proliferation through nicotinic acetylcholine receptor (nAChR)-mediated induction of fibronectin expression. However, the mechanisms by which this agent affects human lung cancer occurrence and progression remain incompletely elucidated. Here, we show that nicotine increases the expression of integrin-linked kinase (ILK), a unique intracellular adaptor and kinase that has been implicated in the regulation of cancer cell growth/survival. This effect was blocked by the α7 nAChR antagonist, α-bungarotoxin, and by α7 nAChR siRNA. Silencing of ILK blocked the stimulatory effect of nicotine on phosphorylation of the PI3-K downstream target Akt and on cell growth. Furthermore, nicotine increased ILK gene promoter activity through inhibition of transcription factor AP-2α protein expression. Silencing of AP-2α enhanced, while exogenous expression of AP-2α attenuated, the stimulatory effect of nicotine on ILK expression. These findings appear to be relevant in vivo since we found that nicotine treatment increased tumor growth in animal models. Collectively, our results demonstrate that nicotine increases ILK gene expression through α7 nAChR-mediated inhibition of AP-2α followed by stimulation of PI3-K/Akt signaling. In turn, this leads to increased NSCLC cell growth. This study unveils a novel mechanism by which nicotine promotes human lung carcinoma cell growth/survival.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1230.
Abstract 1293: Neutrophil gelatinase-associated lipocalin regulates the expression of angiogenic cytokines and VEGF in human breast and ovarian cancer cells
Authors: Yang Jiang , Moses Marsha A.
Published: February 2011

Neutrophil gelatinase-associated lipocalin (NGAL; lipocalin 2) is a small extracellular protein (25 kDa) that is a member of the lipocalin protein family. It has been shown to bind ferric siderophores and to play an important role in iron metabolism. Increased NGAL levels have been reported in a variety of epithelial cancers, including breast, ovary, pancreas, colon, esophagus and thyroid cancer. We have recently reported that NGAL promotes breast cancer progression by inducing the epithelial to mesenchymal transition. In the course of these studies, we observed an increase in microvessel density in NGAL-expressing tumors. In the current study, we explored the potential role of NGAL in tumor angiogenesis by studying its effects on the expression of angiogenic factors. We first determined the levels of vascular endothelial growth factor (VEGF) in NGAL-expressing cells derived from the MCF-7 human breast cancer cell line. Using reverse-transcriptase PCR and ELISA, VEGF was found to be induced by NGAL at both the transcript and protein levels. Consistently, HIF-1α, one of the most important transcriptional regulators of VEGF, was observed to be accumulated in the nuclei of NGAL-expressing cells. Silencing of HIF-1α in NGAL-expressing cells significantly reduced VEGF expression. Consistent with this NGAL-induced VEGF production, there was a significant increase in endothelial cell migration towards the conditioned media collected from NGAL-expressing cells. Interestingly, treatment with a VEGF-neutralizing antibody only partially inhibited the migration-inducing activity of the conditioned media, suggesting that NGAL might induce the secretion of other angiogenic factors. To investigate this possibility, a panel of angiogenic factors was compared between the conditioned media collected from NGAL-expressing cells and control MCF-7 cells using antibody arrays. Several angiogenic cytokines, including IL-6, MCP-1 and GM-CSF, were found to be upregulated with NGAL expression. The effects of NGAL on VEGF and angiogenic cytokine production were also studied in SKOV-3 ovarian cancer cells. In this study, NGAL was silenced in SKOV-3 cells using specific siRNAs. The levels of both VEGF and IL-6 were found to be reduced after NGAL silencing, suggesting that NGAL's control of VEGF and cytokine production may be common among cancer cell types. These results suggest a novel mechanism through which NGAL may promote tumor angiogenesis and progression. (This work is supported by NIH R01 CA118764, P01 CA045548 and The Advanced Medical Foundation.)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1293.
Abstract 1608: Novel lipogenic enzyme ELOVL7 is involved in prostate cancer growth through saturated long-chain fatty acid metabolism
Authors: Tamura Kenji , Makino Asami , Hullin-Matsuda Francoise , Kobayashi Toshihide , Furihata Mutsuo , Chung Suyoun , Ashida Shingo , Miki Tsuneharu , Fujioka Tomoaki , Shuin Taro , Nakamura Yusuke , Nakagawa Hidewaki
Published: February 2011

A number of epidemiologic studies have indicated a strong association between dietary fat intake and prostate cancer development, suggesting that lipid metabolism plays some important roles in prostate carcinogenesis and its progression. In this study, through our genome-wide gene expression analysis of clinical prostate cancer cells, we identified a novel lipogenic gene, ELOVL7, coding a possible long-chain fatty acid elongase, as overexpressed in prostate cancer cells. ELOVL7 expression is regulated by the androgen pathway through SREBP1, as well as other lipogenic enzymes. Knockdown of ELOVL7 resulted in drastic attenuation of prostate cancer cell growth, and it is notable that high-fat diet promoted the growth of in vivo tumors of ELOVL7-expressed prostate cancer. In vitro fatty acid elongation assay and fatty acid composition analysis indicated that ELOVL7 was preferentially involved in fatty acid elongation of saturated very-long-chain fatty acids (SVLFA, C20:0). Lipid profiles showed that knockdown of ELOVL7 in prostate cancer cells affected SVLFAs in the phospholipids and the neutral lipids, such as cholesterol ester. Focusing on cholesterol ester as a source of de novo steroid synthesis, we show that ELOVL7 affected de novo androgen synthesis in prostate cancer cells. These findings suggest that EVOLV7 could be involved in prostate cancer growth and survival through the metabolism of SVLFAs and their derivatives, could be a key molecule to elucidate the association between fat dietary intake and prostate carcinogenesis, and could also be a promising molecular target for development of new therapeutic or preventive strategies for prostate cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1608.
Abstract 17: Preclinical and phase 1 trial results of JI-101, a novel, oral tyrosine kinase inhibitor that selectively targets VEGFR2, EphB4, and PDGFRβ
Authors: Velleca Mark , Miller Linda , Sharma Sunil , Gordon Michael , Brittelli David , Elkin Lisa , Govindarajan Raman , Klein Udo , Mitchell Scott , N Nandhagopal , Qian Dapeng , Qian Xiaobing , Stephen Thomas , Srinivasa Sreesha , Stern Warren
Published: February 2011

Introduction

JI-101 is a highly selective and potent angiogenesis inhibitor with unique EphB4 activity that distinguishes it from other agents in clinical development. We report the preclinical pharmacology and early clinical experience with this compound.

Procedures

Lead optimization was used to identify cgi1842 (now JI-101) with a targeted activity profile that is selective and unique. JI-101 was tested in binding, enzymatic, and cell-based assays for activity against kinase and non-kinase targets. Preclinical in vivo testing for PK, PD, efficacy, and safety pharmacology was performed in mouse, rat, guinea pig, and dog. A Phase I clinical trial is underway: 12 patients have been treated thus far with 28-day oral daily dosing at 100 mg, 200 mg, 400 mg QD or at 200 mg BID. PK, PD, and imaging analyses have been performed.

Data

JI-101 has a MW of 466D and high (<100 nm) potency against VEGFR2, EphB4, and PDGFR in enzymatic and cell-based assays. Broad kinase cross-screening (Ambit KinomeScan, 445 kinases) reveals that JI-101 is highly selective for angiogenic kinases with only 23 kinases demonstrating a Kd 15). JI-101 was co-administered with paclitaxel in a MDA-MB-231 mouse xenotransplant model, resulting in greater efficacy and no increased toxicity. Based on 28-day GLP toxicology studies conducted in rat and dog, a 100 mg qd starting dose (2 × 50 mg capsules) was chosen for Phase I studies. JI-101 appears to be very well-tolerated to date with no dose limiting toxicities. Patients have received 1-9 cycles with most receiving multiple cycles and 5 remain on study at 100 mg (1 pt), 200 mg (1 pt), and 400 mg (3 pt). Manageable hypertension has been seen in all cohorts indicating a possible PD effect. Hand-foot syndrome of short duration was seen at the 400mg dose level. Further clinical details of the PK, PD, tolerability and efficacy of JI-101 will be presented.

Conclusions

JI-101 potently inhibits 3 critical angiogenic kinases (VEGFR2, EphB4, and PDGFR) and is the only such “triple” angiogenesis inhibitor currently in development. JI-101 shows robust preclinical and clinical pharmacology and is well-tolerated to date in human trials. In tumors where EphB4 may play an oncogenic role, such as head and neck cancer, JI-101 may have anti-proliferative effects in addition to anti-angiogenic effects and should be amenable to combination therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 17.
Abstract 2241: Role of tissue markers in the prediction of bone metastases from breast cancer
Authors: Sacanna Emanuele , Ibrahim Toni , Gaudio Michele , Mercatali Laura , Ricci Rossana , Scarpi Emanuela , Ricci Marianna , Serra Patrizia , Serra Luigi , Arienti Chiara , Bravaccini Sara , Amadori Dino
Published: February 2011

Introduction: Bone metastasis is a frequent event in patients with breast cancer. OPG (osteoprotegerin) protects bone from excessive resorption by binding to RANK-L (receptor activator of nuclear factor-kB-ligand) and by preventing it from binding to RANK. These molecules are closely involved in the process of metastasization to bone. CXCL12/SDF-1α, the ligand of CXCR4, is overexpressed in bone, and the local chemokine milieu is now emerging as a key determinant in organ selectivity by tumor cells. We carried out a retrospective analysis to evaluate the potential role of these biological markers in predicting bone metastasization in breast cancer patients.

Materials and methods: OPG and CXCR4 expression was determined by avidin-biotin immunohistochemistry using a polyclonal (H-249, Santa Cruz) and monoclonal (Ab58176, Abcam) antibody, respectively. Immunohistochemical staining was carried out on sections from paraffin-embedded blocks of 40 primary breast cancers. Ten patients (median age of 64 years, range 48-78) were disease-free (DF) at a median of 9.8 (range 6.9-11.5) years, while 30 (median age 67 years, range 42-87) had relapsed. In the latter group, 10 (median age 66 years, range 42-87) had visceral metastases (VM) and 20 (median age 69 years, range 42-87) had bone metastases (BM).

Results: Considering only strong cytoplasmic expression as positive, 22.5% of tumors were positive for OPG and 25% for CXCR4. In particular, OPG was expressed in 20% of tumors in DF patients and in 25% in BM patients. Conversely, CXCR4 was expressed in 10% of tumors in DF patients and in 45% in BM patients. OPG was expressed in only 20% of tumors in VM patients. There was no correlation between OPG and CXCR4 expression, and positivity to at least one of the two markers was observed in 30% of tumors in DF patients, 20% in VM patients and 55% in BM patients. In this last subgroup, there were no statistically significant associations between marker expression and tumor characteristics. Furthermore, no relation was observed between marker expression and disease-free or overall survival in BM patients.

Conclusions: Our preliminary results suggest that cytoplasmic CXCR4 expression in primary breast cancers could play a role in predicting bone metastases. Enrolment of breast cancer patients in a larger study is ongoing to confirm these data and to evaluate the predictive role of RANK and RANK-L in bone metastasization.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2241.
Abstract 2281: Host CXCR2 expression regulates mammary tumor growth, angiogenesis and progression
Authors: Nannuru Kalyan C. , Varney Michelle L. , Singh Rakesh K.
Published: February 2011

Breast cancer is a devastating clinical condition affecting more than 100,000 women in a year and is responsible for more than 40,000 deaths every year. Chemokines and their receptors have been shown to regulate various aspects of tumor development and progression. Constitutive expression of pro-inflammatory chemokines, a hallmark of many human cancers, helps establish a supportive tumor stroma, and, in some cases, directly stimulates tumor proliferation and invasion via receptors on tumor cells. Evidence suggests that CXCR2 regulate multiple functions in leukocytes, endothelial cells (ECs), and/or tumor cells and their precursors and thus play important role in tumor angiogenesis, growth and metastasis. In this study, we investigated the role of host CXCR2 expression in mammary tumor growth, neovascularization and tumor induced osteolysis using a syngenic mouse mammary tumor model. Eight week old BALB/c mice either expressing CXCR2 (mCXCR2 WT) or genomic knock down of CXCR2 (mCXCR2 −/−) were used to study the functional role of host CXCR2 in mammary tumor growth and progression. Orthotopical implantation of mammary tumors in mouse mammary fat pad reveals a significant decrease in tumor growth in mCXCR2 −/− compared to wild type mice. Knock-down of host CXCR2 has significantly reduced tumor-induced neovascularization in orthotopical tumors. Bone is the most common site of metastasis in breast cancer. To access the role of host CXCR2 in tumor-induced osteolysis and tumor growth in bone microenvironment, mammary tumor cells were implanted on calvaria. Tumor-induced osteolysis and osteoclast homing at the tumor-bone interface was significantly reduced in CXCR2 knockout mice compared to wild type mice. These results suggest that host CXCR2 may play a critical role in tumor development and angiogenesis as well as tumor-induced osteolysis. These data demonstrate direct evidence of host CXCR2 in breast cancer progression and metastasis, and will allow development of CXCR2 targeting as a novel approach to breast cancer treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2281.
Abstract 229: Omega-3 fatty acids strongly induce apoptosis in human breast cancer cells in vitro and in vivo via formation of reactive oxygen species and caspase 8 activation
Authors: Wang Pan , Kang Ki-Sung , Yamabe Noriko , Fukui Masayuki , Jay Taylor , Zhu B T.
Published: February 2011

[Hypothesis] The present study sought to investigate the in vitro and in vivo anticancer effect of two representative omega-3 fatty acids, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), with a focus on assessing the role of fatty acid-induced oxidative stress and apoptosis as mechanisms for their anticancer actions.

[Methods] We used in vitro cell culture of MCF-7 human breast cancer cells and in vivo athymic nude mice model to study the effect of omega-3 fatty acids on cancer cell proliferation and the mechanism of the cell death.

[Results] In vitro studies showed that DHA and EPA each strongly reduced the viability and DNA synthesis of MCF-7 human breast cancer cells in culture, and they also promoted cell death via apoptosis. Mechanistically, accumulation of reactive oxygen species and activation of caspase 8 contributed critically to the induction of apoptotic cell death. Co-presence of antioxidants or a selective inhibition or knockdown of caspase 8 each effectively abrogated the cytotoxic effect of DHA. Using athymic nude mice as an in vivo model, we found that feeding animals with 5% fish oil-containing diet for 6 weeks significantly reduced the growth of MCF-7 human breast cancer cells through inhibition of cancer cell proliferation as well as promotion of cell death. Using 3-nitrotyrosine as a parameter, we confirmed that fish oil diet significantly increased oxidative stress in tumor cells in vivo. Analysis of fatty acid content in plasma and tissues showed that feeding animals a 5% fish oil diet significantly increased DHA and EPA levels in both normal and tumorous mammary tissues by 329 and 300%, respectively.

[Conclusion] The results of this investigation clearly showed that omega-3 fatty acids strongly inhibited human breast cancer proliferation both in vitro and in vivo via formation of reactive oxygen species and induction of apoptosis by caspase 8 activation. This study calls for further studies in human subjects to assess the effectiveness of fish oil as a dietary supplement in the treatment of certain types of cancers.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 229.