bluuebunny/crossref_metadata_2025 huggingface dataset:
Liquid in the gastroesophageal segment promotes reflux, but compliance does not: a mathematical modeling study
Authors: Ghosh Sudip K. , Kahrilas Peter J. , Brasseur James G.
Published: August 2008

The mechanical force relationships that distinguish normal from chronic reflux at sphincter opening are poorly understood and difficult to measure in vivo. Our aim was to apply physics-based computer simulations to determine mechanical pathogenesis of gastroesophageal reflux. A mathematical model of the gastroesophageal segment (GES) was developed, incorporating the primary anatomical and physiomechanical elements that drive GES opening and reflux. In vivo data were used to quantify muscle stiffness, sphincter tone, and gastric pressure. The liquid lining the mucosa was modeled as an “effective liquid film” between the mucosa and a manometric catheter. Newton's second law was solved mathematically, and the space-time details of opening and reflux were predicted for systematic variations in gastric pressure increase, film thickness, muscle stiffness, and tone. “Reflux” was defined as “2 ml of refluxate entering the esophagus within 1 s.” GES opening and reflux were different events. Both were sensitive to changes in gastric pressure and sphincter tone. Reflux initiation was extremely sensitive to the liquid film thickness; the protective function of the sphincter was destroyed with only 0.4 mm of liquid in the GES. Compliance had no effect on reflux initiation, but affected reflux volume. The presence of abnormal levels of liquid within the collapsed GES can greatly increase the probability for reflux, suggesting a mechanical mechanism that may differentiate normal reflux from gastroesophageal reflux disease. Compliance does not affect the probability for reflux, but affects reflux volume once it occurs. Opening without reflux suggests the existence of “gastroesophageal pooling” in the distal esophagus, with clinical implications.
Role of endothelial nitric oxide synthase-derived nitric oxide in activation and dysfunction of cerebrovascular endothelial cells during early onsets of sepsis
Authors: Handa Osamu , Stephen Jancy , Cepinskas Gediminas
Published: August 2008

Sepsis-associated encephalopathy is an early manifestation of sepsis, resulting in a diffuse dysfunction of the brain. Recently, nitric oxide (NO) has been proposed to be one of the key molecules involved in the modulation of inflammatory responses in the brain. The aim of this study was to assess the role of NO in cerebrovascular endothelial cell activation/dysfunction during the early onsets of sepsis. To this end, we employed an in vitro model of sepsis in which cultured mouse cerebrovascular endothelial cells (MCVEC) were challenged with blood plasma (20% vol/vol) obtained from sham or septic (feces-induced peritonitis, FIP; 6 h) mice. Exposing MCVEC to FIP plasma for 1 h resulted in increased production of reactive oxygen species and NO as assessed by intracellular oxidation of oxidant-sensitive fluorochrome, dihydrorhodamine 123 (DHR 123), and nitrosation of NO-specific probe, DAF-FM, respectively. The latter events were accompanied by dissociation of tight junction protein, occludin, from MCVEC cytoskeletal framework and a subsequent increase in FITC-dextran (3-kDa mol mass) flux across MCVEC grown on the permeable cell culture supports, whereas Evans blue-BSA (65-kDa mol mass) or FITC-dextran (10-kDa mol mass) flux were not affected. FIP plasma-induced oxidant stress, occludin rearrangement, and MCVEC permeability were effectively attenuated by antioxidant, 1-pyrrolidinecarbodithioic acid (PDTC; 0.5 mM), or interfering with nitric oxide synthase (NOS) activity [0.1 mM nitro-l-arginine methyl ester (l-NAME) or endothelial NOS (eNOS)-deficient MCVEC]. However, treatment of MCVEC with PDTC failed to interfere with NO production, suggesting that septic plasma-induced oxidant stress in MCVEC is primarily a NO-dependent event. Taken together, these data indicate that during early sepsis, eNOS-derived NO exhibits proinflammatory characteristics and contributes to the activation and dysfunction of cerebrovascular endothelial cells.
Formation and function of ceramide-enriched membrane platforms with CD38 during M1-receptor stimulation in bovine coronary arterial myocytes
Authors: Jia Su-Jie , Jin Si , Zhang Fan , Yi Fan , Dewey William L. , Li Pin-Lan
Published: August 2008

CD38 contains an ADP ribosylcyclase domain that mediates intracellular Ca2+ signaling by the production of cyclic ADP-ribose (cADPR), but the mechanisms by which the agonists activate this enzyme remain unclear. The present study tested a hypothesis that a special lipid-raft (LR) form, ceramide-enriched lipid platform, contributes to CD38 activation to produce cADPR in response to muscarinic type 1 (M1) receptor stimulation in bovine coronary arterial myocytes (CAMs). By confocal microscopic analysis, oxotremorine (Oxo), an M1 receptor agonist, was found to increase LR clustering on the membrane with the formation of a complex of CD38 and LR components such as GM1, acid sphingomyelinase (ASMase), and ceramide, a typical ceramide-enriched macrodomain. At 80 μM, Oxo increased LR clustering by 78.8%, which was abolished by LR disruptors, methyl-β-cyclodextrin (MCD), or filipin. With the use of a fluorescence resonance energy transfer (FRET) technique, 15.5 ± 1.9% energy transfer rate (vs. 5.3 ± 0.9% of control) between CD38 and LR component, ganglioside M1 was detected, further confirming the proximity of both molecules. In the presence of MCD or filipin, there were no FRET signals detected. In floated detergent-resistant membrane fractions, CD38 significantly increased in LR fractions of CAMs treated by Oxo. Moreover, MCD or filipin attenuated Oxo-induced production of cADPR via CD38. Functionally, Oxo-induced intracellular Ca2+ release and coronary artery constriction via cADPR were also blocked by LR disruption or ASMase inhibition. These results provide the first evidence that the formation of ceramide-enriched lipid macrodomains is crucial for Oxo-induced activation of CD38 to produce cADPR in CAMs, and these lipid macrodomains mediate transmembrane signaling of M1 receptor activation to produce second messenger cADPR.
Improved tumor targeting of anti–epidermal growth factor receptor Nanobodies through albumin binding: taking advantage of modular Nanobody technology
Authors: Tijink Bernard M. , Laeremans Toon , Budde Marianne , Walsum Marijke Stigter-van , Dreier Torsten , de Haard Hans J. , Leemans C. René , van Dongen Guus A.M.S.
Published: August 2008

The ∼15-kDa variable domains of camelid heavy-chain-only antibodies (called Nanobodies) can easily be formatted as multivalent or multispecific single-chain proteins. Because of fast excretion, however, they are less suitable for therapy of cancer. In this study, we aimed for improved tumor targeting of a bivalent anti–epidermal growth factor receptor (EGFR) Nanobody (αEGFR-αEGFR) by fusion to a Nanobody unit binding to albumin (αAlb). Biodistributions of αEGFR-αEGFR, αEGFR-αEGFR-αAlb (∼50 kDa), αTNF-αTNF-αAlb (control, binding tumor necrosis factor-α), and the ∼150-kDa anti-EGFR antibody cetuximab were compared in A431 xenograft-bearing mice. The proteins were radiolabeled with 177Lu to facilitate quantification. Tumor uptake of 177Lu-αEGFR-αEGFR decreased from 5.0 ± 1.4 to 1.1 ± 0.1 %ID/g between 6 and 72 h after injection. Due to its rapid blood clearance, tumor-to-blood ratios >80 were obtained within 6 h after injection. Blood clearance became dramatically slower and tumor uptake became significantly higher by introduction of αAlb. Blood levels of αEGFR-αEGFR-αAlb were 21.2 ± 2.5, 11.9 ± 0.6, and 4.0 ± 1.4 and tumor levels were 19.4 ± 5.5, 35.2 ± 7.5, and 28.0 ± 6.8 %ID/g at 6, 24, and 72 h after injection, respectively. Tumor uptake was at least as high as for cetuximab (15.5 ± 3.9, 27.1 ± 7.9, and 25.6 ± 6.1 %ID/g) and significantly higher than for αTNF-αTNF-αAlb. αEGFR-αEGFR-αAlb showed faster and deeper tumor penetration than cetuximab. These data show that simple fusion of αEGFR and αAlb building blocks results in a bifunctional Nanobody format, which seems more favorable for therapy as far as pharmacokinetics and tumor deposition are concerned. [Mol Cancer Ther 2008;7(8):2288–97]
Breaching the DNA damage checkpoint via PF-00477736, a novel small-molecule inhibitor of checkpoint kinase 1
Authors: Blasina Alessandra , Hallin Jill , Chen Enhong , Arango Maria Elena , Kraynov Eugenia , Register James , Grant Stephan , Ninkovic Sacha , Chen Ping , Nichols Tim , O'Connor Patrick , Anderes Kenna
Published: August 2008

Checkpoints are present in all phases of the cell cycle and are regarded as the gatekeepers maintaining the integrity of the genome. Many conventional agents used to treat cancer impart damage to the genome and activate cell cycle checkpoints. Many tumors are defective in the tumor suppressor p53 and therefore lack a functional G1 checkpoint. In these tumors, however, the S-G2 checkpoints remain intact and, in response to DNA damage, arrest cell cycle progression allowing time for DNA repair. Checkpoint kinase 1 (Chk1) is a key element in the DNA damage response pathway and plays a crucial role in the S-G2-phase checkpoints. Inhibiting Chk1 represents a therapeutic strategy for creating a “synthetic lethal” response by overriding the last checkpoint defense of tumor cells against the lethal damage induced by DNA-directed chemotherapeutic agents. Chk1 inhibition is consistent with emerging targeted therapies aiming to exploit molecular differences between normal and cancer cells. Adding a Chk1 inhibitor to DNA-damaging cytotoxic therapy selectively targets tumors with intrinsic checkpoint defects while minimizing toxicity in checkpoint-competent normal cells. PF-00477736 was identified as a potent, selective ATP-competitive small-molecule inhibitor that inhibits Chk1 with a Ki of 0.49 nM. PF-00477736 abrogates cell cycle arrest induced by DNA damage and enhances cytotoxicity of clinically important chemotherapeutic agents, including gemcitabine and carboplatin. In xenografts, PF-00477736 enhanced the antitumor activity of gemcitabine in a dose-dependent manner. PF-00477736 combinations were well tolerated with no exacerbation of side effects commonly associated with cytotoxic agents. [Mol Cancer Ther 2008;7(8):2394–404]
Jadomycin B, an Aurora-B kinase inhibitor discovered through virtual screening
Authors: Fu Da-Hua , Jiang Wei , Zheng Jian-Ting , Zhao Gui-Yu , Li Yan , Yi Hong , Li Zhuo-Rong , Jiang Jian-Dong , Yang Ke-Qian , Wang Yanchang , Si Shu-Yi
Published: August 2008

Aurora kinases have emerged as promising targets for cancer therapy because of their critical role in mitosis. These kinases are well-conserved in all eukaryotes, and IPL1 gene encodes the single Aurora kinase in budding yeast. In a virtual screening attempt, 22 compounds were identified from nearly 15,000 microbial natural products as potential small-molecular inhibitors of human Aurora-B kinase. One compound, Jadomycin B, inhibits the growth of ipl1-321 temperature-sensitive mutant more dramatically than wild-type yeast cells, raising the possibility that this compound is an Aurora kinase inhibitor. Further in vitro biochemical assay using purified recombinant human Aurora-B kinase shows that Jadomycin B inhibits Aurora-B activity in a dose-dependent manner. Our results also indicate that Jadomycin B competes with ATP for the kinase domain, which is consistent with our docking prediction. Like other Aurora kinase inhibitors, Jadomycin B blocks the phosphorylation of histone H3 on Ser10 in vivo. We also present evidence suggesting that Jadomycin B induces apoptosis in tumor cells without obvious effects on cell cycle. All the results indicate that Jadomycin B is a new Aurora-B kinase inhibitor worthy of further investigation. [Mol Cancer Ther 2008;7(7):2386–93]
Human endothelial precursor cells express tumor endothelial marker 1/endosialin/CD248
Authors: Bagley Rebecca G. , Rouleau Cecile , St. Martin Thia , Boutin Paula , Weber William , Ruzek Melanie , Honma Nakayuki , Nacht Mariana , Shankara Srinivas , Kataoka Shiro , Ishida Isao , Roberts Bruce L. , Teicher Beverly A.
Published: August 2008

Angiogenesis occurs during normal physiologic processes as well as under pathologic conditions such as tumor growth. Serial analysis of gene expression profiling revealed genes [tumor endothelial markers (TEM)] that are overexpressed in tumor endothelial cells compared with normal adult endothelial cells. Because blood vessel development of malignant tumors under certain conditions may include endothelial precursor cells (EPC) recruited from bone marrow, we investigated TEM expression in EPC. The expression of TEM1 or endosialin (CD248) and other TEM has been discovered in a population of vascular endothelial growth factor receptor 2+/CD31+/CD45−/VE-cadherin+ EPC derived from human CD133+/CD34+ cells. EPC share some properties with fully differentiated endothelial cells from normal tissue, yet reverse transcription-PCR and flow cytometry reveal that EPC express higher levels of endosialin at the molecular and protein levels. The elevated expression of endosialin in EPC versus mature endothelial cells suggests that endosialin is involved in the earlier stages of tumor angiogenesis. Anti-endosialin antibodies inhibited EPC migration and tube formation in vitro. In vivo, immunohistochemistry indicated that human EPC continued to express endosialin protein in a Matrigel plug angiogenesis assay established in nude mice. Anti-endosialin antibodies delivered systemically at 25 mg/kg were also able to inhibit circulating murine EPC in nude mice bearing s.c. SKNAS tumors. EPC and bone marrow–derived cells have been shown previously to incorporate into malignant blood vessels in some instances, yet they remain controversial in the field. The data presented here on endothelial genes that are up-regulated in tumor vasculature and in EPC support the hypothesis that the angiogenesis process in cancer can involve EPC. [Mol Cancer Ther 2008;7(8):2536–46]
New insights into the pharmacology and cytotoxicity of gemcitabine and 2′,2′-difluorodeoxyuridine
Authors: Veltkamp Stephan A. , Pluim Dick , van Eijndhoven Maria A.J. , Bolijn Maria J. , Ong Felix H.G. , Govindarajan Rajgopal , Unadkat Jashvant D. , Beijnen Jos H. , Schellens Jan H.M.
Published: August 2008

In a clinical study with oral gemcitabine (2′,2′-difluorodeoxycytidine, dFdC), 2′,2′-difluorodeoxyuridine (dFdU) was extensively formed and accumulated after multiple oral dosing. Here, we have investigated the in vitro cytotoxicity, cellular uptake, efflux, biotransformation, and nucleic acid incorporation of dFdC and dFdU. Short-term and long-term cytotoxicity assays were used to assess the cytotoxicity of dFdC and dFdU in human hepatocellular carcinoma HepG2, human lung carcinoma A549, and Madin-Darby canine kidney cell lines transfected with the human concentrative or equilibrative nucleoside transporter 1 (hCNT1 or hENT1), or empty vector. Radiolabeled dFdC and dFdU were used to determine cellular uptake, efflux, biotransformation, and incorporation into DNA and RNA. The compounds dFdC, dFdU, and their phosphorylated metabolites were quantified by high-performance liquid chromatography with UV and radioisotope detection. dFdU monophosphate, diphosphate, and triphosphate (dFdU-TP) were formed from dFdC and dFdU. dFdU-TP was incorporated into DNA and RNA. The area under the intracellular concentration-time curve of dFdC-TP and dFdU-TP and their extent of incorporation into DNA and RNA inversely correlated with the IC50 of dFdC and dFdU, respectively. The cellular uptake and cytotoxicity of dFdU were significantly enhanced by hCNT1. dFdU inhibited cell cycle progression and its cytotoxicity significantly increased with longer duration of exposure. dFdU is taken up into cells with high affinity by hCNT1 and phosphorylated to its dFdU-TP metabolite. dFdU-TP is incorporated into DNA and RNA, which correlated with dFdU cytotoxicity. These data provide strong evidence that dFdU can significantly contribute to the cytotoxicity of dFdC. [Mol Cancer Ther 2008;7(8):2415–25]
Anticancer steroid sulfatase inhibitors: synthesis of a potent fluorinated second-generation agent, in vitro and in vivo activities, molecular modeling, and protein crystallography
Authors: Woo L.W. Lawrence , Fischer Delphine S. , Sharland Christopher M. , Trusselle Melanie , Foster Paul A. , Chander Surinder K. , Di Fiore Anna , Supuran Claudiu T. , De Simone Giuseppina , Purohit Atul , Reed Michael J. , Potter Barry V.L.
Published: August 2008

An improved steroid sulfatase inhibitor was prepared by replacing the N-propyl group of the second-generation steroid-like inhibitor (2) with a N-3,3,3-trifluoropropyl group to give (10). This compound is 5-fold more potent in vitro, completely inhibits rat liver steroid sulfatase activity after a single oral dose of 0.5 mg/kg, and exhibits a significantly longer duration of inhibition over (2). These biological properties are attributed to the increased lipophilicity and metabolic stability of (10) rendered by its trifluoropropyl group and also the potential H-bonding between its fluorine atom(s) and Arg98 in the active site of human steroid sulfatase. Like other sulfamates, (10) is expected to be sequestered, and transported by, erythrocytes in vivo because it inhibits human carbonic anhydrase II (hCAII) potently (IC50, 3 nmol/L). A congener (4), which possesses a N-(pyridin-3-ylmethyl) substituent, is even more active (IC50, 0.1 nmol/L). To rationalize this, the hCAII-(4) adduct, obtained by cocrystallization, reveals not only the sulfamate group and the backbone of (4) interacting with the catalytic site and the associated hydrophobic pocket, respectively, but also the potential H-bonding between the N-(pyridin-3-ylmethyl) group and Nε2 of Gln136. Like (2), both (10) and its phenolic precursor (9) are non-estrogenic using a uterine weight gain assay. In summary, a highly potent, long-acting, and nonestrogenic steroid sulfatase inhibitor was designed with hCAII inhibitory properties that should positively influence in vivo behavior. Compound (10) and other related inhibitors of this structural class further expand the armory of steroid sulfatase inhibitors against hormone-dependent breast cancer. [Mol Cancer Ther 2008;7(8):2435–44]
Optimization of antibody binding to FcγRIIa enhances macrophage phagocytosis of tumor cells
Authors: Richards John O. , Karki Sher , Lazar Greg A. , Chen Hsing , Dang Wei , Desjarlais John R.
Published: August 2008

The contribution of Fc-mediated effector functions to the therapeutic efficacy of some monoclonal antibodies has motivated efforts to enhance interactions with Fcγ receptors (FcγR). Although an early goal has been enhanced FcγRIIIa binding and natural killer (NK) cell antibody-dependent cell-mediated cytotoxicity (ADCC), other relevant cell types such as macrophages are dependent on additional activating receptors such as FcγRIIa. Here, we describe a set of engineered Fc variants with diverse FcγR affinities, including a novel substitution G236A that provides selectively enhanced binding to FcγRIIa relative to FcγRIIb. Variants containing this substitution have up to 70-fold greater FcγRIIa affinity and 15-fold improvement in FcγRIIa/FcγRIIb ratio and mediate enhanced phagocytosis of antibody-coated target cells by macrophages. Specific double and triple combination variants with this substitution are simultaneously capable of exhibiting high NK-mediated ADCC and high macrophage phagocytosis. In addition, we have used this unique set of variants to quantitatively probe the relative contributions of individual FcγR to effector functions mediated by NK cells and macrophages. These experiments show that FcγRIIa plays the most influential role for macrophages and, surprisingly, that the inhibitory receptor FcγRIIb has little effect on effector function. The enhancements in phagocytosis described here provide the potential to improve the performance of therapeutic antibodies targeting cancers. [Mol Cancer Ther 2008;7(8):2517–27]
IRC-083927 is a new tubulin binder that inhibits growth of human tumor cells resistant to standard tubulin-binding agents
Authors: Liberatore Anne-Marie , Coulomb Hélène , Pons Dominique , Dutruel Olivier , Kasprzyk Philip G. , Carlson Mark , Nelson Ann Savola , Newman Simon P. , Stengel Chloe , Auvray Pierrïck , Hesry Vincent , Foll Béatrice , Narboux Nadine , Morlais Delphine , Le Moing Mélissa , Bernetiere Sonia , Dellile Raphael , Camara Jose , Ferrandis Eric , Bigg Dennis C. , Prévost Grégoire P.
Published: August 2008

Tubulin is a validated target for antitumor drugs. However, the effectiveness of these microtubule-interacting agents is limited by the fact that they are substrates for drug efflux pumps (P-glycoprotein) and/or by the acquisition of point mutations in tubulin residues important for drug-tubulin binding. To bypass these resistance systems, we have identified and characterized a novel synthetic imidazole derivative IRC-083927, which inhibits the tubulin polymerization by a binding to the colchicine site. IRC-083927 inhibits in vitro cell growth of human cancer cell lines in the low nanomolar range. More interesting, it remains highly active against cell lines resistant to microtubule-interacting agents (taxanes, Vinca alkaloids, or epothilones). Such resistances are due to the presence of efflux pumps (NCI-H69/LX4 resistant to navelbine and paclitaxel) and/or the presence of mutations on β-tubulin and on α-tubulin and β-tubulin (A549.EpoB40/A549.EpoB480 resistant to epothilone B or paclitaxel). IRC-083927 displayed cell cycle arrest in G2-M phase in tumor cells, including in the drug-resistant cells. In addition, IRC-083927 inhibited endothelial cell proliferation in vitro and vessel formation in the low nanomolar range supporting an antiangiogenic behavior. Finally, chronic oral treatment with IRC-083927 (5 mg/kg) inhibits the growth of two human tumor xenografts in nude mice (C33-A, human cervical cancer and MDA-MB-231, human hormone-independent breast cancer). Together, the antitumor effects induced by IRC-083927 on tumor models resistant to tubulin agents support further investigations to fully evaluate its potential for the treatment of advanced cancers, particularly those resistant to current clinically available drugs. [Mol Cancer Ther 2008;7(8):2426–34]
Androgen receptor inactivation contributes to antitumor efficacy of 17α-hydroxylase/17,20-lyase inhibitor 3β-hydroxy-17-(1H-benzimidazole-1-yl)androsta-5,16-diene in prostate cancer
Authors: Vasaitis Tadas , Belosay Aashvini , Schayowitz Adam , Khandelwal Aakanksha , Chopra Pankaj , Gediya Lalji K. , Guo Zhiyong , Fang Hong-Bin , Njar Vincent C.O. , Brodie Angela M.H.
Published: August 2008

We previously reported that our novel compound 3β-hydroxy-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (VN/124-1) is a potent 17α-hydroxylase/17,20-lyase (CYP17) inhibitor/antiandrogen and strongly inhibits the formation and proliferation of human prostate cancer LAPC4 tumor xenografts in severe combined immunodeficient mice. In this study, we report that VN/124-1 and other novel CYP17 inhibitors also cause down-regulation of androgen receptor (AR) protein expression in vitro and in vivo. This mechanism of action seems to contribute to their antitumor efficacy. We compared the in vivo antitumor efficacy of VN/124-1 with that of castration and a clinically used antiandrogen, Casodex, and show that VN/124-1 is more potent than castration in the LAPC4 xenograft model. Treatment with VN/124-1 (0.13 mmol/kg twice daily) was also very effective in preventing the formation of LAPC4 tumors (6.94 versus 2410.28 mm3 in control group). VN/124-1 (0.13 mmol/kg twice daily) and VN/124-1 (0.13 mmol/kg twice daily) + castration induced regression of LAPC4 tumor xenografts by 26.55% and 60.67%, respectively. Treatments with Casodex (0.13 mmol/kg twice daily) or castration caused significant tumor suppression compared with control. Furthermore, treatment with VN/124-1 caused marked down-regulation of AR protein expression, in contrast to treatments with Casodex or castration that caused significant up-regulation of AR protein expression. The results suggest that VN/124-1 acts by several mechanisms (CYP17 inhibition, competitive inhibition, and down-regulation of the AR). These actions contribute to inhibition of the formation of LAPC4 tumors and cause regression of growth of established tumors. VN/124-1 is more efficacious than castration in the LAPC4 xenograft model, suggesting that the compound has potential for the treatment of prostate cancer. [Mol Cancer Ther 2008;7(8):2348–57]
Elesclomol induces cancer cell apoptosis through oxidative stress
Authors: Kirshner Jessica R. , He Suqin , Balasubramanyam Vishwasenani , Kepros Jane , Yang Chin-Yu , Zhang Mei , Du Zhenjian , Barsoum James , Bertin John
Published: August 2008

Elesclomol (formerly STA-4783) is a novel small molecule undergoing clinical evaluation in a pivotal phase III melanoma trial (SYMMETRY). In a phase II randomized, double-blinded, controlled, multi-center trial in 81 patients with stage IV metastatic melanoma, treatment with elesclomol plus paclitaxel showed a statistically significant doubling of progression-free survival time compared with treatment with paclitaxel alone. Although elesclomol displays significant therapeutic activity in the clinic, the mechanism underlying its anticancer activity has not been defined previously. Here, we show that elesclomol induces apoptosis in cancer cells through the induction of oxidative stress. Treatment of cancer cells in vitro with elesclomol resulted in the rapid generation of reactive oxygen species (ROS) and the induction of a transcriptional gene profile characteristic of an oxidative stress response. Inhibition of oxidative stress by the antioxidant N-acetylcysteine blocked the induction of gene transcription by elesclomol. In addition, N-acetylcysteine blocked drug-induced apoptosis, indicating that ROS generation is the primary mechanism responsible for the proapoptotic activity of elesclomol. Excessive ROS production and elevated levels of oxidative stress are critical biochemical alterations that contribute to cancer cell growth. Thus, the induction of oxidative stress by elesclomol exploits this unique characteristic of cancer cells by increasing ROS levels beyond a threshold that triggers cell death. [Mol Cancer Ther 2008;7(8):2319–27]
Mitotic catastrophe cell death induced by heat shock protein 90 inhibitor in BRCA1-deficient breast cancer cell lines
Authors: Zajac Magdalena , Moneo Maria Victoria , Carnero Amancio , Benitez Javier , Martínez-Delgado Beatriz
Published: August 2008

Heat shock protein 90 (Hsp90) is a molecular chaperone involved in folding, assembly, maturation, and stabilization of the client proteins that regulate survival of malignant cells. As previous reports correlate high Hsp90 expression with decreased survival in breast cancer, Hsp90 may be a favorable target for investigational therapy in breast cancer. In our study, we have examined the response of a panel of both BRCA1-null (UACC 3199, HCC 1937, and MBA-MD-436) and BRCA1-wt breast cancer cell lines (MCF-7, MBA-MD-157, and Hs578T) to determine the proteins governing response to Hsp90 inhibitor 17-allyloamino-17-demethoxy-geldanamycin. On treatment with the drug, cells arrested at G2-M phase and entered aberrant mitosis in a BRCA1-dependent manner. Failure to arrest the cells at or before mitosis resulted in formation of micronucleated cells, aberrant segregation of chromosomes, microtubule misalignment, and multicentrosomes, leading in eventual mitotic catastrophe cell death. Our observations show that BRCA1 mediates G2-M transition mainly through chek1 on 17-allyloamino-17-demethoxy-geldanamycin treatment. [Mol Cancer Ther 2008;7(8):2358–66]
High-content imaging characterization of cell cycle therapeutics through in vitro and in vivo subpopulation analysis
Authors: Low Jonathan , Huang Shuguang , Blosser Wayne , Dowless Michele , Burch John , Neubauer Blake , Stancato Louis
Published: August 2008

Although the cycling of eukaryotic cells has long been a primary focus for cancer therapeutics, recent advances in imaging and data analysis allow even further definition of cellular events as they occur in individual cells and cellular subpopulations in response to treatment. High-content imaging (HCI) has been an effective tool to elucidate cellular responses to a variety of agents; however, these data were most frequently observed as averages of the entire captured population, unnecessarily decreasing the resolution of each assay. Here, we dissect the eukaryotic cell cycle into individual cellular subpopulations using HCI in conjunction with unsupervised K-means clustering. We generate distinct phenotypic fingerprints for each major cell cycle and mitotic compartment and use those fingerprints to screen a library of 310 commercially available chemotherapeutic agents. We determine that the cell cycle arrest phenotypes caused by these agents are similar to, although distinct from, those found in untreated cells and that these distinctions frequently suggest the mechanism of action. We then show via subpopulation analysis that these arrest phenotypes are similar in both mouse models and in culture. HCI analysis of cell cycle using data obtained from individual cells under a broad range of research conditions and grouped into cellular subpopulations represents a powerful method to discern both cellular events and treatment effects. In particular, this technique allows for a more accurate means of assessing compound selectivity and leads to more meaningful comparisons between so-called targeted therapeutics. [Mol Cancer Ther 2008;7(8):2455–63]
Relaxin treatment of solid tumors: effects on electric field–mediated gene delivery
Authors: Henshaw Joshua , Mossop Brian , Yuan Fan
Published: August 2008

Pulsed electric fields have been shown to enhance interstitial transport of plasmid DNA (pDNA) in solid tumors in vivo. However, the extent of enhancement is still limited partly due to the collagen component in extracellular matrix. To this end, effects of collagen remodeling on interstitial electrophoresis were investigated by pretreatment of tumor-bearing mice with a recombinant human relaxin (rh-Rlx). In the study, two tumor lines (4T1 and B16.F10) were examined and implanted s.c. to establish two murine models: dorsal skin-fold chamber (DSC) and hind leg. Effects of rh-Rlx on pDNA electrophoresis were measured either directly in the DSC model or indirectly in the hind leg model via reporter gene expression. It was observed that rh-Rlx treatment reduced collagen levels in the hind leg tumors but not in the DSC tumors. The observation correlated with the results from electromobility experiments, where rh-Rlx treatment enhanced transgene expression in 4T1 hind leg tumors but did not increase the electromobility of pDNA in the DSC tumors. In addition, it was observed that pDNA binding to collagen could block its diffusion in collagen gel in vitro. These observations showed that effects of rh-Rlx on the collagen content depended on microenvironment in solid tumors and that rh-Rlx treatment would enhance electric field–mediated gene delivery only if it could effectively reduce the collagen content in collagen-rich tumors. [Mol Cancer Ther 2008;7(8):2566–73]
Analysis of the Association of Preeclampsia with Polymorphisms of the INS, INSR and IRS1 Genes in Mexican Women
Authors: Machorro-Lazo Maria-Victoria , Sanchez-Corona Jose , Martínez-Abundis Esperanza , González-Ortiz Manuel , Galaviz-Hernandez Carlos , Perea Francisco-Javier , Garcia-Zapien Alejandra-Guadalupe , Cruz-Quevedo Edhit-Guadalupe , Salgado-Goytia Lorenzo , Moran-Moguel Maria-Cristina , Flores-Martínez Silvia-Esperanza
Published: August 2008

Background/Aims: It has been proposed that preeclampsia is a metabolic syndrome of pregnancy. The polymorphisms PstI and MaeIII of INS, NsiI of INSR and Ala513Pro and Gly972Arg of IRS1 have been associated with metabolic syndrome; moreover, the products of these genes are functionally contiguous during insulin signaling. The aim of this study was to assess whether these polymorphisms are associated with preeclampsia. Methods: 46 normotensive pregnant women and 43 preeclamptic patients were included in the study to develop a clinical, biochemical and genotypic profile of preeclampsia. Clinical evaluation consisted of measurement of blood pressure, height and weight. Peripheral blood samples were collected for determination of fasting glucose and insulin concentrations and for extraction of genomic DNA. Proteinuria was determined. Polymorphisms were detected using PCR-RFLP. Results: The normotensive and preeclampsia groups did not differ significantly in clinical and biochemical traits, except for systolic and diastolic blood pressure (p < 0.0001). Polymorphisms previously associated with metabolic syndrome in Mexican populations were not associated with preeclampsia in Mexican women (p > 0.05). Conclusion: The lack of an association between preeclampsia and the polymorphisms studied suggests that other genes whose products do not have direct functional interaction with metabolic syndrome or epigenetic factors may play a role in preeclampsia.
Correlation of Quality of Life with Clinical Parameters and Eosinophilic Cation Protein Levels in Children with Allergic Rhinoconjunctivitis
Authors: Yuksel Hasan , Yilmaz Ozge , Sogut Ayhan , Ertan Pelin , Onur Ece
Published: August 2008

Background: Systemic and mucosal roles of eosinophils in the pathogenesis of allergic rhinoconjunctivitis (AR) are known. The aim of this study was to investigate the relationship of clinical parameters and quality of life with eosinophilic cationic protein (ECP) in grass pollen-sensitive children with seasonal AR. Methods: This study included 31 children with AR and 18 healthy controls aged between 5 and 15 years. ECP levels in nasal lavage fluid and serum were measured. AR symptom scores were calculated and the pediatric rhinoconjunctivitis quality of life questionnaire (PRQLQ) was filled in for all patients. Results: Mean serum and nasal ECP levels (ng/ml) were significantly higher in the patient group (p < 0.05 for both). In the AR group, the mean clinical symptom score was 7.3 ± 2.2, while the total PRQLQ score, activity limitation, symptoms and emotional function domains were 2.5 ± 0.9, 3.0 ± 1.1, 2.5 ± 1.2 and 2.6 ± 1.1, respectively. The total clinical symptom score and disease duration showed a significant correlation with the total PRQLQ score (p = 0.00 and 0.003, respectively). However, neither nasal lavage nor serum ECP levels were significantly correlated with symptom score, duration of disease, PRQLQ total score or domains (p > 0.05 for all). Conclusion: These results may indicate the absence of a correlation between clinical status and quality of life and levels of ECP in tissues with allergic inflammation.
Can the Use of HEPA Cleaners in Homes of Asthmatic Children and Adolescents Sensitized to Cat and Dog Allergens Decrease Bronchial Hyperresponsiveness and Allergen Contents in Solid Dust?
Authors: Sulser Claudia , Schulz Gabriele , Wagner Petra , Sommerfeld Christine , Keil Thomas , Reich Andreas , Wahn Ulrich , Lau Susanne
Published: August 2008

Background: Exposure and sensitization to pet allergens are associated with allergic asthma in children. Conflicting data have emerged regarding the potential benefit of air cleaners with respect to a reduction of indoor pet allergens and bronchial hyperresponsiveness (BHR). Methods: In a randomized controlled trial 36 asthmatic children with sensitization to cat and/or dog and significant exposure to cat and/or dog allergen (Fel d 1 and/or Can f 1 >500 ng/g of carpet dust) were included in order to study the effect of high-efficiency particulate arresting (HEPA) air cleaners placed in the living room and bedroom compared with the effect of sham air cleaners. Patients were allocated to two groups: group 1 exposed to active filters and group 2 exposed to sham filters. At month 0, after 6 months and after 12 months, pulmonary function testing and cold air challenge were performed, serum eosinophil cationic protein (ECP) and specific IgE to seven aeroallergens were determined, and carpet dust samples and filters were collected. Major pet allergen concentrations (Fel d 1, Can f 1) were determined in filters and bulk dust samples. Results: Thirty patients completed the study. After 6 and 12 months, there was no significant change in delta FEV1 after cold air challenge, or in the use of medication and serum ECP levels. However, there was a trend in the active group towards an improvement of bronchial hyperresponsiveness, whereas the sham filter group showed a deterioration of BHR. Conclusion: Although HEPA air cleaners retained airborne pet allergens, no effect on disease activity or allergen concentrations in bulk dust samples was observed in this study.
IgE-Mediated Occupational Asthma from Epoxy Resin
Authors: Hannu Timo , Frilander Heikki , Kauppi Paula , Kuuliala Outi , Alanko Kristiina
Published: August 2008

Background: Epoxy resins (ERs) are used in paints and other protective coatings, including flooring materials. Bisphenol A diglycidyl ether (BADGE) ERs (BADGE ERs) account for about 75% of the ERs used world-wide. ERs can cause both immediate and delayed allergic reactions, but immediate reactions are rare. Methods: Occupational asthma (OA) was diagnosed on the basis of a specific challenge test combined with the patient’s history of occupational exposure and respiratory symptoms. Results: A 39-year-old nonsmoking construction worker experienced dyspnea when laying ER-containing floors, but not in other situations. He also presented skin symptoms. IgE-mediated allergy to BADGE ER could be verified with both serum IgE antibodies and skin prick tests. The specific bronchial challenge test with BADGE ER caused an immediate asthmatic reaction. On patch testing, a positive reaction was provoked by BADGE ER. Conclusions: This is the first study on a patient exposed to BADGE ER with IgE-mediated immediate OA, based on a positive inhalation challenge test. If work-related respiratory symptoms develop when handling ERs, the possibility of OA should be recognized.
Lactobacillus rhamnosus GG and Lactobacillus casei Suppress Escherichia coli-Induced Chemokine Expression in Intestinal Epithelial Cells
Authors: Toki Shinji , Kagaya Shinji , Shinohara Miwa , Wakiguchi Hiroshi , Matsumoto Takashi , Takahata Yoshihisa , Morimatsu Fumiki , Saito Hirohisa , Matsumoto Kenji
Published: August 2008

Background: Recently, some strains of lactic acid bacteria (LAB) have been reported to prevent the development of atopic dermatitis and to improve allergic symptoms, especially in young children. However, the mechanisms involved in these effects are not fully understood. Intestinal microbiota play critical roles in the development of host immune development and are recognized and regulated by the host through intestinal epithelial cells (IECs). We thus hypothesized that LAB influence the host immune system through the activation of IECs. To begin testing this hypothesis, chemokine expression in IECs exposed to intestinal bacteria was investigated. Methods: Caco-2 cell monolayers were stimulated with different concentrations of various live or heat-killed intestinal bacteria or bacterial components for up to 3 h. Changes in the gene expressions of various chemokines were measured using quantitative real-time PCR. Results: The expressions of CCL20, CXCL8, CXCL10 and CX3CL1 were strongly induced by nonpathogenic Escherichia coli in a dose-dependent manner and were partially induced by some commensal LAB. In contrast, Lactobacillus rhamnosus GG (LGG) and Lactobacillus casei did not induce these chemokine expressions. In addition, LGG significantly suppressed the expressions of CCL20 and CXCL10 induced by E. coli, peptidoglycan or flagellin when cultured simultaneously. Conclusions:LGG and L. casei markedly suppressed E. coli-induced chemokine expression, presumably through the suppression of the Toll-like receptor-mediated signal transduction pathway, at least in part. The clinical importance of this suppressive effect and the mechanisms involved require further investigation; however, such effects can be used as a marker to identify clinically useful LAB.
Allergic Sensitization to Bovine β-Lactoglobulin: Comparison between Germ-Free and Conventional BALB/c Mice
Authors: Hazebrouck S. , Przybylski-Nicaise L. , Ah-Leung S. , Adel-Patient K. , Corthier G. , Wal J.-M. , Rabot S.
Published: August 2008

Background: The ‘hygiene hypothesis’ suggests that high hygienic standards met in western countries lead to a lack of microbial exposure, thus promoting the development of atopy by preventing the proper maturation of the immune system. Germ-free animals are deprived of the immune stimulation that occurs during postnatal gut colonization by commensal bacteria. Germ-free mice could thereby provide an attractive model for studying the impact of gut microbiota on the development of Th2-mediated disorders such as allergy. Methods: Germ-free and conventional BALB/c mice were sensitized to β-lactoglobulin (BLG), a major cow’s milk allergen, by means of intraperitoneal injections in the presence of incomplete Freund’s adjuvant. Time courses of serum and fecal BLG-specific antibody responses were monitored and cytokine production was assayed in BLG-reactivated splenocytes. Results: Serum BLG-specific IgG1 and IgE concentrations were significantly higher in germ-free mice during the primary immune response and IgE production persisted longer in germ-free mice. Furthermore, secretion of BLG-specific IgA was evidenced only in feces from germ-free mice while, in contrast, fecal IgG1 concentrations were at least 3-fold higher in conventional mice than in germ-free mice. Production of IL-5, IL-10 and IFN-γ was 3-fold enhanced in BLG-reactivated splenocytes from germ-free mice. Conclusion: The absence of gut microbiota significantly affects the BLG-specific immune response in BALB/c mice, thus suggesting that this model might be of interest for further studies exploring the influence of gut colonization by different bacterial strains on the development of an allergic-type sensitization.
Haplotype Analysis of the NADPH Oxidase p22phox Gene in Patients with Bronchial Asthma
Authors: Izakovicova Holla Lydie , Kaňková Kateřina , Znojil Vladimir
Published: August 2008

Background: Superoxide produced by airway epithelial and inflammatory cells may contribute to the pathogenesis of asthma. NADPH oxidase (an enzyme complex made up of 5 subunits) is a major source of superoxide in cells. We sought to elucidate the role of genetic variability in the gene encoding the p22phox subunit of the NADPH oxidase (CYBA, 16q24.3) in asthma susceptibility by means of an association study of haplotypes based on 3 common single nucleotide polymorphisms (SNPs), –930A/G, 242C/T (H72Y) and 640A/G. Methods: SNPs were genotyped by PCR-RFLP methods in a cohort of age- and sex-matched subjects with bronchial asthma (n = 305) and healthy controls (n = 311). Haplotypes were constructed in silico using a Monte Carlo-based algorithm. Furthermore, association of SNPs with sensitization to selected allergens (total 15 tested by skin prick test positivity/negativity) was analyzed. Results: In a single locus analysis, SNPs 242C/T and 640A/G were marginally significantly associated with asthma (p = 0.036 and 0.036, respectively). SNP 640A/G showed a significant association with sensitization to 2 allergens tested (pcorr < 0.02). Haplotype analysis identified a total of 8 haplotypes with population frequencies from 0.07 to 0.22. Distribution of haplotypes significantly differed between cases and controls (omnibus p = 0.017, 10,000 permutations). In the post-hoc analysis, haplotype CYBA3 (–930G/242T/640A) was associated with an increased risk of asthma (pcorr < 0.05; OR = 1.43, 95% CI 1.06–1.93). Conclusions: This study supports the hypothesis that genetic variability in the CYBA gene probably contributes to the susceptibility to bronchial asthma (or its related phenotypes) in our population.
Familial Aggregation of Skin Sensitization to Aeroallergens in a Rural Area in China
Authors: Jin Yongtang , Wang Weiqin , Xu Yingchun , Zhao Jinzhuo , Liu Hao , Xue Saoli
Published: August 2008

Background: The prevalence of allergic sensitization has increased all over the world during last years and allergic sensitization may be related to genetic or environmental factors or to both. The objective of this study was to estimate the prevalence rates of allergic sensitization and assess the familial aggregation of allergic sensitization to aeroallergens in asthma index families. Methods: The skin prick test (SPT) reactivity to 8 aeroallergens and the SPT reactivity-related risk factors were evaluated. The allergic sensitization was based on SPT. Univariate and multivariate logistic regressive models were used to estimate the effect size (odds ratio, OR) of correlation within families with positive SPT. Results: Of 4,213 analyzed subjects in 1,006 asthma index families, 40.9% had at least one positive SPT, 42.3% in parents (father: 43.3%; mother: 42.3%), 43.9% in first offspring, and 36.0% in subsequent offspring. The most common allergens were Dermatophagoides pteronyssinus (DP), cockroach and Dermatophagoides farinae (DF). Allergic sensitization status of both parents and first offspring was related to the risk of allergic sensitization for subsequent offspring. The OR for the positive (father)-positive (first offspring) group was 1.92 (p < 0.01), 3.26 (p < 0.01) for the positive (mother)-positive (first offspring) group, and 2.12 (p < 0.01) for the positive (father)-positive (mother) group, which were adjusted for age, sex, body mass index, history of asthma and smoking status. Conclusions: Our findings supported a significant familial aggregation of allergic sensitization in this Chinese population and highlighted the role of genetic factors in allergic sensitization.
Epidemiology, Characteristics and Distinctiveness of Headaches in Children from Vojvodina, Serbia
Authors: Knežević-Pogančev M.
Published: August 2008

From 1988 to 2004, an investigation was carried out on a population of 30,636 children (50.38% boys and 49.62% girls) in 9 towns of Vojvodina, province in northern Serbia. Idiopathic recurrent headache was found in 27.46% of children aged 3–17 years (26.8% of males and 28.4% of females). Migraine headache was found in 8.63% of children aged 3–17 years (8.0% of males and 9.6% of females). Recurrent headaches showed an occurrence that increased with age in the observed group, from 16.65 to 32.87% in boys and from 14.23 to 40.50% in girls (ages 3–5 and >12 years, respectively). Of the children with migraine, 25.55% had migraine with aura, 67.21% had migraine without aura and 7.23% had other migraine types. Idiopathic recurrent headache was found in children at a mean age of 9 years and 2.5 months (range 3–17 years). The presence of both recurrent and migraine headaches was found to be higher than previously described. In children, these headaches appear to be significantly associated with incomplete families, a poor economic situation and being the second-born child. Children with recurrent headache were diagnosed and followed by a general practitioner in 31.5% of cases, by a pediatrician in 21.6% and by a neurologist or neuropediatrician in the remaining 46.8%. Normal daily activity was compromised in children with recurrent headache for 3.91 days per year.