bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of Tool Geometry on Punching Performance
Authors: Singh U. P. , Klingenberg W. , Urquhart W.
Published: April 2008

In today’s competitive industrial environment, the quality of a product is no longer the major factor which influences purchase by a customer, it is simply a prerequisite for competing in the market. To compete and survive, the necessity is to provide the product at an affordable price, a target that can only be attained through actions such as improving the efficiency of manufacturing operations. In sheet metal industries, the cost of tooling contributes a considerable part to the overall manufacturing cost of a component. It is imperative to keep this cost down by ensuring the tool operates uninterrupted for long periods of time, achievable in one way by reducing the stress on the tool during punching by optimizing the clearance between the die and the punch. This paper deals with this problem using finite element techniques. Having created models of various punch configurations, one is able to analyze the effect of variations in tool geometry on punching force and tool deformation, a parameter highly relevant in assessing performance in terms of the accuracy of machined components. Computed results using FE models are compared against results obtained through experimentation. Due to the nature of the punching process being inherently noisy, consideration was given to noise levels, how they are affected by changes in tool geometry, and the subsequent effect on the workforce considering the fact that reducing stress on the operator could only improve quality and productivity. Finally, some suggestions are given as to how the efficiency of a punching tool and quality of the punched product can be improved.
Activity of Pradofloxacin against
Porphyromonas
and
Prevotella
spp. Implicated in Periodontal Disease in Dogs: Susceptibility Test Data from a European Multicenter Study
Authors: Stephan Bernd , Greife Heinrich A. , Pridmore Andrew , Silley Peter
Published: April 2008

Collaborating veterinarians from five European countries collected subgingival bacterial samples from dogs exhibiting clinical periodontal disease. Sterile endodontic paper points were used for collection of the samples, which were transported to a central laboratory for susceptibility testing. Anaerobic bacteria were isolated and
Porphyromonas
and
Prevotella
isolates identified to the species level; susceptibility to pradofloxacin and metronidazole was determined using the CLSI agar dilution methodology. A total of 630 isolates, 310 of
Porphyromonas
spp. and 320 of
Prevotella
spp., were isolated. Pradofloxacin MIC data for all isolates were in the range of ≤0.016 to 1 μg/ml, the overall MIC
50
was 0.062, and the overall MIC
90
was 0.25 μg/ml. There were no differences in activity against
Porphyromonas
and
Prevotella
isolates or in the pradofloxacin susceptibility distributions from the different European countries. All isolates were within the wild-type distribution and were fully susceptible to pradofloxacin. Metronidazole was also highly active against these strains: 316 of 320
Prevotella
strains (98.8%) and 309 of 310
Porphyromonas
strains (99.7%) were susceptible (MICs of ≤8 μg/ml). However, three
Prevotella
strains had intermediate metronidazole susceptibility (MICs of 16 μg/ml), while one
Prevotella
and one
Porphyromonas
strain were metronidazole resistant (MICs of 128 and 256 μg/ml, respectively). Pradofloxacin, a novel broad-spectrum fluoroquinolone, demonstrates a high degree of antianaerobic activity against strains isolated from clinical cases of periodontal disease and shows activity against metronidazole-resistant isolates. The broad-spectrum activity of pradofloxacin makes it a suitable candidate for the treatment of periodontal disease in dogs.
Susceptibility of
Chlamydia trachomatis
to the Excipient Hydroxyethyl Cellulose: pH and Concentration Dependence of Antimicrobial Activity
Authors: Sater Ali A. Abdul , Ojcius David M. , Meyer Matthew P.
Published: April 2008

Hydroxyethyl cellulose (HEC) is used as a neutral excipient in microbicides used against sexually transmitted pathogens. However, HEC inhibits the infection of cervical epithelial cells by
Chlamydia trachomatis
at pH 5 in a concentration-dependent manner. At pH 7, infection is inversely dependent on the concentration of HEC, possibly due to pH-dependent calcium sequestration.
Local Treatment of Experimental
Pseudomonas aeruginosa
Osteomyelitis with a Biodegradable Dilactide Polymer Releasing Ciprofloxacin
Authors: Kanellakopoulou Kyriaki , Thivaios George C. , Kolia Maria , Dontas Ismini , Nakopoulou Lydia , Dounis Eleftherios , Giamarellos-Bourboulis Evangelos J. , Andreopoulos Andreas , Karagiannakos Panayotis , Giamarellou Helen
Published: April 2008

A biodegradable system of poly-
d
,
l
-dilactide releasing ciprofloxacin was assessed in a
Pseudomonas aeruginosa
osteomyelitis model after inoculation of the test pathogen into the left tibia of 76 New Zealand White rabbits; 31 were controls (group A), and 45 were implanted with the polymer at the infection site (group B). The rabbits were killed on a weekly basis, and cancellous bone was harvested for histopathology and for estimation of bacterial growth and the concentrations of ciprofloxacin. Tibial X ray was performed immediately before the animals were killed. The total number of fistulas with purulent discharge that developed after inoculation of the pathogen was counted, and fistulas with purulent discharge were found in 16 animals in group A (51.6%) and 3 animals in group B (6.7%) (
P
< 0.0001). The animals in group A had a profound loss of body weight compared to the animals in group B. The main radiological finding was the presence of sequestra in 25 animals (80.6%) in group A and 6 animals in group B (13.3%) (
P
< 0.0001). The bacterial load in group B was significantly reduced compared to that in group A, possibly due to the prolonged local antibiotic release at concentrations exceeding even 80 times the MIC for the test pathogen. The histology of animals killed after week 49 revealed a mild inflammatory reaction accompanied by diffuse fibrosis and new bone formation in group A animals and the presence of small polymer particles in group B animals. It is concluded that the system described achieved eradication of the pathogen, accompanied by clinical and radiologically confirmed benefits, so this treatment may be a candidate for the management of difficult orthopedic infections.
Whole-Genome Pyrosequencing of an Epidemic Multidrug-Resistant
Acinetobacter baumannii
Strain Belonging to the European Clone II Group
Authors: Iacono Michele , Villa Laura , Fortini Daniela , Bordoni Roberta , Imperi Francesco , Bonnal Raoul J. P. , Sicheritz-Ponten Thomas , De Bellis Gianluca , Visca Paolo , Cassone Antonio , Carattoli Alessandra
Published: April 2008

The whole-genome sequence of an epidemic, multidrug-resistant
Acinetobacter baumannii
strain (strain ACICU) belonging to the European clone II group and carrying the plasmid-mediated
bla
OXA
-
58
carbapenem resistance gene was determined. The
A. baumannii
ACICU genome was compared with the genomes of
A. baumannii
ATCC 17978 and
Acinetobacter baylyi
ADP1, with the aim of identifying novel genes related to virulence and drug resistance.
A. baumannii
ACICU has a single chromosome of 3,904,116 bp (which is predicted to contain 3,758 genes) and two plasmids, pACICU1 and pACICU2, of 28,279 and 64,366 bp, respectively. Genome comparison showed 86.4% synteny with
A. baumannii
ATCC 17978 and 14.8% synteny with
A. baylyi
ADP1. A conspicuous number of transporters belonging to different superfamilies was predicted for
A. baumannii
ACICU. The relative number of transporters was much higher in ACICU than in ATCC 17978 and ADP1 (76.2, 57.2, and 62.5 transporters per Mb of genome, respectively). An antibiotic resistance island, AbaR2, was identified in ACICU and had plausibly evolved by reductive evolution from the AbaR1 island previously described in multiresistant strain
A. baumannii
AYE. Moreover, 36 putative alien islands (pAs) were detected in the ACICU genome; 24 of these had previously been described in the ATCC 17978 genome, 4 are proposed here for the first time and are present in both ATCC 17978 and ACICU, and 8 are unique to the ACICU genome. Fifteen of the pAs in the ACICU genome encode genes related to drug resistance, including membrane transporters and ex novo acquired resistance genes. These findings provide novel insight into the genetic basis of
A. baumannii
resistance.
Utilization of an Intracellular Bacterial Community Pathway inKlebsiella pneumoniaeUrinary Tract Infection and the Effects of FimK on Type 1 Pilus Expression
Authors: Rosen David A. , Pinkner Jerome S. , Jones Jennifer M. , Walker Jennifer N. , Clegg Steven , Hultgren Scott J.
Published: April 2008

Klebsiella pneumoniaeis an important cause of urinary tract infection (UTI), but little is known about its pathogenesis in vivo. The pathogenesis of theK. pneumoniaecystitis isolate TOP52 was compared to that of the uropathogenicEscherichia coli(UPEC) isolate UTI89 in a murine cystitis model. Bladder and kidney titers of TOP52 were lower than those of UTI89 at early time points but similar at later time points. TOP52, like UTI89, formed biofilm-like intracellular bacterial communities (IBCs) within the murine bladder, albeit at significantly lower levels than UTI89. Additionally, filamentation of TOP52 was observed, a process critical for UTI89 evasion of neutrophil phagocytosis and persistence in the bladder. Thus, the IBC pathway is not specific to UPEC alone. We investigated if differences in type 1 pilus expression may explain TOP52's early defect in vivo. The type 1 pilus operon is controlled by recombinase-mediated (fimE,fimB, andfimX) phase variation of an invertible promoter element. We found thatK. pneumoniaecarries an extra gene of unknown function at the 3′ end of its type 1 operon,fimK, and the genome lacks the recombinasefimX. A deletion mutant offimKwas constructed, and TOP52 ΔfimKhad higher titers and formed more IBCs in the murine cystitis model than wild type. The loss offimKor expression ofE. coli fimXfrom a plasmid in TOP52 resulted in a larger phase-ON population and higher expression levels of type 1 pili and gave TOP52 the ability to form type 1-dependent biofilms. Complementation with pfimKdecreased type 1 pilus expression and biofilm formation of TOP52 ΔfimKand decreased UTI89 biofilm formation. Thus,K. pneumoniaeappears programmed for minimal expression of type 1 pili, which may explain, in part, whyK. pneumoniaeis a less prevalent etiologic agent of UTI than UPEC.
gp96 Is a Human Colonocyte Plasma Membrane Binding Protein for
Clostridium difficile
Toxin A
Authors: Na Xi , Kim Ho , Moyer Mary P. , Pothoulakis Charalabos , LaMont J. Thomas
Published: April 2008

Clostridium difficile
toxin A (TxA), a key mediator of antibiotic-associated colitis, requires binding to a cell surface receptor prior to internalization. Our aim was to identify novel plasma membrane TxA binding proteins on human colonocytes. TxA was coupled with biotin and cross-linked to the surface of HT29 human colonic epithelial cells. The main colonocyte binding protein for TxA was identified as glycoprotein 96 (gp96) by coimmunoprecipitation and mass spectrum analysis. gp96 is a member of the heat shock protein family, which is expressed on human colonocyte apical membranes as well as in the cytoplasm. TxA binding to gp96 was confirmed by fluorescence immunostaining and in vitro coimmunoprecipitation. Following TxA binding, the TxA-gp96 complex was translocated from the cell membrane to the cytoplasm. Pretreatment with gp96 antibody decreased TxA binding to colonocytes and inhibited TxA-induced cell rounding. Small interfering RNA directed against gp96 reduced gp96 expression and cytotoxicity in colonocytes. TxA-induced inflammatory signaling via p38 and apoptosis as measured by activation of BAK (Bcl-2 homologous antagonist/killer) and DNA fragmentation were decreased in gp96-deficient B cells. We conclude that human colonocyte gp96 serves as a plasma membrane binding protein that enhances cellular entry of TxA, participates in cellular signaling events in the inflammatory cascade, and facilitates cytotoxicity.
Perceived anxiety and plasma cortisol concentrations following rock climbing with differing safety rope protocols
Authors: Hodgson C I , Draper N , McMorris T , Jones G , Fryer S , Coleman I
Published: April 2008

Objectives:

To examine how different safety rope protocols impact on subjective anxiety and self-confidence levels and plasma cortisol concentrations and the relationship between subjective states and cortisol during rock climbing.

Methods:

Participants (n = 12) were tested in three climbing conditions that were designed to invoke low, moderate and high physical and mental stress. Plasma cortisol concentrations were collected before and after climbing and participants reported subjective anxiety and self-confidence states for each climb.

Results:

Repeated measures analysis of variance showed significant differences between conditions for somatic anxiety (F2, 22 = 7.74, p = 0.009), self-confidence (F2, 22 = 9.52, p = 0.001) and change in plasma cortisol concentration (F2, 22 = 3.71, p = 0.041). Preplanned polynomial comparisons showed that these were linear effects; somatic anxiety was higher in the higher stress conditions whilst self-confidence was lower. Plasma cortisol concentration change was also linear. Regression analyses showed cubic relationships between plasma cortisol concentration and cognitive anxiety (R2 = 0.452), self-confidence (R2 = 0.281) and somatic anxiety (R2 = 0.268).

Conclusions:

There is a relationship between plasma cortisol concentration and subjective anxiety and self-confidence states during rock climbing. Changes in the way the safety rope is organised can impact on anxiety, cortisol concentration and self-confidence during rock climbing.
Conifer stomate analysis as a paleoecological tool: an example from the Hudson Bay Lowlands
Authors: Hansen Barbara C. S.
Published: April 2008

The identification of conifer stomata in fossil pollen preparations of peat cores from the Hudson Bay Lowlands is used to determine the local presence of conifers in lieu of macrofossil analyses. The differentiation of eight conifer stomate types (Picea type, Larix laricina, Pinus sp., Abies sp., Tsuga mertensiana, Tsuga heterophylla, Chamaecyparis nootkatensis, and Thuja type) is accomplished with a key, diagrammatic stomate illustrations, photographs, and measurements. Results of fossil conifer-stomate analyses indicate that both Picea and Larix arrived locally in the Albany River area of the Hudson Bay Lowlands about 4800 BP. In the Old Man Bog area, Larix arrived earlier, about 6000 BP, but Picea arrived more than 2000 years later (3700 BP). Fossil stomate and pollen results are compared. Key words: conifer stomata, local conifer presence, Hudson Bay Lowlands.
Variation in cone morphology and seed characters in Pinus strobus in Quebec
Authors: Beaulieu Jean , Simon Jean-Pierre
Published: April 2008

To investigate the patterns of variation in white pine (Pinus strobus L.), 10 natural populations from Quebec were compared using the phenotypic variation of cone and seed traits. Eight characters were measured on 10 cones collected from 30 trees in each of the 10 sampled populations. Four populations were from the Ottawa River region and four from the St. Lawrence Lowlands region, while two populations were at the margin of the natural range of the species in Quebec, from the Abitibi region and Anticosti Island. Significant differences among populations were detected for each character. Estimates of repeatability of these traits were relatively high, suggesting substantial genotypic control over them. These estimates were consistent from population to population. Results of multivariate analyses suggest that populations from the Ottawa River region are similar to those from the St. Lawrence region, with populations from these regions diverging only in cone scale length. However, populations from the St. Lawrence region were more distinct from each other than those from the Ottawa River region. The population from Abitibi deviates considerably from the other populations, suggesting habitat-selection pressures acting at the margin of the natural range of the species under continental conditions. Finally, no general geographical trend was detected in the observed variation. Key words: white pine, morphology, cone, seed, variation.
Apical dominance in rhizomes of quackgrass, Elytrigia repens: the effect of auxin, cytokinins, and abscisic acid
Authors: Taylor J. S. , Robertson J. M. , Harker K. N. , Bhalla M. K. , Daly E. J. , Pearce D. W.
Published: April 2008

Experiments were designed to determine the impact of abscisic acid, indole-3-acetic acid, and cytokinins on dormancy of quackgrass (Elytrigia repens (L.) Nevski) rhizome axillary buds using exogenous hormone treatments and analysis of endogenous hormones. Exogenous hormone treatments were applied in solution or in lanolin paste to 5-node segments of rhizome with an apical tip intact or removed. Abscisic acid inhibited bud growth except at concentrations of 0.5 – 1 μg ∙ mL−1 when it stimulated growth: this appeared to be based on an inhibition of apical dominance rather than a stimulation of bud growth per se. Both indole-3-acetic acid and cytokinins stimulated bud growth, indole-3-acetic acid at concentrations of 0.5 – 5 μg ∙ mL−1 and cytokinins at higher concentrations (i.e., 10 – 100 μg ∙ mL−1). Indole-3-acetic acid also increased elongation of the buds, whereas abscisic acid and cytokinins did not. Levels of endogenous hormones were measured in bud samples: indole-3-acetic acid was quantified as its methyl ester by combined gas chromatography – mass spectrometry – selected ion monitoring; abscisic acid was quantified as its methyl ester by gas chromatography – electron capture; and cytokinins were quantified using a soybean callus bioassay. Hormone levels were generally higher in the most active buds of a 5-node section. Abscisic acid was also measured in buds 24 h after sheath leaf removal, a practice known to promote bud sprouting. Sheath leaf removal had no significant effect on abscisic acid levels. Key words: quackgrass, Elytrigia repens, auxins, abscisic acid, cytokinins, apical dominance.
Alterations in Aquatic Plant Community Structure following Liming of an Acidic Adirondack Lake
Authors: Weiher Evan R. , Boylen Charles W. , Bukaveckas Paul A.
Published: April 2008

The changes in the macrophyte community composition of Woods Lake, New York (Adirondack Region), following lake neutralization with calcite are described with reference to the immigration and extirpation of species. Seven new species were found after calcite addition; one of these, Potamogeton epihydrus, became the single most abundant species within the lake. Two species were extirpated from the lake (Sphagnum sp. and Utricularia geminiscapa) within 5 yr following the initial treatment. Overall plant coverage decreased from 47.6 to ~20% of the lake surface area. This decrease was due in large part to the decline of the previously dominant Utricularia purpurea. Expansion of floating-leafed species and a canopy-forming pondweed (P. epihydrus) signifies a shift in community structure towards that characteristic of unperturbed circumneutral lakes. Specific responses of neutralization were often not consistent with the known physiological ecology or biogeographic distribution of species.
Determination of Composition and Vertical Structure of Fish Communities Using in situ Measurements of Acoustic Target Strength
Authors: Barange M. , Hampton I. , Pillar S. C. , Soule M. A.
Published: April 2008

A split-beam echo-sounder was used to estimate in situ acoustic target strengths (TS) of fish from a number of different fish communities on the South African continental shelf. The TS and size distributions (obtained by aimed trawling) were used to describe the size structure and vertical distribution of these communities. TS distributions obtained from several monospecific populations of juvenile Cape hake (Merluccius capensis), the dominant species present, were self-consistent, and there was good correspondence between modes in the TS and length distributions of juvenile hake, adult round herring (Etrumeus whiteheadi), anchovy (Engraulis capensis) recruits, pelagic goby (Sufflogobius bibarbatus), horse mackerel (Trachurus trachurus capensis), and ribbon fish (Lepidopus caudatus). Average TS values for all these species, both per individual and normalized by weight, are presented and compared with published values. The use of TS information in studies of the small-scale community structure and dynamics of fish populations is discussed. It is concluded that the method can be effective on low-density, multispecific assemblages such as those in our study, avoiding many of the pitfalls of conventional net sampling.
Relationships between Common Biological Reference Points Used as Thresholds and Targets of Fisheries Management Strategies
Authors: Mace Pamela M.
Published: April 2008

Relationships between various biological reference points (BRPs) used to define thresholds and targets of fisheries management strategies were examined for a range of combinations of life history characteristics. Rank orderings of the selected BRPs were strongly influenced by the degree of density dependence in the underlying spawning–recruitment (S–R) relationship. The validity of F0.1, Fmax, F20% (the fishing mortality at which spawning per recruit is 20% of the maximum), and other reference fishing mortality rates as approximations to Fmsy or as thresholds of overfishing is highly dependent on life history characteristics, particular the degree of density dependence in the S–R relationship. It is recommended that F40% be adopted as a target fishing mortality rate when the S–R relationship is unknown and that threshold levels of biomass be related to the estimated or assumed degree of density dependence in the S–R relationship. Two new methods of calculating threshold biomass levels are suggested as alternatives to the familiar 20% B0 rule.
Life History Forms of Male Masu Salmon (Oncorhynchus masou) in South Primor'e, Russia
Authors: Tsiger V. V. , Skirin V. I. , Krupyanko N. I. , Kashkin K. A. , Semenchenko A. Yu.
Published: April 2008

In South Primor'e (Russia), mature male masu salmon (Oncorhynchus masou) parr are found at age 0 + to 3 + (9–22 cm). After first maturation, some male parr die, while others survive and mature the following year. Immature males become smolts after one or two winters in rivers and migrate to the sea in spring–early summer. After one winter in the sea, they return as sea-run normal-sized (40–70 cm) males at age R.1 +. Besides mature male parr and normal-sized sea-run males, jacks (28–33 cm) at age R.0 + were found. Such males return to rivers after some months of marine life and do not overwinter in the sea. Some jacks previously spawned as parr, while others did not. All jacks and normal-sized sea-run males die after spawning. Thus, male masu salmon mature as one of four life history forms: (1) male parr, (2) jacks that matured previously as parr, (3) jacks that did not mature previously as parr, and (4) ordinary sea-run males. While mature male parr and sea-run males are ordinary life history forms, both forms of jack are rare.
Long-term Experiments on the Stability of Two Fish Populations in Previously Unexploited Arctic Lakes
Authors: Johnson Lionel
Published: April 2008

Previous research indicated that fish populations in arctic lakes maintain a constant size distribution and abundance in the face of the environmental variability experienced over their recent history. Such stability was tested over 15 seasons in Little Nauyuk and Gavia lakes (Northwest Territories) which contained previously undisturbed populations of Arctic char (Salvelinus alpinus). Initially, the length–frequency distribution of Arctic char in Little Nauyuk Lake was bell shaped, and in Gavia Lake, it could be described by a negative logarithmic expression. Initial sampling of Gavia Lake removed the largest fish, allowing a well-defined mode to develop (cf. Little Nauyuk Lake). In both lakes the modal value remained constant throughout the exploitation phase. When fishing ceased the populations returned to their original state without evident oscillation. The stable state of the dominant population is considered to be one of "least dissipation". It is hypothesized that ecosystem structure depends on countervailing forces, one tending to decelerate energy flow through the ecosystem and the second tending to accelerate it. For ecosystems to exist, the tendency to decelerate energy flow must dominate system behaviour over ecological time. An ecosystem is regarded as a hemeorhetic system, stability seeking through the stabilization of energy flows.
STEADY STATES FOR ONE-DIMENSIONAL CURVATURE FLOWS
Authors: NI YILONG , ZHU MEIJUN
Published: April 2008

We define two conformal structures on S1which give rise to a different view of the affine curvature flow and a new curvature flow, the "Q-curvature flow". The steady states of these flows are studied. More specifically, we prove four sharp inequalities, which state the existence of the corresponding extremal metrics.
LEFT-SYMMETRIC BIALGEBRAS AND AN ANALOGUE OF THE CLASSICAL YANG–BAXTER EQUATION
Authors: BAI CHENGMING
Published: April 2008

We introduce a notion of left-symmetric bialgebra which is an analogue of the notion of Lie bialgebra. We prove that a left-symmetric bialgebra is equivalent to a symplectic Lie algebra with a decomposition into a direct sum of the underlying vector spaces of two Lagrangian subalgebras. The latter is called a parakähler Lie algebra or a phase space of a Lie algebra in mathematical physics. We introduce and study coboundary left-symmetric bialgebras and our study leads to what we call "S-equation", which is an analogue of the classical Yang–Baxter equation. In a certain sense, the S-equation associated to a left-symmetric algebra reveals the left-symmetry of the products. We show that a symmetric solution of the S-equation gives a parakähler Lie algebra. We also show that such a solution corresponds to the symmetric part of a certain operator called "[Formula: see text]-operator", whereas a skew-symmetric solution of the classical Yang–Baxter equation corresponds to the skew-symmetric part of an [Formula: see text]-operator. Thus a method to construct symmetric solutions of the S-equation (hence parakähler Lie algebras) from [Formula: see text]-operators is provided. Moreover, by comparing left-symmetric bialgebras and Lie bialgebras, we observe that there is a clear analogue between them and, in particular, parakähler Lie groups correspond to Poisson–Lie groups in this sense.
"Good" worms and human rights
Authors: Aycock John , Maurushat Alana
Published: April 2008

The extent of Internet censorship in countries like China is regularly tested, but the testing methods used from within a censored country can entail risk for humans. A benevolent worm can be used for testing instead: the worm's self-replication, long the bane of suggested benevolent viruses and worms, is shown to be essential here. We describe the design of this benevolent worm, along with some other related applications for it. A technical, ethical, and legal analysis is provided.
Modeling the CD8+ T Effector to Memory Transition in Adoptive T-Cell Antitumor Immunotherapy
Authors: Rolle Cleo E. , Carrio Roberto , Malek Thomas R.
Published: April 2008

Adoptive T-cell therapy with CD8+ CTLs is often characterized by poor persistence of the transferred T cells and limited effector responses. Improved persistence and therapeutic efficacy have been noted when antigen-activated CD8+ T cells express properties of memory cells. The current study was undertaken to more precisely characterize the development of memory-like CD8+ T cells from short-term CTLs in vitro and upon transfer in vivo, including their antitumor activity. Ovalbumin (OVA)–specific OT-I CTLs acquired phenotypic and functional properties of memory cells 2 to 3 days later either by lowering the concentration of antigen to a level that does not support primary responses and providing a survival signal through transgenic Bcl-2 in vitro or simply by transferring early day 3 CTLs to antigen-free lymphoid-replete mice. In lymphoid-replete mice, established OVA-expressing E.G7 tumor was rejected by short-term CTLs that simultaneously acquired memory-like properties in secondary lymphoid tissues, where tumor antigen level remained low. Collectively, these data indicate that CTLs readily converted to memory-like cells upon lowering antigen to a concentration that selectively supports memory responses and suggest that such conversion predicts successful adoptive immunotherapy. [Cancer Res 2008;68(8):2984–92]
NVP-AUY922: A Novel Heat Shock Protein 90 Inhibitor Active against Xenograft Tumor Growth, Angiogenesis, and Metastasis
Authors: Eccles Suzanne A. , Massey Andy , Raynaud Florence I. , Sharp Swee Y. , Box Gary , Valenti Melanie , Patterson Lisa , de Haven Brandon Alexis , Gowan Sharon , Boxall Frances , Aherne Wynne , Rowlands Martin , Hayes Angela , Martins Vanessa , Urban Frederique , Boxall Kathy , Prodromou Chrisostomos , Pearl Laurence , James Karen , Matthews Thomas P. , Cheung Kwai-Ming , Kalusa Andrew , Jones Keith , McDonald Edward , Barril Xavier , Brough Paul A. , Cansfield Julie E. , Dymock Brian , Drysdale Martin J. , Finch Harry , Howes Rob , Hubbard Roderick E. , Surgenor Alan , Webb Paul , Wood Mike , Wright Lisa , Workman Paul
Published: April 2008

We describe the biological properties of NVP-AUY922, a novel resorcinylic isoxazole amide heat shock protein 90 (HSP90) inhibitor. NVP-AUY922 potently inhibits HSP90 (Kd = 1.7 nmol/L) and proliferation of human tumor cells with GI50 values of approximately 2 to 40 nmol/L, inducing G1-G2 arrest and apoptosis. Activity is independent of NQO1/DT-diaphorase, maintained in drug-resistant cells and under hypoxic conditions. The molecular signature of HSP90 inhibition, comprising induced HSP72 and depleted client proteins, was readily demonstrable. NVP-AUY922 was glucuronidated less than previously described isoxazoles, yielding higher drug levels in human cancer cells and xenografts. Daily dosing of NVP-AUY922 (50 mg/kg i.p. or i.v.) to athymic mice generated peak tumor levels at least 100-fold above cellular GI50. This produced statistically significant growth inhibition and/or regressions in human tumor xenografts with diverse oncogenic profiles: BT474 breast tumor treated/control, 21%; A2780 ovarian, 11%; U87MG glioblastoma, 7%; PC3 prostate, 37%; and WM266.4 melanoma, 31%. Therapeutic effects were concordant with changes in pharmacodynamic markers, including induction of HSP72 and depletion of ERBB2, CRAF, cyclin-dependent kinase 4, phospho-AKT/total AKT, and hypoxia-inducible factor-1α, determined by Western blot, electrochemiluminescent immunoassay, or immunohistochemistry. NVP-AUY922 also significantly inhibited tumor cell chemotaxis/invasion in vitro, WM266.4 melanoma lung metastases, and lymphatic metastases from orthotopically implanted PC3LN3 prostate carcinoma. NVP-AUY922 inhibited proliferation, chemomigration, and tubular differentiation of human endothelial cells and antiangiogenic activity was reflected in reduced microvessel density in tumor xenografts. Collectively, the data show that NVP-AUY922 is a potent, novel inhibitor of HSP90, acting via several processes (cytostasis, apoptosis, invasion, and angiogenesis) to inhibit tumor growth and metastasis. NVP-AUY922 has entered phase I clinical trials. [Cancer Res 2008;68(8):2850–60]
The Regulatory T Cell–Associated Transcription Factor FoxP3 Is Expressed by Tumor Cells
Authors: Ebert Lisa M. , Tan Bee Shin , Browning Judy , Svobodova Suzanne , Russell Sarah E. , Kirkpatrick Naomi , Gedye Craig , Moss Denis , Ng Sweet Ping , MacGregor Duncan , Davis Ian D. , Cebon Jonathan , Chen Weisan
Published: April 2008

FoxP3 is a member of the forkhead family of transcription factors critically involved in the development and function of CD25+ regulatory T cells (Treg). Until recently, FoxP3 expression was thought to be restricted to the T-cell lineage. However, using immunohistochemistry and flow cytometric analysis of human melanoma tissue, we detected FoxP3 expression not only in the tumor infiltrating Treg but also in the melanoma cells themselves. FoxP3 is also widely expressed by established human melanoma cell lines (as determined by flow cytometry, PCR, and Western blot), as well as cell lines derived from other solid tumors. Normal B cells do not express FoxP3; however, expression could be induced after transformation with EBV in vitro and in vivo, suggesting that malignant transformation of healthy cells can induce FoxP3. In addition, a FOXP3 mRNA variant lacking exons 3 and 4 was identified in tumor cell lines but was absent from Treg. Interestingly, this alternative splicing event introduces a translation frame-shift that is predicted to encode a novel protein. Together, our results show that FoxP3, a key regulator of immune suppression, is not only expressed by Treg but also by melanoma cells, EBV-transformed B cells, and a wide variety of tumor cell lines. [Cancer Res 2008;68(8):3001–9]
Apoptotic Activity and Mechanism of 2-Cyano-3,12-Dioxoolean-1,9-Dien-28-Oic-Acid and Related Synthetic Triterpenoids in Prostate Cancer
Authors: Hyer Marc L. , Shi Ranxin , Krajewska Maryla , Meyer Colin , Lebedeva Irina V. , Fisher Paul B. , Reed John C.
Published: April 2008

Synthetic triterpenoids 2-cyano-3, 12-dioxooleana-1, 9-(11)-dien-28-oic acid (CDDO) and CDDO-Me (CDDO-methyl ester) have entered clinical trials for cancer. We determined that CDDO analogues at submicromolar concentrations induce apoptosis of cultured prostate cancer cell lines, LNCaP, ALVA31, Du145, PC3, and PPC1, with lethal dose 50% ∼1 μmol/L for CDDO-Me and an imidazole analogue (CDDO-Im). These compounds induced apoptosis of prostate cancer cells as characterized by cleavage of caspase-3, caspase-7, caspase-8, caspase-9, caspase-10, BID, and poly(ADP)ribose polymerase and by dependence on caspase activity. Moreover, triterpenoid-induced cell death was abolished by caspase-8–targeting small interfering (si) RNA. To explore the mechanism(s) involved in caspase-8 activation, we examined cell surface expression of death receptor (DR)4 and DR5 after triterpenoid treatment. Cell surface DR4 and DR5 expression was significantly up-regulated by CDDO or CDDO-Im but not by CDDO-Me. DR4 and DR5 knockdown with siRNA significantly inhibited apoptosis induced by CDDO and CDDO-Im but had no effect on CDDO-Me–induced killing, suggesting that CDDO and CDDO-Im induce apoptosis by a different mechanism than CDDO-Me. In addition to activating the caspase-8–dependent extrinsic apoptosis pathway, we observed that Bcl-XL overexpression inhibited triterpenoid-mediated killing of prostate cancer cell line Du145, suggesting that the intrinsic pathway (via mitochondria) also participates in triterpenoid-mediated killing. In vivo antitumor activity of CDDO-Me was shown using a Du145 tumor xenograft model in nude rats. Altogether, these findings suggest CDDO and related synthetic triterpenoids should be further evaluated as potential novel therapeutics for hormone refractory prostate cancers. [Cancer Res 2008;68(8):2927–33]
Genetic Variation in the Inhibin Pathway and Risk of Testicular Germ Cell Tumors
Authors: Purdue Mark P. , Graubard Barry I. , Chanock Stephen J. , Rubertone Mark V. , Erickson Ralph L. , McGlynn Katherine A.
Published: April 2008

Gene-knockout studies in mice suggest that INHA, encoding a subunit of gonadotropin-regulating proteins known as inhibins, is a tumor suppressor for testicular stromal cell tumors. It is not known whether genetic variation in the inhibin pathway also influences susceptibility to testicular germ cell tumors (TGCT), the most common testicular cancer in young men. To address this question, we conducted a case-control analysis (577 cases; 707 controls) of single-nucleotide polymorphisms (SNP) in genes in the inhibin pathway among participants in the U.S. Servicemen's Testicular Tumor Environmental and Endocrine Determinants Study. Thirty-eight tagging SNPs in six genes (INHA, INHBA, INHBB, INHBC, INHBE, and SMAD4) were genotyped. Odds ratios (OR) and 95% confidence intervals (CI) relating variant genotypes to TGCT risk were calculated using unconditional logistic regression. Among White subjects, an elevated risk of TGCT was observed for carriers of the T allele of the INHA variant rs2059693 (CT genotype: OR, 1.33; 95% CI, 1.04–1.71; TT: OR, 1.60; 95% CI, 1.01–2.52; Ptrend = 0.008). The association with rs2059693 was stronger for nonseminomas, and for teratomas and teratocarcinomas in particular (N = 58; CT: OR, 1.63; 95% CI, 0.89–2.99; TT: OR, 4.54; 95% CI 2.00–10.3; Ptrend = 0.0008). We found no evidence of association with variants in the other investigated genes. These findings suggest that genetic variation in the INHA locus influences TGCT development. [Cancer Res 2008;68(8):3043–8]
Broad Overexpression of Ribonucleotide Reductase Genes in Mice Specifically Induces Lung Neoplasms
Authors: Xu Xia , Page Jennifer L. , Surtees Jennifer A. , Liu Houchun , Lagedrost Sarah , Lu Young , Bronson Roderick , Alani Eric , Nikitin Alexander Yu. , Weiss Robert S.
Published: April 2008

Ribonucleotide reductase (RNR) catalyzes the rate-limiting step in nucleotide biosynthesis and plays a central role in genome maintenance. Although a number of regulatory mechanisms govern RNR activity, the physiologic effect of RNR deregulation had not previously been examined in an animal model. We show here that overexpression of the small RNR subunit potently and selectively induces lung neoplasms in transgenic mice and is mutagenic in cultured cells. Combining RNR deregulation with defects in DNA mismatch repair, the cellular mutation correction system, synergistically increased RNR-induced mutagenesis and carcinogenesis. Moreover, the proto-oncogene K-ras was identified as a frequent mutational target in RNR-induced lung neoplasms. Together, these results show that RNR deregulation promotes lung carcinogenesis through a mutagenic mechanism and establish a new oncogenic activity for a key regulator of nucleotide metabolism. Importantly, RNR-induced lung neoplasms histopathologically resemble human papillary adenocarcinomas and arise stochastically via a mutagenic mechanism, making RNR transgenic mice a valuable model for lung cancer. [Cancer Res 2008;68(8):2652–60]
Androgen Receptor Overexpression in Prostate Cancer Linked to Purα Loss from a Novel Repressor Complex
Authors: Wang Longgui G. , Johnson Edward M. , Kinoshita Yayoi , Babb James S. , Buckley Michael T. , Liebes Leonard F. , Melamed Jonathan , Liu Xiao-Mei , Kurek Ralf , Ossowski Liliana , Ferrari Anna C.
Published: April 2008

Increased androgen receptor (AR) expression and activity are pivotal for androgen-independent (AI) prostate cancer (PC) progression and resistance to androgen-deprivation therapy. We show that a novel transcriptional repressor complex that binds a specific sequence (repressor element) in the AR gene 5′-untranslated region contains Purα and hnRNP-K. Purα expression, its nuclear localization, and its AR promoter association, as determined by chromatin immunoprecipitation analysis, were found to be significantly diminished in AI-LNCaP cells and in hormone-refractory human PCs. Transfection of AI cells with a plasmid that restored Purα expression reduced AR at the transcription and protein levels. Purα knockdown in androgen-dependent cells yielded higher AR and reduced p21, a gene previously shown to be under negative control of AR. These changes were linked to increased proliferation in androgen-depleted conditions. Treatment of AI cells with histone deacetylase and DNA methylation inhibitors restored Purα protein and binding to the AR repressor element. This correlated with decreased AR mRNA and protein levels and inhibition of cell growth. Purα is therefore a key repressor of AR transcription and its loss from the transcriptional repressor complex is a determinant of AR overexpression and AI progression of PC. The success in restoring Purα and the repressor complex function by pharmacologic intervention opens a promising new therapeutic approach for advanced PC. [Cancer Res 2008;68(8):2678–88]
Down-regulation of Caspase-2 by Rottlerin via Protein Kinase C-δ–Independent Pathway
Authors: Basu Alakananda , Adkins Brett , Basu Chandreyi
Published: April 2008

Protein kinase C-δ (PKCδ) plays an important role in DNA damage–induced apoptosis. We have previously shown that the PKCδ inhibitor rottlerin protects against cisplatin-induced apoptosis acting upstream of caspase-9. In the present study, we have investigated if rottlerin regulates caspase-2 activation. Knockdown of caspase-2 by siRNA inhibited processing of apical caspase-9 and caspase-8, whereas depletion of caspase-9 had little effect on caspase-2 processing. Rottlerin inhibited activation and processing of caspase-9 and caspase-8 and cleavage of poly(ADP)ribose polymerase. We made a novel observation that rottlerin induced down-regulation of caspase-2 but not of caspase-3, caspase-7, caspase-8, or caspase-9. Pharmacologic inhibitors of PKC, such as Gö 6983 and bisindolylmaleimide, or depletion of PKCδ by siRNA had no effect on the down-regulation of caspase-2 by rottlerin. The proteasome inhibitor MG132 reversed caspase-2 down-regulation by rottlerin, whereas calpain inhibitor had no effect. These results suggest that rottlerin induces down-regulation of caspase-2 via PKCδ-independent but ubiquitin proteasome–mediated pathway. Furthermore, down-regulation of caspase-2 by rottlerin can explain its antiapoptotic function during DNA damage–induced apoptosis. [Cancer Res 2008;68(8):2795–802]
The Mitochondrial Uncoupling Protein-2 Promotes Chemoresistance in Cancer Cells
Authors: Derdak Zoltan , Mark Nicholas M. , Beldi Guido , Robson Simon C. , Wands Jack R. , Baffy György
Published: April 2008

Cancer cells acquire drug resistance as a result of selection pressure dictated by unfavorable microenvironments. This survival process is facilitated through efficient control of oxidative stress originating from mitochondria that typically initiates programmed cell death. We show this critical adaptive response in cancer cells to be linked to uncoupling protein-2 (UCP2), a mitochondrial suppressor of reactive oxygen species (ROS). UCP2 is present in drug-resistant lines of various cancer cells and in human colon cancer. Overexpression of UCP2 in HCT116 human colon cancer cells inhibits ROS accumulation and apoptosis after exposure to chemotherapeutic agents. Tumor xenografts of UCP2-overexpressing HCT116 cells retain growth in nude mice receiving chemotherapy. Augmented cancer cell survival is accompanied by altered NH2-terminal phosphorylation of the pivotal tumor suppressor p53 and induction of the glycolytic phenotype (Warburg effect). These findings link UCP2 with molecular mechanisms of chemoresistance. Targeting UCP2 may be considered a novel treatment strategy for cancer. [Cancer Res 2008;68(8):2813–9]
Breast Carcinoma–Associated Fibroblasts and Their Counterparts Display Neoplastic-Specific Changes
Authors: Hawsawi Nahed M. , Ghebeh Hazem , Hendrayani Siti-Faujiah , Tulbah Asma , Al-Eid Maha , Al-Tweigeri Taher , Ajarim Dahish , Alaiya Ayodele , Dermime Said , Aboussekhra Abdelilah
Published: April 2008

It has become clear that the initiation and progression of carcinomas depend not only on alterations in epithelial cells, but also on changes in their microenvironment. To identify these changes, we have undertaken cellular and molecular characterization of carcinoma-associated fibroblasts (CAF) and their tumor counterpart fibroblasts (TCF) isolated from 12 breast cancer patients. Normal breast fibroblasts (NBF) from plastic surgery were used as normal control. We present evidence that both CAFs and TCFs are myofibroblasts and show tumor-associated features. Indeed, the p53/p21 response pathway to γ-rays was defective in 70% CAFs, whereas it was normal in all the TCF and NBF cells. In addition, the basal levels of the p53 and p21 proteins were significantly low in 83% of CAFs and modulated in the majority of TCFs compared with NBFs. Interestingly, both TCFs and CAFs expressed high levels of the cancer marker survivin and consequently exhibited high resistance to cisplatin and UV light. Moreover, most CAFs were positive for the proliferation marker Ki-67 and exhibited high proliferation rate compared with NBFs and TCFs. However, proliferating cell nuclear antigen was highly expressed in both CAFs and TCFs. Using the two-dimensional gel electrophoresis technique, we have also shown that CAF, TCF, and NBF cells present different proteome profiles, with many proteins differentially expressed between these cells. Taken together these results indicate that different genetic alterations can occur in breast CAFs and their corresponding adjacent counterparts, showing the important role that stroma could play in breast carcinogenesis and treatment. [Cancer Res 2008;68(8):2717–25]
Identification of Let-7–Regulated Oncofetal Genes
Authors: Boyerinas Benjamin , Park Sun-Mi , Shomron Noam , Hedegaard Mads M. , Vinther Jeppe , Andersen Jens S. , Feig Christine , Xu Jinbo , Burge Christopher B. , Peter Marcus E.
Published: April 2008

MicroRNAs (miRNA) are small RNA molecules of ∼20 to 22 nucleotides that reduce expression of proteins through mRNA degradation and/or translational silencing. Each known miRNA has a large number of predicted targets. Members of the let-7/miR-98 family of miRNAs are up-regulated at the end of embryonic development. Let-7 is often down-regulated early during cancer development, suggesting that let-7–regulated oncofetal genes (LOG) may become reexpressed in cancer cells. Using comparative bioinformatics, we have identified 12 conserved LOGs that include HMGA2 and IMP-1/CRD-BP. IMP-1 has growth-promoting activities through stabilization of c-myc mRNA. We experimentally confirmed that IMP-1 is a direct let-7 target that promotes cell growth and motility of tumor cells, and we confirmed by proteomics analysis that IMP-1 and HMGA2 are major miRNA targets. Our data suggest that a substantial part of the growth inhibitory activities of let-7 comes from suppressing the expression of IMP-1. LOGs could be novel therapeutic targets and potential biomarkers for cancer treatment. [Cancer Res 2008;68(8):2587–91]