bluuebunny/crossref_metadata_2025_split huggingface dataset:
Novel Antibiotics Targeting Respiratory ATP Synthesis in Gram-Positive Pathogenic Bacteria
Authors: Balemans Wendy , Vranckx Luc , Lounis Nacer , Pop Ovidiu , Guillemont Jérôme , Vergauwen Karen , Mol Selena , Gilissen Ron , Motte Magali , Lançois David , De Bolle Miguel , Bonroy Kristien , Lill Holger , Andries Koen , Bald Dirk , Koul Anil 
Published: May 2012

ABSTRACT

Emergence of drug-resistant bacteria represents a high, unmet medical need, and discovery of new antibacterials acting on new bacterial targets is strongly needed. ATP synthase has been validated as an antibacterial target in
Mycobacterium tuberculosis
, where its activity can be specifically blocked by the diarylquinoline TMC207. However, potency of TMC207 is restricted to mycobacteria with little or no effect on the growth of other Gram-positive or Gram-negative bacteria. Here, we identify diarylquinolines with activity against key Gram-positive pathogens, significantly extending the antibacterial spectrum of the diarylquinoline class of drugs. These compounds inhibited growth of
Staphylococcus aureus
in planktonic state as well as in metabolically resting bacteria grown in a biofilm culture. Furthermore, time-kill experiments showed that the selected hits are rapidly bactericidal. Drug-resistant mutations were mapped to the ATP synthase enzyme, and biochemical analysis as well as drug-target interaction studies reveal ATP synthase as a target for these compounds. Moreover, knockdown of the ATP synthase expression strongly suppressed growth of
S. aureus
, revealing a crucial role of this target in bacterial growth and metabolism. Our data represent a proof of principle for using the diarylquinoline class of antibacterials in key Gram-positive pathogens. Our results suggest that broadening the antibacterial spectrum for this chemical class is possible without drifting off from the target. Development of the diarylquinolines class may represent a promising strategy for combating Gram-positive pathogens.
MK-5172, a Selective Inhibitor of Hepatitis C Virus NS3/4a Protease with Broad Activity across Genotypes and Resistant Variants
Authors: Summa Vincenzo , Ludmerer Steven W. , McCauley John A. , Fandozzi Christine , Burlein Christine , Claudio Giuliano , Coleman Paul J. , DiMuzio Jillian M. , Ferrara Marco , Di Filippo Marcello , Gates Adam T. , Graham Donald J. , Harper Steven , Hazuda Daria J. , McHale Carolyn , Monteagudo Edith , Pucci Vincenzo , Rowley Michael , Rudd Michael T. , Soriano Aileen , Stahlhut Mark W. , Vacca Joseph P. , Olsen David B. , Liverton Nigel J. , Carroll Steven S. 
Published: May 2012

ABSTRACT
HCV NS3/4a protease inhibitors are proven therapeutic agents against chronic hepatitis C virus infection, with boceprevir and telaprevir having recently received regulatory approval as add-on therapy to pegylated interferon/ribavirin for patients harboring genotype 1 infections. Overcoming antiviral resistance, broad genotype coverage, and a convenient dosing regimen are important attributes for future agents to be used in combinations without interferon. In this communication, we report the preclinical profile of MK-5172, a novel P2-P4 quinoxaline macrocyclic NS3/4a protease inhibitor currently in clinical development. The compound demonstrates subnanomolar activity against a broad enzyme panel encompassing major hepatitis C virus (HCV) genotypes as well as variants resistant to earlier protease inhibitors. In replicon selections, MK-5172 exerted high selective pressure, which yielded few resistant colonies. In both rat and dog, MK-5172 demonstrates good plasma and liver exposures, with 24-h liver levels suggestive of once-daily dosing. When administered to HCV-infected chimpanzees harboring chronic gt1a or gt1b infections, MK-5172 suppressed viral load between 4 to 5 logs at a dose of 1 mg/kg of body weight twice daily (b.i.d.) for 7 days. Based on its preclinical profile, MK-5172 is anticipated to be broadly active against multiple HCV genotypes and clinically important resistance variants and highly suited for incorporation into newer all-oral regimens.
An Automated Image-Based Method for Rapid Analysis of Chlamydia Infection as a Tool for Screening Antichlamydial Agents
Authors: Osaka Ichie , Hills Jeffrey M. , Kieweg Sarah L. , Shinogle Heather E. , Moore David S. , Hefty P. Scott 
Published: May 2012

ABSTRACT

A major limitation in the identification of novel antichlamydial compounds is the paucity of effective methods for large-scale compound screening. The immunofluorescence assay is the preferred approach for accurate quantification of the intracellular growth of
Chlamydia
. In this study, an immunofluorescence image-based method (termed image-based automated chlamydial identification and enumeration [iBAChIE]) was customized for fully automated quantification of
Chlamydia
infection using the freely available open-source image analysis software program CellProfiler and the complementary data exploration software program CellProfiler Analyst. The method yielded enumeration of different species and strains of
Chlamydia
highly comparably to the conventional manual methods while drastically reducing the analysis time. The inhibitory capability of established antichlamydial activity was also evaluated. Overall, these data support that iBAChIE is a highly effective tool for automated quantification of
Chlamydia
infection and assessment of antichlamydial activities of molecules. Furthermore, iBAChIE is expected to be amenable to high-throughput screening studies for inhibitory compounds and fluorescently labeled molecules to study host-pathogen interactions.
Inactivation of Mycobacterium tuberculosis
l
,
d
-Transpeptidase Ldt
Mt1
by Carbapenems and Cephalosporins
Authors: Dubée Vincent , Triboulet Sébastien , Mainardi Jean-Luc , Ethève-Quelquejeu Mélanie , Gutmann Laurent , Marie Arul , Dubost Lionel , Hugonnet Jean-Emmanuel , Arthur Michel 
Published: May 2012

ABSTRACT

The structure of
Mycobacterium tuberculosis
peptidoglycan is atypical since it contains a majority of 3→3 cross-links synthesized by
l
,
d
-transpeptidases that replace 4→3 cross-links formed by the
d
,
d
-transpeptidase activity of classical penicillin-binding proteins. Carbapenems inactivate these
l
,
d
-transpeptidases, and meropenem combined with clavulanic acid is bactericidal against extensively drug-resistant
M. tuberculosis
. Here, we used mass spectrometry and stopped-flow fluorimetry to investigate the kinetics and mechanisms of inactivation of the prototypic
M. tuberculosis
l
,
d
-transpeptidase Ldt
Mt1
by carbapenems (meropenem, doripenem, imipenem, and ertapenem) and cephalosporins (cefotaxime, cephalothin, and ceftriaxone). Inactivation proceeded through noncovalent drug binding and acylation of the catalytic Cys of Ldt
Mt1
, which was eventually followed by hydrolysis of the resulting acylenzyme. Meropenem rapidly inhibited Ldt
Mt1
, with a binding rate constant of 0.08 μM
−1
min
−1
. The enzyme was unable to recover from this initial binding step since the dissociation rate constant of the noncovalent complex was low (<0.1 min
−1
) in comparison to the acylation rate constant (3.1 min
−1
). The covalent adduct resulting from enzyme acylation was stable, with a hydrolysis rate constant of 1.0 × 10
−3
min
−1
. Variations in the carbapenem side chains affected both the binding and acylation steps, ertapenem being the most efficient Ldt
Mt1
inactivator. Cephalosporins also formed covalent adducts with Ldt
Mt1
, although the acylation reaction was 7- to 1,000-fold slower and led to elimination of one of the drug side chains. Comparison of kinetic constants for drug binding, acylation, and acylenzyme hydrolysis indicates that carbapenems and cephems can both be tailored to optimize peptidoglycan synthesis inhibition in
M. tuberculosis
.
Facultative Sterol Uptake in an Ergosterol-Deficient Clinical Isolate of Candida glabrata Harboring a Missense Mutation in
ERG11
and Exhibiting Cross-Resistance to Azoles and Amphotericin B
Authors: Hull Claire M. , Parker Josie E. , Bader Oliver , Weig Michael , Gross Uwe , Warrilow Andrew G. S. , Kelly Diane E. , Kelly Steven L. 
Published: May 2012

ABSTRACT

We identified a clinical isolate of
Candida glabrata
(CG156) exhibiting flocculent growth and cross-resistance to fluconazole (FLC), voriconazole (VRC), and amphotericin B (AMB), with MICs of >256, >256, and 32 μg ml
−1
, respectively. Sterol analysis using gas chromatography-mass spectrometry (GC-MS) revealed that CG156 was a sterol 14α-demethylase (Erg11p) mutant, wherein 14α-methylated intermediates (lanosterol was >80% of the total) were the only detectable sterols.
ERG11
sequencing indicated that CG156 harbored a single-amino-acid substitution (G315D) which nullified the function of native Erg11p. In heterologous expression studies using a doxycycline-regulatable
Saccharomyces cerevisiae erg11
strain, wild-type
C. glabrata
Erg11p fully complemented the function of
S. cerevisiae
sterol 14α-demethylase, restoring growth and ergosterol synthesis in recombinant yeast; mutated CG156 Erg11p did not. CG156 was culturable using sterol-free, glucose-containing yeast minimal medium (
glc
YM). However, when grown on sterol-supplemented
glc
YM (with ergosta 7,22-dienol, ergosterol, cholestanol, cholesterol, Δ
7
-cholestenol, or desmosterol), CG156 cultures exhibited shorter lag phases, reached higher cell densities, and showed alterations in cellular sterol composition. Unlike comparator isolates (harboring wild-type
ERG11
) that became less sensitive to FLC and VRC when cultured on sterol-supplemented
glc
YM, facultative sterol uptake by CG156 did not affect its azole-resistant phenotype. Conversely, CG156 grown using
glc
YM with ergosterol (or with ergosta 7,22-dienol) showed increased sensitivity to AMB; CG156 grown using
glc
YM with cholesterol (or with cholestanol) became more resistant (MICs of 2 and >64 μg AMB ml
−1
, respectively). Our results provide insights into the consequences of sterol uptake and metabolism on growth and antifungal resistance in
C. glabrata
.
An Integrated Approach for Identification and Target Validation of Antifungal Compounds Active against Erg11p
Authors: Hoepfner Dominic , Karkare Shantanu , Helliwell Stephen B. , Pfeifer Martin , Trunzer Markus , De Bonnechose Sophie , Zimmerlin Alfred , Tao Jianshi , Richie Daryl , Hofmann Andreas , Reinker Stefan , Frederiksen Mathias , Movva N. Rao , Porter Jeffrey A. , Ryder Neil S. , Parker Christian N. 
Published: May 2012

ABSTRACT

Systemic life-threatening fungal infections represent a significant unmet medical need. Cell-based, phenotypic screening can be an effective means of discovering potential novel antifungal compounds, but it does not address target identification, normally required for compound optimization by medicinal chemistry. Here, we demonstrate a combination of screening, genetic, and biochemical approaches to identify and characterize novel antifungal compounds. We isolated a set of novel non-azole antifungal compounds for which no target or mechanism of action is known, using a screen for inhibition of
Saccharomyces cerevisiae
proliferation. Haploinsufficiency profiling of these compounds in
S. cerevisiae
suggests that they target Erg11p, a cytochrome P450 family member, which is the target of azoles. Consistent with this, metabolic profiling in
S. cerevisiae
revealed a buildup of the metabolic intermediates prior to Erg11p activity, following compound treatment. Further, human cytochrome P450 is also inhibited in
in vitro
assays by these compounds. We modeled the Erg11p protein based on the human CYP51 crystal structure, and
in silico
docking of these compounds suggests that they interact with the heme center in a manner similar to that of azoles. Consistent with these docking observations,
Candida
strains carrying azole-resistant alleles of
ERG11
are also resistant to the compounds in this study. Thus, we have identified non-azole Erg11p inhibitors, using a systematic approach for ligand and target characterization.
Antitubercular Activity of Disulfiram, an Antialcoholism Drug, against Multidrug- and Extensively Drug-Resistant Mycobacterium tuberculosis Isolates
Authors: Horita Yasuhiro , Takii Takemasa , Yagi Tetsuya , Ogawa Kenji , Fujiwara Nagatoshi , Inagaki Emi , Kremer Laurent , Sato Yasuo , Kuroishi Ryuji , Lee YooSa , Makino Toshiaki , Mizukami Hajime , Hasegawa Tomohiro , Yamamoto Ryuji , Onozaki Kikuo 
Published: May 2012

ABSTRACTThe antimycobacterial activities of disulfiram (DSF) and diethyldithiocarbamate (DDC) against multidrug- and extensively drug-resistant tuberculosis (MDR/XDR-TB) clinical isolates were evaluatedin vitro. Both DSF and DDC exhibited potent antitubercular activities against 42 clinical isolates ofM. tuberculosis, including MDR/XDR-TB strains. Moreover, DSF showed remarkable bactericidal activityex vivoandin vivo. Therefore, DSF might be a drug repurposed for the treatment of MDR/XDR-TB.
Different Biosynthetic Pathways to Fosfomycin in Pseudomonas syringae and Streptomyces Species
Authors: Kim Seung Young , Ju Kou-San , Metcalf William W. , Evans Bradley S. , Kuzuyama Tomohisa , van der Donk Wilfred A. 
Published: May 2012

ABSTRACT

Fosfomycin is a wide-spectrum antibiotic that is used clinically to treat acute cystitis in the United States. The compound is produced by several strains of streptomycetes and pseudomonads. We sequenced the biosynthetic gene cluster responsible for fosfomycin production in
Pseudomonas syringae
PB-5123. Surprisingly, the biosynthetic pathway in this organism is very different from that in
Streptomyces fradiae
and
Streptomyces wedmorensis
. The pathways share the first and last steps, involving conversion of phosphoenolpyruvate to phosphonopyruvate (PnPy) and 2-hydroxypropylphosphonate (2-HPP) to fosfomycin, respectively, but the enzymes converting PnPy to 2-HPP are different. The genome of
P. syringae
PB-5123 lacks a gene encoding the PnPy decarboxylase found in the
Streptomyces
strains. Instead, it contains a gene coding for a citrate synthase-like enzyme, Psf2, homologous to the proteins that add an acetyl group to PnPy in the biosynthesis of FR-900098 and phosphinothricin. Heterologous expression and purification of Psf2 followed by activity assays confirmed the proposed activity of Psf2. Furthermore, heterologous production of fosfomycin in
Pseudomonas aeruginosa
from a fosmid encoding the fosfomycin biosynthetic cluster from
P. syringae
PB-5123 confirmed that the gene cluster is functional. Therefore, two different pathways have evolved to produce this highly potent antimicrobial agent.
Implication of Antigenic Conversion of Helicobacter pylori Lipopolysaccharides That Involve Interaction with Surfactant Protein D
Authors: Yokota Shin-ichi , Amano Ken-ichi , Nishitani Chiaki , Ariki Shigeru , Kuroki Yoshio , Fujii Nobuhiro 
Published: May 2012

ABSTRACT

We propose two antigenic types of
Helicobacter pylori
lipopolysaccharides (LPS): highly antigenic epitope-carrying LPS (HA-LPS) and weakly antigenic epitope-carrying LPS (WA-LPS) based on human serum reactivity. Strains carrying WA-LPS are highly prevalent in isolates from gastric cancer patients. WA-LPS exhibits more potent biological activities compared to HA-LPS, namely, upregulation of Toll-like receptor 4 (TLR4) expression and induction of enhanced epithelial cell proliferation. The results of competitive binding assays using monosaccharides and methylglycosides, as well as binding assays using glycosidase-treated LPS, suggested that β-linked
N
-acetyl-
d
-glucosamine and β-linked
d
-galactose residues largely contributed to the highly antigenic epitope and the weakly antigenic epitope, respectively. WA-LPS exhibited greater binding activity to surfactant protein D (SP-D) in a Ca
2+
-dependent manner, and this interaction was inhibited by methyl-β-
d
-galactoside. The biological activities of WA-LPS were markedly enhanced by the addition of SP-D. Lines of evidence suggested that removal of β-
N
-acetyl-
d
-glucosamine residue, which comprises the highly antigenic epitope, results in exposure of the weakly antigenic epitope. The weakly antigenic epitope interacted preferentially with SP-D, and SP-D enhanced the biological activity of WA-LPS.
Development of an Fc-Enhanced Anti–B7-H3 Monoclonal Antibody with Potent Antitumor Activity
Authors: Loo Deryk , Alderson Ralph F. , Chen Francine Z. , Huang Ling , Zhang Wenjun , Gorlatov Sergey , Burke Steve , Ciccarone Valentina , Li Hua , Yang Yinhua , Son Tom , Chen Yan , Easton Ann N. , Li Jonathan C. , Rillema Jill R. , Licea Monica , Fieger Claudia , Liang Tony W. , Mather Jennie P. , Koenig Scott , Stewart Stanford J. , Johnson Syd , Bonvini Ezio , Moore Paul A. 
Published: May 2012

Abstract
Purpose: The goal of this research was to harness a monoclonal antibody (mAb) discovery platform to identify cell-surface antigens highly expressed on cancer and develop, through Fc optimization, potent mAb therapies toward these tumor-specific antigens.
Experimental Design: Fifty independent mAbs targeting the cell-surface immunoregulatory B7-H3 protein were obtained through independent intact cell-based immunizations using human tissue progenitor cells, cancer cell lines, or cell lines displaying cancer stem cell properties. Binding studies revealed this natively reactive B7-H3 mAb panel to bind a range of independent B7-H3 epitopes. Immunohistochemical analyses showed that a subset displayed strong reactivity to a broad range of human cancers while exhibiting limited binding to normal human tissues. A B7-H3 mAb displaying exquisite tumor/normal differential binding was selected for humanization and incorporation of an Fc domain modified to enhance effector-mediated antitumor function via increased affinity for the activating receptor CD16A and decreased binding to the inhibitory receptor CD32B.
Results: MGA271, the resulting engineered anti–B7-H3 mAb, mediates potent antibody-dependent cellular cytotoxicity against a broad range of tumor cell types. Furthermore, in human CD16A-bearing transgenic mice, MGA271 exhibited potent antitumor activity in B7-H3–expressing xenograft models of renal cell and bladder carcinoma. Toxicology studies carried out in cynomolgus monkeys revealed no significant test article-related safety findings.
Conclusions: This data supports evaluation of MGA271 clinical utility in B7-H3–expressing cancer, while validating a combination of a nontarget biased approach of intact cell immunizations and immunohistochemistry to identify novel cancer antigens with Fc-based mAb engineering to enable potent antitumor activity. Clin Cancer Res; 18(14); 3834–45. ©2012 AACR.
Antiplatelet Drug Response Status Does Not Predict Recurrent Ischemic Events in Stable Cardiovascular Patients
Authors: Reny Jean-Luc , Berdagué Philippe , Poncet Antoine , Barazer Isabelle , Nolli Séverine , Fabbro-Peray Pascale , Schved Jean-François , Bounameaux Henri , Mach François , de Moerloose Philippe , Fontana Pierre 
Published: May 2012

Background—
The biological response to antiplatelet drugs has repeatedly been shown to predict the recurrence of major adverse cardiovascular events (MACEs). However, most studies involved coronary artery disease patients with recent vessel injury shortly after the initiation of antiplatelet therapy. Data on stable cardiovascular patients are scarce, and the added predictive value of specific assays (the vasodilator phosphoprotein assay for the clopidogrel response and serum thromboxane B2 for the aspirin response) and aggregation-based assays relative to common predictors has rarely been addressed.

Methods and Results—

Stable cardiovascular outpatients participating in the Antiplatelet Drug Resistances and Ischemic Events (ADRIE) study (n=771) were tested twice, at 2 separate visits, with specific and aggregation-based assays. Follow-up lasted 3 years, and <1% of patients were lost to follow-up. MACEs were adjudicated by an independent committee. Multivariate survival analyses included relevant variables identified in univariate analysis and platelet function test results. The C-index was used to express the prognostic value of various multivariate models. MACEs, the primary end point, occurred in 16% of patients. Hypertension, smoking, older age, and elevated low-density lipoprotein cholesterol were predictive of MACE recurrence, with a C-index of 0.63 (
P
<0.001). Neither the specific nor the aggregation-based assays added significant predictive value for the primary end point.

Conclusions—
Biological antiplatelet drug responsiveness, measured with specific or aggregation-based assays, has no incremental predictive value over common cardiovascular risk factors for MACE recurrence in stable cardiovascular outpatients. These results do not support platelet function testing for MACE risk evaluation in stable cardiovascular patients.

Clinical Trial Registration—

URL:
http://www.clinicaltrials.gov
. Unique identifier: NCT00501423.
The United States Registry for Fibromuscular Dysplasia
Authors: Olin Jeffrey W. , Froehlich James , Gu Xiaokui , Bacharach J. Michael , Eagle Kim , Gray Bruce H. , Jaff Michael R. , Kim Esther S.H. , Mace Pam , Matsumoto Alan H. , McBane Robert D. , Kline-Rogers Eva , White Christopher J. , Gornik Heather L. 
Published: May 2012

Background—
Fibromuscular dysplasia (FMD), a noninflammatory disease of medium-size arteries, may lead to stenosis, occlusion, dissection, and/or aneurysm. There has been little progress in understanding the epidemiology, pathogenesis, and outcomes since its first description in 1938.

Methods and Results—
Clinical features, presenting symptoms, and vascular events are reviewed for the first 447 patients enrolled in a national FMD registry from 9 US sites. Vascular beds were imaged selectively based on clinical presentation and local practice. The majority of patients were female (91%) with a mean age at diagnosis of 51.9 (SD 13.4 years; range, 5–83 years). Hypertension, headache, and pulsatile tinnitus were the most common presenting symptoms of the disease. Self-reported family history of stroke (53.5%), aneurysm (23.5%), and sudden death (19.8%) were common, but FMD in first- or second-degree relatives was reported only in 7.3%. FMD was identified in the renal artery in 294 patients, extracranial carotid arteries in 251 patients, and vertebral arteries in 82 patients. A past or presenting history of vascular events were common: 19.2% of patients had a transient ischemic attack or stroke, 19.7% had experienced arterial dissection(s), and 17% of patients had an aneurysm(s). The most frequent indications for therapy were hypertension, aneurysm, and dissection.

Conclusions—
In this registry, FMD occurred primarily in middle-aged women, although it presents across the lifespan. Cerebrovascular FMD occurred as frequently as renal FMD. Although a significant proportion of FMD patients may present with a serious vascular event, many present with nonspecific symptoms and a subsequent delay in diagnosis.