No abstract

We have evaluated the diagnostic utility of eleven Toxoplasma gondii recombinant antigens (P22 [SAG2], P24 [GRA1], P25, P28 [GRA2], P29 [GRA7], P30 [SAG1], P35, P41 [GRA4], P54 [ROP2], P66 [ROP1], and P68) in immunoglobulin G (IgG) and IgM recombinant enzyme-linked immunosorbent assays (Rec-ELISAs). Following an initial evaluation, six recombinant antigens (P29, P30, P35, P54, P66, and P68) were tested in the IgG and IgM Rec-ELISAs with four groups of samples which span the toxoplasmosis disease spectrum (negative, chronic infection, acute infection, and recent seroconversion). Our results suggest that the combination of P29, P30, and P35 in an IgG Rec-ELISA and the combination of P29, P35, and P66 in an IgM Rec-ELISA can replace the tachyzoite antigen in IgG and IgM serologic tests, respectively. The relative sensitivity, specificity, and agreement for the IgG P29-P30-P35 Rec-ELISA were 98.4, 95.7, and 97.2%, respectively. The resolved sensitivity, specificity, and agreement for the IgM P29-P35-P66 Rec-ELISA were 93.1, 95.0, and 94.5%, respectively. Relative to the tachyzoite-based immunocapture IgM assay, the IgM P29-P35-P66 Rec-ELISA detects fewer samples that contain IgG antibodies with elevated avidity from individuals with an acute toxoplasmosis.

Starting from a rapidly metabolized adamantane 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) inhibitor 22a, a series of E-5-hydroxy-2-adamantamine inhibitors, exemplified by 22d and (+/-)-22f, was discovered. Many of these compounds are potent inhibitors of 11beta-HSD1 and are selective over 11beta-HSD2 for multiple species (human, mouse, and rat), unlike other reported species-selective series. These compounds have good cellular potency and improved microsomal stability. Pharmacokinetic profiling in rodents indicated moderate to large volumes of distribution, short half-lives, and a pharmacokinetic species difference with the greatest exposure measured in rat with 22d. One hour postdose liver, adipose, and brain tissue 11beta-HSD1 inhibition was confirmed with (+/-)-22f in a murine ex vivo assay. Although 5,7-disubstitued-2-adamantamines provided greater stability, a single, E-5-position, polar functional group afforded inhibitors with the best combination of stability, potency, and selectivity. These results indicate that adamantane metabolic stabilization sufficient to obtain short-acting, potent, and selective 11beta-HSD1 inhibitors has been discovered.

We report the discovery and characterization of a novel ribosome inhibitor (NRI) class that exhibits selective and broad-spectrum antibacterial activity. Compounds in this class inhibit growth of many gram-positive and gram-negative bacteria, including the common respiratory pathogens Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, and Moraxella catarrhalis, and are nontoxic to human cell lines. The first NRI was discovered in a high-throughput screen designed to identify inhibitors of cell-free translation in extracts from S. pneumoniae. The chemical structure of the NRI class is related to antibacterial quinolones, but, interestingly, the differences in structure are sufficient to completely alter the biochemical and intracellular mechanisms of action. Expression array studies and analysis of NRI-resistant mutants confirm this difference in intracellular mechanism and provide evidence that the NRIs inhibit bacterial protein synthesis by inhibiting ribosomes. Furthermore, compounds in the NRI series appear to inhibit bacterial ribosomes by a new mechanism, because NRI-resistant strains are not cross-resistant to other ribosome inhibitors, such as macrolides, chloramphenicol, tetracycline, aminoglycosides, or oxazolidinones. The NRIs are a promising new antibacterial class with activity against all major drug-resistant respiratory pathogens.Respiratory tract infections are the number 1 killer worldwide, responsible for over 50 million deaths each year. Although antibacterial therapy has successfully stemmed the tide against infection since the middle of the last century, antibacterial resistance of Streptococcus pneumoniae, the most commonly identified pathogen associated with community-acquired pneumonia, is on the rise (2,4,8,22). A recent worldwide study documents that a significant fraction of S. pneumoniae isolates have reduced susceptibility to penicillin (36%) and macrolides (31%) (5). Although overall rates of resistance to fluoroquinolones are low, these rates were found to be increasing rapidly in Canada (1). In general, pathogenic bacteria continuously evolve mechanisms of resistance to currently used antibacterial agents. The discovery of novel antibacterial classes would be the most powerful way to generate new therapy against these resistant pathogens. Unfortunately, novel antibacterial classes have been difficult to discover, with the oxazolidinones, identified in 1980, being the last example to successfully reach the clinic.The bacterial ribosome is a proven target for antibacterial chemotherapy (6,17,20,21). Since the 1940s, small-molecule ribosome inhibitors such as chloramphenicol, tetracyclines, macrolides, aminoglycosides, and, more recently, oxazolidinones have been used to combat bacterial infections in humans (11). These diverse chemical classes of ribosome inhibitors each bind to a different site on the ribosome, which is not surprising given its large size and complexity. Accordingly, drug resistance to each class generally develops separately, such that resista...

The authors report the development of a high-throughput screen for inhibitors of Streptococcus pneumoniae transcription and translation (TT) using a luciferase reporter, and the secondary assays used to determine the biochemical spectrum of activity and bacterial specificity. More than 220,000 compounds were screened in mixtures of 10 compounds per well, with 10,000 picks selected for further study. False-positive hits from inhibition of luciferase activity were an extremely common artifact. After filtering luciferase inhibitors and several known classes of antibiotics, approximately 50 hits remained. These compounds were examined for their ability to inhibit Escherichia coli TT, uncoupled S. pneumoniae translation or transcription, rabbit reticulocyte translation, and in vitro toxicity in human and bacterial cells. One of these compounds had the desired profile of broad-spectrum biochemical activity in bacteria and selectivity versus mammalian biochemical and whole-cell assays.