Estrogen receptor beta-subtype selective tetrahydrofluorenones: use of a fused pyrazole as a phenol bioisostere.

Synthesis of a series of fused pyrazole tetrahydrofluorenone analogs which are potent, ERbeta subtype selective ligands is described. Analogs possessing subnanomolar ERbeta binding, greater than 100-fold ERbeta-selectivity, and oral bioavailability are reported.

The discovery of tetrahydrofluorenones as a new class of estrogen receptor beta-subtype selective ligands.

Synthesis and derivatization of a series of substituted tetrahydrofluorenone analogs giving potent, ERbeta subtype selective ligands are described. Several analogs possessing ERbeta binding affinities comparable to 17beta-estradiol but with greater than 75-fold selectivity over ERalpha are reported.

Succinic acids as potent inhibitors of plasmid-borne IMP-1 metallo-beta-lactamase.

IMP-1 metallo-beta-lactamase (class B) is a plasmid-borne zinc metalloenzyme that efficiently hydrolyzes beta-lactam antibiotics, including carbapenems, rendering them ineffective. Because IMP-1 has been found in several clinically important carbapenem-resistant pathogens, there is a need for inhibitors of this enzyme that could protect broad spectrum antibiotics such as imipenem from hydrolysis and thus extend their utility. We have identified a series of 2,3-(S,S)-disubstituted succinic acids that are potent inhibitors of IMP-1. Determination of high resolution crystal structures and molecular modeling of succinic acid inhibitor complexes with IMP-1 has allowed an understanding of the potency, stereochemistry, and structure-activity relationships of these inhibitors.

Dicationic 2-fluorenonylcarbapenems: potent anti-MRS agents with improved solubility and pharmacokinetic properties.

The synthesis and biological evaluation of a series of dicationic-substituted 2-fluorenonylcarbapenems is described. This class of compounds showed enhanced water solubility while maintaining potent activity against MRS. Introduction of a 1-beta-methyl substituent was found to improve pharmacokinetics.

Synthesis and SAR of thioester and thiol inhibitors of IMP-1 metallo-beta-lactamase.

Potent thioester and thiol inhibitors of IMP-1 metallo-beta-lactamase have been synthesized employing a solid-phase Mitsunobu reaction as the key step.

Synthesis and activity of 2-(sulfonamido)methyl-carbapenems: discovery of a novel, anti-MRSA 1,8-naphthosultam pharmacophore.

A series of 1beta-methyl carbapenems substituted at the 2-position with lipophilic, acyclic and cyclic (sulfonamido)methyl groups was prepared and evaluated for activity against resistant gram-positive bacteria. From these studies, the 1,8-naphthosultamyl group emerged as a novel, PBP2a-binding, anti-MRSA pharmacophore worthy of further exploration.

Synthesis and properties of 2-(naphthosultamyl)methyl-carbapenems with potent anti-MRSA activity: discovery of L-786,392.

A series of 1beta-methyl-2-(naphthosultamyl)methyl-carbapenems bearing dicationic groups on the naphthosultamyl moiety was prepared and evaluated for activity against resistant gram-positive bacteria. Based on a combination of excellent in vitro antibacterial activity, acceptable mouse acute toxicity, and a desirable fragmentation pattern on beta-lactam ring opening, the analog 2g (L-786,392) was selected for extended evaluation.

Reduced immunotoxicity and preservation of antibacterial activity in a releasable side-chain carbapenem antibiotic.

A carbapenem antibiotic, L-786,392, was designed so that the side chain that provides high-affinity binding to the penicillin-binding proteins responsible for bacterial resistance was also the structural basis for ameliorating immunopathology. Expulsion of the side chain upon opening of the beta-lactam ring retained antibacterial activity while safely expelling the immunodominant epitope. L-786,392 was well tolerated in animal safety studies and had significant in vitro and in vivo activities against methicillin- and vancomycin-resistant Staphylococci and vancomycin-resistant Enterococci.

Antibiotic use in the Australian community, 1990-1995.

OBJECTIVE: To determine the pattern of antibiotic use in the Australian community, 1990-1995, and compare it with the pattern in other developed countries. DESIGN: Survey of data from the national database on drugs dispensed in Australia (1990-1995), an international database on retail drug sales (1985-1994), and Australian prescriber surveys (1994, 1995). MAIN OUTCOME MEASURES: National and international retail sales of oral antibiotics (defined daily doses [DDDs]/1000 population/day) and antibiotic prescriptions dispensed through community pharmacies by drug type; antibiotic prescribing profiles for common conditions. RESULTS: Antibiotic use in Australia remained steady between 1990 and 1995, with an estimated 24.7 DDDs/1000 population/day dispensed through community pharmacies in 1990 and 24.8 DDDs/1000 population/day in 1995. Amoxycillin, although declining in use, remained the most dispensed antibiotic. Compared with the other countries surveyed, Australia had the highest percentage use of tetracyclines, such as doxycycline, and the lowest percentage use of fluoroquinolones. Use of trimethoprim-sulfamethoxazole and flucloxacillin declined in Australia. In new cases of upper respiratory tract infection or pharyngitis, an antibiotic prescription was recorded for 57% of urban patient encounters and 73% of rural patient encounters. CONCLUSIONS: Antibiotic use in Australia is high, as in many other developed countries, but did not increase between 1990 and 1995. The overall profile of antibiotic use in Australia by drug class was similar to that in the United Kingdom. Antibiotics were still commonly prescribed for upper respiratory tract infection (which is usually viral), more commonly by rural than by urban general practitioners.

Scytalidium dimidiatum in Australia.

Six patients with Scytalidium dimidiatum (Hendersonula toruloidea) infections of their skin and nails are presented. Two of these patients originated in tropical areas where the fungus is endemic and four were Australian born. Three of the Australian-born patients had no history of travel to areas where the fungus is endemic. It appears that S. dimidiatum is present in Australia.

Antibiotic resistance among respiratory pathogens in preschool children.

OBJECTIVE: To determine the prevalence of antibiotic resistance among common respiratory pathogens circulating in the community. DESIGN: Survey of common respiratory pathogens isolated from nasal discharges. SETTING: 117 childcare centres and kindergartens in metropolitan Melbourne between May and July 1991-1993 and 42 from sociodemographically matching suburbs in Sydney between May and July, 1993. SUBJECTS: Children aged six years and under with nasal discharge. OUTCOME MEASURES: Resistance to penicillins, erythromycin and tetracycline among isolates of Haemophilus influenzae, Streptococcus pneumoniae, Moraxella (Branhamella) catarrhalis and Staphylococcus aureus. RESULTS: A total of 2286 nasal discharge swabs were collected. Amoxycillin resistance was detected in 99 of 711 isolates of H. influenzae (13.9%) and penicillin resistance in 781 of 834 isolates of M. catarrhalis (93.6%), 342 of 375 isolates of S. aureus (91.2%), and 30 of 781 isolates of S. pneumoniae (3.9%). Of 86 strains of H. influenzae type b isolated, 20 (23.3%) produced beta-lactamase. Penicillin resistance tended to become more common among isolates of H. influenzae and S. pneumoniae during the three-year period. CONCLUSION: Antibiotic resistance, mediated by beta-lactamase or altered penicillin-binding proteins, among respiratory pathogens carried by preschool children was significant and possibly increasing. This highlights the impact of prescribed antibiotics in the community and the folly of prescribing the limited store of antibiotics for viral infections.

Preparation and structure-activity relationships of simplified analogues of the antifungal agent cilofungin: a total synthesis approach.

The echinocandins are a well-known class of lipopeptides characterized by their potent antifungal activity against Candida species. The mechanism of action of the echinocandins is generally thought to be the inhibition of beta-1,3-glucan synthesis, an important structural component in the cell wall of Candida species. Extensive structure-activity studies on the fatty acid side chain of echinocandin B (1) led to the preparation of the clinical candidate cilofungin (4). However, little is known about the cyclic peptide. We now report the preparation, by solid-phase synthesis, of a series of simplified analogs of cilofungin in which the unusual amino acids found in the echinocandins were replaced with more readily accessible natural amino acids. The solid-phase approach to the total synthesis of these analogs allowed us to conveniently explore structural modifications that could not be accomplished by chemical modification of the natural product. The simplest analog 5 showed no biological activity. Structural complexity was then returned to the system in a systematic fashion so as to reapproach the original cilofungin structure. Antifungal activity and the inhibition of beta-1,3-glucan synthesis were monitored at each step of the process, thereby revealing the basic structure-activity relationships of the amino acids and the minimal structural requirements for biological activity in the echinocandin ring system. The results suggests that the 3-hydroxy-4-methylproline residue enhances activity but the L-homotyrosine residue is crucial for both antifungal activity and the inhibition of beta-1,3-glucan synthesis.

Sequences mediating the translation of mouse S16 ribosomal protein mRNA during myoblast differentiation and in vitro and possible control points for the in vitro translation.

The translation of ribosomal protein (r-protein) mRNAs is generally inefficient and regulated during the differentiation of mouse myoblasts into fibers. In this discussion we show that the first 31 nucleotides of the S16 r-protein mRNA, when located at the 5' end of the mRNA, are sufficient to impart the translational properties of an r-protein mRNA to the SV-GALK mRNA, which is normally translated efficiently in both myoblasts and fibers. If the same S16 sequences are located within the interior of the 5'-untranslated region of the SV-GALK mRNA, however, they do not impart the translational properties of an r-protein mRNA to the SV-GALK mRNA. The translation of mouse r-protein mRNAs was examined in vitro to help elucidate the mechanisms controlling their translation. Mouse r-protein mRNAs are inefficiently translated in rabbit reticulocyte extracts, and the same sequences that mediate their inefficient and regulated translation during myoblast differentiation also mediate their inefficient translation in a position-dependent manner in reticulocyte extracts. To determine whether the subpolysomal r-protein mRNAs that are not actively translated in vivo are capable of translation, subpolysomal RNA was translated in reticulocyte extracts. The subpolysomal r-protein mRNAs are just as capable of translation as are polysomal mRNAs. To help identify the initiation factors and/or the steps in the initiation pathway that mediate the inefficient translation of r-protein mRNAs, reticulocyte extracts were supplemented with purified initiation factors. Only eIF-4F, the cap-binding complex, and eIF-3, which is involved in subunit dissociation and interacts with eIF-4F during initiation, stimulated the translation of r-protein mRNA. These experiments, along with m7GDP inhibition studies, suggest that eIF-4F and/or eIF-3, or the steps mediated by these factors, mediate the inefficient translation in reticulocyte extracts and raise the possibility that these steps also control the regulated translation of r-protein mRNAs during myoblast differentiation.

Antioxidant-based inhibitors of leukotriene biosynthesis. The discovery of 6-[1-[2-(hydroxymethyl)phenyl]-1-propen-3-yl]-2,3-dihydro-5- benzofuranol, a potent topical antiinflammatory agent.

The leukotrienes, metabolites of arachidonic acid produced through the action of the enzyme 5-lipoxygenase, are important mediators of immediate hypersensitivity and inflammation. Among the variety of diseases in which the leukotrienes may play a symptomatic or causative role is the dermatological condition psoriasis, a chronic proliferative disease of the skin. This study reports the synthesis and comparative biological activities of various ortho-substituted phenols including 4-methoxyphenols, 6-hydroxy-1,2,3,4-tetrahydrobenzopyrans, 2,3-dihydro-5-benzofuranols, and 5-benzofuranols. The phenols prepared in this study were evaluated for their ability to inhibit the production of leukotriene B4(LTB4) in isolated human polymorphonuclear leukocytes (PMNs) and to inhibit a topical inflammatory response in the topical mouse ear (TME) model. In the former case, when the log IC50 was plotted versus the log of the octanol/water partition coefficient (log P), to eliminate the effect of lipophilicity, the 2,3-dihydro-5-benzofuranol ring system was shown to be more potent than the other ring systems examined throughout the range of partition coefficients studied. The ability to inhibit leukotriene production in vitro in human PMNs can be rationalized on the basis of a model that suggests that the observed inhibition is dependent on the kinetic ability of the inhibitor to reduce a radical species and on the fraction of inhibitor that is partitioned into the cell membrane. While the in vivo antiinflammatory activity as measured by the TME did not correlate with the in vitro data, it was felt that the TME represented a valuable measure of the ability of a compound to penetrate the skin to the site of an ongoing inflammatory response. Of the compounds synthesized in this study, 6-[1-[2-(hydroxymethyl)phenyl]-1-propen-3-yl]-2,3-dihydro-5-benzof uranol (1, L-651896) was chosen for further development.

2,3-Dihydro-5-benzofuranols as antioxidant-based inhibitors of leukotriene biosynthesis.

The enzymes that catalyze the oxidative metabolism of arachidonic acid have provided fertile ground for the development of useful therapeutic agents for nearly a quarter century. Inhibitors of the enzyme cyclooxygenase prevent the formation of the prostaglandins and thromboxanes and are clinically useful antiinflammatories and peripheral analgesics. More recently it has been discovered that the enzyme 5-lipoxygenase is the first step in the formation of a series of biologically important metabolites of arachidonic acid, the leukotrienes. Evidence suggests that an inhibitor of 5-lipoxygenase may be a useful therapeutic agent in the treatment of asthma, immediate hypersensitivity, and inflammation. Various antioxidants have been examined as inhibitors of 5-lipoxygenase in vitro. We were intrigued by recent reports that the 2,3-dihydro-5-benzofuranol ring system maximizes the stereoelectronic effects necessary for efficient hydrogen atom abstraction by peroxyl radicals. In this study we describe the synthesis of over 50 new 2,3-dihydro-5-benzofuranols and their biological evaluation as inhibitors of leukotriene biosynthesis in isolated human polymorphonuclear leukocytes. We show that the 2,3-dihydro-5-benzofuranol ring system, although not a potent inhibitor of leukotriene biosynthesis in itself, can provide a useful template for the design of antioxidant-based inhibitors of leukotriene biosynthesis. Furthermore, within a structural class the potency of a given analogue can be predicted on the basis of its overall calculated lipophilicity (log P). The data are interpreted in terms of a model in which the observed inhibition by this class of inhibitors is dependent on the intrinsic ability of the antioxidant to reduce the enzyme and on the fraction of the inhibitor that is partitioned into the membrane.

Insulin stimulates the translation of ribosomal proteins and the transcription of rDNA in mouse myoblasts.

Insulin stimulates the translation of ribosomal protein (r-protein) mRNAs and the transcription of rDNA in mouse MM14DZ myoblasts. Analysis of the distribution of S16, L18, and L32 r-protein mRNAs in polysome gradients indicates that the increased translation of these mRNAs in insulin-treated myoblasts is due to the recruitment of mRNAs that were not previously being translated. In contrast, the translational efficiencies of beta-actin, c-myc, and p31 mRNAs are not affected by insulin. Hybridization analysis of RNA transcribed in nuclear run-on reactions indicates that insulin also stimulates the transcription of rDNA. Both the increases in r-protein translation and rDNA transcription occur coordinately and are maximal within 15 min of insulin treatment of myoblasts. However, insulin has no effect on the rate of cell division or the steady state levels of r-protein mRNAs. Surprisingly, after myoblasts differentiate into fibers, insulin does not affect the r-protein mRNA translation or rDNA transcription. These experiments indicate that the synthesis of the macromolecular components of ribosomes is tightly and coordinately controlled in myoblasts.

Effective protection against influenza after vaccination with subunit vaccine.

Immunization with an influenza subunit vaccine given in one dose about 1 month to 2 months before onset of an epidemic of influenza afforded from 80% to 90% protection in a double-blind clinical trial which was supported by isolation of virus and serological studies. In the vaccinated group, 20% failed to develop antibodies to the vaccine. Either serology tests or attempted isolation of virus alone would have failed to detect some of the cases.