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Antimicrobial Agents and Chemotherapy, May 2008, p. 1653-1662, Vol. 52, No. 5
0066-4804/08/$08.00+0     doi:10.1128/AAC.01383-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

In Vitro Activities of the Rx-01 Oxazolidinones against Hospital and Community Pathogens

Laura Lawrence, Paul Danese, Joe DeVito, Francois Franceschi, and Joyce Sutcliffe^*

Rib-X Pharmaceuticals, Inc., 300 George Street, New Haven, Connecticut 06511

Received 25 October 2007/ Returned for modification 26 November 2007/ Accepted 22 February 2008

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Rx-01_423 and Rx-01_667 are two members of the family of oxazolidinones that were designed using a combination of computational and medicinal chemistry and conventional biological techniques. The compounds have a two- to eightfold-improved potency over linezolid against serious gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA), multidrug-resistant streptococci, and vancomycin-resistant enterococci. This enhanced potency extends to the coverage of linezolid-resistant gram-positive microbes, especially multidrug-resistant enterococci and pneumococci. Compounds from this series expand the spectrum compared with linezolid to include fastidious gram-negative organisms like Haemophilus influenzae and Moraxella catarrhalis. Like linezolid, the Rx-01 compounds are bacteriostatic against MRSA and enterococci but are generally bactericidal against S. pneumoniae and H. influenzae.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
The frequency of antimicrobial-resistant microbes isolated in hospital and community settings is increasing worldwide (6, 25, 29, 36, 45, 50, 52, 57). Staphylococcus aureus and coagulase-negative staphylococci (CNS) ranked as the top two most frequently isolated pathogens associated with bloodstream infections or found in intensive care units (ICUs) in the United States or North America, respectively, from 2000 to 2002 (25, 29). Methicillin-resistant S. aureus (MRSA) accounted for 59.2%, 55%, and 47.9% of the strains from non-ICU inpatients, ICU patients, and outpatients, respectively, in the United States (50). Multidrug resistance phenotypes (resistance to at least three non-β-lactams) were common among cases of both inpatient MRSA (59.9%) and outpatient MRSA (40.8%) infections (50). Enterococcus faecalis and Enterococcus faecium were the third and sixth most common bloodstream isolates in the United States in 2002, collectively accounting for 10% of bloodstream infections. The majority of E. faecium strains isolated in ICUs in the United States are vancomycin resistant (60 to 76.3%) (25, 57). The need for new therapies that are effective against these resistance organisms would be welcomed by the medical and regulatory communities, especially if the therapy allowed an intravenous-to-oral switch to accommodate the growing number of patients completing their therapies as outpatients.

Community-acquired pneumonia (CAP) is the leading cause of death by infectious disease (2). The frequency of penicillin-resistant pneumococci in the United States was 34.2%, with 21.5% high-level resistance from 1999 to 2000 (13), and in 1998, fluoroquinolone resistance was seen for the first time (41). Macrolide resistance by target modification (Erm methylase or ribosomal mutations) and/or by Mef(A)-mediated efflux was 27.9% in U.S. isolates obtained from 2001 to 2002 (9, 15). Recent work found that the percentage of pneumococcal strains that were resistant to both penicillin and erythromycin is increasing faster than the percentage of strains singly resistant to either antibiotic and that there was an uncommon increase in Streptococcus pneumoniae strains carrying both erm(B) and mef(A) resistance mechanisms, especially in children 14 years of age (8, 9, 16, 34). Thus, commonly prescribed anti-infectives such as β-lactams and the currently used macrolides are no longer reliably effective against multidrug-resistant Streptococcus pneumoniae, the most important bacterial pathogen in respiratory tract infections (RTIs) in children and adults (24).

In addition to pneumococci, two gram-negative microbes, Haemophilus influenzae and Moraxella catarrhalis, are frequently isolated as the causative agents of RTIs, especially in cases of sinusitis and recurrent cases of otitis media. H. influenzae is also the organism most frequently isolated from patients with acute exacerbation of chronic bronchitis (35%) and the second most frequently isolated organism in cases of pneumonia and other RTIs (37). Similarly, M. catarrhalis was most commonly isolated from the nasopharynx of patients with sinusitis (29%) and is considered to be the third most frequently isolated pathogen in cases of RTIs (37). β-Lactamase-mediated resistance to ampicillin in H. influenzae isolates ranged from 15% in New England to 32% in the East South Central region of the United States (37). Notably, clarithromycin-resistant isolates of H. influenzae were observed in cases of sinusitis (36%) and RTIs worldwide (48). In a recent U.S. survey, penicillin-resistant M. catarrhalis isolates were found at a frequency of 91.5%, and sinusitis isolates displayed an erythromycin resistance frequency of 15% (48).

Pneumonia can also be caused by intracellular pathogens including Mycoplasma pneumoniae, Legionella pneumophila, and Chlamydophila pneumoniae (32, 44). More recently, cases of necrotizing pneumonia have been attributed to community-acquired MRSA (19). Although many of the community-acquired MRSA isolates are susceptible to trimethoprim-sulfamethoxazole or clindamycin, the susceptibilities to these drugs are not expected to last as their use increases. Thus, an empirical oral therapy that could treat these important gram-positive and gram-negative RTI pathogens is needed.

We designed a novel class of antibiotics that target the ribosome, a validated target for many marketed antibiotics and highlighted by nature, as many species use ribosome-targeted secondary metabolites to compete in nutrient-sparse environments. The starting point for the design of the Rx-01 family was the atomic resolution structures of linezolid and sparsomycin antibiotics complexed to the large ribosomal subunit (22) (Fig. 1). Analyses of the structures together with proprietary computational tools provided details of the interactions and ways to enhance or modify those interactions, allowing for a >100-fold increase in binding affinity and a significant circumvention of resistance mechanisms (22, 47, 56, 60). In addition, we used computational models to expand the spectrum of the oxazolidinone Rx-01 series to fastidious gram-negative microbes responsible for RTIs (56). In this paper, we examine the microbiological activities of several compounds from this effort.

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FIG. 1. Structures of linezolid and Rx-01 oxazolidinones.

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Bacterial strains. Robert Moellering, Jr. (Beth Israel Deaconess Medical Center, Boston, MA), provided staphylococcal, enterococcal, and pneumococcal isolates, including linezolid-resistant isolates. The isogenic pair of S. aureus isolates containing the erm(B)-cfr gene cluster designated the mlr operon (53) was obtained from Alexander Mankin (University of Illinois, Chicago, IL). A clinical isolate of S. pneumoniae that contains a two-amino-acid deletion in ribosomal protein L4 conferring resistance to macrolides, linezolid, and chloramphenicol (17, 58) was obtained from David Farrell (GR Micro Ltd., London, United Kingdom). Barry Kreiswirth (Public Health Institute, Newark, NJ) provided MRSA isolates with known clonal phenotypes as distinguished by spa lineage (46). Recently isolated U.S. isolates from cases of outpatient skin and soft tissue infections (2005 to 2006) were obtained from Eurofins Medinet (Herndon, VA). Other strain sets of S. pneumoniae or Streptococcus pyogenes with a defined macrolide-resistant mechanism(s) were gifts of David Farrell (GR Micro Ltd., London, United Kingdom). Another set of S. pyogenes strains with distinct emm types was a gift of Debra Bessen (New York Medical College, Valhalla, NY). Bacterial isolates and their relevant phenotypes used in studies to determine frequencies of resistance are listed in Table 1.

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TABLE 1. Bacterial isolates for resistance studies

Antibiotics and susceptibility tests. Rib-X (Rx) compounds and linezolid were synthesized at Rib-X Pharmaceuticals, Inc. The procedure for the synthesis of linezolid was described previously (60). Reference standards were purchased from either Alembic Ltd., Gujarat, India (azithromycin), or Sigma-Aldrich Corp. (St. Louis, MO) (penicillin G potassium salt, levofloxacin, and amoxicillin-clavulanate [Augmentin]). Telithromycin (Ketek) was purified at Rib-X from 800-mg tablets (NDC 0088-2225-41).

Susceptibility testing for bacteria that grow aerobically. For susceptibility testing, all control compounds (except levofloxacin) and Rx compounds were dissolved in 100% dimethyl sulfoxide (DMSO) at 12.8 mg/ml and serially diluted twofold in 100% DMSO to 0.025 mg/ml in 96-well plates (100x stock wells). Levofloxacin was prepared at 12.8 mg/ml in water containing 0.01 M NaOH. A volume of the 100x compound stock of each serial dilution (or 100% DMSO that served as a growth control) was used to make a 10-fold dilution in cation-adjusted Mueller-Hinton broth (CAMHB) (catalog number BBL#212322; Becton Dickinson and Company Diagnostics, Cockeysville, MD). Ten microliters of each 10-fold dilution was added to 90 µl of a 1:200 dilution of each organism at a 0.5 McFarland standard for a final drug concentration range of 128 µg/ml to 0.25 µg/ml of compound. The final concentration of DMSO in CAMHB was 1%, with each well containing an inoculum of 2 x 10⁵ to 7 x 10⁵ CFU/ml, as recommended by the Clinical and Laboratory Standards Institute (CLSI) (10). An aliquot of the growth control from each organism was diluted and streaked onto solid medium (blood or chocolate agar) to quantify inoculum size and to ensure a lack of contamination. Organisms were frozen as stocks using the Microbank bacterial preservation system (Pro-Lab Diagnostics, Austin, TX) and were streaked and passaged onto blood or chocolate agar plates (Hardy Diagnostics, Santa Maria, CA) at least two times to ensure sterility and viability prior to testing. For S. pneumoniae, CAMHB was supplemented with 5% lysed horse blood (Remel, Inc., Lenexa, KS) prepared according to CLSI methods (10); H. influenzae Haemophilus test medium was supplemented with hematin (catalog number H3281; Sigma-Aldrich Corp., St. Louis, MO), NAD (catalog number 43410l; Fluka Chemical Corp., Milwaukee, WI), and yeast extract (catalog number 212750; Becton Dickinson and Company Diagnostics, Cockeysville, MD) according to CLSI guidelines. Control strains (S. pneumoniae ATCC 49619, H. influenzae ATCC 49247, S. aureus ATCC 29213, and E. faecalis ATCC 29212) and performance standards for testing were described previously (10). MICs were determined as the lowest concentration without visible growth following incubation for 20 to 22 h at 35°C.

For the strain set of MRSA isolates with defined clonal phenotypes, testing was done in the laboratory of Barry Kreiswirth (Public Health Research Institute, Newark, NJ) using CLSI methodologies.

Susceptibility testing of C. pneumoniae, L. pneumophila, M. pneumoniae, Mycoplasma hominis, Ureaplasma urealyticum, and Chlamydia trachomatis. Susceptibility testing of M. pneumoniae was based on a broth microdilution method (55) as described previously by Critchley et al. (11). MICs for C. trachomatis and C. pneumoniae grown in McCoy cell monolayers to antimicrobial agents were determined in 96-well flat-bottom microtiter tissue culture plates. An MIC was defined as the lowest concentration of antimicrobial agent that completely inhibited the formation of normal inclusions, which were detected by the immunofluorescence staining techniques described previously by Fenelon et al. and Ridgway et al. (18, 40). Susceptibility tests with L. pneumophila were conducted by broth microdilution according to CLSI guidelines for aerobic bacteria (10) by using 96-well microtiter plates containing concentrations of test antimicrobials (14). The MIC was read as the first well showing no visible growth at 48 h. MIC studies on all atypical microbes (C. pneumoniae, L. pneumophila, M. pneumoniae, M. hominis, C. trachomatis, and U. urealyticum) were performed by GR Micro Ltd., London, United Kingdom, for Rib-X Pharmaceuticals, Inc.

Time-kill methodology. Time-kill studies were performed using 125-ml culture flasks containing 25 ml of CAMHB (or supplemented for susceptibility testing as described above) with approximately 5 x 10⁵ CFU/ml bacteria. Concentrations of four and eight times the predetermined MIC were added to each flask except for the growth control. The number of viable cells (CFU/ml) was quantified at 1, 2, 4, 6, 8, and 24 h after antibiotic addition via serial dilution into sterile saline and plating of 0.1 ml of serial dilutions onto either blood or chocolate agar. Plates were incubated for 20 to 24 h at 35°C.

Determination of spontaneous resistance frequencies (single-step selection). Mutant selection with the strains listed in Table 1 was performed essentially as described previously (49). Briefly, single colonies from a culture incubated overnight on blood or chocolate agar plates were used to inoculate 4 ml of CAMHB or Haemophilus test medium and grown for 16 h at 35°C on a roller drum. Cells were subcultured by dilution to an optical density at 600 nm of 0.05 in fresh medium and grown (with shaking) to late logarithmic phase. Cells were concentrated by centrifugation at room temperature and resuspended in fresh medium, and aliquots were spread onto nonselective agar plates and selective agar plates with compounds at 1x, 2x, 4x, and 8x the predetermined MIC and incubated at 37°C for 48 h.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Antibiotic susceptibilities to gram-positive hospital pathogens. Tables 2 and 3 provide the MIC range and MIC₅₀ and MIC₉₀ values of Rx-01_423 and Rx-01_667 against isolates of S. pneumoniae, methicillin-susceptible S. aureus (MSSA), MRSA, CNS, and vancomycin-susceptible and -resistant enterococci. Both Rx-01_423 and Rx-02_667 were eightfold more potent against this collection of pneumococci than linezolid.

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TABLE 2. Antibacterial activities against nosocomial gram-positive isolates of Streptococcus and Staphylococcus spp.

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TABLE 3. Antibacterial activities against nosocomial gram-positive isolates of enterococci

MIC₉₀ values for Rx-01_423 and Rx-01_667 against MSSA and CNS (including 19/25 oxacillin-resistant isolates) and enterococci (Tables 2 and 3) were within twofold of each other. Rx-01_423 was fourfold more active than linezolid against MSSA and CNS and eightfold more active than linezolid against all enterococci except vancomycin-resistant E. faecium. Rx-01_667 was two- to fourfold more active against MSSA and CNS and 4- to 16-fold more active against enterococci than linezolid. Rx-01_667 was the most active compound tested against both vancomycin-resistant and -susceptible E. faecalis and E. faecium isolates, with MIC₉₀ values of 0.5 to 1 µg/ml. All groups of enterococcal and staphylococcal isolates were resistant to azithromycin and the related ketolide antibiotic telithromycin, as defined by their respective MIC₉₀ results; only the MSSA isolates were susceptible to telithromycin. All staphylococcal and enterococcal isolates except MSSA isolates had MIC₉₀ values indicating resistance to levofloxacin. Quinupristin-dalfopristin (Synercid) was active against all staphylococcal and enterococcal isolates except E. faecalis. The MIC₉₀ values of oxacillin against all groups of enterococci and staphylococci indicated resistance, with MSSA being the exception. All isolates were susceptible to vancomycin except the enterococcal groups designated vancomycin resistant. Both species of vancomycin-resistant enterococci were resistant to gentamicin and ampicillin.

The data for MRSA isolates reported in Table 2 come from a strain set obtained from Robert Moellering, Jr. (Beth Israel Deaconess Medical School, Boston, MA), and consist of 31 isolates isolated in 2003 to 2004 from a single hospital. The MIC₉₀ values of both Rx compounds against this strain set were 4 µg/ml. The MIC₅₀ value as well as the distribution of the MICs indicated that both Rx compounds had better potency than linezolid across the strain set (Table 2 and Fig. 2A). The Rx compounds had MICs equally distributed over 0.5-, 1-, 2-, and 4-µg/ml values, whereas the MIC distribution for linezolid was heavily weighted at 2 µg/ml (n = 21), followed by 4 µg/ml (n = 8).

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FIG. 2. Distributions of MICs of Rx-01 compounds and linezolid against different groups of MRSA isolates. (A) Hospital-acquired MRSA isolates from 2003 to 2004. (B) Clonal MRSA isolates from 1960 to 2001. (C) Outpatient MRSA isolates from 2005 to 2006. White bars, Rx-01_423; black bars, linezolid; striped bars, Rx-01_667.

To ensure that the Rx compounds were active against MRSA isolates with distinct clonal lineages (as determined by spa typing) (46), the compound was evaluated against 56 clones in the laboratory of Barry Kreiswirth (Public Health Research Institute, Newark, NJ). There were 83 isolates in the data set, with some spa phenotypes being represented more than once; isolation dates spanned from 1960 to 2001. In this study, Rx-01_667 was twofold more potent in MIC₅₀ and MIC₉₀ values than linezolid, whereas Rx-01_423 was equivalent to linezolid (MIC₅₀ and MIC₉₀ of 2 and 4 µg/ml, respectively) (Table 4). There is a distinct difference in the distributions of MICs of these three compounds against this panel of MRSA isolates (Fig. 2B), with Rx-01_667, Rx-01_423, and linezolid possessing MICs of 1 µg/ml against 56.6%, 36.1%, and 16.9% of the isolates, respectively. The MIC₅₀ values indicate that >50% of these isolates were resistant to levofloxacin and erythromycin. Twenty-four MSSA isolates representing 19 distinct clones were also evaluated; Rx-01_667 and Rx-01_423 had fourfold and twofold advantages over linezolid (see MIC₉₀ values in Table 4).

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TABLE 4. MICs of Rx-02_423 and Rx-01_667 against MRSA and MSSA isolates

The third set of MRSA isolates used to evaluate the Rx compounds was from 33 cases of outpatient skin and soft tissue infections (emergency rooms and walk-in clinics, etc.) in the United States during 2005 and 2006. Rx-01_667 and Rx-01_423 possessed MICs of 0.5 to 2 µg/ml against 91% and 94% of these isolates, respectively, whereas only 21% of the isolates were susceptible to linezolid at 2 µg/ml (Fig. 2C and Table 4). More than 50% of the MRSA isolates were resistant to erythromycin or levofloxacin, and all strains were resistant to oxacillin (Table 4).

Antibiotic susceptibilities to respiratory tract pathogens. The Rx compounds were tested for activities against common respiratory tract isolates, including S. pneumoniae, S. pyogenes, H. influenzae, and M. catarrhalis (Tables 5 and 6). Rx-01_423 and Rx-01_667 were the most active antimicrobial agents against S. pneumoniae, according to MIC₉₀ values. The activities of the two Rx compounds were not affected by penicillin or macrolide resistance mediated by either target mutations or modifications. MIC₉₀ values of penicillin G against S. pneumoniae ranged from 4 to 8 µg/ml, and for every set of pneumococcal isolates, except the clinical isolates with defined ribosomal mutations (17), the MIC₅₀ values indicate that at least 50% of the isolates were highly resistant to penicillin (MIC 1 µg/ml). The Rx compounds were at least eightfold more active than linezolid against all streptococci including S. pyogenes. Telithromycin and azithromycin were more active than the Rx compounds against macrolide-susceptible S. pyogenes isolates, but the Rx compounds enjoyed a four- to eightfold advantage in MIC over telithromycin and a 64- to 128-fold advantage over azithromycin when susceptibility to macrolide-resistant [mef(A)] group A streptococci was evaluated. Clindamycin susceptibility to known mef- or erm-containing strains was as expected for the defined strain collections.

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TABLE 5. Antibacterial activities against S. pneumoniae respiratory tract pathogens

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TABLE 6. Antibacterial activities against S. pyogenes, M. catarrhalis, and H. influenzae respiratory tract pathogens

Rx-01_423 and Rx-01_667 had MIC₉₀s of 2 and 4 µg/ml and 0.5 and 1 µg/ml against the gram-negative respiratory isolates M. catarrhalis and H. influenzae, respectively. Rx-01_423 was eightfold and fourfold more active than linezolid, respectively, whereas Rx-01_667 was 32-fold and 16-fold more active than linezolid, respectively. All isolates were susceptible to levofloxacin, but the H. influenzae MIC₉₀ value indicated that at least 10% of the isolates were resistant to amoxicillin-clavulanate.

Antibiotic susceptibilities to atypical respiratory tract pathogens. Since atypical bacteria are important causative pathogens for community respiratory tract infections, the Rx-01 oxazolidinones and control compounds were evaluated against clinical isolates of M. pneumoniae, L. pneumophila, C. pneumoniae, C. trachomatis, U. urealyticum, and M. hominis (Table 7). Rx-01_667 is 16-fold more potent than linezolid against M. pneumoniae and C. trachomatis and 2-fold more potent against M. hominis and U. urealyticum. The activities of Rx-0_667 and linezolid against five isolates of C. pneumoniae appeared to be equivalent. Erythromycin had an MIC₉₀ of 2 µg/ml against all the species except M. hominis, where the MIC₉₀ was 256 µg/ml.

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TABLE 7. Antibacterial activities against atypical respiratory tract isolates

Susceptibility of linezolid-resistant isolates to Rx compounds. Rx-01_423 and Rx-01_667 were tested against a panel of clinical isolates of Staphylococcus spp. or Enterococcus spp. that were linezolid resistant. Linezolid resistance was conferred by a mutation in one or more copies of 23S rRNA (either G2576U or U2500A, according to Escherichia coli numbering) or by an uncharacterized mechanism (Table 8). In addition, the susceptibilities of a pair of isogenic S. aureus strains with and without cfr, the gene encoding an A2503 methylase that confers resistance to multiple antibiotics targeting the 50S ribosomal subunit (3, 30, 53), were evaluated. The susceptibility of a clinical isolate of S. pneumoniae (PU1071099) that contains a two-amino-acid deletion in ribosomal protein L4 conferring resistance to macrolides, linezolid, and chloramphenicol was also assessed (17, 58).

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TABLE 8. Susceptibilities of linezolid-resistant strains to Rx compounds

Both Rx compounds were 2-fold to >16-fold more potent than linezolid, with MICs of 0.25 to 64 µg/ml against the clinical staphylococcal strains that were linezolid resistant. The MIC of linezolid increased fourfold when the cfr gene was present in S. aureus, while it rose only twofold for the more potent Rx compounds. The MICs of linezolid were 8 to 64 µg/ml against linezolid-resistant enterococcal strains, whereas Rx-01_423 and Rx-01_667 were 4- to 32-fold more potent, with MIC ranges of 0.5 to 4 µg/ml and 0.25 to 4 µg/ml, respectively. The pneumococcal clinical isolate that is resistant to linezolid via a deletion in rplD (L4 gene) was fully susceptible to both Rx compounds. Although some cross-resistance was apparent, both Rx compounds were consistently more potent than linezolid, especially against linezolid-resistant enterococci, where the majority of linezolid resistance has been reported in clinics (4, 12, 20, 21, 23, 26, 28, 43, 61).

Time-kill studies. Rx-01_423 and Rx-01_667 were evaluated for their in vitro pharmacodynamic effects against three strains of MRSA, one strain of vancomycin-resistant E. faecalis, two pneumococcal isolates, and two H. influenzae isolates (Fig. 3). The MICs for the strains are listed in Table 9. Like linezolid and vancomycin, both Rx compounds were found to be bacteriostatic at 4x or 8x their respective MICs against the three MRSA isolates and the vanB E. faecalis isolate (Fig. 3). A greater-than-3-log kill was observed for both S. pneumoniae isolates and two non-type-B H. influenzae isolates for Rx-01_423. Like azithromycin, Rx-01_667 was bactericidal for three of the four respiratory tract isolates at 4x and 8x their respective MICs.

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FIG. 3. Time-kill kinetics of Rx compounds against three MRSA isolates (67-0, BK 11118, and BK 11512), a vancomycin-resistant E. faecalis isolate, two S. pneumoniae isolates (02J1258 and 02J1175), and two H. influenzae isolates (1100 and A1950). , Rx-01_423 at 4x MIC; , Rx-01_423 at 8x MIC; , Rx-01_667 at 4x MIC; •, Rx-01_667 at 8x MIC; , linezolid at 4x MIC; , linezolid at 8x MIC; , vancomycin at 4x MIC; , vancomycin at 8x MIC; *, azithromycin at 4x MIC; , azithromycin at 8x MIC.

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TABLE 9. MICs of compounds against strains for resistance frequency and time-kill kinetics

Frequency of spontaneous resistance. Table 1 lists the isolates used in resistance emergence studies, and their respective MICs for selected antibiotics can be found in Table 8. Strains were tested for frequency of resistance to Rx-01_423, Rx-01_667, and linezolid. The frequencies for all three compounds were below the limit of detection for the assay (generally <10^–9 to <10^–10) (data not shown). Rifampin, a compound that generally yields resistant mutants at a high frequency in vitro, was included as a positive control; resistance to rifampin for S. aureus ATCC 29213 was 2.3 x 10^–8 mutants/ml. The low frequency of spontaneous resistance to the Rx compounds was observed even when the strain was already linezolid resistant (e.g., E. faecalis ATCC 29212-P5 and E. faecium A6349).

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Rx-01_423 and Rx-01_667 are protein synthesis inhibitors that bind to the 50S ribosomal subunit more tightly than linezolid (Fig. 1). Their binding and spectrum were optimized using the atomic resolution structures of sparsomycin and linezolid complexed to the Haloarcula marismortui 50S region and a computational suite that enriched for H. influenzae activity and drug-like properties (22). We sought a compound that could be used intravenously and orally to treat major gram-positive nosocomial pathogens such as MRSA, S. pneumoniae, and vancomycin-resistant enterococci. A second objective of the program was to expand the spectrum to include fastidious gram-negative bacteria like H. influenzae and M. catarrhalis, thereby facilitating empirical treatment of serious community-acquired pneumonia, including pneumonia caused by MSSA, MRSA, S. pneumoniae, H. influenzae, or M. catarrhalis. Microbiological results for Rx-01_423 and Rx-01_667 demonstrate that both objectives were achieved. In addition, both compounds have good activity against the majority of intracellular bacteria responsible for CAP. The microbiological activities, combined with efficacy (oral and intravenous) in murine infection models (31), led to the evaluation of both compounds in phase I human pharmacokinetic/safety trials. Rx-01_667, known as RX-1741, has currently progressed to phase 2 trials, and its spectrum could make it a useful treatment option for CAP, especially if more MRSA is seen, as well as for serious gram-positive infections.

Linezolid resistance in E. faecalis, E. faecium, S. pneumoniae, and S. aureus has been documented (4, 12, 17, 20, 21, 23, 26, 28, 33, 35, 39, 43, 54, 58). Linezolid resistance is most commonly associated with a G2576T mutation in the gene encoding 23S rRNA. The T2500A mutation in domain V, the peptidyl transferase center, has also been found in an MRSA strain (35). Although linezolid-resistant strains are found relatively infrequently in the clinic, the infections that they cause can be life-threatening, underscoring the importance of readily available treatments. In addition to having two- to eightfold better activity against linezolid-susceptible staphylococci, enterococci, and pneumococci, Rx-01_423 and Rx-01_667 were also more active against linezolid-resistant isolates, especially enterococci and pneumococci. The Rx compounds were eightfold more active against the clinical isolate of S. pneumoniae that had a deletion in the conserved region of ribosomal protein L4 conferring resistance to macrolides, lincosamides, and chloramphenicol (17, 58). Both Rx compounds retained activity even when A2503 was modified by a Cfr rRNA methylase. Although few clinical isolates carrying cfr have been described, the tendency of this gene to reside in tandem with an Erm methylase on a mobilizable element does not bode well; erm genes are widely distributed among bacterial species.

The spontaneous frequency of resistance to either staphylococcal or enterococcal strains, including linezolid-resistant isolates that contain a G2576U mutation in one or more copies of 23S rRNA, was low. Although resistance to linezolid appears infrequently (1), especially in S. aureus, there have been reports of high levels (>20%) of linezolid-resistant enterococci in some hospitals (7, 12), and large surveillance studies of linezolid in the United States are consistent with an increase in the number of linezolid-resistant enterococci (27). With the continued use of linezolid, resistance will likely increase, especially if resistance can be mediated by mobile elements.

Like linezolid, Rx-01 compounds were bacteriostatic against MRSA and enterococci. Cidality was seen with both isolates of S. pneumoniae, including strain 02J1258, a multidrug-resistant isolate (59). Both compounds were bactericidal against H. influenzae 54A1100, but Rx-01_667 was static against H. influenzae A1950, like azithromycin at either 4x or 8x their respective MICs.

Rx-01_423 and Rx-01_667 were designed using knowledge gained from atomic resolution structures of the ribosome and from computational models that enriched for specific gram-negative activity and oral bioavailability (22, 56, 60). Their advantages over linezolid include an expanded spectrum that may permit the empirical treatment of serious CAP, including MRSA, and activity against linezolid-resistant enterococci and pneumococci. With both oral and intravenous dosage forms under development, Rx-01_667 should prove to be a useful addition to the medical armament.

 
This paper provides only a part of the story of the design of the Rx-01 family of antibiotics. We acknowledge the team of scientists including structural biologists, computational and medicinal chemists, biochemists, and pharmacologists whose combined efforts produced the Rx-01 oxazolidinone family.

 
* Corresponding author. Present address: NanoBio Corporation, 2311 Green Rd., Suite A, Ann Arbor, MI 48105. Phone: (734) 302-9128. Fax: (734) 302-9150. E-mail: joyce.sutcliffe{at}nanobio.com

Published ahead of print on 3 March 2008.

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1
1. Anderegg, T. R., H. S. Sader, T. R. Fritsche, J. E. Ross, and R. N. Jones. 2005. Trends in linezolid susceptibility patterns: report from the 2002-2003 worldwide Zyvox Annual Appraisal of Potency and Spectrum (ZAAPS) Program. Int. J. Antimicrob. Agents 26:13-21.[CrossRef][Medline]
2
2. Anderson, R. N., and B. L. Smith. 2005. Deaths: leading causes for 2002. Natl. Vital Stat. Rep. 53:1-89.[Medline]
3
3. Arias, C. A., M. V. Villegas, J. Reyes, J. Moreno, L. Xiong, M. Vallejo, K. Lolans, A. S. Mankin, E. Castaneda, and J. Quinn. 2006. Linezolid resistance in the absence of 23S rRNA mutations in methicillin-resistant Staphylococcus aureus, abstr. C2-1147, p. 125. Abstr. 46th Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
4
4. Auckland, C., L. Teare, F. Cooke, M. E. Kaufmann, M. Warner, G. Jones, K. Bamford, H. Ayles, and A. P. Johnson. 2002. Linezolid-resistant enterococci: report of the first isolates in the United Kingdom. J. Antimicrob. Chemother. 50:743-746.[Abstract/Free Full Text]
5
5. Bayer, A. S., C. Li, and M. Ing. 1998. Efficacy of trovafloxacin, a new quinolone antibiotic, in experimental staphylococcal endocarditis due to oxacillin-resistant strains. Antimicrob. Agents Chemother. 42:1837-1841.[Abstract/Free Full Text]
6
6. Beekmann, S. E., K. P. Heilmann, S. S. Richter, J. Garcia-de-Lomas, and G. V. Doern. 2005. Antimicrobial resistance in Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and group A beta-haemolytic streptococci in 2002-2003. Results of the multinational GRASP Surveillance Program. Int. J. Antimicrob. Agents 25:148-156.[CrossRef][Medline]
7
7. Bonora, M. G., M. Solbiati, E. Stepan, A. Zorzi, A. Luzzani, M. R. Catania, and R. Fontana. 2006. Emergence of linezolid resistance in the vancomycin-resistant Enterococcus faecium multilocus sequence typing C1 epidemic lineage. J. Clin. Microbiol. 44:1153-1155.[Abstract/Free Full Text]
8
8. Brown, S. D., and D. J. Farrell. 2004. Antibacterial susceptibility among Streptococcus pneumoniae isolated from paediatric and adult patients as part of the PROTEKT US study in 2001-2002. J. Antimicrob. Chemother. 54(Suppl. 1):i23-i29.[Abstract]
9
9. Brown, S. D., D. J. Farrell, and I. Morrissey. 2004. Prevalence and molecular analysis of macrolide and fluoroquinolone resistance among isolates of Streptococcus pneumoniae collected during the 2000-2001 PROTEKT US study. J. Clin. Microbiol. 42:4980-4987.[Abstract/Free Full Text]
10
10. Clinical and Laboratory Standards Institute. 2006. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically, Approved standard, 7th ed., vol. 26. CLSI, Wayne, PA.
11
11. Critchley, I. A., M. E. Jones, P. D. Heinze, D. Hubbard, H. D. Engler, A. T. Evangelista, C. Thornsberry, J. A. Karlowsky, and D. F. Sahm. 2002. In vitro activity of levofloxacin against contemporary clinical isolates of Legionella pneumophila, Mycoplasma pneumoniae and Chlamydia pneumoniae from North America and Europe. Clin. Microbiol. Infect. 8:214-221.[CrossRef][Medline]
12
12. Dobbs, T. E., M. Patel, K. B. Waites, S. A. Moser, A. M. Stamm, and C. J. Hoesley. 2006. Nosocomial spread of Enterococcus faecium resistant to vancomycin and linezolid in a tertiary care medical center. J. Clin. Microbiol. 44:3368-3370.[Abstract/Free Full Text]
13
13. Doern, G. V., K. P. Heilmann, H. K. Huynh, P. R. Rhomberg, S. L. Coffman, and A. B. Brueggemann. 2001. Antimicrobial resistance among clinical isolates of Streptococcus pneumoniae in the United States during 1999-2000, including a comparison of resistance rates since 1994-1995. Antimicrob. Agents Chemother. 45:1721-1729.[Abstract/Free Full Text]
14
14. Dunbar, L. M., and D. J. Farrell. 2007. Activity of telithromycin and comparators against isolates of Legionella pneumophila collected from patients with community-acquired respiratory tract infections: PROTEKT years 1-5. Clin. Microbiol. Infect. 13:743-746.[CrossRef][Medline]
15
15. Farrell, D. J., and S. G. Jenkins. 2004. Distribution across the USA of macrolide resistance and macrolide resistance mechanisms among Streptococcus pneumoniae isolates collected from patients with respiratory tract infections: PROTEKT US 2001-2002. J. Antimicrob. Chemother. 54(Suppl. 1):i17-i22.[Abstract]
16
16. Farrell, D. J., S. G. Jenkins, S. D. Brown, M. Patel, B. S. Lavin, and K. P. Klugman. 2005. Emergence and spread of Streptococcus pneumoniae with erm(B) and mef(A) resistance. Emerg. Infect. Dis. 11:851-858.[Medline]
17
17. Farrell, D. J., I. Morrissey, S. Bakker, S. Buckridge, and D. Felmingham. 2004. In vitro activities of telithromycin, linezolid, and quinupristin-dalfopristin against Streptococcus pneumoniae with macrolide resistance due to ribosomal mutations. Antimicrob. Agents Chemother. 48:3169-3171.[Abstract/Free Full Text]
18
18. Fenelon, L. E., G. Mumtaz, and G. L. Ridgway. 1990. The in-vitro antibiotic susceptibility of Chlamydia pneumoniae. J. Antimicrob. Chemother. 26:763-767.[Abstract/Free Full Text]
19
19. Gillet, Y., B. Issartel, P. Vanhems, J. C. Fournet, G. Lina, M. Bes, F. Vandenesch, Y. Piemont, N. Brousse, D. Floret, and J. Etienne. 2002. Association between Staphylococcus aureus strains carrying gene for Panton-Valentine leukocidin and highly lethal necrotising pneumonia in young immunocompetent patients. Lancet 359:753-759.[CrossRef][Medline]
20
20. Gonzales, R. D., P. C. Schreckenberger, M. B. Graham, S. Kelkar, K. DenBesten, and J. P. Quinn. 2001. Infections due to vancomycin-resistant Enterococcus faecium resistant to linezolid. Lancet 357:1179.[CrossRef][Medline]
21
21. Herrero, I. A., N. C. Issa, and R. Patel. 2002. Nosocomial spread of linezolid-resistant, vancomycin-resistant Enterococcus faecium. N. Engl. J. Med. 346:867-869.[Free Full Text]
22
22. Ippolito, J., Z. Kanyo, B. Wimberly, D. Wang, E. Skripkin, J. Devito, B. Freeborn, J. Sutcliffe, E. Duffy, and F. Franceschi. 2005. Structural basis for the binding of Rx-01, a novel oxazolidinone class, to bacterial ribosomes, abstr. F-1254, p. 197. Abstr. 45th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
23
23. Johnson, A. P., L. Tysall, M. W. Stockdale, N. Woodford, M. E. Kaufmann, M. Warner, D. M. Livermore, F. Asboth, and F. J. Allerberger. 2002. Emerging linezolid-resistant Enterococcus faecalis and Enterococcus faecium isolated from two Austrian patients in the same intensive care unit. Eur. J. Clin. Microbiol. Infect. Dis. 21:751-754.[CrossRef][Medline]
24
24. Jones, M. E., R. S. Blosser-Middleton, C. Thornsberry, J. A. Karlowsky, and D. F. Sahm. 2003. The activity of levofloxacin and other antimicrobials against clinical isolates of Streptococcus pneumoniae collected worldwide during 1999-2002. Diagn. Microbiol. Infect. Dis. 47:579-586.[CrossRef][Medline]
25
25. Jones, M. E., D. C. Draghi, C. Thornsberry, J. A. Karlowsky, D. F. Sahm, and R. P. Wenzel. 2004. Emerging resistance among bacterial pathogens in the intensive care unit—a European and North American Surveillance study (2000-2002). Ann. Clin. Microbiol. Antimicrob. 3:14. http://www.ann-clinmicrob.com/content/3/1/14.[CrossRef][Medline]
26
26. Jones, R. N., P. Della-Latta, L. V. Lee, and D. J. Biedenbach. 2002. Linezolid-resistant Enterococcus faecium isolated from a patient without prior exposure to an oxazolidinone: report from the SENTRY Antimicrobial Surveillance Program. Diagn. Microbiol. Infect. Dis. 42:137-139.[CrossRef][Medline]
27
27. Jones, R. N., T. R. Fritsche, H. S. Sader, and J. E. Ross. 2007. LEADER surveillance program results for 2006: an activity and spectrum analysis of linezolid using clinical isolates from the United States (50 medical centers). Diagn. Microbiol. Infect. Dis. 59:309-317.[Medline]
28
28. Jones, R. N., J. E. Ross, T. R. Fritsche, and H. S. Sader. 2006. Oxazolidinone susceptibility patterns in 2004: report from the Zyvox Annual Appraisal of Potency and Spectrum (ZAAPS) Program assessing isolates from 16 nations. J. Antimicrob. Chemother. 57:279-287.[Abstract/Free Full Text]
29
29. Karlowsky, J. A., M. E. Jones, D. C. Draghi, C. Thornsberry, D. F. Sahm, and G. A. Volturo. 2004. Prevalence and antimicrobial susceptibilities of bacteria isolated from blood cultures of hospitalized patients in the United States in 2002. Ann. Clin. Microbiol. Antimicrob. 3:7.[CrossRef][Medline]
30
30. Long, K. S., J. Poehlsgaard, C. Kehrenberg, S. Schwarz, and B. Vester. 2006. The Cfr rRNA methyltransferase confers resistance to phenicols, lincosamides, oxazolidinones, pleuromutilins, and streptograrnin A antibiotics. Antimicrob. Agents Chemother. 50:2500-2505.[Abstract/Free Full Text]
31
31. Luo, X., K. Harrington, H. Jing, E. Burak, and J. Sutcliffe. 2005. In vivo efficacy of designer oxazolidinones in murine infectious models, abstr. F-1260, p. 199. Abstr. 45th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
32
32. Marrie, T. J., M. Poulin-Costello, M. D. Beecroft, and Z. Herman-Gnjidic. 2005. Etiology of community-acquired pneumonia treated in an ambulatory setting. Respir. Med. 99:60-65.[CrossRef][Medline]
33
33. Marshall, S. H., C. J. Donskey, R. Hutton-Thomas, R. A. Salata, and L. B. Rice. 2002. Gene dosage and linezolid resistance in Enterococcus faecium and Enterococcus faecalis. Antimicrob. Agents Chemother. 46:3334-3336.[Abstract/Free Full Text]
34
34. McCormick, A. W., C. G. Whitney, M. M. Farley, R. Lynfield, L. H. Harrison, N. M. Bennett, W. Schaffner, A. Reingold, J. Hadler, P. Cieslak, M. H. Samore, and M. Lipsitch. 2003. Geographic diversity and temporal trends of antimicrobial resistance in Streptococcus pneumoniae in the United States. Nat. Med. 9:424-430.[CrossRef][Medline]
35
35. Meka, V. G., S. K. Pillai, G. Sakoulas, C. Wennersten, L. Venkataraman, P. C. DeGirolami, G. M. Eliopoulos, R. C. Moellering, Jr., and H. S. Gold. 2004. Linezolid resistance in sequential Staphylococcus aureus isolates associated with a T2500A mutation in the 23S rRNA gene and loss of a single copy of rRNA. J. Infect. Dis. 190:311-317.[CrossRef][Medline]
36
36. Moran, G. J., A. Krishnadasan, R. J. Gorwitz, G. E. Fosheim, L. K. McDougal, R. B. Carey, and D. A. Talan. 2006. Methicillin-resistant S. aureus infections among patients in the emergency department. N. Engl. J. Med. 355:666-674.[Abstract/Free Full Text]
37
37. Pfaller, M. A., A. F. Ehrhardt, and R. N. Jones. 2001. Frequency of pathogen occurrence and antimicrobial susceptibility among community-acquired respiratory tract infections in the respiratory surveillance program study: microbiology from the medical office practice environment. Am. J. Med. 111(Suppl. 9A):4S-12S.[CrossRef][Medline]
38
38. Prystowsky, J., F. Siddiqui, J. Chosay, D. L. Shinabarger, J. Millichap, L. R. Peterson, and G. A. Noskin. 2001. Resistance to linezolid: characterization of mutations in rRNA and comparison of their occurrences in vancomycin-resistant enterococci. Antimicrob. Agents Chemother. 45:2154-2156.[Abstract/Free Full Text]
39
39. Rahim, S., S. K. Pillai, H. S. Gold, L. Venkataraman, K. Inglima, and R. A. Press. 2003. Linezolid-resistant, vancomycin-resistant Enterococcus faecium infection in patients without prior exposure to linezolid. Clin. Infect. Dis. 36:E146-E148.[CrossRef][Medline]
40
40. Ridgway, G. L., J. M. Owen, and J. D. Oriel. 1976. A method for testing the antibiotic susceptibility of Chlamydia trachomatis in a cell culture system. J. Antimicrob. Chemother. 2:71-76.[Abstract/Free Full Text]
41
41. Sahm, D. F., J. A. Karlowsky, L. J. Kelly, I. A. Critchley, M. E. Jones, C. Thornsberry, Y. Mauriz, and J. Kahn. 2001. Need for annual surveillance of antimicrobial resistance in Streptococcus pneumoniae in the United States: 2-year longitudinal analysis. Antimicrob. Agents Chemother. 45:1037-1042.[Abstract/Free Full Text]
42
42. Sakoulas, G., H. S. Gold, L. Venkataraman, R. C. Moellering, M. J. Ferraro, and G. M. Eliopoulos. 2003. Introduction of erm(C), into a linezolid- and methicillin-resistant Staphylococcus aureus does not restore linezolid susceptibility. J. Antimicrob. Chemother. 51:1039-1041.[Free Full Text]
43
43. Seedat, J., G. Zick, I. Klare, C. Konstabel, N. Weiler, and H. Sahly. 2006. Rapid emergence of resistance to linezolid during linezolid therapy of an Enterococcus faecium infection. Antimicrob. Agents Chemother. 50:4217-4219.[Abstract/Free Full Text]
44
44. Seenivasan, M. H., V. L. Yu, and R. R. Muder. 2005. Legionnaires' disease in long-term care facilities: overview and proposed solutions. J. Am. Geriatr. Soc. 53:875-880.[CrossRef][Medline]
45
45. Seybold, U., E. V. Kourbatova, J. G. Johnson, S. J. Halvosa, Y. F. Wang, M. D. King, S. M. Ray, and H. M. Blumberg. 2006. Emergence of community-associated methicillin-resistant Staphylococcus aureus USA300 genotype as a major cause of health care-associated blood stream infections. Clin. Infect. Dis. 42:647-656.[CrossRef][Medline]
46
46. Shopsin, B., and B. N. Kreiswirth. 2001. Molecular epidemiology of methicillin-resistant Staphylococcus aureus. Emerg. Infect. Dis. 7:323-326.[Medline]
47
47. Skripkin, E., T. McConnell, B. King, J. DeVito, F. Franceschi, and J. Sutcliffe. 2005. Designer oxazolidinones bind to the 50S peptidyl-transferase region and can overcome ribosome-based linezolid resistance, abstr. F-1255, p. 197. Abstr. 45th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
48
48. Sokol, W. 2001. Epidemiology of sinusitis in the primary care setting: results from the 1999-2000 Respiratory Surveillance Program. Am. J. Med. 111(Suppl. 9A):19S-24S.[CrossRef][Medline]
49
49. Stockdale, M. W., L. Tysall, A. P. Johnson, D. M. Livermore, and N. Woodford. 2004. Low in vitro selection frequencies of enterococcal and staphylococcal mutants resistant to the oxazolidinone AZD2563. Int. J. Antimicrob. Agents 23:88-91.[CrossRef][Medline]
50
50. Styers, D., D. J. Sheehan, P. Hogan, and D. F. Sahm. 2006. Laboratory-based surveillance of current antimicrobial resistance patterns and trends among Staphylococcus aureus: 2005 status in the United States. Ann. Clin. Microbiol. Antimicrob. 5:2.[CrossRef][Medline]
51
51. Tait-Kamradt, A., J. Clancy, M. Cronan, F. Dib-Hajj, L. Wondrack, W. Yuan, and J. Sutcliffe. 1997. mefE is necessary for the erythromycin-resistant M phenotype in Streptococcus pneumoniae. Antimicrob. Agents Chemother. 41:2251-2255.[Abstract]
52
52. Tenover, F. C., L. K. McDougal, R. V. Goering, G. Killgore, S. J. Projan, J. B. Patel, and P. M. Dunman. 2006. Characterization of a strain of community-associated methicillin-resistant Staphylococcus aureus widely disseminated in the United States. J. Clin. Microbiol. 44:108-118.[Abstract/Free Full Text]
53
53. Toh, S. M., L. Xiong, C. A. Arias, M. V. Villegas, K. Lolans, J. Quinn, and A. S. Mankin. 2007. Acquisition of a natural resistance gene renders a clinical strain of methicillin-resistant Staphylococcus aureus resistant to the synthetic antibiotic linezolid. Mol. Microbiol. 64:1506-1514.[CrossRef][Medline]
54
54. Tsiodras, S., H. S. Gold, G. Sakoulas, G. M. Eliopoulos, C. Wennersten, L. Venkataraman, R. C. Moellering, and M. J. Ferraro. 2001. Linezolid resistance in a clinical isolate of Staphylococcus aureus. Lancet 358:207-208.[CrossRef][Medline]
55
55. Waites, K. B., L. B. Duffy, T. Schmid, D. Crabb, M. S. Pate, and G. H. Cassell. 1991. In vitro susceptibilities of Mycoplasma pneumoniae, Mycoplasma hominis, and Ureaplasma urealyticum to sparfloxacin and PD 127391. Antimicrob. Agents Chemother. 35:1181-1185.[Abstract/Free Full Text]
56
56. Wang, D., E. Sherer, and E. Duffy. 2005. A computational suite for the discovery of designer oxazolidinones suitable for intravenous and oral usage, abstr. F-1853, p. 211. Abstr. 45th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
57
57. Wisplinghoff, H., T. Bischoff, S. M. Tallent, H. Seifert, R. P. Wenzel, and M. B. Edmond. 2004. Nosocomial bloodstream infections in US hospitals: analysis of 24,179 cases from a prospective nationwide surveillance study. Clin. Infect. Dis. 39:309-317.[CrossRef][Medline]
58
58. Wolter, N., A. M. Smith, D. J. Farrell, W. Schaffner, M. Moore, C. G. Whitney, J. H. Jorgensen, and K. P. Klugman. 2005. Novel mechanism of resistance to oxazolidinones, macrolides, and chloramphenicol in ribosomal protein L4 of the pneumococcus. Antimicrob. Agents Chemother. 49:3554-3557.[Abstract/Free Full Text]
59
59. Wolter, N., A. M. Smith, D. E. Low, and K. P. Klugman. 2007. High-level telithromycin resistance in a clinical isolate of Streptococcus pneumoniae. Antimicrob. Agents Chemother. 51:1092-1095.[Abstract/Free Full Text]
60
60. Zhou, J., A. Bhattacharjee, S. Chen, Y. Chen, E. Duffy, J. Farmer, J. Goldberg, R. Hanselmann, J. Ippolito, R. Lou, A. Orbin, A. Oyelere, J. Salvino, D. Springer, J. Tran, D. Wang, and Y. Wu. 2005. Design at the atomic level: synthesis of biaryl oxazolidinones as potent orally active antibiotics, abstr. F-1252, p. 197. Abstr. 45th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
61
61. Zurenko, G. E., W. M. Todd, B. Hafkin, B. Meyers, C. Kauffman, J. Bock, J. Slightom, and D. Shinabarger. 1999. Development of linezolid-resistant Enterococcus faecium in two compassionate use program patients treated with linezolid, abstr. 0848, p. 118. Abstr. 39th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.

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Antimicrobial Agents and Chemotherapy, May 2008, p. 1653-1662, Vol. 52, No. 5
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