Besifloxacin is a new fluoroquinolone anti-infective developed for ophthalmic use. Besifloxacin ophthalmic suspension 0.6%(Besivance) was recently approved for the treatment of bacterial conjunctivitis. The objective of this article is to provide a comprehensive overview of microbiological, pharmacokinetic/pharmacodynamic and clinical studies with besifloxacin. Microbiological studies have demonstrated that besifloxacin has wide-spectrum and potent activity against common ocular pathogens, including Gram-negative and Gram-positive pathogens associated with bacterial conjunctivitis, and retained activity against fluoroquinolone-resistant staphylococci and multidrug-resistant strains. In preclinical and human studies, topically applied besifloxacin had a prolonged ocular concentration and minimal systemic exposure. In clinical studies, patients randomized to besifloxacin ophthalmic suspension 0.6% experienced significantly higher rates of clinical resolution and microbial eradication than patients randomized to vehicle. Besifloxacin ophthalmic suspension 0.6% was also found to be as effective and well tolerated as moxifloxacin ophthalmic solution 0.5%. The low minimum inhibitory concentrations and high attainment of pharmacodynamic targets with besifloxacin may contribute to a lower risk for the emergence of bacterial resistance, although further studies are needed. These data indicate that besifloxacin ophthalmic suspension 0.6% is an important new option for the treatment of bacterial conjunctivitis.

Purpose: To assess the impact of benzalkonium chloride (BAK) on the minimum inhibitory concentration (MIC) and mutant prevention concentration (MPC) of gatifloxacin against Gram-positive pathogens in comparison to gatifloxacin and moxifloxacin alone, moxifloxacin plus BAK, and/or levofloxacin. Methods: The MIC was measured following incubation of 10(5) colony-forming units (CFU)/mL of coagulase-negative staphylococci (CNS; n = 20), methicillin-susceptible Staphylococcus aureus (MSSA; n = 20), and methicillin-resistant S. aureus (MRSA; n = 20) with gatifloxacin, levofloxacin, or moxifloxacin. When present, BAK was added from 3.125 mug/mL to 6.25 mug/mL. The MPC was measured following incubation of 10(10) CFU/mL of MRSA (n = 9) and a commercially available MSSA strain with gatifloxacin or moxifloxacin in the absence and presence of BAK at concentrations from 7 mug/mL to 10 mug/mL. Results: CNS was more susceptible to gatifloxacin (MIC(90)= 2 mug/mL) than levofloxacin (MIC(90)= 8 mug/mL) or moxifloxacin (MIC(90)= 4 mug/mL). MSSA was more susceptible to moxifloxacin (MIC(90)= 1 mug/mL) than gatifloxacin (MIC(90)= 4 mug/mL) or levofloxacin (MIC(90)= 4 mug/mL). MRSA were resistant to gatifloxacin, levofloxacin, and moxifloxacin. In the presence of BAK, however, the MIC(90) of gatifloxacin and moxifloxacin against CNS, MSSA, and MRSA was </=0.008 mug/mL. Gatifloxacin and moxifloxacin had similar MPCs against MRSA (>/=4 mug/mL). In the presence of BAK, the MPC of gatifloxacin and moxifloxacin against MRSA ranged from </=0.004 mug/mL to 0.125 mug/mL. Conclusions: BAK substantially lowered the MIC and MPC of gatifloxacin and moxifloxacin against Gram-positive staphylococci compared to gatifloxacin alone, moxifloxacin alone, and/or levofloxacin. These findings suggest that the presence of BAK in the ophthalmic formulation of gatifloxacin (Zymar((R))) may serve to enhance the potency of gatifloxacin and decrease its propensity to select for fluoroquinolone-resistant S. aureus strains.

Objectives Besifloxacin is a new fluoroquinolone in development for ocular use. We investigated its mode of action and resistance in two major ocular pathogens, Streptococcus pneumoniae and Staphylococcus aureus, and in the reference species Escherichia coli. Methods Primary and secondary targets of besifloxacin were evaluated by:(i) mutant selection experiments;(ii) MIC testing of defined topoisomerase mutants; and (iii) inhibition and cleavable complex assays with purified S. pneumoniae and E. coli DNA gyrase and topoisomerase IV enzymes. Results Enzyme assays showed similar besifloxacin activity against S. pneumoniae gyrase and topoisomerase IV, with IC(50) and CC(25) of 2.5 and 1 microM, respectively. In contrast to ciprofloxacin and moxifloxacin, besifloxacin was equally potent against both S. pneumoniae and E. coli gyrases. DNA gyrase was the primary target in all three species, with substitutions observed at positions 81, 83 and 87 in GyrA and 426 and 466 in GyrB (E. coli numbering). Topoisomerase IV was the secondary target. Notably, resistant mutants were not recovered at 4-fold besifloxacin MICs for S. aureus and S. pneumoniae, and S. aureus topoisomerase mutants were only obtained after serial passage in liquid medium. Besifloxacin MICs were similarly affected by parC or gyrA mutations in S. aureus and S. pneumoniae and remained below 1 mg/L in gyrA-parC double mutants. Conclusions Although mutant selection experiments indicated that gyrase is a primary target, further biochemical and genetic studies showed that besifloxacin has potent, relatively balanced activity against both essential DNA gyrase and topoisomerase IV targets in S. aureus and S. pneumoniae.

Clerocidin (CL), a microbial diterpenoid, reacts with DNA via its epoxide group and stimulates DNA cleavage by type II DNA topoisomerases. The molecular basis of CL action is poorly understood. We establish by genetic means that CL targets DNA gyrase in the Gram-positive bacterium Streptococcus pneumoniae, and promotes gyrase-dependent single- and double-stranded DNA cleavage in vitro. CL-stimulated DNA breakage exhibited a strong preference for guanine preceding the scission site (-1 position). Mutagenesis of -1 guanines to A, C or T abrogated CL cleavage at a strong pBR322 site. Surprisingly, for double-strand breaks, scission on one strand consistently involved a modified (piperidine-labile) guanine and was not reversed by heat, salt or EDTA, whereas complementary strand scission occurred at a piperidine-stable -1 nt and was reversed by EDTA. CL did not induce cleavage by a mutant gyrase (GyrA G79A) identified here in CL-resistant pneumococci. Indeed, mutations at G79 and at the neighbouring S81 residue in the GyrA breakage-reunion domain discriminated poisoning by CL from that of antibacterial quinolones. The results suggest a novel mechanism of enzyme inhibition in which the -1 nt at the gyrase-DNA gate exhibit different CL reactivities to produce both irreversible and reversible DNA damage.

Community-acquired pneumonia (CAP) is the cause of substantial morbidity, mortality, and resource utilization worldwide. When choosing an antimicrobial, effective treatment depends on proper patient evaluation and the identification of numerous risk factors, such as recent antibiotic exposure or the presence of comorbidity. Patients without any risk factor should be treated effectively with a narrow spectrum beta-lactam agent, like amoxicillin, or a macrolide. If a risk factor is present, agents with a broader spectrum of activity should be selected for the empirical therapy. The newer-generation quinolones are suitable agents with their excellent in vitro activity and pharmacodynamic-pharmacokinetic properties. They are not only active against susceptible CAP pathogens, but also against the resistant strains. Among the quinolones, gemifloxacin has the best in vitro activity. Its improved bioavailability, pharmacokinetic-pharmacodynamic properties, and safety profile make this agent an excellent option for the treatment of CAP.

Clerocidin (CL), a diterpenoid natural product, alkylates DNA through its epoxide moiety and exhibits both anticancer and antibacterial activities. We have examined CL action in the presence of topoisomerase IV from Streptococcus pneumoniae. CL promoted irreversible enzyme-mediated DNA cleavage leading to single- and double-stranded DNA breaks at specific sites. Reaction required the diterpenoid function: no cleavage was seen using a naphthalene-substituted analogue. Moreover, drug-induced DNA breakage was not observed using a mutant topoisomerase IV (ParC Y118F) unable to form a cleavage complex with DNA. Sequence analysis of 102 single-stranded DNA breaks and 79 double-stranded breaks revealed an overwhelming preference for G at the -1 position, i.e. immediately 5' of the enzyme DNA scission site. This specificity contrasts with that of topoisomerase IV cleavage with antibacterial quinolones. Indeed, CL stimulated DNA breakage by a quinolone-resistant topoisomerase IV (ParC S79F). Overall, the results indicate that topoisomerase IV facilitates selective irreversible CL attack at guanine and that its cleavage complex differs markedly from that of mammalian topoisomerase II which promotes both irreversible and reversible CL attack at guanine and cytosine, respectively. The unique ability to form exclusively irreversible DNA breaks suggests topoisomerase IV may be a key intracellular target of CL in bacteria.

Fluoroquinolones are a class of synthetic antibacterial agents that were approved for ocular therapy in 1991 and have become popular therapy for the treatment and prevention of various ocular infections. These agents are synthetic, broad-spectrum, rapidly bactericidal, and have good penetration into ocular tissues. Their main mechanism of action is the inhibition of bacterial enzymes needed for bacterial DNA synthesis. However, antibiotic resistance occurred swiftly to the earlier fluoroquinolones and better fluoroquinolones were needed. The fourth-generation fluoroquinolones, such as moxifloxacin and gatifloxacin, have enhanced activity against gram-positive bacteria while retaining potent activity against most gram-negative bacteria. These fourth-generation fluoroquinolones have improved penetration into the anterior chamber and have also demonstrated increased in vivo efficacy in several animal models of ocular infections. In addition, topical ophthalmic antibiotic products can deliver antibiotic concentrations directly to the eye that are thousands of times higher than their MICs. This article reviews published data describing the in vitro potency of moxifloxacin and its in vivo activity for treating and preventing experimental ocular infections.

The increased use of fluoroquinolones has led to increasing resistance to these antimicrobials, with rates of resistance that vary by both organism and geographic region. Resistance to fluoroquinolones typically arises as a result of alterations in the target enzymes (DNA gyrase and topoisomerase IV) and of changes in drug entry and efflux. Mutations are selected first in the more susceptible target: DNA gyrase, in gram-negative bacteria, or topoisomerase IV, in gram-positive bacteria. Additional mutations in the next most susceptible target, as well as in genes controlling drug accumulation, augment resistance further, so that the most-resistant isolates have mutations in several genes. Resistance to quinolones can also be mediated by plasmids that produce the Qnr protein, which protects the quinolone targets from inhibition. Qnr plasmids have been found in the United States, Europe, and East Asia. Although Qnr by itself produces only low-level resistance, its presence facilitates the selection of higher-level resistance mutations, thus contributing to the alarming increase in resistance to quinolones.

The increasing resistance of Streptococcus pneumoniae, the most important community respiratory pathogen, to beta-lactams and other first-line antimicrobial agents usually employed for the empirical treatment of lower respiratory tract infections has led to the inclusion, in several current guidelines, of a fluoroquinolone with improved activity against pneumococci as the first choice agent for the management of such infections. The excellent microbiological, pharmacokinetic, and pharmacodynamic characteristics of the new fluoroquinolones (levofloxacin, moxifloxacin, gemifloxacin, and gatifloxacin) have encouraged their growing use, probably contributing to the emergence of fluoroquinolone-resistant pneumococci; although pneumococcal resistance to new fluoroquinolones is currently low, there is still concern about the potential for widespread emergence of resistance to these agents if they become indiscriminately used. Levofloxacin clinical failures have already been reported in the management of patients with pneumococcal community-acquired pneumonia; development of resistance in clinical isolates of S. pneumoniae has prompted a critical reexamination of the newer fluoroquinolones to assess their potency and to preserve their activity. An understanding of the pharmacokinetic and pharmacodynamic properties, allowing selection of the most potent fluoroquinolone, will reduce the opportunity for resistance to develop. Finally, a targeted use of these agents will maintain class efficacy.

Gemifloxacin is a synthetic fluoroquinolone antimicrobial agent exhibiting potent activity against most gram-negative and gram-positive organisms, such as the important community-acquired respiratory pathogens Streptococcus pneumoniae (including multidrug-resistant S. pneumoniae), Haemophilus influenzae , and Moraxella catarrhalis . The agent's mechanism of action involves dual targeting of two essential bacterial enzymes: DNA gyrase and topoisomerase IV. Gemifloxacin was approved by the Food and Drug Administration in April 2003 for treatment of community-acquired pneumonia and acute bacterial exacerbation of chronic bronchitis. The drug has an oral bioavailability of approximately 71%. Approximately 20-35% of gemifloxacin is excreted unchanged in the urine after 24 hours. The elimination half-life of gemifloxacin is 6-8 hours in patients with normal renal function, supporting once-daily dosing. The 24-hour free-drug area under the plasma concentration-time curve:minimum inhibitory concentration ratio (fAUC(0-24):MIC) associated with efficacy, based on results from in vitro and animal models of infection, is approximately 30. With a mean fAUC(0-24) of approximately 3 microg*hour/ml (35% of total AUC(0-24) of 8.4) and a median S. pneumoniae MIC for 90% of tested strains of 0.03, a fAUC(0-24):MIC ratio of 100 would be expected after standard dosing (320 mg once/day). In clinical studies involving both hospitalized and outpatient populations, gemifloxacin has been highly effective in the treatment of community-acquired pneumonia and acute exacerbation of chronic bronchitis. Clinical success rates ranged from 93.9-95.9% in patients with community-acquired pneumonia and 96.1-97.5% in those with acute exacerbation of chronic bronchitis. Gemifloxacin is well tolerated; the frequency of adverse events with this agent is low. Most adverse events are mild-to-moderate in severity, with diarrhea (< 4%), nausea and rash (< 3%), and headache (< 2%) most commonly reported. Drug interactions with gemifloxacin are not common, although absorption is greatly reduced when given with divalent and trivalent cation-containing compounds, such as antacids. Due to its potent activity against many common gram-positive and gram-negative respiratory pathogens, its proven clinical efficacy, and its favorable safety profile, gemifloxacin is a highly effective empiric treatment for community-acquired lower respiratory tract infections.

Quinolones are one of the largest classes of antimicrobial agents used worldwide. This review considers the quinolones that are available currently and used widely in Europe (norfoxacin, ciprofloxacin, ofloxacin, levofloxacin and moxifloxacin) within their historical perspective, while trying to position them in the context of recent and possible future advances based on an understanding of:(1) their chemical structures and how these impact on activity and toxicity;(2) resistance mechanisms (mutations in target genes, efflux pumps);(3) their pharmacodynamic properties (AUC/MIC and Cmax/MIC ratios; mutant prevention concentration and mutant selection window); and (4) epidemiological considerations (risk of emergence of resistance, clonal spread). Their main indications are examined in relation to their advantages and drawbacks. Overall, it is concluded that these important agents should be used in an educated fashion, based on a careful balance between their ease of use and efficacy vs. the risk of emerging resistance and toxicity. However, there is now substantial evidence to support use of the most potent drug at the appropriate dose whenever this is required.

In Streptococcus pneumoniae, an H103Y substitution in the ATP binding site of the ParE subunit of topoisomerase IV was shown to confer quinolone resistance and hypersensitivity to novobiocin when associated with an S84F change in the A subunit of DNA gyrase. We reconstituted in vitro the wild-type topoisomerase IV and its ParE mutant. The ParE mutant enzyme showed a decreased activity for decatenation at subsaturating ATP levels and was more sensitive to inhibition by novobiocin but was as sensitive to quinolones. These results show that the ParE alteration H103Y alone is not responsible for quinolone resistance and agree with the assumption that it facilitates the open conformation of the ATP binding site that would lead to novobiocin hypersensitivity and to a higher requirement of ATP.

Topoisomerase (topo) IV and gyrase are bacterial type IIA DNA topoisomerases essential for DNA replication and chromosome segregation that act via a transient double-stranded DNA break involving a covalent enzyme-DNA "cleavage complex." Despite their mechanistic importance, the DNA breakage determinants are not understood for any bacterial type II enzyme. We investigated DNA cleavage by Streptococcus pneumoniae topo IV and gyrase stabilized by gemifloxacin and other antipneumococcal fluoroquinolones. Topo IV and gyrase induce distinct but overlapping repertoires of double-strand DNA breakage sites that were essentially identical for seven different quinolones and were augmented (in intensity) by positive or negative supercoiling. Sequence analysis of 180 topo IV and 126 gyrase sites promoted by gemifloxacin on pneumococcal DNA revealed the respective consensus sequences: G(G/c)(A/t)A*GNNCt(T/a)N(C/a) and GN4G(G/c)(A/c)G*GNNCtTN(C/a)(preferred bases are underlined; disfavored bases are in small capitals; N indicates no preference; and asterisk indicates DNA scission between -1 and +1 positions). Both enzymes show strong preferences for bases clustered symmetrically around the DNA scission site, i.e.+1G/+4C,-4G/+8C, and particularly the novel -2A/+6T, but with no preference at +2/+3 within the staggered 4-bp overhang. Asymmetric elements include -3G and several unfavored bases. These cleavage preferences, the first for Gram-positive type IIA topoisomerases, differ markedly from those reported for Escherichia coli topo IV (consensus (A/G)*T/A) and gyrase, which are based on fewer sites. However, both pneumococcal enzymes cleaved an E. coli gyrase site suggesting overlap in gyrase determinants. We propose a model for the cleavage complex of topo IV/gyrase that accommodates the unique -2A/+6T and other preferences.

We determined fluoroquinolone microbiological resistance breakpoints for Streptococcus pneumoniae by using genetic instead of pharmacokinetic-pharmacodynamic parameters. The proposed microbiological breakpoints define resistance as the MIC at which >50% of the isolates carry quinolone resistance-determining region mutations and/or, if data are available, when Monte Carlo simulations demonstrate a <90% chance of bacteriological eradication. The proposed microbiological resistant breakpoints are as follows (in micrograms per milliliter): gatifloxacin,>0.25; gemifloxacin,>0.03; levofloxacin,>1; and moxifloxacin,>0.12. Monte Carlo simulations of the once daily 400-mg doses of gatifloxacin and 750-mg doses levofloxacin demonstrated a high level of target attainment (free-drug area under the concentration-time curve from 0 to 24 h/MIC ratio of 30) by using these new genetically derived breakpoints.

OBJECTIVE: To evaluate the microbiology, pharmacokinetic parameters, drug interactions, and results of the available clinical trials of gemifloxacin for the treatment of community-acquired pneumonia (CAP) and acute exacerbation of chronic bronchitis (AECB). DATA SOURCES: MEDLINE (1966-September 2003) was searched for primary and review articles. Data from the manufacturer were also included. Key words included adverse effects, clinical trials, drug interactions, gemifloxacin, and pharmacokinetic parameters. STUDY SELECTION AND DATA EXTRACTION: All articles and product labeling concerning gemifloxacin, a fluoroquinolone antibiotic recently approved by the Food and Drug Administration for treatment of CAP and AECB, were included for review. DATA SYNTHESIS: Compared with currently available fluoroquinolones, gemifloxacin demonstrated improved in vitro activity against Streptococcus pneumoniae (minimum inhibitory concentration for 90% eradication 0.03 microg/mL) and similar activity against gram-negative respiratory pathogens (Haemophilus influenzae, Moraxella catarrhalis) and atypical pathogens such as Chlamydia pneumoniae, Legionella pneumophila, and Mycoplasma pneumoniae. Gemifloxacin, consistent with other available fluoroquinolones, has insufficient activity against methicillin-resistant Staphylococcus aureus to allow clinical use for such infections. Gemifloxacin has adequate bioavailability and a favorable drug interaction profile. Gemifloxacin was comparable to commonly employed nonfluoroquinolone regimens for treatment of CAP and AECB, although the studies were designed to demonstrate equivalence. Gemifloxacin once daily for 5-7 days was well tolerated in controlled and uncontrolled clinical studies. Available clinical data, however, are insufficient to draw clinical or toxicologic distinctions between gemifloxacin and other fluoroquinolones. CONCLUSIONS: Gemifloxacin may be a suitable choice for empiric treatment of CAP or AECB. However, due to the significant history of fluoroquinolone-induced hepatic failure and dermatologic complications, the use of this drug should be closely monitored.

We have examined the antipneumococcal activities of novel quinolone dimers in which ciprofloxacin was tethered to itself or to pipemidic acid by linkage of C-7 piperazinyl rings. Symmetric 2,6-lutidinyl- and trans-butenyl-linked ciprofloxacin dimers (dimers 1 and 2, respectively) and a pipemidic acid-ciprofloxacin dimer (dimer 3) had activities against Streptococcus pneumoniae strain 7785 that were comparable to that of ciprofloxacin, i.e., MICs of 2, 1, and 4 to 8 microg/ml versus an MIC of 1 to 2 microg/ml, respectively. Surprisingly, unlike ciprofloxacin (which targets topoisomerase IV), several lines of evidence revealed that the dimers act through gyrase in S. pneumoniae. First, ciprofloxacin-resistant parC mutants of strain 7785 remained susceptible to dimers 1 to 3, whereas a gyrA mutation conferred a four- to eightfold increase in the dimer MIC but had little effect on ciprofloxacin activity. Second, dimer 1 selected first-step gyrA (S81Y or S81F) mutants (MICs, 8 to 16 microg/ml) that carried wild-type topoisomerase IV parE-parC genes. Third, dimers 1 and 2 promoted comparable DNA cleavage by S. pneumoniae gyrase and topoisomerase IV, whereas ciprofloxacin-mediated cleavage was 10-fold more efficient with topoisomerase IV than with gyrase. Fourth, the GyrA S81F and ParC S79F enzymes were resistant to dimers, confirming that the resistance phenotype is largely silent in parC mutants. Although a dimer molecule could bind very tightly by bridging quinolone binding sites in the enzyme-DNA complex, the greater potency of ciprofloxacin against gyrase and topoisomerase IV suggests that dimers 1 to 3 bind in a monomeric fashion. The bulky C-7 side chain may explain dimer targeting of gyrase and activity against efflux mutants. Tethered quinolones have potential as mechanistic tools and as novel antimicrobial agents.

Quinolones are widely used in the treatment of respiratory infections, in large part because of their activity against Streptococcus pneumoniae and other commonly encountered respiratory tract pathogens. Pneumococcal isolates that are resistant to these "respiratory quinolones" have now begun to emerge. Resistance is attributable to mutations affecting the intracellular targets of these drugs, topoisomerase IV and DNA gyrase; drug efflux contributes to quinolone resistance in some isolates. Most commonly, strains fully resistant to the newer quinolones have one or more mutations affecting DNA gyrase and topoisomerase IV. Although various agents of this class exhibit selectivity in primarily targeting one or the other of these enzymes, the passage of isolates in the presence of any agent can result in selection of mutations affecting both enzymes. Quinolone resistance in S. pneumoniae has arisen in heterogeneous genetic backgrounds but, ominously, has now appeared in strains that are well adapted for regional and global transmission.

This review focuses on the activity of gemifloxacin, a new respiratory fluoroquinolone, against the two most important bacterial pathogens associated with lower respiratory tract infections, namely Streptococcus pneumoniae and Haemophilus influenzae.

Moxifloxacin has enhanced potency against Staphylococcus aureus, lower propensity to select for resistant mutants, and higher bactericidal activity against highly resistant strains than ciprofloxacin. Despite similar activity against purified S. aureus topoisomerase IV and DNA gyrase, it selects for topoisomerase IV mutants, making topoisomerase IV the preferred target in vivo.

With the continuing development of clinical drug resistance among bacteria and the advent of resistance to the recently released agents quinupristin-dalfopristin and linezolid, the need for new, effective agents to treat multi-drug-resistant Gram-positive infections remains important. This review focuses on agents presently in clinical development for the treatment of serious multidrug-resistant staphylococcal, enterococcal and pneumococcal infections, including methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci and penicillin-resistant Streptococcus pneumoniae. Agents to be discussed that affect the prokaryotic cell wall include the antimethicillin-resistant S. aureus cephalosporins BAL9141 and RWJ-54428, the glycopeptides oritavancin and dalbavancin and the lipopeptide daptomycin. Topoisomerase inhibitors include the fluoroquinolones gemifloxacin, sitafloxacin and garenoxacin. Protein synthesis inhibitors are represented by the ketolides telithromycin and cethromycin, the oxazolidinones and the glycylcycline tigecycline. Although each of these compounds has demonstrated antibacterial activity against antibiotic-resistant pathogens, their final regulatory approval will depend on an acceptable clinical safety profile.