<i>In Vitro</i>and<i>In Vivo</i>Antibacterial Evaluation of Cadazolid, a New Antibiotic for Treatment of Clostridium difficile Infections

We studied the molecular mechanisms of linezolid resistance in 9 isolates of toxigenic Clostridium difficile with high linezolid MICs. The activity of linezolid was determined against 891 clinical isolates of toxigenic C. difficile. The MIC 50 and MIC 90 of linezolid were 0.75 g/ml and 1.5 g/ml, respectively. Nine strains (1%) showed high linezolid MICs (6 g/ml to 16 g/ml) and also were resistant to clindamycin, erythromycin, and chloramphenicol. These strains were selected for molecular studies: sequencing of domain V of the 23 rRNA gene, detection of the cfr methyltransferase gene, and sequencing of the ribosomal protein genes rplC and rplD. Molecular relatedness between strains was assessed using PCR ribotyping and MLVA (multilocus variablenumber tandem-repeat analysis) typing. The strains belonged to ribotypes 001 (2/9), 017 (6/9), and 078 (1/9). MLVA showed that strains of ribotype 001 and 017 belonged to the same clonal complex in each ribotype. We did not detect mutations in the 23S rRNA gene. The cfr gene was detected in 7 of 9 strains. Sequencing of cfr amplicons revealed a similarity of 100% to a fragment of transposon Tn6218 of C. difficile, which was annotated as a putative chloramphenicol/florfenicol resistance protein. We were unable to detect mechanisms of resistance to linezolid in the 2 strains belonging to ribotype 001. While the relevance of our results lies in the detection of the cfr gene as a possible mechanism of resistance to linezolid in C. difficile, our findings should be assessed by further investigations to characterize these possible cfr genes and their contribution to linezolid resistance. Linezolid is the first antibiotic in the oxazolidinone group and has been used for more than 10 years to treat Gram-positive infections (1). Resistance to linezolid in Gram-positive bacteria is very uncommon and has been reported mainly in clinical isolates of coagulase-negative staphylococcus, Staphylococcus aureus, Enterococcus faecalis, and Enterococcus faecium, as well as in nonhuman isolates of other Gram-positive bacteria and Gram-negative bacteria (2). Resistance is due to the presence of 1 or more of 3 mechanisms: point mutations in the 23S rRNA gene (central loop of domain V, mainly G2576T), mutations/deletions in ribosomal proteins L3 (rplC gene) and L4 (rplD gene), and methylation of position A2503 of the 23S rRNA gene mediated by an rRNA methyltransferase encoded by the multiresistance gene cfr (chloramphenicol-florfenicol resistance), which is harbored mainly in transferable plasmids (3). This last mechanism also has been associated with resistance to phenicols, lincosamides, oxazolidinones, pleuromutilins, and streptogramin A antibiotics (4).Clostridium difficile is the main cause of antibiotic-associated diarrhea. Although linezolid is not used for the treatment of C. difficile infection (5, 6), some observations suggest that patients treated with linezolid for ventilator-associated pneumonia could be protected against this infection (7).Resistance to linezolid occasionally has bee...

Section: Resultsmentioning

confidence: 99%

Clostridium difficile Isolates with High Linezolid MICs Harbor the Multiresistance Gene cfr

Marı́n

Martín

Alcalá

et al. 2015

“…In this study, recurrence rates were lower with all cadazolid dosages than with vancomycin regardless of the criteria applied for analysis, which in turn resulted in higher sustained clinical response rates in patients receiving cadazolid than in those receiving vancomycin. Should this observation be confirmed by statistically significant results on sustained clinical response in the ongoing phase 3 studies, it could be explained by the inhibitory effect of cadazolid on C. difficile toxin synthesis and spore formation and the preservation of normal gut microbiota (10,11,15).…”

Section: Discussionmentioning

confidence: 99%

“…In vitro, cadazolid demonstrates potent activity against C. difficile, including the BI/NAP1/ 027 strain, with a low propensity for resistance development (10)(11)(12)(13). In cultures of toxigenic C. difficile, cadazolid strongly inhibits de novo formation of toxins A and B, the main virulence factors of C. difficile, and prevents in vitro C. difficile spore formation at sub-growth-inhibitory concentrations (11). In healthy male patients, single and twice-daily (BID) ascending oral doses of 30 to 3,000 mg cadazolid resulted in very low systemic exposure, with the majority of cadazolid being excreted unchanged in the feces (14).…”

mentioning

confidence: 99%

Multicenter, Double-Blind, Randomized, Phase 2 Study Evaluating the Novel Antibiotic Cadazolid in Patients with Clostridium difficile Infection

Louie

Nord

Talbot

et al. 2015

C lostridium difficile infection (CDI), the main cause of nosocomial infectious diarrhea, results from the growth of toxinproducing C. difficile in the colon following disruption of the normal enteric microbiota, usually as a consequence of antibiotic therapy (1). The frequency and severity of CDI have risen over the past decade, with associated increases in morbidity and mortality, especially among the elderly (2, 3). The increase in CDI has been attributed, at least in part, to the epidemic C. difficile BI/NAP1/027 strain, first reported in 2005 (1, 4). Current treatment of CDI includes metronidazole, vancomycin, or fidaxomicin, with cure rates of approximately 86 to 95% (5-7); however, 15 to 40% of patients experience recurrence following clinical cure (6-9). Reducing recurrence rates, together with improving outcomes for those severely affected by CDI, remains a substantial unmet medical need.Cadazolid is a novel, nonabsorbable antibiotic that acts by inhibiting bacterial protein synthesis. In vitro, cadazolid demonstrates potent activity against C. difficile, including the BI/NAP1/ 027 strain, with a low propensity for resistance development (10)(11)(12)(13). In cultures of toxigenic C. difficile, cadazolid strongly inhibits de novo formation of toxins A and B, the main virulence factors of C. difficile, and prevents in vitro C. difficile spore formation at sub-growth-inhibitory concentrations (11). In healthy male patients, single and twice-daily (BID) ascending oral doses of 30 to 3,000 mg cadazolid resulted in very low systemic exposure, with the majority of cadazolid being excreted unchanged in the feces (14). In a human gut model, cadazolid demonstrated narrowspectrum activity, eliminating CDI while having a very limited impact on the normal gut microbiota (15), which, together with preventing the formation of C. difficile spores, may indicate that cadazolid has the potential to reduce CDI recurrence.Here, we report the results of a phase 2 study investigating the efficacy and safety of three oral dosages of cadazolid, with vancomycin as an active reference, in patients with CDI.

“…However, clinical data are needed to confirm the overall low propensity of resistance development of cadazolid. Results of in vitro and in vivo evaluations of cadazolid are reported in a companion article by Locher et al (26). Tubes containing 2 ml of 2-fold serial dilutions of the test antibiotics were inoculated with approximately 5 ϫ 10 6 to 2 ϫ 10 7 CFU.…”

Section: Discussionmentioning

confidence: 99%

Investigations of the Mode of Action and Resistance Development of Cadazolid, a New Antibiotic for Treatment of Clostridium difficile Infections

Locher

Caspers

Bruyère

et al. 2014

Self Cite

Cadazolid is a new oxazolidinone-type antibiotic currently in clinical development for the treatment of Clostridium difficileassociated diarrhea. Here, we report investigations on the mode of action and the propensity for spontaneous resistance development in C. difficile strains. Macromolecular labeling experiments indicated that cadazolid acts as a potent inhibitor of protein synthesis, while inhibition of DNA synthesis was also observed, albeit only at substantially higher concentrations of the drug. Strong inhibition of protein synthesis was also obtained in strains resistant to linezolid, in agreement with low MICs against such strains. Inhibition of protein synthesis was confirmed in coupled transcription/translation assays using extracts from different C. difficile strains, including strains resistant to linezolid, while inhibitory effects in DNA topoisomerase assays were weak or not detectable under the assay conditions. Spontaneous resistance frequencies of cadazolid were low in all strains tested (generally <10؊10 at 2؋ to 4؋ the MIC), and in multiple-passage experiments (up to 13 passages) MICs did not significantly increase. Furthermore, no cross-resistance was observed, as cadazolid retained potent activity against strains resistant or nonsusceptible to linezolid, fluoroquinolones, and the new antibiotic fidaxomicin. In conclusion, the data presented here indicate that cadazolid acts primarily by inhibition of protein synthesis, with weak inhibition of DNA synthesis as a potential second mode of action, and suggest a low potential for spontaneous resistance development.