Repurposing β-Lactams for the Treatment of Mycobacterium kansasii Infections: An In Vitro Study

Abstract

Mycobacterium kansasii (Mkn) causes tuberculosis-like lung infection in both immunocompetent and immunocompromised patients. Current standard therapy against Mkn infection is lengthy and difficult to adhere to. Although β-lactams are the most important class of antibiotics, representing 65% of the global antibiotic market, they have been traditionally dismissed for the treatment of mycobacterial infections, as they were considered inactive against mycobacteria. A renewed interest in β-lactams as antimycobacterial agents has shown their activity against several mycobacterial species, including M. tuberculosis, M. ulcerans or M. abscessus; however, information against Mkn is lacking. In this study, we determined the in vitro activity of several β-lactams against Mkn. A selection of 32 agents including all β-lactam chemical classes (penicillins, cephalosporins, carbapenems and monobactams) with three β-lactamase inhibitors (clavulanate, tazobactam and avibactam) were evaluated against 22 Mkn strains by MIC assays. Penicillins plus clavulanate and first- and third-generation cephalosporins were the most active β-lactams against Mkn. Combinatorial time-kill assays revealed favorable interactions of amoxicillin-clavulanate and cefadroxil with first-line Mkn treatment. Amoxicillin-clavulanate and cefadroxil are oral medications that are readily available, and well tolerated with an excellent safety and pharmacokinetic profile that could constitute a promising alternative option for Mkn therapy.

Keywords: Mycobacterium kansasii; amoxicillin–clavulanate; cefadroxil; nontuberculous mycobacteria; repurposing; β-lactam combinations.

Figure 1

Heat map representation of the activity of thirteen selected β-lactams against Mkn clinical strains. MIC values (mg/L) were calculated in the presence/absence of a fixed 4 mg/L dose of clavulanate and avibactam in Middlebrook 7H9 broth plus ADC. Clavulanate was the most effective β-lactamase inhibitor. Amoxicillin–clavulanate and cefadroxil were the more active beta-lactams. PEN: penicillin; AMP: ampicillin; AMX: amoxicillin. CFX: cefadroxil; CXM: cefuroxime; CFD: cefdinir; CAZ: ceftazidime; CEF: cefepime. IMI: imipenem; MER: meropenem; ERT: ertapenem; DOR: doripenem. AZT: aztreonam. CLV: clavulanate; AVI: avibactam. nd: not determined.

Figure 2

MIC distribution of amoxicillin, cefadroxil and meropenem tested in presence/absence of clavulanate against a panel of Mkn clinical isolates. MIC (mg/L) values were determined in Middlebrook 7H9 broth plus ADC. Clavulanate was added at a fixed 4 mg/L dose. AMX: amoxicillin; CFX: cefadroxil; MER: meropenem; CLV: clavulanate.

Figure 3

Dose-response time-kill assays of β-lactams against Mkn. (A) Amoxicillin, (B) cefadroxil and (C) meropenem were tested alone and in combination with clavulanate against the Mkn ATCC 12478 strain in Middlebrook 7H9 broth plus ADC. Two positive growth control cultures were included: one with no antibiotics (CONTROL) and one containing only clavulanate (CONTROL/CLV). Data represent one experiment of at least two independent experiments performed in duplicate. Clavulanate was added at a fixed dose of 4 mg/L. AMX: amoxicillin; CFX: cefadroxil; MER: meropenem; CLV: clavulanate.

Figure 4

Time-kill assays of amoxicillin–clavulanate in combination with standard therapies against Mkn ATCC 12478. (A) Single drugs. (B) Pairwise combinations of amoxicillin–clavulanate with either rifampicin, ethambutol, isoniazid or clarithromycin. (C) Standard triple therapies against Mkn and quadruple combinations including amoxicillin–clavulanate. Time-kill assays were performed in Middlebrook 7H9 broth plus ADC. MIC values used were: AMX: 8 mg/L; RIF: 0.125 mg/L; EMB: 4 mg/L; CLA: 0.25 mg/L and INH: 8 mg/L. Clavulanate was added at a fixed dose of 4 mg/L. A drug-free positive control culture was included (CONTROL). Data represents one experiment of at least two independent experiments performed in duplicate. AMX/CLV: amoxicillin–clavulanate; RIF: rifampicin, EMB: ethambutol, INH: isoniazid, CLA: clarithromycin.

Figure 5

Time-kill assays of cefadroxil in combination with standard therapies against Mkn ATCC 12478. (A) Single drugs. (B) Pairwise combinations of cefadroxil with either rifampicin, ethambutol, isoniazid or clarithromycin. (C) Standard triple therapies against Mkn and quadruple combinations including cefadroxil. Time-kill assays were performed in Middlebrook 7H9 broth plus ADC. MIC values used were: CFX: 8 mg/L; RIF: 0.125 mg/L; EMB: 4 mg/L; CLA: 0.25 mg/L and INH: 8 mg/L. A positive control with no antibiotics was included (CONTROL). Data represent one experiment of at least two independent experiments performed in duplicate. CFX: cefadroxil; RIF: rifampicin, EMB: ethambutol, INH: isoniazid, CLA: clarithromycin.