Interactions between HMR 3647, a New Ketolide, and Human Polymorphonuclear Neutrophils

Abstract

HMR 3647, a new ketolide, is active upon intracellular pathogens. We previously demonstrated that HMR 3004 (RU 64004), another ketolide, is highly concentrated by human polymorphonuclear neutrophils (PMNs). This prompted us to evaluate whether the presence of a 3-keto group instead of an l-cladinose, a neutral sugar characteristic of erythromycin A derivatives, confers peculiar pharmacokinetic properties with regard to cellular accumulation and efflux. After incubation with the radiolabelled drug, HMR 3647 uptake was determined by a velocity gradient centrifugation technique. HMR 3647 was avidly concentrated by PMNs, without saturation, over a 3-h incubation period, with cellular-to-extracellular concentration ratios of 31 ± 4.2 at 5 min and up to 348 ± 27.1 at 180 min. About 60% of HMR 3647 was located in the granular compartment; less than 6% was associated with the membranes. HMR 3647 gradually egressed from loaded cells placed in drug-free medium. Uptake was dependent on environmental temperature (activation energy, 128 ± 9.4 kJ/mol) but not on extracellular pH. HMR 3647 displayed Michaelis-Menten saturation kinetics with a mean Vmax of 2315 ng/2.5 × 106 PMNs/5 min and a mean Km of 117 mg/liter (144 μM). As already observed with erythromycin A-derived macrolides, extracellular Ca2+ was necessary for optimal uptake of HMR 3647. Interestingly, verapamil increased the uptake of HMR 3647 at 5 min, but this was followed by gradual inhibition at later incubation times, a phenomenon probably related to stimulation of drug efflux. The impact of intracellular accumulation of HMR 3647 on PMN functions was also investigated. In contrast to other erythromycin A derivatives, HMR 3647 only weakly triggered granule exocytosis, but it inhibited superoxide anion production in a time- and concentration-dependent manner, with concentrations which inhibited 50% of control response of 55 (67 μM) (5 min) and 30 (36 μM) (30 min) mg/liter for formyl-methionyl-leucyl-phenylalanine stimulation and 117 (143 μM) (5 min) and 44 (54 μM) (30 min) mg/liter for phorbol myristate acetate stimulation.

P‐Glycoprotein‐Dependent Trafficking of Nanoparticle‐Drug Conjugates

“…In contrast, while P-gp-dependent cell accumulation of macrolide-AuNRs was not observed in P-gp-null COLO 205 cells, accumulation of Azith- and Clarith-AuNR was significantly increased in P-gp(+) J774.2 cells following competitive inhibition, with no significant changes in TriKeto-AuNR accumulation. These findings agree well with previous reports indicating i) macrolide-competitive P-gp binding by verapamil and cyclosporine, [16] ii) diminished recognition of TriKeto (TE-802) by P-gp, [9f] iii) enhanced in vivo efficacy of third-generation tricyclic ketolides, [9d,9e] and iv) P-gp-dependent accumulation/cytotoxicity of free macrolide ligands (Supporting Data, Figure S3). These data suggest that P-gp-dependent trafficking can significantly affect the cellular accumulation of nanoscale drug carriers to which P-gp substrates are appended (e.g.…”

Activity of telithromycin and comparators against isolates of Legionella pneumophila collected from patients with community-acquired respiratory tract infections: PROTEKT Years 1–5

“…Telithromycin is active in vitro against typical community‐acquired respiratory tract infection pathogens, including penicillin‐ and macrolide‐resistant strains of Streptococcus pneumoniae [7]. Telithromycin is concentrated within polymorphonuclear neutrophils [8] and shows high‐level activity against atypical intracellular pathogens, including L. pneumophila [9].…”

Uptake and intracellular activity of ketolide HMR 3647 in human phagocytic and non-phagocytic cells

“…HMR 3647 is strongly accumulated by human polymorphonuclear neutrophils (PMNs), without saturation, and shows a slow efflux. Uptake depends on environmental temperature, not extracellular pH, and also impairs oxidant production by PMNs in a time‐dependent manner [9]. Most intracellular bacteria, however, multiply or survive within macrophages.…”