Unité de pharmacologie cellulaire et moléculaire, Université catholique de Louvain, 73.70 avenue E. Mounier 73, B-1200 Brussels, Belgium
Keywords: L. monocytogenes , listeriosis
Sir,
Listeriosis is one of the potential adverse effects of TNF--neutralizing treatments.1 Glatiramer (copolymer 1; COPAXONE), a mixed, random polymer of Ala, Glu, Lys and Tyr used in the treatment of relapsingremitting multiple sclerosis2 blocks the secretion of TNF-
from IFN-
- and endotoxin-stimulated THP-1 macrophages.3 We have, therefore, examined the influence of glatiramer on the ability of IFN-
to contain Listeria infection and on the activity of ampicillin and moxifloxacin to kill intraphagocytic bacteria in THP-1 macrophages. Glatiramer [CAS Registry no. 147 245-92-9; batch no. 242908102: average molecular weight, 7500 Da (limits, 420016 350); amino acid content (molecular fraction) L-Glu, 0.139; L-Ala, 0.432; L-Tyr, 0.091; L-Lys, 0.338; total amino acid residue content, 87.9%; bacterial endotoxin content, <0.25 endotoxin units/mg] was kindly received from Teva Pharmaceuticals Industries (Petah Tiqua, Israel). All experimental procedures and assay methods have been described in our previous publications.4,5 We used a concentration of glatiramer of 20 mg/L, which was both non-toxic (based on lactate dehydrogenase release) and effective in blocking the production of TNF-
in THP-1 cells.3
Glatiramer (20 mg/L) did not influence the intrinsic antimicrobial activity of ampicillin or moxifloxacin towards L. monocytogenes, based on MIC determinations in broth [0.3±0.1 and 0.5±0.1 mg/L (arithmetic dilutions) for ampicillin and moxifloxacin, respectively]. Unstimulated cells produced only negligible amounts of TNF- and glatiramer did not alter this behaviour. In contrast, the medium of cells exposed to IFN-
(100 units/mL; 24 h) contained 38.3±6.0 ng/L of TNF-
, and this concentration was decreased by
2/3 in the presence of glatiramer. With infected cells, TNF-
production remained low in unstimulated cells and unaffected by the presence of glatiramer, whereas it amounted to 31.6±1.2 ng/L (5 h post-phagocytosis) in IFN-
-stimulated cells (24 h prior to infection). This production was again decreased by 2/3 if glatiramer was present. Glatiramer did not significantly modify the capacity of THP-1 macrophages to phagocytose L. monocytogenes. Figure 1 shows that glatiramer did not modify the growth of intracellular L. monocytogenes compared with untreated cells in the 24 h model (this model uses gentamicin at a concentration of 2 x its MIC to prevent the extracellular growth of L. monocytogenes). No change was seen either in the 5 h model (data not shown). As previously described,6 IFN-
impaired the intracellular growth of L. monocytogenes by
50%, and glatiramer did not modify this effect. When infected cells were exposed to ampicillin for 24 h after phagocytosis, the bacterial load was reduced by
1.7 log compared with the original, post-phagocytosis inoculum. Glatiramer, IFN-
, or the combination of glatiramer and IFN-
did not significantly modify this effect of ampicillin. In the next series of experiments, we examined the activity of moxifloxacin (4 mg/L) using the 5 h model. We observed a decrease in the post-phagocytosis inoculum of 1.34±0.03, 1.26±0.16, 1.31±0.07 and 1.32±0.07 log10 units for cells treated with moxifloxacin alone, glatiramer and moxifloxacin, IFN-
and moxifloxacin, and the combination of glatiramer, IFN-
and moxifloxacin, respectively. In parallel experiments, we examined the influence of glatiramer on the accumulation of moxifloxacin and no effect was seen [apparent cellular to extracellular drug concentration ratios at 2 h of 9.6±2.0 in controls versus 9.4±1.1 in cells exposed to glatiramer (20 mg/L) during the uptake period; similar values were found for cells pre-exposed to glatiramer (20 mg/L) for 24 h]. Glatiramer did not influence the accumulation of three other quinolones (ciprofloxacin, levofloxacin and garenoxacin).
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Acknowledgements
Mrs N. Aguilera provided critical help in some of the experiments described in this paper and Ms M. C. Cambier maintained the cell culture line. S. C. was Boursier of the Belgian Fonds pour la Formation à la Recherche dans l'Industrie et l'Agriculture (F.R.I.A.), is F. V. B. Chercheur Qualifié of the Belgian Fonds National de la Recherche Scientifique (F.N.R.S.). This work was supported by the Belgian Fonds de la Recherche Scientifique Médicale (F.R.S.M.; grant no. 3.4.612.00 F). Aventis Pharma s.a., Brussels, Belgium, provided the sample of glatiramer used in this study.
Footnotes
* Corresponding author. Tel: +32-2-762-21-36; Fax: +32-2-764-73-73; Email: tulkens{at}facm.ucl.ac.be
Present address: Department of Veterinary Science and Microbiology, University of Arizona, 1117 E. Lowell, Bldg. 90, Room 306, Tucson, AZ 85721, USA
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