Research article

In vitro prediction of stop-codon suppression by intravenous gentamicin in patients with cystic fibrosis: a pilot study

Isabelle Sermet-Gaudelus^1,2 , Michel Renouil³ , Anne Fajac⁴ , Laure Bidou^5,6 , Bastien Parbaille^5,6 , Sébastien Pierrot⁷ , Nolwen Davy³ , Elise Bismuth³ , Philippe Reinert⁸ , Gérard Lenoir¹ , Jean François Lesure⁹ , Jean Pierre Rousset^5,6 and Aleksander Edelman^2,10

¹Centre de Ressources et de Compétence en Mucoviscidose, Hôpital Necker-Enfants Malades, AP-HP, Paris, France

²INSERM, U806, Paris, France

³Centre de Ressources et de Compétence en Mucoviscidose, Groupe Hospitalier Sud Réunion, Saint Pierre, France

⁴Service d'Histologie-Biologie Tumorale, Hôpital Tenon, AP-HP, UPRES EA 3499, Université Pierre et Marie Curie, Paris, France

⁵IGM, Université Paris-Sud, UMR 8621, Orsay, France

⁶CNRS, Orsay, France

⁷Service d'ORL, Hôpital Necker-Enfants Malades, AP-HP, Paris, France

⁸Centre de Ressources et de Compétence en Mucoviscidose, Centre Hospitalier Intercommunal, 40 Avenue de Verdun, Créteil, France

⁹Centre de Ressources et de Compétence en Mucoviscidose, Hôpital d'Enfants, Saint Denis, France

¹⁰Université Paris-Descartes, Faculté de Médecine René Descartes, Paris, France

author email corresponding author email

BMC Medicine 2007, 5:5doi:10.1186/1741-7015-5-5

Published:	29 March 2007

Abstract

Background

Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) protein, which acts as a chloride channel activated by cyclic AMP (cAMP). The most frequent mutation found in 70% of CF patients is F508del, while premature stop mutations are found in about 10% of patients. In vitro aminoglycoside antibiotics (e.g. gentamicin) suppress nonsense mutations located in CFTR permitting translation to continue to the natural termination codon. Pharmacologic suppression of stop mutations within the CFTR may be of benefit to a significant number of patients. Our pilot study was conducted to determine whether intravenous gentamicin suppresses stop codons in CF patients and whether it has clinical benefits.

Methods

A dual gene reporter system was used to determine the gentamicin-induced readthrough level of the most frequent stop mutations within the CFTR in the French population. We investigated readthrough efficiency in response to 10 mg/kg once-daily intravenous gentamicin perfusions in patients with and without stop mutations. Respiratory function, sweat chloride concentration, nasal potential difference (NPD) and CFTR expression in nasal epithelial cells were measured at baseline and after 15 days of treatment.

Results

After in vitro gentamicin incubation, the readthrough efficiency for the Y122X mutation was at least five times higher than that for G542X, R1162X, and W1282X. In six of the nine patients with the Y122X mutation, CFTR immunodetection showed protein at the membrane of the nasal epithelial cells and the CFTR-dependent Cl^- secretion in NPD measurements increased significantly. Respiratory status also improved in these patients, irrespective of the gentamicin sensitivity of the bacteria present in the sputum. Mean sweat chloride concentration decreased significantly and normalised in two patients. Clinical status, NPD and sweat Cl^- values did not change in the Y122X patients with no protein expression, in patients with the other stop mutations investigated in vitro and those without stop mutations.

Conclusion

Suppression of stop mutations in the CFTR gene with parenteral gentamicin can be predicted in vitro and is associated with clinical benefit and significant modification of the CFTR-mediated Cl^- transport in nasal and sweat gland epithelium.