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Suppression of CFTR premature termination codons and rescue of CFTR protein and function by the synthetic aminoglycoside NB54

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Abstract

Certain aminoglycosides are capable of inducing “translational readthrough” of premature termination codons (PTCs). However, toxicity and relative lack of efficacy deter treatment with clinically available aminoglycosides for genetic diseases caused by PTCs, including cystic fibrosis (CF). Using a structure-based approach, the novel aminoglycoside NB54 was developed that exhibits reduced toxicity and enhanced suppression of PTCs in cell-based reporter assays relative to gentamicin. We examined whether NB54 administration rescued CFTR protein and function in clinically relevant CF models. In a fluorescence-based halide efflux assay, NB54 partially restored halide efflux in a CF bronchial epithelial cell line (CFTR genotype W1282X/F508del), but not in a CF epithelial cell line lacking a PTC (F508del/F508del). In polarized airway epithelial cells expressing either a CFTR-W1282X or -G542X cDNA, treatment with NB54 increased stimulated short-circuit current (I SC) with greater efficiency than gentamicin. NB54 and gentamicin induced comparable increases in forskolin-stimulated I SC in primary airway epithelial cells derived from a G542X/F508del CF donor. Systemic administration of NB54 to Cftr−/− mice expressing a human CFTR-G542X transgene restored 15–17% of the average stimulated transepithelial chloride currents observed in wild-type (Cftr+/+) mice, comparable to gentamicin. NB54 exhibited reduced cellular toxicity in vitro and was tolerated at higher concentrations than gentamicin in vivo. These results provide evidence that synthetic aminoglycosides are capable of PTC suppression in relevant human CF cells and a CF animal model and support further development of these compounds as a treatment modality for genetic diseases caused by PTCs.

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Abbreviations

AHB:

(S)-4-Amino-2 hydroxybutanoyl

CF:

Cystic fibrosis

CFTR:

Cystic fibrosis transmembrane conductance regulator

I SC :

Short-circuit current

PTC:

Premature termination codons

SPQ:

6-Methoxy-N-(3-sulfopropyl)-quinolinium

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Acknowledgments

The authors thank Kim Keeling for critically reading the manuscript. This study was supported by US National Institute of Health grants 1K23DK075788-01 and 1R03DK084110-01 (to SMR), 1P30DK072482-01A1 and Cystic Fibrosis Foundation grant R464-CR02 (to EJS), US-Israel Binational Science Foundation grant 2006/301 and Mitchel Fund 2012386 (to TB), and Horowitz Fund (grant no. 2010826 to TB and DMB). VB acknowledges the financial support by the Center of Absorption in Science, the Ministry of Immigration Absorption, and the Ministry of Science and Technology, Israel (Kamea Program).

Disclosure statement

SMR serves on the advisory board for PTC Therapeutics, Inc. DMB serves as a consultant for PTC Therapeutics, Inc.

Author information

Authors and Affiliations

  1. Department of Medicine, University of Alabama at Birmingham, Birmingham, AL, USA

    Steven M. Rowe, Peter Sloane, Li Ping Tang & Kyle Backer

  2. Department of Pediatrics, University of Alabama at Birmingham, Birmingham, AL, USA

    Steven M. Rowe

  3. Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, AL, USA

    Steven M. Rowe & David M. Bedwell

  4. Department of Microbiology, University of Alabama at Birmingham, Birmingham, AL, USA

    Jessica Buckley-Lanier & David M. Bedwell

  5. Department of Cell Biology, University of Alabama at Birmingham, Birmingham, AL, USA

    Zsuzsa Bebok

  6. Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, Birmingham, AL, USA

    Steven M. Rowe, Peter Sloane, Li Ping Tang, Kyle Backer, Marina Mazur, Zsuzsa Bebok & David M. Bedwell

  7. The Edith and Joseph Fisher Enzyme Inhibitors Laboratory, Schulich Faculty of Chemistry, Technion-Israel Institute of Technology, Haifa, 32000, Israel

    Igor Nudelman, Valery Belakhov & Timor Baasov

  8. DiscoveryBioMed, Inc., Innovation Depot, Birmingham, AL, USA

    Erik Schwiebert

  9. University of Alabama at Birmingham, MCLM 768, 1500 3rd Ave South, Birmingham, AL, 35294-0005, USA

    Steven M. Rowe

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  1. Steven M. Rowe
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  2. Peter Sloane
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  12. David M. Bedwell
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Corresponding author

Correspondence to Steven M. Rowe.

Additional information

This research was funded by the US National Institute of Health grants 1K23DK075788-01 and 1R03DK084110-01 (to SMR), 1P30DK072482-01A1 and Cystic Fibrosis Foundation grant R464-CR02 (to EJS), US-Israel Binational Science Foundation grant 2006/301 and Mitchel Fund 2012386 (to TB), and Horowitz Fund (grant no. 2010826 to TB and DMB). VB acknowledges the financial support by the Center of Absorption in Science, the Ministry of Immigration Absorption, and the Ministry of Science and Technology, Israel (Kamea Program). We are grateful to Cheryl Owens for assistance with preparation of the manuscript.

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Rowe, S.M., Sloane, P., Tang, L.P. et al. Suppression of CFTR premature termination codons and rescue of CFTR protein and function by the synthetic aminoglycoside NB54. J Mol Med 89, 1149–1161 (2011). https://doi.org/10.1007/s00109-011-0787-6

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  • DOI: https://doi.org/10.1007/s00109-011-0787-6

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