Azithromycin blocks autophagy and may predispose cystic fibrosis patients to mycobacterial infection
Maurizio Renna, … , David C. Rubinsztein, R. Andres Floto
Maurizio Renna, … , David C. Rubinsztein, R. Andres Floto
Published August 1, 2011
Citation Information: J Clin Invest. 2011;121(9):3554-3563. https://doi.org/10.1172/JCI46095.
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Research Article Immunology

Azithromycin blocks autophagy and may predispose cystic fibrosis patients to mycobacterial infection

Abstract

Azithromycin is a potent macrolide antibiotic with poorly understood antiinflammatory properties. Long-term use of azithromycin in patients with chronic inflammatory lung diseases, such as cystic fibrosis (CF), results in improved outcomes. Paradoxically, a recent study reported that azithromycin use in patients with CF is associated with increased infection with nontuberculous mycobacteria (NTM). Here, we confirm that long-term azithromycin use by adults with CF is associated with the development of infection with NTM, particularly the multi-drug-resistant species Mycobacterium abscessus, and identify an underlying mechanism. We found that in primary human macrophages, concentrations of azithromycin achieved during therapeutic dosing blocked autophagosome clearance by preventing lysosomal acidification, thereby impairing autophagic and phagosomal degradation. As a consequence, azithromycin treatment inhibited intracellular killing of mycobacteria within macrophages and resulted in chronic infection with NTM in mice. Our findings emphasize the essential role for autophagy in the host response to infection with NTM, reveal why chronic use of azithromycin may predispose to mycobacterial disease, and highlight the dangers of inadvertent pharmacological blockade of autophagy in patients at risk of infection with drug-resistant pathogens.

Authors

Maurizio Renna, Catherine Schaffner, Karen Brown, Shaobin Shang, Marcela Henao Tamayo, Krisztina Hegyi, Neil J. Grimsey, David Cusens, Sarah Coulter, Jason Cooper, Anne R. Bowden, Sandra M. Newton, Beate Kampmann, Jennifer Helm, Andrew Jones, Charles S. Haworth, Randall J. Basaraba, Mary Ann DeGroote, Diane J. Ordway, David C. Rubinsztein, R. Andres Floto

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Figure 6

Azithromycin blocks intracellular killing of mycobacteria.

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Azithromycin blocks intracellular killing of mycobacteria. (A and B) Hum...
(A and B) Human macrophages were infected with luminescent mycobacteria — M. bovis BCG (A) and M. abscessus (B) — for 2 hours, washed, and exposed to the indicated concentrations of azithromycin (μg/ml) and/or 200 nM rapamycin for 24 hours. Viable intracellular mycobacteria were then assessed by measuring cell-associated luminescence after cell lysis. (C) IFN-γ and TNF-α enhanced intracellular killing of M. abscessus in human macrophages, and this was blocked by azithromycin pretreatment. Results were normalized to levels obtained without cytokine addition. (D) Patient-derived M. abscessus strains were incubated with peripheral blood from healthy subjects pretreated for 24 hours with vehicle alone or increasing concentrations of azithromycin. After 72 hours of incubation at 37°C, viable mycobacteria were quantified after sample lysis by counting CFUs. Addition of azithromycin, while reducing mycobacterial growth (white), prevented any additional autophagy-dependent killing achieved by coincubation with IFN-γ (200 U/ml) or rapamycin (100 nM). (E) Induction of macrolide resistance in M. abscessus. M. abscessus-lux was grown for 6 days in liquid culture (i.e., Middlebrook 7H9 plus ADC enrichment) in the presence of 0.1 μg/ml azithromycin or vehicle alone, washed, and resuspended in RPMI with 10% FCS with the indicated concentrations of azithromycin. Viable mycobacteria were assessed by measuring luminescence after 24 hours. *P < 0.05; **P < 0.005.