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Cysteamine (Lynovex®), a novel mucoactive antimicrobial & antibiofilm agent for the treatment of cystic fibrosis.

Charrier C, Rodger C, Robertson J, Kowalczuk A, Shand N, Fraser-Pitt D, Mercer D, O'Neil D - Orphanet J Rare Dis (2014)

Bottom Line: Any successful therapeutic strategy designed to combat the respiratory pathology of this condition must address the altered lung physiology and recurrent, complex, polymicrobial infections and biofilms that affect the CF pulmonary tract.In all cases, the 'gold standard' therapeutic agents were employed as control/comparator compounds against which the efficacy of cysteamine was compared.The data we present here provides a platform for cysteamine's continued investigation as a novel treatment for this poorly served orphan disease.

View Article: PubMed Central - PubMed

Affiliation: NovaBiotics Ltd, Cruickshank Building, Craibstone, Aberdeen, AB21 9TR, UK. cedric.charrier@gmail.com.

ABSTRACT

Background: There remains a critical need for more effective, safe, long-term treatments for cystic fibrosis (CF). Any successful therapeutic strategy designed to combat the respiratory pathology of this condition must address the altered lung physiology and recurrent, complex, polymicrobial infections and biofilms that affect the CF pulmonary tract. Cysteamine is a potential solution to these unmet medical needs and is described here for the first time as (Lynovex®) a single therapy with the potential to deliver mucoactive, antibiofilm and antibacterial properties; both in oral and inhaled delivery modes. Cysteamine is already established in clinical practice for an unrelated orphan condition, cystinosis, and is therefore being repurposed (in oral form) for cystic fibrosis from a platform of over twenty years of safety data and clinical experience.

Methods: The antibacterial and antibiofilm attributes of cysteamine were determined against type strain and clinical isolates of CF relevant pathogens using CLSI standard and adapted microbiological methods and a BioFlux microfluidic system. Assays were performed in standard nutrient media conditions, minimal media, to mimic the low metabolic activity of microbes/persister cells in the CF respiratory tract and in artificial sputum medium. In vivo antibacterial activity was determined in acute murine lung infection/cysteamine nebulisation models. The mucolytic potential of cysteamine was assessed against DNA and mucin in vitro by semi-quantitative macro-rheology. In all cases, the 'gold standard' therapeutic agents were employed as control/comparator compounds against which the efficacy of cysteamine was compared.

Results: Cysteamine demonstrated at least comparable mucolytic activity to currently available mucoactive agents. Cysteamine was rapidly bactericidal against both metabolically active and persister cells of Pseudomonas aeruginosa and also emerging CF pathogens; its activity was not sensitive to high ionic concentrations characteristic of the CF lung. Cysteamine prevented the formation of, and disrupted established P. aeruginosa biofilms. Cysteamine was synergistic with conventional CF antibiotics; reversing antibiotic resistance/insensitivity in CF bacterial pathogens.

Conclusions: The novel mucolytic-antimicrobial activity of cysteamine (Lynovex®) provides potential for a much needed new therapeutic strategy in cystic fibrosis. The data we present here provides a platform for cysteamine's continued investigation as a novel treatment for this poorly served orphan disease.

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Antimicrobial activity of cysteamine following nebulisation (A) and intra-tracheal dosing (B) in a mouse acute lung infection model. (A) The clinical strain P. aeruginosa EUPPA103 was administered at ~6.5 × 104 CFU/mouse by intranasal injection under temporary inhaled anesthesia. Mice were placed within sealed nebulisation chamber and exposed to cysteamine at 4.2 mg/ml for 5, 10 or 20 minutes (total 1 dose) or tobramycin at 4.2 mg/ml in aqueous solution for 10 minutes via aerosol delivery system 1 hour post-infection. Experimental endpoint was lung tissue burden 25 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue. (B) The clinical strain P. aeruginosa ATCC27853 was administered at 3 × 104 - 1 × 105 cfu/40 μl/mouse by intranasal injection under temporary inhaled anesthesia. Mice were given two doses (5 mg/kg each) of cysteamine or tobramycin delivered intratracheally 10 min and 6 h after infection. Experimental endpoint was lung tissue burden 26 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue.
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Fig8: Antimicrobial activity of cysteamine following nebulisation (A) and intra-tracheal dosing (B) in a mouse acute lung infection model. (A) The clinical strain P. aeruginosa EUPPA103 was administered at ~6.5 × 104 CFU/mouse by intranasal injection under temporary inhaled anesthesia. Mice were placed within sealed nebulisation chamber and exposed to cysteamine at 4.2 mg/ml for 5, 10 or 20 minutes (total 1 dose) or tobramycin at 4.2 mg/ml in aqueous solution for 10 minutes via aerosol delivery system 1 hour post-infection. Experimental endpoint was lung tissue burden 25 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue. (B) The clinical strain P. aeruginosa ATCC27853 was administered at 3 × 104 - 1 × 105 cfu/40 μl/mouse by intranasal injection under temporary inhaled anesthesia. Mice were given two doses (5 mg/kg each) of cysteamine or tobramycin delivered intratracheally 10 min and 6 h after infection. Experimental endpoint was lung tissue burden 26 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue.

Mentions: To assess whether the in vitro antibiofilm and antibacterial data derived in our studies translated in a more clinically relevant, complex biological setting, we employed a well-established mouse model of P. aeruginosa lung infection and determined the impact therein of cysteamine exposure on lung bacterial burdens. We also assessed how well tolerated cysteamine was when delivered via the respiratory route as to date, its safety and toxicological potential have not been determined by this mode of application. Cysteamine was well tolerated and did not cause any adverse effects when administered to mice via two respiratory routes; either by a single dose of cysteamine or tobramycin delivered by nebulisation (4.2 mg/ml for 5 min, 10 min or 20 min) following infection with P. aeruginosa EUPPA103 (Figure 8A), or two doses (5 mg/kg each) delivered intratracheally 10 min and 6 h after infection with P. aeruginosa ATCC27853 (Figure 8B). These experiments provide the first evidence of in vivo antibacterial efficacy of cysteamine in respiratory tissue. In the mice infected with P. aeruginosa EUPPA103, nebulised cysteamine (4.2 mg/ml, 5 – 20 min exposure) significantly reduced bacterial lung burden (LogR 0.94-1.11 cfu/g; P ≤ 0.001) compared to vehicle only treated mice (StatsDirect Kruskal-Wallis test: Squared ranks approximate equality of variance test). Tobramycin (4.2 mg/ml aqueous solution) treated mice also demonstrated statistically significant reduction in lung burden (LogR 1.51 CFU/g; P = 0.0 117) (StatsDirect Kruskal-Wallis test: all pairwise comparisons (Conover-Inman)). A trend to dose dependency for cysteamine was observed, with greatest efficacy demonstrated by 20 min exposure (5.48 log10 CFU/g) compared with 5 min (5.65 log10 CFU/g) and 10 min exposure (5.64 log10 CFU/g). In the mice infected with P. aeruginosa ATCC27853, intratracheal administration of cysteamine reduced bacterial burden in the lungs from 8.17 ± 0.08 log10 CFU/g (control animals) to 7.86 ± 0.18 log10 CFU/g (cysteamine-treated animals).Figure 8


Cysteamine (Lynovex®), a novel mucoactive antimicrobial & antibiofilm agent for the treatment of cystic fibrosis.

Charrier C, Rodger C, Robertson J, Kowalczuk A, Shand N, Fraser-Pitt D, Mercer D, O'Neil D - Orphanet J Rare Dis (2014)

Antimicrobial activity of cysteamine following nebulisation (A) and intra-tracheal dosing (B) in a mouse acute lung infection model. (A) The clinical strain P. aeruginosa EUPPA103 was administered at ~6.5 × 104 CFU/mouse by intranasal injection under temporary inhaled anesthesia. Mice were placed within sealed nebulisation chamber and exposed to cysteamine at 4.2 mg/ml for 5, 10 or 20 minutes (total 1 dose) or tobramycin at 4.2 mg/ml in aqueous solution for 10 minutes via aerosol delivery system 1 hour post-infection. Experimental endpoint was lung tissue burden 25 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue. (B) The clinical strain P. aeruginosa ATCC27853 was administered at 3 × 104 - 1 × 105 cfu/40 μl/mouse by intranasal injection under temporary inhaled anesthesia. Mice were given two doses (5 mg/kg each) of cysteamine or tobramycin delivered intratracheally 10 min and 6 h after infection. Experimental endpoint was lung tissue burden 26 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Fig8: Antimicrobial activity of cysteamine following nebulisation (A) and intra-tracheal dosing (B) in a mouse acute lung infection model. (A) The clinical strain P. aeruginosa EUPPA103 was administered at ~6.5 × 104 CFU/mouse by intranasal injection under temporary inhaled anesthesia. Mice were placed within sealed nebulisation chamber and exposed to cysteamine at 4.2 mg/ml for 5, 10 or 20 minutes (total 1 dose) or tobramycin at 4.2 mg/ml in aqueous solution for 10 minutes via aerosol delivery system 1 hour post-infection. Experimental endpoint was lung tissue burden 25 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue. (B) The clinical strain P. aeruginosa ATCC27853 was administered at 3 × 104 - 1 × 105 cfu/40 μl/mouse by intranasal injection under temporary inhaled anesthesia. Mice were given two doses (5 mg/kg each) of cysteamine or tobramycin delivered intratracheally 10 min and 6 h after infection. Experimental endpoint was lung tissue burden 26 h post-infection. Vehicle was sterile physiological water. The lower limit of detection was approximately 50 cfu/g of tissue.
Mentions: To assess whether the in vitro antibiofilm and antibacterial data derived in our studies translated in a more clinically relevant, complex biological setting, we employed a well-established mouse model of P. aeruginosa lung infection and determined the impact therein of cysteamine exposure on lung bacterial burdens. We also assessed how well tolerated cysteamine was when delivered via the respiratory route as to date, its safety and toxicological potential have not been determined by this mode of application. Cysteamine was well tolerated and did not cause any adverse effects when administered to mice via two respiratory routes; either by a single dose of cysteamine or tobramycin delivered by nebulisation (4.2 mg/ml for 5 min, 10 min or 20 min) following infection with P. aeruginosa EUPPA103 (Figure 8A), or two doses (5 mg/kg each) delivered intratracheally 10 min and 6 h after infection with P. aeruginosa ATCC27853 (Figure 8B). These experiments provide the first evidence of in vivo antibacterial efficacy of cysteamine in respiratory tissue. In the mice infected with P. aeruginosa EUPPA103, nebulised cysteamine (4.2 mg/ml, 5 – 20 min exposure) significantly reduced bacterial lung burden (LogR 0.94-1.11 cfu/g; P ≤ 0.001) compared to vehicle only treated mice (StatsDirect Kruskal-Wallis test: Squared ranks approximate equality of variance test). Tobramycin (4.2 mg/ml aqueous solution) treated mice also demonstrated statistically significant reduction in lung burden (LogR 1.51 CFU/g; P = 0.0 117) (StatsDirect Kruskal-Wallis test: all pairwise comparisons (Conover-Inman)). A trend to dose dependency for cysteamine was observed, with greatest efficacy demonstrated by 20 min exposure (5.48 log10 CFU/g) compared with 5 min (5.65 log10 CFU/g) and 10 min exposure (5.64 log10 CFU/g). In the mice infected with P. aeruginosa ATCC27853, intratracheal administration of cysteamine reduced bacterial burden in the lungs from 8.17 ± 0.08 log10 CFU/g (control animals) to 7.86 ± 0.18 log10 CFU/g (cysteamine-treated animals).Figure 8

Bottom Line: Any successful therapeutic strategy designed to combat the respiratory pathology of this condition must address the altered lung physiology and recurrent, complex, polymicrobial infections and biofilms that affect the CF pulmonary tract.In all cases, the 'gold standard' therapeutic agents were employed as control/comparator compounds against which the efficacy of cysteamine was compared.The data we present here provides a platform for cysteamine's continued investigation as a novel treatment for this poorly served orphan disease.

View Article: PubMed Central - PubMed

Affiliation: NovaBiotics Ltd, Cruickshank Building, Craibstone, Aberdeen, AB21 9TR, UK. cedric.charrier@gmail.com.

ABSTRACT

Background: There remains a critical need for more effective, safe, long-term treatments for cystic fibrosis (CF). Any successful therapeutic strategy designed to combat the respiratory pathology of this condition must address the altered lung physiology and recurrent, complex, polymicrobial infections and biofilms that affect the CF pulmonary tract. Cysteamine is a potential solution to these unmet medical needs and is described here for the first time as (Lynovex®) a single therapy with the potential to deliver mucoactive, antibiofilm and antibacterial properties; both in oral and inhaled delivery modes. Cysteamine is already established in clinical practice for an unrelated orphan condition, cystinosis, and is therefore being repurposed (in oral form) for cystic fibrosis from a platform of over twenty years of safety data and clinical experience.

Methods: The antibacterial and antibiofilm attributes of cysteamine were determined against type strain and clinical isolates of CF relevant pathogens using CLSI standard and adapted microbiological methods and a BioFlux microfluidic system. Assays were performed in standard nutrient media conditions, minimal media, to mimic the low metabolic activity of microbes/persister cells in the CF respiratory tract and in artificial sputum medium. In vivo antibacterial activity was determined in acute murine lung infection/cysteamine nebulisation models. The mucolytic potential of cysteamine was assessed against DNA and mucin in vitro by semi-quantitative macro-rheology. In all cases, the 'gold standard' therapeutic agents were employed as control/comparator compounds against which the efficacy of cysteamine was compared.

Results: Cysteamine demonstrated at least comparable mucolytic activity to currently available mucoactive agents. Cysteamine was rapidly bactericidal against both metabolically active and persister cells of Pseudomonas aeruginosa and also emerging CF pathogens; its activity was not sensitive to high ionic concentrations characteristic of the CF lung. Cysteamine prevented the formation of, and disrupted established P. aeruginosa biofilms. Cysteamine was synergistic with conventional CF antibiotics; reversing antibiotic resistance/insensitivity in CF bacterial pathogens.

Conclusions: The novel mucolytic-antimicrobial activity of cysteamine (Lynovex®) provides potential for a much needed new therapeutic strategy in cystic fibrosis. The data we present here provides a platform for cysteamine's continued investigation as a novel treatment for this poorly served orphan disease.

Show MeSH
Related in: MedlinePlus