Nadine Taleb1,2, Adèle Coriati1, Christian Khazzaka1, Jonathan Bayonne1, Virginie Messier1, Rémi Rabasa-Lhoret1,3,4. 1. 1 Institut de recherches cliniques de Montréal , Montréal, Québec, Canada . 2. 2 Division of Biomedical Sciences, Faculty of Medicine, Université de Montréal , Montréal, Québec, Canada . 3. 3 Montreal Diabetes Research Center , Québec, Canada . 4. 4 Department of Nutrition, Faculty of Medicine, Université de Montréal , Montréal, Québec, Canada .
Abstract
BACKGROUND: Available glucagon formulations are approved for immediate use after reconstitution for severe hypoglycemia emergency treatment. However, they are used in dual-hormone artificial pancreas (insulin and glucagon) studies through subcutaneous infusion pumps over 24 h. Chemical and physical stability of such glucagon use have not been reported in a comprehensive manner. MATERIALS AND METHODS: Recombinant Glucagon DNA (Eli Lilly) was used. Compatibility and sterility of glucagon delivery through subcutaneous pump systems were verified. Glucagon degradation through liquid chromatography with tandem mass spectrometry (LC-MS/MS), fibrillation using intrinsic tryptophan fluorescence shift, and bioactivity through a cell-protein kinase A-based fluorescent bioassay were assessed over a range of different physical conditions (temperature, movement, and air bubbles). RESULTS: Subcutaneous infusion pump systems administered glucagon in sterile conditions and with comparable accuracy to insulin delivery; mean absolute relative difference of actual versus expected weights were 1.2% ± 1.1% for glucagon and 1.1% ± 0.5% for insulin (P = 0.9). In comparison to freshly reconstituted samples, glucagon analyzed through LC-MS/MS was intact at 93.0% ± 7.0% after 24 h (P = 0.42) and 83.04% ± 6.0% after 48 h (P = 0.02) of incubation in pumps at 32°C. Peak wavelengths for Trp fluorescence did not differ from samples exposed to air bubbles or movement whether incubated (in infusion sets for 24 h at 32°) immediately or 24- and 48-h poststorage at 4°C (P = 0.10, 0.70 and 0.80, respectively) and no significant differences in bioactivity (shifts in EC50) were found for the same conditions (P = 0.13, 0.83, and 0.63). CONCLUSION: Available glucagon formulations are chemically and physically stable, as well as compatible with delivery through subcutaneous infusion pumps over 24 h and can be used in long-term clinical trials.
BACKGROUND: Available glucagon formulations are approved for immediate use after reconstitution for severe hypoglycemia emergency treatment. However, they are used in dual-hormone artificial pancreas (insulin and glucagon) studies through subcutaneous infusion pumps over 24 h. Chemical and physical stability of such glucagon use have not been reported in a comprehensive manner. MATERIALS AND METHODS: Recombinant Glucagon DNA (Eli Lilly) was used. Compatibility and sterility of glucagon delivery through subcutaneous pump systems were verified. Glucagon degradation through liquid chromatography with tandem mass spectrometry (LC-MS/MS), fibrillation using intrinsic tryptophan fluorescence shift, and bioactivity through a cell-protein kinase A-based fluorescent bioassay were assessed over a range of different physical conditions (temperature, movement, and air bubbles). RESULTS: Subcutaneous infusion pump systems administered glucagon in sterile conditions and with comparable accuracy to insulin delivery; mean absolute relative difference of actual versus expected weights were 1.2% ± 1.1% for glucagon and 1.1% ± 0.5% for insulin (P = 0.9). In comparison to freshly reconstituted samples, glucagon analyzed through LC-MS/MS was intact at 93.0% ± 7.0% after 24 h (P = 0.42) and 83.04% ± 6.0% after 48 h (P = 0.02) of incubation in pumps at 32°C. Peak wavelengths for Trp fluorescence did not differ from samples exposed to air bubbles or movement whether incubated (in infusion sets for 24 h at 32°) immediately or 24- and 48-h poststorage at 4°C (P = 0.10, 0.70 and 0.80, respectively) and no significant differences in bioactivity (shifts in EC50) were found for the same conditions (P = 0.13, 0.83, and 0.63). CONCLUSION: Available glucagon formulations are chemically and physically stable, as well as compatible with delivery through subcutaneous infusion pumps over 24 h and can be used in long-term clinical trials.
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