Quantitation of the monoclonal antibody Denosumab by bioassay and validated LC methods.

Autor: Perobelli RF; Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, 97105-900 Santa Maria, RS, Brazil., Xavier B; Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, 97105-900 Santa Maria, RS, Brazil., Silveira ARD; Department of Industrial Pharmacy, Federal University of Santa Maria, 97105-900 Santa Maria, RS, Brazil., Remuzzi GL; Department of Industrial Pharmacy, Federal University of Santa Maria, 97105-900 Santa Maria, RS, Brazil., Motta LGJ; Department of Industrial Pharmacy, Federal University of Santa Maria, 97105-900 Santa Maria, RS, Brazil., Dalmora SL; Department of Industrial Pharmacy, Federal University of Santa Maria, 97105-900 Santa Maria, RS, Brazil. Electronic address: sdalmora@terra.com.br.
Jazyk: angličtina
Zdroj: International journal of biological macromolecules [Int J Biol Macromol] 2018 Nov; Vol. 119, pp. 96-104. Date of Electronic Publication: 2018 Jul 23.
DOI: 10.1016/j.ijbiomac.2018.07.120
Abstrakt: The monoclonal antibody Denosumab (DmAb) is clinically used to treat osteoporosis and bone loss. We developed a bioassay based on the ability of DmAb to inhibit the effect of human receptor activator of nuclear factor-κB ligand (RANKL) to stimulate the formation of osteoclasts derived from RAW 264.7 cells. This bioassay was applied in conjunction with size exclusion high-performance liquid chromatography (SE-HPLC) and reversed-phase high-performance liquid chromatography (RP-HPLC) methods, with diode array detection (DAD), validated for the quantitation of this biotechnology-derived medicine. The SE-HPLC(DAD) method was carried out on a TSKGel G2000SW XL column and the mobile phase consisted of potassium phosphate buffer with sodium chloride, pH 7.4. The gradient RP-HPLC(DAD) method was carried out on a Vydac 214TP C 4 column at 60 °C. The mobile phases consisted of 0.1% v/v trifluoroacetic acid (TFA) in water and 0.1% v/v TFA in acetonitrile. Calibration curves were linear over the concentration ranges 6-200 μg mL -1 and 6-300 μg mL -1 for the SE-HPLC(DAD) and RP-HPLC(DAD) methods respectively. The bioassay results correlated with the LC methods results, indicating the capabilities of these methods to quantitate DmAb, which will contribute to ensure the batch-to-batch consistency and efficacy of this biotherapeutic.
(Copyright © 2018 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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