Development and validation of a cell-based methodology to evaluate filgrastim relative affinity to granulocyte...

Filgrastim, a therapeutic protein for the treatment of neutropenia, exerts its biological activity by interacting with the granulocyte colony-stimulating factor receptor. However, the affinity of filgrastim to its receptor is usually overlooked during characterization and comparability tests, since its biological activity is assessed only by in vitro proliferation assays. In this work, we propose the use of a cell-based colorimetric method to determine the relative affinity of filgrastim to its receptor expressed in murine myeloid leukemia cells. After performing a validation exercise, the method proved to be accurate, specific, and linear within an affinity interval of 75 to 130%; precise with a confidence interval of 93.6 to 114.2% and a coefficient of variation of 12.6%. The validation exercise proved that the proposed cell-based method is a viable alternative to evaluate the biological activity of filgrastim via the affinity for its receptor and it is suitable to be included as part of its biological characterization exercises as well as in comparability exercises for products coming from distinct manufacturing processes.