Specific Method Validation and Sample Analysis Approaches for Biocomparability Studies of Denosumab Addressing Method and Manufacture Site Changes

Abstract

Abstract. Manufacturing changes during a biological drug product life cycle occur often; one common change is that of the manufacturing site. Comparability studies may be required to ensure that the changes will not affect the pharmacokinetic properties of the drug. In addition, the bioanalytical method for sample analysis may evolve during the course of drug development. This paper illustrates the scenario of both manufacturing and bioanalytical method changes encountered during the development of denosumab, a fully human monoclonal antibody which inhibits bone resorption by targeting RANK Ligand. Here, we present a rational approach to address the bioanalytical method changes and provide considerations for method validation and sample analysis in support of biocomparability studies. An updated and improved ELISA method was validated, and its performance was compared to the existing method. The analytical performances, i.e., the accuracy and precision of standards and validation samples prepared from both manufacturing formulation lots, were evaluated and found to be equivalent. One of the lots was used as the reference standard for sample analysis of the biocomparability study. This study was sufficiently powered using a parallel design. The bioequivalence acceptance criteria for small molecule drugs were adopted. The pharmacokinetic parameters of the subjects dosed with both formulation lots were found to be comparable.

A simple and robust reporter gene assay for measuring the bioactivity of anti-RANKL therapeutic antibodies

“…Competitive ELISA was reported to determine the bioactivity of anti-RANKL mAbs. 18 However, ELISA only involves the initial step of binding, which is far from the end point of the biological response, so it could not completely represent the MOA of anti-RANKL mAbs. As for bioactivity determination of anti-RANKL mAbs with cell based methods, calcification of porcine valvular interstitial cells was checked by Alizarin red staining, 19 and maturation of patients-derived and mice-derived osteoclast were indicated by tartrate resistant acid phosphatase (TRAP) activity, 20 while both of which are unsuitable for routine bioassay due to the employment of primary cells and the resultant high variability.…”