“Fit-for-Purpose” Method Validation and Application of a Biomarker (C-terminal Telopeptides of Type 1 Collagen) in Denosumab Clinical Studies

Abstract

Abstract. Biomarkers are used to study drug effects, exposure-response relationships, and facilitate early decision making during development. Denosumab, a fully human monoclonal antibody against receptor activator of nuclear factor-κB ligand, profoundly inhibits bone resorption. C-terminal telopeptides of type I collagen (CTx), a bone resorption biomarker, provides early indications of denosumab effectiveness and informs protracted clinical outcomes (e.g., bone mineral density). Because of the dynamic relationship between denosumab and CTx, a precise and robust assay was desired. Thus, we adopted a fit-for-purpose approach to modify and validate a commercial CTx diagnostic kit to meet the intended applications of a quantitative pharmacodynamic biomarker for denosumab development. Seven standards were prepared to replace five calibrators provided in the kit. Three quality controls (QC) and two sample controls were used to characterize and monitor assay performance. Robotic workstations were used for standard and QC preparation and assay execution. Method validation experiments were conducted with rigor and procedures similar to those used for drug bioanalysis. The method demonstrated a linear range of 0.0490-2.34 ng/mL with four-parameter logistic regression. Inter-assay total error of validation samples in serum was ≤26.7%. Extensive tests were conducted on selectivity in sera from target populations, specificity, stability, parallelism, and dilutional linearity. Applications to samples from numerous clinical studies confirmed that the CTx method was reliable, robust, and fit for use as an early indicator of denosumab effectiveness. Refinement supported the confidence for use in pharmacokinetic/pharmacodynamic modeling, dose selections, correlation to clinical effects, and formulation bioequivalence work.KEY WORDS: bone resorption marker; commercial immunoassay kit; method validation; pharmacodynamic (PD) biomarker; serum C-terminal telopeptides of type 1 (CTx).

Nutritional approach for inhibiting bone resorption in institutionalized elderly women with vitamin D insufficiency and high prevalence of fracture

“…therefore, bone biochemical markers are extremely useful to predict the long-term responsiveness to treatment after a few weeks. early changes in bone biochemical markers, particularly those reflecting bone resorption, such as ctX, predict the longterm rate of bone loss as well as the risk of incident fracture (40)(41)(42)(43)(44)(45)(46)(47)(48)(49). the magnitude of the reduction in markers of bone resorption observed in the foregoing study is less than that observed following therapeutic doses of strong inhibitors of bone resorption such as bisphosphonates (44,46,47).…”

Recommendations for Selection and Characterization of Protein Biomarker Assay Calibrator Material

“…In these For exploratory assays, the new lot of calibrator material should show similar results as the previous lot as suggested in Table V. In the event the performance of the new lot of material differs from the reference lot, we recommend obtaining another replacement lot (if available), normalizing the new lot concentration to the original lot using a correction factor (29), or value assignment (23,31,32). For treatment decision-making assays, multiple lots (a minimum of three) should be tested as early as possible and a normalization strategy via value assignment should be considered.…”

Addressing The Challenges of Biomarker Calibration Standards in Ligand-Binding Assays: A European Bioanalysis Forum Perspective

“…Trending of results for biomarker assays is already used by many to assess QC stability and assay robustness [13]. future science group…”

“…Inclusion of such samples affords confidence that comparable results are produced each time the assay is utilized and would detect upward or downward trends in concentration assuming that stability of such samples can be determined. Such an approach is described by Wang et al [13], where both high and low 'authentic samples' (incurred samples) were analyzed in each sample analysis run. Furthermore they were able to demonstrate reliability and robustness for the method despite two different reference material lots being used over 3 years in 12 clinical studies.…”