(M1330-06-39) Regulatory Perspective of Considerations in Endogenous Therapeutic Analyte Bioanalysis

Purpose: Endogenous substances are compounds that are naturally present in the body either because the body produces them, or they are present in the normal diet [1]. Examples of endogenous therapeutics include hormones, neurotransmitters, vitamins, fatty acids, inorganic elements, and others. The accuracy of analyte measurement following the administration of an endogenous therapeutic agent poses a challenge as the exogenous therapeutic analyte and its endogenous counterpart cannot be distinguished. This presentation will highlight the challenges encountered in endogenous therapeutic analyte bioanalysis and share the regulatory perspective of important considerations when developing bioanalytical methods and conducting bioanalysis for endogenous therapeutic analytes during drug development.

Methods: Real case examples encountered during developing and validating bioanalytical methods for endogenous therapeutic analytes used in bioanalysis when conducting bioavailability (BA) and bioequivalence (BE) studies during drug development in support of new drug applications (NDAs), biologics license applications (BLAs), or abbreviated new drug applications (ANDAs) were collected. Practical lessons learned including those from the inspection and compliance perspective were summarized.

Results: Some common issues identified in the collected case examples related to bioanalysis of endogenous therapeutic analytes include: (1) While surrogate matrix was used to prepare calibration standards (CSs) and quality controls (QCs), the endogenous concentration of the therapeutic analyte was not accounted for in quantitation of the incurred study samples; (2) Absence of cross-validation data supporting the accuracy and precision of the therapeutics analyte measurements addressing the potential matrix effect (e.g., absence of parallelism test), potential differences in recovery when a surrogate matrix was employed in the bioanalytical method development and validation instead of the authentic matrix (i.e., human serum); (3) While CSs for a clinical study was prepared in an artificial matrix (e.g., buffer), investigation to demonstrate that the performance in CSs and QCs were comparable to those prepared in authentic matrix (e.g., human serum) was not conducted; (4) Incurred study samples went through a different sample preparation method compared to the CSs and QCs resulting in an uncertainty of the accuracy and precision of the incurred study samples; (5) The stability of the analyte during sample collection and handling was not adequately demonstrated during bioanalytical method development and validation. Deviation for standard procedures of sample handling and processing led to unexpectedly high or low analyte concentrations (i.e., false positive or false negative); (6) Difficulty in comparing drug products where the endogenous concentrations of the therapeutic analyte are relatively high compared with the exogenously administered therapeutic analyte. In addition, during inspection, it was found that a cluster of subjects with unexpectedly high baseline concentrations for endogenous therapeutic analytes were reported without any justification or explanation affecting the outcome of a BE study following baseline correction. The recently published International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) M10 Guidance for Industry: Bioanalytical Method Validation and Sample Analysis [2] outlines practical approaches such as: (1) the surrogate matrix approach, (2) the surrogate analyte approach, (3) the background subtraction approach, and (4) the standard addition approach, that can be utilized to calculate the concentration of the analyte in the study samples when matrices without interference are not available. Based on this guidance and the lessons learned from our collected case examples, practical tips and strategies to consider when developing bioanalytical methods for endogenous therapeutic analytes during drug development will be summarized and discussed.

Conclusion: When the endogenous substances are identical to the therapeutic analyte (i.e., drug) that is being administered, it can be difficult to determine the amount of therapeutic analyte released from the dosage form. This presentation highlights the common challenges encountered, issues identified, and lessons learned related to bioanalysis of endogenous therapeutics analytes and provides practical tips and strategies to consider from a regulatory perspective. It is not an overstatement to say that accurate, precise, and reliable bioanalysis of endogenous therapeutic analytes is pivotal for the success of drug development when they are the primary analyte of interest.

References: 1. U.S. Food and Drug Administration (FDA), Guidance for Industry: Bioequivalence Studies With Pharmacokinetic Endpoints for Drugs Submitted Under an ANDA, August 2021
2. U.S. FDA, Guidance for Industry: M10 Bioanalytical Method Validation and Study Sample Analysis, November 2022

Acknowledgements: Disclaimer: The views expressed are those of the authors and do not reflect official policy of the US FDA. No official endorsement by the FDA is intended or should be inferred. Acknowledgement: Hyeonglim Seo was supported by an appointment to the Oak Ridge Institute for Science and Education (ORISE) Research Participation Program at CDER administered by the ORISE through an agreement between the U.S. Department of Energy and FDA.