Multifaceted Bioanalytical Methods for the Comprehensive Pharmacokinetic and Catabolic Assessment of MEDI3726, an Anti-Prostate-Specific Membrane Antigen Pyrrolobenzodiazepine Antibody–Drug Conjugate

Abstract

Complex biotherapeutic modalities, such as antibody-drug conjugates (ADC), present significant challenges for the comprehensive bioanalytical characterization of their pharmacokinetics (PK) and catabolism in both preclinical and clinical settings. Thus, the bioanalytical strategy for ADCs must be designed to address the specific structural elements of the protein scaffold, linker, and warhead. A typical bioanalytical strategy for ADCs involves quantification of the Total ADC, Total IgG, and Free Warhead concentrations. Herein, we present bioanalytical characterization of the PK and catabolism of a novel ADC. MEDI3726 targets prostate-specific membrane antigen (PMSA) and is comprised of a humanized IgG1 antibody site-specifically conjugated to tesirine (SG3249). The MEDI3726 protein scaffold lacks interchain disulfide bonds and has an average drug to antibody ratio (DAR) of 2. Based on the structural characteristics of MEDI3726, an array of 4 bioanalytical assays detecting 6 different surrogate analyte classes representing at least 14 unique species was developed, validated, and employed in support of a first-in-human clinical trial (NCT02991911). MEDI3726 requires the combination of heavy-light chain structure and conjugated warhead to selectively deliver the warhead to the target cells. Therefore, both heavy-light chain dissociation and the deconjugation of the warhead will affect the activity of MEDI3726. The concentration− time profiles of subjects dosed with MEDI3726 revealed catabolism of the protein scaffold manifested by the more rapid clearance of the Active ADC, while exhibiting minimal deconjugation of the pyrrolobenzodiazepine (PBD) warhead (SG3199).

First-in-Human, Phase 1 Dose-Escalation Study of Biparatopic Anti-HER2 Antibody–Drug Conjugate MEDI4276 in Patients with HER2-positive Advanced Breast or Gastric Cancer

“…ADCs are complex biotherapeutic modalities that require sophisticated bioanalytical methods to properly interrogate their PK and catabolism (49,61). To enable quantitative analysis of changes in DAR over time for MEDI426 and to assess the impact that deacetylation would have on the tubulysin warhead, we employed sophisticated, novel, DAR-sensitive, bioanalytical methods for the assessment of MEDI4276 PK (50).…”

2020 White Paper on Recent Issues in Bioanalysis: BMV of Hybrid Assays, Acoustic MS, HRMS, Data Integrity, Endogenous Compounds, Microsampling and Microbiome ( Part 1 –Recommendations on Industry/Regulators Consensus on BMV of Biotherapeutics by LCMS, Advanced Application in Hybrid Assays, Regulatory Challenges in Mass Spec, Innovation in Small Molecules, Peptides and Oligos)

“…Antibody-drug conjugates (ADC) present significant bioanalytical challenges for ensuring comprehensive PK characterization. In a case study, multiple hybrid assays were developed in order to appropriately characterize the PK properties of a unique ADC, ultimately measuring at least 14 unique species/catabolites [27]. In another case study, this technique was used for ADA impact assessments [28], where hybrid assays were developed to confirm the impact on the PK by ADA and to deconvolute the ADA binding domain.…”

Target-responsive subcellular catabolism analysis for early-stage antibody–drug conjugates screening and assessment

“…After proving TARSC analysis enables ADC catabolism studies in lysosomes, we next asked whether TARSC is applicable to examine the active payloads released from ADCs in intact cells where multiple crucial biological processes including antigen‒antibody binding, intracellular internalization, trafficking and lysosomal degradation can be reflected 12 , 30 . We found that lys-MCC-DM1 did not show dose-dependent increase with the administered concentrations of T-DM1, whereas DM1 increased with the ramped administered concentrations.…”