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  2. Measurement and Mathematical Characterization of Cell-Level Pharmacokinetics of Antibody-Drug Conjugates: A Case Study with Trastuzumab-vc-MMAE

Measurement and Mathematical Characterization of Cell-Level Pharmacokinetics of Antibody-Drug Conjugates: A Case Study with Trastuzumab-vc-MMAE

  • Drug Metab Dispos. 2017 Nov;45(11):1120-1132. doi: 10.1124/dmd.117.076414.
Aman P Singh 1 Dhaval K Shah 2
Affiliations

Affiliations

  • 1 Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, The State University of New York at Buffalo, Buffalo, New York.
  • 2 Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, The State University of New York at Buffalo, Buffalo, New York dshah4@buffalo.edu.
Abstract

The main objective of this work was to understand and mathematically characterize the cellular disposition of a tool antibody-drug conjugate (ADC), trastuzumab-valine-citrulline-monomethyl Auristatin E (T-vc-MMAE). Toward this goal, three different analytical methods were developed to measure the concentrations of different ADC-related analytes in the media and cell lysate. A liquid chromatography-tandem mass spectrometry method was developed to quantify unconjugated drug (i.e., MMAE) concentrations, a forced deconjugation method was developed to quantify total drug concentrations, and an enzyme-linked immunosorbent assay method was developed to quantify total antibody (i.e., trastuzumab) concentrations. Cellular disposition studies were conducted in low-HER2-(GFP-MCF7) and high-HER2-expressing (N87) cell lines, following continuous or 2-hour exposure to MMAE and T-vc-MMAE. Similar intracellular accumulation of MMAE was observed between two cell lines following incubation with plain MMAE. However, when incubated with T-vc-MMAE, much higher intracellular exposures of unconjugated drug, total drug, and total antibody were observed in N87 cells compared with GFP-MCF7 cells. A novel single-cell disposition model was developed to simultaneously characterize in vitro pharmacokinetics of all three analytes of the ADC in the media and cellular space. The model was able to characterize all the data well and provided robust estimates of MMAE influx rate, MMAE efflux rate, and intracellular degradation rate for T-vc-MMAE. ADC internalization and degradation rates, HER2 expression, and MMAE efflux rate were found to be the key parameters responsible for intracellular exposure to MMAE, on the basis of a global sensitivity analysis. The single-cell pharmacokinetics model for ADCs presented here is expected to provide a better framework for characterizing bystander effect of ADCs.

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