Abstract
The main objective of this work was to understand and mathematically characterize the cellular disposition of a tool ADC, trastuzumab-vc-MMAE (T-vc-MMAE). Towards this goal, three different analytical methods were developed to measure the concentrations of different ADC related analytes in the media and cell lysate. An LC-MS/MS method was developed to quantify unconjugated drug (i.e. MMAE) concentrations, a forced deconjugation method was developed to quantify total drug concentrations, and an ELISA 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 2h exposure of MMAE and T-vc-MMAE. No difference was observed in the intracellular accumulation of MMAE between the two cell lines following incubation with MMAE. However, when incubated with T-vc-MMAE, much higher intracellular exposures of unconjugated drug, total drug, and antibody were observed in N87 cells compared to GFP-MCF7 cells. A novel single cell disposition model was developed to simultaneously characterize in vitro PK 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 cell-line specific estimates of MMAE influx rate, MMAE efflux rate, and intracellular degradation rate for T-vc-MMAE. Global Sensitivity Analysis was performed on the model to identify key parameters responsible for the exposure of different analytes in cellular and extracellular space. The single cell PK model for ADCs presented here is expected to provide a better framework for characterizing bystander effect of ADCs.
- The American Society for Pharmacology and Experimental Therapeutics