ICE Bioscience Posters

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The in vivo drug-to-antibody ratio (DAR) value and drug load distribution on an ADC are crucial to the drug efficacy and safety. The evaluation of ADC stability in systemic circulation is usually achieved by characterization of free payload release rate and DAR value. During linker payload screening stage, the payload release rate showed the extent of linker cleavage in plasma, and the change of DAR value gave more information of deconjugation. The commonly used detection methods for DAR value were UV/Vis spectroscopy[2], hydrophobic interaction chromatography (HIC)[3], RP-HPLC[4], et al. Those methods were more compatible with samples with simple components instead of biological samples. There were many challenges in the analysis of DAR value by LC-MS in biological samples.

A highly specific and sensitive LBA-LC-MS method was successfully developed for the DAR value detection of Trastuzumab deruxtecan (T-DXD) in biological samples. The T-DXD was enriched by immunocapture with biotinylated antigen from human plasma and by anti-human IgG Ab from other nonclinical species plasma. After eluting the ADC from the immunocapture beads, 20 mM TECP was add to the elution; a Thermo QE Plus was used to detect the intensity of LC and HC. The intensity of the top three glycosylation HCs were summarized. The average DAR was calculated based on the summarized intensity. The results showed that the DAR values of T-DXD in mouse plasma sample after Day 0, 3 and 7 day incubation were 7.9, 6.8 and 3.6, respectively. The method had been used to evaluate both in vitro and in vivo ADC plasma stability. The workflow of the method can also apply to both conventional conjugated ADCs and site-specific conjugated ADCs, which could help researchers understand stability of ADC much better.