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J Pharm Biomed Anal. 2019 Sep 10;174:441-449. doi: 10.1016/j.jpba.2019.06.017. Epub 2019 Jun 11.

Simultaneous quantification of total antibody and antibody-conjugated drug for XMT-1522 in human plasma using immunocapture-liquid chromatography/mass spectrometry.

Author information

1
Mersana Therapeutics Inc., Cambridge, MA, 02139, USA. Electronic address: LXu@mersana.com.
2
Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA, 19341, USA. Electronic address: ZZhang1@frontagelab.com.
3
Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA, 19341, USA. Electronic address: sxu1@frontagelab.com.
4
Mersana Therapeutics Inc., Cambridge, MA, 02139, USA. Electronic address: jxu@jouncetx.com.
5
Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA, 19341, USA. Electronic address: JLin@frontagelab.com.
6
Mersana Therapeutics Inc., Cambridge, MA, 02139, USA. Electronic address: DLee@mersana.com.

Abstract

XMT-1522, an antibody-drug conjugate (ADC) currently in Phase I clinical development, represents the first Dolaflexin®-based, cleavable ADC with a high drug-antibody ratio (DAR). In this work, a novel immunocapture LC-MS/MS method was successfully developed for the simultaneous quantification of both total antibody and cleavable antibody-conjugated drug auristatin F-hydroxypropylamide (AF-HPA) in human plasma. This method utilized microwave-assisted enzymatic digestion for the total antibody and chemical release of the drug from ADC on a 96-well based immunocapture sample preparation platform. The total antibody and the conjugated drug AF-HPA were separated and subsequently quantified concurrently by LC-MS/MS. The linear range of the standard curve for total antibody was from 50 to 5000 ng/mL and for AF-HPA was from 3.3 to 330 ng/mL. The linearities showed R2 ≥ 0.993 for total antibody and R2 ≥ 0.996 for AF-HPA, respectively. The intra- and inter-day precision and accuracy were well within 15%. The validated method, with the characteristics of high efficiency, great selectivity, free of carryover, short LC-MS/MS time (˜3.5 min) and low sample volume (20 μl), was successfully applied for analyzing Phase 1 cancer patient samples.

KEYWORDS:

Antibody drug conjugate; Auristatin F hydroxypropylamide; Drug-antibody ratio; Immunocapture mass spectrometry; Microwave-assisted enzymatic digestion

PMID:
31220702
DOI:
10.1016/j.jpba.2019.06.017

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