A Purification Strategy Utilizing Hydrophobic Interaction Chromatography to Obtain Homogeneous Species from a Site-Specific Antibody Drug Conjugate Produced by AJICAP™ First Generation.

Yutaka Matsuda,Monica Leung,Brian Mendelsohn,Tatsuya Okuzumi

doi:10.3390/antib9020016

Abstract

In recent years, site-specific antibody drug conjugates (ADC)s have been in great demand because they have an expanded therapeutic index compared with conventional ADCs. AJICAP™ technology is a chemical conjugation platform to obtain site-specific ADCs through the use of a class of Fc-affinity compounds. Promising results from early technology development studies led to further investigation of AJICAP™ ADC materials to obtain site-specific and homogeneous drug antibody ratio (DAR) ADCs. Here we report site-specific conjugation followed by a preparative hydrophobic interaction chromatography (HIC) purification strategy to obtain purified “DAR = 1.0” and “DAR = 2.0” AJICAP™ ADC materials. Optimization of the mobile phase conditions and resin achieved a high recovery rate. In vitro biological assay demonstrated the target selective activity for purified homogeneous DAR ADCs. These results indicate the ability of a HIC purification strategy to provide “DAR = 1.0” and “DAR = 2.0” AJICAP™ ADCs with considerable potency and target selectivity.

Highlights

Recent years have seen rapid growth in the research and development of antibody drug conjugates (ADCs), with 7 ADCs approved for clinical use by the Food and Drug Administration (FDA) and over 85ADCs currently in clinical development [1,2,3]
Preparative hydrophobic interaction chromatography (HIC) purification of the resulting AJICAPTM-ADC was conducted with an AKTA
Preparative HIC purification of the resulting AJICAPTM-ADC was conducted with an AKTA Pure chromatography system

Summary

Introduction

Recent years have seen rapid growth in the research and development of antibody drug conjugates (ADCs), with 7 ADCs approved for clinical use by the Food and Drug Administration (FDA) and over 85ADCs currently in clinical development [1,2,3]. All of the current ADCs on the market have a stochastic distribution of cytotoxic drugs conjugated to multiple sites of the antibody, leading to variation between batches in both the drug to antibody ratio (DAR) and the drug-linker attachment sites [4]. These heterogeneous ADCs produced by nonspecific conjugation techniques may have diminished efficacy or increased toxicity, limiting their therapeutic index compared with a homogeneous ADC [5,6]. Our research group has developed an innovative chemical site-specific conjugation platform termed AJICAPTM first generation [8] This technology enables the installation of reactive moieties compatible with payloads and linkers, such as thiol functional groups to well-defined amino acid residues through the use of

Methods

Results

Conclusion