Abstract
Recent years have shown a tremendous increase and diversification in antibody-based therapeutics with advances in production techniques and formats. The plethora of currently investigated bi- to multi-specific antibody architectures can be harnessed to elicit a broad variety of specific modes of actions in oncology and immunology, spanning from enhanced selectivity to effector cell recruitment, all of which cannot be addressed by monospecific antibodies. Despite continuously growing efforts and methodologies, the identification of an optimal bispecific antibody as the best possible combination of two parental monospecific binders, however, remains challenging, due to tedious cloning and production, often resulting in undesired extended development times and increased expenses. Although automated high throughput screening approaches have matured for pharmaceutical small molecule development, it was only recently that protein bioconjugation technologies have been developed for the facile generation of bispecific antibodies in a ‘plug and play’ manner. In this review, we provide an overview of the most relevant methodologies for bispecific screening purposes—the DuoBody concept, paired light chain single cell production approaches, Sortase A and Transglutaminase, the SpyTag/SpyCatcher system, and inteins—and elaborate on the benefits as well as drawbacks of the different technologies.
Highlights
The human body is continuously exposed to potentially harmful and life-threatening opponents such as pathogens or malignant cells
The general process of bispecific antibody generation applying DuoBodies involves three basic steps: (1) separate production of monospecific antibodies harboring respective mutations in mammalian cell cultures, (2) purification according to standard processes (e.g., Protein-A), (3) (c)Fab-arm exchange (FAE) under tailored laboratory conditions followed by another purification step
The concept of bispecific antibodies for effector cell recruitment dates back to the 1980s, with reports on production [111] and effective CTL retargeting [112] and the design of a universal T cell engager [113]
Summary
The human body is continuously exposed to potentially harmful and life-threatening opponents such as pathogens or malignant cells. The general process of bispecific antibody generation applying DuoBodies involves three basic steps: (1) separate production of monospecific antibodies harboring respective mutations in mammalian cell cultures, (2) purification according to standard processes (e.g., Protein-A), (3) (c)FAE under tailored laboratory conditions followed by another purification step. This typically yields bispecific antibodies with more than 95% heterodimer content. The DuoHexaBody technology offers a broad applicability for bispecific antibody generation with target-mediated enhanced potencies for CDC-mediated effector functions as elegantly demonstrated by Oostindie et al [35]
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