How to choose a bispecific antibody format?

The success of a bispecific antibody (bsAb) therapy largely depends on choosing the appropriate format for the specific therapeutic application. With numerous bispecific formats available, each with its own unique features and advantages, making the right choice can be a complex yet crucial decision. In this article, we will explore the key considerations for choosing a bsAb format, providing insights to help guide you through this intricate process.

Understanding bispecific antibodies

Bispecific antibodies are a class of engineered antibodies designed to target two distinct antigens simultaneously. They possess unique structures that allow them to bind to multiple targets, enabling novel therapeutic strategies in various disease areas. 

BsAbs can be broadly categorized into two main groups: those with Fc regions and those without.^​1​ The Fc region is the tail region of an antibody which interacts with and activates the immune system. Fc-containing BsAbs retain Fc-mediated effector functions such as antibody-dependent cell-mediated cytotoxicity (ADCC), while Fc-lacking BsAbs offer advantages such as enhanced tissue penetration and reduced immunogenicity.^​2​

BsAbs hold immense potential for addressing unmet medical needs and driving innovation in drug development. In the following sections, we will evaluate the key factors to consider when choosing a bsAb format, empowering you to make informed decisions.

Factors to consider when choosing a BsAb format

The antibody format directly influences the antibody’s efficacy, safety, and manufacturability. With numerous formats available, each offering unique advantages and challenges, selecting the most suitable format can significantly impact the success of a therapeutic candidate.

After considering the therapeutic objective and target antigens, it’s essential to assess pharmacokinetics, manufacturing feasibility, and immunogenicity of the antibody under consideration.

Therapeutic objective

Consider the specific disease target(s) and the desired mechanism of action for the bsAb. For example, if the goal is to activate immune cells and enhance tumor cell killing, a bsAb format capable of engaging both T cells and tumor cells may be preferred.^​3​

Target antigens

Evaluate the expression levels and distribution of the target antigens in tissues and cells of interest. Additionally, consider the specificity and affinity required for each target antigen to ensure effective target engagement and therapeutic efficacy.

Pharmacokinetics and pharmacodynamics (PK/PD)

Consider factors such as serum half-life, tissue distribution, and clearance rates. Additionally, evaluate the pharmacodynamic effects of the bsAb, including receptor binding kinetics and downstream signaling pathways.^​4​

Immunogenicity and safety

Assessing the potential immunogenicity of the bsAb format is crucial for ensuring patient safety. Consider strategies to minimize immunogenicity, such as humanization of antibody sequences.^​5​

Manufacturing considerations

Evaluate the feasibility and scalability of manufacturing the chosen bsAb format. Consider factors such as expression system compatibility, purification methods, and downstream processing requirements. Choosing a format that is compatible with current manufacturing capabilities can streamline the production process and reduce development costs.

At evitria, we specialize in navigating you through the intricate process of selecting the optimal bispecific antibody format tailored to your therapeutic objectives. But we don’t just provide guidance; we also handle the production of the required antibodies, ensuring the highest quality in minimal time. With our collaborative methodology and state-of-the-art technology, you can confidently propel your bsAb projects towards clinical triumph.

Common bispecific antibody formats

When selecting the ideal bispecific antibody format for your therapeutic goals, you’ll likely weigh your options among a few main formats. These include the Duobody system by Genmab, which facilitates the creation of bsAbs through a straightforward Fab arm exchange process.^​6​  In Fab arm exchange, two antibody fragments containing different antigen-binding sites are combined under controlled laboratory conditions to allow bonds to form, creating a bispecific molecule.^​7​

Another option is the IgG-scFv format.^​8​ In this format, two single-chain variable fragments (scFvs) from the same antibody are fused to the heavy or light chains of another antibody. scFvs contain the variable regions of both heavy and light chains, which together comprise an antigen-binding site. IgG-scFvs are tetravalent, with two binding sites per antigen (2 + 2 valence). They have variable expression and purification schemes depending on the specific scFv sequence used. 

The Knobs-into-Holes (KIH) format and related techniques like Crossmab^​9​ present additional methods 1 + 1 bispecific generation. In these formats, heavy chains of two different antibodies are modified to introduce distinctive binding sites—a “knob” or “hole”—to promote their heterodimerization. However, these formats can come with their own specific challenges, such as in undesired homodimer formation or improper heavy/light chain pairing.

Bispecific antibody production with evitria

Choosing the right bispecific antibody format is paramount for achieving therapeutic success. By collaborating with experts and utilizing advanced technologies, researchers can navigate this process effectively and accelerate bispecific antibody development.

evitria offers comprehensive support throughout the development journey, from format selection to bispecific antibody production. Utilizing Lonza’s bYlok® bispecific pairing technology, evitria ensures high-quality production of bispecific antibodies, with >95% correct heavy chain-light chain pairing. This technology enables rapid transient production of variants for discovery and non-GMP development studies, accelerating the identification of lead candidates and facilitating earlier access to engineered bispecifics.

Whether you require guidance in format selection or seamless production, evitria is your trusted partner for advancing bispecific antibody projects with confidence. Together, we can shape the future of therapeutic innovation and improve patient outcomes.

Read more about Bispecific Antibodies from Desmond Schofield

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• Development of bispecific antibodies: challenges & solutions
• Bispecific antibody formats
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