Bi- and multi-specific Antibodies (bsAbs)
Target selection
Target selection for bispecific antibodies is driven by the desired mode of action and a detailed understanding of the underlying biology.
Many applications of bsAbs exploit the ability of bsAbs to bring two targets together. Most approved bsAbs for cancer treatment engage both a target on the surface of the cancer cell and an activating molecule on an immune cell, to direct the patient’s own immune cells to kill tumours. In non-oncology fields, the blockbuster drug emicizumab (Hemlibra®) brings together two blood clotting factors for the treatment of haemophilia. Targeted protein degradation is also attracting interest as a new approach to target knockdown: here, a bispecific antibody is used to bring the target together with a naturally occurring ‘tag’ which marks it for degradation by the cell.
However, potential modes of action of bispecific antibodies are highly varied, and not all modes rely
on the ability of the antibody to pull components together.
For example, bispecific antibodies can act as dual inhibitors, knocking down two different targets simultaneously to provide therapeutic synergy.
There is also an increasing interest in the potential of biparatopic binding, i.e., the binding of two non-overlapping epitopes on the same target. Biparatopic binding can have numerous effects, including superior affinity and specificity resulting from avidity effects, promoting antagonism, locking target conformation, bringing about higher-order target clustering and overcoming resistance mechanisms. Jazz Pharmaceutical’s zanidatamab (Ziihera®) binds to two HER2 epitopes to potentiate efficacy and gained FDA approval in November 2024, illustrating the exciting potential of this technology.
"Alphamab Oncology and Chia Tai Tianqing Pharmaceutical Group both have a biparatopic anti-HER2 ADC in phase III clinical trials. Dual target or epitope binding can be used to make ADCs more selective to cancer cells, to modify internalisation or to overcome clinical drug-resistant mutations."
Affinity and valency
Optimisation of the activity of an antibody can require fine tuning of the relative binding strength of the antibody for each of its targets. For example, in the case of T-cell engagers, tuning the strength of tumour-antigen binding can influence the degree of off-target effects, while tuning the strength of binding to the T-cell can influence the potency of the response and the extent of systemic cytokine release. Another exciting approach that is gaining traction is to engineer the binding site to have greater binding affinity at the acidic pH found in the tumour microenvironment, so that the drug is more targeted to the desired site of action (an approach also being applied to ADCs, as in Halozyme’s HTI-1511).
Formats
The range of possible structures for bispecific antibodies is sometimes referred to as the bispecific ‘zoo’, with over 100 formats of varying shapes and sizes having been developed. For some of the choices that must be made, the underlying structure/function relationship is reasonably well-known. For example, the presence of an Fc region has effects including providing a relatively long half-life and the potential for engaging with other elements of the immune system (complement and Fc receptors). For other choices, the effect will be less clear: the geometry of the binding site in the overall structure can influence the binding strength in ways that can be unpredictable.
Novel formats are also being developed with new functionality. For example, the ‘pro-drug’ concept is being extended to bispecific antibodies to make formats which become active only when present at the desired target site, such as the tumour microenvironment. Approaches include having the binding site masked by a moiety which is removed by a tumour-specific protease, or assembly of two ‘hemibodies’ into a functional antibody only active at the target site.
Production and isolation
As with all types of therapeutic antibodies, developability considerations are of critical importance and are increasingly being considered at the earliest stages of lead selection.
Bispecific antibodies generally require assembly of complex asymmetric structures, introducing the additional challenge of making sure that the right chains come together in the right combinations. Incorrect pairing can reduce production efficiency and make isolation of the desired species much more difficult. A range of approaches have been developed to tackle this, often leading to proprietary technology platforms such as Genentech’s knob-into-hole mutations to ensure correct heavy chain pairing; Roche’s CrossMAb® approach to ensure correct light chain-heavy chain pairings or Genmab’s Duobody® technology.
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