Specific Mechanisms of Bispecific Antibodies

Bispecific antibodies (bsAbs) are a class of artificially synthesized antibody molecules that can simultaneously recognize two different antigens or epitopes. They have unique functions and advantages that monoclonal antibodies (mAbs) do not have, such as recruiting and activating immune cells, blocking multiple signaling pathways, enhancing affinity and specificity, etc. However, the development and application of bsAbs also face many challenges, such as the diversity of structure and format, the uncertainty of biological activity and stability, the difficulty of production and purification, etc. There are several main mechanisms of bsAbs in tumor immunotherapy, including immune cell recruitment, dual checkpoint blockade, dual signaling inhibition, co-localized blocking, targeted payload delivery, biparatopic recognition and tumor-targeted immunomodulation, etc.

Fig.1 Examples of obligate mechanisms of action of bsAbs. (Labrijn AF, 2019)

Immune Cell Recruiting

Immune cell recruitment is the most common and successful mechanism of bsAbs in tumor immunotherapy. It utilizes the bispecificity of bsAbs to connect tumor cells and immune cells, thereby achieving specific recognition and killing of tumor cells. The advantage of this mechanism is that it can activate and expand immune cells that cannot recognize tumor antigens, such as T cells, NK cells, macrophages, etc., and avoid damage to normal tissues. Currently, there are various types of bsAbs that achieve immune cell recruitment, among which CD3×CD19 bsAbs are representative, and have achieved remarkable clinical effects in B-cell malignancies such as leukemia and lymphoma.

Dual Checkpoint Blockade

Dual checkpoint blockade is another important mechanism of bsAbs in tumor immunotherapy. It utilizes the bispecificity of bsAbs to block two immune checkpoint molecules simultaneously, thereby enhancing the tumor immune response. Immune checkpoint molecules are a class of molecules that regulate the activity and function of immune cells. They can maintain the balance and tolerance of the immune system under normal conditions, but are exploited by tumor cells to suppress the attack of immune cells under tumor conditions, leading to tumor immune escape. Currently, there are various monoclonal antibodies that target different immune checkpoint molecules for blockade therapy, such as PD1, PDL1, CTLA4, etc., and have achieved some clinical effects. However, single checkpoint blockade therapy also has many limitations and problems, such as low efficacy, high resistance, and large toxic side effects. To solve these problems, scientists have developed a new type of bsAbs that can block two immune checkpoint molecules simultaneously, thereby achieving stronger tumor immune activation and wider applicability.

Dual Signaling Inhibition

Dual signaling inhibition is another effective mechanism of bsAbs in tumor immunotherapy. It utilizes the bispecificity of bsAbs to inhibit two tumor signaling pathways simultaneously, thereby preventing tumor cell growth, proliferation and metastasis. Tumor signaling pathways are a class of molecular networks that regulate the biological behavior of tumor cells. They can maintain the homeostasis and function of cells under normal conditions, but are mutated or overexpressed by tumor cells to gain survival advantage and adapt to stress under tumor conditions, leading to malignant transformation and resistance of tumor cells. Currently, there are various monoclonal antibodies that target different tumor signaling pathways for inhibition therapy, such as EGFR, HER2, VEGF, etc., and have achieved some clinical effects. However, single signaling pathway inhibition therapy also has many limitations and problems, such as low efficacy, high resistance, and large toxic side effects. To solve these problems, scientists have developed a new type of bsAbs that can inhibit two tumor signaling pathways simultaneously, thereby achieving stronger tumor cell killing and wider applicability.

Co-localized Blocking

Co-localized blocking is a novel and effective mechanism of bsAbs in tumor immunotherapy. It utilizes the bispecificity of bsAbs to block two adjacent or identical antigens or epitopes, thereby reducing the escape and resistance of tumor cells. Tumor cells under immune pressure or drug pressure will change the expression or structure of antigens or epitopes to evade the attack of immune cells or the action of drugs, leading to the failure of tumor immunotherapy or targeted therapy. Currently, there are various monoclonal antibodies that target different antigens or epitopes for blockade therapy, such as HER2, EpCAM, CD20, etc., and have achieved some clinical effects. However, single antigen or epitope blockade therapy also has many limitations and problems, such as low efficacy, high resistance, and large toxic side effects. To solve these problems, scientists have developed a new type of bsAbs that can block two adjacent or identical antigens or epitopes simultaneously, thereby achieving stronger tumor cell killing and wider applicability.

Targeted Payload Delivery

The targeted payload delivery concept uses bispecific antibodies (BsAbs) that bind to both a tumor-associated antigen (TAA) on tumor cells and a payload linked by molecules like digoxigenin. The payload, which could be a cytotoxic drug, immunotoxin, or fluorescent dye, is transported to and released inside tumor cells, reducing exposure to normal cells. BsAbs are designed in various structures (e.g., IgG-scFv, Fab-scFv) and can attach payloads covalently or non-covalently. TAAs include HER2, IGF1R, CD22. 

Biparatopic bsAbs

Biparatopic bispecific antibodies (bsAbs) target two distinct epitopes on the same antigen, enhancing binding affinity and avidity compared to monospecific antibodies. This dual targeting improves specificity, reduces off-target effects, and may delay resistance by engaging multiple essential pathways in target cells. Biparatopic bsAbs are created through methods like chemical conjugation (linking two antibody fragments with a chemical linker), hybridoma fusion (merging hybridoma cells to produce bsAbs), genetic engineering (modifying antibody gene DNA), or phage display (selecting antibody fragments on bacteriophages). Various formats like IgG-scFv, dual variable domain-immunoglobulin, bispecific T-cell engager, dual-affinity re-targeting, and Fab-arm exchange offer different benefits in valency, size, flexibility, and stability, influencing their production and clinical use.

Tumor-Targeted Immunomodulators

Tumor-targeted Immunomodulators (TTIMs) are bispecific antibodies (BsAbs) that simultaneously target tumor antigens and immune checkpoints, crucial for T cell regulation. Cancer cells often escape immune detection by interacting with these checkpoints, thereby suppressing T cell activity. TTIMs operate by binding to both a tumor antigen and an immune checkpoint molecule, promoting T cell and cancer cell proximity, enhancing T cell attack. They block checkpoint interactions, reactivating T cell anti-tumor functions, and can trigger immune cell signaling or deliver agents like cytokines or toxins to the tumor, boosting local immune responses or directly attacking cancer cells.

Conclusion

Taken together, bsAbs have unique functions and advantages that monoclonal antibodies do not have, such as recruiting and activating immune cells, blocking multiple signaling pathways, enhancing affinity and specificity, etc., thereby achieving stronger tumor immune activation and wider applicability. However, the development and application of bsAbs also face many challenges, such as the diversity of structure and format, the uncertainty of biological activity and stability, the difficulty of production and purification, etc. Therefore, the future needs to further optimize the design and manufacture of bsAbs, as well as conduct more clinical trials to verify the effectiveness and safety of bsAbs in tumor immunotherapy.

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