Overview of F(ab')2-scFv2

Background of F(ab')2-scFv2

Bispecific antibodies are antibodies that can simultaneously recognize two different antigens or two different epitopes on the same antigen. Bispecific antibodies possess functions that monoclonal antibodies lack, such as redirecting cytotoxic T cells or natural killer cells to eliminate tumor cells, blocking two signaling pathways at the same time, activating two signaling pathways simultaneously, and more. Bispecific antibodies hold promising applications in cancer, autoimmune diseases, infectious diseases, and other fields. They come in various structural forms, which can be categorized based on the presence of Fc fragments and symmetry. Bispecific antibodies without Fc fragments include single-chain variable fragments (scFv), bispecific T cell engagers, bispecific single-domain antibodies (Duobody), etc. Bispecific antibodies with Fc fragments consist of symmetrical IgG-like bispecific antibodies and asymmetrical F(ab')2-scFv2 antibodies.

F(ab')2-scFv2 is an asymmetrical bispecific antibody with Fc fragments, consisting of two F(ab')2 fragments and one scFv2 fragment, which recognize two different antigens respectively. F(ab')2-scFv2 can be generated using four methods: quadroma technology, transfectoma, genetically engineered mice and recombinant DNA technology. F(ab')2-scFv2 exhibits several advantages, such as high affinity, high stability, high penetration and low immunogenicity. However, it also has certain disadvantages, including a complex preparation process, high cost and possible dose dependence. F(ab')2-scFv2 is primarily employed in clinical practice for the treatment of cancer and autoimmune diseases, such as lymphoma, leukemia, breast cancer, rheumatoid arthritis, etc. Currently, there are ongoing clinical trials or approved marketing of several F(ab')2-scFv2 antibodies, including catumaxomab, blinatumomab, emactuzumab, etc. These antibodies achieve therapeutic effects through different mechanisms, such as redirecting immune cells to kill tumor cells, blocking tumor growth factors or signaling pathways, activating macrophages to phagocytose tumor cells, etc.

Structural Features of F(ab')2-scFv2

F(ab')2-scFv2 is an asymmetrical bispecific antibody with Fc fragments, composed of an F(ab')2 fragment and an scFv2 fragment linked by a disulfide bond. The F(ab')2 fragment consists of two Fab fragments, one of which lacks the Fc part. The F(ab')2 fragment possesses bivalency, enabling it to bind to two identical antigen molecules. The scFv2 fragment comprises two scFv fragments, each composed of a heavy chain variable region and a light chain variable region. The scFv2 fragment exhibits monovalency, allowing it to bind to a different antigen molecule. Consequently, the F(ab')2-scFv2 antibody exhibits trivalency, enabling it to bind to two different antigen molecules. It has a molecular weight of approximately 150 kDa.

Fig.1 Schematic Diagram of F(ab')2-scFv2

Clinical Data of F(ab')2-scFv2

F(ab')2-scFv2 is a type of bispecific antibody, comprising of two F(ab')2 fragments and two scFv fragments, capable of simultaneously recognizing two different antigens. This antibody structure possesses a high molecular weight (about 150 kDa), a prolonged serum half-life, low immunogenicity, and minimal renal clearance. F(ab')2-scFv2 antibodies show potential advantages in the treatment of various diseases, especially in tumor immunotherapy.

Currently, several F(ab')2-scFv2 antibodies are undergoing clinical trials, primarily targeting tumor-related antigens, as shown in Table 1. Among them, the most advanced one is RG7828, developed by Roche, which is a bispecific antibody targeting CD20 and CD3 for the treatment of relapsed or refractory B-cell malignancies. This antibody has entered phase III clinical trials, and is currently recruiting patients. The mechanism of action of RG7828 involves inducing T-cell-mediated tumor cell killing by simultaneously binding to CD20-positive tumor cells and CD3-positive T cells. In phase II clinical trials, RG7828 showed good safety and efficacy, with an overall response rate of 43.3% for refractory non-Hodgkin lymphoma (NHL) and 29.5% for chronic lymphocytic leukemia (CLL), along with median durations of response of 11.6 months and 9.7 months, respectively. However, there are currently no F(ab')2-scFv2 antibodies approved for marketing by regulatory authorities.

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