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Updated: Feb 20, 2026
A GPC3-targeting Bispecific Antibody, GPC3-S-Fab, with Potent Cytotoxicity
Published on: July 12, 2018
Generation and Characterization of a Bispecific Antibody Targeting Both PD-1 and c-MET
1Key Laboratory of Metabolism and Molecular Medicine, Ministry of Education, and Department of Biochemistry and Molecular Biology, School of Basic Medical Science, Fudan University, Shanghai, China.
Background:
Bispecific antibodies, BsAbs, are molecules with the ability to bind to two different epitopes on the same or different antigens. c-MET, cellular-mesenchymal to epithelial transition factor, is deregulated in many types of human malignancies. Abnormal c-MET activation in cancer correlates with poor prognosis. PD-1, programmed death-1, is an additional inhibitory receptor expressed by T cells. Blocking the interactions between PD-1 and PD-L1 has emerged as a promising immunotherapy for treating cancer.
Objectives:
The goal of this study was to identify a novel bispecific antibody targeting both c-MET and PD-1 as an anti-cancer therapeutic candidate.
Methods:
The BsAb was produced using 293E expression system and purified by Protein A affinity chromatography. Then the binding specificity and affinity of the BsAb was examined by FACS and biolayer light interferometry. The ability of the BsAb to inhibit the proliferation of tuman cells was measured using the CellTiter 96 Aqueous One Solution Cell Proliferation Assay kit; the potential signaling pathway involved was identified by Western Blot. Cytokine secreted by PHA-L stimulated PBMC was measured by ELISA. Effects of BsAb on PBMC-mediated lysis of MKN45 cells was measured by LDH cytotoxicity assay.
Results:
Based on the original sequences of PD-1 and c-MET mAb, a BsAb gene was designed, cloned into pCEP4 vector for expression in 293E cells. The BsAb was obtained after purification of the cell culture supernatant. It can bind to PD-1 and c-MET simultaneously, the calculated affinity was 11.5 nM for PD-1 and 9.09 nM for c-MET. The BsAb enhanced IFN-γ production over control IgG by 2-3 folds. It also inhibit the c-MET pathway activation and the proliferation of tumor cells significantly, comparable to JnJ-38877605. The BsAb showed dose-dependent cytotoxic activity against MKN45 cells.
Conclusion:
Our results indicated that a novel BsAb recognizing PD-1 and c-MET was successfully generated. It could redirect T cells to kill tumor cells, while retaining its inherent ability to restore T cells and inhibit tumor cells. With this potential, this BsAb could be developed as a therapeutic candidate for the treatment of various solid tumors.
Insights
A novel bispecific antibody (BsAb) targeting both cellular-mesenchymal to epithelial transition factor (c-MET) and programmed death-1 (PD-1) was developed. This dual-targeting BsAb shows potential for treating solid tumors by redirecting T cells and inhibiting tumor growth.
Area of Science:
- Immunology
- Oncology
- Biotechnology
Background:
- Bispecific antibodies (BsAbs) offer dual-targeting capabilities for enhanced therapeutic efficacy.
- Deregulation of c-MET is implicated in numerous human malignancies, correlating with poor prognosis.
- Programmed death-1 (PD-1) blockade is a key strategy in cancer immunotherapy.
Purpose of the Study:
- To identify and characterize a novel bispecific antibody targeting both c-MET and PD-1.
- To evaluate the therapeutic potential of this dual-targeting antibody as an anti-cancer agent.
Main Methods:
- Production and purification of the BsAb using 293E expression system and Protein A affinity chromatography.
- Assessment of binding specificity and affinity to PD-1 and c-MET via FACS and biolayer interferometry.
- Evaluation of anti-tumor activity, including cell proliferation inhibition, signaling pathway modulation, cytokine production, and cytotoxic effects.
Main Results:
- A novel BsAb targeting PD-1 and c-MET was successfully generated with high affinity (11.5 nM for PD-1, 9.09 nM for c-MET).
- The BsAb significantly enhanced IFN-γ production, inhibited c-MET pathway activation, and suppressed tumor cell proliferation.
- Comparable efficacy to existing treatments was observed, with dose-dependent cytotoxic activity against MKN45 cells.
Conclusions:
- A novel bispecific antibody targeting PD-1 and c-MET has been successfully developed.
- This BsAb demonstrates potential to redirect T cells for tumor cell killing and restore T cell function.
- The generated BsAb is a promising therapeutic candidate for various solid tumors.
