Expression and activity identification of bispecific antibody targeting PD-1/CD19
DOI：10.3872/j.issn.1007-385x.2021.04.007
- VernacularTitle:靶向抗PD-1/CD19双特异性抗体的表达与活性鉴定
- Author:
ZHAO Xiaocui
1
;
LI Ranran,
1
;
HU Yali,
1
;
LI Xiangguo
1
;
LI Jing
1
;
LI Feng
1
Author Information
1. Shunhao Cell Biotechnology [Tianjin] Co., Ltd., Tianjin 300308, China
- Publication Type:Journal Article
- Keywords:
bispecific antibody (BsAb);
PD-1;
CD19;
PD-1/PD-L1 interaction;
antibody-dependent cell-mediated cytotoxicity (ADCC);
tumor;
immunotherapy
- From:
Chinese Journal of Cancer Biotherapy
2021;28(4):359-364
- CountryChina
- Language:Chinese
-
Abstract:
[Abstract] Objective: To construct and purify the recombinant bispecific antibody (BsAb) targeting PD-1 and CD19 and evaluate its activity. Methods: With pCAR1 plasmid as the vector, the eukaryotic expression vector of anti-PD-1/CD19 BsAb was constructed by molecular cloning technology, and then transfected into mammalian cell line CHO-S by PEI reagent for transiently expressing antibody. The BsAb was purified by Affinity chromatography and then identified by SDS-PAGE and WB. The blocking activity of BsAb on PD-1/PD-L1 in vitro was detected by Luciferase reporter gene assay. The activity of antibody (BsAb)-dependent cell (PBMC)-mediated cytotoxicity (ADCC) in vitro was evaluated by lactate dehydrogenase (LDH) cytotoxicity assay. Results: The double plasmid eukaryotic expression vector pCAR1-19X3 was successfully constructed, and anti-PD-1/CD19 BsAb was successfully expressed in CHO-S cells, named pCAR1-19X3-TY. pCAR1-19X3-TY could effectively block the binding of PD-1 to its ligand PD-L1 in vitro, and the EC50 based on the dose-response curve was 0.306 μg/ml. ADCC results showed that pCAR1-19X3-TY could mediate the cytotoxicity of PBMC against Raji cells, and the curve showed a linear upward trend; when the effect/target ratio was 50∶1, the target cell lysis rate of pCAR1-19X3-TY was (38.9±0.3)%, which was not significantly different from that of the positive treatment group (46.7±4.9)% (P>0.05), but significantly higher than that of the negative control group (1.2±0.1)% (P<0.05). Conclusion: The recombinant anti-PD-1/CD19 BsAb can effectively block the binding of PD-1 and PD-L1 and activate PBMC mediated cytotoxicity against Raji cells. pCAR1-19X3-TY has the potential application value in the treatment of B-cell malignant tumor.
- Full text:20210407.pdf
