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Analysis of aDR5scFv with Specific Identification and Function
Xiaofeng Cheng1,2, Qingyu Meng2, Chunling Gao1
11 Radiotherapy Department, ChengGong Hospital, Xiamen University , Xiamen, Fujian, China .
Abstract:
Death receptor 5 (DR5) can selectively induce cell death in a wide variety of tumor cells. However, at least certain versions of the recombinant soluble TRAIL (sTRAIL) or anti-DR5 monoclonal antibody (mAb) are also shown to cause apoptosis in normal cells (especially in hepatocytes), hampering its clinical use for cancer therapy. Recently, the development of small recombinant antibody fragments as high-affinity therapeutic reagents with reduced immunogenicity has come under the spotlight. A popular format of engineered recombinant antibody fragment is the single-chain fixed-variable (scFv) molecule, in which the VH and VL regions of the parental antibody are joined by a polypeptide linker. The scFv fragment retains the target specificity and antigen binding affinity of the intact antibody, whereas it can be genetically designed and produced in large quantities by ectopically expressing both VH and VL regions from a single cDNA in cells. In this study, an aDR5scFv was constructed and expressed, and it was conformed so that it could recognize and bind eDR5 specifically. The therapeutic effects on human lung adenocarcinoma cells lines 973 in vitro and in vivo were detected by MTT assay, flow cytometry, hematoxylin and eosin staining, and TUNEL assay. aDR5scFv was able to induce 973 cell apoptosis in an in vitro system. The protein expressions of caspase-3, Bax, and cytochrome c were raised, and aDR5scFv also inhibited tumor growth in mice with its effect as well as with radiotherapy. It is concluded that aDR5scFv could possibly be considered as a novel therapeutic candidate for the treatment of tumors.
Insights
Engineered antibody fragments targeting Death Receptor 5 (DR5) offer a promising cancer therapy. A novel anti-DR5 single-chain variable fragment (aDR5scFv) selectively induced apoptosis in lung cancer cells and inhibited tumor growth in vivo.
Area of Science:
- Oncology
- Immunology
- Molecular Biology
Background:
- Recombinant soluble TRAIL and anti-DR5 monoclonal antibodies show potential for cancer therapy by inducing cell death via Death Receptor 5 (DR5).
- However, these agents can cause apoptosis in normal cells, particularly hepatocytes, limiting their clinical application.
- Engineered antibody fragments, such as single-chain variable fragments (scFv), are emerging as therapeutic candidates due to high affinity, specificity, and reduced immunogenicity.
Purpose of the Study:
- To construct and express a novel anti-DR5 single-chain variable fragment (aDR5scFv).
- To evaluate the therapeutic efficacy of aDR5scFv against human lung adenocarcinoma cells (973) in vitro and in vivo.
- To assess the potential of aDR5scFv as a novel cancer therapeutic agent.
Main Methods:
- Construction and expression of aDR5scFv targeting human DR5 (eDR5).
- In vitro assessment using MTT assay and flow cytometry to detect apoptosis in 973 cells.
- In vivo evaluation in mice using hematoxylin and eosin staining and TUNEL assay to assess tumor growth inhibition and apoptosis, in combination with radiotherapy.
Main Results:
- The constructed aDR5scFv specifically recognized and bound to eDR5.
- aDR5scFv effectively induced apoptosis in 973 lung adenocarcinoma cells in vitro.
- In vivo studies demonstrated that aDR5scFv inhibited tumor growth in mice, with comparable effects to radiotherapy, and increased expression of apoptosis-related proteins like caspase-3, Bax, and cytochrome c.
Conclusions:
- The engineered aDR5scFv exhibits specific binding to eDR5 and potent anti-tumor activity.
- aDR5scFv demonstrates the ability to induce apoptosis in lung cancer cells both in vitro and in vivo.
- This novel aDR5scFv represents a potential new therapeutic candidate for cancer treatment, possibly in combination therapies.
