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A New Efficient Method for Production of Recombinant Antitumor Cytokine TRAIL and Its Receptor-Selective Variant
A V Yagolovich1,2, A A Artykov1,2, D A Dolgikh1,2
1Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, 117997, Russia.
Abstract:
The cytokine TRAIL induces apoptosis in tumor cells of various origin without affecting normal cells. Clinical trials of TRAIL-receptor (DR4 and DR5) agonists (recombinant TRAIL or death receptors antibodies) have largely failed because most human tumors were resistant to them. Currently, a second generation of agents targeted at TRAIL-R with increased efficiency has been developed. To this end, we have developed DR5-B, a variant of TRAIL selectively interacting with DR5. We have developed a new efficient method for production of TRAIL and DR5-B using expression of these proteins in Escherichia coli strain SHuffle B. The proteins were isolated from the cytoplasmic fraction of cells and purified to a high degree of homogeneity using metal-affinity and ion-exchange chromatography. The protein yield was 211 and 173 mg from one liter of cell culture for DR5-B and TRAIL, respectively, which significantly exceeded the results obtained by other methods. DR5-B killed tumor cells of different origin more efficiently and rapidly compared with TRAIL. The resulting preparations can be used for the study of TRAIL signaling pathways and in preclinical and clinical trials as antitumor agents.
Insights
Researchers developed DR5-B, a potent TRAIL variant, to overcome tumor resistance to TRAIL-based therapies. This new agent efficiently kills cancer cells and shows promise for cancer treatment development.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in cancer cells but faces resistance in clinical trials.
- Development of TRAIL-receptor agonists has been hindered by tumor resistance.
- A new generation of TRAIL-targeted agents is needed for improved cancer therapy.
Purpose of the Study:
- To develop a novel TRAIL variant, DR5-B, with enhanced efficacy against resistant tumors.
- To establish an efficient production method for TRAIL and DR5-B using bacterial expression.
- To evaluate the antitumor activity of DR5-B compared to TRAIL.
Main Methods:
- Engineered DR5-B, a TRAIL variant targeting DR5, for selective interaction.
- Utilized Escherichia coli strain SHuffle B for recombinant protein expression.
- Purified proteins using metal-affinity and ion-exchange chromatography.
- Assessed cell killing efficiency and speed of DR5-B versus TRAIL on tumor cells.
Main Results:
- Achieved high yields of DR5-B (211 mg/L) and TRAIL (173 mg/L) from bacterial cultures.
- DR5-B demonstrated superior and faster tumor cell killing compared to native TRAIL.
- Purified proteins were homogeneous and suitable for further studies.
Conclusions:
- DR5-B is a highly effective TRAIL variant with enhanced antitumor activity.
- The bacterial expression system provides an efficient method for producing TRAIL and DR5-B.
- DR5-B holds potential for preclinical and clinical applications as an anticancer agent.
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