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Published on: August 7, 2018
Construction and validation of a novel tumor necrosis factor-related apoptosis-inducing ligand mutant MuR5S4-TR
Yaqin Zhao1, Jianping He1, Xia Liu1
1Department of Medical Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu, China.
Aim:
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can selectively kill tumor cells but has no significant effect on normal cells. However, the use of TRAIL is limited for resistance by more than 50% of the tumor cell lines. It's very important to develop a more efficient form of TRAIL for cancer treatment.
Methods:
The N-terminal in soluble fragments (114-281aa) of TRAIL was redesigned to construct a novel TRAIL mutant-MuR5S4-TR. The Cell Counting Kit-8 method to explore the antitumor effects. The potential mechanisms were also explored.
Results:
Novel TRAIL mutant with cell-penetrating peptides (CPP) like and Second mitochondria-derived activator of caspases (Smac) like structure-MuR5S4-TR was successfully constructed. The prokaryotic expression system was successfully built, and the MuR5S4-TR was purified and reconfirmed by western blot. MuR5S4-TR could enhance the antitumor effects of TRAIL in most of the cancer cell lines significantly, NCI-H460 lung cancer cell line, for instance. After MuR5S4-TR treatment, the expressions of death receptor 4 (DR4), DR5, Caspase-8, and cleaved Caspase-3 were remarkably increased, however, there was no significant difference in X-linked inhibitor of apoptosis expression.
Conclusion:
We constructed a novel TRAIL mutant with CPP-like and Smac-like structure-MuR5S4-TR. The MuR5S4-TR showed significantly stronger antitumor effects than TRAIL in many tumor cell lines. The MuR5S4-TR showed strong antitumor effects both in vitro and in vivo. This preliminary study implies that MuR5S4-TR may be a more efficient form of TRAIL for cancer therapy.
Insights
A novel TRAIL mutant, MuR5S4-TR, was engineered to overcome cancer cell resistance. This enhanced TRAIL variant demonstrates potent antitumor effects, offering a promising new avenue for cancer therapy.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in tumor cells.
- TRAIL resistance in over 50% of cancer cell lines limits its therapeutic efficacy.
- Development of enhanced TRAIL variants is crucial for effective cancer treatment.
Purpose of the Study:
- To construct and characterize a novel TRAIL mutant, MuR5S4-TR, with enhanced anti-cancer properties.
- To investigate the antitumor effects and underlying mechanisms of MuR5S4-TR.
- To evaluate MuR5S4-TR as a potential improved therapeutic agent for cancer.
Main Methods:
- Redesigning the N-terminal fragment of TRAIL to create the MuR5S4-TR mutant.
- Utilizing a prokaryotic expression system for protein production and purification.
- Assessing antitumor activity using the Cell Counting Kit-8 assay and western blot analysis.
Main Results:
- Successfully constructed and purified the novel TRAIL mutant MuR5S4-TR.
- MuR5S4-TR significantly enhanced antitumor effects in various cancer cell lines, including NCI-H460.
- Treatment with MuR5S4-TR upregulated death receptors (DR4, DR5) and apoptosis-related proteins (Caspase-8, cleaved Caspase-3).
Conclusions:
- A novel TRAIL mutant, MuR5S4-TR, incorporating cell-penetrating peptide-like and Smac-like structures was successfully developed.
- MuR5S4-TR exhibits superior antitumor activity compared to native TRAIL in vitro and in vivo.
- MuR5S4-TR represents a promising candidate for more effective cancer therapy.
