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Updated: Aug 11, 2026
Identifying Dysregulated Genes Induced by Kaposi's Sarcoma-associated Herpesvirus (KSHV)
Published on: September 14, 2010
[Construction and identification of recombinant vectors carrying herpes simplex virus thymidine kinase genes
Xiao-Ping Liu1, Bao-Jin Li, Chao Zhang
1Department of Hepatobilliary and Pancreatic Surgery, Shenzhen Hospital, Peking University, Shenzhen, Guangdong 518036, P. R. China. ninliu@163.com
Background & Objective:
Blocking tumor angiogenesis is a promising tumoricidal approach. Herpes simplex virus-thymidine kinase (HSV-tk) suicide gene could effectively damage neoplastic vascular endothelial cells. However, the killing effect of currently used HSV-tk system is non-specific due to employing cytomegalovirus (CMV) as its promoter. Kinase domain insert containing receptor (KDR), a kind of vascular endothelial growth factor receptor, is highly expressed on tumor vascular endothelial cells, but lowly expressed on normal tissues. This study was to construct a recombinant adenovirus containing KDR promoter-mediated HSV-tk suicide gene, and analyze its specific expression on vascular endothelial cells.
Methods:
KDR-tk fragment was subcloned into corresponding restriction sites of vector with directional cloning method using pAdeasy system to construct recombinant adenoviral plasmid containing KDR promoter-controlled HSV-tk gene (AdKDR-tk). After packaged and amplified in 293 cells, the virus was used to infect KDR-expressed human umbilical venous endothelial cells (HUVECs) and KDR-unexpressed liver cancer cell line HepG2. After administration of ganciclovir (GCV), proliferation of gene-transfected HUVECs and HepG2 cells was evaluated by MTT assay.
Results:
According to the results of restriction digestion, reverse transcription-polymerase chain reaction (RT-PCR) and PCR, the length, position and orientation of inserted genes were correct. The titer of the recombinant adenovirus was 1 x 10 (10) pfu/ml. Under infection index of 100, with the concentration of GCV increased from 0 to 50 microg/ml, the survival rates of AdKDR-tk-transfected HUVECs and HepG2 cells decreased from 100% to (28.94+/-5.67)% and (75.45+/-2.91)%, respectively (P<0.01).
Conclusion:
Adenovirus-mediated transfection of KDR promoter-HSV-tk gene could direct endothelial specific gene expression.
Insights
This study developed a targeted gene therapy using adenovirus to deliver the herpes simplex virus-thymidine kinase (HSV-tk) suicide gene. This approach specifically targets tumor vascular endothelial cells, offering a more precise cancer treatment strategy.
Area of Science:
- Oncolytic virotherapy
- Gene therapy
- Molecular oncology
Context:
- Tumor angiogenesis is a critical target for cancer treatment.
- Current suicide gene therapies lack specificity, leading to off-target effects.
- The herpes simplex virus-thymidine kinase (HSV-tk) suicide gene system shows promise for damaging neoplastic vascular endothelial cells.
Purpose:
- To construct a recombinant adenovirus carrying the HSV-tk suicide gene under the control of the Kinase domain insert containing receptor (KDR) promoter.
- To evaluate the specific expression and efficacy of the KDR promoter-mediated HSV-tk suicide gene system in targeting tumor vascular endothelial cells.
Summary:
- A recombinant adenovirus (AdKDR-tk) was successfully constructed, utilizing the KDR promoter to drive HSV-tk expression.
- In vitro studies showed that AdKDR-tk specifically targeted and induced cell death in KDR-expressing human umbilical venous endothelial cells (HUVECs) upon ganciclovir administration.
- Non-KDR-expressing HepG2 cells exhibited significantly higher survival rates, demonstrating the promoter's specificity.
Impact:
- This targeted gene therapy approach offers enhanced specificity for damaging tumor vasculature.
- The KDR promoter-mediated HSV-tk system represents a promising strategy for improving the efficacy and reducing side effects of cancer gene therapy.
- Further research could lead to novel therapeutic strategies for various cancers by exploiting tumor-specific promoters.
