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Herpes simplex virus thymidine kinase/ganciclovir-mediated apoptotic death of bystander cells
W Hamel1, L Magnelli, V P Chiarugi
1Preuss Laboratory Molecular Neuro-oncology, Brain Tumor Research Center, University of California, USA.
Abstract:
An emerging strategy for cancer gene therapy involves the transfer of the herpes simplex virus thymidine kinase (HSV-tk) gene into tumor cells, rendering them susceptible to the cytotoxic effects of ganciclovir. The observation that HSV-tk-expressing cells can also induce cell death in neighboring cells, which do not express HSV-tk, has been called the bystander effect. Gap junction-mediated transfer of cytotoxic molecules to bystander cells may be an important mechanism of bystander cell death, although others have suggested a role for phagocytosis. In this study, we evaluated the mode of cell death in bystander cells. We detected apoptosis in bystander cells and found that bystander cell death could be inhibited by BCL2 expression. We determined that ganciclovir incubations for 10 h were sufficient to induce cell death in most bystander cells cocultured with HSV-tk-expressing cells. During this period, no phagocytosis was detected, although it was obvious at later stages.
Insights
Cancer gene therapy using herpes simplex virus thymidine kinase (HSV-tk) triggers bystander cell death, primarily through apoptosis. This cell death, induced by ganciclovir, can be blocked by BCL2 expression and occurs rapidly, preceding phagocytosis.
Area of Science:
- Oncology
- Molecular Biology
- Gene Therapy
Background:
- Cancer gene therapy utilizes the herpes simplex virus thymidine kinase (HSV-tk) gene to sensitize tumor cells to ganciclovir.
- The HSV-tk system exhibits a bystander effect, where HSV-tk-expressing cells induce death in neighboring non-expressing cells.
- Potential mechanisms for the bystander effect include gap junction-mediated transfer and phagocytosis.
Purpose of the Study:
- To investigate the specific mode of cell death in bystander cells within the HSV-tk/ganciclovir cancer gene therapy model.
- To determine the temporal relationship between cell death mechanisms and the presence of phagocytosis.
Main Methods:
- Co-culture of HSV-tk-expressing cells with non-expressing bystander cells.
- Treatment with ganciclovir and assessment of cell death markers.
- Evaluation of apoptosis using specific assays.
- Monitoring for phagocytosis and inhibition by BCL2 expression.
Main Results:
- Apoptosis was detected in bystander cells following ganciclovir treatment.
- BCL2 expression was found to inhibit bystander cell death.
- Ganciclovir incubation for 10 hours was sufficient to induce significant bystander cell death.
- Phagocytosis was not detected during the initial 10-hour incubation period but was evident at later time points.
Conclusions:
- Apoptosis is a primary mechanism driving bystander cell death in the HSV-tk/ganciclovir system.
- The bystander effect operates rapidly, preceding significant phagocytosis.
- BCL2 can modulate the extent of bystander cell death, offering a potential target for therapeutic intervention.