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[Changes in etoposide-induced apoptosis of HeLa tumor cells transfected with antisense oligonucleotide for BCL-2
Ilona Bednarek1, Daniel Sypniewski, Joanna Solarz
1Zakładu Biotechnologii i Inzynierii Genetycznej Slaskiego Uniwersytetu Medycznego w Katowicach. dribednarek@sum.edu.pl
Unlabelled:
Overexpression of genes involved in proliferation, including genes of BCL family, is often found in cells of most malignant tumors. Currently developed strategies of cancer treatment include trials combining classical chemotherapy with silencing of genes expression using the gene therapy. The aim of the study was to induce silencing of BCL-2 gene expression in HeLa tumor cell line using antisense oligonucleotides (ASOs) technique.
Material And Methods:
Studies were carried out on in vitro HeLa cell cultures treated with etoposide. Cells were transfected by lipofection with ASO targeting BCL-2 mRNA. Effects of BCL-2 silencing were determined by Real-Time RT-PCR, proliferation/cytotoxicity MTT [3-(4,5-dimethylthiazol-2-yl)-2-5-dipheryl tetrazolium bromide] test, and by microscopic detection of apoptotic cells.
Results:
We showed a decrease in BCL-2 mRNA level in cells transfected with anti-BCL-2 ASO at concentration ranging from 50 to 1200 nM. Apoptotic cells were detected more frequently in transfected cultures compared with untreated controls. However, MTT tests did not display significant decrease of cell proliferation in the transfected cultures as compared with cultures treated with etoposide alone.
Conclusions:
1. These results indicate that the studied ASO sequence-specifically decreases BCL-2 expression in HeLa cells in vitro, although its action is limited mostly by transfection efficiency. 2. The use of the anti-BCL-2 oligonucleotide in combination with etoposide results in significant deacrease of proliferation in cell cultures and this phenomenon is a result of synergy between used chemotherapy and gene therapy.
Insights
Antisense oligonucleotides (ASOs) targeting BCL-2 mRNA reduced BCL-2 expression in HeLa cells. Combining ASO therapy with etoposide chemotherapy synergistically decreased tumor cell proliferation.
Area of Science:
- Molecular Biology
- Cancer Research
- Gene Therapy
Context:
- Overexpression of proliferation-associated genes, including the BCL family, is common in malignant tumors.
- Current cancer treatment strategies explore combining chemotherapy with gene silencing.
- Gene therapy offers a novel approach to target specific gene expressions in cancer cells.
Purpose:
- To investigate the efficacy of antisense oligonucleotides (ASOs) in silencing BCL-2 gene expression in the HeLa tumor cell line.
- To evaluate the combined effect of BCL-2 silencing and etoposide chemotherapy on cancer cell proliferation and apoptosis.
Summary:
- The study utilized antisense oligonucleotides (ASOs) targeting BCL-2 mRNA in vitro HeLa cell cultures treated with etoposide.
- BCL-2 mRNA levels decreased in cells transfected with anti-BCL-2 ASO, and apoptosis increased.
- While ASO alone showed limited impact on proliferation, its combination with etoposide resulted in significant proliferation reduction due to synergistic effects.
Impact:
- Demonstrates sequence-specific BCL-2 gene silencing in HeLa cells using ASOs, though limited by transfection efficiency.
- Highlights the synergistic potential of combining gene therapy (anti-BCL-2 ASO) with chemotherapy (etoposide) for enhanced cancer treatment.
- Provides a foundation for developing targeted gene silencing strategies in combination cancer therapies.
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