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Induction of Invasive Transitional Cell Bladder Carcinoma in Immune Intact Human MUC1 Transgenic Mice: A Model for Immunotherapy Development
Published on: October 30, 2013
Response of bladder carcinoma cells to TRAIL and antisense oligonucleotide, Bcl-2 or clusterin treatments
Bo Sun1, Jacob A Moibi, Allan Mak
1Department of Surgery, Cross Cancer Institute, University of Alberta, Edmonton, Alberta, Canada.
Purpose:
Bladder transitional cell carcinoma is the second most common urological malignancy, of which 80% are superficial disease limited to the bladder. Superficial bladder transitional cell carcinoma has a high propensity for recurrence and progression after initial resection, necessitating adjuvant intravesical therapy. TRAIL (tumor necrosis factor-related apoptosis inducing ligand) can selectively induce apoptosis in most tumor cells while sparing normal cells. TRAIL drives not only the death receptor pathway, but also the mitochondrial pathway through Bid. Due to the anti-apoptotic functions of Bcl-2 and clusterin on the mitochondrial apoptotic pathway the effects of down-regulating these proteins were examined in partially TRAIL resistant bladder transitional cell carcinoma cell lines.
Materials And Methods:
Antisense oligonucleotides targeting Bcl-2 and clusterin were used alone or combined with TRAIL and cytotoxicity was examined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolim bromide) proliferation assay. Apoptotic pathway signals were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis/Western blotting after the various combination treatments. All statistical tests were 2 sided.
Results:
Although no direct correlation between TRAIL sensitivity and the relative expression levels of Bcl-2 and clusterin was found in the bladder transitional cell carcinoma cell lines examined, antisense oligonucleotide mediated the down-regulation of Bcl-2 and clusterin, increasing the sensitivity of the partially resistant cells to TRAIL. This was mediated through increased apoptotic signaling of the mitochondrial pathway, as evident by the increased activation of caspase-9 and 3, and cleaved DFF45. There was no benefit of combined antisense oligonucleotide therapy.
Conclusions:
This study provides proof of principle that TRAIL combined with antisense oligonucleotide-Bcl-2 may have potential as a novel future treatment strategy for bladder transitional cell carcinoma.
Insights
Down-regulating Bcl-2 and clusterin with antisense oligonucleotides enhanced TRAIL therapy for bladder cancer. This approach increased cancer cell death via the mitochondrial apoptotic pathway, showing potential for future bladder transitional cell carcinoma treatments.
Area of Science:
- Oncology
- Molecular Biology
- Cancer Therapeutics
Background:
- Bladder transitional cell carcinoma is a common urological malignancy with high recurrence rates after resection.
- Adjuvant intravesical therapy is often necessary for superficial bladder cancer.
- Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in tumor cells via death receptor and mitochondrial pathways.
Purpose of the Study:
- To investigate the potential of down-regulating anti-apoptotic proteins Bcl-2 and clusterin to enhance TRAIL-induced apoptosis in partially TRAIL-resistant bladder transitional cell carcinoma (BCC) cell lines.
- To examine the effects of antisense oligonucleotides targeting Bcl-2 and clusterin on BCC cell sensitivity to TRAIL.
Main Methods:
- Antisense oligonucleotides targeting Bcl-2 and clusterin were used alone and in combination with TRAIL.
- Cytotoxicity was assessed using the MTT proliferation assay.
- Apoptotic signaling was analyzed via Western blotting to detect key proteins in the mitochondrial pathway.
Main Results:
- Antisense oligonucleotides successfully downregulated Bcl-2 and clusterin in BCC cell lines.
- This downregulation increased the sensitivity of partially TRAIL-resistant cells to TRAIL.
- Enhanced apoptosis was mediated by increased activation of caspase-9 and caspase-3, indicating activation of the mitochondrial pathway. No benefit was observed from combined antisense oligonucleotide therapy.
Conclusions:
- Down-regulating Bcl-2 and clusterin via antisense oligonucleotides can overcome partial TRAIL resistance in bladder cancer cells.
- This strategy enhances apoptosis through the mitochondrial pathway, offering a potential novel therapeutic approach for bladder transitional cell carcinoma.
- Combined antisense oligonucleotide therapy did not provide additional benefit.
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