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Adenovirus-mediated p53 gene therapy in pediatric soft-tissue sarcoma cell lines: sensitization to cisplatin and
Hooman Ganjavi1, Matthew Gee, Aru Narendran
1Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada.
Abstract:
Sarcomas, or tumors of connective tissue, represent roughly 20% of childhood cancers. Although the cure rate for sarcomas in general has significantly improved in the last 10 years, there continue to be subgroups that are difficult to treat. High-grade or metastatic soft-tissue sarcomas and rhabdomyosarcomas (RMS) of the extremities remain therapeutic challenges and their prognosis is often poor. The future of sarcoma therapy will likely include molecular approaches including gene/protein expression profiling and gene-based therapy. Most sarcomas harbor defects in the p53 or pRb pathways. The tumor suppressor p53 is central to regulation of cell growth and tumor suppression and restoring wild-type p53 function in pediatric sarcomas may be of therapeutic benefit. Studies with adenoviral-mediated p53 gene transfer have been conducted in many cancer types including cervical, ovarian, prostatic and head and neck tumors. Studies of this approach, however, remain limited in pediatric cancers, including sarcomas. Using three viral constructs containing cDNA for wild-type p53, mutant p53 (C135S) and lacZ, we studied the effect of adenoviral-mediated gene therapy in four pediatric sarcoma cell lines, RD and Rh4 (RMS), Rh1 (Ewing's sarcoma) and A204 (undifferentiated sarcoma). Using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay, we have shown a dose-dependent decrease in cell viability 72 h post-treatment that occurs with Ad-wtp53 but not with Ad-mutp53. Cells treated with Ad-wtp53 show upregulation of the p53 downstream targets, p21(CIP1/WAF1) and bax. Growth curves demonstrate suppression of cell growth over a period of 4 days and cells treated with Ad-wtp53 demonstrate a significant increase in sensitivity to the chemotherapeutic agents, cisplatin and doxorubicin. Our results indicate that restoration of wild-type p53 function in pediatric sarcoma cells could provide a basis for novel approaches to treatment of this disease.
Insights
Restoring wild-type p53 function via adenoviral gene therapy significantly reduced pediatric sarcoma cell viability and enhanced chemotherapy sensitivity. This approach offers a promising new avenue for treating challenging pediatric sarcomas.
Area of Science:
- Oncology
- Molecular Biology
- Gene Therapy
Background:
- Sarcomas constitute 20% of childhood cancers, with high-grade or metastatic subtypes and rhabdomyosarcomas (RMS) presenting significant therapeutic challenges.
- Defects in the p53 or pRb pathways are common in sarcomas, highlighting the tumor suppressor p53's critical role in cell growth regulation and tumor suppression.
- While adenoviral-mediated p53 gene transfer has shown promise in various cancers, its application in pediatric sarcomas remains limited.
Purpose of the Study:
- To investigate the therapeutic potential of adenoviral-mediated gene therapy by restoring wild-type p53 function in pediatric sarcoma cell lines.
- To evaluate the effects of wild-type p53 gene transfer on sarcoma cell viability, downstream target expression, and sensitivity to chemotherapeutic agents.
Main Methods:
- Utilized three viral constructs: Ad-wtp53 (wild-type p53), Ad-mutp53 (mutant p53 C135S), and Ad-lacZ (control).
- Treated four pediatric sarcoma cell lines (RD, Rh4, Rh1, A204) with viral constructs.
- Assessed cell viability using the MTT assay, analyzed p53 downstream targets (p21, bax) via gene expression, and evaluated cell growth suppression and chemotherapy sensitivity (cisplatin, doxorubicin).
Main Results:
- Adenoviral-mediated transfer of wild-type p53 (Ad-wtp53) demonstrated a dose-dependent decrease in sarcoma cell viability, unlike Ad-mutp53.
- Ad-wtp53 treatment led to the upregulation of p53 downstream targets, p21(CIP1/WAF1) and bax.
- Growth curves indicated suppressed cell proliferation, and cells treated with Ad-wtp53 showed significantly increased sensitivity to cisplatin and doxorubicin.
Conclusions:
- Restoration of wild-type p53 function in pediatric sarcoma cells via adenoviral gene therapy is a viable strategy.
- This approach holds promise for developing novel therapeutic strategies for challenging pediatric sarcomas.
- The findings support further investigation into p53-based gene therapy for pediatric sarcoma treatment.
