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Wild-type p53 gene transfection in human cultured sarcomas: effect of CDDP
K Endo1, I Kuratate, M Watanabe
1First Department of Pathology, Faculty of Medicine, Tottori University, Yonago, Tottori, Japan.
Abstract:
We examined the susceptibility of five human bone and soft tissue sarcoma cell lines to transfection with recombinant p53 adenovirus vector (AxCA-p53). Transfection efficiency was more than 90% at 72 h with AxCA-lacZ at a multiplicity of infection (MOI) of 50 in all the cell lines, except for MG-63 (p53 gene mutated) cells. Western blot analysis showed overexpression of both P21/Waf1 and Bax protein in all the cell lines, implying sufficient and successful p53 gene transfection. AxCA-p53 transfection at MOI of 50 resulted in a significant decline of viable cells at 72 h, due to apoptosis, in NY (mutated) and Saos-2 (deletion), but not in the other three lines. The two apoptosis-induced cell lines showed a gradual increase in Bax expression up to 72 h and non-detectable expression of Bcl-XL from 48 h, suggesting the involvement of an apoptosis-inducing mechanism. Pre-treatment with cis-diamminedichloroplatinum (II) (CDDP) at 0.1 microg/ml significantly suppressed tumor cell viability in NY and HuO-3N1 (mutated), but not in the other three lines including HT-1080 carrying the wild-type p53 gene, implying the existence of different mechanisms for the tumor suppressive effect of p53 gene transfection and CDDP. These results indicate that wild-type p53 gene transfection with CDDP is a promising therapy for some, but not all, non-resectable bone-and soft tissue sarcomas, regardless of intrinsic p53 gene status.
Insights
Recombinant p53 adenovirus vector (AxCA-p53) successfully transfected sarcoma cells, inducing apoptosis and reducing viability in some cell lines. This gene therapy, combined with cis-diamminedichloroplatinum (II) (CDDP), shows promise for treating non-resectable sarcomas.
Area of Science:
- Oncology
- Molecular Biology
- Gene Therapy
Background:
- Sarcomas are cancers arising from connective tissues.
- The p53 tumor suppressor gene plays a critical role in cell cycle regulation and apoptosis.
- Gene therapy offers a potential strategy for cancer treatment.
Purpose of the Study:
- To evaluate the efficacy of recombinant p53 adenovirus vector (AxCA-p53) in human bone and soft tissue sarcoma cell lines.
- To investigate the role of p53 gene transfection and cis-diamminedichloroplatinum (II) (CDDP) in inducing apoptosis and suppressing tumor cell viability.
- To explore the mechanisms underlying p53 gene therapy in different sarcoma subtypes.
Main Methods:
- Transfection of five human sarcoma cell lines with AxCA-p53.
- Assessment of transfection efficiency using AxCA-lacZ.
- Western blot analysis to detect P21/Waf1 and Bax protein expression.
- Evaluation of cell viability and apoptosis induction.
- Pre-treatment with CDDP to assess combined therapeutic effects.
Main Results:
- High transfection efficiency (>90%) achieved in most cell lines.
- AxCA-p53 transfection induced apoptosis and reduced viable cells in NY (mutated) and Saos-2 (deletion) cell lines.
- Apoptosis induction correlated with increased Bax and decreased Bcl-XL expression.
- CDDP pre-treatment suppressed tumor cell viability in NY and HuO-3N1 (mutated) cell lines.
- Therapeutic effects of p53 gene transfection and CDDP varied among cell lines, irrespective of p53 gene status.
Conclusions:
- Wild-type p53 gene transfection can induce apoptosis in specific sarcoma cell lines.
- Combined therapy with p53 gene transfection and CDDP shows potential for treating non-resectable sarcomas.
- The effectiveness of this combined therapy may depend on the specific sarcoma subtype and its intrinsic p53 gene status.