Restoration of p53 function in anaplastic Wilms' tumor

Background/Purpose: Recent studies have reported a high incidence of p53 mu tations in anaplastic Wilms' tumors (WT). Restoration of the normal p53 sta te by current gene therapy techniques is thus an attractive potential mode of therapy for this tumor, which is poorly responsive to standard therapy. The purpose of this study is to determine whether gene delivery of normal p 53 is possible and to characterize the subsequent effect of restoring the w ild-type p53 state. Methods: Anaplastic WT RM1 cells (mutant p53) were transduced with replicat ion-deficient adenoviral vectors containing either the wild-type p53 gene ( rAd-p53) or the gene encoding a green fluorescent protein (rAd-GFP). The tr ansduction efficiency of adenovirus for RM1 cells was determined by flow cy tometric analysis of rAd-GFP-transduced cells. The effect of p53 transducti on on cell viability was evaluated using a colorimetric proliferation assay . Apoptosis was evaluated by labeling DNA breaks using a TUNEL assay (Apo-D irect kit). Results: Cells treated with increasing concentrations of viral particles re lative to tumor cells (multiplicity of infection-MOI) showed a dose-depende nt increase in the number of cells transduced. Twenty-four hours after vira l treatment, the percentage of cells transduced for MOIs of 10, 50, 100, an d 500 was 29.5, 60.9, 74.6, and 92.4, respectively; at 48 hours the percent age of cells transduced increased to 70.8, 90.7, 93.7, and 96.3, respective ly. Viral treatment at an MOI of 50 reduced cell proliferation by 10% at 17 hours and 97% at 5 days; at an MOI of 100, the relative reduction in proli feration was 15% and 99.8%, respectively. When assayed, 30% of cells became apoptotic at an MOI of 50, and 48% at an MOI of 100. Conclusions: Highly efficient delivery of the p53 tumor suppressor gene by adenoviral vector to anaplastic WT is possible. Subsequent restoration of t he normal p53 state results in reduced viability and increased apoptosis. G ene replacement of p53 may represent a novel therapeutic agent for anaplast ic Wilms' tumors. J Pediatr Surg 36:43-50. Copyright (C) 2001] by W.B. Saun ders Company.