Therapeutic potential of p53 restoration in spontaneous tumors

Over 80% of p53 mutations found in human cancers are p53 missense mutations. Recent studies have shown that p53 restoration leads to tumor regression in mice with p53 deletions, but the therapeutic efficacy of p53 restoration in tumors containing p53 missense mutations has not been evaluated. Since p53 mutant such as p53R172H has gain-of-function activities and dominant-negative effect that repress wild type p53, the activity of restored wild-type p53 might be compromised by the mutant p53 in tumors. We hypothesized that p53 restoration in tumors with the p53R172H mutation may be less therapeutically effective as p53 restoration in tumors null for p53. I tested this hypothesis by comparison of the therapeutic outcomes of p53 restoration in mice with spontaneous tumors that either lacked p53 or contained the p53R172H mutation. While p53 restoration causes tumor regression in mice lacking p53, the same p53 restoration halts tumor progression in mice with the p53R172H mutation. This phenotypic difference suggests a dominant-negative activity of the mutant p53. Moreover, I showed that the mutant p53 only inhibits part of the activity of the restored wild-type p53 and that the remaining wild-type activity still causes a delay in tumor progression. We conclude that p53 restoration has therapeutic potential in p53R172H tumors via suppression of tumor progression. This knowledge is of critical importance for p53 targeted cancer therapy because many patients with cancers harbor p53 missense mutations rather p53-null mutations. Since p53R172H mutation represents one of the most frequent and potent p53 missense mutations observed in human cancers, the current findings implicates that p53 restoration may be therapeutically important not only in human cancers characterized by loss of p53 alleles but also in those in which p53 missense mutations play an important pathogenetic role. ^

IL -12 inhibits the metastatic potential of osteosarcoma cells and induces tumor regression via a mechanism involving Fas /Fas ligand pathway

Despite multiple changes in the adjuvant chemotherapy regimens used to treat osteosarcoma (OS), the 2-year metastasis-free survival has remained at 65–70% for the past 10 years. Characterizing the molecular determinants that permit metastatic spread of tumor cells is a crucial element in developing new approaches for the treatment of osteosarcoma. Since OS metastasizes almost exclusively to the lung, an organ with constitutive Fas ligand (FasL) expression, we hypothesized that the expression of Fas (CD95, APO-1) by OS cells may play a role in the ability of these cells to form lung metastases. Fas expression was quantified in human SAOS-2 OS cells and selected variants (LM2, LM4, LM5, LM6, LM7). Using northern blot, FACS and RT-PCR analysis, low Fas expression was found to correlate with higher metastatic potential in these cell lines. The highly metastatic LM7 cell line was transfected with the full-length human Fas gene and injected into athymic nude mice. The median number of metastatic nodules per mouse fell from over 200 to 1.1 and the size of the nodules decreased from a range of 0.5–9.0 mm to less than 0.5 mm in the Fas-transfected cell line compared to the native LM7 cell line. Additionally, the subsequent incidence of lung metastases was lower in the Fas-expressing cell line. IL-12 was seen to upregulate Fas expression in the highly metastatic LM sublines in vitro. To visualize the effects of IL-12 in vivo, nude mice were injected with LM7 cells and treated biweekly for 4 weeks with Ad.mIL-12, saline control or Ad.βgal. Lung sections were analyzed via immunchistochemistry for Fas expression. A higher expression of Fas was found in tumors from mice receiving IL-12. To study the mechanism by which IL-12 upregulates Fas, LM7 cells were transfected with a luciferase reporter gene construct containing the full-length human fas promoter. Treatment with IL-12 increased luciferase activity. We therefore conclude that IL-12 influences the metastatic potential of OS cells by upregulating the fas promoter, resulting in increased cell surface Fas expression and susceptibility to Fas-induced cell death. ^