Mice with the p53 tumor suppressor gene knocked out by gene targeting develop normally but have increased susceptibility to spontaneous tumorigenesis. We are characterizing and using nullizygous p53- knockout mice as a spontaneous tumor model to screen for nutritional and chemopreventive agents which can offset the increased risk of tumorigenesis resulting from the loss of tumor suppressor function. We are also analyzing ex vivo tissues and cells from these genetically susceptible mice to define the diet-gene interactions underlying efficacious interventions. We have reported that calorie restriction (CR), a potent inhibitor of many types of rodent tumors, significantly delays spontaneous tumor development in both p53-knockout and wild-type mice. In addition, we showed that while p53 status relates to the rapidity with which tumors develop (p53-knockout mice died, on average, approximately 60 weeks earlier than wild-type mice), the effect of CR, relative to ad libitum feeding, was p53-independent. We have also reported that the chemopreventive agent, dehydroepiandrosterone (DHEA) and its derivative 16-alpha-fluoro-androsten-17-one (8354), but not quercetin, all-trans-retinoic acid or d-limonene, also delay spontaneous tumor development in p53-knockout mice. Furthermore, CR, DHEA and 8354 each suppressed macrophage nitric oxide production, suggesting that a common denominator of interventions which suppress spontaneous tumorigenesis in p53-knockout mice is the ability to decrease oxidative stress. Studies are underway with putative anti-oxidative agents such as piroxicam (non-steroidal anti-inflammatory drug), genistein (a phytoestrogen), beta-carotene, alpha-tocopherol, 4- hydroxyphenylretinamide, and 9-cis-retinoic acid, to further test this hypothesis. Also currently in progress are molecular studies on tissues collected serially from control and treated (with a focus on CR, DHEA- or 8354- treated) mice to characterize the messenger RNA expression of cell cycle-related genes (Waf-1/p21, ras and myc); apoptosis-related genes (Bcl-2, Bax) as well as immunohistochemical markers of proliferation and apoptosis. We are also comparing differential messenger RNA expression in tissues from these mice using the differential display reverse transcription PCR technique. In addition, we are attempting to develop new models of low-dose chemical or hormone-induced lymphoma, prostate and colon tumors by testing the response of hemizygous p53-knockout mice to various regimes of nitrosomethylurea, dimethylhydrazine or testosterone.
