Mechanistic studies of mouse skin tumor promotion by chrysarobin

Since the anthrone chrysarobin oxidizes and generates free radicals, investigations were conducted to assess a possible role for free radicals or reactive oxygen species (ROS) in skin tumor promotion by chrysarobin. Epidermal glutathione levels were not noticeably altered by chrysarobin, nor did a glutathione-depleting agent enhance promotion by chrysarobin. Multiple applications of chrysarobin increased lipid peroxide levels in mouse epidermis two-fold as compared with controls. The antioxidant $\alpha$-tocopherol and the lipoxygenase inhibitor nordihydroguaiaretic acid both inhibited production of lipid peroxides by chrysarobin. The antioxidants $\alpha$-tocopherol acetate and ascorbyl palmitate effectively inhibited promotion and promoter-related effects induced by chrysarobin. Since prooxidant states can lead to increases in intracellular Ca$\sp{2+}$, the effect of two Ca$\sp{2+}$ antagonists, verapamil and TMB-8, on chrysarobin-induced promotion and promoter-related effects were investigated. Both Ca$\sp{2+}$ antagonists inhibited promotion and promoter-related effects induced by chrysarobin, suggesting a possible role for intracellular Ca$\sp{2+}$ alterations in chrysarobin-tumor promotion. Since radical generating compounds are reported to possess the ability to enhance progression of papillomas to squamous cell carcinomas (SCCs), the effects of chrysarobin on papilloma development were tested. Growth kinetics and regression of papillomas generated with limited promotion with chrysarobin were similar to what was reported for the nonradical generating promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) (Aldaz et al., 1991). To test the chrysarobin's ability to enhance progression of pre-existing papillomas to SCCs, tumors were generated by initiation with dimethylbenz (a) anthracene and promotion with TPA. Then mice were treated with chrysarobin, TPA or acetone for 45 weeks. When mice treated with chrysarobin were compared to mice treated continually with TPA with similar numbers of papillomas, the number of papillomas that progressed to SCCs was similar, suggesting that papilloma burden influences the progression of papillomas to SCCs, rather than radical production. In summary, the present study suggests that chrysarobin produces oxidative stress in mouse epidermis as indicated by the generation of lipid peroxides. Antioxidants inhibited production of lipid peroxides and tumor promotion by chrysarobin. Collectively, these data suggest a role for free radicals or ROS in tumor promotion by chrysarobin. ^

Tp53-dependent repression of cell cycle target genes: Global and mechanistic analysis

Most studies of p53 function have focused on genes transactivated by p53. It is less widely appreciated that p53 can repress target genes to affect a particular cellular response. There is evidence that repression is important for p53-induced apoptosis and cell cycle arrest. It is less clear if repression is important for other p53 functions. A comprehensive knowledge of the genes repressed by p53 and the cellular processes they affect is currently lacking. We used an expression profiling strategy to identify p53-responsive genes following adenoviral p53 gene transfer (Ad-p53) in PC3 prostate cancer cells. A total of 111 genes represented on the Affymetrix U133A microarray were repressed more than two fold (p ≤ 0.05) by p53. An objective assessment of array data quality was carried out using RT-PCR of 20 randomly selected genes. We estimate a confirmation rate of >95.5% for the complete data set. Functional over-representation analysis was used to identify cellular processes potentially affected by p53-mediated repression. Cell cycle regulatory genes exhibited significant enrichment (p ≤ 5E-28) within the repressed targets. Several of these genes are repressed in a p53-dependent manner following DNA damage, but preceding cell cycle arrest. These findings identify novel p53-repressed targets and indicate that p53-induced cell cycle arrest is a function of not only the transactivation of cell cycle inhibitors (e.g., p21), but also the repression of targets that act at each phase of the cell cycle. The mechanism of repression of this set of p53 targets was investigated. Most of the repressed genes identified here do not harbor consensus p53 DNA binding sites but do contain binding sites for E2F transcription factors. We demonstrate a role for E2F/RB repressor complexes in our system. Importantly, p53 is found at the promoter of CDC25A. CDC25A protein is rapidly degraded in response to DNA damage. Our group has demonstrated for the first time that CDC25A is also repressed at the transcript level by p53. This work has important implications for understanding the DNA damage cell cycle checkpoint response and the link between E2F/RB complexes and p53 in the repression of target genes. ^

A movement from traditional malarial chemoprophylaxis to the use of intermittent preventive treatment as a method of protection for children diagnosed with sickle-cell disease: A proposal for consideration based on a systematic review of the literature

Approximately 200,000 African children are born with sickle-cell anemia each year. Research has shown that individuals with hemoglobin disorders, particularly sickle-cell anemia, have increased susceptibility to contracting malaria. Currently it is recommended that patients diagnosed with sickle-cell anemia undergo malaria chemoprophylaxis in order to decrease their chances of malarial infection. However, studies have shown that routine administration of these drugs increases the risk of drug resistance and could possibly impair the development of naturally acquired immunity. Clinical trials have shown intermittent preventive treatment (IPT) to be an effective method of protection against malaria. The objective of this report was to review previously conducted clinical trials that study the effects of intermittent preventive treatment on malaria and anemia in infants and children. Based on the review, implications for its appropriateness as a protective measure against malaria for infants and children diagnosed with sickle-cell disease were provided.^ The 18 studies reviewed were randomized controlled trials that focused on IPT’s effect on malaria (7 studies), anemia (1 study), or both (8 studies). In addition to these 16, one study looks at IPT’s effect on molecular resistance to malaria, and another study is a follow-up to a study in order to review IPT’s potential to cause a rebound effect. The 18 th study in this review specifically looks at IPT’s protective efficacy in children with SCA. The studies in this report were restricted to randomized controlled trials that have been performed from 2000 to 2010. Reports on anemia were included to illustrate possible added benefits of the use of IPT specific to burdens associated with SCA other than malaria susceptibility. The outcomes of these studies address several issues of concern involving the administration of IPT: protective efficacy (in reference to age, seasonal versus perennial malaria regions, and overall effectiveness against malaria and anemia), drug resistance, drug rebound effect, drug side-effects, and long-term effects. Overall, these showed that IPT has a significant level of protective efficacy against malaria and/or anemia in children. More specifically, the IPT study evaluating children diagnosed with sickle-cell anemia proved IPT to be a more effective method of protection than traditional chemoprophylaxis. ^