Chemotherapy and radiation therapy for cancer often have severe side effects that limit their efficacy. Because these effects are in part determined by p53-mediated apoptosis, temporary suppression of p53 has been suggested as a therapeutic strategy to prevent damage of normal tissues during treatment of p53-deficient tumors. To test this possibility, a small molecule was isolated for its ability to reversibly block p53-dependent transcriptional activation and apoptosis. This compound, pifithrin-alpha, protected mice from the lethal genotoxic stress associated with anticancer treatment without promoting the formation of tumors. Thus, inhibitors of p53 may be useful drugs for reducing the side effects of cancer therapy and other types of stress associated with p53 induction.

Ionizing radiation (IR) has proven to be a powerful medical treatment in the fight against cancer. Rational and effective use of its killing power depends on understanding IR-mediated responses at the molecular, cellular and tissue levels. Tumour cells frequently acquire defects in the molecular regulatory mechanisms of the response to IR, which sensitizes them to radiation therapy. One of the key molecules involved in a cell's response to IR is p53. Understanding these mechanisms indicates new rational approaches to improving cancer treatment by IR.

p53-dependent apoptosis contributes to the side effects of cancer treatment, and genetic or pharmacological inhibition of p53 function can increase normal tissue resistance to genotoxic stress. It has recently been shown that p53 can induce apoptosis through a mechanism that does not depend on transactivation but instead involves translocation of p53 to mitochondria. To determine the impact of this p53 activity on normal tissue radiosensitivity, we isolated a small molecule named pifithrin-mu (PFTmu, 1) that inhibits p53 binding to mitochondria by reducing its affinity to antiapoptotic proteins Bcl-xL and Bcl-2 but has no effect on p53-dependent transactivation. PFTmu has a high specificity for p53 and does not protect cells from apoptosis induced by overexpression of proapoptotic protein Bax or by treatment with dexamethasone (2). PFTmu rescues primary mouse thymocytes from p53-mediated apoptosis caused by radiation and protects mice from doses of radiation that cause lethal hematopoietic syndrome. These results indicate that selective inhibition of the mitochondrial branch of the p53 pathway is sufficient for radioprotection in vivo.

Large parts of mammalian genomes are transcriptionally inactive and enriched with various classes of interspersed and tandem repeats. Here we show that the tumor suppressor protein p53 cooperates with DNA methylation to maintain silencing of a large portion of the mouse genome. Massive transcription of major classes of short, interspersed nuclear elements (SINEs) B1 and B2, both strands of near-centromeric satellite DNAs consisting of tandem repeats, and multiple species of noncoding RNAs was observed in p53-deficient but not in p53 wild-type mouse fibroblasts treated with the DNA demethylating agent 5-aza-2'-deoxycytidine. The abundance of these transcripts exceeded the level of β-actin mRNA by more than 150-fold. Accumulation of these transcripts, which are capable of forming double-stranded RNA (dsRNA), was accompanied by a strong, endogenous, apoptosis-inducing type I IFN response. This phenomenon, which we named "TRAIN" (for "transcription of repeats activates interferon"), was observed in spontaneous tumors in two models of cancer-prone mice, presumably reflecting naturally occurring DNA hypomethylation and p53 inactivation in cancer. These observations suggest that p53 and IFN cooperate to prevent accumulation of cells with activated repeats and provide a plausible explanation for the deregulation of IFN function frequently seen in tumors. Overall, this work reveals roles for p53 and IFN that are key for genetic stability and therefore relevant to both tumorigenesis and the evolution of species.interferon alpha-beta receptor | RNA sequencing | epigenetic repression | microarray hybridization

To analyze the involvement of p53‐dependent transcriptional activation in normal development and in response to DNA damage in vivo, we created transgenic mice with a lacZ reporter gene under the control of a p53‐responsive promoter. Five independent strains showed similar patterns of transgene expression. In untreated animals, lacZ expression was limited to the developing nervous system of embryos and newborn mice and was strongly decreased in the adult brain. γ‐irradiation or adriamycin treatment induced lacZ expression in the majority of cells of early embryos and in the spleen, thymus and small intestine in adult mice. Transgene expression was p53 dependent and coincided with the sites of strong p53 accumulation. The lacZ‐expressing tissues and early embryos, unlike other adult tissues and late embryos, are characterized by high levels of p53 mRNA expression and respond to DNA damage by massive apoptotic cell death. Analysis of p53‐null mice showed that this apoptosis is p53 dependent. These data suggest that p53 activity, monitored by the reporter lacZ transgene, is the determinant of radiation and drug sensitivity in vivo and indicate the importance of tissue and stage specificity of p53 regulation at the level of mRNA expression.

Ionizing radiation (IR) induces p53-dependent apoptosis in radiosensitive tissues, suggesting that p53 is a determinant of radiation syndromes. In fact, p53-deficient mice survive doses of IR that cause lethal hematopoietic syndrome in wild-type animals. Surprisingly, p53 deficiency results in sensitization of mice to higher doses of IR, causing lethal gastro-intestinal (GI) syndrome. While cells in the crypts of p53-wild-type epithelium undergo prolonged growth arrest after irradiation, continuous cell proliferation ongoing in p53-deficient epithelium correlates with accelerated death of damaged cells followed by rapid destruction of villi and accelerated lethality. p21-deficient mice are also characterized by increased sensitivity to GI syndrome-inducing doses of IR. We conclude that p53/p21-mediated growth arrest plays a protective role in the epithelium of small intestine after severe doses of IR. Pharmacological inhibition of p53 by a small molecule that can rescue from lethal hematopoietic syndrome has no effect on the lethality from gastro-intestinal syndrome, presumably because of a temporary and reversible nature of its action.

Chronic inflammation is known to promote cancer, suggesting that negative regulation of inflammation is likely to be tumor suppressive. We found that p53 is a general inhibitor of inflammation that acts as an antagonist of nuclear factor kappaB (NFkappaB). We first observed striking similarities in global gene expression profiles in human prostate cancer cells LNCaP transduced with p53 inhibitory genetic element or treated with TNF, suggesting that p53 inhibits transcription of TNF-inducible genes that are largely regulated by NFkappaB. Consistently, ectopically expressed p53 acts as an inhibitor of transcription of NFkappaB-dependent promoters. Furthermore, suppression of inflammatory response by p53 was observed in vivo in mice by comparing wild-type and p53 null animals at molecular (inhibition of transcription of genes encoding cytokines and chemokines, reducing accumulation of reactive oxygen species and protein oxidation products), cellular (activation of macrophages and neutrophil clearance) and organismal (high levels of metabolic markers of inflammation in tissues of p53-deficient mice and their hypersensitivity to LPS) levels. These observations indicate that p53, acting through suppression of NFkappaB, plays the role of a general "buffer" of innate immune response in vivo that is well consistent with its tumor suppressor function and frequent constitutive activation of NFkappaB in tumors.

Numerous observations indicate a strong link between chronic inflammation and cancer. This link is supported by substantial experimental evidence indicating mutual negative regulation of NF-κB, the major regulator of inflammation, and p53, the major tumor suppressor. This antagonistic relationship reflects the opposite principles of the physiological responses driven by these transcription factors, which act as sensors and mediators of intrinsic and extrinsic cell stresses, respectively. Constitutive activation of NF-κB, the underlying cause of chronic inflammation, is a common acquired characteristic of tumors. A variety of experimental methods have been used to demonstrate that constitutive activation of NF-κB reduces the tumor suppressor activity of p53, thereby creating permissive conditions for dominant oncogene-mediated transformation. Loss of p53 activity is also a characteristic of the majority of tumors and results in unleashed inflammatory responses due to loss of p53-mediated NF-κB suppression. On the other hand, in natural or pharmacological situations of enforced p53 activation, NF-κB activity, inflammation, and immune responses are reduced, resulting in different pathologies. It is likely that the chronic inflammation that is commonly acquired in various tissues of older mammals leads to general suppression of p53 function, which would explain the increased risk of cancer observed in aging animals and humans. Although the molecular mechanisms underlying reciprocal negative regulation of p53 and NF-κB remain to be deciphered, this phenomenon has important implications for pharmacological prevention of cancer and aging and for new approaches to control inflammation.