The toxicity of ionizing radiation is associated with massive apoptosis in radiosensitive organs. Here, we investigate whether a drug that activates a signaling mechanism used by tumor cells to suppress apoptosis can protect healthy cells from the harmful effects of radiation. We studied CBLB502, a polypeptide drug derived from Salmonella flagellin that binds to Toll-like receptor 5 (TLR5) and activates nuclear factor-kappaB signaling. A single injection of CBLB502 before lethal total-body irradiation protected mice from both gastrointestinal and hematopoietic acute radiation syndromes and resulted in improved survival. CBLB502 injected after irradiation also enhanced survival, but at lower radiation doses. It is noteworthy that the drug did not decrease tumor radiosensitivity in mouse models. CBLB502 also showed radioprotective activity in lethally irradiated rhesus monkeys. Thus, TLR5 agonists could potentially improve the therapeutic index of cancer radiotherapy and serve as biological protectants in radiation emergencies.

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.

Purpose Development of mucositis is a frequent side effect of radiotherapy of patients with head and neck cancer. We have recently reported that bacterial flagellin, an agonist of Toll-like receptor 5 (TLR5), can protect rodents and primates from acute radiation syndrome caused by total body irradiation (Burdelya et al., 2008, Science 320: 226-30). Here we analyzed the radioprotective efficacy of TLR5 agonist under conditions of local, single dose or fractionated radiation treatment. Methods and Materials Mice received either single-dose (10, 15, 20 or 25 Gy) or fractioned irradiation (cumulative dose up to 30 Gy) of the head and neck area with or without subcutaneous injection of pharmacologically optimized flagellin, CBLB502, 30 minutes prior to irradiation. Results CBLB502 significantly reduced the severity of dermatitis and mucositis, accelerated tissue recovery and reduced the extent of radiation induced weight loss in mice after a single dose of 15 or 20 Gy but not from 25 Gy of radiation. CBLB502 was also protective from cumulative doses of 25 and 30 Gy delivered in two (10+15 Gy) or three (3 × 10 Gy) fractions, respectively. While providing protection to normal epithelia, CBLB502 did not affect the radiosensitivity of syngeneic squamous carcinoma SCCVII grown orthotopically in mice. Use of CBLB502 also elicited a radiation independent growth inhibitory effect upon TLR5-expressing tumors demonstrated in the mouse xenograft model of human lung adenocarcinoma A549. Conclusion CBLB502 combines properties of supportive care (radiotherapy adjuvant) and anticancer agent, both mediated via activation of TLR5 signaling in the normal tissues or the tumor, respectively.

Factors that mediate p53 tumor suppressor activity remain largely unknown. In this study we describe a systematic approach to identify downstream mediators of tumor suppressor function of p53, consisting of global gene expression profiling, focused short hairpin RNA (shRNA) library creation, and functional selection of genetic elements cooperating with oncogenic Ras in cell transformation. This approach is based on our finding that repression of gene expression is a major event, occurring in response to p53 inactivation during transformation and immortalization of primary cells. Functional analysis of the subset of genes universally down-regulated in the cells that lacked functional p53 revealed BTG2 as a major downstream effector of p53-dependent proliferation arrest of mouse and human fibroblasts transduced with oncogenic Ras. shRNA-mediated knockdown of BTG2 cooperates with oncogenic Ras to transform primary mouse fibroblasts containing wild-type transcriptionally active p53. Repression of BTG2 results in up-regulation of cyclins D1 and E1 and phosphorylation of Rb and, in cooperation with other oncogenic elements, induces neoplastic transformation of primary human fibroblasts. BTG2 expression was found to be significantly reduced in a large proportion of human kidney and breast carcinomas, suggesting that BTG2 is a tumor suppressor that links p53 and Rb pathways in human tumorigenesis.

p53 tumor suppressor gene controls cell response to a variety of stresses inducing growth arrest or apoptosis in damaged cells. It largely determines the sensitivity of tumor and normal cells to radiation and chemotherapy, and, therefore, de®nes both the ecacy and limitations of anti-cancer treatment. To determine molecular mechanisms of p53-dependent stress response in normal tissues we identi®ed and compared the spectra of radiation-responsive genes in cells of dierent origin and p53 status using a cDNA array hybridization technique. The majority of genes identi®ed were p53-dependent and cell type speci®c. Several of the new p53 responders encode known secreted growth inhibitory factors. This suggests that p53, in addition to its intrinsic antiproliferation activity, can cause`bystander eect' by inducing export of growth suppressive stimuli from damaged cells to neighboring cells. Consistently, a p53-dependent accumulation of factors, which causes growth inhibitory eects in a variety of cell lines, was found after gamma irradiation in the media from established and primary cell cultures and in the urine of irradiated mice. Moreover, p53-dependent factors released by normal human ®broblasts potentiated the cytotoxic eect of a chemotherapeutic drug on co-cultivated tumor cells. This suggests a previously unknown role for normal cells in chemo-and radiation therapy of cancer.