Nuclear Factor κB Inhibitors Alleviate and the Proteasome Inhibitor PS-341 Exacerbates Radiation Toxicity in Zebrafish Embryos

Inflammatory changes are a major component of the normal tissue response to ionizing radiation, and increased nuclear factor κB (NF-κB) activity is an important mediator of inflammatory responses. Here, we used zebrafish embryos to assess the capacity of two different classes of pharmacologic agents known to target NF-κB to modify radiation toxicity in the vertebrate organism. These were proteasome inhibitors, including lactacystin, MG132, and PS-341 (Bortezomib/VELCADE), and direct inhibitors of NF-κB activity, including ethyl pyruvate (EP) and the synthetic triterpenoid CDDO-TFEA (RTA401), among others. The proteasome inhibitors either did not significantly affect radiation sensitivity of zebrafish embryos (MG132, lactacystin) or rendered zebrafish embryos more sensitive to the lethal effects of ionizing radiation (PS-341). Radiosensitization by PS-341 was reduced in fish with impaired p53 expression or function but not associated with enhanced expression of select p53 target genes. In contrast, the direct NF-κB inhibitors EP and CDDO-TFEA significantly improved overall survival of lethally irradiated zebrafish embryos. In addition, direct NF-κB inhibition reduced radiation-induced apoptosis in the central nervous system, abrogated aberrations in body axis development, restored metabolization and secretion of a reporter lipid through the gastrointestinal system, and improved renal clearance compromised by radiation. In contrast to amifostine, EP and CDDO-TFEA not only protected against but also mitigated radiation toxicity when given 1 to 2 hours postexposure. Finally, four additional IκB kinase inhibitors with distinct mechanisms of action similarly improved overall survival of lethally irradiated zebrafish embryos. In conclusion, inhibitors of canonical pathways to NF-κB activation may be useful in alleviating radiation toxicity in patients. [Mol Cancer Ther 2009;8(9):2625-34] Reprinted with permission from the American Association of Cancer Research, “Nuclear factor κB inhibitors alleviate and the proteasome inhibitor PS-341 exacerbates radiation toxicity in zebrafish embryos”, Molecular Cancer Therapy, 2009;8(9), pages 2625-2634

Radiation Therapy for Esophageal Cancer

Esophageal cancer develops in the mucosa of the esophagus and spreads toward the muscle layer. The nonsurgical treatment for localized, deeply invasive esophageal cancer has been external beam radiation therapy (EBRT) and concurrent chemotherapy. Recently, intraluminal brachytherapy showed a strong potential for the improvement of the therapeutic ratio. It was found that the fractionated high dose rate (HDR) brachytherapy offered beneficial palliation for a longer period of time with more durable symptom control. A similar was concluded for advanced unresectable esophageal cancer in previously irradiated patients. HDR brachytherapy may be a useful salvage treatment option for inoperable patients diagnosed with local esophageal cancer. Although better local control can be achieved with higher brachytherapy dose, this increases the risk of acute morbidity and late morbidity, especially in the setting of recurrence cancer. It was found that the moderate dose of EBRT and HDR brachytherapy could give a better local response than EBRT alone

Differential regulation of p53 function by the N-terminal ΔNp53 and Δ113p53 isoforms in zebrafish embryos

<p>Abstract</p> <p>Background</p> <p>The p53 protein family coordinates stress responses of cells and organisms. Alternative promoter usage and/or splicing of p53 mRNA gives rise to at least nine mammalian p53 proteins with distinct N- and C-termini which are differentially expressed in normal and malignant cells. The human N-terminal p53 variants contain either the full-length (FL), or a truncated (ΔN/Δ40) or no transactivation domain (Δ133) altogether. The functional consequences of coexpression of the different p53 isoforms are poorly defined. Here we investigated functional aspects of the zebrafish ΔNp53 ortholog in the context of FLp53 and the zebrafish Δ133p53 ortholog (Δ113p53) coexpressed in the developing embryo.</p> <p>Results</p> <p>We cloned the zebrafish ΔNp53 isoform and determined that ionizing radiation increased expression of steady-state <it>ΔNp53 </it>and <it>Δ113p53 </it>mRNA levels in zebrafish embryos. Ectopic ΔNp53 expression by mRNA injection caused hypoplasia and malformation of the head, eyes and somites, yet partially counteracted lethal effects caused by concomitant expression of FLp53. FLp53 expression was required for developmental aberrations caused by ΔNp53 and for ΔNp53-dependent expression of the cyclin-dependent kinase inhibitor 1A (CDKN1A, p21, Cip1, WAF1). Knockdown of p21 expression markedly reduced the severity of developmental malformations associated with ΔNp53 overexpression. By contrast, forced Δ113p53 expression had little effect on ΔNp53-dependent embryonal phenotypes. These functional attributes were shared between zebrafish and human ΔNp53 orthologs ectopically expressed in zebrafish embryos. All 3 zebrafish isoforms could be coimmunoprecipitated with each other after transfection into Saos2 cells.</p> <p>Conclusions</p> <p>Both alternative N-terminal p53 isoforms were expressed in developing zebrafish in response to cell stress and antagonized lethal effects of FLp53 to different degrees. However, in contrast to Δ113p53, forced ΔNp53 expression itself led to developmental defects which depended, in part, on p21 transactivation. In contrast to FLp53, the developmental abnormalities caused by ΔNp53 were not counteracted by concomitant expression of Δ113p53.</p