Inhibition of nuclear factor kappa-B signaling reduces growth in medulloblastoma in vivo

Abstract Background Medulloblastoma is a highly malignant pediatric brain tumor that requires surgery, whole brain and spine irradiation, and intense chemotherapy for treatment. A more sophisticated understanding of the pathophysiology of medulloblastoma is needed to successfully reduce the intensity of treatment and improve outcomes. Nuclear factor kappa-B (NFκB) is a signaling pathway that controls transcriptional activation of genes important for tight regulation of many cellular processes and is aberrantly expressed in many types of cancer. Methods To test the importance of NFκB to medulloblastoma cell growth, the effects of multiple drugs that inhibit NFκB, pyrrolidine dithiocarbamate, diethyldithiocarbamate, sulfasalazine, curcumin and bortezomib, were studied in medulloblastoma cell lines compared to a malignant glioma cell line and normal neurons. Expression of endogenous NFκB was investigated in cultured cells, xenograft flank tumors, and primary human tumor samples. A dominant negative construct for the endogenous inhibitor of NFκB, IκB, was prepared from medulloblastoma cell lines and flank tumors were established to allow specific pathway inhibition. Results We report high constitutive activity of the canonical NFκB pathway, as seen by Western analysis of the NFκB subunit p65, in medulloblastoma tumors compared to normal brain. The p65 subunit of NFκB is extremely highly expressed in xenograft tumors from human medulloblastoma cell lines; though, conversely, the same cells in culture have minimal expression without specific stimulation. We demonstrate that pharmacological inhibition of NFκB in cell lines halts proliferation and leads to apoptosis. We show by immunohistochemical stain that phosphorylated p65 is found in the majority of primary tumor cells examined. Finally, expression of a dominant negative form of the endogenous inhibitor of NFκB, dnIκB, resulted in poor xenograft tumor growth, with average tumor volumes 40% smaller than controls. Conclusions These data collectively demonstrate that NFκB signaling is important for medulloblastoma tumor growth, and that inhibition can reduce tumor size and viability in vivo. We discuss the implications of NFκB signaling on the approach to managing patients with medulloblastoma in order to improve clinical outcomes.</p

NF-κB-independent sensitization of glioblastoma cells for TRAIL-induced apoptosis by proteasome inhibition

The transcription factor nuclear factor-kappaB (NF-κB) is a key regulator of stress-induced transcriptional activation and has been implicated in mediating primary or acquired apoptosis resistance in various cancers. In the present study, we therefore investigated the role of NF-κB in regulating apoptosis in malignant glioma, a prototypic tumor refractory to current treatment approaches. Here, we report that constitutive NF-κB DNA-binding activity was low or moderate in eight different glioblastoma cell lines compared to Hodgkin's lymphoma cells, known to harbor aberrant constitutive NF-κB activity. Specific inhibition of NF-κB by overexpression of inhibitor of κB (IκB)α superrepressor did not enhance spontaneous apoptosis of glioblastoma cells. Also, overexpression of IκBα superrepressor had no significant impact on apoptosis induced by two prototypic classes of apoptotic stimuli, that is, chemotherapeutic drugs or death-inducing ligands such as TNF-related apoptosis inducing ligand (TRAIL), which are known to trigger NF-κB activation as part of a cellular stress response. Similarly, inhibition of NF-κB by the proteasome inhibitor MG132 did not increase doxorubicin (Doxo)-induced apoptosis of glioblastoma cells, although it prevented DNA binding of NF-κB complexes in response to Doxo. Interestingly, proteasome inhibition significantly sensitized glioblastoma cells for TRAIL-induced apoptosis. These findings indicate that the characteristic antiapoptotic function of NF-κB reported for many cancers is not a primary feature of glioblastoma and thus, specific NF-κB inhibition may not be effective for chemosensitization of glioblastoma. Instead, proteasome inhibitors, which enhanced TRAIL-induced apoptosis in an NF-κB-independent manner, may open new perspectives to increase the efficacy of TRAIL-based regimens in glioblastoma, which warrants further investigation. © 2007 Nature Publishing Group All rights reserved

Mechanisms of proteasome inhibitor-induced cytotoxicity in malignant glioma.

The 26S proteasome constitutes an essential degradation apparatus involved in the consistent recycling of misfolded and damaged proteins inside cells. The aberrant activation of the proteasome has been widely observed in various types of cancers and implicated in the development and progression of carcinogenesis. In the era of targeted therapies, the clinical use of proteasome inhibitors necessitates a better understanding of the molecular mechanisms of cell death responsible for their cytotoxic action, which are reviewed here in the context of sensitization of malignant gliomas, a tumor type particularly refractory to conventional treatments