Activation of NFkappaB is inhibited by curcumin and related enones.

The transcription factor NFkappaB (NFkappaB) is up-regulated in many cancer cells where it contributes to development of the pro-survival, anti-apoptotic state. The natural product curcumin is a known inhibitor of activation of NFkappaB. Enone analogues of curcumin were compared with curcumin for their abilities to inhibit the TNFalpha-induced activation of NFkappaB, using the Panomics' NFkappaB Reporter Stable Cell Line. The enones tested included curcumin analogues that retained the 7-carbon spacer between the aromatic rings, analogues with a 5-carbon spacer, and analogues with a 3-carbon spacer. Inhibitors of NFkappaB activation were identified in all three series, a number of which were more active than curcumin. Enone analogues in the series with the 5-carbon spacer were especially active, including members that contained heterocyclic rings. 1,5-Bis(3-pyridyl)-1,4-pentadien-3-one was the most active analogue, IC50 = 3.4 +/- 0.2 microM. The most active analogues retain the enone functionality, although some analogues devoid of the enone functionality exhibited activity. The activity of the analogues as inhibitors of the activation of NFkappaB did not correlate with their anti-oxidant activity. The data suggest that the abilities of curcumin and analogues to prevent the stress-induced activation of NFkappaB result from the inhibition of specific targets rather than from activity as anti-oxidants.

Expression of angiogenic factors vascular endothelial growth factor and interleukin-8/CXCL8 is highly responsive to ambient glutamine availability: role of nuclear factor-kappaB and activating protein-1.

Vascular endothelial growth factor (VEGF) and interleukin-8/CXCL8 (IL-8) are prominent pro-angiogenic and pro-metastatic proteins that represent negative prognostic factors in many types of cancer. Hypoxia is thought to be the primary environmental cause of VEGF and IL-8 expression in solid tumors. We hypothesized that a lack of nutrients other than oxygen could stimulate the expression of these factors and previously demonstrated that expression of VEGF and IL-8 is responsive to amino acid deprivation. In the present study, we examined the effect of glutamine availability on the expression of these factors as well as the role of transcription factors NFkappaB and activating protein-1 (AP-1) in the response of TSE human breast carcinoma cells to glutamine deprivation. VEGF and IL-8 secretion and mRNA levels were dramatically induced by glutamine deprivation. mRNA stabilization contributed to this response. Glutamine deprivation increased NFkappaB (p65/p50) and AP-1 (Fra-1/c-Jun+JunD) DNA-binding activities. Blocking NFkappaB and AP-1 activation with curcumin as well as expression of dominant inhibitors, inhibitor of nuclear factor-kappaB (IkappaB) super repressor (IkappaBM), and a mutant form of c-Fos (A-Fos) demonstrated that the activation of NFkappaB and AP-1 transcription factors was necessary for the induction of IL-8 expression but dispensable for the induction of VEGF expression. A macro-array containing 111 NFkappaB target genes identified a total of 17 that were up-regulated 2-fold or more in response to glutamine deprivation. These included growth regulated oncogene alpha (GROalpha/GRO1/CXCL1), another neutrophil chemoattractant implicated in tumor angiogenesis and metastasis.

Expression of the pro-angiogenic factors vascular endothelial growth factor and interleukin-8/CXCL8 by human breast carcinomas is responsive to nutrient deprivation and endoplasmic reticulum stress.

BACKGROUND: The expression of pro-angiogenic cytokines, such as vascular endothelial growth factor (VEGF) and interleukin-8/CXCL8 (IL-8), plays an important role in tumor growth and metastasis. Low oxygen tension within poorly-vascularized tumors is thought to be the prime stimulus causing the secretion of VEGF. The expression of IL-8 by solid tumors is thought to be primarily due to intrinsic influences, such as constitutive activation of nuclear factor kappa B (NF-kappaB). However, VEGF expression is responsive to glucose deprivation, suggesting that low concentrations of nutrients other than oxygen may play a role in triggering the pro-angiogenic phenotype. Glucose deprivation causes endoplasmic reticulum (ER) stress and alters gene expression through the unfolded protein response (UPR) signaling pathway. A branch of the UPR, known as the ER overload response (EOR), can cause NF-kappaB activation. Thus, we hypothesized that treatments that cause ER stress and deprivation of other nutrients, such as amino acids, would trigger the expression of angiogenic cytokines by breast cancer cell lines. RESULTS: We found that glutamine deprivation and treatment with a chemical inducer of ER stress (tunicamycin) caused a marked induction of the secretion of both VEGF and IL-8 protein by a human breast adenocarcinoma cell line (TSE cells). Glutamine deprivation, glucose deprivation and several chemical inducers of ER stress increased VEGF and IL-8 mRNA expression in TSE and other breast cancer cell lines cultured under both normoxic and hypoxic conditions, though hypoxia generally diminished the effects of glucose deprivation. Of all amino acids tested, ambient glutamine availability had the largest effect on VEGF and IL-8 mRNA expression. The induction of VEGF mRNA expression, but not IL-8, was sustained and closely corresponded with the upregulated expression of the ER stress-responsive genes glucose-regulated protein 78 (GRP78) and growth arrest and DNA damage inducible gene 153 (GADD153). CONCLUSION: These results suggest that nutrient deprivation within the solid tumor microenvironment might contribute to the activation of a pro-angiogenic phenotype. The angiogenic switch may act to increase blood supply in response to nutrient deprivation as well as hypoxia.