17-Aminogeldanamycin Inhibits Constitutive Nuclear Factor-Kappa B (NF-κB) Activity in Patient-Derived Melanoma Cell Lines

Abstract

Melanoma remains incurable skin cancer, and targeting heat shock protein 90 (HSP90) is a promising therapeutic approach. In this study, we investigate the effect of 17-aminogeldanamycin, a potent HSP90 inhibitor, on nuclear factor-kappa B (NF-κB) activity in BRAFV600E and NRASQ61R patient-derived melanoma cell lines. We performed time-lapse microscopy and flow cytometry to monitor changes in cell confluence and viability. The NF-κB activity was determined by immunodetection of phospho-p65 and assessment of expression of NF-κB-dependent genes by quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, and enzyme-linked immunosorbent assay (ELISA). Constitutive activity of p65/NF-κB was evident in all melanoma cell lines. Differences in its level might be associated with genetic alterations in CHUK, IL1B, MAP3K14, NFKBIE, RIPK1, and TLR4, while differences in transcript levels of NF-κB-inducible genes revealed by PCR array might result from the contribution of other regulatory mechanisms. 17-Aminogeldanamycin markedly diminished the level of phospho-p65, but the total p65 protein level was unaltered, indicating that 17-aminogeldanamycin inhibited activation of p65/NF-κB. This conclusion was supported by significantly reduced expression of selected NF-κB-dependent genes: cyclin D1 (CCND1), C-X-C motif chemokine ligand 8 (CXCL8), and vascular endothelial growth factor (VEGF), as shown at transcript and protein levels, as well as secretion of IL-8 and VEGF. Our study indicates that 17-aminogeldanamycin can be used for efficient inhibition of NF-κB activity and the simultaneous diminution of IL-8 and VEGF levels in the extracellular milieu of melanoma.

Inhibition of Hsp90 Counteracts the Established Experimental Dermal Fibrosis Induced by Bleomycin

“…Interestingly, in the study by Sibinska et al [55], only the lower dose of 17-DMAG (i.e., 10 mg/kg vs. 25 mg/kg) significantly decreased both the bronchoalveolar lavage fluid and serum levels of TGF-β in the bleomycin-induced pulmonary fibrosis. Similarly, inhibition of Hsp90 reduced local or systemic levels of IL-8 in experimental models of various inflammatory and tumorous conditions [72][73][74][75][76]. However, further studies are required to elucidate the mechanisms underlying the suppression of local and systemic inflammatory response mediated by 17-DMAG, as well as the potential link between the humoral or cellular immune response and the progression of dermal fibrosis induced by bleomycin.…”

DNA damage and NF‐κB inactivation implicate glycyrrhizic acid‐induced G 1 phase arrest in hepatocellular carcinoma cells

“…Cyclin D1, encoded by the CCND1 gene, is a cell cycle regulator that mediates cell cycle progression through promoting G 1 /S phase transition (González‐Ruiz et al, 2021). NF‐κB, the p50/p65 heterodimer of transcription factor, may induce CCND1 expression and lead to the upregulation of Cyclin D1 (Hartman et al, 2020; Velho et al, 2012). Suppression of NF‐κB activation has been indicated to trigger G 1 phase arrest through Cyclin D1 downregulation (Kuo et al, 2012).…”

The Chaperone System in Salivary Glands: Hsp90 Prospects for Differential Diagnosis and Treatment of Malignant Tumors

“…Similar results, in which phosphorylated NF-κB is also downregulated as a consequence of Hsp90 inhibition, were reported for prostate carcinoma [ 100 ]. Hsp90 inhibition decreased constitutive and induced NF-κB activity in human myeloid leukemia [ 49 , 101 , 102 , 103 ], chronic lymphocytic leukemia [ 104 , 105 ], primary effusion lymphoma [ 106 ], and melanoma [ 107 ], which led to cytotoxicity; cell-cycle arrest; reduction in proliferation, migration, and invasion; and induced apoptosis. Moreover, blocking NF-κB and Akt survival pathways using the Hsp90 inhibitor potentiates chemotherapeutic drug cytotoxicity in lung cancer [ 108 ].…”