AG490 Promotes HIF-1α Accumulation by Inhibiting Its Hydroxylation

Abstract

AG490 is a tyrphostin originally described as a Janus Activated Kinase (JAK) 2 inhibitor. AG490 also inhibits epidermal growth factor receptor (EGFR) and guanylyl cyclases (GC). More recently, AG490 was associated with oxidative stress protection in experimental acute kidney injury models. We now show that AG490 is also a strong activator of the Hypoxia Inducible Factor (HIF)-1. Under normoxic conditions HIF-1α is degraded through hydroxylation, von Hippel Lindau protein (VHL)-mediated ubiquitin tagging and proteasomal degradation. AG490 increased HIF-1α protein, but not HIF-1α mRNA levels, dose- and time-dependently in cultured endothelial, vascular smooth muscle and kidney proximal tubular epithelial cells. AG490 increased HIF-1α protein half-life, suggesting that HIF-1α protein accumulation resulted from a decreased degradation. In this regard, AG490 prevented HIF-1α hydroxylation and increased HIF-1α protein levels in human renal carcinoma cells expressing VHL, but did not further increase HIF-1α in VHL negative cells. AG490 did not prevent the proteasomal degradation of other proteins. HIF-1α was not upregulated by dominant negative JAK2constructs, tyrphostin AG9, the EGFR inhibitors erbstatin and genistein, the GC inhibitor Ly83583 or cGMP analogues. Finally, AG490 also increased HIF-1α transcriptional activity evidenced by the increased HIF-1α-dependent VEGF expression. In conclusion, AG490 is a novel HIF-1α activator that increases HIF-1α half-life and protein levels through interference with HIF-1α hydroxylation and VHL-mediated degradation. This action may contribute to the cell and tissue protective effects of AG490.

The metabolic bone disease associated with the Hyp mutation is independent of osteoblastic HIF1α expression

“…To test the role of HIF activation on the control of FGF23, the compound AG409 was used, as this analog strongly activates HIF1α in isolated cell systems (Fernandez-Sanchez et al, 2012). In this regard, primary cultures of differentiated osteoblasts/osteocytes and the osteoblastic UMR-106 cell line were treated with AG490, and Fgf23 mRNA expression was examined by qPCR.…”

LPS‐induced inflammation desensitizes hepatocytes to Fas‐induced apoptosis through Stat3 activation—The effect can be reversed by ruxolitinib

“…In contrast, we found that ruxolitinib pre‐treatment with subsequent LPS administration further up‐regulated IL‐6 and IL‐1β expressions, excluding the possibility that apoptosis sensitization effect might occur due to anti‐inflammatory properties of ruxolitinib. These discrepancies might emerge due to differences between Kupffer cells and macrophage cultures or peritoneal macrophages or due to lower specificity of AG490 inhibitor that can also inhibit epidermal growth factor receptor signalling, essential for LPS‐induced signalling, and IL‐6 and TNF‐α production …”

“…These discrepancies might emerge due to differences between Kupffer cells and macrophage cultures or peritoneal macrophages or due to lower specificity of AG490 inhibitor that can also inhibit epidermal growth factor receptor signalling, essential for LPS-induced signalling, and IL-6 and TNF-α production. [35][36][37][38] So far, ruxolitinib has been typically used in chronic models with multiple applications, and we have explored its effect in an acute experimental model. We have confirmed its effect by immunoblotting of pStat3.…”

Cross-talk between HIF and p53 as mediators of molecular responses to physiological and genotoxic stresses

“…An inhibitor of Janus Activated Kinase (JAK2), AG490, prevents HIF-1α hydroxylation and thus interferes with VHL-mediated degradation resulting in increased HIF-1α protein half-life [7]. Another mechanism by which HIF-1α can be rescued from degradation is via interaction with ubiquitin-specific protease 19 (USP19) [8].…”