Inhibition of hypoxia-inducible factor 1α accumulation by glyceryl trinitrate and cyclic guanosine monophosphate

• Published in: Bioscience reports 2020-01, Vol.40 (1)
• Main Authors: Kim, Judy, Barsoum, Ivraym B, Loh, Harrison, Paré, Jean-François, Siemens, D Robert, Graham, Charles H
• Format: Article
• Language: English
• Subjects: Calpain - metabolism; Cancer; Cell Line, Tumor; Cyclic GMP - analogs & derivatives; Cyclic GMP - metabolism; Cyclic GMP - pharmacology; Humans; Hypoxia-Inducible Factor 1, alpha Subunit - genetics; Hypoxia-Inducible Factor 1, alpha Subunit - metabolism; Male; Nitric Oxide - metabolism; Nitric Oxide Donors - pharmacology; Nitroglycerin - pharmacology; Prostatic Neoplasms - genetics; Prostatic Neoplasms - metabolism; Prostatic Neoplasms - pathology; Signal Transduction; Therapeutics & Molecular Medicine; Tumor Hypoxia; Tumor Microenvironment
• ISSN: 0144-8463; 1573-4935
• DOI: 10.1042/BSR20192345

A key mechanism mediating cellular adaptive responses to hypoxia involves the activity of hypoxia-inducible factor 1 (HIF-1), a transcription factor composed of HIF-1α, and HIF-1β subunits. The classical mechanism of regulation of HIF-1 activity involves destabilisation of HIF-1α via oxygen-dependent hydroxylation of proline residues and subsequent proteasomal degradation. Studies from our laboratory revealed that nitric oxide (NO)-mediated activation of cyclic guanosine monophosphate (cGMP) signalling inhibits the acquisition of hypoxia-induced malignant phenotypes in tumour cells. The present study aimed to elucidate a mechanism of HIF-1 regulation involving NO/cGMP signalling. Using human DU145 prostate cancer cells, we assessed the effect of the NO mimetic glyceryl trinitrate (GTN) and the cGMP analogue 8-Bromo-cGMP on hypoxic accumulation of HIF-1α. Concentrations of GTN known to primarily activate the NO/cGMP pathway (100 nM-1 µM) inhibited hypoxia-induced HIF-1α protein accumulation in a time-dependent manner. Incubation with 8-Bromo-cGMP (1 nM-10 µM) also attenuated HIF-1α accumulation, while levels of HIF-1α mRNA remained unaltered by exposure to GTN or 8-Bromo-cGMP. Furthermore, treatment of cells with the calpain (Ca2+-activated proteinase) inhibitor calpastatin attenuated the effects of GTN and 8-Bromo-cGMP on HIF-1α accumulation. However, since calpain activity was not affected by incubation of DU145 cells with various concentrations of GTN or 8-Bromo-cGMP (10 nM or 1 µM) under hypoxic or well-oxygenated conditions, it is unlikely that NO/cGMP signalling inhibits HIF-1α accumulation via regulation of calpain activity. These findings provide evidence for a role of NO/cGMP signalling in the regulation of HIF-1α, and hence HIF-1-mediated hypoxic responses, via a mechanism dependent on calpain.