Hypoxia Promotes Uveal Melanoma Invasion Through Enhanced Notch and MAPK Activation

PLoS One. 2014 Aug 28;9(8):e105372. doi: 10.1371/journal.pone.0105372. eCollection 2014.

Abstract

The transcriptional response promoted by hypoxia-inducible factors has been associated with metastatic spread of uveal melanoma. We found expression of hypoxia-inducible factor 1α (HIF-1α) protein in well-vascularized tumor regions as well as in four cell lines grown in normoxia, thus this pathway may be important even in well-oxygenated uveal melanoma cells. HIF-1α protein accumulation in normoxia was inhibited by rapamycin. As expected, hypoxia (1% pO2) further induced HIF-1α protein levels along with its target genes VEGF and LOX. Growth in hypoxia significantly increased cellular invasion of all 5 uveal melanoma lines tested, as did the introduction of an oxygen-insensitive HIF-1α mutant into Mel285 cells with low HIF-1α baseline levels. In contrast, HIF-1α knockdown using shRNA significantly decreased growth in hypoxia, and reduced by more than 50% tumor invasion in four lines with high HIF-1α baseline levels. Pharmacologic blockade of HIF-1α protein expression using digoxin dramatically suppressed cellular invasion both in normoxia and in hypoxia. We found that Notch pathway components, including Jag1-2 ligands, Hes1-Hey1 targets and the intracellular domain of Notch1, were increased in hypoxia, as well as the phosphorylation levels of Erk1-2 and Akt. Pharmacologic and genetic inhibition of Notch largely blocked the hypoxic induction of invasion as did the pharmacologic suppression of Erk1-2 activity. In addition, the increase in Erk1-2 and Akt phosphorylation by hypoxia was partially reduced by inhibiting Notch signaling. Our findings support the functional importance of HIF-1α signaling in promoting the invasive capacity of uveal melanoma cells in both hypoxia and normoxia, and suggest that pharmacologically targeting HIF-1α pathway directly or through blockade of Notch or Erk1-2 pathways can slow tumor spread.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Line, Tumor
  • Digoxin / pharmacology
  • Enzyme Inhibitors / pharmacology
  • Humans
  • Hypoxia / genetics
  • Hypoxia / metabolism
  • Hypoxia / pathology*
  • Hypoxia-Inducible Factor 1, alpha Subunit / genetics
  • Hypoxia-Inducible Factor 1, alpha Subunit / metabolism
  • MAP Kinase Signaling System / drug effects
  • MAP Kinase Signaling System / physiology*
  • Melanoma / genetics
  • Melanoma / metabolism
  • Melanoma / pathology*
  • Neoplasm Invasiveness / pathology*
  • Proto-Oncogene Proteins c-akt / metabolism
  • Receptors, Notch / genetics
  • Receptors, Notch / metabolism*
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Uveal Neoplasms / genetics
  • Uveal Neoplasms / metabolism
  • Uveal Neoplasms / pathology*

Substances

  • Enzyme Inhibitors
  • HIF1A protein, human
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Receptors, Notch
  • Digoxin
  • Proto-Oncogene Proteins c-akt