Hypoxia is an inadequate oxygen supply to tissues and cells, which can restrict their function. The hypoxic response is primarily mediated by the hypoxia-inducible transcription factors, HIF-1 and HIF-2, which have both overlapping and unique target genes. HIF target gene activation is highly context specific and is not a reliable indicator of which HIF-α isoform is active. For example in some cell lines, the individual HIFs have specific temporal and functional roles: HIF-1 drives the initial response to hypoxia (<24 hours) and HIF-2 drives the chronic response (>24 hours). Here, we review the significance of the HIF switch and the relationship between HIF-1 and HIF-2 under both physiological and pathophysiological conditions.

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Most solid tumors and their metastases experience periods of low oxygen or hypoxia, which is of major clinical significance as it promotes both tumor progression and resistance to therapy. Critical mediators of the hypoxic response are the hypoxia-inducible factors HIF-1α and HIF-2α. The HIFs are nonredundant and regulate both overlapping and unique downstream target genes. Here, we describe a novel mechanism for the switch between HIF-1α– and HIF-2α–dependent transcription during tumor hypoxia caused by the hypoxia associated factor (HAF). HAF is overexpressed in a variety of tumors and its levels are decreased during acute hypoxia, but increased following prolonged hypoxia. We have previously identified HAF as an E3 ubiquitin ligase that binds and ubiquitinates HIF-1α by an oxygen and pVHL-independent mechanism, thus targeting HIF-1α for proteasomal degradation. Here, we show that HAF also binds to HIF-2α, but at a different site than HIF-1α, and increases HIF-2α transactivation without causing its degradation. HAF, thus, switches the hypoxic response of the cancer cell from HIF-1α–dependent to HIF-2α–dependent transcription and activates genes involved in invasion such as MMP9, PAI-1, and the stem cell factor OCT-3/4. The switch to HIF-2α–dependent gene expression caused by HAF also promotes an enriched tumor stem cell population, resulting in highly aggressive tumors in vivo. Thus, HAF, by causing a switch from a HIF-1α– to HIF-2α–dependent response to hypoxia, provides a mechanism for more aggressive growth of tumors under prolonged hypoxia.

We have reported previously that PX-478 (S-2-amino-3-[4 ¶-N,N,-bis(chloroethyl)amino]phenyl propionic acid N-oxide dihydrochloride) has potent antitumor activity against a variety of human tumor xenografts associated with the levels of the hypoxia-inducible factor-1A (HIF-1A) within the tumor. We now report that PX-478 inhibits HIF-1A protein levels and transactivation in a variety of cancer cell lines. Hypoxia-induced vascular endothelial growth factor formation was inhibited by PX-478, whereas baseline levels of vascular endothelial growth factor in normoxia were unaffected. Studies of the mechanism of PX-478 action showed that HIF-1A inhibition occurs in both normoxia and hypoxia and does not require pVHL or p53. In addition, PX-478 decreases levels of HIF-1A mRNA and inhibits translation as determined by 35 S labeling experiments and reporter assays using the 5 ¶ untranslated region of HIF-1A. Moreover, to a lesser extent, PX-478 also inhibits HIF-1A deubiquitination resulting in increased levels of polyubiquitinated HIF-1A. The inhibitory effect of PX-478 on HIF-1A levels is primarily due to its inhibition of translation because HIF-1A translation continues in hypoxia when translation of most proteins is decreased. We conclude that PX-478 inhibits HIF-1A at multiple levels that together or individually may contribute to its antitumor activity against HIF-1A-expressing tumors. [Mol Cancer Ther 2008;7(1):90 -100]