Negative regulation of the hypoxia-inducible transcription factors by IPAS defines a novel anti-angiogenesis therapeutic strategy

Angiogenesis is an essential component of proliferative synovitis leading to joint destruction in rheumatoid arthritis (RA). Recent studies have demonstrated the presence of hypoxia and the accumulation of pro-angiogenic growth factors, including vascular endothelial growth factor (VEGF), in the synovium from patients with RA, indicating a possible involvement of those factors in synovial angiogenesis. The hypoxia-inducible transcription factor-1α (HIF-1α) is a key regulator of VEGF gene expression and angiogenesis under hypoxic conditions [1]. We have recently demonstrated abnormal expression of HIF-1α and VEGF in the synovium from patients with RA, indicating a potential contribution of the HIF-1α –VEGF system to angiogenic processes of synovitis in RA. The precise mechanism of HIF-1α-mediated control of VEGF expression and angiogenesis, however, is largely unknown.

Given this background, we aimed to elucidate the molecular mechanism underlying hypoxia-inducible VEGF expression via HIF-1α, and identified a novel basic helix–loop–helix/Per, Arnt, Sim protein (inhibitory Per, Arnt, Sim domain protein [IPAS]) that has high structural similarity to HIF-1α [2]. In sharp contrast to HIF-1α, IPAS was not capable of activating hypoxia-inducible gene expression. Co-expression of IPAS and HIF-1α resulted in repression of hypoxia-inducible VEGF gene expression, demonstrating a dominant-negative regulatory activity of IPAS on HIF-1α-mediated control of gene expression. In situ hybridization analysis of various mouse tissues showed predominant IPAS expression in the avascular corneal epithelium, correlating with low levels of VEGF gene expression under hypoxic conditions. Strikingly, application of an IPAS antisense oligonucleotide to the mouse cornea induced angiogenesis under normoxic conditions, and unmasked hypoxia-dependent induction of VEGF gene expression in hypoxic cornea cells. Moreover, ectopic expression of IPAS in hepatoma cells impaired induction of genes involved in adaptation to a hypoxic environment, and resulted in retarded tumor growth and tumor vascular density in vivo. Taken together, IPAS, a dominant-negative regulator of HIF-1α function, would define a novel anti-angiogenic mechanism under hypoxic conditions, providing a possible therapeutic strategy for angiogenic diseases including RA.