Abstract A202: Lipocalin2 stabilizes hypoxia inducible factor-1α through the iron delivery into normoxic cancer cells

Abstract

Abstract The level of lipocalin 2 (LCN2), a secreted protein of the lipocalin superfamily, increases in various tumors. Moreover, LCN2 is known to promote tumor progression with the hypoxia inducible factor-1α (HIF-1α)-mediated epithelial mesenchymal transition and angiogenesis. However, the mechanism of LCN2-induced HIF-1α activation is not fully understood. In the present study, we examined the effect of LCN2 on HIF-1α activation in the normoxic cancer cells that HIF-1α is immediately degraded. A mouse melanoma cell line, B16-F1, was treated with recomninant LCN2 protein under normoxic condition. After the treatment, the protein level and nuclear translocation of HIF-1α were examined by western blot and immunohistochemical staining, respectively. Moreover, the activation of HIF-1α was evaluated by reporter gene assay in the cells transfected with the plasmid DNA containing HIF-1α response element. The intracellular iron was quantified by iron colorimetric assay kit. In the B16-F1 cells treated with LCN2, HIF-1α protein was not degraded and its nuclear translocation was observed even under normoxic condition, suggesting that LCN2 stabilizes HIF-1α in normoxic cancer cells. Moreover, the HIF-1α-mediated transcriptional activity in the B16-F1 cells treated with LCN2 was significantly higher than those with non-treatment, suggesting that LCN2 activates HIF-1α in normoxic cancer cells. Next, to clarify the mechanism of LCN2-mediated HIF-1α activation, we hypothesized that LCN2, which is an Fe3+ transport protein, stabilizes HIF-1α by preventing the activity of proline hydroxylase (PHD) as a primary enzyme of HIF-1α degradation, because the increase of intracellular Fe3+ is known to inhibit PHD activity. As a result, the intracellular iron increased in the cells treated with LCN2 protein. Moreover, the level of HIF-1α by co-treatment of LCN2 with FeCl3 was higher than that only by LCN2 treatment. Collectively, LCN2-mediated Fe3+ influx stabilizes HIF-1α in normoxic cancer cells. Citation Format: Susumu Hama, Ibuki Nakamura, Akinori Nishimoto, Takayuki Nishi, Shoko Itakura, Kentaro Kogure. Lipocalin2 stabilizes hypoxia inducible factor-1α through the iron delivery into normoxic cancer cells. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A202.