ARC-111 inhibits hypoxia-mediated hypoxia-inducible factor-1alpha accumulation

ARC-111, a small-molecule topoisomerase I inhibitor, is a potent cytotoxic drug against multiple human cancer cell lines under normoxic conditions (Li et al., Cancer Res 2003; 63:8400-8407). In this study, we explore the potential of ARC-111 as a hypoxia-inducible factor-1alpha inhibitor under hypoxic conditions. The transcription factor, hypoxia-inducible factor-1alpha, is an essential regulator of tumorigenesis and an attractive molecular target for cancer therapy. We demonstrate that ARC-111 specifically inhibits hypoxia-induced accumulation of hypoxia-inducible factor-1alpha, but not other short half-life proteins in multiple human cancer cell lines. ARC-111 inhibits hypoxia-inducible factor-1alpha protein synthesis specifically and does not inhibit protein synthesis globally. We demonstrate that inhibition of hypoxia-inducible factor-1alpha accumulation by ARC-111 is independent of proteasomal degradation. In addition, we demonstrate using topoisomerase I-resistant cell lines that topoisomerase I is required for ARC-111-mediated hypoxia-inducible factor-1alpha inhibition. Experiments performed with nocodazole indicate that ARC-111 inhibits hypoxia-inducible factor-1alpha accumulation in a cell-cycle-independent manner. Analysis of AKT and mammalian target of rapamycin phosphorylation reveals that ARC-111 does not exhibit inhibitory effect on the phosphatidylinositol-3-kinase AKT mammalian target of rapamycin signaling pathway. It has been previously shown that topotecan, a topoisomerase I inhibitor, can also modulate hypoxia-induced hypoxia-inducible factor-1alpha accumulation (Rapisarda et al., Cancer Res 2003; 64:1475-1482). In addition to inhibiting hypoxia-induced accumulation of hypoxia-inducible factor-1alpha, ARC-111 exhibits antiproliferative effects against multiple human cancer cell lines. We demonstrate that topoisomerase I is required for the antiproliferative effects of ARC-111. Antiproliferative effects of ARC-111, however, are oxygen-independent, which is distinguishable from inhibition of hypoxia-inducible factor-1alpha accumulation by ARC-111, which is only observed under hypoxia. The results indicate that inhibiting hypoxia-inducible factor-1alpha accumulation and exhibiting antiproliferation of ARC-111 are through distinct mechanisms of action, which reinforce the potential anticancer effect of ARC-111 on hypoxic tumors.