Effects of transferrin receptor blockade on cancer cell proliferation and hypoxia-inducible factor function and their differential regulation by ascorbate.
Cellular iron is needed for cell survival and hydroxylation of hypoxia-inducible factor-1alpha (HIF-alpha) by prolyl hydroxylases (PHD). One mechanism of iron uptake is mediated by the cell surface transferrin receptor (TfR). Because iron is required for cell growth and suppression of HIF-alpha levels, we tested the effects of the two anti-TfR monoclonal antibodies (mAb) E2.3 and A27.15 on growth of breast cancer cells and induction of HIF-alpha and hypoxia-regulated genes. Treatment with both mAbs together synergistically inhibited cell proliferation in a dose-responsive manner by up to 80% following 8 days of exposure, up-regulated HIF-1alpha and HIF transcription targets, down-regulated TfR expression, and down-regulated cellular labile iron pool by 60%. Because combined treatment with anti-TfR mAbs resulted in the up-regulation of the hypoxia pathway, which may increase tumor angiogenesis, we analyzed the effects of ascorbate on cell viability and HIF-1alpha levels in cells treated with both anti-TfR mAbs together, as ascorbate has been shown to be required by PHD enzymes for full catalytic activity. Ascorbate at physiologic concentrations (25 micromol/L) suppressed HIF-1alpha protein levels and HIF transcriptional targets in anti-TfR mAb-treated cells but did not suppress the antiproliferative effect of the mAbs. These results indicate that the addition of ascorbate increased the activity of the PHD enzymes in down-regulating HIF but not the proliferation of iron-starved anti-TfR mAb-treated cells. The use of anti-TfR mAbs and ascorbate in inhibiting both cell proliferation and HIF-1alpha and angiogenesis under normoxic conditions may be of therapeutic use.
Jones
Dylan T
DT
Cancer Research UK Growth Factor Group, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford, UK.
Trowbridge
Ian S
IS
Harris
Adrian L
AL
D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D001205:Ascorbic Acid / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology* / Q000628:therapy; D048708:Cell Growth Processes / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004357:Drug Synergism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis* / Q000235:genetics; D007501:Iron / Q000378:metabolism; D011990:Receptors, Transferrin / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects
D000911:Antibodies, Monoclonal; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D011990:Receptors, Transferrin; D007501:Iron; D001205:Ascorbic Acid