Novel thioredoxin inhibitors paradoxically increase hypoxia-inducible factor-alpha expression but decrease functional transcriptional activity, DNA binding, and degradation.
OBJECTIVE
Hypoxia-inducible factor-alpha (HIF-alpha) is a transcription factor that regulates the response to hypoxia. HIF-alpha protein is found at high levels in many cancers, and the redox protein thioredoxin-1 (Trx-1) increases both aerobic and hypoxia-induced HIF-alpha. Therefore, Trx-1 and HIF-alpha are attractive molecular targets for novel cancer therapeutics.


METHODS
We investigated whether two novel anticancer drugs AJM290 and AW464 (quinols), which inhibit Trx-1 function, can inhibit the HIF pathway.


RESULTS
Treatment of several cancer cell lines with AJM290 or AW464 prevented the hypoxia-induced increase of vascular endothelial growth factor (VEGF) at subtoxic concentrations. AJM290 and AW464 also decreased VEGF in pVHL mutant renal cell carcinoma cells that constitutively overexpress HIF-alpha protein. They surprisingly up-regulated HIF-alpha expression in breast cancer cell lines in normoxia and hypoxia as well as in pVHL mutant cells. In the MDA-MB-468 breast cancer cell line, the compounds inhibited RNA and protein expression of the HIF-alpha target genes, carbonic anhydrase IX, VEGF, and BNIP3, concordantly with HIF-alpha up-regulation. Both compounds specifically inhibited HIF-alpha-dependent induction of hypoxia regulatory element-luciferase and HIF-1alpha hypoxia regulatory element-DNA binding. To analyze the HIF-1alpha domain inhibited by AJM290, we transfected cells with plasmids expressing a fusion protein of Gal linked to HIF-1alpha or HIF-1alpha COOH-terminal transactivation domain (CAD) with a Gal4-responsive luciferase reporter gene. AJM290 inhibited both the full-length HIF-1alpha and HIF-1alpha CAD transcriptional activity.


CONCLUSIONS
AJM290 and AW464 are inhibitors of HIF-1alpha CAD transcription activity and DNA binding, but they also inhibit degradation of HIF, in contrast to other Trx inhibitors.
Jones
Dylan T
DT
Cancer Research UK Growth Factor Group, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, University of Oxford, UK.
Pugh
Christopher W
CW
Wigfield
Simon
S
Stevens
Malcolm F G
MF
Harris
Adrian L
AL
D000970:Antineoplastic Agents / Q000494:pharmacology; D051792:Basic Helix-Loop-Helix Proteins; D052160:Benzothiazoles / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects; D003512:Cyclohexanones / Q000494:pharmacology*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000378:metabolism*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D007211:Indoles / Q000494:pharmacology*; D007976:Leupeptins / Q000494:pharmacology; D008954:Models, Biological; D050505:Mutant Proteins / Q000378:metabolism; D011489:Protein Denaturation / Q000187:drug effects*; D011994:Recombinant Proteins / Q000378:metabolism; D020218:Response Elements / Q000502:physiology; D013450:Sulfones / Q000494:pharmacology*; D013879:Thioredoxins / Q000037:antagonists & inhibitors*; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D000099162:Endothelial PAS Domain-Containing Protein 1
D000970:Antineoplastic Agents; D051792:Basic Helix-Loop-Helix Proteins; D052160:Benzothiazoles; D003512:Cyclohexanones; D004268:DNA-Binding Proteins; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007211:Indoles; D007976:Leupeptins; D050505:Mutant Proteins; D011994:Recombinant Proteins; D013450:Sulfones; D013879:Thioredoxins; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D000099162:Endothelial PAS Domain-Containing Protein 1; C515225:AJM 290; C515226:AW 464; C497442:HIF1A protein, human; C467484:VEGFA protein, human; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde