Received for publication March 26, 2007.
Revised June 22, 2007.
Accepted for publication June 26, 2007.

The phosphoinositide-dependent kinase-1 inhibitor, OSU-03012, prevents Y-box binding protein-1 (YB-1) from inducing epidermal growth factor receptor (EGFR)

Abstract

The epidermal growth factor receptor (EGFR) is integral to basal-like and Her-2 over-expressing breast cancers. Such tumors are associated with poor prognosis, the majority of which express high levels EGFR. We reported that EGFR expression is induced by the oncogenic transcription factor Y-box binding protein-1 (YB-1) that occurs in a manner dependent upon phosphorylation by Akt. Herein, we questioned whether blocking Akt with OSU-03012, a phosphoinositide-dependent protein kinase-1 (PDK-1) small molecule inhibitor, could prevent YB-1 from binding to the EGFR promoter. MDA-MB-468 and SUM 149 are basal-like breast cancer (BLBC) cells that were used for our studies because they express high levels of activated PDK-1, YB-1 and EGFR compared to the immortalized breast epithelial cell line 184htrt. In these cell lines, YB-1 preferentially bound to the -1kB of the EGFR promoter whereas this did not occur in the 184htrt cells based on chromatin immunoprecipitation. When the cells were exposed to OSU-03012 for 6 hours, YB-1/EGFR promoter binding was significantly attenuated. To further confirm this observation, gel shift assays showed that the drug inhibits YB-1/EGFR promoter binding. The inhibitory effect of OSU-03012 on EGFR was also observed at the mRNA and protein levels. OSU-03012 ultimately inhibited the growth of BLBC in monolayer and soft agar coordinate with the induction of apoptosis. Furthermore, OSU-03012 inhibited the expression of EGFR by 48% in tumor xenografts derived from MDA-MB-435/Her-2 cells. This correlated with loss of YB-1 binding to the EGFR promoter. Hence, we find that OSU-03012 inhibits YB-1 resulting in a loss of EGFR expression in vitro and in vivo.

Key words: NGF/EGF, DNA binding sites, Promoter analysis, Regulation - transcriptional, Oncogenes, Transcription targets

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