Posttranscriptional upregulation of HER3 by HER2 mRNA induces trastuzumab resistance in breast cancer

Xin Li, Yuxiu Xu, Yun Ding, Changfei Li, Hong Zhao, Jiandong Wang, Songdong Meng

HER2 gene amplification generates an enormous number of HER2 transcripts, but the global effects on endogenous miRNA targets including HER family members in breast cancer are unexplored. We generated a HER2-3'UTR expressing vector to test the tumor-promoting properties in HER2 low expressing T47D and MCF7 cells. Through microarray analysis and real-time PCR analysis we identified genes that were regulated by HER2-3'UTR. Positive and negative manipulation of miRNA expression, response element mutational studies and transcript reporter assays were performed to explore the mechanism of competitive sequestration of miR125a/miRNA125b by HER2 3'UTR. To investigate if trastuzumab-induced upregulation of HER3 is also mediated through miRNA de-repression, we used the CRISPR/cas9 to mutate the endogenous HER2 mRNA in HER2 over-expressing Au565 cells. Finally, we looked at cohorts of breast cancer samples of our own and the TCGA to show if HER2 and HER3 mRNAs correlate with each other. The HER2 3'UTR pronouncedly promoted cell proliferation, colony formation, and breast tumor growth. High-throughput sequencing revealed a significant increase in HER3 mRNA and protein levels by the HER2 3'untranslated region (3'UTR). The HER2 3'UTR harboring a shared miR-125a/b response element induced miR-125a/b sequestration and thus resulted in HER3 mRNA derepression. Trastuzumab treatment upregulated HER3 via elevated HER2 mRNA expression, leading to trastuzumab resistance. Depletion of miR-125a/b enhanced the antitumor activity of trastuzumab. Microarray data from HER2-overexpressing primary breast cancer showed significant elevation of mRNAs for predicted miR-125a/b targets compared to non-targets. These results suggest that HER2 3'UTR-mediated HER3 upregulation is involved in breast cell transformation, increased tumor growth, and resistance to anti-HER2 therapy. The combinatorial targeting of HER3 mRNA or miR-125a/b may offer an effective tool for breast cancer therapy.