Abstract A008: HER2Δ16, the oncogenic isoform of HER2, suppresses microRNA-7 to promote breast tumor cell migration and proliferation
Authors: Huynh Felicia C. , Jones Frank E. 
Published: May 2015

Abstract
Of the five different molecular subtypes of breast cancer, HER2-positive patients have the worst overall prognosis and shortest relapse-free survival. We have shown that HER2-positive breast tumors co-express the oncogenic HER2 isoform, HER2Δ16, which is associated with metastasis and trastuzumab resistance. Here we hypothesize that HER2Δ16 represents the transforming form of HER2 and drives tumorigenesis through deregulation of microRNA (miR) expression. We compared global miR expression profiles between parental MCF-7 breast cancer cells (MCF-7/pcDNA) and a MCF-7 cell line with ectopic expression of HER2Δ16 (MCF-7/HER2Δ16) and found altered regulation of multiple miRs. Interestingly, we observed a 4.8 fold suppression of the tumor suppressor microRNA-7 (miR-7) in the MCF-7/HER2Δ16 cells. MiR-7 has been shown to regulate EGFR expression and clinically, the coexpression of HER2 and EGFR in primary breast tumors is associated with significantly worse prognosis than tumors with HER2 or EGFR expression alone.
To determine the impact of miR-7 on EGFR expression and breast tumor cell proliferation and migration, we reestablished miR-7 expression in the highly invasive MCF-7/HER2Δ16 cell line (MCF-7/HER2Δ16/miR-7) and we suppressed miR-7 expression in the non-invasive MCF-7 cell line (MCF-7/miR-7KD). As predicted, reexpression of miR-7 in MCF-7/HER2Δ16 cells dramatically reduced EGFR protein levels, while suppression of miR-7 in MCF-7 cells increased EGFR protein. Furthermore, the reestablished miR-7 expression in MCF-7/HER2Δ16 cells significantly reduced the size and numbers of colonies in a colony formation assay, caused a G1 arrest of the cell cycle and abolished cell migration to the levels observed in noninvasive parental MCF-7 cells. Conversely, inhibition of miR-7 in MCF-7 cells resulted in an increase in colony formation and S-phase of the cell cycle. Taken together, these results indicate that suppression of miR-7 is necessary but not sufficient to promote breast tumor cell migration, while miR-7 directly regulates cellular proliferation.
To investigate the contribution of EGFR to HER2Δ16 tumorigenesis, we silenced EGFR expression using shRNAs (MCF-7/HER2Δ16/EGFRKD). While the MCF-7/HER2Δ16/miR-7 cells inhibited colony formation, the MCF-7/HER2Δ16/EGFRKD cells showed enhanced colony formation. However, suppression of EGFR significantly reduced HER2Δ16-induced cell migration activity. These results indicate that EGFR is an essential component of the HER2Δ16 cell migration pathway, but is dispensable for HER2Δ16 colony formation activity.
Interestingly, reexpression of miR-7 in the MCF-7/HER2Δ16 cell line resulted in a loss of Src kinase phosphorylated activation. Our preliminary results suggest that Src kinase activation in the absence of miR-7 drives HER2Δ16 tumor cell proliferation. We are currently investigating the contribution of miR-7 to HER2Δ16-induced resistance to both trastuzumab and lapatinib.
In summary, HER2Δ16 represents a novel breast oncogene that promotes breast tumor cell proliferation and metastasis through suppression of miR-7 and cooperation with Src kinase. Our data suggests that modulation of miR-7 expression can be implemented as a therapeutic approach for the treatment of the most aggressive and therapeutically refractory HER2-positive metastatic breast tumors.
Citation Format: Felicia C. Huynh, Frank E. Jones. HER2Δ16, the oncogenic isoform of HER2, suppresses microRNA-7 to promote breast tumor cell migration and proliferation. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research: Genetics, Biology, and Clinical Applications; Oct 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2013;11(10 Suppl):Abstract nr A008.