1645PD - Microrna-9 and -224 in trastuzumab resistant HER2 positive breast cancer cells

Date 01 October 2012
Event ESMO Congress 2012
Session Basic Science and Translational Research II
Topics Basic Science
Breast Cancer
Presenter Karen Howe
Authors K. Howe1, A. Eustace2, S. Souahli2, B.C. Browne2, S. Aherne2, N. Barron2, N. Walsh2, J.P. Crown2, N. O'Donovan2
  • 1Molecular Therapeutics For Cancer Ireland, National Institute For Cellular Biotechnology, Molecular Therapeutics for Cancer Ireland, Dublin City University, 9 - Dublin/IE
  • 2National Institute For Cellular Biotechnology,, Molecular Therapeutics for Cancer Ireland, Dublin City University, 9 - Dublin/IE

 

Abstract

HER2 positive breast cancer accounts for approximately 25 % of all breast cancer cases. Trastuzumab, a humanised monoclonal antibody, is an approved established treatment for HER2 positive breast cancer; however, patients that initially respond frequently develop resistance. The aim of this study is to investigate microRNAs in cell line models of acquired and innate trastuzumab resistance. MicroRNA was extracted from the HER2 positive cells; SKBR3, BT474, and the acquired trastuzumab resistant variants SKBR3-T and BT474-T, and from a panel of innate trastuzumab sensitive (BT474, EFM-192A, SKBR3 and MDA-MB-361) or resistant cell lines (UACC-732, JIMT-1, HCC-202, HCC-1954, HCC1569 and MDA-MB-453), in triplicate. MicroRNA profiling was performed on SKBR3 and SKBR3-T using Taqman Low Density Arrays (TLDA). Differentially regulated miRNAs were selected using > 2-fold change and a P-value of < 0.05. Individual quantitative RT-PCR (qRT-PCR) was performed to confirm alterations in miRNAs. Functional studies were carried out using Ambion® Pre-miR miRNA Precursors and Anti-miR miRNA Inhibitors for miR-224 in SKBR3 cells. TLDA analysis identified nine differentially regulated microRNAs in the SKBR3-T cells. Individual qRT-PCR assays confirmed that 5 miRNAs were up-regulated and 4 were down-regulated. MiR-9 was 2.2-fold up-regulated (p = 0.04), while miR-224 was 1.6-fold down-regulated (p = 0.01) in SKBR3-T compared to the SKBR3 cells. Further validation of these targets in the BT474 model showed that miR-224 expression is lost in the resistant cells compared to the parental BT474 cells. MiR-9 is not significantly altered in the BT474-T cells. In the innate resistant models, miR-224 expression is reduced or lost in 4/6 resistant cell lines. Expression of miR-9 does not significantly differ between the innately sensitive and resistant cell lines. Transfection of SKBR3 cells with pre-miR-224 significantly decreases cell proliferation by 23.5% (p = 0.04).

Conclusions

This is the first report of the involvement of miR-9 and miR-224 in trastuzumab resistance in HER2 positive breast cancer. Preliminary functional studies suggest that miR-224 may play a role in regulating cell growth in HER2 positive breast cancer cells.

Disclosure

All authors have declared no conflicts of interest.