Breast Cancer Research and Treatment

, Volume 133, Issue 2, pp 459–471

The tumor microenvironment modulates tamoxifen resistance in breast cancer: a role for soluble stromal factors and fibronectin through β1 integrin

  • Osvaldo Pontiggia
  • Rocio Sampayo
  • Diego Raffo
  • Andrea Motter
  • Ren Xu
  • Mina J. Bissell
  • Elisa Bal de Kier Joffé
  • Marina Simian
Preclinical study

DOI: 10.1007/s10549-011-1766-x

Cite this article as:
Pontiggia, O., Sampayo, R., Raffo, D. et al. Breast Cancer Res Treat (2012) 133: 459. doi:10.1007/s10549-011-1766-x

Abstract

Tamoxifen resistance has been largely attributed to genetic alterations in the epithelial tumor cells themselves, such as overexpression of HER-2/Neu. However, in the clinic, only about 15–20% of cases of HER-2/Neu amplification has actually been correlated to the acquisition of endocrine resistance, suggesting that other mechanisms must be involved as well. Using the epithelial LM05-E and the fibroblastic LM05-F cell lines, derived from the estrogen dependent spontaneous M05 mouse mammary tumor, as well as MCF-7 cells, we analyzed whether soluble stromal factors or extracellular matrix components protected against tamoxifen induced cell death. Involvement of signaling pathways was determined by using specific inhibitors and western blot, and phosphorylation of the estrogen receptor alpha by western blot and immunofluorescence. Soluble factors produced by the fibroblastic cells protect the epithelial tumor cells from tamoxifen-induced cell death through a mechanism that involves EGFR and matrix metalloproteinases upstream of PI3K/AKT. Exogenous fibronectin by itself confers endocrine resistance through interaction with β1 integrin and activation of PI3K/AKT and MAPK/ERK 1/2 pathways. The conferred resistance is reversed by blocking β1 integrin. We show also that treatment with both conditioned medium and fibronectin leads to the phosphorylation of the estrogen receptor at serine-118, suggesting stromal factors as modulators of ER activity. Our results show that the tumor microenvironment can modulate tamoxifen resistance, providing an alternative explanation for why patients become refractory to hormone-therapy.

Keywords

Breast cancer Tamoxifen resistance Estrogen receptor Tumor microenvironment Fibronectin Soluble stromal factors β1 integrin 

Abbreviations

MMP

Matrix metalloproteinase

ER

Estrogen receptor

FN

Fibronectin

FCM

LM05-F conditioned medium

NMC

Non conditioned medium

PI

Propidium iodide

DAPI

4′,6-Diamino-2-phenylindole

EGFR

Epidermal growth factor receptor

pSer-118 ER

Phosphor-serine-118 ER

ECM

Extracellular matrix

E2

Estradiol

PD

PD98059

LY

LY294002

T

4-OH-tamoxifen

Supplementary material

10549_2011_1766_MOESM1_ESM.tif (2.8 mb)
Supplementary Fig. 1LM05-E cells were starved for 48 h and then treated for 1 h with either vehicle, the PI3K/AKT inhibitor LY294002 (LY; 10 μM) (a), the MAPK/ERK inhibitor PD98059 (PD; 10 μM) (b) or the EGFR inhibitor AG1478 (AG; 6.4 μM) (c). At that time they were treated or not for 10 min with FCM. Samples were processed for western blot. As shown the three inhibitors were effective at the concentrations used. d LM05-E cells were plated on FN and treated with 10 nM estradiol (E2) or estradiol plus the PI3K/AKT inhibitor LY294002 (LY) or the MAPK/ERK inhibitor PD98059 (PD), both at a 10 μM concentration. Under these experimental conditions none of the inhibitors induced cell death (TIFF 2913 kb)

Copyright information

© Springer Science+Business Media, LLC. 2011

Authors and Affiliations

  • Osvaldo Pontiggia
    • 1
  • Rocio Sampayo
    • 1
  • Diego Raffo
    • 1
  • Andrea Motter
    • 1
  • Ren Xu
    • 2
    • 3
  • Mina J. Bissell
    • 2
  • Elisa Bal de Kier Joffé
    • 1
  • Marina Simian
    • 1
  1. 1.Área de InvestigacionesInstituto de Oncología “Ángel H. Roffo”Buenos AiresArgentina
  2. 2.Life Sciences DivisionLawrence Berkeley National LaboratoryBerkeleyUSA
  3. 3.Department of Molecular and Biomedical PharmacologyUniversity of KentuckyLexingtonUSA

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