Effect of the EGFR density of breast cancer cells on nuclear importation, in vitro cytotoxicity, and tumor and normal-tissue uptake of [111In]DTPA-hEGF

Hu, Meiduo; Scollard, Deborah; Chan, Conrad; Chen, Paul; Vallis, Katherine; Reilly, Raymond M.

doi:10.1016/j.nucmedbio.2007.06.010

Next »Nuclear Medicine and Biology
Volume 34, Issue 8 , Pages 887-896, November 2007

Effect of the EGFR density of breast cancer cells on nuclear importation, in vitro cytotoxicity, and tumor and normal-tissue uptake of [¹¹¹In]DTPA-hEGF

Part of this work was presented at the Canadian Breast Cancer Research Alliance Reasons for Hope Conference, Montreal, Quebec, May 6–8, 2006.

Meiduo Hu
- Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada M5S 3M2
Deborah Scollard
- Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada M5S 3M2
Conrad Chan
- Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada M5S 3M2
Paul Chen
- Neopharm, Inc., Waukegan, MI 60085-8328, USA
Katherine Vallis
- Radiobiology Research Institute, University of Oxford, Oxford OX3 7LJ, UK
Raymond M. Reilly
- Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada M5S 3M2
- Department of Medical Imaging, University of Toronto, Toronto, Ontario, Canada M5G 1X5
- Toronto General Research Institute, University Health Network, Toronto, Ontario, Canada M5G 2C4
- Corresponding author. Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada M5S 3M2. Tel.: +1 416 946 5522; fax: +1 416 978 8511.

Received 27 April 2007; received in revised form 15 June 2007; accepted 25 June 2007. published online 03 September 2007.

Abstract

Introduction

Our objective was to evaluate the effect of epidermal growth factor receptor(s) (EGFR) density on the importation and nuclear localization of ¹¹¹In-labeled diethylenetriaminepentaacetic acid human epidermal growth factor ([¹¹¹In]DTPA-hEGF) in breast cancer (BC) cells in vitro and in tumor xenografts and normal tissues in vivo in athymic mice, as well as on its cytotoxicity and tumor and normal-tissue distribution.

Methods

The internalization and nuclear importation of [¹¹¹In]DTPA-hEGF were measured in MCF-7, MDA-MB-231, BT-474 and MDA-MB-468 BC cells (10⁴, 2×10⁵, 6×10⁵ and 10⁶ EGFR/cell, respectively). The molecular size (M_r) distribution and immunoreactivity of nuclear radioactivity were characterized. Tumor and normal-tissue uptake of [¹¹¹In]DTPA-hEGF in athymic mice implanted subcutaneously with BC xenografts were compared. Nuclear radioactivity in the tumor, lungs, liver, kidneys, spleen and colon was measured.

Results

There was a direct association between EGFR density and the nuclear localization of [¹¹¹In]DTPA-hEGF in BC cells; nuclear importation approached saturation at 6×10⁵ EGFR/cell. Almost all nuclear radioactivities exhibited an M_r of >100 kDa; immunoreactivity with anti-hEGF, anti-EGFR and anti-importin β₁ antibodies was detected. The efflux of nuclear radioactivity was slowest for MDA-MB-468 cells. Cytotoxicity was correlated with EGFR expression. Uptake was greater in MDA-MB-468 than in MCF-7 xenografts and improved with preinjection of a 100-fold excess of unlabeled DTPA-hEGF. Nuclear importation was higher in liver, kidney and spleen cells than in tumor cells.

Conclusion

[¹¹¹In]DTPA-hEGF is translocated to the nucleus of BC cells complexed with EGFR and importin β₁. Nuclear importation and cytotoxicity are effected by EGFR density. The absence of hepatic and renal toxicities in [¹¹¹In]DTPA-hEGF cannot be explained by a low efficiency of nuclear importation.

Keywords: Indium-111, Epidermal growth factor, Epidermal growth factor receptor, Nuclear importation, Breast cancer, Importin β₁