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Open Access Highly Accessed Research article

Pleurocidin-family cationic antimicrobial peptides are cytolytic for breast carcinoma cells and prevent growth of tumor xenografts

Ashley L Hilchie1, Carolyn D Doucette2, Devanand M Pinto34, Aleksander Patrzykat4, Susan Douglas14 and David W Hoskin125*

Author Affiliations

1 Department of Microbiology & Immunology, Dalhousie University, 5850 College St., Halifax, B3H 4R2, Canada

2 Department of Pathology, Dalhousie University, 5850 College St., Halifax, B3H 4R2, Canada

3 Department of Chemistry, Dalhousie University, 6274 Coburg Rd., Halifax, B3H 4R2, Canada

4 Institute for Marine Biosciences, National Research Council, 1411 Oxford St., Halifax, B3H 3Z1, Canada

5 Department of Surgery, Dalhousie University, 1276 South Park St., Halifax, B3H 4R2, Canada

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Breast Cancer Research 2011, 13:R102  doi:10.1186/bcr3043

Published: 24 October 2011

Abstract

Introduction

Cationic antimicrobial peptides (CAPs) defend against microbial pathogens; however, certain CAPs also exhibit anticancer activity. The purpose of this investigation was to determine the effect of the pleurocidin-family CAPs, NRC-03 and NRC-07, on breast cancer cells.

Methods

MTT (3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide) and acid phosphatase cell-viability assays were used to assess NRC-03- and NRC-07-mediated killing of breast carcinoma cells. Erythrocyte lysis was determined with hemolysis assay. NRC-03 and NRC-07 binding to breast cancer cells and normal fibroblasts was assessed with fluorescence microscopy by using biotinylated-NRC-03 and -NRC-07. Lactate dehydrogenase-release assays and scanning electron microscopy were used to evaluate the effect of NRC-03 and NRC-07 on the cell membrane. Flow-cytometric analysis of 3,3'-dihexyloxacarbocyanine iodide- and dihydroethidium-stained breast cancer cells was used to evaluate the effects of NRC-03 and NRC-07 on mitochondrial membrane integrity and reactive oxygen species (ROS) production, respectively. Tumoricidal activity of NRC-03 and NRC-07 was evaluated in NOD SCID mice bearing breast cancer xenografts.

Results

NRC-03 and NRC-07 killed breast cancer cells, including drug-resistant variants, and human mammary epithelial cells but showed little or no lysis of human dermal fibroblasts, umbilical vein endothelial cells, or erythrocytes. Sublethal doses of NRC-03 and, to a lesser extent, NRC-07 significantly reduced the median effective concentration (EC50) of cisplatin for breast cancer cells. NRC-03 and NRC-07 bound to breast cancer cells but not fibroblasts, suggesting that killing required peptide binding to target cells. NRC-03- and NRC-07-mediated killing of breast cancer cells correlated with expression of several different anionic cell-surface molecules, suggesting that NRC-03 and NRC-07 bind to a variety of negatively-charged cell-surface molecules. NRC-03 and NRC-07 also caused significant and irreversible cell-membrane damage in breast cancer cells but not in fibroblasts. NRC-03- and NRC-07-mediated cell death involved, but did not require, mitochondrial membrane damage and ROS production. Importantly, intratumoral administration of NRC-03 and NRC-07 killed breast cancer cells grown as xenografts in NOD SCID mice.

Conclusions

These findings warrant the development of stable and targeted forms of NRC-03 and/or NRC-07 that might be used alone or in combination with conventional chemotherapeutic drugs for the treatment of breast cancer.