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Anti-tissue factor short hairpin RNA inhibits breast cancer growth in vivo

  • Preclinical study
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Abstract

In breast cancer, there is a correlation between tissue factor (TF) expression, angiogenesis and disease progression. TF stimulates tumour angiogenesis, in part, through up-regulation of vascular endothelial growth factor (VEGF). Therefore, this study aimed to establish whether TF stimulates angiogenesis and tumour progression directly and independent of VEGF up-regulation. Initially, the effects of TF and VEGF were assessed on endothelial cell migration (Boyden chamber) and differentiation (tubule formation on Matrigel). Subsequently, MDA-MB-436 breast cancer cells, which produce high levels of both TF and VEGF (western blot analysis), were established in vivo, following which tumours were treated three times per week for 3 weeks with intra-tumoural injections of either anti-VEGF siRNA, anti-TF shRNA, the two treatments combined, or relevant controls. Both VEGF and TF significantly stimulated endothelial cell migration and tubule formation (P < 0.02). Breast cancer xenografts (MDA-MB-436) treated with TF or VEGF-specific agents demonstrated significant inhibition in tumour growth (VEGFsiRNA 61%; final volume: 236.2 ± 23.2 mm3 vs TFshRNA 89%; 161.9 ± 17.4 mm3 vs combination 93%; 136.3 ± 9.2 mm3 vs control 400.4 ± 32.7 mm3; P < 0.005). Microvessel density (MVD), a measure of angiogenesis, was also significantly inhibited in all groups (MVD in control = 29 ± 2.9; TFshRNA = 18 ± 1.1; VEGFsiRNA = 16.7 ± 1.5; both = 12 ± 2.1; P < 0.004), whereas the proliferative index of the tumours was only reduced in the TFshRNA-treated groups (control = 0.51 ± 0.011; TFshRNA = 0.41 ± 0.014; VEGFsiRNA = 0.49 ± 0.013; both = 0.41 ± 0.004; P < 0.008). These data suggest that TF has a direct effect on primary breast cancer growth and angiogenesis, and that specific inhibition of the TF-signalling pathway has potential for the treatment of primary breast cancer.

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Abbreviations

CCS:

Cumulative Chalkley score

GF:

Growth factor

HuDMEC:

Human dermal microvascular endothelial cells

MVD:

Microvessel density

PECAM:

Platelet endothelial cell adhesion molecule

TF:

Tissue factor

VEGF:

Vascular endothelial growth factor

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Acknowledgments

We gratefully acknowledge the technical support of Jenny Globe, Carmel Nichols and Yvonne Stephenson. This study was supported by Yorkshire Cancer Research (grant number S287).

Conflict of interest

The authors wish to declare a conflict of interest for co-author Dr. Mohammed Amarzguioui. He serves as Research Director for the company SiRNAsense AS, which is currently developing siRNA-based drugs targeting TF.

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Authors and Affiliations

  1. Microcirculation Research Group, Academic Unit of Surgical Oncology, Department of Oncology, Faculty of Medicine, Dentistry and Health, University of Sheffield Medical School, Beech Hill Road, Sheffield, South Yorkshire, S10 2RX, UK

    J. E. Bluff, J. Slattery, M. W. R. Reed, N. J. Brown & C. A. Staton

  2. siRNAsense, University of Oslo, Oslo, Norway

    M. Amarzguioui

Authors
  1. J. E. Bluff
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  2. M. Amarzguioui
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  3. J. Slattery
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  4. M. W. R. Reed
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  5. N. J. Brown
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  6. C. A. Staton
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Corresponding author

Correspondence to C. A. Staton.

Additional information

N. J. Brown and C. A. Staton—Joint senior authors.

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Bluff, J.E., Amarzguioui, M., Slattery, J. et al. Anti-tissue factor short hairpin RNA inhibits breast cancer growth in vivo. Breast Cancer Res Treat 128, 691–701 (2011). https://doi.org/10.1007/s10549-010-1149-8

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  • DOI: https://doi.org/10.1007/s10549-010-1149-8

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