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STC1 expression is associated with tumor growth and metastasis in breast cancer

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Abstract

Stanniocalcin-1 (STC1) is a secreted glycoprotein implicated in several pathologies including retinal degeneration, cerebral ischemia, angiogenesis and inflammation. Aberrant STC1 expression has been reported in breast cancer but the significance of this is not clear. High levels of STC1 expression were found in the aggressive 4T1 murine mammary tumor cells and in the MDA-MB-231 human breast cancer line. To investigate its significance, stable clones with STC1 down-regulation using shRNA were generated in both tumor models. The consequences of STC1 down-regulation on cell proliferation, chemotactic invasion, tumor growth and metastasis were assessed. Down-regulation of STC1 in the 4T1 murine mammary tumor cells had a major impact on mammary tumor growth. This observation was replicated in a second tumor model with the MDA-MB-231 human breast cancer line, with a significant reduction in primary tumor formation and a major inhibition of metastasis as well. Interestingly, in both models, proliferation in vitro was not affected. Subsequent microarray gene expression profiling identified 30 genes to be significantly altered by STC1 down-regulation, the majority of which are associated with known hallmarks of carcinogenesis. Furthermore, bioinformatic analysis of breast cancer datasets revealed that high expression of STC1 is associated with poor survival. This is the first study to show definitively that STC1 plays an oncogenic role in breast cancer, and indicates that STC1 could be a potential therapeutic target for treatment of breast cancer patients.

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Abbreviations

BLI:

Bioluminescence intensity

STC1:

Stanniocalcin-1

shRNA:

Short hairpin RNA

siRNA:

Small interfering RNA

FCS:

Fetal calf serum

qRT-PCR:

Quantitative reverse transcriptase PCR

PCR:

Polymerase chain reaction

GAPDH:

Glyceraldehydes 3-phosphate dehydrogenase

SCID:

Severe combined immunodeficiency

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Acknowledgments

This project was supported by a Project Grant (APP633262) from the National Health and Medical Research Council of Australia, and by an Australian National Breast Cancer Foundation Fellowship to RLA. We thank Dr ZL Ou for the MDA-MB-231HM cells, Dr Joey Lai for microarray analysis and Dr Erdahl Teber for bioinformatics assistance. We also thank Dr Belinda Parker for generating the data in Supplementary Fig. 1a.

Conflict of interest

The authors declare no competing interests and this manuscript does not contain clinical studies or patient data.

Author information

Authors and Affiliations

  1. Cancer Research Unit, Children’s Medical Research Institute, Westmead, NSW, Australia

    Andy C-M Chang, Lily I. Huschtscha & Roger R. Reddel

  2. Faculty of Medicine, University of Sydney, Sydney, NSW, Australia

    Andy C-M Chang, Lily I. Huschtscha & Roger R. Reddel

  3. Metastasis Research Laboratory, Peter MacCallum Cancer Centre, East Melbourne, Australia

    Judy Doherty, Richard Redvers & Christina Restall

  4. Research Division, Peter MacCallum Cancer Centre, Locked Bag #1, A’Beckett Street, Melbourne, VIC, 8006, Australia

    Robin L. Anderson

  5. Sir Peter MacCallum Department of Oncology, The University of Melbourne, VIC, Australia

    Robin L. Anderson

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  1. Andy C-M Chang
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  2. Judy Doherty
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  3. Lily I. Huschtscha
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  4. Richard Redvers
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  6. Roger R. Reddel
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  7. Robin L. Anderson
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Corresponding author

Correspondence to Robin L. Anderson.

Additional information

Andy C-M Chang and Judy Doherty have contributed equally to this work.

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Fig. 1

a Average STC1 and STC2 mRNA levels from 6 individual primary tumors samples generated from three lines of the 4T1 model. RTA relative transcript abundance (arbitrary units) as measured by qRT-PCR and compared to GAPDH levels. b Gene expression of STC1 and STC2 derived from the cDNA microarray dataset of 52 cell lines from Kao et al [22]. Data from three cell lines available in our laboratory are shown (PPTX 69 KB)

Fig. 2

Tumor growth and metastasis of control and STC1 reduced 4T1ch9 tumors. a Primary tumor growth. b Average time to resection of primary tumors. c Visual metastasis score for each mouse. n = 13 for control tumors and n = 11 for STC1 reduced tumors (PPTX 120 KB)

Fig. 3

Hematoxylin and eosin stained sections of primary mammary tumors isolated from SCID mice injected with MDA-MB-231HM-luc non-silencing control (#758) and STC1 knockdown (#339 and #858) clones (PPTX 841 KB)

Fig. 4

Bioluminescence imaging of metastasis to the thorax of mice bearing MDA-MB231HM-luc mammary tumors at 42 days after implantation. Primary tumors were masked to enable imaging of metastases to thoracic region (PPTX 566 KB)

Fig. 5

To validate the microarray data, the expression levels of 11 genes randomly selected were analyzed by qRT-PCR and found to display similar expression profiles to those in the microarray data. STC1 and GAPDH were used as reference. PCR for each gene was done in triplicate reactions (PPTX 65 KB)

Supplementary material 6 (DOCX 17 kb)

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Chang, A.CM., Doherty, J., Huschtscha, L.I. et al. STC1 expression is associated with tumor growth and metastasis in breast cancer. Clin Exp Metastasis 32, 15–27 (2015). https://doi.org/10.1007/s10585-014-9687-9

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  • DOI: https://doi.org/10.1007/s10585-014-9687-9

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