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Characterization of the expression of the pro-metastatic MenaINV isoform during breast tumor progression

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Abstract

Several functionally distinct isoforms of the actin regulatory Mena are produced by alternative splicing during tumor progression. Forced expression of the MenaINV isoform drives invasion, intravasation and metastasis. However, the abundance and distribution of endogenously expressed MenaINV within primary tumors during progression remain unknown, as most studies to date have only assessed relative mRNA levels from dissociated tumor samples. We have developed a MenaINV isoform-specific monoclonal antibody and used it to examine MenaINV expression patterns in mouse mammary and human breast tumors. MenaINV expression increases during tumor progression and to examine the relationship between MenaINV expression and markers for epithelial or mesenchymal status, stemness, stromal cell types and hypoxic regions. Further, while MenaINV robustly expressed in vascularized areas of the tumor, it is not confined to cells adjacent to blood vessels. Altogether, these data demonstrate the specificity and utility of the anti-MenaINV-isoform specific antibody, and provide the first description of endogenous MenaINV protein expression in mouse and human tumors.

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Abbreviations

DCIS:

Ductal carcinoma in situ

EGF:

Epidermal growth factor

EMT:

Epithelial to mesenchymal transition

FFPE:

Formalin fixed paraffin embedded

GFP:

Green fluorescent protein

IP:

Immunoprecipitation

LN:

Lymph node

SMA:

Smooth muscle actin

TMEM:

Tumor microenvironment of metastasis

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Acknowledgments

This work was supported by Department of Defense Breast Cancer Research Program Grants W81XWH-10-1-0040 to SKH and W81XWH-13-1-0031 to MJO, NIH Grant U54-CA112967 to FBG and DAL and GM58801 to FBG, funds from the Ludwig Center at MIT to FBG and NR, the KI NCI Core Grant P30-CA14051, and CA150344 to JC and CA100324 for JJ. JJ, JC and FG are compensated members if the scientific advisory board of MetaStat. No funding was provided by MetaStat for this work. We thank the Histology facilities in the KI Swanson Biotechnology Center for support. We thank Evanthia Roussos for gifts of tumor tissue.

Author contributions

MJO designed and performed experiments, data analysis and prepared the manuscript. SKH developed and validated the INV antibody, designed and performed experiments and completed data analysis. NR performed immunostaining experiments and image analysis. MNM developed MenaINV ELISA, performed experiments and completed data analysis. FBG and DAL guided overall experimental design. JJ and JC contributed human tissue sections and interpretation of them. All authors commented on the manuscript.

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Correspondence to Madeleine J. Oudin.

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Madeleine J. Oudin and Shannon K. Hughes have contributed equally to this work.

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Figure S1

Table with characteristics of human tumors. Table summarizing clinical features of human breast cancer tumors used for immunohistochemical analyses: tumor size, ER, PR or Her2 status, lymph node (LN) positivity, and site of metastasis (PDF 188 kb)

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Oudin, M.J., Hughes, S.K., Rohani, N. et al. Characterization of the expression of the pro-metastatic MenaINV isoform during breast tumor progression. Clin Exp Metastasis 33, 249–261 (2016). https://doi.org/10.1007/s10585-015-9775-5

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  • DOI: https://doi.org/10.1007/s10585-015-9775-5

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