Breast cancer normalization induced by embryonic mesenchyme is mediated by extracellular matrix biglycan

Integrative Biology 2013 5(8):1045-1056.

Bischof AG, Yüksel D, Mammoto T, Mammoto A, Krause S,  Ingber DE

Vascular Biology Program, Boston Children’s Hospital and Harvard Medical School, Boston MA 02115, USA; Graduate Program in Biophysics, Harvard University, Cambridge, MA 02138, USA; Wyss Institute for Biologically Inspired Engineering at Harvard University, Boston, MA 02115; and Harvard School of Engineering and Applied Sciences, Cambridge MA 02138, USA.

 

Abstract

Some epithelial cancers can be induced to revert to quiescent differentiated tissue when combined with embryonic mesenchyme; however, the mechanism of this induction is unknown. Here we combine tissue engineering, developmental biology, biochemistry and proteomics approaches to attack this problem. Using a synthetic reconstitution system, we show that co-culture of breast cancer cells with embryonic mesenchyme from early stage (E12.5-13.5) mammary glands decreases tumor cell proliferation while stimulating acinus differentiation, whereas cancer-associated fibroblasts (CAFs) fail to produce these normalizing effects. When insoluble extracellular matrices (ECMs) were isolated from cultured early stage (E12.5-13.5) embryonic mammary mesenchyme cells or E10 tooth mesenchyme and recombined with mammary tumor cells, they were found to be sufficient to induce breast cancer normalization including enhanced expression of estrogen receptor-α (ER-α). In contrast, ECM from later stage (E14.5) mammary mesenchyme and conditioned medium isolated from mesenchymal cell cultures were ineffective. Importantly, when the inductive ECMs produced by early stage embryonic mammary mesenchyme were scraped from dishes and injected into fast-growing breast tumors in mice, they significantly inhibited cancer expansion. Proteomics analysis of the detergent insoluble ECM material revealed several matrix components that were preferentially expressed in the embryonic ECMs. Analysis of two of these molecules previously implicated in cancer regulation–biglycan and tenascin C–revealed that addition of biglyan can mimic the tumor normalization response, and that siRNA knockdown of its expression in cultured embryonic mesenchyme results in loss of the ECM’s inductive activity. These studies confirm that embryonic mesenchyme retains the ability to induce partial breast cancer reversion, and that its inductive capability resides at least in part in the ECM protein biglycan that it produces.

PMID: 23817524

 

Supplement:

Cancer is generally viewed as an irreversible genetic disease of uncontrolled cell growth and hence, most current treatment options focus on the use of anti-proliferative therapies that can lead to significant systemic toxicity. However, several studies published more than 35 years ago suggested that some epithelial cancers can be induced to ‘normalize’ by suppressing their growth and exhibiting normal differentiated tissue morphology when combined with embryonic mesenchyme1 or extracellular matrix (ECM)2. These results were not pursued as molecular genetics came to the fore in the cancer research field. But in recent years, experiments have revealed that changes in the tumor stroma, including alterations in ECM structure, mechanics, and composition, can actively contribute to cancer progression3, making the microenvironment an important target for development of new therapeutics.

Ingber Donald Elliott-fig1

Figure 1. ECM deposited by embryonic mesenchymal cells induces breast cancer differentiation and formation of hollow mammary acini in 3D culture, whereas ECM produced by cancer fibroblasts does not.

 

In this research article, we experimentally confirmed that embryonic mesenchymal cells, but not adult cancer-associated fibroblasts, can induce normalization of various mouse and human breast cancer cells, as measured by suppressed growth, increased histodifferentiation (lumen and duct formation), and enhanced basement membrane accumulation (Figure 1). In addition, we show that detergent-extracted ECM from cultured embryonic mesenchymal cells is sufficient to induce mammary tumor cell normalization to the same extent as co-cultures with living cells. Early embryonic ECM also restores estrogen receptor-α expression, decreases epithelial cell migration in vitro, and most importantly, it suppresses cancer growth when injected into implanted syngeneic tumors in mice.

These findings provide support for development of a new type of differentiation therapy for solid cancers based on design of biomimetic inductive materials that mimic critical properties of the embryonic mesenchymal ECM. As a first step towards this goal, we performed proteomics analysis identifying 11 ECM proteins that are preferentially expressed in the inductive embryonic mesenchyme ECMs. Further study of two of these proteins led to the discovery that biglycan is a key mediator of this cancer normalization response.

 

References:

  1. DeCosse JJ, Gossens CL, Kuzma JF, Unsworth BR 1973. Breast cancer: induction of differentiation by embryonic tissue. Science 181:1057-8.
  2. Ingber DE, Madri JA, Jamieson JD 1986. Basement membrane as a spatial organizer of polarized epithelia. Exogenous basement membrane reorients pancreatic epithelial tumor cells in vitro. Am J Pathol. 122:129-39.
  3. Bissell MJ, Kenny PA, Radisky DC  2005.  Microenvironmental regulators of tissue structure and function also regulate tumor induction and progression: the role of extracellular matrix and its degrading enzymes. Cold Spring Harb Symp Quant Biol 70:343-56.

 

Acknowledgements: This work was supported by a Department of Defense Breast Cancer

Innovator Award (BC074986 to DEI) and the Wyss Institute for Biologically Inspired Engineering at Harvard University

 

Contact:

Donald Ingber, MD,PhD

Wyss Institute for Biologically Inspired Engineering at Harvard University

3 Blackfan Circle, CLSB 5th Floor

Boston MA 02115

don.ingber@wyss.harvard.edu

www.wyss.harvard.edu

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