**3.1.3 SDC-1 and SDC-4**

440 Breast Cancer – Focusing Tumor Microenvironment, Stem Cells and Metastasis

Fig. 2. Expression of NRP-1, SDC-4 and CSPG4 in breast cancer cells. Cells were grown in standard medium, harvested and then stained with monoclonal antibodies against the

NRP-1 is a 120-130 kDa transmembrane glycoprotein, initially characterized as a neuronal receptor for specific secreted members of the semaphorin family involved in exon repulsation (Kolodkin et al., 1997). A substantial fraction of NRP-1 is a PG with a GAG chain attached (Shintani et al., 2006). In addition to being a receptor for a number of class 3 semaphorins, NRP-1 also serves as a receptor for some members of vascular endothelial growth factor (VEGF), and placental growth factor (PlGF) (Migdal et al., 1998; Soker et al.,

Considerable data support a functional role for NRP-1 in regulating VEGF activities in endothelium. It has been shown that semaphorin-3A competes with VEGF165 binding to NRP-1 and inhibits angiogenesis *in vitro* (Miao et al., 1999). NRP-1 knock-out mice, in addition to neural defects, exhibit transposition of large vessels, disorganized and insufficient capillary

indicated targets. Stained cells were then analyzed by flow cytometry.

1998; Makinen et al., 1999; Wise et al., 1999; Klagsbrun et al., 2002).

**3.1.2 NRP-1** 

SDC-1 is mainly expressed by epithelia and plasma cells. Although there are inconsistent reports (Barbareschi et al., 2003; Tsanou et al., 2004), the expression of SDC-1 is generally down-regulated in malignant tumors, and lower levels of expression have been associated with high metastatic/aggressive potential in many tumors (Nackaerts et al., 1997; Kumar-Singh et al., 1998; Mikami et al., 2001; Numa et al., 2002). SDC-1 has also been shown to act as a tumor suppressor molecule by inhibiting cell growth and inducing apoptosis (Mali et al., 1994; Dhodapkar et al., 1998). Therefore, during tumor development the decrease of SDC-1 expression may be an important step from tumorigenesis to a metastatic phenotype. However, there are conflicting data on the role of SDC-1; both its loss and over-expression in carcinoma cells have been associated with malignant progression (Baba et al., 2006).

SDC-4 is more ubiquitously expressed by most cell types, and little is known about its role in malignancy. Among the four members of the syndecan family, SDC-4 is the only one involved in the formation of fibronectin-induced focal adhesions, in cooperation with β1 integrin receptors (Woods and Couchman, 1994; Woods et al., 2000). SDC-4 has been implicated in cytoskeletal organization and regulation of cell adhesiveness. The migratory capacity of lymphocytes and dendritic cells has been reported to be mediated by SDC-4 (Kaneider et al., 2002; Greene et al., 2003; Feistritzer et al., 2004; Averbeck et al., 2007). Our data suggest a role for relative expression of SDC-1 and SDC-4, low SDC-1 and high SDC-4 expression, in metastatic breast cancer cells (**Figure 3**).

Fig. 3. Relative expression of SDC-1 and SDC-4 in human breast cancer cells using quantitative real-time PCR. Means of three independent experiments (±SD) are shown.

On the Role of Cell Surface Chondroitin Sulfates and

protein).

**5. Conclusion** 

approaches.

**7. References** 

1443-1447.

1, pp, 91-98.

**6. Acknowledgement** 

discussions regarding directions for these studies.

screening. *Ann Oncol,* Vol., No., pp.

Their Core Proteins in Breast Cancer Metastasis 443

patients with aggressive breast cancer falls in line with our data (Wang et al., 2010a; Wang et al., 2010b). However, targeting a core protein may bring in specificity issues as these PGs are also expressed in stroma. Additionally, tumor cells can escape treatment by immune editing and replacing a PG with another one. 3) Targeting a combination of sugar and PG that can be accomplished by simultaneous targeting of the core protein and the polysaccharide, or by developing reagents like mAb specific for the whole entity (polysaccharide and the core

Breast cancer cell surface CS-GAGs and their interaction with P-selectin should be considered as viable targets for the development of novel diagnostic or therapeutic strategies. Our studies suggest that CS-GAGs, their biosynthetic pathway, or the core protein carrying them, can be potential targets in dealing with aggressive breast tumors. However, in order to efficiently block tumor cell dissemination by interrupting Pselectin/CS interaction, targeting any single PG does not seem to be enough, as other PGs can probably compensate and support metastatic processes. In this regard, global targeting of specific CS isomers, or combined targeting of the glycan and the PG, may be effective

The study was supported in part by a pilot project grant through the UAMS Center for Clinical and Translational Research, Award Number 1UL1RR029884 from the National Center for Research Resources, and in part by a UAMS College of Medicine grant (both to BMK). The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. We thank Dr. Thomas Kieber-Emmons for useful

Alini, M & Losa, G A (1991). Partial characterization of proteoglycans isolated from

Autier, P, Boniol, M, Middleton, R, Dore, J F, Hery, C, Zheng, T & Gavin, A (2011).

Averbeck, M, Gebhardt, C, Anderegg, U, Termeer, C, Sleeman, J P & Simon, J C (2007).

Baba, F, Swartz, K, van Buren, R, Eickhoff, J, Zhang, Y, Wolberg, W & Friedl, A (2006).

Barbareschi, M, Maisonneuve, P, Aldovini, D, Cangi, M G, Pecciarini, L, Angelo Mauri, F,

dendritic cell motility. *Exp Dermatol,* Vol. 16, No. 7, pp, 580-589.

neoplastic and nonneoplastic human breast tissues. *Cancer Res,* Vol. 51, No. 5, pp,

Advanced breast cancer incidence following population-based mammographic

Switch in syndecan-1 and syndecan-4 expression controls maturation associated

Syndecan-1 and syndecan-4 are overexpressed in an estrogen receptor-negative, highly proliferative breast carcinoma subtype. *Breast Cancer Res Treat,* Vol. 98, No.

Veronese, S, Caffo, O, Lucenti, A, Palma, P D, Galligioni, E & Doglioni, C (2003).

Therefore, relative expression of certain PGs or modification in their GAG chains may affect tumor aggressive phenotype through promoting survival, growth, and the metastatic capability of tumor cells. P-selectin can bind to CS-GAGs of these PGs and binding to each PG can have different functional consequences. These molecules has been linked to motility, invasion, angiogenesis, and cancer stem cell properties. Therefore, depending on the setting and expression of other molecules, P-selectin interaction may lead to various tumor promoting outcomes.

In studying the role of P-selectin in tumor growth and metastasis in a P-selectin-deficient Rag2-/- background, it was demonstrated that growth of subcutaneously challenged tumor cells were reduced significantly in the absence of P-selectin (Kim et al., 1998). This significantly slower growth rate in P-selectin deficient mice was unexpected because Pselectin is assumed to play a role in leukocytic infiltrates within tumors, which are generally inversely associated with tumor growth (Kreider et al., 1984). These findings, consistent with our hypothesis, demonstrate that the presence of P-selectin ligands on tumor cells and P-selectin-mediated interactions with stroma leads to tumorigenesis and tumor growth promotion.
