Role of CD44s and CD44v6 on human breast cancer cell adhesion, migration, and invasion
Introduction
Breast cancer is one of the most common cancer among American women and world worldwide. In 2007, about 178,480 new cases of invasive breast cancer were diagnosed and approximately 40,460 women died from the disease nationwide (American, 2007). Despite improvements in treatment modalities, there is still a high failure rate mainly due to tumor invasion and metastasis. Breast cancer invasion is a three-step process characterized by altered cellular adhesion, cell motility and invasion of the extracellular matrix. Signals transmitted to the tumor cells from the peritumoral stroma will influence their activity, proliferation, and motility. This is accomplished in part through transmembrane receptors that interact with stromal extracellular matrix molecules. One of these receptors is CD44, which binds to the extracellular matrix component HA (Underhill, 1992). The engagement of CD44 with HA results in intracellular signaling that has been linked to diverse cellular functions such as adhesion, migration, and invasion, which are all important in cancer progression and metastasis (Toole, 2002, Naor et al., 1997).
CD44 (also known as homing cellular adhesion molecule, Hermes antigen, and PGP-1) is an 85–90 kDa transmembrane glycoprotein receptor comprising of four functional domains. The distal extracellular domain is primarily responsible for the binding of HA, while the proximal extracellular domain is the site of alternative splicing for CD44 mRNA that produces different isoforms of CD44. The CD44 transmembrane domain is a single 23 amino acid transmembrane domain typical of single-pass membrane glycoproteins. The intracellular domain interacts with cytoskeletal proteins and other intracellular signaling proteins. The gene encoding CD44 consists of 20 exons. In the standard form (CD44s), 10 of the 20 exons are transcribed. Multiple variant isoforms (CD44v2–v10) arise from alternate mRNA splicing of the remaining 10 exons (Tolg et al., 1993). In contrast to the standard form of CD44, which is usually ubiquitously expressed on epithelial cells and lymphocytes, CD44 variants exhibit tissue specific expression. Some of these variants, in particular splice variants of CD44v6 are associated with aggressive behavior and correlate with poor prognosis in a variety of human malignancies including breast cancer (Lopez et al., 2005, Auvinen et al., 2005, Joensuu et al., 1993).
CD44 specific monoclonal antibodies have been used extensively to understand the function of CD44 in cancer. Several monoclonal antibodies have been developed to recognize distinct epitopes on CD44. Antibody binding to specific epitopes on CD44 can clarify how different epitopes mediate distinct functional effects and influences of HA recognition (Lesley et al., 1993, Peach et al., 1993. Only CD44 antibodies that can block CD44–HA interaction are referred to as blocking antibodies and these are few in number (Zhong et al., 1995). Additionally, it has been established that HA-oligomers composed of at least 6 repeating disaccharides (HAOligo-6) is capable of binding to CD44 efficiently and blocking CD44–HA interaction (Nemec et al., 1987, Knudson et al., 1993).
In this current study we examined the effects of distinct antibodies against CD44s and one of its variants CD44v6 to understand the significance of their expression in mediating breast cancer cell adhesion, invasion, and migration.
Section snippets
Cell culture
Breast cancer cell lines MDA-MB-468, BT483 and MDA-MB-157 were acquired from the American Type Culture Collection (Rockville, MD). Breast cancer cell lines MDA-MB-231, MCF-7, and T47D were kindly provided by Dr. Hsing-Jing Kung (UC Davis). MDA-MB-468, MDA-MB-231, MCF-7, and T47D cells were cultured in Dulbecco's Modified Eagle Medium (GIBCO, Carlsbad, CA) supplemented with 10% fetal bovine serum (Omega Scientific, Tarzana, CA), l-glutamine [2 mM] (GIBCO), Pen Strep [100 μg/mL], and MEM
Expression of CD44s and CD44v6 on breast cancer cell lines
The expression of CD44s and CD44v6 receptor was characterized on 6 breast cancer cells lines including BT483, MDA-MB-157, MDA-MB-231, MDA-MB-468, MCF-7, and T47D. Western blotting of the breast cancer cell lines revealed that they expressed varying amounts of CD44s with a molecular weight between 80 and 90 kDa; the expected size for CD44s. The amount of CD44s varied significantly between the different cell lines. MDA-MB-468 express high levels of CD44s, while MDA-MB-157 and MDA-MB-231 expressed
Discussion
In recent years, numerous investigations have focused on the expression of CD44s and its isoforms in malignancy to further clarify their role in tumor progression and metastasis. We reported previously that in breast cancer, the tumor–stroma interface is enriched in HA, but how this could alter tumor cell functions is not well understood. Therefore, we explored the role of HA receptors, CD44s and CD44v6 on breast cancer cell adhesion, invasion, and migration.
We first characterized the
Conflict of interest statement
The authors declare that there are no conflicts of interest.
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