Hyaluronan-CD44 Interaction with Protein Kinase Cϵ Promotes Oncogenic Signaling by the Stem Cell Marker Nanog and the Production of MicroRNA-21, Leading to Down-regulation of the Tumor Suppressor Protein PDCD4, Anti-apoptosis, and Chemotherapy Resistance in Breast Tumor Cells

Abstract

Multidrug resistance and disease relapse is a challenging clinical problem in the treatment of breast cancer. In this study, we investigated the hyaluronan (HA)-induced interaction between CD44 (a primary HA receptor) and protein kinase C⑀ (PKC⑀), which regulates a number of human breast tumor cell functions. Our results indicate that HA binding to CD44 promotes PKC⑀ activation, which, in turn, increases the phosphorylation of the stem cell marker, Nanog, in the breast tumor cell line MCF-7. Phosphorylated Nanog is then translocated from the cytosol to the nucleus and becomes associated with RNase III DROSHA and the RNA helicase p68. This process leads to microRNA-21 (miR-21) production and a tumor suppressor protein (e.g. PDCD4 (program cell death 4)) reduction. All of these events contribute to up-regulation of inhibitors of apoptosis proteins (IAPs) and MDR1 (multidrug-resistant protein), resulting in anti-apoptosis and chemotherapy resistance. Transfection of MCF-7 cells with PKC⑀ or Nanog-specific small interfering RNAs effectively blocks HA-mediated PKC⑀-Nanog signaling events, abrogates miR-21 production, and increases PDCD4 expression/eIF4A binding. Subsequently, this PKC⑀-Nanog signaling inhibition causes IAP/MDR1 down-regulation, apoptosis, and chemosensitivity. To further evaluate the role of miR-21 in oncogenesis and chemoresistance, MCF-7 cells were also transfected with a specific anti-miR-21 inhibitor in order to silence miR-21 expression and inhibit its target functions. Our results indicate that anti-miR-21 inhibitor not only enhances PDCD4 expression/eIF4A binding but also blocks HA-CD44-mediated tumor cell behaviors. Thus, this newly discovered HA-CD44 signaling pathway should provide important drug targets for sensitizing tumor cell apoptosis and overcoming chemotherapy resistance in breast cancer cells.Chemotherapeutic failure frequently contributes to morbidity in patients diagnosed with solid tumors, such as breast cancers (1-3). Recent studies indicate that oncogenic signaling and tumor cell-specific function are directly involved in chemotherapeutic drug resistance and breast tumor progression (4 -6). A number of studies have aimed at identifying those molecules that are specifically expressed by epithelial tumor cells and correlate with metastatic behavior and chemoresistance. Among such molecules is hyaluronan (HA), 2 a major component in the extracellular matrix of most mammalian tissues (7,8). HA is a nonsulfated, unbranched glycosaminoglycan, consisting of repeating disaccharide units, D-glucuronic acid, and N-acetyl-D-glucosamine (9, 10). The biosynthesis of HA is regulated by three mammalian HA synthase isozymes, HA synthase 1, 2, and 3 (11-14). Abnormal production of HA directly contributes to aberrant cellular processes, such as transformation and metastasis (15). Furthermore, HA is digested into a variety of smaller-sized molecules by various hyaluronidases (16). Activation of extracellular matrix-degrading enzymes, such as the hyaluronidases, appears to be closely associated w...

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“…miR-21 is also mentioned several times in the earlier discussion on pathway targets as being involved in different cancers including pancreas (50), lung (51), and breast (45). Increased miR-21 expression has been shown to be indicative of chemoresistance through 2 pathways.…”

“…PTEN and miR-21, -101, and -449a/b PTEN is a tumor suppressor gene that is known to regulate apoptosis and cell invasion (45). In a recent preliminary report by Sumaiyah et al (46), upregulation of miR-21 in breast cancer cells correlates with decreased expression of PTEN and an increased resistance to the HER2 monoclonal antibody trastuzumab.…”

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