BackgroundOvarian carcinoma (OvCa) is the most lethal gynecological malignancy among women and its poor prognosis is mainly due to metastasis. Chemokine receptor CCR9 is primarily expressed by a small subset of immune cells and its only natural ligand, CCL25, is largely expressed in the thymus, which involutes with age. Other than the thymus, CCL25 is expressed by the small bowel. Interactions between CCL25 and CCR9 have been implicated in leukocyte trafficking to the small bowel, a frequent metastatic site for OvCa cells. The current study shows OvCa tissue and cells significantly express CCR9, which interacts with CCL25 to support carcinoma cell migration and invasion.MethodsRT-PCR and flow cytometry techniques were used to quantify the expression CCR9 by OvCa cells. OvCa tissue microarrays (TMA) was used to confirm CCR9 expression in clinical samples. The Aperio ScanScope scanning system was used to quantify immunohistochemical staining. Cell invasion and migration assays were performed using cell migration and matrigel invasion chambers. Matrix metalloproteinase (MMP) mRNAs were quantified by RT-PCR and active MMPs were quantified by ELISA.ResultsOur results show significantly (p < 0.001) higher expression of CCR9 by mucinous adenocarcinoma, papillary serous carcinoma, and endometriod ovarian carcinoma cases, than compared to non-neoplastic ovarian tissue. Furthermore, CCR9 expression was significantly elevated in OvCa cell lines (OVCAR-3 and CAOV-3) in comparison to normal adult ovarian epithelial cell mRNA. OvCa cells showed higher migratory and invasive potential towards chemotactic gradients of CCL25, which was inhibited by anti-CCR9 antibodies. Expression of collagenases (MMP-1, -8, and -13), gelatinases (MMP-2 and -9), and stromelysins (MMP-3, -10, and -11) by OvCa cells were modulated by CCL25 in a CCR9-dependent fashion.ConclusionsThese results demonstrate both biological significance and clinical relevance of CCL25 and CCR9 interactions in OvCa cell metastasis.

BackgroundChemotherapy heavily relies on apoptosis to kill breast cancer (BrCa) cells. Many breast tumors respond to chemotherapy, but cells that survive this initial response gain resistance to subsequent treatments. This leads to aggressive cell variants with an enhanced ability to migrate, invade and survive at secondary sites. Metastasis and chemoresistance are responsible for most cancer-related deaths; hence, therapies designed to minimize both are greatly needed. We have recently shown that CCR9-CCL25 interactions promote BrCa cell migration and invasion, while others have shown that this axis play important role in T cell survival. In this study we have shown potential role of CCR9-CCL25 axis in breast cancer cell survival and therapeutic efficacy of cisplatin.MethodsBromodeoxyuridine (BrdU) incorporation, Vybrant apoptosis and TUNEL assays were performed to ascertain the role of CCR9-CCL25 axis in cisplatin-induced apoptosis of BrCa cells. Fast Activated Cell-based ELISA (FACE) assay was used to quantify In situ activation of PI3Kp85, AktSer473, GSK-3βSer9 and FKHRThr24 in breast cancer cells with or without cisplatin treatment in presence or absence of CCL25.ResultsCCR9-CCL25 axis provides survival advantage to BrCa cells and inhibits cisplatin-induced apoptosis in a PI3K-dependent and focal adhesion kinase (FAK)-independent fashion. Furthermore, CCR9-CCL25 axis activates cell-survival signals through Akt and subsequent glycogen synthase kinase-3 beta (GSK-3β) and forkhead in human rhabdomyosarcoma (FKHR) inactivation. These results show that CCR9-CCL25 axis play important role in BrCa cell survival and low chemotherapeutic efficacy of cisplatin primarily through PI3K/Akt dependent fashion.

BackgroundCisplatin is more often used to treat ovarian cancer (OvCa), which provides modest survival advantage primarily due to chemo-resistance and up regulated anti-apoptotic machineries in OvCa cells. Therefore, targeting the mechanisms responsible for cisplatin resistance in OvCa cell may improve therapeutic outcomes. We have shown that ovarian cancer cells express CC chemokine receptor-9 (CCR9). Others have also shown that CCL25, the only natural ligand for CCR9, up regulates anti-apoptotic proteins in immature T lymphocytes. Hence, it is plausible that CCR9-mediated cell signals might be involved in OvCa cell survival and inhibition of cisplatin-induced apoptosis. In this study, we investigated the potential role and molecular mechanisms of CCR9-mediated inhibition of cisplatin-induced apoptosis in OvCa cells.MethodsCell proliferation, vibrant apoptosis, and TUNEL assays were performed with or without cisplatin treatment in presence or absence of CCL25 to determine the role of the CCR9-CCL25 axis in cisplatin resistance. In situ Fast Activated cell-based ELISA (FACE) assays were performed to determine anti-apoptotic signaling molecules responsible for CCL25-CCR9 mediated survival.ResultsOur results show interactions between CCR9 and CCL25 increased anti-apoptotic signaling cascades in OvCa cells, which rescued cells from cisplatin-induced cell death. Specifically, CCL25-CCR9 interactions mediated Akt, activation as well as GSK-3β and FKHR phosphorylation in a PI3K-dependent and FAK-independent fashion.ConclusionsOur results suggest the CCR9-CCL25 axis plays an important role in reducing cisplatin-induced apoptosis of OvCa cells.

Breast cancer (BrCa) is one of the most frequently diagnosed cancers and the second leading cause of cancer-related deaths in North American women. Most deaths are caused by metastasis, and BrCa is characterized by a distinct metastatic pattern involving lymph nodes, bone marrow, lung, liver and brain. Migration of metastatic cells share many similarities with leukocyte trafficking, which are regulated by chemokines and their receptors. The current study evaluates the expression and functional role of CCR9, and its only known ligand, CCL25, in BrCa cell migration and invasion. Quantitative immunohistochemical analysis showed that both moderately and poorly differentiated BrCa tissue expressed significantly more (P<0.0001) CCR9 compared to non-neoplastic breast tissue. Interestingly, poorly differentiated BrCa tissue expressed significantly more (P<0.0001) CCR9 compared to moderately differentiated BrCa tissue. Similarly, CCR9 was highly expressed by the aggressive breast cancer cell line (MDA-MD-231) compared to the less aggressive MCF-7. Migration as well as invasion assays were used to evaluate the functional interaction between CCR9 and CCL25 in BrCa cell lines (MDA-MB-231 and MCF-7). Neutralizing CCR9-CCL25 interactions significantly impaired the migration and invasion of BrCa cells. Furthermore, CCL25 enhanced the expression of MMP-1, -9, -11 and -13 active proteins by BrCa cells in a CCR9-dependent fashion. These studies show CCR9 is functionally and significantly expressed by BrCa (poorly > moderately differentiated) tissue and cells as well as that CCL25 activation of this receptor promotes breast tumor cell migration, invasion and MMP expression, which are key components of BrCa metastasis.

Breast cancer (BrCa) is one of the most frequently diagnosed cancers and the second leading cause of cancer-related deaths in the U.S. Most cancer-related deaths are due to metastasis. In addition, BrCa cells readily become resistant to a wide variety of chemotherapeutic agents, which is a major obstacle in the treatment of BrCa. Tumor cell migration and metastasis share similarities with leukocyte trafficking, which is regulated, in part, by chemokines and their receptors. We have previously shown that CCR9 is functionally expressed by BrCa cells and its expression is enhanced by the presence of its ligand, CCL25. This study characterizes the role of CCL25-CCR9 cell-signaling cascades involved in BrCa cell resistance to cisplatin. This drug is often ineffective due to cancer cell resistance and the development of anti-apoptotic mechanisms. In this regard, CCR9-CCL25 interaction plays a role in immature T-cell survival through PI3K and G ai protein-dependent activation of Akt/protein kinase B. Activated Akt phosphorylates multiple downstream targets that are involved in cell survival signaling, which include glycogen synthase kinase (GSK-3), forkhead transcriptional factor (FKHR), and caspase-9. We examined the effect of CCR9-CCL25 interactions on BrCa cell lines MDA-MB-231 and MDA-435, with or without cisplatin treatment. CCR9-CCL25 interactions decreased cisplatin-induced apoptosis. These interactions also modulated the activation of cell signaling cascades involved in cell survival. These studies provide important new information regarding the cellular and molecular mechanisms that mediate BrCa cell survival.

Ovarian cancer (OvCa) is the most lethal among gynecologic malignancies. Recently, chemokines have been shown to function in non-random tumor-cell metastasis to distant organs in a fashion that is similar to chemokine-directed lymphocyte migration. Also, the interactions between CCR9 and its ligand, CCL25, have been implicated in leukocyte trafficking to the small intestines, a frequent metastatic site for OvCa. Previously, our laboratory has shown that ovarian cancer cells strongly express CCR9. We have also shown that CCL25 induces the chemotaxis of OvCa cell lines and increases the invasive potential of OvCa cells. Recent studies in T cells, have shown that CCR9-CCL25 interaction can induce anti-apoptotic pathways, namely through Akt/protein kinase B activation, which is PI3K- and G?i protein-dependent. Activated Akt phosphorylates multiple down stream targets that are involved in cell survival signals, which include glycogen synthase kinase-3? and forkhead transcriptional factor. Cisplatin is a commonly used anticancer drug for the treatment of OvCa. It has been shown that the anti-apoptotic signals of the PI3K-Akt survival pathways are involved in tumor cell cisplatin resistance. Here we characterize the molecular mechanisms of CCL25-CCR9 cell-signaling cascades involved in ovarian cancer survival and show that CCL25-CCR9 interactions mediate cisplatin survival through the activation of anti-apoptosis pathway. These studies will provide novel information regarding the cellular and molecular mechanisms, following CCL25-CCR9 interaction, that modulate ovarian cell metastasis and cisplatin resistance.