[Show abstract][Hide abstract] ABSTRACT: IL-17A is a pro-inflammatory cytokine that is normally associated with autoimmune arthritis and other pro-inflammatory conditions. Recently, IL-17A has emerged as a critical factor in enhancing breast cancer (BC)-associated metastases. We generated immune competent arthritic mouse models that develop spontaneous BC-associated bone and lung metastasis. Using these models, we have previously shown that neutralization of IL-17A resulted in significant reduction in metastasis. However, the underlying mechanism/s remains unknown.
We have utilized two previously published mouse models for this study: 1) the pro-arthritic mouse model (designated SKG) injected with metastatic BC cell line (4T1) in the mammary fat pad, and 2) the PyV MT mice that develop spontaneous mammary gland tumors injected with type II collagen to induce autoimmune arthritis. Mice were treated with anti-IL-17A neutralizing antibody and monitored for metastasis and assessed for pro-inflammatory cytokines and chemokines associated with BC-associated metastasis.
We first corroborate our previous finding that in vivo neutralization of IL-17A significantly reduced metastasis to the bones and lungs in both models. Next, we report that treatment with anti-IL17A antibody significantly reduced the expression of a key chemokine, CXCL12 (also known as stromal derived factor-1 (SDF - 1)) in the bones and lungs of treated mice. CXCL12 is a ligand for CXCR4 (expressed on BC cells) and their interaction is known to be critical for metastasis. Interestingly, levels of CXCR4 in the tumor remained unchanged with treatment. Consequently, protein lysates derived from the bones and lungs of treated mice were significantly less chemotactic for the BC cells than lysates from untreated mice; and addition of exogenous SDF-1 to the lysates from treated mice completely restored BC cell migration. In addition, cytokines such as IL-6 and M-CSF were significantly reduced in the lung and bone lysates following treatment. The data presented suggests that systemic neutralization of IL-17A can block the CXCR4/SDF-1 signaling pathway by reducing the expression of SDF-1 in the metastatic niches and significantly reducing metastasis in both mouse models.
In our model, neutralization of IL-17A regulates SDF-1 expression in the metastatic niches either directly or indirectly via reducing levels of IL-6 and M-CSF.
BMC Cancer 03/2014; 14(1):225. · 3.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The transmembrane glycoprotein Mucin 1 (MUC1) is aberrantly glycosylated and overexpressed in a variety of epithelial cancers, and plays a crucial role in progression of the disease. Tumor-associated MUC1 differs from the MUC1 expressed in normal cells with regard to its biochemical features, cellular distribution, and function. In cancer cells, MUC1 participates in intracellular signal transduction pathways and regulates the expression of its target genes at both the transcriptional and post-transcriptional levels. This review highlights the structural and functional differences that exist between normal and tumor-associated MUC1. We also discuss the recent advances made in the use of MUC1 as a biomarker and therapeutic target for cancer.
Trends in Molecular Medicine 03/2014; · 9.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mucin 1 (MUC1) is a transmembrane mucin glycoprotein that is over-expressed and aberrantly glycosylated in >80% of human pancreatic ductal adenocarcinoma (PDA) and is associated with poor prognosis. To understand the role of MUC1 in PDA, we have recently developed two mouse models of spontaneous PDA, one that expresses full-length human MUC1 transgene (KCM mice) and one that is null for MUC1 (KCKO mice). We have previously reported that KCM mice express high levels of myeloid derived suppressor cells (MDSCs) in their tumors and develop highly aggressive PDA. To further understand the underlying mechanism for high MDSC levels in KCM-tumors, we generated primary cell lines from KCM and KCKO-tumors. In this study, we report that MDSCs derived using KCM cells express significantly higher levels of arginase 1 and inducible nitric oxide synthase (markers associated with immune suppression) and lower levels of CD115 (a marker associated with maturation of myeloid cells) as compared to KCKO-derived MDSCs. Functionally, KCM-derived MDSCs secrete significantly higher levels of urea and nitric oxide (NO) when co-cultured with normal splenic cells as compared to KCKO-derived MDSCs. Data indicates that KCM-derived MDSCs remain immature and are more suppressive as compared to KCKO-derived MDSCs. This was further corroborated in vivo where MDSCs isolated from KCM-tumor-bearing mice retained their immature state and were highly suppressive as compared to MDSCs derived from KCKO-tumor-bearing mice. Finally, we show that KCM cells secrete significantly higher levels of prostaglandin E2 (PGE2), a COX-2 metabolite and a known driver of suppressive MDSCs as compared to KCKO cells. Thus, inhibiting PGE2 with a specific COX-2 inhibitor reverses the immunosuppressive and immature phenotype of KCM-derived MDSCs. This is the first report that clearly suggests a functional role of pancreatic tumor-associated MUC1 in the development of functional MDSCs.
[Show abstract][Hide abstract] ABSTRACT: Vesicular stomatitis virus (VSV) is a promising oncolytic agent against various malignancies. Here, for the first time, we tested VSV in vitro and in vivo in a clinically relevant, immunocompetent mouse model of pancreatic ductal adenocarcinoma (PDA). Our system allows the study of virotherapy against PDA in the context of overexpression (80% PDA patients) or no expression of human mucin 1 (MUC1), a major marker for poor prognosis in patients. In vitro, we tested three VSV recombinants, wild-type VSV, VSV-GFP, and a safe oncolytic VSV-ΔM51-GFP, against five mouse PDA cell lines either expressing human MUC1 or MUC1-null. All viruses demonstrated significant oncolytic abilities independent of MUC1 expression, although VSV-ΔM51-GFP was somewhat less effective in two PDA cell lines. In vivo administration of VSV-ΔM51-GFP resulted in significant reduction of tumor growth for tested mouse PDA xenografts (+MUC1 or MUC1-null), and antitumor efficacy was further improved when the virus was combined with the chemotherapeutic drug gemcitabine. The antitumor effect was transient in all tested groups. The developed system can be used to study therapies involving various OVs and chemotherapeutics, with the goal of inducing tumor-specific immunity while preventing premature virus clearance.
[Show abstract][Hide abstract] ABSTRACT: Breast tumor cells alter their microenvironment in part through the expression of protumor molecules that influence macrophages during tumor progression and metastasis. Macrophage recruitment is stimulated by chemotactic factors, including tumor necrosis factor alpha (TNF-α), which also stimulates the cytotoxic/tumor cell killing macrophage phenotype. Through TNF-α converting enzyme (TACE/ADAM17) activities, breast tumor cells shed membrane-bound proteins, including their TNF receptors (sTNFR1/2), which serve as decoys sequestering TNF-α and preventing TNF-α-driven apoptosis of tumor cells, thereby decreasing TNF-α bioavailability. Here we investigated the levels of sTNFRs shed by breast tumor cells and determined the effects of shed sTNFRs on macrophage migration toward TNF-α. TNF-α and sTNFRs concentrations were measured in murine normal epithelial, stromal, and mammary tumor cells. The migration of murine macrophages towards TNF-α in the presence of tumor derived soluble factors (TDSFs) shed by TACE was determined. TNF-α concentrations secreted by tumor and normal epithelial cells were below the detection limit contrasting with stromal cells, especially macrophages, which expressed higher levels of TNF-α (P<0.001). Regardless of the cell tested, treatment with the TACE inhibitor TAPI-0 led to a significant decrease in sTNFR2 shed (P<0.05). The dose-dependent macrophage migration toward TNF-α prevented by incubation with TDSFs was not observed with TDSFs collected following TAPI-0 treatment (P<0.05). Furthermore, the TNF-α-driven increased pAkt expression in macrophage was inhibited by TACE shed TDSFs (P<0.05). These results highlight the role of tumor-shed sTNFRs in TNF-α -driven macrophage chemotaxis.
Journal of interferon & cytokine research: the official journal of the International Society for Interferon and Cytokine Research 06/2013; · 1.63 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: INTRODUCTION: Breast cancer remains the second leading cause of cancer related deaths for women in the United States. Metastasis is regulated not only by intrinsic genetic changes in malignant cells, but also by the microenvironment, especially those associated with chronic inflammation. We have recently reported that mice that suffer from autoimmune arthritis have significantly increased incidence of bone and lung metastasis and decreased survival associated with breast cancer. In this study, we evaluated the mechanism underlying the increased metastasis. Methods We used two mouse models; one that develops spontaneous autoimmune arthritis (SKG mice) injected with metastatic breast cancer cells (4T1), and second, that develops spontaneous breast cancer (MMTV-PyV MT mice) injected with type II collagen to induce autoimmune arthritis. Mast cell levels and metastasis were monitored. RESULTS: First, we confirm that breast tumor-bearing arthritic mice have significantly higher incidence of bone and lung metastasis than their non-arthritic counterparts. Next, we show increased recruitment of mast cells within the primary tumor of arthritic mice which facilitate metastasis. Next, we report that arthritic mice without any tumors have higher numbers of mast cells in the bones and lungs which may be the underlying cause for the enhanced lung and bone metastasis observed in the arthritic mice. Next, we show that once the tumor cells populate the metastatic niches (bones and lungs), they further increase the mast cell population within the niche and assist in enhancing metastasis. This may primarily be due to the interaction of c-kit receptor present on mast cells and stem cell factor (SCF, the ligand for ckit) expressed on tumor cells. Finally, we show that targeting the SCF/cKit interaction with an anti-ckit antibody reduces the differentiation of mast cells and consequently reduces metastasis. CONCLUSION: This is the first report to show that mast cells may play a critical role in not only remodeling the tumor microenvironment but also the metastatic niche to facilitate efficient metastasis through SCF/cKit interaction in breast cancer with arthritis.
Breast cancer research: BCR 04/2013; 15(2):R32. · 5.87 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: MUC1 (CD227), a membrane tethered mucin glycoprotein, is overexpressed in >60% of human pancreatic cancers (PCs), and is associated with poor prognosis, enhanced metastasis and chemoresistance. The objective of this study was to delineate the mechanism by which MUC1 induces drug resistance in human (BxPC3 and Capan-1) and mouse (KCKO, KCM) PC cells. We report that PC cells that express high levels of MUC1 exhibit increased resistance to chemotherapeutic drugs (gemcitabine and etoposide) in comparison with cells that express low levels of MUC1. This chemo resistance was attributed to the enhanced expression of multidrug resistance (MDR) genes including ABCC1, ABCC3, ABCC5 and ABCB1. In particular, levels of MRP1 protein encoded by the ABCC1 gene were significantly higher in the MUC1-high PC cells. In BxPC3 and Capan-1 cells MUC1 upregulates MRP1 via an Akt-dependent pathway, whereas in KCM cells MUC1-mediated MRP1 upregulation is via an Akt-independent mechanism. In KCM, BxPC3 and Capan-1 cells, the cytoplasmic tail motif of MUC1 associates directly with the promoter region of the Abcc1/ABCC1 gene, indicating a possible role of MUC1 acting as a transcriptional regulator of this gene. This is the first report to show that MUC1 can directly regulate the expression of MDR genes in PC cells, and thus confer drug resistance.
[Show abstract][Hide abstract] ABSTRACT: Oncolytic virus (OV) therapy takes advantage of common cancer characteristics, such as defective type I interferon (IFN) signaling, to preferentially infect and kill cancer cells with viruses. Our recent study (Murphy et al., 2012. J. Virol. 86, 3073-87) found human pancreatic ductal adenocarcinoma (PDA) cells were highly heterogeneous in their permissiveness to vesicular stomatitis virus (VSV) and suggested at least some resistant cell lines retained functional type I IFN responses. Here we examine cellular responses to infection by the oncolytic VSV recombinant VSV-ΔM51-GFP by analyzing a panel of 11 human PDA cell lines for expression of 33 genes associated with type I IFN pathways. Although all cell lines sensed infection by VSV-ΔM51-GFP and most activated IFN-α and β expression, only resistant cell lines displayed constitutive high-level expression of the IFN-stimulated antiviral genes MxA and OAS. Inhibition of JAK/STAT signaling decreased levels of MxA and OAS and increased VSV infection, replication and oncolysis, further implicating IFN responses in resistance. Unlike VSV, vaccinia and herpes simplex virus infectivity and killing of PDA cells was independent of the type I IFN signaling profile, possibly because these two viruses are better equipped to evade type I IFN responses. Our study demonstrates heterogeneity in the type I IFN signaling status of PDA cells and suggests MxA and OAS as potential biomarkers for PDA resistance to VSV and other OVs sensitive to type I IFN responses.
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Controversy exists as to the ability of human gammaherpesviruses to cause or exacerbate breast cancer disease in patients. The difficulty in conducting definitive human studies can be overcome by investigating developing breast cancer in a mouse model. In this study, we utilized mice latently infected with murine gammaherpesvirus 68 (HV-68) to question whether such a viral burden could exacerbate metastatic breast cancer disease using a mouse mammary tumor model. RESULTS: Mice latently infected with HV-68 had a similar primary tumor burden, but much greater metastatic disease, when compared to mock treated mice given the transplantable tumor, 4T1. This was true for lung lesions, as well as secondary tumor masses. Increased expression of pan-cytokeratin and VEGF-A in tumors from HV-68 infected mice was consistent with increased metastatic disease in these animals. Surprisingly, no viral particles could be cultured from tumor tissues, and the presence of viral DNA or RNA transcripts could not be detected in primary or secondary tumor tissues. CONCLUSIONS: Latent HV-68 infection had no significant effect on the size of primary 4T1 mammary tumors, but exacerbated the number of metastatic lung lesions and secondary tumors when compared to mock treated mice. Increased expression of the tumor marker, pan-cytokeratin, and VEGF-A in tumors of mice harboring latent virus was consistent with an exacerbated metastatic disease. Mechanisms responsible for this exacerbation are indirect, since no virus could be detected in cancerous tissues.
[Show abstract][Hide abstract] ABSTRACT: Monoclonal antibodies (mAbs) against tumor-associated antigens are useful anticancer agents. Antibody-dependent cellular cytotoxicity (ADCC) is one of the major mechanisms responsible for initiating natural killer cell (NK)-mediated killing of tumors. However, the regulation of ADCC via NK cells is poorly understood. We have investigated the cytolytic activity of NK cells against pancreatic cancer cells that were coated with an antibody directed against the human tumor antigen, Mucin-1 designated HMFG-2, either alone or conjugated to CpG oligodeoxynucleotide (CpG ODN). Conjugated antibodies were tested for their ability to elicit ADCC in vitro and in vivo against pancreatic cancer cells. NK cells cultured in the presence of immobilized CpG ODN, HMFG-2 Ab, or CpG ODN-conjugated HMFG-2 Ab were able to up-regulate perforin similarly. Interestingly, a significant higher ADCC was observed when CpG ODN-conjugated HMFG-2-coated tumor cells were co-cultured with NK cells compared to unconjugated HMFG-2 Ab or CpG ODN alone. Moreover, MyD88-deficient NK cells can perform ADCC in vitro. Furthermore, intratumoral injections of CpG ODN-conjugated HMFG-2 induced a significant reduction in tumor burden in vivo in an established model of pancreatic tumor in nude mice compared to CpG ODN or the HMFG-2 alone. Depletion of macrophages or NK cells before treatment confirmed that both cells were required for the anti-tumor response in vivo. Results also suggest that CpG ODN and HMFG-2 Ab could be sensed by NK cells on the mAb-coated tumor cells triggering enhanced ADCC in vitro and in vivo.
Cancer Immunology and Immunotherapy 04/2012; · 3.64 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Vesicular stomatitis virus (VSV) is a promising oncolytic agent against a variety of cancers. However, it has never been tested in any pancreatic cancer model. Pancreatic ductal adenocarcinoma (PDA) is the most common and aggressive form of pancreatic cancer. In this study, the oncolytic potentials of several VSV variants were analyzed in a panel of 13 clinically relevant human PDA cell lines and compared to conditionally replicative adenoviruses (CRAds), Sendai virus and respiratory syncytial virus. VSV variants showed oncolytic abilities superior to those of other viruses, and some cell lines that exhibited resistance to other viruses were successfully killed by VSV. However, PDA cells were highly heterogeneous in their susceptibility to virus-induced oncolysis, and several cell lines were resistant to all tested viruses. Resistant cells showed low levels of very early VSV RNA synthesis, indicating possible defects at initial stages of infection. In addition, unlike permissive PDA cell lines, most of the resistant cell lines were able to both produce and respond to interferon, suggesting that intact type I interferon responses contributed to their resistance phenotype. Four cell lines that varied in their permissiveness to VSV-ΔM51 and CRAd dl1520 were tested in mice, and the in vivo results closely mimicked those in vitro. While our results demonstrate that VSV is a promising oncolytic agent against PDA, further studies are needed to better understand the molecular mechanisms of resistance of some PDAs to oncolytic virotherapy.
Journal of Virology 03/2012; 86(6):3073-87. · 5.08 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pancreatic ductal adenocarcinoma (PDA) has one of the worst prognoses of all cancers. Mucin 1 (MUC1), a transmembrane mucin glycoprotein, is a key modulator of several signaling pathways that affect oncogenesis, motility and metastasis. Its expression is known to be associated with poor prognosis in patients. However, the precise mechanism remains elusive. We report a novel association of MUC1 with platelet-derived growth factor-A (PDGFA). PDGFA is one of the many drivers of tumor growth, angiogenesis and metastasis in PDA. Using mouse PDA models as well as human samples, we show clear evidence that MUC1 regulates the expression and secretion of PDGFA. This, in turn, influences proliferation and invasion of pancreatic cancer cells leading to higher tumor burden in vivo. In addition, we reveal that MUC1 overexpressing cells are heavily dependent on PDGFA both for proliferation and invasion, whereas MUC1-null cells are not. Moreover, PDGFA and MUC1 are critical for translocation of β catenin to the nucleus for oncogenesis to ensue. Finally, we elucidate the underlying mechanism by which MUC1 regulates PDGFA expression and secretion in pancreatic cancer cells. We show that MUC1 associates with Hif1-α, a known transcription factor involved in controlling PDGFA expression. Furthermore, MUC1 facilitates Hif1-α translocation to the nucleus. In summary, we have demonstrated that MUC1-induced invasion and proliferation occurs via increased exogenous production of PDGFA. Thus, impeding MUC1 regulation of PDGFA signaling may be therapeutically beneficial for patients with PDA.Oncogene advance online publication, 23 January 2012; doi:10.1038/onc.2011.651.
[Show abstract][Hide abstract] ABSTRACT: Several studies have demonstrated that sites of chronic inflammation are often associated with the establishment and growth of various malignancies. A common inflammatory condition in humans is autoimmune arthritis (AA). Although AA and cancer are different diseases, many of the underlying processes that contribute to the disorders of the joints and connective tissue that characterize AA also affect cancer progression and metastasis. Systemically, AA can lead to cellular infiltration and inflammation of the lungs. Several studies have reported statistically significant risk ratios between AA and breast cancer. Despite this knowledge being available, there has been minimal research linking breast cancer, arthritis, and metastasis associated with breast cancer. Notably both diseases are extremely prevalent in older post-menopausal women.
To establish the novel link between arthritis induced inflammation and secondary metastasis associated with breast cancer, PyV MT mice that spontaneously develop mammary gland carcinoma were injected with Type II collagen (CII) to induce arthritis at 9 and 18 weeks of age for pre-metastatic and metastatic condition. The sites of secondary metastasis and the associated inflammatory microenvironment were evaluated.
A significant increase in breast cancer-associated secondary metastasis to the lungs and bones was observed in the arthritic versus the non-arthritic PyV MT mice along with an increase in primary tumor burden. We report significant increases in the levels of interstitial cellular infiltrates and pro-inflammatory cytokines such as interleukin-17 (IL-17), interleukin-6 (IL-6), Pro- Matrix metallopeptidase 9 (Pro-MMP9), insulin like growth factor-II (GF-II) and macrophage colony stimulating factor (M-CSF) in the arthritic lung and bone milieu as well as in the circulation. These pro-inflammatory cytokines along with the inflammatory microenvironment may be the underlying factors facilitating tumor progression and metastasis in arthritic PyV MT mice. This was further substantiated by treatment with celecoxib, an anti-inflammatory drug + αIL-17 antibody that significantly reduced the secondary metastasis to lung and bone.
The data generated not only reveal the underlying mechanism of high susceptibility to bone and lung metastasis in an arthritic condition but our combination therapies may lead to treatment modalities that will be capable of reducing tumor burden, and preventing relapse and metastasis in arthritic patients with breast cancer.
[Show abstract][Hide abstract] ABSTRACT: The human epithelial mucin MUC1 is a heavily glycosylated transmembrane protein that is overexpressed and aberrantly glycosylated on over 90% of human breast cancers. The altered glycosylation of MUC1 reveals an immunodominant peptide along its tandem repeat (TR) that has been used as a target for tumour immunotherapy. In this study, we used the MUC1 TR peptide as a test antigen to determine whether a plant-expressed human tumour-associated antigen can be successfully expressed in a plant system and whether it will be able to break self-antigen tolerance in a MUC1-tolerant mouse model. We report the expression of MUC1 TR peptide fused to the mucosal-targeting Escherichia coli enterotoxin B subunit (LTB-MUC1) in a plant host. Utilizing a rapid viral replicon transient expression system, we obtained high yields of LTB-MUC1. Importantly, the LTB-MUC1 fusion protein displayed post-translational modifications that affected its antigenicity. Glycan analysis revealed that LTB-MUC1 was glycosylated and a MUC1-specific monoclonal antibody detected only the glycosylated forms. A thorough saccharide analysis revealed that the glycans are tri-arabinans linked to hydroxyprolines within the MUC1 tandem repeat sequence. We immunized MUC1-tolerant mice (MUC1.Tg) with transiently expressed LTB-MUC1, and observed production of anti-MUC1 serum antibodies, indicating breach of tolerance. The results indicate that a plant-derived human tumour-associated antigen is equivalent to the human antigen in the context of immune recognition.
[Show abstract][Hide abstract] ABSTRACT: MUC1 is overexpressed and aberrantly glycosylated in more than 60% of pancreatic ductal adenocarcinomas. The functional role of MUC1 in pancreatic cancer has yet to be fully elucidated due to a dearth of appropriate models. In this study, we have generated mouse models that spontaneously develop pancreatic ductal adenocarcinoma (KC), which are either Muc1-null (KCKO) or express human MUC1 (KCM). We show that KCKO mice have significantly slower tumor progression and rates of secondary metastasis, compared with both KC and KCM. Cell lines derived from KCKO tumors have significantly less tumorigenic capacity compared with cells from KCM tumors. Therefore, mice with KCKO tumors had a significant survival benefit compared with mice with KCM tumors. In vitro, KCKO cells have reduced proliferation and invasion and failed to respond to epidermal growth factor, platelet-derived growth factor, or matrix metalloproteinase 9. Further, significantly less KCKO cells entered the G(2)-M phase of the cell cycle compared with the KCM cells. Proteomics and Western blotting analysis revealed a complete loss of cdc-25c expression, phosphorylation of mitogen-activated protein kinase (MAPK), as well as a significant decrease in nestin and tubulin-α2 chain expression in KCKO cells. Treatment with a MEK1/2 inhibitor, U0126, abrogated the enhanced proliferation of the KCM cells but had minimal effect on KCKO cells, suggesting that MUC1 is necessary for MAPK activity and oncogenic signaling. This is the first study to utilize a Muc1-null PDA mouse to fully elucidate the oncogenic role of MUC1, both in vivo and in vitro.
Cancer Research 05/2011; 71(13):4432-42. · 9.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Increased motility and invasiveness of pancreatic cancer cells are associated with epithelial to mesenchymal transition (EMT). Snai1 and Slug are zinc-finger transcription factors that trigger this process by repressing E-cadherin and enhancing vimentin and N-cadherin protein expression. However, the mechanisms that regulate this activation in pancreatic tumors remain elusive. MUC1, a transmembrane mucin glycoprotein, is associated with the most invasive forms of pancreatic ductal adenocarcinomas (PDA). In this study, we show that over expression of MUC1 in pancreatic cancer cells triggers the molecular process of EMT, which translates to increased invasiveness and metastasis. EMT was significantly reduced when MUC1 was genetically deleted in a mouse model of PDA or when all seven tyrosines in the cytoplasmic tail of MUC1 were mutated to phenylalanine (mutated MUC1 CT). Using proteomics, RT-PCR and western blotting, we revealed a significant increase in vimentin, Slug and Snail expression with repression of E-Cadherin in MUC1-expressing cells compared with cells expressing the mutated MUC1 CT. In the cells that carried the mutated MUC1 CT, MUC1 failed to co-immunoprecipitate with beta-catenin and translocate to the nucleus, thereby blocking transcription of the genes associated with EMT and metastasis. Thus, functional tyrosines are critical in stimulating the interactions between MUC1 and beta-catenin and their nuclear translocation to initiate the process of EMT. This study signifies the oncogenic role of MUC1 CT and is the first to identify a direct role of the MUC1 in initiating EMT during pancreatic cancer. The data may have implications in future design of MUC1-targeted therapies for pancreatic cancer
[Show abstract][Hide abstract] ABSTRACT: Since epithelial mucin 1 (MUC1) is associated with several adenocarcinomas at the mucosal sites, it is pertinent to test the efficacy of a mucosally targeted vaccine formulation. The B subunit of the Vibrio cholerae cholera toxin (CTB) has great potential to act as a mucosal carrier for subunit vaccines. In the present study we evaluated whether a MUC1 tandem repeat (TR) peptide chemically linked to CTB would break self-antigen tolerance in the transgenic MUC1-tolerant mouse model (MUC1.Tg) through oral or parenteral immunizations. We report that oral immunization with the CTB-MUC1 conjugate along with mucosal adjuvant, unmethylated CpG oligodeoxynucleotide (ODN) and interleukin-12 (IL-12) did not break self-antigen tolerance in MUC1.Tg mice, but induced a strong humoral response in wild-type C57BL/6 mice. However, self-antigen tolerance in the MUC1.Tg mouse model was broken after parenteral immunizations with different doses of the CTB-MUC1 conjugate protein and with the adjuvant CpG ODN co-delivered with CTB-MUC1. Importantly, mice immunized systemically with CpG ODN alone and with CTB-MUC1 exhibited decreased tumor burden when challenged with a mammary gland tumor cell line that expresses human MUC1.
Cancer Immunology and Immunotherapy 12/2010; 59(12):1801-11. · 3.64 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Increased motility and invasiveness of pancreatic cancer cells are associated with epithelial to mesenchymal transition (EMT). Snai1 and Slug are zinc-finger transcription factors that trigger this process by repressing E-cadherin and enhancing vimentin and N-cadherin protein expression. However, the mechanisms that regulate this activation in pancreatic tumors remain elusive. MUC1, a transmembrane mucin glycoprotein, is associated with the most invasive forms of pancreatic ductal adenocarcinomas (PDA). In this study, we show that over expression of MUC1 in pancreatic cancer cells triggers the molecular process of EMT, which translates to increased invasiveness and metastasis. EMT was significantly reduced when MUC1 was genetically deleted in a mouse model of PDA or when all seven tyrosines in the cytoplasmic tail of MUC1 were mutated to phenylalanine (mutated MUC1 CT). Using proteomics, RT-PCR and western blotting, we revealed a significant increase in vimentin, Slug and Snail expression with repression of E-Cadherin in MUC1-expressing cells compared with cells expressing the mutated MUC1 CT. In the cells that carried the mutated MUC1 CT, MUC1 failed to co-immunoprecipitate with β-catenin and translocate to the nucleus, thereby blocking transcription of the genes associated with EMT and metastasis. Thus, functional tyrosines are critical in stimulating the interactions between MUC1 and β-catenin and their nuclear translocation to initiate the process of EMT. This study signifies the oncogenic role of MUC1 CT and is the first to identify a direct role of the MUC1 in initiating EMT during pancreatic cancer. The data may have implications in future design of MUC1-targeted therapies for pancreatic cancer.
[Show abstract][Hide abstract] ABSTRACT: To study immunology in breast tumors, we have utilized a mammary gland adenocarcinoma model in which mice develop spontaneous tumors of the mammary gland which are initiated at puberty and express a human tumor antigen, MUC1. MUC1 (CD227) is over-expressed in 90% of human breast cancers and its glycosylation status and pattern of expression in cancer cells is altered. Humoral and cellular responses to MUC1 have been reported in breast cancer patients and therefore, MUC1 is being evaluated as a target for immune intervention. This mouse model of spontaneous breast cancer allows the evaluation of anti-MUC1 immune responses at all stages of the disease. In this report, we review the model as it pertains to a) the development of the tumor, b) MUC1 expression, and the native immune responses against MUC1 as tumors progress, and c) the immune suppressive microenvironment within the developing tumor. Finally, we report our latest findings describing the therapeutic efficacy of adoptively transferred MUC1-specific cytotoxic T lymphocytes (MUC1-CTL) in these mice and discuss ways to increase their effectiveness by agonistic monoclonal antibody against CD137 T cell costimulatory molecule.