Anna Chytil

Vanderbilt University, Нашвилл, Michigan, United States

Are you Anna Chytil?

Claim your profile

Publications (65)474.75 Total impact

  • Cancer Research 08/2015; 75(15 Supplement):4083-4083. DOI:10.1158/1538-7445.AM2015-4083 · 9.33 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Bone Morphogenetic Proteins (BMPs) are secreted cytokines/growth factors belonging to the Transforming Growth Factor β (TGFβ) family. BMP ligands have been shown to be overexpressed in human breast cancers. Normal and cancerous breast tissue display active BMP signaling as indicated by phosphorylated Smads 1, 5 and 9. We combined mice expressing the MMTV.PyMT oncogene with mice having conditional knockout (cKO) of BMP receptor type 1a (BMPR1a) using whey acidic protein (WAP)-Cre and found this deletion resulted in delayed tumor onset and significantly extended survival. Immunofluorescence staining revealed that cKO tumors co-expressed Keratin 5 and mesenchymal cell markers such as Vimentin. This indicates that epithelial-to-mesenchymal (EMT)-like transitions occurred in cKO tumors. We performed microarray analysis on these tumors and found changes that support EMT-like changes. We established primary tumor cell lines and found that BMPR1a cKO had slower growth in vitro and in vivo upon implantation. cKO tumor cells had reduced migration in vitro. We analyzed human databases from TCGA and survival data from microarrays to confirm BMPR1a tumor promoting functions, and found that high BMPR1a gene expression correlates with decreased survival regardless of molecular breast cancer subtype. In conclusion, the data indicate that BMP signaling through BMPR1a functions as a tumor promoter.
    Oncotarget 06/2015; 6(26). DOI:10.18632/oncotarget.4413 · 6.36 Impact Factor

  • Cancer Research 05/2015; 75(9 Supplement):P6-03-04-P6-03-04. DOI:10.1158/1538-7445.SABCS14-P6-03-04 · 9.33 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Metastasis is the most devastating aspect of cancer, however we know very little about the mechanisms of local invasion, the earliest step of metastasis. During tumor growth CD11b+Gr1+ cells, known also as MDSCs, have been shown to promote tumor progression by a wide spectrum of effects that suppress the anti-tumor immune response. In addition to immunosuppression, CD11b+Gr1+ cells promote metastasis by mechanisms that are currently unknown. CD11b+Gr1+ cells localize near fibroblasts, which remodel the ECM and leave tracks for collective cell migration of carcinoma cells. In this study we discovered that CD11b+Gr1+ cells promote invasion of mammary carcinoma cells by increasing fibroblast migration. This effect was directed by secreted factors derived from CD11b+Gr1+ cells. We have identified several CD11b+Gr1+ cell secreted proteins that activate fibroblast migration, including CXCL11, CXCL15, FGF2, IGF-I, IL1Ra, Resistin, and Shh. The combination of CXCL11 and FGF2 had the strongest effect on fibroblast migration that is associated with Akt1 and ERK1/2 phosphorylation. Analysis of subsets of CD11b+Gr1+ cells identified that CD11b+Ly6ChighLy6Glow cells increase fibroblast migration more than other myeloid cell populations. Additionally, tumor-derived CD11b+Gr1+ cells promote fibroblast migration more than splenic CD11b+Gr1+ cells of tumor-bearing mice. While TGFβ signaling in fibroblasts does not regulate their migration toward CD11b+Gr1+ cells, however deletion of TGFβ receptor II on CD11b+Gr1+ cells downregulates CXCL11, Shh, IGF1 and FGF2 resulting in reduced fibroblast migration. These studies show that TGFβ signaling in CD11b+Gr1+ cells promotes fibroblast directed carcinoma invasion and suggests that perivascular CD11b+Ly6ChighLy6Glow cells may be the stimulus for localized invasion leading to metastasis.
    PLoS ONE 01/2015; 10(1):e0117908. DOI:10.1371/journal.pone.0117908 · 3.23 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Postnatal mammary gland development and differentiation occur during puberty and pregnancy. To explore the role of DNA methylation in these processes, we determined the genome-wide DNA methylation and gene expression profiles of CD24(+)CD61(+)CD29(hi), CD24(+)CD61(+)CD29(lo), and CD24(+)CD61(-)CD29(lo) cell populations that were previously associated with distinct biological properties at different ages and reproductive stages. We found that pregnancy had the most significant effects on CD24(+)CD61(+)CD29(hi) and CD24(+)CD61(+)CD29(lo) cells, inducing distinct epigenetic states that were maintained through life. Integrated analysis of gene expression, DNA methylation, and histone modification profiles revealed cell-type- and reproductive-stage-specific changes. We identified p27 and TGFβ signaling as key regulators of CD24(+)CD61(+)CD29(lo) cell proliferation, based on their expression patterns and results from mammary gland explant cultures. Our results suggest that relatively minor changes in DNA methylation occur during luminal differentiation compared with the effects of pregnancy on CD24(+)CD61(+)CD29(hi) and CD24(+)CD61(+)CD29(lo) cells. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
    Stem Cell Reports 01/2015; 4(2). DOI:10.1016/j.stemcr.2014.12.009 · 5.37 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: IntroductionTransforming growth factor beta (TGFß) plays a major role in the regulation of tumor initiation, progression, and metastasis. It is depended on the type II TGFß receptor (TßRII) for signaling. Previously, we have shown that deletion of TßRII in mammary epithelial of MMTV-PyMT mice results in shortened tumor latency and increased lung metastases. In MMTV-Neu mouse model, which is related to human epidermal growth factor 2 (HER2)¿+¿human breast cancer, active TGFß signaling increased the number of circulating tumor cells and metastases. Therefore, the loss-of-function experiments suppressed metastasis in Neu-induced tumors. In the current study, we describe a newly discovered connection between attenuated TGFß signaling and HER2 signaling in mammary tumor progression.Methods All studies were performed on MMTV-Neu mice with and without dominant-negative TßRII (DNIIR) in mammary epithelium. Mammary tumors were analyzed by flow cytometry, immunohistochemistry, and immunofluorescence staining. The levels of secreted proteins were measured by enzyme-linked immunosorbent assay. Whole-lung mount staining was used to quantitate lung metastasis. The Cancer Genome Atlas (TCGA) datasets were used to determine the relevance of our findings to human breast cancer.ResultsAttenuated TGFß signaling led to a delay of HER2+ tumor onset, but increased the number of metastases. The DNIIR tumors were characterized by increased vasculogenesis, vessel leakage, and increased expression of vascular endothelial growth factor (VEGF). During DNIIR tumor progression, both the levels of CXCL1/5 and the number of CD11b+Gr1+ cells and T cells decreased. Analysis of TCGA datasets demonstrated a significant negative correlation between TGFBR2 and VEGF genes expression. Higher VEGFA expression correlated with shorter distant metastasis-free survival only in HER2+ patients with no differences in HER2-, estrogen receptor +/- or progesterone receptor +/- breast cancer patients.Conclusion Our studies provide insights into a novel mechanism by which epithelial TGFß signaling modulates the tumor microenvironment, and by which it is involved in lung metastasis in HER2+ breast cancer patients. The effects of pharmacological targeting of the TGFß pathway in vivo during tumor progression remain controversial. The targeting of TGFß signaling should be a viable option, but because VEGF has a protumorigenic effect on HER2+ tumors, the targeting of this protein could be considered when it is associated with attenuated TGFß signaling.
    Breast cancer research: BCR 10/2014; 16(5):425. DOI:10.1186/s13058-014-0425-7 · 5.49 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: There is growing evidence that generation of adenosine from ATP, which is mediated by the CD39/CD73 enzyme pair, predetermines immunosuppressive and proangiogenic properties of myeloid cells. We have previously shown that the deletion of the TGF-β type II receptor gene (Tgfbr2) expression in myeloid cells is associated with decreased tumor growth, suggesting protumorigenic effect of TGF-β signaling. In this study, we tested the hypothesis that TGF-β drives differentiation of myeloid-derived suppressor cells into protumorigenic terminally differentiated myeloid mononuclear cells (TDMMCs) characterized by high levels of cell-surface CD39/CD73 expression. We found that TDMMCs represent a major cell subpopulation expressing high levels of both CD39 and CD73 in the tumor microenvironment. In tumors isolated from mice with spontaneous tumor formation of mammary gland and conditional deletion of the type II TGF-β receptor in mammary epithelium, an increased level of TGF-β protein was associated with further increase in number of CD39(+)CD73(+) TDMMCs compared with MMTV-PyMT/TGFβRII(WT) control tumors with intact TGF-β signaling. Using genetic and pharmacological approaches, we demonstrated that the TGF-β signaling mediates maturation of myeloid-derived suppressor cells into TDMMCs with high levels of cell surface CD39/CD73 expression and adenosine-generating capacity. Disruption of TGF-β signaling in myeloid cells resulted in decreased accumulation of TDMMCs, expressing CD39 and CD73, and was accompanied by increased infiltration of T lymphocytes, reduced density of blood vessels, and diminished progression of both Lewis lung carcinoma and spontaneous mammary carcinomas. We propose that TGF-β signaling can directly induce the generation of CD39(+)CD73(+) TDMMCs, thus contributing to the immunosuppressive, proangiogenic, and tumor-promoting effects of this pleiotropic effector in the tumor microenvironment.
    The Journal of Immunology 08/2014; 193(6). DOI:10.4049/jimmunol.1400578 · 4.92 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Bone Morphogenetic Protein (BMP) receptors mediate a diverse range of signals to regulate both development and disease. BMP activity has been linked to both tumor promoting and suppressive functions in both tumor cells and their surrounding microenvironment. We sought to investigate the requirement for BMPR2 in stromal fibroblasts during mammary tumor formation and metastasis. We utilized FSP1 (Fibroblast Specific Protein-1) promoter driven Cre to genetically delete BMPR2 in mice expressing the MMTV.PyVmT mammary carcinoma oncogene. We found that abrogation of stromal BMPR2 expression via FSP1 driven Cre resulted in increased tumor metastasis. Additionally, similar to epithelial BMPR2 abrogation, stromal loss of BMPR2 results in increased inflammatory cell infiltration. We proceeded to isolate and establish fibroblast cell lines without BMPR2 and found a cell autonomous increase in inflammatory cytokine secretion. Fibroblasts were co-implanted with syngeneic tumor cells and resulted in accelerated tumor growth and increased metastasis when fibroblasts lacked BMPR2. We observed that the loss of BMPR2 results in increased chemokine expression, which facilitates inflammation by a sustained increase in myeloid cells. The chemokines increased in BMPR2 deleted cells correlated with poor outcome in human breast cancer patients. We conclude that BMPR2 has tumor suppressive functions in the stroma by regulating inflammation.
    Molecular Oncology 08/2014; 9(1). DOI:10.1016/j.molonc.2014.08.004 · 5.33 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Introduction There is a major need to better understand the molecular basis of triple negative breast cancer (TNBC) in order to develop effective therapeutic strategies. Using gene expression data from 587 TNBC patients we previously identified six subtypes of the disease, among which a mesenchymal-stem like (MSL) subtype. The MSL subtype has significantly higher expression of the transforming growth factor beta (TGF-β) pathway-associated genes relative to other subtypes, including the TGF-β receptor type III (TβRIII). We hypothesize that TβRIII is tumor promoter in mesenchymal-stem like TNBC cells. Methods Representative MSL cell lines SUM159, MDA-MB-231 and MDA-MB-157 were used to study the roles of TβRIII in the MSL subtype. We stably expressed short hairpin RNAs specific to TβRIII (TβRIII-KD). These cells were then used for xenograft tumor studies in vivo; and migration, invasion, proliferation and three dimensional culture studies in vitro. Furthermore, we utilized human gene expression datasets to examine TβRIII expression patterns across all TNBC subtypes. Results TβRIII was the most differentially expressed TGF-β signaling gene in the MSL subtype. Silencing TβRIII expression in MSL cell lines significantly decreased cell motility and invasion. In addition, when TβRIII-KD cells were grown in a three dimensional (3D) culture system or nude mice, there was a loss of invasive protrusions and a significant decrease in xenograft tumor growth, respectively. In pursuit of the mechanistic underpinnings for the observed TβRIII-dependent phenotypes, we discovered that integrin-α2 was expressed at higher level in MSL cells after TβRIII-KD. Stable knockdown of integrin-α2 in TβRIII-KD MSL cells rescued the ability of the MSL cells to migrate and invade at the same level as MSL control cells. Conclusions We have found that TβRIII is required for migration and invasion in vitro and xenograft growth in vivo. We also show that TβRIII-KD elevates expression of integrin-α2, which is required for the reduced migration and invasion, as determined by siRNA knockdown studies of both TβRIII and integrin-α2. Overall, our results indicate a potential mechanism in which TβRIII modulates integrin-α2 expression to effect MSL cell migration, invasion, and tumorigenicity.
    Breast cancer research: BCR 07/2014; 16(4):R69. DOI:10.1186/bcr3684 · 5.49 Impact Factor

  • Cancer Research 05/2014; 73(3 Supplement):A16-A16. DOI:10.1158/1538-7445.TIM2013-A16 · 9.33 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Therapeutic interventions that target critical elements of the tumor stromal microenvironment may improve blockades to metastatic progression. Here we report characterization of a mechanism that address this opportunity, based on studies of the polyoma middle T-induced mouse model of mammary carcinoma that exhibits aggressive metastases due to lack of the type II TGF-ß receptor (PyMTmgko mice). We hypothesized that PyMTmgko-activated stroma interacts with carcinoma cells to promote invasion and metastasis. Extracellular matrix associated with PyMTmgko tumors was found to exhibit increased stiffness, fibrillar collagen levels and expression of the collagen crosslinking enzyme lysyl oxidase (LOX), compared to polyoma middle T-induced mammary carcinomas in mice expressing the type II TGF-ß receptor (PyMTfl/fl). Inhibiting LOX activity in PyMTmgko mice had no effect on tumor latency or size, but decreased tumor metastasis by inhibiting tumor cell intravasation. This phenotype was associated with a decrease in keratin 14-positive myoepithelial cells in PyMTmgko tumors following LOX inhibition along with a decrease in focal adhesion formation. Interestingly, the primary source of LOX was determined to be activated fibroblasts. LOX expression was driven at this site by myeloid cell-derived TGF-ß, which is linked to human breast cancer. Our results suggested that stromal expansion in PyMTmgko tumors was elicited by immune cell infiltrates that promoted fibroblast activation, leading to metastatic behavior by modulating phenotypic characteristics of basal epithelial cells. More broadly, these findings argue that microenvironmental changes triggered by epithelial cells can drive tumor pathogenicity and offer important therapeutic targets to inhibit metastasis.
    Cancer Research 07/2013; 73(17). DOI:10.1158/0008-5472.CAN-13-0012 · 9.33 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Introduction Transforming growth factor beta (TGF-β) has a dual role during tumor progression, initially as a suppressor and then as a promoter. Epithelial TGF-β signaling regulates fibroblast recruitment and activation. Concurrently, TGF-β signaling in stromal fibroblasts suppresses tumorigenesis in adjacent epithelia, while its ablation potentiates tumor formation. Much is known about the contribution of TGF-β signaling to tumorigenesis, yet the role of TGF-β in epithelial-stromal migration during tumor progression is poorly understood. We hypothesize that TGF-β is a critical regulator of tumor-stromal interactions that promote mammary tumor cell migration and invasion. Methods Fluorescently labeled murine mammary carcinoma cells, isolated from either MMTV-PyVmT transforming growth factor-beta receptor II knockout (TβRII KO) or TβRIIfl/fl control mice, were combined with mammary fibroblasts and xenografted onto the chicken embryo chorioallantoic membrane. These combinatorial xenografts were used as a model to study epithelial-stromal crosstalk. Intravital imaging of migration was monitored ex ovo, and metastasis was investigated in ovo. Epithelial RNA from in ovo tumors was isolated by laser capture microdissection and analyzed to identify gene expression changes in response to TGF-β signaling loss. Results Intravital microscopy of xenografts revealed that mammary fibroblasts promoted two migratory phenotypes dependent on epithelial TGF-β signaling: single cell/strand migration or collective migration. At epithelial-stromal boundaries, single cell/strand migration of TβRIIfl/fl carcinoma cells was characterized by expression of α-smooth muscle actin and vimentin, while collective migration of TβRII KO carcinoma cells was identified by E-cadherin+/p120+/β-catenin+ clusters. TβRII KO tumors also exhibited a twofold greater metastasis than TβRIIfl/fl tumors, attributed to enhanced extravasation ability. In TβRII KO tumor epithelium compared with TβRIIfl/fl epithelium, Igfbp4 and Tspan13 expression was upregulated while Col1α2, Bmp7, Gng11, Vcan, Tmeff1, and Dsc2 expression was downregulated. Immunoblotting and quantitative PCR analyses on cultured cells validated these targets and correlated Tmeff1 expression with disease progression of TGF-β-insensitive mammary cancer. Conclusion Fibroblast-stimulated carcinoma cells utilize TGF-β signaling to drive single cell/strand migration but migrate collectively in the absence of TGF-β signaling. These migration patterns involve the signaling regulation of several epithelial-to-mesenchymal transition pathways. Our findings concerning TGF-β signaling in epithelial-stromal interactions are important in identifying migratory mechanisms that can be targeted as recourse for breast cancer treatment.
    Breast cancer research: BCR 07/2012; 14(4):R98. DOI:10.1186/bcr3217 · 5.49 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: By crossing LysM-Cre and TGF-β type II receptor (Tgfbr2) floxed mice we achieved specific deletion of Tgfbr2 in myeloid cells (Tgfbr2(MyeKO) mice). S.c.-injected (LLC, EL4-OVA) and implanted (MMTV-PyMT) carcinoma cells grow slower in Tgfbr2(MyeKO) mice. The number of CD45(+) cells in the tumor tissue was the same in both genotypes of mice, but upon analysis, the percentage of T cells (CD45(+)CD3(+)) in the KO mice was increased. By flow cytometry analysis, we did not detect any differences in the number and phenotype of TAMs, CD11b(+)Gr1(+), and DCs in Tgfbr2(MyeKO) compared with Tgfbr2(MyeWT) mice. ELISA and qRT-PCR data showed differences in myeloid cell functions. In Tgfbr2(MyeKO) TAMs, TNF-α secretion was increased, basal IL-6 secretion was down-regulated, TGF-β did not induce any VEGF response, and there was decreased MMP9 and increased MMP2 and iNOS expression. TGF-β did not have any effect on CD11b(+)Gr1(+) cells isolated from Tgfbr2(MyeKO) mice in the regulation of Arg, iNOS, VEGF, and CXCR4, and moreover, these cells have decreased suppressive activity relative to T cell proliferation. Also, we found that DCs from tumor tissue of Tgfbr2(MyeKO) mice have increased antigen-presented properties and an enhanced ability to stimulate antigen-specific T cell proliferation. We conclude that Tgfbr2 in myeloid cells has a negative role in the regulation of anti-tumorigenic functions of these cells, and deletion of this receptor decreases the suppressive function of CD11b(+)Gr1(+) cells and increases antigen-presenting properties of DCs and anti-tumorigenic properties of TAMs.
    Journal of leukocyte biology 06/2012; 92(3):641-51. DOI:10.1189/jlb.1211639 · 4.29 Impact Factor

  • Cancer Research 06/2012; 72(8 Supplement):4306-4306. DOI:10.1158/1538-7445.AM2012-4306 · 9.33 Impact Factor

  • Cancer Research 06/2012; 72(8 Supplement):2456-2456. DOI:10.1158/1538-7445.AM2012-2456 · 9.33 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: We report that IL-17 significantly increases the secretion of CXCL1 and CXCL5 from mammary carcinoma cells, which is downregulated by TGF-β through the type II TGF-β receptor (TβRII). Carcinoma cells with conditional knockout of TβRII (Tgfbr2(KO)) have enhanced sensitivity to IL-17a in the stimulation of chemokine secretion. During polyoma middle T (PyMT) induced tumor progression, levels of Th17 inducing cytokines TGF-β, IL-6, IL-23 were increased in PyMT/Tgfbr2(KO) tumors, which was associated with an increased number of Th17 cells. IL-17 increased the suppressive function of MDSCs on T cells through the upregulation of Arg, IDO, and COX2. Treatment of PyMT/Tgfbr2(KO) mice with anti-IL-17 Ab decreased carcinoma growth and metastatic burden. Analysis of human breast cancer transcriptome databases showed a strong association between IL-17 gene expression and poor outcome in lymph node positive, estrogen receptor negative or luminal B subtypes suggesting potential therapeutic approaches.
    Cancer Discovery 10/2011; 1(5):430-41. DOI:10.1158/2159-8290.CD-11-0100 · 19.45 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Pancreatic ductal adenocarcinoma (PDAC), one of the most lethal neoplasms, is characterized by an expanded stroma with marked fibrosis (desmoplasia). We previously generated pancreas epithelium-specific TGF-β receptor type II (Tgfbr2) knockout mice in the context of Kras activation (mice referred to herein as Kras+Tgfbr2KO mice) and found that they developed aggressive PDAC that recapitulated the histological manifestations of the human disease. The mouse PDAC tissue showed strong expression of connective tissue growth factor (Ctgf), a profibrotic and tumor-promoting factor, especially in the tumor-stromal border area, suggesting an active tumor-stromal interaction. Here we show that the PDAC cells in Kras+Tgfbr2KO mice secreted much higher levels of several Cxc chemokines compared with mouse pancreatic intraepithelial neoplasia cells, which are preinvasive. The Cxc chemokines induced Ctgf expression in the pancreatic stromal fibroblasts, not in the PDAC cells themselves. Subcutaneous grafting studies revealed that the fibroblasts enhanced growth of PDAC cell allografts, which was attenuated by Cxcr2 inhibition. Moreover, treating the Kras+Tgfbr2KO mice with the CXCR2 inhibitor reduced tumor progression. The decreased tumor progression correlated with reduced Ctgf expression and angiogenesis and increased overall survival. Taken together, our data indicate that tumor-stromal interactions via a Cxcr2-dependent chemokine and Ctgf axis can regulate PDAC progression. Further, our results suggest that inhibiting tumor-stromal interactions might be a promising therapeutic strategy for PDAC.
    The Journal of clinical investigation 09/2011; 121(10):4106-17. DOI:10.1172/JCI42754 · 13.22 Impact Factor

  • Cancer Research 07/2011; 71(14):5052-3. DOI:10.1158/0008-5472.CAN-11-0688 · 9.33 Impact Factor

  • Cancer Research 07/2011; 71(8 Supplement):922-922. DOI:10.1158/1538-7445.AM2011-922 · 9.33 Impact Factor

  • Cancer Research 07/2011; 71(8 Supplement):2845-2845. DOI:10.1158/1538-7445.AM2011-2845 · 9.33 Impact Factor

Publication Stats

5k Citations
474.75 Total Impact Points


  • 1995-2015
    • Vanderbilt University
      • • Department of Cancer Biology
      • • Department of Medicine
      Нашвилл, Michigan, United States
  • 2003-2014
    • Gateway-Vanderbilt Cancer Treatment Center
      Clarksville, Tennessee, United States