Transforming growth factor β (TGF-β) plays an important role in tumor initiation and progression, functioning as both a suppressor and a promoter. The mechanisms underlying this dual role of TGF-β remain unclear. TGF-β exerts systemic immune suppression and inhibits host immunosurveillance. Neutralizing TGF-β enhances CD8+ T-cell- and NK-cell-mediated anti-tumor immune responses. It also increases neutrophil-attracting chemokines resulting in recruitment and activation of neutrophils with an antitumor phenotype. In addition to its systemic effects, TGF-β regulates infiltration of inflammatory/immune cells and cancer-associated fibroblasts in the tumor microenvironment causing direct changes in tumor cells. Understanding TGF-β regulation at the interface of tumor and host immunity should provide insights into developing effective TGF-β antagonists and biomarkers for patient selection and efficacy of TGF-β antagonist treatment.
The role of myeloid cells in supporting cancer growth is well established. Most work has focused on myeloid-derived suppressor cells (MDSC) that accumulate in tumor-bearing animals, but tumor-associated neutrophils (TAN) are also known to be capable of augmenting tumor growth. However, little is known about their evolution, phenotype, and relationship to naïve neutrophils (NN) and to the granulocytic fraction of MDSC (G-MDSC).
In the current study, a transcriptomics approach was used in mice to compare these cell types. Our data show that the three populations of neutrophils are significantly different in their mRNA profiles with NN and G-MDSC being more closely related to each other than to TAN. Structural genes and genes related to cell-cytotoxicity (i.e. respiratory burst) were significantly down-regulated in TAN. In contrast, many immune-related genes and pathways, including genes related to the antigen presenting complex (e.g. all six MHC-II complex genes), and cytokines (e.g. TNF-α, IL-1-α/β), were up-regulated in G-MDSC, and further up-regulated in TAN. Thirteen of the 25 chemokines tested were markedly up-regulated in TAN compared to NN, including striking up-regulation of chemoattractants for T/B-cells, neutrophils and macrophages.
This study characterizes different populations of neutrophils related to cancer, pointing out the major differences between TAN and the other neutrophil populations.
Though TGF-β inhibition enhances anti-tumor immunity mediated by CD8+ T cells in several tumor models, it is not always sufficient for rejection of tumors. In the present study, to maximize the anti-tumor effect of TGF-β blockade, we tested the effect of anti-TGF-β combined with an irradiated tumor vaccine in a subcutaneous CT26 colon carcinoma tumor model. The irradiated tumor cell vaccine alone in prophylactic setting significant delayed tumor growth, whereas anti-TGF-β antibodies alone did not show any anti-tumor effect. However, tumor growth was inhibited significantly more in vaccinated mice treated with anti-TGF-β antibodies compared to vaccinated mice without anti-TGF-β suggesting that anti-TGF-β synergistically enhanced irradiated tumor vaccine efficacy. CD8+ T cell-depletion completely abrogated the vaccine efficacy, so protection required CD8+ T cells. Depletion of CD25+ T regulatory cells led to the almost complete rejection of tumors without the vaccine, whereas anti-TGF-β did not change the number of CD25+ T regulatory cells in un-vaccinated and vaccinated mice. Though the abrogation of CD1d-restricted NKT cells, which have been reported to induce TGF-β production by MDSC through an IL-13-IL-4R-STAT6 pathway, partially enhanced anti-tumor immunity regardless of vaccination, abrogation of the NKT cell-IL-13-IL-4R-STAT-6 immunoregulatory pathway did not enhance vaccine efficacy. Taken together, these data indicated that anti-TGF-β enhances efficacy of a prophylactic vaccine in normal individuals despite their not having the elevated TGF-β levels found in cancer patients and that the effect is not dependent on TGF-β solely from CD4+CD25+ T regulatory cells or the NKT cell-IL-13-IL-4R-STAT-6 immunoregulatory pathway.
Transforming growth factor (TGF)-β is a potent immunosuppressive cytokine necessary for cancer growth. Animal and human studies have shown that pharmacologic inhibition of TGF-β slows the growth rate of established tumors and occasionally eradicates them altogether. We observed, paradoxically, that inhibiting TGF-β before exposing animals to tumor cells increases tumor growth kinetics. We hypothesized that TGF-β is necessary for the anti-tumor effects of cytotoxic CD8+ T lymphocytes (CTLs) during the early stages of tumor initiation.
BALB/c mice were pretreated with a blocking soluble TGF-β receptor (sTGF-βR, TGF-β-blockade group, n=20) or IgG2a (Control group, n=20) before tumor inoculation. Tumor size was followed for 6 weeks. In vivo lymphocyte assays and depletion experiments were then performed to investigate the immunological basis of our results. Lastly, animals were pretreated with either sTGF-βR (n=6) or IgG2a (n=6) prior to immunization with an adenoviral vector encoding the human papillomavirus E7 gene (Ad.E7). One week later, flow cytometry was utilized to measure the number of splenic E7-specific CD8+ T cells.
Inhibition of TGF-β before the injection of tumor cells resulted in significantly larger average tumor volumes on days 11, 17, 22, 26 and 32 post tumor-inoculation (p < 0.05). This effect was due to the inhibition of CTLs, as it was not present in mice with severe combined immunodeficiency (SCID) or those depleted of CD8+ T cells. Furthermore, pretreatment with sTGF-βR inhibited tumor-specific CTL activity in a Winn Assay. Tumors grew to a much larger size when mixed with CD8+ T cells from mice pretreated with sTGF-βR than when mixed with CD8+ T cells from mice in the control group: 96 mm3 vs. 22.5 mm3, respectively (p < 0.05). In addition, fewer CD8+ T cells were generated in Ad.E7-immunized mice pretreated with sTGF-βR than in mice from the control group: 0.6% total CD8+ T cells vs. 1.9%, respectively (p < 0.05).
These studies provide the first in vivo evidence that TGF-β may be necessary for anti-tumor immune responses in certain cancers. This finding has important implications for our understanding of anti-tumor immune responses, the role of TGF-β in the immune system, and the future development of TGF-β inhibiting drugs.
Malignant mesothelioma; Tumor immunology; Immune suppression; TGF-β; CD8+ Cytotoxic T cell
Tolerance induction in T cells takes place in most tumors and is thought to account for tumor evasion from immune eradication. Production of the cytokine TGF-β is implicated in immunosuppression, however the cellular mechanism by which TGF-β induces T cell dysfunction remains unclear. Using a transgenic model of prostate cancer, we showed that tumor development was not suppressed by the adaptive immune system, which was associated with heightened TGF-β signaling in T cells from the tumor-draining lymph nodes. Blockade of TGF-β signaling in T cells enhanced tumor antigen-specific T cell responses, and inhibited tumor development. Surprisingly, T cell- but not Treg cell-specific ablation of TGF-β1 was sufficient to augment T cell cytotoxic activity and blocked tumor growth and metastases. These findings reveal that T cell production of TGF-β1 is an essential requirement for tumors to evade immunosurveillance independent of TGF-β produced by tumors.
Many advanced tumors produce excessive amounts of Transforming Growth Factor-β (TGF-β) which, in normal epithelial cells, is a potent growth inhibitor. However, in oncogenically activated cells, the homeostatic action of TGF-β is often diverted along alternative pathways. Hence, TGF-β signaling elicits protective or tumor suppressive effects during the early growth-sensitive stages of tumorigenesis. However, later in tumor development when carcinoma cells become refractory to TGF-β-mediated growth inhibition, the tumor cell responds by stimulating pathways with tumor progressing effects. At late stages of malignancy, tumor progression is driven by TGF-β overload. The tumor microenvironment is a target of TGF-β action that stimulates tumor progression via pro-tumorigenic effects on vascular, immune, and fibroblastic cells. Bone is one of the richest sources of TGF-β in the body and a common site for dissemination of breast cancer metastases. Osteoclastic degradation of bone matrix, which accompanies establishment and growth of metastases, triggers further release of bone-derived TGF-β. This leads to a vicious positive feedback of tumor progression, driven by ever increasing levels of TGF-β released from both the tumor and bone matrix. It is for this reason, that pharmaceutical companies have developed therapeutic agents that block TGF-β signaling. Nonetheless, the choice of drug design and dosing strategy can affect the efficacy of TGF-β therapeutics. This review will describe pre-clinical and clinical data of four major classes of TGF-β inhibitor, namely i) ligand traps, ii) antisense oligonucleotides, iii) receptor kinase inhibitors and iv) peptide aptamers. Long term dosing strategies with TGF-β inhibitors may be ill-advised, since this class of drug has potentially highly pleiotropic activity, and development of drug resistance might potentiate tumor progression. Current paradigms for the use of TGF-β inhibitors in oncology have therefore moved towards the use of combinatorial therapies and short term dosing, with considerable promise for the clinic.
Transforming growth factor-β (TGF-β)
There is little understanding of the impact of tumor-associated neutrophils (TAN) on adaptive immunity to tumors. In this study, we report the results of an investigation of the pathobiological basis for the prognostic significance of neutrophil elastase (NE), a serine protease found in neutrophil granules, in a model of cyclin E-overexpressing breast cancer. We established that NE was expressed by TAN within breast cancer tissues but not by breast cancer cells. NE modulated killing of breast cancer cells by cytotoxic T lymphocytes (CTL) specific for cyclin E-derived HLA-A2 restricted peptide (ILLDWLMEV). Breast cancer cells exhibited striking antigen-specific uptake of NE from the microenvironment that was independent of NE enzymatic activity. Further, NE uptake increased expression of low molecular weight forms of cyclin E and enhanced susceptibility to peptide-specific CTL lysis, suggesting that cyclin E peptides are naturally presented on breast cancer cells. Taken together, our findings reveal a previously unknown mechanism of antitumor adaptive immunity that links cancer cell uptake of an inflammatory mediator to an effective cytolytic response against an important breast cancer antigen.
neutrophil elastase; cyclin E; breast cancer; innate immunity; adaptive immunity
The transforming growth factor-β (TGF-β) signaling pathway plays a pivotal role in diverse cellular processes. TGF-β switches its role from tumor suppressor in normal or dysplastic cells to a tumor promoter in advanced cancers. It is widely believed that Smad-dependent pathway is involved in TGF-β tumor suppressive functions, whereas activation of Smad-independent pathways coupled with the loss of tumor suppressor functions of TGF-β is important for its pro-oncogenic functions. TGF-β signaling has been considered as a very suitable therapeutic target. The discovery of oncogenic actions of TGF-β has generated a great deal of enthusiasm for developing TGF-β signaling inhibitors for the treatment of cancer. The challenge is to identify the group of patients where targeted tumors are not only refractory to TGF-β-induced tumor suppressor functions but also responsive to tumor promoting effects of TGF-β. TGF-β pathway inhibitors including small and large molecules have now entered clinical trials. Preclinical studies with these inhibitors have shown promise in a variety of different tumor models. Here we emphasize on the mechanisms of signaling and specific targets of the TGF-β pathway that are critical effectors of tumor progression and invasion. This report also focuses on the therapeutic intervention of TGF-β signaling in human cancers.
Overexpression of transforming growth factor β1 (TGF-β1) has been linked to immune suppression, tumor angiogenesis, tumor cell migration, tumor cell survival, and tumor cell invasion in many cancers. In the present study, we found abundant expression of TGF-β1 in the microenvironment of four different pathological types of meningioma tumors. TGF-β1 induced invasion in malignant meningioma cells with an associated upregulation of urokinase-type plasminogen activator (uPA), uPAR, cathepsin B, and MMP-9, and this increase in proliferation was coupled with the expression of anti-apoptotic and pro-survival signaling molecules. In addition to the intense immunoreactivity of meningioma tumors to X-linked inhibitor to apoptosis (XIAP), its knockdown abolished the TGF-β1-induced proliferation of these cells. The stimulation of XIAP expression and the activation of pSMAD-2 is mediated by phosphatidylinositol 3-kinase (PI3K)- and MEK-dependent pathways, and the addition of anti-TGF-β1 antibodies prevented their expression with a consequent decrease in invasion. Bicistronic shRNA constructs targeting uPAR and cathepsin B (pUC) quenched TGF-β1-driven invasion and survival of meningioma cells by downregulation of XIAP and pSMAD-2 expression. Animal models with intracranial tumors showed elevated levels of TGF-β1, XIAP and pSMAD-2, and pUC treatment prevented this increased expression. Thus, targeted silencing of TGF-β1-induced signaling by pUC in meningioma would provide new treatment approaches for management of meningioma.
meningioma; TGF-β1; XIAP; shRNA; uPAR and cathepsin B
The skeleton is a preferred site for cancer metastasis. These bone metastases cause dysregulated bone remodeling and the associated morbidity of fractures, pain, hypercalcemia and catastrophic nerve compression syndromes. Transforming growth factor-β (TGF-β) is stored in mineralized bone matrix, and released and activated by osteoclastic bone resorption. Once activated, TGF-β stimulates nearby metastatic tumor cells within the bone microenvironment to secrete factors that further drive osteolytic destruction of the bone. Therefore, TGF-β and its signaling constitute a critical component driving the feed-forward vicious cycle of cancer growth in bone. Moreover, additional pro-tumorigenic activities attributed to TGF-β include activation of epithelial-to-mesenchymal transition, increased tumor cell invasion, enhanced angiogenesis and various immunomodulatory properties. Blocking the TGF-β signaling pathway to interrupt this vicious cycle and manipulate the bone microenvironment offers a promising area for therapeutic intervention to decrease skeletal metastasis and normalize bone homeostatic mechanisms. In this review, preclinical and clinical data are evaluated for the potential use of TGF-β pathway inhibitors in clinical practice to treat bone metastases and its associated comorbidities.
Transforming growth factor-β (TGF-β) elicits both tumor-suppressive and tumor-promoting functions during cancer progression. Here, we describe the tumor-promoting functions of TGF-β and how these functions play a role in cancer progression. Normal epithelial cells undergo epithelial-mesenchymal transition (EMT) through the action of TGF-β, while treatment with TGF-β and fibroblast growth factor (FGF)-2 results in transdifferentiation into activated fibroblastic cells that are highly migratory, thereby facilitating cancer invasion and metastasis. TGF-β also induces EMT in tumor cells, which can be regulated by oncogenic and anti-oncogenic signals. In addition to EMT promotion, invasion and metastasis of cancer are facilitated by TGF-β through other mechanisms, such as regulation of cell survival, angiogenesis, and vascular integrity, and interaction with the tumor microenvironment. TGF-β also plays a critical role in regulating the cancer-initiating properties of certain types of cells, including glioma-initiating cells. These findings thus may be useful for establishing treatment strategies for advanced cancer by inhibiting TGF-β signaling.
Angiogenesis; cancer-initiating cell; EMT; invasion; metastasis; TGF-β
Transforming growth factor (TGF)-β is the prototype in a family of secreted proteins that act in autocrine and paracrine pathways to regulate cell development and function. Normal cells typically coexpress TGF-β receptors and one or more isoforms of TGF-β, thus the synthesis and secretion of TGF-β as an inactive latent complex is considered an essential step in regula-ting the activity of this pathway. To determine whether intracellular activation of TGF-β results in TGF-β ligand–receptor interactions within the cell, we studied pristane-induced plasma cell tumors (PCTs). We now demonstrate that active TGF-β1 in the PCT binds to intracellular TGF-β type II receptor (TβRII). Disruption of the expression of TGF-β1 by antisense TGF-β1 mRNA restores localization of TβRII at the PCT cell surface, indicating a ligand-induced impediment in receptor trafficking. We also show that retroviral expression of a truncated, dominant-negative TβRII (dnTβRII) effectively competes for intracellular binding of active ligand in the PCT and restores cell surface expression of the endogenous TβRII. Analysis of TGF-β receptor–activated Smad2 suggests the intracellular ligand–receptor complex is not capable of signaling. These data are the first to demonstrate the formation of an intracellular TGF-β–receptor complex, and define a novel mechanism for modulating the TGF-β signaling pathway.
receptor; trafficking; intracellular; signal-transduction; plasmacytoma
We investigated the in vivo role of CD69 by analyzing the susceptibility of CD69−/− mice to tumors. CD69−/− mice challenged with MHC class I− tumors (RMA-S and RM-1) showed greatly reduced tumor growth and prolonged survival compared with wild-type (WT) mice. The enhanced anti–tumor response was NK cell and T lymphocyte–mediated, and was due, at least in part, to an increase in local lymphocytes. Resistance of CD69−/− mice to MHC class I− tumor growth was also associated with increased production of the chemokine MCP-1, diminished TGF-β production, and decreased lymphocyte apoptosis. Moreover, the in vivo blockade of TGF-β in WT mice resulted in enhanced anti–tumor response. In addition, CD69 engagement induced NK and T cell production of TGF-β, directly linking CD69 signaling to TGF-β regulation. Furthermore, anti-CD69 antibody treatment in WT mice induced a specific down-regulation in CD69 expression that resulted in augmented anti–tumor response. These data unmask a novel role for CD69 as a negative regulator of anti–tumor responses and show the possibility of a novel approach for the therapy of tumors.
cytokines; homeostasis; apoptosis
Transforming growth factor (TGF)-β is known to be produced by progressor tumors and to immobilize dendritic cells (DCs) within those tumors. Moreover, although TGF-β1 has been shown to promote tumor progression, there is still no direct, in vivo evidence as to whether TGF-β1 is able to directly induce distant metastasis.
To address that issue and investigate the mechanism by which TGF-β1 suppresses DC activity, we subdermally inoculated mouse ears with squamous cell carcinoma cells stably expressing TGF-β1 or empty vector (mock).
The numbers of DCs within lymph nodes draining the resultant TGF-β1-expressing tumors was significantly lower than within nodes draining tumors not expressing TGF-β1. We then injected fluorescently labeled bone marrow-derived dendritic cells into the tumors, and subsequent analysis confirmed that the tumors were the source of the DCs within the tumor-draining lymph nodes, and that there were significantly fewer immature DCs within the nodes draining TGF-β1-expressing tumors than within nodes draining tumors not expressing TGF-β1. In addition, 14 days after tumor cell inoculation, lymph node metastasis occurred more frequently in mice inoculated with TGF-β1 transfectants than in those inoculated with the mock transfectants.
These findings provide new evidence that tumor-derived TGF-β1 inhibits migration of DCs from tumors to their draining lymph nodes, and this immunosuppressive effect of TGF-β1 increases the likelihood of metastasis in the affected nodes.
dendritic cell; migration; transforming growth factor-β1; tumor draining lymph node; lymph node metastasis
Macrophages are widely distributed innate immune cells that play indispensable roles in the innate and adaptive immune response to pathogens and in-tissue homeostasis. Macrophages can be activated by a variety of stimuli and polarized to functionally different phenotypes. Two distinct subsets of macrophages have been proposed, including classically activated (M1) and alternatively activated (M2) macrophages. M1 macrophages express a series of proinflammatory cytokines, chemokines, and effector molecules, such as IL-12, IL-23, TNF-α, iNOS and MHCI/II. In contrast, M2 macrophages express a wide array of anti-inflammatory molecules, such as IL-10, TGF-β, and arginase1. In most tumors, the infiltrated macrophages are considered to be of the M2 phenotype, which provides an immunosuppressive microenvironment for tumor growth. Furthermore, tumor-associated macrophages secrete many cytokines, chemokines, and proteases, which promote tumor angiogenesis, growth, metastasis, and immunosuppression. Recently, it was also found that tumor-associated macrophages interact with cancer stem cells. This interaction leads to tumorigenesis, metastasis, and drug resistance. So mediating macrophage to resist tumors is considered to be potential therapy.
Transforming growth factor (TGF)-β family members regulate multiple aspects of wound repair through effects on cell proliferation, matrix production, and tissue inflammation, but the effects of TGF-β on wound closure itself have been controversial. We found that blocking antibodies to TGF-β enhanced the degree of closure of scratch wounds in primary airway epithelial monolayers, while addition of exogenous TGF-β1 inhibited the degree of closure, suggesting that endogenous activation of TGF-β normally serves as a brake on the degree of wound closure. Although these cells secreted large amounts of TGF-β2 and small amounts of TGF-β1, blockade of TGF-β1 enhanced the degree of wound closure, whereas blockade of TGF-β2 had no effect. TGF-β1 (but not TGF-β2) can be activated by two members of the integrin family, αvβ6 and αvβ8, which are both expressed on airway epithelial cells. Wounding induced activation of TGF-β through effects of both integrins, but antibodies against αvβ8 enhanced the degree of wound closure, whereas antibodies against αvβ6 did not.
airway epithelium; integrin αvβ6; integrin αvβ8; TGF-β; wound closure
Transforming growth factor-β (TGF-β) is a highly pleiotropic cytokine playing pivotal roles in immune regulation. TGF-β facilitates tumor cell survival and metastasis by targeting multiple cellular components. Focusing on its immunosuppressive functions, TGF-β antagonists have been employed for cancer treatment to enhance tumor immunity. TGF-β antagonists exert anti-tumor effects through #1 activating effector cells such as NK cells and cytotoxic CD8+ T cells (CTLs), #2 inhibiting regulatory/suppressor cell populations, #3 making tumor cells visible to immune cells, #4 inhibiting the production of tumor growth factors. This review focuses on the effect of TGF-β on T cells, which are differentiated into effector T cells or newly identified tumor-supporting T cells.
transforming growth factor-β (TGF-β); primary tumor; metastasis; CD8+T cells; IL-17
Upon activation, neutrophils release fibers composed of chromatin and neutrophil proteins termed neutrophil extracellular traps (NETs). NETs trap and kill microbes, activate dendritic cells and T cells, and are implicated in autoimmune and vascular diseases. Given the growing interest in the role of neutrophils in cancer immunoediting and the diverse function of NETs, we searched for NETs release by tumor-associated neutrophils (TANs). Using pediatric Ewing sarcoma (ES) as a model, we retrospectively examined histopathological material from diagnostic biopsies of eight patients (mean ± SD age of 11.5 ± 4.7 years). TANs were found in six patients and in two of those we identified NETs. These two patients presented with metastatic disease and despite entering complete remission after intensive chemotherapy had an early relapse. NETs were not identified in the diagnostic biopsies of two patients with localized disease and two with metastatic disease. This study is the first to show that TANs in ES are activated to make NETs, pointing to a possible role of NETs in cancer.
cancer; cancer immunoediting; Ewing sarcoma; neutrophils; neutrophil extracellular traps
Non-melanoma skin cancers (NMSCs) are the most common neoplasm in organ transplant recipients (OTRs). These cancers are more invasive and metastatic as compared to those developed in normal cohorts. Previously, we have shown that immunosuppressive drug, cyclosporine A (CsA) directly alters tumor phenotype of cutaneous squamous cell carcinomas (SCCs) by activating TGF-β and TAK1/TAB1 signaling pathways. Here, we identified novel molecular targets for the therapeutic intervention of these SCCs. We observed that combined blockade of Akt and p38 kinases-dependent signaling pathways in CsA-promoted human epidermoid carcinoma A431 xenograft tumors abrogated their growth by more than 90%. This diminution in tumor growth was accompanied by a significant decrease in proliferation and an increase in apoptosis. The residual tumors following the combined treatment with Akt inhibitor triciribine and p38 inhibitors SB-203580 showed significantly diminished expression of phosphorylated Akt and p38 and these tumors were less invasive and highly differentiated. Diminished tumor invasiveness was associated with the reduced epithelial–mesenchymal transition as ascertained by the enhanced E-cadherin and reduced vimentin and N-cadherin expression. Consistently, these tumors also manifested reduced MMP-2/9. The decreased p-Akt expression was accompanied by a significant reduction in p-mTOR. These data provide first important combinatorial pharmacological approach to block the pathogenesis of CsA-induced highly aggressive cutaneous neoplasm in OTRs.
OTR; SCC; Cyclosporine A; mTOR; p38; Akt
Transforming growth factor-β (TGF-β) is a potent pleiotropic cytokine that regulates mammalian development, differentiation, and homeostasis in essentially all cell types and tissues. TGF-β normally exerts anticancer activities by prohibiting cell proliferation, and by creating cell microenvironments that inhibit cell motility, invasion and metastasis. However, accumulating evidence indicates the process of tumorigenesis, particularly that associated with metastatic progression, confers TGF-β with oncogenic activities, a functional switch known as the “TGF-β Paradox.” The molecular determinants governing the “TGF-β Paradox” are complex and represent an intense area of investigation by researchers in academic and industrial settings. Recent findings link genetic and epigenetic events in mediating the acquisition of oncogenic activity by TGF-β, as does aberrant alterations within tumor microenvironments. These events coalesce to enable TGF-β to direct metastatic progression via the stimulation of epithelial-mesenchymal transition (EMT), which permits carcinoma cells to abandon polarized epithelial phenotypes in favor of apolar mesenchymal-like phenotypes. Attempts to deconstruct the EMT process induced by TGF-β have identified numerous signaling molecules, transcription factors, and microRNAs operant in mediating the initiation and resolution of this complex transdifferentiation event. Besides its ability to enhance carcinoma cell invasion and metastasis, EMT also engenders transitioned cells with stem-like properties, including the acquisition of self-renewal and tumor-initiating capabilities coupled to chemoresistance. Here we review recent findings that delineate the pathophysiological mechanisms whereby EMT stimulated by TGF-β promotes metastatic progression and disease recurrence in human carcinomas.
Cancer stem cells; Chemoresistance; Epithelial-mesenchymal transition; Integrins; Metastasis; Transforming growth factor-β; Tumor microenvironment
Many advanced tumors produce excess amounts of Transforming Growth Factor-β (TGF-β), which is a potent growth inhibitor of normal epithelial cells. However, in tumors its homeostatic action on cells can be diverted along several alternative pathways. Thus, TGF-β signaling has been reported to elicit a preventative or tumor suppressive effect during the earlier stages of tumorigenesis, but later in tumor development, when carcinoma cells become refractory to TGF-β-mediated growth inhibition, response to TGF-β signaling elicits predominantly tumor progressing effects. This is not a simple switch from suppression to progression, but more like a rheostat, involving multiple complementary and antagonizing activities that slowly tip the balance from one to the other. This review will focus on the multiple activities of TGF-β in regulation of two epithelial tumor types, namely squamous cell carcinoma and breast cancer. Basic findings in current mouse models of cancer are presented, as well as a discussion of the complicating issue of outcome of altered TGFβ signaling depending on genetic variability between mouse strains. This review also discusses the role TGF-β within the tumor microenvironment particularly its ability to polarize the microenvironment towards a pro-tumorigenic milieu.
TGF-β; tumor microenvironment; Breast cancer
TGF-β is a pluripotent cytokine that can have both tumor suppressing and tumor promoting effects on epithelial cells. It is unclear what determines when TGF-β and its signaling pathway act predominantly as a tumor suppressor pathway or as a tumor-promoter pathway and whether TGF-β can have both classes of effects concurrently on a cell. We investigated the effect of TGF-β on anoikis in colorectal cancer cell lines sensitive to TGF-_-mediated growth inhibition to determine if the context of the cells could be one of the factors that would affect whether TGF-β exerts tumor suppressor or oncogene activity on colon cancer cells. We observed variable effects of TGF-_ on anoikis in these cell lines, even though they all are growth-inhibited by TGF-β. Thus, we show that TGF-β has variable effects on anoikis in colon cancer cell lines that likely reflects the effects of concurrent gene mutations in the cancer cells and the activation state of the signaling pathways controlled by these genes.
Transforming Growth Factor Beta; colorectal cancer; apoptosis; anoikis
Tanshinone IIA (Tan IIA) is a diterpene quinone extracted from the root of Salvia miltiorrhiza, a Chinese traditional herb. Although previous studies have reported the anti-tumor effects of Tan IIA on various human cancer cells, the underlying mechanisms are not clear. The current study was undertaken to investigate the molecular mechanisms of Tan IIA's apoptotic effects on leukemia cells in vitro.
The cytotoxicity of Tan IIA on different types of leukemia cell lines was evaluated by the 3-[4,5-dimethylthiazol-2,5]-diphenyl tetrazolium bromide (MTT) assay on cells treated without or with Tan IIA at different concentrations for different time periods. Cellular apoptosis progression with and without Tan IIA treatment was analyzed by Annexin V and Caspase 3 assays. Gene expression profiling was used to identify the genes regulated after Tan IIA treatment and those differentially expressed among the five cell lines. Confirmation of these expression regulations was carried out using real-time quantitative PCR and ELISA. The antagonizing effect of a PXR inhibitor L-SFN on Tan IIA treatment was tested using Colony Forming Unit Assay.
Our results revealed that Tan IIA had different cytotoxic activities on five types of leukemia cells, with the highest toxicity on U-937 cells. Tan IIA inhibited the growth of U-937 cells in a time- and dose-dependent manner. Annexin V and Caspase-3 assays showed that Tan IIA induced apoptosis in U-937 cells. Using gene expression profiling, 366 genes were found to be significantly regulated after Tan IIA treatment and differentially expressed among the five cell lines. Among these genes, CCL2 was highly expressed in untreated U-937 cells and down-regulated significantly after Tan IIA treatment in a dose-dependent manner. RT-qPCR analyses validated the expression regulation of 80% of genes. Addition of L- sulforaphane (L-SFN), an inhibitor of Pregnane × receptor (PXR) significantly attenuated Tan IIA's effects using colony forming assays.
Tan IIA has significant growth inhibition effects on U-937 cells through the induction of apoptosis. And Tan IIA-induced apoptosis might result from the activation of PXR, which suppresses the activity of NF-κB and lead to the down-regulation of CCL2 expression.
Gene expression profiling; apoptosis; CCL2; U-937 cell lines; tanshinone IIA (Tan IIA)
Adoptive cellular immunotherapy has promise as an approach to eradicate established tumors. However, a significant hurdle in the success of cellular immunotherapy involves recently identified mechanisms of immune suppression on cytotoxic T-cells at the effector phase.
Transforming growth factor-β (TGF-β) is one of the most important of these immunosuppressive factors because it affects both T-cell and macrophage functions. We thus hypothesized that systemic blockade of TGF-β signaling combined with adoptive T-cell transfer would enhance the effectiveness of the therapy.
Flank tumors were generated in mice using the OVA-albumin (OA) expressing thymoma cell line, EG7. Splenocytes from transgenic OT-1 mice (whose CD8 T-cells recognize an immunodominant peptide in OA) were activated in vitro and adoptively transferred into mice bearing large tumors in the presence or absence of an orally available TGF-β receptor-I kinase blocker (SM16).
We observed markedly smaller tumors in the group receiving the combination of SM16 chow and adoptive transfer. Additional investigation revealed that TGF-β receptor blockade increased the persistence of adoptively transferred T-cells in the spleen and lymph nodes, increased numbers of adoptively transferred T-cells within tumors, increased activation of these infiltrating T-cells, and altered the tumor microenvironment with a significant increase in TNF-α and decrease in arginase mRNA expression
We found that systemic blockade of TGF-β receptor activity augmented the anti-tumor activity of adoptively transferred T-cells and may thus be a useful adjunct in future clinical trials.
tumor immunology; immunosuppression; TGFβ; Cytotoxic T-cells; cytokines; adoptive transfer
Transforming growth factor-β (TGF-β) normally inhibits breast cancer development by preventing mammary epithelial cell (MEC) proliferation, by inducing MEC apoptosis, and by creating cell microenvironments that maintain MEC homeostasis and prevent their uncontrolled growth and motility. Mammary tumorigenesis elicits dramatic alterations in MEC architecture and microenvironment integrity, which collectively counteract the tumor-suppressing activities of TGF-β and enable its stimulation of breast cancer invasion and metastasis. How malignant MECs overcome the cytostatic actions imposed by normal microenvironments and TGF-β, and how abnormal microenvironments conspire with TGF-β to stimulate the development and progression of mammary tumors remains largely undefined. These knowledge gaps have prevented science and medicine from implementing treatments effective in simultaneously targeting abnormal cellular microenvironments, and in antagonizing the oncogenic activities of TGF-β in developing and progressing breast cancers. c-Abl is a ubiquitously expressed nonreceptor protein tyrosine kinase that essentially oversees all aspects of cell physiology, including the regulation of cell proliferation, migration and adhesion, as well as that of cell survival. Thus, the biological functions of c-Abl are highly reminiscent of those attributed to TGF-β, including the ability to function as either a suppressor or promoter of tumorigenesis. Interestingly, while dysregulated Abl activity clearly promotes tumorigenesis in hematopoietic cells, an analogous role for c-Abl in regulating solid tumor development, including those of the breast, remains controversial. Here, we review the functions of c-Abl in regulating breast cancer development and progression, and in alleviating the oncogenic activities of TGF-β and its stimulation of epithelial-mesenchymal transition during mammary tumorigenesis.
Breast cancer; c-Abl; Epithelial-mesenchymal transition; Metastasis; Signal transduction; Transforming growth factor-β