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The American Journal of Pathology logoLink to The American Journal of Pathology
. 2012 Jun;180(6):2188–2200. doi: 10.1016/j.ajpath.2012.02.031

Role of Cripto-1 during Epithelial-to-Mesenchymal Transition in Development and Cancer

Maria C Rangel 1, Hideaki Karasawa 1, Nadia P Castro 1, Tadahiro Nagaoka 1, David S Salomon 1, Caterina Bianco 1,⁎
PMCID: PMC3378914  PMID: 22542493

Abstract

Epithelial-to-mesenchymal transition (EMT) is a critical multistep process that converts epithelial cells to more motile and invasive mesenchymal cells, contributing to body patterning and morphogenesis during embryonic development. In addition, both epithelial plasticity and increased motility and invasiveness are essential for the branching morphogenesis that occurs during development of the mammary gland and during tumor formation, allowing cancer cells to escape from the primary tumor. Cripto-1, a member of the epidermal growth factor–Cripto-1/FRL-1/Cryptic (EGF/CFC) gene family, together with the transforming growth factor (TGF)-β family ligand Nodal, regulates both cell movement and EMT during embryonic development. During postnatal development, Cripto-1 regulates the branching morphogenesis of the mouse mammary gland and enhances both the invasive and migratory properties of mammary epithelial cells in vitro. Furthermore, transgenic mouse models have shown that Cripto-1 promotes the formation of mammary tumors that display properties of EMT, including the down-regulation of the cell surface adherens junctional protein E-cadherin and the up-regulation of mesenchymal markers, such as vimentin, N-cadherin, and Snail. Interestingly, Cripto-1 is enriched in a subpopulation of embryonal, melanoma, prostate, and pancreatic cancer cells that possess stem-like characteristics. Therefore, Cripto-1 may play a role during developmental EMT, and it may also be involved in the reprogramming of differentiated tumor cells into cancer stem cells through the induction of an EMT program.

The epithelial-to-mesenchymal transition (EMT) is a multistep process during which epithelial cells sequentially lose their epithelial characteristics and acquire motility and invasive properties that are characteristic of a mesenchymal phenotype.1 Cell surface adherens junctional proteins, such as E-cadherin, and tight junction proteins, such as occludins and claudins, mediate epithelial connections between adjacent epithelial cells and are essential for both the maintenance of the epithelial cell phenotype and the preservation of apical-basal polarity. During EMT, E-cadherin and other adherens junction and tight junction proteins are down-regulated and replaced by N-cadherin, providing more transient adhesive and motile properties.1 In addition, reorganization of cytoskeletal elements, such as increases in actin stress fiber formation and in vimentin and smooth muscle actin expression, allows the cells to acquire a more spindle-shaped form and gain a mesenchymal phenotype, exhibiting front-to-back, leading-edge polarity.1 Finally, the cells become devoid of any intercellular contacts, and invade and migrate along specific routes, possibly guided by collagen fibrils and other extracellular matrix proteins to vascular or lymphatic capillaries, where they intravasate. Once they reach their final destination, such as an end-organ metastatic site, extravasating mesenchymal cells reacquire their epithelial phenotype through a process known as mesenchymal-to-epithelial transition (MET).2

Functional loss of E-cadherin is considered a hallmark of EMT and is frequently observed at sites of EMT in both embryonic development and cancer, especially at the leading edge of tumors, inhibiting contact with activated host stromal cells.1 EMT is triggered by a variety of extracellular growth factors, such as transforming growth factor (TGF)-β, fibroblast growth factors (FGFs), Wnts, extracellular matrix proteins, and transcription factors, that cooperate to reprogram epithelial cells into a mesenchymal phenotype.1 Transcription factors that repress E-cadherin expression and induce expression of a mesenchymal gene signature include the zinc finger proteins Snail and Slug, the basic helix loop helix factors Twist1 and Twist2, and the zinc finger E-box–binding homeobox 1 and 2, respectively.3

EMT is critical for gastrulation and neural crest cell formation during embryogenesis and also plays a role in fibrosis and wound healing in adult tissues.1 Aberrant activation of the developmental EMT program contributes to tumor invasion, metastatic dissemination, and acquisition of therapeutic resistance to chemotherapy and radiation. Numerous inducers of EMT during embryonic development and cancer have been identified.4 Cripto-1, a member of the epidermal growth factor–Cripto-1/FRL-1/Cryptic (EGF/CFC) gene family, is an example of a critical EMT inducer during embryonic development that is often reactivated in cancer cells, regulating the multistep program of tumorigenesis.5 In this review article, we will illuminate the role played by Cripto-1 as an EMT regulator during embryogenesis, mammary gland development, and tumorigenesis, specifically in breast cancer progression.

Binding Partners and Intracellular Signaling Pathways Modulated by Cripto-1

Cripto-1 has multiple binding partners and can modulate, in either a positive or a negative manner, a variety of intracellular signaling pathways (Nagaoka et al6 provide an in-depth understating of the Cripto-1 signaling network). Two of the major signaling pathways activated by Cripto-1 are the Nodal/Alk4/Alk7/Smad-2 signaling pathway and the 78-kDa glucose-regulated protein (GRP78)/Glypican-1/c-src/mitogen-activated protein kinase (MAPK)/Akt signaling pathway.6

During embryogenesis, Cripto-1 functions primarily as a coreceptor for the TGF-β family ligands Nodal and growth and differentiation factors (GDFs) 1 and 3, leading to the activation of type I (Alk4/Alk7) serine-threonine kinase receptors and the Activin type II receptor complex, which triggers both the phosphorylation and activation of the Smad-2/Smad-3 intracellular signaling pathway.6 Cripto-1 also interacts with other TGF-β family members, such as TGF-β1 and Activins A and B, interfering with the binding of these ligands to their cognate receptors and attenuating TGF-β1 and Activin A and B signaling in multiple cell lines.6

In addition to functioning as a coreceptor for TGF-β family ligands, Cripto-1 can also bind to the glycosyl-phosphatidyl-inositol–anchored heparan sulfate proteoglycan glypican-1 or to the endoplasmic reticulum protein GRP78 on the cell membrane, and it can induce the activation of a c-src/MAPK/Akt signaling pathway, promoting cell proliferation, motility, and survival.6 Indeed, the activation of a c-src/MAPK/Akt signaling pathway by Cripto-1 in a glypican-1– or GRP78-dependent manner strongly correlates with Cripto-1 oncogenic activity both in vitro and in vivo. Furthermore, Cripto-1 cross talks with several other signaling pathways, including the tyrosine kinase receptor erbB4, Wnt/β-catenin, Notch, caveolin-1, and apelin pathways, indicating that the Cripto-1 signaling pathway might have multiple cellular functions.6

Cripto-1 in EMT during Embryonic Development

EMT performs a critical role during embryonic development, especially in gastrulation, which is a fundamental process for the generation of the three primitive germ layers.7 During gastrulation, ectodermal cells at the primitive streak, because of signaling molecules generated by the Spermann-Mangold organizer, undergo EMT (Figure 1). These cells migrate through the posterior region of the primitive streak into the underlying tissue, producing the primary mesenchyme or undergoing MET and forming the underlying endoderm7 (Figure 1). TGF-β superfamily members, such as Nodal, GDF1, and GDF3 and their coreceptors Cripto-1, Wnt/β-catenin, and FGF, have been implicated in coordinately regulating tissue morphogenesis during gastrulation7 (Figure 1). During early mouse embryonic development, Cripto-1 expression is initially detected in the inner cell mass of the blastocyst and in extra-embryonic trophoblast cells before the gastrulation stage.5,8 In the epiblast before primitive streak formation, asymmetric expression of Cripto-1 occurs in a gradient (ie, in a proximal-distal manner). During the initial stages of gastrulation, localization of Cripto-1 occurs at the nascent primitive streak and then spreads to the embryonic mesoderm. Cripto-1 expression is significantly reduced during the early neural lateral plate stage and disappears in the late neural plate stage.5,8 Cripto-1–null mice are embryonically lethal because of a lack of embryonic mesoderm and a definitive endoderm.5,8 Similarly, knockout mouse mutants of other EGF/CFC family members, such as zebra fish one eyed pinhead (oep) and mouse Cryptic, are characterized by the absence of ventral neuroectoderm and left/right laterality defects resulting from abnormal gastrulation and defects in cell movement.5,8 During gastrulation and lateral plate formation, the Notch1 and canonical Wnt/β-catenin signaling pathways are intimately involved in regulating the downstream expression of Nodal and Cripto-1, respectively.9,10

Figure 1.

Figure 1

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EMT during gastrulation. Gastrulation is a fundamental process for the generation of the three primary germ layers: ectoderm, mesoderm, and endoderm. Ectodermal cells at the primitive streak undergo EMT in response to signaling molecules generated by the Spemann-Mangold organizer. These cells migrate through the primitive streak into the underlying tissue, becoming primary mesoderm. Ingressing cells can also undergo MET and produce the endoderm.

The heart is the first organ to develop during embryogenesis, and cardiac progenitor cells are initially specified during the EMT program at gastrulation.11 Several studies indicate that Cripto-1 and canonical Wnt/β-catenin signaling are essential for cardiomyocyte differentiation. In this regard, Cripto-1–null−/− embryonic stem (ES) cells lose their ability to form beating cardiomyocytes, whereas the cardiomyocyte differentiation program can be rescued by reintroducing a Cripto-1–expressing gene into Cripto-1–null−/− cells.12,13 Furthermore, the G-protein–coupled receptor putative receptor protein related to Angiotensin-type I receptor and its ligand apelin have been identified as downstream targets of Cripto-1 signaling during the cardiomyocyte differentiation of ES cells, promoting mesoderm patterning and cardiac specification.14 Interestingly, mutation analysis in human genomic DNA suggests that a variant of the teratocarcinoma-derived growth factor-1 (TDGF-1) gene encoding the Cripto-1 protein is associated with the development of a ventricular septal defect, which is the most common congenital heart defect.15

In contrast, Liguori et al16 revealed the existence of a stable neural regionalization of the anterior character in Cripto-1–null−/− embryos. Cripto-1–null−/− embryos can still form functional neural stem cells that are able to differentiate and maintain a neural phenotype both in vivo and in vitro.17 Cripto-1–null−/− ES cells spontaneously differentiate into neurons in the absence of retinoic acid, indicating that Nodal and Cripto-1 are negative regulators of neuroectoderm differentiation during embryogenesis.13 Furthermore, disruption of Cripto-1 expression in mouse ES cells can enhance neurogenesis and midbrain dopaminergic differentiation in vitro.18,19 These findings suggest that Cripto-1 is a potential target for Parkinson's disease therapy, and mouse ES cells treated with a Cripto-1–blocking peptide induce enhanced functional recovery in a rat model of Parkinson's disease.20

Whether all of the Cripto-1 activities during embryogenesis are Nodal dependent remains controversial. For instance, null mutants for Cripto-1 and Nodal, although they share several similarities, display significant differences in their phenotypes.5,8 However, Cryptic, a Cripto-1 ortholog, or GDF1 and/or GDF3, which are expressed during embryogenesis, might compensate for Cripto-1 or Nodal loss during embryogenesis. In fact, the Cripto-1/Cryptic double mutant shows striking similarities to the Nodal-null mutant phenotype, indicating that EGF/CFC proteins are required for almost all of Nodal activities during embryonic development.21

On the other hand, several studies have suggested that Cripto-1 can function during embryogenesis independently of Nodal. Cripto-1F78A/F78A mouse embryos, containing the amino acid substitution F78A, which fails to activate the Nodal/Alk4/Smad-2 signaling pathway, can initiate gastrulation movements and pattern anterior/posterior axis formation, unlike Cripto-1–null mice.22 Another study has also shown that Cripto-1 uses a distinct signaling pathway from Nodal in the induction of visceral endoderm (VE) and anterior VE (AVE) differentiation. In fact, extra-embryonic endodermal stem cells can differentiate into VE and AVE by treatment with Nodal or Cripto-1 in chimeric embryos. However, although the differentiation program induced by Nodal is inhibited by SB431542, a selective and potent inhibitor of Alk4/Alk5/Alk7, differentiation of VE and AVE, stimulated by Cripto-1, is unaffected by SB431542 treatment. These results suggest that Cripto-1 can contribute to the differentiation of VE and AVE independently of Nodal.23 Taken together, it would appear that Cripto-1 and Nodal are dependent on each other for signaling during embryogenesis, although different studies have shed light on Cripto-1/Nodal-independent and Nodal/Cripto-1–independent signaling pathways.

Cross Talk between Cripto-1 and Other EMT Regulators in Embryogenesis and Cancer

TGF-β Signaling Pathway

The TGF-β signaling pathway is a critical inducer of EMT in the context of development and during late-stage cancer progression.24 During embryogenesis, TGF-β is required for heart valve formation and palatogenesis, processes that require cell plasticity and EMT.25,26 In vitro studies have shown that TGF-β is a classic inducer of EMT in a variety of systems. In breast cancer, TGF-β can function as a tumor suppressor in the early onset of cancer or as a tumor promoter in later stages, probably through its ability to induce EMT by interacting with the ras, p38 MAPK, and Wnt/β-catenin signaling pathways.24 The TGF-β signaling pathway directly regulates the transcription of classic EMT transcription factors, such as Snail, Twist, and Slug (Figure 2).24 Conversely, TGF-β is a target of the transcription factors Snail and Slug.27 For example, Snail- and Slug-overexpressing MCF-7 cells show an increase in TGF-β expression, enhanced activation of TGF-β signaling, and an increased migratory response, which are significantly reduced after treatment with specific TGF-β inhibitors.27 Nodal positively regulates expression of Snail, Slug, and Twist in Xenopus, possibly through the activation of NF-κB.28 Interestingly, Twist and Slug negatively regulate expression of Cerberus, a Nodal inhibitor, thereby increasing Nodal activity and generating a positive feedback loop28 (Figure 2).

Figure 2.

Figure 2

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Cross talk of Cripto-1 with other EMT signaling pathways. Cripto-1 cooperates with several EMT signaling pathways in the induction of EMT in both embryogenesis and cancer. Cross talk of Cripto-1 with other EMT inducers is described in detail in the text. HIF, hypoxia-inducible factor.

Cripto-1, in addition to mediating Nodal, GDF1, and GDF3 signaling, can modulate signaling of other TGF-β family ligands, including Activin and TGF-β. Cripto-1 binds to TGF-β1 and interferes with the binding of TGF-β1 to the TGF-β type I receptor, interfering with TGF-β signaling and tumor suppressor activity in a variety of cell lines.29,30 Similarly, Cripto-1 forms a complex with Activins A and B, preventing the activation of an Activin type II and Alk4 receptor complex.31,32 Furthermore, the heat shock protein GRP78 interacts with Cripto-1 and potentiates the Cripto-1 inhibitory effects on TGF-β and Activin signaling.33,34 Interestingly, TGF-β family ligands, including TGF-β1 and bone morphogenic protein-4 (BMP-4), can directly regulate Cripto-1 gene expression in both NTERA-2/D1 human embryonal carcinoma (EC) cells and LS174-T human colon cancer cells because of the presence of functional Smad-binding elements within the Cripto-1 promoter region (Table 1).35–41 Surprisingly, although TGF-β1 strongly enhances Cripto-1 mRNA and protein expression, BMP-4 down-regulates Cripto-1 expression and interferes with Cripto-1 oncogenic activity in vitro. In fact, BMP-4 inhibits migration of LS174-T cells and induces differentiation of NTERA-2/D1 cells.35 Therefore, Cripto-1 might play a role in restraining the TGF-β cytostatic program while promoting TGF-β protumorigenic effects in late stages of oncogenic transformation. In fact, TGF-β signaling might promote tumorigenesis, in part, through the direct induction of Cripto-1 expression, which, in turn, attenuates TGF-β cytostatic effects while increasing proliferation, motility, invasion, and EMT of tumor cells (Figure 2 and Table 1). However, the potential cooperation between Cripto-1 and TGF-β signaling in promoting cell motility, invasion, and EMT in advanced tumors requires further investigation.

Table 1.

Regulation of Cripto-1 Expression

Type of regulator Regulators mRNA and/or protein Methods Ref. no.
Positive
 Growth factors TGF-β1 mRNA/protein RT-qPCR, WB 35
Wnt/β-catenin mRNA RT-qPCR 36
 Transcription factors HIF-1α mRNA/protein RT-qPCR, WB 37
Msx2 mRNA/protein Semiquantitative RT-PCR, WB 38
Nanog mRNA Microarray 39
Oct-4 mRNA Microarray 39
Negative
 Growth factors BMP-4 mRNA/protein RT-qPCR, WB 35
 Transcription factors Netrin-1 mRNA/protein RT-qPCR, WB 40
GCNF mRNA Northern blot 41
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GCNF, germ cell nuclear factor; RT-qPCR, quantitative RT-PCR; WB, Western blot.

Canonical Wnt/β-Catenin Signaling Pathway

Similar to Cripto-1, the canonical Wnt/β-catenin signaling pathway is implicated in EMT during development and cancer.41 The Wnt/β-catenin signaling pathway can regulate EMT during gastrulation and cardiac valve formation.41 Cross talk between Wnt/β-catenin and Cripto-1 has been clearly demonstrated during embryonic development (Figure 2). For instance, Cripto-1 expression is lost in β-catenin–knockout embryos.10 Interestingly, both β-catenin– and Cripto-1–knockout embryos show similar phenotypes that are characterized by the absence of an anteroposterior axis. Wnt3-knockout embryos also show down-regulation of Cripto-1 expression, although the levels of Cripto-1 down-regulation are not as dramatic as in the β-catenin–knockout embryos. The direct regulation of Cripto-1 expression by β-catenin is dependent on the presence of T-cell factor/lymphoid enhancer factor–binding elements in the promoter and first intronic region of the human Cripto-1 gene (Table 1).36 Studies in Xenopus have also shown that one of the six Cripto-1 orthologs in Xenopus, FRL-1, directly binds to Wnt11 and, together with the glycosyl-phosphatidyl-inositol–linked proteoglycan Glypican-4, activates the canonical Wnt/β-catenin signaling.42 FRL-1 and Wnt11 are both essential for dorsal axis formation in Xenopus embryos and, therefore, Wnt11- and FRL-1–depleted embryos show similar phenotypes.42

The Wnt/β-catenin pathway also plays a key role during mammary gland development, regulating the formation of the mammary placodes during midgestation (E10) and postnatally by promoting side branching in the virgin mammary gland and alveolar differentiation during pregnancy.43 Multiple Wnt ligands and their receptors are expressed in cells composing the terminal end buds (TEBs), which are the sites of active epithelial growth and migration in the virgin mammary gland.43 Moreover, Wnt1 overexpression in the mouse mammary gland, under the control of the mouse mammary tumor virus (MMTV) promoter, induces epithelial hyperbranching and precocious alveolar differentiation.44 A functional link between Wnt/β-catenin signaling and Cripto-1 in the mammary gland is supported by the observation that Cripto-1 overexpression in the mouse mammary gland is associated with an increase in the activated, dephosphorylated form of β-catenin and an increase in the inactive, phosphorylated form of glycogen synthase kinase-3β.45 Conversely, an increase in Cripto-1 expression has been detected by immunohistochemistry in MMTV-Wnt1 mammary tumors, indicating that Cripto-1 might be a mediator of the effects of activated Wnt/β-catenin signaling in the mammary gland during tumorigenesis.46

In vitro studies have also implicated Wnt/β-catenin signaling in epithelial plasticity and EMT. Activation of the Wnt/β-catenin pathway in breast cancer cells induces the EMT regulators Snail and Twist and up-regulates the mesenchymal marker vimentin (Figure 2).47–49 Conversely, Twist-overexpressing MCF-7 breast cancer cells activate the Wnt/β-catenin signaling pathway and induce expression of CD44 and aldehyde dehydrogenase 1, stem cell markers associated with EMT.50

Brachyury

Brachyury, a T-box transcription factor, is required for both normal gastrulation movements and posterior mesoderm formation during embryogenesis (Figure 1). Brachyury is a direct transcriptional target of the FGF and canonical Wnt/β-catenin signaling pathways (Figure 2).51,52 Moreover, Brachyury regulates the expression of Wnt3a and Wnt8, suggesting that a positive autoregulatory loop between Brachyury and the canonical Wnt/β-catenin signaling pathway might occur during embryogenesis.53 Notably, Cripto-1 is also essential for the up-regulation of Brachyury expression in response to Wnt/β-catenin signaling in P19 EC cells and by Nkx2.5 in cardiomyocyte precursor cells (Figure 2).54,55 In fact, both Brachyury and Nkx2.5 are essential inducers of mesoendodermal cells that are committed to a cardiomyocyte lineage. The embryonic, pluripotential stem cell transcription factor Oct-4 positively regulates Cripto-1 expression in P19 cells, which, in turn, allows Brachyury to respond to Wnt/β-catenin signaling. This finding might be significant, because Brachyury overexpression in tumor cells leads to the acquisition of an EMT phenotype and an increase in both cell migration and invasion.56 Furthermore, Brachyury is overexpressed in various human tumor tissues and cancer cell lines, and has been associated with advanced human lung tumors.56 Because Cripto-1 has been involved in the induction of EMT in cancer, Brachyury might represent a downstream target of Cripto-1 in a feed-forward regulatory loop for controlling the induction of EMT in cancer cells.

Cripto-1 Functions in Mammary Gland Development and Tumorigenesis

The mammary gland is a unique organ because most of its developmental program occurs postnatally. During embryogenesis, mammary glands are specified early during midgestation, with the formation of the mammary milk lines on the ventral surface of the embryo at E8.5.57 Ectodermal cells along the mammary lines aggregate and form mammary placodes or buds at E10.5. The placodes invaginate into the underlying ventral dermal stroma, and epithelial bud cells proliferate and form a branching duct-like tree in the surrounding stromal mesenchyme, producing a rudimentary mammary gland. Postnatally, during puberty at the age of 3 to 4 weeks, the mammary gland begins to elongate with the formation of highly proliferative TEBs that invade the mammary fat pad and produce an extensive network of branching ducts that fill the mammary fat pad.58 During pregnancy, the mammary gland undergoes alveolar development and side branching in preparation for lactation. On cessation of lactation and during involution, the gland regresses to its previous state.

During branching morphogenesis in the postpubescent gland, cap cells at the leading edge of the TEBs exhibit features of epithelial plasticity, acquiring a transient mesenchymal-like phenotype necessary for the generation of an intricate mammary ductal tree.58 Epithelial cell plasticity of the developing mammary gland is influenced by pituitary hormones (eg, prolactin), ovarian hormones (eg, estrogen and progesterone), and locally derived growth factors [eg, amphiregulin, insulin-like growth factor-1 and insulin-like growth factor-2, hepatocyte growth factor (HGF), Wnt4 and Wnt5a, FGFs, colony-stimulating factor (CSF)-1, and TGF-β1], which can be expressed in the epithelial cells, stroma, adipocytes, mast cells, or macrophages that surround the TEBs.58 An analysis of the temporal expression of Cripto-1 in the developing mammary gland indicates that Cripto-1 is expressed during embryonic and postnatal mammary gland development. Strizzi et al40 have found that Cripto-1 can be detected in the mesenchyme surrounding the mammary placodes but not in the epithelial placodes, similar to Msx2, Lef-1, and β-catenin in the Wnt pathway, suggesting that Cripto-1 might be important for mammary mesenchymal development.

Postnatally, Cripto-1 can be detected at low levels in ductal epithelial cells of the virgin mouse mammary gland, and its expression significantly increases during pregnancy and lactation, with its expression also being detected in human milk.59–61 Cripto-1 has been localized in myoepithelial, luminal epithelial, and cap cells of the advancing TEBs of the branching ducts of the virgin mouse mammary gland.60 Because cap cells of TEBs exhibit signs of epithelial plasticity (eg, loss of apical-basal polarity) or secretion of extracellular metalloproteases, activation of Cripto-1 in these cells might contribute to the induction of epithelial plasticity, EMT, and ductal invasion into the mammary fat pad during mammary gland development.

Several studies have clearly shown that Cripto-1 is involved in regulating branching morphogenesis and EMT of the mammary gland both in vitro and in vivo. Kenney et al59 first demonstrated that recombinant Cripto-1 that had been incorporated into Elvax slow-release implants and inserted into the mammary gland of virgin mice promoted a de novo network of branching ducts migrating toward the pellet. These dramatic changes in cell morphological characteristics are also observed in vitro. For example, treatment of cobblestone-like NOG-8 mouse mammary epithelial cell colonies (Figure 3A) with recombinant Cripto-1 for several days can induce scattering of peripheral epithelial cells, which acquire more fibroblastic-like morphological characteristics (Figure 3B). In contrast, untreated control NOG8 cells continue to grow in tight colonies without showing any morphological changes (Figure 3A). In addition, overexpression of Cripto-1 in EpH4 mouse mammary epithelial cells induces the extensive formation of branching, duct-like structures that contain a lumen when grown in a three-dimensional collagen type I matrix (Figure 3, C and D).

Figure 3.

Figure 3

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Effects of Cripto-1 on morphological characteristics and branching morphogenesis of mouse mammary epithelial cells. NOG-8 mouse mammary epithelial cells were grown on plastic as colonies in the absence (A) or presence (B) of recombinant Cripto-1 protein (50 ng/mL) (R&D Systems, Minneapolis, MN) for 8 days. Cripto-1 treatment induces scattering of peripheral epithelial cells, which acquire more fibroblastic-like morphological characteristics (B). EpH4 wild-type (C) and EpH4 Cripto-1 cells (D) were grown in collagen type I matrix for 2 weeks. Cripto-1 overexpression induces the extensive formation of branching duct-like structures that contain a lumen (D) compared with EpH4 control cells (C).

Remarkably, Cripto-1–expressing mammary epithelial cells also show enhanced migration and invasion capacity in vitro compared with control mammary epithelial cells.62 However, this increase in cell motility in response to Cripto-1 overexpression is not unique to mammary epithelial cells, because it has also been demonstrated in a variety of cancer cell lines. For instance, MCF-7 breast cancer cells and Caski human cervical carcinoma cells overexpressing Cripto-1, or LS174-T colon cancer or NTERA-2/D1 EC cells that express endogenous Cripto-1, show a significant increase in their migration and invasion behaviors compared with parental cell lines.8,35,63 Interestingly, Cripto-1 overexpression in Caski cells also induces a more invasive mesenchymal phenotype, as demonstrated by the dramatic increase in vimentin mRNA and protein expression in Cripto-1–Caski cells compared with wild-type control cells.63

Transgenic mouse studies46,64 have confirmed the direct involvement of Cripto-1 in mammary gland branching morphogenesis and tumorigenesis. In fact, nulliparous transgenic female mice expressing the human Cripto-1 transgene in the mammary gland under the transcriptional control of the MMTV promoter (MMTV–Cripto-1 mice) exhibit dramatic morphological changes in the nulliparous virgin mammary gland, specifically displaying a significant increase in lateral secondary and tertiary side branching (compare Figure 4, A and B, with Figure 4, C and D, respectively). Mammary glands from MMTV–Cripto-1 mice also exhibit regions of focal proliferation (Figure 4B) that, on histological analysis, were confirmed to be intraductal epithelial hyperplasias. Aged multiparous MMTV–Cripto-1 mice exhibit pronounced mammary multifocal hyperplasias and papillary adenocarcinomas, suggesting that Cripto-1 requires additional genetic alterations to promote mammary tumor formation, because tumor formation requires multiparity and the tumor frequency is between 40% and 50% in MMTV–Cripto-1 mice.46 Remarkably, mammary tumors from MMTV–Cripto-1 transgenic mice exhibit areas of morphological changes associated with EMT.45 In fact, MMTV–Cripto-1 mammary tumors show a reduction of intercellular adhesion proteins, such as E-cadherin, and an increase in the expression of mesenchymal markers, including N-cadherin and vimentin.

Figure 4.

Figure 4

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Whole mount histological features of mammary glands from nulliparous virgin MMTV–Cripto-1 and wild-type FVB/N mice. Virgin MMTV–Cripto-1 mammary glands (A and B) exhibit more secondary and tertiary ductal side branching compared with wild-type control mammary glands (C and D). Arrows, hyperplastic regions. Original magnification: ×1 (A and C); ×5 (B and D).

Increased expression of several integrins, including integrins α3, α5, β1, β3, and β4, is also detected in MMTV–Cripto-1 mammary tumors, thereby promoting the activation of focal adhesion kinase, which leads to cell spreading. Furthermore, MMTV–Cripto-1 tumors exhibit an increased activation of c-src and Akt intracellular signaling pathways; phosphorylated and inactive glycogen synthase kinase-3β; nuclear, dephosphorylated, and active β-catenin; and enhanced levels of cyclin D1. The zinc-finger transcription factor Snail is also expressed in MMTV–Cripto-1 mammary tumors, suggesting that it might be directly involved in the down-regulation of E-cadherin observed in these lesions (Figure 2). Notably, positive staining for Snail, vimentin, N-cadherin, and cyclin D1 is generally restricted to the undifferentiated areas of the tumor that contain large undifferentiated cells with mesenchyme-like morphological characteristics.45 Markers of EMT could also be detected in uterine leiomyosarcomas that develop in approximately 20% of nulliparous or multiparous MMTV–Cripto-1 mice.65

Transgenic mice that overexpress the human Cripto-1 transgene in the mammary gland under the transcriptional control of the whey acidic protein promoter also develop mammary tumors in multiparous mice that, unlike the adenocarcinomas in MMTV–Cripto-1 mice, display a variety of histological subtypes that resemble the histological characteristics of mammary tumors that arise in transgenic mice overexpressing Wnt1 or activated forms of β-catenin.64,66,67 This finding suggests that Cripto-1 might target a bipotent stem-like or progenitor cell population in whey acidic protein–Cripto-1 mice that can produce multiple histological subtypes, as has been observed in MMTV-Wnt1 and MMTV-activated β-catenin mice.68,69

HC-11 mouse mammary epithelial cells overexpressing Cripto-1 (HC-11/Cripto-1) also undergo EMT, as shown by a decrease in E-cadherin expression and an increase in vimentin, N-cadherin, and Snail expression.45 In addition, signaling molecules that are known to be activated during EMT, such as c-src, focal adhesion kinase, and Akt, are highly phosphorylated in HC-11/Cripto-1 cells compared with HC-11 control cells. Interestingly, the c-src inhibitor PP2 inhibited activation and phosphorylation of c-src, focal adhesion kinase, and Akt in HC-11/Cripto-1 cells and strongly interfered with their migratory behaviors, indicating that activation of c-src by Cripto-1 is critical for mammary epithelial cell migration.45 Intriguingly, Netrin-1, a neural axon guidance protein and a Nodal target gene in embryonic lateral plate mesoderm, antagonized Cripto-1–induced EMT, migration, and the ability of Cripto-1 to attenuate β-casein expression in HC-11/Cripto-1 cells in response to lactogenic hormones (Table 1).80 In fact, Cripto-1 fibroblastic-like cells became more epithelial in their morphological characteristics, suggesting that Netrin-1 might promote MET in the context of Cripto-1 overexpression.80

di Bari et al38 have shown that Cripto-1 is a downstream target of the transcription factor Msx2 (Figure 2 and Table 1). Msx2, like Cripto-1, is expressed at sites of EMT during embryogenesis and in the mammary embryonic mesenchyme, and promotes branching morphogenesis of the mammary gland.81 Overexpression of Msx2 in NMuMG mouse mammary epithelial cells induces EMT through the up-regulation of Cripto-1 expression. Inhibition of Cripto-1 expression, using a specific anti-Cripto-1 small-interfering RNA or blockade of the c-src signaling pathway, converts the mesenchymal phenotype of NMuMG Msx2-expressing cells into more epithelial morphological characteristics.38 Interestingly, Msx2 is co-expressed with Cripto-1 and activated c-src in tissue sections obtained from a small cohort of infiltrating breast carcinomas, indicating that the cross talk between Msx2 and Cripto-1 might be involved in mammary tumor progression and invasion.

Furthermore, Cripto-1 has been overexpressed in a variety of human tumors and human cancer cell lines, whereas low or undetectable levels of expression are detected in normal adult tissues and in non-transformed normal cell lines (Table 2).8,65,70-79 The function of Cripto-1 in regulating EMT in cancer cells might explain the association of its expression with more aggressive behavior in several types of cancers, as shown in Table 2. Increased Cripto-1 expression and decreased E-cadherin expression have been positively associated with tumor progression and poor prognosis in patients with gastric and breast cancers.70,72 A positive correlation between high levels of Cripto-1 expression and prognostic factors (eg, tumor size, depth of invasion, lymph node metastasis, liver metastasis, and TNM stage) has been detected in gastric cancer.70 Furthermore, high levels of Cripto-1 expression and loss of E-cadherin expression represent independent prognostic factors in gastric cancer, suggesting that combined analysis of Cripto-1 and E-cadherin might have a significant value in predicting patient prognosis and in evaluating the metastatic potential of gastric cancer.70 Cripto-1 overexpression is also found in high-grade and poor-prognosis breast tumors compared with low-grade and good-prognosis breast cancers.72

Table 2.

Cripto-1 Expression in Human Cancer Cell Lines and Human Tumors

Tissue Cripto-1 mRNA and/or protein expression⁎
 Clinical correlations
Human cancer cell lines Human tumors
Bladder NA Bladder cancer8 NA
Breast MCF-7, ZR-75-1, T-47D, MDA-MB-231, SkBr3, and MDA-MB-4688,71 Breast cancer8,72,73 High histological grade, PR positive, high Ki–67 index, Nottingham Prognostic Index >5.4, invasive ductal or lobular carcinomas, decreased survival
Cervix NA Cervical carcinoma8 High tumor size, lymphovascular involvement, endometrial and parametrial extension, lymph node metastasis
Colon GEO, DLD-1, CoLo201, Sw620, LS174T, CBS, HT-29, HCT116, and LoVo8 Colon cancer8 Advanced tumor stage, high Ki-67 index, lymphovascular involvement, metachronous metastasis, decreased survival, poor prognosis
Endometrium NA Endometrial carcinoma8,74 High histological grade, advanced surgical stage, myometrial invasion, positive peritoneal washing cytology
Eye OCM-1, OCM-8, 92–1, and OMM-18 Uveal melanoma8 Extrascleral extension, liver metastasis
Gallbladder NA Gallbladder carcinoma8 Papillary and well-differentiated tumors
Lung NA Lung carcinoma8 NA
Nasopharynx NA Nasopharyngeal cancer8 Lymph node metastasis, distant metastasis, advanced clinical stage
Oral cavity SCC-15, HSC-2, HSC-3, HSC-4, HO-1-N1, KOSCC-11, KOSCC-25B, and KOSCC-33A75 Oral squamous cell carcinoma75 Moderately/poorly differentiated tumors
Ovary NIH_OVCAR3, NIH_OVCAR4, and NIH_OVCAR876 Ovarian carcinoma8 Advanced tumor stage
Pancreas NA Pancreatic cancer8 Advanced tumor stage
Prostate DU145, LNCap, and PC38,78 NA NA
Skin ROS184, CON242, COPA159, and C8168,31 Melanoma78 and basal cell carcinoma8 NA
Stomach HSC398 Gastric cancer8,70,79 Advanced tumor stage, poor prognosis, lymph node metastasis, distant metastasis, histological intestinal type, moderately differentiated tumors
Testis NTERA-2 and NCCIT8,31 Seminomas and nonseminomas (embryonal carcinomas, teratocarcinomas, and choriocarcinomas)8 NA
Uterus NA Leiomyosarcoma of the uterus65 NA
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NA, not available; PR, progesterone receptor.

⁎

As detected by using Northern blot, quantitative and semiquantitative RT-PCR, Western blot, or immunohistochemical analysis.

Nodal has been a key regulator of melanoma tumorigenicity, promoting the migration and invasion of melanoma cells.82 Nodal expression correlates with both melanoma progression and advanced breast cancer.82,83 In fact, Nodal depletion in aggressive melanoma cells decreases anchorage-independent growth and epithelial cell plasticity.82 Nodal can positively regulate the expression of the EMT regulators Twist, Snail, and Slug during mesoderm formation in Xenopus (Figure 2).28 Twist and Slug, in turn, negatively affect the expression of Cerberus, a Nodal antagonist (Figure 2).28 A recent report showed that Cripto-1 might also function in human melanomas. De Luca et al78 detected Cripto-1 expression in 43% of primary human cutaneous melanomas and in 57% of melanoma cell lines (Table 2). Notably, Cripto-1 strongly enhanced the motility of melanoma cell lines; blockade of c-src and Nodal signaling with specific inhibitors or interference of Cripto-1 expression using small-interfering RNAs produced a significant inhibition of the growth and the invasive ability of melanoma cells.78 These results suggest that Cripto-1 and Nodal might cooperate in the induction of a more aggressive phenotype in human melanomas and possibly in other tumors, such as breast cancer.

Cripto-1 in Embryonic Cells and CSCs

Stem cells can have two distinct functions: They divide either symmetrically for an uncertain period of time, maintaining their undifferentiated state (self-renewal), or asymmetrically, to generate progenitor, transamplifying cells that can differentiate into distinct cell types throughout early development and in adult tissues.84 In the developing embryo, stem cells can produce all three primary germ layers: ectoderm, endoderm, and mesoderm. In adult organisms, stem cells and progenitor cells can be found in different tissue types and function as an internal repair system, dividing to replenish specialized cells and maintaining normal turnover of regenerative organs, such as blood, skin, and intestinal tissues.84

During the past several years, a large effort has been made to identify new stem cell markers in an attempt to isolate stem cell populations, especially because of their potential as targets for therapeutic intervention. By using human ES cells, a subset of genes has been identified as potential stem cell markers, including Nanog, Oct4, connexin 43 (GJA1), and TDGF-1/Cripto-1.5 Interestingly, analysis across species has found a set of common highly conserved signaling pathways in both mouse and human ES cells, including TDGF-1/Cripto-1, Oct-4, Lefty, Nodal, Sox-2, undifferentiated embryonic cell transcription factor-1, and telomerase reverse transcriptase.5 Remarkably, Loh et al39 found that Cripto-1 is a direct downstream target gene of the ES pluripotential transcription factors Oct-4 and Nanog (Figure 2 and Table 1). Reciprocally, Activin, Nodal, and Cripto-1 are essential in initiating and maintaining the expression of Nanog and Oct-4 (Figure 2).85 Moreover, some of the ES cell markers, such as Oct-4, Sox-2, Kfl4, and c-Myc, can reprogram human fibroblasts to generate pluripotent cells, also known as induced pluripotent stem cells (iPSCs).86 Conversely, human iPSCs express ES cell genes (Sox15, SALL4, Dax1, Zfp296, FGF4, Sox10, and TDGF-1/Cripto-1), exhibit ES growth properties, and can be selectively channeled into differentiated cells from the three germ layers.5 Several mechanisms have regulated the expression of embryonic genes in iPSCs and ES cells, including DNA methylation.87 For example, 23 genes, including Sox15, SALL4, Oct-4, and TDGF-1/Cripto-1, are hypomethylated in iPSCs/ES cells and are, therefore, highly expressed.87 Reciprocally, in all somatic tissues, most cells exhibit hypermethylation of genes, such as Nanog, Oct-4, Stellar, and TDGF-1/Cripto-1.88

In addition to embryonic development and adult regenerative tissues, a subpopulation of cells that share characteristics associated with normal stem cells has been identified in human tumors. These cancer stem cells (CSCs), also called tumor-initiating cells, are identified by their ability to self-renew and initiate new tumors of a similar phenotype when injected in small numbers into immunodeficient mice.89 Moreover, CSCs are proposed to endure in tumors as a distinct population and may lead to tumor relapse and/or metastasis because they are chemoresistant and radioresistant.5,8,89 Similarities between embryonic development and cell transformation during oncogenesis have led to the identification of common signaling pathways, such as the Wnt/β-catenin, Hedgehog, and Notch pathways, suggesting that the inappropriate reactivation of developmental programs in a subpopulation of stem-like cells might drive cell transformation and tumor progression in adult tissues.5 For example, EMT is an essential developmental program that can be inappropriately activated in cancer cells and can induce the formation of migratory mesenchymal cells that possess invasive properties and metastatic potential.90

Furthermore, recent evidence suggests that adult cells that undergo EMT acquire stem cell–like properties.91,92 Al-Hajj et al93 separated tumorigenic from nontumorigenic cancer cells based on the expression of cell surface markers. A small subpopulation of cells characterized by a CD44high/CD24low phenotype was capable of generating tumors in a xenograft model more effectively than a CD44low/CD24high subpopulation of cells. Strikingly, EMT activation in immortalized human mammary epithelial cells by either TGF-β1 exposure or overexpression of the EMT-inducing transcription factors Snail or Twist induces cells to adopt a CD44high/CD24low stem cell–like profile. In addition, this population of CD44high/CD24low cells showed mesenchymal traits and stem cell properties, such as enhanced mammosphere-forming ability and differentiation into cells expressing lineage-specific markers of myoepithelial or luminal epithelial cells.93

These seminal experiments suggest that EMT is sufficient to stimulate a subpopulation of cells to acquire stem cell–like properties. Cripto-1 is an inducer of EMT during embryogenesis, and recent reports94 have shown that Cripto-1 is enriched in a subpopulation of cancer cells with stem-like characteristics, suggesting a link between CSCs and EMT in these cells. For instance, two distinct subpopulations of cells expressing high and low levels of Cripto-1 have been identified in human EC cells.94 These EC cell subpopulations, expressing different levels of Cripto-1 on the cell surface, were isolated by fluorescence-activated cell sorter analysis and demonstrated different in vitro and in vivo oncogenic abilities. The EC cell subpopulation expressing high levels of Cripto-1 formed tumor spheres in serum-free suspension cultures with a higher efficiency than the low Cripto-1–expressing EC cell fraction. In addition, both fractions of EC cells were injected s.c. into nude mice, and the Cripto-1 high EC cell expression fraction produced larger tumors and appeared with a shorter tumor latency period compared with tumors derived from the low Cripto-1–expressing EC cells. Finally, in the high Cripto-1–expressing EC subpopulation of cells, Cripto-1 expression was positively regulated by ES pluripotential stem cell transcription factors Nanog and Oct-4 and by an Smad2/3-dependent signaling pathway involving Activin, Nodal, GDF1, or GDF3.94

Similarly, activation of the Nodal/Cripto-1 signaling pathway has regulated self-renewal and in vivo tumorigenicity of pancreatic CSCs.95 Inhibition of Nodal/Activin/Cripto-1 signaling through either pharmacological inhibition or short hairpin RNA technology reversed the chemoresistance of orthotopically engrafted pancreatic CSCs to gemcitabine,95 providing a novel therapeutic approach to target pancreatic CSCs. In addition, Cripto-1 has been identified in a small subpopulation of stem-like cells in hormone-responsive and nonresponsive prostate tumor cells, and in human melanomas.5 Finally, in the bone marrow hematopoietic niche, Cripto-1 expression in the stroma was essential in regulating the lineage specification of a CD34/GRP78 myeloid progenitor population under hypoxic conditions.96 This finding might be significant, because hypoxia can directly enhance Cripto-1 expression through the binding of hypoxia-inducible factor-1α to hypoxia-responsive elements within the Cripto-1 promoter region (Figure 2 and Table 1).37

A recent report97 highlights the signaling pathways that induce and maintain an EMT program in mammary epithelial cells. The authors analyzed the proteins secreted into the culture medium of immortalized human mammary epithelial cells that either display a CD44high/CD24low cell surface marker profile or were overexpressing the transcription factor Twist using an antibody array. Interestingly, Cripto-1, together with other TGF-β family members, Wnt signaling molecules, and proangiogenic factors, was among the secreted proteins that were detected in mammary epithelial cells undergoing EMT, suggesting the activation of autocrine signaling loops during EMT. Therefore, TGF-β signaling, in collaboration with canonical and noncanonical Wnt signaling pathways, can establish cooperating autocrine and paracrine signaling loops that can induce EMT in mammary epithelial cells. However, in the normal mammary gland, these secreted growth factors fail to induce EMT because of the concomitant secretion of TGF-β/Nodal and Wnt signaling pathway inhibitors, such as Lefty, Cerberus, dickkopf1, secreted frizzled-related proteins, and BMPs.

Conclusions

This review has highlighted the importance of Cripto-1 in regulating cell plasticity and EMT during embryogenesis, mammary gland development, and cancer. Targeting molecular signals that lead to both the EMT switch in tumor cells and the emergence of dedifferentiated cancer cells with stem-like properties is a promising therapeutic strategy. Several studies have shown that pharmacological agents that target Cripto-1 in cancer have therapeutic value.98 Therefore, therapeutic strategies inhibiting the induction of EMT may reduce the formation of cancer stem-like cells and improve clinical outcome.

Footnotes

Supported by the Intramural Research Program, NIH, National Cancer Institute, Center for Cancer Research.

A guest editor acted as editor-in-chief for this manuscript. No person at Thomas Jefferson University was involved in the peer review process or final disposition for this article.

References

  • 1.Kalluri R., Weinberg R.A. The basics of epithelial-mesenchymal transition. J Clin Invest. 2009;119:1420–1428. doi: 10.1172/JCI39104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 2.Chen J., Han Q., Pei D. EMT and MET as paradigms for cell fate switching. J Mol Cell Biol. 2011;4:66–69. doi: 10.1093/jmcb/mjr045. [DOI] [PubMed] [Google Scholar]
  • 3.Peinado H., Olmeda D., Cano A. Snail, Zeb and bHLH factors in tumour progression: an alliance against the epithelial phenotype? Nat Rev Cancer. 2007;7:415–428. doi: 10.1038/nrc2131. [DOI] [PubMed] [Google Scholar]
  • 4.Micalizzi D.S., Farabaugh S.M., Ford H.L. Epithelial-mesenchymal transition in cancer: parallels between normal development and tumor progression. J Mammary Gland Biol Neoplasia. 2010;15:117–134. doi: 10.1007/s10911-010-9178-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Bianco C., Rangel M.C., Castro N.P., Nagaoka T., Rollman K., Gonzales M., Salomon D.S. Role of Cripto-1 in stem cell maintenance and malignant progression. Am J Pathol. 2010;177:532–540. doi: 10.2353/ajpath.2010.100102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Nagaoka T., Karasawa H., Castro N.P., Rangel M.C., Salomon D.S., Bianco C. An evolving web of signaling networks regulated by Cripto-1. Growth Factors. 2012;30:13–21. doi: 10.3109/08977194.2011.641962. [DOI] [PubMed] [Google Scholar]
  • 7.Heisenberg C.P., Solnica-Krezel L. Back and forth between cell fate specification and movement during vertebrate gastrulation. Curr Opin Genet Dev. 2008;18:311–316. doi: 10.1016/j.gde.2008.07.011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.de Castro N.P., Rangel M.C., Nagaoka T., Salomon D.S., Bianco C. Cripto-1: An embryonic gene that promotes tumorigenesis. Future Oncol. 2010;6:1127–1142. doi: 10.2217/fon.10.68. [DOI] [PubMed] [Google Scholar]
  • 9.Krebs L.T., Iwai N., Nonaka S., Welsh I.C., Lan Y., Jiang R., Saijoh Y., O'Brien T.P., Hamada H., Gridley T. Notch signaling regulates left-right asymmetry determination by inducing Nodal expression. Genes Dev. 2003;17:1207–1212. doi: 10.1101/gad.1084703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Morkel M., Huelsken J., Wakamiya M., Ding J., van de Wetering M., Clevers H., Taketo M.M., Behringer R.R., Shen M.M., Birchmeier W. Beta-catenin regulates Cripto- and Wnt3-dependent gene expression programs in mouse axis and mesoderm formation. Development. 2003;130:6283–6294. doi: 10.1242/dev.00859. [DOI] [PubMed] [Google Scholar]
  • 11.Wagner M., Siddiqui M.A. Signal transduction in early heart development (I): cardiogenic induction and heart tube formation. Exp Biol Med (Maywood) 2007;232:852–865. [PubMed] [Google Scholar]
  • 12.Xu C., Liguori G., Adamson E.D., Persico M.G. Specific arrest of cardiogenesis in cultured embryonic stem cells lacking Cripto-1. Dev Biol. 1998;196:237–247. doi: 10.1006/dbio.1998.8862. [DOI] [PubMed] [Google Scholar]
  • 13.Parisi S., D'Andrea D., Lago C.T., Adamson E.D., Persico M.G., Minchiotti G. Nodal-dependent Cripto signaling promotes cardiomyogenesis and redirects the neural fate of embryonic stem cells. J Cell Biol. 2003;163:303–314. doi: 10.1083/jcb.200303010. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.D'Aniello C., Lonardo E., Iaconis S., Guardiola O., Liguoro A.M., Liguori G.L., Autiero M., Carmeliet P., Minchiotti G. G protein-coupled receptor APJ and its ligand apelin act downstream of Cripto to specify embryonic stem cells toward the cardiac lineage through extracellular signal-regulated kinase/p70S6 kinase signaling pathway. Circ Res. 2009;105:231–238. doi: 10.1161/CIRCRESAHA.109.201186. [DOI] [PubMed] [Google Scholar]
  • 15.Wang B., Yan J., Peng Z., Wang J., Liu S., Xie X., Ma X. Teratocarcinoma-derived growth factor 1 (TDGF1) sequence variants in patients with congenital heart defect. Int J Cardiol. 2011;146:225–227. doi: 10.1016/j.ijcard.2009.08.046. [DOI] [PubMed] [Google Scholar]
  • 16.Liguori G.L., Echevarria D., Improta R., Signore M., Adamson E., Martinez S., Persico M.G. Anterior neural plate regionalization in cripto null mutant mouse embryos in the absence of node and primitive streak. Dev Biol. 2003;264:537–549. doi: 10.1016/j.ydbio.2003.08.023. [DOI] [PubMed] [Google Scholar]
  • 17.Liguori G.L., Echevarria D., Bonilla S., D'Andrea D., Liguoro A., Persico M.G., Martinez S. Characterization of the functional properties of the neuroectoderm in mouse Cripto(−/−) embryos showing severe gastrulation defects. Int J Dev Biol. 2009;53:549–557. doi: 10.1387/ijdb.082650gl. [DOI] [PubMed] [Google Scholar]
  • 18.Parish C.L., Parisi S., Persico M.G., Arenas E., Minchiotti G. Cripto as a target for improving embryonic stem cell-based therapy in Parkinson's disease. Stem Cells. 2005;23:471–476. doi: 10.1634/stemcells.2004-0294. [DOI] [PubMed] [Google Scholar]
  • 19.Sonntag K.C., Simantov R., Björklund L., Cooper O., Pruszak J., Kowalke F., Gilmartin J., Ding J., Hu Y.P., Shen M.M., Isacson O. Context-dependent neuronal differentiation and germ layer induction of Smad4−/− and Cripto−/− embryonic stem cells. Mol Cell Neurosci. 2005;28:417–429. doi: 10.1016/j.mcn.2004.06.003. [DOI] [PubMed] [Google Scholar]
  • 20.Lonardo E., Parish C.L., Ponticelli S., Marasco D., Ribeiro D., Ruvo M., De Falco S., Arenas E., Minchiotti G. A small synthetic cripto blocking Peptide improves neural induction, dopaminergic differentiation, and functional integration of mouse embryonic stem cells in a rat model of Parkinson's disease. Stem Cells. 2010;28:1326–1337. doi: 10.1002/stem.458. [DOI] [PubMed] [Google Scholar]
  • 21.Chu J., Shen M.M. Functional redundancy of EGF-CFC genes in epiblast and extraembryonic patterning during early mouse embryogenesis. Dev Biol. 2010;342:63–73. doi: 10.1016/j.ydbio.2010.03.009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.D'Andrea D., Liguori G.L., Le Good J.A., Lonardo E., Andersson O., Constam D.B., Persico M.G., Minchiotti G. Cripto promotes A-P axis specification independently of its stimulatory effect on Nodal autoinduction. J Cell Biol. 2008;180:597–605. doi: 10.1083/jcb.200709090. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Kruithof-de Julio M., Alvarez M.J., Galli A., Chu J., Price S.M., Califano A., Shen M.M. Regulation of extra-embryonic endoderm stem cell differentiation by Nodal and Cripto signaling. Development. 2011;138:3885–3895. doi: 10.1242/dev.065656. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Wendt M.K., Allington T.M., Schiemann W.P. Mechanisms of the epithelial-mesenchymal transition by TGF-beta. Future Oncol. 2009;5:1145–1168. doi: 10.2217/fon.09.90. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Kaartinen V., Haataja L., Nagy A., Heisterkamp N., Groffen J. TGFbeta3-induced activation of RhoA/Rho-kinase pathway is necessary but not sufficient for epithelio-mesenchymal transdifferentiation: implications for palatogenesis. Int J Mol Med. 2002;9:563–570. [PubMed] [Google Scholar]
  • 26.Azhar M., Brown K., Gard C., Chen H., Rajan S., Elliott D.A., Stevens M.V., Camenisch T.D., Conway S.J., Doetschman T. Transforming growth factor Beta2 is required for valve remodeling during heart development. Dev Dyn. 2011;240:2127–2141. doi: 10.1002/dvdy.22702. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27.Dhasarathy A., Phadke D., Mav D., Shah R.R., Wade P.A. The transcription factors Snail and Slug activate the transforming growth factor-beta signaling pathway in breast cancer. PLoS One. 2011;6:e26514. doi: 10.1371/journal.pone.0026514. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28.Zhang C., Klymkowsky M.W. Unexpected functional redundancy between Twist and Slug (Snail2) and their feedback regulation of NF-kappaB via Nodal and Cerberus. Dev Biol. 2009;331:340–349. doi: 10.1016/j.ydbio.2009.04.016. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Shukla A., Ho Y., Liu X., Ryscavage A., Glick A.B. Cripto-1 alters keratinocyte differentiation via blockade of transforming growth factor-beta1 signaling: role in skin carcinogenesis. Mol Cancer Res. 2008;6:509–516. doi: 10.1158/1541-7786.MCR-07-0396. [DOI] [PubMed] [Google Scholar]
  • 30.Gray P.C., Shani G., Aung K., Kelber J., Vale W. Cripto binds transforming growth factor beta (TGF-beta) and inhibits TGF-beta signaling. Mol Cell Biol. 2006;26:9268–9278. doi: 10.1128/MCB.01168-06. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Adkins H.B., Bianco C., Schiffer S.G., Rayhorn P., Zafari M., Cheung A.E., Orozco O., Olson D., De Luca A., Chen L.L., Miatkowski K., Benjamin C., Normanno N., Williams K.P., Jarpe M., LePage D., Salomon D., Sanicola M. Antibody blockade of the Cripto CFC domain suppresses tumor cell growth in vivo. J Clin Invest. 2003;112:575–587. doi: 10.1172/JCI17788. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 32.Gray P.C., Harrison C.A., Vale W. Cripto forms a complex with activin and type II activin receptors and can block activin signaling. Proc Natl Acad Sci U S A. 2003;100:5193–5198. doi: 10.1073/pnas.0531290100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33.Shani G., Fischer W.H., Justice N.J., Kelber J.A., Vale W., Gray P.C. GRP78 and Cripto form a complex at the cell surface and collaborate to inhibit transforming growth factor beta signaling and enhance cell growth. Mol Cell Biol. 2008;28:666–677. doi: 10.1128/MCB.01716-07. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 34.Kelber J.A., Panopoulos A.D., Shani G., Booker E.C., Belmonte J.C., Vale W.W., Gray P.C. Blockade of Cripto binding to cell surface GRP78 inhibits oncogenic Cripto signaling via MAPK/PI3K and Smad2/3 pathways. Oncogene. 2009;28:2324–2336. doi: 10.1038/onc.2009.97. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Mancino M., Strizzi L., Wechselberger C., Watanabe K., Gonzales M., Hamada S., Normanno N., Salomon D.S., Bianco C. Regulation of human Cripto-1 gene expression by TGF-beta1 and BMP-4 in embryonal and colon cancer cells. J Cell Physiol. 2008;215:192–203. doi: 10.1002/jcp.21301. [DOI] [PubMed] [Google Scholar]
  • 36.Hamada S., Watanabe K., Hirota M., Bianco C., Strizzi L., Mancino M., Gonzales M., Salomon D.S. beta-Catenin/TCF/LEF regulate expression of the short form human Cripto-1. Biochem Biophys Res Commun. 2007;355:240–244. doi: 10.1016/j.bbrc.2007.01.143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Bianco C., Cotten C., Lonardo E., Strizzi L., Baraty C., Mancino M., Gonzales M., Watanabe K., Nagaoka T., Berry C., Arai A.E., Minchiotti G., Salomon D.S. Cripto-1 is required for hypoxia to induce cardiac differentiation of mouse embryonic stem cells. Am J Pathol. 2009;175:2146–2158. doi: 10.2353/ajpath.2009.090218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38.di Bari M.G., Ginsburg E., Plant J., Strizzi L., Salomon D.S., Vonderhaar B.K. Msx2 induces epithelial-mesenchymal transition in mouse mammary epithelial cells through upregulation of Cripto-1. J Cell Physiol. 2009;219:659–666. doi: 10.1002/jcp.21712. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Loh Y.H., Wu Q., Chew J.L., Vega V.B., Zhang W., Chen X., Bourque G., George J., Leong B., Liu J., Wong K.Y., Sung K.W., Lee C.W., Zhao X.D., Chiu K.P., Lipovich L., Kuznetsov V.A., Robson P., Stanton L.W., Wei C.L., Ruan Y., Lim B., Ng H.H. The Oct4 and Nanog transcription network regulates pluripotency in mouse embryonic stem cells. Nat Genet. 2006;38:431–440. doi: 10.1038/ng1760. [DOI] [PubMed] [Google Scholar]
  • 40.Strizzi L., Mancino M., Bianco C., Raafat A., Gonzales M., Booth B.W., Watanabe K., Nagaoka T., Mack D.L., Howard B., Callahan R., Smith G.H., Salomon D.S. Netrin-1 can affect morphogenesis and differentiation of the mouse mammary gland. Journal of cellular physiology. 2008;216:824–834. doi: 10.1002/jcp.21462. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 41.Micalizzi D.S., Ford H.L. Epithelial-mesenchymal transition in development and cancer. Future Oncol. 2009;5:1129–1143. doi: 10.2217/fon.09.94. [DOI] [PubMed] [Google Scholar]
  • 42.Tao Q., Yokota C., Puck H., Kofron M., Birsoy B., Yan D., Asashima M., Wylie C.C., Lin X., Heasman J. Maternal Wnt11 Activates the Canonical Wnt Signaling Pathway Required for Axis Formation in Xenopus Embryos. Cell. 2005;120:857–871. doi: 10.1016/j.cell.2005.01.013. [DOI] [PubMed] [Google Scholar]
  • 43.Jarde T., Dale T. Wnt signalling in murine postnatal mammary gland development. Acta Physiol (Oxf) 2012;204:118–127. doi: 10.1111/j.1748-1716.2011.02283.x. [DOI] [PubMed] [Google Scholar]
  • 44.Bocchinfuso W.P., Hively W.P., Couse J.F., Varmus H.E., Korach K.S. A mouse mammary tumor virus-Wnt-1 transgene induces mammary gland hyperplasia and tumorigenesis in mice lacking estrogen receptor-alpha. Cancer Res. 1999;59:1869–1876. [PubMed] [Google Scholar]
  • 45.Strizzi L., Bianco C., Normanno N., Seno M., Wechselberger C., Wallace-Jones B., Khan N.I., Hirota M., Sun Y., Sanicola M., Salomon D.S. Epithelial mesenchymal transition is a characteristic of hyperplasias and tumors in mammary gland from MMTV-Cripto-1 transgenic mice. J Cell Physiol. 2004;201:266–276. doi: 10.1002/jcp.20062. [DOI] [PubMed] [Google Scholar]
  • 46.Wechselberger C., Strizzi L., Kenney N., Hirota M., Sun Y., Ebert A., Orozco O., Bianco C., Khan N.I., Wallace-Jones B., Normanno N., Adkins H., Sanicola M., Salomon D.S. Human Cripto-1 overexpression in the mouse mammary gland results in the development of hyperplasia and adenocarcinoma. Oncogene. 2005;24:4094–4105. doi: 10.1038/sj.onc.1208417. [DOI] [PubMed] [Google Scholar]
  • 47.Reinhold M.I., Kapadia R.M., Liao Z., Naski M.C. The Wnt-inducible transcription factor Twist1 inhibits chondrogenesis. J Biol Chem. 2006;281:1381–1388. doi: 10.1074/jbc.M504875200. [DOI] [PubMed] [Google Scholar]
  • 48.Yook J.I., Li X.Y., Ota I., Hu C., Kim H.S., Kim N.H., Cha S.Y., Ryu J.K., Choi Y.J., Kim J., Fearon E.R., Weiss S.J. A Wnt-Axin2-GSK3beta cascade regulates Snail1 activity in breast cancer cells. Nat Cell Biol. 2006;8:1398–1406. doi: 10.1038/ncb1508. [DOI] [PubMed] [Google Scholar]
  • 49.Gilles C., Polette M., Mestdagt M., Nawrocki-Raby B., Ruggeri P., Birembaut P., Foidart J.M. Transactivation of vimentin by beta-catenin in human breast cancer cells. Cancer Res. 2003;63:2658–2664. [PubMed] [Google Scholar]
  • 50.Li J., Zhou B.P. Activation of beta-catenin and Akt pathways by Twist are critical for the maintenance of EMT associated cancer stem cell-like characters. BMC Cancer. 2011;11:49. doi: 10.1186/1471-2407-11-49. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 51.Schulte-Merker S., Smith J.C. Mesoderm formation in response to Brachyury requires FGF signalling. Curr Biol. 1995;5:62–67. doi: 10.1016/s0960-9822(95)00017-0. [DOI] [PubMed] [Google Scholar]
  • 52.Arnold S.J., Stappert J., Bauer A., Kispert A., Herrmann B.G., Kemler R. Brachyury is a target gene of the Wnt/beta-catenin signaling pathway. Mech Dev. 2000;91:249–258. doi: 10.1016/s0925-4773(99)00309-3. [DOI] [PubMed] [Google Scholar]
  • 53.Martin B.L., Kimelman D. Regulation of canonical Wnt signaling by Brachyury is essential for posterior mesoderm formation. Dev Cell. 2008;15:121–133. doi: 10.1016/j.devcel.2008.04.013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 54.Liu H., Harris T.M., Kim H.H., Childs G. Cardiac myocyte differentiation: the Nkx2.5 and Cripto target genes in P19 clone 6 cells. Funct Integr Genomics. 2005;5:218–239. doi: 10.1007/s10142-005-0140-7. [DOI] [PubMed] [Google Scholar]
  • 55.Marikawa Y., Tamashiro D.A., Fujita T.C., Alarcon V.B. Dual roles of Oct4 in the maintenance of mouse P19 embryonal carcinoma cells: as negative regulator of Wnt/beta-catenin signaling and competence provider for Brachyury induction. Stem Cells Dev. 2011;20:621–633. doi: 10.1089/scd.2010.0209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 56.Fernando R.I., Litzinger M., Trono P., Hamilton D.H., Schlom J., Palena C. The T-box transcription factor Brachyury promotes epithelial-mesenchymal transition in human tumor cells. J Clin Invest. 2010;120:533–544. doi: 10.1172/JCI38379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 57.Robinson G.W. Cooperation of signalling pathways in embryonic mammary gland development. Nat Rev Genet. 2007;8:963–972. doi: 10.1038/nrg2227. [DOI] [PubMed] [Google Scholar]
  • 58.Sternlicht M.D. Key stages in mammary gland development: the cues that regulate ductal branching morphogenesis. Breast Cancer Res. 2006;8:201. doi: 10.1186/bcr1368. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 59.Kenney N., Smith G., Johnson M., Rosemberg K., Salomon D.S., Dickson R. Cripto-1 activity in the intact and ovariectomized virgin mouse mammary gland. Pathogenesis. 1997;1:57–71. [Google Scholar]
  • 60.Kenney N.J., Huang R.P., Johnson G.R., Wu J.X., Okamura D., Matheny W., Kordon E., Gullick W.J., Plowman G., Smith G.H. Detection and location of amphiregulin and Cripto-1 expression in the developing postnatal mouse mammary gland. Mol Reprod Dev. 1995;41:277–286. doi: 10.1002/mrd.1080410302. [DOI] [PubMed] [Google Scholar]
  • 61.Bianco C., Wechselberger C., Ebert A., Khan N.I., Sun Y., Salomon D.S. Identification of Cripto-1 in human milk. Breast Cancer Res Treat. 2001;66:1–7. doi: 10.1023/a:1010648923432. [DOI] [PubMed] [Google Scholar]
  • 62.Wechselberger C., Ebert A.D., Bianco C., Khan N.I., Sun Y., Wallace-Jones B., Montesano R., Salomon D.S. Cripto-1 enhances migration and branching morphogenesis of mouse mammary epithelial cells. Exp Cell Res. 2001;266:95–105. doi: 10.1006/excr.2001.5195. [DOI] [PubMed] [Google Scholar]
  • 63.Ebert A.D., Wechselberger C., Nees M., Clair T., Schaller G., Martinez-Lacaci I., Wallace-Jones B., Bianco C., Weitzel H.K., Salomon D.S. Cripto-1-induced increase in vimentin expression is associated with enhanced migration of human Caski cervical carcinoma cells. Exp Cell Res. 2000;257:223–229. doi: 10.1006/excr.2000.4881. [DOI] [PubMed] [Google Scholar]
  • 64.Sun Y., Strizzi L., Raafat A., Hirota M., Bianco C., Feigenbaum L., Kenney N., Wechselberger C., Callahan R., Salomon D.S. Overexpression of human Cripto-1 in transgenic mice delays mammary gland development and differentiation and induces mammary tumorigenesis. Am J Pathol. 2005;167:585–597. doi: 10.1016/S0002-9440(10)63000-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 65.Strizzi L., Bianco C., Hirota M., Watanabe K., Mancino M., Hamada S., Raafat A., Lawson S., Ebert A., D'Antonio A., Losito S., Normanno N., Salomon D. Development of leiomyosarcoma of the uterus in MMTV-CR-1 transgenic mice. J Pathol. 2007;211:36–44. doi: 10.1002/path.2083. [DOI] [PubMed] [Google Scholar]
  • 66.Miyoshi K., Shillingford J.M., Le Provost F., Gounari F., Bronson R., von Boehmer H., Taketo M.M., Cardiff R.D., Hennighausen L., Khazaie K. Activation of beta-catenin signaling in differentiated mammary secretory cells induces transdifferentiation into epidermis and squamous metaplasias. Proc Natl Acad Sci U S A. 2002;99:219–224. doi: 10.1073/pnas.012414099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 67.Miyoshi K., Rosner A., Nozawa M., Byrd C., Morgan F., Landesman-Bollag E., Xu X., Seldin D.C., Schmidt E.V., Taketo M.M., Robinson G.W., Cardiff R.D., Hennighausen L. Activation of different Wnt/beta-catenin signaling components in mammary epithelium induces transdifferentiation and the formation of pilar tumors. Oncogene. 2002;21:5548–5556. doi: 10.1038/sj.onc.1205686. [DOI] [PubMed] [Google Scholar]
  • 68.Lim E., Wu D., Pal B., Bouras T., Asselin-Labat M.L., Vaillant F., Yagita H., Lindeman G.J., Smyth G.K., Visvader J.E. Transcriptome analyses of mouse and human mammary cell subpopulations reveal multiple conserved genes and pathways. Breast Cancer Res. 2010;12:R21. doi: 10.1186/bcr2560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 69.Teuliere J., Faraldo M.M., Deugnier M.A., Shtutman M., Ben-Ze'ev A., Thiery J.P., Glukhova M.A. Targeted activation of beta-catenin signaling in basal mammary epithelial cells affects mammary development and leads to hyperplasia. Development. 2005;132:267–277. doi: 10.1242/dev.01583. [DOI] [PubMed] [Google Scholar]
  • 70.Zhong X.Y., Zhang L.H., Jia S.Q., Shi T., Niu Z.J., Du H., Zhang G.G., Hu Y., Lu A.P., Li J.Y., Ji J.F. Positive association of up-regulated Cripto-1 and down-regulated E-cadherin with tumour progression and poor prognosis in gastric cancer. Histopathology. 2008;52:560–568. doi: 10.1111/j.1365-2559.2008.02971.x. [DOI] [PubMed] [Google Scholar]
  • 71.Normanno N.Q.C.-F., Gullick W.J., Pedrsico M.G., Yarden Y., Wen D., Plowman G., Kenney N., Johnson G.R., Kim N., Brandt R., Martinez-Lacaci I., Dickson R.B., Salomon D.S. Expression of amphireguli, cripto-1 and heregulin in human breast cancer cells. International Journal of Oncology. 1993;2:903–911. doi: 10.3892/ijo.2.6.903. [DOI] [PubMed] [Google Scholar]
  • 72.Gong Y.P., Yarrow P.M., Carmalt H.L., Kwun S.Y., Kennedy C.W., Lin B.P., Xing P.X., Gillett D.J. Overexpression of Cripto and its prognostic significance in breast cancer: a study with long-term survival. Eur J Surg Oncol. 2007;33:438–443. doi: 10.1016/j.ejso.2006.10.014. [DOI] [PubMed] [Google Scholar]
  • 73.Bianco C., Strizzi L., Mancino M., Rehman A., Hamada S., Watanabe K., De Luca A., Jones B., Balogh G., Russo J., Mailo D., Palaia R., D'Aiuto G., Botti G., Perrone F., Salomon D.S., Normanno N. Identification of cripto-1 as a novel serologic marker for breast and colon cancer. Clin Cancer Res. 2006;12:5158–5164. doi: 10.1158/1078-0432.CCR-06-0274. [DOI] [PubMed] [Google Scholar]
  • 74.Niikura H., Sasano H., Kaga K., Sato S., Yajima A. Expression of epidermal growth factor family proteins and epidermal growth factor receptor in human endometrium. Hum Pathol. 1996;27:282–289. doi: 10.1016/s0046-8177(96)90070-2. [DOI] [PubMed] [Google Scholar]
  • 75.Yoon H.J., Hong J.S., Shin W.J., Lee Y.J., Hong K.O., Lee J.I., Hong S.P., Hong S.D. The role of Cripto-1 in the tumorigenesis and progression of oral squamous cell carcinoma. Oral Oncol. 2011;47:1023–1031. doi: 10.1016/j.oraloncology.2011.07.019. [DOI] [PubMed] [Google Scholar]
  • 76.Casamassimi A., De Luca A., Agrawal S., Stromberg K., Salomon D.S., Normanno N. EGF-related antisense oligonucleotides inhibit the proliferation of human ovarian carcinoma cells. Ann Oncol. 2000;11:319–325. doi: 10.1023/a:1008350811639. [DOI] [PubMed] [Google Scholar]
  • 77.Lawrence M.G., Margaryan N.V., Loessner D., Collins A., Kerr K.M., Turner M., Seftor E.A., Stephens C.R., Lai J., Postovit L.M., Clements J.A., Hendrix M.J. Reactivation of embryonic nodal signaling is associated with tumor progression and promotes the growth of prostate cancer cells. Prostate. 2011 doi: 10.1002/pros.21335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 78.De Luca A., Lamura L., Strizzi L., Roma C., D'Antonio A., Margaryan N., Pirozzi G., Hsu M.Y., Botti G., Mari E., Hendrix M.J., Salomon D.S., Normanno N. Expression and functional role of CRIPTO-1 in cutaneous melanoma. Br J Cancer. 2011;105:1030–1038. doi: 10.1038/bjc.2011.324. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 79.Saeki T., Salomon D.S., Normanno N., Johnson G.R., Gullick W.J., Mandai K., Moriwaki S., Takashima S., Kuniyasu M., Tahara E., Kawami M., Nishiyama M., Toge T. Immunohistochemical detection of cripto-1-, amphiregulin and trasnforming growth factor alpha in human gastric carcinomas and intestinal metaplasias. Int J Onc. 1994;5:215–223. doi: 10.3892/ijo.5.2.215. [DOI] [PubMed] [Google Scholar]
  • 80.Strizzi L., Bianco C., Raafat A., Abdallah W., Chang C., Raafat D., Hirota M., Hamada S., Sun Y., Normanno N., Callahan R., Hinck L., Salomon D. Netrin-1 regulates invasion and migration of mouse mammary epithelial cells overexpressing Cripto-1 in vitro and in vivo. J Cell Sci. 2005;118:4633–4643. doi: 10.1242/jcs.02574. [DOI] [PubMed] [Google Scholar]
  • 81.Satoh K., Ginsburg E., Vonderhaar B.K. Msx-1 and Msx-2 in mammary gland development. J Mammary Gland Biol Neoplasia. 2004;9:195–205. doi: 10.1023/B:JOMG.0000037162.84758.b5. [DOI] [PubMed] [Google Scholar]
  • 82.Topczewska J.M., Postovit L.M., Margaryan N.V., Sam A., Hess A.R., Wheaton W.W., Nickoloff B.J., Topczewski J., Hendrix M.J. Embryonic and tumorigenic pathways converge via Nodal signaling: role in melanoma aggressiveness. Nat Med. 2006;12:925–932. doi: 10.1038/nm1448. [DOI] [PubMed] [Google Scholar]
  • 83.Postovit L.M., Margaryan N.V., Seftor E.A., Kirschmann D.A., Lipavsky A., Wheaton W.W., Abbott D.E., Seftor R.E., Hendrix M.J. Human embryonic stem cell microenvironment suppresses the tumorigenic phenotype of aggressive cancer cells. Proc Natl Acad Sci U S A. 2008;105:4329–4334. doi: 10.1073/pnas.0800467105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 84.Bendall S.C., Stewart M.H., Bhatia M. Human embryonic stem cells: lessons from stem cell niches in vivo. Regen Med. 2008;3:365–376. doi: 10.2217/17460751.3.3.365. [DOI] [PubMed] [Google Scholar]
  • 85.Vallier L., Mendjan S., Brown S., Chng Z., Teo A., Smithers L.E., Trotter M.W., Cho C.H., Martinez A., Rugg-Gunn P., Brons G., Pedersen R.A. Activin/Nodal signalling maintains pluripotency by controlling Nanog expression. Development. 2009;136:1339–1349. doi: 10.1242/dev.033951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 86.Takahashi K., Tanabe K., Ohnuki M., Narita M., Ichisaka T., Tomoda K., Yamanaka S. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell. 2007;131:861–872. doi: 10.1016/j.cell.2007.11.019. [DOI] [PubMed] [Google Scholar]
  • 87.Nishino K., Toyoda M., Yamazaki-Inoue M., Makino H., Fukawatase Y., Chikazawa E., Takahashi Y., Miyagawa Y., Okita H., Kiyokawa N., Akutsu H., Umezawa A. Defining hypo-methylated regions of stem cell-specific promoters in human iPS cells derived from extra-embryonic amnions and lung fibroblasts. PLoS One. 2010;5:e13017. doi: 10.1371/journal.pone.0013017. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 88.De Carvalho D.D., You J.S., Jones P.A. DNA methylation and cellular reprogramming. Trends Cell Biol. 2010;20:609–617. doi: 10.1016/j.tcb.2010.08.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 89.Alison M.R., Lim S.M., Nicholson L.J. Cancer stem cells: problems for therapy? J Pathol. 2011;223:147–161. doi: 10.1002/path.2793. [DOI] [PubMed] [Google Scholar]
  • 90.Thiery J.P., Acloque H., Huang R.Y., Nieto M.A. Epithelial-mesenchymal transitions in development and disease. Cell. 2009;139:871–890. doi: 10.1016/j.cell.2009.11.007. [DOI] [PubMed] [Google Scholar]
  • 91.Mani S.A., Guo W., Liao M.J., Eaton E.N., Ayyanan A., Zhou A.Y., Brooks M., Reinhard F., Zhang C.C., Shipitsin M., Campbell L.L., Polyak K., Brisken C., Yang J., Weinberg R.A. The epithelial-mesenchymal transition generates cells with properties of stem cells. Cell. 2008;133:704–715. doi: 10.1016/j.cell.2008.03.027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 92.May C.D., Sphyris N., Evans K.W., Werden S.J., Guo W., Mani S.A. Epithelial-mesenchymal transition and cancer stem cells: a dangerously dynamic duo in breast cancer progression. Breast Cancer Res. 2011;13:202. doi: 10.1186/bcr2789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 93.Al-Hajj M., Wicha M.S., Benito-Hernandez A., Morrison S.J., Clarke M.F. Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci U S A. 2003;100:3983–3988. doi: 10.1073/pnas.0530291100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 94.Watanabe K., Meyer M.J., Strizzi L., Lee J.M., Gonzales M., Bianco C., Nagaoka T., Farid S.S., Margaryan N., Hendrix M.J., Vonderhaar B.K., Salomon D.S. Cripto-1 is a cell surface marker for a tumorigenic, undifferentiated subpopulation in human embryonal carcinoma cells. Stem Cells. 2010;28:1303–1314. doi: 10.1002/stem.463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 95.Lonardo E., Hermann P.C., Mueller M.T., Huber S., Balic A., Miranda-Lorenzo I., Zagorac S., Alcala S., Rodriguez-Arabaolaza I., Ramirez J.C., Torres-Ruiz R., Garcia E., Hidalgo M., Cebrian D.A., Heuchel R., Lohr M., Berger F., Bartenstein P., Aicher A., Heeschen C. Nodal/Activin signaling drives self-renewal and tumorigenicity of pancreatic cancer stem cells and provides a target for combined drug therapy. Cell Stem Cell. 2011;9:433–446. doi: 10.1016/j.stem.2011.10.001. [DOI] [PubMed] [Google Scholar]
  • 96.Miharada K., Karlsson G., Rehn M., Rorby E., Siva K., Cammenga J., Karlsson S. Cripto regulates hematopoietic stem cells as a hypoxic-niche-related factor through cell surface receptor GRP78. Cell Stem Cell. 2011;9:330–344. doi: 10.1016/j.stem.2011.07.016. [DOI] [PubMed] [Google Scholar]
  • 97.Scheel C., Eaton E.N., Li S.H., Chaffer C.L., Reinhardt F., Kah K.J., Bell G., Guo W., Rubin J., Richardson A.L., Weinberg R.A. Paracrine and autocrine signals induce and maintain mesenchymal and stem cell states in the breast. Cell. 2011;145:926–940. doi: 10.1016/j.cell.2011.04.029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 98.Bianco C., Salomon D.S. Targeting the embryonic gene Cripto-1 in cancer and beyond. Expert Opin Ther Pat. 2010;20:1739–1749. doi: 10.1517/13543776.2010.530659. [DOI] [PMC free article] [PubMed] [Google Scholar]

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