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Metastatic Cancer CellMarina Bacac and Ivan Stamenkovic
Experimental Pathology Unit, Department of Pathology, University of Lausanne,Switzerland; email: Ivan.Stamenkovic@chuv.ch, Marina.Bacac@chuv.ch
Annu. Rev. Pathol. Mech. Dis. 2008. 3:22147
First published online as a Review in Advance onSeptember 17, 2007
The Annual Review of Pathology: Mechanisms ofDisease is online at pathmechdis.annualreviews.org
This articles doi:10.1146/annurev.pathmechdis.3.121806.151523
Copyright c 2008 by Annual Reviews.All rights reserved
1553-4006/08/0228-0221$20.00
Key Words
tumor-host interactions, invasion, adhesion, proteolysis
Abstract
Metastasis is the result of cancer cell adaptation to a tissue microen-vironment at a distance from the primary tumor. Metastatic cance
cells require properties that allow them not only to adapt to a for-eign microenvironment but to subvert it in a way that is conducive
to their continued proliferation and survival. Recent conceptual andtechnological advances have contributed to our understanding o
the role of the host tissue stroma in promoting tumor cell growthand dissemination and have provided new insight into the genetic
makeup of cancers with high metastatic proclivity.
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INTRODUCTION
The ability to metastasize is a hallmark ofmalignant tumors, and metastasis is the prin-
cipal cause of death among cancer patients.It is the single most challenging obstacle to
successful cancer management and may alsobe viewed as the last frontier of cancer re-
search. Metastasis is the process whereby can-cer cells spread throughout the body, estab-
lishing new colonies in organs at a distancefrom the one where the primary tumor orig-
inated. It is well established that metastasis isa complex, multistep process, which although
considered highly inefficient from the cellularpoint of view, virtually constitutes a death sen-
tence for the patient. Its prevention and man-agement are therefore among the key goals in
clinical and basic cancer research.
There are three major routes for tu-mor dissemination: lymphatic vessels, blood
vessels, and serosal surfaces. Tumor metas-tases occurring via these three routes are re-
ferred to as lymphatic, hematogenous, andtranscoelomic, respectively. Epithelial malig-
nancies, or carcinomas, typically begin theirdissemination by the lymphatic route, with
hematogenous metastases occurring at a latertime. In contrast, bone and soft tissue tu-
mors, or sarcomas, preferentially metasta-
size by the hematogenous route, whereastranscoelomic metastasis is the property ofa relatively small group of tumors that in-
cludes mesotheliomas and ovarian carcino-mas. Because of their prevalence, lymphatic
and hematogenous metastases will provide themain focus of the present review.
A tumor cell that initiates a metastaticcolony must (a) detach from the primary
mass, (b) invade the local host tissue stroma,(c) penetrate local lymphatic and blood ves-
sels, (d) survive within the circulation, (e) be-come arrested in capillaries or venules of
other organs, (f) penetrate the correspondingparenchyma, (g) adapt to the newly colonized
milieu or subvert the local microenvironmentto suit its own needs, and (h) divide to form
the new tumor (Figure 1). Although much
has been learned regarding the properties that
such a cell requires, several key questions arestill unresolved. For example, do cells display-
ing metastatic proclivity emerge late in tu-mor progression as a result of multiple mu-
tations and selection, or are they part of thecell population that constitutes early malig-
nant growth? What role does the host mi-croenvironment play in tumor cell dissemina-
tion, and which components of tumor-stromacross talk might provide potential therapeutic
targets?
At least five functions are required for atumor cell to successfully complete the se-
quence of events outlined above. They in-clude interaction with the local microenvi-
ronment, migration, invasion, resistance toapoptosis, and the ability to induce angio-
genesis. All five functions are regulated byadhesion and proteolysis, which together pro-
vide the most fundamental molecular effectormechanisms upon which a metastatic cell re-
lies (Table 1). Adhesion and proteolysis de-termine tumor cell interaction with other cells
and with the extracellular matrix (ECM), helpcreate a path for migration, promote angio-
genesis, and both directly and indirectly trig-ger survival signals.
Transformation results in major pheno-typic changes that affect cell surface receptor
expression, cytoskeletal function, growthfactor and cytokine secretion, proteolytic
enzyme production, and the glycosyltrans-ferase and glycosidase repertoire (Figure 2)
These combined changes alter the way inwhich the transformed cell communicates
with its microenvironment and, by the sametoken, the way in which it is perceived by
normal surrounding cells. They not onlyprovide transformed cells with the ability to
disrupt the barriers that keep their normalcounterparts confined to a defined tissue
compartment, but also with the means tosubject the host tissue microenvironment to
their rules and use its resources for their own
survival, growth, and dissemination. It is in-creasingly clear that tumor cells depend upon
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Basement membranedegradation
Basement membrane
Migration
Growing metastases
Proliferation, angiogenesis,
microenvironment activation
Dormantmetastases
Collagen fibers
Detachment
Intravasation
Extravasation Circulation
Normal epithelia
In situ carcinoma
Invasive carcinoma
No proliferation
Invasion
Figure 1
Principal steps in metastasis. Transformation of normal epithelial cells leads to carcinoma in situ, which,as a result of loss of adherens junctions, evolves toward the invasive carcinoma stage. Following basementmembrane degradation, tumor cells invade the surrounding stroma, migrate and intravasate into blood orlymph vessels, and become transported until they arrest in the capillaries of a distant organ.
their microenvironment to metastasize and
that they even rely on host tissue stromal cellsto provide functions, such as a diversity ofproteolytic activity, that they themselves may
lack. Understanding tumor-host interactionsmay therefore provide a key to understanding
metastasis. A more recently emerging view,based on gene expression profile analysis of
diverse primary and metastatic tumors, is
that metastatic cells may constitute part ofthe early makeup of a malignant tumor. Thisreview highlights our current understanding
of tumor cell properties and host tissueresponses whose combination culminates in
cancer metastasis and discusses the origin ofmetastatic cells in light of recent observations.
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Table 1 Summary of adhesion molecules and proteolytic enzymes discussed in the text that are implicated in tumor
metastasis on the basis of experimental evidence
Mechanism Candidate functions Reference(s)
Adhesion
Cadherins
E-cadherin Promotes cell-cell adhesion; prevents cell detachment; cleaved by MMP-3, MMP-7,
and ADAMs; tumor suppressor
(1, 2, 4, 5)
N-cadherin Promotes migration and invasion; regulates activation of FGFRs (19, 20)Integrins
21, 31 Enhance metastasis in selected experimental models (28, 29)
v3 Promotes migration and invasion; mediates tumor cell-platelet interactions (30, 123)
64 Promotes migration, invasion, and proliferation; cooperates with RTKs (31, 32)
41 Mediates tumor cell adhesion to endothelial cells (126)
Immunoglobulin superfamily
VCAM-1 Mediates tumor cellendothelial cell adhesion (126)
L1 Promotes tumorigenicity and motility (55)
NrCAM Promotes tumorigencity and migration (56)
NCAM Its downregulation is associated with enhanced lymph node metastasis in some tumor
models
(57)
Selectins Mediate tumor cellendothelial interactions and tumor cellplatelet/leukocyte adhesion (119122)
Cell surface proteoglycans
CD44 Mediates interaction with hyaluronan; interacts with RTKs; specific isoforms provide a
scaffold for the assembly of molecular complexes that can promote metastasis
(5860, 62, 63)
Proteolysis
Matrix metalloproteinases
MMP-1 Degrades collagen; promotes invasion (79, 85)
MMP-2 Activates growth factors, including TGF-; promotes invasion; interacts with v3
integrin on the cell surface
(79, 85)
MMP-3 Promotes tuumorigenesis; activates growth factors, including HB-EGF (79, 85, 93)
MMP-7 Promotes cell survival; activates HB-EGF; interacts with CD44 on the cell surface (63, 79, 85)MMP-9 Promotes invasion; enhances angiogenesis; promotes intravasation; activates TGF-;
interacts with CD44 on the cell surface
(62, 79, 85)
MMP-14, -15, -16 Degrade native basement membrane; promote invasion (7779, 85)
Cathepsins Promote tumor growth and invasion; may be expressed by tumor cells or exclusively by
stromal cells
(74)
FUNDAMENTAL MOLECULAREFFECTOR MECHANISMSOF METASTASIS
Adhesion
Detachment, cadherins, and epithelial-
to-mesenchymal transition. Interepithelial
cell interactions are regulated by complex ad-hesion mechanisms, including tight and ad-
herens junctions and desmosomes (1). Trans-
formed and malignant cells that become de-tached from the epithelium display loss of ad-
herens junctions, which, in epithelial cells
are constituted primarily by E-cadherin (12). Like other members of the cadherin fam-ily, E-cadherin displays homophilic binding
specificity. Its adhesive functions are stabi-lized by-catenin, which binds to its cyto-
plasmic domain, providing a link to-cateninand the actin cytoskeleton (3). Experiments
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performedinvitroandinvivohaveshownthat
genetic and antibody-mediated inhibition ofE-cadherin function can alter the phenotype
of epithelial cells from noninvasive to invasive
(4). Conversely, introduction of E-cadherininto E-cadherin-deficient invasive carcinoma
cells abrogated their invasiveness (3). Down-regulation of E-cadherin activity in vivo us-
ing a dominant negative E-cadherin constructin the transgenic Rip-Tag mouse model of
pancreatic islet cell carcinoma resulted in thetransition of a well-differentiated -cell ade-
noma to an invasive carcinoma (5). Crossingthe Rip-Tag mice with mice that maintain
E-cadherin expression in transformed -cellsarrested tumor progression in the adenoma
stage (5). Together, these observations arguethat E-cadherin functions as a tumor suppres-
sor.Although loss of function mutations occur
in E-cadherin, they are not commonly ob-served in malignant tumors (3). Mechanisms
that decrease or abrogate E-cadherin func-tion in carcinoma cells include transcriptional
repression, followed by promoter methyla-tion (6, 7), disruption of cytoskeletal connec-
tions, increased intracellular degradation, andproteolytic cleavage of the extracellular do-
main by matrix metalloproteinases (MMPs)(3). Several of the transcriptional repressors
that control E-cadherin expression in devel-opment, including Snail, Slug, SIP1, Twist,
dEF1, and E12/E47 (812), are implicatedin E-cadherin downregulation in malignant
cells. An alternative mechanism that can in-activate E-cadherin function in tumor cells
is disruption of the link between E-cadherin
and the cytoskeleton. One example is pro-vided by mutations in -catenin that abro-
gate its binding to -catenin and result in anonadhesive phenotype (3). Coordinate re-
ceptor tyrosine kinase (RTK)-integrin sig-naling can also interfere with E-cadherin
function. Activated RTKs and Src family ki-nases induce tyrosine phosphorylation of the
E-cadherin-catenin complex, which is thenrecognized by the Cbl-like E3 ubiquitin lig-
ase and downregulated by endocytosis (13). In
Selectin ligands
CD44v3
MMP-7
N-cadherin
pro-HB-EGF
Glycosaminoglycanchains
ErbB4
MMP-14
FGFR
c-Met
c-Met
MMP-15
CD44inactive
E-cadherin
5 1 3
v
6
1
3 4
BM
Extracellular matrix
ErbB2
Extracellular matrix
Figure 2
Changes of adhesive properties in transformed cells. Normal epithelial cellcommunication with its microenvironment is regulated byE-cadherin-mediated cell-cell interaction and 1-integrin-mediatedadhesion to the basement membrane (BM). Transformation results in thecadherin switch that leads to E-cadherin loss and replacement byN-cadherin, which plays an important role in invasion by regulatingfibroblast growth factor receptor (FGFR) function. Carcinomas frequentlyexpress v3 and 64 integrins, which promote invasion andproliferation, in part through their interactions with receptor tyrosinekinases, including ErbB2 and Met. Transformation also results in changes
in glycosylation of cell surface proteoglycans such as CD44, whichregulates invasion by coordinating MMP-7-mediated heparin-bindingepidermal growth factor (HB-EGF) activation and ErbB4 signaling.Changes in the glycosyltransferase repertoire can also result in thedecoration of cell surface receptors by oligosaccharides that constituteselectin ligands. Several transmembrane MMPs are expressed by tumorcells and mediate BM degradation.
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v-src-transformed cells, this process is de-
pendent upon integrin signaling and fo-cal adhesion kinase (FAK) phosphorylation
(14, 15). Integrin signaling operates through
Snail/Slug to suppress E-cadherin expres-sion and disrupt adherens junctions. In some
epithelial cells, this process may be medi-ated by integrin-linked kinase (16, 17). Fi-
nally, proteolytic cleavage of the extracellu-lar domain of E-cadherin by MMP-3 and
MMP-7 disrupts its ability to promote cellularinteractions (18). Abrogation of E-cadherin-
mediated cell-cell adhesion results in detach-ment of tumor cells from the epithelial cell
layer and affects signaling pathways impli-cated in cell migration and growth, includ-
ing Rho GTPase-mediated modulation of theactin cytoskeleton and the canonical Wnt sig-
naling pathway (3).Loss of E-cadherin in malignant cells
may be replaced by other cadherins, mostcommonly, N-cadherin (Figure 2). This
process, known as the cadherin switch, isassociated with a phenotypic change observed
in vitro known as epithelial-to-mesenchymaltransition (EMT). EMT, defined as the
conversion of epithelial cells to motile,fibroblast-like cells that express mesenchymal
rather than epithelial cell markers, is acommon event during normal embryonic
development and is observed as epithelial cellprogress through the stages of carcinogenesis
in vitro. It is proposed to reflect invasive andmetastatic properties of transformed epithe-
lial cells, but remains somewhat controversialbecause full EMT is difficult to prove in
vivo. Nevertheless, expression of N-cadherin
may make a critical contribution to invasionthrough both its adhesive and signaling
functions. N-cadherin can mediate cell-cellinteractions with N-cadherin-expressing
stromal cells, which may play an importantrole in the ability of tumor cells to direct host
responses. N-cadherin binds to and regulatesthe activation of fibroblast growth factor
receptors (FGFRs), thereby helping assemblethe FGFR-signaling complex, which triggers
downstream signaling pathways, including
phospholipase C-, phosphatidylinosito
3-kinase, and mitogen-activated protein ki-nase (MAPK). The combined action of these
signaling pathways promotes cell survivalmigration, and invasion (19, 20). Similar to
E-cadherin, the extracellular domain of N-cadherin is susceptible to proteolytic cleavage
by MMPs. The N-cadherin cleavage productcan block N-cadherin-mediated cell-cell ad-
hesion, but can also stimulate FGFR signalingon adjacent cells in paracrine fashion (19
20). Cleavage of N-cadherin at a site withinthe transmembrane or cytoplasmic domain
by a -secretase-type protease results in thetranslocation of its carboxy-terminal segment
to the nucleus, where it represses transcrip-tion mediated by the CREB-binding protein
(21).
Interaction with the extracellular matrix
integrins. Tumor cell interactions with theECM are mediated primarily by integrins and
play a key role in tumor invasion and spreadIntegrins form a large family of adhesion re-
ceptors, each member consisting of an anda transmembrane chain (22). In mammals
18 and8 chainsassociateinvariouscombi-nations to give rise to 24 integrins that recog-
nize distinct ECM ligands, with, neverthelesssome degree of overlap (22). When integrins
bind to ECM, they aggregate in the planeof the cell membrane and associate with a
molecular complex composed of adaptor, sig-naling, and cytoskeletal proteins that orches-
trate the organization of actin filaments. Or-ganization of actin filaments into stress fibers
in turn, promotes further integrin aggrega-tion that results in increased ECM binding in
a positive feedback loop. The outcome is anintegrin-mediated assembly of ECM and cy-
toskeletal protein clusters on each side of thecell membrane known as focal adhesions and
ECM contacts (23).Integrins trigger both mechanical and
chemical signals that organize and remodelthe cytoskeleton of the cell, regulate adhesive
versus migratory interactions with the ECM
impart polarity, and control proliferation
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and survival. To exert their effects, integrins
cooperatewith RTKs, thereby jointly control-ling survival and mitogenic pathways. A re-
ductionist view might be that integrins me-
diate cell adhesion and impose positionalcontrol on RTK activity, which together de-
termine whether cells migrate and proliferatein response to cytokines and growth factors.
Although seemingly straightforward, integrinimplication in cellular functions is compli-
cated by their diverse adhesive and signalingproperties that provide them the ability to
affect cell function in a variety of ways thatare often context dependent. Thus, 21 and31 integrins mediate epithelial cell adhe-sion to basal lamina and maintain them in
a quiescent state. These integrins are oftendownregulated in carcinomas and their re-
expression in carcinoma cells can decrease oreven revert the malignant phenotype (2427).
However, both of these integrins can enhancemetastasis in selected experimental models
(28, 29), underscoring the cell context and tu-mor stage dependence of the effects of1 in-
tegrins. By contrast, v3 (30) and 64 (31)integrins are frequently upregulated in car-
cinomas, where they may promote migration,invasion, and proliferation. In addition to par-
ticipating in hemidesmosome organization,the 64 integrin cooperates with epider-
mal growth factor receptor (EGFR), ErbB2,and Met, and is likely to promote the growth
of carcinomas in which activating mutationsof the corresponding growth factor receptor
genes represent the oncogenic driving force(3133). In support of this view, introduction
of the 4 chain into 4-negative breast car-
cinoma cells activates the phosphatidylinosi-tol 3-kinase pathway, which results in activa-
tion of Rac and increased invasiveness of thesecells in vitro (31). The cytoplasmic domain
of 4 also acts as an adaptor and amplifierof proinvasive signals induced by the hepato-
cyte growth factor receptor Met in cells un-dergoing Met-mediated transformation (33).
Both EGF and Met induce phosphorylationof4 and enhance SHC signaling, which dis-
rupts hemidesmosomes and increases epithe-
lial cell migration and carcinoma cell invasion
(32). RTKs may therefore augment the sig-naling functions of 64 at the expense of
its ability to mediate stable adhesion. Simi-larly, cooperation betweenv3 and platelet-
derived growth factor receptor (PDGFR) mayenhance growth and migration of tumor cells
overexpressing PDGF (34). By altering theirintegrin repertoire, neoplastic cells can hone
part of the molecular machinery that under-lies adhesion, migration, survival, and growth
to optimally serve their needs (Figure 2).Upon clustering in focal adhesions, inte-
grins activate several protein tyrosine kinases,central among which is FAK (35, 36). The
adaptor proteins paxillin and talin mediate in-tegrin interaction with FAK, which coordi-
nates many of the key events that constitute
or are related to integrin signaling (36). Acti-vation of FAK is initiated by autophosphory-
lation at Tyr397, which results in a structuralmotif recognized by SH2-domain-containing
proteins, including Src. Binding to FAK acti-vates Src, resulting in phosphorylation of ad-
ditional FAK residues and recruitment of sev-eral signaling adaptor and effector proteins,
guanine nucleotide exchange factors, GTPaseactivating proteins, cytoskeletal adaptors, and
proteolytic enzymes (23, 36, 37). The N-terminal domain of FAK provides a signal-
ing linkage between integrins and RTKs,especially EGFR and PDGFR (38). FAK
thus constitutes a platform for the coordi-nated growth-factor-receptorintegrin signal
exchange, regulation of Rho GTPase activ-ity, and focal complex turnover. Binding of
the GRB2 adaptor protein to FAK generatesan important link to the activation of Ras
and the MAPK/extracellular-signal-regulatedkinase-2 (ERK2) cascade. ERK2 phospho-
rylation can modulate focal contact dynam-ics in motile cells in addition to promoting
both proliferation and survival. Recruitmentof guanine nucleotide exchange factors, in-
cluding p190RhoGEF, may provide a directlink to RhoA activation (39) and provide reg-
ulation of promigratory versus adhesive inter-actions with substrate. Cytoskeletal adaptors,
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including ezrin, and proteolytic enzymes,
including calpain, regulate intracellular link-age of focal contacts to the actin cytoskeleton
and focal contact turnover (40).
Cell migration induced by EGF or PDGFrequires FAK association with both RTKs
and integrin-containing focalcomplexes,con-sistent with the notion that FAK can inte-
grate promigratory signals from integrins andRTKs. These signals culminate in the activa-
tion of Rho GTPases and downstream effec-tors of the MAPK pathway, including ERK
and Jun-amino-terminal kinase ( JNK) (23).ERK and JNK participate in regulating cell
migration by phosphorylating and activatingthe myosin light chain kinase, which induces
contraction of actomyosin fibers (41). JNKin-duces phosphorylation of paxillin, which may
participate in cell migration by facilitating fo-cal adhesion turnover (42). Rho GTPases ac-
tivated by FAK include Rho, Rac, and Cdc42(43, 44). Cdc42 and Rac have both been
implicated in carcinoma invasion, as theypromote actin polymerization at the leading
edge and, consequently, formation of filopo-dia and lamellipodia, respectively (43). Both
GTPases activate the ARP2/3 complex andinduce actinfilament assembly coordinated by
the Wiskott-Aldrich syndrome protein (45).They also activate p21-activatedkinase, which
enhances actin polymerization by activatingLIM kinase. Whereas Cdc42 and Rac pro-
mote actin polymerization at the leading edge,Rho orchestrates the assembly and contrac-
tion of actomyosin fibers, which pulls thetrailing edge forward during migration. At
least two Rho effector molecules, Rho kinase
(ROCK) and mammalian diaphanous, func-tion jointly to induce the assembly of acto-
myosin fibers (46). By inhibiting myosin lightchain phosphatase, ROCK promotes myosin
light chain phosphorylation and actomyosinfiber contraction (47). Rho-ROCK signaling
appears to regulate several aspects of carci-noma dissemination and is required for cancer
cells to invade three-dimensional matrices byamoeboid movement (48). Gene expression
profile comparison between melanoma cells
with low and high colony-forming abilities
in the lung in experimental metastasis assaysidentified RhoC as one of the most robustly
upregulated genes in the highly metastaticvariants (49).
Although FAK is commonly associatedwith coordinating migration signals, there
is abundant evidence to suggest that it canalso promote invasion in both normal and
neoplastic cells (50, 51). Malignant cells fre-quently display elevated FAK levels and activ-
ity(50),which is associated with shape changepodosome formation, and induction of in-
vadopodia (52). In experimental models, theinvasive tumor phenotype was associated with
the accumulation of FAK-Src signaling com-plexes within invadopodia, specialized cel
protrusions enriched in integrins and MMPs
Consistent with these observations, overex-pression of FAK in some tumor cell types was
reported to induce invasion (53, 54). Inter-estingly, the invasion-promoting property of
FAK appears to be distinct from its ability toinduce migration (52).
Immunoglobulin superfamily adhesion
receptors. Adhesion receptors belonging tothe immunoglobulin superfamily have been
implicated in the progression of some carci-nomas. Theadhesion receptorL1 is highlyex-
pressed at the invasive front of colorectal can-cers. L1 is a direct Wnt/-catenin signaling
pathway target in colorectal cancer cells, andin experimental model systems has been ob-
served to enhance motility and tumorigenicity(55).
Similar to L1, the neuronal cell adhe-sion receptor NrCAM, which is also a tar-
get of Wnt signaling, promotes tumorigenic-ity and migration in various tumor cell types
(56). By contrast, downregulation of neuralcell adhesion molecule is associated with en-
hanced lymph node metastasis in the Rip-Tagtransgenic mouse model of pancreatic islet
cell carcinogenesis (57). Like N-cadherinL1 and neural cell adhesion molecule bind
to and activate FGFRs in neurons and tu-
mor cells, thereby participating in modulating
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integrin-mediated cell adhesion to and migra-
tion on the ECM (20).
Cell surface proteoglycan function
changes. Transformation may also result inthe activation and differential glycosylation
of cell surface proteoglycans. One suchproteoglycan relevant to tumor metastasis
is CD44, the principal cell surface receptorfor hyaluronan. In addition to mediating cell
attachment to hyaluronan-coated surfacesand participating in hyaluronan metabolism,
CD44, by virtue of its structural polymor-phism and facultative decoration with a
variety of glycosaminoglycans, can serveas a multipurpose cell surface scaffold that
orchestrates the assembly of complexescomprising various classes of molecules (58).
Thus, CD44 associates with and promotesoligomerization of the ErbB family of
growth factor receptors (59) and Met (60).It also localizes various MMPs and some of
their substrates to the cell surface (6163).Its cytoplasmic domain interacts with the
ezrin-radixin-moesin family of cytoskeletalinteractors (64), including merlin, which is
implicated in tumor metastasis (65). Themechanisms whereby CD44 promotes tumor
invasion and dissemination include enhance-ment of cell migration (66), coordination of
proteolytic activity on the cell surface (62,
63), and enhancement of survival (62, 63, 67).Although it is expressed in most carcinomas,
CD44 promotes invasion and metastasis ina selective manner. By analogy to integrins,
CD44 may be expressed in an inactive confor-mation, and its activation requires, at the very
least, partial desialylation of the extracellulardomain and a critical cell surface expression
density (6871). CD44 appears to be consti-tutively active in fibroblasts and mesenchymal
progenitor cells, and to regulate migrationand adhesion of both cell types. Consistent
with this observation, as well as observationsin mouse model systems, sarcomas may
retain CD44 function and display at leastpartial dependence upon its expression for
migration, invasion, and metastasis (72).
Proteolysis, Invasion,and Tumor-Host Interactions
Following several rounds of division within
the epithelial compartment, generating acarcinoma in situ, malignant cells disrupt the
basement membrane (BM), allowing themto come into direct contact with structural
and cellular components of the stroma. Thisphase can be subdivided into several events
key for subsequent tumor dissemination,including (a) expression by the tumor cells
of the proteolytic arsenal required to de-grade the BM; (b) interaction with stromal
fibroblasts and modification of their functionto better serve the requirements of tumor
cell growth, migration, and survival; (c)
recruitment of leukocytes that may amplifythe stromal reaction and further facilitate
tumor cell dissemination; (d) angiogenesis;and finally (e) intravasation (Figure 3). The
stromal reaction to invading tumor cells isvariable, depending in part upon tumor cell
properties and in part upon the local stromalcomposition. Tumor cells typically produce
mediators that can initiate local stromalcell activation, leading to ECM remodeling
and recruitment of additional stromal cellpopulations, which provides permissive
conditions for tumor growth. The combined
proteolytic machinery of the tumor and acti-vated stromal cells degrades ECM proteins,uncovering cryptic sites that may display pro-
migratory properties and releasing se-questered growth and survival factors,
including insulin-like growth factor-1, trans-forming growth factor- (TGF-), PDGF,
vascular endothelial growth factor (VEGF),fibroblast growth factor, and hepatocyte
growth factor, thereby augmenting theirbioavailability (73).
Recent assessment of the expression pro-file of stroma associated with invasive can-
cer in a transgenic model of multistage car-cinogenesis of the prostate revealed a gene
signature reminiscent of that associated withwound healing (74). Importantly, genes that
were upregulated in the reactive stroma were
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BM
Normal epithelia
VEGFR3
Fibroblast
Monocyte/TAM
Granulocyte
Blood vessel
TGF-
Proliferation/differentiation
EMT/invasion
Recruitment,
proliferation,
and activation
HPCs and EPCs
Release ofsequestered
growth factors
IGF-1, TGF-, PDGF,VEGF, bFGF, HGF/SF
Tumor cell
CAF
Activatedfibroblast
ECM remodeling
MMP-9, uPA,cathepsins
VEGF-A
VEGF-C, VEGF-D
Lymphatic vessel
Lymphangiogenesis
Angiogenesis
EPC recruitmentSDF-1
VEGFR1
Collagen
fibers
Figure 3
Tumor-host interactions. Basement membrane (BM) degradation allows tumor cells to enter into contactwith stromal fibroblasts and alter their phenotype toward that of myofibroblasts (CAFs). Invading tumor
cells secrete numerous growth factors that stimulate angiogenesis, including VEGF-A, which binds toVEGFR1 on endothelial hematopietic precursor cells (HPCs) and endothelial precursor cells (EPCs), aswell as on monocytes/tumor-associated macrophages (TAMs), resulting in their recruitment andactivation. Tumor cells also secrete VEGF-C and -D, which bind and activate VEGFR3 on lymphaticendothelial cells and stimulate lymphangiogenesis. Activated CAFs, TAMs, and tumor cells secretenumerous extracellular matrix (ECM)-degrading enzymes, including matrix metalloproteinases (MMPs),cathepsins, and uPA, whose combined activity releases numerous ECM-sequestered growth factors thatfurther stimulate stromal fibroblast proliferation and tumor cell invasion.
found to have predictive value for both overall
and metastasis-free survival of prostate andbreast carcinoma patients. Several of these
genes were found to be upregulated in tumor-associatedstromal remodelingin studies using
different approaches to address tumor-hostinteractions (75, 76). Together these obser-
vations strongly suggest that the stromal re-sponse to primary carcinoma growth may
hold the key to subsequent development anddissemination.
Disruption of the basement membrane
The first barrier to invasion by carcinoma
cells is constituted by the BM. Conventional
wisdom would have it that a broad range ofMMPs can degrade the various components
of the BM. However, most of the evidencesupporting this view was derived from studies
of tumor invasion of matrigel, which iscomposed of denatured BM but does not
recapitulate its native structure. Recent workhas provided direct evidence that proteolytic
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degradation of the BM is mediated by trans-
membrane MMPs (MT-MMPs), includingMT1-, MT2-, and MT3-MMP (MMP-14,
-15, and -16, respectively), all three of which
are commonly expressed by cancer cells(77). Importantly, the ability to degrade
native BM has been shown to be restrictedto these three MMPs. Neither soluble nor
artificially membrane-tethered MMP-2 andMMP-9, which are commonly associated
with tumor invasion, were able to promotecell penetration of the BM (77). The same
held true for several other secreted MMPs,providing the first clear evidence that native
BM degradation by tumor cells is dependentupon MT-MMP-mediated proteolysis (77).
MT-MMPs display collagenase activityand are capable of degrading collagen IV,
which is a major constitutent of the BM.Their ability to degrade other collagens
and numerous other ECM componentssuggests that MT-MMPs may be implicated
in events that occur beyond BM invasion.Consistent with this view, experiments using
three-dimensional collagen gels showed thatMT1-MMP expression provides tumor cells
with a growth advantage in vitro and in vivo(78). The replicative advantage conferred by
MT1-MMP requires pericellular ECM pro-teolysis, as proliferation is abrogated in tumor
cells suspended in protease-resistant collagengels. In the absence of proteolysis, tumor
cells embedded in physiologically relevantECM matrices adopt a spherical config-
uration and fail to display shape changesand cytoskeletal reorganization required for
three-dimensional growth. These obser-
vations suggest that MT1-MMP regulatesproliferation by controlling cell geometry
within the the three-dimensional ECM (78).In addition to these essential roles in
the early steps of tumor metastasis, MT1-MMP activity provides the principal source
of MMP-2 activation (79) and promotes an-giogenesis (80) by degrading the fibrin ma-
trix that surrounds newly formed blood ves-sels, facilitating endothelial cell penetration of
tumor tissue (81). Consistent with their role
in invasion, MT-MMPs have been observed
to colocalize with integrins to invadopodia(82). MMP-14 has also been shown to bind
and cleave the extracellular domain of CD44,which helps detach tumor cells from the ECM
and promotes migration (83, 84).
Proteolytic events within the extracellular
matrix. The evidence that MT-MMPs me-diate tumor cell disruption of the BM as well
as subsequent ECM invasion raises the ques-
tion as to what role secreted MMPs play intumor metastasis, particularly because the ma-
jority of them appear to be supplied by stro-malcells(79). Several linesof evidencesuggest
that at least some of the secreted MMPs mayhave a robust tumor-initiating effect (79, 85).
However, secreted MMPs may also partici-pate in tumor dissemination (79, 85). Secreted
MMPs can interact with cell surface adhesionreceptors and proteoglycans, leading to co-
operation between adhesive and proteolyticmechanisms (79, 85), by analogy to the well-
established functional relationship betweenintegrins and the urokinase receptor (86).
Hyaluronan-dependent association betweenCD44 and MMP-9, which is cell-context de-
pendent, has been shown to promote pro-teolytic activation of latent TGF- relevant
to both inflammation (87) and cancer (62).In a mouse model of mammary carcinoma
dissemination, the functional CD44/MMP-9/TGF- complex promoted angiogenesis
and survival of metastatic tumor cells (62, 67).Similarly, the cell surface complex comprising
CD44HSPG, proheparin-binding epider-mal growth factor (HB-EGF), MMP-7, and
ErbB4 promotes both normal and malignantcell survival by facilitating MMP-7-mediated
HB-EGF activation and its engagement ofErbB4 (Figure 2) (63). The sum of these ob-
servations suggests that secreted MMPs teth-ered to the surface of stromal or tumor cells
may indirectly participate in metastasis by ac-tivating relevant growth factors, promoting
angiogenesis, and probably further disrupting
structural ECM barriers to invading cells.
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Interaction with fibroblasts and soluble
regulators of tumor-host cross talk. Hav-ing crossed the disrupted BM, tumor cells for
the first time find themselves in direct contact
with stromal fibroblasts. Tumor-derived anddegraded ECM-released growth factors,
including PDGF and TGF-, alter thefibroblast phenotype to one reminiscent
of myofibroblasts (Figure 3) (88). Thesetumor-conditioned stromal fibroblasts are
referred to as carcinoma-associated fibrob-lasts (CAFs), and their contribution to tumor
initiation and growth is now well established(8991). However, the mechanisms whereby
CAFs promote tumor progression are onlybeginning to emerge. CAFs are an abundant
source of proteolytic enzymes, includingMMPs and cathepsins (74, 92, 93), which
may stimulate tumor cell growth and inva-sion at both primary and secondary sites.
Increased deposition of collagen I and IIIas well as de novo expression of tenascin C
may provide additional signals that facilitatetumor invasion and metastasis; by secreting
chemokines such as monocyte chemotacticprotein-1 and cytokines such as interleukin-1
(IL-1), CAFs participate in regulating theinflammatory response to tumor invasion. In
addition, CAF-derived stromal-cell-derivedfactor-1 (SDF-1) has been shown to mediate
bone marrowderived endothelial cell pre-cursor recruitment and to directly increase
tumor cell proliferation (94).Among the soluble factors implicated in
coordinating tumor-host cross talk, TGF-plays a leading role. Although TGF- in-
hibits proliferation of normal epithelial cells
and carcinoma cells at early stages of progres-sion, it stimulates fibroblast growth and ECM
secretion and promotes late-stage carcinomainvasiveness (88). Both transgenic models
(95) and experimental metastasis assays haveshown that TGF- can enhance dissemina-
tion of at least some carcinomas (67). Accord-ingly, soluble TGF-receptor fusion proteins
were observed to reduce metastatic growthof tumor cells injected into immunocompro-
mised mice (96). TGF- can induce EMT
in tumor cells resistant to its cytostatic ef-
fects and, as already discussed, may be a majorplayer in the activation of normal fibroblasts
to display tumor-promoting functions (88).
Migration. ECM remodeling and the pres-ence of activated stroma fibroblasts create
conditions favorable for tumor cell migra-tion. Although the molecular mechanisms
that underlie migration are reasonably wellunderstood (see above), the question as to
how tumor cells actually migrate in a three-
dimensional structure has only recently beenaddressed. Real-time imaging of invading tu-
mor cells in three-dimensional collagen gelshasgiven rise to some surprising observations
Single tumor cells that had detached fromthe original tumor mass were shown to dis-
play two possible migration patterns. Mes-enchymal cells, or malignant epithelial cells
that had undergone EMT, migrate along aclassical scheme that includes protrusion of
the leading edge, formation of focal contactswith the ECM, recruitment of surface pro-
teases to ECM contacts resulting in localizedproteolysis, Rho-mediated contraction of ac-
tomyosin leading to cell contraction, and fi-nally detachment of the trailing edge. How-
ever, other malignant cells display amoeboidmovement through collagen gels. This type
of movement relies on cell deformability andrelatively weak interactions with the ECM
Movement is generated by cortical filamen-tousactin, whereasfocalcontacts,stress fibers
and localized proteolysis at cell-ECM con-tacts are lacking (97). Cells displaying amoe-
boid movement typically circumscribe but donot degrade collagen fibers (97). Although
thistype of movementcharacterizes lymphoidcells, carcinoma cells can adopt it as well
(97).Tumor cells can also migrate in groups
or aggregates (Figure 2). Here again twotypes of movement have been described. One
consists of chain migration, where cells followeach other in single file and form a chain-like
image. It is displayed by neural crest cells
(98) and normal myoblasts (99), but can also
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be observed in melanomas. Lobular invasive
breast carcinoma as well as ovarian carcinomacells often display a chain-type arrangement.
The second type of group movement of
malignant cells is referred to as collectivemigration and invasion and mimics a well-
described phenomenon that occurs duringdevelopment. Following neural tube closure,
cells in the blastoderm or ectoderm migrate insheets(100), andsimilar migration is observed
during branching morphogenesis of mam-mary glands and ducts (101). Malignant cells
also have the ability to aggregate and migrateas a functional unit (97, 102). In contrast to
individual migrating cells, cell-cell adhesionthat occurs in cell aggregates leads to a specific
form of cortical actin filament assembly alongcell junctions that allows the formation of a
larger-sized, multicellular contractile body(97, 102). The cells at the front of the body are
designated path-finding cells and are the onesthat generate traction via pseudopod activity
and expression of clusters of integrins andMMPs within the corresponding invadopodia
(102, 103). Cells in the inner and trailingregions are passively pulled along. Recent
work has shown that the type-1 mucin-likecell surface receptor podoplanin is upregu-
lated at the outer edge of growing tumorsand may promote collective tumor cell mi-
gration (104). Collective movement has beenobserved in several types of carcinoma (97).
Inflammation. Recruitment of leukocytesmay have different effects in different tu-
mor types. Thus, accumulation of myeloidcells, including neutrophils monocytes and
macrophages, is associated with indolent evo-lution in some cancers, but bears a much more
somber prognosis in others (105). Infiltrat-ing tumor-associated macrophages (TAMs)
present antigen and secrete cytokines thatsupport an adaptive antitumor immune re-
sponse. On the other hand, if tumor cells re-sist the immune reponse, which most solid
tumors appear to do succesfully, the TAM-derived chemokine/cytokine repertoire may
promote tumor progression (106). In addition
to CAFs, tumor cells themselves may recruit
hematopoietic precursors and leukocytes.Myeloid precursors as well as monocytes
express receptors for several growth fac-tors/cytokinessecreted by tumorcells, includ-
ing VEGFR1, which binds tumor-derivedVEGF-A and placental growth factor PIGF,
facilitating their recruitment to the tumor mi-croenvironment where they can differentiate
into TAMs and promote tumor growth anddissemination. In a model of skin carcinogen-
esis in the mouse, TAM-derived MMP-9 was
shown to play a key role in promoting tu-mor angiogenesis (107). Macrophages were
also found to play an essential role in tumorintravasation (108), whereas macrophage de-
pletion hasbeen observed to repress late-stagetumor progression and metastasis but not pri-
mary tumor growth (109, 110). Together withCAFs, TAMs and possibly other leukocytes
may supply tumor cells with proinvasive fac-tors that facilitate metastasis (111).
Angiogenesis. One of the prerequisites for
metastatic tumor growth is the inductionof angiogenesis (112, 113). Angiogenesis is
frequently induced by transforming eventsthat promote tumor progression and aug-
ment expression of angiogenic factors. Thus,VEGF-A expression is induced by the MAPK
signaling pathway and hypoxia that accom-panies rapid primary tumor growth. Hypoxia
induces and stabilizes expression of hypoxia-inducible-factor-1, which drives VEGF-A
transcription. VEGF-A is among growthfactors deposited in the ECM and whose
bioavailability, in addition to that of otherproangiogenic factors including basic fibrob-
last growth factor and TGF-, is increased asa result of tissue remodeling (114).
Studies in cancer patients as well as in
mouse models provide evidence that lym-phangiogenesis, defined as the outgrowth
of new lymphatic vessels from preexistingones, promotes metastasis to regional drain-
ing lymph nodes of a tumor (115, 116). Lym-phangiogenesis is induced by tumor-derived
VEGF-C and -D members of the VEGF
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family, which bind to VEGFR3 on the sur-
face of lymphatic endothelial cells. VEGF-Cexpression is regulated, at least in part, by in-
flammatory responses triggered by IL-1 and
tumor necrosis factor signaling. VEGF-Dis the product of an immediate early, Fos-
regulated gene, and its expression may there-fore be regulatedby oncogenicsignaling path-
ways (115, 116).
Intravasation. It would seem logical thatan increased lymphatic and vascular network
may facilitate penetration of the vascular lu-men by invading tumor cells. Carcinomas ini-
tiallyform metastasis in local lymph nodes andonly at a later stage in other organs. There
is an ongoing and as yet unresolved debateas to whether distal hematogenous metastases
in carcinomas develop as a result of vascularinvasion and penetration at the primary tu-
mor site or whether they are derived fromcells that have colonized local lymph nodes.
In the first case, tumor cells would need todegrade vascular BM and irrupt into the cir-
culation. An argument favoring this possibil-ity is that vascular invasion and penetration
by tumor cells is observed by microscopic ex-amination of tissue sections. This is further
supported by experimental approaches using
an in vivo assay (117), where intravasation de-pended upon MMP-9 activity and constituteda rate-limiting step in metastasis. Tumor cell
dissemination from lymph nodes could occurby migration of the cells to efferent lymph
vessels and transport to the vena cava fromwhere hematogenous spread would be possi-
ble. Currently, it would seem plausible thatboth mechanisms might be operational and
that the relative ease of lymph vessel inva-sionand penetration might explain that lymph
nodes are usually the first metastatic site incarcinomas. It is also possible, however, that
colonization of both lymphoid and nonlym-
phoid organs occurs within a comparable timeframe, but owing to local conditions tumor
growth proceeds more rapidly in lymphoidtissues.
Tumor Dissemination
Survival in lymph and blood and interac-
tions with endothelium. Once tumor cells
have penetrated the blood circulation, theyare exposed to shear stress and to interac-
tions with leukocytes that may lead to theirdestruction. It wouldappear, however, thattu-
mor cells are capable of resisting shear stresspossibly aided by platelets and leukocytes, and
that they may rely on some of the mechanismsused by leukocytes to adhere to endothelium
Transformed cells often express an alteredglycosyltranferase repertoire with respect to
normal counterparts. Glycosyl- and sialyl-transferases expressed in many carcinoma
types decorate cell surface receptors witholigosaccharide structures that correspond to
ligands of selectins, a C-type lectin class of
cell surface adhesion molecules that regulateleukocyte endothelial interactions and leuko-
cyte trafficking (118). P-selectin/CD62P isexpressed on the surface of activated platelets
and endothelial cells, and E-selectin/CD62Eis predominantly induced in activated en-
dothelium. L-selectin/CD62L is expressed onthe surface of a broad range of leukocyte
subpopulations.A variety of potentially metastatic tu-
mor cells, particularly those that are mucin
richoften derived from colonexpress se-lectin ligands (119121). Circulating tumorcells that express selectin ligands can become
coated with platelets and leukocytes, creat-ing a microembolus that may obstruct cap-
illaries of various organs (119); they may alsoadhere to activated endothelial cells. In an ex-
perimental metastasis model, B16 melanomacells engineered to synthesize oligosaccha-
rides that constitute selectin ligands displayedan altered pattern of organ colonization
compared with their parental counterparts(122).
Platelets and leukocytes can also inter-act with tumor cells via v3-dependent
adhesion (123). Experimental models sug-gest that tumor-cellplatelet/leukocyte inter-
actions may favor tumor metastasis (120, 121
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124, 125), but whether such a mechanism is
important in human cancer metastasis is sub-ject to debate.
Experimental evidence suggests that inte-
grins and immunoglobulin superfamily adhe-sion molecules are also implicated in tumor
cell adhesion to endothelium. The 41 in-tegrin, associatedprimarily with lymphocytes,
is expressed on a variety of tumor cell types,and its ligand VCAM-1 was shown to support
melanoma cell adhesion to endothelial cells(126).
Leukocyte adhesion to activated en-dothelium typically relies on three sets of
events: selectin-mediated low-affinity inter-actions responsible for leukocyte rolling on
the endothelium; endothelial cell-derivedchemokine-mediated leukocyte activation
that changes leukocyte 2 integrin confor-mation from low to high affinity; and high-
affinity interaction between leukocyte 2integrins and endothelial ICAM-1, which is
required for theleukocyte arrest that precedesand is necessary for diapedesis/extravasation
(118). Despite being larger and having dif-ferent morphological properties than circu-
lating leukocytes, carcinoma cells can makeuse of at least some of the adhesive mecha-
nisms that govern leukocyte trafficking to in-teract with vascular endothelium. However,
it remains to be demonstrated whether theseadhesive events are relevant to human cancer
metastasis.
Random arrest or programmed organ-
specific homing. Intravital microscopy im-ages argue that tumor cells tend to obstruct
capillaries, particularly if they are aggregatedor bound to leukocytes and platelets. Follow-
ing arrest in the capillary bed, they proliferatelocally and disrupt the capillary wall, whereby
they penetrate the local parenchyma (127).It would therefore seem that tumor cell ar-
rest in capillaries and subsequent extravasa-tion are primarily mechanical processes that
could occur in all organs. Organ-specific tu-mor cell homing would then require spe-
cific mechanims. Three nonmutually exclu-
sive candidate mechanisms proposed thus
far are chemokine-receptor-mediated chemo-taxis, the establishment of a metastatic niche,
and a tumor cell genetic program that facili-tates adaptation to a particular microenviron-
ment.
Chemokine-receptor-mediated chemo-taxis. Chemokines are believed to cooperate
with adhesion receptors in determining wheretumor cells arrest and extravasate. Tumor
cells can express a variety of chemokinereceptors, including CXCR4, that serves as
a receptor for CXCL12/SDF. Secretion ofSDF by host tissue stromal fibroblasts is
suggested to promote chemotaxis of tumorcells expressing CXCR4 and to determine, at
least in part, the localization of metastases of
certain tumor types (128).
Preparation of the metastatic microenvi-
ronment. Recent studies have suggested thatby virtue of their cytokine and chemokine
repertoire, tumorsmayhave theability to pre-pare the microenvironment of distant organs
to receive disseminating cells and allow theirproliferation. Tumor and associated stromal
cellderived chemokines can recruit endothe-lial and hematopoietic progenitor cells (EPCs
and HPCs, respectively) to the relevant or-gans prior to tumor cell arrival, which, to-
gether with tumor cellderived deposition offibronectin, appear to precondition the local
microenvironment (129, 130). Targeted inhi-bition of these cells using anti-VEGFR1 neu-
tralizing antibodiessuggeststhattheproposedpreconditioning is necessary for metastatic
spread. The mechanisms that govern HPCrecruitment to potential metastatic sites are
still unclear, as is the manner in which HPCs
render anygiven site permissive for metastatictumor growth. It is possible that recruitment
is initiated by the very first tumor cells thatarrive at a distant site and that local HPC
activity then alters the local microenviron-ment, amplifying tumor cell recruitment and
allowing subsequent division. However, the
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existence of metastatic niches remains to be
confirmed.
Gene expression patterns that determine
organ-specific homing of tumor cells. Pi-oneeringworkusingan experimentalmetasta-
sis approach has shown that repeated cycles ofin vivo passage of tumor cells that develop few
lung tumors following tail vein injection inmice result in selection of cells that preferen-
tially develop lung metastasis (131). As men-tioned above, thegeneexpression profileanal-
ysis of B16 melanoma cells selected for lungcolonization revealed upregulation of numer-
ous genes including RhoC (49), and forcedexpression if RhoC in parental B16 cells re-
sulted in the preferential homing to the lung(49).
To address this issue in human tumors,one study focused on the human breast
carcinoma MDA-MB231 cell line derivedfrom the pleural effusion of a patient with
widespread metastasis. Repeated rounds of invivo passage and selection allowed isolation of
different sublines of MDA-MB231 cells thatpredictably formed tumors in given organs
following intravenous injection (132). Geneexpression profiling of these sublines shows
that they express a specific set of genes thatcorrelates with general metastatic proclivity.
However, the selected cell lines expressed
additional gene signatures that correlatedwith the organ to which they metastasized
(132). Thus, a set of 54 genes was identi-fied that distinguished cell lines displaying
lung tropism (132). Although none of theidentified genes alone could recapitulate the
metastatic phenotype of selected cell lines,combinations of the genes could induce the
poorly metastatic parental MDA-MB231cells to colonize the organ from which the
metastatic cell variants were retrieved.Prior to this study, the same cell line had
been used to identify genes that may be in-volved in promoting bone metastasis (133).
Most of the highly overexpressed genes incell lines derived from and selected for their
ability to induce bone metastases encoded cell
membrane or secreted molecules that were in
some way relevant to the bone microenviron-ment. Expression of any one of these genes in
parental MDA-MB231 cellsfailedto augmenttheir intrinsic metastatic potential to bone
However, the combination of three to four ofthe genes augmented the metastatic activity
of parental cells to levels comparable to thosedisplayed by the most aggressive cell lines ex-
pressing the entire bone metastasis gene set(133). These observations strongly suggest
that cooperation among several genes that en-
code proteins with complementary functionsunderlies the metastatic phenotype. Cells ex-
pressing the required genes were identified inthe initiating tumor cell line, suggesting that
breast cancer cells that display a gene expres-sion signature associated with bone or lung
metastatic proclivity exist in the parental tu-mor cell population.
Establishment of new colonies. Extrava-sation was long thought to be a rate-
limiting step in metastasis. However, RAS-transformed NIH3T3 cells and parenta
counterparts were found to extravasate intothe liver at a comparable rate following in-
jection into the portal vein, whereas only theRAS-transformed cells could form metastatic
growth (134). These observations provideconvincing evidence that the rate-limiting
step is not extravasation, but rather the abilityof the cells to establish themselves in the new
host tissue microenvironment.Once tumor cells have penetrated the
parenchyma of an organ other than the onewhere they originated, they must create a mi-
croenvironment conducive to their survivaland proliferation. The situation is analogous
to that of invading cells of the primary tu-
mor penetrating its BM and entering into di-rect physical contact for the first time with the
stroma. However, the stromal composition ofthe secondary site may differ from that sur-
rounding theprimary tumor, andtheability ofthetumor cells to subvert the local microenvi-
ronment will most likely determine their fate
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Dormancy. Metastases from some human
cancers occur as many as 20 years follow-ing removal of the primary tumor. These
metastatic lesions are believed to be dormant
for an extended period of time and to becomeproliferative once localconditionsbecome ap-
propriately permissive.Dormancy is a concealed state, and as such
does not readily lend itself to direct study. Ex-perimental work, however, has provided ev-
idence for the existence of micrometastasesthat fail to induce angiogenesis and in which
cell proliferation is balanced by cell death be-cause of inadequate blood supply (135). These
small metastatic lesions may therefore consti-tute one source of tumor dormancy. Another
possible source of tumor dormancy are iso-lated tumor cells that arrive at secondary sites
where they may persist for long periods oftime without being able to divide (127). Inter-
estingly, recovery of isolated dormant mam-mary carcinoma cells from liver tissue showed
that they retain tumorigenicity when injectedinto immunocompromised mice (127). These
observations suggest that the host tissue mayprovide permissive or restrictive cues that de-
termine whether metastatic cells may prolif-erate and generate secondary tumors. Recent
evidence suggests that changes in the ECMthat lead to a shift in the equilibrium between
natural stimulators and inhibitors of angio-genesis, many of which are ECM degradation
products, may allow dormant metastatic le-sions to induce an angiogenic switch and de-
velop into full-blown secondary tumors (136).
EARLY EMERGENCE ORLATE-STAGE SELECTIONOF METASTATIC CELLS?
Until recently, the prevailing reasoning was
that metastasis arises from rare tumor cellsthat emerge relatively late in tumor progres-
sion. The genetic makeup of these cells wasbelieved to be the consequence of stochastic
accumulation of mutations that provide themwith all of the properties required for dissem-
ination. This view is supported by the well-
documented correlation between primary tu-
mor size and risk of metastasis. However, theassociation of clinical features such as tumor
grade with metastatic proclivity and the oc-currence of bone micrometastases early in the
evolution of cancer are inconsistent with astrictly stochastic model.
The use of DNA microarray studies toidentify transcriptional signatures that may
distinguish metastatic from primary tumorgrowth has further challenged this view, sug-
gesting that relevant signatures may be de-tected in early-stage primary tumors that are
destined to metastasize. This second viewis supported by observations from several
independent and distinctly designed studies(Figure 4). The first of these studies showed
that the clinical outcome of breast cancer pa-
tients can be predicted by a poor progno-sis gene expression signature present in the
majority of early-stage primary tumors (137,138). This gave rise to the notion that certain
tumors may have the properties required formetastasis from the very beginning. The size
of the primary tumor would then be irrele-vant in terms of the risk of developing metas-
tases, and even small tumors may be expectedto contain cells with metastatic potential.
In a second study, comparison of the geneexpression profile of metastatic adenocarci-
noma lesions to unmatched primary carcino-mas revealed a 17-gene expression signature
thatdistinguishedprimary frommetastatic tu-mors (139). Subsequent analysis revealed that
numerous early-stage primary solid tumors ofdiverse histotypes harbored the same gene ex-
pression signature, suggesting that these tu-mors were most likely associated with metas-
tases and poor clinical outcome.Because the bone marrow is a major hom-
ing site for breast cancer, a third study un-dertook the task of analyzing genomic differ-
ences between single bone marrowderivedmicrometastatic cells and the primary tumor
by comparative genomic hybridization (140).Single viable disseminated breast cancer cells
hadan abundance of chromosomal copy num-ber changes in their genome, with significant
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Metastasis
is an
early event
in tumor
progression
Primary tumors vs. metastatic nodules
17-genesignature
Breast ProstateLung
Good
Poor
The metastatic potential is encoded in the
bulk of primary tumors
(Ramaswamy et al. 2003)
Early-stage primary
invasive breast tumors
Tumor microenvironment
70-genesignature
BreastGood
Poor
Prostate Breast
Good
Poor
Wound-
response
genes
No mets
Mets
Metastatic proclivity is present in early tumors
(vant Veer et al. 2002)
Wound response signature in primary tumors
predicts increased risk of metastasis
and poor outcome
(Bacac et al. 2006, Chang et al. 2005)
In situ Invasive
LCM stroma
Bone metastases vs. primary tumor
Dissemination occurs early during tumorigenesis
(Gangnus et al. 2004)
Short-termculture
CGH
11-genesignature
Metastases vs.primarytumor
Breast ProstateLung
Good
Poor
Stem cellness and cancer
Tumors with stem celllike signatures are
likely to have poor prognosis
(Glinsky et al. 2005)
Bmi/
Bmi+/+Mouse-human
translational
genomics
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intercellular heterogeneitybut more impor-
tantly, they displayed numerous differences incomparison to the matching primary tumors.
These observations led to the conclusion that
metastasis is an early event in malignant tu-mors and that disseminated cells evolve inde-
pendently of the primary tumor.A fourth study addressed the possibility
that cancers with poor prognosis and highmetastatic proclivity may display some prop-
erties that characterize normal stem cells.Comparison of primary and metastatic mouse
prostate cancers with normal stem cells thathad retained or lost their self-renewing po-
tential identifieda common 11-gene signaturethat was then used to probe human cancers
(141143). Expression of this gene signaturein 11 distinct types of primary cancers was
consistently a powerful predictor of a shortinterval to disease recurrence, distant metas-
tasis, and death following therapy. These ob-servations are consistent with the possibility
that cancer cells with high metastatic procliv-ity display features that are at least in part
reminiscent of those of normal stem cells. In-terestingly, a fraction of each of the gene ex-
pression signatures identified in these stud-ies comprised stromal cell transcripts, some
of which were part of the recently describedpredictive stromal gene set (74). This further
underscores the notion that the stroma mayactively participate in determining the ability
of cancer to metastasize (74, 75, 144, 145).The emerging, and still controversial, con-
cept of cancer stem cells suggests that amongthe heterogeneous populations of cells that
compose primary tumors is a small popula-
tion of stem cellresembling malignant cellsthat constitute the driving force of the tumor
and are essential for progression and metasta-sis. In support of this view, eight out of nine
tumor samples that served as a source for theidentification of breast cancerinitiating cells
were derived from metastatic lesions (146).These cells may share attributes of normal
stem cells that are relevant to their natural be-havior and resistance to chemotherapy. Nor-
mal stem cells express multidrug-resistancegenes, which, coupled to their slow prolifer-
ative rate, may play a key role in their abilityto withstand cytotoxic drugs and repopulate
tissues that had been depleted of their nor-mal cells by chemotherapy. Metastatic cancer
lesions are notorious for resistance to con-ventional chemotherapy, at least in part be-
cause of multidrug-resistance gene expres-sion. Whether or not the stem cell connection
turns out to be correct, the lack of respon-siveness of metastatic lesions to conventional
therapy and the increasingly clear evidencethat the stroma is implicated in tumor metas-
tasis warrant a closer look at the molecularmechanisms that govern tumor-host interac-
tions at both primary and metastatic sites.
SUMMARY AND FUTURE
DIRECTIONSRecent studies strongly support the view that
the capacity of a tumor to disseminate is ac-quired at early steps during the multistep pro-
cess of tumorigenesis. They also suggest thatspecific genes uniquely responsible for cancer
cell dissemination and metastatic growth are
Figure 4
Overview of recent microarray and CGH studies that led to the notion that metastasis is an early event intumorigenesis. Validation of signatures identified by Ramaswamy et al. (139) on patient cohorts withearly-stage tumors showed that metastatic proclivity is present early in tumor evolution. Studiesperformed by Gangnus et al. (140) on bone micrometastasis confirmed this notion. More recently,Glinsky et al. (141) proposed that tumors with a stem cell expression signature are likely to have a poorprognosis. Most of these signatures contained stroma-associated genes, consistent with the notion thatthe tumor microenvironment plays an important role in dissemination. This was also suggested by recentstudies focusing on stromal reactions to tumor invasion and injury that showed that a wound-responsesignature in primary tumors predicts increased risk of metastasis.
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unlikely to be discovered. Rather, the com-
bined effect of oncogene signaling and tu-mor suppressor gene loss in the appropriate
cellular environment is likely to determine
whether a cancer cell has the potential to col-onize distant organs. Success of secondary tu-
mor growth is then determined by the natureof the host response and the tumor cells abil-
ity or inability to subvert it.The realization that early cancer har-
bors metastatic potential should warrant pre-ventive treatment of metastatic disease. Our
extensive understanding of the mechanismswhereby cancer cells spread should allow the
development of strategies that can effectivelyblock tumor dissemination. Approaches likely
to meet with success are those that simul-
taneously target several mechanisms uponwhich tumor cell dissemination depends, in-
cluding angiogenesis, proteolysis, and growthfactor signaling. Much more challenging
is the prospect of reversing already estab-lished metastatic growth. The formidable
ability of metastatic lesions to evade cyto-toxic drug effects, at least in part due to
multidrug resistance family gene expressionindicates that we need to look elsewhere
for therapeutic solutions. As metastatic tu-mors require local stroma support for growth
identifying targetable tumor-host interac-tion mechanisms appears to be an appealing
pursuit.
DISCLOSURE STATEMENT
The authors are not aware of any biases that might be perceived as affecting the objectivity ofthis review.
ACKNOWLEDGMENTS
This work was supported by the Fonds de la Recherche Scientifique grant number 3100A0-
105833 and by the National Center of Competence in Research Molecular Oncology.
LITERATURE CITED
1. Perez-MorenoM, JamoraC, Fuchs E. 2003. Stickybusiness: orchestrating cellular signals
at adherens junctions. Cell112:535482. Conacci-Sorrell M, Zhurinsky J, Ben-Zeev A. 2002. The cadherin-catenin adhesion
system in signaling and cancer. J. Clin. Invest. 109:987913. Wheelock MJ, Johnson KR. 2003. Cadherins as modulators of cellular phenotype. Annu
Rev. Cell Dev. Biol. 19:207354. Vleminckx K, Vakaet LJ, Mareel M, Fiers W, van Roy F. 1991. Genetic manipulation of
E-cadherin expression by epithelial tumor cells reveals an invasion suppressor role. Cell66:10719
5. PerlAK,WilgenbusP,DahlU,SembH,ChristoforiG.1998.AcausalroleforE-cadherinin the transition from adenoma to carcinoma. Nature 392:19093
6. Graff JR, Gabrielson E, Fujii H, Baylin SB, Herman JG. 2000. Methylation patterns ofthe E-cadherin 5 CpG island are unstable and reflect the dynamic, heterogeneous loss
of E-cadherin expression during metastatic progression. J. Biol. Chem. 275:2727327. Nass SJ, Herman JG, Gabrielson E, Iversen PW, Parl FF, et al. 2000. Aberrant methy-
lation of the estrogen receptor and E-cadherin 5 CpG islands increases with malignantprogression in human breast cancer. Cancer Res. 60:434648
8. Bolos V, Peinado H, Perez-Moreno MA, Fraga MF, Esteller M, Cano A. 2003. Thetranscription factor Slug represses E-cadherin expression and induces epithelial to mes-
enchymaltransitions: a comparison with Snail andE47repressors.J. Cell Sci. 116:499511
240 Bacac Stamenkovic
8/2/2019 Review2008_metastasisb Cancer Cell
21/29
9. Cano A, Perez-Moreno MA, Rodrigo I, Locascio A, BlancoMJ, et al. 2000. The transcrip-
tion factor Snail controls epithelial-mesenchymal transitions by repressing E-cadherinexpression. Nat. Cell Biol. 2:7683
10. Peinado H, Ballestar E, Esteller M, Cano A. 2004. Snail mediates E-cadherin repression
by the recruitment of the Sin3A/histone deacetylase 1 (HDAC1)/HDAC2 complex. Mol.Cell. Biol. 24:30619
11. Perez-Moreno MA, Locascio A, Rodrigo I, Dhondt G, Portillo F, et al. 2001. A new
role for E12/E47 in the repression of E-cadherin expression and epithelial-mesenchymaltransitions. J. Biol. Chem. 276:2742431
12. Yang J, Mani SA, Donaher JL, Ramaswamy S, Itzykson RA, et al. 2004. Twist, a master
regulator of morphogenesis, plays an essential role in tumor metastasis. Cell117:92739
13. Fujita Y, Krause G, Scheffner M, Zechner D, Leddy HE, et al. 2002. Hakai, a c-Cbl-like
protein, ubiquitinates and induces endocytosis of the E-cadherin complex. Nat. Cell Biol.4:22231
14. Avizienyte E, Frame MC. 2005. Src and FAK signalling controls adhesion fate and theepithelial-to-mesenchymal transition. Curr. Opin. Cell Biol. 17:54247
15. Avizienyte E, Wyke AW, Jones RJ, McLean GW, Westhoff MA, et al. 2002. Src-inducedde-regulation of E-cadherin in colon cancer cells requires integrin signalling. Nat. Cell
Biol. 4:6323816. Novak A, Hsu SC, Leung-Hagesteijn C, Radeva G, Papkoff J, et al. 1998. Cell adhesion
and theintegrin-linked kinase regulate the LEF-1 and-catenin signaling pathways.Proc.
Natl. Acad. Sci. USA 95:437479
17. Tan C, Costello P, Sanghera J, Dominguez D, Baulida J, et al. 2001. Inhibition of integrinlinked kinase (ILK) suppresses -catenin-Lef/Tcf-dependent transcription and expres-
sion of theE-cadherin repressor, Snail, in APC/ human colon carcinoma cells. Oncogene
20:13340
18. Noe V, Fingleton B, Jacobs K, Crawford HC, Vermeulen S, et al. 2001. Release of
an invasion promoter E-cadherin fragment by matrilysin and stromelysin-1. J. Cell Sci.114:11118
19. Cavallaro U, Christofori G. 2004. Cell adhesion andsignalling by cadherinsandIg-CAMsin cancer. Nat. Rev. Cancer4:11832
20. Cavallaro U, Niedermeyer J, Fuxa M, Christofori G. 2001. N-CAM modulates tumour-cell adhesion to matrix by inducing FGF-receptor signalling. Nat. Cell Biol. 3:65057
21. Marambaud P, Wen PH, Dutt A, Shioi J, Takashima A, et al. 2003. A CBP bindingtranscriptional repressor produced by the PS1/-cleavage of N-cadherin is inhibited by
PS1 FAD mutations. Cell114:63545
22. Hynes RO. 2002. Integrins: Bidirectional, allosteric signaling machines. Cell110:67387
23. Giancotti FG, Ruoslahti E. 1999. Integrin signaling. Science 285:102832
24. Ivaska J, Nissinen L, Immonen N, Eriksson JE, Kahari VM, Heino J. 2002. Integrin21 promotes activation of protein phosphatase 2A and dephosphorylation of Akt and
glycogen synthase kinase 3 . Mol. Cell. Biol. 22:13525925. Ivaska J, Reunanen H, Westermarck J, Koivisto L, Kahari VM, Heino J. 1999. Inte-
grin 21 mediates isoform-specific activation of p38 and upregulation of collagen genetranscription by a mechanism involving the 2 cytoplasmic tail. J. Cell Biol. 147:40116
26. Owens DM, Watt FM. 2001. Influence of 1 integrins on epidermal squamous cellcarcinoma formation in a transgenic mouse model: 31, but not 21, suppresses
malignant conversion. Cancer Res. 61:524854
www.annualreviews.org Metastatic Cancer Cell 241
8/2/2019 Review2008_metastasisb Cancer Cell
22/29
27. ZutterMM, Santoro SA,Staatz WD,Tsung YL.1995.Re-expression of the21 integrin
abrogates the malignant phenotype of breast carcinoma cells. Proc. Natl. Acad. Sci. USA92:741115
28. Chan BM, Matsuura N, Takada Y, Zetter BR, Hemler ME. 1991. In vitro and in vivo
consequences of VLA-2 expression on rhabdomyosarcoma cells. Science 251:1600229. Wang H, Fu W, Im JH, Zhou Z, Santoro SA, et al. 2004. Tumor cell 31 integrin and
vascular laminin-5 mediate pulmonary arrest and metastasis. J. Cell Biol. 164:9354130. Gladson CL, Cheresh DA. 1991. Glioblastoma expression of vitronectin and the v3
integrin. Adhesion mechanism for transformed glial cells. J. Clin. Invest. 88:19243231. Mercurio AM, Rabinovitz I. 2001. Towards a mechanistic understanding of tumor
invasionlessons from the 64 integrin. Semin. Cancer Biol. 11:1294132. Mariotti A, Kedeshian PA, Dans M, Curatola AM, Gagnoux-Palacios L, Giancotti FG
2001. EGF-R signaling through Fyn kinase disrupts the function of integrin 64 athemidesmosomes: role in epithelial cell migration and carcinoma invasion. J. Cell Biol
155:4475833. Trusolino L, Bertotti A, Comoglio PM. 2001. A signaling adapter function for 64
integrin in the control of HGF-dependent invasive growth. Cell107:6435434. Schneller M, Vuori K, Ruoslahti E. 1997. v3 integrin associates with activated in-
sulin and PDGF receptors and potentiates the biological activity of PDGF. EMBO J
16:5600735. Hannigan G, Troussard AA, Dedhar S. 2005. Integrin-linked kinase: a cancer therapeutic
target unique among its ILK. Nat. Rev. Cancer5:516336. Schlaepfer DD, Mitra SK. 2004. Multiple connections link FAK to cell motility and
invasion. Curr. Opin. Genet. Dev. 14:9210137. Mitra SK, Schlaepfer DD. 2006. Integrin-regulated FAK-Src signaling in normal and
cancer cells. Curr. Opin. Cell Biol. 18:5162338. Sieg DJ, Hauck CR, Ilic D, Klingbeil CK, Schaefer E, et al. 2000. FAK integrates growth-
factor and integrin signals to promote cell migration. Nat. Cell Biol. 2:2495639. Zhai J, Lin H, Nie Z, Wu J, Canete-Soler R, et al. 2003. Direct interaction of focal
adhesion kinase with p190RhoGEF. J. Biol. Chem. 278:2486573
40. Carragher NO, Westhoff MA, Fincham VJ, Schaller MD, Frame MC. 2003. A novel rolefor FAK as a protease-targeting adaptor protein: regulation by p42 ERK and Src. Curr
Biol. 13:14425041. Klemke RL, Cai S, Giannini AL, Gallagher PJ, de Lanerolle P, Cheresh DA. 1997
Regulation of cell motility by mitogen-activated protein kinase. J. Cell Biol. 137:4819242. Huang C, Rajfur Z, Borchers C, Schaller MD, Jacobson K. 2003. JNK phosphorylates
paxillin and regulates cell migration. Nature 424:2192343. Keely PJ, Westwick JK, Whitehead IP, Der CJ, Parise LV. 1997. Cdc42 and Rac1 induce
integrin-mediated cell motility and invasiveness through PI(3)K. Nature 390:6323644. Raftopoulou M, Hall A. 2004. Cell migration: Rho GTPases lead the way. Dev. Biol
265:233245. Pollard TD, Borisy GG. 2003. Cellular motility driven by assembly and disassembly of
actin filaments. Cell112:4536546. Watanabe N, Kato T, Fujita A, Ishizaki T, Narumiya S. 1999. Cooperation between
mDia1 and ROCK in Rho-induced actin reorganization. Nat. Cell Biol. 1:1364347. Kimura K, Ito M, Amano M, Chihara K, Fukata Y, et al. 1996. Regulation of myosin
phosphatase by Rho and Rho-associated kinase (Rho-kinase). Science 273:2454848. Sahai E, Marshall CJ. 2003. Differing modes of tumour cell invasion have distinct require-
ments for Rho/ROCK signalling and extracellular proteolysis. Nat. Cell Biol. 5:71119
242 Bacac Stamenkovic
8/2/2019 Review2008_metastasisb Cancer Cell
23/29
49. Clark EA, Golub TR, Lander ES, Hynes RO. 2000. Genomic analysis of metastasis
reveals an essential role for RhoC. Nature 406:5323550. Gabarra-Niecko V, Schaller MD, Dunty JM. 2003. FAK regulates biological processes
important for the pathogenesis of cancer. Cancer Metastasis Rev. 22:35974
51. Ilic D, Genbacev O, Jin F, Caceres E, Almeida EA, et al. 2001. Plasma membrane-associated pY397FAK is a marker of cytotrophoblast invasion in vivo and in vitro. Am. J.
Pathol. 159:9310852. Hsia DA, Mitra SK, Hauck CR, Streblow DN, Nelson JA, et al. 2003. Differential reg-
ulation of cell motility and invasion by FAK. J. Cell Biol. 160:7536753. Hauck CR, Sieg DJ, Hsia DA, Loftus JC, Gaarde WA, et al. 2001. Inhibition of focal ad-
hesion kinase expression or activity disrupts epidermal growth factor-stimulated signalingpromoting the migration of invasive human carcinoma cells. Cancer Res. 61:707990
54. Schneider GB,Kurago Z, Zaharias R, GrumanLM, Schaller MD,Hendrix MJ.2002.Ele-vated focal adhesion kinase expression facilitates oral tumor cell invasion. Cancer95:2508
1555. Gavert N, Conacci-Sorrell M, Gast D, Schneider A, Altevogt P, et al. 2005. L1, a novel
target of-catenin signaling, transforms cells and is expressed at the invasive front ofcolon cancers. J. Cell Biol. 168:63342
56. Conacci-Sorrell ME, Ben-Yedidia T, Shtutman M, Feinstein E, Einat P, Ben-Zeev A.2002. Nr-CAM is a target gene of the -catenin/LEF-1 pathway in melanoma and
colon cancer and its expression enhances motility and confers tumorigenesis. Genes Dev.16:205872
57. Perl AK, Dahl U, Wilgenbus P, Cremer H, Semb H, Christofori G. 1999. Reducedexpression of neural cell adhesion molecule induces metastatic dissemination of pancreatic tumor cells. Nat. Med. 5:28691
58. Ponta H, Sherman L, Herrlich PA. 2003. CD44: from adhesion molecules to signalling
regulators. Nat. Rev. Mol. Cell Biol. 4:334559. Sherman LS, Rizvi TA, Karyala S, Ratner N. 2000. CD44 enhances neuregulin signaling
by Schwann cells. J. Cell Biol. 150:10718460. Orian-Rousseau V, Chen L, Sleeman JP, Herrlich P, Ponta H. 2002. CD44 is required
for two consecutive steps in HGF/c-Met signaling. Genes Dev. 16:30748661. Yu Q, Stamenkovic I. 1999. Localization of matrix metalloproteinase 9 to the cell surface
provides a mechanism for CD44-mediated tumor invasion. Genes Dev. 13:354862. Yu Q, Stamenkovic I. 2000. Cell surface-localized matrix metalloproteinase-9 prote-
olytically activates TGF- and promotes tumor invasion and angiogenesis. Genes Dev.14:16376
63. Yu WH, Woessner JFJ, McNeish JD, Stamenkovic I. 2002. CD44 anchors theassembly of
matrilysin/MMP-7 with heparin-binding epidermal growth factor precursor and ErbB4and regulates female reproductive organ remodeling. Genes Dev. 16:30723
64. Tsukita S, Oishi K, Sato N, Sagara J, Kawai A, Tsukita S. 1994. ERM family members asmolecular linkers between the cell surface glycoprotein CD44 and actin-based cytoskele-
tons. J. Cell Biol. 126:39140165. McClatchey AI. 2003. Merlin and ERM proteins: unappreciated roles in cancer develop-
ment? Nat. Rev. Cancer3:8778366. Thomas L, Byers HR, Vink J, Stamenkovic I. 1992. CD44H regulates tumor cell migra-
tion on hyaluronate-coated substrate. J. Cell Biol. 118:9717767. Yu Q, Stamenkovic I. 2004. Transforming growth factor- facilitates breast carcinoma
metastasis by promoting tumor cell survival. Clin. Exp. Metastasis21:23542
www.annualreviews.org Metastatic Cancer Cell 243
8/2/2019 Review2008_metastasisb Cancer Cell
24/29
68. English NM, Lesley JF, Hyman R. 1998. Site-specific de-N-glycosylation of CD44 can
activate hyaluronan binding, and CD44 activation states show distinct threshold densitiesfor hyaluronan binding. Cancer Res. 58:373642
69. Lesley J, Hascall VC, Tammi M, Hyman R. 2000. Hyaluronan binding by cell surfaceCD44. J. Biol. Chem. 275:2696775
70. Skelton TP, Zeng C, Nocks A, Stamenkovic I. 1998. Glycosylation provides both stim-
ulatory and inhibitory effects on cell surface and soluble CD44 binding to hyaluronan.
J. Cell Biol. 140:4314671. Sleeman J, Rudy W, Hofmann M, Moll J, Herrlich P, Ponta H. 1996. Regulated clus-
tering of variant CD44 proteins increases their hyaluronate binding capacity. J. Cell Biol
135:11395072. Weber GF, Bronson RT, Ilagan J, Cantor H, Schmits R, Mak TW. 2002. Absence of the
CD44 gene prevents sarcoma metastasis. Cancer Res. 62:22818673. Liotta LA, Kohn EC. 2001. The microenvironment of the tumour-host interface. Nature
411:3757974. Bacac M, Provero P, Mayran N, Stehle JC, Fusco C, Stamenkovic I. 2006. A mouse
stromal response to tumor invasion predicts prostate and breast cancer patient survival.PLoS One 1:e32
75. Chang HY, Sneddon JB, Alizadeh AA, Sood R, West RB, et al. 2004. Gene expression
signature of fibroblast serum response predicts human cancer progression: similaritiesbetween tumors and wounds. PLoS Biol. 2:e7
76. Allinen M, Beroukhim R, Cai L, Brennan C, Lahti-Domenici J, et al. 2004. Molecularcharacterization of the tumor microenvironment in breast cancer. Cancer Cell6:1732
77. Hotary K, Li XY, Allen E, Stevens SL, Weiss SJ. 2006. A cancer cell metalloprotease
triad regulates the basement membrane transmigration program. Genes Dev. 20:26738678. Hotary KB, Allen ED, Brooks PC, Datta NS, Long MW, Weiss SJ. 2003. Membrane
type I matrix metalloproteinase usurps tumor growth control imposed by the three-dimensional extracellular matrix. Cell114:3345
79. Egeblad M, Werb Z. 2002. New functions for the matrix metalloproteinases in cancer
progression. Nat. Rev. Cancer2:16174
80. Zhou Z, Apte SS, Soininen R, Cao R, Baaklini GY, et al. 2000. Impaired endochondral os-sification and angiogenesis in mice deficient in membrane-type matrix metalloproteinase
I. Proc. Natl. Acad. Sci. USA 97:40525781. Hiraoka N, Allen E, Apel IJ, Gyetko MR, Weiss SJ. 1998. Matrix metalloproteinases
regulate neovascularization by acting as pericellular fibrinolysins. Cell95:3657782. Nakahara H, Howard L, Thompson EW, Sato H, Seiki M, et al. 1997. Transmem-
brane/cytoplasmic domain-mediated membrane type 1-matrix metalloprotease docking
to invadopodia is required for cell invasion. Proc. Natl. Acad. Sci. USA 94:79596483. Kajita M, Itoh Y, Chiba T, Mori H, Okada A, et al. 2001. Membrane-type 1 matrix
metalloproteinase cleaves CD44 and promotes cell migration. J. Cell Biol. 153:89390484. Mori H, Tomari T, Koshikawa N, Kajita M, Itoh Y, et al. 2002. CD44 directs membrane-
type 1 matrix metalloproteinase to lamellipodia by associating with its hemopexin-like
domain. EMBO J. 21:39495985. Stamenkovic I. 2003. Extracellular matrix remodelling: the role of matrix metallopro-
teinases. J. Pathol. 200:4486486. Wei Y, Lukashev M, Simon DI, Bodary SC, Rosenberg S, et al. 1996. Regulation of
integrin function by the urokinase receptor. Science 273:15515587. Teder P, Vandivier RW, Jiang D, Liang J, Cohn L, et al. 2002. Resolution of lung inflam-
mation by CD44. Science 296:15558
244 Bacac Stamenkovic
8/2/2019 Review2008_metastasisb Cancer Cell
25/29
88. Bierie B, Moses HL. 2006.