Download The Biology of Cancer Metastasis: Historical

Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Published OnlineFirst on July 7, 2010 as 10.1158/0008-5472.CAN-10-1040
AACR Centennial Series
Cancer
Research
AACR Centennial Series: The Biology of Cancer Metastasis:
Historical Perspective
James E. Talmadge1 and Isaiah J. Fidler2
Abstract
Metastasis resistant to therapy is the major cause of death from cancer. Despite almost 200 years of study,
the process of tumor metastasis remains controversial. Stephen Paget initially identified the role of host-tumor
interactions on the basis of a review of autopsy records. His “seed and soil” hypothesis was substantiated a
century later with experimental studies, and numerous reports have confirmed these seminal observations. An
improved understanding of the metastatic process and the attributes of the cells selected by this process is
critical for the treatment of patients with systemic disease. In many patients, metastasis has occurred by the
time of diagnosis, so metastasis prevention may not be relevant. Treating systemic disease and identifying
patients with early disease should be our goal. Revitalized research in the past three decades has focused
on new discoveries in the biology of metastasis. Even though our understanding of molecular events that
regulate metastasis has improved, the contributions and timing of molecular lesion(s) involved in metastasis
pathogenesis remain unclear. Review of the history of pioneering observations and discussion of current controversies should increase understanding of the complex and multifactorial interactions between the host and
selected tumor cells that contribute to fatal metastasis and should lead to the design of successful therapy.
Cancer Res; 70(14); 5649–69. ©2010 AACR.
Introduction
Tumor metastasis is a multistage process during which
malignant cells spread from the primary tumor to discontiguous organs (1). It involves arrest and growth in different microenvironments, which are treated clinically with different
strategies depending on the tumor histiotype and metastatic
location. Thus, a single hepatic metastasis may be resected,
whereas bone marrow metastases may be treated with bonetargeting radioisotopes. Because of cellular heterogeneity,
however, therapies have varying efficacy, challenging the oncologist and our understanding of the metastatic process (2).
Nonmalignant cells, under steady-state conditions, proliferate as needed to replace themselves as they age or become
injured. However, this process can go awry, resulting in uncontrolled proliferation and tumor formation, which can be
benign or malignant. Benign tumors are generally slow growing, enclosed within a fibrous capsule, noninvasive, and morphologically resemble their cellular precursor. If a benign
tumor is not close to critical vascular or neural tissue,
prompt diagnosis and treatment frequently result in a cure.
Authors' Affiliations: 1 The University of Nebraska Medical Center,
Transplantation Immunology Laboratory, Omaha, Nebraska; and 2The
University of Texas M.D. Anderson Cancer Center, Department of
Cancer Biology, Cancer Metastasis Research Center, Houston, Texas
Corresponding Author: Isaiah J. Fidler, The University of Texas M.D.
Anderson Cancer Center, 1515 Holcombe Boulevard, Unit 854, Houston,
TX 77030. Phone: 713-792-8580; Fax: 713-745-2174; E-mail: ifidler@
mdanderson.org.
doi: 10.1158/0008-5472.CAN-10-1040
©2010 American Association for Cancer Research.
In contrast, malignant tumors rarely encapsulate, grow rapidly, invade regional tissues, have morphologic abnormalities
such that their tissue of origin may be unrecognizable, and
metastasize. The term “metastasis” was coined in 1829 by
Jean Claude Recamier (1) and is now defined as “the transfer
of disease from one organ or part to another not directly
connected to it” (Fig. 1; ref. 3). Metastasis is the primary clinical challenge as it is unpredictable in onset and it exponentially increases the clinical impact to the host.
In many patients, metastasis has occurred by the time of
diagnosis, although this may not be apparent clinically (4).
Although tumor metastasis can occur early in tumor progression when the primary tumor is small or even undetectable,
most occurs later when the primary tumor is larger. This pattern is supported by the observation that surgical excision of
smaller lesions is often curative and forms the basis for tumor, nodes, metastasis (TNM) staging. When metastasis has
already occurred, strategies targeting tumor cell invasion and
other early aspects of the metastatic process may not be relevant to outcome. The suggestion, based on microarray analysis, that all tumor cells within a malignant tumor are equally
metastatic is likely to be incorrect (5–7). This controversial
conclusion is associated with the low sensitivity of microarray data that support the similarity of primary and metastatic tumor expression signatures, which are similarly due
to the activation of a metastatic genetic program in early
progenitors. It has been suggested that the overgrowth and
dominance within primary and secondary lesions by a single
tumor cell population, with a uniform metastatic signature,
is associated with early metastasis. An extension of this hypothesis is that the late emergence of metastatic clones will
www.aacrjournals.org
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5649
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
Figure 1. Selected events that have influenced the history and progress of cancer metastasis, 1829–1979.
result in divergent expression patterns between primary tumors and metastases, secondary to the masking of metastatic signatures in the primary tumor, by persisting
nonmetastatic clones (8). However, studies of clonal cell lines
derived from late-stage human carcinomas (9) have provided
direct evidence that individual cancer cells, coexisting within
a tumor, differ in their metastatic capability, including ones
that are nonmetastatic, confirming the tumor heterogeneity
shown in preclinical studies with murine (10–13), as well as
human tumors (14). Furthermore, the expression signatures
of tumors derived from cloned weakly and/or nonmetastatic
human cell lines and from their isogenic metastatic counterparts from the same patient tumors differ, although the
expression signature of metastases and that of their corresponding primaries are similar (15).
The pathogenesis of metastasis involves a series of steps,
dependent on both the intrinsic properties of the tumor cells
and the host response (16). In 1889, the English surgeon
Stephen Paget (17) identified the role of host-tumor cell
interactions. He addressed the question, “What is it that
decides what organs shall suffer in a case of disseminated
cancer?” (17). On the basis of a review of autopsy records
from 735 women with fatal breast cancer, he was able to propose an answer. In addition, he remarked on the discrepancy
between the blood supply and frequency of metastasis to
specific organs. This discrepancy included a high incidence
of metastasis to the liver, ovary, and specific bones, and a
low incidence to the spleen. His observations contradicted
the prevailing theory of Virchow (18), that metastasis could
be explained simply by the arrest of tumor-cell emboli in the
vasculature. Paget concluded that “remote organs cannot be
altogether passive or indifferent regarding embolism” and
elaborated upon the “seed and soil” principle, stating: “When
a plant goes to seed, its seeds are carried in all directions, but
they can only live and grow if they fall on congenial soil.” He
also suggested, “All reasoning from statistics is liable to many
5650
Cancer Res; 70(14) July 15, 2010
errors. But, the analogy from other diseases seems to support
what these records have suggested, the dependence of the
seed upon the soil” (17). Forty years later, in 1928, Ewing
challenged the “seed and soil” hypothesis (19). He proposed
that mechanical forces and circulatory patterns between the
primary tumor and the secondary site accounted for organ
specificity. The seminal studies by Fidler and coworkers
(10, 20) conclusively showed that, although tumor cells traffic
through the vasculature of all organs, metastases selectively
develop in congenial organs.
Studies in the late 1970s and early 1980s (10–26) stimulated research into the pathobiology of metastasis, resulting in
extensive research into the local microenvironment, or
“niche,” of the primary tumor and metastatic foci. They also
provided new insight into biological heterogeneity, the metastasis phenotype, and the selection of metastatic variants
during or by the process of metastasis (10). Thus, the studies
of Paget remain a basis for ongoing research and continue to
be cited 120 years after publication (Fig. 2). The “seed and
soil” hypothesis is now widely accepted although the “seed”
may now be identified as a progenitor cell, initiating cell,
cancer stem cell (CSC), or metastatic cell, and the “soil”
discussed as a host factor, stroma, niche, or organ microenvironment (27). Regardless, few currently disagree that the
outcome of metastasis is dependent on interactions between
tumor cells and host tissue. Similar to Paget's seed and soil
hypothesis, Fidler's studies have withstood the test of time
and have been repeatedly verified and validated. These results focused on the process of metastasis being sequential
and selective with stochastic elements, that is, the three
S's (28). This concept was initially controversial and has
provided a lightening rod on which new concepts and
hypotheses have been tested (21). Inarguably, at least to a
pathologist, malignancy is not a characteristic shared by all
cells within a tumor. This may seem obvious; however, recent
literature includes discussions in which the term “malignant”
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
Figure 1. Time Line. Continued.
is used as a synonym for “neoplastic” or “tumorigenic.” Indeed, given the myopia associated with historical literature,
researchers continue to rediscover classic observations, frequently under new synonyms. Unfortunately, these “new”
observations do not always incorporate the detailed understanding of tumor biology and experimental designs critical
to research into malignancy and metastasis. Studies focused
on metastasis require both in vivo and pathologic analyses
because malignancy and metastasis are biologic phenomena
and can only be described based on in vivo analysis. In the
absence of whole animal studies, the potential for misinterpretation is high.
The Pathogenesis of Metastasis
The process of cancer metastasis consists of sequential and
interrelated steps (Fig. 3), each of which can be rate limiting
because a failure at any step may halt the process (16). The
outcome of the process is dependent on both the intrinsic
properties of the tumor cells and the host response, such that
metastasis is a balance of host-tumor cellular interactions that
can vary between patients (29). In principle, the steps or events
required for metastasis are the same for all tumors (Table 1).
Thus, the outgrowth of a metastatic lesion requires that it develop a vascular network, evade the host's immune response
(30), and respond to organ-specific factors that influence
growth (31–33). Once they do, the cells can again invade the
host stroma, penetrate blood vessels, and enter the circulation
to produce secondary metastases, or so-called “metastasis of
metastases” (16, 34, 35). One critical aspect of this process is
the role of the arresting organ. As identified in the seed and
soil hypothesis, some organs can support the survival and
growth of the tumor emboli or limit its survival. Further, blood
flow characteristics and the structure of the vascular system
can also regulate the patterns of metastatic dissemination
www.aacrjournals.org
(36, 37). Autopsy studies in breast and prostate cancer support
an increased number of bone metastases based on blood flow.
Thus, some tumor-organ pairs of metastasis may have a positive relationship with metastasis formation (38) and some may
be associated with blood flow patterns (39).
Only a few cells in a primary tumor are believed to be able
to give rise to a metastasis (22, 40), which is a result of the
elimination of circulating tumor cells that fail to complete all
the steps in the metastatic process. The complexity of this
progression explains, in part, why the metastatic process
was suggested to be inefficient (38). For example, the presence of tumor cells in the circulation does not predict that
metastasis will occur as most of the tumor cells that enter
the blood stream are rapidly eliminated (22). The intravenous
injection of radiolabeled B16 melanoma cells revealed that by
24 hours after injection into the circulation, 0.1% or less of
the cells were still viable, and less than 0.01% of tumor cells
within the circulation survived to produce experimental lung
metastases (22). Observations such as this one raise the question of whether the development of metastases is the result
of fortuitous survival and growth of a few neoplastic cells, or
represents the selective growth of a unique subpopulation of
malignant cells with properties conducive to metastasis.
That is, can all cells growing in a primary tumor produce secondary lesions, or do only specific and unique cells possess
the properties required to enable them to metastasize? Because this question has largely been addressed in rodents,
and clinical studies have been more difficult, debate continues. Recent data show, however, that human neoplasms
are biologically heterogeneous (41–43) and that the process
of clinical metastasis is selective (44, 45).
The presence of tumor cells or emboli distant from the primary tumor does not prove that metastasis has occurred.
Circulating tumor cells are rapidly eliminated, and arrest in
a capillary bed or the marrow is not indicative that a
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5651
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
Figure 2. Selected events that have influenced the history and progress of cancer metastasis, 1980–Present.
metastatic focus will form or has formed. Indeed, cellular arrest during the metastatic process frequently results in cellular apoptosis, dormancy, and, more rarely, a clinically
detectable metastasis. Thus, the process of metastasis is inefficient, seldom exceeding 0.01% (22), whereas the entry of
tumor cells into the circulation is common and more than a
million cells per gram of tumor can be shed daily (46). Tarin
and coworkers documented the low frequency of secondary
foci using peritoneovenous shunts to reduce ascites in ovarian cancer (47). Although millions of tumor cells were directly deposited into the vena cava every 24 hours by the shunt,
patients rarely developed secondary tumors. Similarly, the
finding of epithelial cells within the marrow of patients with
breast cancer has prognostic indications (48). Not all cells
within the marrow form metastatic nodules owing, in part,
to host responses against the tumor cells. Thus, antitumor
host response can contribute to a soil incapable of supporting tumor cell growth, or associated with the development of
tumor dormancy, or both.
The process of tumor initiation, progression, and metastasis is not rapid, and tumor growth rates are generally slow.
Mammography (49, 50) has shown that primary breast tumors have an average doubling time of 157 days (32), varying
from 44 to more than 1,800 days during exponential growth
(51). Thus, the growth of a tumor from initiation to a size of
1 cm (52), which is the limitation of most current imaging
tools, requires an average of 12 years. This finding is significant because a 1-cm tumor has 109 cells and has undergone
at least 30 doublings from tumor initiation to diagnosis (4).
On the basis of these parameters and the observation that a
tumor burden of approximately 1,000 cm3 is generally lethal
(53), the time from diagnosis to mortality represents 10 doubling times from a 1-cm tumor and a shorter time frame.
Thus, three quarters of a tumor's life history has occurred
prior to diagnosis, and metastasis can occur prior to diagno-
5652
Cancer Res; 70(14) July 15, 2010
sis. Similarly, a tumor embolus associated with a surgical
shower would require 10 to 12 years to grow to a 1-cm
metastasis. These timelines are supported by registry data,
which suggest that the median time from tumor resection
to a diagnosis of metastasis for patients with a T1 (<2 cm)
tumor is 35 months versus 19.9 months for patients with a
T3 (>5 cm) tumor (54). These data support the suggestion
that metastasis can occur years prior to diagnosis. The time
required for the development of a malignant tumor and
progression from an adenoma to a malignant carcinoma
may be longer than the progression of an adenoma to a
metastasis, which may be because more mutations and
clonal expansion(s) are required for progression to malignancy as compared with the number required to form a
metastasis (55). The time between the appearance of a small
adenoma and the diagnosis of malignancy may be as long as 20
to 25 years based on studies in patients with familial adenomatous polyposis (56), and confirmed by serial studies of sporadic colorectal cancer that revealed the transition from a
large adenoma to carcinoma takes about 15 years (57). These
estimates are consistent with the doubling times of tumors,
determined by serial radiologic studies and serial measurements of the carcinoembryonic antigen (CEA) serum biomarker (58). This period differs from the observed mean doubling
times of a metastasis, which may be 2 to 4 months (55).
The Organ Microenvironment
Clinical observations of cancer patients and studies in
rodent models of cancer have revealed that some tumor
phenotypes tend to metastasize to specific organs, independent of vascular anatomy, rate of blood flow, and number of tumor cells delivered to an organ. Experimental
data supporting Paget's “seed and soil” hypothesis have
been derived from studies on the invasion and growth of
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
Figure 2. Time Line. Continued.
B16 melanoma metastases in specific organs (20). In these
studies, melanoma cells were injected intravenously into
syngeneic mice with tumor foci development in the lungs
and in fragments of pulmonary or ovarian tissue implanted
intramuscularly. In contrast, metastatic lesions did not develop in implanted renal tissue, or at the site of surgical
trauma, suggesting that metastatic sites are not solely
determined by neoplastic cell characteristics, or host microenvironment (20). Ethical considerations rule out experimental analysis of cancer metastasis in humans, but the
use of peritoneovenous shunts for the palliation of ascites
in women with progressive ovarian cancer has provided an
opportunity to study factors that affect metastatic spread
(47). Human ovarian cancer cells can grow in the peritoneal cavity, either in the ascites fluid or by attaching to
the surface of peritoneal organs. These malignant cells,
however, do not metastasize to other visceral organs.
One (incorrect) explanation for the lack of visceral metastases is that tumor cells cannot gain entrance into the
systemic circulation. Studies by Tarin and colleagues of
metastasis by ovarian tumors in patients whose ascitic
fluid was drained into the venous circulation addressed
this question (47). Clinically, this procedure resulted in
palliation with minimal complications; however, it allowed
the entry of viable cancer cells into the jugular vein. Autopsy findings from 15 patients substantiated the clinical
observations that the shunts did not significantly increase
the risk of metastasis to organs outside the peritoneal
cavity. Indeed, despite continuous entry of millions of tumor cells into the circulation, metastases to the lung, the
first capillary bed encountered, were rare (47). These results provide compelling verification of the “seed and soil”
hypothesis. Organ-specific metastases have also been
shown in studies of cerebral metastasis after injection of
syngeneic tumor cells into the internal carotid artery of
mice. Differences between two mouse melanomas were
www.aacrjournals.org
found on the basis of patterns of brain metastasis such
that the K-1735 melanoma produced lesions in the brain
parenchyma (59) and the B16 melanomas produced meningeal metastases (60). Thus, different sites of tumor
growth within an organ include differential interactions
between metastatic cells and the organ environment, possibly in terms of specific binding to endothelial cells and
responses to local growth factors (61).
Seed and Soil
Since the 1980s, many investigators have contributed to our
understanding of cancer pathogenesis and metastasis, and the
dependency of these processes on the interaction between
cancer and homeostatic factors (62). Our concept of the “seed
and soil” hypothesis consists of three principles. The first principle is that primary neoplasms (and metastases) consist of
both tumor cells and host cells. Host cells include, but are
not limited to, epithelial cells, fibroblasts, endothelial cells,
and infiltrating leukocytes. Moreover, neoplasms are biologically heterogeneous and contain genotypically and phenotypically diverse subpopulations of tumor cells, each of which has
the potential to complete some of the steps in the metastatic
process, but not all. Studies using in situ hybridization and immunohistochemical staining have shown that the expression
of genes and/or proteins associated with proliferation, angiogenesis, cohesion, motility, and invasion vary among different
regions of neoplasms (62, 63). Because of the heterogeneity of
tumors and the tumor infiltration by “normal” host cells, the
search for genes and/or proteins that are associated with
metastasis cannot be conducted by the indiscriminate and
nonselective analysis of tumor tissues.
The second principle is that a focused analysis into the metastatic process is needed to reveal the selection of cells that
succeed in invasion, embolization, survival in the circulation,
arrest in a distant capillary bed, and extravasation into and
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5653
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
Figure 3. The process of cancer metastasis consists of sequential, interlinked, and selective steps with some stochastic elements. The outcome of
each step is influenced by the interaction of metastatic cellular subpopulations with homeostatic factors. Each step of the metastatic cascade is potentially
rate limiting such that failure of a tumor cell to complete any step effectively impedes that portion of the process. Therefore, the formation of clinically
relevant metastases represents the survival and growth of selected subpopulations of cells that preexist in primary tumors.
multiplication within organ parenchyma (Fig. 3). These
successful metastatic cells (“seed”) have been likened to a
decathlon champion who must be proficient in 10 events,
rather than just a few (64). However, some steps in this pro-
cess incorporate stochastic elements. Overall, metastasis favors the survival and growth of a few subpopulations of
cells that preexist within the parent neoplasm. The current
data, especially studies focused on isolated tumor cells,
Table 1. Steps in the metastatic process
Step
1
2
3
4
5
6
7
8
9
10
5654
Description
After the initial transforming event, the growth of neoplastic cells is progressive and frequently slow;
Vascularization is required for a tumor mass to exceed a 1- to 2-mm diameter (200, 201), and the synthesis and secretion
of angiogenesis factors has a critical role in establishing a vascular network within the surrounding host tissue (201);
Local invasion of the host stroma by tumor cells can occur by multiple mechanisms, including, but not limited to,
thin-walled venules and lymphatic channels, both of which offer little resistance to tumor cell invasion (202);
Detachment and embolization of tumor cell aggregates, which may be increased in size via interaction with
hematopoietic cells within the circulation;
Circulation of these emboli within the vascular; both hematologic and lymphatic;
Survival of tumor cells that trafficked through the circulation and arrest in a capillary bed;
Extravasation of the tumor embolus, by mechanisms similar to those involved in the initial tissue invasion;
Proliferation of the tumor cells within the organ parenchyma resulting in a metastatic focus;
Establish vascularization, and defenses against host immune responses; and
Reinitiate these processes for the development of metastases from metastases.
Cancer Res; 70(14) July 15, 2010
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
support a clonal origin for metastases, such that different
metastases originate from different single cells (21, 64). The
third principle is that metastatic development occurs in
specific organs, or microenvironments (“soil”), that are biologically unique. Endothelial cells in the vasculature of different organs express different cell-surface receptors (65) and
growth factors that can support or inhibit the growth of metastatic cells (66). Thus, the outcome of metastasis depends on
multiple interactions (“cross-talk”) of metastasizing cells
with homeostatic mechanisms, such that the therapy of
metastases can target not only the cancer cells, but also
the homeostatic factors that promote tumor-cell growth,
survival, angiogenesis, invasion, and metastasis (67).
As discussed above, the analysis by Stephen Paget showed
that breast tumors preferentially spread to the liver, but not
the spleen. The bone marrow is also a preferential target for
breast cancer metastases, with some patients developing metastases more frequently than others. These observations (68,
69) identify that the site of secondary tumors is dependent on
the tumor cell (“seed”) and the target organ (“soil”). Therefore,
a metastatic focus is established only if the seed can grow in
the soil, that is, if the microenvironment of the target site
is compatible with the properties and requirements of the
disseminated tumor cell. Although this hypothesis was challenged (2), it is now largely accepted that the anatomic architecture of the blood flow is not sufficient to fully describe the
patterns of metastatic tumor spread. There are several molecular and cellular explanations that support the seed and soil
hypothesis, including the observations that endothelial cells
lining blood vessels in different organs express different
adhesion molecules (61); results from in vivo phage display
studies suggest that every vascular bed may have its own
specific molecular “address” (70); and the observation that
tumor cells expressing the corresponding receptor may
use these receptor-ligands to traffic to and arrest in specific
tissues. There is also compelling evidence supporting a role
for chemokines in the homing or chemoattraction of organselective metastasis (71). The CXCR4 receptor seems to have
a role in the bone marrow, lymph nodes, and pulmonary
targeting of metastases from breast tumors that express
its ligand, CXCL12 (72). CXCR4 regulates chemotaxis in vitro
and CXCR4-blocking antibodies have been shown to impair
lung metastasis in SCID mouse xenograft experiments (71).
However, in vivo, CXCR4 activation can also promote the
outgrowth of metastases in specific tissues rather than invasion (73). Recently, an immunohistochemical analysis of
CXCL12 expression in tumor endothelial cells was reported
to have a significant correlation with survival in patients with
brain metastases (74). Additional support for organ-specific
targeting has come from the gene-expression analysis of
breast cancer cells selected for increased metastasis to bone
(75) or lungs (76) that identified genes functionally involved
in organ-specific metastasis.
Cellular Infiltration of Tumors and Metastases
The role of tumor infiltration by leukocytes in tumor growth
and development is complex. Although tumor-associated
www.aacrjournals.org
macrophages (TAM) have tumoricidal activity and can stimulate antitumor T cells, tumor cells can block inflammatory
cell infiltration and the function of infiltrating cells (77).
Tumor-derived molecules can regulate the expansion of myeloid progenitors (Fig. 4), their mobilization from the marrow, chemo-attraction to tumors, and activation, resulting
in the promotion of tumor survival, growth, angiogenesis,
and metastasis. Numerous studies have shown that following
activation, macrophages can kill tumor cells in vitro (78).
Indeed, if a macrophage-activating agent is injected prior
to isolation of macrophages from metastases, they have tumoricidal activity in vitro (78). However, tumor infiltration
by macrophages seems to have predominantly a protumorigenic and/or metastatic phenotype; this was initially
shown in a study in which the macrophage content of
six carcinogen-induced fibrosarcomas was reported to directly correlate with immunogenicity and, inversely, with
metastatic potential (79). Other studies using human breast
carcinomas and melanomas have been equivocal (80). One
clinical study showed that patients with metastatic disease
had a low macrophage infiltration (≤10%) of their primary
neoplasm, whereas 13 patients with benign tumors and 6
out of 31 patients with malignant tumors and no clinical evidence of metastases had a TAM frequency of ≥10% (80).
Thus, most (81–83), but not all studies (84), have shown that
there is no relationship between immunogenicity, metastatic
propensity, and TAM frequency. Macrophages differentiate
from CD34+ bone marrow progenitors following expansion
and commitment and are mobilized from the marrow into
the periphery (56, 85), in which they differentiate into
monocytes and following invasion into a tissue, mature
into macrophages (86). Macrophage infiltration of primary
tumors is regulated, at least in part, by cytokines, growth
factors, and enzymes secreted by the primary tumor. The
primary tumor can also regulate the function of TAMs,
including tumor cell cytotoxicity, which can be tumor
dependent (87).
Clinical studies of macrophage infiltration of tumors have
suggested that it does not correlate with the immunogenicity
and metastatic propensity (81–83, 88); rather, it is associated
with a poor prognosis (89). Thus, a low frequency of macrophage infiltration does not guarantee that a tumor will metastasize as multiple factors are critical (90). Macrophage
infiltration of tumors is regulated by tumor-associated chemoattractants such that host and/or tumor interactions
may result in qualitative (87) or quantitative (91) regulation
of macrophage infiltration, which can be obscured by the
technologies used to assess macrophage infiltration (82, 92).
Macrophages present in the extracellular matrix and capsular
area of tumors can facilitate tumor invasion and are important
during the development of early stage lesions (93). Further,
the proteolytic enzyme secreted by activated macrophages
facilitates tumor invasion and extravasation. Indeed, in vitro
coculture of macrophages with tumor cells can accelerate
tumor growth (94). Macrophage infiltration of some tumor
models has been found to inversely correlate with relapse-free
survival (RFS), microvessel density, and mitotic index (95).
TAMs also express a number of factors (96) that stimulate
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5655
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
Figure 4. Tumors can secrete growth factors and cytokines that result in the expansion and mobilization of myeloid progenitors from the marrow with
trafficking to various extramedullary sites including the spleen, liver, lungs, and primary and metastatic tumor lesions. These committed myeloid progenitors
(CMP) can mature into DCs, MDSCs, and macrophages including TAMs, as well as become activated, or “paralyzed,” within the tumor environment.
These heterogeneous cells include progenitor cells, immature cells, mature, and activated cells. Dependent upon the infiltrating subset and extent of
maturation and activation, these cells can be a critical component and regulator of angiogenesis, vascularogenesis, and tumor regression or growth.
tumor cell proliferation and survival and support angiogenesis, a process essential for tumor growth to a size larger
than the 1 to 2 mm3 size that can occur in the absence of
angiogenesis.
Myeloid-derived suppressor cells (MDSC) have also been
identified in the circulation of tumor-bearing hosts and infiltrating tumors. MDSCs in mice are CD11b+Gr-1+ (97, 98),
whereas the human equivalents, which were originally described in the peripheral blood of squamous cell carcinoma
of the head and neck patients (99), and, more recently, other
cancer histiotypes, are DR−CD11b+ (99–101). Tumor growth
is associated with the expansion of this heterogeneous cellular population, which can inhibit T-cell number and function.
The mechanisms of MDSC immunosuppression are diverse,
including upregulation of reactive oxygen species (ROS), nitric oxide (NO) production, L-arginine metabolism, as well as
secretion of immunosuppressive cytokines. The immunosuppressive functions of MDSCs were initially described in the
late 1970s when they were identified as natural suppressor
(NS) cells and defined as cells without lymphocyte-lineage
markers that could suppress lymphocyte response to immunogens and mitogens (102, 103).
5656
Cancer Res; 70(14) July 15, 2010
Granulocyte-macrophage colony stimulating factor (GMCSF) has been directly associated with MDSC-dependent
T-cell suppression (97), which can be reversed by blocking
antibodies (104). Mice bearing transplantable tumors that
secrete GM-CSF have increased numbers of MDSC and
suppressed T-cell immunity (105), a finding that contrasts
with the adjuvant activity of GM-CSF for tumor vaccines
(106). This difference is associated with GM-CSF levels
such that high levels expand MDSC numbers and reduce
vaccine responses, whereas lower levels augment tumor
immunity. Vascular endothelial growth factor (VEGF) has
also been directly linked with MDSC expansion and tumor
progression (107). VEGF can suppress tumor immunity
(108) via an inhibitory effect on dendritic cell (DC) differentiation (109). Thus, there is a correlation between plasma VEGF levels in cancer patients, a poor prognosis (110),
and VEGF-induced abnormalities in DC differentiation
(111), resulting in an inverse correlation between DC frequency and VEGF expression (112). As expected, neutralizing VEGF antibodies can reverse not only VEGF-induced
defects in DC differentiation (112), but also improve DC
differentiation in tumor-bearing mice (113).
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
Recently, Finke and colleagues (114) reported that administration of sunitinib, a receptor tyrosine kinase inhibitor, resulted in a significant decrease in MDSC within patients with
metastatic renal cell carcinoma (115). Whether this decrease
was associated with a reduction in VEGF levels and neoangiogenesis or as an inhibition of Flt3-mediated expansion of
MDSC remains to be addressed. Regardless of this observation there have been few studies that examined the infiltration of tumors by MDSCs. In one recent rodent study, it was
shown that CCL2 mediated MDSC chemotaxis in vitro and
that migration or chemotaxis of MDSCs could be blocked
with neutralizing CCL2 antibodies (Abs) or by blocking
CCR2. Sunitinib has also been shown to mediate reversal of
MDSC accumulation in renal cell carcinoma (115). These
observations suggest that the regulation of infiltrating myeloid cells has the potential to control the growth of primary
tumors and metastasis (Fig. 5).
Metastatic Heterogeneity
Cells with different metastatic properties have been isolated from the same parent tumor, supporting the hypothesis that not all the cells in a primary tumor can
successfully metastasize (41–43). Two approaches have
been to show that cells within the parent neoplasm differ
in metastatic capacity. In the first approach, metastatic
cells are selected in vivo, such that tumor cells are implanted at a primary site or injected intravenously, metastases allowed to grow, individual lesions collected and
expanded in vitro, and these cells used to repeat the process. This cycle is repeated several times and the function
of the cycled cells compared with that of cells from the
parent tumor to determine whether the selection process
resulted in an enhanced metastatic capacity (116). Several
studies have shown that the increase in metastatic capacity did not result from the adaptation of tumor cells to
preferential growth in a specific organ but, rather, was
due to selection (23, 24, 117). This procedure was originally
used to isolate the B16-F10 line from the wild-type B16
melanoma (116), but has also been successfully used to
produce tumor cell lines with increased metastatic capacity from many murine and human tumors (118, 119).
In the second approach, cells are selected for an enhanced
expression of a phenotype believed to be important to a step
in the metastatic process and are then tested in vivo to determine whether their metastatic potential has changed. This
method has been used to examine properties as diverse as
resistance to T lymphocytes (120), adhesive characteristics
(121), invasive capacity (122, 123), lectin resistance (124),
and resistance to natural killer cells (125). The finding that
a characteristic, typically using a single tumor model with
only one or two variants, is associated with metastasis is often considered as proof that this parameter is directly responsible for metastasis. However, this can be model
dependent, associated with the selective procedure and has
resulted in conclusions and associations that are not reproducible. Clearly, numerous phenotypes are associated, in
part, with the metastatic process, but rarely represent all
the phenotypes critical to the final outcome. Thus, studies
of this nature, as well as comparisons of clonal subpopulations, lend insight into phenotypes that may have a role in
metastasis, but can lead one astray in terms of overall conclusions into the process. This finding is also true of microarray analyses, whereby, the primary tumor can have
multiple cellular phenotypes because of clonal heterogeneity,
as well as the confounding variable of infiltrating “normal”
host cells obfuscating conclusions. As such, conclusions
based on a limited number of tumor samples and/or clones,
into a process as complex as metastasis, have the potential to
be unsupportable.
One criticism of studies into metastatic heterogeneity is
that isolated tumor line(s) may have arisen as a result of
Figure 5. The clonal selection model of metastasis suggests that the cell populations in the primary tumor with all of the genetic prerequisites required for
metastatic capacity are the subpopulations that metastasize. Further, both the cells within the primary tumor and the metastatic lesion(s) can continue
to diversify as the lesions grow.
www.aacrjournals.org
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5657
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
an adaptive rather than a selective process, which was addressed by the analysis of metastatic heterogeneity by Fidler
and Kripke in 1977 using the murine B16 melanoma (10).
A modification of the fluctuation assay of Luria and Delbruck
(126) showed that different tumor cell clones, each derived
from individual cells isolated from the parent tumor, varied
in their ability to form pulmonary nodules following intravenous inoculation into syngeneic mice. Control subcloning
procedures showed that the observed diversity was not a
consequence of the cloning procedure (10). To exclude the
possibility that the metastatic heterogeneity found in the
B16 melanoma might have been introduced as a result of
in vivo or in vitro cultivation, the biologic and metastatic
heterogeneity of a mouse melanoma induced in a C3H/HeN
mouse was also studied (127). This melanoma designated
K-1735 was established in culture and immediately cloned
(128, 129). An experiment similar to that undertaken with
the B16 melanoma confirmed that (10) the clones differed
from each other and the parent tumor in their ability to
produce lung metastases. In addition, the clones differed in
metastatic potential, resulting in metastases that varied in
size and pigmentation. Metastases to the lungs, lymph nodes,
brain, heart, liver, and skin were not melanotic, whereas
those growing in the brain were uniformly melanotic
(59, 128, 129).
An extension of studies into metastatic heterogeneity
assessed immunologic rejection by a normal syngeneic host
(130, 131), as compared with young nude mice (132), a
model in which the immunologic barrier to metastasis is
removed and antigenic metastatic cells are able to successfully complete the process. In these studies, cells of two
clones that did not produce metastases in normal syngeneic
mice produced lung metastasis in the young nude mice.
However, most of the nonmetastatic clones were nonmetastatic in both normal syngeneic and nude recipients. Therefore, the ability to metastasize in syngeneic mice was
primarily not due to immunologic rejection, but by other
deficiencies that prevented completion of a step in the
metastatic cascade.
The observation that preexisting tumor subpopulations
with metastatic properties exist in the primary tumor has
been confirmed by many laboratories using a wide range
of tumors (16, 26, 30, 53, 133–135). In addition, studies using young nude mice as models for metastasis of human
neoplasms have shown that human tumor cell lines and
freshly isolated human tumors, such as colon and renal carcinomas, also contain subpopulations of cells with widely
differing metastatic properties. Talmadge and Fidler undertook a series of studies to address the question of whether
the cells that survive to form metastases possess a greater
metastatic capacity than cells in an unselected neoplasm
(23, 25, 116). In these studies, cell lines derived from spontaneous metastatic foci were found to produce significantly
more metastases than cells from the parent tumor. Studies
with heterogeneous, unselected tumors support the hypothesis that metastasis is a selective process, regulated by
a number of different mechanisms. It should be noted that
metastatic foci can be clonal in origin, yet originate from
5658
Cancer Res; 70(14) July 15, 2010
tumor emboli containing more than a single tumor cell. Tumor emboli composed of heterogeneous tumor cells can
arise if an embolus is derived from a nonhomologous zone
within a primary tumor (136). The zonal composition of a
tumor may play a role in the metastatic process, but also
has the potential to obscure the results of microarray studies. The zonal composition can result in the appearance of
minimal heterogeneity if a sample is obtained from a single
zone within a neoplastic lesion. Thus, as recommended by
Bachtiary and colleagues (137), microarray analyses need to
consider intratumoral heterogeneity and zonal gene expression, such that multiple tumor biopsies should be studied
from discontiguous sites. Moroni and colleagues (138), using human colorectal carcinomas, reported intratumoral
heterogeneity and epidermal growth factor receptor (EGFR)
gene amplification that correlated with protein levels. Experiments using human tumors in nude mice by Pettaway
and colleagues (139) also revealed that orthotopic implementation of heterogeneous human prostate cancer cell lines into
nude mice resulted in rapid local growth and distant metastatic foci with an increased metastatic propensity to various
organ sites including the gastrointestinal tract. The clonal origin of these metastatic foci was documented with karyotypic
markers extending and confirming the prior studies by Fidler
and Talmadge (21).
The Clonal Origin of Metastases
Nowell (140, 141) suggested that acquired genetic variability within developing clones of tumors, coupled with
selection pressures, results in the emergence of new tumor
cell variants that display increased malignancy. This hypothesis suggested that tumor progression toward malignancy is accompanied by heightened genetic instability of
the progressing cells. To test this hypothesis, Cifone and
Fidler measured the mutation rates of paired metastatic
and nonmetastatic cloned lines isolated from four different
murine tumors (142). They found that highly metastatic
cells were phenotypically less stable than their benign
counterparts. Moreover, the spontaneous mutation rate of
the highly metastatic clones was several fold higher than
in nonmetastatic or poorly metastatic clones. These observations were subsequently confirmed and extended with
other murine (143) and human neoplasms (144, 145). Taken together, these studies suggest that metastatic tumor
populations have a greater likelihood of cells undergoing
rapid phenotypic diversification. This genetic instability,
coupled with a Darwinian selection, may result in populations resistant to normal homoeostatic growth controls,
chemotherapeutic intervention, immune attack, and environmental restraints (146). Indeed, this process can be extended by the mutagenic activity of drugs commonly used
to treat tumors (147). This biological heterogeneity is
found both within a single metastasis (intralesional heterogeneity) and among different metastases (interlesional heterogeneity). The heterogeneity reflects two processes: the
selective nature of the metastatic process and the rapid
evolution and phenotypic diversification of clonal tumor
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
growth, resulting from the inherent genetic and phenotypic
instability of clonal tumor cell populations. Like primary
neoplasms, metastases have the potential for a unicellular
or multicellular origin. To determine whether metastases
arise from the same clone, or whether different metastases
are produced from different progenitor cells, Talmadge and
colleagues designed a series of experiments based on the
gamma-irradiation induction of random chromosome
breaks and rearrangements to serve as “markers” (148).
They examined the metastatic phenotype of individually
spontaneous lung metastases that arose from subcutaneously implanted tumors (148). In 10 metastases, all the
chromosomes were normal, making it impossible to establish whether they were of uni- or multicellular origin.
In other lesions, unique karyotypic patterns of abnormal
marker chromosomes were found, indicating that each
metastasis originated from a single progenitor cell.
Subsequent experiments showed that when heterogeneous clumps of different melanoma cell lines were
injected intravenously, the resulting lung metastases were
of unicellular origin (21). These studies indicated that
regardless of whether an embolus was homogeneous
or heterogeneous, metastases originate from a single
proliferating cell. The clonality of metastases has also
been reported for numerous other tumors, including
breast cancer and fibrosarcomas, as well as melanoma (145).
The role of clonal selection (Fig. 6) during the process
of metastasis and the emergence of successive clonal subpopulations has been supported by studies in which individual cells were tagged by unique markers allowing them
to be tracked (149). This tracking has been undertaken
with radiation-induced cytogenetic (chromosomal) markers
(28), which clearly showed clonal selection. This observation was confirmed with the use of drug resistance mar-
kers (150) and X-linked isoenzyme mosaicisms (151).
However, all of these approaches are limited and recent
studies have used markers on the basis of the random insertion of transfected plasmids, which results in the generation of large numbers of uniquely marked cell clones
(139, 152, 153). Clearly, clonal selection and intrahost
metastatic heterogeneity are supported by preclinical
studies (148, 150, 151, 153–155). However, clinical confirmation has been challenging owing to methodological
impediments. Metastatic tissue is generally obtained
asynchronously, relative to primary tumor tissue, and comparisons complicated by tissue availability. Several studies
have successfully compared primary tumors to metastatic
foci within the same patient on the basis of a number of
different phenotypes. Kuukasjarvei and colleagues (44) analyzed the genetic composition of 29 primary breast carcinomas and paired asynchronous metastases. They found
that 69% of the metastases had a high degree of clonality with the corresponding primary tumor, whereas
chromosomal X inactivation patterns supported the remaining metastatic lesions as originating from the
same clone as the primary tumor. It was concluded that
although all metastases were derived from the parent
tumor, metastasis occurred at different times. Studies
using microarray analyses have also addressed this
process and are discussed in the section below entitled
“Microarray analysis of heterogeneity, clonality, and
prognostic potential.”
Results from the microarray and computational analyses have revealed that a small number of genes are differentially expressed between tumors and metastases,
supporting the conclusion that although metastases resemble the primary tumor on a mutational basis, a few
genes differ consistently and are of significant mechanistic
importance.
Figure 6. The immune cells infiltrating tumors can regulate their growth, progression, and metastasis. Tumor regression is associated with infiltration by
mature DCs and cytotoxic T cells (CTL) and type 1 T-helper type 1 bioactivity. Contrasting with this, tumor growth is facilitated via immune-mediated
immunosuppression and neoangiogenesis following infiltration of tumors with immature DCs, MDSCs, plasmacytoid DCs (pDC) M2 macrophages, as well
as T-regulatory (T-reg) cells and a low frequency of CD4 and CD8 effector T cells.
www.aacrjournals.org
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5659
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
Current Controversies in Metastatic Research
Premetastatic niche
Recently, it was suggested that to form a metastatic focus, primary tumors produce factors that induce the formation of an appropriate environment in the soil (organ) prior
to the seeding of metastatic cells. This suggestion led to the
concept of a premetastatic niche, whereby a special, permissive microenvironment in the secondary site is developed (156). The initial study into a premetastatic niche
indicated that vascular endothelial growth factor receptor
1 (VEGFR1+) hematopoietic progenitor cells contribute to
metastatic spread (157). These VEGFR1+ cells were suggested to arrive at a distant organ site, forming a premetastatic niche of clustered bone marrow hematopoietic cells
(157). They were suggested to alter the tissue microenvironment and promote the recruitment of tumor cells by the
upregulation of integrins and cytokines. The blockade of
VEGFR1 cells in vivo has been suspected to abrogate the
formation of premetastatic niches and inhibit the growth
of metastatic foci. The regulation of VLA-4 and fibronectin
by VEGFR1+ cells also helps dictate which organs attract
hematopoietic progenitors and support their adhesion,
thereby promoting premetastatic niches (157). In addition,
this adhesion molecule expression may contribute to survival of the metastatic tumor cells as discussed in the section
on MDSCs. Gao and colleagues showed that angiogenesis is
essential for the growth of micrometastases (158), and that
this process was regulated by bone marrow–derived endothelial progenitor cells (EPC). At the micrometastatic stage,
EPCs traffic to and colonize micrometastases. Accelerating
angiogenesis facilitates the growth into gross metastases. The blockade of EPCs by short hairpin RNA directed
against the Id1 gene was found to inhibit angiogenesis at
the micrometastatic stage and to impair subsequent growth
(158). Furthermore, although the number of EPCs contributing to angiogenesis in metastases remained low (in the
range of 11%), their impact on the development of micrometastases and rapid cell proliferation is critical for progressive growth of metastasis (159).
Thus, experimental evidence supports the formation of
a premetastatic niche, which can facilitate metastasis via
VEGFR1+ bone marrow–derived hematopoietic progenitor
cells (157). These cells express hematopoietic markers,
and chemokines and chemokine receptors (e.g., CXCL12/
CXCR4), promoting either their homing to the target tissue or recruitment and/or attachment of tumor metastatic
cells. This process includes the CXCL12/CXCR4 axis,
which has a critical role in stem cell migration (160). Activation of CXCR4 induces motility, chemotactic response,
adhesion, secretion of MMPs, and release of angiogenic
factors, such as VEGF-A. Thus, disseminating tumor cells,
in order to survive at distant sites, need a supportive microenvironment similar to that found in the primary tumor. One way this may occur is by the mobilization and
recruitment to a premetastatic niche of immune and/or inflammatory cells that can stimulate growth of metastatic
cells (161).
5660
Cancer Res; 70(14) July 15, 2010
Multiple cellular and molecular mechanisms, whereby
tumor cells interact with immune cells, regulate their
number and function, and allow tumor cells to escape
from immunologic recognition and control (Fig. 4). These
immunoregulatory events have important physiological and
pathophysiological significance, not only for host-tumor interactions and clinical outcomes, but also for clinical intervention. Studies to date have focused on the inflammatory
cell infiltration within tumors and draining lymph nodes and,
more recently, subset analyses. T-lymphocytic responses to
tumors are generally associated with a good prognosis; however, vaccine strategies have had little impact to date (162).
This is due, in part, to the escape mechanisms that tumors
use to circumvent immunologic responses. Host-tumor interactions limit lymphocyte infiltration such that the majority of
infiltrating cells are from the innate immune system (163)
and are typically immature or with impaired function (162).
These cells can “paralyze” the induction of T-cell and DC responses (164), in addition to facilitating tumor proliferation
and metastasis (165) via pro-angiogenic activity. Indeed, the
immune cells infiltrating tumors can secrete immunosuppressive enzymes, resulting in lymphocytic apoptosis and
the induction of tolerance (166). Critical questions remain,
including how tumors become refractory to immune intervention strategies to overcome these immunosuppressive
mechanisms.
Epithelial-mesenchymal transitions
Molecular technologies, including DNA, antibody, and proteomic arrays, and short-hairpin RNA libraries, have been
used to identify multiple molecules that contribute to the
metastatic process. This group includes growth factors, cytokines and chemokines, pro-angiogenic factors, extracellular
matrix-remodeling molecules, and, most recently, transcription factors that may regulate cellular changes during tumor
invasion-metastasis. These transcription factors include Slug,
Snail, Goose-coid, Twist, and ZEB1, which are highly expressed by metastatic cells and have been suggested to have
a role in inducing epithelial-mesenchymal transitions (EMT;
refs. 167–170), defined as a rapid and reversible change in
cellular phenotypes. This concept originated from studies into embryologic development, including the change in phenotype and behavior of cells invaginating from the surface into
the interior of an embryo to form the mesoderm during gastrulation (171), changes during renal organogenesis, and the
origin and fate of the neural crest (172). EMT studies have
focused on the invasive behavior and spindling of cultured
cells (173), using explanted embryonic tissue (174). Thus,
the potential relevance of EMT to tumor progression is
largely based on in vitro studies (175), using transformed
epithelial cells and growth factors or extracellular matrix
manipulations. Genes upregulated during tumor progression and metastasis are associated with multiple processes,
including embryogenesis, tissue morphogenesis, and wound
healing (176). In neoplasia, EMT is suggested to result in
the loss of epithelial properties, including cell-cell adhesion
and baso-apical polarity, and the gain of mesenchymal
properties, such as an increased ability to migrate on and
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
invade through extracellular matrix proteins (176). EMT has
also been associated with the loss of expression and/or
mislocalization of proteins involved in the formation of
cell-cell junctions, such as E-cadherin, zonula occludens
(ZO-1) and/or claudin, and the gain of mesenchymal protein expression. Thus, a depression in E-cadherin expression
is frequently observed during cancer progression (171), and
solid cancers are associated with a loss or downregulation
of E-cadherin expression (171, 177).
EMT has been suggested to have a role in the progression
of benign tumor cells to invasive and metastatic cells (171).
This suggestion is largely based on cell culture, microarray,
and histiologic studies. EMT proponents have suggested that
tumors that display malignant histopathologic features
including loose, scattered cancer cells and spindle cell cytology, which are more aggressive based on grade, stage, and
clinical outcome, support the concept that EMT is mechanistically significant (168). Inarguably, carcinomas are histologically heterogeneous in addition to being infiltrated by
mesenchymal cell histiotypes. The histologic identification
of mesenchymal elements in human tumors, identified as
fetal-type fibroblasts and malignant stroma was described
years ago (178). Indeed, a collection of 100 dual-staining
spindle cell breast carcinomas was reported in 1989 (179).
Further, antibodies against keratin 14 and smooth muscle
actin have identified spindle cell elements, which may or
may not be clinically significant (180, 181). Foci with a histologic appearance of EMT are found in human tumors and
this anaplastic appearance, including loss of cell polarity and
lineage-specific or tissue-specific cytology are common features of carcinomas. The morphologic disorder, pleomorphism, and anaplasia as observed by an unskilled observer
may be suggestive of a mesenchymal lineage. The examples
cited as evidence of EMT in clinical neoplasms—including
scattered single cell infiltration by lobular carcinomas of
the breast (171) or diffuse carcinomas of the stomach (182),
spindle cell differentiation in squamous carcinomas, and
blending of sheets of carcinomatous cells into a highly cellular,
desmoplastic stroma without a clear line of demarcation—
potentially represent inappropriate analysis of these lesions.
As discussed below, diffusely infiltrating cells in lobular carcinomas retain their epithelial identity.
The data supporting EMT, on the basis of in vitro observations, have largely used two-dimensional environments that
do not contain host stroma, such as vascular, immune,
endocrine, or neurologic elements. Similarly, studies of cells
in three-dimensional matrices do not recapitulate the host
environment, and the identification of cells in vitro as epithelial or mesenchymal in origin, based on morphology, is subjective. Indeed, studies in which epithelial and mesenchymal
cell markers are used to provide evidence for a switch from
one differentiated cell lineage to another are also suspect as
they relate a marker to a morphologic phenotype, which
itself is claimed to transmute into another cell type and thus
the identification process becomes self-fulfilling. Inappropriate gene expression by tumors is well established, and
genetically unstable cells and infiltrating cells could contribute to shifts in lineage markers. Indeed, it is difficult to
www.aacrjournals.org
define cells on morphologic criteria alone, and the gain or
loss of markers is insufficient to conclude that a cell population has undergone whole-scale gene expression reprogramming. Although the presence of EMT is largely argued
on the basis of evidence from in vitro experiments, the in vivo
data are unclear (183). Whether the progression of a noninvasive tumor into a metastatic tumor involves EMT (177,
183) needs further study. This controversy is extended by
the rarity of EMT-like morphological changes in primary tumors and the observation that metastases are histologically
and molecularly similar to the primary tumor. Thus, the role
of EMT in tumor metastasis is difficult to assess as it is challenging to obtain tumor cells that have activated in the EMT
program for or during metastasis (171).
Microarray analysis of heterogeneity, clonality, and
prognostic potential
Expression profiling has revealed “metastasis signatures”
or poor prognosis gene signatures expressed by neoplastic
cells within individual carcinomas and the potential to predict metastasis (5, 184). This finding has been suggested to
infer that metastatic propensity might be an intrinsic property of a primary tumor, which is developed relatively early in
multistep tumor progression and is, therefore, expressed by
the bulk of neoplastic cells within a tumor. It has also been
proposed (75, 76) that a set of genes may mediate metastasis
to specific organs, such as to the bone marrow or lungs.
Studies by Urquidi and colleagues (9) provided direct proof
that individual cancer cells coexist within a tumor, differ in
metastatic capability, and that metastatic primary tumors
contain tumor subpopulations with variant metastatic proclivities and expression profiles. Montel and colleagues
(185) reported that as the metastatic proclivity of a tumor
increases, the cellular populations within the tumor develop
increasingly variable expression profiles. They suggested, on
the basis of the concept of tumor progression (140), that the
metastatic foci selected during clonal evolution represent the
final stage in the development of a metastasis. However, as
we have reviewed here, tumor progression is a continuum
and does not end with metastasis. It continues during the
growth of primary tumors and metastatic nodule(s), including the development and selection of variants resistant to
chemotherapy. Tumor cells within a metastasis, at least initially, express all of the genetic alterations necessary for the
metastatic process. However, as tumors progress, heterogenic tumor subpopulations develop within a metastasis, including cells with different metastatic capabilities. Gancberg
and colleagues (186) examined pairs of primary and metastatic tumors from 100 breast cancer patients and reported
that 6% had discordant Her2/neu overexpression by the metastatic tissue as compared with the primary tumor. FISH
analysis from 68 of the patients revealed a 7% discordance
between primary and secondary lesions, but from different
patients than those found by immunohistochemistry. In
addition, they examined Her2/neu overexpression from
multiple metastatic lesions in 17 patients, revealing that
18% differed in Her2/neu expression. Suzuki and Tarin (8)
also examined matched, primary breast tumors and their
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5661
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
Table 2. Metastasis timeline
Year
Discovery
1829
1858
1889
Jean Claude Recamier developed the term “metastasis” (1).
Rudolf Virchow suggested that metastatic tumor dissemination was determined by mechanical factors (18).
Stephen Paget proposed the “seed and soil” hypothesis to explain the organs that are afflicted with
disseminated cancer (17).
Makoto Takahashi developed the first murine model of metastasis (203).
James Ewing proposed that metastasis was determined by the anatomy of the vascular and lymphatic channels
that drain the primary tumor (19).
Dale Rex Coman identified the role of cellular adhesiveness to metastasis and the distribution of
embolic tumor cells (204).
Irving Zeidman showed the transpulmonary passage of tumor cell emboli resulting in metastases on the
arterial side (205).
Balduin Lucke studied the organ specificity of tumor growth following intravenous injection of tumor
cells and comparing growth in the liver and lung (206).
Eva Kline showed that cells adapted to ascites growth preexist within the parent tumor and have an
increased malignant phenotype (207).
Gabriel and Tatiana Gasic showed that enzymatic manipulation of cell surfaces can affect metastatic potential (208).
Bernard and Edwin Fisher used 51chromium-labeled tumor cells in quantitative dissemination studies (209).
Bernard and Edwin Fisher distinguished lymphatic from hematogenous metastasis (210).
Isaiah Fidler reported that metastasis can result from the survival of only a few tumor cells using
IUdR-labeled tumor cells (22).
Isaiah Fidler reported the in vivo selection of tumor cells for enhanced metastatic potential (116).
Beppino Giovanella and coworkers showed that human tumors can metastasize in thymic-deficient nude mice (211).
Irwin Bross proposed the metastatic cascade to define a number of sequential events needed for
disseminated cancer (212).
Garth Nicolson proposed that the organ specificity of metastasis is determined by cell adhesion (213).
Peter Nowell proposed the clonal evolution of tumor-cell population (140).
Lance Liotta and Jerome Kleinerman linked invasion and metastasis to the production of proteolytic
enzymes by metastatic cells (214).
Isaiah Fidler and Margaret Kripke showed the metastatic heterogeneity of neoplasms (10).
Lance Liotta showed metastatic potential is correlated with enzymatic degradation of basement
membrane collagen (215).
Ian Hart and Isaiah Fidler showed the organ specificity of metastasis using ectopic organs (20).
James Talmadge and collaborators showed the regulatory role of natural killer cells in tumor metastasis (216).
George Poste and Isaiah Fidler reported that interactions between clonal subpopulations could
stabilize their metastatic properties, whereas a clonal subpopulation is unstable, which, following a short coculture,
results in the emergence of metastatic variants (13).
Karl Hellmann designed first antimetastasis drug clinical trial (Razoxane; 217).
James Talmadge and Sandra Wolman showed that cancer metastases are clonal and that metastases originate
from a single surviving cell (148).
James Talmadge and Isaiah Fidler provided direct evidence that the metastatic process was selective using
spontaneous metastases from multiple metastatic variants (25).
George Poste and Irving Zeidman showed the rapid development of metastatic heterogeneity by clonal
origin metastases (150).
Garth Nicolson and Susan Custead showed that metastasis is not due to adaption to a new organ environment
but rather is a selective process (134).
David Tarin and colleagues reported the evidence of organ-specific metastasis in ovarian carcinoma patients
who were treated with peritoneovenous shunts (47).
James Kozlowski and coworkers showed metastatic heterogeneity of human tumors using immune
incompetent mice (14).
Avraham Raz and coworkers showed tumor heterogeneity for motility and adhesive properties and their
association with metastasis (218).
1915
1929
1944
1952
1952
1955
1962
1965
1966
1970
1973
1973
1975
1975
1976
1976
1977
1980
1980
1980
1981
1982
1982
1982
1982
1982
1984
1984
1984
(Continued on the following page)
5662
Cancer Res; 70(14) July 15, 2010
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
Table 2. Metastasis timeline (Cont'd)
Year
Discovery
1985
1986
1988
1990
Isaiah Fidler addressed the critical role of macrophages in the process of metastasis (219).
Leonard Weiss and coworkers identified the concept of metastatic inefficiency (220).
Patricia Steeg and collaborators identified the first metastasis suppressor gene (221).
Richard Wahl and coworkers showed the ability of 2-deoxy-2[18F]fluoro-D-glucose (FDG) to detect lymph
node metastases with positron emission tomography (PET) scans (222).
Lloyd Culp and collaborators documented the utility of bacterial LacZ as a marker to detect micrometastases
during tumor progression (223).
Judah Folkman and coworkers showed a relationship between metastasis and angiogenesis in patients with
breast cancer (200).
Jo Van Damme reported the role of chemokines in tumor-associated macrophage facilitation of metastasis (225).
Judah Folkman showed that the removal of a malignant primary tumor in mice spurs the growth of remote
tumors, or metastases (226).
Robert Hoffman and colleagues demonstrated visualization of tumor cell invasion and metastasis using green
fluorescent protein expression (227).
David Botstein showed diversity in the gene expression patterns by breast cancer tissues (228).
Irving Weissman and coworkers suggested the role of CSCs in metastasis (229).
Rene Bernards and Robert Weinberg proposed that metastatic potential is determined early in tumorigenesis,
explaining the apparent metastatic molecular signature by most cells in a primary tumor (230).
Jean Paul Thiery and Robert Weinberg proposed that EMT could explain metastatic progression (171, 230).
Christopher Klein and collaborators reported the genetic heterogeneity of single disseminated tumors cells in
minimal residual cancer (231).
Laura Van 't Veer and coworkers reported that a specific gene expression profile of primary breast cancers
was associated with the development of metastasis and a poor clinical outcome (184).
Yibin Kang and colleagues identified a molecular signature associated with metastasis of breast tumors to bone (75).
Michael Clarke and Max Wicha suggest that the ability of breast cancer tumors to metastasize resides in
just a few “breast CSCs” that are highly resistant to chemotherapy (232).
Kent Hunter emphasized the role of genetic susceptibility for the metastatic process (233).
Li Ma and Robert Weinberg identified the first metastasis-promoting micro-RNA (234).
William Harbor and coworkers showed that micro-RNA expression patterns could predict metastatic risk (235).
Joan Massagué and coworkers reported that micro-RNAs can suppress metastases (236).
1990
1991
1992
1994
1997
2000
2001
2002
2002
2002
2002
2003
2003
2006
2007
2008
2008
metastases and concluded that there were limited but statistically significant differences between the primary tumors
and their lymphatic metastases. This study (8) also suggested
that the genetic program for metastasis developed over time,
although occasionally it occurred early during tumor progression, ultimately resulting in heterogeneity both within
and between metastases.
The identification of microarray profiles with clinical relevance has supported the development of prognostic signatures for classifying tumors into risk groups and groups
targeted for differential treatment approaches or no treatment. In breast cancer this approach has been undertaken
by multiple investigators with disparate results. The research strategy used a retrospective analysis of samples
with follow-up, selection of discriminator genes, and the development of a predictive multigene classifier (187). A statistical analysis from the early studies suggested flaws in
this experimental design (188). In retrospect two data sets
are needed: one to develop the classifier (training set), and
another to test the classifier (test or validation set). These
two sets are obtained by splitting the original data set, if it
www.aacrjournals.org
is large enough. Further, a validation using a third independent set is desirable. With this approach multiple genomicprognostic classifiers for breast cancer have been developed,
although the overlap of genes in these signatures has been
low. Nonetheless, the primary determinants of all the signatures are proliferation, ER-status, HER2-status, and, less
prominently, angiogenesis, invasiveness, and apoptosis.
Thus, it is likely that these signatures detect the same biological processes and pathways involved in metastasis.
Cancer Stem Cells
It has been more than a century since Cohnheim (189) proposed the “embryonic theory” of cancer that postulated that
human tumors arise from embryonic cells that persevere in tissues without reaching maturity. This theory has developed into
the CSC hypothesis, which states that cancers develop from a
subset of malignant cells that possess stem cell characteristics,
such that tumors have rare cells with infinite growth potential
(190). It is noted that this characteristic of infinite growth potential is also the definition of a tumor cell. CSCs are suggested
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5663
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
to express characteristics of both stem cells and cancer cells,
and have properties of self-renewal, asymmetric cell division,
resistance to apoptosis, independent growth, tumorigenicity,
and metastatic potential. The CSC theory suggests that tumors
include a population of asymmetrically dividing CSCs that
give rise to rapidly expanding progenitor cells, which eventually differentiate and exhaust their proliferative potential.
The stem cells retain their original phenotype and proliferative
competence, which is referred to as “self-renewal” capacity.
Cell populations enriched for CSCs are hypothesized to display
increased tumorigenic potential in serial transplantation
assays, as compared with the bulk of tumor cells.
The first definitive evidence of a CSC was with acute myeloid leukemia (AML) by Bonnet and Dick in 1997 (191). This
study used the cellular hierarchy observed for hematopoietic
cells as a model to describe the development of AML from a
primitive progenitor cancer cell, now known as the CSC. The
tumor initiating cells were all found to have a distinct surface
phenotype (CD34+/CD38−), irrespective of AML subtypes and/
or the degree of heterogeneity in the tumor from which they
were isolated (191). A CSC origin has now been shown for a
range of “liquid tumors” (192), including B- and T-acute lymphoblastic leukemia (B-ALL and T-ALL, respectively) and arguably with a variety of solid tumors (193).
Given the similarities between normal and CSCs it has been
speculated that CSCs originate from normal self-renewing
stem cells that have accumulated oncogenic mutations.
However, it is suggested that mutations in progenitor or differentiated cells induce dedifferentiation and self-renewal capacity (194, 195). The self-renewal capacity and multipotency
of CSCs is suggested to provide a mechanism for tumor recurrence, frequently observed after surgical excision of the
primary tumor, but presumably not the CSCs. As discussed
above, tumors are heterogeneous such that not all tumor
cells possess the same phenotype including metastatic potential. The CSC theory implies that if CSCs are the only subset of cells capable of initiating new tumor growth, then
CSCs must be involved in the metastatic process. Therefore,
it is posited that as a CSC possesses tumorigenic, invasive,
and migratory characteristics it will also have all of the necessary attributes to result in a metastasis. However, many reports on CSCs are contradictory, varying in how they should
be identified, their characteristics, and correlations between
clinical outcome and tumor CSC status (196). These contradictory observations have made the CSC hypothesis highly
contentious, including the finding that CSCs are tumor cells
with metastatic properties and that the cells within the metastatic lesion may then lose the metastatic phenotype; the
hypotheses are not well supported clinically. Further, it is
not clear whether CSCs are “true” stem cells or represent a
highly malignant cellular subpopulation. The recent publication by Quintana and colleagues showed that by optimizing
xenotransplantation procedures, one can increase the frequency of tumor-initiating human melanoma cells (197). This
finding suggests that, at least in some human cancers,
tumor-initiating cells are frequent, and the question is raised
of whether the CSC concept applies to solid tumors. Thus,
the link between CSCs and metastasis is circumstantial,
5664
Cancer Res; 70(14) July 15, 2010
based on correlations between the frequency of cells required
to form a metastasis, cellular phenotypes, and the expression
of genes associated with “stem cells” or EMT (198). Indeed,
the concept that the rare metastasis-forming cells found in
tumors correspond to the equally rare CSCs seems to represent a new nomenclature for a concept discussed in the
1970s by Fidler and coworkers. Both selected cells and CSCs
have the potential to form a secondary tumor, with a similar
histological architecture as tumor cells in the primary tumor
(199). Further, both stem cells and metastatic cells are critically reliant on the microenvironment (“metastatic niche”)
that they reside in. As Kaplan and colleagues have discussed
such a niche is required for the formation of metastases
(157), and the need to change an organ microenvironment
into an appropriate niche may contribute to the latency
(and potentially failure) of a disseminated tumor cell to grow
into a gross metastasis (190).
Summary and Future Directions
Once a diagnosis of a primary cancer is established, the
urgent question is whether it is localized or metastasized.
Despite improvements in diagnosis, surgical techniques, general patient care, and local and systemic adjuvant therapies,
most deaths from cancer result from metastases that are resistant to conventional therapies. The organ microenvironment, in addition to facilitating metastasis, can also modify
the response of metastatic tumor cells to therapy and reduce
the effectiveness of anticancer agents. In addition, metastatic
cells are genetically unstable with diverse karyotypes, growth
rates, cell-surface properties, antigenicities, immunogenicities, marker enzymes, and sensitivity to various therapeutic
agents resulting in biological heterogeneity. The process of
cancer metastasis is sequential and selective and incorporates stochastic elements. Thus, the growth of metastases represents the endpoint of many events that few tumor cells
survive. Primary tumors consist of multiple subpopulations
of cells with invasive and metastatic properties, with the potential to form a metastasis in a process dependent on the
interplay of tumor cells with host factors. The finding that
different metastases can originate from multiple progenitor
cells contributes to the biological diversity and provides additional challenges to therapeutic intervention. Further, even
within a solitary metastasis of clonal origin, heterogeneity
can rapidly develop. Thus, it is critical to improve our understanding of metastatic cell characteristics that will allow us
to target them for therapeutic intervention. Clearly, the pathogenesis of metastasis depends on multiple interactions between metastatic cells and host homeostatic mechanisms
(Table 2). We posit that the interruption of these interactions
will inhibit or help eradicate metastasis. Clinical efforts have
focused on the inhibition or destruction of tumor cells; however, strategies to treat metastatic tumor cells and modulate
the host microenvironment now offer new treatment approaches. The recent advances in our understanding of the
metastatic process at the cellular and molecular level provide
unprecedented potential for improvement and the development of effective adjuvant therapies.
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
Disclosure of Potential Conflicts of Interest
Grant Support
Acknowledgments
Nebraska Research Initiative Grant “Translation of Biotechnology into the
Clinic” (J. E. Talmadge), Avon-National Cancer Institute Patient Award
CA036727 (J. E. Talmadge), Cancer Center Support Core Grant CA16672, and
Grant 1U54CA143837 (I. J. Fidler) from the National Cancer Institute, National
Institutes of Health.
The authors thank Jill Hallgren, Alice Cole, and Lola López for their expert
assistance in the preparation of this manuscript.
Received 09/29/2009; revised 04/12/2010; accepted 04/25/2010; published
OnlineFirst 07/06/2010.
No potential conflicts of interest were disclosed.
References
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
Recamier JC. Recherches sur le traitement du cancer sur la compression methodique simple ou combinee et sur l'histoire generale
de la meme maladie. 2nd ed. 1829.
Fidler IJ. The pathogenesis of cancer metastasis: the ‘seed and soil’
hypothesis revisited. Nat Rev Cancer 2003;3:453–8.
Dorland WA. Dorland's illustrated medical dictionary. 24th ed.
Philadelphia (PA): Saunders Co.; 1965.
Del Monte U. Does the cell number 10(9) still really fit one gram of
tumor tissue? Cell Cycle 2009;8:505–6.
van de Vijver MJ, He YD, van't Veer LJ, et al. A gene-expression
signature as a predictor of survival in breast cancer. N Engl
J Med 2002;347:1999–2009.
Weigelt B, Glas AM, Wessels LF, et al. Gene expression profiles of
primary breast tumors maintained in distant metastases. Proc Natl
Acad Sci U S A 2003;100:15901–5.
Ramaswamy S, Ross KN, Lander ES, Golub TR. A molecular signature of metastasis in primary solid tumors. Nat Genet 2003;33:
49–54.
Suzuki M, Tarin D. Gene expression profiling of human lymph node
metastases and matched primary breast carcinomas: clinical implications. Mol Oncol 2007;1:172–80.
Urquidi V, Sloan D, Kawai K, et al. Contrasting expression of thrombospondin-1 and osteopontin correlates with absence or presence
of metastatic phenotype in an isogenic model of spontaneous human breast cancer metastasis. Clin Cancer Res 2002;8:61–74.
Fidler IJ, Kripke ML. Metastasis results from preexisting variant cells
within a malignant tumor. Science 1977;197:893–5.
Kripke ML, Gruys E, Fidler IJ. Metastatic heterogeneity of cells from
an ultraviolet light-induced murine fibrosarcoma of recent origin.
Cancer Res 1978;38:2962–7.
Nicolson GL, Brunson KW, Fidler IJ. Specificity of arrest, survival,
and growth of selected metastatic variant cell lines. Cancer Res
1978;38:4105–11.
Poste G, Doll J, Fidler IJ. Interactions among clonal subpopulations
affect stability of the metastatic phenotype in polyclonal populations of B16 melanoma cells. Proc Natl Acad Sci U S A 1981;78:
6226–30.
Kozlowski JM, Hart IR, Fidler IJ, Hanna N. A human melanoma line
heterogeneous with respect to metastatic capacity in athymic nude
mice. J Natl Cancer Inst 1984;72:913–7.
Montel V, Huang TY, Mose E, Pestonjamasp K, Tarin D. Expression
profiling of primary tumors and matched lymphatic and lung metastases in a xenogeneic breast cancer model. Am J Pathol 2005;166:
1565–79.
Poste G, Fidler IJ. The pathogenesis of cancer metastasis. Nature
1980;283:139–46.
Paget S. The distribution of secondary growths in cancer of the
breast. Lancet 1889;133:571–3.
Virchow R. Cellular pathologie, 1858. Nutr Rev 1989;47:23–5.
Ewing J. Neoplastic diseases: a treatise on tumors. 3rd ed.
Philadelphia (PA): WB Saunders; 1928.
Hart IR, Fidler IJ. Role of organ selectivity in the determination of metastatic patterns of B16 melanoma. Cancer Res 1980;40:2281–7.
Fidler IJ, Talmadge JE. Evidence that intravenously derived murine
pulmonary melanoma metastases can originate from the expansion
of a single tumor cell. Cancer Res 1986;46:5167–71.
Fidler IJ. Metastasis: guantitative analysis of distribution and fate of
www.aacrjournals.org
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
34.
35.
36.
37.
38.
39.
40.
41.
42.
43.
44.
tumor embolilabeled with 125I-5-iodo-2'-deoxyuridine. J Natl Cancer Inst 1970;45:773–82.
Talmadge JE, Fidler IJ. Cancer metastasis is selective or random
depending on the parent tumor population. Nature 1978;27:593–4.
Raz A, Hanna N, Fidler IJ. In vivo isolation of a metastatic tumor cell
variant involving selective and nonadaptive processes. J Natl Cancer Inst 1981;66:183–9.
Talmadge JE, Fidler IJ. Enhanced metastatic potential of tumor
cells harvested from spontaneous metastases of heterogeneous
murine tumors. J Natl Cancer Inst 1982;69:975–80.
Fidler IJ. Tumor heterogeneity and the biology of cancer invasion
and metastasis. Cancer Res 1978;38:2651–60.
Langley RR, Fidler IJ. Tumor cell-organ microenvironment interactions in the pathogenesis of cancer metastasis. Endocr Rev 2007;
28:297–321.
Price JE, Aukerman SL, Fidler IJ. Evidence that the process of murine melanoma metastasis is sequential and selective and contains
stochastic elements. Cancer Res 1986;46:5172–8.
Willis RA. The spread of tumors in the human body. London: Butterworths and Company; 1952. pp. 1–447.
Fidler IJ, Gersten DM, Hart IR. The biology of cancer invasion and
metastasis. Adv Cancer Res 1978;28:149–250.
Nicolson GL. Cancer metastasis: tumor cell and host organ properties important in metastasis to specific secondary sites. Biochim
Biophys Acta 1988;948:175–224.
Naito S, Giavazzi R, Fidler IJ. Correlation between the in vitro interaction of tumor cells with an organ environment and metastatic behavior in vivo. Invasion Metastasis 1987;7:16–29.
Price JE, Naito S, Fidler IJ. Growth in an organ microenvironment
as a selective process in metastasis. Clin Exp Metastasis 1988;6:
91–102.
Sugarbaker EV. Cancer metastasis: a product of tumor-host interactions. Curr Probl Cancer 1979;3:1–59.
Weiss L. Principles of metastasis. Orlando (FL): Academic Press;
1985.
Fidler IJ, Kripke ML. Genomic analysis of primary tumors does not
address the prevalence of metastatic cells in the population. Nat
Genet 2003;34:23.
Chambers AF, Groom AC, MacDonald IC. Dissemination and
growth of cancer cells in metastatic sites. Nat Rev Cancer 2002;
2:563–72.
Weiss L. Metastatic inefficiency: causes and consequences. Cancer Rev 1986;3:1–24.
Chen LL, Blumm N, Christakis NA, Barabasi AL, Deisboeck TS.
Cancer metastasis networks and the prediction of progression patterns. Br J Cancer 2009;101:749–58.
Fidler IJ. Biological behavior of malignant melanoma cells correlated to their survival in vivo. Cancer Res 1975;35:218–24.
Chiang AC, Massague J. Molecular basis of metastasis. N Engl
J Med 2008;359:2814–23.
Waghorne C, Thomas M, Lagarde A, Kerbel RS, Breitman ML. Genetic evidence for progressive selection and overgrowth of primary
tumors by metastatic cell subpopulations. Cancer Res 1988;48:
6109–14.
Wang K, Li J, Li S, Bolund L, Wiuf C. Estimation of tumor heterogeneity using CGH array data. BMC Bioinformatics 2009;10:12.
Kuukasjarvi T, Karhu R, Tanner M, et al. Genetic heterogeneity and
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5665
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
45.
46.
47.
48.
49.
50.
51.
52.
53.
54.
55.
56.
57.
58.
59.
60.
61.
62.
63.
64.
65.
66.
67.
68.
69.
5666
clonal evolution underlying development of asynchronous metastasis in human breast cancer. Cancer Res 1997;57:1597–604.
Bissig H, Richter J, Desper R, et al. Evaluation of the clonal relationship between primary and metastatic renal cell carcinoma
by comparative genomic hybridization. Am J Pathol 1999;155:
267–74.
Butler TP, Gullino PM. Quantitation of cell shedding into efferent
blood of mammary adenocarcinoma. Cancer Res 1975;35:512–6.
Tarin D, Price JE, Kettlewell MG, et al. Mechanisms of human tumor
metastasis studied in patients with peritoneovenous shunts. Cancer
Res 1984;44:3584–92.
Kasimir-Bauer S. Circulating tumor cells as markers for cancer risk
assessment and treatment monitoring. Mol Diagn Ther 2009;13:
209–15.
Arnerlöv C, Emdin SO, Lundgren B, et al. Breast carcinoma growth
rate described by mammographic doubling time and S-phase fraction. Correlations to clinical and histopathologic factors in a
screened population. Cancer 1992;70:1928–34.
Shackney SE, McCormack GW, Cuchural GJ, Jr. Growth rate patterns of solid tumors and their relation to responsiveness to therapy: an analytical review. Ann Intern Med 1978;89:107–21.
von Fournier D, Weber E, Hoeffken W, et al. Growth rate of 147
mammary carcinomas. Cancer 1980;45:2198–207.
Peer PG, van Dijck JA, Hendriks JH, Holland R, Verbeek AL.
Age-dependent growth rate of primary breast cancer. Cancer
1993;71:3547–51.
Fidler IJ, Balch CM. The biology of cancer metastasis and implications for therapy. Curr Probl Surg 1987;24:129–209.
Engel J, Eckel R, Kerr J, et al. The process of metastasisation for
breast cancer. Eur J Cancer 2003;39:1794–806.
Jones S, Chen WD, Parmigiani G, et al. Comparative lesion sequencing provides insights into tumor evolution. Proc Natl Acad
Sci U S A 2008;105:4283–8.
Robinson SN, Chavez JM, Blonder JM, et al. Hematopoietic progenitor cell mobilization in mice by sustained delivery of granulocyte colony-stimulating factor. J Interferon Cytokine Res 2005;25:
490–500.
Muto T, Bussey HJ, Morson BC. The evolution of cancer of the colon and rectum. Cancer 1975;36:2251–70.
Staab HJ, Anderer FA, Hornung A, Stumpf E, Fischer R. Doubling
time of circulating cea and its relation to survival of patients with
recurrent colorectal cancer. Br J Cancer 1982;46:773–81.
Price JE, Tarin D, Fidler IJ. Influence of organ microenvironment on
pigmentation of a metastatic murine melanoma. Cancer Res 1988;
48:2258–64.
Schackert G, Fidler IJ. Site-specific metastasis of mouse melanomas and a fibrosarcoma in the brain or meninges of syngeneic animals. Cancer Res 1988;48:3478–84.
Nicolson GL. Organ specificity of tumor metastasis: role of preferential adhesion, invasion and growth of malignant cells at specific
secondary sites. Cancer Metastasis Rev 1988;7:143–88.
Fidler IJ. The organ microenvironment and cancer metastasis.
Differentiation 2002;70:498–505.
Simone NL, Bonner RF, Gillespie JW, Emmert-Buck MR, Liotta LA.
Laser-capture microdissection: opening the microscopic frontier to
molecular analysis. Trends Genet 1998;14:272–6.
Fidler IJ. Critical factors in the biology of human cancer metastasis:
twenty-eighth G.H.A. Clowes memorial award lecture. Cancer Res
1990;50:6130–8.
Pasqualini R, Ruoslahti E. Organ targeting in vivo using phage display peptide libraries. Nature 1996;380:364–6.
Uehara H, Kim SJ, Karashima T, et al. Effects of blocking plateletderived growth factor-receptor signaling in a mouse model of experimental prostate cancer bone metastases. J Natl Cancer Inst
2003;95:458–70.
Fidler IJ. Critical determinants of metastasis. Semin Cancer Biol
2002;12:89–96.
Paget S. The distribution of secondary growths in cancer of the
breast. 1889. Cancer Metastasis Rev 1989;8:98–101.
Fuchs E. In: Braumueller W, editor. Metastasenbildung. Wien:
Sarcom des Uvealtractus; 1882, p. 197–206.
Cancer Res; 70(14) July 15, 2010
70.
71.
72.
73.
74.
75.
76.
77.
78.
79.
80.
81.
82.
83.
84.
85.
86.
87.
88.
89.
90.
91.
92.
93.
Ruoslahti E. Vascular zip codes in angiogenesis and metastasis.
Biochem Soc Trans 2004;32:397–402.
Muller A, Homey B, Soto H, et al. Involvement of chemokine receptors in breast cancer metastasis. Nature 2001;410:50–6.
Huang EH, Singh B, Cristofanilli M, et al. A CXCR4 antagonist
CTCE-9908 inhibits primary tumor growth and metastasis of breast
cancer. J Surg Res 2009;155:231–6.
Zeelenberg IS, Ruuls-Van SL, Roos E. The chemokine receptor
CXCR4 is required for outgrowth of colon carcinoma micrometastases. Cancer Res 2003;63:3833–9.
Andrea S, Emanuela M, Chiara C, et al. CXCL12, CXCR4 and
CXCR7 expression in brain metastases. Cancer Biol Ther 2009;8:
1608–14.
Kang Y, Siegel PM, Shu W, et al. A multigenic program mediating breast cancer metastasis to bone. Cancer Cell 2003;3:
537–49.
Minn AJ, Gupta GP, Siegel PM, et al. Genes that mediate breast
cancer metastasis to lung. Nature 2005;436:518–24.
Talmadge JE, Singh RK, Fidler IJ, Raz A. Murine models to evaluate
novel and conventional therapeutic strategies for cancer. Am
J Pathol 2007;170:793–804.
Sone S, Fidler IJ. Activation of rat alveolar macrophages to the tumoricidal state in the presence of progressively growing pulmonary
metastases. Cancer Res 1981;41:2401–6.
Eccles SA, Alexander P. Macrophage content of tumours in relation
to metastatic spread and host immune reaction. Nature 1974;250:
667–9.
Gauci CL, Alexander P. The macrophage content of some human
tumors. Cancer Lett 1975;1:29–32.
Evans R, Lawler EM. Macrophage content and immunogenicity of
C57BL/6J and BALB/cByJ methylcholanthrene-induced sarcomas.
Int J Cancer 1980;26:831–5.
Key M, Talmadge JE, Fidler IJ. Lack of correlation between the
progressive growth of spontaneous metastases and their content of infiltrating macrophages. J Reticuloendothel Soc 1982;
32:387–96.
Kripke ML, Fisher MS. Immunologic parameters of ultraviolet carcinogenesis. J Natl Cancer Inst 1976;57:211–5.
Pross HF, Kerbel RS. An assessment of intratumor phagocytic and
surface marker-bearing cells in a series of autochthonous and early
passaged chemically induced murine sarcomas. J Natl Cancer Inst
1976;57:1157–67.
Watanabe T, Dave B, Heimann DG, et al. GM-CSF-mobilized peripheral blood CD34+ cells differ from steady- state bone marrow
CD34+ cells in adhesion molecule expression. Bone Marrow Transplant 1997;19:1175–81.
Ross J, Auger M. The biology of the macrophage. In: Burke B,
Lewis C, editors. The macrophage. 2nd ed. Oxford: University
Press; 2002.
Mantovani A. Effects on in vitro tumor growth of murine macrophages isolated from sarcoma lines differing in immunogenicity
and metastasizing capacity. Int J Cancer 1978;22:741–6.
Steele RJ, Eremin O, Brown M, Hawkins RA. A high macrophage
content in human breast cancer is not associated with favourable
prognostic factors. Br J Surg 1984;71:456–8.
Sun XF, Zhang H. Clinicopathological significance of stromal variables: angiogenesis, lymphangiogenesis, inflammatory infiltration,
MMP and PINCH in colorectal carcinomas. Mol Cancer 2006;5:
43–63.
Fidler IJ, Schackert G, Zhang RD, Radinsky R, Fujimaki T. The biology of melanoma brain metastasis. Cancer Metastasis Rev 1999;
18:387–400.
Alexander P, Eccles SA, Gauci CL. The significance of macrophages in human and experimental tumors. Ann N Y Acad Sci
1976;276:124–33.
Donkor MK, Lahue E, Hoke TA, et al. Mammary tumor heterogeneity in the expansion of myeloid-derived suppressor cells. Int Immunopharmacol 2009;9:937–48.
Lin EY, Nguyen AV, Russell RG, Pollard JW. Colony-stimulating factor 1 promotes progression of mammary tumors to malignancy.
J Exp Med 2001;193:727–40.
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
94.
95.
96.
97.
98.
99.
100.
101.
102.
103.
104.
105.
106.
107.
108.
109.
110.
111.
112.
113.
114.
115.
116.
117.
Hagemann T, Robinson SC, Schulz M, et al. Enhanced invasiveness
of breast cancer cell lines upon co-cultivation with macrophages is
due to TNF-α dependent up-regulation of matrix metalloproteases.
Carcinogenesis 2004;25:1543–9.
Hamada I, Kato M, Yamasaki T, et al. Clinical effects of tumorassociated macrophages and dendritic cells on renal cell carcinoma. Anticancer Res 2002;22:4281–4.
Lewis C, Murdoch C. Macrophage responses to hypoxia: implications for tumor progression and anti-cancer therapies. Am J Pathol
2005;167:627–35.
Bronte V, Wang M, Overwijk WW, et al. Apoptotic death of CD8+
T lymphocytes after immunization: induction of a suppressive population of Mac-1+/Gr-1+ cells. J Immunol 1998;161:5313–20.
Kusmartsev S, Gabrilovich DI. STAT1 signaling regulates tumorassociated macrophage-mediated T cell deletion. J Immunol
2005;174:4880–91.
Mirza N, Fishman M, Fricke I, et al. All-trans-retinoic acid improves
differentiation of myeloid cells and immune response in cancer patients. Cancer Res 2006;66:9299–307.
Almand B, Clark JI, Nikitina E, et al. Increased production of immature myeloid cells in cancer patients: a mechanism of immunosuppression in cancer. J Immunol 2001;166:678–89.
Almand B, Resser JR, Lindman B, et al. Clinical significance of defective dendritic cell differentiation in cancer. Clin Cancer Res 2000;
6:1755–66.
Strober S. Natural suppressor (NS) cells, neonatal tolerance, and
total lymphoid irradiation: exploring obscure relationships. Annu
Rev Immunol 1984;2:219–37.
Badger AM, King AG, Talmadge JE, et al. Induction of non-specific
suppressor cells in normal Lewis rats by a novel azaspirane SK&F
105685. J Autoimmun 1990;3:485–500.
Bronte V, Apolloni E, Cabrelle A, et al. Identification of a CD11b(+)/
Gr-1(+)/CD31(+) myeloid progenitor capable of activating or suppressing CD8(+) T cells. Blood 2000;96:3838–46.
Young MR, Lathers DM. Myeloid progenitor cells mediate immune
suppression in patients with head and neck cancers. Int J Immunopharmacol 1999;21:241–52.
Dranoff G. GM-CSF-secreting melanoma vaccines. Oncogene
2003;22:3188–92.
Ohm JE, Carbone DP. VEGF as a mediator of tumor-associated immunodeficiency. Immunol Res 2001;23:263–72.
Gabrilovich D, Ishida T, Oyama T, et al. Vascular endothelial growth
factor inhibits the development of dendritic cells and dramatically
affects the differentiation of multiple hematopoietic lineages in vivo.
Blood 1998;92:4150–66.
Ellis LM, Takahashi Y, Liu W, Shaheen RM. Vascular endothelial
growth factor in human colon cancer: biology and therapeutic implications. Oncologist 2000;5[Suppl 1]:11–5.
Toi M, Kondo S, Suzuki H, et al. Quantitative analysis of vascular
endothelial growth factor in primary breast cancer. Cancer 1996;77:
1101–6.
Gabrilovich DI, Chen HL, Girgis KR, et al. Production of vascular
endothelial growth factor by human tumors inhibits the functional
maturation of dendritic cells. Nat Med 1996;2:1096–103.
Saito H, Tsujitani S, Ikeguchi M, Maeta M, Kaibara N. Relationship
between the expression of vascular endothelial growth factor
and the density of dendritic cells in gastric adenocarcinoma tissue.
Br J Cancer 1998;78:1573–7.
Gabrilovich DI, Ishida T, Nadaf S, Ohm JE, Carbone DP. Antibodies
to vascular endothelial growth factor enhance the efficacy of cancer
immunotherapy by improving endogenous dendritic cell function.
Clin Cancer Res 1999;5:2963–70.
Finke JH, Rini B, Ireland J, et al. Sunitinib reverses type-1 immune
suppression and decreases T-regulatory cells in renal cell carcinoma patients. Clin Cancer Res 2008;14:6674–82.
Ko JS, Zea AH, Rini BI, et al. Sunitinib mediates reversal of myeloidderived suppressor cell accumulation in renal cell carcinoma patients. Clin Cancer Res 2009;15:2148–57.
Fidler IJ. Selection of successive tumour lines for metastasis. Nat
New Biol 1973;242:148–9.
Nicolson GL, Dulski KM. Organ specificity of metastatic tumor
www.aacrjournals.org
118.
119.
120.
121.
122.
123.
124.
125.
126.
127.
128.
129.
130.
131.
132.
133.
134.
135.
136.
137.
138.
139.
140.
141.
142.
colonization is related to organ-selective growth properties of malignant cells. Int J Cancer 1986;38:289–94.
Fidler IJ. General considerations for studies of experimental cancer
metastasis. In: Busch H, editor. Methods of cancer research, Vol.
XV. New York: New Academic Press; 1978, p 399–439.
Brunson KW, Nicolson GL. Selection and biologic properties of malignant variants of a murine lymphosarcoma. J Natl Cancer Inst
1978;61:1499–503.
Fidler IJ, Gersten DM, Budmen MB. Characterization in vivo and
in vitro of tumor cells selected for resistance to syngeneic lymphocyte-mediated cytotoxicity. Cancer Res 1976;36:3160–5.
Briles EB, Kornfeld S. Isolation and metastatic properties of detachment variants of B16 melanoma cells. J Natl Cancer Inst 1978;60:
1217–22.
Hart IR. The selection and characterization of an invasive variant of
the B16 melanoma. Am J Pathol 1979;97:587–600.
Poste G, Doll J, Hart IR, Fidler IJ. In vitro selection of murine B16
melanoma variants with enhanced tissue-invasive properties. Cancer Res 1980;40:1636–44.
Kerbel RS. Immunologic studies of membrane mutants of a highly
metastatic murine tumor. Am J Pathol 1979;97:609–22.
Hanna N. Role of natural killer cells in control of cancer metastasis.
Cancer Metastasis Rev 1982;1:45–64.
Luria SE, Delbruck M. Mutations of bacteria from virus sensitivity to
virus resistance. Genetics 1943;28:491–511.
Kripke ML. Speculations on the role of ultraviolet radiation in the
development of malignant melanoma. J Natl Cancer Inst 1979;63:
541–8.
Fidler IJ, Kripke ML. Metastatic heterogeneity of cells from the K1735 melanoma. In: Grundmann E, editor. Metastatic tumor growth.
Stuttgart: Gustav Fischer Verlag; 1980, p. 71–81.
Fidler IJ, Gruys E, Cifone MA, Barnes Z, Bucana C. Demonstration
of multiple phenotypic diversity in a murine melanoma of recent origin. J Natl Cancer Inst 1981;67:947–56.
Fidler IJ, Gersten DM, Kripke ML. Influence of immune status on the
metastasis of three murine fibrosarcomas of different immunogenicities. Cancer Res 1979;39:3816–21.
Fidler IJ, Kripke ML. Tumor cell antigenicity, host immunity and cancer metastasis. Cancer Immunol Immunother 1980;7:201–5.
Aukerman SL, Price JE, Fidler IJ. Different deficiencies in the prevention of tumorigenic-low-metastatic murine K-1735b melanoma
cells from producing metastases. J Natl Cancer Inst 1986;77:
915–24.
Heppner GH. Tumor heterogeneity. Cancer Res 1984;44:2259–65.
Nicolson GL, Custead SE. Tumor metastasis is not due to adaptation of cells to a new organ environment. Science 1982;215:
176–8.
Poste G. Pathogenesis of metastatic disease: implications for current therapy and for the development of new therapeutic strategies.
Cancer Treat Rep 1986;70:183–99.
Nakamura T, Kuwai T, Kitadai Y, et al. Zonal heterogeneity for gene
expression in human pancreatic carcinoma. Cancer Res 2007;67:
7597–604.
Bachtiary B, Boutros PC, Pintilie M, et al. Gene expression profiling
in cervical cancer: an exploration of intratumor heterogeneity. Clin
Cancer Res 2006;12:5632–40.
Moroni M, Veronese S, Benvenuti S, et al. Gene copy number for
epidermal growth factor receptor (EGFR) and clinical response to
antiEGFR treatment in colorectal cancer: a cohort study. Lancet
Oncol 2005;6:279–86.
Pettaway CA, Pathak S, Greene G, et al. Selection of highly metastatic variants of different human prostatic carcinomas using
orthotopic implantation in nude mice. Clin Cancer Res 1996;2:
1627–36.
Nowell PC. The clonal evolution of tumor cell populations. Science
1976;194:23–8.
Nowell PC. Chromosomal and molecular clues to tumor progression. Semin Oncol 1989;16:116–27.
Cifone MA, Fidler IJ. Increasing metastatic potential is associated
with increasing genetic instability of clones isolated from murine
neoplasms. Proc Natl Acad Sci U S A 1981;78:6949–52.
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5667
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Talmadge and Fidler
143. Hill RP, Chambers AF, Ling V, Harris JF. Dynamic heterogeneity:
rapid generation of metastatic variants in mouse B16 melanoma
cells. Science 1984;224:998–1001.
144. Masramon L, Vendrell E, Tarafa G, et al. Genetic instability and divergence of clonal populations in colon cancer cells in vitro. J Cell
Sci 2006;119:1477–82.
145. Jones TD, Carr MD, Eble JN, et al. Clonal origin of lymph node metastases in bladder carcinoma. Cancer 2005;104:1901–10.
146. Welch DR, Tomasovic SP. Implications of tumor progression on
clinical oncology. Clin Exp Metastasis 1985;3:151–88.
147. Poste G, Greig R. On the genesis and regulation of cellular heterogeneity in malignant tumors. Invasion Metastasis 1982;2:137–76.
148. Talmadge JE, Wolman SR, Fidler IJ. Evidence for the clonal origin of
spontaneous metastases. Science 1982;217:361–3.
149. Talmadge JE, Zbar B. Clonality of pulmonary metastases from the
bladder 6 subline of the B16 melanoma studied by southern hybridization. J Natl Cancer Inst 1987;78:315–20.
150. Poste G, Tzeng J, Doll J, et al. Evolution of tumor cell heterogeneity
during progressive growth of individual lung metastases. Proc Natl
Acad Sci U S A 1982;79:6574–8.
151. Ootsuyama A, Tanaka K, Tanooka H. Evidence by cellular mosaicism for monoclonal metastasis of spontaneous mouse mammary
tumors. J Natl Cancer Inst 1987;78:1223–7.
152. Talmadge C, Tanio Y, Meeker A, Talmadge J, Zbar B. Tumor cells
transfected with the neomycin resistance gene (neo) contain unique
genetic markers useful for identification of tumor recurrence and
metastasis. Invasion Metastasis 1987;7:197–207.
153. Korczak B, Robson IB, Lamarche C, Bernstein A, Kerbel RS. Genetic tagging of tumor cells with retrovirus vectors: clonal analysis of
tumor growth and metastasis in vivo. Mol Cell Biol 1988;8:3143–9.
154. Talmadge JE, Benedict K, Madsen J, Fidler IJ. Development of biological diversity and susceptibility to chemotherapy in murine cancer metastases. Cancer Res 1984;44:3801–5.
155. Wolman SR, McMorrow LE, Fidler IJ, Talmadge JE. Development
and progression of karyotypic variability in melanoma K1735 following X-irradiation. Cancer Res 1985;45:1839–44.
156. Kaplan RN, Rafii S, Lyden D. Preparing the “soil”:the premetastatic
niche. Cancer Res 2006;66:11089–93.
157. Kaplan RN, Riba RD, Zacharoulis S, et al. VEGFR1-positive haematopoietic bone marrow progenitors initiate the pre-metastatic niche.
Nature 2005;438:820–7.
158. Gao D, Nolan DJ, Mellick AS, et al. Endothelial progenitor cells control the angiogenic switch in mouse lung metastasis. Science 2008;
319:195–8.
159. Talmadge JE, Key M, Fidler IJ. Macrophage content of metastatic
and nonmetastatic rodent neoplasms. J Immunol 1981;126:2245–8.
160. Petit I, Jin D, Rafii S. The SDF-1-CXCR4 signaling pathway: a molecular hub modulating neo-angiogenesis. Trends Immunol 2007;
28:299–307.
161. DeNardo DG, Johansson M, Coussens LM. Immune cells as mediators of solid tumor metastasis. Cancer Metastasis Rev 2008;27:11–8.
162. Brigati C, Noonan DM, Albini A, Benelli R. Tumors and inflammatory
infiltrates: friends or foes? Clin Exp Metastasis 2002;19:247–58.
163. Chen Q, Wang WC, Evans SS. Tumor microvasculature as a barrier to
antitumor immunity. Cancer Immunol Immunother 2003;52:670–9.
164. Dvorak HF. Tumors: wounds that do not heal. Similarities between
tumor stroma generation and wound healing. N Engl J Med 1986;
315:1650–9.
165. Sica A, Schioppa T, Mantovani A, Allavena P. Tumour-associated
macrophages are a distinct M2 polarised population promoting tumour progression: potential targets of anti-cancer therapy. Eur
J Cancer 2006;42:717–27.
166. Malmberg KJ. Effective immunotherapy against cancer: a question
of overcoming immune suppression and immune escape? Cancer
Immunol Immunother 2004;53:879–92.
167. Batlle E, Sancho E, Franci C, et al. The transcription factor snail is a
repressor of E-cadherin gene expression in epithelial tumour cells.
Nat Cell Biol 2000;2:84–9.
168. Yang J, Mani SA, Donaher JL, et al. Twist, a master regulator of
morphogenesis, plays an essential role in tumor metastasis. Cell
2004;117:927–39.
5668
Cancer Res; 70(14) July 15, 2010
169. Hartwell KA, Muir B, Reinhardt F, et al. The Spemann organizer
gene, Goosecoid, promotes tumor metastasis. Proc Natl Acad
Sci U S A 2006;103:18969–74.
170. Spaderna S, Schmalhofer O, Wahlbuhl M, et al. The transcriptional
repressor ZEB1 promotes metastasis and loss of cell polarity in
cancer. Cancer Res 2008;68:537–44.
171. Thiery JP. Epithelial-mesenchymal transitions in tumour progression. Nat Rev Cancer 2002;2:442–54.
172. Duband JL, Monier F, Delannet M, Newgreen D. Epitheliummesenchyme transition during neural crest development. Acta Anat
(Basel) 1995;154:63–78.
173. Stoker M, Gherardi E, Perryman M, Gray J. Scatter factor is a fibroblast-derived modulator of epithelial cell mobility. Nature 1987;327:
239–42.
174. Hay ED. An overview of epithelio-mesenchymal transformation.
Acta Anat (Basel) 1995;154:8–20.
175. Greenburg G, Hay ED. Cytodifferentiation and tissue phenotype
change during transformation of embryonic lens epithelium to
mesenchyme-like cells in vitro. Dev Biol 1986;115:363–79.
176. Thiery JP, Sleeman JP. Complex networks orchestrate epithelialmesenchymal transitions. Nat Rev Mol Cell Biol 2006;7:131–42.
177. Thompson EW, Newgreen DF, Tarin D. Carcinoma invasion and
metastasis: a role for epithelial-mesenchymal transition? Cancer
Res 2005;65:5991–5.
178. Delinassios JG, Kottaridis SD, Garas J. Uncontrolled growth of
tumour stromal fibroblasts in vitro. Exp Cell Biol 1983;51:201–9.
179. Wargotz ES, Deos PH, Norris HJ. Metaplastic carcinomas of the
breast. II. Spindle cell carcinoma. Hum Pathol 1989;20:732–40.
180. Dairkee SH, Blayney C, Smith HS, Hackett AJ. Monoclonal antibody that defines human myoepithelium. Proc Natl Acad Sci U S A
1985;82:7409–13.
181. Dairkee SH, Mayall BH, Smith HS, Hackett AJ. Monoclonal
marker that predicts early recurrence of breast cancer. Lancet
1987;1:514.
182. Rosivatz E, Becker I, Specht K, et al. Differential expression of the
epithelial-mesenchymal transition regulators snail, SIP1, and twist
in gastric cancer. Am J Pathol 2002;161:1881–91.
183. Tarin D, Thompson EW, Newgreen DF. The fallacy of epithelial
mesenchymal transition in neoplasia. Cancer Res 2005;65:
5996–6000.
184. van 't Veer LJ, Dai H, van de Vijver MJ, et al. Gene expression profiling predicts clinical outcome of breast cancer. Nature 2002;415:
530–6.
185. Montel V, Mose ES, Tarin D. Tumor-stromal interactions reciprocally modulate gene expression patterns during carcinogenesis and
metastasis. Int J Cancer 2006;119:251–63.
186. Gancberg D, Di LA, Cardoso F, et al. Comparison of HER-2 status
between primary breast cancer and corresponding distant metastatic sites. Ann Oncol 2002;13:1036–43.
187. Pfeffer U, Romeo F, Noonan DM, Albini A. Prediction of breast cancer metastasis by genomic profiling: where do we stand? Clin Exp
Metastasis 2009;26:547–58.
188. Dupuy A, Simon RM. Critical review of published microarray studies
for cancer outcome and guidelines on statistical analysis and reporting. J Natl Cancer Inst 2007;99:147–57.
189. Cohnheim J. Ueber Entzundung und Eiterung. Path Anat Physio
Klin Med 1867;40:1–79.
190. Li F, Tiede B, Massagué J, Kang Y. Beyond tumorigenesis: cancer
stem cells in metastasis. Cell Res 2007;17:3–14.
191. Bonnet D, Dick JE. Human acute myeloid leukemia is organized as
a hierarchy that originates from a primitive hematopoietic cell. Nat
Med 1997;3:730–7.
192. Cox CV, Martin HM, Kearns PR, et al. Characterization of a progenitor cell population in childhood T-cell acute lymphoblastic leukemia. Blood 2007;109:674–82.
193. Hermann PC, Huber SL, Herrler T, et al. Distinct populations of cancer stem cells determine tumor growth and metastatic activity in
human pancreatic cancer. Cell Stem Cell 2007;1:313–23.
194. Pardal R, Clarke MF, Morrison SJ. Applying the principles of stemcell biology to cancer. Nat Rev Cancer 2003;3:895–902.
195. Bachoo RM, Maher EA, Ligon KL, et al. Epidermal growth factor
Cancer Research
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
Cancer Metastasis
196.
197.
198.
199.
200.
201.
202.
203.
204.
205.
206.
207.
208.
209.
210.
211.
212.
213.
214.
215.
216.
receptor and Ink4a/Arf: convergent mechanisms governing terminal
differentiation and transformation along the neural stem cell to astrocyte axis. Cancer Cell 2002;1:269–77.
Kelly PN, Dakic A, Adams JM, Nutt SL, Strasser A. Tumor growth
need not be driven by rare cancer stem cells. Science 2007;317:337.
Quintana E, Shackleton M, Sabel MS, et al. Efficient tumour formation by single human melanoma cells. Nature 2008;456:593–8.
Geiger TR, Peeper DS. Metastasis mechanisms. Biochim Biophys
Acta 2009;1796:293–308.
Brabletz T, Jung A, Spaderna S, Hlubek F, Kirchner T. Opinion: migrating cancer stem cells - an integrated concept of malignant tumour progression. Nat Rev Cancer 2005;5:744–9.
Folkman J. Angiogenesis: initiation and modulation. In: Nicholson
GL, Milas L, editors. Cancer invasion and metastasis: biologic
and therapeutic aspects. New York: Raven Press; 1984, p. 201–9.
Folkman J, Klagsbrun M. Angiogenic factors. Science 1987;235:
442–7.
Liotta LA. Tumor invasion and metastases-role of the extracellular
matrix: Rhoads Memorial Award lecture. Cancer Res 1986;46:1–7.
Takahashi M. An experimental study of metastasis. J Pathol Bacteriol 1915;20:1–13.
Coman DR. Decreased mutual adhesiveness, a property of cells
from squamous cell carcinomas. Cancer Res 1944;4:625–9.
Zeidman I. BUSS JM. Transpulmonary passage of tumor cell emboli. Cancer Res 1952;12:731–3.
Lucke B, Breedis C, Woo ZP, Berwick L, Nowell P. Differential
growth of metastatic tumors in liver and lung: experiments with rabbit V2 carcinoma. Cancer Res 1952;12:734–8.
Klein E. Immediate transformation of solid into ascites tumors;studies on a mammary carcinoma of an inbred mouse strain. Exp Cell
Res 1955;8:213–25.
Gasic G, Gasic T. Removal of sialic acid from the cell coat in tumor
cells and vascular endothelium, and its effects on metastasis. Proc
Natl Acad Sci U S A 1962;48:1172–7.
Fisher B, Fisher ER. The organ distribution of disseminated 51Crlabeled tumor cells. Cancer Res 1967;27:412–20.
Fisher B, Fisher ER. The interrelationship of hematogenous and
lymphatic tumor cell dissemination. Surg Gynecol Obstet 1966;
122:791–8.
Giovanella BC, Yim SO, Morgan AC, Stehlin JS, Williams LJ, Jr.
Brief communication: metastases of human melanomas transplanted in “nude” mice. J Natl Cancer Inst 1973;50:1051–3.
Bross ID, Viadana E, Pickren JW. The metastatic spread of myeloma and leukemias in men. Virchows Arch A Pathol Anat Histol 1975;
365:91–101.
Nicolson GL, Winkelhake JL. Organ specificity of blood-borne tumour metastasis determined by cell adhesion? Nature 1975;255:
230–2.
Liotta LA, Kleinerman J, Catanzaro P, Rynbrandt D. Degradation of
basement membrane by murine tumor cells. J Natl Cancer Inst
1977;58:1427–31.
Liotta LA, Tryggvason K, Garbisa S, et al. Metastatic potential correlates with enzymatic degradation of basement membrane collagen. Nature 1980;284:67–8.
Talmadge JE, Meyers KM, Prieur DJ, Starkey JR. Role of NK cells in
www.aacrjournals.org
217.
218.
219.
220.
221.
222.
223.
224.
225.
226.
227.
228.
229.
230.
231.
232.
233.
234.
235.
236.
tumour growth and metastasis in beige mice. Nature 1980;284:
622–4.
Herman EH, Witiak DT, Hellmann K, Waravdekar VS. Biological
properties of ICRF-159 and related bis(dioxopiperazine) compounds. Adv Pharmacol Chemother 1982;19:249–90.
Volk T, Geiger B, Raz A. Motility and adhesive properties of highand low-metastatic murine neoplastic cells. Cancer Res 1984;44:
811–24.
Fidler IJ. Macrophages and metastasis-a biological approach to
cancer therapy. Cancer Res 1985;45:4714–26.
Weiss L, Mayhew E, Rapp DG, Holmes JC. Metastatic inefficiency
in mice bearing B16 melanomas. Br J Cancer 1982;45:44–53.
Steeg PS, Bevilacqua G, Kopper L, et al. Evidence for a novel gene
associated with low tumor metastatic potential. J Natl Cancer Inst
1988;80:200–4.
Wahl RL, Kaminski MS, Ethier SP, Hutchins GD. The potential of
2-deoxy-2[18F]fluoro-D-glucose (FDG) for the detection of tumor involvement in lymph nodes. J Nucl Med 1990;31:1831–5.
Lin WC, Pretlow TP, Pretlow TG, Culp LA. Bacterial lacZ gene as a
highly sensitive marker to detect micrometastasis formation during
tumor progression. Cancer Res 1990;50:2808–17.
Weidner N, Semple JP, Welch WR, Folkman J. Tumor angiogenesis
and metastasis-correlation in invasive breast carcinoma. N Engl
J Med 1991;324:1–8.
Opdenakker G, Van DJ. Chemotactic factors, passive invasion and
metastasis of cancer cells. Immunol Today 1992;13:463–4.
O'Reilly MS, Holmgren L, Shing Y, et al. Angiostatin: a novel angiogenesis inhibitor that mediates the suppression of metastases by a
Lewis lung carcinoma. Cell 1994;79:315–28.
Chishima T, Miyagi Y, Wang X, et al. Cancer invasion and micrometastasis visualized in live tissue by green fluorescent protein expression. Cancer Res 1997;57:2042–7.
Perou CM, Sorlie T, Eisen MB, et al. Molecular portraits of human
breast tumours. Nature 2000;406:747–52.
Reya T, Morrison SJ, Clarke MF, Weissman IL. Stem cells, cancer,
and cancer stem cells. Nature 2001;414:105–11.
Bernards R, Weinberg RA. A progression puzzle. Nature 2002;
418:823.
Klein CA, Blankenstein TJ, Schmidt-Kittler O, et al. Genetic heterogeneity of single disseminated tumour cells in minimal residual cancer. Lancet 2002;360:683–9.
Dontu G, Al-Hajj M, Abdallah WM, Clarke MF, Wicha MS. Stem cells
in normal breast development and breast cancer. Cell Prolif 2003;
36[Suppl 1]:59–72.
Hunter K. Host genetics influence tumour metastasis. Nat Rev Cancer 2006;6:141–6.
Ma L, Teruya-Feldstein J, Weinberg RA. Tumour invasion and metastasis initiated by microRNA-10b in breast cancer. Nature 2007;
449:682–8.
Worley LA, Long MD, Onken MD, Harbour JW. Micro-RNAs associated with metastasis in uveal melanoma identified by multiplexed
microarray profiling. Melanoma Res 2008;18:184–90.
Tavazoie SF, Alarcon C, Oskarsson T, et al. Endogenous human
microRNAs that suppress breast cancer metastasis. Nature 2008;
451:147–52.
Cancer Res; 70(14) July 15, 2010
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.
5669
Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-1040
AACR Centennial Series: The Biology of Cancer Metastasis:
Historical Perspective
James E. Talmadge and Isaiah J. Fidler
Cancer Res Published OnlineFirst July 7, 2010.
Updated version
E-mail alerts
Reprints and
Subscriptions
Permissions
Access the most recent version of this article at:
doi:10.1158/0008-5472.CAN-10-1040
Sign up to receive free email-alerts related to this article or journal.
To order reprints of this article or to subscribe to the journal, contact the AACR Publications
Department at [email protected].
To request permission to re-use all or part of this article, contact the AACR Publications
Department at [email protected].
Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 2010 American Association for Cancer
Research.