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Endocrine-Related Cancer (2003) 10 23–42
REVIEW
Endometrial cancer: experimental models
useful for studies on molecular aspects of
endometrial cancer and carcinogenesis
G Vollmer
Molecular Cell Physiology and Endocrinology, Institute of Zoology, Dresden University of Technology,
Mommsenstr. 13, 01062 Dresden, Germany
(Requests for offprints should be addressed to G Vollmer; Email: Guenter.Vollmer@mailbox.tu-dresden.de)
Abstract
There is deﬁnitely a need for the development of new drugs for the treatment and cure of endometrial
cancer. In addition there are various new drugs or phyto-remedies under development which are
intended for use in the treatment and prevention of breast cancer, for the treatment of menopausal
symptoms and for hormone replacement therapy. The efﬁcacy of novel drugs targeting steroid
receptors in endometrial cancers has to be evaluated and the safety of other endocrine measures on
endometrial cancers or on endometrial carcinogenesis has to be assessed. For these experimental
purposes ﬁve main classes of experimental models are available: spontaneous endometrial
tumorigenesis models in inbred animals (Donryu rats, DA/Han rats, BDII/Han rats), inoculation tumors
from chunks of tumors (rat EnDA-tumor, human EnCa 101 tumor) or from inoculated tumor cell lines
(rat RUCA-I cells, human Ishikawa and ECC-1 cells), developmental estrogenic exposure or chemical
carcinogen exposure of CD-1 and ICR mice, transgenic approaches such as mice heterozygous
regarding the tumor suppressor gene PTEN (pten + / − -mice) and endometrial tumor cell lines cultured
under conditions promoting in vivo-like morphology and functions e.g. cell culture on reconstituted
basement membrane. Although the number of models is comparatively small, most aspects related
to functions of estrogenic or gestagenic substances are assessable, particularly if various
experimental models are combined. Whereas models based on human endometrial adenocarcinoma
cells are widely used, the properties and advantages of animal-derived models have mainly been
ignored so far.
Endocrine-Related Cancer (2003) 10 23–42
Introduction
Endometrial cancer is the most frequently diagnosed malignancy of the female genital tract. Incidence rates of endometrial cancer are described as 10 to 25 women per
100 000 with a clear geographic variation between e.g.
European (Spain, UK, France) and North American (USA
and Canada) countries, with a slightly higher incidence in
North America (Parazzini et al. 1991). In 1998, an estimate
of 36 000 new cases and 6300 deaths attributable to endometrial cancer were reported in the USA (Podratz et al.
1998) resulting in an overall frequency which is only
exceeded by breast, lung and colon cancer (Li et al. 1999).
However, the mortality rate of endometrial cancer if compared with other cancers is low and the prognosis is excellent if it is detected in the early stages (Creasman 1997).
The present knowledge of the role of endocrine factors in
the etiology of endometrial adenocarcinomas, their role in
the regulation of tumor growth, invasion and metastasis of
endometrial adenocarcinoma cells, as well as the potential
value of established and novel endocrine manipulations for
prevention and treatment of endometrial adenocarcinomas
have been exhaustively reviewed in a recent review article
(Emons et al. 2000).
In the concluding remarks of the same paper it was stated
that neither progestin treatment, the major hormonal therapy
of endometrial carcinoma, nor cytotoxic chemotherapy
showed substantial beneﬁts in the adjuvant setting. Therefore, future research activities must evaluate new compounds
and new treatment strategies.
The endometrium is a classical hormone-dependent
tissue and most of the endometrial adenocarcinomas are
hormone-dependent tumors. A high percentage of these
tumors express the estrogen receptor(s) and/or progesterone
receptors. Investigators have, therefore, aimed at targeting
steroid hormone receptors by the development of novel sub-
Endocrine-Related Cancer (2003) 10 23–42
1351-0088/03/010–023  2003 Society for Endocrinology Printed in Great Britain
Online version via http://www.endocrinology.org
Vollmer: Endometrial cancer
stances e.g. new antiestrogens, selective estrogen receptor
modulators (SERMs) and aromatase inhibitors. For testing
and characterization of these new substances appropriate
models have to be available.
A further need for endometrial tumor models stems from
safety considerations. Estrogens not only act as tumor promoters but also as carcinogens. This has consequences for
the approval of endocrine active substances intended for use
in tumor therapy e.g. breast cancer, tumor prevention or hormone replacement therapy. The concerns are that substances
which are beneﬁcial in one organ might harm another, e.g.
tamoxifen, a common therapy in breast cancer, might
increase the risk for endometrial cancer (Emons et al. 2000).
Finally, plenty of phytoestrogens are recommended either as
semipure substances or, as in the case of soy or red clover
extracts, for use as supportive measures in hormone replacement therapy as well as for use in lifestyle medicine. None
of these substances has been exhaustively investigated
regarding its safety (Fugh-Berman & Kronenberg 2001, Balk
et al. 2002, Burton & Wells 2002).
These examples clearly demonstrate the need for experimental endometrial tumor models. The aim of this paper
is to review the knowledge of endometrial tumor models.
Particular emphasis is given to their applicability in vivo and
to their potential responsiveness to natural as well as synthetic estrogens and progestins. The latter feature is a prerequisite for the process of development and evaluation of
endocrine active substances.
Spontaneous endometrial
adenocarcinoma of rats
Longevity studies revealed that female animals of four rat
strains die with an uncommonly high incidence rate for endometrial adenocarcinoma if kept to their natural life end. Due
to their clinical, pathophysiological, histological and biochemical features they all represent excellent experimental
models for endometrial carcinogenesis. Their potential as
experimental tumor models has so far not been appreciated
by the scientiﬁc community.
Han:Wistar rats
Already in 1981 Deerberg et al. published a report showing
that virgin Han:Wistar rats die with an incidence rate of 39%
from tumors of the uterus if kept to their natural life end. A
total of 35% of these tumors were endometrial adenocarcinomas. This rat strain and the pathogenesis of its endometrial
adenocarcinomas have not been investigated any further.
Donryu rats
Female Donryu rats have been presented as a hormonedependent endometrial cancer model, an issue discussed in
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detail below. This model is not well characterized regarding
histopathological properties and metastatic potential. Genetically, in spontaneous endometrial cancers point mutations of
the K-ras locus have been found recurrently (Tanoguchi et
al. 1999). The latter mutation also has a rather frequent
occurrence in human endometrial adenocarcinoma (Boyd &
Risinger 1991, Semczuk et al. 1998).
Female animals of the Donryu strain are characterized
by a similar mortality rate for endometrial adenocarcinoma
as Han:Wistar rats and exhibit an incidence rate of 35.1%
for endometrial adenocarcinoma and a total of proliferative
lesions of approximately 60% (Nagaoka et al. 1990). The
pathogenesis of endometrial adenocarcinoma in this rat strain
is characterized by some features which are discussed as risk
factors contributing to endometrial carcinogenesis in humans.
Donryu rats are characterized by an imbalanced (increased)
estradiol/progesterone rate which at the age of 12 months is
increased almost ﬁvefold if compared, for example, with
aged Fischer-344 rats (Nagaoka et al. 1990). In humans anovulatory cycles reﬂecting the estrogen/progesterone imbalance are regarded as a typical risk factor for endometrial
adenocarcinoma (Barakat et al. 1997).
It has been proposed that endometrial carcinogenesis in
Donryu rats is hormone dependent (Nagaoka et al. 1994).
However, this conclusion has been drawn from experimental
observations which should not necessarily be interpreted as
hormone dependency. The ﬁrst piece of evidence derives
from the above mentioned estrogen/progesterone imbalance
in the serum of these animals. The other piece of evidence
has been deduced from comparative studies with Fischer-344
rats which have a normal cyclicity and almost no advanced
histological changes such as hyperplastic or neoplastic
lesions (Nagaoka et al. 1994) or alterations in the proliferative capacity of the uterine epithelium (Ando-Lu et al. 1998).
Further, reproductive experience reduced the incidence of
endometrial and mammary carcinoma. This ﬁnding is analogous to the situation discussed for humans (Terry et al.
1999), although it only becomes apparent in Donryu rats after
three periods of gestation. One- or twofold experiences were
ineffective if compared with nulliparous animals (Nagaoka
et al. 2000). However, the conclusive experiments proving
hormone dependency, namely comparative studies in normal
cyclic and ovariectomized animals are missing.
The overall yield of endometrial adenocarcinoma could
be increased and the onset of endometrial adenocarcinoma
could be considerably accelerated if animals were intraperitoneally treated four times with a combination of estradiol
diproprionate and N-methyl-N-nitrosourea (Nagaoka et al.
1993) or if they received a single intra-uterine administration
of N-ethyl-N′-nitro-N-nitrosoguanidine via the vagina
(Ando-Lu et al. 1994). High fat diets also led to an earlier
onset of endometrial carcinogenensis which was paralleled
by an earlier onset of imbalance in estrogen/progesterone
levels (Nagaoka et al. 1995). Transplacental administration
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Endocrine-Related Cancer (2003) 10 23–42
of diethylstilbestrol on days 17 and 19 at a dose of 0.1 mg/
kg led to an increase in lesions of the female reproductive
tract of Donryu female offspring, including an increase in
endometrial adenocarcinoma (Kitamura et al. 1999).
Although the overall process of carcinogenesis is accelerated
by these chemical carcinogens or the tumor promoting estrogen, it is questionable whether results from studies applying
this approach can be interpreted more accurately than studies
using the spontaneous protocol. Treatment with chemical
carcinogens induces mutations which unpredictably add to
genetic alterations already present in Donryu rats, like the
frequent mutation of the Ki-ras locus (Tanoguchi et al.
1999).
So far two therapeutic studies have been described using
this model in the situation of spontaneous endometrial carcinogenesis. Treatment with dietary indole-3-carbinol
reduced the incidence rate of endometrial carcinogenesis
considerably. The study showed a clear correlation between
the induction of estradiol 2-hydroxylation and a more normalized estrogen/progesterone level (Kojima et al. 1994). On
a mechanistic level this means chemoprevention is due to
induction of estradiol 2-hydroxylase. The other chemoprevention study was performed on N-ethyl-N′-nitro-Nnitrosoguanidine-treated Donryu rats. Tamoxifen treatment
signiﬁcantly reduced the number of proliferative lesions in
the uteri which also was limited to a few hyperplastic lesions
(Yoshida et al. 1998). The authors claimed that tamoxifen
acted as an antiestrogen, because of decreased serum estradiol levels. However, in none of the papers were any comments on estrogen receptor involvement made. Receptor
involvement is crucial for any antiestrogenic function. In
addition, the ﬁnding mentioned above does not reﬂect the
situation in humans. For the human situation it is well
accepted that tamoxifen increases the relative risk for endometrial carcinogenesis (ACOG Committee Opinion 1996,
Love et al. 1999), a calculation from epidemiological ﬁndings substantiated by the stimulation of proliferation in cultured endometrial adenocarcinoma cells by tamoxifen
(Gusberg 1994, Creasman 1997).
DA/Han rats
DA/Han rats are characterized by an inbred background.
Speciﬁc gene alterations are not yet documented. The overwhelming majority of tumors are moderately to well differentiated which makes the tumor phenotypically similar to most
of the human endometrial adenocarcinomas. This model
exhibits a highly metastatic phenotype (Fig. 1), but has not
so far been used for intervention studies, because the tumor
can be transplanted into syngenic animals. There it gives rise
to metastasizing endometrial adenocarcinomas which by
histopathological and biochemical means (expression of
estrogen receptor α (ER-α)) are undistinguishable from the
parental tumor (Horn et al. 1993, 1994).
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Female DA/Han animals die from endometrial adenocarcinoma with an incidence rate of > 60% if kept to their natural life end at 24–27 months of age. Ovariectomy prior to
onset of cyclicity prevents endometrial carcinogenesis
(Deerberg et al. 1985). This has to be interpreted as a clear
indication of hormone dependency of the carcinogenic process. Sixty-three percent of all of the developing endometrial
adenocarcinomas of the DA/Han rats metastasize into the
lung, thereby using a lymphogenic pathway (Fig. 1; F Deerberg, unpublished observations). Lymphatic invasion is a
characteristic of type II endometrial carcinoma (Emons et al.
2000). Distant metastases are rare in cases of human endometrial cancer (Cook et al. 1999); however, among those few
cases the lung has been described as a frequent metastases
site (Bouros et al. 1996).
So far, these spontaneously occurring tumors have not
been used for experimental purposes. However, from these
tumors a serially transplantable tumor called EnDA (Horn et
al. 1993) and a cell line called RUCA-I (Schu¨tze et al. 1992)
which also gives rise to hormone-sensitive endometrial cancers if inoculated into syngenic rats or athymic nude mice
(see below) have been established (Fig. 2).
BDII/Han rats
In terms of an endometrial cancer model the female rats of
this strain are unique worldwide. Since being ﬁrst described
in 1987 (Deerberg & Kaspareit 1987) it took almost a decade
before they were used for cancer treatment experiments, and
an additional ﬁve more years before investigators started to
unravel their genetic peculiarities which contribute to the
high incidence of endometrial cancer. This tumor model is
genetically well characterized (discussed in detail below),
although some important genetic information is still missing
e.g. mutation frequency of some tumor suppressor genes (e.g.
APC, PTEN). Most histopathological properties known for
human endometrial adenocarcinoma, e.g. tumor grading, are
also found in this spontaneous animal model for endometrial
adenocarcinoma (Deerberg & Kaspareit 1987). Furthermore,
this model has been investigated for the occurrence of precancerous lesions and their immunohistochemical properties.
In rat endometrial tissue the same precancerous lesions have
been found which are also detectable in human specimens.
In addition, as in hyperplastic and displastic human endometrium, tenascin-C, a stromal marker for proliferative epithelial disease, is strongly up-regulated in similar lesions of
rat endometrial tissue (Vollmer et al. 1990, 1991, Sasano et
al. 1993). These ﬁndings illustrate the large similarities in
cell biological aspects in the pathogenesis of endometrial
cancers in humans and rats. In experimental setups for the
understanding of the molecular mechanisms of endometrial
carcinogenesis this model is of high value and should be considered as one of the most important in vivo models. There
are several experimental pieces of evidence that endometrial
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Vollmer: Endometrial cancer
Figure 1 Endometrial carcinogenesis and lung metastasis of DA/Han rats. In the upper panel the overall incidence of
endometrial carcinogenesis of DA/Han rats, dependent on age, is given. The lower panel shows the percentage of those tumors
developing lung metastases. ECA, endometrial carcinoma.
carcinogenesis of female BDII/Han animals is hormone
dependent and therefore represents an endometrial tumor
model for spontaneous hormonal carcinogenesis.
If female animals of this strain are kept to their natural
life end (around 27 months of age) they die from endometrial
carcinoma or metastases with an incidence rate of > 90%.
The overwhelming majority (87%–97%) of all carcinomas in
the various experimental groups were endometrial adenocarcinomas. The small remaining group consisted of anaplastic
carcinomas, adenosquamous carcinomas and squamous cell
carcinomas (Deerberg & Kaspareit 1987). This powerful
study was set up with 50 animals per experimental group
26
initially. In parallel experimental groups it was demonstrated
that neither maintenance of animals in a germ-free environment nor feeding of a puriﬁed diet reduced the incidence rate
of tumors. The group of retired breeders also exhibited an
incidence rate of > 90% but was characterized by a signiﬁcantly longer life span. This ﬁnding is a clear indication of
hormonal involvement in the carcinogenic process. Numbers
of pregnancies are regarded as a life style factor inversely
correlated to endometrial carcinogenesis in humans (Emons
et al. 2000 and references therein). But even more importantly, if BDII/Han rats are ovariectomized prior to estrous
cyclicity, they die from causes other than endometrial
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Endocrine-Related Cancer (2003) 10 23–42
Figure 2 The EnDA/RUCA model. This ﬁgure shows the experimental possibilities of the syngenic EnDA/RUCA model
consisting of spontaneous endometrial carcinogenesis of DA/Han rats, of the EnDA inoculation tumor and of cultured and/or
inoculated RUCA-I cells.
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27
Vollmer: Endometrial cancer
carcinoma, the incidence rate for this tumor decreasing to
0%. This is the strongest piece of evidence in favor of hormone-dependent endometrial carcinogenesis in BDII/Han
rats (Deerberg & Kaspareit 1987). This latter ﬁnding was
further substantiated in a more recent study in which life long
treatment with the progestin, melengestrol acetate, in three
different doses (0.1, 0.2 and 0.4 mg/kg/day) completely suppressed endometrial carcinogenesis (Deerberg et al. 1995).
At the highest dose glucocorticoid-like side effects were
observed.
Only recently has the hormonal carcinogenesis of inbred
BDII/Han rats been recognized as a model system to study
genes involved in endometrial carcinogenesis. Using them as
models for the understanding of genetic elements involved
in the ontogeny of endometrial adenocarcinoma in general,
genetic changes occurring in the course of endometrial carcinogenesis could be demonstrated in crossbreeding experiments applying comparative genomic hybridization (Helou et
al. 2001). The most common aberration was ampliﬁcation of
the proximal region of rat chromosome 4. The genes Cdk6
(cyclin dependent kinase 6) and Met (hepatocyte growth
factor receptor) were found to be located in the core of each
ampliﬁed region and ampliﬁed most recurrently and at the
highest level among the genes tested (Walentinsson et al.
2001). These data suggested that up-regulation of Cdk6 and/
or Met contributes to the development of endometrial cancers
in BDII/Han rats. The human homolog of Cdk6 has been
postulated to be an important player in cell cycle control.
Evidence exists that Cdk6 provides the link between growth
factor stimulation and onset of cell cycle progression
(Meyerson & Harlow 1994). However, the only documentation of Cdk6 overexpression in association with Cdk6
ampliﬁcation comes from human gliomas (Costello et al.
1997). The Met protoocogene encodes a transmembrane
growth factor which binds the cytokine hepatocyte growth
factor/scatter factor (HGF/SF). The highest levels of this
tyrosine kinase receptor are found in epithelial tissues. The
ligand, in contrast, is predominantly expressed in mesenchymal tissues suggesting a paracrine mode of action
(Gherardi & Stoker 1991). The major effect of HGF/SF function mediated by Met is thought to be epithelial cell proliferation and motility (Sugawara et al. 1997), including endometrial epithelium as well (Wagatsuma et al. 1998). Further,
it has been shown that HGF/SF can stimulate invasion of
endometrial adenocarcinoma cell lines if they express MET
(Bae-Jump et al. 1999).
Extra copies of chromosomal regions were found for
chromosome 6 and eight cancer-related genes were predicted
to be located in this chromosomal region of BDII/Han rats.
These genes comprised the N-myc protooncogene, apolipoprotein B, the DEAD box gene, ornithine decarboxylase, proopiomelanocortin, ribonucleotide reductase, M2 polypeptide
and syndecan. Ampliﬁcation of N-myc was by far the highest
suggesting that Mycn ampliﬁcation and overexpression con-
28
tributes to the development of this hormone-dependent tumor
(Karlsson et al. 2001). In addition, three major chromosomal
regions representing multiple susceptibility genes involved in
the development of endometrial adenocarcinoma in rat could
be identiﬁed (Roshani et al. 2001). Loss of heterozygosity
analysis revealed three major losses of heterozygosity
regions on chromosome 10 (Behboudi et al. 2001). By radiation hybrid mapping and single and dual color ﬂuorescence
in situ hybridization techniques a detailed chromosomal map
of the proximal part of chromosome 10 could be established.
With this approach the regional localization of 14 genes,
most of them cancer related, and of 5 microsatellite markers
could be determined (Behboudi et al. 2002).
Finally, in crossbreeding experiments of female BDII/
Han rats with males of two strains with a low incidence of
endometrial adenocarcinoma, three chromosomal regions,
which give rise to susceptibility for endometrial adenocarcinoma, could be identiﬁed (Roshani et al. 2001). This is a clear
indication that endometrial carcinogenesis of BDII/Han rats
exhibits heritable features. Interestingly, the genes affecting
susceptibility to endometrial adenocarcinoma were different
in the two crosses, suggesting that genes behind the susceptibility in BDII/Han animals may interact with various genes
in different genetic backgrounds. Data in the literature suggest that this feature of heritability of endometrial carcinogenesis in BDII/Han rats is shared by at least two forms of
human endometrial cancer (Sandles et al. 1992, Gruber &
Thompson 1996, Lynch et al. 1996). Whether or not this
heritability of endometrial carcinogenesis resembles the heritable phenotype in human endometrial adenocarcinoma of
patients with hereditary non-polyposis colorectal cancer
(HNPCC) remains open. The latter phenotype would, at least,
require microsatellite instability, mutation or alteration of
promotor methylation of mismatch repair genes and
mutations of the pten locus (Kuismanen et al. 2002, Whelan
et al. 2002, Zhou et al. 2002a,b). A corresponding genetic
analysis of endometrial adenocarcinoma of BDII/Han rats is
still missing.
Mouse models
Mouse models historically have proved to be particularly
useful in studying the effects of developmental exposure to
hormones, particularly estrogens. This topic will be discussed
in more detail below. In addition, the ICR mouse model
which responds to chemical carcinogens by induction of
endometrial cancer is described.
Transgenic mouse models are emerging tools for carcinogenesis in general. In the context of endometrial carcinogenesis pten + / − transgenic mice have to be discussed. These
animals exhibit a phenotype strongly resembling human
Cowden syndrome, a syndrome associated with high incidence of breast and endometrial neoplasia (Stambolic et al.
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Endocrine-Related Cancer (2003) 10 23–42
2000). This particular mouse strain, therefore, has to be discussed as a potential endometrial cancer model.
Developmental hormone (estrogen) exposure
In various mouse and rat strains estrogens increase the incidence of tumorigenesis in several organs (Liehr 2001 and
references therein), including the uterus, particularly during
developmental exposure. In humans, epidemiologists have
recognized prolonged estrogen action or estrogen use unopposed by gestagens as a risk factor for endometrial carcinogenesis (Key & Pike 1988, Weiderpass et al. 1999). From all
these data the question has been raised whether estrogens act
as carcinogens (Boyd 1996). From mechanistic and molecular studies more and more pieces of evidence have accumulated to strengthen this hypothesis (reviewed by Liehr 2000,
2001).
The neonatal mouse has been proposed as a model for
hormonal carcinogenesis of the endometrium (Newbold et al.
1990). If outbred mice are treated neonatally (days 1–5) they
will develop endometrial adenocarcinoma in a dosedependent manner and dependent on the strength of the estrogen. In the initial paper, Newbold et al. (1990) showed that
diethylstilbestrol and derivatives thereof are more potent than
estradiol. This model has been further characterized in terms
of cellular differentiation and gene expression in the epithelium (Yoshida et al. 2000). In the meantime, this model
has been successfully applied to test for the developmental
toxicity of tamoxifen (Newbold et al. 1997), catecholestrogens (Newbold & Liehr 2000) and genistein (Newbold et al.
2001). All these substances induce endometrial adenocarcinoma if injected in neonatal mice during postnatal days 1–
5. Whether this model can be replaced by transgenic mice
overexpressing ER and which exhibit an accelerated onset of
endometrial carcinogenesis (Couse et al. 1997) remains to be
elucidated, because these tumors are characterized by a more
aggressive phenotype and may therefore be less representative of the human situation than endometrial carcinogenesis
in wild-type animals.
In summary, CD-1 mice appear to be an excellent model
to test for developmental toxicity of estrogens, thereby evaluating the capacity of test substances to induce hormonedependent endometrial cancers.
Chemical carcinogen-induced
endometrial cancers in ICR mice
ICR mice respond to treatment with N-methyl-N-nitrosourea
(NMU) or estradiol with the development of endometrial
cancers within 30 weeks. This process is signiﬁcantly accelerated if ICR mice are treated with both agents simultaneously
(Niwa et al. 1991). The enhancing effects on endometrial
carcinogenesis initiated by NMU are not only detectable for
estradiol but also for estrone and estriol (Niwa et al. 1993).
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In comparison to human endometrial carcinogenesis, histopathological examinations revealed that these tumors
develop from various preneoplastic, hyperplastic lesions,
resembling the human situation (Niwa et al. 1991). Interestingly, the carcinogen and estradiol induce different preneoplastic lesions. Estradiol induces glandular hyperplasia, while
adenomatous hyperplasia is induced by NMU. For the development of atypical hyperplasia the cooperative action of
estradiol and NMU is favorable (Niwa et al. 1996).
Whereas the histopathology of NMU-induced endometrial tumors in ICR mice resembles the situation described
for endometrial carcinogenesis, the limited genetic analysis
described for the mouse model is different to human endometrial carcinogenesis. In specimens of human endometrial
cancer the mutation of various Ras loci (Boyd & Risinger
1991, Ignar-Trowbridge et al. 1992) and the p53 gene
(Risinger et al. 1992) are found quite often. These kinds of
mutations are very rare in NMU-initiated endometrial tumors
in ICR mice (Murase et al. 1995).
With regard to steroid receptor expression and protein
levels this model is poorly characterized. In a short-time protocol in ovariectomized ICR mice, animals respond to estrogen treatment by up-regulation of steady state mRNA levels
of the proto-oncogenes fos and jun (Niwa et al. 1998).
The NMU-induced and estradiol-enhanced endometrial
carcinogenesis proved to be particularly useful for treatment
studies. Several hormonal and traditional treatment procedures were tested leading with one exception to inhibition
or prevention of all tumors in the ICR model. Carcinogenesis
induced by a combination of estradiol and NMU was
inhibited by gestagen treatment (medroxyprogesterone acetate; Niwa et al. 1995) or by treatment with the antiestrogen,
toremifen (Niwa et al. 2002). In contrast, treatment with the
selective estrogen receptor modulator, tamoxifen, stimulated
endometrial carcinogenesis in NMU-treated animals with or
without the combined treatment with estradiol (Niwa et al.
1998).
Prevention of endometrial carcinogenesis in the above
described mouse model including a reduction of hyperplastic
foci was detected following treatment of animals with danazol (Niwa et al. 2000) or following treatment with traditional
Japanese or Chinese remedies such as extracts from Glycyrrhiza radix (Niwa et al. 1999) and Juzen-taiho-to (Niwa
et al. 2001). Shimotsu-to was identiﬁed as the active principle in Juzen-taiho-to (Lian et al. 2002). Finally, prevention
studies with isoﬂavones revealed that both genistein and
daidzein exhibited an inhibitory effect on endometrial carcinogensis in the estradiol/NMU ICR mouse model, particularly on the indices of atypical endometrial hyperplasia (Lian
et al. 2001).
In summary, NMU-induced endometrial adenocarcinomas in ICR mice are particularly sensitive to various hormonal treatment procedures. The limitation of this model is
its poor characterization of hormone responsiveness at a
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Vollmer: Endometrial cancer
molecular level. Neither the levels nor the qualities of
expressed steroid hormone receptors are known nor is there
any information on hormonally triggered signal transduction
cascades or responsive genes except for fos and jun.
Heterozygous mouse models with pten +/−
-mice as an example
PTEN/MMAC1/TEP1 has been detected as one of the most
commonly mutated tumor suppressor genes in human cancer
(Li et al. 1997, Steck et al. 1997). A signiﬁcant rate of PTEN
mutations has also been reported for human endometrial adenocarcinoma (Risinger et al. 1997, 1998, Tashiro et al. 1997).
This mutation has been found in up to 80% of cases of human
endometrial adenocarcinoma, and is already detectable in 33–
55% of the precancerous lesions (Latta & Chapman 2002,
Konopka et al. 2002). In addition to a mutation promoter,
hypermethylation is an alternative way to inactivate tumor
suppressor genes. There is one report in the literature showing
that the frequency of promoter hypermethylation of the PTEN
tumor suppressor gene in endometrial adenocarcinoma is
around 19% (Salvesen et al. 2001).
Although the importance of a functional PTEN protein
has been known for several years, the association of PTEN
expression with a prognosis is not yet clear. It appears as if
loss of PTEN expression is associated with metastatic disease
(Salvesen et al. 2002) and may serve as an independent prognostic marker for patients who undergo postoperative chemotherapy (Kanamori et al. 2002).
In order to understand the biological role of this dominant tumor suppressor gene, transgenic PTEN knock-out
mice have been created. A PTEN − / − mutation is lethal presumably due to defective chorio-allantoic development
(Suzuki et al. 1998).
Surprisingly, the mutation of one allele is sufﬁcient to
cause neoplasia in multiple organ systems in these pten + / −
-mice (Podsypanina et al. 1999). A particularly high incidence has also been detected in breast as well as in endometrial neoplasia and preneoplasia, strongly resembling human
Cowden syndrome (Stambolic et al. 2000). Indeed, in the
human situation and in addition to the occurrence of PTEN
mutations in sporadic tumor, germ-line mutations are
believed to cause related autosomal dominant hamartoma
syndromes, among them Cowden syndrome (Liaw et al.
1997, Eng 1998). Although the phenotype detectable in
pten + / − -mice in terms of the development of Cowden syndrome and the association with the high incidence of development of precancerous and cancerous lesions of breast and
endometrium is strikingly similar to the human situation,
some differences do exist. Unlike the human situation, in
pten + / − -mice neoplasia of the skin and brain were notably
absent, whereas the observed changes in the endometrium
were very consistent (Podsypanina et al. 1999).
30
Endometrial carcinoma, except colon carcinoma itself, is
the most common malignancy in patients with hereditary
non-polyposis colon cancer (HNPCC). This disease is characterized by germ-line mutations in mismatch repair genes
and by microsatellite instability (Fishel et al. 1993, Leach et
al. 1993, Peltoma¨ki et al. 1993, Bronner et al. 1994, Nicolaides et al. 1994, Papadopoulos et al. 1994). Surprisingly,
patterns of microsatellite instability differ between endometrial and colorectal tumors from patients with HNPCC
(Kuismanen et al. 2002).
To learn more about the involvement of these genes in
the pathogenesis of endometrial carcinogenesis, tumor specimens from patients with HNPCC and an established mutation
record of either the MLH1 or MSH2 missmatch repair gene
locus were subjected to analysis of the PTEN locus. Mutation
of the mismatch repair loci and PTEN gene were very
common in HNPCC, with PTEN mutation presumably preceding mutations of the mismatch repair loci (Zhou et al.
2002b). Interestingly, crosses of pten + / − - with mlh1 − /
−
-mice were characterized by an accelerated endometrial
tumorigenesis (Wang et al. 2002).
In summary, endometrial cancers from pten + / − -mice
with or without mlh1 − / − may serve as an excellent model
for hereditary endometrial cancer, because of the strong
resemblance with human Cowden disease and human
HNPCC. In addition, Stambolic et al. (2000) proposed
pten + / − -mice as a model for endometrial adenocarcinomas
which develop in women with unopposed estrogen stimulation. These patients rather commonly suffer from loss of heterozygosity at the PTEN locus and/or mutations in all stages
of endometrial hyperplasia (Risinger et al. 1997, 1998,
Tashiro et al. 1997).
Whether pten + / − -mice can serve as a hormonedependent cancer model or if they are suitable as models for
hormonal treatment protocols remains open, because no data
are available on these issues. However, despite the lack of
the latter information it has to be stated that pten + / − -mice
represent a promising new endometrial cancer model,
because of the similarity to hereditary human endometrial
cancer.
Inoculation tumors
These tumors are derived by inoculating chunks of tumors or
deﬁned numbers of cultured cells e.g. subcutaneously or into
the fat pad of syngenic animals, athymic nude mice or rats.
It is the scope of this paper to focus primarily on steroid
hormone dependent or at least steroid hormone sensitive in
vivo models. For this reason in the following sections evidence for hormone responsiveness from in vitro data is
brieﬂy summarized prior to the review of in vivo application
data of either model.
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Endocrine-Related Cancer (2003) 10 23–42
The RUCA-I/EnDA model
From a spontaneous endometrial adenocarcinoma a transplantable tumor called EnDA-tumor (Horn et al. 1993) and
a cell line called RUCA-I (Schu¨tze et al. 1992) have been
derived. These models have primarily been used to test novel
antiestrogens, potential agonistic activities of phytoestrogens
and o,p′-DDT as an example of an endocrine-disrupting
chemical.
To test for antiestrogenic activities of antiestrogens
chunks of EnDA-tumors or RUCA-I cells have been inoculated into syngenic DA/Han rats or athymic nude rats (Fig.
2; Horn et al. 1993, Vollmer & Schneider 1996). Tumor
growth from chunks of EnDA-tumors at the ectopic site as
well as formation of lymphogenic and pulmonary metastasis
are estrogen-sensitive processes, because the tumor growth
rate at the ectopic site, as well as the weight of ipsilateral
axillary lymph nodes and the number of lung metastases is
reduced in ovariectomized animals if compared with intact
animals. These effects most likely are mediated by the ERα,
which unlike the progesterone receptor is expressed in these
tumors. This experimental model has been successfully
applied to study, for example, the antiestrogenic activity and
function of ZK 119010 in an endometrial-derived experimental tumor model (Horn et al. 1993, 1994). As indicated above
an endometrial adenocarcinoma cell line, the RUCA-I cell
line, has been established from this transplantable EnDAtumor. This cell line is characterized by the expression of the
ERα and by its estrogen responsiveness in vitro and in vivo
(Fig. 2). If RUCA-I cells are cultured on reconstituted basement membrane (matrigel) they respond to treatment with
estradiol by stimulation of proliferation and alteration of gene
expression e.g. by the upregulation of complement C3
expression (Vollmer et al. 1995a) which is the major estrogen responsive gene in the rat uterus (Sundstro¨m et al. 1989),
as well as by down-regulation of ﬁbronectin expression
(Vollmer et al. 1995b) and up-regulation of clusterin gene
expression (Wu¨nsche et al. 1998). In vivo, if inoculated subcutaneously into the hind limb of syngenic DA/Han rats
RUCA-I cells form estrogen-sensitive metastasizing adenocarcinomas which by histological and biochemical means are
indistinguishable from tumors arising from chunks of EnDAtumors. Tumor growth and metastasis in both experimental
setups are very fast. After approximately 30–35 days animals
are clinically ill from the metastatic disease if 0.5–1 × 106
cells are inoculated at the ectopic site (Horn et al. 1994,
Vollmer & Schneider 1996).
In this transplantation approach RUCA-I cells at the inoculation site as well as metastases of these cells at lymphogenic
sites and in the lung react very sensitively to antiestrogen treatment. The weight of the primary tumor at the ectopic site, the
weight of the axillary ipsilateral lymph nodes as well as the
number of lung metastases were found to be reduced compared
www.endocrinology.org
with control animals in response to antiestrogen treatment of
inoculated animals (Vollmer & Schneider 1996). In
ectopically grown tumors, in lymph node metastases and in the
normal uterus (the latter organ was analyzed for control
purposes) expression of estrogen-dependent genes was downregulated signiﬁcantly following treatment of animals with the
pure antiestrogen ICI 182,780 (Wu¨nsche et al. 1998), amongst
these genes were matrix metalloproteinases (Tu¨shaus L,
Hopert AC & Vollmer G, unpublished observations).
Inoculation of RUCA-I cells into ovariectomized DA/
Han rats for the purpose of screening for estrogenic properties of phytoestrogens and endocrine-disrupting chemicals in
an endometrial-derived tumor model led to unexpected
results. Although tumor weight increased following oral
application of ethinyl estradiol in a 28-day treatment protocol, the weight of the tumor tissue remained almost unaffected following treatment of animals with a selected phytoestrogen, genistein (Diel et al. 2001) or a representative
industrial chemical o,p′-DDT (Diel et al., in preparation).
Unlike ethinyl estradiol-treated animals, there was no signiﬁcant alteration of gene expression detectable in tumor tissues of animals treated with genistein or o,p′-DDT. Control
investigations in the normal rat uterus showed that this organ
responded very sensitively to treatment with ethinyl estradiol, genistein and o,p′-DDT, both with an increase in tissue
weight as well as up-regulation of estrogen-dependent gene
expression (Diel et al. 2001).
In conclusion, RUCA-I cells if inoculated into syngenic
DA/Han rats represent a very sensitive estrogen-responsive
tumor model for the purpose of antiestrogen screening and
testing in an endometrial-derived tumor model. The RUCA-I
cell-derived model is of limited value for testing of agonistic
estrogenic properties of e.g. suspected phytoestrogens or
endocrine-disrupting chemicals. Experimentally, the EnDA/
RUCA-I model shows a unique feature: it is possible to work
in syngenic animals which means it is not necessary to use
athymic nude mice or rats having an imperfect immune
system which have to be kept in germ-free surroundings.
Ishikawa cells
Hormonal responsiveness of Ishikawa cells
in vitro
Since their ﬁrst description (Nishida et al. 1985) Ishikawa
cells, a human endometrial adenocarcinoma cell line expressing the estrogen and the progesterone receptors, are the most
widespread human endometrial-derived cell culture model.
Later on, applying the limited dilution method eighteen different clones of Ishikawa cells were established (Nishida et
al. 1996), ﬁfteen of them being estrogen receptor positive.
However, in terms of population doubling time, plating efﬁciency or saturation density there was no signiﬁcant
31
Vollmer: Endometrial cancer
difference among the clones (Nishida et al. 1996). In addition
to the expression of functional estrogen receptors (Holinka
et al. 1986a, b) and progesterone receptors (Lessey et al.
1996), Ishikawa cells express the androgen receptor (Lovely
et al. 2000) and the receptor for aryl hydrocarbons (Wormke
et al. 2000). There exists a large body of evidence for the
hormonal responsiveness of Ishikawa cells in vitro. In summary, the evidence for hormonal responsiveness is based
mainly on hormonal modiﬁcation of proliferation, cellular
functions or gene expression (for summary see Table 1 and
references therein).
In vitro the cells are in use for the elucidation of
molecular mechanisms of hormone action e.g. in drug
development, in the drug discovery process, for testing of
potential agonistic functions of antiestrogens or selective
estrogen receptor modulators in an endometrial-derived
model (Labrie et al. 2001), in studies of ligand independent
activation of the estrogen receptor, in anchorage independent tumor growth (Holinka et al. 1989), in studies on
factors controlling hormonal receptivity (Appa Rao et al.
2001), in environmental toxicology studies on the function
of phytoestrogens (Markiewicz et al. 1993, Liu et al. 2001,
Frigo et al. 2002) and endocrine-disrupting chemicals
(Bergeron et al. 1999) in an endometrial model, in paracrine cell/cell-interaction studies (Yang et al. 2001, Arnold
et al. 2002), in studies on signaling cross-talk
(Ignar-Trowbridge et al. 1995, Wormke et al. 2000), and
others (Kanishi et al. 2000).
Involvement of a switch in the angiogenic pathways
during tumorigenesis has become a recent focus of interest.
The participation of the ERα in this process was studied in
Ishikawa cells following overexpression of the ERα protein.
Overexpression of ERα not only signiﬁcantly inhibited the
growth of xenografted cells, but also down-regulated vascular endothelial growth factor expression in tumor xenografts,
resulting in a decreased vascularization of the tumors and the
inhibition of the angiogenic agent integrin αvβ3 (Ali et al.
2000). These experimental ﬁndings provide evidence in favor
of the assumption that high levels of ERα may be beneﬁcial
in the control of endometrial cancer due to its inhibitory
effects on angiogenic pathways.
Finally, endometrial carcinoma tumorgenicity could be
suppressed following introduction of chromosome 18 into
Ishikawa cells (Yamada et al. 1995). This chromosome is
known to harbor the presumptive tumor suppressor gene
DCC and therefore experimental evidence is provided for the
hypothesis of DCC being a candidate for an endometrial carcinoma tumor suppressor gene.
In summary, Ishikawa cells represent a combined in
vitro/in vivo human endometrial tumor model which is particularly suitable for the study of hormonal growth control.
In addition, it may be useful in characterizing organ speciﬁcity of natural and synthetic estrogens, like phytoestrogens
or endocrine-disrupting chemicals.
Enca101 tumor and ECC-1 cells
Ishikawa cells as an in vivo tumor model
Because of their hormone responsiveness in vitro Ishikawa
endometrial adenocarcinoma cells have also been developed
as an estrogen sensitive in vivo tumor model (Nishida et al.
1986). As mentioned above, eighteen subclones were isolated, all of these subclones were found being transplantable
into athymic nude mice (Nishida et al. 1996). This model has
mainly been used for two purposes: (1) to elucidate general
mechanisms in tumor biology, particularly towards the
understanding of estrogen and progesterone function in endometrial cancer and (2) as an endometrial model for hormonal
treatment of endometrial cancer.
The Ishikawa endometrial adenocarcinoma model proved
to be useful for studies on the regulation of growth control
by steroid hormones in endometrial adenocarcinoma in vivo
(Gong et al. 1994). The most important result of this study
was that 4-hydroxytamoxifen, like estradiol, stimulates endometrial tumor growth in vivo. This effect most likely
coincides with alteration of levels of expression of transforming growth factor (TGF)-α and TGF-β. In a similar
approach following inoculation of Ishikawa cells into the fat
pad of nude mice it could be demonstrated that raloxifen
exhibits a growth stimulatory potential in this assay
(Barsalou et al. 2002).
32
ECC-1 cells as an in vitro model
ECC-1 endometrial adenocarcinoma cells (Satyaswaroop &
Tabibzadeh 1991) have been derived from a transplantable
endometrial adenocarcinoma called EnCa101 which was
established immediately after the technique of tumor growth
in athymic nude mice became available (Satyaswaroop et al.
1981). Most importantly, ECC-1 cells form tumors with
glandular structures if inoculated into athymic nude mice
(Satyaswaroop & Tabibzadeh 1991).
The EnCA101 tumor and the ECC-1 cell line respond
to estrogen treatment (Satyaswaroop et al. 1983) and are
particularly rich in progesterone receptors (Clarke &
Satyaswaroop 1985). In addition, tumors respond to
tamoxifen treatment by stimulation of growth (Jordan et al.
1991, Tonetti et al. 1998), a feature which from epidemiological studies is expected to increase the risk for sporadic
human endometrial cancer during breast cancer therapy with
tamoxifen (Emons et al. 2000 and references therein). There
are many pieces of experimental evidence proving the
responsiveness of this model to treatment with sex steroid
hormones (for summary see Table 2). For these reasons the
EnCA101/ECC-1 tumor is the most widely used in vivo
human endometrial adenocarcinoma model.
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Endocrine-Related Cancer (2003) 10 23–42
Table 1 Hormonal responsiveness of Ishikawa cells
Receptor
Ligand(s)
Functional parameter
Regulation
Reference
ER
ER
ER
ER
ER
ER
ER
E2
E2
4-OHT
4-OHT
E2
EM652, EM800
E2
Stimulation
Stimulation
Stimulation
Stimulation
Stimulation
No regulation
Stimulation
Holinka et al. (1986a,b)
Holinka et al. (1989)
Anzai et al. (1989)
Croxtall et al. (1990)
Holinka et al. (1986c)
Labrie et al. (2001)
Fujimoto et al. (1996b)
ER
Resveratrol
E2, Tam
Down-regulation
Down-regulation
No regulation
Up-regulation
Bhat & Pezzuto (2001)
ER
ER
ER
ER
E2
Tam
E2
ER
AR
PR
PR
E2
DHT, T
P
MPA
PR
PR
PR
P; E2 + P
P
P
Proliferation
Proliferation, colony formation
Proliferation
Proliferation
Alkaline phosphatase
Alkaline phosphatase
Migration through basement
membrane
E2 induced alkaline phosphatase
ERα
ERβ
Proliferation, plasminogen
activator
Glycogen metabolism, gelatinase
Glycogen metabolism, gelatinase
Growth, cyclin D1, invasiveness,
MMP-1, -7, -9
AF2
Alkaline phosphatase
Growth
Growth
P27Kip
PAI-1
Estrogen sulfotransferase
α1ß1 integrin
PR
PR
MPA
MPA
PR
P
Sex hormone binding globulin
8 products from differential
display RT-PCR
VEGFs
PR
P
Osteopontin
Up-regulation
Down-regulation
Up-regulation
Stimulation
Up-regulation
Stimulation
Suppression
Up-regulation
Up-regulation
Up-regulation
Up-regulation
Upregulation
Up-/down-regulation
(gene dependent)
Down-regulation of E2
induced induction
Up-regulation
Hochner-Celnikier et al.
(1997)
Mizumoto et al. (2002)
Sakamoto et al. (2002)
Markiewicz & Gurpide (1997)
Holinka & Gurpide (1992)
Shiozawa et al. (2001)
Fujimoto et al. (1996a)
Falany & Falany (1996)
Lessey et al. (1996),
Castelbaum et al. (1997)
Misao et al. (1998)
Sakata et al. (1998)
Fujimoto et al. (1999)
Appa Rao et al. (2001)
Abbreviations: ER, estrogen receptor; E2, estradiol; 4-OHT, 4-hydroxytamoxifen; Tam, tamoxifen; DHT, dihydrotestosterone; T,
testosterone; P, progesterone; MPA, medroxyprogesterone acetate; MMP, matrix metalloproteinase; PAI, plasminogen activator
inhibitor; VEGF, vascular endothelial growth factor.
EnCa101 and ECC-1 cells-derived tumors
In terms of hormonal regulation of tumor growth, inoculation
of EnCa101 tumors or ECC-1 endometrial adenocarcinoma
cells into nude mice appears to be the most complete model.
It is responsive to estrogens and progestins and it grows following tamoxifen stimulation. In addition, the EnCA101
tumors exhibit another feature which is highly advantageous
in experimental cancer research: it can be used as a multi-site
transplantation model thereby saving animals (Heitjan et al.
2002). In these tumors the most interesting features of hormonal inﬂuences on endometrial tumor growth can be
studied, e.g. testing of novel estrogen receptor antagonists,
treatment of tamoxifen-stimulated tumor growth – a major
concern as indicated above (ACOG Committee Opinion
1996, Love et al. 1999) – and characterizing the effects of
progestins, these being the adjuvant therapy for endometrial
carcinoma for decades (Emons et al. 2000 and references
therein).
www.endocrinology.org
Once the estrogen dependency of tumor growth of
EnCa101 tumors (Satyaswaroop et al. 1983, Jordan et al.
1991) and their estrogen-responsiveness as measured by gene
expression, particularly of progesterone receptor (Clarke et
al. 1987) and of c-fos, (Sakakibara et al. 1992) had been
established, in a ﬁrst series of larger experiments the effects
of antiestrogens and progestins on the growth of these tumors
were investigated. In fact, for progestins schedules for progestin administration were designed in this model (Mortel et al.
1990).
For antiestrogens it could be shown that almost all antiestrogens tested stimulated tumor growth although to a lesser
degree than tamoxifen (Gottardis et al. 1990) with the exception of the pure antagonist ICI 164,384 which was not only
void of tumor growth stimulatory activity but also blocked
tamoxifen-induced tumor growth.
During combination treatments with tamoxifen and progestins, resistance of EnCA101 tumors to treatment became
apparent. This resistance was attributed to a desensitization
33
Vollmer: Endometrial cancer
Table 2 Hormonal responsiveness of EnCa101 tumors and ECC-1 cells
Cell line/
tumor
Receptor
Ligand
Functional parameter
Regulation
Reference
EnCa101
EnCa101
EnCa101
ECC-1
ER
ER
ER
ER
ER
ECC-1
ER
Growth in vivo
Growth in vivo
Growth in vivo
Growth in vivo
Growth in vivo
Growth in vivo
Growth in vivo
Proliferation in vitro
Increase
Increase
Increase
Increase
No effect
Stimulation
No effect
Stimulation
Satyaswaroop et al. (1993)
Jordan et al. (1991)
Jordan et al. (1989)
Dardes et al. (2002a)
ECC-1
E2
Tam
E2/Tam
E2
GW5638
E2
Tam, Ral
E2
EnCa101
ECC-1
EnCa101
ECC-1
ECC-1
ECC-1
ER
ER
ER
ER
ER
ER
E2
E2, BPA
E2, Tam
E2
E2
E2, Ral
ICI182,780
E2
E2
Progesterone receptor
Progesterone receptor
c-fos
Cathepsin D
GREB1
ER-α
ER-α
pS2, VEGF
HER2/neu
Stimulation
Stimulation
Induction
Induction
Induction
Down-regulation
Degradation
Induction
Down-regulation
Dardes et al. (2002b)
Bergeron et al. (1999), Castro-Rivera
et al. (1999), Dardes et al. (2002b)
Clarke et al. (1987)
Bergeron et al. (1999)
Sakakibara et al. (1992)
Castro-Rivera et al. 1999
Ghosh et al. (2000)
Dardes et al. (2002b)
Abbreviations: ER, estrogen receptor; E2, estradiol; Ral, raloxifen; Tam, tamoxifen; BPA, bisphenol A; VEGF, vascular
endothelial growth factor; pS2, presenelin-2.
following down-regulation of the progesterone receptor
(Satyaswaroop et al. 1992). Based on the results of another
study in which progesterone levels, progesterone receptor
levels and rates of tumor growth were assessed, it was predicted that intermittent progestin administration may result
in better control of endometrial cancer growth in the nude
mouse model (Mortel et al. 1990).
The EnCa101 tumor model is characterized by another
feature – the estrogen-inducible expression of aromatase.
Aromatase expression has been reported to be up-regulated
in endometrial adenocarcinoma, particularly in the stromal
tissue compartment (Watanabe et al. 1995, Sasano et al.
1996). Analyses of aromatase (Cyp19) gene polymorphism
revealed that these polymorphisms might represent one genetic risk factor for endometrial cancer development (Berstein
et al. 2001). Despite the clear up-regulation of aromatase
activity in endometrial cancer, the use of aromatase inhibitors
in a clinical setting has not been exhaustively investigated
and the clinical outcome is questionable (Rose et al. 2000,
Berstein et al. 2002).
Estrogen-inducible aromatase activity, however, is
interesting for general tumor biology because the role of
androgens in the growth of endometrial carcinoma is poorly
understood. The question of androgen sensitive endometrial
tumor growth was addressed in this particular tumor model.
There was little detectable growth promoting activity of
androgens which was lower by far than that of estrogens.
Surprisingly, there was no difference in the effects of
androgens on the growth of EnCa101 tumors if the effects of
aromatizable and non-aromatizable androgens were compared (Legro et al. 2001).
34
In summary, the EnCa101/ECC-1 tumor model is the
most complete endometrial tumor model because the effects
of estrogens, progestins and androgens can be assessed due
to the stable and, in part, estrogen inducible expression of the
estrogen receptor, the progesterone receptor and aromatase.
These concluding remarks have recently been substantiated
by studies on the characterization of some novel antiestrogens: the tamoxifen analog GW5638 and the piperidin
derivative ERA-923. These substances, like tamoxifen, block
growth of breast cancer cells and, in addition, more effectively block EnCa101 endometrial tumor growth
(Greenberger et al. 2001, Dardes et al. 2002a).
Differentiation of endometrial
adenocarcinoma cells in vitro
Endometrial adenocarcinomas differ in their ability to form
glandular structures; however most of the sporadic occurring
endometrial adenocarcinomas are well to moderately well
differentiated. In conventional cell culture on plastic in the
presence of charcoal stripped fetal calf serum or in the presence of a serum-free deﬁned medium, endometrial adenocarcinoma cells acquire a polygonal cell shape with a low
degree of differentiation, and in a more narrow view cannot
be regarded as correlated to the relatively high differentiated
status of endometrial adenocarcinoma cells in vivo.
For cells derived from a variety of organs, particularly
for normal and malignant mammary cells, a huge body of
evidence has accumulated (for review see Hansen & Bissell
2000) that culturing of glandular cells on reconstituted
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Endocrine-Related Cancer (2003) 10 23–42
Alteration of
gene expression
Adenocarcinoma cell
(dedifferentiated)
basement
membrane
TN-C rich
Stromal
matrix
Adenocarcinoma cell on
basement membrane
(differentiated)
Processes regulated by basement membrane
Reference
Cellular morphology and ultrastructure
Satyaswaroop & Tabibzadeh. (1991), Hopfer et al. (1996),
Behrens et al.(1996), Pinelli et al. (1998)
Proliferation
Tan et al. (1999)
Hormone responsiveness
Vollmer et al. (1995a,b), Wünsche et al. (1998)
Gene expression in general
Strunck et al. (1996)
Expression of integrins
Strunck and Vollmer (1996)
Expression of cytokeratins
Pastor U, Strunck E & Vollmer G, unpublished observations
Mitochondrial gene expression
Strunck E & Vollmer G, unpublished observations
icb-1 expression (differentiation marker)
Treeck et al. (1998)
Expression of phosphatases (L3-PSP)
Strunck et al. (2001)
Proteases (MMP2, MMP13)
Tüshaus L, Holpert A C & Vollmer G, in preparation
Figure 3 Responses of endometrial adenocarcinoma cells to contact with reconstituted basement membrane. The upper part is
a schematic drawing of the situation of an endometrial adenocarcinoma cell grown in conventional cell culture on plastic
(dedifferentiated) and on reconstituted basement membrane (differentiated). This ﬁgure also indicates the presence of the
stromal matrix underneath the basement membrane, which in cases of endometrial adenocarcinoma is particularly rich in
tenascin-C (TN-C). The lower part shows the processes regulated by the basement membrane. icb-1, induced by contact to
basement membrane; L3-PSP, L3-phosphoserine phosphatase; MMP, matrix metalloproteinase.
basement membrane induces differentiation processes,
thereby increasing physiological responsiveness considerably
if compared with cells kept under conventional cell culture
conditions. The evidence for morphological and functional
differentiation of human endometrial adenocarcinoma cells
of Ishikawa (Pinelli et al. 1998), of the ECC-1 cell line
(Satyaswaroop et al. 1991) and of the RUCA-I rat endometrial adenocarcinoma cells (Vollmer et al. 1995a) is summarized in Fig. 3. The most important observation is that the
morphological differentiation strongly correlates with functional differentiation, the latter being most importantly evidenced by the induction of hormone responsiveness.
With the culturing of endometrial adenocarcinoma cells
on reconstituted basement membrane a more pronounced
www.endocrinology.org
physiological phenotype of these cells is achieved, allowing
the assessment of physiological in vivo functions in an in
vitro model. This assay can be modiﬁed into an in vitro
invasion assay for the assessment of hormonal inﬂuences on
tumor cell invasion through basement membranes (Fujimoto
et al. 1996b). Finally, a modiﬁcation of this assay has been
described by reconstituting the natural situation even further
and adding conditioned medium of endometrial stromal cells
or cultured endometrial stromal cells to the system. In this
way two important parameters in endometrial tumor growth
can be assessed: the contribution of paracrine cellular interactions to tumor growth and the hormonal regulation of cells
of the two tissue compartments in a single cell culture dish
(Arnold et al. 2002).
35
Vollmer: Endometrial cancer
Conclusions
There is deﬁnitely a need for the development of substances
for the treatment and cure of endometrial cancer. In addition
there are various new drugs or phyto-remedies under development which are intended to be used in the treatment and
prevention of breast cancer, for the treatment of menopausal
symptoms and for hormone replacement therapy. The efﬁcacy of novel drugs targeting steroid receptors in endometrial
cancers has to be evaluated and the safety of other endocrine
measures on endometrial cancers or on endometrial carcinogenesis has to be assessed. For these purposes a relatively
small number of animal models or combined in vivo/in vitro
tumor models, based on human endometrial adenocarcinoma
cells, are available. However, these models alone or in combination cover all needs for drug development or testing of
drug safety in relation to endometrial cancer. While models
consisting of human endometrial adenocarcinoma cells transplanted to athymic nude mice are commonly used, the potential of animal models consisting of spontaneous endometrial
carcinogenesis or transplantation of rat endometrial adenocarcinoma cells to syngenic animals is largely ignored. First
of all, with these models treatment protocols for chronic
treatment of both primary tumors and metastases can be
established. And even if the inoculation approach is used
there is still a considerable advantage because syngenic animals can be used. Thus the use of the cost intensive athymic
nude animal, a more artiﬁcial model which has to be kept
in germ-free surroundings because of its imperfect immune
system, can be avoided.
In the future, the number of studies using transgenic animals and therapy approaches in these animals will deﬁnitely
increase. The advantage of these tools is that the genetic
modiﬁcation leading to the carcinogenic process is more precisely deﬁned. In this way single genes or corresponding
signal transduction pathways can be targeted in quite a selective way.
Acknowledgements
The author is grateful to Dr Deerberg for giving authorization
for the use of unpublished data on lung metastasis of endometrial adenocarcinoma in DA/Han rats.
The author wishes to thank Drs Susanne Starcke, Jannette Wober and Oliver Zierau for critical reading of the
manuscript and for their helpful suggestions. In addition,
special thanks are due to Dr Oliver Zierau for his assistance
with the art work.
This paper was supported by the Deutsche Forschungsgemeinschaft Vo410/5-4, Vo410/6-1 and Vo410/6-3.
Dedication
This paper is dedicated to honour the work of the recently
deceased Dr Friedrich Deerberg, formerly of the German
36
Institute for Laboratory Animals. Dr Friedrich Deerberg systematically investigated spontaneous carcinogenesis in inbred
laboratory animals and developed experimental animal
models useful for studying cancer treatment and prevention.
Amongst these models are two spontaneous hormonedependent endometrial cancer models which are exhaustively
discussed in this paper. Personally, the author is indebted to
Dr Deerberg for supplying him with the DA/Han and BD/
Han rat endometrial cancer models and the cell lines developed thereof.
References
ACOG Committee Opinion 1996 Tamoxifen and endometrial
cancer. International Journal of Gynecology and Obstetrics 53
197–199.
Ali SH, O’Donnell AL, Balu D, Pohl MB, Seyler MJ, Mohamed S,
Mousa S & Dandona P (2000) Estrogen receptor-α in the
inhibition of cancer growth and angiogenesis. Cancer Research
60 7094–7098.
Ando-Lu J, Takahashi M, Imai S, Ishihara R, Kitamura T, Iijima
T, Takano S, Nishiyama K, Suzuki K & Maekawa A 1994
High-yield induction of uterine endometrial adenocarcinomas in
Donryu rats by a single intra-uterine administration of
N-ethyl-N′-nitro-N-nitrosoguanidine via the vagina. Japanese
Journal of Cancer Research 85 789–793.
Ando-Lu J, Sasahara K, Nishiyama K, Takano S, Takahashi M,
Yoshida M & Maekawa A 1998 Strain-differences in
proliferative activity of uterine endometrial cells in Donryu and
Fischer 344 rats. Experimental and Toxicologic Pathology 50
185–190.
Anzai Y, Holinka CF, Kuramoto H & Gurpide E 1989 Stimulatory
effects of 4-hydroxytamoxifen on proliferation of human
endometrial adenocarcinoma cells (Ishikawa line). Cancer
Research 49 2362–2365.
Appa Rao KB, Murray MJ, Fritz MA, Meyer WR, Chambers AF,
Truong PR & Lessey BA 2001 Osteopontin and its receptor
ανβ3 integrin are coexpressed in the human endometrium during
menstrual cycle but regulated differentially. Journal of Clinical
Endocrinology and Metabolism 86 4991–5000.
Arnold JT, Lessey BA, Seppa¨la¨ M & Kaufman DG 2002 Effect of
normal endometrial stroma on growth and differentiation in
Ishikawa endometrial adenocarcinoma cells. Cancer Research 62
79–88.
Bae-Jump V, Segreti EM & Vandermolen D 1999 Hepatocyte
growth factor (HGF) induces invasion of endometrial carcinoma
cell lines in vitro. Gynecological Oncology 73 265–272.
Balk JL, Whiteside DA, Naus G, DeFerrari E & Roberts JM 2002
A pilot study of the effects of phytoestrogen supplementation on
postmenopausal endometrium. Journal of the Society for
Gynecological Investigation 9 238–242.
Barakat RR, Park RC, Grigsby PW, Muss HD & Norris HJ 1997
Corpus: epithelial tumors. In Principles and Practice of
Gynecologic Oncology, edn 2, pp 859–896. Eds WJ Hoskins,
CA Perez & RC Young. Philadelphia: Lippincott-Raven.
Barsalou A, Dayan G, Anghel SI, Alaoui-Jamali M, Van de Velde
P & Mader S 2002 Growth-stimulatory and transcription
activation properties of raloxifene in human endometrial
Ishikawa cell. Molecular and Cellular Endocrinology 190
65–73.
www.endocrinology.org
Endocrine-Related Cancer (2003) 10 23–42
Behboudi A, Levan G, Hedrich HJ & Klinga-Levan K 2001 High
density marker loss of heterozygosity analysis of rat
chromosome 10 in endometrial adenocarcinoma. Genes
Chromosomes Cancer 32 330–341.
Behboudi A, Roshani L, Kost-Alimova M, Sjostrand E,
Montelius-Alatalo K, Rohme D, Klinga-Levan K & Stahl K
2002 Detailed chromosomal and radiation hybrid mapping in the
proximal part of rat chromosome 10 and gene order comparison
with mouse and human. Mammalian Genome 13 302–309.
Behrens P, Meissner C, Hopfer H, Schumann J, Tan MI,
Ellerbrake N, Strunck E & Vollmer G 1996 Laminin mediates
basement membrane induced differentiation of HEC1B
endometrial adenocarcinoma cells. Biochemical and Cellular
Biology 74 875–886.
Bergeron RM, Thompson TB, Leonard LS, Pluta L & Gaido KW
1999 Estrogenicity of bisphenol A in human endometrial
carcinoma cell line. Molecular and Cellular Endocrinology 150
179–187.
Berstein LM, Imyanitov EN, Suspitsin EN, Grigoriev MY, Sokolov
EP, Togo A, Hanson KP, Poroshina TE, Vasiljev DA,
Kovalevskij AY & Gamajunova VB 2001 CYP19 gene
polymorphism in endometrial cancer patients. Journal of Cancer
Research and Clinical Oncology 127 135–138.
Berstein L, Maximov S, Gershfeld E, Meshkova I, Gamajunova V,
Tsyrlina E, Larionov A, Kovalevskij A & Vasilyev D 2002
Neoadjuvant therapy of endometrial cancer with aromatase
inhibitor letrozole: endocrine and clinical effects. European
Journal of Gynecology and Reproductive Biology 105 161–165.
Bhat KPL & Pezzuto JM 2001 Resveratrol exhibits cytostatic and
antiestrogenic properties with human endometrial
adenocarcinoma (Ishikawa) cells. Cancer Research 61 6137–
6144.
Bouros D, Papadakis K, Siafakas N & Fuller AF Jr 1996 Patterns
of pulmonary metastasis from uterine cancer. Oncology 53 360–
363.
Boyd J 1996 Estrogen as a carcinogen: the genetics and molecular
biology of human endometrial carcinoma. Progress in Clinical
and Biological Research 394 151–173.
Boyd J & Risinger JI 1991 Analysis of oncogene alterations in
human endometrial carcinoma: prevalence of ras mutations.
Molecular Carcinogenesis 4 189–195.
Bronner CE, Baker SM, Morrison PT, Warren G, Smith LG,
Lescoe MK, Kane M, Earabino C, Lipford J & Lindblom A
1994 Mutation in the DANN mismatch repair gene homologue
hMLH1 is associated with hereditary non-polyposis colon
cancer. Nature 368 258–261.
Burton JL & Wells M 2002 The effect of phytoestrogens on the
female genital tract. Journal of Clinical Pathology 55 401–407.
Castelbaum AJ, Ying L, Somkuti SG, Sun J, Ilesami OA & Lessey
BA 1997 Characterization of integrin expression in a well
differentiated endometrial adenocarcinoma cell line (Ishikawa).
Journal of Clinical Endocrinology and Metabolism 82 136–142.
Castro-Rivera E, Wormke M & Safe S 1999 Estrogen and aryl
hydrocarbon responsiveness of ECC1 endometrial cancer cells.
Molecular and Cellular Endocrinology 150 11–21.
Clarke CL & Satyaswaroop PG 1985 Photoafﬁnity labeling of the
progesterone receptor from human endometrial carcinoma.
Cancer Research 45 5417–5420.
Clarke CL, Feil PD & Satyaswaroop PG 1987 Progesterone
receptor regulation by 17beta-estradiol in human endometrial
carcinoma grown in nude mice. Endocrinology 121 1642–1648.
Cook AM, Lodge N & Blake P 1999 Stage IV endometrial
www.endocrinology.org
carcinoma: a 10 year review of patients. British Journal of
Radiology 72 485–488.
Costello JF, Plass C, Arap W, Chapman VM, Held WA, Berger
MS, Su Huang HJ & Cavenee WK 1997 Cyclin-dependent
kinase 6 (CDK6) ampliﬁcation in human gliomas identiﬁed
using two dimensional separation of genomic DNA. Cancer
Research 57 1250–1254.
Couse JF, Davis VL, Hanson RB, Jefferson WN, McLachlan JA,
Bullock BC, Newbold RR & Korach KS 1997 Accelerated onset
of uterine tumors in transgenic mice with aberrant expression of
the estrogen receptor after neonatal exposure to diethylstilbestrol.
Molecular Carcinogenesis 19 236–242.
Creasman WT 1997 Endometrial cancer: incidence, prognostic
factors, diagnosis and treatment. Seminars in Oncology 24 SI
140–SI 150.
Croxtall JD, Elder MG & White JO 1990 Hormonal control of
proliferation in the Ishikawa endometrial adenocarcinoma cell
line. Journal of Steroid Biochemistry 35 665–669.
Dardes RC, O’Reagan RM, Gajdos C, Robinson SP, Bentrem D,
De Los Reyes A & Jordan VC 2002a Effects of a new clinically
relevant antiestrogen (GW5638) related to tamoxifen on breast
and endometrial cancer growth in vivo. Clinical Cancer
Research 8 1995–2001.
Dardes RC, Scha¨fer, JM, Pearce ST, Osipo C, Chen B & Jordan
VC 2002b Regulation of estrogen target genes and growth by
selective estrogen-receptor modulators in endometrial cancer
cells. Gynecological Oncology 85 498–506.
Deerberg F & Kaspareit J 1987 Endometrial carcinoma in BDII/
Han rats: model of a spontaneous hormone-dependent tumor.
Journal of the National Cancer Institute 78 1245–1251.
Deerberg F, Rehm S & Pittermann W 1981 Uncommon frequency
of adenocarcinoma of the uterus in virgin Han:Wistar rats.
Veterinary Pathology 18 707–713.
Deerberg F, Rehm S & Rapp KG 1985 Survival data and
age-associated spontaneous diseases in two strains of rats.
In The Contribution of Laboratory Animal Science to the
Welfare of Man and Animals, pp 171–179. Eds J Archibald,
J Ditchﬁeld & HC Rowsell. Stuttgart and New York:
Fisher-Verlag.
Deerberg F, Pohlmeyer G, Lo¨rcher K & Petrow V 1995 Total
suppression of spontaneous endometrial carcinoma in BDII/Han
rats by melengestrol acetate. Oncology 52 319–325.
Diel P, Smolnikar K, Schulz T, Laudenbach-Leschowski U,
Michna H & Vollmer G 2001 Phytoestrogens and
carcinogenesis – differential effects of genistein in experimental
models of normal and malignant rat endometeium. Human
Reproduction 16 997–1006.
Emons G, Fleckenstein G, Hinney B, Huschmand A & Heyl W
2000 Hormonal interactions in endometrial cancer.
Endocrine-Related Cancer 7 227–242.
Eng C 1998 Genetics of Cowden syndrome: through the looking
glass of oncology. International Journal of Oncology 12 701–
710.
Falany JL & Falany CN 1996 Regulation of estrogen
sulfotransferase in human endometrial adenocarcinoma cells by
progesterone. Endocrinology 137 1395–1405.
Fishel R, Lescoe MK, Rao MR, Copeland NG, Jenkins NA, Garber
J, Kane M & Kolodner R 1993 The human mutator gene
homolog MSH2 and its association with hereditary nonpolyposis
colon cancer. Cell 75 1215–1225.
Frigo DE, Duong BN, Melnik LI, Schief LS, Collins-Burrow BM,
Pace DK, McLachlan JA & Burrow ME 2002 Flavonoid
37
Vollmer: Endometrial cancer
phytochemicals regulate activator protein-1 signal transduction
pathways in endometrial and kidney stable cell lines. Journal of
Nutrition 132 1848–1853.
Fugh-Berman A & Kronenberg F 2001 Red clover (Trifolium
pratense) for menopausal women: current state of knowledge.
Menopause 8 333–337.
Fujimoto J, Hori M, Ichigo S & Tamaya 1996a Sex steroids
regulate the expression of plasminogen activator inhibitor-1
(PAI-1) and its mRNA in uterine endometrial cancer cells.
Journal of Steroid Biochemistry and Molecular Biology 59 1–8.
Fujimoto J, Hori M, Ichigo S, Morishita S & Tamaya T 1996b
Estrogen activates migration potential of endometrial cancer
cells through basement membrane. Tumour Biology 17 48–57.
Fujimoto J, Sakaguchi H, Hirose R, Ichigo S & Tamaya 1999
Progestins suppress estrogen-induced expression of vascular
endothelia growth factor (VEGF) subtypes in uterine endometrial
cancer cells. Cancer Letters 141 63–71.
Gherardi E & Stoker M 1991 Hepatocyte growth factor-scatter
factor: mitogen, motogen and met. Cancer Cells 3 227–232.
Ghosh MG, Thompson DA & Weigel RJ 2000 PDZK1 and
GREB1 are estrogen regulated genes expressed in
hormone-responsive breast cancer. Cancer Research 60 6367–
6375.
Gong Y, Murphy LC & Murphy LJ 1994 Hormonal regulation of
proliferation and transforming growth factors gene expression in
human endometrial adenocarcinoma xenografts. Journal of
Steroid Biochemistry and Molecular Biology 50 13–19.
Gottardis MM, Ricchio ME, Satyaswaroop PG & Jordan VC 1990
Effect of steroidal and nonsteroidal antiestrogens on the growth
of tamoxifen-stimulated human endometrial carcinoma
(EnCa101) in athymic mice. Cancer Research 50 3189–3192.
Greenberger LM, Annable T, Collins KI, Komm BS, Lyttle
CR, Miller CP, Satyaswaroop PG, Zhang Y & Frost P 2001
A new antiestrogen, 2-(4-hydroxy-phenyl)-3-methyl-1[4-(2-piperidin-1-yl-ethoxy)-benzyl]-1H-indol-5-ol hydrochloride
(ERA-923), inhibits the growth of tamoxifen-sensitive and
-resistant tumors and is devoid of uterotropic effects in mice and
rats. Clinical Cancer Research 7 3166–3177.
Gruber SB & Thompson WD 1996 A population-based study of
endometrial cancer and familial risk in younger women. Cancer
and steroid hormone study group. Cancer, Epidemiology,
Biomarkers & Prevention 5 411–417.
Gusberg SB 1994 Estrogen and endometrial cancer: an epilogue a`
la recherche du temps perdu. Gynecology and Oncology 52 3–9.
Hansen RK & Bissell MJ 2000 Tissue architecture and breast
cancer: the role of extracellular matrix and steroid hormones.
Endocrine-Related Cancer 7 95–113.
Heitjan DF, Derr JA & Satyaswaroop PG 1992 The multi-site
tumour transplantation model for human endometrial carcinoma:
a statistical evaluation. Cell Proliferation 25 193–203.
Helou K, Walentinsson A, Beckmann B, Johansson A, Hedrich HJ,
Szpirer C, Klinga-Levan K & Levan G 2001 Analysis of genetic
changes in rat endometrial carcinomas by means of comparative
genomic hybridization. Cancer Genetics and Cytogenetics 127
118–127.
Hochner-Celnikier D, Greenﬁeld C, Finci-Yeheskel, Milwidsky A,
Gutman A, Goldman-Wohl D, Yagel S & Mayer M 1997
Tamoxifen exerts oestrogen-agonistic effects on proliferation and
plasminogen activation, but not on gelatinase activity, glycogen
metabolism and p53 protein expression, in cultures of
oestrogen-responsive human endometrial adenocarcinoma cells.
Molecular Human Reproduction 3 1019–1027.
38
Holinka CF & Gurpide E 1992 Growth-promoting effects of
progesterone in human endometrial cancer cell line
(Ishikawa-Var I). Journal of Steroid Biochemistry and Molecular
Biology 43 635–640.
Holinka CF, Hata H, Kuramoto H & Gurpide E 1986a Responses
to estradiol in a human endometrial adenocarcinoma cell line
(Ishikawa). Journal of Steroid Biochemistry 24 85–89.
Holinka CF, Hata H, Gravanis A, Kuramoto H & Grupide E 1986b
Effects of estradiol on proliferation of endometrial
adenocarcinoma cells (Ishikawa cells). Journal of Steroid
Biochemistry 25 781–786.
Holinka CF, Hata H, Kuramoto H & Gurpide E 1986c Effects of
steroid hormones and antisteroids on alkaline phosphatase
activity in human endometrial cancer cells (Ishikawa line).
Cancer Research 46 2771–2774.
Holinka CF, Anzai Y, Hata H, Kimmel N, Kuramoto H & Gurpide
E 1989 Proliferation and responsiveness to estrogen of human
endometrial cancer cells under serum-free culture conditions.
Cancer Research 49 32078–33301.
Hopfer H, Rinehart CA Jr, Kaufman DG & Vollmer G 1996
Basement membrane induced differentiation of HEC-1B(L)
endometrial adenocarcinoma cells affects both morphology and
gene expression. Biochemical Cell Biology 74 165–177.
Horn DW, Vollmer G, Deerberg F & Schneider MR 1993 The
EnDA endometrial adenocarcinoma: an oestrogen-sensitive,
metastasizing in vivo tumour model of the rat. Journal of Cancer
Research and Clinical Oncology 119 450–456.
Horn DW, Vollmer G, von Angerer E & Schneider MR 1994
Effect of the nonsteroidal antiestrogen ZK 119,010 on growth
and metastasis of the EnDA endometrial carcinoma.
International Journal of Cancer 58 426–429.
Ignar-Trowbridge D, Risinger JI, Dent GA, Kohler M, Berchuk A,
McLachlan JA & Boyd J 1992 Mutations of the Ki-ras oncogene
in endometrial carcinoma. American Journal of Obstetrics and
Gynecology 167 227–232.
Ignar-Trowbridge DM, Pimentel M, Teng CT, Korach KS &
McLachlan JA 1995 Cross talk between peptide growth factor
and estrogen receptor signaling systems. Environmental Health
Perspectives 103 (Suppl. 7) 35–38.
Jordan VC, Gottardis MM, Robinson SP & Friedl A 1989
Immune-deﬁcient animals to study ‘hormone-dependent’ breast
and endometrial cancer. Journal of Steroid Biochemistry 34 169–
176.
Jordan VC, Gottardis MM & Satyaswaroop PG 1991
Tamoxifen-stimulated growth of human endometrial carcinoma.
Annals of the New York Acadamy of Sciences 622 439–446.
Kanamori Y, Kigawa J, Itamochi H, Sultana H, Suzuki M, Ohwada
M, Kanamura T, Sugiyama T, Kikuchi Y, Kita T, Fujiwara K &
Terakawa N 2002 Pten expression is associated with prognosis
for patients with advanced endometrial carcinoma undergoing
postoperative chemotherapy. International Journal of Cancer
100 686–689.
Kanishi Y, Kobayashi Y, Noda S, Ishizuka B & Saito B 2000
Differential growth inhibitory effect of melatonin on two
endometrial cancer cell lines. Journal of Pineal Research 28
227–233.
Karlsson A, Helou K, Walentinsson A, Hedrich HJ, Szpirer C &
Levan G 2001 Ampliﬁcation of Mycn, Ddx1, Rrm2 and Odc1 in
rat uterine endometrial carcinomas. Genes Chromosomes Cancer
31 345–356.
Key TJA & Pike MC 1988 The dose–effect relationship between
‘unopposed’ estrogens and endometrial mitotic rate: its central
www.endocrinology.org
Endocrine-Related Cancer (2003) 10 23–42
role in explaining and predicting endometrial cancer risk. British
Journal of Cancer 57 202–212.
Kitamura T, Nishimura S, Sasahara K, Yoshida M, Ando J,
Takahashi M, Shirai T & Maekawa A 1999 Transplacental
administration of diethylstilbestrol (DES) causes lesions in
female reproductive organs of Donryu rats, including
endometrial neoplasia. Cancer Letters 141 219–228.
Kojima T, Tanaka T & Mori H 1994 Chemoprevention of
spontaneous endometrial cancer in female Donryu rats by dietary
indole-3-carbinol. Cancer Research 54 1446–1449.
Konopka B, Paszko Z, Janiec-Jankowska A & Golulda M 2002
Assessment of the quality and frequency of mutations occurence
in PTEN gene in endometrial carcinomas and hyperplasias.
Cancer Letters 178 43–51.
Kuismanen SA, Moisio AL, Schweizer P, Truninger K, Salovaara
R, Arola J, Butzow R, Jiricny J, Nystro¨m-Lahti M & Peltoma¨ki
P 2002 Endometrial and colorectal tumors from patients with
hereditary nonpolyposis colon cancer display different patterns
of microsatellite instability. American Journal of Pathology 160
1953–1958.
Labrie F, Labrie C, Belanger A, Simard J, Giguere V, Tremblay
A & Tremblay G 2001 EM-652 (SCH57068), a pure SERM
having complete antiestrogenic activity in the mammary gland
and endometrium. Journal of Steroid Biochemistry and
Molecular Biology 79 213–225.
Latta E & Chapman WB 2002 PTEN mutations and evolving
concepts in endometrial neoplasia. Current Opinion in Obstetrics
and Gynecology 14 59–65.
Leach FS, Nicolaides NC, Wei YF, Liu B, Jen J, Parsons R,
Peltomaki P, Sistonen P, Aaltonen LA & Nystrom-Lahti M 1993
Mutation of a mutS homolog in hereditary nonpolyposis colon
cancer. Cell 75 1215–1225.
Legro RS, Kunselman AR, Miller AS & Satyaswaroop PG 2001
Role of androgens in the growth of endometrial carcinoma: an in
vivo animal model. American Journal of Obstetrics and
Gynecology 184 303–308.
Lessey BA, Ilesanmi AO, Castelbaum AJ, Somkuti SG, Chwalisz
K & Satyaswaroop P 1996 Characterization of functional
progesterone receptor in an endometrial adenocarcinoma cell line
(Ishikawa): progesterone-induced expression of the α1 integrin.
Journal of Steroid Biochemistry and Molecular Biology 59 31–
39.
Li C, Samsiol G & Iosif C 1999 Quality of life in endometrial
cancer survivors. Maturitas 31 227–236.
Li J, Yen C, Liaw D, Podsypanina K, Bose S, Wang SL, Puc J,
Miliaresis C, Rodgers L, McCombie R, Bigner SH, Giovanella
BC, Ittmann M, Tycko B, Hibshoosh H, Wigler MH & Parsons
R 1997 PTEN, a putative protein tyrosine phosphatase gene
mutated in human brain, breast, and prostate cancer. Science 275
1943–1947.
Lian Z, Niwa K, Tagami K, Hashimoto M, Gao J, Yokuyama Y,
Mori H & Tamaya T 2001 Preventive effects of isoﬂavones,
genistein and daidzein, on estradiol-17beta-related endometrial
carcinogenesis in mice. Japanese Journal of Cancer Research 92
726–734.
Lian Z, Niwa K, Gao J, Tagami K, Hashimoto M, Yokoyama Y,
Mori H & Tamaya T 2002 Shimotsu-to is the agent in
Juzen-taiho-to responsible for the prevention of endometrial
carcinogenesis in mice. Cancer Letters 182 19–26.
Liaw D, Marsh DJ, Li J, Dahia PL, Wang SI, Zheng Z, Bose S,
Call KM, Tsou HC, Paecocke M, Eng C & Parsons R 1997
Germline mutations of the PTEN gene in Cowden disease, an
www.endocrinology.org
inherited breast and thyroid cancer syndrome. Nature Genetics
16 64–67.
Liehr JG 2000 Is estradiol a genotoxic mutagenic carcinogen?
Endocrine Reviews 21 40–54.
Liehr JG 2001 Genotoxicity of the steroidal estrogens oestrone and
oestradiol: possible mechanisms of uterine and mammary cancer
development. Human Reproduction Update 7 273–281.
Liu J, Burdette JE, Xu H, Gu C, Van Breemen RB, Bhat KPL,
Booth N, Constantinou AI, Pezzuto JM, Fong HHS, Farnsworth
NR & Bolton JL 2001 Evaluation of estrogenic activity of plant
extracts for the potential treatment of menopausal symptoms.
Journal of Agriculture and Food Chemistry 49 2472–2479.
Love CDB, Muir BB, Scrimgeou JB, Leonard RCF, Dillon P &
Dixon JM 1999 Investigation of endometrial abnormalities in
asymptomatic women treated with tamoxifen and an evaluation
of the role of endometrial screening. Journal of Clinical
Oncology 17 2050–2054.
Lovely LP, Appa Rao KB, Gui Y & Lessey BA 2000
Characterization of androgen receptors in a well-differentiated
endometrial adenocarcinoma cell line (Ishikawa). Journal of
Steroid Biochemistry and Molecular Biology 74 235–241.
Lynch HT, Krush AJ, Larsen AL & Magnuson CW 1966
Endometrial carcinoma: multiple primary malignancies,
constitutional factors, and heredity. American Journal of Medical
Science 252 381–390.
Markiewicz & Gurpide 1997 Estrogenic and progestagenic
acitivities of physiologic and synthetic androgens, as measured
by in vitro bioassays. Methods and Findings in Experimental
and Clinical Pharmacology 19 215–222.
Markiewicz L, Garey J, Adlercreutz H & Gurpide E 1993 In vitro
bioassay of non-steroidal phytoestrogens Journal of Steroid
Biochemistry and Molecular Biology 45 399–405.
Meyerson M & Harlow E 1994 Identiﬁcation of G1 kinase activity
for cdk6, a novel cyclin D partner. Molecular and Cellular
Biology 14 2077–2086.
Misao R, Nakanishi Y, Fujimoto J & Tamaya T 1998 Effect of
medroxyprogesterone acetate on sex hormone-binding globulin
mRNA expression in the human endometrial cancer cell line
Ishikawa. European Journal of Endocrinology 138 574–582.
Mizumoto H, Saito T, Ashihara K, Nishimura M, Tanaka R &
Kudo R 2002 Acceleration of invasive activity via matrix
metalloproteinases by transfection of the estrogen receptor-alpha
gene in endometrial carcinoma cells. International Journal of
Cancer 100 401–406.
Mortel R, Zaino RJ & Satyaswaroop PG 1990 Designing a
schedule of progestin administration in the control of
endometrial carcinoma growth in the nude mouse model.
American Journal of Obstetrics and Gynecology 162 928–934.
Murase T, Niwa K, Morishita S, Itoh N, Mori H, Tanaka T &
Tamaya T 1995 Rare occurrence of p53 and ras gene mutations
in preneoplastic and neoplastic mouse endometrial lesion
induced by N-methyl-N-nitrosourea. Cancer Letters 92 223–227.
Nagaoka T, Onodera H, Matsushima A, Todate A, Shibutani M,
Ogasawara H & Maekawa A 1990 Spontaneous uterine
adenocarcinomas in aged rats and their relation to endocrine
imbalance. Journal of Cancer Research and Clinical Oncology
116 623–628.
Nagaoka T, Takeuchi M, Onodera H, Mitsumori K, Lu J &
Maekawa A 1993 Experimental induction of uterine
adenocarcinoma in rats by estrogen and N-methyl-N-nitrosourea.
In Vivo 7 525–530.
39
Vollmer: Endometrial cancer
Nagaoka T, Takeuchi, M, Onodera H, Matsushima Y, Ando-Lu
J & Maekawa A 1994 Sequential observation of spontaneous
endometrial adenocarcinoma development in Donryu rats.
Toxicological Pathology 22 261–269.
Nagaoka T, Onodera H, Hayashi Y & Maekawa A 1995 Inﬂuence
of high-fat diets on the occurence of spontaneous uterine
endometrial adenocarcinomas in rats. Teratogenesis,
Carcinogenesis and Mutagenesis 15 167–177.
Nagaoka T, Takegawa K, Takeuchi M & Maekawa A 2000 Effects
of reproduction on spontaneous development of endometrial
adenocarcinomas and mammary tumors in Donryu rats. Japanese
Journal of Cancer Research 91 375–382.
Newbold RR & Liehr JG 2000 Induction of uterine
adenocarcinoma in CD-1 mice by catechol estrogens. Cancer
Research 60 235–237.
Newbold RR, Bullock BC & McLachlan JA 1990 Uterine
adenocarcinoma in mice following developmental treatment with
estrogens: a model for hormonal carcinogenesis. Cancer
Research 50 7677–7681.
Newbold RR, Jefferson WN, Padilla-Burgos E & Bullock BC 1997
Uterine carcinoma in mice treated neonatally with tamoxifen.
Carcinogenesis 18 2293–2298.
Newbold RR, Banks EP, Bullock B & Jefferson WN 2001 Uterine
adenocarcinoma in mice treated neonatally with genistein.
Cancer Research 61 4325–4328.
Nicolaides NC, Papadopoulos N, Liu B, Wie YF, Carter KC,
Ruben SM, Rosen CA, Haseltine WA, Fleischmann RD &
Fraser CM 1994 Mutations of two PMS homologues in
hereditary nonpolyposis colon cancer. Nature 371 75–80.
Nishida M, Kasahara K, Kaneko M, Iwasaki H & Hayashi K 1985
Establishment of a new human endometrial adenocarcinoma cell
line, Ishikawa cells, containing estrogen and progesterone
receptors. Nippon Sanka Fujinka Gakkai Zasshi 37 1103–1111.
Nishida M, Kasahara K, Tsuji T, Kaneko M & Iwasaki H 1986
Mechanisms of the antitumor action of gestagens on endometrial
cancer. Nippon Sanka Fujinka Gakkai Zasshi 38 2214–2222.
Nishida M, Kasahara K, Oki A, Satoh T, Arai Y & Kubo T 1996
Establishment of eighteen clones of Ishikawa cells. Human Cell
9 109–116.
Niwa K, Tanaka T, Mori H, Yokoyama Y, Furui T, Mori H &
Tamaya T 1991 Rapid induction of endometrial carcinoma in
ICR mice treated with N-methyl-N-nitrosourea and
17beta-estradiol. Japanese Journal of Cancer Research 82 1391–
1396.
Niwa K, Murase T, Furui T, Morishita S, Mori H, Tanaka T, Mori
H & Tamaya T 1993 Enhancing effects of estrogens on
endometrial carcinogenesis initiated by N-methyl-N-nitrosoeurea
in ICR mice. Japanese Journal of Cancer Research 84 951–955.
Niwa K, Morishita S, Murase T, Itoh, N, Tanaka T, Mori H &
Tamaya T 1995 Inhibitory effects of medroxyprogesterone
acetate on mouse endometrial carcinogenesis. Japanese Journal
of Cancer Research 86 724–729.
Niwa K, Morishita S, Murase T, Mudigdo A, Tanaka T, Mori H &
Tamaya T 1996 Chronological observation of mouse endometrial
carcinogenesis induced by N-methyl-N-nitrosourea and
17beta-estradiol. Cancer Letters 104 115–119.
Niwa K, Morishita S, Hashimoto M, Itoh T, Fujimoto J, Mori H &
Tamaya T 1998 Effects of tamoxifen on endometrial
carcinogenesis in mice. Japanese Journal of Cancer Research 89
502–509.
Niwa K, Hashimoto M, Morishita S, Ykoyama Y, Mori H &
Tamaya T 1999 Preventive effects of Glycyrrhizae radix extract
40
on estrogen-related endometrial carcinogenesis in mice. Japanese
Journal of Cancer Research 90 726–732.
Niwa K, Hashimoto M, Morishita S, Yokoyama Y, Lian Z, Tagami
K, Mori H & Tamaya T 2000 Preventive effects of danazol on
endometrial carcinogenesis in mice. Cancer Letters 158 133–
139.
Niwa K, Hashimoto M, Morishita S, Lian Z, Tagami K, Mori H &
Tamaya T 2001 Preventive effects of Juzen-taiho-to on
N-methyl-N-nitrosourea and estradiol-17beta-induced
endometrial carcinogenesis in mice. Carcinogenesis 22 587–591.
Niwa K, Hashimoto M, Lian Z, Gao J, Tagami K, Yokoyama Y,
Mori H & Tamaya T 2002 Inhibitory effects of toremifene on
N-methyl-N-nitrosourea and estradiol-17beta-induced
endometrial carcinogenesis in mice. Japanese Journal of Cancer
Research 93 626–635.
Papadopoulos N, Nicolaides NC, Wei YF, Ruben SM, Carter KC,
Rosen CA, Haseltine WA, Fleischmann RD, Fraser CM &
Adams MD 1994 Mutation of mutL homolog in hereditary colon
cancer. Science 263 1625–1629.
Parazzini F, La Cecchia C, Bocciolone L & Francheschi S 1991
The epidemiology of endometrial cancer. Gynecology and
Oncology 41 1–16.
Peltoma¨ki P, Lothe RA, Aaltonen LA, Pylkkanen L, Nystrom-Lahti
M, Seruca M, David L, Holm R, Ryberg D & Haugen A 1993
Microsatellite instability associated with tumors that characterize
the hereditary non-polyposis colorectal carcinoma syndrome.
Cancer Research 53 5853–5855.
Pinelli DM, Drake J, Williams MC, Cavanagh D & Becker JL
1998 Hormonal modulation of Ishikawa cells during
three-dimensional growth in vitro. Journal of the Society for
Gynecological Investigation 5 217–223.
Podratz KC, Mariani A & Webb MJ 1998 Staging and therapeutic
value of lymphadenectomy in endometrial cancer (Editorial).
Gynecology and Oncology 70 163–164.
Podsypanina K, Ellenson LH, Nemes A, Gu J, Tamura M, Yamada
K, Cordon-Cardo C, Catoretti G & Fisher PE 1999 Mutation of
Pten/Mmac1 in mice causes neoplasia in multiple organ systems.
PNAS 96 1563–1568.
Risinger JI, Dent GA, Ignar-Trowbridge D, McLachlan JA, Tsao
MS, Senterman M & Boyd J 1992 p53 gene mutations in human
endometrial carcinoma. Molecular Carcinogenesis 5 250–253.
Risinger JI, Hayes AK, Berchuk A & Barrett JC 1997 PTEN/
MMAC1 mutations in endometrial cancers. Cancer Research 57
4736–4738.
Risinger JI, Hayes K, Maxwell GL, Carney ME, Dodge RK,
Barrett JC & Berchuk A 1998 PTEN mutation in endometrial
cancers is associated with favorable clinical and pathologic
characteristics. Clinical Cancer Research 4 3005–3010.
Rose PG, Brunetto VL, VanLe L, Bell J, Walker JL & Lee RB
2000 A phase II trial of anastrozole in advanced recurrent or
persistent endometrial carcinoma: a Gynecologic Oncology
Group study. Gynecological Oncology 78 212–216.
Roshani L, Wedekind D, Szpirer J, Taib Z, Szpirer C, Beckmann
B, Rivie`re M, Hedrich HJ & Klinga-Levan K 2001 Genetic
identiﬁcation of multiple susceptibility genes involved in the
development of endometrial carcinoma in a rat model.
International Journal of Cancer 94 795–799.
Sakakibara K, Kan NC & Satyaswaroop PG 1992 Both
17beta-estradiol and tamoxifen induce c-fos messenger
ribonucleic acid expression in human endometrial carcinoma
grown in nude mice. American Journal of Obstetrics and
Gynecology 166 206–212.
www.endocrinology.org
Endocrine-Related Cancer (2003) 10 23–42
Sakamoto T, Eguchi H, Omoto Y, Ayabe T, Mori H & Hayashi S
2002 Estrogen receptor-mediated effects of tamoxifen on human
endometrial cancer cells. Molecular and Cellular Endocrinology
192 93–104.
Sakata M, Kurachi H, Morishige K, Ogura K, Yamaguchi M,
Nishio Y, Ikegami H, Miyake A & Murata Y 1998 Messenger
RNA differential display reverse-transcriptase-polymerasechain-reaction analysis of a progesterone-suppressive gene in a
human endometrial-cancer cell line. International Journal of
Cancer 78 125–129.
Salvesen HB, MacDonald N, Ryan A, Jacobs IJ, Lynch ED,
Akslen LA & Das S 2001 PTEN methylation is associated with
advanced stage and microsatellite instability in endometrial
carcinomas. International Journal of Cancer 91 22–26.
Salvesen HB, Stefansson I, Kalvenes MB, Das S & Akslen LA
2002 Loss of PTEN expression is associated with metastatic
disease in patients with endometrial carcinoma. Cancer 94
2185–2191.
Sandles LG, Shulman LP, Elias S, Photopulos GJ, Smiley LM,
Posten WM & Simpson JL 1992 Endometrial carcinoma: genetic
analysis suggesting heritable site-speciﬁc uterine cancer.
Gynecological Oncology 47 167–171.
Sasano H, Kaga K, Sato S, Yajima A, Nagura H & Harada N 1996
Aromatase cytochrome P450 gene expression in endometrial
carcinoma. British Journal Cancer 74 1541–1544.
Sasano H, Nagura H, Watanabe K, Ito K, Tsuiki A, Sato S, Yajima
A, Kusakabe M & Sakakura T 1993 Tenascin expression in
normal and abnormal human endometrium. Modern Pathology 6
323–326.
Satyaswaroop PG & Tabibzadeh SS 1991 Extracellular matrix and
the patterns of differentiation of human endometrial carcinomas
in vitro and in vivo. Cancer Research 51 5661–5666.
Satyaswaroop PG, Zaino RJ & Mortel R 1983 Human endometrial
adenocarcinoma transplanted into nude mice: growth regulation
by estradiol. Science 219 58–60.
Satyaswaroop PG, Clarke CL, Zaino RJ & Mortel R 1992
Apparent resistance in human endometrial carcinoma during
combination treatment with tamoxifen and progestin may result
from desensitization following downregulation of tumor
progesterone receptor. Cancer Letters 62 107–114.
Schu¨tze N, Kraft V, Deerberg F, Winking H, Knuppen R &
Vollmer G 1992 Functions of estrogens and antiestrogens in the
rat endometrial adenocarcinoma cell lines RUCA-I and
RUCA-II. International Journal of Cancer 52 941–949.
Semczuk A, Berbec H, Kostuch M, Cybulski M, Wojcierowski J &
Baranowski W 1998 K-ras gene point mutations in human
endometrial carcinomas: correlation with clinicopathological
features and patients outcome. Journal of Cancer Research and
Clinical Oncology 124 695–700.
Shiozawa T, Horiuchi A, Kato K, Obinata M, Konishi I, Fujii S &
Nikaido T 2001 Up-regulation of p27Kip1 by progestins is
involved in the growth suppression of the normal and malignant
human endometrial glandular cells. Endocrinology 142 4182–
4188.
Stambolic V, Tsao M-S, Macpherson D, Suzuki A, Chapman
WB & Mak TW 2000 High incidence of breast and endometrial
neoplasia resembling human Cowden syndrome in pten + / −
mice. Cancer Research 60 3605–3611.
Steck P, Pershouse MA, Jasser SA, Yung WK, Lin H, Ligon AH,
Langford LA, Baumgard ML, Hattier T, Davis T, Frye C, Hu R,
Swedlund B, Teng DH & Tavtigian SV 1997 Identiﬁcation of a
candidate tumour suppresor gene, MMAC1, at chromosome
www.endocrinology.org
10q23.3 that is mutated in multiple advanced cancers. Nature
Genetics 15 356–362.
Strunck E & Vollmer G 1996 Variants of integrin beta 4 subunit in
human endometrial adenocarcinoma cells: mediators of
ECM-induced differentiation? Biochemical and Cellular Biology
74 867–873.
Strunck E, Hopert AC & Vollmer G 1996 Basement membrane
regulates gene expression in HEC1B(L) endometrial
adenocarcinoma cells. Biochemical and Biophysical Research
Communications 221 346–350.
Strunck E, Frank K, Tan MI & Vollmer G 2001 Expression of
L-3-phosphoserine phosphatase is regulated by reconstituted
basement membrane. Biochemical and Biophysical Research
Communications 281 747–753.
Suguwara J, Fukaya T, Murakami T, Yoshida H & Yajima A 1997
Hepatocyte growth factor stimulated proliferation, migration, and
lumen formation of human endometrial epithelial cells in vitro.
Biology of Reproduction 57 936–942.
Sundstro¨m SA, Komm BS, Ponce-de-Leon H, Yi Z, Teuscher C &
Lyttle CR 1989 Estrogen regulation of tissue-speciﬁc expression
of complement C3. Journal of Biological Chemistry 264 16941–
16947.
Suzuki A, de la Pompa JL, Stambolic V, Elia AJ, Sasaki T, del
Barco Barrantes I, Ho A, Wakeham A, Itie A, Khoo W,
Fukumoto M & Mak TW 1998 High cancer susceptibility and
embryonic lethality associated with mutations of the PTEN
tumor suppresssor gene in mice. Current Biology 8 1169–1178.
Tan MI, Strunck E, Scholzen T, Gerdes J & Vollmer G 1999
Extracellular matrix regulates steady-state mRNA levels of the
proliferation associated protein Ki-67 in endometrial cancer
cells. Cancer Letters 140 145–152.
Tanoguchi K, Yaegashi N, Jiko K, Maekawa A, Sato S & Yajima
A 1999 K-ras point mutations in spontaneously occurring
endometrial adenocarcinomas in Donryu rats. Tohoku Journal of
Experimental Medicine 189 87–93.
Tashiro H, Blazes MS, Wu R, Cho KR, Bose S, Wang SI, Li J,
Parsons R & Ellenson LH 1997 Mutations in PTEN are frequent
in endometrial carcinoma but rare in other common
gynecological malignancies. Cancer Research 57 3935–3940.
Terry P, Baron JA, Weiderpass E, Yuen J, Lichtenstein P & Nyren
O 1999 Lifestyle and endometrial cancer risk: a cohort study
from the Swedish twin registry. International Journal of Cancer
82 38–42.
Tonetti, DA, O’Reagan R, Tanjore S, England G & Jordan VC
1998 Antiestrogen stimulated human endometrial cancer growth:
laboratory and clinical considerations. Journal of Steroid
Biochemistry and Molecular Biology 65 181–189.
Treeck O, Strunck E & Vollmer G 1998 A novel basement
membrane-induced gene identiﬁed in the human endometrial
adenocarcinoma cell line (HEC1B) FEBS Letters 425 426–430.
Vollmer G & Schneider MR 1996 The rat endometrial
adenocarcinoma cell line RUCA-I: a novel hormone-responsive
in vivo/in vitro tumor model. Jourmal of Steroid Biochemistry
and Molecular Biology 58 103–115.
Vollmer G, Siegal GP, Chiquet-Ehrismann R, Lightner VA,
Arnholt H & Knuppen R 1990 Tenascin expression in the
human endometrium and in endometrial adenocarcinomas.
Laboratory Investigations 62 725–730.
Vollmer G, Deerberg F, Siegal GP & Knuppen R 1991 Altered
tenascin expression during spontaneous endometrial
carcinogenesis in the BDII/Han rat. Virchows Archive of B Cell
Pathology Including Molecular Pathology 60 83–99.
41
Vollmer: Endometrial cancer
Vollmer G, Ellerbrake N, Hopert A-C, Wu¨nsche W & Knuppen R
1995a Extracellular matrix induces hormone responsiveness in
RUCA-I rat endometrial adenocarcinoma cells. Journal of
Steroid Biochemistry and Molecular Biology 52 259–269.
Vollmer G, Hopert AC, Ellerbrake N, Wu¨nsche W & Knuppen R
1995b Fibronectin is an estrogen repressed protein in RUCA-I
rat endometrial adenocarcinoma cell. Journal of Steroid
Biochemistry and Molecular Biology 54 141–139.
Wagatsuma S, Konno R, Sato S & Yajima A 1998 Tumor
angiogenesis, hepatocyte growth factor, and c-Met expression in
endometrial carcinoma. Cancer 82 520–530.
Walentinsson A, Helou K, Wallenius V, Hedrich HJ, Szpirer C &
Levan G 2001 Independent ampliﬁcation of two gene clusters on
chromosome 4 in rat endometrial cancer: identiﬁcation and
molecular characterization. Cancer Research 61 8263–8273.
Wang H, Douglas W, Lia M, Edelmann W, Kucherlapati R,
Podsypanina K, Parsons R & Ellenson LH 2002 DNA mismatch
repair deﬁciency accelerates endometrial tumorigenesis in Pten
heterozygous mice. American Journal of Pathology 160 1481–
1486.
Watanabe K, Sasano, H, Harada N, Ozaki M, Niikura H, Sato S &
Yajima A 1995 Aromatase in human endometrial carcinoma and
hyperplasia. Immunohistochemical, in situ hybridization, and
biochemical studies. American Journal of Pathology 146 491–
500.
Weiderpass E, Adami H-O, Baron JA, Magnusson C, Bergstro¨m R,
Lindgren A, Correia N & Persson I 1999 Risk of endometrial
cancer following estrogen replacement with or without
progestins. Journal of the National Cancer Institute 91 1131–
1137.
Whelan AJ, Babb S, Mutch DG, Rader J, Herzog TJ, Todd C,
Ivanovich JL & Goodfellow PJ 2002 MSI in endometrial
carcinoma: absence of MLH1 promoter methylation is associated
with increased familial risk for cancers. International Journal of
Cancer 99 697–704.
42
Wormke M, Castro-Rivera E, Chen I & Safe S 2000 Estrogen and
aryl hydrocarbon receptor expression and crosstalk in human
Ishikawa endometrial cells. Journal of Steroid Biochemistry and
Molecular Biology 72 197–207.
Wu¨nsche W, Tenniswood MP, Hopert A-C, Schneider MR &
Vollmer G 1998 Estrogenic regulation of clusterin mRNA in
normal and malignant endometrial tissue. International Journal
of Cancer 76 684–688.
Yamada H, Sasaki M, Honda T, Wake N, Boyd J, Oshimura M &
Barrett JC 1995 Suppression of endometrial carcinoma cell
tumorigenicity by human chromosome 18. Genes Chromosome
Cancer 13 18–24.
Yang S, Fang Z, Gurates B, Tamura M, Miller J, Ferrer K &
Bulun SE 2001 Stromal PRs mediate induction of
17beta-hydroxysteroid dehydrogenase type 2 expression in
human endometrial epithelium: a paracrine mechanism for
inactivation of E2. Molecular Endocrinology 15 2093–2105.
Yoshida A, Newbold RR & Dixon D 2000 Abnormal
differentiation and p21 expression of endometrial epithelial cells
following developmental exposure to diethylstilbestrol (DES).
Toxicological Pathology 28 237–245.
Yoshida M, Kudoh K, Katsuda S, Takahashi M, Ando J &
Maekawa A 1998 Inhibitory effect of uterine endometrial
carcinogenesis in Donryu rats by tamoxifen. Cancer Letters 11
43–51.
Zhou CP, Loukola A, Salovaara R, Nystro¨m-Lahti M, Peltoma¨ki P,
de la Chapelle A, Aaltonen LA & Eng C 2002a PTEN
mutational spectra, expression levels, and subcellular localization
in microsatellite stable and unstable colorectal cancers. American
Journal of Pathology 161 439–447.
Zhou XP, Kuismanen S, Nystrom-Lahti M, Peltoma¨ki P & Eng C
2002b Distinct PTEN mutational spectra in hereditary
non-polyposis colon cancer syndrome-related endometrial
carcinomas compared to sporadic microsatellite unstable tumors.
Human Molecular Genetics 11 445–450.
www.endocrinology.org