Adipose Tissue-Derived Mesenchymal Stromal/Stem Cells, Obesity and the Tumor Microenvironment of Breast Cancer

Abstract

Breast cancer is the most frequently diagnosed cancer and a common cause of cancer-related death in women. It is well recognized that obesity is associated with an enhanced risk of more aggressive breast cancer as well as reduced patient survival. Adipose tissue is the major microenvironment of breast cancer. Obesity changes the composition, structure, and function of adipose tissue, which is associated with inflammation and metabolic dysfunction. Interestingly, adipose tissue is rich in ASCs/MSCs, and obesity alters the properties and functions of these cells. As a key component of the mammary stroma, ASCs play essential roles in the breast cancer microenvironment. The crosstalk between ASCs and breast cancer cells is multilateral and can occur both directly through cell–cell contact and indirectly via the secretome released by ASC/MSC, which is considered to be the main effector of their supportive, angiogenic, and immunomodulatory functions. In this narrative review, we aim to address the impact of obesity on ASCs/MSCs, summarize the current knowledge regarding the potential pathological roles of ASCs/MSCs in the development of breast cancer, discuss related molecular mechanisms, underline the possible clinical significance, and highlight related research perspectives. In particular, we underscore the roles of ASCs/MSCs in breast cancer cell progression, including proliferation and survival, angiogenesis, migration and invasion, the epithelial–mesenchymal transition, cancer stem cell development, immune evasion, therapy resistance, and the potential impact of breast cancer cells on ASCS/MSCs by educating them to become cancer-associated fibroblasts. We conclude that ASCs/MSCs, especially obese ASCs/MSCs, may be key players in the breast cancer microenvironment. Targeting these cells may provide a new path of effective breast cancer treatment.

Full text

Citation: Ritter, A.; Kreis, N.-N.;
Hoock, S.C.; Solbach, C.; Louwen, F.;
Yuan, J. Adipose Tissue-Derived
Mesenchymal Stromal/Stem Cells,
Obesity and the Tumor
Microenvironment of Breast Cancer.
Cancers 2022,14, 3908. https://
doi.org/10.3390/cancers14163908
Academic Editors: Daniele Generali
and Ida Paris
Received: 15 July 2022
Accepted: 9 August 2022
Published: 12 August 2022
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations.
Copyright: © 2022 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
cancers
Review
Adipose Tissue-Derived Mesenchymal Stromal/Stem Cells,
Obesity and the Tumor Microenvironment of Breast Cancer
Andreas Ritter * , Nina-Naomi Kreis , Samira Catharina Hoock, Christine Solbach , Frank Louwen
and Juping Yuan *
Obstetrics and Prenatal Medicine, Gynecology and Obstetrics, University Hospital Frankfurt, J. W.
Goethe-University, Theodor-Stern-Kai 7, D-60590 Frankfurt, Germany
*Correspondence: andreas.ritter@kgu.de (A.R.); yuan@em.uni-frankfurt.de (J.Y.);
Tel.: +49-69-6301-83297 (A.R.); +49-69-6301-5819 (J.Y.); Fax: +49-69-6301-6144 (A.R.); +49-69-6301-84183 (J.Y.)
Simple Summary:
Adipose tissue is the major microenvironment of breast cancer. Adipose tissue-
derived mesenchymal stromal/stem cells (ASCs/MSCs) are key players in adipose tissue. ASCs/MSCs,
particularly in the obese state, are critical in remodeling the tumor microenvironment and promoting
breast cancer progression. In this review, we have addressed the impact of obesity on ASCs/MSCs,
summarized the crosstalk between ASCs/MSCs and breast cancer cells, discussed related molecular
mechanisms, and highlighted related research perspectives.
Abstract:
Breast cancer is the most frequently diagnosed cancer and a common cause of cancer-related
death in women. It is well recognized that obesity is associated with an enhanced risk of more aggres-
sive breast cancer as well as reduced patient survival. Adipose tissue is the major microenvironment
of breast cancer. Obesity changes the composition, structure, and function of adipose tissue, which
is associated with inflammation and metabolic dysfunction. Interestingly, adipose tissue is rich in
ASCs/MSCs, and obesity alters the properties and functions of these cells. As a key component
of the mammary stroma, ASCs play essential roles in the breast cancer microenvironment. The
crosstalk between ASCs and breast cancer cells is multilateral and can occur both directly through
cell–cell contact and indirectly via the secretome released by ASC/MSC, which is considered to
be the main effector of their supportive, angiogenic, and immunomodulatory functions. In this
narrative review, we aim to address the impact of obesity on ASCs/MSCs, summarize the current
knowledge regarding the potential pathological roles of ASCs/MSCs in the development of breast
cancer, discuss related molecular mechanisms, underline the possible clinical significance, and high-
light related research perspectives. In particular, we underscore the roles of ASCs/MSCs in breast
cancer cell progression, including proliferation and survival, angiogenesis, migration and invasion,
the epithelial–mesenchymal transition, cancer stem cell development, immune evasion, therapy
resistance, and the potential impact of breast cancer cells on ASCS/MSCs by educating them to
become cancer-associated fibroblasts. We conclude that ASCs/MSCs, especially obese ASCs/MSCs,
may be key players in the breast cancer microenvironment. Targeting these cells may provide a new
path of effective breast cancer treatment.
Keywords:
ASCs/MSCs; obesity; breast cancer; tumor microenvironment; cancer-associated
fibroblasts
;
cancer-associated stem cells; epithelial–mesenchymal transition; therapy resistance
1. Introduction
The prevalence of obesity has tripled during the last decades, posing a major challenge
to the entire society and health care systems worldwide [
1
–
3
]. Obesity, a condition of
increased adiposity resulting from an imbalance between food intake and energy expen-
diture [
4
], is categorized according to body mass index (BMI)
≥
30 kg/m
2
[
5
]. Obesity
associates with multiple disorders, including diabetes, hypertension, and cardiovascular
Cancers 2022,14, 3908. https://doi.org/10.3390/cancers14163908 https://www.mdpi.com/journal/cancers

Cancers 2022,14, 3908 2 of 35
diseases [
6
]. In addition, it is characterized by an increased incidence of cancer in various
organs, such as the colon, rectum, kidney, pancreas, gallbladder, liver, thyroid, breast, ovary,
and endometrium [
7
–
9
]. Obesity associates with inflammation and metabolic dysfunction,
which greatly promote cancer development. Among various adipose tissue cell types,
adipose tissue-derived mesenchymal stromal/stem cells (ASCs), belonging to mesenchy-
mal stromal/stem cells (MSCs), are dramatically altered during obesity progression [
10
].
Obesity-associated ASCs are of crucial importance, contributing to the establishment of
the breast cancer microenvironment and promoting the progression of breast cancer. In
this narrative review, we have summarized the current knowledge regarding the potential
pathological roles of obese ASCs/MSCs in the development of breast cancer, addressed the
possible impact of obesity on ASCs/MSCs, discussed the intertwined relationship between
ASCs and breast cancer cells, explored associated molecular mechanisms, and highlighted
related research perspectives.
2. Method
A search was performed for original articles and reviews published between January
2000 and April 2022 in PubMed with a focus on ASCs/MSCs and breast cancer by using
the following search terms (or combination of terms): ASCs, MSCs, obesity, breast cancer,
cancer microenvironment, proliferation, invasion, metastasis, cancer progression, and
therapy resistance. Only English-language and full-text articles were included.
3. Obesity and Breast Cancer
Among the malignancies, breast cancer is the most frequently diagnosed cancer and a
common cause of cancer-related death in women. It originates from deregulation of normal
growth pathways in mammary epithelial cells due to genetic mutations or epigenetic modi-
fications [
11
]. Breast cancer is composed of distinct subtypes and is highly heterogeneous
in both molecular and clinical terms. Based on the expression of the estrogen receptor (ER),
progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), as well
as a proliferation marker Ki67, breast cancer can be divided into four intrinsic molecular
subtypes: luminal A (ER
+
, PR
+
, HER2
−
, Ki67
Low
), luminal B (ER
+
, PR
+
, HER
+/−
, Ki67
High
),
HER2-enriched (ER
−
, PR
−
, HER2
+
), and basal-like subtype (ER
−
, PR
−
, HER2
−
), which
largely resembles triple-negative breast cancer (TNBC) and comprises approximately 15%
of all breast cancer cases [
12
]. Among these subtypes, TNBC, typically more aggressive,
is associated with a higher rate of relapse and poor prognosis owing to the development
of metastases in distant organs such as brain, liver, bone, and lungs [
13
,
14
]. The therapy
of breast cancer consists of surgical resection, radiation, chemotherapy, and other thera-
peutic options, including antihormone therapy, signaling pathway targeting, DNA repair
inhibition, aberrant epigenetic reversion, and immuno-oncology therapeutics [
15
]. Despite
advanced therapeutic options, breast cancer remains one of the most common causes of
female death [
16
], largely due to therapy resistance and metastases to brain, liver, lung, or
bone. It is therefore necessary to explore the molecular mechanisms of cancer progression
and pave new ways that offer more effective breast cancer treatment.
Obese women with breast cancer have larger tumors and an enhanced risk of metas-
tasis that contributes to a 30% increased risk of death [
17
–
19
]. Specifically, obesity as-
sociates with enhanced risk of more aggressive breast cancer as well as reduced sur-
vival of postmenopausal breast cancer patients [
20
,
21
]. While numerous meta-analyses
have consistently shown positive associations between obesity and risk of hormone-
receptor-positive (ER
+
and PR
+
) breast cancer [
22
–
24
], growing evidence suggests that
abdominal obesity, also known as central obesity, may increase the risk for TNBC in pre-
menopausal women
[25–27]
. Moreover, compared to lean patients, obese patients respond
worse to therapy, particularly when diagnosed with TNBC, contributing to the overall
worse prognosis [27].
The dramatic increase in the prevalence of obesity, combined with the fact that over
75% of new cases of breast cancer occur in postmenopausal women [
28
], represents a

Cancers 2022,14, 3908 3 of 35
pressing challenge in breast cancer prevention, treatment, and survival. Despite being a
field of intensive research, the molecular mechanisms underlying the association of obesity
with breast cancer are still incomplete.
Adipose Tissue: Microenvironment, ASCs and Obesity
Although breast cancer initiation is largely driven by acquired genetic alterations, the
tumor microenvironment (TME) is crucial in its progression. Breast cancer cells are mainly
surrounded by mammary adipose tissue and intermingled with a repertoire of stromal
cells such as ASCs, fibroblasts, endothelial, and immune cells. Breast cancer is able to
change the adjacent adipose tissue by stimulating the transcription of genes associated
with tumor growth, stemness, and progression [
29
,
30
]. In turn, adipose tissue cells, both
adipocytes as well as stromal cells, promote cancer progression by secreting growth factors,
cytokines, chemokines, and pro-migratory extracellular matrix (ECM) components [
31
,
32
].
Thus, breast-cancer-educated adipocytes and stromal cells, together with soluble factors
as well as insoluble ECM proteins secreted by adipocytes, stromal cells, and cancer cells,
constitute the TME [
15
,
33
]. Of importance, the TME is dynamic, as a multitude of stromal
cell types, such as endothelial progenitors, immune cells, fibroblasts, ASCs, and MSCs, are
recruited to develop the TME [
34
,
35
]. The TME is not only critical for cancer progression,
but also for coordinating cancer plasticity and immune escape [15].
Obesity changes the landscape of adipose tissue. In the early stages of adipose tissue
expansion, adipocyte hypertrophy generates a local hypoxia that contributes to increased
secretion of adipokines, inflammatory cytokines, and lipid metabolites. These changes fur-
ther reduce the metabolic flexibility of adipocytes, increase the rate of apoptosis, and recruit
more inflammatory cells, including lymphocytes and macrophages, in adipose tissue [36].
The chronic hypoxia observed in obese adipose tissue results in chronic inflammation, en-
doplasmic reticulum (ER) stress, and an alteration of the ECM. In fact, obese adipose tissue
is characterized by elevated saturated fatty acids released by obesity-associated lipolysis
that induce macrophage activation via toll-like receptor 4 (TLR4) stimulating nuclear factor
kappa B (NF-kB) signaling and inflammation in adipose tissues [
37
]. This further activates
the transcription of pro-inflammatory genes including interleukin 6 (IL6), IL1
β
, and tumor
necrosis factor
α
(TNF
α
) in adipocytes and stromal cells, causing local and systemic inflam-
mation [
38
]. Accordingly, obesity is characterized by increased levels of circulating factors
including insulin, insulin-like growth factor 1 (IGF-1), leptin, and inflammatory cytokines
such as IL6 and TNF
α
[
39
]. Obese adipose tissue is further marked by increased infiltration
of immune cells, cellular stress, hypoxia, insulin resistance, glucose intolerance, adipocyte
hyperplasia and hypertrophy, reduced angiogenesis, and impaired tissue homeostasis [
10
].
In particular, obesity increases the number of myofibroblasts in mammary adipose tissue
that deposits a more fibrillary and stiffer ECM, correlating with the malignant behavior of
mammary epithelial cells and with inflammation [
40
]. Thus, various cell types of obese
adipose tissue communicate with breast cancer cells, promote breast cancer progression,
reshape metabolism in the TME, and suppress anti-tumor immunity [
41
–
43
]. Collectively,
obesity changes the composition, structure, and function of adipose tissue, which associates
with inflammation and metabolic dysfunction and promotes breast cancer development.
Interestingly, adipose tissue, a mesodermal-derived organ, is the richest source of
MSCs in the human body [
44
]. MSCs are multipotent cells involved in the maintenance
of tissue homeostasis, regulation of local immune response, and regeneration of damaged
tissues [
45
]. ASCs, like other MSCs in the body, have many characteristics in common with
MSCs, including morphology, extensive proliferation potential, and the ability to undergo
multi-lineage differentiation
in vitro
[
32
,
46
–
48
]. ASCs are able to produce a large variety of
growth factors and have immunomodulatory properties [
10
,
49
,
50
]. Moreover, ASCs are
anti-inflammatory because of their multiple capabilities, such as inhibiting the activation of
natural killer cells, impairing cytotoxicity processes, reducing the proliferation of B cells,
decreasing immunoglobulin production, and suppressing B cell functions [49].

Cancers 2022,14, 3908 4 of 35
However, obesity alters most of the enumerated functions of ASCs, which in turn
affect their surrounding cells in adipose tissue [
10
,
51
]. In this context, the interaction
between ASCs and breast cancer cells could have a crucial role in malignant progres-
sion
[52–54]
. In fact, ASCs have been shown to impact breast cancer progression by
secreting cytokines/chemokines and other regulatory factors influencing angiogenesis, mi-
gration, and invasion of breast cancer cells [
53
,
55
,
56
]. Additionally, breast-cancer-associated
ASCs interfere with the immunomodulatory function of natural killer cells [
57
]. These
data suggest that the communication between obesity-associated ASCs, immune cells, and
breast cancer cells may modify cellular compartments, leading to the co-evolution of cancer
cells and their microenvironment [58].
4. Crosstalk between ASCs and Breast Cancer Cells
The crosstalk between ASCs and breast cancer cells can occur directly via cell–cell
interaction and indirectly via the secretome released by the cells. Cancer cells secrete
numerous chemotaxis signals [
59
], which recruit ASCs from local adipose tissues as well as
MSCs from bone marrow into malignant tissue [
60
]. Cancer-educated ASCs/MSCs may
differentiate into cancer-associated ASCs/MSCs or cancer-associated fibroblasts (CAFs) [
61
].
These ASCs/MSCs in turn promote breast cancer progression [
62
]. In particular, the
ASC/MSC secretome, which is composed of a large number of secreted proteins, peptides
and extracellular vesicles (EVs), is considered to be the main effector of their regenerative,
tropic, trophic, angiogenic, and immunomodulatory functions.
Regarding the indirect manner, both breast cancer cells and ASCs/MSCs are potent in
secreting a large number of soluble bioactive factors [
61
,
63
]. In particular, MSCs have the
ability to migrate into malignant areas and stimulate cancer development by secreting a
range of paracrine factors such as chemokines C-X-C ligand 1 (CXCL1), CXCL2, CXCL5,
CXCL7, and CXCL12/stromal-cell-derived factor 1 (SDF1); cytokines such as IL6, IL8, and
transforming growth factor
β
(TGF
β
); and growth factors including epidermal growth
factor (EGF), insulin-like growth factor 1 (IGF1), and vascular endothelial growth factor
(VEGF) [
58
,
61
,
64
]. It has been shown that MSCs facilitate angiogenesis by paracrine
secretion of angiogenic growth factors such as platelet-derived growth factor (PDGF) and
VEGF [
58
,
65
]. Our own studies also demonstrate that ASCs isolated from subcutaneous
as well as visceral adipose tissue released numerous cytokines, chemokines, and growth
factors involved in inflammation, angiogenesis, and cell migration and proliferation, such
as IL6, IL8, TNF
α
, CXCL1/2/3, CXCL5, monocyte chemotactic and activating factor (CCL2),
and EGF [32,48,66].
Moreover, both breast cancer cells and MSC/ASC-derived EVs are essential for the
crosstalk between MSCs/ASCs and breast cancer cells [
63
,
67
,
68
]. EVs are a heterogeneous
group of membrane-bound vesicles released from cells by invagination and budding. They
facilitate cell-to-cell interactions via contact with neighboring cells or internalization by
recipient cells, which includes fusion with membrane and endocytosis [
69
]. According
to the biogenesis, biophysical properties, and function, EVs can be classified into three
main subtypes, namely exosomes (30–150 nm), microvesicles (MVs) (50–1000 nm), and
apoptotic blebs (1000–5000 nm) [
70
]. Among these EVs, exosomes and MVs are of particular
importance in cell–cell communication [
70
]. Both of them contain lipids, proteins, and
genetic material, such as DNA, messenger RNA (mRNA), microRNA (miRNA), and long
non-coding RNAs (lncRNA) that can be delivered to and reprogram the recipient cells [
71
].
Studies have shown that MSC/ASC-EVs exert both inhibitory and promoting effects in
several situations and different stages of breast cancer. Through the transfer of various
tumor-related factors, EVs promote proliferation, angiogenesis, metastasis, and drug re-
sistance of malignant tumor cells [
72
,
73
], as shown in breast cancer cells stimulated with
Her2-loaded EVs [
72
]. These data suggest that ASCs/MSCs may secrete molecules that act
in concert with the secretome of breast cancer cells to remodel the microenvironment.
The direct crosstalk between ASCs and breast cancer cells is strictly dependent on
their close contact that is established in the TME. Interestingly, tunneling nanotubes (TNTs)

Cancers 2022,14, 3908 5 of 35
have emerged as a new important means of cell–cell communication. TNTs are thin mem-
brane protrusions that connect cells over long distances allowing the exchange of various
cellular components, including organelles, proteins, calcium ions, viruses, and bacteria [
74
].
Notably, TNTs are able to connect multiple cells forming functional cellular networks [
75
].
TNTs are therefore considered as novel bridges of intercellular communication in physio-
logical and pathological cell processes [
76
]. Interestingly, MSCs have been shown to form
TNTs and transfer mitochondria and other components to target cells [
77
–
79
]. This occurs
under both physiological and pathological conditions, where cells are under stress, leading
to changes in cellular energy metabolism and functions [
76
]. In this context, it is feasible to
hypothesize that the protective role of ASCs/MSCs in breast cancer cell survival may be
partially mediated through the formation of TNTs, in particular, when breast cancer cells
are under stress from chemotherapy or radiotherapy.
Moreover, the activation of several signaling pathways requires direct cell–cell contact
via their membrane-bound ligands and receptors, such as the canonical Notch signal-
ing pathway [
80
]. Notch signaling is linked to the maintenance of breast cancer stem
cells [
81
] and induction of epithelial-to-mesenchymal transition (EMT) resulting in an
increase in migration and invasion of breast cancer cells [
82
]. In fact, direct co-culture
of obese ASCs enhanced Notch signaling in ER
+
breast cancer cells co-responsible for
radiation resistance [83].
Finally, cell–cell fusion, a process that merges the lipid bilayers of two different cells,
plays a crucial role during embryonic development as well as in tissue regeneration [
84
,
85
].
Studies also provide evidence that cell–cell fusion is closely related to cancer development
and metastasis [
86
]. Although this highly regulated process is not yet fully understood,
bone-marrow-derived cells were reported to be able to fuse to cancer cells, and the fused
hybrids acquired more malignant characteristics and enhanced self-renewal ability [
87
].
In line with this observation, MSCs were reported to fuse with diverse malignant cells to
promote proliferation and metastasis, including with lung cancer cells [
88
], liver cancer
cells [
89
], and gastric cancer cells [
90
]. In particular, it was demonstrated that MSCs were
fused with breast cancer cells and promoted their metastatic capacity [
91
]. Recently, is has
been revealed that ASCs are able to fuse spontaneously with breast cancer cells, where
breast cancer stem cell (CSC) markers CD44
+
CD24
−
/
low
EpCAM
+
are enriched in this
fused population [
92
]. These studies suggest cell fusion as a direct interaction between
ASCs/MSCs and cancer cells. Further investigations are needed to explore the molecular
mechanisms by which ASCs/MSCs and malignant cells are able to fuse and how this
process promotes malignancy.
In sum, as illustrated in Figure 1, the crosstalk between ASCs/MSCs and breast cancer
cells is multilateral and majorly mediated by indirect patterns such as the secretion of
soluble bioactive factors and EVs released by ASCs as well as breast cancer cells. Although
observed mainly
in vitro
, the interaction of breast cancer cells with ASCs/MSCs may
be supported by direct cell–cell contacts including the formation of TNTs, binding of
membrane-bound ligands to receptors, and cell–cell fusion. These communications may
reshape the TME and fuel breast cancer progression and therapy resistance.

Cancers 2022,14, 3908 6 of 35
Cancers 2022, 14, 3908 6 of 35
Figure 1. Simplified model representing the crosstalk between ASCs/MSCs and breast cancer cells.
The communication between ASCs/MSCs and breast cancer cells may occur directly via cell–cell
contact, namely TNTs, cell fusion, and the binding of membrane-bound ligands to receptors, or in-
directly via released soluble bioactive factors such as cytokines, chemokines, and growth factors,
and EVs including exosomes and microvesicles. EV, extracellular vesicles; ASCs, adipose tissue-
derived mesenchymal stromal/stem cells; MSCs, mesenchymal stromal/stem cells; mRNA, messen-
ger RNA; miRNA, microRNA; lncRNA, long non-coding RNA; TNTs, tunneling nanotubes.
5. Mutual Interaction between ASCs/MSCs and Breast Cancer Cells
The communication between MSCs/ASCs and breast cancer cells has been an inten-
sive research focus. The related studies are mostly performed using in vitro models to
investigate the effects of ASCs/MSCs or their conditioned medium on proliferation, sur-
vival, migration, and invasion of breast cancer cell lines. Breast cancer cell lines are classi-
fied based on the status of three important cell surface receptors conventionally used for
breast cancer subtyping, ER, PR, and HER2 [93]. Most studies used the following breast
cancer cell lines: low metastatic breast cancer cell line BT474 (ER+, PR+, HER2+), MCF-7
(ER+, PR+, HER2−) and T47D (ER+, PR+, HER2−), metastatic breast cancer cell lines HCC1954
(ER−, PR−, HER2+, with wild type breast cancer gene 1 (BRCA1)), SKBR3 (ER−, PR−, HER2+,
with wild type BRCA1), and MDA-MB-453 (ER−, PR−, HER2+, with wild type BRCA1), and
highly metastatic breast cancer cell lines MDA-MB-231 (triple negative, with wild type
BRCA1), MDA-MB-468 (triple negative, with wild type BRCA1), MDA-MB-436 (triple
negative, mutated BRCA1), and SUM149 (triple negative, mutated BRCA1) [93]. Regard-
ing ASCs/MSCs, while tumor adjacent cells [94] or “cancer-educated” MSCs [95] were re-
cently used, most of the studies employed ASCs/MSCs isolated from non-breast sources,
including abdominal adipose tissue, bone marrow, and peripheral blood [32,58,96]. It is
well-known that ASCs/MSCs from different tissues and organs have distinct tran-
scriptomic, biochemical, and secretory profiles, as well as biologic functions in tissue-spe-
cific homeostasis, immune modulation, and vasculogenesis/angiogenesis [97]. This, to-
gether with other diversities, such as different BMI, varied donor age, variable ASC/MSC
passages, and individual experiment settings, often leads to inconclusive results with
Figure 1.
Simplified model representing the crosstalk between ASCs/MSCs and breast cancer
cells. The communication between ASCs/MSCs and breast cancer cells may occur directly via cell–
cell contact, namely TNTs, cell fusion, and the binding of membrane-bound ligands to receptors,
or indirectly via released soluble bioactive factors such as cytokines, chemokines, and growth
factors, and EVs including exosomes and microvesicles. EV, extracellular vesicles; ASCs, adipose
tissue-derived mesenchymal stromal/stem cells; MSCs, mesenchymal stromal/stem cells; mRNA,
messenger RNA; miRNA, microRNA; lncRNA, long non-coding RNA; TNTs, tunneling nanotubes.
5. Mutual Interaction between ASCs/MSCs and Breast Cancer Cells
The communication between MSCs/ASCs and breast cancer cells has been an in-
tensive research focus. The related studies are mostly performed using
in vitro
models
to investigate the effects of ASCs/MSCs or their conditioned medium on proliferation,
survival, migration, and invasion of breast cancer cell lines. Breast cancer cell lines are
classified based on the status of three important cell surface receptors conventionally used
for breast cancer subtyping, ER, PR, and HER2 [
93
]. Most studies used the following
breast cancer cell lines: low metastatic breast cancer cell line BT474 (ER
+
, PR
+
, HER2
+
),
MCF-7 (ER
+
, PR
+
, HER2
−
) and T47D (ER
+
, PR
+
, HER2
−
), metastatic breast cancer cell
lines HCC1954 (ER
−
, PR
−
, HER2
+
, with wild type breast cancer gene 1 (BRCA1)), SKBR3
(ER
−
, PR
−
, HER2
+
, with wild type BRCA1), and MDA-MB-453 (ER
−
, PR
−
, HER2
+
, with
wild type BRCA1), and highly metastatic breast cancer cell lines MDA-MB-231 (triple
negative, with wild type BRCA1), MDA-MB-468 (triple negative, with wild type BRCA1),
MDA-MB-436 (triple negative, mutated BRCA1), and SUM149 (triple negative, mutated
BRCA1) [
93
]. Regarding ASCs/MSCs, while tumor adjacent cells [
94
] or “cancer-educated”
MSCs [
95
] were recently used, most of the studies employed ASCs/MSCs isolated from
non-breast sources, including abdominal adipose tissue, bone marrow, and peripheral
blood [
32
,
58
,
96
]. It is well-known that ASCs/MSCs from different tissues and organs have
distinct transcriptomic, biochemical, and secretory profiles, as well as biologic functions in
tissue-specific homeostasis, immune modulation, and vasculogenesis/angiogenesis [
97
].
This, together with other diversities, such as different BMI, varied donor age, variable
ASC/MSC passages, and individual experiment settings, often leads to inconclusive results

Cancers 2022,14, 3908 7 of 35
with breast-cancer-supportive and -suppressive functions [
58
,
98
], which may not reflect
the situation in vivo in breast cancer tissue.
5.1. ASCs/MSCs Influence Breast Cancer and Related Molecular Mechanisms
Much attention has been paid to elucidating how ASCs/MSCs impact breast cancer
cells as well as their TME (Table 1). Although their exact roles are not yet completely
understood, ASCs/MSCs are described as both pro- or anti-tumorigenic, depending on
the type and source of ASCs/MSCs, the use of breast cancer cell lines, and the
in vitro
or
in vivo
models. The studies concerning the anti-tumorigenic impact of MSCs are limited.
MSCs have been reported to exert their negative impact on breast cancer by impairing
angiogenesis via secretion of exosomes [
99
], reducing migration and invasion via the release
of tissue inhibitor of metalloproteinase (TIMPs) [
100
], and decreasing breast tumor growth
via down regulation of the STAT3 signaling pathway [
101
]. Nevertheless, the majority of
studies report pro-tumorigenic effects of ASCs/MSCs on breast cancer cells, which are
multilayered, as depicted in Table 1. The related molecular mechanisms are discussed
in detail.
Table 1. Functional alterations of breast cancer cells induced by ASCs/MSCs.
ASC/MSC Source Study Design Functions and Molecular Mechanisms Ref.
Breast cancer promoting effects of ASCs/MSCs derived from human tissues
Human ASCs derived from
visceral and subcutaneous
adipose tissue
MCF7, MDA-MB-231 BC
cell lines and MCF10A
in vitro
Direct co-culture of ASCs promoted proliferation of BC
cells with an upregulation of AURKA,PLK1,BCL6, IL6,
and IL8, whereas indirect co-culture led to EMT of BC
cells via STAT3 and ERK signaling.
[32]
Human ASCs obtained from
ATCC
MCF7 and BT474 BC cell lines
in vitro
Supernatant of ASCs increased BC cell proliferation and
radiotherapy resistance by IGF1 secretion. BC cells
overexpressed IGF1R upon radiotherapy.
[
102
]
Human BM-MSCs
MCF7, T47D, and
SK-Br-3 BC cell lines
in vitro
BM-MSC supernatant increased proliferation of BC cells
independent of IL6 and VEGF, but both signaling
proteins stimulated migration by the activation of
MAPK, AKT, and p38 MAPK.
[
103
]
Human BC-derived MSCs MCF7 BC cell line in vitro
Mammary MSCs increased proliferation and cisplatin
resistance of MCF7 cells by triggering the IL6/STAT3
pathway.
[
104
]
Human MSCs from primary
BC tissue
Co-transplantation BC
xenograft mouse model
of MCF7 and MSCs in vivo
and in vitro
Mammary MSCs promoted BC proliferation and
mammosphere formation via EGF/EGFR/AKT
signaling.
[
105
]
Human ASCs from adipose
tissues
MCF-7, BT-474, T-47D, and
4T1 BC cell lines in vitro
and in vivo
PDGF-D secreted by ASCs stimulated tumor growth
in vivo, mammosphere formation in vitro, and EMT in
BC cells.
[
106
]
Human MSCs from
supraclavicular lymph node
(LN-MSCs) and liver
(Lv-MSCs)
MDA-MB-231, –436, –468,
MCF7 BC cell lines, and
MCF10A cells in vitro
and in vivo
The engulfment of MSCs by BC cells increased the gene
expression of WNT5A,MSR1,ELMO1,IL1RL2,ZPLD1,
and SIRPB1. This further increased BC cell migration,
invasion, and mammosphere formation
in vitro
and the
tumor metastasis in vivo.
[
107
]
Human ASCs from facial or
abdominal liposuction MCF7 BC cell line in vitro
ASCs co-cultured with MCF7 stimulated EMT in BC
cells. The data also suggest that EMT was induced by
the cross-interactions with the TGFβ/Smad and
PI3K/AKT pathways.
[
108
]
Human ASCs isolated from
SAT via bariatric surgery, and
mammary ASCs from
subcutaneous breast
preadipocytes
MCF7 and SUM149 BC cell
lines in vitro, and orthotopic
grafting of 4T1 cells into the
mammary fat pad in vivo
Both ASCs subtypes suppressed the cytotoxicity of
cisplatin and paclitaxel. Depletion of ASCs by D-CAN, a
proapoptotic peptide targeting specific ASCs, reduced
spontaneous BC lung metastases in a mouse allograft
model and a BC xenograft model, when combined with
cisplatin treatment.
[
109
]

Cancers 2022,14, 3908 8 of 35
Table 1. Cont.
ASC/MSC Source Study Design Functions and Molecular Mechanisms Ref.
Human ASCs isolated from
breast adipose tissues of
breast cancer patients and
normal individuals
underwent cosmetic
mammoplasty surgery
Breast tissue and BC tissue
samples in vitro
ASCs isolated from breast cancer patients displayed
elevated levels of IL10 and TGF
β
1, and the supernatant
stimulated the expression of IL4, TGFβ1, IL10, CCR4,
and CD25 in PBLs.
[
110
]
Human ASCs isolated from
breast tumor (T-MSC) and
normal breast adipose tissue
(N-MSC)
Breast tissue and BC tissue
samples
in vitro
, PBLs
in vitro
The TME altered the secretome of T-MSCs with
increased secretion of TGFβ, PGE2, IDO, VEGF, and
lowered secretion of MMP2/9 compared to N-MSCs.
T-MSCs also stimulated the proliferation of PBLs.
[
111
]
Human ASCs isolated from
normal breast adipose tissue
(nASCs) or that of a woman
with breast cancer (cASCs)
Breast tissue and BC tissue
samples in vitro, B cells and
Tregs in vitro
nASCs reduced proliferation of B cells in direct
co-culture, and the TNF
α+
/IL10
+
B cells ratio decreased
in all co-cultures with ASCs, to a barely significantly
higher extent in cASCs. nASCs shifted the cytokine
profile of B cells toward an anti-inflammatory profile.
[
112
]
Human ASCs isolated from
the breast adipose tissue of
reduction mammoplasty
patients with different BMI
MCF7 and SUM159PT BC cell
lines and HMEC breast cell
line in vitro
Supernatant of all analyzed ASCs stimulated
proliferation, migration, and invasion of breast cancer
cells and increased the number of lipid droplets in their
cytoplasm. This was mechanistically associated with the
upregulated expression of the fatty acid receptor CD36,
presenting the capacity of ASCs to induce metabolic
reprogramming via CD36-mediated fatty acid uptake.
[
113
]
Human primary
subcutaneous pre-adipocytes
(pre-hASCs, Lonza)
MCF7, T47D, ZR-75-1,
SK-BR-3 BC cell lines and
murine 3T3-L1 pre-adipocytes
in vitro
Conditioned medium of ASCs stimulated proliferation
and migration of MCF7, T47D, SK-BR-3, and ZR-75-1
cells. Additionally, supernatant of ASCs upregulated the
expression of S100A7 and its knockdown abrogated the
tumorigenic effect of ASCs on the tested breast cancer
cells.
[
114
]
Breast cancer promoting effects of ASCs/MSCs derived from murine tissue
Murine MSCs derived from
spontaneous lymphomas,
mouse bone marrow, and
mouse ears
Syngeneic tumor
transplantation mouse model
in vivo
TNFαdependent monocyte/macrophage recruitment
led to increased tumor volume upon co-injection with
MSCs, associated with CCR2 dependent
immunosuppression of neutrophils, monocytes, and
macrophages.
[
115
]
Murine BM-MSCs and MSCs
isolated from murine lung
cancers
4T1 BC mouse model
in vivo
BM-MSCs and MSCs from lung cancers were able to
recruit CXCR2
+
neutrophils into the tumor by TNF
α
via
activation of CXCL1, CXCL2, and CXCL5 and promoted
tumor metastasis.
[
116
]
Murine BM-MSCs
Murine mammary cancer
cell lines PyMT-Luc,
17LC3-Luc and LLC in vitro
Secretion of CXCL5 by BM-MSCs increased, but without
significance, while proliferation of murine BC cell lines
was unchanged, whereas CXCL1 and CXCL5 promoted
BC cell migration.
[
117
]
Murine and human BM-MSCs
4T1 BC mouse model
in vivo and in vitro
Both types of BM-MSCs stimulated 4T1 BC cell
proliferation in vivo and in vitro upon direct cell–cell
contact. BM-MSCs also promoted vessel formation of
HUVECs in vitro and in vivo in DU145 tumors via
TGFβ, VEGF, and IL6 release.
[
118
]
Murine ASCs isolated from
abdominal cavity
4T1 BC mouse cell line
in vitro and CT26 murine
colon cancer cell line
in vitro
Co-culture of ASCs induced stemcellrelated genes in
cancer cells such as SOX2,NANOG,ALDH1, and
ABCG2. ASCs accelerated tumor growth. Secretion of
IL6 regulated stemcellrelated genes and activated
JAK2/STAT3 in murine cancer cells.
[
119
]
Breast cancer promoting effects of obese ASCs/MSCs
Human ASCs isolated from
breast cancer tissue of lean
and obese patients
Human BC patient-derived
xenograft cells in vivo
Adipsin secreted by obese ASCs stimulated factor B and
C3a, which induced BC proliferation and expression of
CSC genes CD44,CXCR4,SNAI2,SNAI1,ZEB1, and
BMI1.
[
120
]

Cancers 2022,14, 3908 9 of 35
Table 1. Cont.
ASC/MSC Source Study Design Functions and Molecular Mechanisms Ref.
Human lean and obese ASCs
isolated from abdominal lipo-
aspirates of subcutaneous
adipose tissue
MCF7, ZR75, or T47D BC
cell lines in vitro and
MCF7 xenograft mouse model
in vivo
Leptin secreted from obese ASCs enhanced BC
proliferation and increased the expression of EMT and
metastasis-related genes such as Serpine1,MMP2, and
IL6.
[
121
]
Human lean (ln) and obese
(ob) ASCs from abdominal
lipo-
aspirates of subcutaneous
adipose tissue
MCF7 and MDA-MB-231
BC cell lines in vitro
Increased proliferation of BC cells by leptin expression
via estrogen stimulation and increased protein levels of
CDKN2A, GSTP1, PGR, and ESR1 in BC cells
co-cultured with ob-ASCs.
[
122
]
Human and murine ASCs
isolated from
lean and obese individuals
Tumor and stromal cell
transplantation in a mammary
mouse xenograft
model in vivo and MCF7
BC cell line in vitro
Obese ASCs secreted higher levels of IGF1, promoting
tumor growth and metastasis, which could be partially
ameliorated by weight loss.
[
123
]
Human lean and obese ASCs
from abdominal lipoaspirates
of subcutaneous adipose
tissue
BT20, MDA-MB-231,
MDA-MB-468, MCF7, and
HCC1806 BC cell lines
in vitro
and patient-derived xenograft
mouse model
Obesity increased the tumorigenic capacity of ASCs
indicated by increased EMT genes Serpine1,SNAI2,
and TWIST1. This effect was likely mediated via
leptin, since its knockdown led to reduced
pro-metastatic effects of obese ASCs.
[
124
]
Human ASCs isolated from
lipoaspirate of subcutaneous
adipose tissue from lean and
obese patients.
MCF7, T47D, and ZR-75 BC
cell lines in vitro
Obese ASCs induced a cancer-stem-like phenotype in
BC cells with elevated gene expression of Notch1,Notch3,
DLL1, and JAG2. This led to radioresistance
and reduced oxidative stress after radiation in
co-cultured BC cells mediated by leptin.
[83]
Human lean and obese ASCs
derived from mammary
adipose tissue
MDA-MB231 BC cell line
and MCF10AT1 in vitro
Obese ASCs activated BC cell migration more effectively
compared to lean ASCs by direct co-culture. Obese
ASCs had an increased potential for ECM remodeling.
[
125
]
Human lean and obese ASCs
from abdominal lipoaspirates
of subcutaneous adipose
tissue
MCF7 BC cell line in vitro
The known CAF markers NG2, ACTA2, VEGF, FAP, and
FSP were elevated in obese ASCs. Obese ASCs were
more potent in inducing the gene expression of
pro-tumorigenic factors in BC cells including Serpin1,
CCL5, TARC (CCL17), IL24, IL6, IGFBP3, adiponectin, and
leptin.
[
126
]
Human lean and obese ASCs
isolated from elective
liposuction
MCF7 BC cell line
in vitro
and
patient-derived mammary
xenograft (PDX)
mouse model in vivo
The increased tumor growth rate observed in
obese-ASCs-enriched PDX tumors was leptin
dependent. The increased metastatic capacity was leptin
independent and was associated with increased gene
expression of Serpine1 and ABCB1 in tumor cells.
[
127
]
Abbreviations: ASCs, adiposetissue-derived mesenchymal stromal/stem cells; MSCs, mesenchymal stromal/stem
cells; BM-MSCs, bonemarrow-derived mesenchymal stromal/stem cells; IL6, interleukin 6; EMT, epithelial-
to-mesenchymal transition; BC, breast cancer; STAT3, signal transducer and activator of transcription 3; ERK,
extracellular-signal regulated kinase; IGF1, insulin-like growth factor 1; IGF1R, insulin-like growth factor 1
receptor; VEGF, vascular endothelial growth factor; MAPK, mitogen-activated protein kinase; AKT, protein kinase
B; EGF, epithelial growth factor; EGFR, epithelial growth factor receptor; PDGF-D, platelet-derived growth factor
D; WNT5A, wingless/integrated 5a; MSR1, macrophage scavenger receptor types I; ELMO1, engulfment and
cell motility protein 1; IL1RL2, interleukin 1 receptor like 2; AURKA, Aurora kinase A; PLK1, Polo-like kinase
1; BCL6, B-cell lymphoma 6; SAT, subcutaneous adipose tissue; CDKN2A, cyclin-dependent kinase inhibitor
2A; GSTP1, glutathione S-transferase P; ABCB1, ATP-binding cassette subfamily B member 1; ZPLD1, zona
pellucida-like domain-containing 1; SIRPB1, signal-regulatory protein beta-1; TGF
β
, transforming growth factor
β
; Smad, suppressor of mothers against decapentaplegic family member; PI3K, phosphoinositide 3-kinase; CCR4,
C-C motif chemokine receptor 4; PBL, peripheral blood lymphocytes; MMP, matrix metalloprotease; PGE2,
prostaglandin E2; IDO, indoleamine 2,3-dioxygenase; TNF
α
, tumor necrosis factor
α
; CXCL1, C-X-C motif
chemokine ligand 1; HUVEC, human umbilical vein endothelial cell; CD44, cluster of differentiation 44; SNAI,
snail family transcriptional repressor; ZEB1, zinc finger E-box binding homebox 1; BMI, body mass index; PGR,
progesterone receptor; ESR1, estrogen receptor 1; ob, obese; ln, lean; DLL1, delta-like canonical Botch ligand 1;
JAG2, jagged canonical Notch ligand 2; IGFBP3, insulin-like growth factor binding protein 3; JAK2, Janus kinase.
5.1.1. Promoting Proliferation and Survival
ASCs/MSCs secrete various growth factors including IGF1 and EGF, and numerous
cytokines such as leptin, IL6, adipsin, and TNF
α
, which stimulate proliferation and sur-

Cancers 2022,14, 3908 10 of 35
vival of breast cancer cells [
67
,
102
,
103
,
116
,
120
,
121
]. In particular, obesity is associated with
an increase of IL6 in the circulation, reinforcing systemic inflammation [
128
]. Interest-
ingly, increased IL6 was correlated with poor prognosis, progression, and migration of
ER-positive breast cancer [
129
]. IL6 was shown
in vitro
to activate the signal transducer
and activator of transcription 3 (STAT3)/protein kinase B (AKT)/mitogen-activated protein
kinase (MAPK) pathways, triggering proliferation of both triple negative and triple positive
breast cancer cell lines [
103
,
104
,
130
]. TNF
α
, another important inflammatory cytokine
released by adipose stromal cells, including ASCs/MSCs, was increased in the TME of pa-
tients with obesity, causing adipose tissue inflammation and inhibiting apoptosis of TNBC
cells [
131
,
132
]. This elevated level of TNF
α
in individuals with obesity might establish a
positive feedback loop, since TNF
α
was shown to activate ASCs/MSCs and stimulate their
secretion of multiple cytokines, such as chemokine (C-C motif) ligand 5 (CCL5), CXCL1,
CXCL2, and CXCL5, which significantly enhanced tumor growth and metastasis [
115
,
116
].
Morbid obesity, defined as BMI equal to or greater than 40 [
133
], is associated with hy-
perleptinemia and increased leptin impairs the negative feedback mechanism between
the adipose tissue and neurons in the hypothalamus [
134
,
135
]. In support of this notion,
ASCs isolated from obese individuals secreted significantly higher levels of leptin that
stimulated the proliferation of low and high malignant breast cancer cells [
121
,
122
]. This is
linked to leptin receptor activation, which triggers multiple pathways, such as Janus kinase
(JAK) and MAPK, with the expression of downstream target genes involved in cell cycle
progression and proliferation, including cyclin D1 (CCND1),VEGF, and proto-oncogene
C-Fos (FOS), transcription factor AP-1 subunit jun (JUN), and transcription factor AP-1
subunit JunB (JUNB) [
136
]. Adipsin is another adipokine upregulated in ASCs derived
from obese patients, which stimulates the cell surface receptor complement C3a receptor
1 (C3aR) and the cleavage of factor B, leading to proliferation of breast cancer cells [
120
].
Moreover, obesity is associated with increased levels of circulating IGF1, also secreted
by ASCs/MSCs [
137
]. Breast cancer cells express IGF1 receptors, and binding of IGF1
activated phosphoinositide 3-kinase (PI3K) and MAPK pathways, promoting cancer cell
proliferation [
138
–
140
]. Similarly, serum levels of hepatocyte growth factor (HGF) were
elevated, which is secreted by stromal cells, including ASCs, during obesity [
123
], and its
receptor, tyrosine-protein kinase Met (c-Met), is expressed on breast cancer cells [
126
]. Thus,
increased expression of HGF promoted c-Met-induced cell proliferation and subsequent
progression of breast cancer [
141
,
142
]. In addition, ASCs/MSCs are capable of modulating
the metabolism of breast cancer cells by stimulating, for example, the upregulation of
cluster of differentiation 36 (CD36), a fatty acid receptor, leading to an increased prolifer-
ation rate [
113
], or the upregulation of S100 calcium-binding protein A7, involved in cell
cycle regulation [
114
]. Moreover, numerous studies demonstrate that cancer cells are able
to educate ASCs/MSCs, resulting in an altered gene and protein expression [
143
]. The
interaction with cancer cells increased the release of tumor-promoting cytokines such as
CCLs, CXCLs, SDF, and EGF [
105
,
117
]; angiogenesis factors including VEGF, angiopoietins,
EGF, galectin-1, IGF1, and keratinocyte growth factor (KGF) [
102
,
115
,
118
,
144
]; and EMT
inducers such as TGF
β
, platelet-derived growth factor D (PDGF-D), and stem cell factor
(SCF) [
106
,
118
,
145
]. These data strongly suggest that ASCs/MSCs promote proliferation
and survival of breast cancer cells by releasing diverse bioactive factors activating various
signaling pathways.
5.1.2. Stimulating Tumor Angiogenesis
ASCs/MSCs are regarded as an important cell type influencing vascular repair mecha-
nisms [
146
] and as inducers of neovascularization [
147
]. The proposed models by which
ASCs/MSCs facilitate these functions are diverse, including direct cell–cell contact [
148
],
differentiation into endothelial cells (ECs) [
147
], and paracrine signaling [
149
]. The molecu-
lar mechanisms depend on the release of angiogenic factors such as angiopoietin 1 (Ang1),
Ang2, VEGF, TGF
β
, SCF, and von-Willebrand factor (vWF); lipids including fatty acids,
phospholipids, ceramide, and sphingolipids; microRNAs such as miR-181b-5p, miR-494,

Cancers 2022,14, 3908 11 of 35
miR-125a, and miR-210; and signaling molecules including wingless/integrated 3a (Wnt3a),
Wnt4, and matrix metalloprotease (MMP) inducer in ECM [
146
,
147
,
149
]. Obesity influences
the direct cell–cell interaction, and the paracrine signaling and differentiation ability of
ASCs/MSCs [
10
,
50
,
150
]. Consistently, it was shown that obese ASCs/MSCs had deficient
angiogenic properties [
151
,
152
], and were not able to promote VEGF expression and tube
formation of injured human umbilical vein endothelial cells (HUVECs) [
153
]. Moreover,
EVs secreted by obese ASCs lost their capacity to stimulate angiogenesis in endothelial
cells, possibly by a significantly reduced expression of miR-126, leading to an upregulation
of sprouty-related EVH1 domain containing 1 (Spred1) and an inhibition of extracellular-
signal regulated kinase (ERK1/2) essential for endothelial cell angiogenesis [
154
]. A re-
cent high-throughput sequencing analysis presents a more complex picture [
155
]. This
study analyzed secreted EVs from obese and lean ASCs, and 83 miRNAs were found to
be significantly deregulated in obese ASCs, with significant implications in angiogene-
sis [
155
]. Clinical data provide further evidence that obesity is associated with resistance to
anti-VEGF therapies, enlarged tumor size and increased vascularization in breast cancer
patients [
156
–
158
]. This could be explained on several levels, including increased IL6 and
other inflammatory cytokines released by ASCs [
66
], macrophages, and adipocytes, which
were shown to trigger resistance toward anti-VEGF therapy [
156
]. Moreover, an increased
secretion of IL1
β
, which is also significantly elevated in obese ASCs [
159
], was identified
to trigger an NLR family CARD-domain-containing protein 4 (NLRC4)-dependent up-
regulation of angiopoetin-like 4 (ANGPTL4), which is a known angiogenic factor in the
TME of breast cancer [
158
]. Its genetic knockout prevented obesity-induced enhanced
angiogenesis in mice [
158
]. Leptin is described as another major driver in the context of
obesity-induced angiogenesis [
157
], which stimulates the expression of VEGF by activating
the hypoxia-inducible factor 1-alpha HIF1
α
and NFkB pathways [
160
]. Interestingly, inhibi-
tion of leptin signaling also decreased the vascular endothelial growth factor receptor 2
(VEGFR-2) expression levels in endothelial cells and breast cancer cells [161]. Collectively,
ASCs/MSCs display a pro-angiogenic phenotype in the TME through diverse signaling.
5.1.3. Escape of Immune Response
To escape anti-tumor immunity, cancer cells exploit cell-intrinsic pathways associated
with resistance to immune cell-mediated attack and avoid recognition by anti-tumor im-
mune cells [
162
–
164
]. Cancer cells may also enhance immunosuppression of the TME by reg-
ulating the expression or secretion of immunosuppressive molecules, including cytokines
and chemokines. On the one hand, this intercellular communication network effectively
inhibits immune effector cells, including T-cells, natural killer (NK) cells, and dendritic cells
(DCs). On the other hand, it promotes the functions and/or the recruitment of immunosup-
pressive cells such as regulatory T-cells (Tregs) and tumor-associated macrophages [
165
,
166
].
Cancer cell escape from the immune response is mediated mainly by paracrine and au-
tocrine stimulation in the TME by a variety of growth factors and cytokines, including
TGFβ, basic fibroblast growth factor (bFGF), VEGF, PDGF, and ILs [167,168].
ASCs/MSCs in the TME are well-known for their exceptional immunomodulatory
capacity. They have a low expression of major histocompatibility complex (MHC) class I,
and expression of class II MHC molecules is completely absent [
169
], which helps these
cells evade immune recognition. Moreover, ASCs/MSCs are capable of modulating the
immune response by suppressing lymphocytes proliferation, inhibiting differentiation of
monocyte-derived immature DCs, and reducing the cytotoxic activity of NK cells [
170
,
171
].
Their functions are supported both by direct cell–cell interaction and by paracrine signaling
through the release of multiple cytokines and other soluble factors [
171
,
172
]. Intriguingly,
cancer cells were shown to exploit the immunomodulatory capacity of ASCs/MSCs. The
supernatant of ASCs, which were isolated from breast cancer tissue, was reported to
upregulate a panel of anti-inflammatory cytokines, such as IL4, IL10, CCR4, CD25, and
TGF
β
in peripheral blood lymphocytes (PBLs) and to increase the number of Tregs, which
could establish an anti-inflammatory reaction in the TME [
110
]. It was also reported that

Cancers 2022,14, 3908 12 of 35
breastcancer-educated MSCs enhanced the proliferation of PBLs by higher secretion of
TGF
β
, prostaglandin (PGE2), indoleamine 2,3-dioxygenase (IDO), and VEGF [
111
]. In
line with these results, the co-culture of cancer associated ASCs with T-cells expanded the
CD25
+
FOXP3
+
CD73
+
CD39
+
Treg population and increased the release of immune suppres-
sive cytokines IL10, IL17, and TGF
β
[
173
]. In addition, indirect co-culture of ASCs with
activated PBLs reduced the number of killer cell lectin-like receptor K1 (NKG2D
+
) and
CD69
+
NK cells [
57
]. Interestingly, reduced proliferation of B-cells and TNF
α+
/IL10
+
cells
was observed only in direct co-culture but not in indirect co-culture experiments [
112
], sug-
gesting the importance of direct cell–cell contact. These data demonstrate that ASCs/MSCs
greatly contribute to the immune escape by affecting the proliferation and function of
diverse immune cells such as PBLs and T- and B-cells in the TME.
Obesity is a key factor influencing the immunomodulatory capacity of ASCs/MSCs,
mainly by altering the cytokine secretion profile with a loss of anti-oxidant molecules such
as glutamate-cysteine ligase (GCL), peroxiredoxin-5 (Prdx5) and Prdx6, as well as a loss of
functions of the tissue development regulators including Ang, Angptl4, follistatin-related
protein 3 (Fstl3), and placental growth factor (PLGF) [
174
]. In contrast, obesity triggers the
secretion of cytokines involved in osteoporosis, negative vessel remodeling, and inflamma-
tion with increased leukemia inhibitory factor (LIF), IL1
β
, CCL2, leptin, interferon gamma
(IFN
γ
), IL6, and TNF
α
[
66
,
121
,
159
,
174
,
175
]. This switch in the secretome of ASCs/MSCs
contributes to adipose tissue inflammation in patients with morbid obesity [
10
], as high-
lighted by multiple investigations [
175
–
178
]. The
in vitro
co-culture of obese ASCs with
mononuclear cells enhanced monocyte and Th17 activation [
178
] and programmed cell
death ligand 1 (PD-L1) expression, decreased the cytokine secretion in Th1 cells, and
reduced the cytolytic activity in Th1 cells dependent on the elevated IFN
γ
release of
ASCs [
175
]. It was also shown that obese ASCs lost their ability to regulate the polarization
of M1/M2 macrophages
in vitro
and
in vivo
[
176
], which was associated with a four-fold
higher concentration of TNF
α
in the supernatant of obese ASCs [
176
]. Accordingly, it
was reported that co-culture with obese ASCs promoted a pro-inflammatory phenotype
in murine macrophages and microglial cells through increased expression of genes in-
volved in inflammation, altered nitric oxide activity, and impaired phagocytosis [
177
].
Strikingly, Benaige et al. found that obese ASCs induced a different phenotypic switch
in macrophages with pro- and anti-inflammatory features, leading to a tumor-associated
macrophage phenotype [
179
]. These authors also reported that co-cultured macrophages
secreted survivin, stimulating the progression of cancer cells [
179
]. Beyond ASCs/MSCs’
secretome, ASC-orchestrated ECM regulation is crucial in restricting access of immune
cells to cancer, by generating a physical barrier to tumor infiltration, inhibition of cytotoxic
response, and the drug diffusion [
180
,
181
]. In conclusion, the immunosuppressive features
of cancer-educated ASCs/MSCs, particularly obese ASCs/MSCs, may greatly contribute to
the immune evasion of breast cancer cells.
5.1.4. Inducing EMT, Migration and Invasion
EMT is the trans-differentiation process that causes epithelial cells to lose their epithe-
lial characteristics, such as cell junctions and apical-basal polarity, and acquire mesenchy-
mal features, promoting cell motility and invasion [
182
]. ASCs/MSCs release cytokines
and growth factors such as TGF
β
, EGF, PDGF, and HGF, which trigger EMT [
183
,
184
].
The main signaling pathways that induce EMT include the TGF
β
, Wnt/
β
-catenin, and
Notch pathways [
185
]. All these pathways converge on the expression and activation of
the transcriptional factors such as Snail, Slug, Twist-related protein (TWIST), forkhead
box C1 (FOXC1), FOXC2, and zinc finger E-box binding homebox 1/2 (ZEB1/2). These
transcription factors suppress the expression of adherens junction and integrin proteins,
which causes tumor cells to lose their polarity and dissociate from adjacent cells and the
basal membrane [
186
,
187
]. While Snail, Slug, and ZEB2 are able to directly repress the
E-cadherin promoter, TWIST1, FOXC2, and ZEB1 possess an indirect molecular mecha-
nism, which disrupts cell polarity and gives rise to the mesenchymal phenotype [
186
,
188
].

Cancers 2022,14, 3908 13 of 35
In particular, TWIST1 is able to promote transformation of normal mammary epithelial
cells into mesenchymal-like cells with increased expression of vimentin, N-cadherin, and
fibronectin [
189
]. Moreover, ZEB1 also plays an important role in EMT regulation in
breast cancer cells [
190
], dramatically increasing the metastatic rate, plasticity, and therapy
resistance of breast cancer [191].
MSCs/ASCs are potent in promoting the EMT process of breast cancer cells and foster
their migration and invasion through various pathways. S1007A is a protein that has been
shown to be strongly upregulated in breast cancer cells treated with conditioned medium
from ASCs, and this upregulation was associated with markedly increased migration [
114
],
possibly by inducing EMT as reported in cervical cancer cells [
192
]. MSCs/ASCs are capable
of reshaping the TME by secreting lots of MMPs, including MMP1, MMP2, MMP3, MMP8,
MMP9, MMP10, and MMP13 [
193
–
195
], which degrade the tumor-associated ECM. The
proteolytic degradation of ECM generates bioactive matrikines and releases matrix-bound
VEGF, supporting the growth, migration, and metastasis of cancer cells [
196
]. The engulf-
ment of MSCs by breast cancer cells is another cellular mechanism by which cancer cells
promote their migration, invasion, metastasis, and self-renewal capacity [
107
]. The process
increased the gene expression of oncogenic factors such as cellular tumor antigen p53 (p53),
WNT5A, Myc proto-oncogene protein c (c-MYC), TGF
β
,and cell-membrane-associated
genes including macrophage scavenger receptor types I and II (MSR1), engulfment and
cell motility protein 1 (ELMO1), interleukin 1 receptor-like 2 (IL1RL2), zona pellucida-like
domain-containing protein 1 (ZPLD1), and signal-regulatory protein beta-1 (SIRPB1) [
107
].
In fact, low expression of WNT5A and MSR1 was linked to reduced metastasis and longer
cancer-free survival of breast cancer patients [
107
,
197
]. In addition, it was reported that
breast cancer cells treated with ASC supernatant upregulated their fatty acid receptor CD36,
which is associated with migration and invasion [113].
Interestingly, various reports showed a reinforced effect of obese ASCs/MSCs on
the malignancy of breast cancers. This could be attributed to several aspects. First, this
enhanced effect could be induced by an increased secretion of cytokines, as shown for
IGF1 [
123
], which stimulated the invasiveness of breast cancer cells by activating its down-
stream targets ERK, serine/threonine-protein kinase mTOR, and STAT3 [
137
], and for
leptin, with the expression and activation of its various effector genes [
83
,
124
]. Second,
obese ASCs were described to induce the invasion of breast cancer cells more efficiently
compared to lean ASCs by direct cell–cell contact, helping the generation of traction forces
within the TME and releasing various MMPs [
125
]. Third, obesity is associated with an
enhanced de-differentiation of ASCs into CAFs, which changes significant parts of their
secretome toward a cancer-supporting phenotype [
126
,
198
]. The changed secretome might
explain the changes observed in constitutively active ER
+
breast cancer cell lines upon
co-culture with obese ASCs [
127
]. In conclusion, ASCs/MSCs have the ability to support
breast cancer cell migration through multiple pathways, in particular through promoting
EMT and reshaping the ECM, which is abused by cancer cells and fueled by morbid obesity
on various molecular levels to increase cancer migration, invasion, and metastasis.
5.1.5. Raising Cancer-Associated Stem Cells
The term cancer stem cell (CSC) characterizes a subpopulation of cancer cells with an
intrinsic self-renewal and tumorigenic capacity, mirrored by their significant role in tumor
development, therapy resistance, relapse, and metastasis [
199
]. The pathways responsible
for establishing a CSC phenotype are diverse and differ among cancer entities [
200
]. In
breast cancer cells, the most important pathways for this process are STAT, Hedgehog,
protein kinase C (PKC), MAPK, Notch, and Hippo, with hundreds of downstream genes re-
sponsible for enhanced stemness [
201
]. Obesity increases systemic levels as well as cellular
secretion of many cytokines and adipokines including leptin, IL6, TNF
α
, IGF1, fatty acid
binding protein 4 (FABP4), and resistin, which are all involved in regulating the pathways
associated with the development of CSCs in breast cancer tissue [
202
]. In accordance with
this, Sabol et al. showed that patient-derived xenograft (PDX) tumors co-cultured with

Cancers 2022,14, 3908 14 of 35
obese ASCs increased the formation of metastases and the number of CD44
+
CD24
−
breast
cancer stem cells in a severely immunodeficient (SCID) mouse model [
124
]. Remarkably,
the stable knockdown of leptin in obese ASCs led to a significant reduction in circulating
CSCs [
124
], suggesting leptin as a key factor to induce CSCs by obese ASCs. Furthermore,
leptin was reported to stimulate the secretion of TGF
β
, which leads to the activation of
SMAD family member 2 (Smad2), Smad3, and Smad4 transcription factors, the repression
of cadherin 1 (CDH1) coding for E-cadherin, and an increased CSC phenotype in breast can-
cer cells [
203
]. Moreover, leptin-induced TGF
β
was shown to trigger de-differentiation of
stromal cells in the TME, including fibroblasts, ASCs, and MSCs, toward a cancer-associated
phenotype [
204
], which altered their cytokine secretion pattern and in turn increased TGF
β
secretion [
205
]. Resistin, another adipokine with an increased secretion in obese adipose
tissue [
206
], was highly associated with the transcription of pluripotency genes such as
aldehyde dehydrogenase 1 family member A1 (ALDH1A1), ITGA4, protein lin-28 homolog
B (LIN28B), smoothened homolog (SMO), and sirtuin 1 (SIRT1) in low malignant breast
cancer cells and non-carcinogenic breast epithelial cells [
207
]. Finally, the obese adipose
tissue is characterized by systemic and local chronic inflammation with a highly increased
level of circulated IL6 [
208
], which is also secreted by ASCs in obese adipose tissue [
66
].
This elevated IL6 level was associated with the activation of the JAK2/STAT3 signaling
cascade and increased levels of SRY-Box transcription factor 2 (SOX2), Nanog homeobox
(Nanog), ALDH1A1, and ATP-binding cassette subfamily G member 2 (ABCG2) genes in
breast cancer cells
in vitro
and
in vivo
[
119
], which was completely prevented by blocking
the cellular IL6 signaling [
119
]. These data strongly suggest that ASCs/MSCs have the
potential to stimulate breast cancer stemness, which is significantly enhanced by factors
associated with obesity.
5.1.6. Facilitating Therapy Resistance
ASCs/MSCs reshape the TME, promote EMT, and support the generation of CSCs,
which are associated with radio- and chemotherapy resistance [
199
,
209
]. The co-culture
of ER
+
breast cancer cells with obese ASCs activated many pathways such as leptin, IL6,
Notch, and jagged canonical Notch ligand 2 (JAG2), which mediated radiation resistance
in ER
+
breast cancer cells [
83
]. Blocking either leptin or IL6 from the culture medium
prevented this radiotherapy resistance [
83
]. Beside the secretion of cytokines, the direct
co-culture of breast cancer cells with ASCs/MSCs activated TGF
β
/Smad, PI3K/AKT, and
MAPK signaling, leading to the induction of chemotherapy resistance after 72 h even in
a time frame before EMT occurred [
32
,
108
]. Interestingly, an
in vivo
experiment showed
that specific depletion of ASCs within the TME by selective peptide targeting D-CAN,
consisting of ASC binding domain and a pro-apoptotic domain, resulted in decreased cis-
platin and paclitaxel resistance in a human breast cancer xenograft model [
109
]. In support
of this observation, another study reported that ASC removal increased the efficiency of
cisplatin and suppressed obesity-induced EMT in obese mice with prostate cancer [
210
]. In
addition, ASCs/MSCs were also reported to increase the chemotherapy resistance of other
cancer entities, including colorectal [
211
], ovarian [
212
–
214
], lung [
215
], squamous cell car-
cinoma [
216
], and acute myeloid leukemia [
217
], by modulating PDGF-BB [
213
], X-linked
inhibitor of apoptosis (XIAP) [
218
], Notch [
217
], STAT3 [
101
], Hedgehog [
214
], and p53
signaling [
211
]. These data highlight the general role of ASCs/MSCs in rendering cancer
cells resistant to radio- and chemotherapy by activating diverse signaling, in particular, by
generating EMT and CSCs, and by reshaping the TME.
In sum, ASCs/MSCs, particularly in the obese state, promote the development of
breast cancer by facilitating cell proliferation and survival, EMT, migration and invasion, an-
giogenesis, CSC formation, immune escape, and therapy resistance (Figure 2). While most
reports emphasize supportive effects for breast cancer cells, ASCs/MSCs have also been
shown to have an anti-tumorigenic function [
101
,
219
]. Secreted factors of human MSCs
isolated from umbilical cords were shown to suppress tumor progression and increase
radiosensitivity through downregulating intra-tumoral STAT3 signaling in a xenograft

Cancers 2022,14, 3908 15 of 35
mouse model and in breast cancer cell lines [
101
]. Additionally, ASC supernatant was
reported to be able to induce cell death with increased caspase-3/7 activity, whereas this
was associated with the augmentation of stemness in breast cancer cells [
219
]. Further
studies are needed to clarify the relationship between ASC/MSC and breast cancer cells.
5.2. Breast Cancer Cells Educate ASCs/MSCs and Related Molecular Mechanisms
While ASCs/MSCs influence breast cancer cells, numerous investigations show that
breast cancer as well as its TME “educate” their surrounding cells, including fibroblasts
(FBs) and ASCs/MSCs, toward pro-tumorigenic phenotypes [
220
]. The most precisely char-
acterized cells are FBs. As depicted in Table 2, multiple cancer-associated fibroblast (CAF)
phenotypes have been identified during the last decade as key components of the TME with
implications in tumor growth, therapy resistance, metastasis, ECM remodeling, and im-
mune tolerance [
34
,
205
,
221
]. Interestingly, ASCs/MSCs are morphologically indistinguish-
able from fibroblasts. These two cell types share many common features including their
surface marker composition, proliferation pattern, differentiation capacity, immunomodula-
tion property, and, to some extent, even their gene expression profiles [
222
,
223
]. The major
difference between these two cell types seems to be their methylation profile. While the
general methylation patterns of MCSs are maintained in long-term culture and aging [
224
],
the methylation of fibroblasts seems to decrease with aging or prolonged culture [
225
].
Indeed, ASCs/MSCs have been proposed to be immature FBs and one of the sources
for FBs [222].
Recently, increasing evidence highlights that ASCs/MSCs are educated and
de-differentiated by cancer cells and the TME, fueling malignancy and therapy resis-
tance [
226
,
227
]. Cancer-cell-secreted factors and direct cancer cell–ASC/MSC contacts
induce a pro-tumorigenic population of ASCs/MSCs, named cancer-associated MSCs
(
CA-MSCs
) [
143
]. CA-MSCs have the ability to differentiate into multiple cell lineages,
such as fibroblasts and adipocytes, suggesting that MSCs may play a key role in the
generation of most stromal components of the TME. A number of reports have demon-
strated that CA-MSCs differentiate into CAFs and cancer-associated adipocytes (CAAs)
in the presence of malignant cells [
105
,
228
]. While the exact mechanisms underlying the
de-differentiation of CA-MSC are not yet clear, this switch resulted in a highly secretory
phenotype with increased secretion of bone morphogenetic protein (BMP2), BMP4, and
IL6 [
229
]. In line with this observation, there was evidence suggesting that cancer-released
TGF
β
was able to activate the Smad signaling pathway in MSCs, which drove differ-
entiation into a cancer-associated phenotype [
230
]. In other tumor entities, including
lymphomas [
115
], lung [
215
], and gastric cancer [
231
,
232
], IL6, IL8, IL17, IL23, and TNF
α
secreted by monocytes, macrophages, neutrophils, and non-MSC stromal cells were shown
to be capable of promoting malignant transition of ASCs/MSCs, which was associated
with significantly increased metastatic rates and tumor growth [
115
,
215
,
227
,
231
,
232
]. Other
molecular mechanisms proposed for CAF activation include Notch/Eph-ephrin signaling,
ECM composition in the TME, DNA damage, physiological stress, inflammatory stimuli,
RTK ligands, and TGF
β
-mediated signaling [
34
]. Moreover, the primary cilium, a sensory
organelle with an exceptionally high receptor density [
233
], was shown to play a critical
role in the de-differentiation process of adipose progenitors toward a CAF phenotype
by mediating TGF
β
signaling [
234
]. All these studies suggest that diverse pathways are
responsible for the activation of CAFs, and the TME is likely the major player in triggering
the de-differentiation of ASCs/MSCs into different CAFs, depending on their cellular
context and the tumor entity.

Cancers 2022,14, 3908 16 of 35
Cancers 2022, 14, 3908 15 of 35
In sum, ASCs/MSCs, particularly in the obese state, promote the development of
breast cancer by facilitating cell proliferation and survival, EMT, migration and invasion,
angiogenesis, CSC formation, immune escape, and therapy resistance (Figure 2). While
most reports emphasize supportive effects for breast cancer cells, ASCs/MSCs have also
been shown to have an anti-tumorigenic function [101,219]. Secreted factors of human
MSCs isolated from umbilical cords were shown to suppress tumor progression and in-
crease radiosensitivity through downregulating intra-tumoral STAT3 signaling in a xen-
ograft mouse model and in breast cancer cell lines [101]. Additionally, ASC supernatant
was reported to be able to induce cell death with increased caspase-3/7 activity, whereas
this was associated with the augmentation of stemness in breast cancer cells [219]. Further
studies are needed to clarify the relationship between ASC/MSC and breast cancer cells.
Figure 2. Schematic representation of potential effect of ASCs/MSCs on breast cancer cells and re-
lated molecular mechanisms. ASCs/MSCs may promote breast cancer cell proliferation and sur-
vival, EMT, migration, and invasion; CSC formation; angiogenesis; immune evasion; and therapy
resistance. ASCs, adipose-tissue-derived mesenchymal stromal/stem cells; MSCs, mesenchymal
stromal/stem cells; IL6, interleukin 6; EMT, epithelial-to-mesenchymal transition; STAT3, signal
transducer and activator of transcription 3; ERK, extracellular-signal regulated kinase; IGF1, insulin-
like growth factor 1; VEGF, vascular endothelial growth factor; MAPK, mitogen-activated protein
kinase; AKT, protein kinase B; CCL2, monocyte chemotactic and activating factor; EGF, epithelial
growth factor; PDGF-D, platelet-derived growth factor D; Wnt, wingless/integrated; TGFβ,
Figure 2.
Schematic representation of potential effect of ASCs/MSCs on breast cancer cells and related
molecular mechanisms. ASCs/MSCs may promote breast cancer cell proliferation and survival, EMT,
migration, and invasion; CSC formation; angiogenesis; immune evasion; and therapy resistance.
ASCs, adipose-tissue-derived mesenchymal stromal/stem cells; MSCs, mesenchymal stromal/stem
cells; IL6, interleukin 6; EMT, epithelial-to-mesenchymal transition; STAT3, signal transducer and
activator of transcription 3; ERK, extracellular-signal regulated kinase; IGF1, insulin-like growth factor
1; VEGF, vascular endothelial growth factor; MAPK, mitogen-activated protein kinase; AKT, protein
kinase B; CCL2, monocyte chemotactic and activating factor; EGF, epithelial growth factor; PDGF-D,
platelet-derived growth factor D; Wnt, wingless/integrated; TGF
β
, transforming growth factor
β
;
PI3K, phosphoinositide 3-kinase; CCR4, C-C motif chemokine receptor 4; TNF
α
, tumor necrosis
factor α; CD25, cluster of differentiation 25; SNAI, snail family transcriptional repressor; ZEB1, zinc
finger E-box binding homebox 1; CAF, cancer-associated fibroblast; CSC, cancer stem cell; JAK2, Janus
kinase 2; FABP4, fatty acid binding protein 4; PKC, protein kinase C; HGF, hepatocyte growth factor;
MMP, matrix metalloprotease; bFGF, basic fibroblast growth factor; vWF, von-Willebrand factor.

Cancers 2022,14, 3908 17 of 35
Table 2. Subtypes of CAFs in different cancer entities.
Fibroblast/CAF Source Study Design Functions and Molecular Mechanisms Ref.
Impact of cancer cells on the fibroblast phenotype
FBs and CAFs isolated from
surgical explantation and
human BM-MSCs obtained
from AOU Meyer Hospital
(Florence)
Co-culture experiments with
FBs, CAFs, and BM-MSCs
with PC3, DU145, and LNCaP
prostate cancer cell lines
in vitro
Prostate cancer cells secreted TGFβ1 and recruited
BM-MSCs into the TME. This in turn led to an elevated
secretion of TGFβ1 in cancer-educated BM-MSCs.
Blocking TGFβ1 reduced the recruitment of BM-MSCs
into the tumor as well as their trans-differentiation.
[
230
]
Pancreatic ductal
adenocarcinoma (PDAC)
tissue
Single-cell RNA sequencing to
characterize CAF
subpopulations in PDAC
The analysis revealed intertumoral heterogeneity
between CAFs, ductal cancer cells, and immune cells in
extremely dense and loose types of PDACs. A highly
metabolic active subtype (meCAFs) was identified.
Patients with abundant meCAFs had a significantly
increased risk for metastasis and poor prognosis. These
patients, however, showed a highly increased response
to immunotherapy.
[
235
]
Murine normal pancreatic and
cancer tissue
Single-cell RNA sequencing to
characterize CAF
subpopulations (normal vs.
pancreatic cancer tissue)
The analysis revealed a landscape of CAFs in pancreatic
cancer during in vivo tumor development. The
LRRC15+CAF lineage was shown to be
TGFβ-dependent and correlated with a poor patient
outcome treated with immunotherapy in multiple solid
tumor entities.
[
236
]
Human and murine PDAC
resection specimens and
normal pancreas tissue
Single-cell RNA sequencing to
characterize CAF
subpopulations (human and
murine)
The analysis from neoplastic and TME of human and
mouse PDAC tumors displayed already described
myCAFs and iCAFs with distinct gene expression
profiles. It further revealed a novel subtype that
expressed MHC class II and CD74 called
“antigen-presenting CAFS (apCAFs)”. These cells
activated antigen-specific CD4+T cells. These
immunomodulatory CAFs were likely associated with a
reduced immune response of PDAC tumors.
[
237
]
Human breast and BC tissue
Single-cell RNA sequencing to
characterize CAF
subpopulations (normal vs.
BC tissue)
The analysis identified different CAF subpopulations in
BC tissue. CAF-S1 (CD29, FAP, α-SMA, PDGFRβ, FSP1,
and CXCL12) was analyzed in detail. These cells
induced an immunosuppressive TME by retaining
CD4+CD25+T cells through the signaling of OX40L,
PD-L2, and JAM2, and increased CD25+FOXP3+T
lymphocytes, and B7H3, DPP4, and CD73 signaling.
[
238
]
Human BC tissue and
metastatic lymph nodes tissue
(LN)
Single-cell RNA sequencing to
characterize CAF
subpopulations (BC and LN
tissue) and co-culture
experiments with MCF7,
MDA-MB-231, and T47D
The analysis identified four CAF subpopulations in LN.
Two had a myCAF gene expression pattern, CAF-S1 and
CAF-S4, accumulated in LN and correlated with cancer
cell invasion. CAF-S1 stimulated cancer cell migration
by stimulating EMT, through CXCL12 and TGFβ
signaling. CAF-S4 induced cancer cell invasion through
Notch signaling. Patients with a high ratio of CAF-S4
cells were prone to develop late distant metastases.
[
239
]
Murine BC tissue and normal
mammary fat pad tissue
Single-cell RNA sequencing to
characterize CAF
subpopulations (BC compared
to pancreatic cancer tissue)
The study identified six CAF subpopulations in a
triple-negative syngeneic breast cancer mouse model.
Among these six subpopulations, myCAFs, iCAFs, and
apCAFs were found to exist in BC cancers and PDAC.
The subtype expressing MHC class II proteins similar to
apCAFs were also found in normal breast/pancreas
tissues, indicating that this specific subtype is not TME
induced. The comparison to a pancreatic tumor model
suggested that similar phenotypes exist in both cancer
entities without a TME-specific subtype.
[
240
]

Cancers 2022,14, 3908 18 of 35
Table 2. Cont.
Fibroblast/CAF Source Study Design Functions and Molecular Mechanisms Ref.
Murine and human BC tissue
and normal mammary fat pad
tissue
Single-cell RNA sequencing to
characterize CAF
subpopulations (murine,
human BC tissue vs. normal
mammary fat pad tissue) and
co-culture experiments with
human MDA-MB-231 as well
as murine 4T1 and EO771.
A negative selection strategy was used to analyze 768
single-cell RNA sequencing transcriptome data of
mesenchymal cells in a BC mouse model. In this
approach, three distinct CAF subpopulations were
defined. These populations were named
“vascular”-CAFs, “matrix”-CAFs and
“development”-CAFs. The found gene signatures were
further verified on the transcriptional and protein levels
in various experimental cancers. Human tumors and
every CAF gene profile were correlated with distinctive
molecular functions.
[
241
]
Normal breast, BC tissue
samples, and metastatic
lymph nodes obtained from
surgery
Comparison of multiple
genome transcriptomic RNA
sequencings
These approaches revealed that most of the described
cancer hallmark signaling pathways were significantly
upregulated in triple-negative breast cancer with a
highly enriched CAF population. BGN, a soluble
secreted protein, was upregulated in CAFs compared to
normal cancer-adjacent fibroblasts (NAFs). The
expression was negatively associated with CD8+T cells
and poor prognostic outcomes.
[
242
]
Human primary bladder
tumor tissues and adjacent
normal mucosae tissues
Single-cell RNA sequencing to
characterize CAF
subpopulations (bladder
cancer tissue vs. normal
mucosae tissue)
iCAFs were identified as poor prognostic marker with
potent pro-proliferation capacities, and their
immunoregulatory function in the TME of bladder
cancer was further deciphered. The LAMP3+dendritic
cell subgroup might be able to recruit regulatory T cells,
which could be a step toward an immunosuppressive
TME.
[
243
]
Abbreviations: FB, fibroblast; BC, breast cancer; CAF, cancer-associated fibroblast; apCAF, antigen presenting
cancer-associated fibroblast; meCAF, metabolic active subtype cancer-associated fibroblast; iCAF, inflammatory
cancer-associated fibroblast; myCAF, myofibroblast cancer-associated fibroblast; BM-MSCs, bone-marrow-derived
mesenchymal stromal/stem cells; PDAC, pancreatic ductal adenocarcinoma; TME, tumor microenvironment;
CD29, cluster of differentiation 29;
α
-SMA, smooth muscle actin; PDGFR
β
, platelet-derived growth factor receptor
beta; IL6, interleukin 6; BGN, biglycan; LAMP3, lysosomal-associated membrane protein 3; FSP1, fibroblast-
specific protein-1; CXCL1, C-X-C motif chemokine ligand 1; JAM2, junctional adhesion molecule 2; DPP4,
dipeptidyl peptidase 4; B7H3/CD273, cluster of differentiation 273.
5.2.1. CAF Subtypes and De-Differentiation of MSCs/ASCs into CAFs
Single-cell RNA sequencing is a tool to classify multiple different subtypes of CAFs,
with a specific gene signature for many tumor entities [
235
,
244
]. In general, the following
three major phenotypes have been described: myCAFs (myofibroblast-like CAFs) with
a high expression of smooth muscle actin (
α
SMA), TGF
β
signaling and the capacity to
remodel the ECM [
235
]; iCAFs (inflammatory CAFs), defined by an increased secretion of
inflammatory cytokines, chemokines, and the complement complex [
236
]; apCAFs (antigen-
presenting), featured as a cell type able to induce T-cell receptor ligation in CD4
+
T cells in
an antigen-dependent manner and express CD74- and MHC-class-II-related genes [237].
Strikingly, a recent study was able to recapitulate the de-differentiation of human
ASCs into similar phenotypes in a pancreatic cancer stromal-rich xenograft model [
245
].
ASCs were shown to de-differentiate into three major subpopulations, myCAF, iCAF, and
apCAF [
245
], demonstrating that ASCs/MSCs are also susceptible to gaining malignant
phenotypes in an
in vivo
mouse model [
245
]. Furthermore, a computational analysis
of single-cell gene expression from pan-cancer biopsies could recapitulate the transition
of ASCs into CAFs expressing COL11A1 [
246
]. This phenotypical switch appears to be
dependent on the interaction between the TME and ASCs.
In vitro
experiments showed
that the direct co-culture of ASCs with human pancreatic cancer cells led to the gene
expression profile of myCAFs, whereas the indirect co-culture with the same cells induced
an iCAF gene expression pattern [
247
]. In line with this observation, the direct co-culture

Cancers 2022,14, 3908 19 of 35
with low malignant breast cancer cells and TNBCs was shown to stimulate the transition
toward a myCAF-like phenotype driven by TGFβ/Smad3 signaling [94].
Interestingly, obesity seems to fuel this de-differentiation process. It was reported that
obese ASCs expressed significantly higher levels of myCAF-associated genes, including
actin alpha 2 (ACTA2), fatty-acid-binding protein 1 (FAP1), fibroblast-specific protein-1
(FSP1), and chondroitin sulfate proteoglycan (NG2), compared to lean control ASCs [
126
].
These cells also displayed an increased secretion of pro-tumorigenic cytokines, such as
TARC (CCL17), CCL5, IL24, and IL6 [
126
]. This could be of clinical relevance, since
breast cancers from obese women have an elevated incidence of desmoplasia, and these
desmoplastic tumors are described as highly fibrillar collagen enriched with an increased
number of CAFs [
40
,
248
], which can be further deciphered into different CAF subtypes
with individual roles inside the TME [
249
,
250
]. In fact, subtypes of cancer associated
ASCs/CAFs have been identified in breast cancer, as demonstrated in Table 3.
Table 3. Subtypes of cancer associated ASCs/CAFs in breast cancer.
Fibroblast/CAF Source Study Design Functions and Molecular Mechanisms Ref.
Human adipose progenitors
(APs) isolated from adipose
tissue and breast-APs (B-APs)
isolated from breast adipose
tissue
MCF-7 and T47D cell lines
in vitro
Primary cilia of APs were required for de-differentiation
of APs into CAFs stimulated by breast cancer cells.
Inhibition of cilia stopped the malignant transition of
APs. Primary cilia mediated TGFβ1 signaling to APs.
[
234
]
Human lean and obese ASCs
from abdominal lipoaspirates
of subcutaneous adipose
tissue
MCF7 cell line in vitro
Co-culture of breast cancer cells with lean and obese
ASCs induced a CAF-like phenotype with elevated gene
expression of NG2, ACTA2, VEGF, FAP, and FSP. This
cancer-educated phenotype was enhanced in obese
ASCs compared to lean counterparts. Obese ASCs were
more potent in inducing the expression of
pro-tumorgenic factors in breast cancer cells including
Serpin1, CCL5, TARC, IL24, IL6, IGFBP3, adiponectin, and
leptin.
[
126
]
Human
adipocytes/pre-adipocytes
isolated from breast cancer
tissue or reduction
mammoplasty
Co-culture with murine
3T3-F442A pre-adipocytes cell
line, murine 4T1 breast cancer
cell line, human breast cancer
cell line SUM159PT in vitro
Co-culture of breast cancer cells with mature adipocytes
or pre-adipocytes led to enhanced secretion of
fibronectin and collagen I. This was associated with
enhanced migration/invasion and the expression of
known CAF marker FSP1. The de-differentiation
process was triggered by the reactivation of the
Wnt/β-catenin pathway in response to Wnt3a.
[
251
]
Human ASCs isolated from
unprocessed subcutaneous
adipose tissue
MDA-MB-231 and MCF7 cell
lines and supernatant,
in vitro
ASCs were de-differentiated in response to supernatant
of breast cancer cells, shown by the expression of
ACTA2, SDF1, CCL5, and tenascin-C, mediated by
TGFβ1/Smad3.
[94]
Immortalized human
AD-MSC cell line ASC52telo
(ATCC)
Capan-1 and MIAPaCa-2
human PDAC cell lines and
stroma-rich cell-derived
xenograft (Sr-CDX) mouse
model in vitro/in vivo
The SR-CDX model resembled the PDAC phenotype
induced by CAFs with accelerated tumor growth,
stromal cell proliferation, chemoresistance, and dense
stroma. Single-cell RNA sequencing revealed that the
CAFs in the TME were derived from the transplanted
AD-MSCs, which de-differentiated into known and
unknown CAF subtypes.
[
245
]
Data sets from multiple
pan-cancer biopsy tissues
Single-cell RNA sequencing
data sets from multiple cancer
biopsies to recapitulate ASC
de-differentiation process
in vitro
This analysis revealed that CAFs originated from a
particular subset of ASCs present in the stroma vascular
fraction of normal adipose tissue. The transition stages
of ASCs were recapitulated toward a cance-associated
phenotype by using a rich pancreatic cancer dataset. At
the endpoint of this transition process, the cells
presented the following upregulated genes: MMP11,
COL11A1, C1QTNF3, CTHRC1, COL12A1, COL10A1,
COL5A2, THBS2, AEBP1, LRRC15, and ITGA11.
[
246
]

Cancers 2022,14, 3908 20 of 35
Table 3. Cont.
Fibroblast/CAF Source Study Design Functions and Molecular Mechanisms Ref.
Immortalized human
AD-MSC cell line ASC52telo
(ATCC)
Capan-1, SUIT-2, and
MIAPaCa-2 human PDAC cell
lines and stroma-rich
cell-derived xenograft
(Sr-CDX) mouse model
in vitro
AD-MSCs acted as precursors for CAFs in vitro.
AD-MSCs could be induced into myCAFS and iCAFs
upon co-culture with PDAC cells. Direct co-culture led
to a myCAF phenotype, whereas indirect co-culture
induced an iCAF gene expression pattern.
[
247
]
Human ASCs (ADSC-GM)
from Lonza
MDA-MB-231 breast cancer
cell line and HUVECs in vitro
EVs from MDA-MB-231 converted ASCs into a
myCAF-like phenotype, with increased VEGF and ECM
remodeling, and partly driven by MAPK signaling.
[
252
]
Abbreviations: AP, adipose progenitors; ASCs, adipose-tissue-derived mesenchymal stromal/stem cells; MSCs,
mesenchymal stromal/stem cells; BM-MSCs, bone-marrow-derived mesenchymal stromal/stem cells; IL6, inter-
leukin 6; CAF, cancer-associated fibroblast; myCAF, myofibroblast cancer-associated fibroblast; EMT, epithelial-to-
mesenchymal transition; VEGF, vascular endothelial growth factor; MAPK, mitogen-activated protein kinase; AKT,
protein kinase B; EGF, epithelial growth factor; BC, breast cancer; C1QTNF3, complement C1q tumor necrosis-
factor-related protein 3; CTHRC1, collagen triple helix repeat-containing protein 1; THBS2, thrombospondin-2;
AEBP1, adipocyte enhancer-binding protein 1; LRRC15, leucine-rich repeat-containing protein 15; ACTA2, actin
alpha 2; MMP, matrix metallopeptidase.
5.2.2. De-Differentiation of MSCs/ASCs into myCAFs Remodeling ECM
The myCAF subpopulation is characterized by an upregulation of genes involved in
smooth muscle contraction, focal adhesion, ECM organization, and collagen formation [
237
].
Additionally, myCAFs are associated with gene expression of ACTA2, transgelin (TAGLN),
myosin light chain 9 (MYL9), tropomyosin 1/2 (TPM1/2), FAP,FSP1, and platelet-derived
growth factor receptor beta (PDGFR
β
) [
253
]. This subpopulation was shown to be essential
for fibrosis in the TME, causing increased density and stiffness [
254
]. This highly fibrotic
ECM decreased T-cell infiltration, and was associated with a hypoxia-induced metabolic
switch, further suppressing the immune response in the TME [
254
]. As a consequence,
myCAFs stimulated tumor proliferation, migration, and invasion [
253
]. In the TME of
breast cancer, myCAFs were shown to promote an immunosuppressive environment by
attracting and retaining CD4
+
CD25
+
T-cells through the ligands tumor necrosis factor
receptor superfamily member 4 (OX40L), PD-L2, and the adhesion molecule junctional
adhesion molecule B (JAM2). Additionally, they were able to increase the number of
CD25
+
FOXP3
+
T-cells through dipeptidyl peptidase 4 (DPP4), CD73, and B7H3 (cluster
of differentiation 276) signaling [
238
]. As a direct effect on breast cancer, these cells were
reported to trigger EMT by activating the CXCL12 and TGF
β
pathways in breast cancer
cells [
239
]. This significantly increased EMT process was associated with enhanced tumor
migration and lymph-node metastasis [
253
]. In addition, a single-cell transcriptomic
analysis comparing tumor-derived FBs and normal tissue-resident FBs revealed that about
79% of CAFs exhibited a myCAF phenotype with high gene levels of
α
SMA in 4T1 murine
breast tumors [
240
]. Another study further defined these
α
SMA-positive cells into matrix
CAFs (mCAFs), with a specific function in matrix remodeling [
241
]. Interestingly, a high
prevalence of mCAFs was observed at the invasive front of cancers and a low abundance
in the cancer core [
241
]. Similar results were shown for ASCs de-differentiated into the
myCAF phenotype. These cells promoted TME fibrosis, desmoplasia and chemoresistance
in a stroma-rich xenograft mouse model [
245
] and enhanced breast cancer cell invasion
in vitro
[
94
]. Given that obesity was reported to stimulate the de-differentiation of ASCs into
an cancer-associated phenotype [
126
], this might explain that obesity fuels the malignant
progression of breast cancer by reshaping its TME.
5.2.3. De-Differentiation of MSCs/ASCs into iCAFs with Secretion of Soluble Factors
and Exosomes
Another important CAF subpopulation is iCAFs with a low
α
SMA expression and
high cytokine production as well as secretion [
242
], found in breast cancer and in pancreas
ductal adenocarcinoma (PDAC) [
240
,
253
]. This extraordinary secretory activity is related to
a high gene expression of important signaling regulators, such as PDGFR
α
, dermatopontin

Cancers 2022,14, 3908 21 of 35
(DPT), C-type lectin domain family 3 member B (CLEC3B), collagen type XIV alpha 1 chain
(COL14A1), lymphocyte antigen 6c1 (Ly6c1), hyaluronan synthase 1 (HAS1), HAS2,IL6,IL8,
IL11,CXCL1,CXCL2, and CCL2 [
240
,
253
]. The data from single-cell RNA sequencing of
PDAC and bladder urothelial carcinoma tissue revealed that the cytokine-cytokine receptor
interaction pathway was significantly enriched in iCAFs [
243
,
255
]. Increased gene levels
of VEGF,FGF,FGF7,IGF1, and IGF2, known for their proliferation-promoting effects in
endothelial, fibroblasts, and cancer cells, were also detected [
243
]. Indeed, the supernatant
of iCAFs isolated from bladder urothelial carcinoma tissue promoted proliferation of
tumor cells [
243
]. These cells were also capable of suppressing the immune response
by interfering with the activity of CD8
+
T-cells, CD4
+
T-cells, Tregs, NK cells, mast cells,
myeloid cells, and neutrophils through the secretion of various cytokines, such as CXCL1,
CXCL12, CXCL16, IL6, IL8, IL11, IL33, LIF, PGE2, PVR cell adhesion molecule (PVR),
podoplanin (PDPN), DPP4, PD1, PD2, and TGF
β
[
243
,
255
]. This immunosuppressive
function could be associated with the poor response to immunotherapy in fibrotic cancers
with a high number of iCAFs [
254
]. Additionally, pharmacologic blockade or depletion
of LIF, a key paracrine factor from iCAFs [
250
], was shown to reduce the progression of
PDAC in a mouse model by modulating cancer cell differentiation and the EMT status [
256
].
Interestingly, the function of iCAFs was not limited to paracrine signaling. These cells
expressed the genes HAS1 and HAS2 responsible for the synthesis of hyaluronan, which
is a major component of the ECM [
237
], and its expression has been shown to correlate
with low immune response and poor prognosis in multiple cancer entities, including breast
cancer [
257
,
258
]. Furthermore, many factors released from iCAFs into the TME facilitate
tumor growth, angiogenesis, and metastasis [
259
], though further
in vivo
and
in vitro
evidence for these functions is needed. Similar to myCAFs, the de-differentiation process of
ASCs toward an iCAF phenotype could be recapitulated in the stroma-rich xenograft mouse
model, which was found to be connected to increased tumor growth [
245
]. In accordance,
a recent study showed that MSCs were able to de-differentiate into inflammatory cancer-
associated cells by activating the IL1
α
/ETS-related transcription factor Elf-3 (Elf3)/yes1-
associated transcriptional regulator (YAP) signaling axis [
260
]. These data strongly suggest
that cancer-cell-educated ASCs/MSCs are capable of undergoing de-differentiation into
iCAFs as well as myCAFs [245,260]; the latter may be driven by morbid obesity [126].
Collectively, these data highlight that breast cancer cells are able to de-differentiate
ASCs/MSCs into different CAF subtypes (Figure 3). Multilateral communication between
ASCs/MSCs, breast cancer cells, and the components of the TME promote breast cancer
progression by activating various signaling pathways via paracrine signaling or direct
cell–cell contact.

Cancers 2022,14, 3908 22 of 35
Cancers 2022, 14, 3908 22 of 35
Figure 3. Simplified model showing that breast cancer cells induce de-differentiation of MSCs/ASCs
into at least two distinct CAF subtypes. The de-differentiation process of MSCs/ASCs in the TME of
breast cancer is triggered by multiple factors including cytokines TGFβ, IL6, IL8, IL17, IL23, and
TNFα, DNA damage, cellular stress, direct cell–cell contact and inflammatory stimuli. This malig-
nant transformation shifts ASCs/MSCs into several cancer-supportive populations including two
typical phenotypes: myCAFs promoting tumor growth, EMT, migration, invasion, and metastasis,
and iCAFs mediating immune evasion, tumor growth, angiogenesis, and metastasis. ASCs, adipose
tissue-derived mesenchymal stromal/stem cells; MSCs, mesenchymal stromal/stem cells; TGFβ,
transforming growth factor β; IL6, interleukin 6; EMT, epithelial-to-mesenchymal transition; TNFα,
tumor necrosis factor α; ECM, extracellular matrix; TME, tumor microenvironment; CD8, cluster of
differentiation 8; myCAF, myofibroblast cancer-associated fibroblast; iCAF, inflammatory cancer-
associated fibroblast; ACTA2, smooth muscle actin, TAGLN, transgelin; MYL9, myosin light chain
9; TPM, tropomyosin; FAP, fibroblast activation protein; FSP1, fibroblast-specific protein-1; PDG-
FRβ, platelet-derived growth factor receptor beta; LIF, leukemia inhibitory factor; CXCL, chemo-
kines C-X-C ligand; HAS, hyaluronan synthase; CCL, monocyte chemotactic and activating factor;
COL14A1, collagen type XIV alpha 1 chain.
Figure 3.
Simplified model showing that breast cancer cells induce de-differentiation of MSCs/ASCs
into at least two distinct CAF subtypes. The de-differentiation process of MSCs/ASCs in the TME
of breast cancer is triggered by multiple factors including cytokines TGF
β
, IL6, IL8, IL17, IL23, and
TNF
α
, DNA damage, cellular stress, direct cell–cell contact and inflammatory stimuli. This malignant
transformation shifts ASCs/MSCs into several cancer-supportive populations including two typical
phenotypes: myCAFs promoting tumor growth, EMT, migration, invasion, and metastasis, and
iCAFs mediating immune evasion, tumor growth, angiogenesis, and metastasis. ASCs, adipose
tissue-derived mesenchymal stromal/stem cells; MSCs, mesenchymal stromal/stem cells; TGF
β
,
transforming growth factor
β
; IL6, interleukin 6; EMT, epithelial-to-mesenchymal transition; TNF
α
,
tumor necrosis factor α; ECM, extracellular matrix; TME, tumor microenvironment; CD8, cluster of

Cancers 2022,14, 3908 23 of 35
differentiation 8; myCAF, myofibroblast cancer-associated fibroblast; iCAF, inflammatory cancer-
associated fibroblast; ACTA2, smooth muscle actin, TAGLN, transgelin; MYL9, myosin light chain 9;
TPM, tropomyosin; FAP, fibroblast activation protein; FSP1, fibroblast-specific protein-1; PDGFR
β
,
platelet-derived growth factor receptor beta; LIF, leukemia inhibitory factor; CXCL, chemokines C-X-
C ligand; HAS, hyaluronan synthase; CCL, monocyte chemotactic and activating factor; COL14A1,
collagen type XIV alpha 1 chain.
6. Clinical Significance
ASCs/MSCs, in particular obese ASCs/MSCs, may contribute significantly to breast
cancer development through several mechanisms, including remodeling the TME (“the soil
for the seed”), promoting EMT, and inducing CSCs that cause clinical complications such
as therapy resistance, cancer relapse, and metastasis. Moreover, mammary ASCs/MSCs
may de-differentiate into CAFs, distribute at the interface between blood vessels and
breast cancer cells, contribute to increased tumor interstitial fluid pressure, and represent a
physical barrier to several drugs [
251
]. In fact, altered ECM induced by ASCs/MSCs/CAFs
may induce tissue stiffness and increased tension, which have been associated with poor
outcome in patients with many solid tumors [
261
]. Importantly, the immunosuppressive
and poorly accessible TME drastically limits the potential of effective therapeutics.
ASCs/MSCs/CAFs may also provide new therapeutic opportunities. Overcoming
immunosuppression of the TME and suppressing the development of CSCs by targeting
cancer-associated ASCs/MSCs/CAFs are of decisive importance for the effective treatment
of breast cancer. Indeed, pre-clinical studies targeting ASCs by a killer peptide D-CAN
showed promising results with a significantly reduced EMT and cancer progression in
prostate cancer mouse models [
210
]. Moreover, a protein inducing apoptosis in CAFs
and angiogenic endothelial cells by targeting a novel site of integrin
α
v
β
3 displayed a
strong reduction in intra-tumoral levels of EGF, IGF1, PDGF, collagen, and angiogenic
vessels in an orthotopic xenograft model [
262
]. Consequently, malignant progression
was highly decreased with reduced cancer cell proliferation, metastasis, tumor growth,
and resistance to chemotherapy [
262
]. Other CAF targeted therapies include compounds
against
α
FAP (myCAFs) [
263
], dasatinib/imatinib (PDGFR inhibitor, myCAFs) [
264
], galu-
nisertib/vactosertib (TGF
β
RI inhibitor, myCAFs) [
265
], and ruxolitinib (JAK signaling
inhibitor, iCAFs) [
249
]. Many of these inhibitors have been already analyzed in completed
clinical trials [
265
]. In addition, ASCs/MSCs might also be an exceptional tool to deliver
chemotherapeutics, demonstrated by a recent study where paclitaxel-loaded ASCs reduced
breast tumor growth [
266
]. These interesting data highlight that targeting cancer-educated
ASCs/MSCs/CAFs in the TME may pave a novel path to effectively combat malignancies,
including breast cancer.
7. Conclusions and Perspectives
Recent data clearly suggest that ASCs/MSCs, in particular obese ASCs/MSCs, play
key roles in remodeling the TME and supporting breast cancer development. Much work
remains. First, further investigations are required to unravel the complex crosstalk between
ASCs/MSCs, breast cancer cells, and other stromal cells. In this context, breast cancer
cells grown in 3D and co-cultured with ASCs/MSCs or other stromal cells will be useful
to represent a more physiological morphology with prevalent cell junctions and polarity,
and to resemble a more physiological phenotype in cell proliferation, gene expression,
and differentiation [
267
]. Second, in-depth analyses are necessary to explore how obesity
remodels the TME, affects the communication between breast cancer cells and ASCs/MSCs,
and potentiates breast cancer cells to educate ASCs/MSCs into CAFs. Third, given the
heterogeneity of ASCs/MSCs, additional work is required to identify and adequately clas-
sify various subpopulations that may have different functions in breast cancer progression,
especially the subpopulation that is able to raise CSCs from breast cancer cells. This will
be crucial to understand how ASCs/MSCs contribute to cancer development and may
lead to the identification of new therapeutic targets or biomarkers as well as the use of

Cancers 2022,14, 3908 24 of 35
ASCs/MSCs as therapeutic tools. In particular, the use of ASCs/MSCs as a. therapeu-
tic tool could benefit from the in-depth characterization of different subtypes, as human
umbilical-cord-derived MSCs have been discussed for their anti-tumorigenic effect [
58
,
101
].
Fourth, the future challenge is to elucidate the detailed molecular mechanisms
in vivo
by
which obese ASCs/MSCs promote tumor growth, induce EMT, facilitate angiogenesis,
raise CSCs, and fuel breast cancer metastasis. Finally, inhibition of the crosstalk between
ASCs/MSCs and breast cancer cells could be an attractive strategy in cancer therapy. As
ASCs/MSCs migrate toward cancer sites, it will also be interesting to develop ASCs/MSCs
as a targeted anticancer therapy through genetic modification or engineering.
Author Contributions:
Conceptualization: J.Y. and F.L. Manuscript writing, original draft: A.R. and
J.Y.; correction and modification: N.-N.K., S.C.H., C.S. and F.L. Table preparation: A.R. and N.-N.K.
Figure preparation: A.R., J.Y. and N.-N.K. All authors have read and agreed to the published version
of the manuscript.
Funding:
The work was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research
Foundation)—project number 413992926 (A.R.); 438690235 (J.Y.).
Conflicts of Interest: The authors declare no conflict of interest.
References
1.
Di Cesare, M.; Bentham, J.; Stevens, G.A.; Zhou, B.; Danaei, G.; Lu, Y.; Bixby, H.; Cowan, M.J.; Riley, L.M.; Hajifathalian, K.; et al.
Trends in adult body-mass index in 200 countries from 1975 to 2014: A pooled analysis of 1698 population-based measurement
studies with 19.2 million participants. Lancet 2016,387, 1377–1396. [CrossRef]
2.
Bixby, H.; Bentham, J.; Zhou, B.; Di Cesare, M.; Paciorek, C.J.; Bennett, J.E.; Taddei, C.; Stevens, G.A.; Rodriguez-Martinez, A.;
Carrillo-Larco, R.M.; et al. Rising rural body-mass index is the main driver of the global obesity epidemic in adults. Nature
2019
,
569, 260–264. [CrossRef]
3.
Wang, L.M.; Zhou, B.; Zhao, Z.P.; Yang, L.; Zhang, M.; Jiang, Y.; Li, Y.C.; Zhou, M.G.; Wang, L.H.; Huang, Z.J.; et al. Body-mass
index and obesity in urban and rural China: Findings from consecutive nationally representative surveys during 2004–18. Lancet
2021,398, 53–63. [CrossRef]
4. Vaisse, C.; Reiter, J.F.; Berbari, N.F. Cilia and Obesity. Cold Spring Harb. Perspect. Biol. 2017,9, a028217. [CrossRef]
5.
Sung, H.; Siegel, R.L.; Torre, L.A.; Pearson-Stuttard, J.; Islami, F.; Fedewa, S.A.; Sauer, A.G.; Shuval, K.; Gapstur, S.M.; Jacobs,
E.J.; et al. Global patterns in excess body weight and the associated cancer burden. CA Cancer J. Clin.
2019
,69, 88–112. [CrossRef]
6.
Malik, V.S.; Willet, W.C.; Hu, F.B. Nearly a decade on—Trends, risk factors and policy implications in global obesity. Nat. Rev.
Endocrinol. 2020,16, 615–616. [CrossRef] [PubMed]
7.
Bhaskaran, K.; Douglas, I.; Forbes, H.; dos-Santos-Silva, I.; Leon, D.A.; Smeeth, L. Body-mass index and risk of 22 specific cancers:
A population-based cohort study of 5.24 million UK adults. Lancet 2014,384, 755–765. [CrossRef]
8.
Donohoe, C.L.; Lysaght, J.; O’Sullivan, J.; Reynolds, J.V. Emerging Concepts Linking Obesity with the Hallmarks of Cancer. Trends
Endocrinol. Metab. 2017,28, 46–62. [CrossRef]
9.
Quail, D.F.; Dannenberg, A.J. The obese adipose tissue microenvironment in cancer development and progression. Nat. Rev.
Endocrinol. 2019,15, 139–154. [CrossRef] [PubMed]
10.
Louwen, F.; Ritter, A.; Kreis, N.N.; Yuan, J. Insight into the development of obesity: Functional alterations of adipose-derived
mesenchymal stem cells. Obes. Rev. 2018,19, 888–904. [CrossRef]
11. Hanahan, D. Hallmarks of Cancer: New Dimensions. Cancer Discov. 2022,12, 31–46. [CrossRef]
12.
Hammerl, D.; Smid, M.; Timmermans, A.M.; Sleijfer, S.; Martens, J.W.M.; Debets, R. Breast cancer genomics and immuno-
oncological markers to guide immune therapies. Semin. Cancer Biol. 2018,52, 178–188. [CrossRef]
13. Foulkes, W.D.; Smith, I.E.; Reis, J.S. Triple-Negative Breast Cancer. N. Engl. J. Med. 2010,363, 1938–1948. [CrossRef] [PubMed]
14.
Zeichner, S.B.; Terawaki, H.; Gogineni, K. A Review of Systemic Treatment in Metastatic Triple-Negative Breast Cancer. Breast
Cancer-Basic 2016,10, 25–36. [CrossRef] [PubMed]
15.
Lin, H.J.; Liu, Y.G.; Lofland, D.; Lin, J.Y. Breast Cancer Tumor Microenvironment and Molecular Aberrations Hijack Tumoricidal
Immunity. Cancers 2022,14, 285. [CrossRef]
16.
Barzaman, K.; Karami, J.; Zarei, Z.; Hosseinzadeh, A.; Kazemi, M.H.; Moradi-Kalbolandi, S.; Safari, E.; Farahmand, L. Breast
cancer: Biology, biomarkers, and treatments. Int. Immunopharmacol. 2020,84, 106535. [CrossRef] [PubMed]
17.
Protani, M.; Coory, M.; Martin, J.H. Effect of obesity on survival of women with breast cancer: Systematic review and meta-analysis.
Breast Cancer Res. Treat. 2010,123, 627–635. [CrossRef]
18.
Chan, D.S.; Norat, T. Obesity and breast cancer: Not only a risk factor of the disease. Curr. Treat. Options Oncol.
2015
,16, 22.
[CrossRef] [PubMed]

Cancers 2022,14, 3908 25 of 35
19.
Ewertz, M.; Jensen, M.B.; Gunnarsdóttir, K.; Højris, I.; Jakobsen, E.H.; Nielsen, D.; Stenbygaard, L.E.; Tange, U.B.; Cold, S. Effect
of obesity on prognosis after early-stage breast cancer. J. Clin. Oncol. Off. J. Am. Soc. Clin. Oncol.
2011
,29, 25–31. [CrossRef]
[PubMed]
20.
Agurs-Collins, T.; Ross, S.A.; Dunn, B.K. The Many Faces of Obesity and Its Influence on Breast Cancer Risk. Front. Oncol.
2019
,
9, 765. [CrossRef]
21.
Garcia-Estevez, L.; Cortes, J.; Perez, S.; Calvo, I.; Gallegos, I.; Gema, B. Obesity and Breast Cancer: A Paradoxical and Controversial
Relationship Influenced by Menopausal Status. Front. Oncol. 2021,11, 705911. [CrossRef] [PubMed]
22.
Renehan, A.G.; Tyson, M.; Egger, M.; Heller, R.F.; Zwahlen, M. Body-mass index and incidence of cancer: A systematic review
and meta-analysis of prospective observational studies. Lancet 2008,371, 569–578. [CrossRef]
23.
Afshin, A.; Forouzanfar, M.H.; Reitsma, M.B.; Sur, P.; Estep, K.; Lee, A.; Marczak, L.; Mokdad, A.H.; Moradi-Lakeh, M.; Naghavi,
M.; et al. Health Effects of Overweight and Obesity in 195 Countries over 25 Years. N. Engl. J. Med.
2017
,377, 13–27. [CrossRef]
[PubMed]
24.
Horn-Ross, P.L.; Canchola, A.J.; Bernstein, L.; Neuhausen, S.L.; Nelson, D.O.; Reynolds, P. Lifetime body size and estrogen-
receptor-positive breast cancer risk in the California Teachers Study cohort. Breast Cancer Res. 2016,18, 132. [CrossRef]
25.
Millikan, R.C.; Newman, B.; Tse, C.K.; Moorman, P.G.; Conway, K.; Smith, L.V.; Labbok, M.H.; Geradts, J.; Bensen, J.T.; Jackson,
S.; et al. Epidemiology of basal-like breast cancer. Breast Cancer Res. Treat. 2008,109, 123–139. [CrossRef]
26.
Harris, H.R.; Willett, W.C.; Terry, K.L.; Michels, K.B. Body Fat Distribution and Risk of Premenopausal Breast Cancer in the
Nurses’ Health Study II. JNCI-J. Natl. Cancer Inst. 2011,103, 273–278. [CrossRef]
27.
Pierobon, M.; Frankenfeld, C.L. Obesity as a risk factor for triple-negative breast cancers: A systematic review and meta-analysis.
Breast Cancer Res. Treat. 2013,137, 307–314. [CrossRef]
28.
Loi, S.; Milne, R.L.; Friedlander, M.L.; McCredie, M.R.; Giles, G.G.; Hopper, J.L.; Phillips, K.A. Obesity and outcomes in
premenopausal and postmenopausal breast cancer. Cancer Epidemiol. Biomark. Prev. 2005,14, 1686–1691. [CrossRef] [PubMed]
29.
Sturtz, L.A.; Deyarmin, B.; van Laar, R.; Yarina, W.; Shriver, C.D.; Ellsworth, R.E. Gene expression differences in adipose tissue
associated with breast tumorigenesis. Adipocyte 2014,3, 107–114. [CrossRef] [PubMed]
30.
Anjanappa, M.; Cardoso, A.; Cheng, L.J.; Mohamad, S.; Gunawan, A.; Rice, S.; Dong, Y.; Li, L.; Sandusky, G.E.; Srour, E.F.; et al.
Individualized Breast Cancer Characterization through Single-Cell Analysis of Tumor and Adjacent Normal Cells. Cancer Res.
2017,77, 2759–2769. [CrossRef] [PubMed]
31.
Balaban, S.; Shearer, R.F.; Lee, L.S.; van Geldermalsen, M.; Schreuder, M.; Shtein, H.C.; Cairns, R.; Thomas, K.C.; Fazakerley, D.J.;
Grewal, T.; et al. Adipocyte lipolysis links obesity to breast cancer growth: Adipocyte-derived fatty acids drive breast cancer cell
proliferation and migration. Cancer Metab. 2017,5, 1. [CrossRef] [PubMed]
32.
Ritter, A.; Friemel, A.; Fornoff, F.; Adjan, M.; Solbach, C.; Yuan, J.; Louwen, F. Characterization of adipose-derived stem cells from
subcutaneous and visceral adipose tissues and their function in breast cancer cells. Oncotarget 2015,6, 34475–34493. [CrossRef]
33.
Lim, B.; Woodward, W.A.; Wang, X.P.; Reuben, J.M.; Ueno, N.T. Inflammatory breast cancer biology: The tumour microenviron-
ment is key. Nat. Rev. Cancer 2018,18, 526. [CrossRef]
34.
Sahai, E.; Astsaturov, I.; Cukierman, E.; DeNardo, D.G.; Egeblad, M.; Evans, R.M.; Fearon, D.; Greten, F.R.; Hingorani, S.R.;
Hunter, T.; et al. A framework for advancing our understanding of cancer-associated fibroblasts. Nat. Rev. Cancer
2020
,20,
174–186. [CrossRef]
35. Hass, R. Role of MSC in the Tumor Microenvironment. Cancers 2020,12, 2107. [CrossRef] [PubMed]
36.
Gutiérrez-Cuevas, J.; Sandoval-Rodriguez, A.; Meza-Rios, A.; Monroy-Ramírez, H.C.; Galicia-Moreno, M.; García-Bañuelos, J.;
Santos, A.; Armendariz-Borunda, J. Molecular Mechanisms of Obesity-Linked Cardiac Dysfunction: An Up-Date on Current
Knowledge. Cells 2021,10, 629. [CrossRef]
37.
Huang, S.; Rutkowsky, J.M.; Snodgrass, R.G.; Ono-Moore, K.D.; Schneider, D.A.; Newman, J.W.; Adams, S.H.; Hwang, D.H.
Saturated fatty acids activate TLR-mediated proinflammatory signaling pathways. J. Lipid Res.
2012
,53, 2002–2013. [CrossRef]
[PubMed]
38.
Howe, L.R.; Subbaramaiah, K.; Hudis, C.A.; Dannenberg, A.J. Molecular pathways: Adipose inflammation as a mediator of
obesity-associated cancer. Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res. 2013,19, 6074–6083. [CrossRef] [PubMed]
39.
Xu, L.; Kitade, H.; Ni, Y.; Ota, T. Roles of Chemokines and Chemokine Receptors in Obesity-Associated Insulin Resistance and
Nonalcoholic Fatty Liver Disease. Biomolecules 2015,5, 1563–1579. [CrossRef]
40.
Seo, B.R.; Bhardwaj, P.; Choi, S.; Gonzalez, J.; Eguiluz, R.C.A.; Wang, K.; Mohanan, S.; Morris, P.G.; Du, B.; Zhou, X.K.; et al.
Obesity-dependent changes in interstitial ECM mechanics promote breast tumorigenesis. Sci. Transl. Med.
2015
,7, 301ra130.
[CrossRef] [PubMed]
41.
Olson, O.C.; Quail, D.F.; Joyce, J.A. Obesity and the tumor microenvironment. Science
2017
,358, 1130–1131. [CrossRef] [PubMed]
42.
Ringel, A.E.; Drijvers, J.M.; Baker, G.J.; Catozzi, A.; García-Cañaveras, J.C.; Gassaway, B.M.; Miller, B.C.; Juneja, V.R.; Nguyen,
T.H.; Joshi, S.; et al. Obesity Shapes Metabolism in the Tumor Microenvironment to Suppress Anti-Tumor Immunity. Cell
2020
,
183, 1848–1866.e26. [CrossRef] [PubMed]
43.
Jin, M.Z.; Jin, W.L. The updated landscape of tumor microenvironment and drug repurposing. Signal Transduct. Target. Ther.
2020,5, 166. [CrossRef] [PubMed]
44.
Zuk, P.A.; Zhu, M.; Mizuno, H.; Huang, J.; Futrell, J.W.; Katz, A.J.; Benhaim, P.; Lorenz, H.P.; Hedrick, M.H. Multilineage cells
from human adipose tissue: Implications for cell-based therapies. Tissue Eng. 2001,7, 211–228. [CrossRef]

Cancers 2022,14, 3908 26 of 35
45.
Galland, S.; Stamenkovic, I. Mesenchymal stromal cells in cancer: A review of their immunomodulatory functions and dual
effects on tumor progression. J. Pathol. 2020,250, 555–572. [CrossRef]
46.
Wagner, W.; Wein, F.; Seckinger, A.; Frankhauser, M.; Wirkner, U.; Krause, U.; Blake, J.; Schwager, C.; Eckstein, V.; Ansorge,
W.; et al. Comparative characteristics of mesenchymal stem cells from human bone marrow, adipose tissue, and umbilical cord
blood. Exp. Hematol. 2005,33, 1402–1416. [CrossRef]
47.
Kern, S.; Eichler, H.; Stoeve, J.; Klüter, H.; Bieback, K. Comparative analysis of mesenchymal stem cells from bone marrow,
umbilical cord blood, or adipose tissue. Stem Cells 2006,24, 1294–1301. [CrossRef]
48.
Ritter, A.; Friemel, A.; Roth, S.; Kreis, N.N.; Hoock, S.C.; Safdar, B.K.; Fischer, K.; Möllmann, C.; Solbach, C.; Louwen, F.; et al.
Subcutaneous and Visceral Adipose-Derived Mesenchymal Stem Cells: Commonality and Diversity. Cells
2019
,8, 1288. [CrossRef]
49.
Strong, A.L.; Burow, M.E.; Gimble, J.M.; Bunnell, B.A. Concise review: The obesity cancer paradigm: Exploration of the
interactions and crosstalk with adipose stem cells. Stem Cells 2015,33, 318–326. [CrossRef]
50.
Ritter, A.; Louwen, F.; Yuan, J. Deficient primary cilia in obese adipose-derived mesenchymal stem cells: Obesity, a secondary
ciliopathy? Obes. Rev. 2018,19, 1317–1328. [CrossRef]
51.
Ritter, A.; Friemel, A.; Kreis, N.N.; Hoock, S.C.; Roth, S.; Kielland-Kaisen, U.; Brüggmann, D.; Solbach, C.; Louwen, F.; Yuan, J.
Primary Cilia Are Dysfunctional in Obese Adipose-Derived Mesenchymal Stem Cells. Stem Cell Rep.
2018
,10, 583–599. [CrossRef]
52.
Li, W.; Xu, H.; Qian, C. c-Kit-Positive Adipose Tissue-Derived Mesenchymal Stem Cells Promote the Growth and Angiogenesis of
Breast Cancer. BioMed Res. Int. 2017,2017, 7407168. [CrossRef]
53.
Bielli, A.; Scioli, M.G.; Gentile, P.; Agostinelli, S.; Tarquini, C.; Cervelli, V.; Orlandi, A. Adult adipose-derived stem cells and breast
cancer: A controversial relationship. Springerplus 2014,3, 345. [CrossRef]
54.
Koellensperger, E.; Bonnert, L.-C.; Zoernig, I.; Marmé, F.; Sandmann, S.; Germann, G.; Gramley, F.; Leimer, U. The impact of
human adipose tissue-derived stem cells on breast cancer cells: Implications for cell-assisted lipotransfers in breast reconstruction.
Stem Cell Res. Ther. 2017,8, 121. [CrossRef]
55.
Wang, Y.; Liu, J.; Jiang, Q.; Deng, J.; Xu, F.; Chen, X.; Cheng, F.; Zhang, Y.; Yao, Y.; Xia, Z.; et al. Human Adipose-Derived
Mesenchymal Stem Cell-Secreted CXCL1 and CXCL8 Facilitate Breast Tumor Growth By Promoting Angiogenesis. Stem Cells
2017,35, 2060–2070. [CrossRef]
56.
Zhao, Y.; Zhang, X.; Zhao, H.; Wang, J.; Zhang, Q. CXCL5 secreted from adipose tissue-derived stem cells promotes cancer cell
proliferation. Oncol. Lett. 2018,15, 1403–1410. [CrossRef]
57.
Bahrami, B.; Hosseini, A.; Talei, A.-R.; Ghaderi, A.; Razmkhah, M. Adipose Derived Stem Cells Exert Immunomodulatory Effects
on Natural Killer Cells in Breast Cancer. Cell J. 2017,19, 137–145.
58.
Lee, H.Y.; Hong, I.S. Double-edged sword of mesenchymal stem cells: Cancer-promoting versus therapeutic potential. Cancer Sci.
2017,108, 1939–1946. [CrossRef]
59.
Krueger, T.E.G.; Thorek, D.L.J.; Denmeade, S.R.; Isaacs, J.T.; Brennen, W.N. Concise Review: Mesenchymal Stem Cell-Based Drug
Delivery: The Good, the Bad, the Ugly, and the Promise. Stem Cells Transl. Med. 2018,7, 651–663. [CrossRef]
60.
Kidd, S.; Spaeth, E.; Dembinski, J.L.; Dietrich, M.; Watson, K.; Klopp, A.; Battula, V.L.; Weil, M.; Andreeff, M.; Marini, F.C. Direct
evidence of mesenchymal stem cell tropism for tumor and wounding microenvironments using
in vivo
bioluminescent imaging.
Stem Cells 2009,27, 2614–2623. [CrossRef]
61.
Karnoub, A.E.; Dash, A.B.; Vo, A.P.; Sullivan, A.; Brooks, M.W.; Bell, G.W.; Richardson, A.L.; Polyak, K.; Tubo, R.; Weinberg, R.A.
Mesenchymal stem cells within tumour stroma promote breast cancer metastasis. Nature
2007
,449, 557–563. [CrossRef] [PubMed]
62.
Yu, P.F.; Huang, Y.; Xu, C.L.; Lin, L.Y.; Han, Y.Y.; Sun, W.H.; Hu, G.H.; Rabson, A.B.; Wang, Y.; Shi, Y.F. Downregulation of CXCL12
in mesenchymal stromal cells by TGFβpromotes breast cancer metastasis. Oncogene 2017,36, 840–849. [CrossRef] [PubMed]
63.
Blache, U.; Horton, E.R.; Xia, T.; Schoof, E.M.; Blicher, L.H.; Schönenberger, A.; Snedeker, J.G.; Martin, I.; Erler, J.T.; Ehrbar, M.
Mesenchymal stromal cell activation by breast cancer secretomes in bioengineered 3D microenvironments. Life Sci. Alliance
2019
,
2, e201900304. [CrossRef]
64.
El-Haibi, C.P.; Karnoub, A.E. Mesenchymal stem cells in the pathogenesis and therapy of breast cancer. J. Mammary Gland Biol.
Neoplasia 2010,15, 399–409. [CrossRef]
65.
Papaccio, F.; Paino, F.; Regad, T.; Papaccio, G.; Desiderio, V.; Tirino, V. Concise Review: Cancer Cells, Cancer Stem Cells, and
Mesenchymal Stem Cells: Influence in Cancer Development. Stem Cells Transl. Med. 2017,6, 2115–2125. [CrossRef]
66.
Ritter, A.; Kreis, N.-N.; Roth, S.; Friemel, A.; Jennewein, L.; Eichbaum, C.; Solbach, C.; Louwen, F.; Yuan, J. Restoration of primary
cilia in obese adipose-derived mesenchymal stem cells by inhibiting Aurora A or extracellular signal-regulated kinase. Stem Cell
Res. Ther. 2019,10, 255. [CrossRef]
67.
Guo, Y.; Zhai, Y.; Wu, L.; Wang, Y.; Wu, P.; Xiong, L. Mesenchymal Stem Cell-Derived Extracellular Vesicles: Pleiotropic Impacts
on Breast Cancer Occurrence, Development, and Therapy. Int. J. Mol. Sci. 2022,23, 2927. [CrossRef]
68.
Dong, M.; Liu, Q.; Xu, Y.; Zhang, Q. Extracellular Vesicles: The Landscape in the Progression, Diagnosis, and Treatment of
Triple-Negative Breast Cancer. Front. Cell Dev. Biol. 2022,10, 842898. [CrossRef]
69.
Abels, E.R.; Breakefield, X.O. Introduction to Extracellular Vesicles: Biogenesis, RNA Cargo Selection, Content, Release, and
Uptake. Cell. Mol. Neurobiol. 2016,36, 301–312. [CrossRef]
70.
Doyle, L.M.; Wang, M.Z. Overview of Extracellular Vesicles, Their Origin, Composition, Purpose, and Methods for Exosome
Isolation and Analysis. Cells 2019,8, 727. [CrossRef]

Cancers 2022,14, 3908 27 of 35
71.
Kletukhina, S.; Neustroeva, O.; James, V.; Rizvanov, A.; Gomzikova, M. Role of Mesenchymal Stem Cell-Derived Extracellular
Vesicles in Epithelial-Mesenchymal Transition. Int. J. Mol. Sci. 2019,20, 4813. [CrossRef]
72.
Ciravolo, V.; Huber, V.; Ghedini, G.C.; Venturelli, E.; Bianchi, F.; Campiglio, M.; Morelli, D.; Villa, A.; Della Mina, P.; Menard,
S.; et al. Potential role of HER2-overexpressing exosomes in countering trastuzumab-based therapy. J. Cell. Physiol.
2012
,227,
658–667. [CrossRef]
73.
Tourneur, L.; Mistou, S.; Schmitt, A.; Chiocchia, G. Adenosine receptors control a new pathway of Fas-associated death domain
protein expression regulation by secretion. J. Biol. Chem. 2008,283, 17929–17938. [CrossRef]
74.
Abounit, S.; Zurzolo, C. Wiring through tunneling nanotubes–from electrical signals to organelle transfer. J. Cell Sci.
2012
,125,
1089–1098. [CrossRef]
75.
Ariazi, J.; Benowitz, A.; De Biasi, V.; Den Boer, M.L.; Cherqui, S.; Cui, H.; Douillet, N.; Eugenin, E.A.; Favre, D.; Goodman, S.; et al.
Tunneling Nanotubes and Gap Junctions-Their Role in Long-Range Intercellular Communication during Development, Health,
and Disease Conditions. Front. Mol. Neurosci. 2017,10, 333. [CrossRef]
76.
Luchetti, F.; Carloni, S.; Nasoni, M.G.; Reiter, R.J.; Balduini, W. Tunneling nanotubes and mesenchymal stem cells: New insights
into the role of melatonin in neuronal recovery. J. Pineal Res. 2022,73, e12800. [CrossRef]
77.
Spees, J.L.; Olson, S.D.; Whitney, M.J.; Prockop, D.J. Mitochondrial transfer between cells can rescue aerobic respiration. Proc.
Natl. Acad. Sci. USA 2006,103, 1283–1288. [CrossRef]
78.
Liu, K.; Ji, K.; Guo, L.; Wu, W.; Lu, H.; Shan, P.; Yan, C. Mesenchymal stem cells rescue injured endothelial cells in an
in vitro
ischemia-reperfusion model via tunneling nanotube like structure-mediated mitochondrial transfer. Microvasc. Res.
2014
,
92, 10–18. [CrossRef]
79.
Islam, M.N.; Das, S.R.; Emin, M.T.; Wei, M.; Sun, L.; Westphalen, K.; Rowlands, D.J.; Quadri, S.K.; Bhattacharya, S.; Bhattacharya,
J. Mitochondrial transfer from bone-marrow-derived stromal cells to pulmonary alveoli protects against acute lung injury. Nat.
Med. 2012,18, 759–765. [CrossRef]
80.
Kopan, R.; Ilagan, M.X. The canonical Notch signaling pathway: Unfolding the activation mechanism. Cell
2009
,137, 216–233.
[CrossRef]
81.
Yuan, X.; Zhang, M.; Wu, H.; Xu, H.; Han, N.; Chu, Q.; Yu, S.; Chen, Y.; Wu, K. Expression of Notch1 Correlates with Breast
Cancer Progression and Prognosis. PLoS ONE 2015,10, e0131689. [CrossRef]
82.
Shao, S.; Zhao, X.; Zhang, X.; Luo, M.; Zuo, X.; Huang, S.; Wang, Y.; Gu, S.; Zhao, X. Notch1 signaling regulates the epithelial-
mesenchymal transition and invasion of breast cancer in a Slug-dependent manner. Mol. Cancer 2015,14, 28. [CrossRef]
83.
Sabol, R.A.; Villela, V.A.; Denys, A.; Freeman, B.T.; Hartono, A.B.; Wise, R.M.; Harrison, M.A.A.; Sandler, M.B.; Hossain, F.; Miele,
L.; et al. Obesity-Altered Adipose Stem Cells Promote Radiation Resistance of Estrogen Receptor Positive Breast Cancer through
Paracrine Signaling. Int. J. Mol. Sci. 2020,21, 2722. [CrossRef] [PubMed]
84.
Aguilar, P.S.; Baylies, M.K.; Fleissner, A.; Helming, L.; Inoue, N.; Podbilewicz, B.; Wang, H.; Wong, M. Genetic basis of cell-cell
fusion mechanisms. Trends Genet. TIG 2013,29, 427–437. [CrossRef] [PubMed]
85.
Alvarez-Dolado, M.; Martínez-Losa, M. Cell fusion and tissue regeneration. Adv. Exp. Med. Biol.
2011
,713, 161–175. [CrossRef]
[PubMed]
86.
Zhou, X.; Merchak, K.; Lee, W.; Grande, J.P.; Cascalho, M.; Platt, J.L. Cell Fusion Connects Oncogenesis with Tumor Evolution.
Am. J. Pathol. 2015,185, 2049–2060. [CrossRef]
87.
Dittmar, T.; Nagler, C.; Niggemann, B.; Zänker, K.S. The dark side of stem cells: Triggering cancer progression by cell fusion. Curr.
Mol. Med. 2013,13, 735–750. [CrossRef] [PubMed]
88.
Xu, M.H.; Gao, X.; Luo, D.; Zhou, X.D.; Xiong, W.; Liu, G.X. EMT and acquisition of stem cell-like properties are involved in
spontaneous formation of tumorigenic hybrids between lung cancer and bone marrow-derived mesenchymal stem cells. PLoS
ONE 2014,9, e87893. [CrossRef] [PubMed]
89.
Li, H.; Feng, Z.; Tsang, T.C.; Tang, T.; Jia, X.; He, X.; Pennington, M.E.; Badowski, M.S.; Liu, A.K.; Chen, D.; et al. Fusion of HepG2
cells with mesenchymal stem cells increases cancer-associated and malignant properties: An
in vivo
metastasis model. Oncol.
Rep. 2014,32, 539–547. [CrossRef]
90.
Xue, J.; Zhu, Y.; Sun, Z.; Ji, R.; Zhang, X.; Xu, W.; Yuan, X.; Zhang, B.; Yan, Y.; Yin, L.; et al. Tumorigenic hybrids between
mesenchymal stem cells and gastric cancer cells enhanced cancer proliferation, migration and stemness. BMC Cancer
2015
,15, 793.
[CrossRef]
91.
Melzer, C.; von der Ohe, J.; Hass, R. Enhanced metastatic capacity of breast cancer cells after interaction and hybrid formation
with mesenchymal stroma/stem cells (MSC). Cell Commun. Signal. 2018,16, 2. [CrossRef]
92.
Chan, Y.W.; So, C.; Yau, K.L.; Chiu, K.C.; Wang, X.; Chan, F.L.; Tsang, S.Y. Adipose-derived stem cells and cancer cells fuse to
generate cancer stem cell-like cells with increased tumorigenicity. J. Cell. Physiol. 2020,235, 6794–6807. [CrossRef] [PubMed]
93.
Dai, X.; Cheng, H.; Bai, Z.; Li, J. Breast Cancer Cell Line Classification and Its Relevance with Breast Tumor Subtyping. J. Cancer
2017,8, 3131–3141. [CrossRef]
94.
Jotzu, C.; Alt, E.; Welte, G.; Li, J.; Hennessy, B.T.; Devarajan, E.; Krishnappa, S.; Pinilla, S.; Droll, L.; Song, Y.H. Adipose tissue
derived stem cells differentiate into carcinoma-associated fibroblast-like cells under the influence of tumor derived factors. Cell.
Oncol. 2011,34, 55–67. [CrossRef] [PubMed]

Cancers 2022,14, 3908 28 of 35
95.
Sai, B.Q.; Dai, Y.F.; Fan, S.Q.; Wang, F.; Wang, L.J.; Li, Z.; Tang, J.Q.; Wang, L.; Zhang, X.N.; Zheng, L.L.; et al. Cancer-educated
mesenchymal stem cells promote the survival of cancer cells at primary and distant metastatic sites via the expansion of bone
marrow-derived-PMN-MDSCs. Cell Death Dis. 2019,10, 941. [CrossRef] [PubMed]
96.
Najar, M.; Fayyad-Kazan, H.; Faour, W.H.; Badran, B.; Journe, F.; Lagneaux, L. Breast cancer cells and bone marrow mesenchymal
stromal cells: A regulated modulation of the breast tumor in the context of immune response. Inflamm. Res.
2017
,66, 129–139.
[CrossRef] [PubMed]
97.
Cho, K.A.; Park, M.; Kim, Y.H.; Woo, S.Y.; Ryu, K.H. RNA sequencing reveals a transcriptomic portrait of human mesenchymal
stem cells from bone marrow, adipose tissue, and palatine tonsils. Sci. Rep. 2017,7, 17114. [CrossRef] [PubMed]
98.
Lin, W.; Huang, L.; Li, Y.; Fang, B.; Li, G.; Chen, L.; Xu, L. Mesenchymal Stem Cells and Cancer: Clinical Challenges and
Opportunities. BioMed Res. Int. 2019,2019, 2820853. [CrossRef] [PubMed]
99.
Lee, J.K.; Park, S.R.; Jung, B.K.; Jeon, Y.K.; Lee, Y.S.; Kim, M.K.; Kim, Y.G.; Jang, J.Y.; Kim, C.W. Exosomes derived from
mesenchymal stem cells suppress angiogenesis by down-regulating VEGF expression in breast cancer cells. PLoS ONE
2013
,
8, e84256. [CrossRef] [PubMed]
100.
Clarke, M.R.; Imhoff, F.M.; Baird, S.K. Mesenchymal stem cells inhibit breast cancer cell migration and invasion through secretion
of tissue inhibitor of metalloproteinase-1 and -2. Mol. Carcinog. 2015,54, 1214–1219. [CrossRef] [PubMed]
101.
He, N.; Kong, Y.; Lei, X.; Liu, Y.; Wang, J.; Xu, C.; Wang, Y.; Du, L.; Ji, K.; Wang, Q.; et al. MSCs inhibit tumor progression
and enhance radiosensitivity of breast cancer cells by down-regulating Stat3 signaling pathway. Cell Death Dis.
2018
,9, 1026.
[CrossRef] [PubMed]
102.
Yang, H.Y.; Qu, R.M.; Lin, X.S.; Liu, T.X.; Sun, Q.Q.; Yang, C.; Li, X.H.; Lu, W.; Hu, X.F.; Dai, J.X.; et al. IGF-1 from adipose-derived
mesenchymal stem cells promotes radioresistance of breast cancer cells. Asian Pac. J. Cancer Prev. APJCP
2014
,15, 10115–10119.
[CrossRef] [PubMed]
103.
De Luca, A.; Lamura, L.; Gallo, M.; Maffia, V.; Normanno, N. Mesenchymal stem cell-derived interleukin-6 and vascular
endothelial growth factor promote breast cancer cell migration. J. Cell. Biochem. 2012,113, 3363–3370. [CrossRef] [PubMed]
104.
Xu, H.; Zhou, Y.; Li, W.; Zhang, B.; Zhang, H.; Zhao, S.; Zheng, P.; Wu, H.; Yang, J. Tumor-derived mesenchymal-stem-cell-secreted
IL-6 enhances resistance to cisplatin via the STAT3 pathway in breast cancer. Oncol. Lett.
2018
,15, 9142–9150. [CrossRef] [PubMed]
105.
Yan, X.-L.; Fu, C.-J.; Chen, L.; Qin, J.-H.; Zeng, Q.; Yuan, H.-F.; Nan, X.; Chen, H.-X.; Zhou, J.-N.; Lin, Y.-L.; et al. Mesenchymal
stem cells from primary breast cancer tissue promote cancer proliferation and enhance mammosphere formation partially via
EGF/EGFR/Akt pathway. Breast Cancer Res. Treat. 2012,132, 153–164. [CrossRef]
106.
Devarajan, E.; Song, Y.H.; Krishnappa, S.; Alt, E. Epithelial-mesenchymal transition in breast cancer lines is mediated through
PDGF-D released by tissue-resident stem cells. Int. J. Cancer 2012,131, 1023–1031. [CrossRef] [PubMed]
107.
Chen, Y.-C.; Gonzalez, M.E.; Burman, B.; Zhao, X.; Anwar, T.; Tran, M.; Medhora, N.; Hiziroglu, A.B.; Lee, W.; Cheng, Y.-H.; et al.
Mesenchymal Stem/Stromal Cell Engulfment Reveals Metastatic Advantage in Breast Cancer. Cell Rep.
2019
,27, 3916–3926.e15.
[CrossRef] [PubMed]
108.
Wu, S.; Wang, Y.; Yuan, Z.; Wang, S.; Du, H.; Liu, X.; Wang, Q.; Zhu, X. Human adipose-derived mesenchymal stem cells promote
breast cancer MCF7 cell epithelial-mesenchymal transition by cross interacting with the TGF-
β
/Smad and PI3K/AKT signaling
pathways. Mol. Med. Rep. 2019,19, 177–186. [CrossRef] [PubMed]
109.
Su, F.; Wang, X.; Pearson, T.; Lee, J.; Krishnamurthy, S.; Ueno, N.T.; Kolonin, M.G. Ablation of Stromal Cells with a Targeted
Proapoptotic Peptide Suppresses Cancer Chemotherapy Resistance and Metastasis. Mol. Ther. Oncolytics
2020
,18, 579–586.
[CrossRef] [PubMed]
110.
Razmkhah, M.; Jaberipour, M.; Erfani, N.; Habibagahi, M.; Talei, A.R.; Ghaderi, A. Adipose derived stem cells (ASCs) isolated
from breast cancer tissue express IL-4, IL-10 and TGF-
β
1 and upregulate expression of regulatory molecules on T cells: Do they
protect breast cancer cells from the immune response? Cell. Immunol. 2011,266, 116–122. [CrossRef] [PubMed]
111.
Sepehr, K.S.; Razavi, A.; Hassan, Z.M.; Fazel, A.; Abdollahpour-Alitappeh, M.; Mossahebi-Mohammadi, M.; Yekaninejad, M.S.;
Farhadihosseinabadi, B.; Hashemi, S.M. Comparative immunomodulatory properties of mesenchymal stem cells derived from
human breast tumor and normal breast adipose tissue. Cancer Immunol. Immunother. 2020,69, 1841–1854. [CrossRef]
112.
Mehdipour, F.; Razmkhah, M.; Rezaeifard, S.; Bagheri, M.; Talei, A.-R.; Khalatbari, B.; Ghaderi, A. Mesenchymal stem cells
induced anti-inflammatory features in B cells from breast tumor draining lymph nodes. Cell Biol. Int.
2018
,42, 1658–1669.
[CrossRef]
113.
Zaoui, M.; Morel, M.; Ferrand, N.; Fellahi, S.; Bastard, J.P.; Lamazière, A.; Larsen, A.K.; Béréziat, V.; Atlan, M.; Sabbah, M. Breast-
Associated Adipocytes Secretome Induce Fatty Acid Uptake and Invasiveness in Breast Cancer Cells via CD36 Independently of
Body Mass Index, Menopausal Status and Mammary Density. Cancers 2019,11, 2012. [CrossRef]
114.
Sakurai, M.; Miki, Y.; Takagi, K.; Suzuki, T.; Ishida, T.; Ohuchi, N.; Sasano, H. Interaction with adipocyte stromal cells induces
breast cancer malignancy via S100A7 upregulation in breast cancer microenvironment. Breast Cancer Res.
2017
,19, 70. [CrossRef]
115.
Ren, G.; Zhao, X.; Wang, Y.; Zhang, X.; Chen, X.; Xu, C.; Yuan, Z.R.; Roberts, A.I.; Zhang, L.; Zheng, B.; et al. CCR2-dependent
recruitment of macrophages by tumor-educated mesenchymal stromal cells promotes tumor development and is mimicked by
TNFα.Cell Stem Cell 2012,11, 812–824. [CrossRef]
116.
Yu, P.F.; Huang, Y.; Han, Y.Y.; Lin, L.Y.; Sun, W.H.; Rabson, A.B.; Wang, Y.; Shi, Y.F. TNF
α
-activated mesenchymal stromal cells
promote breast cancer metastasis by recruiting CXCR2(+) neutrophils. Oncogene 2017,36, 482–490. [CrossRef]

Cancers 2022,14, 3908 29 of 35
117.
Halpern, J.L.; Kilbarger, A.; Lynch, C.C. Mesenchymal stem cells promote mammary cancer cell migration
in vitro
via the CXCR2
receptor. Cancer Lett. 2011,308, 91–99. [CrossRef]
118.
Zhang, T.; Lee, Y.W.; Rui, Y.F.; Cheng, T.Y.; Jiang, X.H.; Li, G. Bone marrow-derived mesenchymal stem cells promote growth and
angiogenesis of breast and prostate tumors. Stem Cell Res. Ther. 2013,4, 70. [CrossRef]
119.
Wei, H.-J.; Zeng, R.; Lu, J.-H.; Lai, W.-F.T.; Chen, W.-H.; Liu, H.-Y.; Chang, Y.-T.; Deng, W.-P. Adipose-derived stem cells promote
tumor initiation and accelerate tumor growth by interleukin-6 production. Oncotarget 2015,6, 7713–7726. [CrossRef]
120.
Goto, H.; Shimono, Y.; Funakoshi, Y.; Imamura, Y.; Toyoda, M.; Kiyota, N.; Kono, S.; Takao, S.; Mukohara, T.; Minami, H.
Adipose-derived stem cells enhance human breast cancer growth and cancer stem cell-like properties through adipsin. Oncogene
2019,38, 767–779. [CrossRef]
121.
Strong, A.L.; Ohlstein, J.F.; Biagas, B.A.; Rhodes, L.V.; Pei, D.T.; Tucker, H.A.; Llamas, C.; Bowles, A.C.; Dutreil, M.F.; Zhang,
S.; et al. Leptin produced by obese adipose stromal/stem cells enhances proliferation and metastasis of estrogen receptor positive
breast cancers. Breast Cancer Res. 2015,17, 112. [CrossRef] [PubMed]
122.
Strong, A.L.; Strong, T.A.; Rhodes, L.V.; Semon, J.A.; Zhang, X.; Shi, Z.; Zhang, S.; Gimble, J.M.; Burow, M.E.; Bunnell, B.A. Obesity
associated alterations in the biology of adipose stem cells mediate enhanced tumorigenesis by estrogen dependent pathways.
Breast Cancer Res. BCR 2013,15, R102. [CrossRef] [PubMed]
123.
Hillers, L.E.; D’Amato, J.V.; Chamberlin, T.; Paderta, G.; Arendt, L.M. Obesity-Activated Adipose-Derived Stromal Cells Promote
Breast Cancer Growth and Invasion. Neoplasia 2018,20, 1161–1174. [CrossRef] [PubMed]
124.
Sabol, R.A.; Bowles, A.C.; Côté, A.; Wise, R.; O’Donnell, B.; Matossian, M.D.; Hossain, F.M.; Burks, H.E.; Del Valle, L.; Miele,
L.; et al. Leptin produced by obesity-altered adipose stem cells promotes metastasis but not tumorigenesis of triple-negative
breast cancer in orthotopic xenograft and patient-derived xenograft models. Breast Cancer Res. 2019,21, 67. [CrossRef]
125.
Ling, L.; Mulligan, J.A.; Ouyang, Y.; Shimpi, A.A.; Williams, R.M.; Beeghly, G.F.; Hopkins, B.D.; Spector, J.A.; Adie, S.G.; Fischbach,
C. Obesity-associated Adipose Stromal Cells Promote Breast Cancer Invasion Through Direct Cell Contact and ECM Remodeling.
Adv. Funct. Mater. 2020,30, 1910650. [CrossRef]
126.
Strong, A.L.; Pei, D.T.; Hurst, C.G.; Gimble, J.M.; Burow, M.E.; Bunnell, B.A. Obesity Enhances the Conversion of Adipose-Derived
Stromal/Stem Cells into Carcinoma-Associated Fibroblast Leading to Cancer Cell Proliferation and Progression to an Invasive
Phenotype. Stem Cells Int. 2017,2017, 9216502. [CrossRef]
127.
Sabol, R.A.; Beighley, A.; Giacomelli, P.; Wise, R.M.; Harrison, M.A.A.; O’Donnnell, B.A.; Sullivan, B.N.; Lampenfeld, J.D.;
Matossian, M.D.; Bratton, M.R.; et al. Obesity-Altered Adipose Stem Cells Promote ER
+
Breast Cancer Metastasis through
Estrogen Independent Pathways. Int. J. Mol. Sci. 2019,20, 1419. [CrossRef]
128. Kwon, H.; Pessin, J.E. Adipokines mediate inflammation and insulin resistance. Front. Endocrinol. 2013,4, 71. [CrossRef]
129.
Esquivel-Velázquez, M.; Ostoa-Saloma, P.; Palacios-Arreola, M.I.; Nava-Castro, K.E.; Castro, J.I.; Morales-Montor, J. The role of
cytokines in breast cancer development and progression. J. Interferon Cytokine Res. 2015,35, 1–16. [CrossRef]
130.
Manore, S.G.; Doheny, D.L.; Wong, G.L.; Lo, H.-W. IL-6/JAK/STAT3 Signaling in Breast Cancer Metastasis: Biology and Treatment.
Front. Oncol. 2022,12, 866014. [CrossRef]
131.
Iyengar, N.M.; Gucalp, A.; Dannenberg, A.J.; Hudis, C.A. Obesity and Cancer Mechanisms: Tumor Microenvironment and
Inflammation. J. Clin. Oncol. Off. J. Am. Soc. Clin. Oncol. 2016,34, 4270–4276. [CrossRef]
132.
Pileczki, V.; Braicu, C.; Gherman, C.D.; Berindan-Neagoe, I. TNF-
α
gene knockout in triple negative breast cancer cell line induces
apoptosis. Int. J. Mol. Sci. 2012,14, 411–420. [CrossRef]
133.
Kitahara, C.M.; Flint, A.J.; de Gonzalez, A.B.; Bernstein, L.; Brotzman, M.; MacInnis, R.J.; Moore, S.C.; Robien, K.; Rosenberg, P.S.;
Singh, P.N.; et al. Association between Class III Obesity (BMI of 40–59 kg/m
2
) and Mortality: A Pooled Analysis of 20 Prospective
Studies. PLoS Med. 2014,11, e1001673. [CrossRef]
134.
Obradovic, M.; Sudar-Milovanovic, E.; Soskic, S.; Essack, M.; Arya, S.; Stewart, A.J.; Gojobori, T.; Isenovic, E.R. Leptin and
Obesity: Role and Clinical Implication. Front. Endocrinol. 2021,12, 585887. [CrossRef]
135.
Gruber, T.; Pan, C.; Contreras, R.E.; Wiedemann, T.; Morgan, D.A.; Skowronski, A.A.; Lefort, S.; De Bernardis Murat, C.; Le
Thuc, O.; Legutko, B.; et al. Obesity-associated hyperleptinemia alters the gliovascular interface of the hypothalamus to promote
hypertension. Cell Metab. 2021,33, 1155–1170. [CrossRef]
136.
Andò, S.; Catalano, S. The multifactorial role of leptin in driving the breast cancer microenvironment. Nat. Rev. Endocrinol.
2012
,
8, 263–275. [CrossRef]
137.
Ianza, A.; Sirico, M.; Bernocchi, O.; Generali, D. Role of the IGF-1 Axis in Overcoming Resistance in Breast Cancer. Front. Cell Dev.
Biol. 2021,9, 641449. [CrossRef]
138.
Christopoulos, P.F.; Msaouel, P.; Koutsilieris, M. The role of the insulin-like growth factor-1 system in breast cancer. Mol. Cancer
2015,14, 43. [CrossRef]
139.
Creighton, C.J.; Casa, A.; Lazard, Z.; Huang, S.; Tsimelzon, A.; Hilsenbeck, S.G.; Osborne, C.K.; Lee, A.V. Insulin-like growth
factor-I activates gene transcription programs strongly associated with poor breast cancer prognosis. J. Clin. Oncol. Off. J. Am. Soc.
Clin. Oncol. 2008,26, 4078–4085. [CrossRef]
140. Pollak, M. Insulin and insulin-like growth factor signalling in neoplasia. Nat. Rev. Cancer 2008,8, 915–928. [CrossRef]
141.
Edakuni, G.; Sasatomi, E.; Satoh, T.; Tokunaga, O.; Miyazaki, K. Expression of the hepatocyte growth factor/c-Met pathway is
increased at the cancer front in breast carcinoma. Pathol. Int. 2001,51, 172–178. [CrossRef]

Cancers 2022,14, 3908 30 of 35
142.
Bell, L.N.; Ward, J.L.; Degawa-Yamauchi, M.; Bovenkerk, J.E.; Jones, R.; Cacucci, B.M.; Gupta, C.E.; Sheridan, C.; Sheridan, K.;
Shankar, S.S.; et al. Adipose tissue production of hepatocyte growth factor contributes to elevated serum HGF in obesity. Am. J.
Physiol. Endocrinol. Metab. 2006,291, E843–E848. [CrossRef]
143.
Guillaume, V.G.J.; Ruhl, T.; Boos, A.M.; Beier, J.P. The Crosstalk Between Adipose-Derived Stem or Stromal Cells (ASC) and
Cancer Cells and ASC-Mediated Effects on Cancer Formation and Progression—ASCs: Safety Hazard or Harmless Source of
Tropism? Stem Cells Transl. Med. 2022,11, 394–406. [CrossRef] [PubMed]
144.
Burns, J.S.; Kristiansen, M.; Kristensen, L.P.; Larsen, K.H.; Nielsen, M.O.; Christiansen, H.; Nehlin, J.; Andersen, J.S.; Kassem, M.
Decellularized matrix from tumorigenic human mesenchymal stem cells promotes neovascularization with galectin-1 dependent
endothelial interaction. PLoS ONE 2011,6, e21888. [CrossRef] [PubMed]
145.
Xu, H.; Li, W.; Luo, S.; Yuan, J.; Hao, L. Adipose derived stem cells promote tumor metastasis in breast Cancer cells by stem cell
factor inhibition of miR20b. Cell. Signal. 2019,62, 109350. [CrossRef]
146.
Watt, S.M.; Gullo, F.; van der Garde, M.; Markeson, D.; Camicia, R.; Khoo, C.P.; Zwaginga, J.J. The angiogenic properties of
mesenchymal stem/stromal cells and their therapeutic potential. Br. Med. Bull. 2013,108, 25–53. [CrossRef]
147.
Shi, W.; Xin, Q.; Yuan, R.; Yuan, Y.; Cong, W.; Chen, K. Neovascularization: The Main Mechanism of MSCs in Ischemic Heart
Disease Therapy. Front. Cardiovasc. Med. 2021,8, 633300. [CrossRef]
148.
Loibl, M.; Binder, A.; Herrmann, M.; Duttenhoefer, F.; Richards, R.G.; Nerlich, M.; Alini, M.; Verrier, S. Direct cell-cell contact
between mesenchymal stem cells and endothelial progenitor cells induces a pericyte-like phenotype
in vitro
.BioMed Res. Int.
2014,2014, 395781. [CrossRef]
149. Maacha, S.; Sidahmed, H.; Jacob, S.; Gentilcore, G.; Calzone, R.; Grivel, J.-C.; Cugno, C. Paracrine Mechanisms of Mesenchymal
Stromal Cells in Angiogenesis. Stem Cells Int. 2020,2020, 4356359. [CrossRef]
150.
Matsushita, K.; Dzau, V.J. Mesenchymal stem cells in obesity: Insights for translational applications. Lab. Investig.
2017
,97,
1158–1166. [CrossRef]
151.
Pérez, L.M.; Bernal, A.; San Martín, N.; Gálvez, B.G. Obese-derived ASCs show impaired migration and angiogenesis properties.
Arch. Physiol. Biochem. 2013,119, 195–201. [CrossRef]
152.
Juntunen, M.; Heinonen, S.; Huhtala, H.; Rissanen, A.; Kaprio, J.; Kuismanen, K.; Pietiläinen, K.H.; Miettinen, S.; Patrikoski, M.
Evaluation of the effect of donor weight on adipose stromal/stem cell characteristics by using weight-discordant monozygotic
twin pairs. Stem Cell Res. Ther. 2021,12, 516. [CrossRef] [PubMed]
153.
Conley, S.M.; Hickson, L.J.; Kellogg, T.A.; McKenzie, T.; Heimbach, J.K.; Taner, T.; Tang, H.; Jordan, K.L.; Saadiq, I.M.; Woollard,
J.R.; et al. Human Obesity Induces Dysfunction and Early Senescence in Adipose Tissue-Derived Mesenchymal Stromal/Stem
Cells. Front. Cell Dev. Biol. 2020,8, 197. [CrossRef]
154.
Togliatto, G.; Dentelli, P.; Gili, M.; Gallo, S.; Deregibus, C.; Biglieri, E.; Iavello, A.; Santini, E.; Rossi, C.; Solini, A.; et al. Obesity
reduces the pro-angiogenic potential of adipose tissue stem cell-derived extracellular vesicles (EVs) by impairing miR-126 content:
Impact on clinical applications. Int. J. Obes. 2016,40, 102–111. [CrossRef] [PubMed]
155.
Eirin, A.; Meng, Y.; Zhu, X.-Y.; Li, Y.; Saadiq, I.M.; Jordan, K.L.; Tang, H.; Lerman, A.; van Wijnen, A.J.; Lerman, L.O. The
Micro-RNA Cargo of Extracellular Vesicles Released by Human Adipose Tissue-Derived Mesenchymal Stem Cells Is Modified by
Obesity. Front. Cell Dev. Biol. 2021,9, 660851. [CrossRef] [PubMed]
156.
Incio, J.; Ligibel, J.A.; McManus, D.T.; Suboj, P.; Jung, K.; Kawaguchi, K.; Pinter, M.; Babykutty, S.; Chin, S.M.; Vardam, T.D.; et al.
Obesity promotes resistance to anti-VEGF therapy in breast cancer by up-regulating IL-6 and potentially FGF-2. Sci. Transl. Med.
2018,10, eaag0945. [CrossRef]
157.
Sánchez-Jiménez, F.; Pérez-Pérez, A.; de la Cruz-Merino, L.; Sánchez-Margalet, V. Obesity and Breast Cancer: Role of Leptin.
Front. Oncol. 2019,9, 596. [CrossRef]
158.
Kolb, R.; Kluz, P.; Tan, Z.W.; Borcherding, N.; Bormann, N.; Vishwakarma, A.; Balcziak, L.; Zhu, P.; Davies, B.S.; Gourronc, F.; et al.
Obesity-associated inflammation promotes angiogenesis and breast cancer via angiopoietin-like 4. Oncogene
2019
,38, 2351–2363.
[CrossRef]
159.
Oñate, B.; Vilahur, G.; Camino-López, S.; Díez-Caballero, A.; Ballesta-López, C.; Ybarra, J.; Moscatiello, F.; Herrero, J.; Badimon,
L. Stem cells isolated from adipose tissue of obese patients show changes in their transcriptomic profile that indicate loss in
stemcellness and increased commitment to an adipocyte-like phenotype. BMC Genom. 2013,14, 625. [CrossRef]
160.
Gonzalez-Perez, R.R.; Xu, Y.; Guo, S.; Watters, A.; Zhou, W.; Leibovich, S.J. Leptin upregulates VEGF in breast cancer via canonic
and non-canonical signalling pathways and NFκB/HIF-1αactivation. Cell. Signal. 2010,22, 1350–1362. [CrossRef]
161.
Newman, G.; Gonzalez-Perez, R.R. Leptin–cytokine crosstalk in breast cancer. Mol. Cell. Endocrinol.
2014
,382, 570–582. [CrossRef]
[PubMed]
162.
Khong, H.T.; Restifo, N.P. Natural selection of tumor variants in the generation of “tumor escape” phenotypes. Nat. Immunol.
2002,3, 999–1005. [CrossRef] [PubMed]
163.
Dunn, G.P.; Bruce, A.T.; Ikeda, H.; Old, L.J.; Schreiber, R.D. Cancer immunoediting: From immunosurveillance to tumor escape.
Nat. Immunol. 2002,3, 991–998. [CrossRef] [PubMed]
164.
Terry, S.; Savagner, P.; Ortiz-Cuaran, S.; Mahjoubi, L.; Saintigny, P.; Thiery, J.P.; Chouaib, S. New insights into the role of EMT in
tumor immune escape. Mol. Oncol. 2017,11, 824–846. [CrossRef]
165.
Spranger, S.; Gajewski, T.F. Impact of oncogenic pathways on evasion of antitumour immune responses. Nat. Rev. Cancer
2018
,18,
139–147. [CrossRef] [PubMed]

Cancers 2022,14, 3908 31 of 35
166.
Bejarano, L.; Jord
¯
ao, M.J.C.; Joyce, J.A. Therapeutic Targeting of the Tumor Microenvironment. Cancer Discov.
2021
,11, 933–959.
[CrossRef]
167. Mortezaee, K. Immune escape: A critical hallmark in solid tumors. Life Sci. 2020,258, 118110. [CrossRef] [PubMed]
168.
Garrido, F.; Aptsiauri, N. Cancer immune escape: MHC expression in primary tumours versus metastases. Immunology
2019
,158,
255–266. [CrossRef]
169.
McIntosh, K.; Zvonic, S.; Garrett, S.; Mitchell, J.B.; Floyd, Z.E.; Hammill, L.; Kloster, A.; Di Halvorsen, Y.; Ting, J.P.; Storms,
R.W.; et al. The immunogenicity of human adipose-derived cells: Temporal changes
in vitro
.Stem Cells
2006
,24, 1246–1253.
[CrossRef] [PubMed]
170.
Russell, K.A.; Chow, N.H.; Dukoff, D.; Gibson, T.W.; LaMarre, J.; Betts, D.H.; Koch, T.G. Characterization and Immunomodulatory
Effects of Canine Adipose Tissue- and Bone Marrow-Derived Mesenchymal Stromal Cells. PLoS ONE
2016
,11, e0167442.
[CrossRef] [PubMed]
171.
Ceccarelli, S.; Pontecorvi, P.; Anastasiadou, E.; Napoli, C.; Marchese, C. Immunomodulatory Effect of Adipose-Derived Stem
Cells: The Cutting Edge of Clinical Application. Front. Cell Dev. Biol. 2020,8, 236. [CrossRef] [PubMed]
172.
Leto Barone, A.A.; Khalifian, S.; Lee, W.P.A.; Brandacher, G. Immunomodulatory Effects of Adipose-Derived Stem Cells: Fact or
Fiction? BioMed Res. Int. 2013,2013, 383685. [CrossRef] [PubMed]
173.
Fakhimi, M.; Talei, A.R.; Ghaderi, A.; Habibagahi, M.; Razmkhah, M. Helios, CD73 and CD39 Induction in Regulatory T Cells
Exposed to Adipose Derived Mesenchymal Stem Cells. Cell J. 2020,22, 236–244. [CrossRef]
174.
Ayaz-Guner, S.; Alessio, N.; Acar, M.B.; Aprile, D.; Özcan, S.; Di Bernardo, G.; Peluso, G.; Galderisi, U. A comparative study
on normal and obese mice indicates that the secretome of mesenchymal stromal cells is influenced by tissue environment and
physiopathological conditions. Cell Commun. Signal. 2020,18, 118. [CrossRef] [PubMed]
175.
Eljaafari, A.; Pestel, J.; Le Magueresse-Battistoni, B.; Chanon, S.; Watson, J.; Robert, M.; Disse, E.; Vidal, H. Adipose-Tissue-Derived
Mesenchymal Stem Cells Mediate PD-L1 Overexpression in the White Adipose Tissue of Obese Individuals, Resulting in T Cell
Dysfunction. Cells 2021,10, 2645. [CrossRef] [PubMed]
176.
Zhu, X.-Y.; Klomjit, N.; Conley, S.M.; Ostlie, M.M.; Jordan, K.L.; Lerman, A.; Lerman, L.O. Impaired immunomodulatory capacity
in adipose tissue-derived mesenchymal stem/stromal cells isolated from obese patients. J. Cell. Mol. Med.
2021
,25, 9051–9059.
[CrossRef] [PubMed]
177.
Harrison, M.A.A.; Wise, R.M.; Benjamin, B.P.; Hochreiner, E.M.; Mohiuddin, O.A.; Bunnell, B.A. Adipose-Derived Stem Cells
from Obese Donors Polarize Macrophages and Microglia toward a Pro-Inflammatory Phenotype. Cells
2020
,10, 26. [CrossRef]
[PubMed]
178.
Eljaafari, A.; Robert, M.; Chehimi, M.; Chanon, S.; Durand, C.; Vial, G.; Bendridi, N.; Madec, A.M.; Disse, E.; Laville, M.; et al.
Adipose Tissue-Derived Stem Cells From Obese Subjects Contribute to Inflammation and Reduced Insulin Response in Adipocytes
Through Differential Regulation of the Th1/Th17 Balance and Monocyte Activation. Diabetes
2015
,64, 2477–2488. [CrossRef]
[PubMed]
179.
Benaiges, E.; Ceperuelo-Mallafré, V.; Madeira, A.; Bosch, R.; Núñez-Roa, C.; Ejarque, M.; Maymó-Masip, E.; Huber-Ruano, I.;
Lejeune, M.; Vendrell, J.; et al. Survivin drives tumor-associated macrophage reprogramming: A novel mechanism with potential
impact for obesity. Cell. Oncol. 2021,44, 777–792. [CrossRef] [PubMed]
180.
Fang, M.; Yuan, J.; Peng, C.; Li, Y. Collagen as a double-edged sword in tumor progression. Tumour Biol.
2014
,35, 2871–2882.
[CrossRef]
181.
Henke, E.; Nandigama, R.; Ergün, S. Extracellular Matrix in the Tumor Microenvironment and Its Impact on Cancer Therapy.
Front. Mol. Biosci. 2020,6, 160. [CrossRef] [PubMed]
182. Hay, E.D. An overview of epithelio-mesenchymal transformation. Acta Anat. 1995,154, 8–20. [CrossRef]
183. Xu, J.; Lamouille, S.; Derynck, R. TGF-β-induced epithelial to mesenchymal transition. Cell Res. 2009,19, 156–172. [CrossRef]
184.
Ogunwobi, O.O.; Liu, C. Hepatocyte growth factor upregulation promotes carcinogenesis and epithelial-mesenchymal transition
in hepatocellular carcinoma via Akt and COX-2 pathways. Clin. Exp. Metastasis 2011,28, 721–731. [CrossRef] [PubMed]
185.
Buyuk, B.; Jin, S.; Ye, K. Epithelial-to-Mesenchymal Transition Signaling Pathways Responsible for Breast Cancer Metastasis. Cell.
Mol. Bioeng. 2022,15, 1–13. [CrossRef] [PubMed]
186.
Moreno-Bueno, G.; Portillo, F.; Cano, A. Transcriptional regulation of cell polarity in EMT and cancer. Oncogene
2008
,27,
6958–6969. [CrossRef]
187. Thiery, J.P.; Lim, C.T. Tumor dissemination: An EMT affair. Cancer Cell 2013,23, 272–273. [CrossRef]
188.
Battula, V.L.; Evans, K.W.; Hollier, B.G.; Shi, Y.; Marini, F.C.; Ayyanan, A.; Wang, R.Y.; Brisken, C.; Guerra, R.; Andreeff, M.; et al.
Epithelial-mesenchymal transition-derived cells exhibit multilineage differentiation potential similar to mesenchymal stem cells.
Stem Cells 2010,28, 1435–1445. [CrossRef] [PubMed]
189.
Mani, S.A.; Guo, W.; Liao, M.J.; Eaton, E.N.; Ayyanan, A.; Zhou, A.Y.; Brooks, M.; Reinhard, F.; Zhang, C.C.; Shipitsin, M.; et al.
The epithelial-mesenchymal transition generates cells with properties of stem cells. Cell
2008
,133, 704–715. [CrossRef] [PubMed]
190.
Wu, H.-T.; Zhong, H.-T.; Li, G.-W.; Shen, J.-X.; Ye, Q.-Q.; Zhang, M.-L.; Liu, J. Oncogenic functions of the EMT-related transcription
factor ZEB1 in breast cancer. J. Transl. Med. 2020,18, 51. [CrossRef] [PubMed]
191.
Lüönd, F.; Sugiyama, N.; Bill, R.; Bornes, L.; Hager, C.; Tang, F.; Santacroce, N.; Beisel, C.; Ivanek, R.; Bürglin, T.; et al. Distinct
contributions of partial and full EMT to breast cancer malignancy. Dev. Cell 2021,56, 3203–3221. [CrossRef] [PubMed]

Cancers 2022,14, 3908 32 of 35
192.
Tian, T.; Hua, Z.; Wang, L.Z.; Wang, X.Y.; Chen, H.Y.; Liu, Z.H.; Cui, Z.M. S100A7 promotes the metastasis and epithelial-
mesenchymal transition on HeLa and CaSki cells. Zhonghua Fu Chan Ke Za Zhi 2018,53, 99–105. [CrossRef] [PubMed]
193.
Lozito, T.P.; Jackson, W.M.; Nesti, L.J.; Tuan, R.S. Human mesenchymal stem cells generate a distinct pericellular zone of MMP
activities via binding of MMPs and secretion of high levels of TIMPs. Matrix Biol. 2014,34, 132–143. [CrossRef]
194.
Casson, J.; O’Kane, S.; Smith, C.-A.; Dalby, M.J.; Berry, C.C. Interleukin 6 Plays a Role in the Migration of Magnetically Levitated
Mesenchymal Stem Cells Spheroids. Appl. Sci. 2018,8, 412. [CrossRef]
195.
Strong, A.L.; Semon, J.A.; Strong, T.A.; Santoke, T.T.; Zhang, S.; McFerrin, H.E.; Gimble, J.M.; Bunnell, B.A. Obesity-associated
dysregulation of calpastatin and MMP-15 in adipose-derived stromal cells results in their enhanced invasion. Stem Cells
2012
,30,
2774–2783. [CrossRef]
196.
Winkler, J.; Abisoye-Ogunniyan, A.; Metcalf, K.J.; Werb, Z. Concepts of extracellular matrix remodelling in tumour progression
and metastasis. Nat. Commun. 2020,11, 5120. [CrossRef]
197.
Zeng, R.; Huang, J.; Zhong, M.-Z.; Li, L.; Yang, G.; Liu, L.; Wu, Y.; Yao, X.; Shi, J.; Wu, Z. Multiple Roles of WNT5A in Breast
Cancer. Med. Sci. Monit. 2016,22, 5058–5067. [CrossRef]
198.
Suarez, N.G.; Fernandez-Marrero, Y.; Torabidastgerdooei, S.; Annabi, B. EGCG Prevents the Onset of an Inflammatory and Cancer-
Associated Adipocyte-like Phenotype in Adipose-Derived Mesenchymal Stem/Stromal Cells in Response to the Triple-Negative
Breast Cancer Secretome. Nutrients 2022,14, 1099. [CrossRef]
199.
Clara, J.A.; Monge, C.; Yang, Y.; Takebe, N. Targeting signalling pathways and the immune microenvironment of cancer stem
cells—A clinical update. Nat. Rev. Clin. Oncol. 2020,17, 204–232. [CrossRef]
200.
Jing, N.; Gao, W.-Q.; Fang, Y.-X. Regulation of Formation, Stemness and Therapeutic Resistance of Cancer Stem Cells. Front. Cell
Dev. Biol. 2021,9, 641498. [CrossRef]
201.
Yousefnia, S.; Forootan, F.S.; Forootan, S.S.; Esfahani, M.H.N.; Gure, A.O.; Ghaedi, K. Mechanistic Pathways of Malignancy in
Breast Cancer Stem Cells. Front. Oncol. 2020,10, 452. [CrossRef] [PubMed]
202.
Soni, S.; Torvund, M.; Mandal, C.C. Molecular insights into the interplay between adiposity, breast cancer and bone metastasis.
Clin. Exp. Metastasis 2021,38, 119–138. [CrossRef] [PubMed]
203.
Mishra, A.K.; Parish, C.R.; Wong, M.-L.; Licinio, J.; Blackburn, A.C. Leptin signals via TGFB1 to promote metastatic potential and
stemness in breast cancer. PLoS ONE 2017,12, e0178454. [CrossRef]
204.
Khan, Z.; Marshall, J.F. The role of integrins in TGF
β
activation in the tumour stroma. Cell Tissue Res.
2016
,365, 657–673.
[CrossRef]
205.
Gieniec, K.A.; Butler, L.M.; Worthley, D.L.; Woods, S.L. Cancer-associated fibroblasts—Heroes or villains? Br. J. Cancer
2019
,121,
293–302. [CrossRef]
206.
Giandalia, A.; Alibrandi, A.; Giorgianni, L.; Lo Piano, F.; Consolo, F.; Longo Elia, G.; Asztalos, B.; Cucinotta, D.; Squadrito, G.;
Russo, G.T. Resistin levels and inflammatory and endothelial dysfunction markers in obese postmenopausal women with type 2
diabetes mellitus. Diabetol. Metab. Syndr. 2021,13, 98. [CrossRef]
207.
Avtanski, D.; Garcia, A.; Caraballo, B.; Thangeswaran, P.; Marin, S.; Bianco, J.; Lavi, A.; Poretsky, L. Resistin induces breast
cancer cells epithelial to mesenchymal transition (EMT) and stemness through both adenylyl cyclase-associated protein 1
(CAP1)-dependent and CAP1-independent mechanisms. Cytokine 2019,120, 155–164. [CrossRef]
208.
Kern, L.; Mittenbühler, M.J.; Vesting, A.J.; Ostermann, A.L.; Wunderlich, C.M.; Wunderlich, F.T. Obesity-Induced TNF
α
and
IL-6 Signaling: The Missing Link between Obesity and Inflammation-Driven Liver and Colorectal Cancers. Cancers 2018,11, 24.
[CrossRef]
209. Smith, B.N.; Bhowmick, N.A. Role of EMT in Metastasis and Therapy Resistance. J. Clin. Med. 2016,5, 17. [CrossRef]
210.
Su, F.; Ahn, S.; Saha, A.; DiGiovanni, J.; Kolonin, M.G. Adipose stromal cell targeting suppresses prostate cancer epithelial-
mesenchymal transition and chemoresistance. Oncogene 2019,38, 1979–1988. [CrossRef]
211.
Ohtsuka, T.; Liu, X.F.; Koga, Y.; Kitajima, Y.; Nakafusa, Y.; Ha, C.W.; Lee, S.W.; Miyazaki, K. Methylation-induced silencing of ASC
and the effect of expressed ASC on p53-mediated chemosensitivity in colorectal cancer. Oncogene
2006
,25, 1807–1811. [CrossRef]
[PubMed]
212.
Nowicka, A.; Marini, F.C.; Solley, T.N.; Elizondo, P.B.; Zhang, Y.; Sharp, H.J.; Broaddus, R.; Kolonin, M.; Mok, S.C.; Thompson,
M.S.; et al. Human omental-derived adipose stem cells increase ovarian cancer proliferation, migration, and chemoresistance.
PLoS ONE 2013,8, e81859. [CrossRef]
213.
Raghavan, S.; Snyder, C.S.; Wang, A.; McLean, K.; Zamarin, D.; Buckanovich, R.J.; Mehta, G. Carcinoma-Associated Mesenchymal
Stem Cells Promote Chemoresistance in Ovarian Cancer Stem Cells via PDGF Signaling. Cancers
2020
,12, 2063. [CrossRef]
[PubMed]
214.
Cascio, S.; Chandler, C.; Zhang, L.; Sinno, S.; Gao, B.; Onkar, S.; Bruno, T.C.; Vignali, D.A.A.; Mahdi, H.; Osmanbeyoglu, H.U.; et al.
Cancer-associated MSC drive tumor immune exclusion and resistance to immunotherapy, which can be overcome by Hedgehog
inhibition. Sci. Adv. 2021,7, eabi5790. [CrossRef]
215.
Fregni, G.; Quinodoz, M.; Möller, E.; Vuille, J.; Galland, S.; Fusco, C.; Martin, P.; Letovanec, I.; Provero, P.; Rivolta, C.; et al.
Reciprocal modulation of mesenchymal stem cells and tumor cells promotes lung cancer metastasis. EBioMedicine
2018
,29,
128–145. [CrossRef]
216.
Scherzed, A.; Hackenberg, S.; Froelich, K.; Kessler, M.; Koehler, C.; Hagen, R.; Radeloff, A.; Friehs, G.; Kleinsasser, N. BMSC
enhance the survival of paclitaxel treated squamous cell carcinoma cells
in vitro
.Cancer Biol. Ther.
2011
,11, 349–357. [CrossRef]

Cancers 2022,14, 3908 33 of 35
217.
Takam Kamga, P.; Bassi, G.; Cassaro, A.; Midolo, M.; Di Trapani, M.; Gatti, A.; Carusone, R.; Resci, F.; Perbellini, O.; Gottardi,
M.; et al. Notch signalling drives bone marrow stromal cell-mediated chemoresistance in acute myeloid leukemia. Oncotarget
2016,7, 21713–21727. [CrossRef]
218.
Castells, M.; Milhas, D.; Gandy, C.; Thibault, B.; Rafii, A.; Delord, J.P.; Couderc, B. Microenvironment mesenchymal cells protect
ovarian cancer cell lines from apoptosis by inhibiting XIAP inactivation. Cell Death Dis. 2013,4, e887. [CrossRef]
219.
de Miranda, M.C.; Ferreira, A.d.F.; de Melo, M.I.A.; Kunrath-Lima, M.; Goes, A.M.d.; Rodrigues, M.A.; Gomes, D.A.; Faria,
J.A.Q.A. Adipose-derived stem/stromal cell secretome modulates breast cancer cell proliferation and differentiation state towards
aggressiveness. Biochimie 2021,191, 69–77. [CrossRef]
220.
Li, J.J.; Tsang, J.Y.; Tse, G.M. Tumor Microenvironment in Breast Cancer-Updates on Therapeutic Implications and Pathologic
Assessment. Cancers 2021,13, 4233. [CrossRef]
221.
Chen, Y.; McAndrews, K.M.; Kalluri, R. Clinical and therapeutic relevance of cancer-associated fibroblasts. Nat. Rev. Clin. Oncol.
2021,18, 792–804. [CrossRef] [PubMed]
222.
Soundararajan, M.; Kannan, S. Fibroblasts and mesenchymal stem cells: Two sides of the same coin? J. Cell. Physiol.
2018
,233,
9099–9109. [CrossRef] [PubMed]
223.
Ugurlu, B.; Karaoz, E. Comparison of similar cells: Mesenchymal stromal cells and fibroblasts. Acta Histochem.
2020
,122, 151634.
[CrossRef]
224.
Schellenberg, A.; Lin, Q.; Schüler, H.; Koch, C.M.; Joussen, S.; Denecke, B.; Walenda, G.; Pallua, N.; Suschek, C.V.; Zenke, M.; et al.
Replicative senescence of mesenchymal stem cells causes DNA-methylation changes which correlate with repressive histone
marks. Aging 2011,3, 873–888. [CrossRef]
225.
Koch, C.M.; Suschek, C.V.; Lin, Q.; Bork, S.; Goergens, M.; Joussen, S.; Pallua, N.; Ho, A.D.; Zenke, M.; Wagner, W. Specific
age-associated DNA methylation changes in human dermal fibroblasts. PLoS ONE 2011,6, e16679. [CrossRef]
226.
Liang, W.; Chen, X.; Zhang, S.; Fang, J.; Chen, M.; Xu, Y.; Chen, X. Mesenchymal stem cells as a double-edged sword in tumor
growth: Focusing on MSC-derived cytokines. Cell. Mol. Biol. Lett. 2021,26, 3. [CrossRef]
227.
Li, W.; Yang, J.; Zheng, P.; Li, H.; Zhao, S. The Origins and Generation of Cancer-Associated Mesenchymal Stromal Cells: An
Innovative Therapeutic Target for Solid Tumors. Front. Oncol. 2021,11, 723707. [CrossRef]
228.
Wu, Q.; Li, B.; Li, Z.; Li, J.; Sun, S.; Sun, S. Cancer-associated adipocytes: Key players in breast cancer progression. J. Hematol.
Oncol. 2019,12, 95. [CrossRef] [PubMed]
229.
McLean, K.; Gong, Y.; Choi, Y.; Deng, N.; Yang, K.; Bai, S.; Cabrera, L.; Keller, E.; McCauley, L.; Cho, K.R.; et al. Human ovarian
carcinoma–associated mesenchymal stem cells regulate cancer stem cells and tumorigenesis via altered BMP production. J. Clin.
Investig. 2011,121, 3206–3219. [CrossRef]
230.
Barcellos-de-Souza, P.; Comito, G.; Pons-Segura, C.; Taddei, M.L.; Gori, V.; Becherucci, V.; Bambi, F.; Margheri, F.; Laurenzana, A.;
Del Rosso, M.; et al. Mesenchymal Stem Cells are Recruited and Activated into Carcinoma-Associated Fibroblasts by Prostate
Cancer Microenvironment-Derived TGF-β1. Stem Cells 2016,34, 2536–2547. [CrossRef]
231.
Li, W.; Zhang, X.; Wu, F.; Zhou, Y.; Bao, Z.; Li, H.; Zheng, P.; Zhao, S. Gastric cancer-derived mesenchymal stromal cells trigger M2
macrophage polarization that promotes metastasis and EMT in gastric cancer. Cell Death Dis.
2019
,10, 918. [CrossRef] [PubMed]
232.
Kamdje, A.H.N.; Etet, P.F.S.; Tagne, R.S.; Vecchio, L.; Lukong, K.E.; Krampera, M. Tumor Microenvironment Uses a Reversible
Reprogramming of Mesenchymal Stromal Cells to Mediate Pro-tumorigenic Effects. Front. Cell Dev. Biol.
2020
,8, 545126.
[CrossRef]
233.
Anvarian, Z.; Mykytyn, K.; Mukhopadhyay, S.; Pedersen, L.B.; Christensen, S.T. Cellular signalling by primary cilia in develop-
ment, organ function and disease. Nat. Rev. Nephrol. 2019,15, 199–219. [CrossRef] [PubMed]
234.
Peraldi, P.; Ladoux, A.; Giorgetti-Peraldi, S.; Dani, C. The Primary Cilium of Adipose Progenitors Is Necessary for Their
Differentiation into Cancer-Associated Fibroblasts that Promote Migration of Breast Cancer Cells In Vitro. Cells
2020
,9, 2251.
[CrossRef] [PubMed]
235.
Wang, Y.; Liang, Y.; Xu, H.; Zhang, X.; Mao, T.; Cui, J.; Yao, J.; Wang, Y.; Jiao, F.; Xiao, X.; et al. Single-cell analysis of pancreatic
ductal adenocarcinoma identifies a novel fibroblast subtype associated with poor prognosis but better immunotherapy response.
Cell Discov. 2021,7, 36. [CrossRef] [PubMed]
236.
Dominguez, C.X.; Müller, S.; Keerthivasan, S.; Koeppen, H.; Hung, J.; Gierke, S.; Breart, B.; Foreman, O.; Bainbridge, T.W.;
Castiglioni, A.; et al. Single-Cell RNA Sequencing Reveals Stromal Evolution into LRRC15(+) Myofibroblasts as a Determinant of
Patient Response to Cancer Immunotherapy. Cancer Discov. 2020,10, 232–253. [CrossRef] [PubMed]
237.
Elyada, E.; Bolisetty, M.; Laise, P.; Flynn, W.F.; Courtois, E.T.; Burkhart, R.A.; Teinor, J.A.; Belleau, P.; Biffi, G.; Lucito, M.S.; et al.
Cross-Species Single-Cell Analysis of Pancreatic Ductal Adenocarcinoma Reveals Antigen-Presenting Cancer-Associated Fibrob-
lasts. Cancer Discov. 2019,9, 1102–1123. [CrossRef] [PubMed]
238.
Costa, A.; Kieffer, Y.; Scholer-Dahirel, A.; Pelon, F.; Bourachot, B.; Cardon, M.; Sirven, P.; Magagna, I.; Fuhrmann, L.; Bernard,
C.; et al. Fibroblast Heterogeneity and Immunosuppressive Environment in Human Breast Cancer. Cancer Cell
2018
,33, 463–479.
[CrossRef] [PubMed]
239.
Pelon, F.; Bourachot, B.; Kieffer, Y.; Magagna, I.; Mermet-Meillon, F.; Bonnet, I.; Costa, A.; Givel, A.-M.; Attieh, Y.; Barbazan, J.; et al.
Cancer-associated fibroblast heterogeneity in axillary lymph nodes drives metastases in breast cancer through complementary
mechanisms. Nat. Commun. 2020,11, 404. [CrossRef]

Cancers 2022,14, 3908 34 of 35
240.
Sebastian, A.; Hum, N.R.; Martin, K.A.; Gilmore, S.F.; Peran, I.; Byers, S.W.; Wheeler, E.K.; Coleman, M.A.; Loots, G.G. Single-Cell
Transcriptomic Analysis of Tumor-Derived Fibroblasts and Normal Tissue-Resident Fibroblasts Reveals Fibroblast Heterogeneity
in Breast Cancer. Cancers 2020,12, 1307. [CrossRef] [PubMed]
241.
Bartoschek, M.; Oskolkov, N.; Bocci, M.; Lövrot, J.; Larsson, C.; Sommarin, M.; Madsen, C.D.; Lindgren, D.; Pekar, G.; Karlsson,
G.; et al. Spatially and functionally distinct subclasses of breast cancer-associated fibroblasts revealed by single cell RNA
sequencing. Nat. Commun. 2018,9, 5150. [CrossRef] [PubMed]
242.
Zheng, S.; Zou, Y.; Tang, Y.; Yang, A.; Liang, J.-Y.; Wu, L.; Tian, W.; Xiao, W.; Xie, X.; Yang, L.; et al. Landscape of cancer-
associated fibroblasts identifies the secreted biglycan as a protumor and immunosuppressive factor in triple-negative breast
cancer. OncoImmunology 2022,11, 2020984. [CrossRef] [PubMed]
243.
Chen, Z.; Zhou, L.; Liu, L.; Hou, Y.; Xiong, M.; Yang, Y.; Hu, J.; Chen, K. Single-cell RNA sequencing highlights the role of
inflammatory cancer-associated fibroblasts in bladder urothelial carcinoma. Nat. Commun. 2020,11, 5077. [CrossRef] [PubMed]
244.
Galbo, P.M., Jr.; Zang, X.; Zheng, D. Molecular Features of Cancer-associated Fibroblast Subtypes and their Implication on Cancer
Pathogenesis, Prognosis, and Immunotherapy Resistance. Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res.
2021
,27, 2636–2647.
[CrossRef] [PubMed]
245.
Miyazaki, Y.; Oda, T.; Inagaki, Y.; Kushige, H.; Saito, Y.; Mori, N.; Takayama, Y.; Kumagai, Y.; Mitsuyama, T.; Kida, Y.S. Adipose-
derived mesenchymal stem cells differentiate into heterogeneous cancer-associated fibroblasts in a stroma-rich xenograft model.
Sci. Rep. 2021,11, 4690. [CrossRef] [PubMed]
246.
Zhu, K.; Cai, L.; Cui, C.; de los Toyos, J.R.; Anastassiou, D. Single-cell analysis reveals the pan-cancer invasiveness-associated
transition of adipose-derived stromal cells into COL11A1-expressing cancer-associated fibroblasts. PLoS Comput. Biol.
2021
,
17, e1009228. [CrossRef]
247.
Miyazaki, Y.; Oda, T.; Mori, N.; Kida, Y.S. Adipose-derived mesenchymal stem cells differentiate into pancreatic cancer-associated
fibroblasts in vitro. FEBS Open Bio 2020,10, 2268–2281. [CrossRef] [PubMed]
248.
Hillers-Ziemer, L.E.; Kuziel, G.; Williams, A.E.; Moore, B.N.; Arendt, L.M. Breast cancer microenvironment and obesity:
Challenges for therapy. Cancer Metastasis Rev. 2022. [CrossRef] [PubMed]
249.
Geng, X.; Chen, H.; Zhao, L.; Hu, J.; Yang, W.; Li, G.; Cheng, C.; Zhao, Z.; Zhang, T.; Li, L.; et al. Cancer-Associated Fibroblast
(CAF) Heterogeneity and Targeting Therapy of CAFs in Pancreatic Cancer. Front. Cell Dev. Biol. 2021,9, 655152. [CrossRef]
250.
Chen, P.Y.; Wei, W.F.; Wu, H.Z.; Fan, L.S.; Wang, W. Cancer-Associated Fibroblast Heterogeneity: A Factor That Cannot Be Ignored
in Immune Microenvironment Remodeling. Front. Immunol. 2021,12, 671595. [CrossRef] [PubMed]
251.
Bochet, L.; Lehuédé, C.; Dauvillier, S.; Wang, Y.Y.; Dirat, B.; Laurent, V.; Dray, C.; Guiet, R.; Maridonneau-Parini, I.; Le Gonidec,
S.; et al. Adipocyte-derived fibroblasts promote tumor progression and contribute to the desmoplastic reaction in breast cancer.
Cancer Res. 2013,73, 5657–5668. [CrossRef] [PubMed]
252.
Song, Y.H.; Warncke, C.; Choi, S.J.; Choi, S.; Chiou, A.E.; Ling, L.; Liu, H.Y.; Daniel, S.; Antonyak, M.A.; Cerione, R.A.; et al. Breast
cancer-derived extracellular vesicles stimulate myofibroblast differentiation and pro-angiogenic behavior of adipose stem cells.
Matrix Biol. J. Int. Soc. Matrix Biol. 2017,60–61, 190–205. [CrossRef]
253.
Mao, X.; Xu, J.; Wang, W.; Liang, C.; Hua, J.; Liu, J.; Zhang, B.; Meng, Q.; Yu, X.; Shi, S. Crosstalk between cancer-associated
fibroblasts and immune cells in the tumor microenvironment: New findings and future perspectives. Mol. Cancer
2021
,20, 131.
[CrossRef]
254.
Jiang, H.; Hegde, S.; DeNardo, D.G. Tumor-associated fibrosis as a regulator of tumor immunity and response to immunotherapy.
Cancer Immunol. Immunother. 2017,66, 1037–1048. [CrossRef] [PubMed]
255.
Barrett, R.L.; Puré, E. Cancer-associated fibroblasts and their influence on tumor immunity and immunotherapy. Elife
2020
,
9, e57243. [CrossRef] [PubMed]
256.
Shi, Y.; Gao, W.; Lytle, N.K.; Huang, P.; Yuan, X.; Dann, A.M.; Ridinger-Saison, M.; DelGiorno, K.E.; Antal, C.E.; Liang, G.; et al.
Targeting LIF-mediated paracrine interaction for pancreatic cancer therapy and monitoring. Nature
2019
,569, 131–135. [CrossRef]
[PubMed]
257.
Schwertfeger, K.L.; Cowman, M.K.; Telmer, P.G.; Turley, E.A.; McCarthy, J.B. Hyaluronan, Inflammation, and Breast Cancer
Progression. Front. Immunol. 2015,6, 236. [CrossRef] [PubMed]
258.
Caon, I.; Bartolini, B.; Parnigoni, A.; Caravà, E.; Moretto, P.; Viola, M.; Karousou, E.; Vigetti, D.; Passi, A. Revisiting the hallmarks
of cancer: The role of hyaluronan. Semin. Cancer Biol. 2020,62, 9–19. [CrossRef] [PubMed]
259.
Shiga, K.; Hara, M.; Nagasaki, T.; Sato, T.; Takahashi, H.; Takeyama, H. Cancer-Associated Fibroblasts: Their Characteristics and
Their Roles in Tumor Growth. Cancers 2015,7, 2443–2458. [CrossRef]
260.
Tran, L.L.; Dang, T.; Thomas, R.; Rowley, D.R. ELF3 mediates IL-1
α
induced differentiation of mesenchymal stem cells to
inflammatory iCAFs. Stem Cells 2021,39, 1766–1777. [CrossRef]
261.
Calvo, F.; Ege, N.; Grande-Garcia, A.; Hooper, S.; Jenkins, R.P.; Chaudhry, S.I.; Harrington, K.; Williamson, P.; Moeendarbary, E.;
Charras, G.; et al. Mechanotransduction and YAP-dependent matrix remodelling is required for the generation and maintenance
of cancer-associated fibroblasts. Nat. Cell Biol. 2013,15, 637–646. [CrossRef] [PubMed]
262.
Sharma, M.; Turaga, R.C.; Yuan, Y.; Satyanarayana, G.; Mishra, F.; Bian, Z.; Liu, W.; Sun, L.; Yang, J.; Liu, Z.-R. Simultaneously
targeting cancer-associated fibroblasts and angiogenic vessel as a treatment for TNBC. J. Exp. Med.
2021
,218, e20200712.
[CrossRef] [PubMed]

Cancers 2022,14, 3908 35 of 35
263.
Bughda, R.; Dimou, P.; D’Souza, R.R.; Klampatsa, A. Fibroblast Activation Protein (FAP)-Targeted CAR-T Cells: Launching an
Attack on Tumor Stroma. ImmunoTargets Ther. 2021,10, 313–323. [CrossRef]
264.
Haubeiss, S.; Schmid, J.O.; Mürdter, T.E.; Sonnenberg, M.; Friedel, G.; van der Kuip, H.; Aulitzky, W.E. Dasatinib reverses
cancer-associated fibroblasts (CAFs) from primary lung carcinomas to a phenotype comparable to that of normal fibroblasts. Mol.
Cancer 2010,9, 168. [CrossRef] [PubMed]
265.
Saw, P.E.; Chen, J.; Song, E. Targeting CAFs to overcome anticancer therapeutic resistance. Trends Cancer
2022
,8, 527–555.
[CrossRef]
266.
Scioli, M.G.; Artuso, S.; D’Angelo, C.; Porru, M.; D’Amico, F.; Bielli, A.; Gentile, P.; Cervelli, V.; Leonetti, C.; Orlandi, A.
Adipose-derived stem cell-mediated paclitaxel delivery inhibits breast cancer growth. PLoS ONE
2018
,13, e0203426. [CrossRef]
267.
Luca, A.C.; Mersch, S.; Deenen, R.; Schmidt, S.; Messner, I.; Schäfer, K.L.; Baldus, S.E.; Huckenbeck, W.; Piekorz, R.P.; Knoefel,
W.T.; et al. Impact of the 3D microenvironment on phenotype, gene expression, and EGFR inhibition of colorectal cancer cell lines.
PLoS ONE 2013,8, e59689. [CrossRef]