As most tumours present self antigens, peripheral toler-
ance has an important role in contributing to immune
evasion by tumours. In addition, the overproduction of
immunosuppressive cytokines, including transforming
growth factor-β (TGFβ), by tumour cells and tumour-
infiltrating leukocytes also contributes to an immuno-
suppressive microenvironment. Many studies indicate
that TGFβ can promote cancer metastasis through
effects on the tumour microenvironment by enhanc-
ing tumour cell invasion and by inhibiting the func-
tion of immune cells1,2. These findings have promoted
interest in targeting TGFβ and its signalling pathway
in patients with cancer. However, such targeting of
TGFβ could result in adverse effects in normal tissues,
as TGFβ-induced signalling is also involved in many
homeostatic processes (FIG. 1). For example, TGFβ can
function as a tumour suppressor to prevent tumorigen-
esis; however, overproduction of TGFβ is frequently
associated with tumour metastasis and poor prognosis
in patients with cancer (FIG. 1). Although the molecu-
lar mechanisms behind this dichotomy of TGFβ func-
tions are not fully elucidated, progress has been made
in understanding the role of TGFβ in different stages of
cancer. This topic has recently been reviewed1,3–5 and is
not discussed here.
This Review focuses on the tumour-promoting
properties of TGFβ, which prevent effective antitumour
immune responses once cancer has been established
in the host. A successful immune response requires
the proper activation and maturation of antigen-
presenting cells (APCs) of the innate immune system
that present antigen to adaptive immune cells. TGFβ
can suppress or alter the activation, maturation and
differentiation of both innate and adaptive immune
cells, including natural killer (NK) cells, dendritic cells
(DCs), macrophages, neutrophils, and CD4+ and CD8+
T cells6. A dampened innate immune response leads
to poor adaptive immunity, resulting in persistence of
the tumour. In addition, TGFβ has an important role
in the differentiation and induction of natural and
induced regulatory T (TReg) cells, which also contrib-
ute to the tolerizing environment. Furthermore, in
the presence of interleukin-6 (IL-6), TGFβ induces
the differentiation of IL-17-producing CD4+ T helper
17 (TH17) cells and CD8+ cytotoxic T cells; however,
the role of IL-17-producing cells in tumour biology
still remains controversial, given that these cells can
have both tumour-promoting and antitumour activi-
ties7. As we discuss, many recent discoveries have been
made towards understanding the biological effects of
TGFβ on different immune cells, although multiple
areas require further investigation. Finally, there is
compelling evidence to support targeting TGFβ with
inhibitors to enhance antitumour immunity in patients
Effects of TGFβ on innate immune cells
NK cells. NK cells are innate lymphoid cells that have an
important role in the antitumour response by recogniz-
ing and directly killing primary tumours and metastases
to the lungs8, as well as rapidly producing chemokines
and cytokines crucial for these functions. For example,
*Yale University School of
Medicine, 300 Cedar Street,
TAC S-569, PO BOX 208011,
New Haven, Connecticut
‡Present address: Gladstone
Institute of Virology and
Immunology, 1650 Owens
Street, San Francisco,
California 94158, USA, and
Department of Microbiology
and Immunology, University
of California, San Francisco,
California 94143-1230, USA.
§Present address: Veteran’s
Health Care System and
Center, 950 Campbell
Avenue, West Haven,
Connecticut 06516, USA.
Correspondence to R.A.F.
The polarization of immune cells in the
tumour environment by TGFβ
Richard A. Flavell*, Shomyseh Sanjabi*‡, Stephen H. Wrzesinski*§ and
Abstract | Transforming growth factor-β (TGFβ) is an immunosuppressive cytokine produced
by tumour cells and immune cells that can polarize many components of the immune system.
This Review covers the effects of TGFβ on natural killer (NK) cells, dendritic cells,
macrophages, neutrophils, CD8+ and CD4+ effector and regulatory T cells, and NKT cells in
animal tumour models and in patients with cancer. Collectively, many recent studies favour
the hypothesis that blocking TGFβ-induced signalling in the tumour microenvironment
enhances antitumour immunity and may be beneficial for cancer therapy. An overview of the
current drugs and reagents available for inhibiting TGFβ-induced signalling and their phase
in clinical development is also provided.
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Premalignant stateMalignant progressionMetastasis
TGF??= tumour suppressor
TGF??= tumour promoter
TGF??= tumour supporter
? Tumorigenic inflammation
? Production of stromal-
The lack of responsiveness of
mature lymphocytes in the
periphery to specific self
antigens. These mechanisms
can control potentially
self-reactive lymphocytes that
have escaped central tolerance
or prevent immune responses
to specialized self proteins that
were not present during
establishment of central
tolerance. Peripheral tolerance
is associated with suppression
of self-reactive antibody
production by B cells and
inhibition of self-reactive
effector cells, such as T helper
cells and cytotoxic T
(MDSCs). A subset of immature
CD11b+GR1+ cells (which
include precursors of
dendritic cells and myeloid
cells) that are produced in
response to various
These cells have been shown to
antigen-specific CD8+ T cell
interferon-γ (IFNγ) production by NK cells is impor-
tant for stimulating effector CD4+ TH1 cells that are
required for clearing tumours. TGFβ attenuates IFNγ
production by NK cells and their lytic activity9,10. These
might be direct effects of TGFβ or might result indirectly
from cell–cell contact between NK cells and regulatory
T (TReg) cells producing this cytokine11. In support of
a direct effect, TGFβ can suppress IFNγ production
through SMAD3, a transcription factor downstream
from TGFβ-induced signalling, which results in sup-
pression of T-bet, a transcription factor required for
Targeted killing by NK cells requires stimulating
the NK cell activating receptors natural killer group 2,
member D (NKG2D), NKp46, NKp44 and NKp30
(REF. 13). It has been shown that exogenously admin-
istered TGFβ inhibits NKp30 and NKG2D expression,
leading to decreased ability of NK cells to kill target
cells14. TGFβ also decreases expression of NKG2D by
NK cells and CD8+ T cells from patients with glioma
who have a high tumour burden15. In patients with lung
and colorectal cancer, the downregulation of NKG2D
expression has been associated with increased serum
levels of TGFβ16. Furthermore, recent studies of iso-
lated NK cells from healthy donors have shown that
platelet-derived TGFβ results in downregulation of
NKG2D expression, causing a decrease in IFNγ pro-
duction and of the degranulation functions that are
essential for tumour destruction by these cells17. Finally,
surface-bound TGFβ on myeloid-derived suppressor cells
(MDSCs) can inhibit NK cell cytolytic activity in an
orthotopic liver cancer model18. These observations
indicate that TGFβ has immunosuppressive effects on
NK cell killing functions in patients with cancer and,
therefore, might be a target for enhancing NK cell-
mediated antitumour immune responses.
Dendritic cells. DCs are APCs that have a crucial role
in the initial activation and subsequent regulation of
immune responses19. In addition to activating T cell-
mediated adaptive immunity, DCs can also activate NK
cells20. DCs can present antigen in an immunogenic
or tolerogenic manner, and so they have an important
role in determining the host response to tumours19,21.
DC activation involves the upregulation of MHC and
co-stimulatory molecule expression, alteration in
motility and the formation of dendrites to increase the
surface area for antigen presentation and interaction
with lymphocytes22. Non-activated or immature DCs
can still present antigen but, in the absence of proper
co-stimulation, this results in T cell tolerance23,24. In
the presence of immune-inhibiting signals, such as
IL-10 and/or corticosteroids, DCs can induce tolerance
by T cell deletion and/or the activation and induction
of TReg cells25,26. So, DCs can induce either immunity or
peripheral tolerance and are an essential determinant
of antitumour immunity.
TGFβ affects DC biology in several ways: TGFβ can
immobilize DCs, thereby interfering with their migra-
tion and the transport of antigen to draining lymph
nodes for presentation to adaptive immune cells, and it
might also directly induce DC apoptosis27–29. Tumour-
infiltrating DCs both secrete and respond to TGFβ,
in either an autocrine or paracrine manner, by down-
regulating expression of MHC class II molecules, the
co-stimulatory molecules CD40, CD80 and CD86, and
production of tumour necrosis factor (TNF), IFNα,
IL-12 and CC-chemokine ligand 5 (CCL5)6,30. These
immature or tolerogenic DCs promote the formation
of TReg cells that potently inhibit the function of other
T cells31,32. Activated DCs can also activate both natu-
ral and induced TReg cells33–36; interestingly, the capac-
ity of DCs to induce both types of TReg cell is greatly
Figure 1 | The yin and yang of TGFβ in tumour development, maintenance and metastasis formation. Before
epithelial cells transform into a malignant tumour, transforming growth factor-β (TGFβ) functions as a tumour suppressor
by blocking the expression of stromal-derived mitogens and suppressing pro-tumorigenic inflammation. Furthermore,
TGFβ supports the cytostasis, terminal differentiation and apoptosis of premalignant cells, which have either an
overexpressed oncogene or suppressed tumour-suppressor gene. Once the epithelial cells become fully malignant, TGFβ
has the opposite effect by blocking the antitumour immune response through support for the activity of regulatory cells
and through direct inhibition of effector cell mechanisms from clearing the established tumour, as described in the main
text and summarized in FIG. 2. Once a tumour is established, TGFβ further supports the formation of metastases to several
sites, including bone and lung tissues. Additional non-immune mechanisms, outside the scope of this Review, that support
tumorigenesis and metastasis formation are addressed in REFS 1, 3.
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(TAMs). An important
component of the tumour
microenvironment. These cells
differentiate from circulating
blood monocytes that have
infiltrated tumours. These cells
can have positive or negative
effects on tumorigenesis (that
is, tumour promotion or
A classically activated
macrophage that is stimulated
by Toll-like receptor ligands
(such as lipopolysaccharide)
and IFNγ and that expresses,
among others, inducible
nitric-oxide synthase and
An alternatively activated
macrophage that is stimulated
by IL-4 or IL-13 and that
expresses arginase 1, the
mannose receptor CD206 and
the IL-4 receptor α-chain.
increased by TGFβ and IL-10 (REFS 37,38). So, in the
context of these immunosuppressive cytokines in the
tumour microenvironment, DCs take up tumour cells,
become tolerogenic TGFβ-secreting cells and promote
the induction of tumour-specific TReg cells in both mice
and humans39–42. Therefore, a growing body of evidence
indicates that TReg cells are induced by TGFβ produced
by DCs; as TReg cells are one of the main obstacles to
successful antitumour immunity, this indicates that the
TGFβ pathway might be targeted to enhance antitumour
immunity in patients with cancer.
Macrophages. The mass of most solid tumours is made
up of a large number of macrophages and, in general,
high numbers of tumour-associated macrophages (TAMs)
correlate with poor cancer prognosis; however, depend-
ing on the type and environment of the tumour, TAMs
may also have antitumour activity43,44. Macrophages are
a heterogeneous population and are typically defined as
being of a classically activated M1 macrophage pheno-
type or alternatively activated M2 macrophage pheno-
type, similar to the CD4+ TH1 versus TH2 cell paradigm.
M1 macrophages are induced by IFNγ and other pro-
inflammatory stimuli and are efficient at presenting anti-
gens, producing pro-inflammatory cytokines, activating
TH1 cell responses, and in general mediating antitumour
responses. By contrast, M2 macrophages with various
phenotypes are induced by IL-10, immune complexes,
glucocorticoids, IL-4 and IL-13 that are involved in
remodelling and repair of damaged tissue, parasite resist-
ance, immune regulation and/or tumour promotion45.
MDSCs are a heterogeneous population of immature
DCs, macrophages, granulocytes and other myeloid cells
in early stages of their differentiation and have proper-
ties similar to those that have been described for M2
macrophages. In late stages of tumour development,
TAMs and MDSCs can produce TGFβ and are classically
involved in cancer progression and metastasis46. It is not
clear whether TGFβ is directly involved in converting
TAMs from an M1 to M2 phenotype or whether by pro-
ducing large amounts of TGFβ TAMs may contribute
to the general immunosuppressive tumour microenvi-
ronment. In skin cancer, TGFβ-mediated recruitment
of macrophages into tumours has an important role
in immune escape, as it converts a regressing tumour
into a progressing tumour47. It is suggested that TGFβ-
recruited TAMs are highly phagocytic and can compete
with DC function, thereby markedly decreasing the abil-
ity of DCs to present tumour antigens to the adaptive
immune system47. Interestingly, the M2 phenotype of
TAMs is mediated by the inhibitory nuclear factor-κB
(NF-κB) subunit p50 and it is thought that the cytokine
milieu of the tumour microenvironment is necessary
to maintain the phenotype48–50. During tumour pro-
gression, the gradual inhibition of NF-κB activity in
TAMs correlates with their switch from an M1 to M2
phenotype51. Peritoneal macrophages from tumour-
bearing hosts produce increased levels of TGFβ, are less
differentiated and have deficiencies in inflammatory
cytokine production owing to decreased expression of
NF-κB and CCAAT/enhancer binding protein (C/
EBP) transcription factors52. In this model, the tumour-
derived factors TGFβ and prostaglandin E2 individually
and additively downregulate NF-κB and C/EBP expres-
sion. There is also evidence that tumour-infiltrating
MDSCs secrete high levels of TGFβ, which upregulates
CD206 (a deactivation marker characteristic of M2
macrophages) expression in an autocrine manner53. It
is unclear how TGFβ production is induced in MDSCs
and macrophages, but the mechanism might involve
IL-13 and glucocorticoids54,55. It is conceivable that the
excess amount of TGFβ in the tumour microenviron-
ment may contribute to the alternative activation of M2
macrophages by downregulating NF-κB expression.
Neutrophils. Neutrophils are short-lived polymor-
phonuclear leukocytes with potent antimicrobial and
inflammatory capacities. Despite their known func-
tion as professional phagocytes, the role of neutrophils
in tumour progression has been controversial and has
received little attention compared with that of macro-
phages. Initial studies characterizing the effect of TGFβ
on the control of inflammatory responses showed that
it was a potent chemotactic factor for neutrophils, pro-
moting their migration but not degranulation or acti-
vation6. Subsequent studies showed that neutrophil
migration could also be indirectly affected by TGFβ
through regulating the expression of adhesion mol-
ecules in the endothelium56,57 and that TGFβ could
inhibit neutrophil cytotoxicity, suggesting that TGFβ
might influence human neutrophil activity in vivo58.
Recently, the contradictory role of neutrophils in both
tumour suppression and tumour promotion by directly
or indirectly controlling tumour growth, angiogenesis
and metastasis (reviewed in REF. 59) was re-evaluated
in terms of the characterization of different types of
tumour-associated neutrophil (TAN) with polarized
N1 or N2 phenotypes60. These polarized populations
are similar to those that have been described for mac-
rophages; their phenotypic development is influenced
by the microenvironment and seems to be controlled
by TGFβ in the tumour proximity. N2 neutrophils are
characterized by an expression profile that promotes
tumour angiogenesis and metastasis61–63. Depletion of
this N2 neutrophil subpopulation in untreated tumour-
bearing mice was sufficient to inhibit tumour growth,
even when CD8+ T cells were absent, highlighting the
immunosuppressive potential of N2 cells60–62. Under
TGFβ-inhibiting conditions, as well as in response to
certain activation signals, neutrophils acquire an anti-
tumour N1 phenotype that promotes tumour cell death
and inhibits tumour growth59,60,64,65. The lack of systemic
effects on neutrophil polarization during TGFβ neutrali-
zation experiments indicated that the effect is mainly
intratumoral, characterized by increased numbers of N1
TANS that express activating chemokines and cytokines,
as well as by changes in endothelial adhesion molecule
expression60. Interestingly N1 and N2 neutrophils were
shown to control the activation status of CD8+ T cells.
This interplay seemed to be reciprocal as activated
CD8+ T cells could control the activation and migra-
tion of neutrophils to the tumour microenvironment as
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well66. Clearly a re-evaluation of the role of neutrophils
in tumour immunology, as well as characterization of
the subpopulation polarized by TGFβ, may be required
to design more effective immunotherapies.
Effects of TGFβ on effector T cells
CD8+ T cells. CD8+ T cells are a crucial component of
antitumour immunity, as tumour antigen-specific cyto-
toxic T lymphocytes (CTLs) have an important role
in the cytolytic killing of tumour cells. Several studies
have shown a direct correlation between the frequency
of CTLs in tumour infiltrating lymphocytes (TILs) and
the overall survival of cohorts of patients with cancer;
in particular, a high ratio of intratumoral activated
cytotoxic CD8+ T cells to TReg cells leads to improved
prognosis67–70. TGFβ-induced signalling in tumour-
specific CTLs dampens their function and frequency
in the tumour71, and blocking TGFβ-induced signal-
ling on CD8+ T cells results in more rapid tumour sur-
veillance and the presence of many more CTLs at the
tumour site72. The co-engagement of the T cell receptor
(TCR) and TGFβ receptor has been shown to increase
CD103 expression on TILs, which might be required for
the activity and retention of CTLs in the tumour micro-
environment; although the function of this integrin on
CTLs is not fully understood73.
Several experimental protocols have been used to
render CD8+ T cells unresponsive to TGFβ. In a model
where a dominant-negative form of TGFβ receptor II
(TGFβRII) is expressed by both CD4+ and CD8+ T cells,
a strong antitumour immune response was associated
with the proliferation and increased activity of tumour-
specific CTLs74,75. Similarly, when tumour-specific CD8+
T cells are rendered unresponsive to TGFβ-induced sig-
nalling by transduction with a similar TGFβRII domi-
nant-negative construct before adoptive transfer, these
cells infiltrate into tumours, secrete cytokines such as
IFNγ and can successfully kill tumour cells76,77.
In some tumour models, systemic blockade of TGFβ,
using a monoclonal antibody or kinase inhibitors to block
downstream signalling, prevents tumour recurrence by
affecting the activity of various cell types, including an
increase in the cytotoxic activity of CTLs78–81. However,
the inhibition of TGFβ using a monoclonal antibody
alone is not always sufficient to promote tumour rejec-
tion in all animal tumour models. In such models, the
combination of a TGFβ-specific antibody with a vac-
cine resulted in a synergistic improvement in the inhi-
bition of tumour growth that is mediated by increased
number and activity of CD8+ T cells82–84. It is speculated
that in these models, the source of inhibitory TGFβ is
the immune system itself and not the tumour, because
the antibody-mediated blockade is effective at enhanc-
ing antitumour immune responses even when antibody
is administered with a prophylactic vaccine before injec-
tion of tumour cells82. This effect of TGFβ is consistent
with the recent finding that TGFβ is responsible for the
apoptosis of short-lived effector T cells that comprise
more than 90% of the effector pool after immuniza-
tion with Listeria monocytogenes85. TGFβ promotes the
apoptosis of these effector T cells by downregulating
the expression of B cell lymphoma 2 (BCL-2), which
opposes the survival function of IL-15 on the short-lived
effector T cell population85. It is possible that blocking
TGFβ-induced signalling with the neutralizing antibody
during administration of the tumour vaccine inhibits the
apoptosis of tumour-specific short-lived effector CD8+
T cells and therefore prevents the termination of CTL
TGFβ-mediated inhibition of CTL function dur-
ing antitumour immunity might be through several
mechanisms. TGFβ directly inhibits CTL function by
suppressing the expression of several cytolytic genes,
including the genes encoding granzyme A, granzyme B,
IFNγ and FAS ligand71. TGFβ also attenuates the effec-
tor function of antigen-specific memory CD8+ T cells
by inhibiting T-bet expression, resulting in inhibition of
IFNγ production86. TGFβ might also block TCR signal-
ling of TILs by upregulating the expression of SPRED1
(sprouty-related, EVH1 domain containing 1), which
is an inhibitor of the RAS–MAPK (mitogen-activated
protein kinase) pathway87. Interestingly, TGFβ can also
influence CD8+ T cell-mediated antitumour immunity
by inducing IL-17 production by CD8+ T cells, although
the effect of IL-17 on tumour growth versus immune
surveillance remains controversial88–91.
CD4+ T cells. CD4+ T cells are central orchestrators of
adaptive immunity; however, their role in antitumour
immune responses has largely been overlooked, mainly
owing to the lack of MHC class II expression by most
forms of solid cancer. TGFβ has been shown to have
effects on all subsets of CD4+ effector T cells by control-
ling the expression of master transcriptional regulators in
these cells. TGFβ inhibits T-bet and GATA-binding pro-
tein3 (GATA3) expression (which determine CTL, TH1
and TH2 cell differentiation), whereas it promotes fork-
head box P3 (FOXP3) and retinoic acid receptor-related
orphan receptor-γt (RORγt) expression (which determine
TReg and TH17 cell differentiation, respectively) (reviewed
in REF. 92). The role of CD4+ T cells in tumour biology has
been classically studied in the context of TReg cells, which
are covered in a separate section of this Review. This sec-
tion focuses on the specific role of TH cell subpopulations
in the control of antitumour immune responses and how
TGFβ in the tumour microenvironment could influence
the polarization of each subset.
Early studies trying to understand the mechanism of
tumour-induced immunosuppression93 identified TGFβ
as one of the main inhibitors of immune responses in the
tumour microenvironment. Tumour-derived TGFβ was
shown to inhibit TH1 cell responses by shifting infiltrat-
ing T cells towards a TH2 cell phenotype94, and hence
TGFβ promoted a less efficient antitumour immune
response. However, later studies comparing the effi-
cacy of TH1 and TH2 effector cell subsets in mediating
antitumour immunity showed that both TH1 and TH2
cells increased the CTL-mediated antitumour response,
although TH1 cells secreting IFNγ seemed to be more
effective by promoting APC activation95,96. Studies using
TCR-transgenic mice further support the requirement
for CD4+ T cells to activate memory CTLs in vivo97, and
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interestingly show tumour eradication by CD4+ T cells
even in cases where tumours were resistant to CD8+
T cell-mediated rejection98. These findings suggested a
potential benefit of TH cells in cancer immunotherapy
and started a search for the most effective antitumour
CD4+ effector T cell population.
Contradictory reports regarding the role of IL-17 in
cancer have made it difficult to conclude whether or
not T cells expressing this cytokine would be beneficial
against tumours99,100. Accumulation of TH17 cells in the
tumour microenvironment has been reported in several
types of cancer, as well as the expression of IL-6, IL-1β
and TGFβ by tumour cells, which are key cytokines
controlling TH17 cell differentiation and prolifera-
tion101,102. TH17-polarized tumour-specific CD4+ T cells
were shown to be more efficient than TH1-polarized
cells in tumour rejection after adoptive transfer, and
this efficiency was probably dependent on IFNγ rather
than IL-17 production90. Similar observations from
transferring CD8+IL-17+ cells, which then become
IFNγ-producing cells, were reported; however, some
discrepancies have been found regarding the role of
IFNγ in these models, as the use of lymphopenic hosts
promotes loss of a TH17 cell phenotype and acquisition
of a TH1 cell phenotype in the transferred cells, poten-
tially masking the real effects of IL-17 in controlling
Recent findings indicate that the differentiation
state of T cells, in terms of naive versus effector or
memory cells, might also be important for mounting
more efficient antitumour responses as single transfers
of naive CD4+ T cells were able to eradicate established
tumours independently of CD8+ T cells, NK cells and
Growing evidence suggests that control of the
cytokines expressed in the tumour microenvironment
can promote tumour eradication by controlling TReg and
TH17 cell polarization in the tumour. Exogenous admin-
istration of IL-2 in tumour-bearing mice increased TReg
cell frequencies and decreased TH17 cell frequencies in
the tumour106, whereas antagonizing the effects of TGFβ
by administering IL-7 has been shown to be useful for
the promotion of TH17 cells107. A complete understand-
ing of the dynamic cytokine network, including the role
of TGFβ, in controlling T cell polarization in tumours, as
well as characterization of the molecular signals mediat-
ing TH cell differentiation, is crucial for dissecting the
beneficial use of TH cells in future immunotherapies
Effects of TGFβ on regulatory cells
CD4+ TReg cells. TReg cells are an immunosuppressive
T cell population that express the transcription fac-
tor FOXP3 and can suppress antitumour immune
responses108. These cells form a heterogeneous popu-
lation containing at least two distinct subsets known
as natural TReg (nTReg) cells and adaptive or induced
TReg (iTReg) cells108. nTReg cells develop in the thymus,
express the IL-2 receptor α-chain (CD25) and maintain
self tolerance in an antigen-independent manner. iTReg
cells, by contrast, develop in the periphery in response
to self or tumour antigens and express variable levels of
CD25 (REF. 108). Although nTReg cells and iTReg cells have
been identified as separate subsets of TReg cells, their
phenotype and function have not been fully established
in tumour-bearing animal models and patients with
cancer. TGFβ could be involved in generating TReg cells
in vivo and this cytokine may help subsets of TReg cells
to suppress effector cell function in the tumour micro-
environment (reviewed in REF. 2 and covered in more
detail in the section detailing effects of TGFβ on effec-
tor cells). Large numbers of TReg cells in patients with
cancer can be inversely correlated with survival109,110.
Although the precise mechanism (or mechanisms)
causing increased numbers of TReg cells in malignan-
cies is unknown, TGFβ — as well as other tumour-
produced chemical mediators working together with
this cytokine, such as prostaglandin E2 and H-ferritin
— has been implicated in inducing TReg cells111,112. In
addition, the production of CCL22 by TAMs surround-
ing tumours might mediate TReg cell trafficking to the
tumour bed through CCR4 (REF. 113). Recently, IL-23
production in the tumour microenvironment has been
implicated in promoting the proliferation of intratu-
moral TReg cells, as these cells express IL-23R, have evi-
dence of STAT3 activation (downstream of IL-23R) and
are decreased in number in tumour-bearing animals
treated with a blocking antibody specific for IL-23R26.
IL-23 may complement the effects of TGFβ, which
also seems to increase the number of intratumoral
Recently, a new regulatory T cell subtype has
been identified that can be induced and expanded
in mice bearing orthotopic liver, lung and melanoma
tumours114. Unlike the conventional TReg cells described
above, this regulatory subtype lacks expression of
CD25 and FOXP3. Instead, the cells express the IL-2R
β-chain (also known as CD122), IL-10, TGFβ1 and the
early activation marker CD69 (REF. 114). Activation of
CD69 with an agonist antibody results in high levels
of membrane-bound TGFβ expression by these cells
through the activation of extracellular signal-regulated
kinase (ERK), and this might contribute to the ability
of this regulatory T cell subset to suppress CD4+ T cell
proliferation and promote the growth of established
tumours114. Although these results suggest that another
subset of regulatory T cells associated with TGFβ pro-
duction can suppress the antitumour immune response,
the role of these cells in patients with cancer remains
to be determined.
TGFβ, in combination with IL-2, is required for the
conversion of naive T cells to iTReg cells in vitro115,116.
Furthermore, the induction of TReg cells by TGFβ might
be a mechanism by which tumours escape the antitu-
mour immune response as several tumours can produce
TGFβ2. Blockade of TGFβ-induced signalling with anti-
bodies or genetic manipulation leads to decreased num-
bers of iTReg cells in some tumour-bearing animals40,117.
Therefore, targeting TGFβ-induced signalling in the
tumour microenvironment could attenuate the immu-
nosuppressive effects of iTReg cells, resulting in increased
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Increased suppressive activity
? Migration and/or recruitment
? NF-?B activation and cytokine
? M1 to M2 phenotype differentiation
Decreased tumour cell killing
? IFN? production
? NKp30 and NKG2D expression
Decreased antigen presentation
? Cytokine production
? Induction of TReg cell
? Production of reactive
? N1 to N2 phenotype
CD8+ regulatory T cells. CD8+ T cells can become sup-
pressor cells similar to CD4+ TReg cells, and TGFβ can
induce CD8+ T cells to express FOXP3 (REFS 118,119).
CD8+ regulatory T cells are induced under immuno-
suppressive conditions such as the tumour microen-
vironment120–123. The role of tumour-infiltrating CD8+
regulatory T cells is less well understood than that of
CD4+ TReg cells. It was recently shown that infiltration of
CD8+ T cells into the immunosuppressive microenviron-
ment of prostate tumours can convert tumour-specific
CD8+ effector T cells into regulatory cells, and that this
regulatory activity could be blocked by a TGFβ-specific
antibody121. In another recent study, CD8+CD25+FOXP3+
regulatory T cells were isolated from colorectal cancer tis-
sue and shown to have suppressive activity ex vivo. In this
study, TGFβ and IL-6 induced the generation of CD8+
regulatory T cells in a synergistic manner122. However,
as these CD8+ regulatory T cells constitute only a small
number of total CD8+ T cells in vivo, more investigation
is needed to understand fully how they are induced and
what is their clinical relevance in patients with cancer.
NKT cells. NKT cells are a heterogeneous subset of
T cells that also have properties of NK cells, and thus
bridge the innate and adaptive immune responses. Unlike
other T cells that recognize MHC class I-presented pep-
tides, NKT cells recognize self and foreign glycolipids
presented by the nonclassical MHC class I molecule
CD1d124. There are two main subtypes of NKT cell that
have opposing roles in the antitumour immune response:
invariant NKT cells (iNKT cells; also known as type I
NKT cells) and type II NKT cells.
iNKT cells are defined by their use of a semi-invari-
ant TCR involving Vα14Jα18 in mice and Vα24Jα18 in
humans, and they respond to α-galactosylceramide,
resulting in increased antitumour responses through
IFNγ production that activates CD8+ T cells and NK
cells125,124. Defects in iNKT cells have been identified in
patients with cancer in later stages of the disease and
increased numbers of circulating and intratumoral
iNKT cells have been associated with improved prog-
nosis126. Targeting iNKT cells with activating agents is
being evaluated in clinical trials (reviewed in REF. 125).
TGFβ has been implicated in suppressing iNKT cells in
patients with cancer and a recent evaluation of these cells
from patients with metastatic melanoma and renal cell
carcinoma suggested that blocking TGFβ in vitro could
enhance iNKT cell activation ex vivo127.
Type II NKT cells have diverse repertoires of TCRs
and, in contrast to iNKT cells, suppress the antitumour
response through several mechanisms, including TGFβ
production124. In a mouse fibroblast tumour model, this
subset of NKT cells can express high levels of IL-13,
leading to the production of TGFβ by MDSCs, which
in turn results in attenuated antitumour responses by
CD8+ effector T cells78. However, in a different tumour
model, evaluating antibody-mediated blockade of TGFβ
combined with a peptide vaccine against a lung cancer
tumour line, results suggested that the IL-13 pathway,
enhanced by type II NKT cells, might not be the mecha-
nism behind the enhanced antitumour activity observed
in vaccinated mice83. These results suggest a more com-
plex interplay, which is yet to be determined, between
TGFβ, type II NKT cells and effector immune cells
responsible for the antitumour response.
Successful cancer immunotherapy depends on overcom-
ing the immunosuppressive milieu in the tumour micro-
environment in patients with cancer. TGFβ has a crucial
immunosuppressive role in both the innate and the adap-
tive arms of the immune response (FIGS 2,3). In terms of
the innate immune response, TGFβ inhibits IFNγ pro-
duction by NK cells causing dampened CD4+ TH1 cell
responses. It downregulates expression of the activating
receptor NKG2D on NK cells resulting in decreased
cytolytic activity and overall poor antitumour responses.
TGFβ also influences the presentation of tumour anti-
gens to the adaptive immune system by decreasing DC
migration and promoting DC apoptosis in some tumour
models. In general, TGFβ inhibits DC maturation and
cytokine production, thereby promoting a tolerogenic
environment. In addition, TGFβ produced by tolerogenic
DCs contributes to TReg cell differentiation. TGFβ can also
favour the differentiation of M2 versus M1 macrophages
by inhibiting NF-κB activation. TAMs are a subtype of
M2 cells that are recruited to the tumour by TGFβ and
also produce high levels of TGFβ. TAMs in the tumour
Figure 2 | Effects of TGFβ on innate immune cells. Transforming growth factor-β
(TGFβ) has an inhibitory effect on innate immunity in the tumour microenvironment
through several pathways. It inhibits natural killer (NK) cell function by downregulating
production of interferon-γ (IFNγ) and expression of the activating receptors NKp30 and
natural killer group 2, member D (NKG2D), thereby decreasing NK cell killing activity. In
the presence of TGFβ, dendritic cells (DCs) acquire a tolerogenic phenotype
characterized by decreased migration, maturation and cytokine production and
increased apoptosis. They also gain the ability to induce regulatory T (TReg) cell
differentiation. TGFβ can also convert N1 neutrophils to a N2 phenotype, which is less
cytotoxic. Similarly, TGFβ can promote the recruitment of M2 over M1 macrophages and
of tumour-associated macrophages (TAMs), and it can decrease cytokine production by
these macrophages by inhibiting nuclear factor-κB (NF-κB) activity.
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? GATA3 and NFAT
? T-bet, STAT4
? T-bet, IFN? and
iTReg cell nTReg cell
microenvironment compete with DCs for antigen uptake
but cannot properly present antigen. TGFβ also promotes
the differentiation of N1 to N2 neutrophils, which, similar
to M2 macrophages, are less cytotoxic. So, blocking TGFβ
can induce an expression profile in the tumour microen-
vironment that promotes better antigen uptake and pres-
entation, resulting in more robust priming and activation
of the adaptive antitumour immune response.
In terms of the adaptive immune response, TGFβ can
also directly dampen the function of CD8+ and CD4+
T cells while promoting the recruitment and differen-
tiation of regulatory T cells at the tumour bed (FIG. 3).
This cytokine inhibits the cytotoxic function of tumour
specific CTLs and promotes apoptosis of the short-lived
effector CD8+ T cells. TGFβ also controls the differ-
entiation of several key CD4+ T cell subsets in tumour
immunology, including TH1, TH17 and TReg cell subpopu-
lations. Importantly, the effect of TGFβ on the differ-
entiation of CD4+ T cells is influenced by the cytokine
milieu in the tumour microenvironment, which indi-
cates that modulating the relative abundance of such
factors might promote antitumour responses in vivo.
It is well documented that both TGFβ and regulatory
T cells have key roles in suppressing the antitumour
immune response; however, the precise contributions
of TGFβ and different regulatory T cell subsets in sup-
pressing effector cell function are still being evaluated
(FIG. 4). For example, although TGFβ can induce CD8+
T cells to become regulatory cells expressing FOXP3,
the precise role of CD8+FOXP3+ T cells in antitumour
immunity remains unclear. TGFβ is also implicated in
suppressing antitumour iNKT cell function; however,
the interplay between TGFβ and immunosuppressive
type II NKT cells is less clear. Given that TGFβ can
actively modulate inflammation and tolerance induc-
tion in the many ways described above, TGFβ blockade
might enhance antitumour immunity through effects on
numerous components of the immune response.
Targeting TGFβ-induced signalling for
immunotherapy of cancer [Au: pls edit to 1 line]
The immunosuppressive effects of TGFβ on immune
cell subsets leading to dampened antitumour immune
responses as described above strongly support the devel-
opment of TGFβ inhibitors to treat patients with cancer.
Several inhibitors of TGFβ-induced signalling, summa-
rized in TABLE 1, are in various stages of development,
targeting several steps in the TGFβ-induced signalling
cascade (FIG. 5). Although most of these approaches are in
preclinical studies, several clinical trials (clinicaltrials.gov
identifier: NCT00761280 and clinicaltrials.gov identifier:
NCT00383292) have evaluated TGFβ inhibition in patients
with cancer using an antibody (GC1008; Cambridge
Antibody Technology/Genzyme), blocking oligonucle-
otides (trabedersen (AP12009; Antisense Pharma)), small
molecule inhibitors (LY373636 and LY2157299 (both Eli
Lilly)) and an allogeneic vaccine approach with a tumour
cell line transfected with an antisense construct against
TGFβ2, belagenpumatucel-L (NovaRx Corporation)128133.
The results from these trials evaluating TGFβ blockade
alone indicate that blockade of TGFβ may enhance antitu-
mour immune responses in patients with cancer, although
Figure 3 | Effects of TGFβ on effector T cells. Transforming growth factor-β (TGFβ) differentially regulates the
survival, differentiation, proliferation and apoptosis of T cell subsets. Among the T helper (TH) cell subpopulations, both
TH1 and TH2 cells can mediate antitumour responses; however, TH1 cells seem to be more efficient. Both natural
regulatory T (nTReg) cell and induced TReg (iTReg) cell populations inhibit antitumour immune responses. Within the tumour
microenvironment, TGFβ can promote tumour growth by the maintenance of TReg cell and differentiation of iTReg cell
subpopulations. TGFβ can also inhibit TH1 cell and cytotoxic T lymphocyte (CTL) functions by downregulating T-bet and
interferon-γ (IFNγ) expression and probably promoting a shift towards TH2 cell differentiation. CTLs are potent antitumour
effector cells. TGFβ could also inhibit tumour immune surveillance by the induction of apoptosis in short-lived effector
CTLs. The role of TH17 cells in tumour biology is still controversial and requires further characterization. FOXP3, forkhead
box P3; GATA3, GATA-binding protein 3; NFAT, nuclear factor of activated T cells; RORγt, retinoic acid receptor-related
orphan receptor-γt; STAT4, signal transducer and activator of transcription 4.
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Immune suppression mechanism?
Type I NKT cell
CD4+ T cell
? Effector cell
? IFN? and cytotoxicity
further immunological endpoints — including evaluat-
ing the effects of TGFβ blockade by these approaches
on effector T cell to TReg cell ratios in the tumour bed,
NK cells, NKT cells and MSDCs — have been limited
or missing in the most recent studies. For example,
peripheral mononuclear cells isolated from patients with
high-grade gliomas receiving trabedersen had increased
antitumour activity against gliomas when stimulated
with IL-2 ex vivo132, which indicates that this approach
might enhance antitumour responses in patients receiving
trabedersen. However, no further immunological char-
acterization of this approach has been published so far.
Another example is the evaluation of the antisense vac-
cine approach in patients with lung cancer who received
belagenpumatucel-L. Initial studies reported increased
serum levels of IFN-γ or immunoglobulins specific for
the administered vaccine cells in a cohort of patients
enrolled in these trials130. However, tumour antigen-spe-
cific immune responses, as well as evaluation of the effec-
tor and regulatory immune cell compartments, in patients
receiving these therapies were not further addressed. In
fact, the most recently published Phase II trial with this
vaccine lacked immunological endpoint analyses, focus-
ing instead on clinical endpoints for patients with lung
cancer stratified according to different levels of circulat-
ing tumour cells131. Without further characterization of
the immunological effects on patients with cancer receiv-
ing anti-TGFβ therapies, it may be difficult to interpret
the importance of the limited clinical efficacy shown in
the most recent trial data. Nevertheless, additional trials
evaluating the small molecule inhibitors, antibody-medi-
ated blockade and infusion of CTLs transfected with the
dominant-negative TGFβRII are ongoing, with additional
immunological endpoints to be evaluated in the last trial,
including CTL survival and function (TABLE 1).
Although systemic blockade of TGFβ has been well
tolerated in preclinical studies, given the pleiotropic
effects of this cytokine, one potential concern of this type
of therapy is the development of autoimmune toxicities in
humans. This could be particularly problematic if TGFβ
blockade is used in combination with other immune-
activating agents, such as cytotoxic T lymphocyte antigen
4 (CTLA-4)- or programmed cell death 1 (PD1)-specific
antibodies (which are also being evaluated as single agents
in clinical trials and have shown several autoimmune
toxicities)134. Other potential toxicities of blocking TGFβ
might result from the cytokine’s homeostatic functions
in other tissues outside of the immune system, including
angiogenesis and the development of musculoskeletal tis-
sues, and TGFβ could also potentially increase the risk of
developing new malignancies.
The manipulation of local TGFβ sources in the
tumour should be considered in the future as a strategy
to inhibit the dominant immunosuppressive intratu-
moral environment while promoting antitumour immu-
nity. Challenges to this approach include being able to
target the tumour microenvironment with TGFβ inhibi-
tors without affecting TGFβ function in the rest of the
Figure 4 | Effects of TGFβ on regulatory cells. Within the tumour microenvironment, transforming growth factor-β
(TGFβ) has been implicated in recruiting natural regulatory T (nTReg) cells, as well as converting CD4+ effector T cells to
induced TReg (iTReg) cells. These TReg cells can express cell surface-bound TGFβ and can inhibit effector cells, including
natural killer (NK) cells and CD8+ T cells, in the tumour microenvironment by cell–cell contact to dampen the antitumour
response. Type I NKT cells, which are responsible for recruiting effector immune cells to the tumour through the
production of large amounts of interferon-γ (IFNγ), can be suppressed by intratumoral TGFβ, whereas type II NKT cells
support increased TGFβ production by myeloid-derived suppressor cells (MDSCs) through the generation of
interleukin-13 (IL-13). CD8+ regulatory T cells have been observed in lung tumours and these might result from the
production of IL-10 by antigen-presenting cells, leading to increased TGFβ production in the tumour microenvironment.
The precise immunosuppressive mechanisms of CD8+ regulatory T cells in regulating the antitumour immune response
have yet to be identified. CTL, cytotoxic T lymphocyte.
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Table 1 | Summary of TGFβ-induced signalling inhibitors being evaluated as immunotherapies
Company or Mechanism of Stage of
Summary of results
Antisense Pharma Antisense
and Phase III
Safe and well-tolerated in patients with high-grade
glioma. 7 out of24 patients with stable disease and 2 out
of 24 patients with complete response 25 months after
initiating therapy (REF. 132 and clinicaltrials.gov identifier:
Decreases TGFβ1 secretion by and subsequent
proliferation of lung, colon and prostate cancer cell lines141
AP11014 Antisense Pharma Antisense
Phase IIISafe but ineffective at improving scarring after eye surgery
when compared with placebo142
Phase II May prevent excessive post operative scarring for
glaucoma surgery; when added to an angiotensin
I-converting enzyme inhibitor [AU:OK?] can arrest
diabetic nephropathy in rats143
for all isoforms of
for all isoforms of
Phase ISafe and well-tolerated in patients with advanced
melanoma and kidney cancer144
1D11R&D SystemsPreclinical Pretreatment of mice engrafted with a syngeneic breast
cancer cell line suppresses breast cancer metastases to
TR1 and MT1 ImClone Systems/
PreclinicalEnhances antitumour responses against murine mammary
and colon cancer cell lines by increasing CTL and NK cell
activity (both cytotoxic activity and IFNγ production),
increasing the number of tumour-infiltrating CD8+ T cells
and decreasing numbers of tumor-infiltrating TReg cells and
splenic MDSCs in treated mice146. TReg cells obtained from
treated mice also showed decreased suppression of naive
T cell proliferation ex vivo146
Eli LillySmall molecule
inhibitors of TGFβRI
PreclinicalMost have not been assessed in animal models;
nanoparticle delivery of Ly364947 resulted in antitumour
activity against human pancreatic and gastric xenografts in
Ly573636Eli Lilly Small molecule
inhibitor of TGFβRI
Phase IIPhase II trial in patients with advanced stage melanoma
is ongoing (clinicaltrials.gov NCT0038329); results
Ly2157299Eli LillySmall molecule
inhibitor of TGFβRI
Phase ISafe and well-tolerated in patients with colon cancer,
prostate cancer, adrenal gland cancer, breast cancer and
No in vivo data available; inhibition of TGFβ signalling has
been established in vitro149,150
inhibitors of TGFβRI
inhibitors of TGFβRI
PreclinicalInhibits growth of multiple myeloma (SD-093) and glioma
(SD208) in vivo151,152
Ki26894Kirin Brewery CompanySmall molecule
inhibitor of TGFβRI
PreclinicalInhibits breast cancer metastasis and increases survival in a
preclinical mouse model153
Sm16Biogen IdecSmall molecule
inhibitor of TGFβRI
PreclinicalIn vivo effects include inhibiting mouse mesothelioma
tumour growth and recurrence following resection in a
Bind to SMADs to inhibit TGFβ-mediated gene expression
in in vitro luciferase reporter assays156
University of WisconsinInteracting peptide
P144 and P17NeoMPS/Digna Biotech14mer peptide
binding to TGFβRI
Preclinical Administration of both peptides with the adjuvant
molecules polyI:C and agonistic CD40-specific antibody
increased antitumour activity against a lymphoma cell
line in mice by increasing NK cell, tumour-specific CTL and
DC activity while suppressing MDSCs and inhibiting TGFβ
production by TReg cells157
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host to maintain systemic homeostatic processes. This
might require new delivery systems, as well as effective
TGFβ-specific drugs that have minimal systemic toxici-
ties for the recipient.
Given that TGFβ affects the activity and differentiation
of numerous immune cell types, it is unclear which of
the effects of TGFβ is most important in the tumour
microenvironment. This is, in part, due to the pleio-
tropic nature of TGFβ and the contextual and combi-
natorial effects that this cytokine can have at a range
of biological concentrations on different cell types at
various stages of their development. Therefore, there
remain several questions regarding the basic biology
of this cytokine as well as the best strategy to modu-
late this pathway, alone and in combination with other
pathways, to enhance antitumour immunity in patients
Although it is thought that tumour cells are an impor-
tant source of TGFβ in the tumour microenvironment,
immune cells themselves might in fact produce more
of this cytokine, which is produced by effector T cells,
regulatory T cells, APCs and MDSCs. Identifying the
most relevant source of TGFβ and the exact cells that
can respond to this cytokine is important, as localized
immunotherapy in the tumour microenvironment might
be safer than systemic therapies that could interfere with
the systemic homeostatic functions of TGFβ. In addition,
TGFβ is expressed and synthesized as an inactive latent
form, which cannot bind to its receptor. TGFβ becomes
activated by interacting with molecules such as plasmin,
matrix metalloproteinases, reactive oxygen species,
thrombospondin 1, αVβ6 integrin and αVβ8 integrin6,92.
Notably, the cells that can activate TGFβ may be different
from those that produce this potent cytokine, and so this
activation step provides a means for TGFβ to integrate
signals from several cell types92. We know little about
how TGFβ is activated in a tumour-bearing environment
and whether tolerogenic DCs, TAMs or MDSCs have
a greater capacity to activate TGFβ than their immu-
nogenic counterparts. Therefore, a precise understand-
ing of the mechanisms by which immunosuppressive
cell subsets work alone and together, and their specific
involvement in producing and/or activating TGFβ, may
improve cancer immunotherapies.
Further studies are also warranted to evaluate
the effect of increasing innate and adaptive immune
responses in a tumour-bearing host. For example, inhibi-
tion of TGFβ offers a means to manipulate T cell polari-
zation in vivo that can change an immunosuppressive
environment into a more antitumour environment once
a tumour has established in the host, as is the case in the
treatment of patients with cancer when these types of
therapies are generally being considered. Once the exact
role of IL-17 and TH17 cells in antitumour immunity has
been defined, modulation of TGFβ levels might also be
used to alter the ratio between TReg cells and TH17 cells in
the tumour microenvironment, as high concentrations
of TGFβ may favour Treg cells, whereas low concentra-
tions of TGFβ may lead to TH17 cell differentiation135.
In addition, blocking TGFβ-induced signalling in com-
bination with tumour vaccines promotes antitumour
immunity that is mediated, in part, by CD8+ T cells82–
84,136, which could lead to a long-term response with
Table 1 (cont.) | Summary of TGFβ-induced signalling inhibitors being evaluated as immunotherapies
Drug (trade name) Company or
Mechanism of Stage of
Summary of results
in an antisense-
NovaRx CorporationAllogeneic tumour
cell vaccine in which
the tumour cells
have been modified
to express a TGFβ2
Phase I and IINo significant toxicities observed in patients with
cancer. When stratified for circulating tumour cells,
patients with few circulating tumour cells had
increased median survival compared with patients
with a greater number of circulating tumour
Soluble TBR2-FcGenzymeStabilized soluble
PreclinicalLifetime exposure was tolerated in mice; decreased
incidence of metastasis formation in a metastatic
melanoma model and an inducible transgenic
breast cancer model158
PlasmidKagawa UniversityPlasmid DNA
fused to human IgG
PreclinicalAdministration to tumour-draining lymph nodes in
mice inhibited the growth of implanted lymphomas
and melanomas, as well as melanoma lung
Baylor College of
the TGFβ DNR
ex vivo and infused
Phase I trial currently
enrolling for lung
NCT00889954 ) and
In preclinical studies DNR-transfected EBV-specific
CTLs were resistant to antiproliferative effects
of exogenously administered TGFβ and had
enhanced cytolytic activity in vitro as well as
enhanced antitumour activity against EBV-positive
lymphomas engrafted in SCID mice160. Phase I
clinical trials evaluating the safety of this approach
in lung cancer and lymphoma patients are ongoing
CTL, cytotoxic T lymphocyte; DC, dendritic cell; DNR, dominant-negative receptor; EBV, Epstein–Barr virus; IFNγ, interferon-γ; MDSC, myeloid-derived suppressor
cells; NK, natural killer; polyI:C, polyinosinic–polycytidylic acid; R, receptor; SCID, severe combined immunodeficient; TGFβ, transforming growth factor-β;
TReg, regulatory T.
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SMAD2 and 3
SMAD2 and 3
SMAD2 and 3
Additional areas for future research related to the
development of agents that efficiently block TGFβ and
its activity include pharmacodynamic profiling of tissue
TGFβ concentrations and the optimization of strategies
to block the most appropriate TGFβ-dependent signal-
ling pathways. For example, the blockade of SMAD-
independent pathways of TGFβ-dependent signalling,
including MAPKs, Rho GTPases and phosphoinositide
3-kinases that are involved in tumour progression and
metastasis, could lead to new strategies to enhance anti-
tumour immunity (FIG. 5). Mutations in SMAD2, SMAD3
and SMAD4 lead to cancer progression137,138, indicating
that the tumour-suppressor properties of TGFβ involve
SMAD-dependent signalling and, therefore, that SMAD-
dependent pathways may not be ideal therapeutic targets.
Future studies analysing the contribution of SMAD-
independent and SMAD-dependent TGFβ-mediated
signalling pathways in controlling antitumour immune
responses in addition to their role in tumour parenchy-
mal cells are required. Understanding the intricate signal-
ling pathways controlled by TGFβ, in both tumour and
immune cells, as well as the mechanism (or mechanisms)
leading to its opposing effects in tumour biology, could
lead to new strategies against cancer139.
Furthermore, identifying the ideal timing of TGFβ
blockade in the host, if used in combination with vac-
cines, cytokine therapies (IL-2 and IL-15) or other
immune-activating antibodies (such as CTLA4-, PD1- or
PDL1-specific blocking antibodies) would be informa-
tive140. Finally, designing optimal methods to deliver the
most effective TGFβ inhibitor to the tumour microenvi-
ronment and the evaluation of exposing expanded T cell
subsets to these drugs ex vivo to enhance adoptive T cell-
based immunotherapies are all areas requiring further
So far, the clinical trials evaluating blockade of TGFβ
in patients with cancer do not show a clear clinical ben-
efit. Therefore, larger studies are warranted to clarify the
toxicity and efficacy of these strategies. In addition, the
optimal dose and timing of TGFβ blockade, as well as the
ideal combination of this approach with other immuno-
therapies, remain unknown. These questions are cur-
rently being addressed in ongoing preclinical studies and
are likely to be the focus of future clinical trials.
Figure 5 | Targets for inhibiting TGFβ and downstream signalling events. The
transforming growth factor-β (TGFβ)-dependent signalling pathway depends on type I
and type II serine-threonine kinase receptors and transcription factors known as
SMADs. The dimeric bioactive ligand binds to a TGFβ type II receptor (TGFβRII), which
in turn phosphorylates and activates TGFβRI. Once TGFβRI is activated, it
phosphorylates the receptor SMADs (SMAD2 and SMAD3), which promotes their
interaction with the common mediator SMAD (SMAD4) and translocation to the
nucleus. The inhibitory SMAD7 negatively regulates TGFβ-induced signalling by
competing with SMAD2 AND SMAD3 for interaction with TGFβRI or SMAD4. Current
TGFβ signalling inhibitors (listed in TABLE 1 and shown in the figure) include ligand,
receptor and SMAD antagonists. However, additional SMAD-independent pathways
have been reported to be induced in response to TGFβ, including the activation of
mitogen-activated protein kinase (MAPK), Rho-like GTPase and phosphoinositide
3-kinase (PI3K) signalling pathways; a complete understanding of these alternative
pathways could potentially indicate additional downstream molecules that could be
targeted in future therapeutic approaches. Figure adapted from REF. 39.
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R.A.F. is an investigator of the Howard Hughes Medical
Institute. This work is supported by a post-doctoral fellow-
ship grant from the Cancer Research Institute (S.S.), the NCI-
funded Yale SPORE in Skin Cancer (P50 CA121974) through
a Yale Skin SPORE Career Development Award (S.H.W.) and
a post-doctoral fellowship from PEW Charitable Trust: PEW
Latin American Fellow Program in Biomedical Sciences
(P.L.L.). Additional support from NIH grants CA121974 and
DK051665 (R.A.F.) and JDRF grant 32-2008-352 (R.A.F.).
Competing interests statement
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CD69 | FOXP3 | IFNγ | IL-2R β-chain | IL-6 | IL-10 | IL-13 |
RORγt | SMAD2 | SMAD3 | SMAD4 | TGFβ | TGFβRII
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About the author
Richard A. Flavell is Sterling Professor of Immunobiology at Yale
University School of Medicine, Connecticut, USA, and an inves-
tigator of the Howard Hughes Medical Institute. He received his
B.Sc. (Honors) in 1967 and Ph.D. in 1979 in biochemistry from
the University of Hull, UK, and carried out postdoctoral work in
Amsterdam, The Netherlands (1970–1972) and Zurich, Switzerland
(1972–1973). Before accepting his current position in 1988, he was an
assistant professor at the University of Amsterdam, The Netherlands
(1974–1979); then Head of the Laboratory of Gene Structure and
Expression at the National Institute for Medical Research, Mill Hill,
London, UK (1979–1982); and subsequently President and Chief
Scientific Officer of Biogen Research Corporation, Cambridge,
Massachusetts, USA (1982–1988). He is a fellow of the Royal Society,
and a member of the National Academy of Sciences and the Institute
of Medicine. He uses transgenic and gene-targeted mice to study T cell
tolerance and activation in immunity and autoimmunity, apoptosis
and the regulation of T cell differentiation.
Shomyseh Sanjabi is an assistant investigator at The Gladstone
Institute of Virology and Immunology and an assistant professor
of microbiology and immunology at the University of California,
San Francisco, USA. She received her Ph.D. from the University of
California, Los Angeles, USA, under the mentorship of S. Smale,
where she studied the mechanism of selectivity among members of
the nuclear factor-κB (NF-κB) family of transcription factors. During
her postdoctoral training with R. Flavell at Yale University, she studied
the effects of transforming growth factor-β (TGFβ)-induced signal-
ling on T cell biology. The current focus of her laboratory is to study
the role of TGFβ-induced signalling in memory T cell development
and in chronic viral infections.
Stephen H. Wrzesinski received an M.D. and a Ph.D. in microbiol-
ogy and immunology from the State University of New York Upstate
Medical University, New York, USA, where he studied the mecha-
nisms behind the pathogenesis of human T cell leukaemia virus type 1
(HTLV-1) tax oncoprotein under the mentorship of G. Feuer. He sub-
sequently completed an internal medicine residency at Dartmouth–
Hitchcock Medical Center, New Hampshire, USA, and a medical
oncology fellowship at the Yale Cancer Center, Connecticut, USA.
During his postdoctoral training with R. Flavell at Yale University,
he evaluated combinations of TGFβ inhibition with cytokine and
antibody therapies against melanoma. He is an assistant professor of
medicine at Yale University and West Haven Veteran’s Administration
Cancer Center, Connecticut, USA, and his research interests include
cancer immunotherapy development.
Paula Licona-Limón obtained her Ph.D at the Universidad Nacional
Autónoma de México, Mexico City, Mexico, and has investigated
the signalling pathway of TGFβ-related proteins in the regulation of
thymocyte differentiation. Her current work as a postdoctoral fellow
in the Flavell laboratory is focused on characterizing the molecular
mechanisms controlling TGFβ-dependent signals in T cells during
steady-state and pathological conditions.
??During early tumour formation, transforming growth factor-β
(TGFβ) can function as a tumour suppressor to prevent tumori-
genesis; however, overproduction of TGFβ in established tumours
is often associated with tumour metastasis and poor prognosis in
patients with cancer.
??The tumour-promoting effects of TGFβ may be due to the
immunosuppressive effects it has on both innate and adaptive
??TGFβ inhibits natural killer cell function by inhibiting cytokine
production and downregulating the expression of activating
??TGFβ inhibits the maturation of, and cytokine production and
proper antigen presentation by, dendritic cells, while promoting
regulatory T (TReg) cell differentiation and an overall tolerogenic
state. TGFβ also promotes an M2 state for macrophages and an N2
state for neutrophils.
??TGFβ directly dampens the function of CD4+ and CD8+ effector
T cells and promotes the survival of TReg cells. Depending on the
cytokine milieu, TGFβ greatly affects the differentiation of several
key CD4+ T cell subsets in tumour immunology.
??There is great interest in targeting TGFβ-induced signalling
for immunotherapy of cancer; however, the dosing, timing and
combination of this approach with other immunotherapies to
achieve the most successful antitumour effect need to be further
000 The polarization of immune cells in the
tumour environment by TGFβ
Richard A. Flavell, Shomyseh Sanjabi, Stephen
H. Wrzesinski and Paula Licona-Limón
A comprehensive description of the many ways in
which TGFβ can inhibit antitumour immune responses
through effects on innate and adaptive immune cells
and how these immunosuppressive effects could be
targeted for the benefit of patients with cancer.
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