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Blocking TGF-β Signaling To Enhance The Efficacy Of Immune Checkpoint Inhibitor

Authors Bai X, Yi M, Jiao Y, Chu Q, Wu K 

Received 22 July 2019

Accepted for publication 25 October 2019

Published 11 November 2019 Volume 2019:12 Pages 9527—9538

DOI https://doi.org/10.2147/OTT.S224013

Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 2

Editor who approved publication: Dr Sanjay Singh



Xianguang Bai,1,2,* Ming Yi,2,* Ying Jiao,2 Qian Chu,2 Kongming Wu2,3

1Medical School, Pingdingshan University, Pingdingshan, Henan, People’s Republic of China; 2Department of Oncology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, People’s Republic of China; 3Department of Oncology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Kongming Wu; Qian Chu
Department of Oncology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, People’s Republic of China
Email [email protected]; [email protected]

Abstract: During malignant transformation, a growing body of mutations accumulate in cancer cells which not only drive cancer progression but also endow cancer cells with high immunogenicity. However, because one or multiple steps in cancer-immunity cycle are impaired, anti-cancer immune response is too weak to effectively clear cancer cells. Therefore, how to restore robust immune response to malignant cells is a hot research topic in cancer therapeutics field. In the last decade, based on the deeper understanding of cancer immunity, great signs of progress have been made in cancer immunotherapies especially immune checkpoint inhibitors (ICIs). ICIs could block negative immune co-stimulatory pathways and reactivate tumor-infiltrating lymphocytes (TILs) from exhausted status. ICIs exhibit potent anti-cancer effect and have been approved for the treatment of numerous cancer types. Parallel with durable and effective tumor control, the actual response rate of ICIs is unsatisfactory. Although a subset of patients benefit from ICIs treatment, a large proportion of patients show primary or acquired resistance. Previously intensive studies indicated that the efficacy of ICIs was determined by a series of factors including tumor mutation burden, programmed death ligand-1 (PD-L1) expression, and TILs status. Recently, it was reported that transforming growth factor-beta (TGF-β) signaling pathway participated in cancer immune escape and ICI resistance. Concurrent TGF-β blockade might be a feasible strategy to enhance the efficacy of immunotherapy and relieve ICI resistance. In this mini-review, we summarized the latest understanding of TGF-β signaling pathway and cancer immunity. Besides, we highlighted the synergistic effect of TGF-β blockade and ICIs.

Keywords: immunotherapy, immune checkpoint inhibitor, PD-1, PD-L1, TGF-β, tumor immune microenvironment, tumor infiltrating lymphocyte

Introduction

Host immunity could recognize and clear non-self immunogenic materials. Theoretically, neoantigens or tumor-associated antigens generated during oncogenesis could initiate anti-cancer immune attack. The robust anti-cancer immune response is usually described as cancer-immunity cycle model.1 Firstly, cancer cells-derived neoantigens or tumor-associated antigens are captured by dendritic cells (DCs). After antigen processing, DCs present cancer antigens with major histocompatibility complex (MHC) molecules to naïve T cells in peripheral lymphoid organs. Following the priming and activation, T cells could specifically recognize cancer antigens. Then, primed T cells traffic and infiltrate into tumor beds. Tumor-infiltrating lymphocytes (TILs) could directly eliminate tumor cells which further release more tumor antigens and upregulate the magnitude of anti-cancer immune response.1 However, this series of stepwise procedures tend to be interrupted by several factors such as downregulated MHC on tumor cells,2 immune editing,3 as well as increased immune checkpoints.4,5 As a result, malignant cells escape from immune attack and eventually develop into visible tumor mass.

Cancer immunotherapy is aiming to launch a self-sustaining cancer-immunity cycle which could self-amplify and self-propagate with minimized treatment-related auto-inflammation.6 Immune checkpoints such as programmed death 1 (PD-1),7 cytotoxic T lymphocyte antigen 4 (CTLA-4),8 lymphocyte activation gene 3 (LAG-3),9 as well as T-cell immunoglobulin and mucin-domain containing-3 (TIM-3)10 are vital factors maintaining pro-tumor immune microenvironment, which are also regarded as ideal targets for cancer immunotherapy. However, anti-cancer immune response is a cyclic and stepwise process.11 The actual effect of anti-cancer immune elimination is determined by upstream immune editing (depletion of cancer cell sub-clones with T cell targets), downstream immunosuppressive tumor microenvironment including antigenic modulation and immune inhibitory cytokines especially transforming growth factor-beta (TGF-β) in tumor beds.1214 It is generally believed that the upregulated immune checkpoints on cancer cells are rate-limiting steps in cancer-immunity cycle.1,15 Nevertheless, the frequent ICIs resistance indicate that PD-1- or CTLA-4-targeted monotherapy could not completely counteract immunosuppression in the tumor microenvironment.16 A comprehensive framework containing multiple factors would be meaningful to remove adverse factors and amplify the whole anti-cancer immunity.17

TGF-β is a versatile molecule which could bi-directionally regulate the initiation and progression of cancer.1820 Besides, TGF-β has a multifaceted influence on tumor immune microenvironment.21 Increasing evidence suggests that the excessive secretion of TGF-β in tumor closely relates to increased pro-tumor immune elements, restrained tumor-killing effect of TIL, as well as limited infiltration of immune effector cells.2224 TGF-β might be an evaluable target for cancer treatment and the dual-blockade of TGF-β/immune checkpoints would have a synergistic effect.

Immune Checkpoints In Tumor Microenvironment

T cell activation is a complex process containing two signals.25 The first activation signal is the specific binding of antigenic peptide-MHC complex on antigen presentation cell (APC) and T cell receptor (TCR) on naïve T cell.25 The second activation signal is also known as co-stimulatory signal which refers to the interaction between co-stimulatory molecules on APC and corresponding receptors on T cell.26 Simultaneous stimulations from first and second signals are the prerequisite of optimal T cell activation. In the absence of co-stimulatory, T cells are prone to be unresponsive to antigenic materials (anergic T cells).26 Besides, some negative co-stimulatory (also termed as co-inhibitory) signals participate in T cell activation as well. Under the physiology condition, co-inhibitory signals maintain peripheral tolerance and prevent anti-immune diseases via counterbalancing co-stimulatory signals.27,28 In tumor immune microenvironment, upregulated immune checkpoints blunt effector T cells and protect cancer cells from immune killing.29 Among all immune checkpoint pathways, PD-1/PD-L1 and CTLA-4/B7 (CD80/CD86) pathways are most well-studied. Multiple agents targeting PD-1, PD-L1, and CTLA-4 have been applied in clinic.3032

PD-1-PD-L1 Signaling Pathway

PD-1 molecule consists of an extracellular IgV-like domain, a transmembrane domain, and an intracellular tail. The cytoplasmic tail of PD-1 contains an immunoreceptor tyrosine-based inhibitory motif (ITIM) and an immunoreceptor tyrosine-based switch motif (ITSM) which are the core structures for immune inhibitory function.33 PD-1 is widely expressed on multiple immune cells such as activated T cells, DCs, and natural killer cells (NKs).3436 The expression pattern of PD-L1 is different from PD-1. PD-L1 is constitutively expressed on immune cells including B cells, T cells, macrophages, DCs, and mesenchymal stem cells.37 Besides, the expression of PD-L1 could be transiently induced by a panel of cytokines especially interferon-gamma (IFN-γ).38,39 Apart from immune cells, a broad range of non-immune cells express PD-L1 as well.35 The upregulation of PD-L1 in tumor cells could be attributed to two factors. Firstly, some oncogenic pathways contribute to PD-L1 overexpression.40 Moreover, locally pre-existed inflammation leads to PD-L1 upregulation as a feedback termed as adaptive immune resistance.41

PD-1 molecule could transduce the signal of PD-L1 when it is cross-linked with TCR. After the stimulation from TCR, the binding of PD-1 and PD-L1 activates this immune inhibitory pathway.42 The tyrosine residues of ITIM and ITSM are phosphorylated and recruit SHP1/2, which could counteract the TCR/CD3- or CD28-mediated phosphorylation.43 PD-1/PD-L1 signaling not only blocks TCR/CD3 pathway by dephosphorylating the core components of TCR downstream signaling such as Zap-70 and Lck but also inhibits PI3K-Akt and Ras-MEK-ERK pathways in T cells.44 The suppressed PI3K and MAPK pathways result in reduced glycolysis/amino acid metabolism and increased fatty acid oxidation. These metabolic changes of T cells propel the differentiation of T cells towards regulatory T cells (Tregs) and exhausted T cells but hamper the differentiation towards effector T cells and memory T cells.42 Besides T cell differentiation, PD-1 signaling could interrupt cell cycle process of T cells via inhibiting PI3K and MAPK pathways.42

CTLA-4-B7 Signaling Pathway

Similarly to PD-1, CTLA-4 molecule contains an extracellular IgV-like domain, a transmembrane domain, and an intracellular tail as well.45 CTLA-4 is upregulated on activated T cells but rarely expressed on naïve T cells.46 Moreover, CTLA-4 is constitutively expressed on Tregs.47 Due to the similar molecular structure as well as higher affinity and avidity, CTLA-4 could competitively antagonize the binding between CD28 and B7.48 CTLA-4 inhibits T cell activation by multiple manners. Firstly, CTLA-4 could recruit phosphatases such as PP2A to reverse TCR/CD3 mediated phosphorylation of downstream proteins.49 Secondly, CTLA-4 downregulates the transcription of IL-2 which is the core cytokine for T cells activation and proliferation.49 Besides, CTLA-4 induces the abundance of B7 molecules on T cells by transendocytosis.47 Moreover, CTLA-4-B7 signaling pathway could induce the generation of several immune inhibitory components such as indoleamine-2, 3-dioxygenase (IDO) and TGF-β.50,51

TGF-β Signaling Pathway And Cancer

Multiple cancers possess a TGF-β-enriched tumor microenvironment. Numerous components of tumor microenvironment including cancer cells, fibroblasts, macrophages, and platelets could secret TGF-β.52

The Structure Of TGF-β

The TGF-β family contains three members: TGF-β1, β2, and β3. All of the three cytokines are synthesized as precursors consisting of a signal peptide, a latency-associated peptide (LAP), a C-terminal fragment.53 Under the guidance of signal peptide, the TGF-β precursor is translocated to endoplasmic reticulum which is further assembled to a dimer by inter-chain disulfide bonds.54 After furin-mediated cleavage, the disulfide-linked C-terminal fragment is non-covalently associated with the disulfide-linked N-terminal LAP, which eventually form the small latent complex.54 The LAP domain folds around C-terminal fragment (mature TGF-β), blocking access of TGF-β to corresponding receptor.54 The active TGF-β homodimer is released from small latent complex by (1) extracellular protease cleavage; (2) in the assistance of latent TGF-β binding protein, separating active TGF-β by cell contraction-derived and integrin-mediated tension; (3) with the help of GARP on the cells such as Tregs or macrophages, releasing active TGF-β by cell contraction-derived and integrin-mediated tension.52,55 All the three TGF-β isoforms are highly homologous (71–79% sequence identity in C-terminal fragment TGF-β regions) and have the similar functions in vitro.54 In vivo studies showed that the predominantly immunity-related isoform was TGF-β1 and TGF-β1 deficiency led to embryonic lethal or severe multi-organ inflammation.56

TGF-β Signaling Pathway

TGF-β signal is transduced by TGF-β receptor complex which consists of a TGF-βI receptor homodimer (TGF-βRI) and a TGF-βII receptor homodimer (TGF-βRII).57 Firstly, extracellular TGF-β binds to TGF-βRII homodimer which further complex with TGF-βRI homodimer.57 Following TGF-β engagement, TGF-βRII homodimer phosphorylates the intracellular domain of TGF-βRI.57 Notably, the transduction of TGF-β2 is usually with the assistance of co-receptor β-glycan (also termed as TGF-βRIII).57 The engagement of TGF-β receptor complex recruits receptor Smad (R-Smad) molecules Smad2 and 3 to the intracellular domain of TGF-βRI.58 Subsequently, Smad2 and 3 are phosphorylated which then form a trimeric complex with Smad4.58 The trimeric Smad complex could translocate to nuclear and regulate gene expression.58 Besides, phosphorylated Smad 2 and 3 could also form a trimeric complex with TIF1γ to regulate the expression of targeting genes.54 Moreover, it was reported that some other members of Smad family including Smad 1 and 5 might participate in TGF-β signaling pathway in partial cells such as Th17 cells.59 Apart from classic Smad pathway, TGF-β signal could also be transduced by some Smad-independent pathways such as PI3K, MAPK, and Rho GTPase pathways.6062 It has been well-established that the exact downstream signaling pathway of TGF-β signal is context-dependent.54

TGF-β Pathway And Cancer Progression

The role of TGF-β pathway is bi-directional for cancer.20 For pre-malignant cells, TGF-β acts as a tumor suppressor via suppressing cell proliferation and promoting cell apoptosis.63 However, for advanced cancer, TGF-β promotes cancer metastasis and induces pro-tumor immune microenvironment.63 In the tumor microenvironment, by paracrine and autocrine ways, overexpressed TGF-β could regulate the functions of cancer cells and stromal cells.64 The increased TGF-β is contributed by cancer cells especially cancer stem cells, Tregs, Bregs, tumor-associated macrophages (TAMs), cancer-associated fibroblasts (CAFs), as well as myeloid-derived suppressor cells (MDSCs).64 Notably, Stanford et al found that in postpartum breast cancers, dying cancer cells could be engulfed by neighboring macrophages (the process is also known as efferocytosis) via receptor tyrosine kinase MerTK.65 As a result, this efferocytosis enhanced the transcription of multiple wound-healing cytokines including TGF-β.65 So far, the mechanism by which the production of TGF-β is increased in CAFs is not clear yet. Previous studies indicated multiple components in the tumor microenvironment such as bone marrow-derived mesenchymal stem cells participated in the transformation of normal fibroblasts to CAFs and enhanced the secretion of TGF-β of CAFs.66 Besides, some cancer-specific miRNA expression pattern elevated the TGF-β generation in CAFs as well. Tanaka et al reported that in esophageal cancers, increased extracellular miR-27 was an unfavorable predictive factor for prognosis. After miR-27a/b transfection, normal fibroblasts exhibited more CAFs-associated markers including increased expression of α-smooth muscle actin and TGF-β.67

As a growth-inhibitory cytokine, TGF-β could effectively suppress cell proliferation by increasing the activity of cyclin-dependent kinase inhibitors such as p15INK4, p21CIP1, and p27KIP1.6870 Simultaneously, TGF-β could downregulate the expression of Myc.71 Besides, TGF-β inhibits tumorigenic inflammation and maintains immune homeostasis.63 Therefore, TGF-β exhibits tumor-suppressing function during the early stage of carcinogenesis.52 However, persistent TGF-β exposure introduces selective pressure and a subset of cancer cells could overcome TGF-β-related tumor-suppressing effect and eventually develop to advanced tumor.63 Actually, the transformation of TGF-β from cancer promotor to cancer suppressor occurs by two approaches.63 Firstly, partial cancer cells acquire mutations in TGF-β signaling pathway and abrogate TGF-β-mediated tumor-suppressing effect.52 In the same time, TGF-β signaling in stromal cells promotes cancer progression by inhibiting immune surveillance and promoting the secretion of some carcinogenic cytokines such as IL-11.63 Besides, oncogenes or tumor suppressors interact with TGF-β signaling and switch TGF-β function.72 For some cancer cells with intact TGF-β signaling pathway, some oncogenic pathways could counteract the pro-apoptosis effect of TGF-β.52 As a result, cancer cells undergo non-lethal epithelial–mesenchymal transition (EMT) and obtain the increased capabilities of migration and distant colonization.52

Notably, overexpressed TGF-β in the tumor microenvironment is highly related with hypoxia. Stephen et al found that breathing supplementary oxygen could effectively relieve regional hypoxia and decrease TGF-β abundance in tumors.73 This treatment using supplementary oxygen convert immunosuppressive and TGF-β-enriched tumor microenvironment to normal microenvironment, which is a promising adjuvant strategy to restore robust anti-cancer immune response.74

The Effect Of TGF-β Signaling Pathway On Immunity

TGF-β signaling pathway has a substantial influence on various immune cells which not only participates in immune cell differentiation but also regulates the activity of immune components (Figure 1).75 Generally, TGF-β acts as an inflammation-inhibitory factor.

Figure 1 TGF-β signaling pathway and its role in tumor immune microenvironment. TGF-β signal is transduced by TGF-β receptor complex which consists of TGF-βI receptors and TGF-βII receptors (TGF-βRI and TGF-βRII). Firstly, extracellular TGF-β binds to TGF-βRII homodimer which further complex with TGF-βRI homodimer. Following TGF-β engagement, TGF-βRII homodimer phosphorylates the intracellular domain of TGF-βRI. The engagement of TGF-β receptor complex recruits receptor Smad (R-Smad) molecules Smad2 and 3 to the intracellular domain of TGF-βRI. Subsequently, Smad2 and 3 are phosphorylated which then form a trimeric complex with Smad4. The trimeric Smad complex could translocate to nuclear and regulate gene expression. Besides, phosphorylated Smad2 and 3 could also form a trimeric complex with TIF1γ to regulate the expression of targeting genes. Apart from classic Smad pathway, TGF-β signal could also be transduced by some Smad-independent pathways such as PI3K, MAPK, and Rho GTPase pathways. TGF-β signaling pathway has a substantial influence on various immune cells including downregulating the cytotoxicity of effector T cells and NKs, promoting the apoptosis of effector T cells, inducing the differentiation towards Tregs, hampering the antigens presentation of DCs. Abbreviations: NK, natural killer cell; CAF, cancer-associated fibroblast; MHC, major histocompatibility complex; IDO, indoleamine-2, 3-dioxygenase; Id1, inhibitor of differentiation 1; Treg, regulatory T cell.

Th1 Cell

Th1 cell is a vital player in anti-cancer immunity and the differentiation of Th1 cells needs APC-derived cytokine IL-12.76 IL-12 could induce Th1 cell to generate abundant IFN-γ and the secretion of IFN-γ could be self-propagated as a positive feedback loop.77 Besides, IL-12 upregulates the expression of key transcription factor T-bet which is the other determinant for Th1 cell differentiation.76 TGF-β could effectively hamper Th1 cell differentiation. Firstly, TGF-β reduces the expression of the receptor of IL-12 on Th0 cells and downregulates the sensitivity of Th0 cells to IL-12.78 Secondly, TGF-β downregulates the level of T-bet in Th1 cells.79 Thirdly, TGF-β inhibits the production of IL-12 of NK.80 Although most previous studies showed that TGF-β suppressed the Th1 differentiation, some studies also indicated that TGF-β could also induce Th1 cell differentiation in some certain background such as in the presence of IFN-γ or IL-4.81 However, the significance of TGF-β-mediated Th1 cell differentiation under certain condition needs further exploration.

CD8+ T Cell

In addition to Th1 cell differentiation, TGF-β could inhibit T cell proliferation by Smad signaling pathway. Previous studies showed TGF-β suppressed CD4+ and CD8+ T cell proliferation via downregulating the transcription of IL-2 and c-Myc, as well as upregulating p21Cip1 and p27Kip1.8284 Besides, TGF-β directly inhibits the cytotoxic activity of CD8+ T cells.85 The undermined lytic function is related to TGF-β-Smad signaling-mediated downregulation of granzyme A, B, perforin, Fas ligand, and IFN-γ.14

Treg

CD4+ CD25+ Foxp3+ Treg cell is a vital subpopulation of T cells which maintains immune homeostasis and participates in cancer immune evasion.86 TGF-β could suppress immune response by regulating Treg cells. The results of mouse experiments indicated that mice lacking TGF-β1 or harboring TGF-βRII deficiency in T cells had decreased Foxp3+ Treg cells in peripheral blood.87,88 In the presence of IL-2, TGF-β promotes the differentiation of naïve T cells towards Treg cells.89 Upon TCR engagement, Smad3 interacts with the enhancer region of Foxp3 (CNS1).90 Besides, Smad3 could upregulate Foxp3 transcription by forming enhanceosome complex with NFATc2 and CREB.91 The level of Foxp3+ Treg cells is remarkably reduced in Smad3 knockout mice.92 Although Smad2 could not directly interact with CNS1, T cells lacking Smad2 showed reduced Foxp3 expression.93 The effect of Smad2 on Foxp3 level was proposed to depend on the interaction between Smad2 and Smad3.54 Apart from Smad pathway, TGF-β could indirectly promote Foxp3 expression by counteracting Foxp3 transcription inhibitory factor such as Gfi-1.94

DC

DC is the key component of cancer-immunity cycle which captures antigenic materials from cancer cells and activates cancer-specific lymphocytes. TGF-β suppresses the antigen presentation of DC by decreasing the expression of MHCII.95 Hypersecretion of TGF-β in tumor microenvironment propels the transformation of DC towards immature myeloid cell phenotype which exhibits potent immune inhibitory effect.96 This transformation is attributed to multiple reasons. Firstly, TGF-β could upregulate the level of inhibitor of differentiation 1 which inhibits the differentiation of DC and proliferation of CD8+ T cells.96 Moreover, TGF-β promotes the formation of regulatory phenotype DC by inducing IDO in plasmacytoid DC and chemokine (C-C motif) ligand 22 (CCL22) chemokine in myeloid DC.97,98

Macrophage

TAM is the important source of TGF-β in tumor microenvironment.99 Besides, TAM activates TGF-β from its precursor via extracellular integrin αvβ8 and matrix metalloproteinase 14.99 It has been reported that TGF-β could inhibit NF-κB signaling-mediated inflammation response by promoting the degeneration of MYD88.100 Besides, TGF-β inhibits the inflammation phenotype macrophages by the interaction between inhibitory molecule Smad7 and TNF signaling pathway, which might contribute to immunosuppressive tumor microenvironment.52

NK

NKs could rapidly respond to a broad range of pathogen challenges, detect, and kill malignant cells. It is notable that NKs kill cancer cells independent on priming and activation processes. Besides, NKs enhance anti-cancer effect of adaptive immunity by secreting cytokines such as IFN-γ and tumor necrosis factor-α (TNF-α).101 TGF-β substantially suppresses the functions of NKs by blocking the expression of IFN-γ.80 Moreover, TGF-β downregulates the levels of NKG2D and NKp30 on NKs, which mediates the recognition of stressed cells.102

CAF

The ratio of CAF is usually increased in the microenvironment of advanced cancers. CAFs generate multiple materials including fibroblast activation protein, α-smooth muscle actin, extracellular matrix proteins (e.g., type I collagen and fibronectin), and various cytokines.103 Actually, CAF is the main producer of TGF-β for multiple cancer types. Previous study showed that increased TGF-β secreted by CAF and other cells suppressed anti-tumor immune response.104

Besides, CAF could be activated by TGF-β as well. The transcriptome analysis of patients undergoing ICI treatment showed that TGF-β-activated CAF gene program was highly correlated with ICI resistance.22 However, anti-TGF-β treatment could effectively convert T cell exclusion and enhance the efficacy of ICIs.22 The exact mechanisms by which TGF-β-activated CAFs lead to cancer immune escape are attributed to three reasons. Apart from the contribution of CAF to the upregulation of TGF-β, TGF-β propels the morphological alterations in CAFs which is helpful to the release of active TGF-β from latent complex.52 Moreover, activated TGF-β signaling pathway hampers the infiltration of T cells by remodeling the matrix architecture in the tumor stroma.52,105

Dual Blockade Of TGF-β And Immune Checkpoint

In tumor microenvironment, cancer cells usually hijack multiple immune inhibitory pathways to escape immune surveillance. Dysregulated TGF-β signaling pathway impaired multiple processes in anti-cancer immune response including antigen presentation, T cell infiltration, and tumor-killing activity. Therefore, ICI monotherapy might fail to restore robust anti-cancer immune response (Figure 2). Feun et al reported the results of NCT02658019 and found the baseline plasma TGF-β levels were significantly correlated to the poor outcomes of pembrolizumab-treated advanced hepatocellular carcinoma patients.106 Given the potentially synergistic effect of TGF-β pathway and immune checkpoint in inducing immune tolerance, a series of studies were conducted to explore the efficacy of combination strategy of TGF-β inhibitor and ICI for cancer treatment (Table 1).107

Table 1 Clinical Trials Of Dual Blockade Of TGF-β And Immune Checkpoint

Figure 2 The synergistic effect of TGF-β pathway and immune checkpoint in inducing immune tolerance. Dysregulated TGF-β signaling pathway impaired multiple processes in anti-cancer immune response including antigen presentation, T cell infiltration, and tumor-killing activity. Hyperactive TGF-β signaling together with increased PD-1/PD-L1 signal axis undermine anti-cancer immune response.

TGF-β Inhibitor Plus ICI

By establishing human microsatellite stable-like colorectal cancers in mice, Tauriello et al found that ICI monotherapy could not effectively eliminate cancer cells.104 However, additional TGF-β blockade remarkably enhanced the anti-cancer effect of ICI.104 The further exploration indicated that TGF-β could induce immune suppression by promoting the formation of T cell-exclusion phenotype tumors which were prone too resistant to ICI treatment.104 Simultaneously, Mariathasan et al noticed the similar phenomenon in patients with metastatic urothelial cancer.22 For metastatic urothelial cancer patients receiving atezolizumab treatment, the treatment response was highly related with the activity of TGF-β signaling pathway in fibroblasts, especially in patients with abundant CD8+ T cells enriched in peritumoral stroma rather than tumor center (immune excluded phenotype).22 Researchers recapitulated the immune-excluded phenotype tumors with EMT6 mouse mammary and MC38 mouse colon carcinoma model. Although neither following atezolizumab nor TGF-β blockade could reduce tumor burden, the combination therapy of atezolizumab and TGF-β blockade potently eradicated cancer cells in the two models.22 Further investigation suggested that the synergistic effect of dual-blockade was attributed to enhanced T cell infiltration into tumor center and anti-tumor immune response.22

In addition to increased T cell infiltration, Terabe et al found that blocking TGF-β1 and TGF-β2 significantly elevated cancer vaccine-induced Th1-type immune response, upregulated IFN-γ production, and increased T-bet expression of tumor-infiltrating CD8+ T cells.108 Besides, TGF-β blockade could enhance the treatment effect of cancer vaccine and anti-PD-1 antibody in mice models.108 Similarly, Chen et al found that TGF-β secreted by MDSCs was highly associated with PD-1/PD-L1 inhibitor resistance by inducing PD-1 upregulation on CD8+ T cells in tumor bed.109 TGF-β blockade significantly promoted the lytic function of tumor antigen-specific CD8+ T cells in vivo and in vitro.109

Fusion Antibody Simultaneously Targeting TGF-β And Immune Checkpoint

In 2018, Lan et al designed a bifunctional fusion protein (M7824) which consisted of a PD-L1 monoclonal antibody and the extracellular domains of human TGFβR-II.110,111 M7824 could simultaneously block cancer cell extrinsic and intrinsic immunosuppressive pathways.110 Under the guidance of PD-L1 monoclonal antibody moiety, M7824 could preciously locate into tumor microenvironment and further neutralize the abundant TGF-β. In vitro study showed that partial M7824 would be internalized after incubation with 293 cells ectopically expressing PD-L1.110 In vivo study M7824 exhibited potent tumor control effect superior to isotype control, trap control, as well as PD-L1 monoclonal antibody.110 Besides, M7824 treatment effectively prolonged survival time in mice bearing EMT6 or MC38 carcinomas.110 Subsequent investigation showed that M7824 elevated both quantity and immune ability of tumor-infiltrating lymphocytes such as CD8+ T cells, NKs, DCs, and macrophages.110 Later, Ravi et al reported another bifunctional fusion protein targeting CTLA-4 and TGFβR-II.112 This bifunctional fusion protein could inhibit the differentiation of Tregs and Th17 cells, and increase tumor-specific IFN-γ+ effector and memory cells.112 The anti-cancer effect of anti-CTLA4-TGFβR-II fusion protein was superior to CTLA-4 or PD-1 inhibitors.112

Conclusion

The biologic effects of TGF-β signaling pathway are highly context-dependent. In addition to the pleiotropic function as a tumor promoter or suppressor, TGF-β pathway could induce immune suppression and participate in immune homeostasis or immune evasion. For normal tissues especially for ones consistently exposure to antigenic materials, intact TGF-β pathway could decrease the risk of inflammation-related malignant transformation. However, for advanced tumor tissues, hyperactive TGF-β pathway undermines immune surveillance and promotes tumor immune escape. TGF-β pathway broadly inhibits multiple anti-cancer producers including T cell priming and activation, immunosuppressive lymphocyte differentiation, cytotoxic function of effectors. Thus, additional TGF-β blockade might effectively enhance the ICI therapy for TGF-β-enriched tumors. We believe that the dual blockade of TGF-β and immune checkpoint would be a promising strategy in clinical practice in the future.

Acknowledgments

This study was supported by the National Natural Science Foundation of China (grant nos. 81572608 and 81874120), the Wuhan Science and Technology Bureau (grant no. 2017060201010170), and Henan Provincial Key Science and Technology Research Projects (192102310087).

Author Contributions

All authors contributed to data analysis, drafting or revising the article, gave final approval of the version to be published, and agree to be accountable for all aspects of the work.

Disclosure

The authors report no conflicts of interest in this work.

References

1. Chen DS, Mellman I. Oncology meets immunology: the cancer-immunity cycle. Immunity. 2013;39:1–10. doi:10.1016/j.immuni.2013.07.012

2. Yoshihama S, Vijayan S, Sidiq T, Kobayashi KS. NLRC5/CITA: a key player in cancer immune surveillance. Trends Cancer. 2017;3:28–38. doi:10.1016/j.trecan.2016.12.003

3. Vinay DS, Ryan EP, Pawelec G, et al. Immune evasion in cancer: mechanistic basis and therapeutic strategies. Semin Cancer Biol. 2015;35(Suppl):S185–S198. doi:10.1016/j.semcancer.2015.03.004

4. Long J, Lin J, Wang A, et al. PD-1/PD-L blockade in gastrointestinal cancers: lessons learned and the road toward precision immunotherapy. J Hematol Oncol. 2017;10:146. doi:10.1186/s13045-017-0511-2

5. Yi M, Jiao D, Xu H, et al. Biomarkers for predicting efficacy of PD-1/PD-L1 inhibitors. Mol Cancer. 2018;17:129. doi:10.1186/s12943-018-0864-3

6. Karasaki T, Nagayama K, Kuwano H, et al. An immunogram for the cancer-immunity cycle: towards personalized immunotherapy of lung cancer. J Thorac Oncol. 2017;12:791–803. doi:10.1016/j.jtho.2017.01.005

7. Liu B, Song Y, Liu D. Recent development in clinical applications of PD-1 and PD-L1 antibodies for cancer immunotherapy. J Hematol Oncol. 2017;10:174. doi:10.1186/s13045-017-0541-9

8. El Dika I, Khalil DN, Abou-Alfa GK. Immune checkpoint inhibitors for hepatocellular carcinoma. Cancer. 2019;125(19):3312–3319. doi:10.1002/cncr.32076

9. Marin-Acevedo JA, Dholaria B, Soyano AE, et al. Next generation of immune checkpoint therapy in cancer: new developments and challenges. J Hematol Oncol. 2018;11:39. doi:10.1186/s13045-018-0582-8

10. Naoum GE, Morkos M, Kim B, Arafat W. Novel targeted therapies and immunotherapy for advanced thyroid cancers. Mol Cancer. 2018;17:51. doi:10.1186/s12943-018-0786-0

11. Yi M, Qin S, Zhao W, et al. The role of neoantigen in immune checkpoint blockade therapy. Exp Hematol Oncol. 2018;7:28. doi:10.1186/s40164-018-0120-y

12. Bose S, Panda AK, Mukherjee S, Sa G. Curcumin and tumor immune-editing: resurrecting the immune system. Cell Div. 2015;10:6. doi:10.1186/s13008-015-0012-z

13. Garrido F, Aptsiauri N, Doorduijn EM, Garcia Lora AM, van Hall T. The urgent need to recover MHC class I in cancers for effective immunotherapy. Curr Opin Immunol. 2016;39:44–51. doi:10.1016/j.coi.2015.12.007

14. Thomas DA, Massague J. TGF-beta directly targets cytotoxic T cell functions during tumor evasion of immune surveillance. Cancer Cell. 2005;8:369–380. doi:10.1016/j.ccr.2005.10.012

15. Predina J, Eruslanov E, Judy B, et al. Changes in the local tumor microenvironment in recurrent cancers may explain the failure of vaccines after surgery. Proc Natl Acad Sci U S A. 2013;110:E415–E424. doi:10.1073/pnas.1211850110

16. Li X, Shao C, Shi Y, Han W. Lessons learned from the blockade of immune checkpoints in cancer immunotherapy. J Hematol Oncol. 2018;11:31.

17. Chen DS, Mellman I. Elements of cancer immunity and the cancer-immune set point. Nature. 2017;541:321–330. doi:10.1038/nature21349

18. Derynck R, Akhurst RJ, Balmain A. TGF-beta signaling in tumor suppression and cancer progression. Nat Genet. 2001;29:117–129.

19. Calon A, Tauriello DV, Batlle E. TGF-beta in CAF-mediated tumor growth and metastasis. Semin Cancer Biol. 2014;25:15–22. doi:10.1016/j.semcancer.2013.12.008

20. Metelli A, Salem M, Wallace CH, et al. Immunoregulatory functions and the therapeutic implications of GARP-TGF-beta in inflammation and cancer. J Hematol Oncol. 2018;11:24. doi:10.1186/s13045-018-0570-z

21. Yang L, Pang Y, Moses HL. TGF-beta and immune cells: an important regulatory axis in the tumor microenvironment and progression. Trends Immunol. 2010;31:220–227. doi:10.1016/j.it.2010.04.002

22. Mariathasan S, Turley SJ, Nickles D, et al. TGFbeta attenuates tumour response to PD-L1 blockade by contributing to exclusion of T cells. Nature. 2018;554:544–548. doi:10.1038/nature25501

23. Polanczyk MJ, Walker E, Haley D, Guerrouahen BS, Akporiaye ET. Blockade of TGF-beta signaling to enhance the antitumor response is accompanied by dysregulation of the functional activity of CD4(+)CD25(+)Foxp3(+) and CD4(+)CD25(-)Foxp3(+) T cells. J Transl Med. 2019;17:219. doi:10.1186/s12967-019-1967-3

24. Ungefroren H. Blockade of TGF-beta signaling: a potential target for cancer immunotherapy? Expert Opin Ther Targets. 2019;23:1–15.

25. Bretscher PA. A two-step, two-signal model for the primary activation of precursor helper T cells. Proc Natl Acad Sci U S A. 1999;96:185–190. doi:10.1073/pnas.96.1.185

26. Appleman LJ, Boussiotis VA. T cell anergy and costimulation. Immunol Rev. 2003;192:161–180. doi:10.1034/j.1600-065X.2003.00009.x

27. Tivol EA, Borriello F, Schweitzer AN, et al. Loss of CTLA-4 leads to massive lymphoproliferation and fatal multiorgan tissue destruction, revealing a critical negative regulatory role of CTLA-4. Immunity. 1995;3:541–547. doi:10.1016/1074-7613(95)90125-6

28. Nishimura H, Nose M, Hiai H, Minato N, Honjo T. Development of lupus-like autoimmune diseases by disruption of the PD-1 gene encoding an ITIM motif-carrying immunoreceptor. Immunity. 1999;11:141–151. doi:10.1016/S1074-7613(00)80089-8

29. Munn DH, Bronte V. Immune suppressive mechanisms in the tumor microenvironment. Curr Opin Immunol. 2016;39:1–6. doi:10.1016/j.coi.2015.10.009

30. Valecha GK, Vennepureddy A, Ibrahim U, et al. Anti-PD-1/PD-L1 antibodies in non-small cell lung cancer: the era of immunotherapy. Expert Rev Anticancer Ther. 2017;17:47–59. doi:10.1080/14737140.2017.1259574

31. Liu SY, Wu YL. Ongoing clinical trials of PD-1 and PD-L1 inhibitors for lung cancer in China. J Hematol Oncol. 2017;10:136. doi:10.1186/s13045-017-0506-z

32. Snyder A, Makarov V, Merghoub T, et al. Genetic basis for clinical response to CTLA-4 blockade in melanoma. N Engl J Med. 2014;371:2189–2199. doi:10.1056/NEJMoa1406498

33. Zhang X, Schwartz JC, Guo X, et al. Structural and functional analysis of the costimulatory receptor programmed death-1. Immunity. 2004;20:337–347. doi:10.1016/S1074-7613(04)00051-2

34. Agata Y, Kawasaki A, Nishimura H, et al. Expression of the PD-1 antigen on the surface of stimulated mouse T and B lymphocytes. Int Immunol. 1996;8:765–772. doi:10.1093/intimm/8.5.765

35. Keir ME, Butte MJ, Freeman GJ, Sharpe AH. PD-1 and its ligands in tolerance and immunity. Annu Rev Immunol. 2008;26:677–704. doi:10.1146/annurev.immunol.26.021607.090331

36. Yamazaki T, Akiba H, Iwai H, et al. Expression of programmed death 1 ligands by murine T cells and APC. J Immunol. 2002;169:5538–5545. doi:10.4049/jimmunol.169.10.5538

37. Yi M, Yu S, Qin S, et al. Gut microbiome modulates efficacy of immune checkpoint inhibitors. J Hematol Oncol. 2018;11:47. doi:10.1186/s13045-018-0592-6

38. Langereis JD, Pickkers P, de Kleijn S, et al. Spleen-derived IFN-gamma induces generation of PD-L1(+)-suppressive neutrophils during endotoxemia. J Leukoc Biol. 2017;102:1401–1409. doi:10.1189/jlb.3A0217-051RR

39. Zerdes I, Matikas A, Bergh J, Rassidakis GZ, Foukakis T. Genetic, transcriptional and post-translational regulation of the programmed death protein ligand 1 in cancer: biology and clinical correlations. Oncogene. 2018;37:4639–4661. doi:10.1038/s41388-018-0303-3

40. Xue S, Hu M, Iyer V, Yu J. Blocking the PD-1/PD-L1 pathway in glioma: a potential new treatment strategy. J Hematol Oncol. 2017;10:81. doi:10.1186/s13045-017-0455-6

41. Taube JM, Anders RA, Young GD, et al. Colocalization of inflammatory response with B7-h1 expression in human melanocytic lesions supports an adaptive resistance mechanism of immune escape. Sci Transl Med. 2012;4:127ra37. doi:10.1126/scitranslmed.3003689

42. Bardhan K, Anagnostou T, Boussiotis VA. The PD1: PD-L1/2Pathway from discovery to clinical implementation. Front Immunol. 2016;7:550. doi:10.3389/fimmu.2016.00550

43. Riley JL. PD-1 signaling in primary T cells. Immunol Rev. 2009;229:114–125. doi:10.1111/imr.2009.229.issue-1

44. Patsoukis N, Brown J, Petkova V, et al. Selective effects of PD-1 on Akt and Ras pathways regulate molecular components of the cell cycle and inhibit T cell proliferation. Sci Signal. 2012;5:ra46. doi:10.1126/scisignal.2002796

45. Teft WA, Kirchhof MG, Madrenas J. A molecular perspective of CTLA-4 function. Annu Rev Immunol. 2006;24:65–97. doi:10.1146/annurev.immunol.24.021605.090535

46. Hegel JK, Knieke K, Kolar P, Reiner SL, Brunner-Weinzierl MC. CD152 (CTLA-4) regulates effector functions of CD8+ T lymphocytes by repressing Eomesodermin. Eur J Immunol. 2009;39:883–893. doi:10.1002/eji.200838770

47. Walker LS, Sansom DM. The emerging role of CTLA4 as a cell-extrinsic regulator of T cell responses. Nat Rev Immunol. 2011;11:852–863. doi:10.1038/nri3108

48. Thompson CB, Allison JP. The emerging role of CTLA-4 as an immune attenuator. Immunity. 1997;7:445–450. doi:10.1016/S1074-7613(00)80366-0

49. Mitsuiki N, Schwab C, Grimbacher B. What did we learn from CTLA-4 insufficiency on the human immune system? Immunol Rev. 2019;287:33–49. doi:10.1111/imr.2019.287.issue-1

50. Zhao Y, Yang W, Huang Y, et al. Evolving roles for targeting CTLA-4 in cancer immunotherapy. Cell Physiol Biochem. 2018;47:721–734. doi:10.1159/000490025

51. Munn DH, Sharma MD, Mellor AL. Ligation of B7-1/B7-2 by human CD4+ T cells triggers indoleamine 2,3-dioxygenase activity in dendritic cells. J Immunol. 2004;172:4100–4110. doi:10.4049/jimmunol.172.7.4100

52. Batlle E, Massague J. Transforming growth factor-beta signaling in immunity and cancer. Immunity. 2019;50:924–940. doi:10.1016/j.immuni.2019.03.024

53. Gleizes PE, Munger JS, Nunes I, et al. TGF-beta latency: biological significance and mechanisms of activation. Stem Cells. 1997;15:190–197. doi:10.1002/stem.150190

54. Travis MA, Sheppard D. TGF-beta activation and function in immunity. Annu Rev Immunol. 2014;32:51–82. doi:10.1146/annurev-immunol-032713-120257

55. Kelly A, Houston SA, Sherwood E, Casulli J, Travis MA. Regulation of Innate and Adaptive Immunity by TGFbeta. Adv Immunol. 2017;134:137–233.

56. Kulkarni AB, Huh CG, Becker D, et al. Transforming growth factor beta 1 null mutation in mice causes excessive inflammatory response and early death. Proc Natl Acad Sci U S A. 1993;90:770–774. doi:10.1073/pnas.90.2.770

57. Kang JS, Liu C, Derynck R. New regulatory mechanisms of TGF-beta receptor function. Trends Cell Biol. 2009;19:385–394. doi:10.1016/j.tcb.2009.05.008

58. Shi Y, Massague J. Mechanisms of TGF-beta signaling from cell membrane to the nucleus. Cell. 2003;113:685–700. doi:10.1016/S0092-8674(03)00432-X

59. Lee Y, Awasthi A, Yosef N, et al. Induction and molecular signature of pathogenic TH17 cells. Nat Immunol. 2012;13:991–999. doi:10.1038/ni.2416

60. Xie F, Ling L, van Dam H, Zhou F, Zhang L. TGF-beta signaling in cancer metastasis. Acta Biochim Biophys Sin (Shanghai). 2018;50:121–132. doi:10.1093/abbs/gmx123

61. Xiao Y, Song YJ, Song B, et al. TGF-beta/MAPK signaling mediates the effects of bone marrow mesenchymal stem cells on urinary control and interstitial cystitis after urinary bladder transplantation. Am J Transl Res. 2017;9:1193–1202.

62. Heldin CH, Moustakas A. Signaling Receptors for TGF-beta Family Members. Cold Spring Harb Perspect Biol. 2016;8. doi:10.1101/cshperspect.a022053

63. David CJ, Massague J. Contextual determinants of TGFbeta action in development, immunity and cancer. Nat Rev Mol Cell Biol. 2018;19:419–435. doi:10.1038/s41580-018-0007-0

64. Ahmadi A, Najafi M, Farhood B, Mortezaee K. Transforming growth factor-beta signaling: tumorigenesis and targeting for cancer therapy. J Cell Physiol. 2019;234:12173–12187. doi:10.1002/jcp.27955

65. Stanford JC, Young C, Hicks D, et al. Efferocytosis produces a prometastatic landscape during postpartum mammary gland involution. J Clin Invest. 2014;124:4737–4752. doi:10.1172/JCI76375

66. Wen S, Niu Y, Yeh S, Chang C. BM-MSCs promote prostate cancer progression via the conversion of normal fibroblasts to cancer-associated fibroblasts. Int J Oncol. 2015;47:719–727. doi:10.3892/ijo.2015.3060

67. Tanaka K, Miyata H, Sugimura K, et al. miR-27 is associated with chemoresistance in esophageal cancer through transformation of normal fibroblasts to cancer-associated fibroblasts. Carcinogenesis. 2015;36:894–903. doi:10.1093/carcin/bgv067

68. Hannon GJ, Beach D. p15INK4B is a potential effector of TGF-beta-induced cell cycle arrest. Nature. 1994;371:257–261. doi:10.1038/371257a0

69. Polyak K, Lee MH, Erdjument-Bromage H, et al. Cloning of p27Kip1, a cyclin-dependent kinase inhibitor and a potential mediator of extracellular antimitogenic signals. Cell. 1994;78:59–66. doi:10.1016/0092-8674(94)90572-X

70. Reynisdottir I, Polyak K, Iavarone A, Massague J. Kip/Cip and Ink4 Cdk inhibitors cooperate to induce cell cycle arrest in response to TGF-beta. Genes Dev. 1995;9:1831–1845. doi:10.1101/gad.9.15.1831

71. Chen CR, Kang Y, Siegel PM, Massague J. E2F4/5 and p107 as Smad cofactors linking the TGFbeta receptor to c-myc repression. Cell. 2002;110:19–32. doi:10.1016/S0092-8674(02)00801-2

72. Zheng X, Liu Q, Yi M, Qin S, Wu K. The regulation of cytokine signaling by retinal determination gene network pathway in cancer. Onco Targets Ther. 2018;11:6479–6487.

73. Hatfield SM, Kjaergaard J, Lukashev D, et al. Immunological mechanisms of the antitumor effects of supplemental oxygenation. Sci Transl Med. 2015;7:277ra30. doi:10.1126/scitranslmed.aaa1260

74. Hatfield SM, Kjaergaard J, Lukashev D, et al. Systemic oxygenation weakens the hypoxia and hypoxia inducible factor 1alpha-dependent and extracellular adenosine-mediated tumor protection. J Mol Med (Berl). 2014;92:1283–1292. doi:10.1007/s00109-014-1189-3

75. Liu M, Li S, Li MO. TGF-beta control of adaptive immune tolerance: a break from treg cells. Bioessays. 2018;40:e1800063. doi:10.1002/bies.201800063

76. O’Garra A, Gabrysova L, Spits H. Quantitative events determine the differentiation and function of helper T cells. Nat Immunol. 2011;12:288–294. doi:10.1038/ni.2003

77. Schoenborn JR, Wilson CB. Regulation of interferon-gamma during innate and adaptive immune responses. Adv Immunol. 2007;96:41–101.

78. Gorham JD, Guler ML, Fenoglio D, Gubler U, Murphy KM. Low dose TGF-beta attenuates IL-12 responsiveness in murine Th cells. J Immunol. 1998;161:1664–1670.

79. Gorelik L, Constant S, Flavell RA. Mechanism of transforming growth factor beta-induced inhibition of T helper type 1 differentiation. J Exp Med. 2002;195:1499–1505. doi:10.1084/jem.20012076

80. Laouar Y, Sutterwala FS, Gorelik L, Flavell RA. Transforming growth factor-beta controls T helper type 1 cell development through regulation of natural killer cell interferon-gamma. Nat Immunol. 2005;6:600–607. doi:10.1038/ni1197

81. Tofukuji S, Kuwahara M, Suzuki J, et al. Identification of a new pathway for Th1 cell development induced by cooperative stimulation with IL-4 and TGF-beta. J Immunol. 2012;188:4846–4857. doi:10.4049/jimmunol.1103799

82. Brabletz T, Pfeuffer I, Schorr E, et al. Transforming growth factor beta and cyclosporin A inhibit the inducible activity of the interleukin-2 gene in T cells through a noncanonical octamer-binding site. Mol Cell Biol. 1993;13:1155–1162. doi:10.1128/MCB.13.2.1155

83. Genestier L, Kasibhatla S, Brunner T, Green DR. Transforming growth factor beta1 inhibits Fas ligand expression and subsequent activation-induced cell death in T cells via downregulation of c-Myc. J Exp Med. 1999;189:231–239. doi:10.1084/jem.189.2.231

84. Wolfraim LA, Walz TM, James Z, Fernandez T, Letterio JJ. p21Cip1 and p27Kip1 act in synergy to alter the sensitivity of naive T cells to TGF-beta-mediated G1 arrest through modulation of IL-2 responsiveness. J Immunol. 2004;173:3093–3102. doi:10.4049/jimmunol.173.5.3093

85. Zhang N, Bevan MJ. TGF-beta signaling to T cells inhibits autoimmunity during lymphopenia-driven proliferation. Nat Immunol. 2012;13:667–673. doi:10.1038/ni.2319

86. Fang P, Li X, Dai J, et al. Immune cell subset differentiation and tissue inflammation. J Hematol Oncol. 2018;11:97. doi:10.1186/s13045-018-0637-x

87. Marie JC, Letterio JJ, Gavin M, Rudensky AY. TGF-beta1 maintains suppressor function and Foxp3 expression in CD4+CD25+ regulatory T cells. J Exp Med. 2005;201:1061–1067. doi:10.1084/jem.20042276

88. Marie JC, Liggitt D, Rudensky AY. Cellular mechanisms of fatal early-onset autoimmunity in mice with the T cell-specific targeting of transforming growth factor-beta receptor. Immunity. 2006;25:441–454. doi:10.1016/j.immuni.2006.07.012

89. Chen W, Jin W, Hardegen N, et al. Conversion of peripheral CD4+CD25- naive T cells to CD4+CD25+ regulatory T cells by TGF-beta induction of transcription factor Foxp3. J Exp Med. 2003;198:1875–1886. doi:10.1084/jem.20030152

90. Schlenner SM, Weigmann B, Ruan Q, Chen Y, von Boehmer H. Smad3 binding to the foxp3 enhancer is dispensable for the development of regulatory T cells with the exception of the gut. J Exp Med. 2012;209:1529–1535. doi:10.1084/jem.20112646

91. Xu L, Kitani A, Strober W. Molecular mechanisms regulating TGF-beta-induced Foxp3 expression. Mucosal Immunol. 2010;3:230–238. doi:10.1038/mi.2010.7

92. Takimoto T, Wakabayashi Y, Sekiya T, et al. Smad2 and Smad3 are redundantly essential for the TGF-beta-mediated regulation of regulatory T plasticity and Th1 development. J Immunol. 2010;185:842–855. doi:10.4049/jimmunol.0904100

93. Lu L, Wang J, Zhang F, et al. Role of SMAD and non-SMAD signals in the development of Th17 and regulatory T cells. J Immunol. 2010;184:4295–4306. doi:10.4049/jimmunol.0903418

94. Zhu J, Davidson TS, Wei G, et al. Down-regulation of Gfi-1 expression by TGF-beta is important for differentiation of Th17 and CD103+ inducible regulatory T cells. J Exp Med. 2009;206:329–341. doi:10.1084/jem.20081666

95. Nandan D, Reiner NE. TGF-beta attenuates the class II transactivator and reveals an accessory pathway of IFN-gamma action. J Immunol. 1997;158:1095–1101.

96. Papaspyridonos M, Matei I, Huang Y, et al. Id1 suppresses anti-tumour immune responses and promotes tumour progression by impairing myeloid cell maturation. Nat Commun. 2015;6:6840. doi:10.1038/ncomms7840

97. Hanks BA, Holtzhausen A, Evans KS, et al. Type III TGF-beta receptor downregulation generates an immunotolerant tumor microenvironment. J Clin Invest. 2013;123:3925–3940. doi:10.1172/JCI65745

98. Pallotta MT, Orabona C, Volpi C, et al. Indoleamine 2,3-dioxygenase is a signaling protein in long-term tolerance by dendritic cells. Nat Immunol. 2011;12:870–878. doi:10.1038/ni.2077

99. Kelly A, Gunaltay S, McEntee CP, et al. Human monocytes and macrophages regulate immune tolerance via integrin alphavbeta8-mediated TGFbeta activation. J Exp Med. 2018;215:2725–2736. doi:10.1084/jem.20171491

100. Lee YS, Park JS, Kim JH, et al. Smad6-specific recruitment of Smurf E3 ligases mediates TGF-beta1-induced degradation of MyD88 in TLR4 signalling. Nat Commun. 2011;2:460. doi:10.1038/ncomms1469

101. Hu W, Wang G, Huang D, Sui M, Xu Y. Cancer immunotherapy based on natural killer cells: current progress and new opportunities. Front Immunol. 2019;10:1205. doi:10.3389/fimmu.2019.01205

102. Castriconi R, Cantoni C, Della Chiesa M, et al. Transforming growth factor beta 1 inhibits expression of NKp30 and NKG2D receptors: consequences for the NK-mediated killing of dendritic cells. Proc Natl Acad Sci U S A. 2003;100:4120–4125. doi:10.1073/pnas.0730640100

103. Xing F, Saidou J, Watabe K. Cancer associated fibroblasts (CAFs) in tumor microenvironment. Front Biosci (Landmark Ed). 2010;15:166–179. doi:10.2741/3613

104. Tauriello DVF, Palomo-Ponce S, Stork D, et al. TGFbeta drives immune evasion in genetically reconstituted colon cancer metastasis. Nature. 2018;554:538–543. doi:10.1038/nature25492

105. Chakravarthy A, Khan L, Bensler NP, Bose P, De Carvalho DD. TGF-beta-associated extracellular matrix genes link cancer-associated fibroblasts to immune evasion and immunotherapy failure. Nat Commun. 2018;9:4692. doi:10.1038/s41467-018-06654-8

106. Feun LG, Li YY, Wu C, et al. Phase 2 study of pembrolizumab and circulating biomarkers to predict anticancer response in advanced, unresectable hepatocellular carcinoma. Cancer. 2019;125:3603–3614. doi:10.1002/cncr.32339

107. Vanpouille-Box C, Formenti SC. Dual transforming growth factor-beta and programmed death-1 blockade: a strategy for immune-excluded tumors? Trends Immunol. 2018;39:435–437. doi:10.1016/j.it.2018.03.002

108. Terabe M, Robertson FC, Clark K, et al. Blockade of only TGF-beta 1 and 2 is sufficient to enhance the efficacy of vaccine and PD-1 checkpoint blockade immunotherapy. Oncoimmunology. 2017;6:e1308616. doi:10.1080/2162402X.2017.1308616

109. Chen X, Wang L, Li P, et al. Dual TGF-beta and PD-1 blockade synergistically enhances MAGE-A3-specific CD8(+) T cell response in esophageal squamous cell carcinoma. Int J Cancer. 2018;143:2561–2574. doi:10.1002/ijc.31730

110. Lan Y, Zhang D, Xu C, et al. Enhanced preclinical antitumor activity of M7824, a bifunctional fusion protein simultaneously targeting PD-L1 and TGF-beta. Sci Transl Med. 2018;10:eaan5488. doi:10.1126/scitranslmed.aan5488

111. David JM, Dominguez C, McCampbell KK, et al. A novel bifunctional anti-PD-L1/TGF-beta Trap fusion protein (M7824) efficiently reverts mesenchymalization of human lung cancer cells. Oncoimmunology. 2017;6:e1349589. doi:10.1080/2162402X.2017.1349589

112. Ravi R, Noonan KA, Pham V, et al. Bifunctional immune checkpoint-targeted antibody-ligand traps that simultaneously disable TGFbeta enhance the efficacy of cancer immunotherapy. Nat Commun. 2018;9:741. doi:10.1038/s41467-017-02696-6

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