Lung cancer is the leading cause of cancer-related death world-wide. Recently, immune checkpoint inhibitors (ICIs) targeting programmed cell death-1 (PD-1)/programmed cell death-ligand 1 (PD-L1) and cytotoxic T-lymphocyte antigen 4 (CTLA-4) axis have become the most promising treatment in several kinds of cancer, especially in lung cancer.[2–5] Although an unprecedented durable response rate has been observed in immunotherapies, the majority of patients do not benefit from the treatment and some patients relapse after a period of response. According to the most typical concept of practicing clinicians, mechanisms of resistance are divided into primary, adaptive, and acquired resistance. Primary resistance is defined as a clinical condition in which the tumor does not respond to immunotherapy strategies. The mechanism underlying the lack of response to immunotherapy may include adaptive immune resistance. Considering the evolutionary nature of immune/cancer cell interactions, this may be manifested as primary resistance, mixed reaction or acquired resistance. The definition of acquired resistance is a clinical scenario where a tumor initially responds to immunotherapy but recurs and progresses over time. At present, data are lacking about mechanisms involved in resistance to ICIs. Enhanced understanding of molecular and immunologic mechanisms of ICI response (and resistance) will not only identify novel biomarkers for prediction and/or prognosis, but ultimately guide optimal clinical combination of ICI. Here, we review the emerging data identifying novel mechanisms of innate and acquired resistance to ICI [Figure 1] and strategies to overcome cancer resistance.
Mechanisms of Resistance to ICIs
Primary and adaptive resistance
Antigen presentation and recognition
The most immediate cause of the tumor's non-response to ICIs is lack of recognition by T cells because of absence of tumor antigen. The process of distinguishing tumor cells from normal cells depends on T-cell recognition of tumor-specific or tumor-associated antigens. Clinical studies show that neoantigens delivered through dendritic cells (DCs) or other means induce potent antigen-specific T cell response and favorable clinical responses in prostate cancer, melanoma, and glioblastoma patients.[10,11] Combined with mass spectrometry and exome sequencing, the immunogenicity of neoantigens can be used to evaluate active T-cell responses.
Genetic instability due to alterations in DNA repair and replication genes can increase immunogenicity through high tumor mutational burden (TMB) with subsequent neoantigen formation. It was found that melanoma patients respond better to PD-1 therapy if tumor cells are enriched with BRCA2 mutations, which is an important homologous recombination DNA repair gene. Alterations in additional DNA damage response genes have recently shown correlation with high TMB and improved clinical outcomes to ICIs in urothelial cancer. Furthermore, tumors with deficiencies in DNA mismatch repair genes leading to microsatellite instability demonstrated high mutational burden with enhanced response to ICIs in a variety of tumors.
Besides, antigen processing, presentation, and immune escape can also be affected by epigenetic modifications in tumor cells which change the expression of immune-related genes.[16,17] For example, histone deacetylase (HDAC) inhibitors have been reported to increase major histocompatibility complex (MHC) and tumor antigen expression, and shift gene expression to a proapoptotic milieu in cancer cells. This suggests that reversing epigenetic modifications in tumor cells may enhance immune recognition and response.
T cell priming and activation
Abnormal Wnt/β-catenin signaling pathway can also lead to immunotherapy resistance. High levels of β-catenin in mice were associated with reduced CD103+ DC in tumor microenvironment. The possible mechanism is that the abnormal WNT/β-catenin signaling pathway induces the expression of transcription inhibitor activating transcription factor 3, which inhibits the expression of CCL4 gene, a chemokine of CD103+ DC, thereby reducing the infiltration of CD103+ DC. The lack of antigen presenting cells (APCs) leads to the dysfunction of initial T cell activation and the decrease of infiltrating T cells, which ultimately affects the immune response. Among human melanomas shown to have a poorly infiltrated phenotype, those containing mutations affecting the β-catenin pathway lacked a CD103+ DC immune signature and were insensitive to anticancer immunotherapies. In addition, the accumulation of CD103+ cross-presenting DCs in mouse tumors was shown to be dependent on the activation of intra-tumoral natural killer (NK) cells secreting the DC chemo-attractants chemokine (C-C motif) ligand (CCL) 5 and lymphotactin. In several human-derived cancer cell lines, the presence of intra-tumoral CCL5 and lymphotactin transcripts is closely correlated with that of gene signatures of both NK cells and CD103+ DCs, and the presence of these cell populations is associated with favorable overall survival (OS).
T cell specific antigen recognition provides the first signal of T cell activation, and the second signal comes from the interaction between the synergistic stimulus molecules expressed by APC and the corresponding receptors or ligands on the surface of T cells, the most important of which is the co-stimulatory molecule CD28-B7. Recent studies have shown that PD-1 inhibitor activated T cells still need the co-stimulation signal of CD28 to promote their proliferation and differentiation into killer T-cells. Trials in mice found that blocking the interaction between CD28 and B7, or knocking out the CD28 gene, prevented T cells from responding to PD-1 treatment. The binding of B7 molecules on its surface with CTLA-4 can lead to the apoptosis of antigen-specific T cells, and the secretion of interleukin (IL)-10 induces T helper 2 type response, thus inducing antigen-specific immune tolerance.
Many negative regulatory factors in tumor immune microenvironment, such as IL-10, vascular endothelial growth factor (VEGF), and transforming growth factor β (TGF-β), can lead to the maturation disorder and dysfunction of DCs, thus affecting the efficacy of immunotherapy. IL-10 and TGF-β can drive the differentiation of monocytes into M2-like tumor-associated macrophages (TAMs), which amongst their other suppressive actions, can also compete with local DCs for tumor antigens and consequently inhibit T cell priming. In addition, IL-10 and TGF-β can limit local T cell priming through the suppression of both DC function and the proliferative capacity of T cells. In addition, the TGF-β-driven activation of fibroblasts gives rise to a specific phenotype of immunomodulatory cancer-associated fibroblasts (CAFs). Through the release of TGF-β and IL-6, CAFs suppress the proliferation and trafficking capacity of antigen-presenting DCs, thereby interfering with tumor-directed T cell priming. In oral squamous cell carcinoma, tumor-secreted VEGF may promote the tumor immunologic escape by inhibiting the differentiation of immature DC from peripheral blood monocyte cells and increasing the levels of dysfunctional mature DC.
T cell trafficking and tumor infiltration
Through the tight regulation of the local chemokine- and cytokine-gradient, CAFs also limit the attraction of T cells to the TME.[30,31] Moreover, TGF-β CAFs can remodel the composition of the extracellular matrix (ECM), resulting in a dense ECM network that poses a physical barrier to T cell infiltration. Furthermore, CAFs can suppress the anti-tumor T cell response in the TME itself, through the up-regulation of immune checkpoint ligands on their cell surfaces.
Chemokines regulate immune cell trafficking in tumors and are implicated in tumor development, progression, and angiogenesis. Most tumors shape local chemokine networks to promote their growth by recruiting stromal cells like TAMs, myeloid-derived suppressor cells (MDSC), and regulatory T cells (Treg), all associated with poor patient prognosis. Several recent studies have investigated the involvement of chemokines in T-cell recruitment to tumors. Tumor cells secrete ligands CCL5, CCL7, and C-X-C motif chemokine ligand 8, and bind to the receptor C-C motif chemokine receptor (CCR) 1 or C-X-C chemokine receptor 2 expressed on the MDSCs subtype to recruit MDSCs into the tumor microenvironment. Tregs selectively over-express the chemokine CCR4, and its specific ligand CCL12 is produced by tumor cells and tumor microenvironment to further recruit Tregs and suppress host immune response.[35,36]In vitro experiments, anti-CCR4 monoclonal antibody inhibits Treg recruitment as well as promotes antibody-dependent cell-mediated cytotoxicity, further reducing the Treg population.
In addition to promoting angiogenesis, VEGF functions as an immunosuppressive cytokine and is associated with resistance to ICIs. VEGF levels were found to be higher in anti-PD-1 therapy non-responders compared with responders. In mouse models, VEGF impeded commitment of lymphoid progenitors, reducing progression to the T-cell lineage. Additionally, VEGF signaling reduces trafficking and extravasation of cytolytic T lymphocyte (CTLs) into the TME while it promotes infiltration of Tregs through a selective endothelium. Furthermore, VEGF increases expression of inhibitory receptors, contributing to CTL exhaustion. Oncogenic signaling through the mitogen-activated protein kinase (MAPK) pathway results in the production of VEGF, IL-1, and IL-8, which inhibits the recruitment and function of T cells.BRAF gene is an important transduction factor of MAPK signaling pathway. In the mouse model, BRAF inhibitor significantly increased the amount of chimeric antigen receptor T cells (CAR-T)/T-cell receptor engineered T-cell infiltration and enhanced its anti-tumor effect.
T-cell killing activity within the tumor microenvironment
The interferon-γ (IFN-γ) pathway is emerging as a key player in host immune response.[43–46] It plays a dual role in anti-tumor immune response. IFN-γ is produced by tumor-specific T cells, it induces an effective anti-tumor immune response by the following ways: (1) its direct anti-proliferative and pro-apoptotic effects on tumor cells; (2) enhancing the expression of MHC and other molecules to increase tumor antigen presentation; and (3) recruiting other immune cells. However, continuous IFN-γ exposure can cause immunoediting of cancer cells, which leads to immune escape.[48,49] One of the mechanisms by which tumor cells escape the effects of IFN-γ is to down-regulate the expression of or mutate related molecules involved in the IFN-γ signaling pathway, such as the downstream signaling molecules of the IFN-γ receptor: Janus kinase (JAK) 1/2 and the signal transducer and activators of transcription (STATs), and so on. It was reported that mutations or epigenetic silencing of molecules in this pathway lead to the loss of the anti-tumor effects both in cell line and animal models. Analysis of patients who did not respond to CTLA-4 antibody showed the high-frequency mutations of JAK2, IFN-γ receptor 1 and 2, interferon regulatory factor 1 (IRF1), and inhibited amplification of genes in this pathway, such as suppressor of cytokine signaling 1 and protein inhibitor of activated STAT 4. Mutations in this pathway also lead to the loss of PD-L1 expression upon IFN-γ exposure, resulting in tumor cells being genetically negative for inducible PD-L1 expression. In this case, it would be useless to block PD-L1 or PD-1 with antibodies. These patients may be the ones with initial resistance to anti-PD-1 treatment.[44,52]
Constitutively expressed inhibitory ligands such as PD-L1 on the surface of tumor cells may significantly inhibit the anti-tumor T cell response. Several studies have revealed a correlation between loss of phosphatase and tensin homolog (PTEN) in cancer and the up-regulation of PD-L1, implicating the role of PD-L1 in tumor immune evasion. PTEN is a tumor suppressor that negatively regulates the phosphoinositide 3-kinase (PI3K)/AKT pathway. The deletion of PTEN increases the expression of immunosuppressive cytokines, resulting in decrease of T cell infiltration and inhibition of autophagy in tumors, thereby reducing T-cell-mediated cell death. The loss of PTEN relates with inferior outcomes with PD-1 inhibitor therapy. In murine models, selective PI3K β inhibitor improved the efficacy of anti-PD-1 and anti-CTLA-4 antibodies. This pathway is responsible for the regulation of cellular processes such as proliferation and survival. PD-L1 expression was also up-regulated in lung squamous cell carcinoma following the simultaneous depletion of PTEN and serine-threonine kinase 11.
Other mechanisms that have been shown to have a role in the constitutive up-regulation of PD-L1 include the transcription factor IRF-1 and mutations in the epidermal growth factor receptor (EGFR). IRF-1 is responsible for the regulation of cell proliferation, apoptosis, and immunity. The knockdown of IRF-1 using small interfering RNA resulted in the decrease in transcription and translation of PD-L1 in a lung carcinoma cell line. Similarly, activation of the EGFR pathway resulted in the increased expression of PD-L1 in lung cancer cell lines and tissue.[57,58] Increased expression of markers of T-cell exhaustion, such as PD-1 and forkhead box P3 (FoxP3), was also observed in the tumor microenvironment. PD-1 blockade increased cytotoxic T-cell numbers as well as effector T-cell function, highlighting the role of the PD-1/PD-L1 axis in immune evasion and its manipulation as a therapeutic strategy. The increased expression of PD-L1 in tumor microenvironment resulted in decreased function of cytotoxic T cells and apoptosis, thus providing an immune escape mechanism for tumor cells.
Indoleamine 2,3-dioxygenase (IDO) was initially viewed as a promising target and biomarker, as high levels of IDO expression were associated with shorter progression free survival (PFS) and poorer outcomes. IDO induces inflammation within the tumor microenvironment, depletes the tryptophan required by cytotoxic T-cells, and induces the conversion of naïve T-cells to Tregs, thereby promoting a tolerogenic and immunosuppressive state. Unfortunately, recent clinical trials with IDO inhibitors have failed to show added efficacy on top of ICIs, raising the needs to re-evaluate its mechanisms and clinical trial design.
TAMs include M1 macrophages that promote anti-tumor immunity and M2 macrophages that promote tumor genesis and development.[60–62] An association was proved between higher frequencies of TAMs and poor prognosis in many tumors. In the mouse model of lung adenocarcinoma, it is possible to inhibit tumor growth by depleting M2 macrophages through inactivation of CCL2 and/or CCR2 signal transduction. Similar results have been shown in various murine tumor models.[64,65] Macrophages were suggested to play an important role in mediating therapeutic resistance.[65–67] It has been reported that macrophages directly inhibit T cell response in hepatocellular carcinoma by B7-H1 and in ovarian carcinoma by B7-H4. In order to overcome the potential resistance mechanism of macrophages, the researchers found that by blocking colony stimulating factor 1 receptor (CSF-1R) in a mouse model of pancreatic cancer, the frequency of TAMs decreased, and the production of IFN-γ increased, leading to inhibition of tumor growth. Importantly, blocking PD-1 or CLTA-4 alone did not significantly inhibit tumor growth.[70,71] However, combined treatment with CSF-1R blocker and PD-1 or CTLA-4 antibody can promote tumor elimination. Similarly, a synergistic effect of CSF-1R inhibitor and adoptive cell therapy (ACT) therapy has been shown in melanoma models. These data suggest that CSF-1R blockade induced TAMs death, thus enhancing the efficacy of immunotherapy.
MDSCs are an important part of the tumor microenvironment and can inhibit the T cell effect and proliferation.[73–76] More and more studies have demonstrated that the number of invasive MDSCs is associated with poor prognosis of patients and decreased efficacy of immunotherapy, including immune checkpoint therapy, adoptive T cell therapy, and DC vaccination. In addition, studies have shown that high expression of PI3Kγ in MDSCs can increase inflammatory mediators and immunosuppressive factors. It was demonstrated that PI3Kγ inhibitor (TG100-115 or IPI-549) can inhibit tumor growth in tumor-bearing mice. Indeed, in several tumors, selective PI3Kγ inhibitors with ICIs promoted tumor regression and increased survival.[81,82] The researchers also demonstrated that the combination of PI3K inhibitor and PD-1 inhibitor improved tumor regression and survival in tumor-bearing mice. In another study, compared with the combined treatment of two drugs (CTLA-4 inhibitor and PD-1 inhibitor), the triple-combination therapy (PI3Kγ inhibitor, CTLA-4 inhibitor, and PD-1 inhibitor) showed stronger anti-tumor effect in tumor-bearing mice. Therefore, it is reasonable to speculate that PI3Kγ inhibitor can promote the anti-tumor immune response and reverse the immune resistance caused by MDSCs.
Tregs play an important role in maintaining self-tolerance by secreting certain inhibitory cytokines such as IL-10, IL-35, TGF-β, or direct cell contact to suppress effector T cell (Teff) responses.[83–86] Many studies indicate that several tumors are infiltrated by Tregs.[87–89]A large number of animal experiments have shown that the consumption of Tregs in tumor microenvironment can enhance or restore anti-tumor immunity.[90,91] The response to anti-CTLA-4 treatment was related to the ratio of Teff to Tregs, and the higher the ratio, the better the response to treatment.[92,93] A retrospective study reported that the high expression of FoxP3+ Tregs was associated with better prognosis in patients treated with anti-CTLA-4 therapy. These data suggest that when immunotherapy fails to increase Teff and/or decrease Tregs to improve the ratio of Teff to Treg, tumors are likely to develop resistance to treatment, both initially and during disease recurrence.
At present, ICI treatment demonstrates a durable long-term response, but some patients develop tumor progression after remission, despite receiving continued therapy. Acquired resistance may be caused by the following reasons, such as changes in signaling pathways, tumor antigen presentation defects, and some new mutations.
Loss of antigen-presenting machinery components such as beta-2-microglobulin (β2M) and human leukocyte antigen (HLA) is an important mechanism to avoid antigen processing and presentation by tumors. β2M is necessary for HLA class I molecules to fold and transport to the cell surface, and its genetic deficiency would result in lack of CD8+ T cell recognition. It was reported that some patients who initially responded to immunotherapies with IL-2 or tumor-infiltrating lymphocyte (TIL) ACT may develop acquired resistance through loss of β2M.[95,96] In patients treated with PD-1 antibody, a new homozygous truncation mutation of β2M occurs in drug-resistant cells, resulting in lack of surface expression of HLA class I. Direct loss of the gene encoding the HLA-C∗08:02 class I molecule was recently described in a colorectal tumor that progressed after initial response to T-cell transfer therapy. These T cells were HLA-C∗08:02-restricted TILs, and thus this mutation directly allowed immune evasion by tumor cells. These examples together reinforce that immunotherapy failure was attributed to acquired defects in HLA class I molecules.
In the other two recurrence cases, a copy number neutral functional deletion mutation was found in JAK1 or JAK2, concurrent with loss of heterozygosity in β2M. These mutations in tumor cells lead to decreased sensitivity to IFN-γ, ultimately preventing IFN-γ-induced cell growth arrest. Upon exposure to IFN-γ (produced by activated T cells), JAK1/2 becomes activated and subsequently phosphorylates a tyrosine residue present on STATs. This JAK/STAT signaling pathway is responsible for cell proliferation, differentiation, cell migration, and apoptosis. IFN-γ also results in the up-regulation of PD-L1 on tumor cells, thus inactivating anti-tumor T cells. Some reports have described up-regulated expression of other T cell checkpoints at the time of acquired resistance, including T cell immunoglobulin mucin 3 (TIM-3), lymphocyte activation gene 3 (LAG3), and V-domain immunoglobulin suppressor of T cell activation (VISTA). By comparing the pathology before treatment with PD-1 inhibitor and the biopsy after drug resistance of uterine leiomyoma, it was found that drug-resistant tumor cells had unique double allelic PTEN gene deletion, and PD-1-positive T cell infiltration was reduced, suggesting that MAPK and PTEN gene mutation may be one of the mechanisms of acquired resistance by ICIs. TIM-3 is highly expressed in the tumor microenvironment, which can promote the apoptosis of Teffs, mediate the proliferation of Tregs and inhibitory cells from bone marrow, and inhibit the functions of NK cells and DC cells, thus promoting the tumor immune escape. It was shown that the acquired resistance of anti-PD-1 monoclonal antibody in the treatment of non-small cell lung cancer (NSCLC) patients is due to the high expression of TIM-3 on cytotoxic CD8+ T cells and its binding to galectin-9 on MDSCs. Studies have shown increased TIM-3 expression in TIL after the resistance with PD-1/PD-L1 inhibitors. The binding degree of T cells to PD-1 antibody was also correlated with TIM-3 expression. Sequential TIM-3 blocker therapy significantly improved the median survival of mouse models resistant to PD-1/PD-L1 inhibitors, and increased the number of tumor infiltrating CD8+ T cells. Analysis of immune cells in pleural effusion of patients with lung adenocarcinoma resistant to PD-1/PD-L1 inhibitors showed that the expression level of TIM-3 in T cells was significantly higher than that of patients who did not receive PD-1/PD-L1 inhibitors or just underwent surgery. All the above studies suggested that abnormal activation of TIM-3 is one of important resistant mechanisms, and blocking the TIM3 signaling pathway is expected to improve the efficacy of PD-1/PD-L1 inhibitors.
LAG3 is an immune checkpoint receptor that suppresses T-cell activity when up-regulated. LAG3 (CD223) is a transmembrane protein receptor that has been identified on activated T-cells, regulatory T-cells, B-cells, NK cells, and plasmacytoid DCs. As with self-antigen, continued exposure to tumor antigens, such as in the tumor microenvironment, causes up-regulation of LAG3 and its inhibitory actions lead to T-cell exhaustion, thus rendering these cells ineffective at directing attacks towards tumors cells. VISTA suppresses the activity of T cells and works as an immune checkpoint regulator. In murine models, tumors that were found to express VISTA had an increased rate of growth due to the T cell suppression. Tumor cells that express PD-1 or CTLA-4 and treated with anti PD-1/CTLA-4 antibodies were found to up-regulate VISTA to avoid immune surveillance. However, VISTA was found in almost all of NSCLCs and may be used as a new target of therapy.
Strategies to Reverse ICIs Resistance
Due to the complex resistant mechanisms of immunotherapy, there is still no standardized solution to this problem. Immunotherapy combined with immunotherapy, chemotherapy, radiotherapy, and targeted therapy is a strategy for the treatment of various cancers, including lung cancer. However, there are challenges related to identifying the appropriate population for combination therapy, balancing synergistic anti-tumor effects, and understanding adverse reactions associated with these treatments.
Combining with other ICIs
In theory, inhibition of CTLA-4 should increase the number of Teffs and reduce Tregs abundance in the tumor microenvironment, while anti-PD-1/PD-L1 therapy should prevent Teffs from being inhibited by malignant and stromal cells. CTLA-4 and PD-1 inhibitors can enhance the killing ability of T cells in different stages. The impact of combining these two therapies was first explored in melanoma and NSCLC.[106,107]
To date, immune checkpoints with promising results are being investigated as potential targets. These include immune checkpoints other than CTLA-4 or PD-1/PD-L1 antibodies. For example, TIGIT protein plays an important role in tumor immune escape and exists on a variety of immune cells, including Tregs, CD8+ T cells, and NK cells. In several mouse models of cancer, combining anti-PD-1 and anti-TIGIT antibodies increased immune responses.[109,110]
LAG-3 is expressed on Teffs, Tregs, and DCs.[111,112] Increased PD-1 and LAG-3 lead to T cell exhaustion and tolerance to self and tumor antigens. In a pre-clinical model, inhibiting the expression of PD-1 and LAG-3 eliminates the tumor. It was demonstrated that the overall response rate of combination of LAG-3 and PD-L1 inhibition was 13% in patients relapsing after anti-PD1 monotherapy. Blocking other immune checkpoints, such as B7S1, NKG2A, CD39, CD73, or the innate immune checkpoint CD47, also enhances anti-tumor responses induced by anti-PD-1/PD-L1 antibodies.
Combining with chemotherapy
Previous studies have shown that traditional chemotherapy drugs may activate the anti-tumor immune response in a variety of ways, including the induction of immunogenic tumor cell death, or disrupting tumor cells to evade immune surveillance. For example, tumor cells treated with anthracyclines activate TIL and DC through purine receptors or toll-like receptor 4, respectively. This changes these cells into “immunogenic tumor cells” and activates the anti-tumor immune response. Gemcitabine can also activate this immune effect by inducing apoptosis of tumor cells, enhancing the cross presentation of CD8+ T cells, and repairing antigen presentation defects in tumor invasive DCs.[118,119] Therefore, the combination of ICIs and chemotherapy may work together synergistically, but this effect is closely related to the type, dose, and regimen (simultaneous or sequential) of the chemotherapy drugs being used. KEYNOTE 189 and KEYNOTE 407, respectively explored the efficacy and safety of pembrolizumab combined with chemotherapy in first-line treatment in non-squamous and squamous NSCLC populations without sensitizing EGFR or ALK mutations, and this regimen has been approved by the FDA as first-line treatment.[120,121]
Combining with radiotherapy
Radiotherapy has direct cytotoxic effects on tumor cells by releasing tumor antigens and mediating anti-tumor response. In addition, ionizing radiation can induce the expression of MHC molecules, immune-activated chemokines and cytokines, and enhance the diversity of T-cell receptor in tumor-specific T cells.[122,123] Conventional radiotherapy combined with PD-1 inhibitors stimulates CD8+ T-cell mediated tumor clearance.[124,125] Stereotactic body radiation therapy (SBRT), an emerging radiotherapy technique, may also enhance anti-tumor effects. An animal model study investigating melanoma and kidney cancer found that PD-1 inhibitor combined with SBRT could make the primary tumor disappear and result in abscopal effect, which is known as eliciting anti-tumor activity in non-irradiated distant tumors.[126,127] Pembro-RT study showed that pembrolizumab combined with SBRT significantly improved the median PFS and objective response rate (ORR) in advanced stage NSCLC patients, compared with those treated with pembrolizumab monotherapy. These results suggest a potential combination of ICIs with radiotherapy as a first-line treatment method.
Combining with targeted therapy
Targeted therapy not only kills tumor cells, but also induces immune effect in tumor cells, host immune system, and tumor microenvironment, which has synergistic anti-cancer effects when combined with immunotherapy. Previous studies have shown that the anti-tumor immune response and tumor angiogenesis pathways have mutual influence. VEGF increases the immunosuppressive cell populations (such as Tregs and MDSCs), limits the maturation of APCs and the function of Teffs, and impedes invasion and migration of tumor-specific T cells and other immune effector cells into the tumor microenvironment. Therefore, VEGF inhibitors enhance the anti-tumor immune effect by antagonizing these multiple pathways. ICIs can also mediate tumor angiogenesis that leads to tumor necrosis and mass invasion of mononuclear cells into the tumor. Pre-clinical and clinical studies have confirmed that immunotherapy may have a synergistic effect with VEGF inhibitors, such as bevacizumab. IMPOWER150 evaluated atezolizumab combined with chemotherapy and bevacizumab as first-line treatment for metastatic non-squamous NSCLC. Results from this study revealed that the median PFS and OS of the combined treatment group were significantly longer than the PFS and OS observed in the group that only receiving chemotherapy and bevacizumab.
EGFR mutations are present in approximately 50% of NSCLC patients with Asian descent. Pre-clinical studies have shown that EGFR mutations can up-regulate PD-L1 expression in tumor cells through the phosphorylated extracellular signal-regulated kinase 1/2/p-c-Jun pathway, which induces apoptosis of T cells. Conversely, EGFR tyrosine kinase inhibitors (EGFR-TKIs) reverse the suppressive effects of the EGFR-PD-1/PD-L1 axis on T cells and increase expression levels of IFN-γ. Expression of PD-L1 in EGFR-TKI resistant cell lines was significantly higher, while PD-1 inhibitor could reduce the activity of EGFR-TKI resistant cell lines. Combining PD-1 inhibitor and EGFR-TKI may enhance CD8+ T cell activities, increase DC recruitment, and enhance ability of the immune system to kill tumor cells. Therefore, studies investigating the combination of PD-1/PD-L1 inhibitor and EGFR-TKI in the treatment of NSCLC are gradually being carried out. Preliminary results of the NCT02088112 study showed that the ORR of EGFR sensitive mutant NSCLC patients without TKI treatment before reached 77.8% to 80.0%, after durvalumab and gifitinib combined therapy, suggesting that this combination is worth further exploration.
BRAF mutations occur in approximately 5% to 10% of all human malignancies. Patients with BRAF V600E mutation substantially benefit from a combination treatment of small molecules that inhibit BRAF and MEK1. Targeted therapy against BRAF gene acts on the tumor microenvironment to promote anti-tumor immune response, by increasing antigen secretion, enhancing HLA expression, and promoting T cell infiltration.[134,135] A recent trial demonstrated the efficacy of combining BRAF and MEK inhibitors with atezolizumab in previously untreated metastatic melanoma patients with BRAF V600E mutations. Other trials found that BRAF inhibitors combined with ICIs such as PD-1 and CTLA-4 antibodies have prolonged clinical benefits.[133,137]
Combining with cytokines
Another strategy used by the tumor environment is immunosuppressive cytokines that avoid immune surveillance. For example, IDO, a tryptophan-metabolizing enzyme produced by cancer cells, TAMs and MDSCs, diminishes T cell anticancer functions and enhances Treg activity.[112,134] In a variety of tumors, the overexpression of IDO allows for the tumor evading immune monitoring and killing, and is associated with a poor prognosis.[138,139] Thus, the IDO inhibitor can slow tumor growth and enhance the anti-tumor immune effect, which may have a synergistic effect along with immunotherapy. Pre-clinical models where both IDO and PD-1 were inhibited have demonstrated successful results. At present, the combination of ICIs and an IDO inhibitor has been carried out successively, as shown in KEYNOTE-654.
Other immune factor antagonists have also been gradually applied into treatment with immunotherapy. In murine models and clinical studies, antibodies blocking co-activated molecules such as CD137, CD134, CD357, and CD40 have obtained long-term survival and these blockers can enhance the anti-tumor efficacy of the PD-1 antibody.[142–144] Another area of interest is to inhibit TAM activity. Colony stimulating factor 1 (CSF-1) is a cytokine secreted by macrophage, maintaining M2 polarization and inducing TAM proliferation. An anti-CSF-1R antibody reprograms TAM polarization and works synergistically with checkpoint blockade in pancreatic cancer.
Combining with epigenetic modifiers
Epigenetic modifications on the DNA of tumor cells may change the expression of immune-related genes, affecting antigen processing, presentation, and immune evasion.[16,17] Demethylating agents may enable re-expression of immune related genes and have potential therapeutic effects, especially when combined with immunotherapy. HDAC inhibitors increase expression of MHC and tumor-associated antigens, which work in synergy along with adoptive cell transfer therapy to improve anti-tumor responses in a murine melanoma model. In addition, hypomethylating agents increase CD80 expression on tumor cells and the number of tumor-infiltrating CD8+ T cells in a lymphoma model. These pre-clinical data suggest that it is possible to reverse epigenetic changes in cancer cells, which may enhance immune recognition and response to immunotherapy.
Combining with cancer vaccines
Cancer vaccines exhibit both antigenicity and immunogenicity. New tumor antigen epitopes are specifically expressed by related gene mutations in the tumor cells without immunologic tolerance, but with high immunogenicity. New tumor antigens of T cells effectively enhance anti-tumor ability. Two recent studies based on RNA and peptide were designed and produced individualized tumor vaccines used for patients with advanced melanoma. Results revealed that the tumor was effectively controlled, demonstrating the feasibility, safety, and immunogenicity of a personalized tumor vaccine.[147,148] Moreover, vaccines achieved better clinical results with checkpoint immunotherapy of PD-1.[149,150] So, the personalized tumor vaccine may open new opportunities for targeted immunotherapy for patients with malignant tumors.
Combining with CAR-T
CAR-T expresses antigen receptor of target cells on T cell surfaces through gene transfection technology. These cells recognize the antigen on the surface of tumor cells in an antigen specific and non-HLA dependent manner and have the ability to kill cells that are being targeted. Recent studies have shown a high success rate for the treatment of hematologic malignancies using CAR-T, particularly B-cell acute lymphoblastic leukemia and non-Hodgkin lymphoma. To translate this success in liquid cancers to solid cancers, CAR-T treatment will have to overcome two major obstacles. First, “on target/off tumor” toxicities must be improved by identifying new targets or by developing new targeting strategies such as dual targeting or rapid exchange of the target by using universal linkers.[153,154] Second, in solid tumors, the tumor microenvironment affects the activation of CAR-T cells and inhibits their anti-tumor function.[155,156] Researchers found that the combination of PD-1/PD-L1 antibody and CAR-T may result in resistance to the suppressive tumor microenvironment. In addition, the researchers designed CAR-T cells targeting both VEGF-2 and IL-2 to enhance the infiltration of CAR-T cells into the tumor. While both have serious immune-related adverse events, it will be interesting to see if combining ICIs may facilitate the use of CAR-T cell therapy in the treatment of solid tumors.
Recently, ICI has become a new breakthrough in the cancer treatment field. However, both primary and secondary resistance to these inhibitors poses great challenges to clinical practice. In-depth study investigating the mechanisms of resistance associated with ICIs and an exploration of how to overcome this resistance can not only increase our understanding of immunotherapy, but also greatly benefit the treatment of patients through the use of more accurate and personalized therapies. Uncovering predictive biomarkers, screening for the dominant population, exploring the use of ICIs combined with other treatments and the development of novel ICIs are effective strategies that may aid in overcoming resistance.
This work was supported by a grant from the Shanghai Jiao Tong University 2011 System Biomedical Collaborative Innovation Center Project (No. 15ZH4009).
Conflicts of interest
1. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin
2018; 68:394–424. doi: 10.3322/caac.21492.
2. Reck M, Rodríguez-Abreu D, Robinson AG, Hui R, Csőszi T, Fülöp A, et al. Pembrolizumab versus chemotherapy for PD-L1–positive non–small-cell lung cancer. N Engl J Med
2016; 375:1823–1833. doi: 10.1056/NEJMoa1606774.
3. Borghaei H, Paz-Ares L, Horn L, Spigel DR, Steins M, Ready NE, et al. Nivolumab versus docetaxel in advanced nonsquamous non-small-cell lung cancer. N Engl J Med
2015; 373:1627–1639. doi: 10.1056/NEJMoa1507643.
4. Hellmann MD, Paz-Ares L, Bernabe Caro R, Zurawski B, Kim SW, Carcereny Costa E, et al. Nivolumab plus ipilimumab in advanced non–small-cell lung cancer. N Engl J Med
2019; 381:2020–2031. doi: 10.1056/NEJMoa1910231.
5. Rittmeyer A, Barlesi F, Waterkamp D, Park K, Ciardiello F, von Pawel J, et al. Atezolizumab versus docetaxel in patients with previously treated non-small-cell lung cancer (OAK): a phase 3, open-label, multicentre randomised controlled trial. Lancet
2017; 389:255–265. doi: 10.1016/S0140-6736(16)32517-X.
6. Sharma P, Hu-Lieskovan S, Wargo JA, Ribas A. Primary, adaptive, and acquired resistance to cancer immunotherapy. Cell
2017; 168:707–723. doi: 10.1016/j.cell.2017.01.017.
7. Gubin MM, Zhang X, Schuster H, Caron E, Ward JP, Noguchi T, et al. Checkpoint blockade cancer immunotherapy targets tumour-specific mutant antigens. Nature
2014; 515:577–581. doi: 10.1038/nature13988.
8. Kantoff PW, Higano CS, Shore ND, Berger ER, Small EJ, Penson DF, et al. Sipuleucel-T immunotherapy for castration-resistant prostate cancer. N Engl J Med
2010; 363:411–422. doi: 10.1056/NEJMoa1001294.
9. Ott PA, Hu Z, Keskin DB, Shukla SA, Sun J, Bozym DJ, et al. An immunogenic personal neoantigen vaccine for patients with melanoma. Nature
2017; 547:217–221. doi: 10.1038/nature22991.
10. Keskin DB, Anandappa AJ, Sun J, Tirosh I, Mathewson ND, Li S, et al. Neoantigen vaccine generates intratumoral T cell responses in phase Ib glioblastoma trial. Nature
2019; 565:234–239. doi: 10.1038/s41586-018-0792-9.
11. Hilf N, Kuttruff-Coqui S, Frenzel K, Bukur V, Stevanović S, Gouttefangeas C, et al. Actively personalized vaccination trial for newly diagnosed glioblastoma. Nature
2019; 565:240–245. doi: 10.1038/s41586-018-0810-y.
12. Yadav M, Jhunjhunwala S, Phung QT, Lupardus P, Tanguay J, Bumbaca S, et al. Predicting immunogenic tumour mutations by combining mass spectrometry and exome sequencing. Nature
2014; 515:572–576. doi: 10.1038/nature14001.
13. Hugo W, Zaretsky JM, Sun L, Song C, Moreno BH, Hu-Lieskovan S, et al. Genomic and transcriptomic features of response to anti-PD-1 therapy in metastatic melanoma. Cell
2016; 165:35–44. doi: 10.1016/j.cell.2016.02.065.
14. Teo MY, Seier K, Ostrovnaya I, Regazzi AM, Kania BE, Moran MM, et al. Alterations in DNA damage response and repair genes as potential marker of clinical benefit from PD-1/PD-L1 blockade in advanced urothelial cancers. J Clin Oncol
2018; 36:1685–1694. doi: 10.1200/jco.2017.75.7740.
15. Le DT, Durham JN, Smith KN, Wang H, Bartlett BR, Aulakh LK, et al. Mismatch repair deficiency predicts response of solid tumors to PD-1 blockade. Science
2017; 357:409–413. doi: 10.1126/science.aan6733.
16. Karpf AR, Jones DA. Reactivating the expression of methylation silenced genes in human cancer. Oncogene
2002; 21:5496–5503. doi: 10.1038/sj.onc.1205602.
17. Kim HJ, Bae SC. Histone deacetylase inhibitors: molecular mechanisms of action and clinical trials as anti-cancer drugs. Am J Transl Res
18. Vo DD, Prins RM, Begley JL, Donahue TR, Morris LF, Bruhn KW, et al. Enhanced antitumor activity induced by adoptive T-cell transfer and adjunctive use of the histone deacetylase inhibitor LAQ824. Cancer Res
2009; 69:8693–8699. doi: 10.1158/0008-5472.Can-09-1456.
19. Spranger S, Bao R, Gajewski TF. Melanoma-intrinsic β-catenin signalling prevents anti-tumour immunity. Nature
2015; 523:231–235. doi: 10.1038/nature14404.
20. Hu-Lieskovan S, Homet Moreno B, Ribas A, Excluding T. Cells: is β-catenin the full story? Cancer Cell
2015; 27:749–750. doi: 10.1016/j.ccell.2015.05.014.
21. Böttcher JP, Bonavita E, Chakravarty P, Blees H, Cabeza-Cabrerizo M, Sammicheli S, et al. NK cells stimulate recruitment of cDC1 into the tumor microenvironment promoting cancer immune control. Cell
2018; 172:1022–1037.e14. doi: 10.1016/j.cell.2018.01.004.
22. Chen L, Diao L, Yang Y, Yi X, Rodriguez BL, Li Y, et al. CD38-mediated immunosuppression as a mechanism of tumor cell escape from PD-1/PD-L1 blockade. Cancer Discov
2018; 8:1156–1175. doi: 10.1158/2159-8290.CD-17-1033.
23. Kamphorst AO, Wieland A, Nasti T, Yang S, Zhang R, Barber DL, et al. Rescue of exhausted CD8 T cells by PD-1-targeted therapies is CD28-dependent. Science
2017; 355:1423–1427. doi: 10.1126/science.aaf0683.
24. Willemen Y, Van den Bergh JM, Lion E, Anguille S, Roelandts VA, Van Acker HH, et al. Engineering monocyte-derived dendritic cells to secrete interferon-α enhances their ability to promote adaptive and innate anti-tumor immune effector functions. Cancer Immunol Immunother
2015; 64:831–842. doi: 10.1007/s00262-015-1688-2.
25. Jain S, Ward MM, O’Loughlin J, Boeck M, Wiener N, Chuang E, et al. Incremental increase in VEGFR1+
hematopoietic progenitor cells and VEGFR2+
endothelial progenitor cells predicts relapse and lack of tumor response in breast cancer patients. Breast Cancer Res Treat
2012; 132:235–242. doi: 10.1007/s10549-011-1906-3.
26. Oishi S, Takano R, Tamura S, Tani S, Iwaizumi M, Hamaya Y, et al. M2 polarization of murine peritoneal macrophages induces regulatory cytokine production and suppresses T-cell proliferation. Immunology
2016; 149:320–328. doi: 10.1111/imm.12647.
27. Fu C, Jiang A. Dendritic cells and CD8 T cell immunity in tumor microenvironment. Front Immunol
2018; 9:3059doi: 10.3389/fimmu.2018.03059.
28. Harryvan TJ, Verdegaal EME, Hardwick JCH, Hawinkels LJAC, van der Burg SH. Targeting of the cancer-associated fibroblast-T-cell axis in solid malignancies. J Clin Med
2019; 8:1989doi: 10.3390/jcm8111989.
29. Eckert AW, Wickenhauser C, Salins PC, Kappler M, Bukur J, Seliger B. Clinical relevance of the tumor microenvironment and immune escape of oral squamous cell carcinoma. J Transl Med
2016; 14:85doi: 10.1186/s12967-016-0828-6.
30. Mariathasan S, Turley SJ, Nickles D, Castiglioni A, Yuen K, Wang Y, et al. TGFβ attenuates tumour response to PD-L1 blockade by contributing to exclusion of T cells. Nature
2018; 554:544–548. doi: 10.1038/nature25501.
31. Tauriello DVF, Palomo-Ponce S, Stork D, Berenguer-Llergo A, Badia-Ramentol J, Iglesias M, et al. TGFβ drives immune evasion in genetically reconstituted colon cancer metastasis. Nature
2018; 554:538–543. doi: 10.1038/nature25492.
32. Chakravarthy A, Khan L, Bensler NP, Bose P, De Carvalho DD. TGF-β-associated extracellular matrix genes link cancer-associated fibroblasts to immune evasion and immunotherapy failure. Nat Commun
2018; 9:4692doi: 10.1038/s41467-018-06654-8.
33. Lakins MA, Ghorani E, Munir H, Martins CP, Shields JD. Cancer-associated fibroblasts induce antigen-specific deletion of CD8 + T cells to protect tumour cells. Nat Commun
2018; 9:948doi: 10.1038/s41467-018-03347-0.
34. Highfill SL, Cui Y, Giles AJ, Smith JP, Zhang H, Morse E, et al. Disruption of CXCR2-mediated MDSC tumor trafficking enhances anti-PD1 efficacy. Sci Transl Med
2014; 6:237ra67doi: 10.1126/scitranslmed.3007974.
35. Sugiyama D, Nishikawa H, Maeda Y, Nishioka M, Tanemura A, Katayama I, et al. Anti-CCR4 mAb selectively depletes effector-type FoxP3+CD4+ regulatory T cells, evoking antitumor immune responses in humans. Proc Natl Acad Sci U S A
2013; 110:17945–17950. doi: 10.1073/pnas.1316796110.
36. Gil M, Komorowski MP, Seshadri M, Rokita H, McGray AJ, Opyrchal M, et al. CXCL12/CXCR4 blockade by oncolytic virotherapy inhibits ovarian cancer growth by decreasing immunosuppression and targeting cancer-initiating cells. J Immunol
2014; 193:5327–5337. doi: 10.4049/jimmunol.1400201.
37. Chang DK, Sui J, Geng S, Muvaffak A, Bai M, Fuhlbrigge RC, et al. Humanization of an anti-CCR4 antibody that kills cutaneous T-cell lymphoma cells and abrogates suppression by T-regulatory cells. Mol Cancer Ther
2012; 11:2451–2461. doi: 10.1158/1535-7163.MCT-12-0278.
38. Chen PL, Roh W, Reuben A, Cooper ZA, Spencer CN, Prieto PA, et al. Analysis of immune signatures in longitudinal tumor samples yields insight into biomarkers of response and mechanisms of resistance to immune checkpoint blockade. Cancer Discov
2016; 6:827–837. doi: 10.1158/2159-8290.CD-15-1545.
39. Ohm JE, Gabrilovich DI, Sempowski GD, Kisseleva E, Parman KS, Nadaf S, et al. VEGF inhibits T-cell development and may contribute to tumor-induced immune suppression. Blood
2003; 101:4878–4886. doi: 10.1182/blood-2002-07-1956.
40. Motz GT, Santoro SP, Wang LP, Garrabrant T, Lastra RR, Hagemann IS, et al. Tumor endothelium FasL establishes a selective immune barrier promoting tolerance in tumors. Nat Med
2014; 20:607–615. doi: 10.1038/nm.3541.
41. Voron T, Colussi O, Marcheteau E, Pernot S, Nizard M, Pointet AL, et al. VEGF-A modulates expression of inhibitory checkpoints on CD8+ T cells in tumors. J Exp Med
2015; 212:139–148. doi: 10.1084/jem.20140559.
42. Liu C, Peng W, Xu C, Lou Y, Zhang M, Wargo JA, et al. BRAF inhibition increases tumor infiltration by T cells and enhances the antitumor activity of adoptive immunotherapy in mice. Clin Cancer Res
2013; 19:393–403. doi: 10.1158/1078-0432.Ccr-12-1626.
43. Gao J, Shi LZ, Zhao H, Chen J, Xiong L, He Q, et al. Loss of IFN-gamma pathway genes in tumor cells as a mechanism of resistance to anti-CTLA-4 therapy. Cell
2016; 167:397–404.e9. doi: 10.1016/j.cell.2016.08.069.
44. Shin DS, Zaretsky JM, Escuin-Ordinas H, Garcia-Diaz A, Hu-Lieskovan S, Kalbasi A, et al. Primary resistance to PD-1 blockade mediated by JAK1/2 mutations. Cancer Discov
2017; 7:188–201. doi: 10.1158/2159-8290.CD-16-1223.
45. Zaretsky JM, Garcia-Diaz A, Shin DS, Escuin-Ordinas H, Hugo W, Hu-Lieskovan S, et al. Mutations associated with acquired resistance to PD-1 blockade in melanoma. N Engl J Med
2016; 375:819–829. doi: 10.1056/NEJMoa1604958.
46. Li LY, Zhang HR, Jiang ZL, Chang YZ, Shao CZ. Overexpression of dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin in dendritic cells protecting against aspergillosis. Chin Med J
2018; 131:2575–2582. doi: 10.4103/0366-6999.244103.
47. Platanias LC. Mechanisms of type-I- and type-II-interferon-mediated signalling. Nat Rev Immunol
2005; 5:375–386. doi: 10.1038/nri1604.
48. Benci JL, Xu B, Qiu Y, Wu TJ, Dada H, Twyman-Saint Victor C, et al. Tumor interferon signaling regulates a multigenic resistance program to immune checkpoint blockade. Cell
2016; 167:1540–1554.e12. doi: 10.1016/j.cell.2016.11.022.
49. Shankaran V, Ikeda H, Bruce AT, White JM, Swanson PE, Old LJ, et al. IFNgamma and lymphocytes prevent primary tumour development and shape tumour immunogenicity. Nature
2001; 410:1107–1111. doi: 10.1038/35074122.
50. Darnell JE Jr, Kerr IM, Stark GR. Jak-STAT pathways and transcriptional activation in response to IFNs and other extracellular signaling proteins. Science
1994; 264:1415–1421. doi: 10.1126/science.8197455.
51. Dunn GP, Bruce AT, Sheehan KC, Shankaran V, Uppaluri R, Bui JD, et al. A critical function for type I interferons in cancer immunoediting. Nat Immunol
2005; 6:722–729. doi: 10.1038/ni1213.
52. Shin DS, Ribas A. The evolution of checkpoint blockade as a cancer therapy: what's here, what's next. Curr Opin Immunol
2015; 33:23–35. doi: 10.1016/j.coi.2015.01.006.
53. Parsa AT, Waldron JS, Panner A, Crane CA, Parney IF, Barry JJ, et al. Loss of tumor suppressor PTEN function increases B7-H1 expression and immunoresistance in glioma. Nat Med
2007; 13:84–88. doi: 10.1038/nm1517.
54. Peng W, Chen JQ, Liu C, Malu S, Creasy C, Tetzlaff MT, et al. Loss of PTEN promotes resistance to T cell-mediated immunotherapy. Cancer Discov
2016; 6:202–216. doi: 10.1158/2159-8290.Cd-15-0283.
55. Xu C, Fillmore CM, Koyama S, Wu H, Zhao Y, Chen Z, et al. Loss of Lkb1 and Pten leads to lung squamous cell carcinoma with elevated PD-L1 expression. Cancer Cell
2014; 25:590–604. doi: 10.1016/j.ccr.2014.03.033.
56. Lee SJ, Jang BC, Lee SW, Yang YI, Suh SI, Park YM, et al. Interferon regulatory factor-1 is prerequisite to the constitutive expression and IFN-gamma-induced upregulation of B7-H1 (CD274). FEBS Lett
2006; 580:755–762. doi: 10.1016/j.febslet.2005.12.093.
57. Akbay EA, Koyama S, Carretero J, Altabef A, Tchaicha JH, Christensen CL, et al. Activation of the PD-1 pathway contributes to immune escape in EGFR-driven lung tumors. Cancer Discov
2013; 3:1355–1363. doi: 10.1158/2159-8290.CD-13-0310.
58. Chen N, Fang W, Zhan J, Hong S, Tang Y, Kang S, et al. Upregulation of PD-L1 by EGFR activation mediates the immune escape in EGFR-Driven NSCLC: implication for optional immune targeted therapy for NSCLC patients with EGFR mutation. J Thorac Oncol
2015; 10:910–923. doi: 10.1097/JTO.0000000000000500.
59. Prendergast GC, Malachowski WP, DuHadaway JB, Muller AJ. Discovery of IDO1 inhibitors: from bench to bedside. Cancer Res
2017; 77:6795–6811. doi: 10.1158/0008-5472.CAN-17-2285.
60. Chanmee T, Ontong P, Konno K, Itano N. Tumor-associated macrophages as major players in the tumor microenvironment. Cancers (Basel)
2014; 6:1670–1690. doi: 10.3390/cancers6031670.
61. Biswas SK, Mantovani A. Macrophage plasticity and interaction with lymphocyte subsets: cancer as a paradigm. Nat Immunol
2010; 11:889–896. doi: 10.1038/ni.1937.
62. Hu W, Li X, Zhang C, Yang Y, Jiang J, Wu C. Tumor-associated macrophages in cancers. Clin Transl Oncol
2016; 18:251–258. doi: 10.1007/s12094-015-1373-0.
63. Fritz JM, Tennis MA, Orlicky DJ, Lin H, Ju C, Redente EF, et al. Depletion of tumor-associated macrophages slows the growth of chemically induced mouse lung adenocarcinomas. Front Immunol
2014; 5:587doi: 10.3389/fimmu.2014.00587.
64. Ries CH, Cannarile MA, Hoves S, Benz J, Wartha K, Runza V, et al. Targeting tumor-associated macrophages with anti-CSF-1R antibody reveals a strategy for cancer therapy. Cancer Cell
2014; 25:846–859. doi: 10.1016/j.ccr.2014.05.016.
65. Ruffell B, Chang-Strachan D, Chan V, Rosenbusch A, Ho CM, Pryer N, et al. Macrophage IL-10 blocks CD8+ T cell-dependent responses to chemotherapy by suppressing IL-12 expression in intratumoral dendritic cells. Cancer Cell
2014; 26:623–637. doi: 10.1016/j.ccell.2014.09.006.
66. De Palma M, Lewis CE. Macrophage regulation of tumor responses to anticancer therapies. Cancer Cell
2013; 23:277–286. doi: 10.1016/j.ccr.2013.02.013.
67. Ruffell B, Coussens LM. Macrophages and therapeutic resistance in cancer. Cancer Cell
2015; 27:462–472. doi: 10.1016/j.ccell.2015.02.015.
68. Kuang DM, Zhao Q, Peng C, Xu J, Zhang JP, Wu C, et al. Activated monocytes in peritumoral stroma of hepatocellular carcinoma foster immune privilege and disease progression through PD-L1. J Exp Med
2009; 206:1327–1337. doi: 10.1084/jem.20082173.
69. Kryczek I, Zou L, Rodriguez P, Zhu G, Wei S, Mottram P, et al. B7-H4 expression identifies a novel suppressive macrophage population in human ovarian carcinoma. J Exp Med
2006; 203:871–881. doi: 10.1084/jem.20050930.
70. Le DT, Lutz E, Uram JN, Sugar EA, Onners B, Solt S, et al. Evaluation of ipilimumab in combination with allogeneic pancreatic tumor cells transfected with a GM-CSF gene in previously treated pancreatic cancer. J Immunother
2013; 36:382–389. doi: 10.1097/CJI.0b013e31829fb7a2.
71. Zhu Y, Knolhoff BL, Meyer MA, Nywening TM, West BL, Luo J, et al. CSF1/CSF1R blockade reprograms tumor-infiltrating macrophages and improves response to T-cell checkpoint immunotherapy in pancreatic cancer models. Cancer Res
2014; 74:5057–5069. doi: 10.1158/0008-5472.Can-13-3723.
72. Mok S, Koya RC, Tsui C, Xu J, Robert L, Wu L, et al. Inhibition of CSF-1 receptor improves the antitumor efficacy of adoptive cell transfer immunotherapy. Cancer Res
2014; 74:153–161. doi: 10.1158/0008-5472.Can-13-1816.
73. Talmadge JE, Gabrilovich DI. History of myeloid-derived suppressor cells. Nat Rev Cancer
2013; 13:739–752. doi: 10.1038/nrc3581.
74. Wesolowski R, Markowitz J, Carson WE 3rd. Myeloid derived suppressor cells - a new therapeutic target in the treatment of cancer. J Immunother Cancer
2013; 1:10doi: 10.1186/2051-1426-1-10.
75. Yang L, DeBusk LM, Fukuda K, Fingleton B, Green-Jarvis B, Shyr Y, et al. Expansion of myeloid immune suppressor Gr+CD11b+ cells in tumor-bearing host directly promotes tumor angiogenesis. Cancer Cell
2004; 6:409–421. doi: 10.1016/j.ccr.2004.08.031.
76. Yang L, Huang J, Ren X, Gorska AE, Chytil A, Aakre M, et al. Abrogation of TGF beta signaling in mammary carcinomas recruits Gr-1+CD11b+ myeloid cells that promote metastasis. Cancer Cell
2008; 13:23–35. doi: 10.1016/j.ccr.2007.12.004.
77. Solito S, Falisi E, Diaz-Montero CM, Doni A, Pinton L, Rosato A, et al. A human promyelocytic-like population is responsible for the immune suppression mediated by myeloid-derived suppressor cells. Blood
2011; 118:2254–2265. doi: 10.1182/blood-2010-12-325753.
78. Meyer C, Cagnon L, Costa-Nunes CM, Baumgaertner P, Montandon N, Leyvraz L, et al. Frequencies of circulating MDSC correlate with clinical outcome of melanoma patients treated with ipilimumab. Cancer Immunol Immunother
2014; 63:247–257. doi: 10.1007/s00262-013-1508-5.
79. Kodumudi KN, Weber A, Sarnaik AA, Pilon-Thomas S. Blockade of myeloid-derived suppressor cells after induction of lymphopenia improves adoptive T cell therapy in a murine model of melanoma. J Immunol
2012; 189:5147–5154. doi: 10.4049/jimmunol.1200274.
80. Laborde RR, Lin Y, Gustafson MP, Bulur PA, Dietz AB. Cancer vaccines in the world of immune suppressive monocytes (CD14(+)HLA-DR(lo/neg) cells): the gateway to improved responses. Front Immunol
2014; 5:147doi: 10.3389/fimmu.2014.00147.
81. Kaneda MM, Messer KS, Ralainirina N, Li H, Leem CJ, Gorjestani S, et al. PI3Kgamma is a molecular switch that controls immune suppression. Nature
2016; 539:437–442. doi: 10.1038/nature19834.
82. De Henau O, Rausch M, Winkler D, Campesato LF, Liu C, Cymerman DH, et al. Overcoming resistance to checkpoint blockade therapy by targeting PI3Kgamma in myeloid cells. Nature
2016; 539:443–447. doi: 10.1038/nature20554.
83. Rudensky AY. Regulatory T cells and Foxp3. Immunol Rev
2011; 241:260–268. doi: 10.1111/j.1600-065X.2011.01018.x.
84. Oida T, Zhang X, Goto M, Hachimura S, Totsuka M, Kaminogawa S, et al. CD4+CD25- T cells that express latency-associated peptide on the surface suppress CD4+CD45RBhigh-induced colitis by a TGF-beta-dependent mechanism. J Immunol
2003; 170:2516–2522. doi: 10.4049/jimmunol.170.5.2516.
85. Sundstedt A, O’Neill EJ, Nicolson KS, Wraith DC. Role for IL-10 in suppression mediated by peptide-induced regulatory T cells in vivo. J Immunol
2003; 170:1240–1248. doi: 10.4049/jimmunol.170.3.1240.
86. Sakaguchi S, Yamaguchi T, Nomura T, Ono M. Regulatory T cells and immune tolerance. Cell
2008; 133:775–787. doi: 10.1016/j.cell.2008.05.009.
87. Woo EY, Yeh H, Chu CS, Schlienger K, Carroll RG, Riley JL, et al. Cutting edge: regulatory T cells from lung cancer patients directly inhibit autologous T cell proliferation. J Immunol
2002; 168:4272–4276. doi: 10.4049/jimmunol.168.9.4272.
88. Ormandy LA, Hillemann T, Wedemeyer H, Manns MP, Greten TF, Korangy F. Increased populations of regulatory T cells in peripheral blood of patients with hepatocellular carcinoma. Cancer Res
2005; 65:2457–2464. doi: 10.1158/0008-5472.Can-04-3232.
89. Chaudhary B, Elkord E. Regulatory T cells in the tumor microenvironment and cancer progression: role and therapeutic targeting. Vaccines (Basel)
2016; 4:28doi: 10.3390/vaccines4030028.
90. Linehan DC, Goedegebuure PS. CD25+ CD4+ regulatory T-cells in cancer. Immunol Res
2005; 32:155–168. doi: 10.1385/ir:32:1-3:155.
91. Viehl CT, Moore TT, Liyanage UK, Frey DM, Ehlers JP, Eberlein TJ, et al. Depletion of CD4+CD25+ regulatory T cells promotes a tumor-specific immune response in pancreas cancer-bearing mice. Ann Surg Oncol
2006; 13:1252–1258. doi: 10.1245/s10434-006-9015-y.
92. Quezada SA, Peggs KS, Curran MA, Allison JP. CTLA4 blockade and GM-CSF combination immunotherapy alters the intratumor balance of effector and regulatory T cells. J Clin Invest
2006; 116:1935–1945. doi: 10.1172/jci27745.
93. Simpson TR, Li F, Montalvo-Ortiz W, Sepulveda MA, Bergerhoff K, Arce F, et al. Fc-dependent depletion of tumor-infiltrating regulatory T cells co-defines the efficacy of anti-CTLA-4 therapy against melanoma. J Exp Med
2013; 210:1695–1710. doi: 10.1084/jem.20130579.
94. Hamid O, Schmidt H, Nissan A, Ridolfi L, Aamdal S, Hansson J, et al. A prospective phase II trial exploring the association between tumor microenvironment biomarkers and clinical activity of ipilimumab in advanced melanoma. J Transl Med
2011; 9:204doi: 10.1186/1479-5876-9-204.
95. D’Urso CM, Wang ZG, Cao Y, Tatake R, Zeff RA, Ferrone S. Lack of HLA class I antigen expression by cultured melanoma cells FO-1 due to a defect in B2m gene expression. J Clin Invest
1991; 87:284–292. doi: 10.1172/JCI114984.
96. Restifo NP, Marincola FM, Kawakami Y, Taubenberger J, Yannelli JR, Rosenberg SA. Loss of functional beta 2-microglobulin in metastatic melanomas from five patients receiving immunotherapy. J Natl Cancer Inst
1996; 88:100–108. doi: 10.1093/jnci/88.2.100.
97. Tran E, Robbins PF, Lu YC, Prickett TD, Gartner JJ, Jia L, et al. T-cell transfer therapy targeting mutant KRAS in cancer. N Engl J Med
2016; 375:2255–2262. doi: 10.1056/NEJMoa1609279.
98. Makker A, Goel MM, Mahdi AA, Bhatia V, Das V, Agarwal A, et al. PI3K/Akt/mTOR signaling & its regulator tumour suppressor genes PTEN & LKB1 in human uterine leiomyomas. Indian J Med Res
2016; 143:S112–S119. doi: 10.4103/0971-5916.191808.
99. Gorman JV, Colgan JD. Regulation of T cell responses by the receptor molecule Tim-3. Immunol Res
2014; 59:56–65. doi: 10.1007/s12026-014-8524-1.
100. Limagne E, Richard C, Thibaudin M, Fumet JD, Truntzer C, Lagrange A, et al. Tim-3/galectin-9 pathway and mMDSC control primary and secondary resistances to PD-1 blockade in lung cancer patients. Oncoimmunology
2019; 8:e1564505doi: 10.1080/2162402X.2018.1564505.
101. Koyama S, Akbay EA, Li YY, Herter-Sprie GS, Buczkowski KA, Richards WG, et al. Adaptive resistance to therapeutic PD-1 blockade is associated with upregulation of alternative immune checkpoints. Nat Commun
2016; 7:10501doi: 10.1038/ncomms10501.
102. Catakovic K, Klieser E, Neureiter D, Geisberger R. T cell exhaustion: from pathophysiological basics to tumor immunotherapy. Cell Commun Signal
2017; 15:1doi: 10.1186/s12964-016-0160-z.
103. Nowak EC, Lines JL, Varn FS, Deng J, Sarde A, Mabaera R, et al. Immunoregulatory functions of VISTA. Immunol Rev
2017; 276:66–79. doi: 10.1111/imr.12525.
104. Villarroel-Espindola F, Yu X, Datar I, Mani N, Sanmamed M, Velcheti V, et al. Spatially resolved and quantitative analysis of VISTA/PD-1H as a novel immunotherapy target in human non-small cell lung cancer. Clin Cancer Res
2018; 24:1562–1573. doi: 10.1158/1078-0432.Ccr-17-2542.
105. Wei SC, Levine JH, Cogdill AP, Zhao Y, Anang NAS, Andrews MC, et al. Distinct cellular mechanisms underlie anti-CTLA-4 and anti-PD-1 checkpoint blockade. Cell
2017; 170:1120–1133.e17. doi: 10.1016/j.cell.2017.07.024.
106. Hellmann MD, Ciuleanu TE, Pluzanski A, Lee JS, Otterson GA, Audigier-Valette C, et al. Nivolumab plus Ipilimumab in lung cancer with a high tumor mutational burden. N Engl J Med
2018; 378:2093–2104. doi: 10.1056/NEJMoa1801946.
107. Larkin J, Hodi FS, Wolchok JD. Combined nivolumab and ipilimumab or monotherapy in untreated melanoma. N Engl J Med
2015; 373:1270–1271. doi: 10.1056/NEJMc1509660.
108. Manieri NA, Chiang EY, Grogan JL. TIGIT: a key inhibitor of the cancer immunity cycle. Trends Immunol
2017; 38:20–28. doi: 10.1016/j.it.2016.10.002.
109. Hung AL, Maxwell R, Theodros D, Belcaid Z, Mathios D, Luksik AS, et al. TIGIT and PD-1 dual checkpoint blockade enhances antitumor immunity and survival in GBM. Oncoimmunology
2018; 7:e1466769doi: 10.1080/2162402X.2018.1466769.
110. Zhang Q, Bi J, Zheng X, Chen Y, Wang H, Wu W, et al. Blockade of the checkpoint receptor TIGIT prevents NK cell exhaustion and elicits potent anti-tumor immunity. Nat Immunol
2018; 19:723–732. doi: 10.1038/s41590-018-0132-0.
111. Lui Y, Davis SJ. LAG-3: a very singular immune checkpoint. Nat Immunol
2018; 19:1278–1279. doi: 10.1038/s41590-018-0257-1.
112. Popovic A, Jaffee EM, Zaidi N. Emerging strategies for combination checkpoint modulators in cancer immunotherapy. J Clin Invest
2018; 128:3209–3218. doi: 10.1172/JCI120775.
113. Li J, Lee Y, Li Y, Jiang Y, Lu H, Zang W, et al. Co-inhibitory molecule B7 superfamily member 1 expressed by tumor-infiltrating myeloid cells induces dysfunction of anti-tumor CD8(+) T cells. Immunity
2018; 48:773–786.e5. doi: 10.1016/j.immuni.2018.03.018.
114. André P, Denis C, Soulas C, Bourbon-Caillet C, Lopez J, Arnoux T, et al. Anti-NKG2A mAb is a checkpoint inhibitor that promotes anti-tumor immunity by unleashing both T and NK cells. Cell
2018; 175:1731–1743.e13. doi: 10.1016/j.cell.2018.10.014.
115. Sade-Feldman M, Yizhak K, Bjorgaard SL, Ray JP, de Boer CG, Jenkins RW, et al. Defining T cell states associated with response to checkpoint immunotherapy in melanoma. Cell
2018; 175:998–1013.e20. doi: 10.1016/j.cell.2018.10.038.
116. Hay CM, Sult E, Huang Q, Mulgrew K, Fuhrmann SR, McGlinchey KA, et al. Targeting CD73 in the tumor microenvironment with MEDI9447. Oncoimmunology
2016; 5:e1208875doi: 10.1080/2162402x.2016.1208875.
117. Liu X, Liu L, Ren Z, Yang K, Xu H, Luan Y, et al. Dual targeting of innate and adaptive checkpoints on tumor cells limits immune evasion. Cell Rep
2018; 24:2101–2111. doi: 10.1016/j.celrep.2018.07.062.
118. Pfirschke C, Engblom C, Rickelt S, Cortez-Retamozo V, Garris C, Pucci F, et al. Immunogenic chemotherapy sensitizes tumors to checkpoint blockade therapy. Immunity
2016; 44:343–354. doi: 10.1016/j.immuni.2015.11.024.
119. Galluzzi L, Buque A, Kepp O, Zitvogel L, Kroemer G. Immunological effects of conventional chemotherapy and targeted anticancer agents. Cancer Cell
2015; 28:690–714. doi: 10.1016/j.ccell.2015.10.012.
120. Gandhi L, Rodriguez-Abreu D, Gadgeel S, Esteban E, Felip E, De Angelis F, et al. Pembrolizumab plus chemotherapy in metastatic non-small-cell lung cancer. N Engl J Med
2018; 378:2078–2092. doi: 10.1056/NEJMoa1801005.
121. Shen K, Cui J, Wei Y, Chen X, Liu G, Gao X, et al. Effectiveness and safety of PD-1/PD-L1 or CTLA4 inhibitors combined with chemotherapy as a first-line treatment for lung cancer: a meta-analysis. J Thorac Dis
2018; 10:6636–6652. doi: 10.21037/jtd.2018.11.72.
122. Martinov T, Fife BT. Fractionated radiotherapy combined with PD-1 pathway blockade promotes CD8 T cell-mediated tumor clearance for the treatment of advanced malignancies. Ann Transl Med
2016; 4:82doi: 10.3978/j.issn.2305-5839.2016.01.13.
123. Sharabi AB, Nirschl CJ, Kochel CM, Nirschl TR, Francica BJ, Velarde E, et al. Stereotactic radiation therapy augments antigen-specific PD-1-mediated antitumor immune responses via cross-presentation of tumor antigen. Cancer Immunol Res
2015; 3:345–355. doi: 10.1158/2326-6066.CIR-14-0196.
124. Deng L, Liang H, Burnette B, Weicheslbaum RR, Fu YX. Radiation and anti-PD-L1 antibody combinatorial therapy induces T cell-mediated depletion of myeloid-derived suppressor cells and tumor regression. Oncoimmunology
2014; 3:e28499doi: 10.4161/onci.28499.
125. Deng L, Liang H, Burnette B, Beckett M, Darga T, Weichselbaum RR, et al. Irradiation and anti-PD-L1 treatment synergistically promote antitumor immunity in mice. J Clin Invest
2014; 124:687–695. doi: 10.1172/jci67313.
126. Siva S, MacManus MP, Martin RF, Martin OA. Abscopal effects of radiation therapy: a clinical review for the radiobiologist. Cancer Lett
2015; 356:82–90. doi: 10.1016/j.canlet.2013.09.018.
127. Park SS, Dong H, Liu X, Harrington SM, Krco CJ, Grams MP, et al. PD-1 restrains radiotherapy-induced abscopal effect. Cancer Immunol Res
2015; 3:610–619. doi: 10.1158/2326-6066.CIR-14-0138.
128. Theelen WSME, Peulen HMU, Lalezari F, van der Noort V, de Vries JF, Aerts JGJV, et al. Effect of pembrolizumab after stereotactic body radiotherapy vs pembrolizumab alone on tumor response in patients with advanced non-small cell lung cancer: results of the PEMBRO-RT phase 2 randomized clinical trial. JAMA Oncol
2019; 5:1276–1282. doi: 10.1001/jamaoncol.2019.1478.
129. Ott PA, Hodi FS, Buchbinder EI. Inhibition of immune checkpoints and vascular endothelial growth factor as combination therapy
for metastatic melanoma: an overview of rationale, preclinical evidence, and initial clinical data. Front Oncol
2015; 5:202doi: 10.3389/fonc.2015.00202.
130. Socinski MA, Jotte RM, Cappuzzo F, Orlandi F, Stroyakovskiy D, Nogami N, et al. Atezolizumab for first-line treatment of metastatic nonsquamous NSCLC. N Engl J Med
2018; 378:2288–2301. doi: 10.1056/NEJMoa1716948.
131. Gibbons DL, Chow LQ, Kim DW, Kim SW, Yeh T, Song X, et al. 57O Efficacy, safety and tolerability of MEDI4736 (durvalumab [D]), a human IgG1 anti-programmed cell death-ligand-1 (PD-L1) antibody, combined with gefitinib (G): a phase I expansion in TKI-naïve patients (pts) with EGFR mutant NSCLC. J Thorac Oncol
2016; 11: (4 Supplement): S79doi: 10.1016/S1556-0864(16)30171-X.
132. Morris V, Kopetz S. BRAF inhibitors in clinical oncology. F1000Prime Rep
2013; 5:11doi: 10.12703/P5-11.
133. Eggermont A, Robert C, Ribas A. The new era of adjuvant therapies for melanoma. Nat Rev Clin Oncol
2018; 15:535–536. doi: 10.1038/s41571-018-0048-5.
134. Chowdhury PS, Chamoto K, Honjo T. Combination therapy
strategies for improving PD-1 blockade efficacy: a new era in cancer immunotherapy. J Intern Med
2018; 283:110–120. doi: 10.1111/joim.12708.
135. Sullivan RJ, Hamid O, Gonzalez R, Infante JR, Patel MR, Hodi FS, et al. Atezolizumab plus cobimetinib and vemurafenib in BRAF-mutated melanoma patients. Nat Med
2019; 25:929–935. doi: 10.1038/s41591-019-0474-7.
136. Amaria RN, Prieto PA, Tetzlaff MT, Reuben A, Andrews MC, Ross MI, et al. Neoadjuvant plus adjuvant dabrafenib and trametinib versus standard of care in patients with high-risk, surgically resectable melanoma: a single-centre, open-label, randomised, phase 2 trial. Lancet Oncol
2018; 19:181–193. doi: 10.1016/S1470-2045(18)30015-9.
137. Ribas A, Lawrence D, Atkinson V, Agarwal S, Miller WH, Carlino MS, et al. Combined BRAF and MEK inhibition with PD-1 blockade immunotherapy in BRAF-mutant melanoma. Nat Med
2019; 25:936–940. doi: 10.1038/s41591-019-0476-5.
138. Austin CJ, Rendina LM. Targeting key dioxygenases in tryptophan–kynurenine metabolism for immunomodulation and cancer chemotherapy. Drug Discov Today
2015; 20:609–617. doi: 10.1016/j.drudis.2014.11.007.
139. Spranger S, Koblish HK, Horton B, Scherle PA, Newton R, Gajewski TF. Mechanism of tumor rejection with doublets of CTLA-4, PD-1/PD-L1, or IDO blockade involves restored IL-2 production and proliferation of CD8(+) T cells directly within the tumor microenvironment. J Immunother Cancer
2014; 2:3doi: 10.1186/2051-1426-2-3.
140. Zhu M, Dancsok AR, Nielsen TO. Indoleamine dioxygenase inhibitors: clinical rationale and current development. Curr Oncol Rep
2019; 21:2doi: 10.1007/s11912-019-0750-1.
141. Paz-Ares L, Rubin S, Zhao Y, Xu L, Samkari A, Awad M. 195TiP A phase 3, randomized, double-blind study of epacadostat plus pembrolizumab vs pembrolizumab as first-line therapy for metastatic non-small cell lung cancer (mNSCLC) expressing high PD-L1 levels (ECHO-305/KEYNOTE-654). J Thoracic Oncol
2018; 13: (4 Supplement): S116–S117. doi: 10.1016/S1556-0864(18)30468-4.
142. Wei H, Zhao L, Li W, Fan K, Qian W, Hou S, et al. Combinatorial PD-1 blockade and CD137 activation has therapeutic efficacy in murine cancer models and synergizes with cisplatin. PLoS One
2013; 8:e84927doi: 10.1371/journal.pone.0084927.
143. Lu L, Xu X, Zhang B, Zhang R, Ji H, Wang X. Combined PD-1 blockade and GITR triggering induce a potent antitumor immunity in murine cancer models and synergizes with chemotherapeutic drugs. J Transl Med
2014; 12:36doi: 10.1186/1479-5876-12-36.
144. Zippelius A, Schreiner J, Herzig P, Müller P. Induced PD-L1 expression mediates acquired resistance to agonistic anti-CD40 treatment. Cancer Immunol Res
2015; 3:236–244. doi: 10.1158/2326-6066.CIR-14-0226.
145. Héninger E, Krueger TE, Lang JM. Augmenting antitumor immune responses with epigenetic modifying agents. Front Immunol
2015; 6:29doi: 10.3389/fimmu.2015.00029.
146. Wang LX, Mei ZY, Zhou JH, Yao YS, Li YH, Xu YH, et al. Low dose decitabine treatment induces CD80 expression in cancer cells and stimulates tumor specific cytotoxic T lymphocyte responses. PLoS One
2013; 8:e62924doi: 10.1371/journal.pone.0062924.
147. Granot-Matok Y, Kon E, Dammes N, Mechtinger G, Peer D. Therapeutic mRNA delivery to leukocytes. J Control Release
2019; 305:165–175. doi: 10.1016/j.jconrel.2019.05.032.
148. Sahin U, Derhovanessian E, Miller M, Kloke BP, Simon P, Löwer M, et al. Personalized RNA mutanome vaccines mobilize poly-specific therapeutic immunity against cancer. Nature
2017; 547:222–226. doi: 10.1038/nature23003.
149. Bialkowski L, Van der Jeught K, Bevers S, Tjok Joe P, Renmans D, Heirman C, et al. Immune checkpoint blockade combined with IL-6 and TGF-beta inhibition improves the therapeutic outcome of mRNA-based immunotherapy. Int J Cancer
2018; 143:686–698. doi: 10.1002/ijc.31331.
150. Kos S, Lopes A, Preat V, Cemazar M, Lampreht Tratar U, Ucakar B, et al. Intradermal DNA vaccination combined with dual CTLA-4 and PD-1 blockade provides robust tumor immunity in murine melanoma. PLoS One
2019; 14:e0217762doi: 10.1371/journal.pone.0217762.
151. Davila ML, Brentjens R, Wang X, Rivière I, Sadelain M. How do CARs work?: Early insights from recent clinical studies targeting CD19. Oncoimmunology
2012; 1:1577–1583. doi: 10.4161/onci.22524.
152. D’Aloia MM, Zizzari IG, Sacchetti B, Pierelli L, Alimandi M. CAR-T cells: the long and winding road to solid tumors. Cell Death Dis
2018; 9:282doi: 10.1038/s41419-018-0278-6.
153. Chen N, Li X, Chintala NK, Tano ZE, Adusumilli PS. Driving CARs on the uneven road of antigen heterogeneity in solid tumors. Curr Opin Immunol
2018; 51:103–110. doi: 10.1016/j.coi.2018.03.002.
154. Lu YJ, Chu H, Wheeler LW, Nelson M, Westrick E, Matthaei JF, et al. Preclinical evaluation of bispecific adaptor molecule controlled folate receptor CAR-T cell therapy with special focus on pediatric malignancies. Front Oncol
2019; 9:151doi: 10.3389/fonc.2019.00151.
155. Gay F, D’Agostino M, Giaccone L, Genuardi M, Festuccia M, Boccadoro M, et al. Immuno-oncologic approaches: CAR-T cells and checkpoint inhibitors. Clin Lymphoma Myeloma Leuk
2017; 17:471–478. doi: 10.1016/j.clml.2017.06.014.
156. Lim WA, June CH. The principles of engineering immune cells to treat cancer. Cell
2017; 168:724–740. doi: 10.1016/j.cell.2017.01.016.
157. Liu J, Zhou G, Zhang L, Zhao Q. Building potent chimeric antigen receptor T cells with CRISPR genome editing. Front Immunol
2019; 10:456doi: 10.3389/fimmu.2019.00456.
158. Chinnasamy D, Yu Z, Kerkar SP, Zhang L, Morgan RA, Restifo NP, et al. Local delivery of interleukin-12 using T cells targeting VEGF receptor-2 eradicates multiple vascularized tumors in mice. Clin Cancer Res
2012; 18:1672–1683. doi: 10.1158/1078-0432.CCR-11-3050.