Moreover, basal levels of autophagy are necessary to maintain a normal number of peripheral B cells and their survival after ligand lipopolysaccharide LPS stimulation that drives plasmablast differentiation and specific IgM and IgG production [ 94 ].

Autophagy-related genes, especially, Atg5 is essential for B cell development [ 44 ]. The autophagy-deficient B cells lack the ability to produce antibodies and cytokines. Unexpectedly, it has been reported that unfractionated splenocytes produce a higher level of antibodies and cytokines than purified B cells, DRibbles stimulation upregulates CD40L expression on macrophages, resulting in increased level of CD40 on B cells.

Moreover, macrophages are able to enhance the antigen presentation function of B cells for specific T cell stimulation [ 95 ]. During autophagy induction, LC3 molecules link to nascent autophagosome membranes in memory B cells, contributing to LC3 abundance and increasing the percentage of autophagosome-containing cells in memory B cells compared with naive B cells, which can explain why the memory B cells have a longer lifespan, and the lack of autophagy in B cells does not affect primary alloantibody responses, but affects secondary alloantibody production [ 96 ].

Therefore, pharmacological inhibitors of autophagy impair antibody recall response. However, Atg5-deficient DCs exhibited unimpaired production of IL, IL-6, and TNF-α [ 98 ].

Autophagy is important for the presentation of cytosolic antigens on MHCII and efficient cross-presentation of soluble antigen. A study revealed that Atg5 deficiency in DCs impaired antigen presentation through the MHCII pathway.

TAMs density is significantly higher than the surrounding normal tissues. Generally, TAMs first originate from monocytes that are recruited into tumors by chemoattractants, including chemokines and cytokines released from both tumor cells and stromal cells.

When monocytes are stimulated to differentiate into macrophages, colony-stimulating factor 1 CSF1 increases the expression and phosphorylation status of ULK1, thus contributing to increased induction of autophagy [ ]. CSF2 is also able to promote monocyte survival and differentiation into macrophages.

The differentiation signal helps Beclin1 release from Bcl-2 by activating JNK and blocks Atg5 cleavage, thus stimulating autophagy, whereas blockade of autophagy has an inhibitory effect on CSF2-induced monocyte differentiation into macrophages [ ].

Autophagy also plays a key role in macrophage polarization. The inhibition of macrophage autophagy promotes M1 polarization, resulting in increased pro-inflammatory cytokine secretion. Then, M1 macrophages stimulate a Th1 response against intracellular microorganisms and tumor cells by activating immune responses, whereas the induction of autophagy promotes M2 polarization.

In the tumor microenvironment, autophagy is triggered by binding of IL-6 and CCL2 to interleukin 6 receptor IL-6R and CCR2, respectively, which is essential for macrophage polarizaton to the M2 phenotype, resulting in increased anti-inflammatory cytokine secretion, which promotes the fading of inflammation as well as tissue repair and remodeling, but M2 macrophages are immunosuppressive cells [ , , , ].

Targeting the autophagy which regulates macrophage polarization toward the M1 phenotype should be a promising anti-tumor strategy. Autophagy has a negative effect on the development of neutrophils. Deficient autophagy indicates an increased proliferation rate in the neutrophil precursor cells of the bone marrow and accelerates the process of neutrophil differentiation, resulting in the accumulation of mature neutrophils in the bone marrow, blood, spleen, and lymph nodes.

Pharmacological inhibition of p38 MAPK or mTORC1 can induce autophagy in neutrophilic precursor cells and block their differentiation [ ]. However, autophagy is required for neutrophil-mediated inflammation.

Autophagy deficiency in neutrophils leads to reduced nicotinamide adenine dinucleotide phosphate NADPH oxidase-mediated ROS production and further contribute to reduced degranulation [ ]. Recent studies have revealed that autophagic activity is also required for the release of neutrophil extracellular traps NETs , representing a distinct form of active neutrophil death, namely NETosis.

NET formation requires both autophagic activity and ROS production. Inhibition of the mTOR pathway accelerates the rate of NET release and stimulates ROS production following neutrophil stimulation with the bacteria-derived peptide formyl-Met-Leu-Phe fMLP [ ].

Tumor-derived neutrophils exhibit much higher levels of LC3-II and have more autophagosomes than their counterparts from blood. There is evidence that enhancement of neutrophil autophagy in hepatocellular carcinoma HCC is correlated with the release of matrix metalloproteinase-9 MMP9 and oncostatin M OSM , but is unrelated to the deactivation of mTOR signaling, which could contribute to the advanced migration of tumor cells.

Neutrophils have the highest expression of IgA Fc receptor FcRI CD89 of all cell types. Co-culturing of tumor cells, neutrophils, and IgA results in significant changes in the cell morphology of tumor cells, which is associated with high LC3-II expression in autophagosomes, but cell apoptosis remains constant.

These phenomena suggest that autophagy participates in the process of activated neutrophil combat against tumor cells [ ].

MDSCs are immune-suppressive cells and their accumulation and suppressive activity are driven by inflammation. It was reported that high mobility group box 1 HMGB1 can promote the survival of MDSCs by inducing autophagy [ ].

In addition, glycolytic metabolism has an essential impact on MDSCs. Glycolysis prevents the AMPK-ULK1 signaling activation and autophagy formation to enhance autophagy-mediated partial liver-enriched activator protein LAP expression, which in turn promotes granulocyte colony-stimulating factor G-CSF and granulocyte macrophage colony-stimulating factor GM-CSF expression and supports MDSCs development in tumors [ ].

Furthermore, MDSCs are identified to induce AMPK phosphorylation, stimulate autophagy and increase the anti-apoptotic factors MCL-1 and BCL-2, which promotes Multiple Myeloma MM progression [ ].

Autophagy is closely intertwined with inflammatory and immune responses, and cytokines may help mediate this interaction. Autophagy has been shown to regulate, and be regulated by, a wide range of cytokines, and autophagy activation can promote or inhibit the secretion of cytokines to control tumor development Fig.

The relationship between autophagy and cytokines. Autophagy activation can promote or inhibit the secretion of cytokines to control tumor development.

IL-2 boosts autophagy induction by promoting ATG5-beclin1-HMGB1 complex formation, however, autophay suppresses NF-κB-mediated IL-2 production. IL-6 exerts anti-autophagic effects by activating p-STAT3 and reducing the protein levels of LC3-II and Beclin 1, in addition, IL-6 promotes autophagy AMPK activation and mTORC1 inhibition, and Akt activation.

Mutually, autophagy promotes the release of IL-6 by activating NF-κB pathway. TNF represses autophagy by decreasing lysosomal acidification, and autophagy inhibits TNF-α expression through blocking p38MAPK phosphorylation and TRAF6 expression. TGF-β has been demonstrated to activate autophagy by Smads and JNK signal pathways, but autophagy decreases mature TGF-β protein levels as a result of increased degradation.

In this figure, IL and TGF-β enhance tumor development, other cytokines suppress tumor development. The two main pro-inflammatory cytokines are IL-1α and IL-1β. Inhibition of IL-1 expression in tumor cells can induce upregulation of p21 and p53, leading to suppression of tumor growth [ ].

Intracellular IL-1β is targeted by autophagosomes. Autophagy has dual effects on inflammasome activation and IL-1β secretion. However, the negative effect is predominant under stable conditions, whereas only the negative role of autophagy in IL-1α activation has been reported [ , ].

It was reported that Atg5-deficient macrophages secrete reduced amounts of IL-1β upon autophagy stimulation, restricted T cells activation, and cytokine production [ 98 ].

The production and release of mature IL-1β requires two distinct signals. The first signal is the interaction between TLR4 and TLR3 and LPS, which activates NF-κB-dependent transcription of the IL-1β gene and secreting minimal amounts of mature IL-1β.

The second signal is the potassium-proton ionophore, ATP, which induces greatly increased levels of extracellular IL-1β that is dependent on caspase-1 [ ]. Both IL-1β and IL-1α can induce autophagy; however, autophagy might limit their secretion, indicating that autophagy represents a negative feedback mechanism for controlling IL-1β and IL-1α secretion [ ].

IFN includes type I, type II, and Type III IFN. Type I IFN includes IFN-α and IFN-β, which are secreted by mononuclear phagocytes and fibroblasts, respectively.

However, the effect of IFN-α depends on the specific targeted cell type. IFN-α can inhibit autophagy when combined with lymphocyte co-culture, although it contributes to greater MHC-1 increases [ 87 ]. It can induce autophagy in various cell types, including epithelial cells, immune cells, and tumor cells.

On the other hand, IFN-γ rapidly and consistently leads to the upregulation of MHCI on the surface and induces autophagy [ 78 , ]. Conversely, IFN-γ is considered as a regulator to Th2 responses, and deletion of IFN-γ gene strongly triggers Th2 cytokines secretion in inflammatory diseases [ ].

Type III IFNs IFN-λs share some common characteristics and therapeutic benefits with type I IFNs, but the effects on cellular autophagy are different from type I IFNs.

IFN-λ1, the main type III IFNs produced by hepatocytes during acute HCV infection, can suppress HCV-induced autophagy indicated by decreased conversion of LC3β-I to LC3β-II amounts, decreased autophagosome formation, and decreased expression of autophagy-related genes ATG5 and GABARAP [ ].

In vitro, IL-6 exerts anti-autophagic effects by activating the phosphorylation of STAT3 at Tyr and reduces the protein levels of LC3-II and Beclin 1.

Treatment with a STAT3 inhibitor can reverse the inhibitory effect of IL-6 on autophagy, as activated STAT3 binds to the promoter of Bcl-2 and leads to its overexpression, which in turn reacts with Beclin1 to inhibit the formation of the Beclin1-VPSAtgp complex to decrease autophagy [ ].

For instance, IL-6 inhibits the formation of IFN-γ and starvation-induced autophagosomes in virulent M. tuberculosis H37Rv-infected macrophages by indicating the decreased LC-II and Beclin1.

In vivo, IL-6 trans-signaling promotes autophagy by stimulating a robust increase in lysosomes but not autophagosomes. The process is dependent on IL-6R expression.

Autophagy is accelerated when IL-6 is complexed with soluble IL-6R and thereafter locates to gp on cellular membranes. The stimulation of autophagy by IL-6 is regulated via multiple complementary mechanisms, including two main signals: one is activating AMPK, which inhibits mTORC1; the other is activating Akt, then phosphorylating STAT3 at S and activating Atg4c and mTORC2, ultimately leading to autophagy-related enzyme production to induce autophagy [ ].

In lung cancer patients, cachexia is prevalent. There is a positive correlation between IL-6 trans-signaling-induced autophagy in the tumor and weight loss [ ]. Mutually, autophagy promotes the release of IL-6 by HBV X protein HBx -induced autophagy and activates the NF-κB pathway.

Autophagy inhibition abrogates NF-κB activation and IL-6 production [ ]. The binding between ATG5, HMGB1 and Beclin1 is essential for ILinduced autophagy. Autophagy inhibitors or knockdown of ATG5 and Beclin1 can block ILinduced autophagy and switch ILinduced proliferation to apoptosis [ ].

High-dose IL-2 HDIL-2 alone increases serum levels of IFN-γ, IL-6, and IL and translocates HMGB1 from the nucleus to the cytosol in hepatocytes. Then, the interaction between HMGB1 and Beclin1 boosts autophagy.

However, the effects could be inhibited by combining with autophagy inhibitor Chloroquine CQ , which inhibits autophagy by blocking acidification of the lysosome, preventing fusion with the autophagosome.

In tumor cells, CQ increases autophagic vacuoles and LC3-II levels, inhibits oxidative phosphorylation and ATP production, and promotes apoptosis, suggesting that the combination of IL-2 with CQ promotes anti-tumor effects, increases long-term survival, decreases toxicity associated with vascular leakage, and enhances immune cell proliferation and infiltration in the liver and spleen [ , ].

IL is important for immune responses and anti-tumor activity. However, the induction of autophagy attenuated the growth-inhibitory effect of IL on hepatoma cells, indicating that restraining autophagy by inhibitors or silencing Beclin1 could enhance ILmediated anti-tumor effects.

IL is produced by activated inflammatory cells, therefore, autophagy decreases the release of IL by inhibiting inflammation. The PI3K pathway promotes phosphorylation of p70S6K through the activation of Akt and mTORC1 [ ]. Furthermore, IL-4 and IL signaling also activate PI3K signaling to activate mTORC1 in macrophage cells, and Th2 cytokines IL-4, IL and IL exert autophagy inhibition in most environments [ ].

During monocytes—DCs differentiation and DCs survival, cytoprotective autophagy responses are essential for counteracting ILtriggered apoptosis, but IL can strongly inhibit starvation-induced autophagy and decrease Bcl-2 levels, which indicates increased levels of Beclin-1, LC3, and mature autophagosomes and results in restricting DCs growth.

IL not only kills nascent antigen-presenting DCs but also specifically skews the growth of DCs toward non-antigen-presenting monocytes—macrophages, and the autophagy inhibitor 3-methyladenine 3-MA restricted DCs differentiation by prompting apoptosis [ ]. Autophagy is likely to have dual effects on IL production, but the mechanisms require further exploration [ , ].

TNF-α inhibited autophagy via disrupting the autophagic flux by decreasing lysosomal acidification, but it was reported that the increased amount of LC3-II protein level, associated with the increased of P62 protein level, without altering P62 mRNA levels.

Inhibition of autophagy and promotion of lysosomal proteolysis accelerate TNF-α-induced death by increasing oxidative stress and toxicity because LPS-induced TNF-α mRNA expression is further significantly enhanced by pretreatment with Bafilomycin A1, implying autophagy plays an inhibitory role in the expression of TNF-α.

Autophagy causes the inhibition of p38MAPK phosphorylation and TRAF6 expression, which are required for the expression of TNF-α [ , ].

In immune evasion, TGF-β exerts suppressive effects directly on effector cells including cytotoxic cells and indirectly promotes the differentiation of regulatory T cells. In the tumor immunosuppressive microenvironment, TGF-β can inhibit NK cells to diminish targeted cell lysis and IFN-γ production.

However, NK cells in the tumor microenvironment may restore their activity by TGF-β blockade with anti-TGF-β antibodies and small molecule inhibitors of TGF-β signaling [ ]. Furthermore, inhibition of autophagy increases mature TGF-β protein levels without inducing TGF-β mRNA expression, indicating that the increase of mature TGF-β protein levels is a result of decreased degradation rather than increased synthesis [ ].

Blocking TGF-β in the co-culture diminishes targeted cells autophagy in a dose-dependent manner, indicating that TGF-β might be responsible for autophagy induction [ 87 , ]. Recently, TGF-β has been demonstrated to activate autophagy in certain HCC and breast cancer cells, which undergo cell cycle arrest and apoptosis in response to TGF-β.

In those malignant cells, TGF-β stimulates the expression of mRNA transcripts of several autophagy-related genes, such as Beclin1, Atg5, Atg7, and death-associated protein kinase Dapk , and induces accumulation of autophagosomes and activation of autophagic flux.

Up regulation of these genes is regulated by the Smad and non-Smad signal transduction pathways, including ERK, JNK, p38MAPK, and PI3K.

Meanwhile, autophagy potentiates the induction of the proapoptotic Bcl-2 family protein Bim and contributes to Bim-mediated apoptosis in hematopoietic cells. TGF-β could also induce directly proapoptotic genes, Bim and Bmf, in a p38 MAPK and Smads-dependent manner, indicating a functional link between autophagy and apoptosis [ , ].

Inhibitor of autophagy, such as 3-MA as a PI3K inhibitor, can block autophagic degradation of proteins and enhance LPS-induced secretion of IL Moreover, knockdown of either Beclin1 or Atg7 enhances secretion of IL at the transcriptional level.

In autophagy-deficient cells, IL secretion is directly regulated by IL-1 signaling and is dependent on the generation of ROS because ROS promoted activation of the inflammasomes, and the secretion of IL-1 and IL-1R1 is known to activate the NF-κB pathway, then stimulate the production of IL as well as TNF-α [ ].

Immunotherapeutic strategies aimed at boosting antitumor immunity are promising candidates for the treatment of tumors. However, the clinical outcomes of these immunotherapeutic strategies have been less effective than anticipated. Immune tolerance to these tumors is still a major impediment in cancer immunotherapy.

As immunologic tolerance molecules, Indoleamine 2,3 dioxygenase IDO , CTLA-4 and PD-1 can regulate tumor immune tolerance through autophagy pathways.

Therefore, understanding the relationship between autophagy and tumor immune tolerance is important for developing tumor immunotherapy strategies.

IDO is produced by tumor cells, tumor-associated MDSCs and TAMs. Autophagy can inhibit the inflammation-mediated expression IDO production by suppressing inflammation [ , , ].

General control nonderepressible 2 GCN2 can be triggered by IDO-mediated tryptophan Trp deficiency, which is recognized as an important effector of the IDO pathway, resulting in auto-phosphorylation and activation of kinase activity that inhibits the translation initiation factor 2α eIF2α , blocking protein synthesis and arresting cell growth.

GCN2 is essential for inflammatory carcinogenesis [ ]. When autophagy is induced by IDO or GCN2, it protect organisms from fatal inflammation disease; therefore, IDO1-GCN2-autophagy signals may be a common circuit induced in human inflammatory disease, which could be potentially targeted for therapeutic benefit [ ].

Furthermore, IDO inhibits a tryptophan sufficiency signal, resulting in the inhibition of mTOR, leading to autophagy via LC3 production, and translational blockade via s6K inactivation. Tryptophan and the experimental agent 1-methyl-D-tryptophan D-1MT, as a mimetic of Trp functionally reverse the effects of IDO on mTOR and autophagy in the sufficiency pathway, but do not affect GCN2 [ , ].

PD-1 acts as a T-cell inhibitory checkpoint molecule and suppresses anti-tumor immunity by developing a T-cell tolerance, inhibiting T cell proliferation, and hindering the recognition of tumor cells via interaction with PD-L1 on the surface of tumor cells. Sigma1 inhibitor has been identified to induce degradation of PD-L1 and suppress the functional interaction of PD-1 and PD-L1 in a co-culture of T-cells and tumor cells via autophagy.

The numbers of T cells and B cells are reduced due to the increased numbers of Tregs and mesenchymal stem cells, and antigen-specific T cells may be defective.

Moreover, because of their inability to proliferate, the capacity of these cells to consume cytokines is reduced, which results in enhanced Th1, Th2, and Th17 cytokines. The relationship between autophagy and tumor immune tolerance.

IDO is thought to potently inhibit effective anti-tumor immunity, drive immunologic tolerance and promote the development of tumor by suppressing cytotoxic T cell responses and inflammatory dendritic cell maturation, magnifying tolerogenic APCs and Tregs generation.

IDO triggers autophagy by inhibiting a tryptophan sufficiency signal, resulting in the inhibition of mTOR, and autophagy can inhibit the inflammation-mediated expression IDO production by suppressing inflammation.

CTLA-4 is an effective therapeutic target in tumor patients. Autophagy induction can improve anti-CTLA-4 curative effects, autophagy activation can restore the expression of CTLA-4 as well as suppressor function, CTLA4 engagement inhibits autophagy by constraining the transcription of LC-3β and the formation of autophagosomes.

PD-1 acts as a T-cell inhibitory checkpoint molecule and suppresses anti-tumor immunity by developing a T-cell tolerance, inhibiting T cells proliferation, and hindering the recognition of tumor cells via interaction with PD-L1 on the surface of tumor cells, and tumor cell-intrinsic PD-L1 can suppress autophagy by activating mTORC1 signaling and inhibiting mTORC2 signaling.

As an immune tolerance checkpoint, CTLA-4 is an effective therapeutic target in tumor patients. It was confirmed that in human melanomas, the expression of the key autophagosome component LC3-β and other autophagy activators are available to suppress primary resistance to CTLA-4 blockade through decreasing MAGE-A protein levels and blocking the MAGE-TRIM28 complex, which indicates that autophagy induction can improve anti-CTLA-4 curative effects [ ].

In conclusion, IDO is thought to potently inhibit effective anti-tumor immunity, drive immunologic tolerance and promote the development of tumor. Therefore, autophagy is regulated by a series of tumor immune tolerance molecules and it also plays a key role in regulating tumor immune tolerance Fig.

The immune system plays a dominant role in tumor treatment by identifying and killing tumor cells during different stages of tumor development. Accumulating studies show that autophagy could up-regulate and down-regulate the immune response by influencing cells and the release of cytokines, which has provided targets and enlightenment for tumor immunotherapy [ 44 ].

As a new generation of anti-tumor therapeutics, tumor immunotherapy plays a predominant role in suppressing tumor development and will continue to progress. So far, antibody targeting therapy synergy with autophagy has been frequently reported. For instance, in ovarian cancer models, MORAB farletuzumab , a humanized monoclonal antibody against folate receptor alpha FRα , has displayed a notable anti-tumor effect through antibody-dependent cellular cytotoxicity by sustaining late-stage autophagy, and when protein and organelle turnover overwhelm the capacity of the cell, which contributes to type II programmed cell death or autophagic death [ , ].

In addition, CD73 enzymes play a pivotal role in generating an immunosuppressed and pro-angiogenic niche to support tumor development.

Pharmacological blockade of CD73 with MEDI an Anti-CD73 Ab increases antigen presentation and autophagy, resulting in enhanced lymphocyte activation and a greater release of proinflammatory Th1 cytokines [ , ].

Recently, therapies aiming at autophagy to enhance the immune responses and anti-tumor effects of immunotherapy have become the prospective strategies, with enhanced angtigen presentation and higher sensitivity to CTLs [ 65 ]. Radiotherapy and chemotherapy might provoke autophagy; therefore, combining immunotherapy with radiotherapy or chemotherapy produces better treatment effects.

Radiation or chemotherapy-induced autophagy has been reported to redistribute mannosephopsphate receptor MPR with its ligands to the autophagosomes via clathrin-coated vesicles.

Low pH in autophagosomes leads to the release of the MPR cargo. Empty MPRs are transported back to the tumor cell surface. Receptors bind to granzyme B GrzB, one such ligand of MPR produced by activated CTLs, which renders tumor cells more susceptible to CTLs killing and potentiates the effect of immunotherapy [ , ].

In addition, autophagy plays a role in antigen processing for MHCI and MHCII presentation. Moreover, SYK augments OxLDL-induced autophagy and MHCII expression in macrophages.

In recent studies, DCs-based vaccines have shown promising therapeutic effects in promoting tumor immunotherapy by boosting antigen presentation. For example, lactosylated N-Alkyl polyethylenimine coated superparamagnetic iron oxide SPIO nanoparticles-induced autophagy can enhance the vaccine functions of DCs by inducing DCs maturation [ ].

Furthermore, Shikonin-induced autophagy can directly contribute to damage-associated molecular patterns DAMPs upregulation and DCs activation, DCs vaccine preparations need the pretreatment of CQ, which will enhance the anti-metastatic effect of shikonin [ ].

Additionally, autophagy can also improve the efficacy of DNA vaccines by synthetizing intracellular vaccine-encoded tumor antigen [ ]. Researchers have observed anti-tumor and anti-metastatic activity of pencoding DNA vaccines, which is a promising strategy for tumor immunotherapy [ ]. However, it has been reported that hypoxia-induced autophagy has attenuated the effects of immunotherapy by impairing CTLs-mediated tumor cell lysis associated with the hypoxia-dependent phosphorylation of STAT3 pSTAT3.

The first mechanism is that the HIF-1α-dependent intrinsic signaling pathway is activated, which phosphorylates Src kinase in the Tyr residue pSrc , leading to STAT3 phosphorylation of the Tyr residue.

When autophagy is blocked, p62 is accumulated and in turn accelerates the delivery of pSTAT3 to the UPS for selective degradation [ , ]. Hypoxia-induced autophagy also degrades NK-derived GrzB and impairs NK-mediated killing, as accumulated HIF-2α transfers to the nucleus and induces the expression of the autophagy sensor ITPR1, leading to the impairment of NK-mediated killing and decreased immunotherapy effects [ , ].

Many studies indicate that autophagy inhibition in tumors can be viewed as an approach to improve anti-tumor immunotherapies. HDIL-2 alone has been found to be an efficient immunotherapy method in an advanced murine metastatic liver tumor model. IL-2 inhibits tumor growth by enhancing immune cell proliferation and infiltration in the liver and spleen; however, the anti-tumor effects of HDIL-2 immunotherapy were significantly heightened when coupled with administration of autophagy inhibitor CQ [ ].

Similarly, in renal cell carcinoma, CQ is also used to improve HDILmediated anti-tumor immunity by enhancing DCs, T-cells and NK cells and limiting ATP production through inhibition of oxidative phosphorylation and promotion of apoptosis [ ].

Another selective PI3K inhibitor, 3-MA acts on Vps34 and PI3Kγ and significantly enhances ILinduced apoptosis in oral squamous cellcarcinomas OSCC , which demonstrates the combination of autophagy inhibitors and IL is a promising approach for tumor immunotherapy [ ].

Autophagy is required for the maintenance of metabolic and genetic homeostasis in eukaryotic organisms, which is involved with various ATG protein complexes regulated by several signaling pathways.

Autophagy plays a dual role in tumor cell growth, which is dependent on the properties of the tumor and cell types. Therefore, when and how autophagy can be pro-survival and pro-death should be carefully interpreted in the future. In the tumor microenvironment, autophagy is an important regulator of immune responses by sustaining homeostasis, activation, and biological functions of immune cells.

However, the autophagy-mediated regulation of the immune system might strengthen or attenuate the effects of immunotherapy Fig. Therefore, whether we should try to enhance or inhibit autophagy in anti-tumor immunotherapy remains to be explored. Many studies have demonstrated that the optimal combination of autophagy-based inducer or inhibitor with various therapeutic strategies, including chemotherapy, radiotherapy, immunotherapy, and gene therapy may be a more efficient approach by eliciting tumor cell death.

Furthermore, many findings confirmed that both autophagy activators and inhibitors are being preclinical studies and potential to cure various tumors in the future Tables 1 and 2. Nevertheless, only a few autophagy inhibitive agents are being applied to strengthen the anti-tumor effects of immunotherapy in preclinical studies Table 3.

Even though, studies about the application of autophagy inhibitors or enhancers alone in clinical treatment have not yet been published, importantly, several clinical trials have been shown that autophagy inhibitors hydroxychloroquine HCQ and CQ, autophagy activators aspirin ASA combined with other antineoplastic drugs significantly improve the therapeutic effect in tumors Table 4.

In the future, efforts should be focused on how to regulate autophagy to strengthen innate and adaptive immune responses and overcome anti-tumor immune resistance in immunotherapy for tumors.

The applications of autophagy for tumor immunotherapy. There is a complicated interaction between autophagy and immune system. Autophagy can enhance immune response by ensuring the inhibitory action of CTL, B cell, Mø, NKT and DC on tumor cells and the release of immunoreactive cytokines, like IL-1, IL-2, IL-6, IL, IL, TNF-α and IFN-γ, resulting in enhanced anti-tumor immunotherapy effects and repressed tumor development.

In addition, autophagy can also reduce immune response by recruiting immunosuppressive Tregs and promoting IL and TGF-β production, contributing to attenuated anti-tumor immunotherapy effects and accelerated tumor development.

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We tested whether TLR5 host receptors are necessary to elicit the anti-tumor response observed in mice treated with CBLB in combination with ICT treatments Fig.

Kaplan—Meier survival curve from two independent experiments are shown in Fig. All mice under vehicle control died by the end of the study Supplementary Fig. All ICT- or CBLBtreated mice in both cohort groups were dead by week 6 Supplementary Fig. Consistent with previous results Fig.

These results indicate that TLR5 host receptors are necessary for anti-tumor responses observed in mice treated with CBLB and ICT treatments.

Mice that showed complete tumor regression were re-challenged by orthotopic injections of 4T1 FUGW-FL cells into the opposite left fourth mammary fat pad without any additional therapy Supplementary Table 5. Figure 3a shows overall survival from the re-challenge experiment.

One re-challenged mouse was excluded from the survival curve shown in Fig. Furthermore, one-week post re-challenge, this mouse showed comparable bioluminescent signal to other re-challenge mice Supplementary Fig. However, no bioluminescent signals or palpable tumor were detected thereafter during the first three weeks of the experiment, indicating that the mouse likely rejected 4T1 FUGW-FL tumor implantation.

Nonetheless, overall, these results indicated that the observed curative effects were likely due to acquired memory for anti-tumor immunity. All mice were injected at week 0 with 4T1 FUGW-FL tumor cells and tumors were allowed to grow without therapeutic intervention.

All tumor-naïve, tumor challenged-mice died by week 6. Only three tumor survivor mice died due to tumor burden: Supplementary Fig. and, Supplementary Table 5. All other tumor survivor mice were tumor-free for at least 60 weeks post orthotopic tumor re-challenge.

To expand on our understanding of the mechanisms of response and characterize changes elicited by ICT and CBLB therapies alone or in combination, we assayed 32 peripheral blood-borne cytokines from aged-matched tumor-free mice healthy mice, no tumor implantation and 4T1 FUGW-FL tumor-bearing mice under our treatment cohorts: tumor-bearing, vehicle control; tumor-bearing, treatment failure; and tumor-bearing, treatment responders Supplementary Fig.

Of note, tumor-free healthy mice showed variable expression of cytokines between weeks 3 and 10 of the study, likely reflecting physiological changes in cytokine expression Fig.

a Heatmap of 32 peripheral blood cytokines of tumor-free mice and mice challenged with 4T1 FUGW-FL tumors three weeks post orthotopic tumor implantation: tumor-free mice healthy mice, no tumor implanted ; tumor-bearing mice, vehicle PBS control non-survivors , tumor-bearing mice, failed treatment non-survivors , and tumor-bearing mice, long-term survivors.

b Volcano plot highlighting detectable statistical difference between peripheral blood cytokines three weeks post orthotopic tumor challenge. c Heatmap of 32 peripheral blood cytokines of mice challenged with 4T1 FUGW-FL tumors 5 to 7 weeks post tumor implantation: tumor-bearing mice, vehicle PBS control non-survivors at week 5; tumor-bearing mice, failed treatment non-survivors , weeks 5 to 7; and tumor-bearing mice, long-term survivors during weeks 6 and 7.

d Volcano plot highlighting detectable statistical difference between peripheral blood cytokines taken between 5 and 7 weeks post orthotopic tumor challenge. e Heatmap of 32 peripheral blood cytokines of mice re-challenged with 4T1 FUGW-FL tumor: tumor naïve mice, tumor-bearing non-survivors , tumor survivor mice, re-challenge failure non-survivors and tumor survivor mice, re-challenge survivor long-term survivors taken three weeks post orthotopic 4T1 FUGW-FL tumor implantation.

f Volcano plot highlighting detectable statistical difference between peripheral blood-bourne cytokines taken three weeks post orthotopic tumor re-challenge. Four profiles emerged from our analysis.

First, it is noteworthy that levels of granulocyte-colony stimulating factor G-CSF , a cytokine involved in the proliferation and differentiation of granulocytes and neutrophils 58 and associated with poorer survival in cervical, non-small cell lung cancer, colon, melanoma, and skin cancers 59 , 60 , 61 , 62 , 63 , showed a statistical increase in tumor-bearing mice, vehicle control compared with tumor-free mice three weeks post 4T1- FUGW-FL tumor implantation Fig.

Although there was no statistical difference between tumor-bearing mice that failed treatment non-survivors , and tumor-bearing mice, treatment responders long-term survivors three weeks post 4T1 FUGW-FL tumor implantation Fig.

Consistent with these results, tumor-bearing mice, treatment responders also showed a statistical decrease during weeks 5 through 7 when compared with tumor-bearing mice, vehicle control Fig. Second, CXCL5, a chemokine involved in recruitment of myeloid derived suppressor cells MDSCs to the tumor microenvironment 64 , 65 , 66 , 67 and associated with poor survival in renal, liver, pancreatic, and cervical cancer 68 , showed a statistical decrease in tumor-bearing mice that responded to treatment long-term survivors compared with tumor-bearing mice that failed treatment non-survivors three weeks post 4T1 FUGW-FL tumor implantation Fig.

By contrast, there was no statistical difference between tumor-bearing mice, vehicle control, compared with tumor-bearing mice that failed treatment non-survivors Fig.

Fourth, although not statistically significance based on two-way ANOVA tests, tumor-bearing mice that responded to treatment showed an overall increase trend for immune-activating cytokines compared to tumor-bearing mice that failed treatment during weeks 5 through 7 post tumor implantation: GM-CSF fold 76 , IL-2 fold 77 , IL 8-fold 78 , IFN-γ 7-fold 79 , and CCL3 four-fold Of note, CXCL1, a chemokine associated with tumor immune suppression 81 , showed a 4-fold increase over tumor-bearing mice that failed treatment Supplementary Table 7.

IL, a cytokine involved in immune suppression through inhibition of antigen-presenting cells APC and the activation of immune suppressive T-reg cells 82 , showed a two-fold decreased in mice that responded to treatment compared with mice that failed treatment Supplementary Table 7.

Interestingly, IL antagonists are currently being explore as anti-tumor immune therapy in combination with TLR agonists and other immunostimulatory treatments Finally, tumor-bearing mice that responded to treatment long-term survivors showed statistical increases in IL-1α, IL, CXCL5, and IL p40 when compared to tumor-free control mice, indicating a distinctive immune activating systemic cytokine profile ten weeks post 4T1 FUGW-FL tumor implantation Supplementary Fig.

Tumor re-challenged and tumor-naïve mice cytokines were assayed three weeks post 4T1 FUGW-FL implantation. Mice that were tumor-free for at least 60 weeks post re-challenge tumor survivor mice, re-challenge survivor revealed a distinctive cytokine profile from those mice that were re-challenged, but developed tumors tumor survivor mice, re-challenge failure Fig.

Interestingly, similar to previous results with challenge mice Fig. Interestingly, re-challenge mice that did not survive tumor survivor mice, re-challenge failure also showed a statistical increase when compared to tumor naïve mice, tumor bearing Fig.

Moreover, levels of G-CSF also trended lower in long-term survivors tumor survivor mice, re-challenge survivor fold decreased when compared with tumor-naïve, tumor-bearing mice non-survivors Supplementary Table 8 , but these values fell shy of statistical significance two-way ANOVA.

Mice that were re-challenged, but did not survive tumor survivor mice, re-challenge failure , showed similar levels of G-CSF to tumor-naïve, tumor-bearing mice at this time point Supplementary Table 8. In addition, four profiles emerged from our analysis. First, cytokines that were upregulated in both re-challenge failure mice and re-challenge survivor mice, but with a far greater increase in re-challenge survivors: LIF 5-fold compared with fold , CXCL1 3-fold compared with fold , IL-2 7-fold compared with fold , IL-7 fold compared with fold , IL p70 fold compared with fold , CCL4 3-fold compared with fold , CCL11 3-fold compared with fold , CCL3 3-fold compared with fold , IL-1α 4-fold compared with fold , IL fold compared with fold , IL-1β 3-fold compared with 6-fold , CXCL2 8-fold compared with fold , IL-4 fold compared with fold , IL-9 4-fold compared with 7-fold , and M-CSF fold compared with fold Supplementary Table 8.

Second, cytokines that were downregulated in both groups, but to a greater extent in the re-challenge failure cohort: IL-3 0. Third, cytokines that showed differential regulation between the two cohort: IL p40 0.

Fourth, cytokines that did not change in one population, but did in another: re-challenge survivor mice upregulated IL fold , CXCL9 fold , IFN-γ 9-fold , CCL5 three-fold , whereas re-challenge failure mice did not upregulate these cytokines greater than two-fold.

CXCL5 showed a 0. IL showed a two-fold increase in mice that failed to survive, but no change in long-term survivors Supplementary Table 8. Taken together, these results showed a distinctive adaptive immune-activating cytokine profile in those mice that survived the re-challenge experiment.

To further elucidate the effects of CBLB and ICT therapies alone or in combination, we explored changes to the tumor immune microenvironment in response to these therapies and inquired if changes to immune cells infiltrate correlated with increased likelihood of survival.

We used a unique strategy based on bioluminescence signal as a prognostic tool for survival outcomes of mice that were euthanized at three weeks post 4T1 FUGW-FL tumor implantation for tumor extraction and subsequent tumor immune infiltrate flow cytometry profiling, in effect at a time before outcomes for individual mice would be known, but bioluminescence signal would be predictive.

Tumor volume and BLI measurements have been useful tools to appraise tumor progression in pre-clinical models 84 , 85 , 86 , Furthermore, BLI measurements have been used to evaluate treatment efficacy in various animal models 88 , Hence, we leveraged overall survival data and tumor progression measurements Fig.

There was also little correlation between Log BLI and tumor burden, suggesting additional information could be provided by each measurement. The predictive diagnostic accuracy of BLI measurements for survival outcomes indicated that mice with higher BLI measurements had decreased likelihood of survival and that mice with lower BLI measurements had increased likelihood of survival.

ROC curves comparing sensitivity and specificity for overall survival by a bioluminescence total photon flux at week 3, b tumor volume at week 3, and c tumor-volume slope between weeks 2 and 3. d Treatment diagram for euthanized mice used for tumor immune profiling. We then proceeded to examine changes to the tumor immune microenvironment in response to treatments at three weeks.

Myeloid and lymphoid cells were identified as described in Supplementary Table However, tumor samples taken from treated mice showed a trend toward increased variability in the number of myeloid cells Supplementary Fig.

This trend could reflect ongoing immune responses in treated mice. Indeed, overall changes to the tumor immune landscape in treated mice are heterogeneous and may point to immune activation. MDSCs are known to suppress T cell activation and proliferation, impair natural killer NK cells functions, induce immune suppressive T reg cells, and promote pro-tumor inflammatory states by regulating cross talk between tumor cells, mast cells and macrophages Overall, these results point to a switch in the state of the tumor immune microenvironment from immune suppression to immune activation in mice with lower BLI signals, which elicits a subsequent curative anti-tumor response.

a Myeloid top panel and lymphoid bottom panel tumor immune infiltrate profile of vehicle and treated mice, assigned a Log BLI measurement taken 3 weeks post 4T1 FUGW-FL tumor implantation and prior to tumor extraction. b Volcano plot highlighting the correlation between immune infiltrate and Log BLI.

Vertical line demarks negative and positive Spearman r correlation values. Finally, we further pursued to corroborate the diagnostic potential of Log BLI signal by comparing the blood-borne cytokine profile of long-term survivor mice Fig.

Of note, CXCL5, previously identified as detrimental for survival Fig. However, it is possible that sample size limited the ability to detect statistical differences. Indeed, because bacterial flagellin is the native ligand for TLR5, overall, our studies provide further mechanistic insight into linkages between ICT response and microbiota 98 , 99 , , Taken together, these results pointed to a new therapeutic strategy that harnessed both innate and adaptive components of the immune system to elicit a lasting antitumor response.

On the one hand, bacterial derived-flagellin has been shown to elicit a targeted antitumor response by binding to TLR5, initiating a cascade of signals that produce a pro-inflammatory response via activation of the transcription factor NF-kB Herein, we first showed that in vitro CBLB, a potent activator of the NF-kB signaling pathways, was sufficient to elicit a TLR5-mediated immunogenic cytokine response in tumor cells.

Given that deficiencies in antigen presentation underlie many mechanisms of resistance against ICT, it is possible to hypothesize that a potent activator of innate immunity may modulate tumor homeostasis, shifting the tumor microenvironment state from immune suppression to immune activation Fig.

Several lines of evidence support this model. First, only treatment with either flagella or CBLB in combination with ICT increased survival in mice bearing highly ICT-refractory 4T1 and BF10 tumors, whereas monotherapies of flagellin, CBLB, or ICT did not show significant curative effects.

Second, the peripheral blood cytokine profiles of 4T1 tumor-bearing mice that responded to treatment and showed complete tumor regression reflected a concerted antitumor response.

While it has been shown that mice bearing tumor cells lacking TLR5 fail to respond to treatment with flagellin 26 , it remains to be studied whether flagella and CBLB can also act on the tumor cells to elicit a curative immune response in the context of combination treatment with ICT.

Furthermore, it remains to be elucidated whether host TLR5 receptors mostly act on components of innate immunity axis or could they also directly act on components of adaptive immunity to elicit durable immune responses in the context of ICT Fourth, nearly all survivor mice that were re-challenged rejected the same tumor, implying an adaptive memory response against tumor cells.

Fifth, the peripheral cytokine profiles of those mice that were re-challenged and rejected the tumor aligned with a strong adaptive immune-activating response. Finally, changes in the tumor immune infiltrate amongst mice with increased likelihood of survival were consistent with immune activity leading to a curative anti-tumor response.

Potent activation of innate immunity through TLR5 agonists in combination with ICT treatments targeting PD-1 and CTLA-4 leads to systemic increases in immune activating cytokine IL, and reduction in immune suppressive cytokines G-CSF and CXCL5.

The remarkable increase of blood-borne G-CSF protein levels in tumor-bearing mice that either failed treatment or served as tumor-bearing untreated controls suggested that G-CSF may be explored as a potential biomarker.

Conversely, low serum level of G-CSF in tumor-bearing mice that responded to treatment or developed long-term tumor immunity suggested utility as a predictive marker for treatment response.

These results further raise caution for the use of G-CSF to prevent neutropenia in cancer patients. Although a recent meta-analysis study showed some benefit of supportive G-CSF therapy in overall survival of patients receiving chemotherapy, data also show an increased risk of developing secondary malignancies G-CSF therapy in the context of ICT remains to be explored.

In conclusion, the success of immune checkpoint therapy in eliciting long lasting curative responses against various types of cancers in subsets of patients make worthwhile efforts to expand the number of patients that respond to this type of treatment. Activators of TLR5, such as flagellin and CBLB, in combination with ICT may provide new therapeutic opportunities for previously unresponsive patients.

Salmonella typhimurium flagellin FLA-ST was purchased from Invivogen. CBLB was a gift from Cleveland Biolabs, Inc.

Monoclonal antibodies 9D9 anti-CTLA-4 and RPM anti-PD-1 were purchased from BioX Cell and maintained in 6. After two weeks, isolated cell colonies were imaged to confirm reporter gene expression, and bioluminescent colonies were harvested and expanded.

Reporter cells were continuously cultured in the presence of 0. BF10 parental cells were cultured according to ATCC protocols Data were imported into Excel Microsoft Corp. The normalized results from repeated experiments were averaged for each time point, and the results graphed as normalized photon flux versus time, with the y -axis on a log2 scale.

Positive error bars represent standard error of the mean for repeated experiments. All animal procedures were approved by the Institutional Animal Care and Use Committee IACUC of the University of Texas M.

Anderson Cancer Center; protocol RN Anderson Cancer Center. Animals were allowed at least one week to acclimate to the animal facility before start of experiments. Mice that reached end point moribund condition or having one tumor measurement in the sagittal or axial plane greater than 1.

Anderson Cancer Center IACUC euthanasia protocols. Flagellin, CBLB, 9D9 anti-CTLA-4 , and RPM anti-PD-1 were suspended in filtered PBS; filtered PBS was used as a vehicle control.

Flagellin or CBLB was administered every two days for two weeks. All mice sorted into different treatment groups showed detectable levels of bioluminescence signal during week 1 and week 2 post 4T1 FUGW-FL tumor cell implantation, confirming the presence of tumors prior to commencement of treatment Supplementary Figs.

ICT was administered on days 1, 3, 5, and 8 of treatment. The mice were imaged using the PerkinElmer IVIS Spectrum Imaging System weekly beginning one week after orthotopic injection of 4T1 FUGW-FL cells into the mammary fat pad.

Mice were imaged ten minutes after injection with d -luciferin Tail samples were processed using DirectPCR Lysis Reagent Tail Viagen Biotech Inc , followed by PCR Supplementary Fig.

The genomic region containing the Tlr5 gene was amplified using the following primers and PCR protocol:. Three days post subcutaneous injection of BF10 cells into the posterior right flank, each mouse was randomly sorted into a group receiving treatment with vehicle control, CBLB only, ICT only 9D9 plus RPM , or CBLB combined with ICT.

CBLB, 9D9, and RPM were suspended in filtered PBS, and filtered PBS was used as a vehicle control. CBLB was administered every two days for two weeks. The supernatant was assayed using Mouse Cytokine Antibody Array C series RayBiotech.

Sera from mice in experiments 6, 7, and 8 Supplementary Table 2 were obtained by submandibular sampling. Sera from mice in experiment 9 Supplementary Table 2 were obtained by saphenous sampling.

Serum samples were analyzed at the Antibody-Based Proteomics Core at Baylor College of Medicine, Houston, TX. The core used the Milliplex Mouse Plex Cytokine Panel Millipore , which included the following cytokines: G-CSF, GM-CSF, CCL11, IFN-γ, IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-9, IL, IL p40 , IL p70 , IL, IL, IL, CXCL10, CXCL1-like, LIF, CXCL5, CCL2, M-CSF, CXCL9, CCL3, CCL4, CXCL2, CCL5, TNF-α, VEGF and appropriate controls and calibration standards.

For cytokines with above-range values, we reported the highest observed value for the corresponding cytokine, whereas for below-range values, we reported the lowest value on the standard curve divided by half , Tumors and spleens were removed with scissors or forceps and weighted.

Nylon mesh was rinsed several times with media. Fluorescence minus one FMO controls were used where indicated to distinguish between positively and negatively stained cells for FoxP3, Ly6G, and Ly6C. Flow cytometry was performed using an LSRII cytometer Becton Dickinson.

Subsequent analysis was performed utilizing FlowJo Statistical analyses were performed using GraphPad Prism version 8. Overall survival was assessed using Kaplan—Meier curves Log-rank Mantel—Cox test and Gehan—Breslow—Wilcoxon tests were used to compare survival rates between treatments.

ROC curves were calculated for predictive analysis. Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article.

All data generated or analyzed during this study are included in this published article and its supplementary information files, including Supplementary Data 1. Ribas, A. Cancer immunotherapy using checkpoint blockade.

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Other research discovered that the best way to avoid adverse immune system changes and help the body to recover after intense exercise was to consume carbohydrates during or after. The authors of the paper suggest that between 30 and 60 grams of carbs every hour during physical activity can help maintain normal immune function.

In addition to a balanced diet and regular exercise, scientists have found evidence of other factors that may affect the response of the immune system. Getting outside in the sunlight may benefit the immune system. Researchers discovered that sunlight energizes infection-fighting T cells that play a key part in immunity.

A study uncovered that anticipating a happy or funny event increased levels of endorphins and other hormones that induce a state of relaxation. Chronic stress can suppress the response of the immune system and its ability to fight disease; therefore, reducing stress may help to prevent infections and other disorders.

Singing in a choir for 1 hour was reported to reduce stress, improve mood, and increase the levels of immune proteins in individuals with cancer and their caregivers.

The study findings demonstrate that something as simple as singing can help reduce the stress-related suppression of the immune system. Research indicated that individuals who were lonely produced higher levels of proteins related to inflammation in response to stress than those who felt they were socially connected.

Although many questions remain about the function of the immune system, it is clear that consuming a healthy diet, regularly exercising, getting adequate sleep, and reducing stress will go a long way to ensuring your immunity is maintained.

People with a weak immune system are more likely to get infections and to have severe symptoms. Get some tips on how to stay healthy here. The immune system defends the body from invaders such as viruses, bacteria, and foreign bodies.

Find out how it works, what can go wrong, and how to…. A strong immune system helps a person stay healthy by fighting off bacteria and viruses. In this article, we look at foods that can help to boost the…. In this article, we describe types of foods that may weaken the immune system and others that may help support it.

Learn more here. Many people take supplements to strengthen their immune systems. But what is the evidence for this, and what are the limits? My podcast changed me Can 'biological race' explain disparities in health? Why Parkinson's research is zooming in on the gut Tools General Health Drugs A-Z Health Hubs Health Tools Find a Doctor BMI Calculators and Charts Blood Pressure Chart: Ranges and Guide Breast Cancer: Self-Examination Guide Sleep Calculator Quizzes RA Myths vs Facts Type 2 Diabetes: Managing Blood Sugar Ankylosing Spondylitis Pain: Fact or Fiction Connect About Medical News Today Who We Are Our Editorial Process Content Integrity Conscious Language Newsletters Sign Up Follow Us.

Medical News Today. Health Conditions Health Products Discover Tools Connect. Human Biology. Nervous system Cardiovascular system Respiratory system Digestive system Immune system. Tips for a healthy immune system. By Hannah Nichols on January 25, — Fact checked by Honor Whiteman.

Share on Pinterest Our immune system protects us from infection and disease, but is there a way we can enhance the way it functions? Can the immune system be boosted?

Share on Pinterest The immune system contains many different cell types that respond to different microbes. Weakened immune system. Impact of lifestyle on immune response. Share on Pinterest Many factors, including diet, exercise, and sleep, can impact immune response.

Diet and the immune system. Share on Pinterest Eating a healthful, balanced diet is important for maintaining immune function. Exercise and the immune system.

Other immune response factors. Share on Pinterest Being outside in the sunshine has been shown to benefit the immune system. Share this article. Latest news Ovarian tissue freezing may help delay, and even prevent menopause.

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Related Coverage. How to stay healthy with a weak immune system. Medically reviewed by Shilpa Amin, M. How the immune system works. Medically reviewed by Meredith Goodwin, MD, FAAFP. The best foods for boosting your immune system. Medically reviewed by Katherine Marengo LDN, R. What to eat and avoid to maintain a strong immune system.

Medically reviewed by Kim Rose-Francis RDN, CDCES, LD. Do supplements really benefit the immune system? Flentie, K. A bioluminescent transposon reporter-trap identifies tumor-specific microenvironment-induced promoters in Salmonella for conditional bacterial-based tumor therapy.

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Download references. We acknowledge support from the Gerald Dewey Dodd, Jr. Endowed Distinguished Chair at The University of Texas MD Anderson Cancer Center. Department of Cancer Systems Imaging, University of Texas MD Anderson Cancer Center, Houston, TX, , USA.

Caleb Gonzalez, Sarah Williamson, Seth T. Chonnam National University Medical School, Gwangju, South Korea. You can also search for this author in PubMed Google Scholar. designed experiments. and S. performed experiments. analyzed data. gifted a reagent. wrote the manuscript. All authors edited the manuscript.

Correspondence to David Piwnica-Worms. The University of Texas MD Anderson Cancer Center has filed a patent application on compounds and methods described in this report C.

The remaining authors declare no competing interests. Communications Biology thanks the anonymous reviewers for their contribution to the peer review of this work. Primary Handling Editors: Shitao Li and Zhijuan Qiu. Open Access This article is licensed under a Creative Commons Attribution 4.

Reprints and permissions. Gonzalez, C. TLR5 agonists enhance anti-tumor immunity and overcome resistance to immune checkpoint therapy. Commun Biol 6 , 31 Download citation. Received : 20 June Accepted : 23 December Published : 12 January Anyone you share the following link with will be able to read this content:.

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Skip to main content Thank you for visiting nature. nature communications biology articles article. Download PDF. Subjects Breast cancer Cancer immunotherapy Cancer therapeutic resistance Chemokines Immunoediting.

Abstract Primary and adaptive resistance to immune checkpoint therapies ICT represent a considerable obstacle to achieving enhanced overall survival.

Introduction Immune checkpoint therapies ICT have opened new therapeutic venues against cancer with lasting curative effects 1 , 2.

Results NF-κB activation of 4T1 cells in vitro To evaluate flagella- and CBLBmediated NF-κB activation of 4T1 mammary carcinoma cells, we stably transfected 4T1 cells with a κB5:IκBɑ-FLuc transcriptional reporter comprised of a concatenated κB5 promoter region, followed by the bioluminescent IκBɑ-FLuc fusion reporter gene 50 , Full size image.

Table 1 Treatment doses and delivery routes. Full size table. Discussion 4T1 mammary carcinoma is a robust murine model to study human triple negative breast cancer, which is highly invasive, metastatic, and resistant to immune check point therapies 96 , Methods Reagents Salmonella typhimurium flagellin FLA-ST was purchased from Invivogen.

Mice All animal procedures were approved by the Institutional Animal Care and Use Committee IACUC of the University of Texas M. Mouse endpoint protocol Mice that reached end point moribund condition or having one tumor measurement in the sagittal or axial plane greater than 1.

Administration of flagellin, CBLB, and immune checkpoint therapy Flagellin, CBLB, 9D9 anti-CTLA-4 , and RPM anti-PD-1 were suspended in filtered PBS; filtered PBS was used as a vehicle control. Bioluminescence imaging in vivo The mice were imaged using the PerkinElmer IVIS Spectrum Imaging System weekly beginning one week after orthotopic injection of 4T1 FUGW-FL cells into the mammary fat pad.

Administration of CBLB and ICT Three days post subcutaneous injection of BF10 cells into the posterior right flank, each mouse was randomly sorted into a group receiving treatment with vehicle control, CBLB only, ICT only 9D9 plus RPM , or CBLB combined with ICT.

Cytokine profile in vivo Sera from mice in experiments 6, 7, and 8 Supplementary Table 2 were obtained by submandibular sampling. Luminex multiplex quantitative analysis Serum samples were analyzed at the Antibody-Based Proteomics Core at Baylor College of Medicine, Houston, TX.

Tumor immune infiltrate profile Tumors and spleens were removed with scissors or forceps and weighted. Flow cytometry Flow cytometry was performed using an LSRII cytometer Becton Dickinson.

Statistics and reproducibility Statistical analyses were performed using GraphPad Prism version 8. Reporting summary Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article.

Data availability All data generated or analyzed during this study are included in this published article and its supplementary information files, including Supplementary Data 1.

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