Therapeutic induction of regulatory, cytotoxic CD8+ T cells in multiple sclerosis

In the setting of autoimmunity, one of the goals of successful therapeutic immune modulation is the induction of peripheral tolerance, a large part of which is mediated by regulatory/suppressor T cells. In this report, we demonstrate a novel immunomodulatory mechanism by an FDA-approved, exogenous peptide-based therapy that incites an HLA class I-restricted, cytotoxic suppressor CD8+ T cell response. We have shown previously that treatment of multiple sclerosis (MS) with glatiramer acetate (GA; Copaxone) induces differential up-regulation of GA-reactive CD8+ T cell responses. We now show that these GA-induced CD8+ T cells are regulatory/suppressor in nature. Untreated patients show overall deficit in CD8+ T cell-mediated suppression, compared with healthy subjects. GA therapy significantly enhances this suppressive ability, which is mediated by cell contact-dependent mechanisms. CD8+ T cells from GA-treated patients and healthy subjects, but not those from untreated patients with MS, exhibit potent, HLA class I-restricted, GA-specific cytotoxicity. We further show that these GA-induced cytotoxic CD8+ T cells can directly kill CD4+ T cells in a GA-specific manner. Killing is enhanced by preactivation of target CD4+ T cells and may depend on presentation of GA through HLA-E. Thus, we demonstrate that GA therapy induces a suppressor/cytotoxic CD8+ T cell response, which is capable of modulating in vivo immune responses during ongoing therapy. These studies not only explain several prior observations relating to the mechanism of this drug but also provide important insights into the natural immune interplay underlying this human immune-mediated disease.     

CD8+ T cells specific for cryptic apoptosis-associated epitopes exacerbate experimental autoimmune encephalomyelitis

The autoimmune immunopathology occurring in multiple sclerosis (MS) is sustained by myelin-specific and -nonspecific CD8⁺ T cells. We have previously shown that, in MS, activated T cells undergoing apoptosis induce a CD8⁺ T cell response directed against antigens that are unveiled during the apoptotic process, namely caspase-cleaved structural proteins such as non-muscle myosin and vimentin. Here, we have explored in vivo the development and the function of the immune responses to cryptic apoptosis-associated epitopes (AEs) in a well-established mouse model of MS, experimental autoimmune encephalomyelitis (EAE), through a combination of immunization approaches, multiparametric flow cytometry, and functional assays. First, we confirmed that this model recapitulated the main findings observed in MS patients, namely that apoptotic T cells and effector/memory AE-specific CD8⁺ T cells accumulate in the central nervous system of mice with EAE, positively correlating with disease severity. Interestingly, we found that AE-specific CD8⁺ T cells were present also in the lymphoid organs of unprimed mice, proliferated under peptide stimulation in vitro, but failed to respond to peptide immunization in vivo, suggesting a physiological control of this response. However, when mice were immunized with AEs along with EAE induction, AE-specific CD8⁺ T cells with an effector/memory phenotype accumulated in the central nervous system, and the disease severity was exacerbated. In conclusion, we demonstrate that AE-specific autoimmunity may contribute to immunopathology in neuroinflammation.Subject terms: Cell death and immune response, Immunological disorders     

Discovery and Function of B-Cell IgD Low (BDL) B Cells in Immune Tolerance

It is now appreciated that in addition to their role in humoral immunity, B cells also exert regulatory mechanisms that lead to attenuation of inflammatory responses. The concept of B-cell regulation became well recognized when mice deficient in B cells due to genetic disruption were shown to be refractory to recovery from the signs of experimental autoimmune encephalomyelitis (EAE), the mouse model of multiple sclerosis. This seminal study spurred the search for B-cell regulatory phenotypes and mechanisms of action. Our approach was to utilize differential B-cell depletion with anti-CD20 to retain B cells whose presence were required to achieve EAE recovery. Utilizing flow cytometry, adoptive cell therapy and genetic approaches, we discovered a new B-cell subset that, upon adoptive transfer into B cell-deficient mice, was sufficient to promote EAE recovery. This B-cell subset is IgM⁺, but due to low/negative IgD cell surface expression, it was named B-cell IgD low (BD_L). Mechanistically, we found that in the absence of BD_L, the absolute cell number of CD4⁺ Foxp3⁺ T regulatory cells (Treg), essential for immune tolerance, was significantly reduced. Furthermore, we found that BD_L expression of glucocorticoid-induced tumor necrosis factor ligand (GITRL) was essential for induction of Treg proliferation and maintenance of their homeostasis. Thus, we have identified a new B-cell subset that is critical for immunological tolerance through interactions with Treg.     

Essential Role of NK Cells in IgG Therapy for Experimental Autoimmune Encephalomyelitis

Intravenous immunoglobulin has long been used in treating autoimmune diseases, although mechanisms remain uncertain. Activating Fcγ receptors are receptors of IgG and reported to be essential in intravenous immunoglobulin (IVIG) therapy. Therefore, we hypothesized natural killer (NK) cells, which express abundant activating Fcγ receptors, are the potential cellular target. In experimental autoimmune encephalomyelitis (EAE), we demonstrated that IgG suppressed disease development in intact, but not in NK cell depleted mice. Adoptive transfer of IgG-treated NK cell could protect mice against EAE, and suppressed interferon γ and interleukin 17 production. The percentage of CD4⁺Foxp3⁺ regulatory T cells was significantly increased. The increase of regulatory T cells was also observed in IgG-treated EAE mice but not in NK cell depleted mice. In vitro experiments confirmed that IgG-treated NK cells enhanced regulatory T cell induction from naïve CD4⁺ T cells. Interestingly, cells from draining lymph nodes produced more interleukin 2 after the adoptive transfer of IgG-treated NK cells. We neutralized interleukin 2 and the induction of CD4⁺Foxp3⁺ T cells by IgG-treated NK cells was significantly reduced. To our knowledge, we identified for the first time the critical role of NK cells in the mechanism of IgG-induced induction of Treg cells in treatment of autoimmunity.     

Targeting the Shift from M1 to M2 Macrophages in Experimental Autoimmune Encephalomyelitis Mice Treated with Fasudil

We observed the therapeutic effect of Fasudil and explored its mechanisms in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). Fasudil, a selective Rho kinase (ROCK) inhibitor, was injected intraperitoneally at 40 mg/kg/d in early and late stages of EAE induction. Fasudil ameliorated the clinical severity of EAE at different stages, and decreased the expression of ROCK-II in spleen, accompanied by an improvement in demyelination and inhibition of inflammatory cells. Fasudil mainly inhibited CD4⁺IL-17⁺ T cells in early treatment, but also elevated CD4⁺IL-10⁺ regulatory T cells and IL-10 production in late treatment. The treatment of Fasudil shifted inflammatory M1 to anti-inflammatory M2 macrophages in both early and late treatment, being shown by inhibiting CD16/32, iNOS, IL-12, TLR4 and CD40 and increasing CD206, Arg-1, IL-10 and CD14 in spleen. By using Western blot and immunohistochemistry, iNOS and Arg-1, as two most specific markers for M1 and M2, was inhibited or induced in splenic macrophages and spinal cords of EAE mice treated with Fasudil. In vitro experiments also indicate that Fasudil shifts M1 to M2 phenotype, which does not require the participation or auxiliary of other cells. The polarization of M2 macrophages was associated with the decrease of inflammatory cytokine IL-1β, TNF-α and MCP-1. These results demonstrate that Fasudil has therapeutic potential in EAE possibly through inducing the polarization of M2 macrophages and inhibiting inflammatory responses.     

Ectonucleotidase CD38 Demarcates Regulatory,Memory-Like CD8+ T Cells with IFN-γ-Mediated Suppressor Activities

Regulatory CD8⁺ T cells are critical for self-tolerance and restricting excessive immune responses. The variety of immune functions they fulfill, the heterogeneity of their phenotype, and the mechanism of action are still poorly understood. Here we describe that regulatory CD8⁺ T cells exhibiting immunosuppressive actions in vitro and in vivo are recognized as CD38^high T cells and present in naive mice. CD38 is a glycosylated membrane protein with ectonucleotidase properties. CD8⁺CD38^high (CD44⁺CD122⁺CD62L^high) lymphocytes suppress CD4⁺ effector T-cell proliferation in an antigen-non specific manner via IFN-γ. While direct cell-to-cell contact is needed for this suppressor activity, it is independent of membrane-bound TGF-β and granzyme B release. IL-15 potentiates the suppressive activity of CD8⁺CD38^high T cells and controls their survival and expansion. In humans CD8⁺CD38^high T cells inhibit CD4⁺ effector T cell proliferation. In vivo, CD8⁺CD38^high, but not CD8⁺CD38⁻ T cells mitigate murine experimental autoimmune encephalomyelitis (EAE) by reducing the clinical score and delaying disease occurrence. EAE suppression is enhanced by pre-treatment of CD8⁺CD38^high T cells with IL-15. These findings add evidence that the expression of ectoenzyme receptor family members positively correlates with suppressor functions and identifies CD8⁺CD38^high T cells as potential inhibitors of excessive immune responses.     

Human amniotic epithelial cells suppress relapse of corticosteroid-remitted experimental autoimmune disease

Background aimsMultiple sclerosis (MS) is considered to be a T-cell–mediated disease. Although MS remits with corticosteroid treatment, the disease relapses on discontinuation of therapy. Human amniotic epithelial cells (hAEC) from the placenta are readily accessible in large quantities and have anti-inflammatory properties. Previously we reported that hAEC given near disease onset ameliorated clinical signs and decreased myelin oligodendrocyte glycoprotein (MOG)-specific immune responses in MOG-induced experimental autoimmune encephalomyelitis (EAE), an experimental MS model.MethodsTo examine the therapeutic effect of hAEC in a clinically relevant setting, we first treated MOG peptide–induced EAE mice with a corticosteroid, prednisolone, in drinking water to induce remission. hAEC were then infused intravenously into the remitted mice. Anti-MOG antibodies in serum were detected by enzyme-linked immunoassay. Splenocyte proliferation was assessed by ³H-thymidine incorporation. Immune cell subpopulations in spleens and lymph nodes and secreted cytokines in splenocyte culture were quantified by flow cytometry. Central nervous system histology was examined with the use of hematoxylin and eosin, Luxol fast blue and immunostaining.ResultsWith cessation of prednisolone treatment, hAEC delayed EAE relapse for 7 days, and, after another 7 days, largely remitted disease in six of eight responder mice. Splenocyte proliferation was suppressed, anti-MOG_35–55 antibodies in serum were decreased and interleukin-2 and interleukin-5 production by splenocytes were elevated after hAEC treatment. In the central nervous system, hAEC-treated mice had decreased demyelination and fewer macrophages in the inflammatory infiltrates. hAEC treatment also increased CD4⁺CD25⁺FoxP3⁺ regulatory T cells in inguinal lymph nodes.ConclusionsThese data demonstrate that the therapeutic effects of hAEC after corticosteroid treatment in an MS model probably are the consequence of peripheral immunoregulation. We suggest that hAEC may have potential as a cell therapy for remitted MS.     

Type II monocytes modulate T cell-mediated central nervous system autoimmune disease

Treatment with glatiramer acetate (GA, copolymer-1, Copaxone), a drug approved for multiple sclerosis (MS), in a mouse model promoted development of anti-inflammatory type II monocytes, characterized by increased secretion of interleukin (IL)-10 and transforming growth factor (TGF)-beta, and decreased production of IL-12 and tumor necrosis factor (TNF). This anti-inflammatory cytokine shift was associated with reduced STAT-1 signaling. Type II monocytes directed differentiation of T(H)2 cells and CD4+CD25+FoxP3+ regulatory T cells (T(reg)) independent of antigen specificity. Type II monocyte-induced regulatory T cells specific for a foreign antigen ameliorated experimental autoimmune encephalomyelitis (EAE), indicating that neither GA specificity nor recognition of self-antigen was required for their therapeutic effect. Adoptive transfer of type II monocytes reversed EAE, suppressed T(H)17 cell development and promoted both T(H)2 differentiation and expansion of T(reg) cells in recipient mice. This demonstration of adoptive immunotherapy by type II monocytes identifies a central role for these cells in T cell immune modulation of autoimmunity.     

Modulation of outward K+ conductance is a post-activational event in rat T lymphocytes responsible for the adoptive transfer of experimental allergic encephalomyelitis

Experimental allergic encephalomyelitis (EAE) is an accepted animal model for the human demyelinating disease multiple sclerosis. The continuously propagated line of Lewis rat T helper lymphocytes (GP1 T cells), specific for the encephalitogenic 68–86 sequence of guinea pig myelin basic protein (GPMBP), mediates the adoptive transfer of EAE into normal syngeneic Lewis rats. Because mitogenic activation of T cells can increase K⁺ conductance, this study investigated changes in the outwardly rectifying K⁺ conductance in GP1 T cells following activation with the encephalitogen, GPMBP. Using the gigohm-seal whole-cell variation of the patch clamp technique, GP1 T cells were studied during a 3-day culture with GPMBP and throughout the subsequent 10 days, as cells progressed through both GPMBP-induced activation (EAE transfer activity) and proliferation responses, finally reverting to the resting state. Resting GP1 T cells exhibited peak K⁺ conductances around 2 nS, while GPMBP-induced activation resulted in 5- to 10-fold increases in peak K⁺ conductance, which temporally coincided with the optimal period of EAE transfer activity. During and immediately after the optimal period for EAE transfer, 20-mV depolarizing shifts in the voltage dependence of both activation and inactivation developed, abruptly reversing to resting values as cells reverted to the resting state. Accompanying the depolarizing shifts were a slowing of the K⁺ current activation kinetics and an acceleration of the deactivation kinetics. These results indicate that the K⁺ conductance in GP1 rat T helper cells is modulated over the full time course of GPMBP-induced cellular responses and that K⁺ channels should be optimally available during the period of adoptive EAE transfer, preceding disease manifestation.     

Regulatory T cell induction during Plasmodium chabaudi infection modifies the clinical course of experimental autoimmune encephalomyelitis

Background

Experimental autoimmune encephalomyelitis (EAE) is used as an animal model for human multiple sclerosis (MS), which is an inflammatory demyelinating autoimmune disease of the central nervous system characterized by activation of Th1 and/or Th17 cells. Human autoimmune diseases can be either exacerbated or suppressed by infectious agents. Recent studies have shown that regulatory T cells play a crucial role in the escape mechanism of Plasmodium spp. both in humans and in experimental models. These cells suppress the Th1 response against the parasite and prevent its elimination. Regulatory T cells have been largely associated with protection or amelioration in several autoimmune diseases, mainly by their capacity to suppress proinflammatory response.

Methodology/Principal Findings

In this study, we verified that CD4⁺CD25⁺ regulatory T cells (T regs) generated during malaria infection (6 days after EAE induction) interfere with the evolution of EAE. We observed a positive correlation between the reduction of EAE clinical symptoms and an increase of parasitemia levels. Suppression of the disease was also accompanied by a decrease in the expression of IL-17 and IFN-γ and increases in the expression of IL-10 and TGF-β1 relative to EAE control mice. The adoptive transfer of CD4⁺CD25⁺ cells from P. chabaudi-infected mice reduced the clinical evolution of EAE, confirming the role of these T regs.

Conclusions/Significance

These data corroborate previous findings showing that infections interfere with the prevalence and evolution of autoimmune diseases by inducing regulatory T cells, which regulate EAE in an apparently non-specific manner.     

CD40 Activation Rescues Antiviral CD8+ T Cells from PD-1-Mediated Exhaustion

The intrahepatic immune environment is normally biased towards tolerance. Nonetheless, effective antiviral immune responses can be induced against hepatotropic pathogens. To examine the immunological basis of this paradox we studied the ability of hepatocellularly expressed hepatitis B virus (HBV) to activate immunologically naïve HBV-specific CD8⁺ T cell receptor (TCR) transgenic T cells after adoptive transfer to HBV transgenic mice. Intrahepatic priming triggered vigorous in situ T cell proliferation but failed to induce interferon gamma production or cytolytic effector function. In contrast, the same T cells differentiated into cytolytic effector T cells in HBV transgenic mice if Programmed Death 1 (PD-1) expression was genetically ablated, suggesting that intrahepatic antigen presentation per se triggers negative regulatory signals that prevent the functional differentiation of naïve CD8⁺ T cells. Surprisingly, coadministration of an agonistic anti-CD40 antibody (αCD40) inhibited PD-1 induction and restored T cell effector function, thereby inhibiting viral gene expression and causing a necroinflammatory liver disease. Importantly, the depletion of myeloid dendritic cells (mDCs) strongly diminished the αCD40 mediated functional differentiation of HBV-specific CD8⁺ T cells, suggesting that activation of mDCs was responsible for the functional differentiation of HBV-specific CD8⁺ T cells in αCD40 treated animals. These results demonstrate that antigen-specific, PD-1-mediated CD8⁺ T cell exhaustion can be rescued by CD40-mediated mDC-activation.     

Measurement of CD8+ and CD4+ T Cell Frequencies Specific for EBV LMP1 and LMP2a Using mRNA-Transfected DCs

An EBV-specific cellular immune response is associated with the control of EBV-associated malignancies and lymphoproliferative diseases, some of which have been successfully treated by adoptive T cell therapy. Therefore, many methods have been used to measure EBV-specific cellular immune responses. Previous studies have mainly used autologous EBV-transformed B-lymphoblastoid cell lines (B-LCLs), recombinant viral vectors transfected or peptide pulsed dendritic cells (DCs) as stimulators of CD8⁺ and CD4⁺ T lymphocytes. In the present study, we used an interferon-γ (IFN-γ) enzyme-linked immunospot (ELISPOT) assay by using isolated CD8⁺ and CD4⁺ T cells stimulated with mRNA-transfected DCs. The frequency of latent membrane protein 1 (LMP1)-specific IFN-γ producing CD4⁺ T cells was significantly higher than that of LMP2a. The frequency of IFN-γ producing CD4⁺ T cells was significantly correlated with that of CD8⁺ T cells in LMP1-specific immune responses (r = 0.7187, Pc < 0.0001). To determine whether there were changes in LMP1- or LMP2a-specific immune responses, subsequent peripheral blood mononuclear cells (PBMCs) samples were analyzed. Significant changes were observed in 5 of the 10 donors examined, and CD4⁺ T cell responses showed more significant changes than CD8⁺ T cell responses. CD8⁺ and CD4⁺ T cells from EBV-seropositive donors secreted only the Th1 cytokines IFN-γ, TNF-α, and IL-2, while Th2 (IL-4) and Th17 (IL-17a) cytokines were not detected. CD4⁺ T cells secreted significantly higher cytokine levels than did CD8⁺ T cells. Analysis of EBV-specific T cell responses using autologous DCs transfected with mRNA might provide a comprehensive tool for monitoring EBV infection and new insights into the pathogenesis of EBV-associated diseases.     

T cell precursor frequency differentially affects CTL responses under different immune conditions

Generation of effective CTL responses is the goal of many vaccination protocols. However, to what extant T cell precursor frequencies will generate a CD8⁺ CTL response has not been elucidated properly. In this study, we employed a model system, in which naive CD4⁺ and CD8⁺ T cells derived from ovalbumin (OVA)-specific TCR transgenic OT II and OT I mice were used for adoptive transfer into wild-type, Ia^b−/− gene knockout and transgenic RIP-mOVA mice, and assessed OVA-pulsed DC (DC_OVA)-stimulated CD8⁺ CTL responses in these mice. We demonstrated that (i) a critical threshold exists above which T cells precursor frequency cannot enhance the CTL responses in wild-type C57BL/6 mice, (ii) increasing CD8⁺ T cell precursors is required to generate CTL responses but with functional memory defect in absence of CD4⁺ T cell help, and (iii) increasing CD4⁺ and CD8⁺ T cell precursors overcomes immune suppression to DC_OVA-stimulated CD8⁺ CTL responses in transgenic RIP-mOVA mice with OVA-specific self immune tolerance. Taken together, these findings may have important implications for optimizing immunotherapy against cancer.     

CD8+ cytotoxic T lymphocytes in cancer immunotherapy: A review

CD8⁺ cytotoxic T lymphocytes (CTLs) are preferred immune cells for targeting cancer. During cancer progression, CTLs encounter dysfunction and exhaustion due to immunerelated tolerance and immunosuppression within the tumor microenvironment (TME), with all favor adaptive immune-resistance. Cancer-associated fibroblasts (CAFs), macrophage type 2 (M2) cells, and regulatory T cells (Tregs) could make immunologic barriers against CD8 ⁺ T cell-mediated antitumor immune responses. Thus, CD8 ⁺ T cells are needed to be primed and activated toward effector CTLs in a process called tumor immunity cycle for making durable and efficient antitumor immune responses. The CD8 ⁺ T cell priming is directed essentially as a corroboration work between cells of innate immunity including dendritic cells (DCs) and natural killer (NK) cells with CD4 ⁺ T cells in adoptive immunity. Upon activation, effector CTLs infiltrate to the core or invading site of the tumor (so-called infiltrated–inflamed [I–I] TME) and take essential roles for killing cancer cells. Exogenous reactivation and/or priming of CD8 ⁺ T cells can be possible using rational immunotherapy strategies. The increase of the ratio for costimulatory to coinhibitory mediators using immune checkpoint blockade (ICB) approach. Programmed death-1 receptor (PD-1)–ligand (PD-L1) and CTL-associated antigen 4 (CTLA-4) are checkpoint receptors that can be targeted for relieving exhaustion of CD8 ⁺ T cells and renewing their priming, respectively, and thereby eliminating antigen-expressing cancer cells. Due to a diverse relation between CTLs with Tregs, the Treg activity could be dampened for increasing the number and rescuing the functional potential of CTLs to induce immunosensitivity of cancer cells.     

Proliferation-Linked Apoptosis of Adoptively Transferred T Cells after IL-15 Administration in Macaques

The adoptive transfer of antigen-specific effector T cells is being used to treat human infections and malignancy. T cell persistence is a prerequisite for therapeutic efficacy, but reliably establishing a high-level and durable T cell response by transferring cultured CD8⁺ T cells remains challenging. Thus, strategies that promote a transferred high-level T cell response may improve the efficacy of T cell therapy. Lymphodepletion enhances persistence of transferred T cells in mice in part by reducing competition for IL-15, a common γ-chain cytokine that promotes T cell memory, but lymphodepleting regimens have toxicity. IL-15 can be safely administered and has minimal effects on CD4⁺ regulatory T cells at low doses, making it an attractive adjunct in adoptive T cell therapy. Here, we show in lymphoreplete macaca nemestrina, that proliferation of adoptively transferred central memory-derived CD8⁺ effector T (T_CM/E) cells is enhanced in vivo by administering IL-15. T_CM/E cells migrated to memory niches, persisted, and acquired both central memory and effector memory phenotypes regardless of the cytokine treatment. Unexpectedly, despite maintaining T cell proliferation, IL-15 did not augment the magnitude of the transferred T cell response in blood, bone marrow, or lymph nodes. T cells induced to proliferate by IL-15 displayed increased apoptosis demonstrating that enhanced cycling was balanced by cell death. These results suggest that homeostatic mechanisms that regulate T cell numbers may interfere with strategies to augment a high-level T cell response by adoptive transfer of CD8⁺ T_CM/E cells in lymphoreplete hosts.     

Mobilization of Regulatory T Cells in Response to Carotid Injury Does Not Influence Subsequent Neointima Formation

Aim

T cells have been attributed an important role in modulating repair responses following vascular injury. The aim of this study was to investigate the role of different T cell subsets in this context.

Methods and Results

A non-obstructive collar was introduced to inflict carotid artery injury in mice and subsequent activation of immune cells in draining lymph nodes and spleen were studied by flow cytometry. Carotid artery injury of wild type mice was associated with mobilization of both Th1 type CD4⁺IFNγ⁺ and regulatory CD4⁺CD25⁺FoxP3⁺ T cells in draining lymph nodes. Studies using FoxP3-green fluorescent protein (GFP) transgenic C57/Bl6 mice demonstrated scattered presence of regulatory T cells in the adventitial tissue of injured arteries as well as a massive emigration of regulatory T cells from the spleen in response to carotid injury. However, deletion of antigen presentation to CD4+ T cells (H2⁰ mice), as well as deletion of regulatory T cells (through treatment with blocking anti-CD25 antibodies), did not affect neointima formation. Also deletion of antigen presentation to CD8⁺ T cells (Tap1⁰ mice) was without effect on carotid collar-induced neointima formation.

Conclusion

The results demonstrate that carotid artery injury is associated with mobilization of regulatory T cells. Depletion of regulatory T cells does not, however, influence the subsequent repair processes leading to the formation of a neointima. The results also demonstrate that lack of CD8⁺ T cells does not influence neointima formation in presence of functional CD4⁺ T cells and B cells.     

Galectins and their ligands: negative regulators of anti-tumor immunity

Cytotoxic CD8⁺ T cells are major players of anti-tumor immune responses, as their functional activity can limit tumor growth and progression. Data show that cytotoxic T cells efficiently control the proliferation of tumor cells through major histocompatibility complex class I-mediated mechanisms; nevertheless, the presence of tumor-infiltrating CD8⁺ T cells in lesional tissue does not always correlate with better prognosis and increased survival of cancer patients. Similarly, adoptive transfer of tumor-specific cytotoxic T cells has only shown marginal improvement in life spans of patients with metastatic disease. In this report, we discuss experimental evidence showing that expression of tumor-derived galectins, galectin (Gal)-1, Gal-3 and Gal-9, and concomitant presence of their ligands on the surface of anti-tumor immunocytes directly compromise anti-tumor CD8⁺ T cell immune responses and, perhaps, undermine the promise of adoptive CD8⁺ T cell immunotherapy. Furthermore, we describe novel strategies designed to counteract Gal-1-, Gal-3- and Gal-9-mediated effects and highlight their targeting potential for creating more effective anti-tumor immune responses. We believe that Gal and their ligands represent an efficacious targeted molecular paradigm that warrants clinical evaluation.     

Combining Regulatory T Cell Depletion and Inhibitory Receptor Blockade Improves Reactivation of Exhausted Virus-Specific CD8+ T Cells and Efficiently Reduces Chronic Retroviral Loads

Chronic infections with human viruses, such as HIV and HCV, or mouse viruses, such as LCMV or Friend Virus (FV), result in functional exhaustion of CD8⁺ T cells. Two main mechanisms have been described that mediate this exhaustion: expression of inhibitory receptors on CD8⁺ T cells and expansion of regulatory T cells (Tregs) that suppress CD8⁺ T cell activity. Several studies show that blockage of one of these pathways results in reactivation of CD8⁺ T cells and partial reduction in chronic viral loads. Using blocking antibodies against PD-1 ligand and Tim-3 and transgenic mice in which Tregs can be selectively ablated, we compared these two treatment strategies and combined them for the first time in a model of chronic retrovirus infection. Blocking inhibitory receptors was more efficient than transient depletion of Tregs in reactivating exhausted CD8⁺ T cells and reducing viral set points. However, a combination therapy was superior to any single treatment and further augmented CD8⁺ T cell responses and resulted in a sustained reduction in chronic viral loads. These results demonstrate that Tregs and inhibitory receptors are non-overlapping factors in the maintenance of chronic viral infections and that immunotherapies targeting both pathways may be a promising strategy to treat chronic infectious diseases.     

Peripheral Blood T Cell Dynamics Predict Relapse in Multiple Sclerosis Patients on Fingolimod

Background

Fingolimod efficiently reduces multiple sclerosis (MS) relapse by inhibiting lymphocyte egress from lymph nodes through down-modulation of sphingosine 1-phosphate (S1P) receptors. We aimed to clarify the alterations in peripheral blood T cell subsets associated with MS relapse on fingolimod.

Methods/Principal Findings

Blood samples successively collected from 23 relapsing-remitting MS patients before and during fingolimod therapy (0.5 mg/day) for 12 months and 18 healthy controls (HCs) were analysed for T cell subsets by flow cytometry. In MS patients, the percentages of central memory T (CCR7⁺CD45RO⁺) cells (TCM) and naïve T (CCR7⁺CD45RO^-) cells decreased significantly, while those of effector memory T (CCR7^-CD45RA^-) and suppressor precursor T (CD28^-) cells increased in both CD4⁺T and CD8⁺T cells from 2 weeks to 12 months during fingolimod therapy. The percentages of regulatory T (CD4⁺CD25^highCD127^low) cells in CD4⁺T cells and CCR7^-CD45RA⁺T cells in CD8⁺T cells also increased significantly. Eight relapsed patients demonstrated greater percentages of CD4⁺TCM than 15 non-relapsed patients at 3 and 6 months (p=0.0051 and p=0.0088, respectively). The IL17-, IL9-, and IL4-producing CD4⁺T cell percentages were significantly higher at pre-treatment in MS patients compared with HCs (p<0.01 for all), while the IL17-producing CD4⁺T cell percentages tended to show a transient increase at 2 weeks of fingolimod therapy (p^corr=0.0834).

Conclusions

The CD4⁺TCM percentages at 2 weeks to 12 months during fingolimod therapy are related to relapse.