Local Immunosuppression Research Articles

Single-cell RNA-sequencing of human spleens reveals an IDO-1+ Tolerogenic Dendritic Cell subset in pancreatic cancer patients that is absent in normal individuals.

Local and systemic immunosuppression are prominent features of pancreatic cancer, rendering anti-tumor effector cells inactive and immunotherapeutic approaches ineffective. The spleen, an understudied point of antigen-presentation and T cell priming in humans, holds particular importance in pancreatic cancer due to its proximity to the developing tumor. As main effectors of antigen presentation, dendritic cells display antigens to lymphocytes, thereby bridging the innate and adaptive immune response. While tumor-infiltrating anti-inflammatory dendritic cells have been described, splenic dendritic cells have historically just been considered to stimulate the anti-tumor immune response. Here, we describe, for the first time, the presence of an immunosuppressive, tolerogenic IDO1+ dendritic cell subset in the spleens of pancreatic cancer patients that likely contributes to systemic immunosuppression that is associated with pancreatic ductal adenocarcinoma. Network analysis of scRNA seq data reveals extensive communication networks between the identified tolerogenic DC cluster and numerous immune cell populations in the spleen. Interactions with innate and adaptive immune cells suggest a broad influence on leukocyte trafficking and immune regulation within the spleen microenvironment. The identification of signaling pathways involving AHR and IDO-1, CCL19, NECTIN2, CLEC2D, and others elucidates potential mechanisms underlying the immunosuppressive functions of this cell type.

TMIC-87. ICAM-1 REGULATE EXTRACELLULAR VESICLE-MEDIATED IMMUNOSUPPRESSIVE MONOCYTE IN HUMAN GLIOBLASTOMA CELL

Abstract Glioblastoma is the most common malignant primary brain tumor and is universally fatal. Treatment involves maximal safe resection with adjuvant chemoradiation. Poor prognosis in glioblastoma is multifactorial, and local and systemic immunosuppression is an important factor. Extracellular vesicles (EVs) are small, membrane-bound particles released by cells into the external environment. They play a key role in intercellular communication. In glioblastomas, EVs regulate immunosuppression through induction of myeloid-derived suppressor cells (MDSCs) in a process dependent on the PD-1/PD-L1 and IL-6 axes. Primary cilia are microtubule-based organelles that play crucial roles in various cellular processes and signal transduction cascades. We recently showed that primary cilia modulate IL-6 expression and consequently, MDSC induction. Comparative proteomic analysis of EVs derived from human glioblastoma cells devoid of cilia showed loss of ICAM-1 expression compared to control glioblastoma cells. ICAM-1 (Intercellular Adhesion Molecule-1) is a cell surface glycoprotein that mediates cell-cell adhesion and immune responses. In this study, we showed that depletion of primary cilia in human glioblastoma cells result in increased PD-L1 but loss of IL-6 and ICAM-1. RNAi-meidated loss of ICAM-1 was synergistic with primary cilia depletion in preventing MDSC inductionn normal human monocytes exposed to glioblastoma-derived EVs. Exogenous IFN-g was sufficient to restore IL-6 levels and MDSC induction despite loss of primary cilia and ICAM-1 suggesting that primary cilia act upstream of IFN-g, IL-6 and ICAM-1 in glioblastoma-mediated MDSC induction. This study is the first to connect primary cilia signal transduction to IL-6 and ICAM-1 expression and to MDSC induction. The evidence suggests a potential parallel primary cilia regulatory cascade independent of the PD-L1 axis that controls tumor-mediated immunosuppression. These results may inform novel therapeutic strategies aimed at increasing efficacy of immunotherapy in the glioblastoma and other neoplastic processes.

CNSC-66. STRUCTURAL AND ELECTROPHYSIOLOGICAL CONNECTIVITY IN GLIOBLASTOMA IS MEDIATED BY TUMOR AND IMMUNE DERIVED CYTOKINES

Abstract BACKGROUND In glioblastomas (GBM), the pleiotropic cytokine interleukin-6 (IL-6) upregulates PD-L1 expression. This results in local immunosuppression favorable to tumor progression, and may account for the limited efficacy of immune checkpoint inhibition. Additionally, IL-6 increases synaptogenesis during development, and contributes to neuronal network remodeling following focal brain or spinal cord injury. Therefore, the aim was to determine whether IL-6 contributes to GBM-induced circuit remodeling from a structural and electrophysiological perspective. METHODS In magnetoencephalography (MEG)-based intratumoral regions of High- (HFC) and Low-Functional Connectivity (LFC) from GBM patients, the expression of IL-6 and GAP43 was compared using bulk and single-cell RNA sequencing and immunofluorescence staining in biological replicates (n=44). In mouse postnatal cortical neurons and primary patient-derived GBM cell cocultures, the expression of Ki67, GAP43, tumor connectivity and neuronal activity were monitored after exogenous application of recombinant human IL-6 (rIL-6) or microglial depletion using PLX5622. Results. IL-6 and GAP43 were overexpressed in HFC compared to LFC regions. In neuron-GBM cocultures, the tumour proliferation index was significantly increased by rIL-6 and decreased by PLX5622. Morphologically, the number of microtubes per tumor cell was significantly increased by rIL-6 and decreased by PLX5622. The expression of GAP43 was inconstantly increased by rIL-6, suggesting the regulation of other pathways involved in tumour microtube formation. Multielectrode array recordings showed a significant increase or decrease in the neuronal activity, following the addition of rIL-6 or microglial depletion, respectively. CONCLUSIONS IL-6-signaling increases proliferation, structural and electrophysiological connectivity in gliomas and may contribute to neuronal network hyper-excitability and synchrony. Microglial depletion provided opposite effects. Additional experiments are mandatory to determine whether the effects of IL-6 on GBM cells depend on the presence of microglial cells or not. These data will be critical to design new therapeutic options, likely to improve the oncological and functional outcomes of patients treated for GBM.

Oncolytic virus and CAR-T cell therapy in solid tumors.

Adoptive immunotherapy with T cells, genetically modified to express a tumor-reactive chimeric antigen receptor (CAR), is an innovative and rapidly developing life-saving treatment for cancer patients without other therapeutic opportunities. CAR-T cell therapy has proven effective only in hematological malignancies. However, although by now only a few clinical trials had promising outcomes, we predict that CAR-T therapy will eventually become an established treatment for several solid tumors. Oncolytic viruses (OVs) can selectively replicate in and kill cancer cells without harming healthy cells. They can stimulate an immune response against the tumor, because OVs potentially stimulate adaptive immunity and innate components of the host immune system. Using CAR-T cells along with oncolytic viruses may enhance the efficacy of CAR-T cell therapy in destroying solid tumors by increasing the tumor penetrance of T cells and reducing the immune suppression by the tumor microenvironment. This review describes recent advances in the design of oncolytic viruses and CAR-T cells while providing an overview of the potential combination of oncolytic virotherapy with CAR-T cells for solid cancers. In this review, we will focus on the host-virus interaction in the tumor microenvironment to reverse local immunosuppression and to develop CAR-T cell effector function.

Chimeric antigen receptor T-cell therapy in patients with malignant glioma-From neuroimmunology to clinical trial design considerations.

Clinical trials evaluating chimeric antigen receptor (CAR) T-cell therapy in patients with malignant gliomas have shown some early promise in pediatric and adult patients. However, the long-term benefits and safety for patients remain to be established. The ultimate success of CAR T-cell therapy for malignant glioma will require the integration of an in-depth understanding of the immunology of the central nervous system (CNS) parenchyma with strategies to overcome the paucity and heterogeneous expression of glioma-specific antigens. We also need to address the cold (immunosuppressive) microenvironment, exhaustion of the CAR T-cells, as well as local and systemic immunosuppression. Here, we discuss the basics and scientific considerations for CAR T-cell therapies and highlight recent clinical trials. To help identify optimal CAR T-cell administration routes, we summarize our current understanding of CNS immunology and T-cell homing to the CNS. We also discuss challenges and opportunities related to clinical trial design and patient safety/monitoring. Finally, we provide our perspective on future prospects in CAR T-cell therapy for malignant gliomas by discussing combinations and novel engineering strategies to overcome immuno-regulatory mechanisms. We hope this review will serve as a basis for advancing the field in a multiple discipline-based and collaborative manner.

Targeting the LMP1-ALIX axis in EBV+ nasopharyngeal carcinoma inhibits immunosuppressive small extracellular vesicle secretion and boosts anti-tumor immunity.

Immunotherapy has revolutionized the therapeutical regimen for nasopharyngeal carcinoma (NPC), yet its response rate remains insufficient. Programmed death-ligand 1 (PD-L1) on small extracellular vesicles (sEVs) mediates local and peripheral immunosuppression in tumors, and the mechanism of PD-L1 loading into these vesicles is garnering increasing attention. Latent membrane protein 1 (LMP1), a key viral oncoprotein expressed in Epstein-Barr virus (EBV)-positive NPC, contributes to remodeling the tumor microenvironment. However, the precise mechanisms by which LMP1 modulates tumor immunity in NPC remain unclear. Here, we aimed to investigate the roles and regulatory mechanisms of LMP1 and sEV PD-L1 in NPC immune evasion. We analyzed the impact of LMP1 on tumor-infiltrating lymphocyte abundance in NPC tissues and humanized tumor-bearing mouse models using multiplex immunofluorescence (mIF) and flow cytometry, respectively. Transmission electron microscopy and nanoparticle tracking analysis were employed to characterize sEVs. Immunoprecipitation-mass spectrometry was utilized to identify proteins interacting with LMP1. The regulatory effects of sEVs on tumor microenvironment were assessed by monitoring CD8+ T cell proliferation and interferon-γ (IFN-γ) expression via flow cytometry. Furthermore, the expression patterns of LMP1 and downstream regulators in NPC were analyzed using mIF and survival analysis. High LMP1 expression in NPC patient specimens and mouse models was associated with restricted infiltration of CD8+ T cells. Additionally, LMP1 promoted sEV PD-L1 secretion, leading to inhibition of CD8+ T cell viability and IFN-γ expression in vitro. Mechanistically, LMP1 recruited apoptosis-linked gene 2-interacting protein X (ALIX) through its intracellular domain and bound PD-L1 through its transmembrane domain, thereby facilitating the loading of PD-L1 into ALIX-dependent sEVs. Disruption of ALIX diminished LMP1-induced sEV PD-L1 secretion and enhanced the anti-tumor immunity of CD8+ T cells both in vitro and in vivo. Moreover, increased expression levels of LMP1 and ALIX were positively correlated with enhanced immunosuppressive features and worse prognostic outcomes in NPC patients. Our findings uncovered the mechanism by which LMP1 interacts with ALIX and PD-L1 to form a trimolecular complex, facilitating PD-L1 loading into ALIX-dependent sEV secretion pathway, ultimately inhibiting the anti-tumor immune response in NPC. This highlights a novel target and prognostic marker for NPC immunotherapy.

Vitreoretinal Surgery in Pediatric Patients with Vitreoretinal Pathology Secondary to Uveitis

In pediatric uveitis, as with all forms of uveitis, the primary treatment modality involves achieving systemic and local immunosuppression through pharmacotherapy. However, due to the challenge of articulating symptoms by patients in the younger age group, resulting in delayed presentation to clinicians, or the tendency for uveitis to manifest more quietly in this age group, pediatric uveitis cases can be diagnosed late, unfortunately presenting with secondary complications at the time of diagnosis. In these patients, after appropriate immunosuppression is achieved, vitreoretinal surgery may be necessary to address vitreoretinal complications. Additionally, in cases where etiology cannot be determined by non-invasive methods, or when there is inadequate response or no response to treatment, rapid progression of inflammation threatening the optic nerve and macula, or suspicion of malignancy, diagnostic vitreous sampling or chorioretinal biopsy via vitreoretinal surgery may be pursued. The aim of this review is to evaluate the role and significance of vitreoretinal surgery in pediatric uveitic eyes.

TIMP1 mediates astrocyte-dependent local immunosuppression in brain metastasis acting on infiltrating CD8+ T cells.

Immunotherapies against brain metastases have shown clinical benefits when applied to asymptomatic patients, but they are largely ineffective in symptomatic cases for unknown reasons. Here we dissect the heterogeneity in metastasis-associated astrocytes using scRNAseq and report a population that blocks the antitumoral activity of infiltrating T cells. This pro-tumoral activity is mediated by the secretion of TIMP1 from a cluster of pSTAT3+ astrocytes that acts on CD63+ CD8+ T cells to modulate their function. Using genetic and pharmacologic approaches in mouse and human brain metastasis models, we demonstrate that combining immune checkpoint blockade antibodies with the inhibition of astrocyte-mediated local immunosuppression may benefit patients with symptomatic brain metastases. We further reveal that the presence of TIMP1 in liquid biopsies provides a biomarker to select patients for this combined immunotherapy. Overall, our findings demonstrate an unexpected immunomodulatory role for astrocytes in brain metastases with clinical implications.

Abstract A045: Disrupting local immunosuppression through combination myCAF/myeloid targeting in pancreatic cancer

Abstract Immunotherapy has revolutionized clinical care for many cancers, yet the approach has not been fully realized in immunologically cold tumors like Pancreatic Ductal Adenocarcinoma (PDAC). One striking hallmark of PDAC is local immunosuppression (LIS) which leads to cytotoxic T cell inactivation and exclusion. There is great need for an increased understanding of the cellular crosstalk within tumors and the identification of stromal and immune populations involved in shaping the tumor microenvironment (TME). Oncogenic KRAS activation in tumor cells promotes the invasion and proliferation of tumor-supporting stromal cells, while excluding cancer-targeted cytotoxic T cells. LIS is mediated by multiple subtypes of cancer-associated fibroblasts (CAFs) and myeloid cells resident within the tumor parenchyma. Multiple prior attempts to reverse LIS in PDAC by targeting individual stromal cell populations have been unsuccessful, alluding to the complexity of stromal crosstalk within the TME. The stromal diversity of PDAC complicates investigating paracrine cascades involving multiple cell types. To decipher diverse drug effects on altering the TME, we employ in vivo studies in mouse models recapitulating the human disease, as well as a novel tumor explant model that enables the short-term culture of intact human or murine PDAC. Importantly, PDAC explants maintain their histopathological architecture and cellular diversity over time. This medium-throughput platform allows for testing of multiple drugs and mechanistic hypotheses in the native PDAC TME. We show in preliminary data that Smoothened inhibition (SMOi) decreases proliferation and activity of myCAFs, but provokes the expansion of CD11b-positive myeloid cells in vivo. Thus, we hypothesize that LIS in PDAC is maintained by a delicate balance between myCAFs and myeloid cells, preventing effective T cell invasion. Single cell RNA-seq data comparing ctrl vs. SMOi-treated murine PDAC elucidates stromal subpopulations involved in the LIS phenotype and guides the identification of myeloid subtypes emerging after SMOi. Strikingly, we demonstrated that simultaneous SMOi and targeting myeloid cells via anti-Gr1 or CCR1 inhibition (CCR1i) significantly elevates cytotoxic T cell numbers up to 20x within the TME. We are currently investigating whether the activity of these T cells may be further potentiated through combination with immunomodulatory agents. By testing various treatment combination in the same TME, we will identify the best synergistic effects for future immunotherapy approaches in human PDAC. In summary, we are elucidating the complex mechanism behind LIS in PDAC by employing our novel explant culture system alongside in vivo studies. We aim to develop a translatable regimen to neutralize LIS, reactivating the cytotoxic T cells in the tumor periphery to invade, proliferate, and attack cancer cells. Citation Format: Marie C Hasselluhn, Lukas Vlahos, Dafydd Thomas, Quin T Waterbury, Alvaro Curiel Garcia, Amanda R Decker-Farrell, Tanner C Dalton, Stephen A Sastra, Carmine F Palermo, Timothy C Wang, Andrea Califano, Kenneth P Olive. Disrupting local immunosuppression through combination myCAF/myeloid targeting in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A045.

P106 ENZYMES INVOLVED IN THE CATABOLISM OF TRYPTOPHAN IN THE PROGNOSIS OF BLADDER CANCER

Bladder cancer (BC) ranks third among the most common carcinomas in Brazil. Tumor immunology opens up perspectives for treatment, where the enzyme indoleamine 2,3-dioxygenase-1 (IDO1) stands out for degrading the amino acid tryptophan, initiating a cascade that culminates in the generation of catabolites (derived from kynurenine). These catabolites, in different associations, determine degrees of local immunosuppression. These catabolites are generated by other enzymes that have not yet been studied in BC. We analyzed the expression of enzymes involved in tryptophan catabolism in order to correlate them with the stage, recurrence, and progression of BC. Analysis of published microarray databases was performed using the NCBI (National Center of Biotechnology Information) and accessed through GEO Datasets (Gene Expression Omnibus). The descriptor “Bladder cancer” was used. Series that provided analysis of normal bladder tissue (Normal) versus BC, including cases of non-muscle invasive BC (CBNMI) and muscle invasive BC (CBMI), recurrence and progression. Statistical analysis was performed using the GEO2R and ROC curve was used. The transcripts were: Tryptophan 2,3-dioxygenase (TDO2), Indoleamine 2,3-dioxygenase (IDO1), Kynurenine Formamidase (AFMID), Kynureninase (KNYU), Kynurenine-oxoglutarate-transaminase 1 (KYAT1), and Kynurenine Monooxygenase (KMU). GSE13507 series was selected, with ten Normal samples and 165 BC samples, including 104 CBNMI and 61 CBMI cases. CBNMI was predicted by KYNU, AFMID, and KYAT1 (AUC of 0.625, 0.589, and 0.638, respectively; p<0.05), while for CBMI were, IDO1, TDO2, and KMO (AUC of 0.662, 0.633, and 0.601, respectively; p<0.05). No enzyme was found to be important in predicting progression; however, the IDO1 enzyme was predictive of recurrence (AUC of 0.366; p<0.05). A relationship between the two IDO1/KYNU was established, which improved the prediction of stage for CBNMI (AUC of 0.733, p<0.05), non-progression (AUC of 0.619, p<0.05), and non-recurrence (AUC of 0.626, p<0.05). The expression of enzymes involved in tryptophan catabolism may aid in the prognosis of BC. It is likely that these enzymes are involved in tumor immunomodulation, which will be explored in future studies.

Impact of Skin Exposure to Benzo[a]pyrene in Rat Model: Insights into Epidermal Cell Function and Draining Lymph Node Cell Response.

The skin is a direct target of the air pollutant benzo[a]pyrene (BaP). While its carcinogenic qualities are well-studied, the immunotoxicity of BaP after dermal exposure is less understood. This study examines the immunomodulatory effects of a 10-day epicutaneous BaP application, in environmentally/occupationally relevant doses, by analyzing ex vivo skin immune response (skin explant, epidermal cells and draining lymph node/DLN cell activity), alongside the skin's reaction to sensitization with experimental hapten dinitrochlorobenzene (DNCB). The results show that BaP application disrupts the structure of the epidermal layer and promotes immune cell infiltration in the dermis. BaP exposure led to oxidative stress in epidermal cells, characterized by decreased reduced glutathione and increased AHR and Cyp1A1 expression. Production and gene expression of proinflammatory cytokines (TNF, IL-1β) by epidermal cells decreased, while IL-10 response increased. Decreased spontaneous production of IFN-γ and IL-17, along with unchanged IL-10, was observed in DLC cells, whereas ConA-stimulated production of these cytokines was elevated. Local immunosuppression caused by BaP application seems to reduce the skin's response to an additional stimulus, evidenced by decreased effector activity of DLN cells three days after sensitization with DNCB. These findings provide new insight into the immunomodulatory effects and health risks associated with skin exposure to BaP.

Cervical Cancer Evades the Host Immune System through the Inhibition of Type I Interferon and CXCL9 by LIF.

Cervical cancer is a viral-associated tumor caused by the infection with the human papilloma virus. Cervical cancer is an immunogenic cancer that expresses viral antigens. Despite being immunogenic, cervical cancer does not fully respond to immune checkpoint inhibitors (ICI). LIF is a crucial cytokine in embryo implantation, involved in maternal tolerance that acts as an immunomodulatory factor in cancer. LIF is expressed in cervical cancer and high levels of LIF is associated with poor prognosis in cervical cancer. We evaluated the impact of LIF on the immune response to ICI using primary plasmocytoid dendritic cells (pDC) and macrophage cultures, syngeneic animals and patient-derived models that recapitulate the human tumor microenvironment. We found that the viral proteins E6 and E7 induce the expression of LIF via the NFκB pathway. The secreted LIF can then repress type I interferon expressed in pDCs and CXCL9 expressed in tumor-associated macrophages. Blockade of LIF promotes the induction of type I interferon and CXCL9 inducing the tumor infiltration of CD8 T cells. This results in the sensitization of the tumor to ICI. Importantly, we observed that patients with cervical cancer expressing high levels of LIF tend to be resistant to ICI. Our data show that the HPV virus induces the expression of LIF to provide a selective advantage to the tumor cell by generating local immunosuppression via the repression of type I interferon and CXCL9. Combinatory treatment with blocking antibodies against LIF and ICI could be effective against cervical cancer expressing high levels of LIF.

A subpopulation of human bone marrow erythroid cells displays a myeloid gene expression signature similar to that of classic monocytes.

Erythroid cells, serving as progenitors and precursors to erythrocytes responsible for oxygen transport, were shown to exhibit an immunosuppressive and immunoregulatory phenotype. Previous investigations from our research group have revealed an antimicrobial gene expression profile within murine bone marrow erythroid cells which suggested a role for erythroid cells in innate immunity. In the present study, we focused on elucidating the characteristics of human bone marrow erythroid cells through comprehensive analyses, including NanoString gene expression profiling utilizing the Immune Response V2 panel, a BioPlex examination of chemokine and TGF-beta family proteins secretion, and analysis of publicly available single-cell RNA-seq data. Our findings demonstrate that an erythroid cell subpopulation manifests a myeloid-like gene expression signature comprised of antibacterial immunity and neutrophil chemotaxis genes which suggests an involvement of human erythroid cells in the innate immunity. Furthermore, we found that human erythroid cells secreted CCL22, CCL24, CXCL5, CXCL8, and MIF chemokines. The ability of human erythroid cells to express these chemokines might facilitate the restriction of immune cells in the bone marrow under normal conditions or contribute to the ability of erythroid cells to induce local immunosuppression by recruiting immune cells in their immediate vicinity in case of extramedullary hematopoiesis.

Endogenous stem cell mobilization and localized immunosuppression synergistically ameliorate DSS-induced Colitis in mice

BackgroundStem cell therapy is a promising alternative for inflammatory diseases and tissue injury treatment. Exogenous delivery of mesenchymal stem cells is associated with instant blood-mediated inflammatory reactions, mechanical stress during administration, and replicative senescence or change in phenotype during long-term culture in vitro. In this study, we aimed to mobilize endogenous hematopoietic stem cells (HSCs) using AMD-3100 and provide local immune suppression using FK506, an immunosuppressive drug, for the treatment of inflammatory bowel diseases.MethodsReactive oxygen species (ROS)-responsive FK506-loaded thioketal microspheres were prepared by emulsification solvent-evaporation method. Thioketal vehicle based FK506 microspheres and AMD3100 were co-administered into male C57BL6/J mice with dextran sulfate sodium (DSS) induced colitis. The effect of FK506-loaded thioketal microspheres in colitis mice were evaluated using disease severity index, myeloperoxidase activity, histology, flow cytometry, and gene expression by qRT-PCR.ResultsThe delivery of AMD-3100 enhanced mobilization of HSCs from the bone marrow into the inflamed colon of mice. Furthermore, targeted oral delivery of FK506 in an inflamed colon inhibited the immune activation in the colon. In the DSS-induced colitis mouse model, the combination of AMD-3100 and FK506-loaded thioketal microspheres ameliorated the disease, decreased immune cell infiltration and activation, and improved body weight, colon length, and epithelial healing process.ConclusionThis study shows that the significant increase in the percentage of mobilized hematopoietic stem cells in the combination therapy of AMD and oral FK506 microspheres may contribute to a synergistic therapeutic effect. Thus, low-dose local delivery of FK506 combined with AMD3100 could be a promising alternative treatment for inflammatory bowel diseases.

Tacrolimus-loaded Drug Delivery Systems in Vascularized Composite Allotransplantation: Lessons and Opportunities for Local Immunosuppression.

Long-term systemic immunosuppression is needed for vascularized composite allotransplantation (VCA). The high rate of acute rejection episodes in the first posttransplant year, the development of chronic rejection, and the adverse effects that come along with this treatment, currently prevent a wider clinical application of VCA. Opportunistic infections and metabolic disturbances are among the most observed side effects in VCA recipients. To overcome these challenges, local immunosuppression using biomaterial-based drug delivery systems (DDS) have been developed. The aim of these systems is to provide high local concentrations of immunosuppressive drugs while reducing their systemic load. This review provides a summary of recently investigated local DDS with different mechanisms of action such as on-demand, ultrasound-sensitive, or continuous drug delivery. In preclinical models, ranging from rodent to porcine and nonhuman primate models, this approach has been shown to reduce systemic tacrolimus (TAC) load and adverse effects, while prolonging graft survival. Localized immunosuppression using biomaterial-based DDS represents an encouraging approach to enhance graft survival and reduce toxic side effects of immunosuppressive drugs in VCA patients. Preclinical models using TAC-releasing DDS have demonstrated high local immunosuppressive effects with a low systemic burden. However, to reduce acute rejection events in translational animal models or in the clinical reality, the use of additional low-dose systemic TAC treatment may be envisaged. Patients may benefit through efficient graft immunosuppression and survival with negligible systemic adverse effects, resulting in better compliance and quality of life.

The Fate of RPE Cells Following hESC-RPE Patch Transplantation in Haemorrhagic Wet AMD: Pigmentation, Extension of Pigmentation, Thickness of Transplant, Assessment for Proliferation and Visual Function-A 5 Year-Follow Up.

(1) Background: We reviewed a stem cell-derived therapeutic strategy for advanced neovascular age-related macular degeneration (nAMD) using a human embryonic stem cell-derived retinal pigment epithelium (hESC-RPE) monolayer delivered on a coated, synthetic basement membrane (BM)-the patch-and assessed the presence and distribution of hESC-RPE over 5 years following transplantation, as well as functional outcomes. (2) Methods: Two subjects with acute vision loss due to sub-macular haemorrhage in advanced nAMD received the hESC-RPE patch. Systematic immunosuppression was used peri-operatively followed by local depot immunosuppression. The subjects were monitored for five years with observation of RPE patch pigmentation, extension beyond the patch boundary into surrounding retina, thickness of hESC-RPE and synthetic BM and review for migration and proliferation of hESC-RPE. Visual function was also assessed. (3) Results: The two study participants showed clear RPE characteristics of the patch, preservation of some retinal ultrastructure with signs of remodelling, fibrosis and thinning on optical coherence tomography over the 5-year period. For both participants, there was evidence of pigment extension beyond the patch continuing until 12 months post-operatively, which stabilised and was preserved until 5 years post-operatively. Measurement of hESC-RPE and BM thickness over time for both cases were consistent with predefined histological measurements of these two layers. There was no evidence of distant RPE migration or proliferation in either case beyond the monolayer. Sustained visual acuity improvement was apparent for 2 years in both subjects, with one subject maintaining the improvement for 5 years. Both subjects demonstrated initial improvement in fixation and microperimetry compared to baseline, at year 1, although only one maintained this at 4 years post-intervention. (4) Conclusions: hESC-RPE patches show evidence of continued pigmentation, with extension, to cover bare host basement membrane for up to 5 years post-implantation. There is evidence that this represents functional RPE on the patch and at the patch border where host RPE is absent. The measurements for thickness of hESC-RPE and BM suggest persistence of both layers at 5 years. No safety concerns were raised for the hypothetical risk of RPE migration, proliferation or tumour formation. Visual function also showed sustained improvement for 2 years in one subject and 5 years in the other subject.

Exosomal PD-L1 in cancer and other fields: recent advances and perspectives.

PD-1/PD-L1 signaling is a key factor of local immunosuppression in the tumor microenvironment. Immune checkpoint inhibitors targeting PD-1/PD-L1 signaling have achieved tremendous success in clinic. However, several types of cancer are particularly refractory to the anti-PD-1/PD-L1 treatment. Recently, a series of studies reported that IFN-γ can stimulate cancer cells to release exosomal PD-L1 (exoPD-L1), which possesses the ability to suppress anticancer immune responses and is associated with anti-PD-1 response. In this review, we introduce the PD-1/PD-L1 signaling, including the so-called 'reverse signaling'. Furthermore, we summarize the immune treatments of cancers and pay more attention to immune checkpoint inhibitors targeting PD-1/PD-L1 signaling. Additionally, we review the action mechanisms and regulation of exoPD-L1. We also introduce the function of exoPD-L1 as biomarkers. Finally, we review the methods for analyzing and quantifying exoPD-L1, the therapeutic strategies targeting exoPD-L1 to enhance immunotherapy and the roles of exoPD-L1 beyond cancer. This comprehensive review delves into recent advances of exoPD-L1 and all these findings suggest that exoPD-L1 plays an important role in both cancer and other fields.

Immunosuppressive microvesicles-mimetic derived from tolerant dendritic cells to target T-lymphocytes for inflammation diseases therapy

The utilization of extracellular vesicles (EV) in immunotherapy, aiming at suppressing peripheral immune cells responsible for inflammation, has demonstrated significant efficacy in treating various inflammatory diseases. However, the clinical application of EV has faced challenges due to their inadequate targeting ability. In addition, most of the circulating EV would be cleared by the liver, resulting in a short biological half-life after systemic administration. Inspired by the natural microvesicles (MV, as a subset of large size EV) are originated and shed from the plasma membrane, we developed the immunosuppressive MV-mimetic (MVM) from endotoxin tolerant dendritic cells (DC) by a straightforward and effective extrusion approach, in which DC surface proteins were inherited for providing the homing ability to the spleen, while αCD3 antibodies were conjugated to the MVM membranes for specific targeting of T cells. The engineered MVM carried a large number of bioactive cargos from the parental cells, which exhibited a remarkable ability to promote the induction of regulatory T cells (Treg) and polarization of anti-inflammatory M2 macrophages. Mechanistically, the elevated Treg level by MVM was mediated due to the upregulation of miR-155-3p. Furthermore, it was observed that systemic and local immunosuppression was induced by MVM in models of sepsis and rheumatoid arthritis through the improvement of Treg and M2 macrophages. These findings reveal a promising cell-free strategy for managing inflammatory responses to infections or tissue injury, thereby maintaining immune homeostasis.

A Roadmap of CAR-T-Cell Therapy in Glioblastoma: Challenges and Future Perspectives.

Glioblastoma (GBM) is the most common primary malignant brain tumor, with a median overall survival of less than 2 years and a nearly 100% mortality rate under standard therapy that consists of surgery followed by combined radiochemotherapy. Therefore, new therapeutic strategies are urgently needed. The success of chimeric antigen receptor (CAR) T cells in hematological cancers has prompted preclinical and clinical investigations into CAR-T-cell treatment for GBM. However, recent trials have not demonstrated any major success. Here, we delineate existing challenges impeding the effectiveness of CAR-T-cell therapy for GBM, encompassing the cold (immunosuppressive) microenvironment, tumor heterogeneity, T-cell exhaustion, local and systemic immunosuppression, and the immune privilege inherent to the central nervous system (CNS) parenchyma. Additionally, we deliberate on the progress made in developing next-generation CAR-T cells and novel innovative approaches, such as low-intensity pulsed focused ultrasound, aimed at surmounting current roadblocks in GBM CAR-T-cell therapy.

Adjuvant PD-1 Checkpoint Inhibition in Early Cutaneous Melanoma: Immunological Mode of Action and the Role of Ultraviolet Radiation.

Melanoma ranks as the fifth most common solid cancer in adults worldwide and is responsible for a significant proportion of skin-tumor-related deaths. The advent of immune checkpoint inhibition with anti-programmed death protein-1 (PD-1) antibodies has revolutionized the adjuvant treatment of high-risk, completely resected stage III/IV melanoma. However, not all patients benefit equally. Current strategies for improving outcomes involve adjuvant treatment in earlier disease stages (IIB/C) as well as perioperative treatment approaches. Interfering with T-cell exhaustion to counteract cancer immune evasion and the immunogenic nature of melanoma is key for anti-PD-1 effectiveness. Yet, the biological rationale for the efficacy of adjuvant treatment in clinically tumor-free patients remains to be fully elucidated. High-dose intermittent sun exposure (sunburn) is a well-known primary risk factor for melanomagenesis. Also, ultraviolet radiation (UVR)-induced immunosuppression may impair anti-cancer immune surveillance. In this review, we summarize the current knowledge about adjuvant anti-PD-1 blockade, including a characterization of the main cell types most likely responsible for its efficacy. In conclusion, we propose that local and systemic immunosuppression, to some extent UVR-mediated, can be restored by adjuvant anti-PD-1 therapy, consequently boosting anti-melanoma immune surveillance and the elimination of residual melanoma cell clones.