Impaired Antigen Presentation Research Articles

Leishmania donovani modulates host miRNAs regulating cholesterol biosynthesis for its survival

Cholesterol reduction by intracellular protozoan parasite Leishmania donovani (L. donovani), causative agent of leishmaniasis, impairs antigen presentation, pro-inflammatory cytokine secretion and host-protective membrane-receptor signaling in macrophages. Here, we studied the miRNA mediated regulation of cholesterol biosynthetic genes to understand the possible mechanism of L. donovani-induced cholesterol reduction and therapeutic importance of miRNAs in leishmaniasis. System-scale genome-wide microtranscriptome screening was performed to identify the miRNAs involved in the regulation of expression of key cholesterol biosynthesis regulatory genes through miRanda3.0. 11 miRNAs out of 2823, showing complementarity with cholesterol biosynthetic genes were finally selected for expression analysis. These selected miRNAs were differentially regulated in THP-1 derived macrophages and in primary human macrophages by L. donovani. Correlation of expression and target validation through luciferase assay suggested two key miRNAs, hsa-miR-1303 and hsa-miR-874-3p regulating the key genes hmgcr and hmgcs1 respectively. Inhibition of hsa-mir-1303 and hsa-miR-874-3p augmented the expression of targets and reduced the parasitemia in macrophages. This study will also provide the platform for the development of miRNA-based therapy against leishmaniasis.

Combination of MHI148 Targeted Photodynamic Therapy and STING Activation Inhibits Tumor Metastasis and Recurrence.

Metastasis and recurrence are notable contributors to mortality associated with breast cancer. Although immunotherapy has shown promise in mitigating these risks after conventional treatments, its effectiveness remains constrained by significant challenges, such as impaired antigen presentation by dendritic cells (DCs) and inadequate T cell infiltration into tumor tissues. To address these limitations, we developed a multifunctional nanoparticle platform, termed GM@P, which consisted of a hydrophobic shell encapsulating the photosensitizer MHI148 and a hydrophilic core containing the STING agonist 2'3'-cGAMP. This design elicited robust type I interferon responses to activate antitumor immunity. The GM@P nanoparticles loaded with MHI148 specifically targeted breast cancer cells. Upon exposure to 808 nm laser irradiation, the MHI148-loaded nanoparticles produced toxic reactive oxygen species (ROS) to eradicate tumor cells through photodynamic therapy (PDT). Notably, PDT stimulated immunogenic cell death (ICD) to foster the potency of antitumor immune responses. Furthermore, the superior photoacoustic imaging (PAI) capabilities of MHI148 enabled the simultaneous visualization of diagnostic and therapeutic procedures. Collectively, our findings uncovered that the combination of PDT and STING activation facilitated a more conducive immune microenvironment, characterized by enhanced DC maturation, infiltration of CD8+ T cells, and proinflammatory cytokine release. This strategy stimulated local immune responses to augment systemic antitumor effects, offering a promising approach to suppress tumor growth, inhibit metastasis, and prevent recurrence.

Targeting BCL9/BCL9L enhances antigen presentation by promoting conventional type 1 dendritic cell (cDC1) activation and tumor infiltration

Conventional type 1 dendritic cells (cDC1) are the essential antigen-presenting DC subset in antitumor immunity. Suppressing B-cell lymphoma 9 and B-cell lymphoma 9-like (BCL9/BCL9L) inhibits tumor growth and boosts immune responses against cancer. However, whether oncogenic BCL9/BCL9L impairs antigen presentation in tumors is still not completely understood. Here, we show that targeting BCL9/BCL9L enhanced antigen presentation by stimulating cDC1 activation and infiltration into tumor. Pharmacological inhibition of BCL9/BCL9L with a novel inhibitor hsBCL9z96 or Bcl9/Bcl9l knockout mice markedly delayed tumor growth and promoted antitumor CD8+ T cell responses. Mechanistically, targeting BCL9/BCL9L promoted antigen presentation in tumors. This is due to the increase of cDC1 activation and tumor infiltration by the XCL1-XCR1 axis. Importantly, using single-cell transcriptomics analysis, we found that Bcl9/Bcl9l deficient cDC1 were superior to wild-type (WT) cDC1 at activation and antigen presentation via NF-κB/IRF1 signaling. Together, we demonstrate that targeting BCL9/BCL9L plays a crucial role in cDC1-modulated antigen presentation of tumor-derived antigens, as well as CD8+ T cell activation and tumor infiltration. Targeting BCL9/BCL9L to regulate cDC1 function and directly orchestrate a positive feedback loop necessary for optimal antitumor immunity could serve as a potential strategy to counter immune suppression and enhance cancer immunotherapy.

Tumor-associated NK cells drive MDSC-mediated tumor immune tolerance through the IL-6/STAT3 axis.

Apart from their killer identity, natural killer (NK) cells have integral roles in shaping the tumor microenvironment. Through immune gene deconvolution, the present study revealed an interplay between NK cells and myeloid-derived suppressor cells (MDSCs) in nonresponders of immune checkpoint therapy. Given that the mechanisms governing the outcome of NK cell-to-myeloid cell interactions remain largely unknown, we sought to investigate the cross-talk between NK cells and suppressive myeloid cells. Upon contact with tumor-experienced NK cells, monocytes and neutrophils displayed increased expression of MDSC-related suppressive factors along with increased capacities to suppress T cells. These changes were accompanied by impaired antigen presentation by monocytes and increased ER stress response by neutrophils. In a cohort of patients with sarcoma and breast cancer, the production of interleukin-6 (IL-6) by tumor-infiltrating NK cells correlated with S100A8/9 and arginase-1 expression by MDSCs. At the same time, NK cell-derived IL-6 was associated with tumors with higher major histocompatibility complex class I expression, which we further validated with b2m-knockout (KO) tumor mice models. Similarly in syngeneic wild-type and IL-6 KO mouse models, we then demonstrated that the accumulation of MDSCs was influenced by the presence of such regulatory NK cells. Inhibition of the IL-6/signal transducer and activator of transcription 3 (STAT3) axis alleviated suppression of T cell responses, resulting in reduced tumor growth and metastatic dissemination. Together, these results characterize a critical NK cell-mediated mechanism that drives the development of MDSCs during tumor immune escape.

Human macrophages infected with Egyptian Rousette bat-isolated Marburg virus display inter-individual susceptibility and antiviral responsiveness

Marburg virus (MARV) is a highly pathogenic filovirus and a causative agent of sporadic zoonotic viral hemorrhagic fever outbreaks with high case fatality rates. In humans, filoviruses like MARV and Zaire Ebola virus (EBOV) target, among others, innate immune cells like dendritic cells and macrophages (MΦs). Filovirus-infected dendritic cells display impaired maturation and antigen presentation, while MΦs become hyper-activated and secrete proinflammatory cytokines and chemokines. Our current understanding of human macrophage responses to MARV remains limited. Here, we used human monocyte-derived macrophages (moMΦs) to address how their phenotype, transcriptional profile, and protein expression change upon an in vitro infection with a bat isolate of MARV. Confirming its tropism for macrophages, we show that MARV induces notable shifts in their transcription distinct from responses induced by lipopolysaccharide (LPS), marked by upregulated gene expression of several chemokines, type I interferons, and IFN-stimulated genes. MARV infection also elicited pronounced inter-individually different transcriptional programs in moMΦs, the induction of Wnt signaling-associated genes, and the downregulation of multiple biological processes and molecular pathways.

Preventing kidney failure in hemolytic anemias with a novel heme and hemoglobin scavenger

Chronic hemolytic conditions, seen in genetic and acquired hemolytic diseases, result in increased levels of circulatory hemoglobin (Hb) released from erythrocytes, free heme, and iron. Cell-free Hb's toxicity arises from the Hb dissociation into dimers that can traverse tissue barriers, impacting crucial compartments like arterial smooth muscle cells and renal glomeruli. Chronic hemolysis leads to vasoconstriction, renal injury, and activates pathways via Hb oxidation and heme-ligation of TLR4 on endothelial cells, as well as leukocytes, affecting adhesion pathways. Hb and heme exposure to the immune system causes phagocyte dysfunction, inflammation suppression, and impaired antigen presentation. Kidneys are particularly susceptible to Hb toxicity due to Hb and dimer penetration into glomeruli and tubules, disrupting filtration forces, elevating the glomerular filtration rate (GFR), and causing acute kidney injury (AKI). To combat this issue, our group developed a novel Hb and heme scavenger based on apohemoglobin-haptoglobin (apoHb-Hp), functioning as both a hemopexin (Hpx) and Hp mimetic to scavenge cell-free Hb and heme during hemolytic states. Our study utilized the dorsal window chamber model in Golden Syrian hamsters, employing FITC-Sinistrin for non-invasive GFR measurement, avoiding the need for blood and urine sampling throughout time to estimate GFR. Two experiments were conducted, volumetric exchange groups with 20% of the animal’s blood volume either with Hb at different concentrations or a combination ApoHb-Hp and Hb. Results showed a significant acute decrease of 35% in half-life of the Hb group at 1.2g/dL and 5g/dL after two hours, and further decrease is seen of 45% at 24hrs. In contrast, the ApoHb-Hp group experienced a 30% decrease at 2hrs but reverted back to baseline at 24hrs, signifying Hb's predominant effect on the acute decrease rather than the volume change. Furthermore, the ApoHb-Hp group demonstrated a quicker return to baseline compared to the Hb group after the 24-hour mark, suggesting prolonged Hb exposure's lasting impact on kidney filtration rate, potentially leading to irreversible damage. Further research is required to comprehend these findings fully and explore ApoHb-Hp's protective role in mitigating Hb-induced kidney damage. Title of the project: Bioengineering a Dual Function Protein Construct to Detoxify Heme and HemoglobinProject number: R01HL159862Title of the project: Bioengineering a novel therapeutic protein complex to minimize the effects of medical device induced hemolysis.Project number: R01HL162120. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 109: Identification of tumor-intrinsic and extrinsic mechanisms of T cell exclusion in estrogen receptor-positive (ER+) breast cancer through integration of bulk and single-cell transcriptome and spatial analysis

Abstract T cells are generally present in low numbers in estrogen receptor-positive (ER+) breast cancer (BC) potentially due to limited antigens or impaired antigen presentation, however defined mechanisms are not fully understood. This study explores the interplay between estrogen receptor signaling (ERS), antigen presentation machinery (APM), and T cell signaling (TC) using diverse datasets, including BC cell lines, TCGA, METABRIC, and clinical trial bulk transcriptomes, single-cell RNA-sequencing of ER+ primary BC, and single-cell spatial multi-plex cyclic immunofluorescence (CyCIF) of primary BC patients. Co-expression analysis of both computationally and manually curated genes in TCGA and METABRIC data unveiled distinct 29-gene ERS and 225-gene combined APM/TC modules with strong inverse correlations in ER+ BC. This inverse correlation was confirmed in independent cohorts. ERS and APM/TC modules were down- and up-regulated, respectively, in response to aromatase inhibitor therapy in ER+ BC patients from the ACOSOG Z1301B trial who demonstrated clinical benefit. Analysis of published single-cell RNA-sequencing data of 20 primary ER+ BC was performed to elucidate cell types that express individual genes in the modules. It confirmed that high ERS activity is associated with reduced T cells, and identified distinct cell types contributing to each module, showing higher ERS activity in tumor cells and higher APM/TC activity in stromal and immune cells. At the cell type level, we found distinct contributions and precise insights of individual genes. The prevalence of ERS activity in tumor cells prompted investigation to determine if the ERS and APM/TC modules were preserved as tumor-intrinsic signaling. Analysis of 54 BC cell lines indicated partially preserved module correlations in cancer cell lines and revealed that the APM/TC module is a contribution of both tumor-intrinsic (e.g., complement and type-I IFN-response), and tumor-extrinsic (e.g., lymphocyte activation) factors. CyCIF analysis of 29 ER+ BC samples was performed to bridge gene signatures with spatial biology and validated that higher ERS activity correlates with lower T cell infiltration. The spatial pattern of T cells varied from immune desert to stromally bound to fully inflamed tumors. Neighborhood analysis revealed increased frequency of CD8 T cells around a tumor cell is associated with elevated MHC I and II in broad immune cells and macrophage subsets, respectively. pTBK1+ macrophages inversely correlated with CD8 T cell frequency and MHC expression in neighboring cells, suggesting an immunosuppressive role. This study uncovers robust and tightly regulated ERS and APM/TC signaling in ER+ patients, unraveling molecular intricacies and offering potential insights for targeted therapeutic interventions. Citation Format: Kenichi Shimada, Yvonne X. Cui, Daniel Michaud, Kelly F. Zheng, Jonathan Goldberg, Ricardo Pastorello, Lukas Kania, Sandra S. McAllister, Sara M. Tolaney, Adrienne Waks, Rinath Jeselsohn, Peter K. Sorger, Judith Agudo, Jennifer L. Guerriero, Elizabeth A. Mittendorf. Identification of tumor-intrinsic and extrinsic mechanisms of T cell exclusion in estrogen receptor-positive (ER+) breast cancer through integration of bulk and single-cell transcriptome and spatial analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 109.

Abstract 6751: In situ vaccination with Flt3l gene modified CD103+ type 1 conventional dendritic cells synergizes with anti-PD-1 checkpoint blockade in murine models of non-small cell lung cancer

Abstract A major hurdle in treatment of Non-Small Cell Lung Cancer (NSCLC) with anti-PD-1 immune checkpoint blockade (ICB) therapy is a lack of response (primary resistance) and relapse after an initial response (acquired resistance). Recent studies reveal that responses to PD-1/PD-L1 blockade are associated with high tumor mutational burden (TMB), increased CD8+ T cell infiltration and high baseline PD-L1 expression within the tumor microenvironment (TME), while impaired tumor antigen presentation and the immunosuppressive TME have been associated with resistance to ICB. One approach to overcome anti-PD-1 resistance is to intratumorally vaccinate NSCLC tumors with gene modified conventional dendritic cells (cDC), specifically the type I conventional DC (cDC1) lineage. Recent studies have established that generation of an anti-tumor immune response driven by CD8+ T cells requires the cross presentation of tumor associated antigens and that cDC1s are the primary cross presenting APC subtype in vivo, which can license CD8+ T cells to initiate an adaptive anti-tumor immune response. In addition, previous studies have shown that intratumoral administration of the FMS-like tyrosine kinase 3 ligand (FLT3L) protein can expand endogenous CD103+ cDC1s in the TME and augment anti-tumor immune responses to ICB therapy. Here, we engineered murine CD103+ cDC1s to constitutively secrete soluble FLT3L (FLT3L_cDC1) and performed in situ vaccination studies on anti-PD1 resistant murine models of NSCLC with LKB1-deficiency and elevated TMB that better represents human disease. In situ vaccination with FLT3L_cDC1 enhances anti-tumor efficacy compared to non-modified cDC1 vaccination and synergizes with anti-PD-1 ICB to inhibit tumor growth. FLT3L_cDC1 + anti-PD-1 combination therapy induces significant activation and expansion T cells and cDC1s within the TME. Furthermore, combination therapy significantly increases DC progenitor numbers within the tumor draining lymph node, including DC progenitors that are committed to the cDC1 lineage. Our data suggests in situ vaccination with FLT3L_cDC1 may represent a promising strategy to potentiate the efficacy of ICB and improve outcomes for patients with primary resistance PD-1/PD-L1 monotherapy. Citation Format: Jensen W. Abascal, Raymond J. Lim, Ramin Salehi-Rad, Zhe Jing, Michael S. Oh, William P. Crosson, Bitta P. Kahangi, Edgar Perez Reyes, Camelia Dumitras, Diana Reyimjan, Jessie Zhu, Linh M. Tran, Manash Paul, Kostyantyn Krysan, Bin Liu, Steven M. Dubinett. In situ vaccination with Flt3l gene modified CD103+ type 1 conventional dendritic cells synergizes with anti-PD-1 checkpoint blockade in murine models of non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6751.

GENERATION OF TOLEROGENIC DENDRITIC CELLS UNDER THE PERSISTENT INFLAMMATION STIMULATION.

Immunosuppression, commonly accompanied by persistent inflammation, is a key feature in the later phase of sepsis. However, the pathophysiological mechanisms underlying this phenomenon remain unclear. Dendritic cells (DCs), specifically tolerogenic DCs (tolDCs), play a crucial role in this process by regulating immune responses through inducing T cell anergy and releasing anti-inflammatory cytokines. Nevertheless, the existing cell models are inadequate for investigating tolDCs during the immunosuppressive phase of sepsis. Therefore, this study aimed to develop a novel in vitro model to generate tolDCs under chronic inflammatory conditions. We have successfully generated tolDCs by exposing them to sublethal lipopolysaccharide (LPS) for 72 h while preserving cell viability. Considering that IL-10-induced tolDCs (IL-10-tolDCs) are well-established models, we compared the immunological tolerance between LPS-tolDCs and IL-10-tolDCs. Our findings indicated that both LPS-tolDCs and IL-10-tolDCs exhibited reduced expression of maturation markers, whereas their levels of inhibitory markers were elevated. Furthermore, the immunoregulatory activities of LPS-tolDCs and IL-10-tolDCs were found to be comparable. These dysfunctions include impaired antigen presenting capacity and suppression of T cell activation, proliferation, and differentiation. Notably, compared with IL-10-tolDCs, LPS-tolDCs showed a reduced response in maturation and cytokine production upon stimulation, indicating their potential as a better model for research. Overall, in comparison with IL-10-tolDCs, our data suggest that the immunological dysfunctions shown in LPS-tolDCs could more effectively elucidate the increased susceptibility to secondary infections during sepsis. Consequently, LPS-tolDCs have emerged as promising therapeutic targets for ameliorating the immunosuppressed state in septic patients.

LATS1/2 loss promote tumor immune evasion in endometrial cancer through downregulating MHC-I expression.

LATS1/2 are frequently mutated and down-regulated in endometrial cancer (EC), but the contributions of LATS1/2 in EC progression remains unclear. Impaired antigen presentation due to mutations or downregulation of the major histocompatibility complex class I (MHC-I) has been implicated in tumor immune evasion. Herein, we elucidate the oncogenic role that dysregulation of LATS1/2 in EC leads to immune evasion through the down-regulation of MHC-I. The mutation and expression as well as the clinical significance of LATS1/2 in EC was assessed in the TCGA cohort and our sample cohort. CRISPR-Cas9 was used to construct knockout cell lines of LATS1/2 in EC. Differentially expressed genes were analyzed by RNA-seq.The interaction between LATS1/2 and STAT1 was verified using co-immunoprecipitation and GST pull-down assays. Mass spectrometry, in vitro kinase assays, ChIP-qPCR, flow cytometry, immunohistochemistry, immunofluorescence and confocal microscopy were performed to investigate the regulation of LATS1/2 on MHC-I through interaction with and phosphorylate STAT1. The killing effect of activated PBMCs on EC cells were used to monitor anti-tumor activity. Here, we demonstrate that LATS1/2 are frequently mutated and down-regulated in EC. Moreover, LATS1/2 loss was found to be associated with a significant down-regulation of MHC-I, independently of the Hippo-YAP pathway. Instead, LATS1/2 were found to directly interact with and phosphorylate STAT1 at Ser727, a crucial transcription factor for MHC-I upregulation in response to interferon-gamma (IFN-γ) signaling, to promote STAT1 accumulating and moving into the nucleus to enhance the transcriptional activation of IRF1/NLRC5 on MHC-I. Additionally, the loss of LATS1/2 was observed to confer increased resistance of EC cells to immune cell-mediated killing and this resistance could be reversed by over-expression of MHC-I. Our findings indicate that dysregulation of LATS1/2 in EC leads to immune evasion through the down-regulation of MHC-I, leading to the suppression of infiltrating activated CD8 + T cells and highlight the importance of LATS1/2 in IFN-γ signaling-mediated tumor immune response, suggesting that LATS1/2 is a promising target for immune checkpoint blockade therapy in EC.

Predicting Immunotherapy Outcomes in Glioblastoma Patients through Machine Learning.

Glioblastoma is a highly aggressive cancer associated with a dismal prognosis, with a mere 5% of patients surviving beyond five years post diagnosis. Current therapeutic modalities encompass surgical intervention, radiotherapy, chemotherapy, and immune checkpoint inhibitors (ICBs). However, the efficacy of ICBs remains limited in glioblastoma patients, necessitating a proactive approach to anticipate treatment response and resistance. In this comprehensive study, we conducted a rigorous analysis involving two distinct glioblastoma patient cohorts subjected to PD-1 blockade treatments. Our investigation revealed that a significant portion (60%) of patients exhibit persistent disease progression despite ICB intervention. To elucidate the underpinnings of resistance, we characterized the immune profiles of glioblastoma patients with continued cancer progression following anti-PD1 therapy. These profiles revealed multifaceted defects, encompassing compromised macrophage, monocyte, and T follicular helper responses, impaired antigen presentation, aberrant regulatory T cell (Tregs) responses, and heightened expression of immunosuppressive molecules (TGFB, IL2RA, and CD276). Building upon these resistance profiles, we leveraged cutting-edge machine learning algorithms to develop predictive models and accompanying software. This innovative computational tool achieved remarkable success, accurately forecasting the progression status of 82.82% of the glioblastoma patients in our study following ICBs, based on their unique immune characteristics. In conclusion, our pioneering approach advocates for the personalization of immunotherapy in glioblastoma patients. By harnessing patient-specific attributes and computational predictions, we offer a promising avenue for the enhancement of clinical outcomes in the realm of immunotherapy. This paradigm shift towards tailored therapies underscores the potential to revolutionize the management of glioblastoma, opening new horizons for improved patient care.

Immune escape of head and neck cancer mediated by the impaired MHC-I antigen presentation pathway.

Tumor immune evasion is a hallmark of Head and Neck Cancers. The advent of immune checkpoint inhibitors (ICIs) in the first-line setting has transformed the management of these tumors. Unfortunately, the response rate of Head and Neck Squamous Cell Carcinomas (HNSCC) to ICIs is below 15%, regardless of the human papillomavirus (HPV) status, which might be partially related with impaired antigen presentation machinery (APM). Mechanistically, HNSCC cells are usually defective in the expression of MHC-I associated APM, while this transcriptional pathway is critical for the activation of tumor-killing effector T-cells. To specifically illuminate the phenomenon and seek for therapeutic strategies, this review summarizes the most recently identified role of genetic and functional dysregulation of the MHC-I pathway, specifically through changes at the genetic, epigenetic, post-transcriptional, and post-translational levels, which substantially contributes to HNSCC immune escape and ICI resistance. Several treatment modalities can be potentially exploited to restore APM signaling in tumors, which improves anti-tumor immunity through the activation of interferons, vaccines or rimantadine against HPV and the inhibition of EGFR, SHP-2, PI3K and MEK. Additionally, the combinatorial use of radiotherapy or cytotoxic agents with ICIs can synergize to potentiate APM signaling. Future directions would include further dissection of MHC-I related APM signaling in HNSCC and whether reversing this inhibition in combination with ICIs would elicit a more robust immune response leading to improved response rates in HNSCC. Therapeutic approaches to restore the MHC-I antigen presentation machinery in Head and Neck Cancer. (Red color texts represent the according strategies and the outcomes).

Bexmarilimab-induced macrophage activation leads to treatment benefit in solid tumors: The phase I/II first-in-human MATINS trial

Macrophage Clever-1 contributes to impaired antigen presentation and suppression of anti-tumor immunity. This first-in-human trial investigates the safety and tolerability of Clever-1 blockade with bexmarilimab in patients with treatment-refractory solid tumors and assesses preliminary anti-tumor efficacy, pharmacodynamics, and immunologic correlates. Bexmarilimab shows no dose-limiting toxicities in part I (n= 30) and no additional safety signals in part II (n= 108). Disease control (DC) rates of 25%-40% are observed in cutaneous melanoma, gastric, hepatocellular, estrogen receptor-positive breast, and biliary tract cancers. DC associates with improved survival in a landmark analysis and correlates with high pre-treatment intratumoral Clever-1 positivity and increasing on-treatment serum interferon γ (IFNγ) levels. Spatial transcriptomics profiling of DC and non-DC tumors demonstrates bexmarilimab-induced macrophage activation and stimulation of IFNγ and Tcell receptor signaling selectively in DC patients. These data suggest that bexmarilimab therapy is well tolerated and show that macrophage targeting can promote immune activation and tumor control in late-stage cancer.

PD-1 Expression By Dendritic Cells Is a Key Regulator of T-Cell Immunity in Cancer

Programmed cell death 1 (PD-1) is an inhibitory receptor expressed on a variety of immune cells and a therapeutic target of checkpoint immunotherapy for cancer patients. PD-1 engagement by PD-L1 or PD-L2 can inhibit activation and expansion of tumor antigen-specific T cells. PD-1 expression in tumor-associated macrophages (TAM) correlates with impaired phagocytosis and antigen presenting function and enhanced immunosuppression. PD-1 is also expressed in dendritic cells (DC) and has been shown to correlate negatively with the outcome of DC-based cancer vaccines. On DC, PD-1 can interact with PD-L1 in cis when both of these receptors are expressed on the same cell surface. DCs are the major antigen presenting cells for cross-presenting tumor antigens to T cells, and PD-L1 upregulation protects them from killing by cytotoxic T lymphocytes, yet dampens the antitumor responses. Very little is known about the specific role of PD-1 on DCs function. In the present study, we sought to understand the role of PD-1 on DCs in the context of cancer. We generated mice with conditional targeting of the Pdcd1 gene (encoding for PD-1) and crossed them with CD11cCre ( Pdcd1 f/fCD11cCre) or Clec9Cre ( Pdcd1 f/fClec9Cre) resulting in specific deletion of PD-1 in DC cells. We used the MC17 fibrosarcoma syngeneic mouse model to investigate the tentative implications of DC-specific PD-1 ablation on anti-tumor responses. We found larger tumor sizes and weights in tumor-bearing Pdcd1 f/fCD11cCre mice compared to their Pdcd1 wt/wtCD11cCre control counterparts. Pdcd1 f/fCD11cCre tumor-bearing mice had significantly higher fractions of DCs in spleens and tumor-draining lymph nodes (tdLNs) but only minimal differences in their activation state and checkpoint marker expression. Similarly, CD4 + and CD8 + T cells from spleens and tdLNs were comparable in percentages and activation states. However, characterization of the immune cells at the tumor site, indicated that Pdcd1 f/fCD11cCre tumor-bearing mice had a more immunosuppressive tumor microenvironment with increased fractions of CD11b +Ly6C hiLy6G -monocytic myeloid derived suppressor cells (M-MDSCs) compared to control tumor-bearing mice. Moreover, flow cytometric analysis revealed that intratumoral M-MDSC in Pdcd1 f/fCD11cCre mice displayed enhanced expression of PD-L1 and CD38, consistent with a more activated and more immunosuppressive phenotype. Characterization of tumor infiltrating CD4 + and CD8 + T cells also revealed vast differences in composition and activation states. Specifically, Pdcd1 f/fCD11cCre mice had larger populations of intratumoral CD3 + T cells, with a greater CD4/CD8 T cell ratio, and higher percentages of CD4 + T regulatory cells. Conversely, intratumoral CD8 + T cells were reduced, and were characterized by diminished expansion of CD8 + T effector cells, and a less activated phenotype as determined by lower expression of CD25, CD69, and GITR. In syngeneic tumor experiments with B16 melanoma expressing ovalbumin (B16-Ova), Pdcd1 f/fCD11cCre mice and Pdcd1 f/fClec9Cre mice similarly had greater tumor sizes and weights compared to their control counterparts. These results indicate that selective ablation of PD-1 in DCs cells confers a more immunosuppressive tumor microenvironment by promoting T regulatory cell expansion and diminishing CD8 + T effector cell activation compromising anti-tumor responses.

Global and cell type-specific immunological hallmarks of severe dengue progression identified via a systems immunology approach.

Severe dengue (SD) is a major cause of morbidity and mortality. To define dengue virus (DENV) target cells and immunological hallmarks of SD progression in children's blood, we integrated two single-cell approaches capturing cellular and viral elements: virus-inclusive single-cell RNA sequencing (viscRNA-Seq 2) and targeted proteomics with secretome analysis and functional assays. Beyond myeloid cells, in natural infection, B cells harbor replicating DENV capable of infecting permissive cells. Alterations in cell type abundance, gene and protein expression and secretion as well as cell-cell communications point towards increased immune cell migration and inflammation in SD progressors. Concurrently, antigen-presenting cells from SD progressors demonstrate intact uptake yet impaired interferon response and antigen processing and presentation signatures, which are partly modulated by DENV. Increased activation, regulation and exhaustion of effector responses and expansion of HLA-DR-expressing adaptive-like NK cells also characterize SD progressors. These findings reveal DENV target cells in human blood and provide insight into SD pathogenesis beyond antibody-mediated enhancement.

Immunobiology of cancer stem cells and their immunoevasion mechanisms.

Cancer stem cells (CSCs) defined as a small fraction of cells within malignancies have been isolated from tumors with different histological origins with stem related characteristics such as self-replicating potential, tumorigenesis, and therapy resistance. The dynamic communication between CSCs and tumor microenvironment particularly immune cells orchestrates their fate and plasticity as well as the patient outcome. According to recent evidence, it has been reported that they harness different immunological pathways to escape immunosurveillance and express aberrantly immunomodulatory agents or decreased levels of factors forming antigen presenting machinery (APM), subsequently followed by impaired antigen presentation and suppressed immune detection. As effective therapies are expected to be able to eradicate CSCs, mechanistic understanding of such interactions can provide insights into causes of therapy failure particularly in immunotherapy. Also, it can contribute to enhance the practical interventions against CSCs and their immunomodulatory features resulting in CSCs eradication and improving patient clinical outcome. The aim of this review is to explain the present knowledge regarding the immunobiology of CSCs and the immunoevasion mechanisms they use.

Clinical, Immunological, and Genetic Findings in Iranian Patients with MHC-II Deficiency: Confirmation of c.162delG RFXANK Founder Mutation in the Iranian Population.

Major histocompatibility complex class II (MHC-II) deficiency is a rare inborn error of immunity (IEI). Impaired antigen presentation to CD4 + T cells results in combined immunodeficiency (CID). Patients typically present with severe respiratory and gastrointestinal tract infections at early ages. Hematopoietic stem cell transplantation (HSCT) is the only curative therapy. We describe the clinical, immunologic, and genetic features of eighteen unrelated Iranian patients with MHC-II deficiency. Consanguinity was present in all affected families. The median age at the initial presentation was 5.5months (range 7days to 18years). The main symptoms included failure to thrive, persistent diarrhea, and pneumonia. Autoimmune and neurologic features were also documented in about one-third of the patients, respectively. Thirteen patients carried RFXANK gene mutations, two carried RFX5 gene mutations, and three carried a RFXAP gene mutation. Six patients shared the same RFXANK founder mutation (c.162delG); limited to the Iranian population and dated to approximately 1296years ago. Four of the patients underwent HSCT; three of them are alive. On the other hand, nine of the fourteen patients who did not undergo HSCT had a poor prognosis and died. MHC-II deficiency is not rare in Iran, with a high rate of consanguinity. It should be considered in the differential diagnosis of CID at any age. With the limited access to HSCT and its variable results in MHC-II deficiency, implementing genetic counseling and family planning for the affected families are mandatory. We are better determined to study the c.162delG RFXANK heterozygous mutation frequency in the Iranian population.

Tumor immune evasion throughloss of MHC class-I antigen presentation.

CD8 T cells recognize cancers when they detect antigenic peptides presented on a tumor's surface MHC-I molecules. Since MHC-I antigen presentation is not essential for cell growth or survival, many cancers inactivate this pathway, and thereby escape control by CD8 T cells. Such immune evasion allows cancers to progress and also become resistant to CD8 T- cell-based immunotherapies, such as checkpoint blockade. Here, we review recent findings about the various different mechanisms that cancers use to impair antigen presentation, the consequence of such changes, and, in some cases, the potential to reverse these defects.

Single-cell RNA sequencing uncovers heterogeneous transcriptional signatures in tumor-infiltrated dendritic cells in prostate cancer

Prostate cancer (PCa) is one of the two solid malignancies in which a higher T cell infiltration in the tumor microenvironment (TME) corresponds with a worse prognosis for the tumor. The inability of T cells to eliminate tumor cells despite an increase in their number reinforces the possibility of impaired antigen presentation. In this study, we investigated the TME at single-cell resolution to understand the molecular function and communication of dendritic cells (DCs) (as professional antigen-presenting cells). According to our data, tumor cells stimulate the migration of immature DCs to the tumor site by inducing inflammatory chemokines. Many signaling pathways such as TNF-α/NF-κB, IL2/STAT5, and E2F up-regulated after DCs enter the tumor location. In addition, some molecules such as GPR34 and SLCO2B1 decreased on the surface of DCs. The analysis of molecular and signaling alterations in DCs revealed some suppression mechanisms of tumors, such as removing mature DCs, reducing the DC's survival, inducing anergy or exhaustion in the effector T cells, and enhancing the differentiation of T cells to Th2 and Tregs. In addition, we investigated the cellular and molecular communication between DCs and macrophages in the tumor site and found three molecular pairs including CCR5/CCL5, CD52/SIGLEC10, and HLA-DPB1/TNFSF13B. These molecular pairs are involved in the migration of immature DCs to the TME and disrupt the antigen-presenting function of DCs. Furthermore, we presented new therapeutic targets by the construction of a gene co-expression network. These data increase our knowledge of the heterogeneity and the role of DCs in PCa TME.

Distinct mechanisms of dysfunctional antigen-presenting DCs and monocytes by single-cell sequencing in multiple myeloma.

Multiple myeloma (MM) is an incurable plasma cell malignancy with the hallmark of immunodeficiency, including dysfunction of T cells, NK cells, and APCs. Dysfunctional APCs have been reported to play a key role in promoting MM progression. However, the molecular mechanisms remain elusive. Here, single-cell transcriptome analysis of dendritic cells (DC) and monocytes from 10 MM patients and three healthy volunteers was performed. Both DCs and monocytes were divided into five distinct clusters, respectively. Among them, monocyte-derived DCs (mono-DC) were shown to develop from intermediate monocytes (IM) via trajectory analysis. Functional analysis showed that, compared with healthy controls, conventional DC2 (cDC2), mono-DC, and IM of MM patients exhibited impaired antigen processing and presentation capacity. Moreover, reduced regulon activity of interferon regulatory factor 1 (IRF1) was found in cDC2, mono-DC and IM of MM patients according to single-cell regulatory network inference and clustering (SCENIC) analysis, while the downstream mechanisms were distinct. Specifically in MM patients, cathepsin S (CTSS) was markedly downregulated in cDC2, major histocompatibility complex (MHC) class II transactivator (CIITA) was significantly decreased in IM, in addition both CTSS and CIITA were downregulated in mono-DC based on differentially expressed genes analysis. In vitro study validated that knockdown of Irf1 downregulated Ctss and Ciita respectively in mouse DC cell line DC2.4 and mouse monocyte/macrophage cell line RAW264.7, which ultimately inhibited proliferation of CD4+ T cells after being cocultured with DC2.4 or RAW264.7 cells. This current study unveils the distinct mechanisms of cDC2, IM, and mono-DC function impairment in MM, offering new insight into the pathogenesis of immunodeficiency.