Activation Of Dendritic Cells Research Articles

Abstract 6735: A systemically administered non-nucleotide STING agonist in albumin nanoformulation with potent antitumor activity and low toxicity

Abstract Background: Stimulator of interferon genes (STING), a dimeric transmembrane adapter protein, plays a pivotal role in regulating tumor immune microenvironment and has been explored as a therapeutic strategy against tumors. Most STING agonists were tested in clinical trials using intratumoral injection with limited systemic efficacy. Several systemically administered STING agonists have progressed to clinical trials, with their efficacy yet to be determined, while their systemic toxicity may limit their application. Our study aimed to develop a novel systemically administrated non-nucleotide STING agonist formulated within albumin nanoparticles, demonstrating potent antitumor activity and low systemic toxicity. Methods & Results: We synthesized a novel non-nucleotide small molecule STING dimer ZSA-51D. The acid form of ZSA-51D exhibited a high binding affinity (EC 50: 1.3 nM) to human STING. ZSA-51D demonstrated potent STING activation in THP1-BlueTM ISG cells, with an EC50 of 5.1 nM. To enable systemic delivery, ZSA-51D was encapsulated in albumin nanoparticles (nano ZSA-51D) with a particle size of 115 nm. Surprisingly, nano ZSA-51D exhibited a tenfold increase in STING activation (EC50 of 0.44 nM) compared to free ZSA-51D. Furthermore, nano ZSA-51D demonstrated sevenfold greater activation of bone marrow-derived dendritic cells (EC 50 of 3.5 nM), and eightfold enhanced repolarization of bone marrow-derived macrophages (EC 50 of 4.2 nM) from M2 to M1 compared to free ZSA-51D. The in-vivo anticancer efficacy was evaluated on MC-38 and KPC 6620 xenograft in C57BL/6 mice using intravenous administration of ZSA-51D (1 mg/kg) or nano ZSA-51D, in combination with PD-1 antibody (α-PD1, 100 μg) for 5 dose every 3 days. The results revealed that nano ZSA-51D exhibited superior anticancer effects compared to free ZSA-51D. Remarkedly, nano ZSA-51D, in combination with α-PD1, completely eradicated cancer in the MC-38 xenograft model. These cured mice completely rejected the rechallenge with MC38 cells 120 days after the initial tumor inoculation, indicating potent and long-term anticancer immune memory for anticancer activity. In addition, we performed the toxicity evaluation of nano ZSA-51D and free ZSA-51D at the same dose regime as the efficacy study. Nano ZSA-51D shows no hematological and liver toxicity. However, free ZSA-51D has a sever local inflammation after injections, whereas nano ZSA-51D has no local toxicity. Conclusion: In this study, we developed a novel systemically administered non-nucleotide sting agonist formulated within albumin nanoparticles. Nano ZSA-51, in combination with α-PD1, demonstrated superior anticancer efficacy and low toxicity, holding the potential for future clinical trials. The significantly enhanced STING activation and in vivo efficacy of nano ZSA-51D warrant further investigation into its underlying mechanisms. Citation Format: Jinsong Tao, Hongyi Zhao, Zhongwei Liu, Hanning Wen, Chengyi Li, Qiuxia Li, Miao He, Bo Wen, Wei Gao, Duxin Sun. A systemically administered non-nucleotide STING agonist in albumin nanoformulation with potent antitumor activity and low toxicity [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 6735.

Abstract 2599: Trastuzumab deruxtecan reprograms immune response by stimulating cGAS-STING pathway in gastric cancer cells

Abstract Background: The combination approach of antibody-drug conjugates and immune checkpoint inhibitors (ICIs) is being tested to enhance efficacy and overcome resistance in solid tumors. Although trastuzumab deruxtecan (T-DXd) is approved in HER2-positive gastric cancer (GC), its impact on the GC immune microenvironment remains elusive. This study explores how T-DXd modulates the immune response, providing essential insights on the development of T-DXd and immunotherapy combinations. Methods: Comet assays and immunofluorescence assessed DNA damage. RNA-seq delineated the impact of T-DXd on GC cells (NCI-N87 and SNU-484). In vivo complex of enzyme assays and immunostaining of topoisomerase 1-DNA covalent cleavage complex (TOP1cc) were used to determine the action mechanism of T-DXd. RT-qPCR, and ELISA measured cGAS-STING cytosolic dsDNA sensing pathway activation. Further validation included cGAS knockdown and 2’3’-cGAMP addition to confirm the role of cGAS in T-DXd-induced promotion of inflammatory response. In coculture experiments, we utilized THP-1-derived dendritic cells (TDDCs) and PBMCs from healthy donors to prove the effect of T-DXd on immune cells. FACS analyses probed the activation markers of TDDCs and CD8+ T cells. Results: T-DXd-induced DNA damage upregulated PD-L1 expression in GC cells in an IRF1-dependent manner. RNA-seq revealed gene signatures associated with increased antitumor immune response, including proinflammatory response and type-I IFN, following T-DXd treatment. Mechanistically, T-DXd co-localized TOP1cc with γ-H2AX-positive micronuclei, activating the cGAS-STING pathway and triggering a robust IFN-I response. Knockdown of cGAS in cancer cells abolished the T-DXd-induced IFN-I response. T-DXd enhanced HLA-DR and CD86 expression in TDDCs, particularly when cocultured with GC cells. The physical interaction between cancer cells and TDDCs, along with the presence of cGAS in cancer cells, was found to be essential for the upregulation of dendritic cell (DC) activation marker by T-DXd. Furthermore, T-DXd elevated CD69, Granzyme B, and IFNG expression in CD8+ T cells exclusively under coculture setting with NCI-N87. Conclusion: T-DXd exerts immunostimulatory effects in GC cells through the activation of the cGAS-STING pathway, leading to a reprogramming of DC and CD8+ T cell phenotypes. This provides a molecular rationale for combining T-DXd with ICIs for the treatment of patients with HER2-positive tumors. Citation Format: Kyoung-Seok Oh, Ah-Rong Nam, Ju-Hee Bang, Yoojin Jeong, Sea Young Choo, Hyo Jung Kim, Su In Lee, Jae-Min Kim, Jeesun Yoon, Tae-Yong Kim, Do-Youn Oh. Trastuzumab deruxtecan reprograms immune response by stimulating cGAS-STING pathway in gastric cancer cells [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 2599.

Abstract 1364: Characterization of CHS-1000, an Fc-modified anti-ILT4 monoclonal antibody for reprogramming suppressive myeloid cells in solid tumors

Abstract Tumor resident immunosuppressive myeloid cells play a significant role in inducing resistance to cancer immunotherapy and are associated with poor prognosis in several solid tumors. The leukocyte immunoglobulin-like receptor B2 (LILRB2), also known as ILT4, is primarily expressed on myeloid cells and is a negative regulator of myeloid cell activation. ILT4 and its primary ligands (HLA-G, HLA-A) are widely expressed in various solid tumors and are associated with disease progression, tumor metastasis and poor clinical outcome. Abrogating inhibitory signals mediated through ILT4 and its ligands is a promising strategy to reprogram suppressive myeloid cells and enhance efficacy of immune checkpoint therapies. Here we report IND-enabling preclinical characterization of CHS-1000, a novel recombinant humanized Fc-modified IgG1 antibody with attenuated Fc-gamma receptor (FcγR) binding, that specifically targets immunoglobulin-like transcript 4 (ILT4). CHS-1000 selectively binds to human ILT4 with high affinity and potently blocks ILT4 interactions with its primary ligands (HLA-A, HLA-G). Functionally, CHS-1000 promotes re-polarization of suppressive myeloid cells to a pro-inflammatory phenotype and enhances activation of dendritic cells and T cells. Combinations of CHS-1000 with PD-1 inhibitors or other immunotherapy agents hold particular promise for enhanced anti-tumor activity. These preclinical characterization data support clinical development of CHS-1000 in combination with toripalimab (a commercial stage anti-PD-1 antibody) as an immunotherapy treatment for patients with solid tumors. Citation Format: Narendiran Rajasekaran, Xiaoguang Wang, Scott Klakamp, Sruthi Ravindranathan, Suzanna Tatarewicz, Julia Du, Brian Nguyen, Kate Widmann, Nessa Hawkins, Daniel J. Chin, Theresa LaVallee, Varun Kapoor. Characterization of CHS-1000, an Fc-modified anti-ILT4 monoclonal antibody for reprogramming suppressive myeloid cells in solid tumors [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 1364.

Abstract 2966: Senescent cancer cell vaccines induce cytotoxic T cell responses targeting primary tumors and disseminated tumor cells

Abstract Diverse conventional cancer therapies, including genotoxic agents and cell cycle or cell division-targeting drugs, can trigger cancer cell senescence. Senescent cells (SnCs) induced by cancer therapy often display unrepaired chromosomal damage but persist indefinitely. SnCs remain metabolically active and display distinct alterations in transcription, secretion, and membrane protein expression/presentation. While initially viewed as a favorable outcome and likely tumor-suppressive mechanism, therapy-induced senescence has more recently been recognized as contributing to resistance, recurrence and metastasis. Adverse effects may be mediated in part by release of inflammatory mediators as components of the senescence-associated secretory phenotype (SASP). Recent studies have argued for reconsidering the potential benefits of therapy-induced senescence, especially in modulating the tumor microenvironment to potentiate anti-tumor immunity and potentiate immunotherapy. We have explored the potential for SnCs to serve as vaccines, building upon prior findings where we demonstrated the immunogenic senescence triggered by the combination of ionizing radiation (IR) and the PARP1/2 inhibitor veliparib. We find that SnCs are avidly taken up by type I conventional, CD11c+/CD103+ dendritic cells (DCs), promoting their activation and maturation. Thereby, SnCs promote DC function in T cell priming and stimulation. The capacity of SnCs to activate DCs depends on how the SnCs are formed, with our results confirming IR+veliparib induces high immunogenicity and implicating a distinct SASP in these effects. On the other hand, blocking STING signaling limits SnC activation of DCs. In vivo experiments with the murine tumor models CT26 and 4T1 suggest that both SnCs and SnC-pulsed DCs serve as effective anti-tumor vaccines. They not only confer protective immunity against tumor engraftment but also act as therapeutic vaccines, inhibiting both primary and disseminated tumor growth, and synergize with radiotherapy and immune checkpoint blockade. While direct administration of tumor-derived SnCs to patients may raise safety concerns, pulsing autologous DCs with tumor SnCs may help drive anti-tumor immune response, offering a promising path toward personalized cancer immunotherapy. Citation Format: Yue Liu, Joanna Pagacz, Donald J. Wolfgeher, Stephen J. Kron. Senescent cancer cell vaccines induce cytotoxic T cell responses targeting primary tumors and disseminated tumor cells [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 2966.

Abstract 2459: ADT-030 is a novel dual-acting RAS/β-catenin inhibitor that potentiates antitumor immunity

Abstract ADT-030 is a novel indene identified from a screening campaign and chemically optimized for drug-like properties that acts as a dual inhibitor of RAS and β-catenin signaling. Here we demonstrate that ADT-030 binds RAS with high affinity and inhibits MAPK/AKT signaling with high potency and selectivity in KRAS mutant cells to inhibit proliferation and induce apoptosis. ADT-030 also inhibits phosphodiesterase 10A (PDE10) that is overexpressed in cancer cells to selectively activate cyclic guanosine monophosphate (cGMP)/protein kinase G (PKG) signaling, which induces phosphorylation and degradation of the oncogenic pool of β-catenin. ADT-030 exhibited cytotoxicity to a variety of murine cancer cell lines in vitro, and inhibited tumor growth in syngeneic immunocompetent mice when orally administered. The antitumor activity of ADT-030 was diminished in mice that were immunodeficient or depleted of CD8+ T cells, indicating a critical role of the host immune system. In a metastatic breast tumor mouse model (4T1), ADT-030 not only delayed the growth of the primary tumor but also reduced tumor colonization in the lungs, leading to increased survival. CyTOF and scRNAseq analyses provided a plethora of information revealing the impact of ADT-030 on the immune milieu in the tumor microenvironment. Notably, ADT-030 selectively induced apoptosis of myeloid-derived suppressor cells (MDSCs) while CD8+ T cells were not affected. Consequently, ADT-030 treatment resulted in a marked reduction of MDSCs, which was accompanied by increased tumor infiltration of effector CD8+ T cells. ADT-030 treatment of cancer cells led to immunogenic cell death and enhanced dendritic cell activation. Furthermore, the ability of ADT-030 to augment the antitumor activity of immune checkpoint blockade therapy underlines its potential as a desirable immune-potentiating agent to improve the outcome of cancer immunotherapy. ADT-030 represents a development candidate ready for IND-enabling safety assessment. Citation Format: Md Yeashin Gazi, Ogacheko Okoko, Xin Wang, Caitlin Brandle, Xi Chen, Khalda Fadlalla, Adam Keeton, Yulia Maxuitenko, Catherine Hedrick, Huidong Shi, Gary Piazza, Gang Zhou. ADT-030 is a novel dual-acting RAS/β-catenin inhibitor that potentiates antitumor immunity [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 2459.

Abstract 97: A E2F6 ceRNA network suppresses dendritic cell function, via PBX1/IL-10 signaling, in ovarian cancer

Abstract It is reported that long-term use of estrogen could increase the risk of ovarian cancer. However, the role of estrogen in immunoevasion is not fully explored. We have previously demonstrated that estrogen-mediated upregulation of E2F6, and, c-Kit, by epigenetic silencing of miR-193a, and a competing endogenous (ceRNA) mechanism. In this study, we found that PBX1, a transcriptional activator of the immunosuppressive cytokine, IL-10, is also a target of miR-193a. Importantly, overexpression of the E2F6 3’UTR upregulates both E2F6 and, PBX1, as well as IL10 in ovarian cancer cell lines, suggesting that ceRNA mechanism exists between E2F6 and PBX1. These phenomena are further supported by our stochastic simulation of the estrogen-mediated E2F6 ceRNA network on the distribution of E2F6 and PBX1 mRNA in cancer cells, which is consistent with the TCGA ovarian cancer RNA-Seq dataset. Importantly, monocyte-derived dendritic cell activation of T-cell function was inhibited by pretreatment of conditioned media derived from ovarian cancer cells overexpressing E2F6 3’UTR; such inhibition was rescueable by an anti-IL-10 antibody. Clinically, IL10 level was higher in ovarian cancer patients with higher E2F6 and PBX1, and in ovarian cancer cell lines overexpressed with E2F6 3’UTR. Taken together, these results showed that E2F6 could suppress anti-tumor immune response of dendritic cell, E2F6 ceRNA network. Epigenetic intervention in restoring the expression of miR-193a may be able to enhance anti-tumor immune response against ovarian cancer. Citation Format: Michael W.Y. Chan, Yin-Chen Chen, Ching-Wen Lin, Frank Cheng, Ching-Cher Sanders Yan, Chao-Ping Hsu, Yu-Min Chuang, Jie-Ting Low, Xiaojing Ma, Yao-Ting Huang, Chia-Bin Chang, Chin Li, Hung-Cheng Lai, Shu-Fen Wu, Shih-Hsun Hung, Je-Chiang Tsai. A E2F6 ceRNA network suppresses dendritic cell function, via PBX1/IL-10 signaling, in ovarian 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 97.

Abstract 481: Proteomimetic polymers limit tumor growth in multiple models via potent antigen-specific T cell activation

Abstract Background: Patient-specific cancer vaccines derived from tumor antigen have been explored as a promising therapeutic strategy, however, challenges delivering vaccine components in a coordinated fashion to elicit antitumor responses remain. To overcome these, we utilize a novel nanoplatform called the Protein-Like Polymer (PLP), which allows for sustained and targeted delivery of tumor antigens in conjunction with adjuvants. Methods: PLPs containing peptide antigens were synthesized via ring-opening metathesis polymerization (ROMP) and characterized. A library of compounds with different sidechain linkage chemistries, degrees of polymerization (DP), and inclusion/exclusion of Oligo-ethylene glycol (OEG) were made to determine design rules for immune activation. Cell uptake and functional assays using payload-specific T Cells were conducted. Immunization in three independent tumor models was done to show generalizability. Ability of PLPs to co-deliver adjuvants was tested by electrostatically coupling small molecule STING agonist, 2’3’ cGAMP. Results: Conjugating peptide antigens to the polymer via a cleavable disulfide linkage, which reduces intracellularly in APCs, resulted in increased endosomal localization, higher levels of induced T cell proliferation, cytokine production, and expression of activation markers in CTLs and APCs. Incorporating a diluent amount of OEG side chains reduced enzymatic degradation while increasing immunogenicity and uptake. Additionally, increasing the DP, and therefore the density of antigen side chains, further improved vaccine efficacy and resistance to proteolysis. Antigen-PLP conjugates enhanced dendritic cell activation and T-cell response only when paired with cells from their cognate system, with no activity in immune cells not expressing receptors for the payload demonstrating antigen-specificity. Mice bearing established B16F10, MC38, or TC-1 tumors treated with PLPs containing gp100, adpgk, or E7 respectively all showed increased survival times, reduced tumor burden with corresponding changes in immune cell profiles, and immunological memory upon rechallenge. Impressively, mice treated with STING-PLP complexes had significantly smaller tumors vs control at day 14 (0.038g vs 0.76g; p <0.0001) and allowed for subcutaneous administration of 2’3’ cGAMP, which traditionally requires intratumoral injection. Studies on the effects of vaccinating with pools of neoantigens multiplexed onto one PLP are ongoing. Conclusion: This work validates the ability of PLPs to overcome major limitations in cancer vaccine development. The modularity of the platform allows for complex nano-architectures including systems capable of delivering challenging compounds, ie small molecule STING agonists, subcutaneously through electrostatic coupling, highlighting its potential to revolutionize cancer vaccinology. Citation Format: Max Wang, Miran Choi, Bin Zhang, Nathan Gianneschi. Proteomimetic polymers limit tumor growth in multiple models via potent antigen-specific T cell activation [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 481.

Abstract 4252: Evaluation of NXI-101 in non-small cell lung cancer (NSCLC) using patient-derived organotypic tumor spheroids (PDOTS): Targeting a novel tumorigenic pathway

Abstract ONCOKINE-1 (OK-1), a soluble protein that is released by tumor cells, is implicated in the resistance of immunotherapy. Here we evaluated the efficacy of NXI-101, a monoclonal antibody that binds and neutralizes OK-1, in a model of patient-derived organotypic tumor spheroids (PDOTS) derived from non-small cell lung carcinoma (NSCLC) patients. NSCLC PDOTS were treated with NXI-101 and the effects of the drug were evaluated on day 3 post-treatment. Cytotoxicity was measured by Hoechst/Propidium Iodide staining and quantified by measuring the area-weighted percentage of cell death. Significant cytotoxicity was observed in 2 out of 10 patient tumors (20%) as a monotherapy and 4 out of 10 (40%) when used in combination with atezolizumab. The therapeutic response for both treatments was positively correlated (p<0.05) with the level of OK-1 expression, determined by immunohistochemistry using tissue sections taken on the day of tumor specimen collection. NXI-101 alone and in combination with atezolizumab had immunomodulatory effects that were associated with cytotoxic response. For example, transcriptomic analysis, Nanostring IO 360 Panel, indicated that the drug treatments resulted in the inhibition of TGF-β pathways and stimulated a number of immune pathways in 3 of the 4 responsive PDOTS. These pathways include interferon-α, JAK-STAT, and dendritic cell activation markers. We also observed evidence of activation of these same pathways in 2 of the 6 non-responsive PDOTS. These data support the prospects of NXI-101 as a first-in-class therapeutic for the treatment of NSCLC when used alone or in combination with an immune checkpoint inhibitor. Moreover, the findings highlight the PDOTS platform's substantial value in capturing patient-specific tumor response variability and providing robust support for the mechanistic evaluation of cancer therapeutics. Citation Format: Julia Vail, Hyung Gun Maeng, Je Woong Jang, Kyungjin Boo, Je-In Youn, Alyssa Martin, Aaron Goldman, Kyoung Wan Yoon, Michael A. Perricone. Evaluation of NXI-101 in non-small cell lung cancer (NSCLC) using patient-derived organotypic tumor spheroids (PDOTS): Targeting a novel tumorigenic pathway [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 4252.

Changes in the innate immune response to SARS-CoV-2 with advancing age in humans

BackgroundAdvancing age is a major risk factor for respiratory viral infections. The infections are often prolonged and difficult to resolve resulting hospitalizations and mortality. The recent COVID-19 pandemic has highlighted this as elderly subjects have emerged as vulnerable populations that display increased susceptibility and severity to SARS-CoV-2. There is an urgent need to identify the probable mechanisms underlying this to protect against future outbreaks of such nature. Innate immunity is the first line of defense against viruses and its decline impacts downstream immune responses. This is because dendritic cells (DCs) and macrophages are key cellular elements of the innate immune system that can sense and respond to viruses by producing inflammatory mediators and priming CD4 and CD8 T-cell responses.ResultsWe investigated the changes in innate immune responses to SARS-CoV-2 as a function of age. Our results using human PBMCs from aged, middle-aged, and young subjects indicate that the activation of DCs and monocytes in response to SARS-CoV-2 is compromised with age. The impairment is most apparent in pDCs where both aged and middle-aged display reduced responses. The secretion of IL-29 that confers protection against respiratory viruses is also decreased in both aged and middle-aged subjects. In contrast, inflammatory mediators associated with severe COVID-19 including CXCL-8, TREM-1 are increased with age. This is also apparent in the gene expression data where pathways related host defense display an age dependent decrease with a concomitant increase in inflammatory pathways. Not only are the inflammatory pathways and mediators increased after stimulation with SARS-CoV-2 but also at homeostasis. In keeping with reduced DC activation, the induction of cytotoxic CD8 T cells is also impaired in aged subjects. However, the CD8 T cells from aged subjects display increased baseline activation in accordance with the enhanced baseline inflammation.ConclusionsOur results demonstrate a decline in protective anti-viral immune responses and increase in damaging inflammatory responses with age indicating that dysregulated innate immune responses play a significant role in the increased susceptibility of aged subjects to COVID-19. Furthermore, the dysregulation in immune responses develops early on as middle-aged demonstrate several of these changes.Graphical abstract

Helicobacter pylori glycan biosynthesis modulates host immune cell recognition and response.

The pathogenic bacterium Helicobacter pylori has evolved glycan-mediated mechanisms to evade host immune defenses. This study tests the hypothesis that genetic disruption of H. pylori glycan biosynthesis alters immune recognition and response by human gastric epithelial cells and monocyte-derived dendritic cells. To test this hypothesis, human cell lines were challenged with wildtype H. pylori alongside an array of H. pylori glycosylation mutants. The relative levels of immune response were measured via immature dendritic cell maturation and cytokine secretion. Our findings indicate that disruption of lipopolysaccharide biosynthesis diminishes gastric cytokine production, without disrupting dendritic cell recognition and activation. In contrast, variable immune responses were observed in protein glycosylation mutants which prompted us to test the hypothesis that phase variation plays a role in regulating bacterial cell surface glycosylation and subsequent immune recognition. Lewis antigen presentation does not correlate with extent of immune response, while the extent of lipopolysaccharide O-antigen elaboration does. The outcomes of this study demonstrate that H. pylori glycans modulate the host immune response. This work provides a foundation to pursue immune-based tailoring of bacterial glycans towards modulating immunogenicity of microbial pathogens.

Lipoteichoic acids influence cell shape and bacterial division of Streptococcus suis serotype 2, but play a limited role in the pathogenesis of the infection

Streptococcus suis serotype 2 is a major swine pathogen and a zoonotic agent, causing meningitis in both swine and humans, responsible for substantial economic losses to the swine industry worldwide. The pathogenesis of infection and the role of bacterial cell wall components in virulence have not been fully elucidated. Lipoproteins, peptidoglycan, as well as lipoteichoic acids (LTA) have all been proposed to contribute to virulence. In the present study, the role of the LTA in the pathogenesis of the infection was evaluated through the characterisation of a mutant of the S. suis serotype 2 strain P1/7 lacking the LtaS enzyme, which mediates the polymerization of the LTA poly-glycerolphosphate chain. The ltaS mutant was confirmed to completely lack LTA and displayed significant morphological defects. Although the bacterial growth of this mutant was not affected, further results showed that LTA is involved in maintaining S. suis bacterial fitness. However, its role in the pathogenesis of the infection appears limited. Indeed, LTA presence reduces self-agglutination, biofilm formation and even dendritic cell activation, which are important aspects of the pathogenesis of the infection caused by S. suis. In addition, it does not seem to play a critical role in virulence using a systemic mouse model of infection.

Biobran/MGN-3, an Arabinoxylan Rice Bran, Exerts Anti-COVID-19 Effects and Boosts Immunity in Human Subjects.

Corona Virus Disease 19 (COVID-19) has been a major pandemic impacting a huge population worldwide, and it continues to present serious health threats, necessitating the development of novel protective nutraceuticals. Biobran/MGN-3, an arabinoxylan rice bran, is a potent immunomodulator for both humans and animals that has recently been demonstrated to protect against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro. We here investigate Biobran/MGN-3's potential to enhance an antiviral immune response in humans. Peripheral blood mononuclear cells (PBMCs) derived from eight subjects taking Biobran/MGN-3 (age 55-65 years) and eight age-matched control subjects were stimulated with irradiated SARS-CoV-2 virus and then subjected to immuno-phenotyping and multiplex cytokine/chemokine assays. Results showed that PBMCs from subjects supplemented with Biobran/MGN-3 had significantly increased activation of plasmacytoid dendritic cells (pDCs) coupled with increased IFN-α secretion. We also observed higher baseline expression of HLA-DR (human leukocyte antigen-DR isotype) on dendritic cells (DCs) and increased secretion of chemokines and cytokines, as well as a substantial increase in cytotoxic T cell generation for subjects taking Biobran/MGN-3. Our results suggest that Biobran/MGN-3 primes immunity and therefore may be used for boosting immune responses against SARS-CoV-2 infections and other diseases, particularly in high-risk populations such as the elderly.

Modern Methods of Obtaining Immune Dendritic Cells With Anti-Tumor Potential

Dendritic cells (DCs) initiate and shape both innate and adaptive immune responses. They specialize in presenting antigens to naïve T cells, thereby directing T cell immune responses and contributing significantly to the maintenance of antitumor immunity. In both human and animal bodies, these cells are present in limited quantities, posing challenges in their procurement. Hence, the quest for obtaining DCs with antitumor properties in vitro from progenitor cells for clinical or experimental use remains pertinent. This research aims to consolidate existing studies on deriving immune DCs from progenitor cells for application in anticancer therapy. Analysis of published reports reveals that monocytes from peripheral blood, mononuclear cells from bone marrow, and cord blood can serve as precursor cells of immune DCs. Protocols for generating immature DCs from progenitor cells involve the addition of various combinations of cytokines to the culture, including granulocyte-macrophage colony-stimulating factor, interleukin-4, and other cytokines. The extensive range of cytokines and conditions influencing the differentiation and functional activity of DCs results in considerable heterogeneity in the phenotypic and functional characteristics of these cells. Sources of tumor antigen for DC-based vaccines encompass tumor lysates, indivi­dual tumor proteins, peptides, and tumor cells in a state of immunogenic apoptosis. This paper delves into the use of maturation factors and cryopreservation as integral stages in obtaining immune DCs. A comprehensive understanding of the parameters involved in obtaining immune DCs is imperative for the development of DC-based vaccines to unleash their full antitumor potential.

Overcoming neutrophil-induced immunosuppression in postoperative cancer therapy: Combined sialic acid-modified liposomes with scaffold-based vaccines

Immunotherapy is a promising approach for preventing postoperative tumor recurrence and metastasis. However, inflammatory neutrophils, recruited to the postoperative tumor site, have been shown to exacerbate tumor regeneration and limit the efficacy of cancer vaccines. Consequently, addressing postoperative immunosuppression caused by neutrophils is crucial for improving treatment outcomes. This study presents a combined chemoimmunotherapeutic strategy that employs a biocompatible macroporous scaffold-based cancer vaccine (S-CV) and a sialic acid (SA)-modified, doxorubicin (DOX)-loaded liposomal platform (DOX@SAL). The S-CV contains whole tumor lysates as antigens and imiquimod (R837, Toll-like receptor 7 activator)-loaded PLGA nanoparticles as immune adjuvants for cancer, which enhance dendritic cell activation and cytotoxic T cell proliferation upon localized implantation. When administered intravenously, DOX@SAL specifically targets and delivers drugs to activated neutrophils in vivo, mitigating neutrophil infiltration and suppressing postoperative inflammatory responses. In vivo and vitro experiments have demonstrated that S-CV plus DOX@SAL, a combined chemo-immunotherapeutic strategy, has a remarkable potential to inhibit postoperative local tumor recurrence and distant tumor progression, with minimal systemic toxicity, providing a new concept for postoperative treatment of tumors.

Curcumin and Berberine Arrest Maturation and Activation of Dendritic Cells Derived from Lupus Erythematosus Patients.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease recognized by elevated activity of autoimmune cells, loss of tolerance, and decreased regulatory T cells producing inhibitory cytokines. Despite many efforts, the definitive treatment for lupus has not been fully understood. Curcumin (CUR) and berberine (BBR) have significant immunomodulatory roles and anti-inflammatory properties that have been demonstrated in various studies. This study aimed to investigate the anti-inflammatory properties of CUR and BBR on human monocyte-derived dendritic cells (DCs) with an special focus on the maturation and activation of DCs. Human monocytes were isolated from the heparinized blood of SLE patients and healthy individuals, which were then exposed to cytokines (IL-4 and GM-CSF) for five days to produce immature DCs. Then, the obtained DCs were characterized by FITC-uptake assay and then cultured in the presence of CUR, BBR, or lipopolysaccharide (LPS) for 48 h. Finally, the maturation of DCs was analyzed by the level of maturation using flow cytometry or real-time PCR methods. The results showed promising anti-inflammatory effects of CUR and BBR in comparison with LPS, supported by a significant reduction of not only co-stimulatory and antigen-presenting factors such as CD80, CD86, CD83, CD1a, CD14, and HLA-DR but also inflammatory cytokines such as IL-12. CUR and BBR could arrest DC maturation and develop a tolerogenic DC phenotype that subsequently promoted the expression of inhibitory cytokines and reduced the secretion of proinflammatory markers.

Impact of Nanoheater Subcellular Localization on the Antitumor Immune Efficacy of Magnetic Hyperthermia

Magnetic nanoparticles (MNPs)-mediated magnetic hyperthermia therapy is garnering attention as a promising modality for tumor remission. This therapy combines localized nano-scale heating and reactive oxygen species-related immunologic effects. A critical aspect in optimizing this therapy involves understanding how nanoheaters’ subcellular localizations influence the therapeutic efficacy and immunological effects of magnetic hyperthermia. Herein, we investigate these effects by focusing on the strategic localization of ferrimagnetic vortex-domain iron oxide (FVIO) nanorings within lysosomes and the cytoplasm of hepatocellular carcinoma (HCC) cells. The comparative analysis revealed that lysosomal-localized magnetic hyperthermia markedly increased apoptosis in Hepa1–6 cells and effectively induced immunogenic cell death (ICD). Mechanistically, lysosomal-localized magnetic hyperthermia led to Bid-associated apoptosis and caspase-1-dependent interleukin-1β (IL-1β) secretion, likely due to enhanced lysosomal membrane permeabilization. Moreover, it reduced calreticulin degradation and increased its presence at the cell membrane, thereby boosting ICD efficiency. In contrast, cytoplasmic-localized magnetic hyperthermia showed comparatively lesser impact than its lysosomal-localized counterpart. In vivo experiments reinforced the exceptional efficacy of lysosomal-localized magnetic hyperthermia, which notably enhanced dendritic cells activation and T cells infiltration in both tumors and lymph nodes, thereby triggering a robust antitumor immune response. Additionally, lysosomal-localized magnetic hyperthermia improved tumor vaccine efficacy and demonstrated the potential for sustaining long-term immune responses in antitumor treatments. This study highlights the crucial role of subcellular positioning of magnetic nanoheaters in determining the efficacy of antitumor immunological responses in magnetic hyperthermia therapy, offering valuable insights for future cancer treatment strategies.

Evaluation of different types of adjuvants in a malaria transmission-blocking vaccine

Adjuvants are critical components for vaccines, which enhance the strength and longevity of the antibody response and influence the types of immune response. Limited research has been conducted on the immunogenicity and protective efficacy of various adjuvants in malaria transmission-blocking vaccines (TBVs). In this study, we formulated a promising TBV candidate antigen, the P. berghei ookinete surface antigen PSOP25, with different types of adjuvants, including the TLR4 agonist monophosphoryl lipid A (MPLA), the TLR9 agonist cytosine phosphoguanosine oligodeoxynucleotides (CpG ODN 1826) (CpG), a saponin adjuvant QS-21, aluminum hydroxide (Alum), and two combination adjuvants MPLA + QS-21 and QS-21 + CpG. We demonstrated that adjuvanted vaccines results in elevated elicited antibody levels, increased proliferation of plasma cells, and efficient formation of germinal centers (GCs), leading to enhanced long-term protective immune responses. Furthermore, CpG group exhibited the most potent inhibition of ookinete formation and transmission-blocking activity. We found that the rPSOP25 with CpG adjuvant was more effective than MPLA, QS-21, MPLA + QS-21, QS-21 + CpG adjuvants in dendritic cells (DCs) activation and differentiation. Additionally, the CpG adjuvant elicited more rubust immune memory response than Alum adjuvant. CpG and QS-21 adjuvants could activate the Th1 response and promote the secretion of IFN-γ and TNF-α. PSOP25 induced a higher number of Tfh cells in splenocytes when combined with MPLA, CpG, and QS-21 + CpG; and there was no increase in these cell populations when PSOP25 was administered with Alum. In conclusion, CpG may confer enhanced efficacy for the rPSOP25 vaccine, as evidenced by the ability of the elicited antisera to induce protective immune responses and improved transmission-blocking activity.

Protein-assisted synthesis of chitosan-coated minicells enhance dendritic cell recruitment for therapeutic immunomodulation within pulmonary tumors

The efficacy of cancer therapies is significantly compromised by the immunosuppressive tumor milieu. Herein, we introduce a previously unidentified therapeutic strategy that harnesses the synergistic potential of chitosan-coated bacterial vesicles and a targeted chemotherapeutic agent to activate dendritic cells, thereby reshaping the immunosuppressive milieu for enhanced cancer therapy. Our study focuses on the protein-mediated modification of bacterium-derived minicells with chitosan molecules, facilitating the precise delivery of Doxorubicin to tumor sites guided by folate-mediated homing cues. These engineered minicells demonstrate remarkable specificity in targeting lung carcinomas, triggering immunogenic cell death and releasing tumor antigens and damage-associated molecular patterns, including calreticulin and high mobility group box 1. Additionally, the chitosan coating, coupled with bacterial DNA from the minicells, initiates the generation of reactive oxygen species and mitochondrial DNA release. These orchestrated events culminate in dendritic cell maturation via activation of the stimulator of interferon genes signaling pathway, resulting in the recruitment of CD4+ and CD8+ cytotoxic T cells and the secretion of interferon-β, interferon-γ, and interleukin-12. Consequently, this integrated approach disrupts the immunosuppressive tumor microenvironment, impeding tumor progression. By leveraging bacterial vesicles as potent dendritic cell activators, our strategy presents a promising paradigm for synergistic cancer treatment, seamlessly integrating chemotherapy and immunotherapy.

Detection of nitric oxide-mediated metabolic effects using real-time extracellular flux analysis.

Dendritic cell (DC) activation is marked by key events including: (I) rapid induction and shifting of metabolism favoring glycolysis for generation of biosynthetic metabolic intermediates and (II) large scale changes in gene expression including the upregulation of the antimicrobial enzyme inducible nitric oxide synthase (iNOS) which produces the toxic gas nitric oxide (NO). Historically, acute metabolic reprogramming and NO-mediated effects on cellular metabolism have been studied at specific timepoints during the DC activation process, namely at times before and after NO production. However, no formal method of real time detection of NO-mediated effects on DC metabolism have been fully described. Here, using Real-Time Extracellular Flux Analysis, we experimentally establish the phenomenon of an NO-dependent mitochondrial respiration threshold, which shows how titration of an activating stimulus is inextricably linked to suppression of mitochondrial respiration in an NO-dependent manner. As part of this work, we explore the efficacy of two different iNOS inhibitors in blocking the iNOS reaction kinetically in real time and explore/discuss parameters and considerations for application using Real Time Extracellular Flux Analysis technology. In addition, we show, the temporal relationship between acute metabolic reprogramming and NO-mediated sustained metabolic reprogramming kinetically in single real-time assay. These findings provide a method for detection of NO-mediated metabolic effects in DCs and offer novel insight into the timing of the DC activation process with its associated key metabolic events, revealing a better understanding of the nuances of immune cell biology.

Exploiting Leishmania-Primed Dendritic Cells as Potential Immunomodulators of Canine Immune Response.

Dendritic cells (DCs) capture pathogens and process antigens, playing a crucial role in activating naïve T cells, bridging the gap between innate and acquired immunity. However, little is known about DC activation when facing Leishmania parasites. Thus, this study investigates in vitro activity of canine peripheral blood-derived DCs (moDCs) exposed to L. infantum and L. amazonensis parasites and their extracellular vesicles (EVs). L. infantum increased toll-like receptor 4 gene expression in synergy with nuclear factor κB activation and the generation of pro-inflammatory cytokines. This parasite also induced the expression of class II molecules of major histocompatibility complex (MHC) and upregulated co-stimulatory molecule CD86, which, together with the release of chemokine CXCL16, can attract and help in T lymphocyte activation. In contrast, L. amazonensis induced moDCs to generate a mix of pro- and anti-inflammatory cytokines, indicating that this parasite can establish a different immune relationship with DCs. EVs promoted moDCs to express class I MHC associated with the upregulation of co-stimulatory molecules and the release of CXCL16, suggesting that EVs can modulate moDCs to attract cytotoxic CD8+ T cells. Thus, these parasites and their EVs can shape DC activation. A detailed understanding of DC activation may open new avenues for the development of advanced leishmaniasis control strategies.