Antigen-presenting Cells Research Articles

Ro5-4864, a translocator protein ligand, regulates T cell-mediated inflammatory responses in skin.

Translocator protein (TSPO) is a mitochondrial outer membrane protein expressed on a variety of immune cells, including macrophages, dendritic cells, and T cells, in addition to neurons and steroid-producing cells. Previous studies of TSPO ligands have suggested that TSPO is involved in multiple cellular functions, including steroidogenesis, immunomodulation, and cell proliferation. Currently, there are limited reports on the effects of TSPO or TSPO ligands on T cell-mediated immune responses. We here investigated the involvement of TSPO/TSPO ligand in T cell responses using a 2,4-dinitro-1-fluorobenzene (DNFB)-induced contact hypersensitivity (CH) model. Treatment with Ro5-4864, a TSPO ligand, during DNFB sensitization reduced the number and activation status of CD4+ and CD8+ T cells in draining lymph nodes and alleviated skin inflammation after DNFB challenge. Adoptive transfer of Ro5-4864-treated mouse-derived DNFB-sensitized T cells to naïve mice inhibited CH responses after DNFB challenge. Ro5-4864-treated sensitized T cells showed lower proliferative responses when stimulated with DNFB-pulsed antigen-presenting cells compared to control-treated sensitized T cells. Ro5-4864 also suppressed cell proliferation, as well as adenosine triphosphate and lactate production, during T cell activation. Moreover, the inhibitory effects of Ro5-4864 on T cell responses were conserved in TSPO-deficient cells. Our results suggest that Ro5-4864 inhibits CH responses by suppressing energy metabolism, at least via glycolysis, to reduce the T cell primary response in a TSPO-independent manner.

Emerging Cationic Nanovaccines

Cationic vaccines of nanometric sizes can directly perform the delivery of antigen(s) and immunomodulator(s) to dendritic cells in the lymph nodes. The positively charged nanovaccines are taken up by antigen-presenting cells (APCs) of the lymphatic system often originating the cellular immunological defense required to fight intracellular microbial infections and the proliferation of cancers. Cationic molecules imparting the positive charges to nanovaccines exhibit a dose-dependent toxicity which needs to be systematically addressed. Against the coronavirus, mRNA cationic nanovaccines evolved rapidly. Nowadays cationic nanovaccines have been formulated against several infections with the advantage of cationic compounds granting protection of nucleic acids in vivo against biodegradation by nucleases. Up to the threshold concentration of cationic molecules for nanovaccine delivery, cationic nanovaccines perform well eliciting the desired Th 1 improved immune response in the absence of cytotoxicity. A second strategy in the literature involves dilution of cationic components in biocompatible polymeric matrixes. Polymeric nanoparticles incorporating cationic molecules at reduced concentrations for the cationic component often result in an absence of toxic effects. The progress in vaccinology against cancer involves in situ designs for cationic nanovaccines. The lysis of transformed cancer cells releases several tumoral antigens, which in the presence of cationic nanoadjuvants can be systemically presented for the prevention of metastatic cancer. In addition, these local cationic nanovaccines allow immunotherapeutic tumor treatment.

Viscoelastic synthetic antigen-presenting cells for augmenting the potency of cancer therapies.

The use of synthetic antigen-presenting cells to activate and expand engineered T cells for the treatment of cancers typically results in therapies that are suboptimal in effectiveness and durability. Here we describe a high-throughput microfluidic system for the fabrication of synthetic cells mimicking the viscoelastic and T-cell-activation properties of antigen-presenting cells. Compared with rigid or elastic microspheres, the synthetic viscoelastic T-cell-activating cells (SynVACs) led to substantial enhancements in the expansion of human CD8+ T cells and to the suppression of the formation of regulatory T cells. Notably, activating and expanding chimaeric antigen receptor (CAR) T cells with SynVACs led to a CAR-transduction efficiency of approximately 90% and to substantial increases in T memory stem cells. The engineered CAR T cells eliminated tumour cells in a mouse model of human lymphoma, suppressed tumour growth in mice with human ovarian cancer xenografts, persisted for longer periods and reduced tumour-recurrence risk. Our findings underscore the crucial roles of viscoelasticity in T-cell engineering and highlight the utility of SynVACs in cancer therapy.

Bayesian metamodeling of early T-cell antigen receptor signaling accounts for its nanoscale activation patterns.

T cells respond swiftly, specifically, sensitively, and robustly to cognate antigens presented on the surface of antigen presenting cells. Existing microscopic models capture various aspects of early T-cell antigen receptor (TCR) signaling at the molecular level. However, none of these models account for the totality of the data, impeding our understanding of early T-cell activation. Here, we study early TCR signaling using Bayesian metamodeling, an approach for systematically integrating multiple partial models into a metamodel of a complex system. We inform the partial models using multiple published super-resolution microscopy datasets. Collectively, these datasets describe the spatiotemporal organization, activity, interactions, and dynamics of TCR, CD45 and Lck signaling molecules in the early-forming immune synapse, and the concurrent membrane alterations. The resulting metamodel accounts for a distinct nanoscale dynamic pattern that could not be accounted for by any of the partial models on their own: a ring of phosphorylated TCR molecules, enriched at the periphery of early T cell contacts and confined by a proximal ring of CD45 molecules. The metamodel suggests this pattern results from limited activity range for the Lck molecules, acting as signaling messengers between kinetically-segregated TCR and CD45 molecules. We assessed the potential effect of Lck activity range on TCR phosphorylation and robust T cell activation for various pMHC:TCR association strengths, in the specific setting of an initial contact. We also inspected the impact of localized Lck inhibition via Csk recruitment to pTCRs, and that of splicing isoforms of CD45 on kinetic segregation. Due to the inherent scalability and adaptability of integrating independent partial models via Bayesian metamodeling, this approach can elucidate additional aspects of cell signaling and decision making.

Effect of Probiotic Lactobacillus Casei on Secretory Immunoglobulin A (S-IgA) Levels in Rats Norvegicus Strain Wistar Post Ovariectomy

Estrogen promotes the growth of Lactobacillus colonies in the vagina, converting glycogen in superficial cells into lactic acid, resulting in a low vaginal pH that inhibits pathogenic bacteria. Post-menopause, Lactobacillus growth declines, leading to thinner secretions, higher vaginal pH, and a weakened immune system. This study investigates the effect of Lactobacillus casei extract on secretory immunoglobulin A (IgA) levels, vaginal acidity, and beneficial bacteria in the vaginal mucosa. An experimental post-test-only control group design was employed with 24 female rats aged 8-10 weeks, weighing 180-220 grams. Following ovariectomy and a 14-day acclimation, the rats were divided into four groups: a control group (K) with no treatment and three treatment groups (P1, P2, P3) receiving Lactobacillus casei extract at doses of 2ml, 2.25ml, and 2.5ml, respectively, administered orally for 14 days. On day 15, vaginal acidity was measured using a swab method, and IgA levels were assessed via ELISA; gram-positive staining was performed for bacterial identification. One-way ANOVA revealed that Lactobacillus casei extract significantly increased IgA levels and reduced vaginal acidity (p = 0.003 and p = 0.000). The reduction in acidity is attributed to Lactobacillus converting glycogen into lactic acid, maintaining vaginal acidity, while S-IgA on the mucosal surface acts as an opsonin and antigen-presenting cell to inhibit pathogenic growth. The study concludes that Lactobacillus casei extract can elevate secretory IgA levels and normalize vaginal pH.

MAIT cells: Conserved watchers on the wall.

MAIT cells are innate-like T cells residing in barrier tissues such as the lung, skin, and intestine. Both the semi-invariant T cell receptor of MAIT cells and the restricting element MR1 are deeply conserved across mammals, indicating non-redundant functions linked to antigenic specificity. MAIT cells across species concomitantly express cytotoxicity and tissue-repair genes, suggesting versatile functions. Accordingly, MAIT cells contribute to antibacterial responses as well as to the repair of damaged barrier tissues. MAIT cells recognize riboflavin biosynthetic pathway-derived metabolites, which rapidly cross epithelial barriers to be presented by antigen-presenting cells. Changes in gut ecology during intestinal inflammation drive the expansion of strong riboflavin and MAIT ligand producers. Thus, MAIT cells may enable real-time surveillance of microbiota dysbiosis across intact epithelia and provide rapid and context-dependent responses. Here, we discuss recent findings regarding the origin and regulation of MAIT ligands and the role of MAIT cells in barrier tissues. We speculate on the potential reasons for MAIT cell conservation during evolution.

Immunoglobin attenuates fulminant myocarditis by inhibiting overactivated innate immune response.

Fulminant myocarditis (FM) is a myocardial inflammatory disease that can result from either viral diseases or autoimmune diseases. In this study, we have determined the treatment effects of immunomodulatory drugs on FM. FM was induced in A/JGpt mice by intraperitoneal administration of coxsackievirus B3, after which immunoglobins were administered daily by intraperitoneal injection. On the seventh day, the cardiac structure and function were determined using echocardiography and cardiac catheterisation. Single-cell RNA sequencing (scRNA-seq) was performed to evaluate CD45+ cells in the heart. Immunoglobin, a typical immunomodulatory drug, dramatically reduced mortality and significantly improved cardiac function in mice with FM. ScRNA-seq revealed that immunoglobin treatment effectively modulated cardiac immune homeostasis, particularly by attenuating overactivated innate immune responses. At the cellular level, immunoglobin predominantly targeted Plac8+ monocytes and S100a8+ neutrophils, suppressing their proinflammatory activities, and enhancing antigen processing and presentation capabilities, thereby amplifying the efficiency and potency of the immune response against the virus. Immunoglobin benefits are mediated by the modulation of multiple signalling pathways, including relevant receptors on immune cells, direction of inflammatory cell chemotaxis, antigen presentation and anti-viral effects. Subsequently, Bst2-ILT7 ligand-receptor-mediated cellular interactions manipulated by immunoglobin were further confirmed in vivo. Immunoglobin treatment significantly attenuated FM-induced cardiac inflammation and improved cardiac function by inhibiting overactivated innate immune responses.

The complex role of immune cells in antigen presentation and regulation of T-cell responses in hepatocellular carcinoma: progress, challenges, and future directions.

Hepatocellular carcinoma (HCC) is a prevalent form of liver cancer that poses significant challenges regarding morbidity and mortality rates. In the context of HCC, immune cells play a vital role, especially concerning the presentation of antigens. This review explores the intricate interactions among immune cells within HCC, focusing on their functions in antigen presentation and the modulation of T-cell responses. We begin by summarizing the strategies that HCC uses to escape immune recognition, emphasizing the delicate equilibrium between immune surveillance and evasion. Next, we investigate the specific functions of various types of immune cells, including dendritic cells, natural killer (NK) cells, and CD8+ T cells, in the process of antigen presentation. We also examine the impact of immune checkpoints, such as cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and the pathways involving programmed cell death protein 1 (PD-1) and programmed death ligand 1 (PD-L1), on antigen presentation, while taking into account the clinical significance of checkpoint inhibitors. The review further emphasizes the importance of immune-based therapies, including cancer vaccines and CAR-T cell therapy, in improving antigen presentation. In conclusion, we encapsulate the latest advancements in research, propose future avenues for exploration, and stress the importance of innovative technologies and customized treatment strategies. By thoroughly analyzing the interactions of immune cells throughout the antigen presentation process in HCC, this review provides an up-to-date perspective on the field, setting the stage for new therapeutic approaches.

Single cell transcriptional analysis of human adenoids identifies molecular features of airway microfold cells.

The nasal, oropharyngeal, and bronchial mucosa are primary contact points for airborne pathogens like Mycobacterium tuberculosis (Mtb), SARS-CoV-2, and influenza virus. While mucosal surfaces can function as both entry points and barriers to infection, mucosa-associated lymphoid tissues (MALT) facilitate early immune responses to mucosal antigens. MALT contains a variety of specialized epithelial cells, including a rare cell type called a microfold cell (M cell) that functions to transport apical antigens to basolateral antigen-presenting cells, a crucial step in the initiation of mucosal immunity. M cells have been extensively characterized in the gastrointestinal (GI) tract in murine and human models. However, the precise development and functions of human airway M cells is unknown. Here, using single-nucleus RNA sequencing (snRNA-seq), we generated an atlas of cells from the human adenoid and identified 16 unique cell types representing basal, club, hillock, and hematopoietic lineages, defined their developmental trajectories, and determined cell-cell relationships. Using trajectory analysis, we found that human airway M cells develop from progenitor club cells and express a gene signature distinct from intestinal M cells. Surprisingly, we also identified a heretofore unknown epithelial cell type demonstrating a robust interferon-stimulated gene signature. Our analysis of human adenoid cells enhances our understanding of mucosal immune responses and the role of M cells in airway immunity. This work also provides a resource for understanding early interactions of pathogens with airway mucosa and a platform for development of mucosal vaccines.

Acquired immunostimulatory phenotype of migratory CD103+ DCs promotes alloimmunity following corneal transplantation.

After transplantation, Th1-mediated immune rejection is the predominant cause of graft failure. Th1 cell sensitization occurs through complex and context-dependent interaction among antigen-presenting cell subsets, particularly CD11b+ DCs (DC2) and CD103+ DCs (DC1). This interaction necessitates further investigation in the context of transplant immunity. We used well-established preclinical models of corneal transplantation and identified distinct roles of migratory CD103+ DC1 in influencing the outcomes of the grafted tissue. In recipients with uninflamed corneal beds, migratory CD103+ DC1 demonstrate a tolerogenic phenotype that modulates the immunogenic capacity of CD11b+ DC2 primarily mediated by IL-10, suppressing alloreactive CD4+ Th1 cells via the PD-L1/PD-1 pathway and enhancing Treg-mediated tolerance via αvβ8 integrin-activated TGF-β1, thus facilitating graft survival. Conversely, in recipients with inflamed and vascularized corneal beds, IFN-γ produced by CD4+ Th1 cells induced migratory CD103+ DC1 to adopt an immunostimulatory phenotype, characterized by the downregulation of regulatory markers, including αvβ8 integrin and IL-10, and the upregulation of IL-12 and costimulatory molecules CD80/86, resulting in graft failure. The adoptive transfer of ex vivo induced tolerogenic CD103+ DC1 (iDC1) effectively inhibited Th1 polarization and preserved the tolerogenic phenotype of their physiological counterparts. Collectively, our findings underscore the essential role played by CD103+ DC1 in modulating host alloimmune responses.

Tobacco smoke exposure is a driver of altered oxidative stress response and immunity in head and neck cancer.

Exposomes are critical drivers of carcinogenesis. However, how they modulate tumor behavior remains unclear. Extensive clinical data link cigarette smoke as a key exposome that promotes aggressive tumors, higher rates of metastasis, reduced response to chemoradiotherapy, and suppressed anti-tumor immunity. We sought to determine whether smoke itself can modulate aggressive tumor behavior in head and neck squamous cell carcinoma (HNSCC) through reprogramming the cellular reductive state. Using established human and murine HNSCC cell lines and syngeneic mouse models, we utilized conventional western blotting, steady state and flux metabolomics, RNA sequencing, quantitative proteomics and flow cytometry to analyze the impact of smoke exposure on HNSCC tumor biology. Cigarette smoke persistently activated Nrf2 target genes essential for maintenance of the cellular reductive state and survival under conditions of increased oxidative stress in HNSCC regardless of HPV status. In contrast to e-cigarette vapor, conventional cigarette smoke mobilizes cellular metabolism toward oxidative stress adaptation, resulting in development of cross-resistance to cisplatin. In parallel, smoke exposure modulates both expression of PDL1 and the secretory phenotype of HNSCC cells through activation of NF-κB resulting in an altered tumor immune microenvironment (TIME) in syngeneic mouse models and altered PBMC differentiation that includes downregulated expression of antigen presentation and costimulatory genes in myeloid cells. Cigarette smoke exposome is a potent activator of the Nrf2 pathway and is a likely primary trigger for the tripartite phenotype of aggressive HNSCC consisting of: 1) reduced chemotherapy sensitivity, 2) enhanced metastatic potential and 3) suppressed anti-tumor immunity. The smoke exposome drives aggressive tumor behavior, treatment resistance and suppressed immunity through coordinated metabolic reprogramming. Successfully targeting this adaptation is critical to improving survival in smokers with head and neck cancer.

Development of Ribityllumazine Analogue as Mucosal-Associated Invariant T Cell Ligands.

Mucosal-associated invariant T (MAIT) cells are a subset of innate-like T cells abundant in human tissues that play a significant role in defense against bacterial and viral infections and in tissue repair. MAIT cells are activated by recognizing microbial-derived small-molecule ligands presented by the MHC class I related-1 protein. Although several MAIT cell modulators have been identified in the past decade, potent and chemically stable ligands remain limited. Herein, we carried out a structure-activity relationship study of ribityllumazine derivatives and found a chemically stable MAIT cell ligand with a pteridine core and a 2-oxopropyl group as the Lys-reactive group. The ligand showed high potency in a cocultivation assay using model cell lines of antigen-presenting cells and MAIT cells. The X-ray crystallographic analysis revealed the binding mode of the ligand to MR1 and the T cell receptor, indicating that it forms a covalent bond with MR1 via Schiff base formation. Furthermore, we found that the ligand stimulated proliferation of human MAIT cells in human peripheral blood mononuclear cells and showed an adjuvant effect in mice. Our developed ligand is one of the most potent among chemically stable MAIT cell ligands, contributing to accelerating therapeutic applications of MAIT cells.

Cancer Vaccines: Recent Insights and Future Directions

The field of cancer immunotherapy has seen incredible advancements in the past decades. mRNA-based cancer vaccines generating de novo T cell responses, particularly against tumor-specific antigens (TSAs), have demonstrated promising clinical outcomes and overcome diverse challenges. Despite the high potential of neoantigens to provide personalized immunotherapies through their tumor specificity and immunogenicity, challenges related to the scarcity of immunogenic neoepitopes have prompted continuous research towards finding new tumor-associated antigens (TAAs) and broader therapeutic frameworks, which may now learn from the genuine successes obtained with neoantigens. As an example, human endogenous retroviruses (HERVs) have emerged as potential alternatives to tumor neoantigens due to their high tumoral expression and ability to elicit both T cell reactivity and B cell responses associated with the efficacy of existing immunotherapies. This review aims to assess the status and limitations of TSA-directed mRNA cancer vaccines and the lessons that can be derived from these and checkpoint inhibitor studies to guide TAA vaccine development. We expect that shared B cell, CD4 and CD8 T cell antigen presentation will be key to stimulate continuous T cell expansion and efficacy for tumors that do not contain pre-existing tertiary lymphoid structures. When these structures are present in highly mutated tumors, the current checkpoint-based immunotherapies show efficacy even in immune privileged sites, and vaccines may hold the key to broaden efficacy to more tumor types and stages.

Stimulator of interferon gene facilitates recruitment of effector CD8 T cells that drive neurofibromatosis type 1 nerve tumor initiation and maintenance.

Plexiform neurofibromas (PNFs) are benign nerve tumors driven by loss of the NF1 tumor suppressor in Schwann cells. PNFs are rich in immune cells, but whether immune cells are necessary for tumorigenesis is unknown. We show that inhibition of stimulator of interferon gene (STING) reduces plasma CXCL10, tumor T cell and dendritic cell (DC) recruitment, and tumor formation. Further, mice lacking XCR-1+ DCs showed reduced tumor-infiltrating T cells and PNF tumors. Antigen-presenting cells from tumor-bearing mice promoted CD8+ T cell proliferation in vitro, and PNF T cells expressed high levels of CCL5, implicating T cell activation. Notably, tumors and nerve-associated macrophages were absent in Rag1-/-; Nf1f/f; DhhCre mice and adoptive transfer of CD8+ T cells from tumor-bearing mice restored PNF initiation. In this setting, PNF shrunk upon subsequent T cell removal. Thus, STING pathway activation contributes to CD8+ T cell-dependent inflammatory responses required for PNF initiation and maintenance.

Effect and mechanism of allergen-specific immunotherapy on small airway dysfunction in children with asthma.

The purpose of this study was to investigate the effectiveness of allergen-specific immunotherapy (AIT) on small airway dysfunction (SAD) and the underlying mechanism with a special focus on basophils. Sixty-five children with mild to moderate asthma who were under regular inhaled corticosteroid (ICS) treatment for more than 1 year but whose FEF75 remained below 65% of the predicted value and had positive results for serum Der p or Der f were enrolled. Children with asthma underwent house dust mite (HDM) subcutaneous immunotherapy (SCIT) treatment for 1 year. Clinical symptoms and lung function were evaluated every 3 months during HDM SCIT treatment. Basophil activation test (BAT) was carried out before and after HDM SCIT treatment. RNA sequencing was performed in isolated basophils from peripheral blood after 6 months of HDM SCIT treatment, followed by GO term and KEGG pathway enrichment analysis between patients with and without HDM SCIT treatment. HDM AIT treatment ameliorated clinical symptoms while concurrently improved lung function parameters, such as FEV1/FVC, FEF75, FEF50 and MMEF (p < .05). It is worth noting that FEF75 values showed a highly significant, gradual and persistent increase (from 49.55 ± 1.27% at baseline to 71.89 ± 2.64% after 1 year of therapy) and 22 of 35 patients no longer had SAD after 1 year of treatment. BAT results revealed that AIT treatment significantly reduced basophil activity to the inhalant allergen mixtures containing HDM in vitro challenge from baseline. GO term and KEGG pathway enrichment analysis of basophils revealed that downregulated genes were mainly involved in immune cell activation, antigen presentation, and Th2 cell differentiation. Our study demonstrated that HDM AIT not only improved SAD related lung function parameters, but also reduced basophil activity. RNA sequencing revealed the inhibition of phagocytosis and the phagosome pathway in basophils which may affect the polarization of Th2 cell differentiation after HDM AIT.

DRAK2 regulates myosin light chain phosphorylation in T cells.

Death-associated protein kinase-related apoptosis-inducing kinase-2 (DRAK2 or STK17B) is a serine/threonine kinase expressed in T cells. Drak2-deficient (Drak2-/-) mice respond effectively to tumors and pathogens while displaying resistance to T cell-mediated autoimmune disease. However, the molecular mechanisms by which DRAK2 impacts T cell function remain unclear. Gaining further insight into the function of DRAK2 in T cells will shed light on differentially regulated pathways in autoreactive and pathogen-specific T cells, which is critical for improving autoimmune therapies. Here, we demonstrate that DRAK2 contributes to activation of myosin light chain (MLC) in both murine and human T cells. In the absence of Drak2, the amount of polymerized actin was decreased, suggesting that DRAK2 modulates actomyosin dynamics. We further show that myosin-dependent T cell functions, such as migration, T cell receptor microcluster accumulation, and conjugation to antigen presenting cells are decreased in the absence of Drak2. These findings reveal that DRAK2 plays an important role in regulating MLC activation within T cells.

Single-Dose Physically Cross-Linked Hyaluronic Acid and Lipid Hybrid Nanoparticles Containing Cyclic Guanosine Monophosphate-Adenosine Monophosphate Eliminate Established Tumors.

Activating the STING pathway in the cytosol of tumor-infiltrating antigen-presenting cells (APCs) represents a promising strategy to elicit potent antitumor immune responses for cancer therapy. However, STING agonists are mostly small hydrophilic molecules that suffer from rapid clearance and poor cytosolic delivery following systemic administration. While various nanoparticles have been developed to promote cytosolic delivery, they often exhibit premature drug release during circulation. Alternatively, stable nanoparticles with sustained release during circulation have poor cytosolic delivery. In this study, we have developed physically cross-linked hyaluronic acid (HA) and lipid hybrid nanoparticles containing cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), denoted as HLHC, to address these challenges. The HLH delivery system has sustained drug release due to multiple lipid layers physically cross-linked by HA. HLHC efficiently delivers cGAMP to the cytosol of APCs, inducing more IFNβ than cGAMP and liposomal cGAMP. HLH also improves the drug circulation time and biodistribution to the tumor compared with the liposomal formulation and free drug. Strikingly, a single dose of HLHC, but not liposomal cGAMP or free cGAMP, elicits potent antitumor immunity and regresses established MC38 tumors. A single dose of HLHC even regresses established B16F10 tumors upon combination with αPD-L1. Moreover, cured animals were protected from rechallenge with the same tumor cells. HLHC represents an efficient strategy to address delivery challenges associated with STING agonists and may have broad applications for the delivery of drugs acting in the cytosol.

A dual-STING-activating nanosystem expands cancer immunotherapeutic temporal window.

Stimulator of interferon genes (STING) is a promising antitumor target via bridging innate and adaptive immunity, yet the transient nature of immune signal transduction renders small-molecule agonists susceptible to short time effectiveness. Here, we report a dual-STING-activating micelle system (D-SAM) to dynamically program STING kinetics. Mechanistically, the natural ligand cGAMP encapsulated in D-SAM initiates STING signaling, while the pH-sensitive polymeric agonist PC7A disassembled from micelle shell buffers lysosomal protons and retards STING degradation. This prolonged STING activity facilitates dendritic cell (DC) antigen presentation and extends cytotoxic T lymphocyte priming. D-SAM improves efficacy over single soluble or delivered agonists against established, metastatic, and recurring murine tumors. Specific depletion of STING in DCs or blockade of CD8+ Tcell infiltration abrogates therapeutic effects. The feasibility of immune modulation is further validated in resected human patient tissues. This work underscores the temporal rhythm of STING as crucial for mounting a potent and enduring antitumor immune response.

Unlocking Intracellular Protein Delivery by Harnessing Polymersomes Synthesized at Microliter Volumes using Photo-PISA.

Efficient delivery of therapeutic proteins and vaccine antigens to intracellular targets is challenging due to generally poor cell membrane permeation and endolysosomal entrapment causing degradation. Herein, these challenges are addressed by developing an oxygen-tolerant photoinitiated polymerization-induced self-assembly (Photo-PISA) process, allowing for the microliter-scale (10µL) synthesis of protein-loaded polymersomes directly in 1536-well plates. High-resolution techniques capable of analysis at a single particle level are employed to analyze protein encapsulation and release mechanisms. Using confocal microscopy and super-resolution stochastic optical reconstruction microscopy (STORM) imaging, their ability to deliver proteins into the cytosol following endosomal escape is subsequently visualized. Lastly, the adaptability of these polymersomes is exploited to encapsulate and deliver a prototype vaccine antigen, demonstrating its ability to activate antigen-presenting cells and support antigen cross-presentation for applications in subunit vaccines and cancer immunotherapy. This combination of ultralow volume synthesis and efficient intracellular delivery holds significant promise for unlocking the high throughput screening of a broad range of otherwise cost-prohibitive or early-stage therapeutic protein and vaccine antigen candidates that can be difficult to obtain in large quantities. The versatility of this platform for rapid screening of intracellular protein delivery can result in significant advancements across the fields of nanomedicine and biomedical engineering.

Anti-cancer immune effect of human colorectal cancer neoantigen peptide based on MHC class I molecular affinity screening

BackgroundTumor antigen peptide vaccines have shown remarkable efficacy, safety, and reliability in recent studies. However, the screening process for immunopotent antigenic peptides is cumbersome, limiting their widespread application. Identifying neoantigen peptides that can effectively trigger an immune response is crucial for personalized cancer treatment.MethodsWhole exome sequencing was performed on patient-derived colon cancer cells to predict 9-amino-acid (9aa) neoantigen peptides. In vitro simulation of endogenous antigen presentation by antigen-presenting cells (dendritic cells) to CD8+ T cells was conducted, aiming to activate the CD8+ immune response to the predicted antigens. The immunological effects of each neoantigen were assessed using flow cytometry and ELISpot assays, while the relationship between neoantigen immunogenicity and MHC molecular affinity was examined.Results1. Next-generation sequencing (NGS) predicted 9-amino acid (9aa) neoantigen peptides for subsequent immunological analysis.2. Higher mDC Levels in Experimental Group: CD11c+CD83+ mature dendritic cells (mDCs) were 96.6% in the experimental group, compared to 0.051% in the control group. CD80 fluorescence intensity was also significantly higher in the experimental group, confirming a greater mDC presence.3. Neoantigen Peptides Promote CD4+, CD8+ T, and NK Cell Proliferation: After 14 days, flow cytometry showed higher percentages of CD4+ T (37.41% vs 7.8%), CD8+ T (16.67% vs 14.63%), and NK cells (33.09% vs 7.81%) in the experimental group, indicating that the neoantigen peptides induced proliferation of CD4+, CD8+ T cells, and NK cells. 4. The results, analyzed using two-way ANOVA, showed that the standardized T-value for HLA molecular affinity variation in the 1-4 range (Group B) was significantly higher than for ≤1 (Group A, p &amp;lt; 0.0001) and &amp;gt;4 (Group C, p &amp;lt; 0.05). Regarding HLA-allele genotypes, HLA-Type 1 had a significantly higher standardized T-value than HLA-Type 2 (p &amp;lt; 0.05) and HLA-Type 3 (p &amp;lt; 0.0001). HLA-Type 1 was identified as the allele associated with the highest T-value.Conclusion1. The most immunogenic neoantigens typically exhibit an MHC molecular affinity variation between 1 and 4, indicating that stronger immunogenicity correlates with higher MHC molecular affinity variation. 2. Each patient's HLA molecules were classified into Types 1, 2, and 3, with Type 1 showing the highest binding capacity for neoantigens. Our findings indicate that the most immunogenic neoantigens were associated with HLA Type 1. 3. Neoantigen peptides were shown to activate the proliferation of both CD8+ T-cells and induce proliferation of CD4+ T-cells and NK cells. 4. Variation in MHC molecular affinity and HLA neoantigen genotype are anticipated to serve as valuable variables for screening highly immunogenic neoantigens, facilitating more efficient preparation of effective polypeptide tumor vaccines.