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Administration of neutralizing antibodies (nAbs) has proved to be effective by providing immediate protection against SARS‐CoV‐2. However, dual strategies combining virus neutralization and immune response stimulation to enhance specific cytotoxic T cell responses, such as dendritic cell (DC) cross‐priming, represent a promising field but have not yet been explored. Here, a broadly nAb, TNT, are first generated by grafting an anti‐RBD biparatopic tandem nanobody onto a trimerbody scaffold. Cryo‐EM data show that the TNT structure allows simultaneous binding to all six RBD epitopes, demonstrating a high‐avidity neutralizing interaction. Then, by C‐terminal fusion of an anti‐DNGR‐1 scFv to TNT, the bispecific trimerbody TNTDNGR‐1 is generated to target neutralized virions to type 1 conventional DCs (cDC1s) and promote T cell cross‐priming. Therapeutic administration of TNTDNGR‐1, but not TNT, protects K18‐hACE2 mice from a lethal SARS‐CoV‐2 infection, boosting virus‐specific humoral responses and CD8+ T cell responses. These results further strengthen the central role of interactions with immune cells in the virus‐neutralizing antibody activity and demonstrate the therapeutic potential of the Fc‐free strategy that can be used advantageously to provide both immediate and long‐term protection against SARS‐CoV‐2 and other viral infections. [ABSTRACT FROM AUTHOR]

Naturally occurring cases of monogenic type 1 diabetes (T1D) help establish direct mechanisms driving this complex autoimmune disease. A recently identified de novo germline gain-of-function (GOF) mutation in the transcriptional regulator STAT3 was found to cause neonatal T1D. We engineered a novel knock-in mouse incorporating this highly diabetogenic human STAT3 mutation (K392R) and found that these mice recapitulated the human autoimmune diabetes phenotype. Paired single-cell TCR and RNA sequencing revealed that STAT3-GOF drives proliferation and clonal expansion of effector CD8+ cells that resist terminal exhaustion. Single-cell ATAC-seq showed that these effector T cells are epigenetically distinct and have differential chromatin architecture induced by STAT3-GOF. Analysis of islet TCR clonotypes revealed a CD8+ cell reacting against known antigen IGRP, and STAT3-GOF in an IGRP-reactive TCR transgenic model demonstrated that STAT3-GOF intrinsic to CD8+ cells is sufficient to accelerate diabetes onset. Altogether, these findings reveal a diabetogenic CD8+ T cell response that is restrained in the presence of normal STAT3 activity and drives diabetes pathogenesis.

The tumor microenvironment plays a critical role in modulating immune responses associated with tumorigenesis, tumor progression, and metastasis. Dendritic cells (DC) play a key role in preventing and progression of metastatic neoplasia by driving and restoring dysfunctional immune systems and obliterating immunosuppression, thus obstructing tumor evasion. In this review, we will discuss the functions of tumor-infiltrating DC in anti-tumor resistance, prevention of tumor recurrence, and immunosuppression. We will also describe DC metabolism, differentiation, and plasticity, which are essential for its function. Cancers like Lymphomas may be able to corrupt immune surveillance by reducing natural killer cell numbers. Thus, interactions between lymphoma and DC with reference to cytotoxicity may be an important event, likely to be mediated via activation with interferon-γ (IFN-γ) and Toll like receptors (TLR) ligands. Mechanisms of DC-mediated cytotoxicity and the role of apoptosis and death receptors, including the role played by nitric oxide, etc., are of immense significance. We will also look into the molecular mechanisms in the tumor microenvironment, reduced drug sensitivity, and tumor relapse, as well as methods for combating drug resistance and focusing on immunosuppressive tumor networks. We will address how DC mediated cytotoxicity in combination with drugs affects tumor growth and expansion in relation to checkpoint inhibitors and regulatory T cells. Innovative approaches for therapeutic modulation of this immunosuppressive adoptive DC immunotherapy will be highlighted, which is necessary for future personalized therapeutic applications.

The innate immune system detects pathogens and initiates adaptive immune responses. Inflammasomes are central components of the innate immune system, but whether inflammasomes provide sufficient signals to activate adaptive immunity is unclear. In intestinal epithelial cells (IECs), inflammasomes activate a lytic form of cell death called pyroptosis, leading to epithelial cell expulsion and the release of cytokines. Here, we employed a genetic system to show that simultaneous antigen expression and inflammasome activation specifically in IECs is sufficient to activate CD8+ T cells. By genetic elimination of direct T cell priming by IECs, we found that IEC-derived antigens were cross-presented to CD8+ T cells. However, cross-presentation of IEC-derived antigen to CD8+ T cells only partially depended on IEC pyroptosis. In the absence of inflammasome activation, cross-priming of CD8+ T cells required Batf3+ dendritic cells (conventional type one dendritic cells [cDC1]), whereas cross-priming in the presence of inflammasome activation required a Zbtb46+ but Batf3-independent cDC population. These data suggest the existence of parallel inflammasome-dependent and inflammasome-independent pathways for cross-presentation of IEC-derived antigens.

Safe and effective T cell vaccines are needed for the treatment or prevention of cancers as well as infectious agents where vaccines for neutralizing antibodies have performed poorly. Recent research highlights an important role for tissue-resident memory T cells (TRM cells) in protective immunity and the role of a subset of dendritic cells that are capable of cross-priming for the induction of TRM cells. However, efficient vaccine technologies that operate through cross-priming and induce robust CD8+ T cell responses are lacking. We developed a platform technology by genetically engineering the bovine papillomavirus L1 major capsid protein to insert a polyglutamic acid/cysteine motif in place of wild-type amino acids in the HI loop. Virus-like particles (VLPs) are formed by self-assembly in insect cells infected with a recombinant baculovirus. Polyarginine/cysteine-tagged antigens are linked to the VLP by a reversible disulfide bond. The VLP possesses self-adjuvanting properties due to the immunostimulatory activity of papillomavirus VLPs. Polyionic VLP vaccines induce robust CD8+ T cell responses in peripheral blood and tumor tissues. A prostate cancer polyionic VLP vaccine was more efficacious than other vaccines and immunotherapies for the treatment of prostate cancer in a physiologically relevant murine model and successfully treated more advanced diseases than the less efficacious technologies. The immunogenicity of polyionic VLP vaccines is dependent on particle size, reversible linkage of the antigen to the VLP, and an interferon type 1 and Toll-like receptor (TLR)3/7-dependent mechanism. [ABSTRACT FROM AUTHOR]

Broadly neutralizing monoclonal antibodies protect against infection with HIV-1 in animal models, suggesting that a vaccine that elicits these antibodies would be protective in humans. However, it has not yet been possible to induce adequate serological responses by vaccination. Here, to activate B cells that express precursors of broadly neutralizing antibodies within polyclonal repertoires, we developed an immunogen, RC1, that facilitates the recognition of the variable loop 3 (V3)-glycan patch on the envelope protein of HIV-1. RC1 conceals non-conserved immunodominant regions by the addition of glycans and/or multimerization on virus-like particles. Immunization of mice, rabbits and rhesus macaques with RC1 elicited serological responses that targeted the V3-glycan patch. Antibody cloning and cryo-electron microscopy structures of antibody-envelope complexes confirmed that immunization with RC1 expands clones of B cells that carry the anti-V3-glycan patch antibodies, which resemble precursors of human broadly neutralizing antibodies. Thus, RC1 may be a suitable priming immunogen for sequential vaccination strategies in the context of polyclonal repertoires.

Malaria remains a significant cause of morbidity and mortality worldwide, particularly in infants and children. Some studies have reported that exposure to malaria antigens in utero results in the development of tolerance, which could contribute to poor immunity to malaria in early life. However, the effector T cell response to pathogen-derived antigens encountered in utero, including malaria, has not been well characterized. Here, we assessed the frequency, phenotype, and function of cord blood T cells from Ugandan infants born to mothers with and without placental malaria. We found that infants born to mothers with active placental malaria had elevated frequencies of proliferating effector memory fetal CD4+ T cells and higher frequencies of CD4+ and CD8+ T cells that produced inflammatory cytokines. Fetal CD4+ and CD8+ T cells from placental malaria-exposed infants exhibited greater in vitro proliferation to malaria antigens. Malaria-specific CD4+ T cell proliferation correlated with prospective protection from malaria during childhood. These data demonstrate that placental malaria is associated with the generation of proinflammatory malaria-responsive fetal T cells. These findings add to our current understanding of fetal immunity and indicate that a functional and protective pathogen-specific T cell response can be generated in utero.

CD8+ T cells belonging to the adaptive immune system play key roles in defending against viral infections and cancers. The current CD8+ T cell-based immunotherapy has emerged as a superior therapeutic avenue for the eradication of tumor cells and long-term prevention of their recurrence in hematologic malignancies. It is believed that an effective adaptive immune response critically relies on the help of the innate compartment. Invariant natural killer T (iNKT) cells are innate-like T lymphocytes that have been considered some of the first cells to respond to infections and can secrete a large amount of diverse cytokines and chemokines to widely modulate the innate and adaptive immune responders. Like CD8+ T cells, iNKT cells also play an important role in defense against intracellular pathogenic infections and cancers. In this review, we will discuss the CD8+ T-cell immunity contributed by iNKT cells, including iNKT cell-mediated cross-priming and memory formation, and discuss recent advances in our understanding of the mechanisms underlying memory CD8+ T-cell differentiation, as well as aging-induced impairment of T-cell immunity. [ABSTRACT FROM AUTHOR]

Transgenerational plasticity in plants is an increasingly recognized phenomenon, yet it is mostly unclear whether transgenerational plasticity is relevant to both the fitness of the plant and its interacting species. Using monoclonal strains of the giant duckweed (Spirodela polyrhiza) and its native herbivore, the waterlily aphid (Rhopalosiphum nymphaeae), we assessed whether pre-treating plants with copper excess, both indoors and outdoors, induces transgenerational plasticity in plant defences that alter plant and herbivore fitness. Outdoors, copper pre-treatment tended to increase plant growth rates under recurring copper excess. Indoors, copper pre-treatment either increased or decreased plant growth rates under recurring conditions, depending on the plant genotype. Copper pre-treatment induced anthocyanins that protected plants against copper toxicity, and these elevated levels were transgenerationally retained. Copper pre-treatment also transgenerationally increased the levels of 12-oxo-phytodienoic acid (OPDA), a jasmonate precursor. Nevertheless, aphids grew up to 50% better when the plants were pre-treated with copper. The increased aphid growth was likely caused by transgenerationally elevated OPDA levels, as aphids grew better when jasmonates were externally applied to plants. Taken together, this study shows that transgenerational plasticity is relevant to both plant and herbivore fitness, which highlights the role of transgenerational plasticity in plant evolution and species interactions., (© 2025 The Author(s). Plant, Cell & Environment published by John Wiley & Sons Ltd.)

CD8+ T cells belonging to the adaptive immune system play key roles in defending against viral infections and cancers. The current CD8+ T cell-based immunotherapy has emerged as a superior therapeutic avenue for the eradication of tumor cells and long-term prevention of their recurrence in hematologic malignancies. It is believed that an effective adaptive immune response critically relies on the help of the innate compartment. Invariant natural killer T (iNKT) cells are innate-like T lymphocytes that have been considered some of the first cells to respond to infections and can secrete a large amount of diverse cytokines and chemokines to widely modulate the innate and adaptive immune responders. Like CD8+ T cells, iNKT cells also play an important role in defense against intracellular pathogenic infections and cancers. In this review, we will discuss the CD8+ T-cell immunity contributed by iNKT cells, including iNKT cell-mediated cross-priming and memory formation, and discuss recent advances in our understanding of the mechanisms underlying memory CD8+ T-cell differentiation, as well as aging-induced impairment of T-cell immunity.

Hepatic metastases are a poor prognostic factor of colorectal carcinoma (CRC) and new strategies to reduce the risk of liver CRC colonization are highly needed. Herein, we used mouse models of hepatic metastatization to demonstrate that the continuous infusion of therapeutic doses of interferon-alpha (IFNα) controls CRC invasion by acting on hepatic endothelial cells (HECs). Mechanistically, IFNα promoted the development of a vascular antimetastatic niche characterized by liver sinusoidal endothelial cells (LSECs) defenestration extracellular matrix and glycocalyx deposition, thus strengthening the liver vascular barrier impairing CRC trans-sinusoidal migration, without requiring a direct action on tumor cells, hepatic stellate cells, hepatocytes, or liver dendritic cells (DCs), Kupffer cells (KCs) and liver capsular macrophages (LCMs). Moreover, IFNα endowed LSECs with efficient cross-priming potential that, along with the early intravascular tumor burden reduction, supported the generation of antitumor CD8+ T cells and ultimately led to the establishment of a protective long-term memory T cell response. These findings provide a rationale for the use of continuous IFNα therapy in perioperative settings to reduce CRC metastatic spreading to the liver.

Tumor immunosurveillance requires tumor cell-derived molecules to initiate responses through corresponding receptors on antigen presenting cells (APCs) and a specific effector response designed to eliminate the emerging tumor cells. This is supported by evidence from immunodeficient individuals and experimental animals. Recent discoveries suggest that adjuvanticity of tumor-derived heat shock proteins (HSPs) and double-stranded DNA (dsDNA) are necessary for tumor-specific immunity. There is also the obligatory early transfer of tumor antigens to APCs. We argue that tumor-derived HSPs deliver sufficient chaperoned antigen for cross-priming within the quantitative limits set by nascent tumors. In contrast to late-stage tumors, we are only just beginning to understand the unique interactions of the immune system with precancerous/nascent neoplastic cells, which is important for improved cancer prevention measures. Tumor-derived heat shock proteins (HSPs) and double-stranded DNA (dsDNA) are self-derived mammalian molecules that can provide adjuvanticity necessary to initiate adaptive immune responses. We propose that the receptors for HSPs and dsDNA, which are CD91 and stimulator of interferon genes (STING), respectively, may play a crucial role in cancer immunosurveillance; their loss, dysfunction, or inhibition allows for an increased incidence of cancer in mice and humans. Tumor antigens chaperoned by HSPs are crosspresented by dendritic cells via CD91 to prime T cell responses. Estimates of the amount of chaperoned antigen and HSPs satisfies the quantitative limits available in nascent, emerging tumors and we argue that this strongly supports such a mechanism as a likely important crosspresenting pathway in cancer immunosurveillance. [ABSTRACT FROM AUTHOR]

Microbial systems have been synthetically engineered to deploy therapeutic payloads in vivo 1,2 . With emerging evidence that bacteria naturally home in on tumours 3,4 and modulate antitumour immunity 5,6 , one promising application is the development of bacterial vectors as precision cancer vaccines 2,7 . Here we engineered probiotic Escherichia coli Nissle 1917 as an antitumour vaccination platform optimized for enhanced production and cytosolic delivery of neoepitope-containing peptide arrays, with increased susceptibility to blood clearance and phagocytosis. These features enhance both safety and immunogenicity, achieving a system that drives potent and specific T cell-mediated anticancer immunity that effectively controls or eliminates tumour growth and extends survival in advanced murine primary and metastatic solid tumours. We demonstrate that the elicited antitumour immune response involves recruitment and activation of dendritic cells, extensive priming and activation of neoantigen-specific CD4 + and CD8 + T cells, broader activation of both T and natural killer cells, and a reduction of tumour-infiltrating immunosuppressive myeloid and regulatory T and B cell populations. Taken together, this work leverages the advantages of living medicines to deliver arrays of tumour-specific neoantigen-derived epitopes within the optimal context to induce specific, effective and durable systemic antitumour immunity., Competing Interests: Competing interests A.R., J.I., T.D. and N.A. have filed a provisional patent application with the US Patent and Trademark Office related to this work. The other authors declare no competing interests., (© 2024. The Author(s).)

Cancer vaccination drives the generation of anti-tumor T cell immunity and can be enhanced by the inclusion of effective immune adjuvants such as type I interferons (IFNs). Whilst type I IFNs have been shown to promote cross-priming of T cells, the role of individual subtypes remains unclear. Here we systematically compared the capacity of distinct type I IFN subtypes to enhance T cell responses to a whole-cell vaccination strategy in a pre-clinical murine model. We show that vaccination in combination with IFNβ induces significantly greater expansion of tumor-specific CD8+ T cells than the other type I IFN subtypes tested. Optimal expansion was dependent on the presence of XCR1+ dendritic cells, CD4+ T cells, and CD40/CD40L signaling. Therapeutically, vaccination with IFNβ delayed tumor progression when compared to vaccination without IFN. When vaccinated in combination with anti-PD-L1 checkpoint blockade therapy (CPB), the inclusion of IFNβ associated with more mice experiencing complete regression and a trend in increased overall survival. This work demonstrates the potent adjuvant activity of IFNβ, highlighting its potential to enhance cancer vaccination strategies alone and in combination with CPB.

Cancer vaccination drives the generation of anti-tumor T cell immunity and can be enhanced by the inclusion of effective immune adjuvants such as type I interferons (IFNs). Whilst type I IFNs have been shown to promote cross-priming of T cells, the role of individual subtypes remains unclear. Here we systematically compared the capacity of distinct type I IFN subtypes to enhance T cell responses to a whole-cell vaccination strategy in a pre-clinical murine model. We show that vaccination in combination with IFNβ induces significantly greater expansion of tumor-specific CD8+ T cells than the other type I IFN subtypes tested. Optimal expansion was dependent on the presence of XCR1+ dendritic cells, CD4+ T cells, and CD40/CD40L signaling. Therapeutically, vaccination with IFNβ delayed tumor progression when compared to vaccination without IFN. When vaccinated in combination with anti-PD-L1 checkpoint blockade therapy (CPB), the inclusion of IFNβ associated with more mice experiencing complete regression and a trend in increased overall survival. This work demonstrates the potent adjuvant activity of IFNβ, highlighting its potential to enhance cancer vaccination strategies alone and in combination with CPB. [ABSTRACT FROM AUTHOR]

Understanding how viruses subvert host immunity and persist is essential for developing strategies to eliminate infection. T cell exhaustion during chronic viral infection is well described, but effects on antibody-mediated effector activity are unclear. Herein, we show that increased amounts of immune complexes generated in mice persistently infected with lymphocytic choriomeningitis virus (LCMV) suppressed multiple Fcγ-receptor (FcγR) functions. The high amounts of immune complexes suppressed antibody-mediated cell depletion, therapeutic antibody-killing of LCMV infected cells and human CD20-expressing tumors, as well as reduced immune complex-mediated cross-presentation to T cells. Suppression of FcγR activity was not due to inhibitory FcγRs or high concentrations of free antibody, and proper FcγR functions were restored when persistently infected mice specifically lacked immune complexes. Thus, we identify a mechanism of immunosuppression during viral persistence with implications for understanding effective antibody activity aimed at pathogen control.

Dendritic cells (DCs) that orchestrate mucosal immunity have been studied in mice. Here we characterized human gut DC populations and defined their relationship to previously studied human and mouse DCs. CD103(+)Sirpα(-) DCs were related to human blood CD141(+) DCs and to mouse intestinal CD103(+)CD11b(-) DCs and expressed markers of cross-presenting DCs. CD103(+)Sirpα(+) DCs aligned with human blood CD1c(+) DCs and mouse intestinal CD103(+)CD11b(+) DCs and supported the induction of regulatory T cells. Both CD103(+) DC subsets induced the TH17 subset of helper T cells, while CD103(-)Sirpα(+) DCs induced the TH1 subset of helper T cells. Comparative analysis of transcriptomes revealed conserved transcriptional programs among CD103(+) DC subsets and identified a selective role for the transcriptional repressors Bcl-6 and Blimp-1 in the specification of CD103(+)CD11b(-) DCs and intestinal CD103(+)CD11b(+) DCs, respectively. Our results highlight evolutionarily conserved and divergent programming of intestinal DCs.

Many current cancer vaccine strategies suffer from the inability to mount a CD8 T cell response that is strong enough to overcome the low immunogenicity of tumors. Viruses naturally possess the sizes, geometries, and physical properties for which the immune system has evolved to recognize, and mimicking those properties with nanoparticles can produce robust platforms for vaccine design. Using the nonviral E2 core of pyruvate dehydrogenase, we have engineered a viral-mimicking vaccine platform capable of encapsulating dendritic cell (DC)-activating CpG molecules in an acid-releasable manner and displaying MHC I-restricted SIINFEKL peptide epitopes. Encapsulated CpG activated bone marrow-derived DCs at a 25-fold lower concentration in vitro when delivered with the E2 nanoparticle than with unbound CpG alone. Combining CpG and SIINFEKL within a single multifunctional particle induced ∼3-fold greater SIINFEKL display on MHC I by DCs over unbound peptide. Importantly, combining CpG and SIINFEKL to the E2 nanoparticle for simultaneous temporal and spatial delivery to DCs showed increased and prolonged CD8 T cell activation, relative to free peptide or peptide-bound E2. By codelivering peptide epitopes and CpG activator in a particle of optimal DC-uptake size, we demonstrate the ability of a noninfectious protein nanoparticle to mimic viral properties and facilitate enhanced DC activation and cross-presentation.

Cerebral malaria is a devastating complication of Plasmodium falciparum infection. Its pathogenesis is complex, involving both parasite- and immune-mediated events. CD8(+) T cells play an effector role in murine experimental cerebral malaria (ECM) induced by Plasmodium berghei ANKA (PbA) infection. We have identified a highly immunogenic CD8 epitope in glideosome-associated protein 50 that is conserved across rodent malaria species. Epitope-specific CD8(+) T cells are induced during PbA infection, migrating to the brain just before neurological signs manifest. They are functional, cytotoxic and can damage the blood-brain barrier in vivo. Such CD8(+) T cells are also found in the brain during infection with parasite strains/species that do not induce neuropathology. We demonstrate here that PbA infection causes brain microvessels to cross-present parasite antigen, while non-ECM-causing parasites do not. Further, treatment with fast-acting anti-malarial drugs before the onset of ECM reduces parasite load and thus antigen presentation in the brain, preventing ECM death. Thus our data suggest that combined therapies targeting both the parasite and host antigen-presenting cells may improve the outcome of CM patients.

Due to inherent differences in cellular composition and metabolic behavior with host cells, tumor-harbored bacteria can discriminatorily affect tumor immune landscape. However, the mechanisms by which intracellular bacteria affect antigen presentation process between tumor cells and antigen-presenting cells (APCs) are largely unknown. The invasion behavior of attenuated Salmonella VNP20009 (VNP) into tumor cells is investigated and an attempt is made to modulate this behavior by modifying positively charged polymers on the surface of VNP. It is found that non-toxic chitosan oligosaccharide (COS) modified VNP (VNP@COS) bolsters the formation of gap junction between tumor cells and APCs by enhancing the ability of VNP to infect tumor cells. On this basis, a bacterial biohybrid is designed to promote in situ antigen cross-presentation through intracellular bacteria induced gap junction. This bacterial biohybrid also enhances the expression of major histocompatibility complex class I molecules on the surface of tumor cells through the incorporation of Mdivi-1 coupled with VNP@COS. This strategic integration serves to heighten the immunogenic exposure of tumor antigens; while, preserving the cytotoxic potency of T cells. A strategy is proposed to precisely controlling the function and local effects of microorganisms within tumors., (© 2024 Wiley‐VCH GmbH.)

Proteasomes are complex macromolecular structures existing in various forms to regulate a myriad of cellular processes. Besides degrading unwanted or misfolded proteins (proteostasis), distinct immune functions were ascribed for the immunoproteasome and thymoproteasome (TPr) complexes. For instance, antigen degradation during ongoing immune responses mainly relies on immunoproteasome activity, whereas intrathymic CD8 T-cell development requires peptide generation by the TPr complex. Despite these substantial differences, the functional contribution of the TPr to peripheral T-cell immunity remains ill-defined. We herein explored whether the use of mesenchymal stromal cells (MSCs) engineered to exhibit altered proteasomal activity through de novo expression of the TPr complex can be exploited as a novel anti-cancer vaccine capable of triggering potent CD8 T-cell activation. Phenotypic and molecular characterization of MSC-TPr revealed a substantial decrease in MHCI (H2-Kb and H2-Dd) expression along with elevated secretion of various chemokines (CCL2, CCL9, CXCL1, LIX, and CX3CL1). In parallel, transcriptomic analysis pinpointed the limited ability of MSC-TPr to present endogenous antigens, which is consistent with their low expression levels of the peptide-loading proteins TAP, CALR, and PDAI3. Nevertheless, MSC-TPr cross-presented peptides derived from captured soluble proteins. When tested for their protective capacity, MSC-TPr triggered modest anti-tumoral responses despite efficient generation of effector memory CD4 and CD8 T cells. In contrast, clodronate administration prior to vaccination dramatically enhanced the MSC-TPr-induced anti-tumoral immunity clearly highlighting a refractory role mediated by phagocytic cells. Thus, our data elute to a DC cross-priming-dependant pathway in mediating the therapeutic effect of MSC-TPr.

Proteasomes are complex macromolecular structures existing in various forms to regulate a myriad of cellular processes. Besides degrading unwanted or misfolded proteins (proteostasis), distinct immune functions were ascribed for the immunoproteasome and thymoproteasome (TPr) complexes. For instance, antigen degradation during ongoing immune responses mainly relies on immunoproteasome activity, whereas intrathymic CD8 T-cell development requires peptide generation by the TPr complex. Despite these substantial differences, the functional contribution of the TPr to peripheral T-cell immunity remains ill-defined. We herein explored whether the use of mesenchymal stromal cells (MSCs) engineered to exhibit altered proteasomal activity through de novo expression of the TPr complex can be exploited as a novel anti-cancer vaccine capable of triggering potent CD8 T-cell activation. Phenotypic and molecular characterization of MSC-TPr revealed a substantial decrease in MHCI (H2-Kb and H2-Dd) expression along with elevated secretion of various chemokines (CCL2, CCL9, CXCL1, LIX, and CX3CL1). In parallel, transcriptomic analysis pinpointed the limited ability of MSC-TPr to present endogenous antigens, which is consistent with their low expression levels of the peptide-loading proteins TAP, CALR, and PDAI3. Nevertheless, MSC-TPr cross-presented peptides derived from captured soluble proteins. When tested for their protective capacity, MSC-TPr triggered modest anti-tumoral responses despite efficient generation of effector memory CD4 and CD8 T cells. In contrast, clodronate administration prior to vaccination dramatically enhanced the MSC-TPr-induced anti-tumoral immunity clearly highlighting a refractory role mediated by phagocytic cells. Thus, our data elute to a DC cross-priming-dependant pathway in mediating the therapeutic effect of MSC-TPr. [ABSTRACT FROM AUTHOR]

Although plasmacytoid dendritic cells (pDCs) have been shown to play a critical role in generating viral immunity and promoting tolerance to suppress antitumor immunity, whether and how pDCs cross-prime CD8 T cells in vivo remain controversial. Using a pDC-targeted vaccine model to deliver antigens specifically to pDCs, we have demonstrated that pDC-targeted vaccination led to strong cross-priming and durable CD8 T cell immunity. Surprisingly, cross-presenting pDCs required conventional DCs (cDCs) to achieve cross-priming in vivo by transferring antigens to cDCs. Taking advantage of an in vitro system where only pDCs had access to antigens, we further demonstrated that cross-presenting pDCs were unable to efficiently prime CD8 T cells by themselves, but conferred antigennaive cDCs the capability of cross-priming CD8 T cells by transferring antigens to cDCs. Although both cDC1s and cDC2s exhibited similar efficiency in acquiring antigens from pDCs, cDC1s but not cDC2s were required for cross-priming upon pDC-targeted vaccination, suggesting that cDC1s played a critical role in pDC-mediated cross-priming independent of their function in antigen presentation. Antigen transfer from pDCs to cDCs was mediated by previously unreported pDC-derived exosomes (pDCexos), that were also produced by pDCs under various conditions. Importantly, all these pDCexos primed naive antigen-specific CD8 T cells only in the presence of bystander cDCs, similarly to cross-presenting pDCs, thus identifying pDCexo-mediated antigen transfer to cDCs as a mechanism for pDCs to achieve cross-priming. In summary, our data suggest that pDCs employ a unique mechanism of pDCexo-mediated antigen transfer to cDCs for cross-priming. [ABSTRACT FROM AUTHOR]

Primary biliary cirrhosis (PBC) is characterized by an intense biliary inflammatory CD4(+) and CD8(+) T cell response. Very limited information on autoantigen-specific cytotoxic T lymphocyte (CTL) responses is available compared with autoreactive CD4(+) T cell responses. Using peripheral blood mononuclear cells (PBMCs) from PBC, we identified an HLA-A2-restricted CTL epitope of the E2 component of pyruvate dehydrogenase (PDC-E2), the immunodominant mitochondrial autoantigen. This peptide, amino acids 159-167 of PDC-E2, induces specific MHC class I-restricted CD8(+) CTL lines from 10/12 HLA-A2(+) PBC patients, but not controls, after in vitro stimulation with antigen-pulsed dendritic cells (DCs). PDC-E2-specific CTLs could also be generated by pulsing DCs with full-length recombinant PDC-E2 protein. Furthermore, using soluble PDC-E2 complexed with either PDC-E2-specific human monoclonal antibody or affinity-purified autoantibodies against PDC-E2, the generation of PDC-E2-specific CTLs, occurred at 100-fold and 10-fold less concentration, respectively, compared with soluble antigen alone. Collectively, these data demonstrate that autoantibody, helper, and CTL epitopes all contain a shared peptide sequence. The finding that autoantigen-immune complexes can not only cross-present but also that presentation of the autoantigen is of a higher relative efficiency, for the first time defines a unique role for autoantibodies in the pathogenesis of an autoimmune disease.

Regulatory T cells (Tregs) prevent autoimmunity but limit antitumor immunity. The canonical NF-κB signaling pathway both activates immunity and promotes thymic Treg development. Here, we report that mature Tregs continue to require NF-κB signaling through IκB-kinase β (IKKβ) after thymic egress. Mice lacking IKKβ in mature Tregs developed scurfy-like immunopathology due to death of peripheral FoxP3+ Tregs. Also, pharmacological IKKβ inhibition reduced Treg numbers in the circulation by ∼50% and downregulated FoxP3 and CD25 expression and STAT5 phosphorylation. In contrast, activated cytotoxic T lymphocytes (CTLs) were resistant to IKKβ inhibition because other pathways, in particular nuclear factor of activated T cells (NFATc1) signaling, sustained their survival and expansion. In a melanoma mouse model, IKKβ inhibition after CTL cross-priming improved the antitumor response and delayed tumor growth. In conclusion, prolonged IKKβ inhibition decimates circulating Tregs and improves CTL responses when commenced after tumor vaccination, indicating that IKKβ represents a druggable checkpoint.

The successful generation of T cell-mediated immunity for the treatment of cancer has been a major focal point of research. One of the critical strategies of cancer immunotherapy is to efficiently activate antigen-specific CD8 T cells in the immunosuppressive tumor environment. Here, we used transgenic OT-I/CD45.2/Rag−/− mice as a source of effector CD8 T cells to determine whether irradiation combined with adoptive T cell transfer therapy could improve T cell proliferation and effector function in murine tumor models. Local irradiation combined with adoptive T cell therapy showed a synergistic effect on tumor growth inhibition in mice. Mechanistically, irradiation increased the release of tumor-associated antigens, which facilitated cross-presentation of tumor-associated antigens by dendritic cells and the priming of antigen-specific T lymphocytes. Additionally, irradiation enhanced the homing of the antigen-specific T cells to tumor tissues via the increased release of CCL5, CXCL9, and CXCL11 from tumor cells. Moreover, irradiation enhanced the proliferation and effector function of both adoptively transferred T cells and endogenous antigen-specific T cells. Our findings provide evidence to support that local irradiation enhanced the therapeutic efficacy of adoptive T cell therapy for cancer, indicating that the combination of radiotherapy and adoptive T cell therapy may be a promising strategy for tumor treatment.

Epizootics of Koi herpesvirus (KHV) cause mass mortality in koi carp (Cyprinus rubrofuscus) and common carp (Cyprinus carpio) worldwide. Rapid and accurate virus detection technology is crucial for preventing pathogen spread and minimizing damage. Although several diagnostic assays have been developed for KHV, the analytical and diagnostic performance of the detection methods has not been evaluated. In this study, we developed and validated the diagnostic performance of two molecular diagnostic assays, cross-priming amplification-based lateral flow assay (CPA-LFA) and TaqMan probe-based real-time polymerase chain reaction (PCR). To detect KHV, primers and probe were designed based on the thymidine kinase (TK) genes. The detection limits of developed CPA-LFA and real-time PCR assays were determined to be 675.69 copies/μL and 8.384 copies/μL, respectively. The diagnostic sensitivity and specificity of the developed assay were determined using fish samples (n = 179). CPA-LFA was found to be 93.67% and 100%, respectively, and real-time PCR was found to be 100% and 100%, respectively. Therefore, the newly developed CPA-LFA and real-time PCR assays accurately and rapidly detect KHV. CPA-LFA is particularly suitable for point-of-care diagnosis because of its simple diagnostic process, and real-time PCR analysis is most suitable for precise diagnosis because it can detect low viral loads., Competing Interests: Declaration of Competing Interest The author declares no conflicts of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Activation of naive CD8-positive T lymphocytes is mediated by dendritic cells that cross-present MHC class I (MHC-I)-associated peptides derived from exogenous Ags. The most accepted mechanism involves the translocation of Ags from phagosomes or endolysosomes into the cytosol, where antigenic peptides generated by cytosolic proteasomes are delivered by the transporter associated with Ag processing (TAP) to the endoplasmic reticulum, or an endocytic Ag-loading compartment, where binding to MHC-I occurs. We have described an alternative pathway where cross-presentation is independent of TAP but remains dependent on proteasomes. We provided evidence that active proteasomes found within the lumen of phagosomes and endolysosomal vesicles locally generate antigenic peptides that can be directly loaded onto trafficking MHC-I molecules. However, the mechanism of active proteasome delivery to the endocytic compartments remained unknown. In this study, we demonstrate that phagosome-associated LC3A/B structures deliver proteasomes into subcellular compartments containing exogenous Ags and that autophagy drives TAP-independent, proteasome-dependent cross-presentation., (Copyright © 2024 by The American Association of Immunologists, Inc.)

Cross-presentation by type 1 DCs (cDC1) is critical to induce and sustain antitumoral CD8 T cell responses to model antigens, in various tumor settings. However, the impact of cross-presenting cDC1 and the potential of DC-based therapies in tumors carrying varied levels of bona-fide neoantigens (neoAgs) remain unclear. Here we develop a hypermutated model of non-small cell lung cancer in female mice, encoding genuine MHC-I neoepitopes to study neoAgs-specific CD8 T cell responses in spontaneous settings and upon Flt3L + αCD40 (DC-therapy). We find that cDC1 are required to generate broad CD8 responses against a range of diverse neoAgs. DC-therapy promotes immunogenicity of weaker neoAgs and strongly inhibits the growth of high tumor-mutational burden (TMB) tumors. In contrast, low TMB tumors respond poorly to DC-therapy, generating mild CD8 T cell responses that are not sufficient to block progression. scRNA transcriptional analysis, immune profiling and functional assays unveil the changes induced by DC-therapy in lung tissues, which comprise accumulation of cDC1 with increased immunostimulatory properties and less exhausted effector CD8 T cells. We conclude that boosting cDC1 activity is critical to broaden the diversity of anti-tumoral CD8 T cell responses and to leverage neoAgs content for therapeutic advantage., (© 2024. The Author(s).)

Cross-presentation, exogenous antigen presentation onto major histocompatibility complex class I molecules on antigen presenting cells, is crucially important for inducing antigen-specific cellular immune responses for cancer immunotherapy and for the treatment of infectious diseases. One strategy to induce cross-presentation is cytosolic delivery of an exogenous antigen using fusogenic or endosomolytic molecule-introduced nanocarriers. Earlier, we reported liposomes modified with pH-responsive polymers to achieve cytosolic delivery of an antigen. Polyglycidol-based or polysaccharide-based pH-responsive polymers can provide liposomes with delivery performance of antigenic proteins into cytosol via membrane fusion with endosomes responding to acidic pH, leading to induction of cross-presentation. Mannose residue was introduced to pH-responsive polysaccharides to increase uptake selectivity to antigen presenting cells and to improve cross-presentation efficiency. However, direct introduction of mannose residue into pH-responsive polysaccharides suppressed cytoplasmic delivery performance of liposomes. To avoid such interference, for this study, mannose-containing glycans were incorporated separately into pH-responsive polysaccharide-modified liposomes. Soybean agglutinin-derived glycopeptide was used as a ligand for lectins on antigen presenting cells. Incorporation of glycopeptide significantly increased the cellular uptake of liposomes by dendritic cell lines and increased cross-presentation efficiency. Liposomes incorporated both glycopeptide and pH-responsive polysaccharides exhibited strong adjuvant effects in vitro and induced the increase of dendritic cells, M1 macrophages, and effector T cells in the spleen. Subcutaneous administration of these liposomes induced antigen-specific cellular immunity, resulting in strong therapeutic effects in tumor-bearing mice. These results suggest that separate incorporation of glycopeptides and pH-responsive polysaccharides into antigen-loaded liposomes is an effective strategy to produce liposome-based nanovaccines to achieve antigen cross-presentation and induction of cellular immunity towards cancer immunotherapy.

Tumor subunit vaccines have great potential in personalized cancer immunotherapy. They are usually administered with adjuvant owing to their low immunogenicity. Cholera toxin (CT) is a biological adjuvant with diverse biological functions and a long history of use. Our earlier study revealed that a CT-like chimeric protein co-delivered with murine granulocyte-macrophage colony stimulating factor (mGM-CSF) and prostate cancer antigen epitope could co-stimulate dendritic cells (DCs) and enhance cross presentation of tumor epitope. To further study the molecular mechanism of CT-like chimeric protein in cross presentation, major histocompatibility complex class I (MHC I)-restricted epitope 257-264 of ovalbumin (OVAT) was used as a model antigen peptide in this study. Recombinant A subunit and pentameric B subunit of CT protein were respectively genetically constructed and purified. Then both assembled into AB5 chimeric protein in vitro. Three different chimeric biomacromolecules containing mGM-CSF and OVAT were constructed according to the different fusion sites and whether the endoplasmic reticulum (ER) retention sequence was included. It was found that A2 domain and B subunit of CT were both available for loading epitopes and retaining GM1 affinity. The binding activity of GM1 was positively correlated with antigen endocytosis. Once internalized, DCs became mature and cross-presented antigen. KDEL helped the whole molecule to be retained in the ER, and this improved the cross presentation of antigen on MHC I molecules. In conclusion, hexameric CT-like chimeric protein with dual effects of GM1 affinity and ER retention sequence were potential in improvement of cross presentation. The results laid a foundation for designing personalized tumor vaccine based on CT-like chimeric protein molecular structure., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Influenza virus is known to cause respiratory tract infections of varying severity in individuals of all ages. The EasyNAT Rapid Flu assay is a newly developed in vitro diagnostic test that employs cross-priming isothermal amplification (CPA) to detect and differentiate influenza A and B viruses in human nasopharyngeal (NP) swabs. The aim of this study is to determine the performance characteristics of the EasyNAT Rapid Flu assay for rapid detection of influenza virus. The limit of detection (LOD) and cross-reactivity of the EasyNAT Rapid Flu assay were assessed. The clinical performance of the assay was evaluated using NP swab samples that were tested with real-time reverse-transcription polymerase chain reaction (RT-PCR) and Xpert Xpress Flu/RSV assay. The LOD for the detection of influenza A and B using the EasyNAT Rapid Flu assay was found to be 500 copies/mL. Furthermore, the assay exhibited no cross-reactivity with other common respiratory viruses tested. For the 114 NP swab samples tested for influenza A using both the EasyNAT Rapid Flu assay and real-time RT-PCR, the two assays demonstrated a high level of agreement ( κ = 0.963, P < 0.001), with a positive percentage agreement (PPA) of 97.7% and a negative percentage agreement (NPA) of 98.6%. Similarly, for the 43 NP swab samples tested for influenza A and B using both the EasyNAT Rapid Flu assay and Xpert Xpress Flu/RSV assay, the two assays showed a high level of agreement ( κ = 0.933, P < 0.001), with the overall rate of agreement (ORA) of 97.7% for influenza A and 100% for influenza B. The EasyNAT Rapid Flu assay demonstrates excellent performance in the detection of influenza A, highlighted by its strong agreement with RT-PCR-based assays.IMPORTANCEThe newly developed EasyNAT Rapid Flu assay is an innovative cross-priming isothermal amplification-based method designed for detecting influenza A and B viruses at point-of-care settings. This study aims to thoroughly assess the analytical and clinical performance of the assay, offering valuable insights into its potential advantages and limitations. The findings of this research hold significant implications for clinical practice., Competing Interests: The authors declare no conflict of interest.

Most birds are monomorphic species and breeds, which makes it difficult to determine their gender by appearances, especially the pigeon is a farm animal with economic interest in meat production, ornamentals, sports, and experimental animals. Until now, the available methods for determining the gender of pigeons have mainly consisted of endoscopy, laparoscopy, karyotyping, polymerase chain reaction (PCR), and other similar techniques. Nonetheless, these methods have notable limitations, such as high expenses, invasiveness, and time-consuming procedures, which hinder their practicality for efficiently determining the gender of pigeons. Therefore, an easy, accurate, sensitive, on-site, affordable, and applicable rapid identification of the gender of the pigeon is widely needed for the owner of the pigeon. The purpose of this study was to develop and evaluate the efficacy of Cross-priming amplification (CPA) combined with an immune-chromatographic strip (CPA-strip) for gender identification of the pigeon. The methodology was optimized through various experimental trials. Subsequently, ten samples collected from pigeons were subjected to analysis using the optimized CPA-strip assay, and the results indicated that all female samples were accurately detected. In contrast, the blood samples collected from chickens and ducks were negative when tested with the CPA-strip assay. In conclusion, our study demonstrates the successful establishment of an immune-chromatographic CPA-strip assay for the on-site gender determination of pigeons with high accuracy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

The efficacy of radiotherapy has not yet achieved optimal results, partially due to insufficient priming and infiltration of effector immune cells within the tumor microenvironment (TME), which often exhibits suppressive phenotypes. In particular, the infiltration of X-C motif chemokine receptor 1 (XCR1)-expressing conventional type-1 dendritic cells (cDC1s), which are critical in priming CD8 + cytotoxic T cells, within the TME is noticeably restricted. Hence, we present a facile methodology for the efficient fabrication of a calcium phosphate hydrogel loaded with X-C motif chemokine ligand 1 (XCL1) to selectively recruit cDC1s. Manganese phosphate microparticles were also loaded into this hydrogel to reprogram the TME via cGAS-STING activation, thereby facilitating the priming of cDC1s propelled specific CD8 + T cells. They also polarize tumor-associated macrophages towards the M1 phenotype and reduce the proportion of regulatory cells, effectively reversing the immunosuppressive TME into an immune-active one. The yielded XCL1@CaMnP gel exhibits significant efficacy in enhancing the therapeutic outcomes of radiotherapy, particularly when concurrently administered with postoperative radiotherapy, resulting in an impressive 60 % complete response rate. Such XCL1@CaMnP gel, which recruits cDC1s to present tumor antigens generated in situ, holds great potential as a versatile platform for enhanced cancer treatment through modulating the immunosuppressive TME., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The induction of a robust CD8 + T cell response is critical for the success of an antiviral vaccine. In this study, we incorporated a STING agonist (SA) 2'3'-cGAMP into a previously developed exosome-based CVB3 viral myocarditis vaccine (Exo-VP1) to enhance its ability to induce CD8 + T cell responses and immunoprotection. Our results showed that compared to free SA adjuvant, exosome-mediated co-delivery (Exo SA -VP1) significantly enhanced SA uptake by dendritic cells (DCs) and more potently stimulated DC maturation. Immunization of mice showed that the Exo SA -VP1 vaccine-induced higher levels of CVB3-specific T cell proliferation and cytotoxicity, significantly increased the percentage of IFN-γ + CD8 + rather than CD4 + T cells, effectively reduced cardiac viral loads, attenuated myocarditis and improved survival in mice compared to the previous Exo-VP1 vaccine. Further investigation showed that Exo SA -VP1 significantly increased both the percentage and antigen cross-presentation capacity of splenic CD8 + DCs. Depletion of these CD8 + DCs by cytochrome C administration nearly abolished the advantage of Exo SA -VP1 in dominantly inducing IFN-γ + CD8 + cytotoxic T lymphocyte (CTL) production in immunized mice. Taken together, our results demonstrated the potential of Exo SA -VP1 as a promising candidate for anti-CVB3 vaccines and provide insights into immune-enhancing strategies aiming at augmenting antigen cross-presentation by DCs and enhancing potent CTL responses., Competing Interests: Declaration of competing interest The authors declared no conflict of interest., (Copyright © 2024. Published by Elsevier B.V.)

The successful generation of T cell-mediated immunity for the treatment of cancer has been a major focal point of research. One of the critical strategies of cancer immunotherapy is to efficiently activate antigen-specific CD8 T cells in the immunosuppressive tumor environment. Here, we used transgenic OT-I/CD45.2/Rag−/− mice as a source of effector CD8 T cells to determine whether irradiation combined with adoptive T cell transfer therapy could improve T cell proliferation and effector function in murine tumor models. Local irradiation combined with adoptive T cell therapy showed a synergistic effect on tumor growth inhibition in mice. Mechanistically, irradiation increased the release of tumor-associated antigens, which facilitated cross-presentation of tumor-associated antigens by dendritic cells and the priming of antigen-specific T lymphocytes. Additionally, irradiation enhanced the homing of the antigen-specific T cells to tumor tissues via the increased release of CCL5, CXCL9, and CXCL11 from tumor cells. Moreover, irradiation enhanced the proliferation and effector function of both adoptively transferred T cells and endogenous antigen-specific T cells. Our findings provide evidence to support that local irradiation enhanced the therapeutic efficacy of adoptive T cell therapy for cancer, indicating that the combination of radiotherapy and adoptive T cell therapy may be a promising strategy for tumor treatment. [ABSTRACT FROM AUTHOR]

Cross-presentation of dead cell-associated antigens by conventional dendritic cells type 1 (cDC1s) is critical for CD8+ T cells response against many tumors and viral infections. It is facilitated by DNGR-1 (CLEC9A), an SYK-coupled cDC1 receptor that detects dead cell debris. Here, we report that DNGR-1 engagement leads to rapid activation of CBL and CBL-B E3 ligases to cause K63-linked ubiquitination of SYK and terminate signaling. Genetic deletion of CBL E3 ligases or charge-conserved mutation of target lysines within SYK abolishes SYK ubiquitination and results in enhanced DNGR-1-dependent antigen cross-presentation. We also find that cDC1 deficient in CBL E3 ligases are more efficient at cross-priming CD8+ T cells to dead cell-associated antigens and promoting host resistance to tumors. Our findings reveal a role for CBL-dependent ubiquitination in limiting cross-presentation of dead cell-associated antigens and highlight an axis of negative regulation of cDC1 activity that could be exploited to increase anti-tumor immunity. [Display omitted] • CBL and CBL-B E3 ligases are recruited to the activated DNGR-1 receptor complex • CBL E3 ligases ubiquitinate SYK to terminate DNGR-1 signaling • Loss of CBL E3 ligases increases phagosomal rupture induced by DNGR-1 signals • This promotes enhanced cross-presentation of dead cell-associated antigens Henry et al. show that signaling by DNGR-1 in cDC1 is under stringent negative feedback regulation by CBL E3 ligases. The latter ubiquitinate the key adaptor SYK to terminate DNGR-1 signaling. cDC1s lacking CBL family E3 ligases are more potent at cross-presentation of dead cell-associated antigens. [ABSTRACT FROM AUTHOR]

Dendritic cells (DCs) play a central role in the regulation of the balance between CD8 T cell immunity vs. tolerance to tumor antigens. Cross-priming, a process which DCs activate CD8 T cells by cross-presenting exogenous antigens, plays a critical role in generating anti-tumor CD8 T cell immunity. However, there are compelling evidences now that the tumor microenvironment (TME)-mediated suppression and modulation of tumor-infiltrated DCs (TIDCs) impair their function in initiating potent anti-tumor immunity and even promote tumor progression. Thus, DC-mediated cross-presentation of tumor antigens in tumor-bearing hosts often induces T cell tolerance instead of immunity. As tumor-induced immunosuppression remains one of the major hurdles for cancer immunotherapy, understanding how DCs regulate anti-tumor CD8 T cell immunity in particular within TME has been under intensive investigation. Recent reports on the Batf3-dependent type 1 conventional DCs (cDC1s) in anti-tumor immunity have greatly advanced our understanding on the interplay of DCs and CD8 T cells in the TME, highlighted by the critical role of CD103+ cDC1s in the cross-priming of tumor antigen-specific CD8 T cells. In this review, we will discuss recent advances in anti-tumor CD8 T cell cross-priming by CD103+ cDC1s in TME, and share perspective on future directions including therapeutic applications and memory CD8 T cell responses.

Dendritic cells (DCs) are professional antigen-presenting cells that play a key role in shaping adaptive immunity. DCs have a unique ability to sample their environment, capture and process exogenous antigens into peptides that are then loaded onto major histocompatibility complex class I molecules for presentation to CD8 + T cells. This process, called cross-presentation, is essential for initiating and regulating CD8 + T cell responses against tumors and intracellular pathogens. In this review, we will discuss the role of DCs in cancer immunity, the molecular mechanisms underlying antigen cross-presentation by DCs, the immunosuppressive factors that limit the efficiency of this process in cancer, and approaches to overcome DC dysfunction and therapeutically promote antitumoral immunity., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The conventional type 1 dendritic cells (cDC1) play a pivotal role in protective immunity against pathogens and cancer. However, their low frequency in the blood and tissues limits their use in immune therapy. We have recently described a method to vaccinate against neoantigens that are induced in tumor cells by targeted delivery of a TAP siRNA to dendritic cells using a TLR9 binding CpG oligonucleotide. Since TLR9 is also expressed in immune suppressive myeloid populations TLR9 targeting could reduce the effectiveness of this approach. Here, we describe a modular multivalent antibody platform to target the TAP siRNA to resident Clec9a expressing cDC1 and show that it leads to selective and sustained TAP downregulation in cDC1 and inhibits tumor growth in mice more effectively than CpG targeted siRNA. To induce DC maturation an agonistic CD40 antibody was administered to the siRNA treated mice. To obviate the need for a second drug formulation and reduce the risk of toxicity, we exploited the multivalent nature of this targeting platform to co-deliver the TAP siRNA and a DC maturation agent, a CpG containing oligonucleotide, to cDC1 in vivo and show that it was more effective than Clec9a targeting of TAP siRNA in combination with CD40 antibody. This study describes a way to manipulate the function of cDC1 cells in vivo using a broadly applicable antibody-based targeting platform to deliver multiple biological agents to specific cells in vivo to potentiate (immune) therapy and to probe the biology of specific cell types in their natural settings., (© 2024. The Author(s).)

Importance: In this study, we successfully established a new One-Pot method, named TB One-Pot, for detecting Mtb in sputum by combining CRISPR-cas12b-mediated trans-cleavage with cross-priming amplification (CPA). Our study evaluated the diagnostic performance of TB One-Pot in clinical sputum samples for tuberculosis. The findings provide evidence for the potential of TB One-Pot as a diagnostic tool for tuberculosis., Competing Interests: The authors declare no conflict of interest.

Antigen cross-priming of CD8 + T cells is a critical process necessary for the effective expansion and activation of CD8 + T cells endowed with the ability to recognize and destroy tumor cells. The cross-presentation of tumor antigens to cross-prime CD8 + T cells is mainly mediated, if not only, by a subset of professional antigen-presenting cells termed type-1 conventional dendritic cells (cDC1). The demise of malignant cells can be immunogenic if it occurs in the context of premortem stress. These ways of dying are termed immunogenic cell death (ICD) and are associated with biochemical features favoring cDC1 for the efficient cross-priming of tumor antigens. Immunosurveillance and the success of immunotherapies heavily rely on the ability of cytotoxic immune cells, primarily CD8 + T cells and NK cells, to detect and eliminate tumor cells through mechanisms collectively known as cytotoxicity. Recent studies have revealed the significance of NK- and CTL-mediated cytotoxicity as a prominent form of immunogenic cell death, resulting in mechanisms that promote and sustain antigen-specific immune responses. This review focuses on the mechanisms underlying the cross-presentation of antigens released during tumor cell killing by cytotoxic immune cells, with an emphasis on the role of cDC1 cells. Indeed, cDC1s are instrumental in the effectiveness of most immunotherapies, underscoring the significance of tumor antigen cross-priming in contexts of immunogenic cell death. The notion of the potent immunogenicity of cell death resulting from NK or cytotoxic T lymphocyte (CTL)-mediated cytotoxicity has far-reaching implications for cancer immunotherapy., (© 2023 The Authors. Immunological Reviews published by John Wiley & Sons Ltd.)

NKT cells are unconventional T cells whose biological role is incompletely understood. Similar to TH cells, activated NKT cells can cause dendritic cell (DC) maturation, which is required for effective CTL responses. However, it is unclear whether and how NKT cells affect CTLs downstream of the DC maturation phase. This is partially due to the lack of techniques to conditionally deplete NKT cells in vivo. To overcome this problem, we have developed two approaches for this purpose in mice: the first is based on mixed bone marrow chimeras where Jα18 knockout and depletable CD90 congenic bone marrow is combined, and the second used PLZFCre × iDTR bone marrow chimeras, which target innate-like T cells. Using these tools, we found that NKT cell depletion at 20 h, that is, after initial DC activation, did not render CTLs helpless, as CD40L signaling by non-NKT cells sufficed. Instead, NKT cell depletion even augmented CD8 T cell expansion and cytotoxicity by mechanisms distinct from reduced STAT6 signaling. These findings revealed a negative feedback loop by which NKT cells control CTL cross-priming downstream of DC maturation. The techniques described in this study expand the toolbox to study NKT cells and other unconventional T cell subsets in vivo and uncovered a hidden immunoregulatory mechanism., (Copyright © 2023 by The American Association of Immunologists, Inc.)

The major histocompatibility complexes of vertebrates play a key role in the immune response. Antigen-presenting cells are loaded on MHC I molecules, which mainly present endogenous antigens; when MHC I presents exogenous antigens, this is called cross-presentation. The discovery of cross-presentation provides an important theoretical basis for the study of exogenous antigens. Cross-presentation is a complex process in which MHC I molecules present antigens to the cell surface to activate CD8 + T lymphocytes. The process of cross-representation includes many components, and this article briefly outlines the origins and development of MHC molecules, gene structures, functions, and their classical presentation pathways. The cross-presentation pathways of MHC I molecules, the cell lines that support cross-presentation, and the mechanisms of MHC I molecular transporting are all reviewed. After more than 40 years of research, the specific mechanism of cross-presentation is still unclear. In this paper, we summarize cross-presentation and anticipate the research and development prospects for cross-presentation.