InVivoMAb rat IgG1 isotype control, anti-horseradish peroxidase

Enterovirus D68 (EV-D68) causes respiratory illness in children. It also causes severe paralysis called acute flaccid myelitis (AFM), which has become a global health threat. Here, we generated an mRNA vaccine expressing virus-like particles (VLPs) of EV-D68. We found that the mRNA vaccine elicited potent neutralizing antibodies against EV-D68 in the blood, and the neutralizing titer was superior to that of the inactivated whole virion (IWV) vaccine. The mRNA vaccine showed protective effects against intranasal challenge with EV-D68, and antisera from the vaccinated mice prevented the paralysis caused by EV-D68 infection in neonatal mice. Moreover, the mRNA vaccine induced neutralizing antibodies in the respiratory tract, which is the entry site for EV-D68. Additionally, it attenuated infection with coxsackievirus B3 (CVB3), which belongs to another enterovirus group, via CD8+ T cell responses. In conclusion, our results suggest that this mRNA vaccine is a promising candidate for EV-D68 prevention.

This study investigates the functions of chromobox 6 (CBX6) in esophageal squamous cell carcinoma (ESCC) and delves into its functional mechanisms. The bioinformatics insights suggested that CBX6 was overexpressed in ESCC and linked to dismal prognosis. Cbx6 knockdown was induced in mouse mEC25 cells. This procedure curbed the proliferation and migration of mEC25 cells and reduced exhaustion of the co-cultured CD8+ T cells. In vivo, Cbx6 knockdown in mEC25 cells reduced tumorigenesis while enhancing immune activity in mice. Further experiments showed that CBX6 reduced CD8+ T cell cytotoxicity by secreting C-C motif chemokine ligand 8 (CCL8) and promoting monocarboxylate transporter 4 (MCT4)-mediated lactate transport. Regarding the mechanism, CBX6 regulated the expression of SWI/SNF related BAF chromatin remodeling complex subunit D1 (Smarcd1) to modulate chromatin remodeling, thus promoting transcription of Ccl8 and Slc16a3 (encoding MCT4). Smarcd1 overexpression restored metabolic activity in mEC25 cells, reduced activity of co-cultured CD8+ T cells, and promoted tumorigenesis in vivo. Tissue microarrays analysis suggested that CBX6 and SMARCD1 were linked to immunosuppression and poor prognosis in clinical samples. In conclusion, this study suggests that CBX6 induces CD8+ T cell exhaustion and tumor development in ESCC through SMARCD1-mediated CCL8 secretion and lactate efflux.

Prenatal maternal immune activation (MIA) has been implicated in autism spectrum disorder (ASD) pathogenesis, with interleukin-6 (IL-6) identified as a key inflammatory mediator. We investigated the therapeutic potential of IL-6 inhibition in an MIA mouse model induced by Toxoplasma gondii soluble tachyzoite antigen (STAg). Adult MIA offspring received systemic administration of the IL-6-neutralizing antibody (MP5-20F3) or isotype control, followed by behavioral assessments one week later. Open field and elevated plus maze tests revealed heightened anxiety-like behaviors in the STAg offspring, which were largely reversed by IL-6 inhibition. Reciprocal social interaction tests showed diminished sociability in the STAg offspring, which was partially restored by IL-6 inhibition. However, core ASD-like features, including impaired social preference and recognition in the three-chamber test, as well as increased repetitive behaviors, remained resistant to IL-6 inhibition. These findings demonstrate that STAg-induced MIA elicits anxiety-like and ASD-like phenotypes in adult offspring, with IL-6 playing an important role in anxiety-like behaviors and social interaction deficits. Systemic IL-6 inhibition partially ameliorates behavioral abnormalities. This study suggests that IL-6-targeted therapies may address a subset of ASD-related symptoms, and comprehensive strategies are needed for broader efficacy.

The ERBB receptor family is widely implicated in epithelial malignancies, yet the functional and immunological significance of ERBB3 in head and neck squamous cell carcinoma (HNSCC) remains insufficiently characterized. In this study, we employed single-cell and bulk transcriptomic analyses to comprehensively map the expression patterns and prognostic value of ERBB family members in HNSCC, identifying ERBB3 as a tumor-specific marker predominantly enriched in malignant epithelial clusters. Notably, high ERBB3 expression was paradoxically associated with favorable overall survival, prompting further mechanistic investigation. Functional assays in SCC9 cells demonstrated that ERBB3 promotes proliferation, colony formation, and invasion. Immune profiling revealed that ERBB3-high tumors displayed enhanced communication with B cell subsets, particularly involving immunosuppressive signals such as TNFRSF13B and IL4R. Flow cytometry analysis in a 4NQO-induced mouse model showed that ERBB3 inhibition reduced CCDC50⁺ B cells while restoring MHC-II expression, indicating a shift toward immune activation. These findings highlight a dual role of ERBB3 in HNSCC, acting both as an oncogenic contributor and an immune-modulatory regulator, and position ERBB3 as a promising context-dependent therapeutic target.

Targeted head and neck irradiation (HNI) used for cancer therapy causes mucosal damage and immune dysregulation, leaving cancer patients highly susceptible to infections of the oral mucosa. Oropharyngeal candidiasis (OPC) is an opportunistic infection caused by Candida albicans, a commensal fungus found in up to 80% of the population at any given time. High colonization levels of C. albicans are known to worsen damage caused by HNI. The interleukin-23 (IL-23)-T-helper 17 (Th17) axis is a central and non-redundant mediator of immunity to OPC, and anti-cytokine biologics targeting IL-23 have come into widespread clinical use for treating various autoimmune conditions. Here, we sought to understand the consequences of IL-23 deficiency in the setting of HNI, taking advantage of a mouse model of radiation-induced oral mucositis (OM) which we combined with fungal infection. Surprisingly, mice lacking IL-23 did not show increased signs of injury to the oral mucosa when subjected to HNI. However, in mice subjected to HNI and exposed to oral C. albicans, damage to the oral mucosa was markedly exacerbated, accompanied by substantially increased fungal susceptibility when IL-23 was absent. Thus, IL-23-driven control of fungal infections is needed to mitigate susceptibility to OM in the high proportion of individuals who carry C. albicans in the mouth.IMPORTANCEIL-23 plays key roles in expanding the T-helper 17 (Th17) cell subset during differentiation and promoting expression of cytokines, including the subset-defining cytokine IL-17 (IL-17A). While the effector cytokines produced by Th17 cells are required for host defense against extracellular microbes, especially Candida albicans, these can also be pathogenic during inflammatory and autoimmune disorders including psoriasis, psoriatic arthritis, and ankylosing spondylitis, among others. While IL-23 and IL-17 are similarly required for protection against oropharyngeal candidiasis (OPC), they can exert divergent functions in other forms of immune-mediated inflammation; for example, anti-IL-17 blockade or Il17ra gene deficiency is linked to inflammatory bowel disease, whereas loss of IL-23 is protective in this setting. We previously showed that head and neck irradiation (HNI) induces IL-17 expression in oral tissue and that IL-17 receptor (IL-17R) signaling is required for resistance against mucosal damage and OPC. In contrast, we show here that loss of IL-23 does not affect oral mucosal injury caused by HNI. However, lack of IL-23 magnifies HNI-induced susceptibility to OPC due to insufficient levels of antimicrobial peptides. Clinically, these findings suggest that patients receiving treatments that target IL-23 may not need to discontinue therapy should they require HNI, but that screening for oral C. albicans may be useful to help limit the risk of developing severe OM and its attendant adverse events.

Iron loading increases infection risk in being a nutrient for invading siderophilic bacteria and by modulating immune functions including the expression of the immune checkpoint regulator T-cell immunoglobulin-and-mucin-containing-domain-3 (TIM-3). TIM-3 affects specific immune cell functions including T-helper cell differentiation but also T cell dysfunction, and immune exhaustion. Given the prevalence of iron overload specifically in patients at higher risk for infection such as those suffering from hemo-oncological diseases, we investigated TIM-3's role in immune control of bacterial sepsis.

B-1 cells are innate-like immune cells abundant in serosal cavities with antibodies enriched in bacterial recognition, yet their existence in humans has been controversial1-3. The CD5+ B-1a subset expresses anti-inflammatory molecules including IL-10, PDL1 and CTLA4 and can be immunoregulatory4-6. Unlike conventional B cells that are continuously replenished, B-1a cells are produced early in life and maintained through self-renewal7. Here we show that the transcription factors TCF1 and LEF1 are critical regulators of B-1a cells. LEF1 expression is highest in fetal and bone marrow B-1 progenitors, whereas the levels of TCF1 are higher in splenic and peritoneal B-1 cells than in B-1 progenitors. TCF1-LEF1 double deficient mice have reduced B-1a cells and defective B-1a cell maintenance. These transcription factors promote MYC-dependent metabolic pathways and induce a stem-like population upon activation, partly via IL-10 production. In the absence of TCF1 and LEF1, B-1 cells proliferate excessively and acquire an exhausted phenotype with reduced IL-10 and PDL1 expression. Furthermore, adoptive transfer of B-1 cells lacking TCF1 and LEF1 fails to suppress brain inflammation. These transcription factors are also expressed in human chronic lymphocytic leukaemia B cells and in a B-1-like population that is abundant in pleural fluid and circulation of some patients with pleural infection. Our findings define a TCF1-LEF1-driven transcriptional program that integrates stemness and regulatory function in B-1a cells.

Cancer cachexia (CC), a syndrome of skeletal muscle and adipose wasting, reduces responsiveness to therapies and increases mortality. There are no approved treatments for CC, which may relate to discordance between preclinical models and human CC. To address the need for clinically relevant models of lung CC, we generated inducible, lung epithelial cell-specific KrasG12D/+ (G12D) mice. G12D mice develop CC over a protracted time course and phenocopy tissue and tumor, cellular, mutational, transcriptomic, and metabolic characteristics of human lung CC. G12D mice demonstrate early loss of adipose, a phenotype that was apparent across numerous models of CC and translates to patients with lung cancer. Tumor-released factors promote adipocyte lipolysis, a driver of adipose wasting in CC, and adipose wasting was inversely related to tumor burden. Thus, G12D mice model key features of human lung CC and highlight a role for early tumor metabolic reprogramming of adipose tissue in CC.

Bacteria of the genus Shigella replicate in intestinal epithelial cells and cause shigellosis, a severe diarrheal disease that resolves spontaneously in most healthy individuals. During shigellosis, neutrophils are abundantly recruited to the gut and have long been thought to be central to Shigella control and pathogenesis. However, how shigellosis resolves remains poorly understood due to the longstanding lack of a tractable and physiological animal model. Here, using our newly developed Nlrc4-/-Casp11-/- mouse model of shigellosis, we unexpectedly find no major role for neutrophils in limiting Shigella or in disease pathogenesis. Instead, we uncover an essential role for macrophages in the host control of Shigella. Macrophages respond to Shigella via Toll-like receptors (TLRs) to produce IL-12, which then induces IFN-γ, a cytokine that is essential to control Shigella replication in intestinal epithelial cells. Collectively, our findings reshape our understanding of the innate immune response to Shigella.

Subclinical parasitemia constitutes the predominant proportion of Plasmodium spp. infections in hyperendemic regions of the world. Elevated levels of serum interleukin-10 (IL-10) are observed in both acute symptomatic and chronic subclinical Plasmodium spp. infections. The role of IL-10 in acute infection has been extensively studied; however, the role of sustained elevated levels of IL-10 in chronic subclinical plasmodial infections remains to be determined. We investigated the role of IL-10 in a long-term subclinical and patent Plasmodium chabaudi chabaudi-AS (Pc) infection using mice lacking humoral immunity (µMT-/- mice). Pc-infected µMT-/- mice exhibit a long-term (99 days) chronic infection, with microscopic levels of parasitemia and without any outward signs of disease. We found that chronically infected mice have slightly elevated levels of tumor necrosis factor α (TNFα) and interferon-γ (IFNγ), and high levels of IL-10 in the circulation. The source of IL-10 was CD4+ T cells. We found that elevated IL-10 levels were mechanistically linked to subclinical Plasmodium infection by blocking IL-10 signaling. Anti-IL-10R resulted in a marked, albeit transient, reduction of the parasitemia that was accompanied by a robust pro-inflammatory response and death of chronically infected µMT-/- mice. A similar outcome was observed in infected µMT-/- mice after CD4+ T cell depletion with anti-CD4 antibody. CD4-depleted infected µMT-/- mice exhibited reduced IL-10 and rapid weight loss, succumbing to infection by day 6 after CD4 neutralization. Our results showed that IL-10 from CD4+ T cells limits immune-mediated pathology in chronic subclinical Pc infection in µMT-/- mice by protecting against excessive inflammatory responses to blood-stage parasites.

Wild-type interleukin (IL)-2 induces anti-tumor immunity and toxicity, predominated by vascular leak syndrome (VLS) leading to edema, hypotension, organ toxicity, and regulatory T cell (Treg) expansion. Efforts to uncouple IL-2 toxicity from its potency have failed in the clinic. We hypothesize that IL-2 toxicity is driven by cytokine release syndrome (CRS) followed by VLS and that coupling IL-2 with IL-10 will ameliorate toxicity. Our data, generated using human primary cells, mouse models, and non-human primates, suggest that coupling of these cytokines prevents toxicity while retaining cytotoxic T cell activation and limiting Treg expansion. In syngeneic murine tumor models, DK210 epidermal growth factor receptor (EGFR), an IL-2/IL-10 fusion molecule targeted to EGFR via an anti-EGFR single-chain variable fragment (scFV), potently activates T cells and natural killer (NK) cells and elicits interferon (IFN)γ-dependent anti-tumor function without peripheral inflammatory toxicity or Treg accumulation. Therefore, combining IL-2 with IL-10 uncouples toxicity from immune activation, leading to a balanced and pleiotropic anti-tumor immune response.

Gastric cancers are classified into four molecular subtypes according to The Cancer Genome Atlas (TCGA) classification: Epstein-Barr virus-positive (EBV-positive), microsatellite instability-high (MSI-H), chromosomal instability (CIN), and genomically stable (GS). Unlike MSI-H gastric cancer, GS and CIN subtypes exhibit immunologically inert microenvironments and demonstrate minimal response to immune checkpoint blockade (ICB), necessitating novel strategies to overcome immunotherapy resistance.

Immunotherapy targeting negative immune checkpoint regulators to enhance the anti-tumour immune response holds promise in the treatment of TNBC. V-domain Ig suppressor of T-cell activation (VISTA) is an immune checkpoint molecule, known to be upregulated and involved in modulating tumour immunity in TNBC. However, how VISTA affects immune response and its therapeutic potential in TNBC remains unclear.

The lack of T cells in tumors is a major hurdle to successful immune checkpoint therapy (ICT). Therefore, therapeutic strategies promoting T cell recruitment into tumors are warranted to improve the treatment efficacy. Here, we report that Fc-optimized anti-cytotoxic T lymphocyte antigen 4 (CTLA-4) antibodies are potent remodelers of tumor vasculature that increase tumor-associated high endothelial venules (TA-HEVs), specialized blood vessels supporting lymphocyte entry into tumors. Mechanistically, this effect is dependent on the Fc domain of anti-CTLA-4 antibodies and CD4+ T cells and involves interferon gamma (IFNγ). Unexpectedly, we find that the human anti-CTLA-4 antibody ipilimumab fails to increase TA-HEVs in a humanized mouse model. However, increasing its Fc effector function rescues the modulation of TA-HEVs, promotes CD4+ and CD8+ T cell infiltration into tumors, and sensitizes recalcitrant tumors to programmed cell death protein 1 (PD-1) blockade. Our findings suggest that Fc-optimized anti-CTLA-4 antibodies could be used to reprogram tumor vasculature in poorly immunogenic cold tumors and improve the efficacy of ICT.

The SARS-CoV-2 pandemic highlighted the potential of mRNA vaccines in rapidly responding to emerging pathogens. However, immunity induced by conventional mRNA vaccines wanes quickly, requiring frequent boosters. Self-amplifying RNA (saRNA) vaccines, which extend antigen expression via self-replication, offer a promising strategy to induce more durable immune responses. In this study, we developed an saRNA vaccine encoding Zika virus (ZIKV) membrane and envelope proteins and evaluated its efficacy in mice. A single vaccination elicited strong humoral and cellular immune responses and reduced viral loads but only for 28 days. By day 84, antibody titers and T cell responses had significantly declined, resulting in reduced efficacy. To address this, we evaluated agonist antibodies targeting the T cell costimulatory molecules OX40 and 4-1BB. Coadministration of agonist antibodies enhanced CD8+ T cell responses to vaccination, resulting in sustained immunity and reduced viral loads at day 84. Depletion and passive transfer studies verified that long-term antiviral immunity was primarily CD8+ T cell dependent, with minimal contributions from antibody responses. These findings suggest that agonists targeting members of the tumor necrosis receptor superfamily, such as OX40 and 4-1BB, might enhance the durability of saRNA vaccine-induced protection, addressing a key limitation of current mRNA vaccine platforms.

The liver is an important metastatic organ that contains many innate immune cells, yet little is known about their role in anti-metastatic defense. We investigated how invariant natural killer T (iNKT) cells influence colorectal cancer-derived liver metastasis using different models in immunocompetent mice. We found that hepatic iNKT cells promote metastasis by creating a supportive niche for disseminated cancer cells. Mechanistically, iNKT cells respond to disseminating cancer cells by producing the fibrogenic cytokines interleukin-4 (IL-4) and IL-13 in a T cell receptor-independent manner. Selective abrogation of IL-4 and IL-13 sensing in hepatic stellate cells prevented their transdifferentiation into extracellular matrix-producing myofibroblasts, which hindered metastatic outgrowth of disseminated cancer cells. This study highlights a novel tumor-promoting axis driven by iNKT cells in the initial stages of metastasis.

IKZF2 (Helios) is a transcription factor that is selectively expressed by Tregs and is essential for preserving the function and stability of Tregs in the tumor microenvironment (TME), where it suppresses the anti-tumor immune response. Targeted IKZF2 degradation by small molecules represents a promising strategy for the development of a new class of cancer immunotherapy. Herein, we describe the discovery of PVTX-405, a potent, effective, highly selective, and orally efficacious IKZF2 molecular glue degrader. PVTX-405 degrades IKZF2 (DC50 = 0.7 nM and Dmax = 91%) while sparing other CRBN neo-substrates. Degradation of IKZF2 by PVTX-405 increases production of inflammatory cytokine IL-2 and reduces the suppressive activity of Tregs, leading to an increase in Teff cell proliferation. Once-daily oral administration of PVTX-405 as single agent significantly delays the growth of MC38 tumors in a syngeneic tumor model using humanized CRBN mice. PVTX-405 in combination with anti-PD1 or anti-LAG3 significantly increases animal survival compared to anti-PD1 or anti-LAG3 alone. Together, these results demonstrate that PVTX-405 is a promising IKZF2 degrader for clinical development for the treatment of human cancers.

The rapid onset of innate immune defenses is critical for early control of viral replication in an infected host and yet it can also lead to irreversible tissue damage, especially in the respiratory tract. Sensitive regulators must exist that modulate inflammation, while controlling the infection. In the present study, we identified acetylcholine (ACh)-producing B cells as such early regulators. B cells are the most prevalent ACh-producing leukocyte population in the respiratory tract demonstrated with choline acetyltransferase (ChAT)-green fluorescent protein (GFP) reporter mice, both before and after infection with influenza A virus. Mice lacking ChAT in B cells, disabling their ability to generate ACh (ChatBKO), but not those lacking ChAT in T cells, significantly, selectively and directly suppressed α7-nicotinic-ACh receptor-expressing interstitial, but not alveolar, macrophage activation and their ability to secrete tumor necrosis factor (TNF), while better controlling virus replication at 1 d postinfection. Conversely, TNF blockade via monoclonal antibody treatment increased viral loads at that time. By day 10 of infection, ChatBKO mice showed increased local and systemic inflammation and reduced signs of lung epithelial repair despite similar viral loads and viral clearance. Thus, B cells are key participants of an immediate early regulatory cascade that controls lung tissue damage after viral infection, shifting the balance toward reduced inflammation at the cost of enhanced early viral replication.