bluuebunny/crossref_metadata_2025 huggingface dataset:
ILC2-driven innate immune checkpoint mechanism antagonizes NK cell anti-metastatic function in the lung.
Authors: Halim Timotheus You Fu , Schuijs Martijn , Png Shaun , Richard Arianne , Tsyben Anastasia , Hamm Gregory , Stockis Julie , Garcia Celine , Pinaud Silvain , Nichols Ashley , Romero-Ros Xavier , Shields Jacqueline , Cohen E. Suzanne , McKenzie Andrew , Goodwin Richard , Su Jing , Eldridge Matthew , Riedel Angela , Serrao Eva , Brindle Kevin , Marioni John
Published: February 2023

Metastasis constitutes the primary cause of cancer-related deaths, with the lung being a commonly affected organ. We found that activation of lung-resident group 2 innate lymphoid cells (ILC2s) orchestrated suppression of natural killer (NK) cell-mediated innate antitumor immunity, leading to increased lung metastases and mortality. Using multiple models of lung metastasis, we show that interleukin (IL)-33-dependent ILC2 activation in the lung is involved centrally in promoting tumor burden. ILC2-driven innate type 2 inflammation is accompanied by profound local suppression of interferon-γ production and cytotoxic function of lung NK cells. ILC2-dependent suppression of NK cells is elaborated via an innate regulatory mechanism, which is reliant on IL-5-induced lung eosinophilia, ultimately limiting the metabolic fitness of NK cells. Single-cell and bulk transcriptomic analysis of lung NK cells reveals that post-transcriptional regulation likely accounts for IL-33-ILC2 dependent suppression of NK cell function. Using mass-spectrometry imaging (MSI) of the lungs of 13 C-Glucose infused mice, and metabolite profiling of ex vivo lung cultures, we find that IL-33 drives localized depletion of glucose via ILC2-dependent recruitment of highly glycolytic eosinophils, which can directly suppress lung NK cell function. Therapeutic targeting of IL-33 or IL-5 reversed NK cell suppression and alleviated cancer burden. Thus, we reveal an important function of IL-33 and ILC2s in promoting tumor metastasis via their capacity to suppress innate type 1 immunity.
Global inhibition of the interaction of NK inhibitory receptors with MHC-I augments coordinated innate and adaptive immunity against cancer metastasis.
Authors: Panda Abir Kumar , Sinha Surajit , Natarajan Kannan , Hernandez Jonathan M. , Margulies David H. , Shevach Ethan M.
Published: February 2023

Natural Killer (NK) cells are innate lymphocytes involved in the first line of immune defense and T cell adaptive immunity against viral infection and cancer, including metastasis. While on patrol, NK cells contact other cells and recognize MHC-I Ags via stochastically expressed MHC-I–specific inhibitory receptors (Ly49s in mice and KIRs in humans) that prevent NK cell activation via cytoplasmic ITIM. The binding site on MHC-Class-I for Ly49 inhibitory receptors is distinct from that for TCRs. The loss of MHC-I expression on tumor cells (“missing self”) abrogates inhibitory signals, resulting in NK activation. Global inhibition of the NK inhibitory receptor interactions in vivo by a pan-anti-MHC-I monoclonal antibody markedly activated IFNg-producing NK cells, independent of Fc receptors. NK cell-derived IFNg, along with other cytokines (MCSF & FLT3L), primed APC to induce IL-12/-15/-18/-21 cytokine cascades and enhanced levels of MHC-I and MHC-II expression that further drove the proliferation of NK cells and memory phenotype (MP) T cells. The global disruption of NK cell/MHC-I interactions significantly enhanced Th1 type signature transcription factors (Tbet & Eomes), cytokines (IFNg & Granzyme B), and chemokine receptors (CXCR3) on NK and MP T cells. Administration of pan-anti-MHC-I to unmanipulated mice profoundly augmented innate and adaptive immunity against viral infection, PD1-resistant transplanted tumors, and successfully constrained lung and liver metastasis, and protected the animals from tumor burden. Moreover, in vitro enhancement of human NK cell proliferation by a pan-anti-HLA Mab suggests that pan-anti-HLA could be a promising therapeutic strategy against chronic viral infection and cancer metastasis.
CD122-SELECTIVE IL-2 COMPLEXES TREAT OVARIAN CARCINOMAS, INDUCE TREG FRAGILITY AND PROMOTE T CELL STEM CELLS
Authors: Deng Yilun , Drerup Justin Michael , Zhang Xinyue , Reyes Ryan M , Mendez Jenny , Raeber Miro E , Garcia Myrna G , Pardon Alvaro S , Gupta Harshita B. , Boyman Onur , Curiel Tyler J
Published: February 2023

Regulatory T cells (Treg) engage IL-2 by high affinity CD25, and anti-tumor effector T cells (Teff) use intermediate affinity CD122. We studied IL-2 complexes (IL-2c) that selectively activate CD122+ Teff over CD25+ Tregs. We found IL-2c but not aPD-L1 potently inhibits ID8agg, an aggressive mouse ovarian cancer (OC) model. IL-2c decreased ascites Treg functional markers (CD25, GranzymeB) while upregulating them on Teffs. IL-2c inhibited Treg suppressive function in ascites but not TDLN. Ascites Tregs after IL-2c showed a fragile phenotype (increased PD-1, T-bet, and IFNg with maintained FoxP3) known to contribute to better immunotherapy response. These data suggest IL-2c inhibits Treg in the ascites through inducing fragile Tregs. Neither CD8+ T cells nor gd T cells is required for IL-2c induced Treg fragility. Studies testing other populations are ongoing. CD8+TCF-1+T cell stem cells (TCSC) improve immune checkpoint blockade in many cancer types. In ID8agg tumors, IL-2c induces CD8+TCF-1+ TCSC with tumor inhibitory capacity when transferred into Rag KO mice. tSNE analysis revealed IL-2c-induced TCSC are CXCR5+PD-1− express CCR2 CXCR3 and produce TNFa, while aPD-L1 induced TCSC are CXCR5+ and PD-1+, consistent with previous reports. These data suggest IL-2c induces beneficial CD8+TCSC distinct from aPD-L1. Although aPD-L1 is not efficacious alone in ID8agg, it significantly promoted IL-2c efficacy. Mice treated with IL-2c+αPD-L1 are resistant to tumor re-challenge even 6 weeks after final dose, suggesting durable immune memory. In sum, IL-2c is a novel OC immunotherapy that targets distinct immune pathways simultaneously, including inhibiting Tregs, boosting TCSCs, and inducing immune memory when combined with aPD-L1.
CaMKK2 deletion causes increased accumulation of CD4+ TILs and an improved ratio of effector memory TILs to Tregs
Authors: Chakraborty Molly Antara , Tomaszewski William H , Polania Jessica Waibl , Racioppi Luigi , Sanchez-Perez Luis A , Gunn Michael D , Sampson John H
Published: February 2023

Objective

Glioblastoma (GBM) is the most lethal primary brain tumor in adults. Although immunotherapy has been successful in some cancers, it has shown limited success in GBM. This is partially due to poor T cell infiltration, and because many T cells found in tumors are either exhausted or Tregs. Re-analysis of publicly available data has shown that CaMKK2 is more highly expressed in advanced brain tumors, and high CaMKK2 expression results in worse survival outcomes in GBM. Additionally, in survival studies that we performed, CaMKK2−/− mice showed an impressive survival phenotype that was dependent on the presence of CD8+ tumor infiltrating lymphocytes (TILs). The objective of our study was to investigate what is different about T cells found in the TME of CaMKK2−/− mice, and if this can explain the observed survival benefit.

Methods

We performed flow cytometry to determine the identities of T cells found in the tumors of CaMKK2−/− and WT mice.

Results/Conclusions

Our experiments show that there was a greater percentage of CD4+ TILs relative to CD8+ TILs in CaMKK2−/− mice, and this increase in percentage was due to increased accumulation of CD4+ TILs. Additionally, we found that there was greater accumulation of CD4+ and CD8+ effector memory TILs and a greater ratio of effector memory TILs to Tregs in CaMKK2−/− mice. The increased accumulation of the CD8+ effector memory TILs is particularly interesting because it is not simply explained by an increased accumulation of cells, as is the case with the CD4+ TILs. The increased accumulation of CD4+ TILs and greater ratio of effector memory TILs to Tregs is expected to have an anti-tumor effect. These findings make CaMKK2 a promising target to treat GBM.
Designing vaccines to combat cancer – an Indigenous approach
Authors: Sika-Paotonu Dianne
Published: February 2023

Pacific Peoples living in Aotearoa/New Zealand and the Pacific Region remain disproportionately affected by some cancers, with the growing burden of cancer globally remaining a significant health concern. Immunotherapy based therapeutic approaches hold some potential towards addressing inequitable cancer outcomes, treatment and management efforts for vulnerable communities, but requires appropriate inclusion and engagement with Pacific communities as part of the research process, consultation and understanding of the cultural context and clinical setting for vulnerable communities. This work sought to apply the use of an Indigenous Pacific framework to promote and support the potential applicability of immunotherapy based cancer research work within vulnerable communities adversely impacted by inequitable cancer outcomes. The Indigenous Pacific Kakala research framework was applied within this setting and supported engagement and research translation efforts and dissemination for researchers and Pacific communities in Aotearoa/New Zealand that was applicable, respectful, meaningful and appropriate to the cultural context for Pacific Peoples. This work highlights the importance of incorporating Indigenous frameworks to support Immunology based research work that is relevant, applicable and also appropriate within cultural settings.
The Impact of Advanced Age on Vaccine-Induced Tissue Immune Recall Responses
Authors: Scordo Julia M , Piergallini Tucker J , Headley Colwyn A , Reuter Nicole , Hodara Vida L , Gonzalez Olga , Giavedoni Luis D , Papin James F , Turner Joanne
Published: February 2023

Infectious diseases account for one-third of deaths in individuals over the age of 65. Vaccines are a primary tool to combat infection, yet they are less effective in the aged population. Many have focused on vaccine-induced peripheral blood responses in old age; however, work from our lab demonstrates that immune responses to vaccination and infection in the periphery and local infection site (tissue) differ. To improve health outcomes in our aged population, we must study both systemic and tissue vaccine responses. We do so using a model of Mycobacterium bovis BCG vaccination and subsequent tuberculin skin test (TST) eight weeks after vaccination in adult and aged baboons. Vaccination generates BCG-specific immune cells that are recruited to the skin upon TST challenge, which we can study in skin biopsies reflecting tissue-specific responses. Three days or one week after TST we examine BCG-specific responses in skin biopsies. We also determine BCG-induced responses in the blood to compare tissue and systemic responses. We find increased oxidation and decreased production of immune proteins in aged baboon skin at the site of TST challenge, despite no alterations in antigen-specific peripheral blood mononuclear cell (PBMC) responses between age groups. Interestingly, cell migration assays show increased migration of PBMCs from aged BCG-vaccinated baboons, reinforcing the notion that systemic responses are not altered and that study of the tissue environment is important to understand responses to antigenic challenge with older age.
Computational models of Immunity – Pertussis Boost (CMI-PB): Engaging the broader scientific community to develop predictive models of Tdap booster vaccination.
Authors: Soldevila Ferran C , Shinde Pramod , Kojima Mari , Overton James A , Ha Brendan , Greenbaum Jason , Ay Ferhat , Kleinstein Steven H , Grant Barry J , Peters Bjoern
Published: February 2023

Whooping cough is a contagious disease caused by Bordetella Pertussis. The introduction of an inactivated whole bacteria vaccine (wP) in the 1940s led to a dramatic decrease in cases. However, due to vaccine related side-effects, this was substituted by the acellular vaccine (aP) in the 90’s which resulted in rise in cases. In order to determine if there are differences between those primed with aP vs. wP in infancy, we created a model using longitudinal samples from aP boosted individuals as a proxy for antigen encounter in vivo. In previous research, we observed substantial variability in magnitude of vaccine-induced responses between individuals regardless of their priming. Yet, we identified a systematic increase of Th2 polarization of T cells at day 7, a higher expression of CCL3 in PBMC on day 3 post-boost, and an increase of IgG4 antibody polarization in aP individuals. It is not clear how these different observations are interlinked.

We hope to address this question in our project, Computational Models of Immunity – Pertussis Boost (CMI-PB). We are establishing a website that will release omics data (10 aP vs 10 wP infancy primed donors) and engage students with educational materials, and help organize a yearly contest with the broader scientific community to help develop predictive models of immunity. This study will identify the key factors responsible for the differences between aP and wP primed individuals, while also providing a model for general vaccinology.
Integrin αvβ3 limits TLR signaling in plasmacytoid dendritic cells to prevent autoimmunity.
Authors: Lorant Alina K , Lacy-Hulbert Adam
Published: February 2023

SLE is a chronic, systemic autoimmune disease affecting millions of people worldwide. SLE is caused by a breakdown of tolerance to self-derived nucleic acids (NA) and characterized by the presence of autoantibodies to DNA/RNA. Plasmacytoid dendritic cells (pDCs) are highly tuned to respond to NA and express a range of receptors for RNA and DNA, including TLRs 7 and 9. Following TLR engagement, pDCs produce high levels of type I interferon (IFN-I), which promotes loss of tolerance, implicating pDCs in development of autoimmunity. However, relatively little is known about how innate immune signaling is regulated in pDCs, and particularly how they distinguish self-derived from foreign NA.

TLRs 7 and 9 are both localized in endosomal compartments, and their NA ligands must be internalized and processed before they can bind. One source of these NA is dying cells and loss of their receptors is associated with SLE. We studied the role of an apoptotic cell receptor, Integrin αvβ3, in TLR signaling of pDCs and found that αvβ3 limits IFN-I production. αvβ3-knockout pDCs show increased TLR signaling and produce more IFN-I and inflammatory cytokines when stimulated in culture. In mice without αvβ3, there is a systemic increase in circulating ISGs and IFN-I after TLR7 stimulation. This corresponds with an increase in pDC activation and a decrease in marginal zone B cells, indicating elevated IFN-I production.

These data indicate αvβ3 has a pDC-intrinsic role in limiting TLR signaling, which we propose is through activation of non-canonical autophagy. Based on these data, we hypothesize that co-receptors involved in capture and internalization of nucleic acids provide contextual ‘cues’ to regulate TLR signaling and prevent autoimmunity.
Myd88 activation in immune cells governs distinct disease manifestations in primary Sjögren’s syndrome
Authors: Kiripolsky Jeremy , Kasperek Eileen M , Zhu Chengsong , Li Quan-Zhen , Wang Jia , Yu Guan , Kramer Jill M
Published: February 2023

Studies in mice and humans identify Myd88 activation as an important driver of autoimmune disease, although the role of this adapter molecule in primary Sjögren’s syndrome (pSS) is poorly understood. In pSS, patients exhibit exocrine gland dysfunction along with serious systemic disease manifestations. Although Myd88-dependent signaling networks are essential for pSS disease pathogenesis, these pathways remain incompletely understood in the context of pSS. The objective of this study was to establish the contribution of Myd88 activation in immune cells to local and systemic pSS manifestations. To this end, we generated a novel conditional knockout pSS mouse model lacking Myd88 in the hematopoietic compartment. We found that ablation of Myd88 in immune cells altered inflammatory mediators in salivary tissue, but did not affect the degree of lymphocytic infiltration in exocrine glands or salivary production. Nephritis was diminished and pulmonary inflammation was increased in mice lacking Myd88 in hematopoietic cells. Finally, we found total IgM titers and autoreactive IgM and IgG were attenuated in pSS mice lacking Myd88 in immune cells. Thus, we demonstrated that Myd88 activation in the hematopoietic compartment is essential for distinct aspects of disease. This study establishes a rationale for design of targeted therapeutics that modulate Myd88 activation in a tissue-specific manner to ameliorate salivary inflammation and systemic disease manifestations characteristic of pSS.
The enzymatic function of IDO2 is not required for its proinflammatory role in driving autoantibodies and disease in preclinical arthritis
Authors: Mandik-Nayak Laura , Merlo Lauren MF , Peng Weidan , DuHadaway James B , Montgomery James D , Prendergast George C , Muller Alexander J
Published: February 2023

IDO2 is one of two closely related tryptophan catabolizing enzymes induced under inflammatory conditions. Although originally thought to be functionally redundant with IDO1, recent studies using IDO1 and IDO2 genetically deficient mice demonstrated a specific pathogenic role for IDO2, but not IDO1, in mediating autoantibody production and inflammatory joint disease in the KRN model of autoimmune arthritis. In humans, there are two single nucleotide polymorphisms in IDO2 that severely impair enzymatic function: Y359X, a nonsense mutation that leads to a stop codon and R248W, a missense mutation that disrupts substrate recognition. These inactivating polymorphisms are common, with only 25% of normal individuals expressing a homozygous wild-type (wt) configuration of the gene conferring full catalytic potential. This, together with IDO2’s weak enzymatic activity, suggests IDO2 may have a non-enzymatic function. To determine if the enzymatic activity of IDO2 was required for its pro-arthritic effect, we generated IDO2 knock-in mice that express catalytically inactive IDO2 (R248W or Y359X) and bred them onto the KRN model. Strikingly, R248W KRN mice developed high levels of autoantibodies and robust arthritis, indistinguishable from mice expressing wt IDO2. In contrast, Y359X KRN mice developed attenuated disease that was correlated with reduced stability of the truncated IDO2 protein, consistent with the effect of IDO2 deficiency in this model. These data confirm IDO2 as an important mediator of autoimmune arthritis and demonstrate that tryptophan catabolism is not the mechanism by which IDO2 drives disease. Taken together, our studies suggest a completely novel function for IDO2, independent of its enzymatic activity.
MDA5 expression impacts immune cell activation during T1D development
Authors: Blum Samuel I , Burg Ashley R , Chen Yi-Guang , Tse Hubert M
Published: February 2023

Coxsackievirus B3 (CVB3) infection has been associated with triggering Type 1 diabetes (T1D), potentially due to direct β-cell damage and bystander activation of autoreactive T cells. CVB3-infection of Non-Obese Diabetic (NOD) mice accelerates T1D onset, in part due to melanoma differentiation-associated protein 5 (MDA5)-dependent antiviral responses. However, the role of MDA5 on immune cells during T1D remains unclear. By using mice that contain an in-frame deletion within the helicase 1 domain of MDA5 (ΔHel1) or lack MDA5 expression (KO), we can test the hypothesis that reduced MDA5 expression impairs IFN-α/β synthesis, immune cell activation, and T1D onset. NOD and KO mice develop T1D equally, but ΔHel1 mice exhibit delayed spontaneous and CVB3-accelerated T1D. During spontaneous T1D, 12-week-old ΔHel1 mice had a significant reduction in TNF+ F4/80+ macrophages (MΦ) (1.6-fold), perforin+ CD8+ T cells (13.1-fold), and IFNγ+ CD4+ T cells (6.1-fold), compared to NOD and KO mice. ΔHel1 mice had a significant reduction in pancreatic IFN-α (2.2-fold) and IFN-β (3.1-fold) levels compared to NOD mice following CVB3-infection. CVB3-infected ΔHel1 mice had a significant decrease in pancreatic TNF+ F4/80+ MΦ (2-fold) compared to NOD and KO mice. However, ΔHel1 and KO mice had reduced pancreatic perforin+ CD8+ T cells (2.5-fold) and IFNγ+ CD4+ T cells (1.8-fold) compared to NOD mice following CVB3 infection. The ΔHel1 mutation decreased basal and CVB3-responsive MΦ and T cells within the pancreata thereby delaying T1D onset. Yet, loss of MDA5 expression failed to delay T1D indicating MDA5-dependant responses may be required to prevent T1D. Future studies will determine if T1D onset is dictated by MDA5 dsRNA binding and/or ATPase activity.
Abnormal C5 production and activation contributes to the mortality of COVID-19 patients
Authors: Fang Zhou , Li Xiang , Cheng Tzu Hsuan , Desikan Shreya , Worah Samrat , Foronjy Robert , Salvani Jerome , Kaiser Ryan , Bendo Audree , Dimaculangan Dennis , Wlody David , Velickovic Ivan , Diaz Geraldine , Santana Jeans , White Seth , Shevde Ketan , Zhang Ming
Published: February 2023

Elucidating the pathogenesis of COVID-19 is vital for treating the disease. Recent clinical studies have suggested that the complement system may play an important role in the course of infection. We investigated the correlation between complement C5, a critical factor in the complement common pathway, and the mortality of COVID-19. Thirty-seven COVID-19 patients were consented for this study. Serial blood samples were collected at different time points from 27 patients who recovered during hospitalization, and 10 patients who did not survive during hospitalization. Circulating native C5 was analyzed by Western Blot to detect α-chain and β-chain of intact C5. The activated C5a levels were detected by an ELISA kit. The levels of C5 β-chain and C5a were significantly higher in the non-survivors than in the survivors. Multivariate regression analyses showed that abnormal levels of C5 β-chain and C5a could independently predict the mortality. Therefore, our results suggest that in COVID-19 patients, the abnormal C5 production and activation may contribute to the systemic inflammation thus affect survival.
Identification of a unique TCR repertoire, consistent with a superantigen selection process in Children with Multi-system Inflammatory Syndrome
Authors: Porritt Rebecca A , Paschold Lisa , Rivas Magali Noval , Cheng Mary Hongying , Bahar Ivet , Binder Mascha , Arditi Moshe
Published: February 2023

Multisystem Inflammatory Syndrome in Children (MIS-C), a hyperinflammatory syndrome associated with SARS-CoV-2 infection, shares many clinical features with toxic shock syndrome, which is triggered by bacterial superantigens. The superantigen specificity for binding different Vβ-chains results in Vβ-skewing, whereby T cells with specific Vβ-chains and diverse antigen specificity are overrepresented in the TCR repertoire. Here, we characterized the TCR repertoire of MIS-C patients and found a profound expansion of TCR Beta Variable gene (TRBV)11-2. Furthermore, TRBV11-2 skewing was remarkably correlated with MIS-C severity and serum cytokine levels. Further analysis of TRBJ gene usage and CDR3 length distribution of MIS-C expanding TRBV11-2 clones revealed extensive junctional diversity, indicating a superantigen-mediated selection process for TRBV expansion. In silico modelling indicates that polyacidic residues in TCR Vβ11-2 engage in strong interactions with the superantigen-like motif of SARS-CoV-2 spike glycoprotein. Overall, our data indicate that the immune response in MIS-C is consistent with superantigenic activation.
Uncovering the underlying immune perturbations that determine long-term severity of chronic virus and Mycobacterium tuberculosis coinfection
Authors: Xu Wenxi , Snell Laura M. , Guo Mengdi , Boukhaled Giselle , Macleod Bethany L. , Li Ming , Tullius Michael V. , Guidos Cynthia J. , Tsao Ming-Sound , Divangahi Maziar , Horwitz Marcus A. , Liu Jun , Brooks David G.
Published: February 2023

Chronic viral infections increase severity of Mycobacterium tuberculosis (Mtb) coinfection, yet how they alter the pulmonary microenvironment to foster coinfection and worsen disease severity is unclear. We developed a coinfection model in mice with chronic lymphocytic choriomeningitis virus and Mtb coinfection that recapitulated the central clinical manifestations of coinfection, including increased Mtb burden, extra-pulmonary dissemination and heightened mortality. These long-term disease consequences were not due to chronic virus-induced immunosuppression or exhaustion, but instead were determined by early alterations in immune surveillance of Mtb coinfection. Mechanistically, increased chronic virus induced TNFα production initially arrested pulmonary Mtb growth, impeding dendritic cell mediated antigen transportation to the lung-draining lymph nodes (LNs) and allowing bacterial sanctuary. The inhibited antigen arrival to LNs delayed CD4 T cell priming, allowing Mtb to replicate to higher set-points before T cell mediated control could be initiated. Once primed, Mtb-specific CD4 T cell differentiation skewed away from the Th1 responses associated with Mtb control, and instead toward Th17 differentiation. The elevated IL17 increased pulmonary neutrophil influx that decreased the long-term survival of coinfected mice. Therapeutically correcting the timing of CD4 T cell priming re-established CD4 Th1 over Th17 dominance, diminished pulmonary neutrophilia, and enabled enhanced Mtb control in the presence of chronic viral coinfection. Thus, Mtb co-opts TNFα from the chronic inflammatory environment to subvert immune-surveillance, avert early immune function and foster long-term coinfection.
Respiratory syncytial virus-induced high mobility group box 1 secretion modulates innate immune response to promote inflammation
Authors: Hosakote Yashoda Madaiah , Kempaiah Kempaiah Rayavara , Garofalo Roberto P , Kurosky Alexander
Published: February 2023

Respiratory syncytial virus (RSV) is the single most common human pathogen responsible for ~ 60% of all acute lower respiratory tract infections in young children, and is a major cause of severe respiratory morbidity and mortality in the elderly, and the immunocompromised. Yet, no vaccine or efficacious treatment is available. Although the mechanisms of RSV-induced acute airway disease are not fully understood, lung inflammatory and immune events are key. Our studies show that RSV induces increased extracellular secretion of high-mobility group box 1 (HMGB1), a prominent damage-associated molecular pattern family member that alerts the immune system to potential damage and trigger immediate response. The goal of this study is to investigate the molecular mechanisms of RSV-induced HMGB1 release in the activation of innate immune response in promoting lung inflammation. Studies were conducted in BALB/c mice as well as human lung epithelial cells including A549 and normal small alveolar epithelial cells. To determine the role of HMGB1 in the activation of innate immune signaling, BALB/c mice and airway epithelial cells were infected with RSV in the presence or absence of recombinant HMGB1 or HMGB1 monoclonal antibody and measured RIG-I-dependent interferon-mediated innate antiviral immune response. Our studies demonstrated an important role of HMGB1 in cellular protection and the activation of innate immune signaling cascades to promote inflammation. Our studies indicate that HMGB1 acts as a signaling molecule to induce an increased inflammatory response in the lung, thus contributing to RSV pathogenesis. Modulation of proinflammatory function of HMGB1 may attenuate RSV-induced lung inflammation.
LUBAC suppresses IL-21-induced apoptosis in CD40-activated murine B cells and promotes germinal center B cell survival and the T-dependent antibody response
Authors: Xu Zhenming , Wang Jingwei , Li Tianbao , Zan Hong , Rivera Carlos Eduardo , Yan Hui
Published: February 2023

B cell activation by Tfh cells, i.e., through CD154 engagement of CD40 and IL-21, and survival within GCs are crucial for the T-dependent Ab response. LUBAC, composed of HOIP, SHARPIN and HOIL-1, catalyzes linear ubiquitination (Linear M1-Ub) to mediate NF-kB activation and cell survival induced by TNF receptor superfamily members, which include CD40. As shown here, B cells expressing the Sharpin null mutation cpdm (Sharpincpdm) could undergo proliferation, CSR and SHM in response to immunization by a T-dependent Ag, but were defective in survival within GCs, enrichment of a high-affinity BCR mutant and production of specific Abs. Sharpincpdm B cells stimulated in vitro with CD154 displayed normal proliferation and differentiation, marginally impaired NF-kB activation and survival, but markedly exacerbated death triggered by IL-21. While activating the mitochondria-dependent apoptosis pathway in both Sharpin+/+ and Sharpincpdm B cells, IL-21 induced Sharpincpdm B cells to undergo sustained activation of caspase 9 and caspase 8 of the mitochondria-dependent and -independent pathway, respectively, and ultimately caspase 3 in effecting apoptosis. These were associated with loss of caspase 8 inhibitor cFLIP and reduction in cFLIP Linear M1-Ub, which interferes with cFLIP poly-ubiquitination at Lys48 and degradation. Finally, the viability of Sharpincpdm B cells was rescued by caspase inhibitors, but virtually abrogated – together with Linear M1-Ub and cFLIP levels – by a small molecule HOIP inhibitor. Thus, LUBAC controls cFLIP expression and inhibits the effects of caspase 8 and IL-21-activated caspase 9, thereby suppressing apoptosis of CD40 and IL-21-activated B cells and promoting GC B cell survival.
Elucidating the role of ATM in BER and MMR during B cell CSR
Authors: Sible Emily , Attaway Mary , Chwat Tzippora , Fiorica Giuseppe , Rahman Sadia , Velez Kristel , Vuong Bao
Published: February 2023

Secondary immunoglobulin isotypes are produced by class switch recombination (CSR), which requires AID-dependent DNA deamination of intronic switch (S) regions within the immunoglobulin heavy chain gene. Non-canonical repair of deaminated DNA by mismatch repair (MMR) or base excision repair (BER) creates staggered DNA breaks that promote recombination between S regions. ATM-dependent phosphorylation of AID at serine-38 (pS38-AID) promotes its interaction with APE1, a BER protein, suggesting that ATM regulates CSR through BER. However, pS38-AID may also play a role in MMR during CSR, although the mechanism remains unknown. To examine whether ATM participates in MMR-mediated CSR, ATM−/− mice were bred to mice deficient for the MMR gene MSH2. Surprisingly, the predicted Mendelian frequencies of ATM−/− MSH2−/− adult mice were not obtained. To bypass the observed lethality, ATM was conditionally deleted on an MSH2−/− background using a floxed ATM allele [ATMF] and B cell-specific Cre recombinase expression. These ATMd/dMSH2−/− mice and B cells display a hypomorphic CSR phenotype comparable to ATM−/− B cells, which suggests that ATM may function genetically upstream of MSH2 and implicates a role for ATM in both MMR and BER.
Genetic regulation of nitric oxide-mediated metabolic reprogramming in dendritic cells
Authors: Snyder Julia Priscilla , Krementsov Dimitry N , Amiel Eyal
Published: February 2023

Dendritic cell (DC) activation is characterized by a rapid “metabolic switch” towards increased glycolysis that supports maturation. Sustained glycolysis post-activation is a requirement for survival in DC subsets that express inducible nitric oxide synthase (Nos2, iNOS), due to the production of nitric oxide (NO) which acts as an inhibitor of mitochondrial respiration. Long-term metabolic reprogramming in activated DCs has almost exclusively been studied in the strain C57BL/6J (B6), which experience loss of mitochondrial function due to NO accumulation. To assess the conservation of NO-driven metabolic regulation in DCs, we generated and compared DCs from B6 mice to the wild-derived genetically diverse PWD/PhJ (PWD) strain. We show attenuated iNOS induction, lowered NO production, and intact mitochondrial respiration in activated PWD DCs. To reduce genetic variability in our model, we generated DCs from a congenic mouse strain containing a genetic interval from PWD—including Nos2—on a B6 background. Activated DCs from the congenic strain produce NO at lower levels than B6, but NO production is not completely ablated as it is for pharmacologic or genetic interventions, thus this model allows for the subtle interrogation of NO-mediated metabolic reprogramming. We show that glucose-dependent carbon flux through the TCA cycle is altered in DCs with restrained NO production compared to B6 DCs, and we demonstrate that these differences are driven by NO. These studies provide a more comprehensive understanding of NO-mediated metabolic reprogramming in the murine myeloid lineage and establish a natural genetic model for investigating the contribution of NO in regulating the interplay between DC metabolism and immune function.
A role of aryl hydrocarbon receptor (AhR) in mesenchymal stem cells
Authors: Jeon Myung-Shin , Lkhagva-Yondon Enkhmaa , Oh Ye-Na
Published: February 2023

Aryl hydrocarbon receptor (AhR), a ligand-induced transcription factor, is known to be involved in the balance between inflammatory and suppressive/regulatory T cell activity. Mesenchymal stem cells (MSCs) possess immunomodulatory activities, including suppression of T- and B-cell activation. MSC-mediated immunosuppression occurs via nitric oxide (NO) in mice. Here, we isolated MSCs from Ahr−/− bone marrow in mice and found less immunosuppressive function compared to the wild type (WT)-MSCs. When MSCs are co-cultured with T cells in the presence of anti-CD3 and anti-CD28 antibodies, the AhR-deficient MSCs less inhibited T-cell proliferation, CD25 and CD122 expression and cytokine productions such as IFN-γ, IL-17A compared with wild type MSCs. In addition, we found AhR-deficient MSCs produce less inducible nitric oxide synthase (iNOS) and NO compared with wild type MSCs, when MSCs were stimulated with IFN-γ, TNF-α, and IL-1β. Since MSCs has been used for the treatment of graft-vs-host disease (GvHD), we injected the WT and AhR-deficient MSCs into the GvHD mice. The AhR-deficient MSCs showed less therapeutic efficacy compared to the WT MSCs. We think that the AhR is involved in the production of NO via regulating iNOS expression which is required for the therapeutic efficacy of MSCs.
Oral mucosal IL-22/STAT3 signaling licenses IL-17-mediated immunity to oral candidiasis
Authors: Aggor Felix E.Y. , Break Timothy , Trevejo-Nunez Giraldina , Whibley Natasha , Bailey Rachel D. , Kaplan Daniel H , Naglik Julian R. , Shan Wei , Shetty Amol C. , McCracken Carrie , Durum Scott K. , Biswas Partha S. , Bruno Vincent M. , Kolls Jay K , Lionakis Michail S , Gaffen Sarah L.
Published: February 2023

Oropharyngeal candidiasis (OPC) is an opportunistic infection of the oral mucosa caused by the commensal fungus Candida albicans. IL-17 and IL-22 both mediate antifungal immunity yet activate distinct downstream signaling pathways. While much is known about IL-17-dependent immunity in OPC, the activities of IL-22 are less well delineated. We show that induction of Il22 is independent of Dectin-1, CARD9 and aryl hydrocarbon receptor (AhR) and is driven by IL-23 and the C. albicans pore forming peptide candidalysin. Despite similar induction requirements and cellular sources, IL-22 and IL-17 function non-redundantly during OPC and exert opposing roles in neutrophil recruitment. The IL-22 and IL-17 receptors are required in anatomically distinct locations; loss of IL-22RA1 in the oral basal epithelial layer (BEL) but not the suprabasal epithelial layer (SEL) causes susceptibility to OPC, whereas IL-17RA is needed in the SEL. Our data reveal that IL-22 is a major activator of STAT3 in the BEL during OPC. Moreover, loss of STAT3 in the BEL but not the SEL renders mice susceptible to OPC. Transcriptional profiling of RNASeq data linked IL-22/STAT3 to oral epithelial cell proliferation and survival, but also, unexpectedly, to driving an IL-17 gene signature. We show that IL-22 acts on the BEL to replenish the IL-17RA-expressing SEL, thereby restoring the ability of the oral epithelium to respond to IL-17. Consequently, IL-22 signaling in BEL ‘licenses’ IL-17R signaling in the oral epithelium, revealing spatially distinct yet cooperative activities of IL-22 and IL-17 in oral candidiasis. This work also suggests that oral thrush in Jobs’ syndrome patients may be caused by STAT3 impairments in the oral epithelium, not just Th17 cells.
The immunomodulating enzyme indoleamine 2,3-dioxygenase 2 (IDO2) as a target for therapy in autoimmune disease
Authors: Merlo Lauren MF , Bowers Jessica , Stefanoni Tony , Gordon Gary V , Getts Robert , Mandik-Nayak Laura
Published: February 2023

The enzyme indoleamine 2,3-dioxygenase 2 (IDO2) is an important mediator of inflammatory autoimmune responses. Unlike its better-known counterpart, IDO1, which is thought to act primarily in an immunosuppressive manner, IDO2 acts to promote inflammation via a B-cell intrinsic mechanism. Prior work has demonstrated that genetic knockout of IDO2 can inhibit both autoantibody production and joint inflammation in the KRN.g7 murine model of arthritis. IDO2 played a similar role in driving inflammation in collagen induced arthritis and contact hypersensitivity models. Given the clear role of IDO2 in mediating autoimmunity, we have worked to find novel ways to target IDO2 in vivo. In addition to previous research successfully targeting the intracellular IDO2 protein with a novel monoclonal antibody, we have recently described an innovative, targeted DNA nanostructure-based siRNA-delivery system to specifically inhibit IDO2 function in B cells. Treatment of KRN mice with this B cell-directed anti-IDO2 siRNA nanotherapy reduced autoantibodies and attenuated arthritis, mimicking the phenotype seen with genetic knockouts. These data provided preclinical proof-of-concept that IDO2 could be effectively targeted as a treatment strategy in murine arthritis and suggested a similar strategy could be used target IDO2 in human RA. Similar to our findings using murine B cells, we found that IDO2 expression was elevated in B cells isolated from RA patients and could be silenced using anti-IDO2 siRNA in vitro. These data provide the first indication for the relevance of IDO2 in human RA and suggest nanoparticle delivery systems are a potential therapeutic strategy to inhibit IDO2 in RA patient B cells.
A high Bacteroides/Clostridium ratio is associated with the ability of male gut microbiota transplants to suppress lupus in female BWF1 mice
Authors: Harder James W , Ma Jing , Alard Pascale , Kosiewicz Michele M
Published: February 2023

Systemic lupus erythematosus (SLE) is associated with microbiota dysbiosis. Our experiments conducted from 2014–16 using NZBxNZW F1 (BWF1) mice showed that gut microbiota composition differed between adult female and male mice, and male cecal transplants into female mice suppressed disease. In 2017, our BWF1 colony was moved to a new campus facility with identical water, food and bedding. Although disease incidence/kinetics were similar between the two facilities, male cecal transplants could not suppress disease in female recipients in the new facility. Several changes were made in 2018, including water source, rooms in the facility and disinfecting procedures. Transplant experiments conducted after the changes (2019–2020) recapitulated the results from our original animal facility, i.e., male cecal transplants could suppress disease in female mice. Microbiota composition in female and male BWF1 mice was compared for the three different periods. We found that Bacteroides abundance was high in male mice, and Clostridium was very low during the periods when male cecal transplants suppressed disease. Alistipes was also present during periods when male cecal transplants worked, and absent when they did not work. These same patterns were also found in females receiving male cecal transplants, with a high Bacteroides/Clostridium ratio and Alistipes presence correlating with disease suppression. These data suggest that Bacteroides and Clostridium may have opposing effects on disease suppression that may also require the presence of Alistipes. Our findings further highlight how changes in environmental conditions can have dramatic effects on microbiota composition that can directly influence the immune response and disease.
Orally-delivered microbial extracellular vesicle induces anti-inflammatory activity in mice
Authors: Argueta Shannon , Cartwright Adam , Ramani Kritika , Cormack Taylor , Chernac Fabian Romano , Hilliard-Barth Kristie , Alami Aula , Raghunathan Divya , Jalan Mihika , Caffry Will , Keats Jake , Invally Krutika , Wang Bin , Kravitz Valeria , Rommel Tyler , Ganguly Tanmoy , Ponichtera Holly , Bodmer Mark , Itano Andrea
Published: February 2023

The small intestinal axis is a network of anatomical and functional connections linking the small intestine with the rest of the body. It senses external signals in the gut lumen and translates them into systemic immune effects. We have previously shown that an oral microbial drug candidate induces anti-inflammatory activity in preclinical models of inflammation by acting directly on host cells without colonization of the gut or modulation of the microbiome. We now extend these observations to EDP2939, a bacterial extracellular vesicle (EV), that has potent anti-inflammatory activity in preclinical models. EVs are non-replicating bacterial membrane vesicles with approximately 1/1000th the volume of the parent cell. EDP2939 was orally delivered and gut-restricted in distribution, it acts by modulation of innate and adaptive immunity within the small intestine to attenuate systemic inflammatory responses.

We observed significantly decreased ear swelling and inflammation in a delayed-type hypersensitivity model, showing that EDP2939 modulates systemic inflammatory responses. Activity of EDP2939 is dependent upon both TLR2 signaling and the presence of local immune cells. In vitro results show TLR2 agonism and induction of anti-inflammatory cytokine responses in immune cells by EVs. This is the first report of striking anti-inflammatory effects of an orally delivered microbial extracellular vesicle. EDP2939 induces broad-based resolution of inflammation across multiple pathways via a novel mechanism of systemic pharmacology without systemic exposure. EVs are particularly effective at engaging host cells in the gut to modulate distal inflammation. These data point to oral EVs as a new class of immunotherapeutic drugs.
Treatment of Autoimmune and Inflammatory Skin Diseases Using Skin-Targeting Bifunctional Antibodies: A Localized Immunomodulation Approach
Authors: Mande Purvi , Borthakur Susmita , Rios Daniel , Halvey Patrick , Boisvert Angela , Rowe Michael , Agrawal Anisha , Cianci Mike , Borah Minasri , Viney Joanne L , Kis-Toh Katalin , Higginson-Scott Nathan , Otipoby Kevin L , Mascanfroni Ivan
Published: February 2023

Current treatment approaches for autoimmune conditions comprise primarily of systemic immunosuppressants or cytokine blockade. Concentration of therapeutic molecules to the tissues that are the sites of autoimmune and inflammatory diseases is a promising approach with the potential to induce therapeutic benefit and avert risks associated with systemic immunotherapies.

Pandion Therapeutics is developing a bifunctional antibody platform that can drive localized immune modulation. The platform combines a “tether antibody” that targets a tissue of choice and “an effector end” that activates specific regulatory immune pathways to restore immune-homeostasis.

Here we report the engineering of a skin-tethered PD-1 agonist and a skin-tethered CD39 that inhibit T cell activation and function and deplete local ATP, respectively, modulating different arms of the immune system in a tissue specific manner. These skin-tethered immune effectors were assessed for drug-like properties in biophysical assays and in vitro and in vivo assays for target binding, cellular activity and tissue specific-localization. Moreover, these bifunctionals were tested in pathway-relevant preclinical models such as Vitiligo and Contact Hypersensitivity. Strikingly, a selective accumulation of the tethered bifunctionals to the skin was observed and correlated with a tether-dependent efficacy compared to a non-tether control.

We believe that this therapeutic approach has the potential to drive the resolution of cutaneous inflammation, providing an opportunity for developing new targeted therapies for autoimmune and inflammatory skin diseases.
Alphataxin, a Small-molecule Drug that Elevates Tumor-infiltrating CD4+ T Cells, in Combination with Anti-PD-1 Therapy, Suppresses Murine Renal Cancer and Metastasis
Authors: Bristow Cynthia L , Reeves Mary Ann B. , Winston Ronald
Published: February 2023

By promoting the cytotoxic function of CD8+ T cells, immune checkpoint inhibitor therapy, e.g. programmed cell death protein-1 (PD-1), effectively inhibits tumor growth in renal cell carcinoma. Yet, as many as 87% of cancer patients do not respond to immune checkpoint therapy. The function of cytotoxic CD8+ T cells integrally relies on CD4+ T helper cells. Remarkably, despite advances in immunotherapy, there are no pharmaceutical treatments that increase circulating CD4+ T cell counts. Nor has there been much attention given to tumor-infiltrating CD4+ T cells. The orally available small-molecule drug Alphataxin targets a pathway that regulates the number of circulating CD4+ T cells. We aimed to examine how Alphataxin affected tumor growth in a murine model of renal cell carcinoma. Alphataxin, in combination with anti-PD-1 antibody, significantly elevated the ratio of circulating and tumor-infiltrating CD4+/CD8+ T cells. In one study, following orthotopic implantation of syngeneic renal cell carcinoma, combination treatment resulted in 100% regression of tumor growth. Moreover, in mice implanted orthotopically with one log more tumor cells, doubling Alphataxin dose in combination treatment led to 100% regression in one-third of mice and 81% suppression of tumor growth in the remaining two-thirds of mice. Lung metastasis was evident in all mice except combination-treated mice. Orally available Alphataxin, the first and only drug developed to increase CD4+ T cells, in combination with anti-PD-1, is a powerful therapeutic method that provides long-term remission in renal cell carcinoma and potentially other T cell-responsive cancers by increasing the number of CD4+ tumor-infiltrating T cells.