bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phosphoproteomics reveals GITR agonism differentially regulates inflammatory vs. regulatory T cell subsets.
Authors: Davis Andrew , Cunningham Lori , Stokes Matthew , Nelson Alissa , Abell Kathryn , Lewis Michael , Scialdone Roy , Bui Than , Alonzo Emily , Freeman Gordon J , Klein Sarah 
Published: February 2023

Abstract
Triggering the co-stimulatory receptor glucocorticoid-induced TNFR-related protein (GITR) on T cells has been identified as a promising cancer immunotherapeutic strategy, yet the signaling events by which GITR agonism induces antitumor immunity are not well understood. GITR is expressed on activated CD4+ and CD8+ T cells, and is constitutively expressed on regulatory T cells (Tregs). While GITR appears to act as a conventional co-stimulatory receptor in effector CD4+ and CD8+ T cells, the role of GITR on Tregs remains controversial. The goal of this research was to utilize a phosphoproteomics approach to identify and compare GITR-mediated intracellular signaling events in induced-Tregs (iTregs), CD4+ T cells, and CD8+ cytotoxic T cells. We used the GITR agonist monoclonal antibody DTA-1 to stimulate the T cell subsets and confirmed GITR signaling via NFκB pathway activation and phospho-JNK induction. The phosphoproteomics data revealed different protein phosphorylation patterns among T cell subsets. For example, we observed phospho-p38 MAPK induction in the iTreg subset but not in the CD4+ T cells or CD8+ cytotoxic T cells. We also observed additional phospho-proteins induced by GITR triggering in the p38 pathway in the iTreg subset including phospho-MKK3, phospho-MKK6, phospho-MAPKAPK-2, and phospho-ATF-2. Induction of p38 MAPK signaling is functionally important as inhibition of p38 abrogated GITR-induced proliferation of iTregs. These data demonstrate that GITR agonism induces distinct phospho-proteome profiles between T cell subsets, and this differential signaling leads to unique responses that will need to be considered when targeting GITR with immunotherapeutic strategies.
PD1 Blockade of Tumor-primed CD4 T Cells Instigates Protective Immunity Against Tumor Rechallenge.
Authors: Falo Louis D , Hao Xingxing , Chen Guo , Carey Cara D , Liu Zuqiang , Bartlett David , Storkus Walter J , Falo Louis D , You Zhaoyang 
Published: February 2023

Abstract
Blockade of immune checkpoint PD1 in cancer patients results in remarkable therapeutic benefits. Revealing mechanisms underlying this revolutionary therapy is critical to identifying biomarkers for predicting responses to this treatment and developing novel PD1 blockade combined therapeutic regimens for broader clinical efficacy. Rejuvenation of preexisting intratumoral ‘exhausted’ T cells and recruitment of peripheral antitumor T cells into tumors have been suggested to explain how PD1 blockade works, but mechanisms of action of anti-PD1 therapy remain to be fully delineated. In a clinically relevant mouse model of spontaneous metastatic breast cancer 4T1.2-Neu, we show that effective and durable anti-PD1 therapy depends on systemic immunity, including type 1 conventional dendritic cells (cDC1: CD8α+/CD103+DC), B cells, CD4 and CD8 T cells. Increased circulating and/or tumor-infiltrating IL12+CD103+DC and IL2+CD4/CD8 T cells are associated with the response to anti-PD1 therapy. PD1 blockade results in tumor-primed CD44+IL2+CD4T cells in the periphery regardless of cDC1. In the breast cancer model with adoptive cell transfer (ACT), PD1 blockade-driven CD44+CD4+T cells infiltrate into tumors and ignite protective immunity against new tumors in an IL2-dependent and tumor-specific manner. In a distinct ACT model using enforced IL2-expressing melanoma antigen specific TRP1-transgenic (Tg) CD4 T cells, IL2+TRP1-Tg CD4 T cells control the growth of PD1-resistant endogenous BP or B16 melanoma. These data suggest the prominent role of PD1 blockade on the effectiveness of tumor-primed CD4T cells in tumor-specific immunity.
ELISA detection of human endogenous retroviral protein HERV-WL in saliva of cancer patients and normal individuals
Authors: Tsiagbe Vincent K , Nemati-Rad Azadeh , Bortnik Volha , Fatahzadeh Mahnaz , Li Yu 
Published: February 2023

Abstract
Previous studies revealed that a novel Human Endogenous Retrovirus (HERV-WL) was overexpressed in tissues from patients with certain aggressive human cancers, using immunohistochemistry techniques. ELISA detection also showed overexpression of HERV-WL in saliva of patients with certain aggressive cancers. To further explore these findings, saliva samples obtained from 12 patients with various cancers (1 with breast & colon cancer, 1, with colon cancer, 1 with multiple myeloma, 1 with prostrate cancer, 1 with renal carcinoma, and 7 with squamous cell carcinoma, were compared to saliva from 5 normal individuals. The studies showed significantly higher HERV-WL protein expression in saliva samples from all the 12 cancer patients (2–157 ug/ml), while HERV-WL protein expression was undetectable or less than 0.5 μg/ml in saliva from normal individuals. These findings warrant further research into the use of HERV-WL expression in saliva as a non-invasive assessment of responsiveness of certain cancers patients to therapy.
BATF3/MYC axis drives hyper-proliferation of TET2 deficient CAR T cells
Authors: Jain Nayan , Zhao Zeguo , Iyer Archana , Lopez Michael , Feucht Judith , Koche Richard , Yang Julie , Zhan Yingqian , Sadelain Michel 
Published: February 2023

Abstract
A recent report described a patient presenting a dominant chimeric antigen receptor (CAR) T cell clone at the height of anti-tumor response wherein a TET2 allele was disrupted by the integration of the lentiviral 4-1BB CAR vector (Fraietta et. al. Nature 2018). The enhanced proliferative ability conferred by this chance TET2 disruption opened up the possibility of generating TET2 edited CAR T cells for an enhanced anti-tumor efficacy. We modeled and evaluated loss of TET2 in human T cells engineered to express different CAR receptors in a murine model of human acute lymphoblastic leukemia (ALL). Our results indicate that CAR design plays a critical role in determining anti-tumor efficacy and early stage T cell phenotype upon TET2 disruption. We further observe that TET2 edited CAR T cells, over time, enter a hyper-proliferative state with a near total loss of effector function. The frequency with which TET2 edited T cells achieve hyper-proliferation is dependent on the signaling properties of the CAR receptor. This state is in contrast to the canonical exhaustion state wherein loss of proliferative ability precedes loss of effector function. This hyper-proliferative state shares some transcriptional features with T cell leukemia/lymphoma with sustained upregulation of cell cycle associated factors. Exome analysis identified point mutations and chromosomal aberrations in hyper-proliferative cells, but they were not conserved across different populations. Chromatin accessibility analysis revealed that loss of TET2 sets an epigenetic state that allows for sustained elevated levels of BATF3 which in turn drives a MYC-dependent proliferative program in TET2 deficient CAR T cells.
GD2 CAR-T Cells exhibit strong cytolytic potency against glioma stem cells
Authors: Hayes Heather B , Logun Meghan T , Chvatal Stacie A , Mueller Katherine P , Piscopo Nicole J , Das Amritava , Saha Krishanu , Brat Daniel J , Karumbaiah Lohitash 
Published: February 2023

Abstract
Glioblastoma (GBM) is an aggressive brain cancer without effective treatments. CAR-T cells targeted to tumor-associated antigens offer promise for treating GBM. Here, we used cellular impedance assays to compare the cytolytic potency and kinetics of conventional viral vs non-viral CRISPR engineered GD2 CAR-T cells against glioma stem cells (GSC), a subpopulation of glioblastoma cells.
Patient-derived N08 GSCs were plated at 50k cells/well on 96-well plates, and impedance was continuously monitored on the Maestro Z impedance platform (Axion BioSystems). GD2 CAR-T cells were engineered using either retroviral transduction (RV) or non-viral CRISPR editing (NV). At 48 hours, GD2 CAR-T cells were added at Effector:Target ratios of 0.1:1, 1:1, and 10:1. Comparisons were made to mCherry T cells (mCh) as a control. Impedance and cytolysis were monitored up to 7 days.
RV and NV GD2 CAR-T cells caused decreases in impedance consistent with T cell-mediated lysis of GSCs, whereas mCh T cells induced little change. NV CAR-T cells exhibited faster killing kinetics compared to RV CAR-T cells. The time to 50% cytolysis (KT50) was significantly shorter for NV vs RV CAR-T cells at 1:1 and 10:1 E:T ratios.
Cytotoxic function was validated with flow cytometry and cytokine analysis at 7 days. All T cells exhibited chronic activation measured by CD69 and CD137 upregulation. Importantly, NV CAR-T cells exhibited less exhaustion, as measured by PD1 and LAG3 expression.
Both RV and NV GD2 CAR-T cells effectively cytolyzed GSCs, with NV CAR-T cells exhibiting more potent and efficient killing. The high potency, fast kinetics, and reduced exhaustion of NV CRISPR GD2 CAR-T cells offer great clinical promise for treating GBM.
The Nanobody JK36 circumvents the epitope overlapping commonly observed between therapeutic anti-CD38 antibodies and commercially available analytical mouse antibodies
Authors: Lacaille Vashti , Kapinsky Michael 
Published: February 2023

Abstract
Therapeutic anti-CD38 antibodies have been demonstrated to elicit immune-mediated elimination of plasma cells and other CD38-expressing cells. Epitope overlapping with commercially available analytical monoclonal mouse anti-CD38 antibodies limits the possibility to study surface CD38 expression in the presence of these therapeutic antibodies. Nanobodies, derived from heavy-chain antibodies that naturally occur in llamas and other Camelids, can circumvent these undesired interferences. Nanobodies are single variable domain antibody fragments (VHH) that often expose a long complementarity-determining region 3 (CDR3). This feature allows them to recognize hidden epitopes, e.g. in molecular cavities, that are inaccessible to the CDRs in conventional analytical and therapeutic antibodies. In a model of anti-CD38 occupation with a therapeutic antibody, we have evaluated JK36, an anti-CD38 nanobody recognizing a cryptic epitope. Normal whole blood samples were pre-incubated with Daratumumab, a therapeutic humanized anti-CD38 mouse antibody. Binding of the Rabbit-JK36 (Rb-JK36) construct was compared to the binding observed with the conventional mouse antibodies, clones LS198-4-3 and T16. LS198-4-3 and T16 antibodies partially or completely failed to label the CD38 epitope, in case of being masked with Daratumumab. In contrast, the Rb-JK36 staining pattern and intensity remained unchanged, confirming that Rb-JK36 binds to an epitope which is not masked by Daratumumab. This study confirms that JK36 is a well-suited probe for research studies on CD38 expression in the presence of therapeutic or commonly used analytical anti-CD38 antibodies, opening new methodical approaches in plasma cell immunotherapy research.
Blockade of checkpoint ILT3/LILRB4/gp49B binding to fibronectin ameliorates autoimmune disease in BXSB/Yaa mice
Authors: Su Mei-Tzu , Inui Masanori , Endo Shota , Takai Toshiyuki 
Published: February 2023

Abstract
Extracellular matrix (ECM) not only provides a supporting framework for each tissue, but also elicits cellular responses by providing an inductive substrate. Fibronectin (FN), a major ECM macromolecule, is composed of multiple modules that interact with ECM-associated proteins, plasma proteins, and cell-surface proteins to ensure its modulatory roles. Here we revealed that FN is identified as a physiological ligand of the immunoglobulin (Ig)-like transcript 3/leukocyte Ig-like receptor B4 (ILT3/LILRB4) which serves as an immune checkpoint for autoimmune and cancer diseases. Human LILRB4 and the murine ortholog, gp49B, bound FN with sub-micromolar affinities as assessed by bio-layer interferometry. The major LILRB4-binding site in FN was located at the N-terminal 30-kDa module (FN30). Blockade of B4–FN binding with gp49 antibody or a recombinant FN30-Fc fusion protein reduced the secretion of pathogenic IgG autoantibodies, thereby reducing PAS-positive glomerular deposition in lupus-prone BXSB/Yaa mice. Compared with the control group, activation-induced cytidine deaminase (AID) expression in splenic activated B cells of FN30-Fc-treated BXSB/Yaa mice was decreased, indicating gp49B-FN30 signaling contributes in the class switch recombination (CSR) of Ig genes. These results suggested that gp49B–FN interplay is critical for pathogenic autoantibodies-mediated glomerulonephritis in the BXSB/Yaa lupus mouse model.
Neutrophil-specific Socs3 deficiency induces brain-targeted experimental autoimmune encephalomyelitis with increased cerebellar neutrophils
Authors: Turbitt William J , Yan Zhaoqi , Yang Wei , Buckley Jessica , Zhou Lianna , Qin Hongwei , Benveniste Etty 
Published: February 2023

Abstract
Multiple sclerosis (MS) is associated with a dysregulated JAK/STAT signaling pathway. Suppressors of cytokine signaling (SOCS) negatively regulate the JAK/STAT pathway. MS patients with disease affecting the cerebellum display rapid progression and poor prognosis. Previous data from our laboratory demonstrates a severe, brain-targeted form of experimental autoimmune encephalomyelitis (EAE) in mice lacking Socs3 in myeloid cells (Socs3ΔLysM), with increased cerebellar infiltration of primed neutrophils. Here, we induced EAE in mice lacking Socs3 specifically in neutrophils (Socs3ΔLy6G; n=26) and Ly6G−/+ control mice (n=13), and examined disease progression, neutrophil populations and CD4+ T-cell polarization at disease peak. Ly6G−/+ mice exhibited clinical signs of classical EAE, whereas Socs3ΔLy6G mice exhibited clinical signs of brain-targeted EAE, similar to Socs3ΔLysM mice. An analysis of the cerebellar cell infiltrate in Socs3ΔLy6G mice demonstrated an increased cellularity (p=0.005), which was comprised of both an increase in the percentage (p=0.020) and total number (p&lt;0.001) of cerebellar neutrophils. Neutrophils from Socs3ΔLy6G mice exhibited a primed phenotype with increased surface expression of CD11b (p=0.084) and reduced expression of CD62L (p&lt;0.001). Additionally, Socs3ΔLy6G mice exhibited an increase in the percent of cerebellar CD4+ T-cells (p=0.018) and a shift toward IFN-γ-producing Th1 CD4+ T-cells (p=0.012). These data conclusively document that neutrophil-specific deficiency of Socs3 is sufficient to induce brain-targeted EAE. Future studies will identify targets to dampen the role of neutrophils in brain-targeted autoimmune neuroinflammation.
METABOLIC AND STRUCTURAL MUSCULAR CHANGES DRIVEN BY CHRONIC DISEASE-LIKE IFN-G EXPOSURE
Authors: Fenimore John Maxwell , Springer Danielle , Valencia Julio C , Edmonston Elijah , Young Howard A 
Published: February 2023

Abstract
Cardiovascular myopathies and fatigue have been associated with chronic inflammatory responses and activated cell-mediated immunity. We report aberrant cardiovascular activity in the form of reduced ejection fraction in vivo in male mice with persistent low-level expression of IFN-g (ARE mice). ARE mice have a replacement with random nucleotides in the 3 prime UTR of the IFN-g gene that serves to remove a regulatory element (AU Rich) that is integral at controlling the stability of IFN-g mRNA. ARE mice had increased cellular and mineral, infiltrates into heart tissue and mitochondria linked metabolic changes that were concomitantly observed following a stress-exacerbated decrease in heart function, with or without exposure to IFN-a. Our data also demonstrates an increase in glucose expenditure and increased general fatigue of ARE mice as well as structural and ultrastructural changes in the heart in male mice. We also observe a decrease in the transcripts and a reversible decrease in function of aerobic respiratory components in the muscle of male mice exposed to IFN-g. Notably, we observe a decrease in X chromosome linked pyruvate dehydrogenase, a key component in pyruvate facilitated entry to the TCA cycle, and other downstream mitochondrial genes. Furthermore, the in vivo data revealed an increase in factors associated with fatigue, such as lactic acid production and changes in expression of genes associated with an increase inanaerobic respiration in cardiac musculature. These results indicate that the chronic expression of IFN-g results in a model for male biased heart failure understress in an autoimmune environment, like the idiopathic cardiomyopathy found in male Systemic Lupus Erythematosus patients.
Aberrant CD39 levels and regulation mark COVID-19 late stage
Authors: Longhi Maria Serena , Wang Na , Kalbasi Ahmadreza , Vuerich Marta , Manickas-Hill Zachary J. , Hecht Jonathan , Shaefi Shahzad , Robson Simon C. 
Published: February 2023

Abstract
Previous studies have reported immune cell exhaustion and perturbed T-cell immunity in novel-coronavirus-disease-19 (COVID-19). Heightened levels of CD39, an ectonucleotidase key to immunoregulation, are detected in CD4 and CD8 T-cells during viral infection where they correlate with worse disease outcome.
We performed a comprehensive transcriptome analysis of T-cell related gene pathways in PBMCs from 18 COVID-19 patients (6 convalescent, 6 with moderate disease and 6 with severe disease) and 6 healthy controls (HC); and in formalin-fixed autoptic tissue, obtained from the spleen, lung, kidney, liver and heart of 5 COVID-19 cases and 2 controls. Analysis was performed using nCounter CAR-T characterization panel (NanoString).
Our data show that in COVID-19 tissues there is a marked decrease in TCR diversity that is also present in PBMCs of severe patients, when compared with patients with moderate disease, convalescents and HC. A similar pattern is also noted when analyzing PBMCs for TCR signaling, Th1, Th17 and Treg-related transcripts. Heightened CD39 transcripts are detected in lung, liver and spleen of COVID-19 patients as well as in PBMCs obtained from patients with severe disease. A similar trend is observed for STAT-3, and HIF-1alpha, transcription factors that regulate CD39. Notably, CD39-AS RNA, a lncRNA that regulates CD39 at the post-transcriptional level, is markedly reduced in severe COVID-19 patients.
In conclusion, our data indicate systemic and local alterations in T-cell immunity in COVID-19. Aberrantly high levels of CD39 and related transcription factors and concomitant decreases in CD39-AS RNA in the late stage of disease suggest a role for CD39 and purinergic signaling in COVID-19 progression.
A role for PD1-PDL1/2 regulatory axis in suppressing T cell independent memory B cell generation and activation.
Authors: Spurrier M. Ariel , Jennings-Gee Jamie E , Daly Christina A , Haas Karen M 
Published: February 2023

Abstract
T cell-independent type 2 antigens (TI-2 Ags), including polysaccharides present on the surface of encapsulated bacteria, are limited in their capacity to generate a recall response. The failure of native polysaccharide-based vaccine-elicited antibody responses to boost following either revaccination or natural infection is a barrier to establishing protection against infectious diseases and is a poorly understood facet of humoral immunity. Previous work has indicated a role for antigen-specific IgG in suppressing TI-2 antibody recall responses, but questions remain as to how TI-2 memory B cell formation and reactivation is regulated. In the current study, we investigated the role of the PD1-PDL regulatory axis in regulating B cell recall responses to TI-2 Ags. Using adoptive transfers of B cells from VHB1–8 transgenic mice in which a fraction of B cells express specificity for NP, we demonstrate B cell-intrinsic PD-1 expression suppresses memory B cell (Bmem) generation as well as Bmem reactivation in response to NP-Ficoll. B cell-extrinsic PDL1 and PDL2 both contribute to suppression of Bmem generation and reactivation. B cell-extrinsic PDL1 and Bmem-intrinsic PDL2 expression play key roles in PD-1-mediated suppression of Bmem reactivation and antibody production. In summary, this work highlights a key role for PD-1 and its ligands in regulating the formation and functional reactivation of TI-2 Ag-specific memory B cells and reveals new avenues for improving TI-2 Ag-based vaccine responses.
Neonatal B cell receptor induces a STAT-dependent negative regulator
Authors: Sakai Jiro , Akkoyunlu Mustafa 
Published: February 2023

Abstract
Neonates and infants have impaired immune responses to immunization compared to adults, resulting in high susceptibility to microbial infection. B lymphocytes have a central role in humoral immunity to immunization and infection, and signals via B cell receptors (BCRs) are essential for the development and maintenance of B lymphocytes. Neonatal B cells, however, do not respond to BCR activation as robustly as adult cells do.
We have identified that signal transducer and activator of transcription (STAT)-related signaling pathways were uniquely activated in neonatal B cells following BCR cross-linking. Among the STAT proteins, we found that STAT3 and STAT5 were highly phosphorylated in neonatal B cells compared to adult B cells in response to BCR activation. Using small molecule inhibitors and small interfering RNA, we identified that STAT5-induced IL-6 activates STAT3 in an autocrine or paracrine manner. Moreover, IL-6-induced STAT3 activation leads to the production of anti-inflammatory cytokine IL-10.
Neonatal spleen is known to contain higher numbers of IL-10 producing Breg cells than adult spleen and IL-10 is known to act as a negative regulator of T cell functions including survival and cytokine production. Our studies unveiled the pathways involved in the production of IL-10 in BCR-activated neonatal cells. We showed that BCR-activated neonatal cells secrete IL-6 in a STAT5-dependent manner. Subsequently, IL-6 signaling through STAT3 leads to the production of IL-10 from B cells.
Marginal zone B cells have higher basal BCR signaling that correlates with altered mobility and spatial organization of IgM-BCRs
Authors: Abraham Libin , Scurll Joshua M , Tafteh Reza , Lu Henry Y , Falcão Rebeca Cardim , Chou Keng C , Coombs Daniel C , Gold Michael R 
Published: February 2023

Abstract
Marginal zone (MZ) B cells exist in a partially-activated ‘primed’ state but the molecular basis for this priming is not fully understood. We found that MZ B cells exhibit greater antigen-independent ‘tonic’ BCR signaling than naïve follicular (FO) B cells. Because BCR signaling output is dependent on BCR spatial organization and BCR-BCR interactions, we hypothesized that the increased tonic BCR signaling in MZ B cells is due to altered lateral mobility and nanoscale organization of BCRs. Single-particle tracking showed that surface IgM-BCRs on MZ B cells have higher diffusion coefficients and decreased confinement compared to IgM-BCRs on FO B cells. In contrast, the mobility and confinement of IgD-BCRs was similar on MZ and FO B cells. To assess BCR nanoscale organization, we used dSTORM and a novel graph-based clustering algorithm. This revealed that IgM-BCRs were more dispersed and less clustered on MZ B cells than on FO B cells, whereas IgD-BCR spatial organization was similar on the two cell populations. Importantly, 3-color STED imaging revealed that phospho-CD79 nanoclusters overlapped to a much greater extent with IgM-BCRs than with IgD-BCRs on FO B cells, and that this IgM-pCD79 overlap was even greater in MZ B cells. MZ B cells also exhibited greater pCD79 signaling in response to membrane-bound antigens than FO B cells. Thus, MZ B cells have greater tonic and antigen-dependent BCR signaling, which correlates with altered IgM-BCR mobility and organization