bluuebunny/crossref_metadata_2025_split huggingface dataset:
Thousands of new antigens are recognized in mice via endogenous antibodies after being cured of a B78 melanoma via immunotherapy
Authors: Hoefges Anna , Erbe-Gurel Amy K , McIlwain Sean J , Melby Andrew S , Xu Angie , Mathers Nicholas , Rakhmilevich Alexander L , Hank Jacquelyn A , Baniel Claire , Pinapati Richard , Garcia Brad , Patel Jigar , Morris Zachary S , Ong Irene M , Sondel Paul M 
Published: January 2023

Abstract
Antibodies can play an important role in innate and adaptive immune responses against cancer. Using a high-density peptide array, we assessed potential protein-targets for antibodies activated in mice cured of their melanoma through a combined immunotherapy regimen. Our goal was to determine the linear peptide sequences recognized by anti-tumor antibodies produced in mice cured of their melanoma with immunotherapy.
Mice with GD2-expressing syngeneic B78 melanoma were treated with a combination immunotherapy capable of inducing an “in situ vaccine” effect (ISV), enabling mice to be cured of their tumors with long-term immune memory. Naïve and immune sera were collected from these mice. Using flow cytometry, immune sera showed strong antibody-binding against B16 (parental cell line of B78 without GD2 expression). These sera were then used on a Nimble Therapeutics’ peptide-array (either whole proteome or a curated list of ~650 proteins) to determine specific antibody-binding sites, and data were analyzed using a probabilistic model. Using the “curated list” peptide array, proteins were selected if the protein was bound in immune sera but not bound in the sera from naïve or non-responding tumor-bearing mice. When focusing on the whole mouse proteome data, thousands of peptides were targeted by 2 or more mice and exhibited strong antibody binding only by immune sera.
We are continuing to refine our analytical methods and are further investigating all identified proteins. These peptides may be new targets for antibody-based or cellular therapies and some of the tumor-specific endogenous antibodies that we have identified may be used as biomarkers to predict response to our ISV regimen and potentially other immunotherapy treatments.
Multi-dimensional analysis of tumor-specific and bystander CD8+ tumor infiltrating lymphocytes with TetTCR-SeqHD
Authors: Schonnesen Alexandra A , Parker Patrik , Ma Ke-Yue , McKinney Jennifer , Flemming Declan , Fox Matthew , Eckhardt Gail , Jiang Ning 
Published: January 2023

Abstract

Objective
Antigen specific CD8+ T cells were isolated from primary tumor tissue and analyzed for on the single cell level to determine tumor-specific signatures for further study.

Methods
Primary tumor, metastatic lesions, adjacent healthy tissue and peripheral blood were from collected from patients at Dell Seton Medical Center at the University of Texas. Matched tumor and healthy samples underwent exome and RNA sequencing to allow prediction of tumor-specific epitopes. We then synthesized DNA-barcoded tetramers for all predicted tumor associated-epitopes and an HLA-matched library of exogenous epitopes. Tetramer stained CD8+ T cells were then sorted and analyzed by TetTCR-SeqHD, a technology that we recently developed to link TCR antigen specificity, TCR sequence, gene expression, and phenotype of single T cells.

Results
We analyzed ~2900 tumor infiltrating lymphocytes (TILs) and T cells from adjacent healthy tissue and observed enrichment for cytotoxic effector and tissue resident memory cells within the tumor versus the healthy tissues. We also note an enrichment of bystander cells, specifically those reactive to Epstein Barr Virus epitopes, within the cytotoxic cluster. Finally, we analyze the distribution and differential expression between tumor-specific lymphocytes and bystander T cells, noting reduced expression of CD57 and CD49d in the tumor-specific TILs, as previously described.

Conclusions
We validate the use of TetTCR-SeqHD for gaining insight into phenotypic and gene expression differences between bystander and tumor-specific TILs. We identified markers for future exploration and demonstrate key differences between shared clonal populations within healthy and cancerous tissues.
USP22 stabilizes FoxP3 to maintain Regulatory T cell function through a dual mechanism resulting in tumor immune evasion
Authors: Montauti Elena , Zhang Yusi , Cortez Jessica , Marson Alexander , Fang Deyu 
Published: January 2023

Abstract
Aggressive cancers are less sensitive to standard cancer treatments available, reducing overall survival rates and increasing relapse percentage in cancer patients. As a result, there is an increasing need for alternative therapeutics that will prolong the survival of patients. Within the last decade, methods to attenuate tumor immune evasion have become a centerpiece of tumor therapies. However, a major hurdle in tumor immunotherapy is the immunosuppression mediated by Regulatory T (Treg) cells, which can promote tumor progression through the suppression of tumor immune evasion. Therefore, the ability to modulate Treg function in the context of cancer could lead to more effective therapies. Here, we identify the ubiquitin-specific peptidase 22 (USP22), a member of the deubiquitination module of the SAGA chromatin modifying complex, as a specific regulator of Foxp3, the lineage-specifying transcription factor of Tregs. Treg-specific ablation of Usp22 in mice reduced Foxp3 at both the transcriptional and post-translational level, creating defects in their suppressive function and leading to spontaneous autoimmunity. Importantly, USP22 ablation protected against tumor growth in multiple cancer models. Collectively, our findings reveal a previously underappreciated physiological function of USP22 in maintaining Treg stability and identify USP22 as a potential target for cancer immunotherapy.
Human influenza reactivity to conserved influenza hemagglutinin and its relation to age and repeat vaccination
Authors: Moritzky Savannah A , Sant Andrea J 
Published: January 2023

Abstract
The first exposures to influenza have been shown to bias the immune system towards a certain strain which is then boosted by subsequent infection and vaccination throughout life. The impact of repeated stimulation of hemagglutinin (HA) epitopes of the CD4 T cell repertoire is unknown. We are interested in the CD4 T cells specific to conserved regions of HA as the conserved stalk region is a target for new vaccine designs and antibodies to HA are a correlate of protection from infection. We are also interested in how CD4 T cell phenotypes after repeated antigen exposure from conserved HA epitopes could change the CD4 T cell repertoire. In order to probe these questions, we analyzed PBMCs collected from healthy human donors using cytokine ELISpots and flow cytometry. We created overlapping peptide pools containing conserved or unique regions of HA, to compare differences in cytokine secretion, abundance, and phenotypes of CD4 T cells elicited to different regions of HA. Our hypothesis is that CD4 T cells specific to repeatedly stimulated epitopes will have a higher frequency and different phenotypes compared to unique epitopes. We have found that there is a robust circulating CD4 T cell response to influenza antigens. The reactivity to conserved regions of HA varies between individuals as well as between strains of influenza in healthy human donors and in some donors, there are CD4 T cells that are biased towards conserved regions of HA. Those CD4 T cells to conserved regions of the stalk could be recalled after vaccination with stalk-based vaccine strategies. We plan to further examine the phenotypes of those CD4 T cells and to determine if decade of birth and vaccination history will impact the abundance of CD4 T cells to conserved HA regions.
Immune tolerance limits effective immunity to epitopes targeted by universal influenza vaccines
Authors: McGargill Maureen Ann , Pillai Meenu R , Labombarde Jocelyn , Lin Chun-Yang , Crawford Jeremy Chase , Chang Ti-Cheng , Keating Rachael , Lewis Carlessia , Guthmiller Jenna , Li Quan-Zhen , Wilson Patrick C , Thomas Paul Glyndwr 
Published: January 2023

Abstract
A universal influenza vaccine could save millions of lives in the event of a deadly pandemic. It is not clear why antibodies specific for conserved regions of influenza viruses are so rare. One possibility is that these antibodies have a higher potential to cross-react to self-proteins, and therefore B cells that generate these antibodies are deleted through tolerance mechanisms. Therefore, we examined whether cross-reactive influenza antibodies had a higher potential to be autoreactive than antibodies specific for one subtype of influenza. We previously demonstrated that H3N2-vaccinated mice treated with a low dose of rapamycin had more cross-reactive influenza antibodies and were better protected against subsequent lethal infections of multiple subtypes. Thus, we utilized rapamycin to increase the frequency of influenza cross-reactive antibodies, and tested whether these antibodies were more reactive to self-proteins than strain-specific antibodies. We found that mice with increased levels of cross-reactive influenza antibodies also had more IgM antibodies specific for self-antigens. Importantly, the increase in autoreactive IgM antibodies correlated with increased susceptibility to disease in mouse models of multiple sclerosis, lupus, and Guillian-Barre Syndrome. Together, our results suggest that influenza cross-reactive antibodies have the potential to be autoreactive. These data have important implications for developing universal influenza vaccines designed to generate durable influenza cross-reactive antibodies.
Regression of murine breast tumors after hetIL-15 monotherapy correlates with a novel population of intratumoral dendritic cells, in addition to increased infiltration of T, NK and cDC1 cells
Authors: Pavlakis George N , Karaliota Sevasti , Stellas Dimitri , Bergamaschi Cristina , Stravokefalou Vasiliki , Nagy Bethany , Felber Barbara K. 
Published: January 2023

Abstract

Background
We studied mechanisms of therapeutic efficacy of the heterodimeric IL-15 (hetIL-15) of human, macaque or mouse in several mouse tumor models, including the murine EO771 orthotopic breast cancer model in syngeneic C57BL/6 mice.

Methods
The effects of hetIL-15 on immune cells were compared in tumors, lymph nodes and spleen by flow cytometry, immunohistochemistry (IHC), gene expression profiling (Nanostring) and metabolic profiling of tumor infiltrating T cells.

Results
hetIL-15 administration resulted in tumor regression in 40% of the treated animals and resistance to tumor rechallenge, inducing long term immunological memory. Activated NK and CD8+ T cells were increased in hetIL-15 treated tumors. hetIL-15 administration resulted in increased intratumoral infiltration of conventional Dendritic cells (cDC1) and also induced a new population of intratumoral DC, (F4/80high DCs), which were correlated negatively with the tumor size. Phenotypic profiling of F4/80high DCs identified expression of several cDC1 specific markers, including CD103, IRF8 and XCR1. tSNE analysis clustered F4/80high DCs close to cDC1s and macrophages.

Conclusion
Complete eradication of EO771 tumors by hetIL-15 treatment correlates with different tumor infiltrating immune cell types. We propose that the anti-cancer activity of hetIL-15 in primary tumors is orchestrated by the interplay of CD8+T cells, cDC1 and a novel subset of antigen presenting cells with a distinct phenotypic profile. Effective hetIL-15 treatment leads to development of long term anti-tumor memory. hetIL-15 supported a favorable metabolic profile of intratumoral effector lymphocytes, important for their function.
Glycolipid-loaded nanoparticles harness iNKT cells for tumor immunotherapy
Authors: Shute Travis J , Dudley Elizabeth , Lai Andrew , Salas Briana , Angel Daniel , Nash Kelly , Leadbetter Elizabeth 
Published: January 2023

Abstract
Invariant natural killer T cells (iNKT) have a well-documented role in anti-tumor immunity through their release of proinflammatory cytokines and cytotoxic compounds. As iNKT cells can have direct and indirect killing effects on tumor cells, we propose a novel strategy for activating iNKT cells, via a PLGA nanoparticle delivery platform, to promote anti-tumor immune responses. Poly-lactic-co-glycolic acid (PLGA) nanoparticles can be reproducibly loaded with an iNKT cell glycolipid agonist, alpha-galactosylceramide (αGalCer), and a tumor associated antigen, ovalbumin (OVA). These dual-loaded PLGA nanoparticles rapidly activate iNKT cells in vivo to produce IFNgamma. Furthermore, in an in vivo model of melanoma, using B16F10-OVA cells, both prophylactic and therapeutic administration of nanoparticles containing αGalCer and OVA led to decreased tumor cell growth and increased survival. Ongoing studies are extending these concepts to PLGA nanoparticles loaded with αGalCer plus an immunogenic peptide from the naturally expressed melanocyte protein glycoprotein 100, gp100 25-33. This novel delivery system provides a platform with tremendous potential to harness iNKT cells for cancer immunotherapy purposes and as part of combinational therapies with other approaches such as checkpoint inhibitors.
De novo DNA methylation programs regulate CAR T-cell exhaustion
Authors: Zebley Caitlin C , Petersen Christopher T. , Prinzing Brooke , Bell Matthew , Fan Yiping , Crawford Jeremy Chase , Houke Haley , Haydar Dalia , Yi Zhongzhen , Nguyen Phuong , DeRenzo Christopher , Lazzarotto Cicera , Tsai Shengdar , Miller Shondra , Langfitt Deanna , Gottschalk Stephen , Krenciute Giedre , Youngblood Benjamin 
Published: January 2023

Abstract
Chimeric antigen receptor (CAR) T-cell therapy is revolutionizing cancer immunotherapy for patients with B cell malignancies. While clinical trials have demonstrated the curative potential of this approach, a significant and well-established limitation is the heightened contraction and transient persistence that CAR T-cells experience during prolonged antigen exposure. Building upon our prior observation that deleting the de novo DNA methyltransferase 3a (Dnmt3a) prevents T cell exhaustion in the prototypical model of LCMV-induced T cell exhaustion, we assessed the role of DNMT3A programming in the dysfunction of human CAR T-cells. Deletion of DNMT3A in multiple human CAR T-cell systems resulted in a striking preservation of the CAR T-cell’s ability to proliferate and mount an effector response during chronic antigen exposure. Whole genome methylation profiling of DNMT3A KO CAR T-cells established an atlas of epigenetically regulated genes targeted during CAR T-cell dysfunction. Cross-reference of our published murine exhaustion methylation profiles with our newly identified human CAR T-cell methylation atlas revealed conservation of epigenetically regulated exhaustion-associated genes. Using a novel epigenetic-based bioinformatic tool that predicts human T-cell differentiation we further documented the preserved developmental plasticity of the DNMT3A KO CAR T-cells. Lastly, analysis of publicly available RNAseq expression data from CD19-CAR T-cell products prior to infusion into CLL patients demonstrated that DNMT3A programming is significantly coupled to clinical outcome. Collectively our results demonstrate that de novo DNA methylation programming is a key factor limiting T-cell based immunotherapy.
Development of a high affinity and specific CHIKV-E2 monoclonal antibody for Chikungunya diagnosis
Authors: Kim Jaemoo , Yang Jihyun , Kim Young Bong , Lee Hee-Jung , Kim Sehyun , Poo Haryoung 
Published: January 2023

Abstract
Chikungunya fever is a vector-borne viral disease transmitted to humans by Chikungunya virus (CHIKV)-infected mosquitoes. There have been many outbreaks of CHIKV infection in world. Local transmission of chikungunya virus was emerged in the America continents at 2013. The virus poses ongoing risks to global health. To control CHIKV infection, it is important to improve the existing CHIKV diagnostic approaches to allow for the detection of low CHIKV quantity and to correctly discriminate CHIKV infections from those due to other mosquito-transmitted viruses, including Zika virus (ZIKV), Japanese encephalitis virus (JEV), and Dengue virus (DENV). Here, we produced monoclonal antibodies (mAbs) against the CHIKV envelope 2 (CHIKV-E2) protein and compared their affinity and specificity with commercially available mAbs using enzyme-linked immunosorbent assays (ELISA). Two anti-CHIKV-E2 mAbs, 19-1 and 21-1, showed higher binding affinities to CHIKV-E2 protein compared with the commercial mAbs. Especially, the 19-1 mAb had the highest binding affinity to inactivated CHIKV. Moreover, the 19-1 mAb had very little cross-reactivity with other mosquito-borne viruses, such as ZIKV, JEV, and DENV. These results suggest that the newly anti-CHIKV-E2 mAb, 19-1, could be applied for a CHIKV diagnostic approaches.
Transcriptional and clonal characterization of B cell plasmablast diversity following primary and secondary natural DENV infection
Authors: Waickman Adam T , Gromowski Gregory D , Li Tao , Siegfried Hayden , Victor Kaitlin , Kuklis Caitlin , Davidson Edgar , Srikiatkhachorn Anon , Ellison Damon , Rothman Alan , Jarman Richard G , Currier Jeffrey R , Friberg Heather L 
Published: January 2023

Abstract
Antibody-mediated humoral immunity is thought to play a central role in mediating the immunopathogenesis of acute DENV infection, but limited data are available on the molecular-level diversity, specificity, and functionality of the antibody response elicited by primary or secondary DENV infection. In order to close this functional gap in our understanding of DENV-specific humoral immunity, we utilized high-throughput single cell RNA sequencing to investigate B cells circulating in both primary and secondary natural DENV infections. We captured full-length paired immunoglobulin receptor sequence data from 9,027 B cells from a total of 6 subjects, including 2,717 plasmablasts. Unexpectedly, we found a high proportion of the DENV-elicited plasmablasts expressing IgA, principally in individuals with primary DENV infections. These IgA class-switched cells were extensively hypermutated and appeared to be derived from memory B cells, even in individuals with a serologically confirmed primary DENV infection. Utilizing a combination of conventional proteomics and high-throughput shotgun mutagenesis, we determined that DENV-reactive IgA class-switched antibodies represent a significant fraction of DENV-reactive and DENV-neutralizing Igs generated in response to DENV infection, and that they exhibit a comparable epitope specificity to DENV-reactive IgG antibodies. These results identify a heretofore unappreciated role for DENV-reactive IgA in the humoral response to DENV infection, and may offer insight into the differential pathogenesis associated with primary and secondary DENV infections.
Placental myeloid cells protect against Zika virus vertical transmission in a murine model
Authors: Winkler Clayton W , Evans Alyssa B. , Carmody Aaron B. , Peterson Karin E. 
Published: January 2023

Abstract
The ability of Zika virus (ZIKV) to cross the placenta and infect the fetus is a key mechanism by which ZIKV causes microcephaly. How the virus crosses the placenta and the role of the immune response in this process remain unclear. In the current study, we examined how ZIKV infection affected the immune cells within the placenta and developing fetus and whether these cells influenced virus vertical transmission (VTx). We found myeloid cells were elevated in the placenta of pregnant mice infected with ZIKV, primarily at the end of pregnancy, as well as in the fetus several days before birth. These cells, which included maternal monocytes/macrophages, neutrophils and fetal myeloid cells contained high levels of viral RNA suggesting they may be infected and contributing to viral replication and VTx. However, depletion of monocyte/macrophage myeloid cells from the dam during ZIKV infection resulted in increased ZIKV infection in the fetus. Myeloid cells were not depleted in the fetus in this experiment, so the increased virus infection was not the result of an impaired fetal myeloid response. Collectively, these data suggest that monocyte/macrophage myeloid cells in the placenta play a significant role in inhibiting ZIKV VTx to the fetus, possibly through phagocytosis of virus or virus infected cells.
Temporal analysis of the plasma membrane proteome after vaccinia virus infection sheds light on virus strategies to evade the immune response
Authors: Depierreux Delphine M , Altenburg Arwen F , Soday Lior , Fletcher-Etherington Alice , Ferguson Brian J , Weekes Michael P , Smith Geoffrey L 
Published: January 2023

Abstract
Vaccinia virus (VACV) is a poxvirus and the vaccine used to protect against variola virus and related orthopoxviruses. VACV encodes multiple proteins whose function is to evade the host immune response, contributing to an immune suppressive environment at the site of infection. There is, however, an incomplete understanding of how this immune suppression by VACV is consistent with a strong induction of immunological memory. There is especially little known about the relationship between NK cells and VACV, such as which ligands trigger NK activation and whether VACV uses strategies to interfere with the NK cell response. The purposes of this study is to clarify these open questions.
Here, we performed a quantitative proteomic analysis of membrane-associated proteins from VACV-infected cells at various time points. Using tandem mass-tag spectrometry we detected about 70 viral proteins and 900 membrane-associated host proteins. Analysis revealed that VACV infection rapidly altered the expression of proteins with immune functions, downregulated the surface expression of plexins, ephrins, cadherins and selectively downregulated MHC-I proteins. Moreover, VACV prevented the upregulation of stress-induced ligands and death receptors. Finally, the kinetics of expression of VACV proteins at the surface of the infected cell were analysed.
Collectively, these data constitute a valuable resource for the study of VACV and its interaction with the host immune response, provide insights into the interaction of NK cells with VACV, suggest novel VACV immune evasion mechanisms and shed light on which host factors are required to raise a protective immunological memory.
Latent gammaherpesvirus infection licenses age-associated B cells for pathogenicity during EAE and MS
Authors: Mouat Isobel , Allanach Jessica , Shanina Iryna , Vorobeychik Galina , Horwitz Marc S 
Published: January 2023

Abstract
Epstein-Barr virus (EBV) has long been implicated in multiple sclerosis (MS), though its mechanism of contribution remains unknown. Age-associated B cells (ABCs) are known to expand and persist following viral infection and are increased in MS patients. We hypothesize that EBV infection expands the number of ABCs and skews the population towards a Th1 inflammatory phenotype, leading them to function pathogenically in MS. To explore how the EBV-primed ABC population contributes to MS we have utilized the in vivo models of EBV, gammaherpesvirus 68 (gHV68), and MS, and MOG35–55 experimental autoimmune encephalomyelitis (EAE). We observe that ABCs display distinct phenotypes during gHV68 infection and EAE: ABCs secrete anti-viral IFNg during gHV68 infection and IL10 during EAE. Intriguingly, latent gHV68 infection prior to EAE results in a heterogeneous ABC phenotype, with IL10 and IFNg-secreting subsets. As IFNg is pathogenic during EAE and MS, and IL10 is protective, the gHV68-EAE ABCs display a more pathogenic phenotype compared to EAE alone, corresponding to the enhanced clinical course during gHV68-EAE. Knocking out ABCs results in an amelioration of disease in gHV68-EAE, but not EAE alone, further substantiating that gHV68 infection drives ABCs towards pathogenicity. In MS patients we observe an increased number and altered inflammatory phenotype of circulating ABCs compared to age and sex-matched healthy controls. These findings indicate that gHV68 and EBV prime ABCs to contribute pathogenically during EAE and MS and suggest that ABCs may be a therapeutic target.