bluuebunny/crossref_metadata_2025_split huggingface dataset:
Particulate Delivery of Innate Immune Agonist for Cancer Immunotherapy
Authors: Watkins Rebekah , Tiet Pamela , Junkins Robert , Gallovic Matthew , Chen Ning , Johnson Brandon , Bachelder Eric , Ainslie Kristy , Ting Jenny P.-Y. 
Published: January 2023

Abstract
Immunotherapies have dramatically improved cancer patient survival; however, certain forms of cancer still rely on conventional chemotherapies and radiation. We have designed a novel cancer treatment that utilizes an immunotherapeutic to augment the adaptive immune system. Many FDA-approved immunotherapies have several drawbacks including high cost, short half-life, and flu-like side effects leading to patient noncompliance. To overcome these difficulties, we have utilized pathogen-associated molecular patterns (PAMP) to stimulate an innate immune response and induce a potent cytokine response. However, the receptor for the PAMP is located in the cytosol, and the PAMP cannot transverse the cell membrane. To overcome this, we have designed a polymeric microparticle (MP) to deliver the PAMP (PAMP MPs) into the cytosol. The polymeric MP is composed of a dextran-based, biodegradable polymer. It has several critical advantages over other FDA-approved delivery vehicles, such as liposomes, including greater stability, higher encapsulation efficiency (EE) of hydrophilic cargo, ease and reproducibility of production, and ease of end point sterilization. Treatment with PAMP MPs resulted in reduced tumor burden compared to mice treated with the soluble PAMP in a model of melanoma, suggesting that the MPs increase the biological activity. Intratumoral treatment with PAMP MPs resulted in increased natural killer cells and professional antigen presenting cells in the tumor environment. In conclusion, our PAMP MPs resulted in significant anti-tumor efficacy in an aggressive murine model of melanoma, and the data suggest that this anti-tumor function is due to an increased immune response within the tumor.
Expression optimization and functional testing of a scorpion toxin/antibody fragment fusion protein, ACDClx, for the immunotherapeutic treatment of glioblastoma
Authors: Cook Rebecca , Diamos Andrew , Mason Hugh , Sirianni Rachael , Mor Tsafrir , Blattman Joseph 
Published: January 2023

Abstract
The prognosis for patients with glioblastoma (GBM), a deadly form of brain tumor with a median survival of &lt;15 months, has not substantially improved since the 1970s. To overcome the myriad of treatment obstacles for GBM, we sought to design a new therapeutic capable of selectively enhancing T cell recognition and killing of all GBM cells while preserving healthy tissue. For this purpose, we have designed, expressed, and performed initial testing of the novel bispecific fusion protein, anti-CD3/chlorotoxin (ACDClx). ACDClx is a fusion of anti-CD3 heavy and light variable regions with a non-toxic, GBM-selective scorpion-derived peptide, chlorotoxin, which has been shown to bind to 100% of GBM cells with no selectivity for any healthy tissue types, including brain. ACDClx was expressed in leaves of N. benthamiana, purified, and tested for the capacity to selectively activate T cells. Initial experiments showed that ACDClx activates T cells, measured via calcium flux and CD69 expression, only if T cells, GBM cells, and ACDClx were all present. Mock purified protein in the presence GBM cells and T cells did not result in T cell activation, nor did ACDClx alone. Before further testing in vitro and in vivo, we sought to increase the production of ACDClx, the expression of which is significantly hampered due to its six disulfide bonds. To this end, we have produced 7 new constructs (5 for N. benthamiana, 2 for E. coli), 5 of which include a unique fusion partner for ACDClx to enhance solubility. Ongoing work includes evaluation of the most efficient construct, followed by evaluation in an immunocompetent tumor model of GBM.
Priming with percutaneous bacillus Calmette-Guerin (BCG) prior to intravesical BCG treatment safely improves BCG-specific response in patients with bladder cancer
Authors: Ji Niannian , Mukherjee Neelam , Morales Edwin E. , Tomasini Maggie E. , Hurez Vincent , Curiel Tyler J. , Abate Getahun , Hoft Daniel F. , Zhao Xiang-Ru , Sourindra Maiti , Cooper Laurence J.N. , Svatek Robert S. 
Published: January 2023

Abstract
Bacillus Calmette-Guérin (BCG) is the gold standard immunologic agent for patients with high-grade non-muscle invasive bladder cancer (NMIBC). Nevertheless, responses to BCG are heterogeneous and limited options exist when BCG therapy fails. Preclinical data in animal models of bladder cancer (BC) suggests that priming with percutaneous BCG vaccine prior to intravesical BCG instillation can enhance BCG-specific immunity and improve outcome. To study the safety, immunogenicity and preliminary efficacy of this approach, we administered percutaneous BCG 21 days prior to intravesical BCG instillation in “prime patients” with NMIBC (n=13) in a prospective single-arm clinical trial. Immune responses and clinical outcomes were monitored and compared to a contemporaneous cohort of “control patients” (n=9) receiving intravesical BCG without prior percutaneous BCG. Priming was well tolerated and no grade ≥3 adverse events were observed. Compared to control, prime patients had improvement in both local and systemic measures of BCG-specific immunity, scored by increased post-BCG urinary IL-8 and IL-17A and increased circulating CD4, CD8, and γδ T cell proliferation and effector function in response to BCG. Furthermore, ex vivo cytotoxicity of circulating NK and γδ T cells against RT4 BC was significantly increased in prime patient after BCG treatment and was mediated in part by NKG2D, suggesting an important role of innate effector immune mechanisms underlying BCG’s antitumor activity. Remarkably, no prime patients progressed whereas 3 out of 9 control patients progressed to muscle-invasive disease and underwent cystectomy. Thus, BCG priming safely enhances BCG-specific immunity and could improve outcome for patients with NMIBC.
Agonist Redirected Checkpoint (ARC), TIM3-Fc-OX40L, for Cancer Immunotherapy
Authors: Fromm George , de Silva Suresh , Patel Arpita , Johannes Kellsey , Hornblower Josiah , Schreiber Taylor H. 
Published: January 2023

Abstract
Combination immunotherapy with bispecific abs, linked scFv’s or T cell engagers have not shown that both checkpoint blockade and TNFR activation can be achieved with a single molecule, likely due to a loss of target avidity. Fusion proteins incorporating the extracellular domain of type I membrane proteins (eg. Enbrel) or type II membrane proteins (eg. OX40L-Fc), linked via an antibody Fc domain, are both functional despite the ECDs being in opposite orientation. We report the generation of a two-sided fusion protein (ARC) incorporating the ECD of TIM3 and the ECD of OX40L, adjoined by a central Fc domain.
The TIM3 end of the ARC binds GAL9 and phosphatidylserine (PS) in vitro and in vivo, and OX40L binds OX40 on the surface of primary T cells. TIM3-Fc-OX40L activates NFκB signaling in the absence of Fc receptor cross-linking, stimulates IL2/TNFa secretion with the superantigen SEB, and increases the in vitro killing of GAL9+ tumor cells. In vivo, TIM3-Fc-OX40L stimulated expansion of antigen-specific CD4 and CD8 T cells in mice adoptively transferred with OT-I/OT-II cells (+ova/alum). Live cell imaging has provided direct mechanistic support for TIM3-Fc-OX40L tethering T cells to tumor cells and eliciting cytotoxic activity (tumor killing). Finally, the therapeutic activity of TIM3-Fc-OX40L in established murine MC38, B16.F10 and CT26 tumors was superior to either TIM3 blocking antibody, OX40 agonist antibody or combination antibody therapy. These data demonstrate feasibility of a novel chimeric fusion protein platform, providing checkpoint blockade and TNF superfamily costimulation in a single molecule, which is unique with this molecule compared to antibody approaches, since all TIM3 ligands can be targeted simultaneously.
Progress in the development of a cell culture system to study the immune responses of reef-building corals.
Authors: Garrison Keith E. , Kuo Catherine 
Published: January 2023

Abstract
Previous work in the laboratory has focused on using fragments of aquaculture-raised specimens of the stony coral Acropora and Montipora. We are interested in these species because Acropora has an expanded immune gene repertoire in comparison to the closely-related anemone species Nematostella, especially in the number of TLR and IL-1R genes. However, space restrictions and maintaining acceptable water chemistry parameters pose a challenge to working with samples of stony corals in this form. In addition, coral fragments are complex holobionts that contain a mixture of coral cells, photosynthetic zooxanthelle, and bacteria, all on a calcium carbonate skeletal matrix. We seek a simplified model system of coral cells growing in isolation to facilitate performing the virus isolation, propagation, and controlled infection experiments that we think are necessary to advance our understanding of coral disease and immune responses. A number of protocols exist to propagate coral cells in a mixture of artificial seawater and typical mammalian cell growth medium. We compared the results of mechanical versus enzymatic isolation of coral tissues from the underlying skeletal matrix. We performed microscopic assessments of the coral and endosymbiont cells immediately following isolation and again after one week in culture. The endosymbiont zooxanthelle are easily visually distinguishable, and appear to persist longer under our culture conditions than the coral cells. It is unclear whether this is due to an improved ability of the zooxanthelle to live independently, or increased susceptibility of the coral cells to pathogen infections not eliminated by antibiotics in the culture medium.
CD8+ single positive and CD4+CD8+ double positive cells constitute a significant proportion of the total lymphocytes in the anterior chamber of the normal porcine eye
Authors: Maslanka Tomasz , Socha Piotr , Dąbrowski Michał , Ziółkowska Natalia , Ziółkowski Hubert 
Published: January 2023

Abstract
This study investigated whether CD4− and CD8-expressing cells are present in the anterior chamber of the normal porcine eye. Intact porcine globes were obtained from local slaughterhouses. They did not show any abnormalities or signs of disease. The cornea was incised and then aqueous humor (AH) from both globes of the same animal was collected. The lymphocyte population in peripheral blood was gated on the basis of forward and side scatter properties. The positioning of this gate served as a reference for setting the lymphocyte gate in AH samples. The number of lymphocytes in the anterior chamber was extremely low; only about 5000 events in the lymphocyte gate were collected for the entire volume of AH samples. Single positive (SP) CD8+ cells constituted almost 40% of the total AH lymphocyte population. In contrast, the mean percentage of SP CD4+ cells in the lymphocyte pool was only 1.77. It is well known that the peripheral blood lymphocyte (PBL) population in pigs contains a large share of CD4+(high)CD8+(low) double positive (DP) cells. Likewise, a large contribution of these cells was found in the anterior chamber, namely their mean percentage was 17.45. However, in contrast to their PBL counterpart, the majority of AH DP cells exhibited low expression of CD4. The results suggest that the anterior chamber in swine is an element in the normal migratory pathway of SP CD4+, SP CD8+ and DP cells. However, the trafficking of these cells into this compartment seems to be relatively poor. Moreover, the study demonstrated a limited presence of SP CD4+ cells in the anterior chamber compared to SP CD8+ and DP cells. Such selective homing of SP CD8+ and CD4+ cells to the anterior chamber may be involved in the induction and maintenance of ocular immune privilege.
Classification the WC1 gene family in Sus scrofa and evaluation of individual SRCR domain affinity for Mycobacterium bovis and Leptospira spp.
Authors: Le Page Lauren Ann , Buck Jessica , Boisvert Natalie , Gillespie Alexandria , Hudgens Edward , Hsu Haoting , Baldwin Cynthia L. , Telfer Janice C. 
Published: January 2023

Abstract
Unlike αβ T cells, γδ T cells respond to a variety non-proteinaceous molecules independent of MHC presentation. γδ T cells are less susceptible to immune evasion, making them an attractive target for next generation vaccines, however, little is known about the best way to prime them. WC1, a member of the group B Scavenger Receptor Cysteine Rich (SRCR) superfamily, is expressed exclusively on γδ T cells in swine and ruminants. Previous work in our lab has determined that there are 13 genes encoding for WC1 in cattle. We have also shown that WC1 functions as hybrid co-receptor and pattern recognition receptor for the γδ TCR. WC1+ γδ T cells share a restriction in TCR gene usage, yet respond to different pathogens based on which WC1 molecule(s) they express. This can be attributed to the ability of the expressed WC1 molecule to recognize and directly bind whole pathogens via its SRCR domains. Because WC1 is expressed as a multigene array, we hypothesize that each WC1 gene has co-evolved with a different set of pathogens. Swine belong to the same order as cattle, Artiodactyl, and are susceptible to infection with many of the same pathogens. The current swine assembly contains two predicted WC1 proteins, neither of which had been confirmed with cDNA evidence. Prior to this study only one full-length cDNA transcript had been successfully amplified. Using 5′/3′ RACE PCR and RT-PCR, we have obtained full length cDNA transcripts for seven WC1 genes with the SRCR domain patterns of a1-[b-c-d-e-d′] or d1-[b-c-d-e-d′]. Through bacterial pull-down assays, we have shown that multiple SRCR domains from different swine WC1 genes bind to vaccine strain Leptospira spp, and freshly grown Pasteur and Danish strains of Mycobacterium bovis.
Evolutionary modification of the VLR-based adaptive immune system in jawless vertebrates: Functional implications
Authors: Das Sabyasachi , Li Jianxu , Hirano Masayuki , Rast Jonathan , Cooper Max D. 
Published: January 2023

Abstract
The three types of variable lymphocyte receptor genes, VLRA, VLRB and VLRC, in the extant jawless vertebrates, encode antigen receptors, the remarkably diverse repertoire of which is generated by insertion of neighboring leucine rich repeat (LRR) sequences into the incomplete germline genes. In both lampreys and hagfish, the B-cell like VLRB+ cells differentiate into VLRB-secreting plasma cells, whereas the αβ and γδ T cell-like VLRA+ and VLRC+ cells express their VLR products solely as cell surface proteins. However, in comparative studies of lampreys and hagfish, we find that remarkable functional differences have evolved in these lymphocyte lineages. In contrast with their striking predominance in lampreys, the VLRB+ cells constitute a minor lymphocyte population in hagfish, and the VLRC+ cells are predominate. Notably the germline VLRB gene in hagfish contains a short non-coding intervening sequence, whereas the VLRB genes in sea lampreys and Japanese lampreys have very long intervening sequences which contain multiple transposable elements that may influence VLRB expression. In keeping with the relative low numbers of hagfish VLRB+ cells, antibody responses to a model immunogen, sheep erythrocytes, are much less robust in hagfish than in lampreys. Thus, even though the fundamental genetic program for differentiation of two prototypic T-like lymphocyte lineages and one B-like lineage is conserved in both jawless and jawed vertebrates, the genetic programs used for fine tuning of the VLR-based immunity have undergone notable independent evolutionary changes in lampreys and hagfish over the past ~480 million years.
Probiotic microbes evoke different responses from bovine BAL in vitro
Authors: Eicher Susan D. , Chitko-McKown Carol G. , Bryan Keith A. 
Published: January 2023

Abstract
Probiotics are used for enteric health. How they interact with upper respiratory immune cells is not known. We aimed to determine how microbes affect functions and markers of bronchoalveolar lung lavage cells (BAL). BAL were isolated from lungs of calves at slaughter. Treatments were a Lactobacillus probiotic (US), a commercially available probiotic (PB), Lactobacillus animalis (LA-51), Propionibacterium freudenreichii (PF-24), Enterococcus faecium (CH-212), E. faecium (SF-273), E. faecium (M-74), Bifidobacterium animalis ssp lactis (BB-12), Bacillus subtilis (EB-15 ), and Bacillus amyloliquifaciens (ZM-16). Microbial cells and BAL were incubated 1:1 for 1 h at 37° C and 5% CO2, then control (CNT), CD14 (FITC) and CD205 (RPE), CD18 (Alexa Fluor 647), and E. coli bioparticles (Molecular Probes) were added. Differences = P ≤ 0.05. Mean fluorescence of phagocytosis of E. coli bioparticles was less than CNT for BB-12 and EB-15, but percentage (%) of cells phagocytizing was not different. Oxidative burst of M-74 was less than CH-212, BB-12, EB-15, PB, and US, but none were different than CNT. The % of cells with oxidative burst was greater for PF-24 than for LA-51, BB-12, ZM-16, PB, and CNT. CD 14 mean fluorescence was least for M-74 compared with all other microbes and CNT, but % of cells expressing CD14 was greatest on PF-24 and US compared with EB-15 and BB-12, but not CNT. DC-205 mean fluorescence was greatest for M-74 compared with LA-51, PF-24, CH-212, SF-273, CNT, and US. Percentage of cell expressing DC-205 and CD18 was not different from controls. PF-24 and M-74 were most immunomodulatory compared with controls. M-74 appeared to suppress and PF-24 to increase leukocyte functions, showing that probiotic bacteria differentially modulate BAL.
Decoding the Feline Leukocyte Antigen MHC class I system via SMRT sequencing
Authors: Hess Paul R. , Holmes Jennifer C. , Dickey Allison N. , Scholl Elizabeth H. , Thorne Jeffrey L. 
Published: January 2023

Abstract
CD8+ T-cell cytotoxicity is an important component of anti-viral immune responses and is restricted by MHC class I peptide presentation. Investigating virus-specific CTL in cats is impeded by the rudimentary understanding of the Feline Leukocyte Antigen class I (FLAI) system. As polygeny and polymorphisms are inherent MHC features, defining functional loci and cataloging alleles are the first steps in characterizing FLAI. Additionally, identification of prevalent alleles will permit discovery of immunodominant CTL responses. Previously, using conventional sequencing of class I hypervariable regions, we defined three class Ia loci, FLA-E, -H and -K, and found allele sharing, even in our small sample of cats. Because of high FLAI homology, partial-length sequences did not allow unambiguous assignment of all alleles to their originating locus. We hypothesized that the PacBio SMRT sequencing platform would allow us to efficiently obtain full-length genotyping of all class Ia loci in multi-cat cohorts. Consensus building of FLAI contigs provided complete MHC sequences with high accuracy, greater depth and sensitivity, and in a fraction of the time of traditional cloning and Sanger techniques. In sequencing 17 cats (including 2 reference individuals), we clarified locus assignments, identified &gt;40 novel classical class I alleles and discovered several common allele supergroups (FLA-E*003, -E*009, -K*002, and -K*003). Also, for the first time, we confirmed the identify and expression of 2 class Ib loci (FLA-J, -L). This work brings the current total of known FLAI alleles to ~100 across 3 class Ia and ~35 across 2 class Ib loci. This new dataset should accelerate feline anti-viral research and facilitate allotransplantation in cats.
Development and Characterization of Chicken CD127-Specific Antibodies
Authors: Sun Zhifeng , Zhao Hongyan , Kim Woohyun , Panebra Alfredo , Lillehoj Hyun S. , Xianyu Zhezi , Gu Changqin , Yan Xianghe , Li Charles 
Published: January 2023

Abstract
Research in avian immunology has been significantly hampered by lack of effective immunological reagents in birds cross-reactive with mammalian orthologs and lack of sensitive assays for a long time. To better serve the avian immunology community, monoclonal and polyclonal antibodies specific for avian molecules of Clusters of Differentiation (CD) are being developed to promote thorough research on avian infection and immunity. In this study, chicken CD127 antibodies were developed and characterized. CD127, also named interleukin-7 receptor (IL-7R), is expressed on various cell types, including naive and memory T cells as well as many other types of immune cells, and plays a critical role in differentiation and activation of T lymphocytes. An extracellular region of chicken CD127 gene of 678 bp was cloned in pET28a(+) vector and expressed in BL21-AI™ E. coli. The expressed CD127 protein was around 30 kDa and crossed-reacted with monoclonal antibodies specific for human CD127 (Clone G-11). The protein was then immunized in BALB/c mice. After 3 immunizations, murine sera were found to detect a protein of 30 kDa expressed in E. coli, and react to the protein expressed on chicken T cell line (CU205) by flow cytometry as well. Flow cytometric analysis of cells from two-week-old chickens showed that CD127-positive populations were detected in spleen and thymus with such murine sera as polyclonal antibodies, and detected in thymus when using anti-human Clone G-11. Availability of these antibodies for CD127 and other CD molecules will dramatically facilitate the commercialization of poultry immune reagents, greatly benefit the avian immunology community and promote poultry health and production.
Immunomediator expression profiling in two beluga whale (Delphinapterus leucas) clinical cases
Authors: Hofstetter Amelia R. , Van Bonn William , Sacco Randy E. 
Published: January 2023

Abstract
Cytokines and other immunomediators can be biomarkers of inflammation. Quantitative real-time PCR (qPCR) has been used to examine cytokine gene expression in beluga whale (Delphinapterus leucas) peripheral blood mononuclear cells (PBMC). Thus, qPCR-based immunomediator assays could supplement clinical data from routine blood draws, revealing altered immune status. We compared immunomediator expression between beluga whales facing immune challenges and healthy companions at two aquaria. In both case studies, mitogens stimulated increased IL-1β, Cox2 and TNF-α message in PBMC from the abnormal whale over the control, but decreased IL-10 mRNA. This supports further elucidation of beluga PBMC immunomediator profiles for use as immune biomarkers.