bluuebunny/crossref_metadata_2025 huggingface dataset:
Anti-α4β7 mAb prevents MAdCAM-1-mediated activation and HIV replication in CD4+ T cells
Authors: Waliszewski Mia , Nawaz Fatima , Cicala Claudia , Ray Jocelyn , Olowojesiku Ronke , Perrone Ian , Wei Danlan , Jelicic Katija , Van Ryk Donald , Ansari Aftab Ahmed , Fauci Anthony S , Arthos James
Published: January 2023

HIV-1 preferentially infects α4β7+ CD4+ T cells, causing irreversible depletion of these cells in gut-associated lymphoid tissue (GALT) during the acute phase of infection. α4β7 mediates homing of CD4+ T cells to GALT through an interaction with mucosal addressin cell adhesion molecule-1 (MAdCAM-1), expressed on gut endothelial venules. In a rhesus macaque model of HIV transmission, administration of an anti-α4β7 mAb protected GALT from SIV infection. The mechanism(s) underlying this protection are poorly understood. We hypothesized that the anti-α4β7 mAb prevents MAdCAM mediated co-stimulation in a manner that could contribute to the protection that we observed. We determined that costimulation of CD4+ T cells through MAdCAM could support both proliferation of CD4+ T cells and viral replication in a manner that was inhibited by the anti-α4β7 mAb. In addition, MAdCAM costimulation induced the up-regulation of Ki-67, CD38 and CXCR5. Interestingly, we found that MAdCAM could induce the expression of CD45RO on naïve CD4+ T cells in the absence of proliferation, suggesting that this mode of costimulation is unique. These results indicate that costimulation of CD4+ T cells via MAdCAM provides a signal that promotes replication of HIV. Moreover, an anti-α4β7 mAb that interferes with the binding of MAdCAM to α4β7 inhibits both proliferation and viral replication, providing a potential mechanism for the capacity of this mAb to protect rhesus macaques from infection by SIV.
Delay in CD8 T-cell priming impairs viral control after vaginal LCMV transmission
Authors: Khan Shahzada , Woodruff Erik M. , Trapecar Martin , Sanjabi Shomyseh
Published: January 2023

Parameters required for eliciting CD8-T-cell-mediated immunity in the tolerogenic environment of female reproductive tract (FRT) are not well understood. We established a mouse model of intravaginal infection with acute lymphocytic choriomeningitis virus (LCMV) and showed that, compared to systemic infection, CD8-T-cell-mediated protective immune response was delayed in the FRT. After i.vag. LCMV infection, live virus is transmitted to the draining iliac lymph node (iLN) from the lower female reproductive tract (LFRT), which helps to prime virus-specific CD8 T cells in the iLN and restricts further systemic spread of the virus. However, T-cell priming in the iLN of i.vag. mice is delayed compared to that in i.p. infected mice, causing a delay in the appearance of effector CD8 T cells in the LFRT. This delayed CD8 T-cell response in LFRT provides LCMV a window of opportunity to replicate robustly in this immune privileged site. Mechanistically, the delay in CD8 T-cell priming is due to suboptimal activation of LFRT migratory DCs, which require further maturation upon migration into the iLN. As a potential cause of the suboptimal DC activation, we found that LFRT expresses limited steady-state levels of viral RNA-sensing pattern-recognition-receptors and elicits minimal induction of interferons and other inflammatory mediators required for anti-viral immunity. These findings provide a mechanistic explanation for the cause of the delay in CD8 T-cell activation after intravaginal viral infection and emphasize the notion that successful vaccination against RNA viruses may require adjuvants that elicit a stronger innate immune response than that provided by live viruses.
Inflammatory signals control human MAIT cell effector function
Authors: Slichter Chloe K , McDavid Andrew , Miller Hannah , Finak Greg , Seymour Brenda , McNevin John , Diaz Gabriela , Czartoski Julie , McElrath Margaret J , Gottardo Raphael , Prlic Martin
Published: January 2023

Mucosal-associated invariant T (MAIT) cells recognize bacterial metabolites presented on MR1 via their semi-invariant T cell receptor (TCR). MAIT cells are a potent effector population located in tissues that are constantly exposed to the microbiome. This raises the question how MAIT cell activation is regulated to avoid unwanted responses to microbiome-derived metabolites. To address this we dissected the MAIT cell response to TCR- and cytokine-mediated signals. We found that stimulating the MAIT TCR in the absence of inflammatory cytokines was not sufficient to induce effector function such as granzyme B and interferon-g expression. Inflammatory signals acting directly on MAIT cells were absolutely necessary to allow for TCR-mediated acquisition of effector function. Importantly, MAIT cells are not generally hyporesponsive to TCR-mediated signals as TCR stimulation in the absence of inflammation elicited a distinct chemokine response.

Single-cell gene expression analysis of MAIT cells isolated from healthy human mucosal tissue revealed increased expression of inflammatory genes compared to their counterparts in the blood, indicating that provided the proper signals mucosal MAIT cells are poised to respond rapidly to infection. Importantly, MAIT cells in the mucosa do not produce inflammatory cytokines directly ex-vivo, despite expressing higher levels of inflammatory transcript.

We propose a novel model in which the strict dependence of TCR-mediated signals on pro-inflammatory cytokines to induce effector function represents a MAIT cell-intrinsic mechanism to prevent unwanted responses to commensal bacteria-derived bacterial metabolites, while allowing potent effector responses following infection.
Hvem expression in innate lymphoid cells mediates host defense against intestinal infection
Authors: Seo Goo-Young , Shui Jr-Wen , Takahashi Daisuke , Kim Pyeung-Hyeun , Kronenberg Mitchell
Published: January 2023

The herpes virus entry mediator (HVEM), a member of the tumor necrosis factor receptor super family (TNFRSF14), has diverse functions in augmenting or inhibiting the immune response. We previously reported that epithelial HVEM signaling at mucosal barriers provides host defense against pathogenic bacteria, but the role of HVEM expression in innate lymphoid cells (ILC) in the mucosal immune system was unknown. To address this issue, we exposed RORgt-Cre × Hvemfl/fl mice with Yersinia enterocolitica (Y. enterocolitica), a small intestine infection model. We found increased bacterial translocation and decreased survival at an early time point. In these mice, Hvem is deleted in NK cells, ILC3 and in all T lymphocytes. Ifng−/− mice also showed reduced survival and increased bacterial translocation early after infection with Y. enterocolitica, demonstrating the importance of this cytokine for host defense. Interestingly, ILC from infected RORgt-Cre × Hvemfl/fl mice produced less IFNg after infection, but IFNg production by CD4+ T cells was not affected. Furthermore, in CD4-Cre × Hvemfl/fl mice, which deleted HVEM only in CD4+ T cells but not in ILCs, were not more susceptible to Y. enterocolitica. Additionally, we found that depletion of ILC from Rag1-deficient mice resulted in increased bacterial translocation and decreased body weight at an early time point. Taken together, our data indicate first, that IFNg production by ILCs, such as NK cells and ILC3, is important for host defense during Y. enterocolitica infection. Second, although the activation of ILCs is regulated by cytokines, our data show that HVEM, a member of the TNF super family, also is required for their ability to respond by producing IFNg early after infection.
Electronic sorting of immune cell subpopulations based on highly plastic genes
Authors: Wang Pingzhang , Han Wenling , Ma Dalong
Published: January 2023

The publicly available omic big data provide a great and valuable resource to drive big-data-based discoveries. Previously, we used immunologically relevant transcriptome data from microarrays to analyze gene co-expression, gene plasticity (GPL) and biomarker evaluation for human and mouse immune cells, and have built two immunological databases, ImmuCo (http://immuco.bjmu.edu.cn/) and ImmuSort (http://immusort.bjmu.edu.cn/), for gene co-expression and gene plasticity analyses, respectively.

In the current study, we addressed a brand new method for the first time, electronic sorting, which surveys big data samples to do virtual analysis and to sort immune cell subpopulations based on highly plastic genes at mRNA level. We also called this process either ‘virtual sorting’ or ‘in silico sorting’, because no actual immune cells were sorted as they are in the fluorescence-activated cell sorting (FACS) process. Through virtual sorting of human Tregs and cytokine-producing T helper cells, our results strongly supported the feasibility, effectiveness and predictability of this method. For example, we found that human CD4+FOXP3hi T cells were highly expressive of certain known molecules such as CD25 and CTLA4, and that this process of investigation did not require isolating or inducing these immune cells in vitro.

We made a systematic virtual analysis of all of the potential human CD4+ T cell subpopulations, and potential signature genes or marker molecules for these subpopulations were identified. Besides CD4+ T cells, the methods in this study are also suitable for other immune cells, non-immune cells and even for cancers for studies on gene expression and regulation.
Kinetic study of B cell-depletion with a novel mAb anti-mouse CD20, clone SA271G2.
Authors: Moyron-Quiroz Juan Enrique , Lin Leo , Oida Takatoku , Garcia-Mojica Salvador , Yang Xifeng
Published: January 2023

CD20 is a member of the membrane-spanning 4A family (MS4A), involved on the transmembrane calcium-influx, activation and proliferation of B cells. CD20 is first expressed on Pre-B cells and its expression is sustained throughout the different stages of the B cell differentiation, with the exception of plasma cells. CD20 is also expressed by many malignant cells from the B cell-lineage, making it an important target for immunotherapy.

We developed a novel antibody anti-mouse CD20 (clone SA271G2, rat IgG2b, κ), useful to deplete B cells in vivo. A single intravenous dose of antibody (0.25mg) depleted more than 90% of the B cells from peripheral blood, spleen and lymph nodes; while in the bone marrow, only mature B cells (CD19hi B220hi IgD+ IgMhi IL-7Ra−), but not immature B cells (CD19int B220lo IgD− IgM−/int IL-7Ra+), were eliminated.

The B cell depletion was sustained for over 20 days; then a gradual return of the B cell population was observed, and was fully recovered around day 40. During this time, no noticeable effect of this antibody on CD20− cells was observed.

SA271G2 is a novel and useful tool to study the role of B cells on in vivo models of disease, mechanisms of the immune response and immunotherapy; with the advantages of allowing the use of only wild type mice, eliminating the need of genetically modified animals, and of having a fully reversible effect on B cell depletion.
Immunomodulatory effects of repository corticotropin injection (H.P. Acthar® gel) on the MRL/lpr model of lupus
Authors: Higgins Paul , Decker Dima , Becker Patrice
Published: January 2023

Repository corticotropin injection (RCI) is indicated during exacerbations or as maintenance therapy for systemic lupus erythematosus (SLE), yet studies on the mechanisms of action have been limited. This study was performed to evaluate the effects of RCI on the MRL/lpr mouse model of lupus. Mice were administered RCI at 160, 400, and 800 U/kg s.c. every other day beginning on week 13 (after onset of clinical symptoms) until week 23. Urine and blood were collected for analysis of proteinuria and anti-dsDNA IgG antibody. In-life assessments were made weekly for lymphadenopathy and skin lesions. Kidneys and spleens were harvested for assessment of histopathology and leukocyte phenotyping performed by FACS. The results demonstrate that RCI at 400 and 800 U/kg significantly inhibited the development of proteinuria compared to placebo (p < 0.01, Friedman test). Development of lymphadenopathy (at 400 and 800 U/kg, p < 0.0001 and p < 0.001 Friedman test, respectively) and skin lesions (at 160 and 400 U/kg, p < 0.0001 and p < 0.05 Friedman test) were both reduced by drug treatment. RCI also significantly inhibited the generation of anti-dsDNA IgG antibodies at week 22 by 74%, 85%, and 90% at 160, 400 and 800 U/kg, respectively. Summed kidney histopathology scores were significantly reduced 67% and 74% at the 400 U/kg and 800 U/kg doses, respectively. Frequencies of specific spleen cell phenotypes were affected by RCI treatment. Immature B cells increased whereas transitional, marginal zone and germinal center B cells decreased. B220+ double negative T cell effector cells were also decreased. These results demonstrate the efficacy of RCI in the MRL/lpr lupus model and describe its immunomodulatory activity against specific lymphocyte subsets.
Kallikrein-Kinin System suppresses Type I Interferon (IFN) responses: a novel mechanism of IFN regulation.
Authors: Sriram Uma , Fernandes Nicole C. , Lee Michael Hweemoon , Didukh Marta , Zuluaga-Ramirez Viviana , Potula Raghava , Gallucci Stefania
Published: January 2023

Kallikrein-Kinin System (KKS) that comprises of kallikreins (klks), bradykinins (bk), angiotensin converting enzyme (ACE), and many other molecules, have classically known to be involved in a variety of physiological processes, including coagulation, angiogenesis and control of blood pressure. The role of KKS in regulating immune responses has come to light recently. Recent reports indicate that KKS is involved in lupus and giving exogenous klks can ameliorate lupus in mice. We have reported earlier the expression of a Type I IFN signature by dendritic cells (DCs) from the B6.Sle1, 2, 3 congenic model of lupus-prone mice. In this study, we directly asked the question if exogenous KKS regulates IFN induced responses. We used Flt3L induced bone-marrow-derived DCs from normal C57BL/6 (B6) mice and PBMC from normal human donors for in vitro studies. We stimulated the cells with recombinant IFN-α, IFN-β, CpG (TLR9 ligand), or R848 (TLR7 ligand) to induce IFN responses, and further treated with KKS candidates [Lys-bk, Hyp-bk, klk]. We analyzed the IFN induced responses by Interferon Stimulated Gene (ISG) expression and cytokine production. KKS candidates suppressed IFN or TLR induced IRF7 gene expression and CXCL10 production in both mouse DCs and human PBMCs. KKS candidates also suppressed IFN or TLR induced IL-6. We also used HOE140, a bk receptor 2 antagonist, and observed that blocking bk signaling increased the IFN responses in vitro. In vivo administration of Hyp-bk after IFN induction with CpG strongly decreased IRF7 and CXCL10 gene expression in B6 splenocytes, confirming our in vitro observations. Our results suggest a novel pathway of Type I IFN regulation by the KKS and opens up a new direction for designing drug targets for lupus.
Increases in the numbers of cells with the phenotype of myeloid-derived suppressor and regulatory T cells in children with acute lymphoblastic leukemia
Authors: SALEM MOHAMED LABIB , Abdou Said , El-Shanshory Mohamed , Attia Mohamed , Zidan Abdel-Aziz , Sobhy Shymaa , Zidan Mona
Published: January 2023

Background

Acute lymphoblastic leukemia (ALL) is the most common cancer diagnosed in children. The precise mechanisms behind the relapse in this disease are not clearly known. One possible mechanism could be the accumulation of regulatory cells including myeloid-derived suppressor cells (MDSCs) and T regulatory cells (Tregs) which we and others have reported to mediate suppression of anti-tumor immune responses. Few previous studies investigated the roles of Treg cells in ALL, while no studies reported the emergence of MDSCs in ALL.

Aim

Therefore, we aimed to analyze the frequencies and phenotype of these cells in a group of Egyptian B-ALL pediatric patients (n=45).

Materials and methods

MDSCs were identified as Lin− HLA-DR− CD33+CD11b+; and Treg cells as CD4+CD25+CD127−/low using multiparametric flow cytometer.

Results

We found significant increases in the numbers of MDSCs and Treg cells in B-ALL patients as compared to healthy control volunteers. B-ALL patients showed significant increased numbers of MDSCs and Tregs before chemotherapy when compared to healthy volunteers. The numbers of MDSCs were more increased while the numbers of Treg cells were more decreased in patients during induction of chemotherapy. The highest increase in the numbers of MDSCs was observed in patients after induction of chemotherapy, while Treg cells showed the most significant decreased numbers after induction of chemotherapy.

Conclusion

Our results indicate that B-ALL patients harbor both MDSCs and Treg cells, opening a new avenue to investigate the mechanism mediating the emergence of these cells in larger numbers of patients at different treatment stages.
Ovarian cancer ascites-activated neutrophils suppress T cell proliferation in a contact-dependent mechanism
Authors: Singel Kelly L , Khan ANM Nazmul H , Emmons Tiffany R , Mayor Paul C , Moysich Kirsten B , Odunsi Kunle , Segal Brahm H
Published: January 2023

Ovarian cancer (OC) is often diagnosed at advanced stages. Necrosis is a hallmark of advanced cancer and releases DAMPs that activate innate immune responses. It is well known that CD8+ T cells mediate antitumor immunity and immunosuppressive myeloid cells can impair T cell responses. In contrast, there is limited knowledge on the role of activated neutrophils in the tumor microenvironment. In patients with newly diagnosed OC, ~90% of ascites cells were inflammatory (CD45+) with varying proportions of neutrophils, monocytes and lymphocytes. Although patient variability existed, neutrophils comprised ~15% of CD45+ cells and the neutrophil:CD8+ T cell ratio was 1:1. In ex vivo studies, cell-free ascites (CFA) attracted normal donor neutrophils (NDN). Neutrophil extracellular traps (NETs) are a distinct mode of death, and characterized by extracellular release of DNA, histones, and granular products. We found that CFA induced NDN to generate NETs ex vivo. In co-culture studies, CD3/CD28-stimulated normal donor T cells were incubated with media; purified NDN 1:1; CFA; or NDN + CFA. After 72h, T cell proliferation was measured by [3H] thymidine incorporation. While CFA or NDN alone did not suppress T cell proliferation, CFA-activated NDN completely suppressed proliferation. Suppression did not affect T cell viability or induce apoptosis, and was not reversed by the addition of arginine, N-acetyl-L-cysteine or IL-2. Suppression required T cell contact with NDN. These results support a model in which ascites-activated neutrophils suppress T cell responses, thereby abrogating antitumor immunity. Further studies will identify ascites constituents that activate neutrophils and mechanisms for neutrophil-mediated T cell suppression.
Regulatory T cells modulate tumor progression and clearance
Authors: Kumar Sanjay , Stokes James A , Gunn Karyn Scissum , Singh Udai P , Manne Upender , Ponnazhagan Selvarangan , Mishra Manoj K
Published: January 2023

Regulatory T (Treg) cells, a distinct lineage of T cells, are required to maintain immunological tolerance. However, the molecular mechanism(s) by which Treg cells modulate tumor growth and clearance needs further investigation. Thus, the goal of this study is to analyze the absolute number and role of Treg cells during tumorigenesis. To achieve this goal, tumor induction studies were performed on C57/B6 mice using transgenic adenocarcinoma mouse prostate (TRAMP) cell lines C1, C2 and C3. Interestingly, TRAMP C1 and C2 are tumorigenic while TRAMP C3 fail to form tumor. C57/B6 mice treated with different tumorigenic concentrations of TRAMP (C1, C2, and C3) cells. The experimental design also included appropriate controls. Developing tumors, spleen and draining lymph nodes were dissected out and single cell suspensions were prepared. Cells were stained with the specific antibodies such as NK cells, NKT cells, CD4, CD8, macrophages, B cells and Treg cells and analyzed using a 13-color flowcytometer. Preliminary data revealed that development of tumors by TRAMP C1 and TRAMP C2 cells and non-tumorigenesis by TRAMP C3 cells was dependent on number of Treg cells in tumor microenvironment. Therefore, the data suggest that the number of Treg cells present in tumor microenvironment may have a direct impact on tumor progression and clearance. Thus, modulating the number of Treg cells in tumor microenvironment may be a successful therapeutic strategy to control tumor progression
CD45 phosphatase regulates the fate of myeloid cells in tumor microenvironment by inhibiting STAT3 activity
Authors: Kumar Vinit , Cheng Pingyan , Condamine Thomas , Mony Sridevi , Languino Lucia , McCaffrey Judith , Hockstein Neil , Guarino Michael , Masters Gregory , Penman Emily , Denstman Fred , Xu George , Altieri Dario , Du Hong , Yan Cong , Gabrilovich Dmitry I.
Published: January 2023

The abnormal differentiation of myeloid cells is one of the key features of altered immune response in cancer. Myeloid derived suppressor cells (MDSC) and tumor associated macrophages (TAM) are the major components of immune suppressive network. MDSC accumulate in large numbers in peripheral lymphoid organs and inside tumors. However, in contrast to peripheral lymphoid organs such as spleen, inside tumors, monocytic MDSC rapidly differentiate to TAM with potent immune suppressive activity. We have demonstrated that this phenomenon is mediated by dramatic down-regulation of STAT3 activity in MDSC in tumor site. In cancer patients, MDSC in tumor site also had substantially lower level of pSTAT3 than MDSC in peripheral blood. These results were unexpected, since high level of STAT3 activity is a hallmark of MDSC present in blood and lymphoid organs. The mice with constitutively active STAT3 had shown the expansion of MDSC and down-regulation of macrophage population in tumor microenvironment, emphasizing the crucial role of STAT3 in myeloid cell differentiation. Hypoxia is an intricate part of tumor microenvironment. The decreased STAT3 activity was caused by hypoxia and affected all myeloid cells but was not observed in tumor cells. We further demonstrate that the up-regulation of CD45 tyrosine phosphatase activity in MDSC exposed to hypoxia in tumor site was responsible for down-regulation of STAT3 activity. Moreover, the positive regulation of CD45 activity in hypoxia was mediated by the disruption of CD45 protein dimerization via sialylation. This suggests a novel role of STAT3 in the regulation of myeloid cells in tumors with potential implications for therapeutic targeting.
The transcription factor GLI2 modulates CD40L expression in the tumor microenvironment
Authors: Han Weiguo , Jackson David A , Matissek Stephan , Misurelli Jason , Elsawa Sherine F
Published: January 2023

The interaction between tumor cells and the tumor microenvironment (TME) is essential for cancer cell growth and survival. Cytokines play an important role in mediating this crosstalk. Although this interaction is clearly established, the molecular mechanisms mediating this remain elusive. In previous work, we have shown that the transcription factor GLI2 modulates the expression and secretion of IL-6 in the TME. In this study, we screened for novel cytokine targets for GLI2 in bone marrow stromal cells and identified CD40L as a novel target of GLI2. GLI2 overexpression in HS-5, L87, L88 and Saka cells increased CD40L expression by real-time PCR. Conversely, GLI2 knockdown resulted in reduced CD40L expression. Similar results were obtained when screening for protein levels by western blotting. Using a mutant of GLI2 that lacks an N-terminal transcriptional repressor domain, we found an increase in CD40L beyond that observed with full-length GLI2. Bioinformatics analysis of a 2000 bp CD40L promoter region identified 2 candidate GLI binding sites in the CD40L promoter region. Using expression, luciferase and chromatin immunoprecipitation assays, we demonstrate that GLI2 binds to CD40L promoter. Furthermore, GLI2 modulates CD40L promoter activity. We were able to rescue the effect of GLI2 knockdown using an RNAi resistant GLI2 cDNA. Finally, we found that signaling through the CCR3-PI3K-AKT mediates GLI2-CD40L. Taken together, our data identify a novel mechanism regulating CD40L in stromal cells in the TME. Because of the importance of CD40L in malignant B cells, understanding the mechanism of its regulation is of great importance for the development of novel therapies aimed at targeting the malignant bone marrow TME.
Absence of Grail in CD8+ T cells confers spontaneous tumor regression.
Authors: Nurieva Roza I , Wang Junmei , Haymaker Cara , Alekseev Andrei , Bernatchez Chantale , Neelapu Sattva S.
Published: January 2023

T cell tolerance is the main obstacle to successful cancer immunotherapy; thus, it is of high priority to develop strategies to break immune tolerance to malignancy. Here we report that expression of the E3 ubiquitin ligase Grail is significantly upregulated in CD8+T cells infiltrated into lymphoma tumors and Grail-deficiency confers tumor rejection. Importantly, therapeutic transfer of Grail-deficient CD8+ T cells was sufficient to reject established tumors. Mechanistically, loss of Grail enhanced anti-tumor reactivity and survival of CD8+ T cells. Unexpectedly, Grail ablation was associated with ability of CD8+ T cells to evade regulatory T cell-mediated suppression. In addition, Grail deficient CD8+ T cells exhibited increased IL-21R expression and subsequently are hyper-responsive to IL-21 treatment. Finally, CD8+ T cells isolated from human lymphoma patients expressed high levels of Grail and lower levels of IL-21R compared with normal donors. Altogether, our data demonstrate that Grail is a crucial factor that controls CD8+ T cell anti-tumor activity.
Lymphoma Galectin-1 Expression Drives CD20 Immunotherapy Resistance
Authors: Lykken Jacquelyn M , Horikawa Mayuka , Minard-Colin Veronique , Kamata Masahiro , Miyagaki Tomomitsu , Poe Jonathan C , Tedder Thomas F
Published: January 2023

Non-Hodgkin’s lymphoma is the most commonly diagnosed hematologic cancer of adults in the United States, with the vast majority deriving from malignant B lymphocytes that express cell surface CD20. CD20 immunotherapy (Rituximab) is widely used to treat Non-Hodgkin’s lymphoma, even though the initial effectiveness of Rituximab varies widely amongst patients and typically wanes over time. The mechanisms through which lymphomas initially resist or gain resistance to immunotherapy are not well-established. To address this, a preclinical mouse model system was developed to comprehensively identify lymphoma transcriptomic changes that confer resistance to CD20 immunotherapy. The collection of spontaneous primary and familial lymphomas revealed that sensitivity to CD20 immunotherapy was not regulated by differences in CD20 expression, prior exposure to CD20 immunotherapy, nor serial in vivo passage. An unbiased forward exome screen of these primary lymphomas was used to validate the utility of this expansive lymphoma cohort, which revealed that increased lymphoma galectin-1 expression strongly correlated with resistance to immunotherapy. Genetically induced lymphoma galectin-1 expression ablated antibody-dependent lymphoma phagocytosis in vitro and lymphoma sensitivity to CD20 immunotherapy in vivo. Human NHLs also express elevated galectin-1 compared with non-malignant lymphocytes, demonstrating the ability of this preclinical model system to identify molecular targets that could be relevant to human therapy. This study therefore established a powerful preclinical model system that permits the comprehensive identification of the dynamic lymphoma molecular network that drives resistance to immunotherapy.
Effects of Yupingfeng formula on anti-lung cancer via increasing immune memory in lung metastasis mouse model
Authors: Xu Zihang , Hu Jiemiao , Mitra Abhisek , Xia Xueqing , Zhu Shiguo , Li Shulin , Chen Xiao
Published: January 2023

Background

Yupingfeng(YPF), a classic herbal Chinese formula, has been used effectively for the cure and prevention of respiratory tract diseases. However, the role of YPF in lung cancer which is the most severe lung disease with no substantial therapies, remains unknown. Therefore, this study was carried out to elucidate the anti-lung cancer role of YPF in lung metastasis mouse model.

Methods

C57/BL6 mice were inoculated with LLC-Fgl2-GFP tumor cells intraosseously, then were divided into two groups. One group was YPF group, and the other was control group. YPF (580 mg/ml)or water were gavaged daily for 30 days. The primary leg tumor diameters were measured weekly. 31 days after inoculation the whole control group were euthanized due to excessive primary leg tumor burden. After India ink pulmonary perfusion of these mice, lots of lung metastases were observed. By striking contrast, the whole YPF group survived tumor free. We then used the same cell line and cell numbers to re-challenge these mice. 21 days after inoculation, the primary leg tumor could be detected. But 7 days later, the primary tumor started shrinking until tumor free again, suggesting YPF could activate the immune memory.

Results

The control group died of excessive primary leg tumor burden and lung metastases, while the YPF group were tumor free, and remained tumor free after being re-challenged with the same tumor cells. Our results suggested that YPF could significantly increase the survival rate of tumor bearing mice, compared to the control group.

In conclusion

These data support YPF can prolong mice’s survival time by enhancing immune memory in lung metastasis mouse model, which might be a possible mechanism for its therapeutic effect on tumor free mice.
Gene electrotransfer of plasmids encoding cytokines as an effective immunotherapy approach for melanoma
Authors: Heller Richard , Lundberg Cathryn M , Burcus Niculina , Edelblute Chelsea , Guo Siqi
Published: January 2023

Gene electrotransfer (GET) of plasmids encoding cytokines has been shown to generate a potent anti-tumor effect. Effective protocols have been developed delivering the plasmids directly into tumors. To obtain an effective immune response, it is critical to achieve a balance between transgene expression and tissue damage. GET is a reliable and effective physical method for in vivo delivery of plasmid DNA. It is particularly attractive for use in anti-cancer cytokine therapy as it does not incur the side effects associated with conventional protein cytokine therapy. Delivery of plasmids encoding cytokines directly to tumors has been shown by our lab and others to induce not only local immune response, but a systemic one as well. The positive responses were directly related to the ability to achieve the appropriate expression profile following delivery of the plasmid. Interestingly, the electrotransfer parameters were dependent on which cytokine was being delivered. For plasmid encoding IL-12 the correct parameters were 1300 V/cm and 100 us. For IL-15, the appropriate parameters were 600 V/cm and 5 ms. The procedure included injecting the plasmid DNA into established B16.F10 melanoma tumors of C57BL/6 mice and electric pulses applied three times over a one week period. To enhance this immune response, current work is further analyzing the immune response to determine if there is a marker that would indicate appropriate delivery of the plasmid. In addition, combination therapies are being explored to evaluate if the response could be augmented. A more complete understanding of the process and identifying other agents that could further augment this approach would be an important step.
Expression of a CD200-targeted chimeric costimulatory receptor enhances T cell effector function and adoptive immunotherapy of disseminated leukemia
Authors: Oda Shannon K , Daman Andrew W , Garcia Nicolas M , Tan Xiaoxia , Greenberg Philip D
Published: January 2023

CD200 (OX2) is an immunosuppressive surface protein upregulated in leukemia and other malignancies, with expression associated with inhibition of antitumor T cell responses and decreased overall survival of patients. Our lab has developed several T cell receptors (TCR) to target intracellular antigens upregulated in malignancies and we questioned if the efficacy of TCR gene therapy could be improved by co-opting CD200 inhibition with a CD200R-CD28 immunomodulatory fusion protein (IFP) that could potentially convert CD200/CD200R signaling into a costimulatory signal. T cells transduced with CD200R-CD28 exhibited enhanced proliferation, accumulation, and effector function in response to CD200+ target cells in vitro. Repeated restimulation with irradiated CD200+ tumor cells enriched the cells transduced with the IFP compared to wild type T cells. The CD200R-CD28 transduced T cells also exhibited enhanced killing of CD200+ compared to CD200− tumor cells. In adoptive therapy of disseminated leukemia, CD200R-CD28-transduced tumor-specific T cells eradicated otherwise lethal disease without a requirement for IL-2 administration to sustain in vivo persistence. Comparison of IFPs with CD28, 4-1BB, and CD28/4-1BB dual costimulatory domains suggested the CD28 signaling domain provides the most significant enhancement of anti-tumor activity. Additionally, modification of the IFP ectodomain length revealed the importance of optimizing the tumor-T cell distance for costimulatory signaling. Transduction of human primary T cells with the equivalent human IFP also resulted in increased proliferation and cytokine production in response to CD200+ leukemia cells, supporting clinical translation of this strategy.
Site-Specific Immunomodulators (SSIs) are novel immunotherapies for cancer
Authors: Zhang Monan Angela , Bosiljcic Momir , Kalyan Shirin , Anderson Rebecca , Bazett Mark , Luk Beryl , Wong Natalie , Mullins David W , Gunn Hal
Published: January 2023

The immune system recognizes and eliminates malignant cells, and maintains an intricate balance between immune activation and cancer. Historically, spontaneous tumor regression has been observed in some patients experiencing acute microbial infections. Building upon this, it was discovered that repeated subcutaneous injection of novel immunotherapies called Site-Specific Immunomodulators (SSIs), derived from killed bacteria known to cause infection in a particular organ or tissue, may be effective at treating cancer growing in the same site. We hypothesize that SSIs stimulate the body’s immune system to reverse immune suppression and dysfunction in the tumor microenvironment, enabling effective anti-cancer immune responses. To test this hypothesis, we evaluated the efficacy of a lung-specific SSI, QBKPN, in preclinical lung cancer models. Repeated subcutaneous administration of QBKPN significantly reduced lung tumor burden at day 16 post-inoculation (p<0.0001) in the Lewis Lung Carcinoma model, improving median survival by 10 days (p<0.005). Similar results were obtained using the aggressive and poorly-immunogenic B16 lung cancer model. Anticancer efficacy was associated with site-specific infiltration of newly recruited monocytes and neutrophils to the lung and mobilization of antigen presenting cells in the lung-draining lymph node. These data complement the positive results from our compassionate use clinical data, and provide strong evidence that SSIs may improve therapeutic outcomes for cancer patients. QBKPN is currently studied in a Phase 2a clinical trial of non-small cell lung cancer in collaboration with the BC Cancer Agency (Trial NCT02256852).
Delayed dosing intervals for quadrivalent human papillomavirus vaccine do not reduce antibody avidity
Authors: Brady Allison M , Walter Emmanuel B , Markowitz Lauri E , Unger Elizabeth R , Panicker Gitika
Published: January 2023

The quadrivalent HPV vaccine (HPV4) is recommended as a 3 dose schedule at 0, 2, 6 months for girls and boys. However, delays in dosing intervals frequently occur. We previously found that delayed intervals in girls resulted in similar or higher antibody (Ab) titers following series completion compared to the recommended schedule. We used archived specimens from that study to determine if delayed dosing intervals affected Ab avidity(a measure of how strongly Abs bind). The importance of avidity is unknown for HPV; however, avidity has been associated with clearance of other pathogens. We tested residual serum from 261 consenting healthy females, ages 9 through 18. Avidity index (AI) was determined just prior to and one month following dose 3 of HPV4 for each participant. Data were analyzed by dosing intervals, classified as:(1) on time dose 2 and 3, (2) delayed dose 2 and on time dose 3, (3) on time dose 2 and delayed dose 3, or (4) delayed dose 2 and 3. Avidity was measured using a modified multiplex ELISA, and AI was calculated as a ratio of IgG Ab bound in the treated and untreated sample. Overall, mean AI was highest for HPV16 and lowest for HPV6. As expected, AI did not differ between groups 1 & 3 or groups 2 & 4 after dose 2, however for most types mean AI was significantly higher for groups with delayed dose 2. Mean AI was higher post-dose 3 in all delayed dosing groups compared to the group 1 for all types. There was a weak to moderate correlation between overall AI and Ab titers for all types (HPV6r=0.27, HPV11 r=0.32, HPV16 r=0.25, HPV18 r=0.44; all p<0.0001). Our findings suggest longer intervals between doses result in higher antibody avidity after a complete 3-doses series, providing further evidence that delayed dosing of HPV4 does not hinder the immune response.
Vaccination with Embryonic Stem Cells Protects against Lung Cancer
Authors: Richie Jamaal , Al-Rayyan Numan , Mitchell Robert , Eaton John , Yaddanapudi Kavitha
Published: January 2023

The antigenic similarity between tumors and embryos has been appreciated for many years and reflects the expression of embryonic gene products by cancer cells and/or cancer-initiating stem cells. Taking advantage of this similarity, we have tested a prophylactic lung cancer vaccine composed of allogeneic murine embryonic stem cells (ESC). Naïve C57BL/6 mice were vaccinated with ESC along with a source of granulocyte macrophage-colony stimulating factor (GM-CSF). Vaccinated mice were protected against subsequent challenge with implantable Lewis lung carcinoma (LLC). ESC-induced anti-tumor immunity was not due to a non-specific “allo-response” as vaccination with allogeneic murine embryonic fibroblasts did not protect against tumor outgrowth. Vaccine efficacy was associated with robust tumor-reactive primary and memory CD8+ T effector responses, Th1 cytokine response, higher intratumoral CD8+T effector/CD4+CD25+Foxp3+ T regulatory cell ratio, and reduced myeloid derived suppressor cells in the spleen. Prevention of tumorigenesis was found to require a CD8-mediated cytotoxic T lymphocyte (CTL) response because in vivo depletion of CD8+ T lymphocytes completely abrogated the protective effect of vaccination. To overcome the limitations of using whole ESC as a vaccine we modified our vaccination approach using exosomes derived from ESC. Our preliminary results thus far indicate that ESC-derived exosomes as a vaccine is effective against implantable lung tumors in mice. Further refinement of this novel vaccine strategy and identification of shared ESC/tumor antigens may lead to immunotherapeutic options for lung cancer patients and, potentially represent a step toward the development of prophylactic cancer vaccines.
Production of DNA, Protein and DNA/Protein vaccines against Leishmania infantum and L.tropica
Authors: Yazar Suleyman , Aksit Abdussamed , Karaca Serkan , Ulusoy Hasan Basri , Cam Mustafa , Hokelek Murat , Kuk Salih
Published: January 2023

Leishmaniasis is a neglected tropical disease caused by obligate intracellular protozoan Leishmania parasites. Especially cutaneous form of leishmaniasis has an increasing importance in Turkey because of immigration.

The aims of this study that is supported by the Scientific and Technological Research Council of Turkey (TUBITAK project number 213S149) are to produce Leishmania infantum and L.tropica activated C kinase receptor (LACK) gene based DNA and protein vaccines and to investigate the effectiveness of these vaccines by prime-boost method alone and combined with two adjuvants in-vivo on mice model.

Methodological steps in this study are to culture L.infantum and L.tropica promastigotes in NNN medium, total genomic DNA isolation from culture, cloning activated C kinase receptor (LACK) gene with specific primers, ligation of PCR product into plasmid, amplifying recombinant plasmid in competent cells, producing endotoxin free DNA vaccine, producing endotoxin free LACK recombinant protein, combining DNA and protein vaccines with MPL-SE and CpG-ODN adjuvants and to investigate the effectiveness of this vaccines in-vivo on mice model.

As a result L.infantum and L.tropica LACK gene based DNA and protein vaccines was produced and ready to immunization.

Th-1 and Th-2 immunologic responses are going to be investigated after two week intervals three time vaccination against L.tropica and L.infantum infections on BALB/c mice.
NKp46 expression identifies canine CD3 negative lymphocytes as natural killer cells with phenotypic and functional similarity to those in humans.
Authors: Foltz Jennifer A , Somanchi Srinivas S , Bishop Erin E , Yang Yanwen , Lee Dean A
Published: January 2023

Canines are a unique model to study adoptive immunotherapy for cancer as they spontaneously develop many cancers with homology to human disease. However canine natural killer (NK) cells are poorly defined, rendering the model less useful for study of innate immune responses. NKp46 (NCR1) is an activating receptor expressed on NK cells and is considered a species-wide marker for NK cells, but its’ expression profile has not been demonstrated in canine. Therefore, we set out to determine whether NKp46 defines canine NK cells phenotypically and functionally. To this end, we generated a monoclonal mouse anti-canine NKp46 antibody by immunizing mice with L-cells expressing a canine NKp46 fusion protein. Canine peripheral blood cells were cultured in vitro with feeder cells expressing membrane bound human IL-21. The resultant cells were assessed for expression of NKp46, T-cell, B-cell, and macrophage markers. Expression of other NK cell genes were analyzed by RT-PCR. Cytotoxicity assays were conducted using a calcein-release assay with cancer cell lines. NKp46 is expressed on CD3− primary and cultured cells. Cultured CD3−/NKp46+ canine lymphocytes displayed spontaneous cytotoxicity against canine tumors and expressed NK cell activating receptors. CD3−/NKp46− (null) cultured cells were evident and had intermediate cytotoxicity between CD3+ and CD3−/NKp46+ cells. NKp46 MFI was inducible in these null cells following activation. We are currently investigating the cytokine, chemokine, and transcription factor profiling of CD3−/NKp46+ cells. We conclude that canine CD3−/NKp46+ cells are NK cells with phenotypic and functional similarity to human NK cells. In addition, canines have peripheral blood NK cell precursors that lack NKp46.
Antibody passive immunization against veterinary infectious disease
Authors: Levings Randall
Published: January 2023

The administration of antibodies for passive immunization against infectious agents has recently received increased attention due its use in human treatment for high consequence diseases (e.g., Ebola). The desire for alternatives to antimicrobial pharmaceutical use in food and companion animals has also sparked interest in this product category for veterinary species. Newtechnologies applied to veterinary passive immunization include host-species origin, chimeric, and ‘veticized’ monoclonal antibodies, as well as plantibodies. However, historic husbandry and veterinary practices, and the availability/low cost of food animal (by-)products, have resulted in continued extensive use of more conventional technologies. Injectable polyclonal serum and immunoglobulin specific for viruses, bacteria and toxins are commercially available. Oral colostrum and related products to treat failure of passive transfer in newborns are used in multiple species. Heterologous-species oral antibodies (including chicken IgY for mammalian use) for prevention of enteric disease is yet another category of product. The wide variety of immunoglobulin sources, species immunized, explicit or implied claims, regulations, and human health impacts for this category of materials provide opportunities for expansion and practical use of comparative immunology information.
Development and characterization of monoclonal antibodies specific for chicken IL-8
Authors: Kim Woohyun , Lillehoj Hyun S , Jeong Misun , Min Wongi , Sullivan Yvonne B , Kakach Laura , LaBresh Joanna W
Published: January 2023

Interleukin-8/CXCL8 (IL-8) is a CXC-family chemokine produced by fibroblasts and other cell types, including epithelial cells, endothelial cells, neutrophils, and macrophages. Given that IL-8 attracts lymphocytes to sites of tissue damage, it plays a role in the inflammatory response and wound healing. Chicken chemotactic and angiogenic factor, encoded by the 9E3/CEF4 gene, was identified and has been studied as a human IL-8 homologue for over than 20 years. However, detecting and quantifying of bioactive chemokine/cytokine in avian species is limited by the lack of specific antibodies and reliable bioassays although the use of mAbs has become routine in the mammalian research and diagnosis. Thus, we develop and characterize mouse mAbs against chIL-8 to further define its immunological and biological properties. Two mouse mAbs against chIL-8 were generated using recombinant chIL-8 protein produced in Escherichia coli. These were confirmed to display specific binding with not only recombinants, but also native chIL-8 by Western blot analysis, enzyme-linked immunosorbent assay, and immunocytochemistry. Inhibition of IL-8-induced chemotaxis of peripheral blood lymphocytes, proliferation of chicken macrophage cells and expression of alpha smooth-muscle actin in chicken embryonic fibroblast cells by both antibodies indicate that both antibodies are capable of blocking chIL-8 bioactivity and will be useful reagents for future investigations of inflammation in poultry.
Efficacy of H5 subunit vaccines to protect poultry against highly pathogenic North American H5N2 avian influenza virus
Authors: Sylte Matthew Jon , Torchetti Mia K. , Killian Mary L. , Spackman Erica , Pantin-Jackwood Mary J. , Kapczynski Darrell R. , Suarez David L. , Swayne David E.
Published: January 2023

Between December 2014 and June 2015, North America experienced the H5 highly pathogenic avian influenza (HPAI) epizootic, the largest recorded animal disease outbreak to date where over 48 million commercial poultry (e.g., chickens and turkeys) were euthanized or died from viral exposure. Soon after the epizootic began, the U.S. Department of Agriculture began testing the efficacy of different vaccines as a possible future control strategy. It was unknown whether heterologous H5 vaccines would affect infection and shedding of HPAI H5 virus in experimentally infected commercial chickens and turkeys. Three vaccine technologies were evaluated for efficacy: 1) inactivated reverse genetic laboratory-generated low pathogenicity avian influenza (LPAI) H5 virus encoding H5 hemagglutinin (HA) gene (rgH5), 2) recombinant turkey herpes virus encoding H5 HA (rHVT-H5), and 3) recombinant replication-deficient alphavirus RNA particle vaccine encoding H5 HA (RP-H5). Among all experiments, the rgH5 vaccine (clade 2.3.4.4) gave the best results in preventing mortality and reducing shedding of the North American clade 2.3.4.4 HPAI H5N2 challenge virus in chickens and turkeys. The rHVT-H5 (Clade 2.2) and RP-H5 (clade 2.3.4.4) were most efficacious as a priming vaccine, followed by booster vaccinations with rgH5-inactivated or RP-H5. Heterologous prime (rHVT-H5) and boosting (RP-H5) provided 100% protection in turkeys challenged with HPAI H5N2, whereas a single dose of RP-H5 provided partial protection. Results of these studies led to the conditional approval for RP-H5 vaccine use in poultry as a control measure for future H5 HPAI outbreaks.