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Advanced Drug Delivery Reviews May 2017The once nascent field of immunoengineering has recently blossomed to include approaches to deliver and present biomolecules to program diverse populations of... (Review)
Review
The once nascent field of immunoengineering has recently blossomed to include approaches to deliver and present biomolecules to program diverse populations of lymphocytes to fight disease. Building upon improved understanding of the molecular and physical mechanics of lymphocyte activation, varied strategies for engineering surfaces to activate and deactivate T-Cells, B-Cells and natural killer cells are in preclinical and clinical development. Surfaces have been engineered at the molecular level in terms of the presence of specific biological factors, their arrangement on a surface, and their diffusivity to elicit specific lymphocyte fates. In addition, the physical and mechanical characteristics of the surface including shape, anisotropy, and rigidity of particles for lymphocyte activation have been fine-tuned. Utilizing these strategies, acellular systems have been engineered for the expansion of T-Cells and natural killer cells to clinically relevant levels for cancer therapies as well as engineered to program B-Cells to better combat infectious diseases.
Topics: Animals; Antigen Presentation; Antigens, Surface; B-Lymphocytes; Cell Engineering; Humans; Killer Cells, Natural; Lymphocyte Activation; Lymphocytes; T-Lymphocytes
PubMed: 28501510
DOI: 10.1016/j.addr.2017.05.005 -
Deutsche Medizinische Wochenschrift... Sep 2022Autoimmune haemolytic anemia (AIHA) is defined as the immune-mediated destruction of red blood cells. In most cases, antibodies that target surface antigens on...
Autoimmune haemolytic anemia (AIHA) is defined as the immune-mediated destruction of red blood cells. In most cases, antibodies that target surface antigens on erythrocytes lead to their premature degradation in the spleen or, less commonly, in the liver. The term includes a heterogenous group of diseases, which differ largely in pathophysiology and treatment. The two most common entities are warm AIHA and cold AIHA. Diagnostic testing involves the analysis of haemolytic markers like lactate dehydrogenase, haptoglobin and unconjugated bilirubin as well as a hemoglobin and reticulocytes. In case of a haemolytic anemia, further testing like a blood smear and a direct antiglobulin test should follow. As diagnostic testing and treatment of AIHA are complex, affected patients should always be referred to a hematologist.In warm AIHA, mainly IgG autoantibodies bind to their antigen on the erythrocyte surface at body temperature, leading to their premature destruction in the spleen. First line treatment options include the administration of steroids which mitigate the destruction of red blood cells by macrophages in the spleen. In contrast, IgM autoantibodies in cold AIHA lead to intravasal agglutination of erythrocytes and complement activation. The IgM antibodies have their highest affinity below body temperature which is why patients experience symptoms mainly in cold-exposed body areas. Although the IgM antibodies dissolve at body temperature, the complement-loaded erythrocytes are destroyed in the liver. Therapeutic options include protection from cold and immunosuppressive agents or complement inhibition.
Topics: Anemia, Hemolytic, Autoimmune; Antigens, Surface; Autoantibodies; Bilirubin; Haptoglobins; Humans; Immunoglobulin G; Immunoglobulin M; Immunosuppressive Agents; Lactate Dehydrogenases
PubMed: 36126922
DOI: 10.1055/a-1767-8281 -
Methods in Molecular Biology (Clifton,... 2023Cell dissociation is an important technique for the study of tissue phenotypes. The method chosen to harvest cells from solid tissues profoundly influences the types of...
Cell dissociation is an important technique for the study of tissue phenotypes. The method chosen to harvest cells from solid tissues profoundly influences the types of cells recovered. Methodology also shapes any biases that are introduced that can act upon cell surface protein phenotypes or gene expression. Here we describe examples of cell surface phenotypic changes and typical yields, under 4 different isolation conditions (enzymatic/non-enzymatic), using the axolotl spleen, and the regenerating limb. We describe simple methods for evaluating the liberation of viable cells and the downstream characterization of cell diversity using a live-cell flow cytometry approach. Of note, the cellular composition of dissociated cells and surface antigen detection vary with each condition. TrypLE and "no enzyme" protocols give the highest surface marker expression, but poor liberation of non-immune cells in the blastema. Liberase-DH and Liberase-TL have alternative neutral proteases and both give acceptable dissociation of diverse cell types in the blastema. Liberase-TL provides the highest yield of all cell sizes and a larger non-immune fraction. Matching dissociation times between limb blastemas and spleens, we demonstrate the effect of "over-digestion" in soft tissues. In the spleen, the Liberase enzyme cocktails produced the lowest yields, worst viability, and the greatest loss of immune cell surface markers, when compared with non-enzymatic and TrypLE dissociation. These examples provide a template for optimizing protocols for individual tissues while achieving the balance between cell recovery and the mitigation of cellular changes appropriate for downstream applications such as single-cell RNA sequencing and flow cytometry.
Topics: Animals; Urodela; Flow Cytometry; Antigens, Surface; Membrane Proteins; Cell Survival
PubMed: 36272089
DOI: 10.1007/978-1-0716-2659-7_25 -
Neuro-oncology Jun 2021Immunotherapy with chimeric antigen receptor (CAR) T cells is actively being explored for pediatric brain tumors in preclinical models and early phase clinical studies....
BACKGROUND
Immunotherapy with chimeric antigen receptor (CAR) T cells is actively being explored for pediatric brain tumors in preclinical models and early phase clinical studies. At present, it is unclear which CAR target antigens are consistently expressed across different pediatric brain tumor types. In addition, the extent of HLA class I expression is unknown, which is critical for tumor recognition by conventional αβTCR T cells.
METHODS
We profiled 49 low- and high-grade pediatric brain tumor patient-derived orthotopic xenografts (PDOX) by flow analysis for the expression of 5 CAR targets (B7-H3, GD2, IL-13Rα2, EphA2, and HER2), and HLA class I. In addition, we generated B7-H3-CAR T cells and evaluated their antitumor activity in vitro and in vivo.
RESULTS
We established an expression hierarchy for the analyzed antigens (B7-H3 = GD2 >> IL-13Rα2 > HER2 = EphA2) and demonstrated that antigen expression is heterogenous. All high-grade gliomas expressed HLA class I, but only 57.1% of other tumor subtypes had detectable expression. We then selected B7-H3 as a target for CAR T-cell therapy. B7-H3-CAR T cells recognized tumor cells in an antigen-dependent fashion. Local or systemic administration of B7-H3-CAR T cells induced tumor regression in PDOX and immunocompetent murine glioma models resulting in a significant survival advantage.
CONCLUSIONS
Our study highlights the importance of studying target antigen and HLA class I expression in PDOX samples for the future design of immunotherapies. In addition, our results support active preclinical and clinical exploration of B7-H3-targeted CAR T-cell therapies for a broad spectrum of pediatric brain tumors.
Topics: Animals; Antigens, Surface; B7 Antigens; Brain Neoplasms; Child; Humans; Mice; Receptors, Chimeric Antigen; T-Lymphocytes; Xenograft Model Antitumor Assays
PubMed: 33320196
DOI: 10.1093/neuonc/noaa278 -
Journal of Translational Medicine Oct 2023Chimeric antigen receptor NK (CAR-NK) cell therapy is one of the most promising immunotherapies. Although it has shown a significant therapeutic effect in hematologic...
BACKGROUND
Chimeric antigen receptor NK (CAR-NK) cell therapy is one of the most promising immunotherapies. Although it has shown a significant therapeutic effect in hematologic malignancies, few successes have been obtained in solid tumors including esophageal squamous cell carcinoma (ESCC). The major reasons are lack of specific cell surface antigens and complex tumor microenvironment. Here we identify CD22, a well-known tumor surface marker in hematologic malignancies, is expressed in ESCC, possibly serving as a potential target of CAR-NK cell therapy.
METHODS
The expression of 13 tumor cell surface antigens used clinically was analyzed in patients from The Cancer Genome Atlas (TCGA) database. Also, mRNA expression were detected in 2 ESCC cell lines and 2 patients samples by qCPR. Then according to Venn diagram, CD22 was selected for further investigation. Following this, the expression of CD22 by immunofluorescence (IF) in ESCC cell lines and by immunohistochemistry (IHC) in 87 cases of human ESCC samples was detected respectively. On the basis of H-score results, the correlation between CD22 expression and clinical parameters was analyzed. As a proof, the efficacy of CD22-targeted CAR-NK cells against ESCC cell lines was performed by a real-time cell analyzer (RTCA) platform.
RESULTS
KYSE-140 and KYSE-150 cell lines displayed surface expression of CD22. IHC showed an 80.46% (70/87) positive rate in ESCC patient samples. Among these, cell membranous expression of CD22 was observed in 27.59% (24/87) patient samples. Through chi-square test, expression of CD22 in ESCC was associated with lymph node metastasis while it was no related to the depth of tumor invasion and clinical stage. Engineered CD22-targeted CAR-NK cells exhibited inhibitory growth capability against ESCC cell lines (p < 0.0001).
CONCLUSIONS
CD22 is a potential tumor surface antigen capable of being targeted by CAR-NK cells in ESCC. And potential therapeutics for ESCC may be developed based on immune cells expressing anti-CD22 CAR. The study also indicates that CD22 CAR-NK cells could be used in other cancers and more in vivo experiments are needed.
Topics: Humans; Esophageal Squamous Cell Carcinoma; Esophageal Neoplasms; Carcinoma, Squamous Cell; Biomarkers, Tumor; Killer Cells, Natural; Hematologic Neoplasms; Antigens, Surface; Cell- and Tissue-Based Therapy; Cell Line, Tumor; Tumor Microenvironment; Sialic Acid Binding Ig-like Lectin 2
PubMed: 37817249
DOI: 10.1186/s12967-023-04409-8 -
Signal Transduction and Targeted Therapy May 2023Current attempts in vaccine delivery systems concentrate on replicating the natural dissemination of live pathogens, but neglect that pathogens evolve to evade the...
Current attempts in vaccine delivery systems concentrate on replicating the natural dissemination of live pathogens, but neglect that pathogens evolve to evade the immune system rather than to provoke it. In the case of enveloped RNA viruses, it is the natural dissemination of nucleocapsid protein (NP, core antigen) and surface antigen that delays NP exposure to immune surveillance. Here, we report a multi-layered aluminum hydroxide-stabilized emulsion (MASE) to dictate the delivery sequence of the antigens. In this manner, the receptor-binding domain (RBD, surface antigen) of the spike protein was trapped inside the nanocavity, while NP was absorbed on the outside of the droplets, enabling the burst release of NP before RBD. Compared with the natural packaging strategy, the inside-out strategy induced potent type I interferon-mediated innate immune responses and triggered an immune-potentiated environment in advance, which subsequently boosted CD40 DC activations and the engagement of the lymph nodes. In both H1N1 influenza and SARS-CoV-2 vaccines, rMASE significantly increased antigen-specific antibody secretion, memory T cell engagement, and Th1-biased immune response, which diminished viral loads after lethal challenge. By simply reversing the delivery sequence of the surface antigen and core antigen, the inside-out strategy may offer major implications for enhanced vaccinations against the enveloped RNA virus.
Topics: Humans; Antigens, Viral; COVID-19 Vaccines; Influenza A Virus, H1N1 Subtype; COVID-19; SARS-CoV-2; Vaccination; Antigens, Surface; Antibodies
PubMed: 37221173
DOI: 10.1038/s41392-023-01414-7 -
Brain Research Bulletin Oct 2022Astrocytes are the main support cells of the central nervous system. They also participate in neuroimmune reactions. In response to pathological and immune stimuli,... (Review)
Review
Astrocytes are the main support cells of the central nervous system. They also participate in neuroimmune reactions. In response to pathological and immune stimuli, astrocytes transform to reactive states characterized by increased release of inflammatory mediators. Some of these molecules are neuroprotective and inflammation resolving while others, including reactive oxygen species (ROS), nitric oxide (NO), matrix metalloproteinase (MMP)- 9, L-glutamate, and tumor necrosis factor α (TNF), are well-established toxins known to cause damage to surrounding cells and tissues. We hypothesized that similar to microglia, the brain immune cells, reactive astrocytes can release a broader set of diverse molecules that are potentially neurotoxic. A literature search was conducted to identify such molecules using the following two criteria: 1) evidence of their expression and secretion by astrocytes and 2) direct neurotoxic action. This review describes 14 structurally diverse molecules as less-established astrocyte neurotoxins, including C-X-C motif chemokine ligand (CXCL)10, CXCL12/CXCL12(5-67), FS-7-associated surface antigen ligand (FasL), macrophage inflammatory protein (MIP)- 2α, TNF-related apoptosis inducing ligand (TRAIL), pro-nerve growth factor (proNGF), pro-brain-derived neurotrophic factor (proBDNF), chondroitin sulfate proteoglycans (CSPGs), cathepsin (Cat)B, group IIA secretory phospholipase A (sPLA-IIA), amyloid beta peptides (Aβ), high mobility group box (HMGB)1, ceramides, and lipocalin (LCN)2. For some of these molecules, further studies are required to establish either their direct neurotoxic effects or the full spectrum of stimuli that induce their release by astrocytes. Only limited studies with human-derived astrocytes and neurons are available for most of these potential neurotoxins, which is a knowledge gap that should be addressed in the future. We also summarize available evidence of the role these molecules play in select neuropathologies where reactive astrocytes are a key feature. A comprehensive understanding of the full spectrum of neurotoxins released by reactive astrocytes is key to understanding neuroinflammatory diseases characterized by the adverse activation of these cells and may guide the development of novel treatment strategies.
Topics: Amyloid beta-Peptides; Antigens, Surface; Astrocytes; Brain-Derived Neurotrophic Factor; Cathepsins; Ceramides; Chemokines; Chondroitin Sulfate Proteoglycans; Glutamic Acid; HMGB Proteins; Humans; Inflammation Mediators; Ligands; Lipocalins; Macrophage Inflammatory Proteins; Microglia; Neurotoxicity Syndromes; Neurotoxins; Nitric Oxide; Phospholipases A2, Secretory; Reactive Oxygen Species; Tumor Necrosis Factor-alpha
PubMed: 35988785
DOI: 10.1016/j.brainresbull.2022.08.015 -
Methods in Molecular Biology (Clifton,... 2022Massive parallel sequencing technology has greatly increased the breadth and depth of transcriptomic data that can be captured from P. falciparum samples. This has...
Massive parallel sequencing technology has greatly increased the breadth and depth of transcriptomic data that can be captured from P. falciparum samples. This has revolutionized in vitro studies but uptake has been slower in the analysis of clinical samples. The principal barriers are the removal of contaminating white blood cells in a malaria endemic setting and preservation of the RNA. We provide here detailed methods for the collection of purified infected erythrocytes and the preservation and extraction of RNA. We also provide methods for assessing and addressing contaminating RNA from erythroid cells, and a protocol for RNAseq library preparation optimized to maximize yield from low amounts of parasite mRNA. Finally, we provide some examples of RNAseq library characteristics that may fail quality control for other species but are in fact satisfactory for P. falciparum RNAseq.
Topics: Antigens, Surface; Erythrocytes; Humans; Malaria, Falciparum; Plasmodium falciparum; Protozoan Proteins; RNA, Messenger
PubMed: 35881347
DOI: 10.1007/978-1-0716-2189-9_15 -
Expert Review of Vaccines Feb 2021Transmission-blocking vaccines (TBV) prevent community spread of malaria by targeting mosquito sexual stage parasites, a life-cycle bottleneck, and will be used in... (Review)
Review
INTRODUCTION
Transmission-blocking vaccines (TBV) prevent community spread of malaria by targeting mosquito sexual stage parasites, a life-cycle bottleneck, and will be used in elimination programs. TBV rely on herd immunity to reduce mosquito infections and thereby new infections in both vaccine recipients and non-recipients, but do not provide protection once an individual receives an infectious mosquito bite which complicates clinical development.
AREAS COVERED
Here, we describe the concept and biology behind TBV, and we provide an update on clinical development of the leading vaccine candidate antigens. Search terms 'malaria vaccine,' 'sexual stages,' 'transmission blocking vaccine,' 'VIMT' and 'SSM-VIMT' were used for PubMed queries to identify relevant literature.
EXPERT OPINION
Candidates targeting zygote surface antigen Pfs25, and its orthologue Pvs25, induced functional activity in humans that reduced mosquito infection in surrogate assays, but require increased durability to be useful in the field. Candidates targeting gamete surface antigens Pfs230 and Pfs48/45, respectively, are in or nearing clinical trials. Nanoparticle platforms and adjuvants are being explored to enhance immunogenicity. Efficacy trials require special considerations, such as cluster-randomized designs to measure herd immunity that reduces human and mosquito infection rates, while addressing human and mosquito movements as confounding factors.
Topics: Animals; Antibodies, Protozoan; Antigens, Surface; Humans; Immunity, Herd; Malaria Vaccines; Malaria, Falciparum; Malaria, Vivax; Mosquito Control; Mosquito Vectors; Plasmodium falciparum; Plasmodium vivax
PubMed: 33478283
DOI: 10.1080/14760584.2021.1878028 -
ACS Applied Bio Materials Oct 2021The expression patterns of surface antigens are associated with the differentiation status and functional characteristics of mammalian cells. To analyze the surface...
The expression patterns of surface antigens are associated with the differentiation status and functional characteristics of mammalian cells. To analyze the surface antigen expression pattern in a high-throughput manner, antibody microarrays have been developed by several groups, including ours. This analysis can be performed using cell-binding assays on microarrays; moreover, this approach has advantages over conventional flow cytometry (FCM). Unlike FCM, the microarray-based method cannot evaluate the concurrent expression of more than two surface antigens on a single cell, and therefore, it cannot be used for cell subset analysis. To overcome this drawback, we prepared an antibody microarray with spots presenting co-immobilized multiple antibodies together with spots presenting each antibody separately. The co-immobilized spots are expected to be reactive for every surface antigen specific to the co-immobilized antibodies. In addition, the concept of an algebra of sets is incorporated into the derivation of quantitative data regarding cell subsets. Here, taking cell subsets with respect to two surface antigens as the simplest example, antibody microarrays were prepared and initially subjected to validation studies to verify the accuracy of cell-binding assays. Quantitative subset analysis was performed using antibody microarrays prepared using the anti-CD13 and anti-CD49f antibodies. For model populations that consisted of discrete subsets, THP-1, HL-60, CCRF-CEM, and Ramos cell lines were used because they were found by FCM to have a singular phenotype, that is, CD13CD49f, CD13CD49f, CD13CD49f, and CD13CD49f, respectively. Five populations were prepared by mixing these cells at various ratios and analyzed for their subsets using microarrays. The results showed that the experimentally determined abundance ratios of the four model subsets were in good agreement with the predetermined abundance ratios, which provided the proof of principle for the new method in the quantitative subset analysis.
Topics: Animals; Antibodies; Antigens, Surface; Flow Cytometry; Integrin alpha6; Mammals; Microarray Analysis
PubMed: 35006690
DOI: 10.1021/acsabm.1c00898