Ligand Antibody Research Articles

Fluorine-18-Labeled Antibody Ligands for PET Imaging of Amyloid-β in Brain.

Antibodies are attractive as radioligands due to their outstanding specificity and high affinity, but their inability to cross the blood–brain barrier (BBB) limits their use for CNS targets. To enhance brain distribution, amyloid-β (Aβ) antibodies were fused to a transferrin receptor (TfR) antibody fragment, enabling receptor mediated transport across the BBB. The aim of this study was to label these bispecific antibodies with fluorine-18 and use them for Aβ PET imaging. Bispecific antibody ligands RmAb158-scFv8D3 and Tribody A2, both targeting Aβ and TfR, were functionalized with trans-cyclooctene (TCO) groups and conjugated with 18F-labeled tetrazines through an inverse electron demand Diels–Alder reaction performed at ambient temperature. 18F-labeling did not affect antibody binding in vitro, and initial brain uptake was high. Conjugates with the first tetrazine variant ([18F]T1) displayed high uptake in bone, indicating extensive defluorination, a problem that was resolved with the second and third tetrazine variants ([18F]T2 and [18F]T3). Although the antibody ligands’ half-life in blood was too long to optimally match the physical half-life of fluorine-18 (t1/2 = 110 min), [18F]T3-Tribody A2 PET seemed to discriminate transgenic mice (tg-ArcSwe) with Aβ deposits from wild-type mice 12 h after injection. This study demonstrates that 18F-labeling of bispecific, brain penetrating antibodies is feasible and, with further optimization, could be used for CNS PET imaging.

Measurement of Integrin Activation and Conformational Changes on the Cell Surface by Soluble Ligand and Antibody Binding Assays.

Soluble ligand and conformation-dependent antibody binding assay of integrins on the cell surface is an effective approach to evaluate the activation status of integrins in live cells. The ligands or antibodies are usually labeled with biotin or a fluorescent dye and incubated with integrin-expressing cells in suspension. The cell-bound ligands and antibodies are then detected by flow cytometry. Here we describe the detailed protocols of soluble ligand or antibody binding assay for αIIbβ3, αVβ3, α5β1, and αLβ2 integrins that are transiently or stably expressed in the model cell lines such as HEK293 or CHO-k1 cells.

Long-acting antibody ligand mimetics for HER4-selective agonism

Neuregulin protein 1 (NRG1) is a large (> 60–amino-acid) natural peptide ligand for the ErbB protein family members HER3 and HER4. We developed an agonistic antibody modality, termed antibody ligand mimetics (ALM), by incorporating complex ligand agonists such as NRG1 into an antibody scaffold. We optimized the linker and ligand length to achieve native ligand activity in HEK293 cells and cardiomyocytes derived from induced pluripotent stem cells (iPSCs) and used a monomeric Fc-ligand fusion platform to steer the ligand specificity toward HER4-dominant agonism. With the help of selectivity engineering, these enhanced ALM molecules can provide an antibody scaffold with increased receptor specificity and the potential to greatly improve the pharmacokinetics, stability, and downstream developability profiles from the natural ligand approach. This ligand mimetic design and optimization approach can be expanded to apply to other cardiovascular disease targets and emerging therapeutic areas, providing differentiated drug molecules with increased specificity and extended half-life.

Design and site-directed immobilization of single-chain Fv antibody to polystyrene latex beads via material-binding peptides and application to latex turbidimetric assay

In this study, immobilization of single-chain Fv (scFv) antibodies on the surfaces of polystyrene (PS) latex beads via material-binding peptides was investigated for sensitive immuno-turbidimetric assay of C-reactive protein (CRP). Anti-CRP scFvs fused with polystyrene-binding peptide (PS-tag) and poly(methylmethacrylate)-binding peptide (PMMA-tag) were over-expressed in Escherichiacoli cells and recovered in the active form following refolding. The beads with PMMA-tag-fused scFv (scFv-PM) were successfully suspended with sufficient dispersion at pH 8.0. Three types of alternative scFv-PMs with a penta-asparatic acid tag (D5-tag) introduced at different positions were then designed. All of the D5-tagged scFv-PMs were successfully immobilized on the surfaces of beads with no significant change in the diameter of the latex beads at pH levels ranging from 6.0 to 8.0. According to the results of turbidimetric assay for the detection of CRP, 13ng/ml of CRP was detectable using beads with D5-tagged scFv-PMs at 400ng/cm3, and no turbidity change was observed in the absence of antigen. When the density of scFv-PM was 250ng/cm2, which was 63% of the maximum density, the beads were dispersed well and reactive with the antigen at a concentration range comparable to those with D5-tagged scFv-PMs. These results indicate that controlling charge density on the surface of beads after site-directed immobilization is definitely important in order to maintain high levels of dispersion and reactivity. Thus, the usefulness of the scFv-PM as well as D5-tagged scFv-PMs developed in the present study should be significant when used as ligand antibodies in the preparation of immuno-latex beads.

Extracellular Vesicles as Biomarkers in Cancer Immunotherapy.

Simple SummaryExtracellular vesicles (EVs) are small particles found throughout the body. EVs are released by living cells and contain cargo representing the cell of origin. In recent years, EVs have gained attention in cancer research. Since the cargo found inside EVs can be traced back to the cell of origin, EVs shed from cancer cells, in particular, may be used to better describe and characterize a patient’s tumor. EVs have been found and isolated from a variety of bodily fluids, including blood, saliva, and amniotic fluid, and therefore offer a non-invasive way of also diagnosing and monitoring patients before, during, and after cancer immunotherapy. The aim of this review article was to summarize some of the recent work conducted in this field and the challenges we face moving forward in utilizing EVs for cancer diagnostic and therapeutic purposes in cancer immunotherapy in the clinical setting.Extracellular vesicles (EVs), including exosomes and microvesicles, are membrane-bound vesicles secreted by most cell types during both physiologic conditions as well in response to cellular stress. EVs play an important role in intercellular communication and are emerging as key players in tumor immunology. Tumor-derived EVs (TDEs) harbor a diverse array of tumor neoantigens and contain unique molecular signature that is reflective of tumor’s underlying genetic complexity. As such they offer a glimpse into the immune tumor microenvironment (TME) and have the potential to be a novel, minimally invasive biomarker for cancer immunotherapy. Immune checkpoint inhibitors (ICI), such as anti- programmed death-1(PD-1) and its ligand (PD-L1) antibodies, have revolutionized the treatment of a wide variety of solid tumors including head and neck squamous cell carcinoma, urothelial carcinoma, melanoma, non-small cell lung cancer, and others. Typically, an invasive tissue biopsy is required both for histologic diagnosis and next-generation sequencing efforts; the latter have become more widespread in daily clinical practice. There is an unmet need for noninvasive or minimally invasive (e.g., plasma-based) biomarkers both for diagnosis and treatment monitoring. Targeted analysis of EVs in biospecimens, such as plasma and saliva could serve this purpose by potentially obviating the need for tissue sample. In this review, we describe the current challenges of biomarkers in cancer immunotherapy as well as the mechanistic role of TDEs in modulating antitumor immune response.

The Vitamin D Receptor in Osteoblast-Lineage Cells Is Essential for the Proresorptive Activity of 1α,25(OH)2D3 In Vivo.

We previously reported that daily administration of a pharmacological dose of eldecalcitol, an analog of 1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3], increased bone mass by suppressing bone resorption. These antiresorptive effects were found to be mediated by the vitamin D receptor (VDR) in osteoblast-lineage cells. Using osteoblast-lineage-specific VDR conditional knockout (Ob-VDR-cKO) mice, we examined whether proresorptive activity induced by the high-dose 1α,25(OH)2D3 was also mediated by VDR in osteoblast-lineage cells. Administration of 1α,25(OH)2D3 (5 μg/kg body weight/day) to wild-type mice for 4 days increased the number of osteoclasts in bone and serum concentrations of C-terminal crosslinked telopeptide of type I collagen (CTX-I, a bone resorption marker). The stimulation of bone resorption was concomitant with the increase in serum calcium (Ca) and fibroblast growth factor 23 (FGF23) levels, and decrease in body weight. This suggests that a toxic dose of 1α,25(OH)2D3 can induce bone resorption and hypercalcemia. In contrast, pretreatment of wild-type mice with neutralizing anti-receptor activator of NF-κB ligand (RANKL) antibody inhibited the 1α,25(OH)2D3-induced increase of osteoclast numbers in bone, and increase of CTX-I, Ca, and FGF23 levels in serum. The pretreatment with anti-RANKL antibody also inhibited the 1α,25(OH)2D3-induced decrease in body weight. Consistent with observations in mice conditioned with anti-RANKL antibody, the high-dose administration of 1α,25(OH)2D3 to Ob-VDR-cKO mice failed to significantly increase bone osteoclast numbers, serum CTX-I, Ca, or FGF23 levels, and failed to reduce the body weight. Taken together, this study demonstrated that the proresorptive, hypercalcemic, and toxic actions of high-dose 1α,25(OH)2D3 are mediated by VDR in osteoblast-lineage cells.

Targeting respiratory diseases using miRNA inhibitor based nanotherapeutics: Current status and future perspectives

MicroRNAs (miRNAs) play a fundamental role in the developmental and physiological processes that occur in both animals and plants. AntagomiRs are synthetic antagonists of miRNA, which prevent the target mRNA from suppression. Therapeutic approaches that modulate miRNAs have immense potential in the treatment of chronic respiratory disorders. However, the successful delivery of miRNAs/antagomiRs to the lungs remains a major challenge in clinical applications. A range of materials, namely, polymer nanoparticles, lipid nanocapsules and inorganic nanoparticles, has shown promising results for intracellular delivery of miRNA in chronic respiratory disorders. This review discusses the current understanding of miRNA biology, the biological roles of antagomiRs in chronic respiratory disease and the recent advances in the therapeutic utilization of antagomiRs as disease biomarkers. Furthermore our review provides a common platform to debate on the nature of antagomiRs and also addresses the viewpoint on the new generation of delivery systems that target antagomiRs in respiratory diseases.

Research progress of immune checkpoint inhibitors in clinical diagnosis and treatment of non-small cell lung cancer

Lung cancer is currently the malignant tumor with the highest morbidity and mortality in the world, and the main type is non-small cell lung cancer. Immune checkpoint inhibitor is a landmark discovery in the history of cancer treatment, which rewrites the history of cancer treatment, and improves the medical treatment of advanced tumors by a big step forward. The article summarizes the research progress of therapeutic drugs against anti-programmed cell death protein and programmed cell death protein ligand antibodies in the clinical diagnosis and treatment of non-small cell lung cancer. The principle of drug action, the differences in the diagnosis and treatment of non-small cell lung cancer in different clinical stages, and future research directions are discussed to provide the usage guidelines of immune checkpoint inhibitors for clinical oncologists.

Synthetic nanobodies as angiotensin receptor blockers

There is considerable interest in developing antibodies as functional modulators of G protein-coupled receptor (GPCR) signaling for both therapeutic and research applications. However, there are few antibody ligands targeting GPCRs outside of the chemokine receptor group. GPCRs are challenging targets for conventional antibody discovery methods, as many are highly conserved across species, are biochemically unstable upon purification, and possess deeply buried ligand-binding sites. Here, we describe a selection methodology to enrich for functionally modulatory antibodies using a yeast-displayed library of synthetic camelid antibody fragments called "nanobodies." Using this platform, we discovered multiple nanobodies that act as antagonists of the angiotensin II type 1 receptor (AT1R). Following angiotensin II infusion in mice, we found that an affinity matured nanobody antagonist has comparable antihypertensive activity to the angiotensin receptor blocker (ARB) losartan. The unique pharmacology and restricted biodistribution of nanobody antagonists may provide a path for treating hypertensive disorders when small-molecule drugs targeting the AT1R are contraindicated, for example, in pregnancy.

Dynamic polarization shifting from M1 to M2 macrophages in reduced osteonecrosis of the jaw-like lesions by cessation of anti-RANKL antibody in mice.

Denosumab-related osteonecrosis of the jaw (DRONJ), which mainly occurs in cancer patients receiving anti-receptor activator NF-kappaB ligand (RANKL) antibody, reduces oral health-related quality of life. However, the exact mechanisms of and definitive treatment strategies for DRONJ remain unknown. We hypothesized that cessation of denosumab heals and/or ameliorates DRONJ, since it is a protein-based antibody agent, although stopping denosumab should be avoided in clinical situations. Therefore, the aims of this study were: 1) to create a healing and/or amelioration murine model of DRONJ-like lesions induced by chemotherapy/anti-RANKL antibody (mAb) combination therapy and tooth extraction; and 2) to investigate histopathology and immunopathology in the extraction sockets by comparing the murine model of DRONJ-like lesions with the amelioration/healing model of DRONJ-like lesions. Eight-week-old, female C57B/6J mice received chemotherapeutic drug (cyclophosphamide: CY) and mAb combination therapy (CY/mAb) with tooth extraction. Open wounds were sustained in CY/mAb-treated mice at 2 and 4 weeks post-extraction. Impaired socket healing was diagnosed as CY/mAb-related ONJ-like lesions at 3 weeks post-extraction in this study. Next, mAb was discontinued for 2 and 4 weeks after diagnosis of CY/mAb-related ONJ-like lesions. mAb cessation for 2 weeks induced partial osseous wound healing and significantly improved soft tissue wound healing of the extraction sockets. Anti-angiogenesis and normal lymphangiogenesis with CY/mAb combination therapy was not changed by mAb discontinuation. However, mAb cessation for 2 weeks significantly increased the number of CD38+F4/80+ M1 and CD163+F4/80+ M2 macrophages, which significantly increased the M2/M1 ratio in the connective tissue of extraction sockets. No direct effects of mAb on macrophages were noted both in vivo and in vitro. Therefore, the developed healing and/or amelioration murine model of CY/mAb-related ONJ-like lesions is a useful tool to investigate the histopathology and immunopathology of DRONJ in humans. Dynamic polarization shifting from M1 to M2 macrophages induced by mAb cessation may play an important role in wound healing, rather than angiogenesis and lymphangiogenesis, in DRONJ.

Identifying the preferable rheumatoid arthritis subgroups for intervention with the anti-RANKL antibody denosumab to reduce progression of joint destruction

ObjectivesTo clarify which rheumatoid arthritis (RA) patients benefit most from the anti-receptor activator of nuclear factor-κB ligand antibody denosumab to reduce the progression of joint destruction.MethodsWe pooled patient data from...

Sustained anti-osteoporotic action of risedronate compared to anti-RANKL antibody following discontinuation in ovariectomized mice

Bisphosphonates and the anti-receptor activator of nuclear factor-kappa B ligand (RANKL) antibody denosumab are effective anti-resorptive drugs commonly prescribed for osteoporosis. Both drugs may, however, have intolerable side effects; so, it is critical to examine their residual efficacy such as maintenance of bone mass following cessation. Therefore, we compared the changes in bone histology following discontinuation of the aminobisphosphonate risedronate and anti-RANKL antibody in ovariectomized (OVX) mice. Twelve-week-old female C57BL/6 N mice were OVX or sham operated. Four weeks after surgery, mice were treated with vehicle, a single injection of anti-RANKL antibody (5 mg/kg), or risedronate (5 μg/kg/day, s.c.) for 4 weeks (the treatment period), followed by vehicle treatment for an additional 4 weeks (discontinuation period). The lumbar spine and proximal tibia were evaluated by micro-computed tomography. In addition, the lumbar spine, proximal tibia, and the femoral shaft were examined by bone histomorphometry. After 4 weeks of discontinuation, OVX mice initially treated with the anti-RANKL antibody exhibited a trend of bone loss associated with increased turnover in both trabecular and cortical bones, although the difference was not significant. By contrast, OVX mice treated with risedronate exhibited maintained or even increased bone mass and suppressed bone turnover. Patients discontinuing denosumab should be carefully monitored for recurrent osteoporosis symptoms, and a replacement drug should be considered.

Functional anomaly of CD8 T cells at earlier differentiation stages during HIV infection as established by analysis of synaptic interface and degranulation pattern

Abstract It is thought that HIV infection could lead to a global defect in CD8+ T cell function. However, the effect of the infection on CD8+ T cells at various stages of differentiation has not been well studied. To evaluate the functional activity of CD8+ T cells from HIV infected people at various differentiation stages, we assessed their ability to form synaptic interfaces using planar lipid bilayers containing anti-CD3 antibody and soluble ICAM-1 ligand. We analyzed the dynamics of the T cell/bilayer interface formation, size of adhesion area, and kinetics of T-cell degranulation, parameters that are linked to the efficiency of CD8+ T cell functional activity. Unexpectedly, we observed that purified naïve CD8+ T cells (TN) and transitional memory (TTM) from HIV-infected individuals developed mature synapses whereas the same subsets from HIV uninfected individuals did not. While the kinetics of degranulation of naïve T cells was slower, both TN and TTM cells reached a similar extent of degranulation. The fraction and kinetics of CD27-CD45RO+ T cell degranulation were similar to that observed for TTM cells. Treatment with ART led to increase in the fraction of degranulating CD27-CD45RO+ T cells. As opposed to memory cells from healthy donors, a sizable fraction of CD27-CD45RO+ T cells from infected individuals develop lamellopodia and a larger adhesion area after synapse formation. These data suggest that less differentiated CD8+ T cells from HIV-infected people revealed functional anomaly that is likely mediated by aberrant functions of integrins and cytoskeleton causing changes in T-cell plasticity as a result of continuous inflammation during chronic HIV infection.

Nanoparticle interactions with immune cells dominate tumor retention and induce T cell-mediated tumor suppression in models of breast cancer.

The factors that influence nanoparticle fate in vivo following systemic delivery remain an area of intense interest. Of particular interest is whether labeling with a cancer-specific antibody ligand ("active targeting") is superior to its unlabeled counterpart ("passive targeting"). Using models of breast cancer in three immune variants of mice, we demonstrate that intratumor retention of antibody-labeled nanoparticles was determined by tumor-associated dendritic cells, neutrophils, monocytes, and macrophages and not by antibody-antigen interactions. Systemic exposure to either nanoparticle type induced an immune response leading to CD8+ T cell infiltration and tumor growth delay that was independent of antibody therapeutic activity. These results suggest that antitumor immune responses can be induced by systemic exposure to nanoparticles without requiring a therapeutic payload. We conclude that immune status of the host and microenvironment of solid tumors are critical variables for studies in cancer nanomedicine and that nanoparticle technology may harbor potential for cancer immunotherapy.

Binding-Induced Stabilization Measured on the Same Molecular Protein Substrate Using Single-Molecule Magnetic Tweezers and Heterocovalent Attachments.

Binding-induced mechanical stabilization plays key roles in proteins involved in muscle contraction, cellular mechanotransduction, or bacterial adhesion. Because of the vector nature of force, single-molecule force spectroscopy techniques are ideal for measuring the mechanical unfolding of proteins. However, current approaches are still prone to calibration errors between experiments and geometrical variations between individual tethers. Here, we introduce a single-molecule assay based on magnetic tweezers and heterocovalent attachment, which can measure the binding of the substrate-ligand using the same protein molecule. We demonstrate this approach with protein L, a model bacterial protein which has two binding interfaces for the same region of kappa-light chain antibody ligands. Engineered molecules with eight identical domains of protein L between a HaloTag and a SpyTag were exposed to repeated unfolding-refolding cycles at forces up to 100 pN for several hours at a time. The unfolding behavior of the same protein was measured in solution buffers with different concentrations of antibody ligands. With increasing antibody concentration, an increasing number of protein L domains became more stable, indicative of ligand binding and mechanical reinforcement. Interestingly, the dissociation constant of the mechanically reinforced states coincides with that measured for the low-avidity binding interface of protein L, suggesting a physiological role for the second binding interface. The molecular approach presented here opens the road to a new type of binding experiments, where the same molecule can be exposed to different solvents or ligands.

Glycolysis and Oxidative Phosphorylation Play Critical Roles in Natural Killer Cell Receptor-Mediated Natural Killer Cell Functions.

Natural killer (NK) cells are innate lymphocytes that directly kill tumor and pathogen-infected cells upon activation by cytokines and NK cell receptors (NKRs) without previous sensitization. It is known that cell metabolism affects the differentiation and effector functions of immune cells. For instance, interleukin−2 and interleukin−15 treatment increases glycolysis and oxidative phosphorylation (OXPHOS) in NK cells to support their effector functions. However, little is known about the metabolic reprogramming of human NK cells upon their activation by NKRs. In this study, we investigated the metabolism of NK cells stimulated via NKRs. We found that NK cells upregulated glycolysis and OXPHOS in response to anti-CD16 antibody or NKG2D ligand engagement. Inhibition of either glycolysis or OXPHOS impaired NK cell production of interferon-γ. Interestingly, inhibition of glycolysis but not OXPHOS decreased NK cell killing and dampened NK cell degranulation and Fas ligand expression, suggesting that glycolysis is more critical for NKR-activated cell cytotoxicity. Thus, our study provides insight into understanding the metabolic requirements underlying different effector functions of human NK cells.

Luminescent Nanocomposites for Biomedical Applications

In this work, we pursue a concept of the multimodal drug preparation based on LuAG:Ce3+ nanoparticles. The functionality studied in this article is focused on the bioimaging of living cells via cathodoluminescence (CL) of scintillating nanoparticles attached to the cell surface. LuAG:Ce3+ scintillating nanoparticles were used as a core; amorphous or mesoporous SiO2 was used as a shell encapsulating the core to increase the surface reactivity; anti-Fas ligand (FasL) antibody was then attached to the outer shell in order to specifically bond the nanocomposite to the cell receptors; finally, polyethylene glycol molecules were attached to increase the biocompatibility of the nanocomposites. CL bioimaging was tested on yeasts Saccharomyces cerevisiae and leukocytes. It was found that the nanocomposites are able to attach to the cells via sorption or bonding to the receptors. The prepared nanocomposite shows stable luminescence under 10-keV electron irradiation, thus making a prospective candidate for this application and future multimodal drug design.

P063 B-cell receptor signalling in lymphoid tissues may be regulated by CEACAM1

Abstract Background It has been recently shown that the carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) expressed in T cells may regulate immune responses in the gut. Moreover, it has also been reported that the treatments with either an agonistic monoclonal antibody (mAb) or natural ligands for this molecule can suppress colitis severity in murine models of inflammatory bowel diseases (IBD). On the other hand, in addition to T cells, B cells are also an important population in the gut-associated lymphoid tissues (GALT) that orchestrate mucosal homeostasis. However, the role of CEACAM1 in B cells has not been elucidated. Methods We analysed primary B-cell subsets in the lymphoid tissues of wild-type C57BL6 mice as well as a murine B-cell line, A20, to determine the expressions and functions of CEACAM1. Results FACS analysis of the lymphocyte subsets isolated from secondary lymphoid tissues such as spleen, mesenteric lymph nodes and Peyer’s patches of C57BL6 revealed higher expression level of CEACAM1 on B-cell surface than that of T cells. Bone marrow analysis showed that such CEACAM1 expression was increased during maturation and differentiation process of B cells. When isolated splenic B cells were stimulated with LPS, anti-CD40 or anti-μ chain Abs in the presence of agonistic anti-CEACAM1 mAb, the usual increased cytokine productions (such as IL-4 and IL-5 by activation via B cell receptor (BCR) signalling) were specifically suppressed by CEACAM1 signalling rather than B-cell activations via either TLR4 or CD40 signalling. Immunofluorescent studies using confocal microscopy revealed co-localisation of CEACAM1 and BCR when B cells were activated with anti-μ chain Ab. Given these results, A20 cells were transfected with CEACAM1 cDNA. Biochemical analysis showed that an inducible overexpression of CEACAM1 suppressed the BCR signalling in these cells when compared with that of vector alone-transfected control. Moreover, the overexpression of CEACAM1 in these cells resulted in reduced expressions of activation markers such as CD69, CD80, CD86, MHC-I and -II on the cell surface. These observations were associated with decreased Ca2+ influx and suppressed cytokine production by the overexpression of CEACAM1 after BCR signal activation. Conclusion These results suggest that CEACAM1 can regulate B-cell activation and differentiation specifically via BCR signalling in the lymphoid tissues. Therefore, this molecule can be a therapeutic target in IBD by regulating of both T-cell and B-cell activation in GALT.

Zwitterionic Phosphodiester-Substituted Neoglycoconjugates as Ligands for Antibodies and Acute Phase Proteins.

Zwitterionic modifications of glycans, such as phosphorylcholine and phosphoethanolamine, are known from a range of prokaryotic and eukaryotic species and are recognized by mammalian antibodies and pentraxins; however, defined saccharide ligands modified with these zwitterionic moieties for high-throughput studies are lacking. In this study, we prepared and tested example mono- and disaccharides 6-substituted with either phosphorylcholine or phosphoethanolamine as bovine serum albumin neoglycoconjugates or printed in a microarray format for subsequent assessment of their binding to lectins, pentraxins, and antibodies. C-Reactive protein and anti-phosphorylcholine antibodies bound specifically to ligands with phosphorylcholine, but recognition by concanavalin A was abolished or decreased as compared with that to the corresponding nonzwitterionic compounds. Furthermore, in array format, the phosphorylcholine-modified ligands were recognized by IgG and IgM in sera of either non-infected or nematode-infected dogs and pigs. Thereby, these new compounds are defined ligands which allow the assessment of glycan-bound phosphorylcholine as a target of both the innate and adaptive immune systems in mammals.

Sensitization to cell death induced by soluble Fas ligand and agonistic antibodies with exogenous agents: A review

Specific binding of either soluble Fas ligand extracellular domain (sFasL) or agonistic anti-Fas receptor extracellular-domain monoclonal-antibodies (FasR-mAb) can trigger apoptotic death of emerging harmful cells in the human body. However, the efficient cell-death induction through the action of these executors are often prevented by the resistance mechanisms equipped with the target cells. Hence, strengthening their cell-death inducing activity by sensitization with the help of exogenous agents will contribute to the development of advanced treatment strategies for many serious diseases caused by impaired cell death, including cancers and autoimmune diseases. This review gives an overview focusing on the sensitization of the cell-death induction via either sFasL- or FasR-mAb-primed signal transduction system with exogenous agents. In the beginning section, the structural and functional characteristics of cell-death induction using these soluble agonistic proteins were briefly introduced. In the following sections, the studies on the sensitization of Fas signaling system with the exogenous agents, classified into two groups, were investigated, based on an extensive survey of the relevant literatures. First, the sensitization with non-cytokine agents was described, where the effects of representative low molecular-weight clinical anticancer drugs were highlighted. Then, the potency of exogenous cytokine agents was depicted, while centering on the sensitization with interferon-γ. The survey revealed that the agents examined here were effective for the sensitization against various malignant tumors-derived and other types of cells by upregulating pro-apoptotic molecular machinery and/or downregulating anti-apoptotic factors. However, in the demanding instances, this strategy still remained dysfunctional in completing the target cell-killing process due to resistance mechanisms, such as overexpression of intracellular inhibitory proteins. Finally, it is proposed that the sensitization of cell-death induction with exogenous agents, combined with empowerment regarding the targeting specificity by protein engineering techniques, is a promising approach to potentiate the soluble agonists for translating them into clinical protein pharmaceuticals.