scholarly journals Human Coagulation Factor X-Adenovirus Type 5 Complexes Poorly Stimulate an Innate Immune Response in Human Mononuclear Phagocytes

2014 ◽  
Vol 89 (5) ◽  
pp. 2884-2891 ◽  
Author(s):  
Karsten Eichholz ◽  
Franck J. D. Mennechet ◽  
Eric J. Kremer
Keyword(s):  
Coagulation Factor ◽  
Human Adenovirus ◽  
Adenovirus Type ◽  
Natural Host ◽  
Innate Immune ◽  
Mononuclear Phagocytes ◽  
Factor X ◽  
Dynamic Interactions

ABSTRACTOne of the first lines of host defense against many viruses in vertebrates is the innate immune system, which detects pathogen-associated molecular patterns (PAMPs) using pathogen recognition receptors (PRR). The dynamic interactions between pathogens and hosts create, in some cases, species-specific relationships. Recently, it was shown that murine factor X (mFX)-armored human adenovirus (HAd) stimulated a mFX-Toll-like receptor 4 (TLR4)-associated response in mouse macrophagesin vitroandin vivo. Given the importance of studies using animals to better understand host-pathogen interactions, we asked if human FX (hFX)-armored HAd type 5 (HAd5) was capable of activating innate immune sensors in primary human mononuclear phagocytes. To this end, we assayed human mononuclear phagocytes for their ability to be stimulated by hFX-armored HAd5 via a TLR/NF-κB pathway, in particular, a TLR4 pathway. In our hands, we found no significant interaction, activation, or maturation of human mononuclear phagocytes caused by the presence of hFX-armored HAd5.IMPORTANCEAnimals, and mice in particular, are often used as informative and powerful surrogates for how pathogens interact with natural host systems. When possible, extended and targeted studies in the natural host can then be performed. Our data will help us understand the differences in preclinical testing in mice and clinical use in humans in order to improve treatment for HAd diseases and Ad vector effectiveness.

scholarly journals Biodistribution and retargeting of FX-binding ablated adenovirus serotype 5 vectors

Blood ◽  
2010 ◽  
Vol 116 (15) ◽  
pp. 2656-2664 ◽  
Author(s):  
Raul Alba ◽  
Angela C. Bradshaw ◽  
Lynda Coughlan ◽  
Laura Denby ◽  
Robert A. McDonald ◽  
...  
Keyword(s):  
Coagulation Factor ◽  
Adenovirus Type ◽  
Factor X ◽  
High Affinity ◽  
Serotype 5 ◽  
Adenoviral Transduction ◽  
High Doses ◽  
Immunohistochemical Studies

AbstractA major limitation for adenoviral transduction in vivo is the profound liver tropism of adenovirus type 5 (Ad5). Recently, we demonstrated that coagulation factor X (FX) binds to Ad5-hexon protein at high affinity to mediate hepatocyte transduction after intravascular delivery. We developed novel genetically FX-binding ablated Ad5 vectors with lower liver transduction. Here, we demonstrate that FX-binding ablated Ad5 predominantly localize to the liver and spleen 1 hour after injection; however, they had highly reduced liver transduction in both control and macrophage-depleted mice compared with Ad5. At high doses in macrophage-depleted mice, FX-binding ablated vectors transduced the spleen more efficiently than Ad5. Immunohistochemical studies demonstrated transgene colocalization with CD11c+, ER-TR7+, and MAdCAM-1+ cells in the splenic marginal zone. Systemic inflammatory profiles were broadly similar between FX-binding ablated Ad5 and Ad5 at low and intermediate doses, although higher levels of several inflammatory proteins were observed at the highest dose of FX-binding ablated Ad5. Subsequently, we generated a FX-binding ablated virus containing a high affinity Ad35 fiber that mediated a significant improvement in lung/liver ratio in macrophage-depleted CD46+ mice compared with controls. Therefore, this study documents the biodistribution and reports the retargeting capacity of FX binding-ablated Ad5 vectors in vitro and in vivo.

scholarly journals Manipulating Adenovirus Hexon Hypervariable Loops Dictates Immune Neutralisation and Coagulation Factor X-dependent Cell Interaction In Vitro and In Vivo

2015 ◽  
Vol 11 (2) ◽  
pp. e1004673 ◽  
Author(s):  
Jiangtao Ma ◽  
Margaret R. Duffy ◽  
Lin Deng ◽  
Rachel S. Dakin ◽  
Taco Uil ◽  
...  
Keyword(s):  
Coagulation Factor ◽  
Cell Interaction ◽  
Factor X ◽  
Dependent Cell

scholarly journals In Vitro and In Vivo Evaluation of Human Adenovirus Type 49 as a Vector for Therapeutic Applications

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1483
Author(s):  
Emily A. Bates ◽  
John R. Counsell ◽  
Sophie Alizert ◽  
Alexander T. Baker ◽  
Natalie Suff ◽  
...  
Keyword(s):  
Viral Vector ◽  
Ex Vivo ◽  
Human Adenovirus ◽  
Adenovirus Type ◽  
Cell Types ◽  
In Vivo Evaluation ◽  
In Vivo Biodistribution ◽  
Adenovirus Type 5

The human adenovirus phylogenetic tree is split across seven species (A–G). Species D adenoviruses offer potential advantages for gene therapy applications, with low rates of pre-existing immunity detected across screened populations. However, many aspects of the basic virology of species D—such as their cellular tropism, receptor usage, and in vivo biodistribution profile—remain unknown. Here, we have characterized human adenovirus type 49 (HAdV-D49)—a relatively understudied species D member. We report that HAdV-D49 does not appear to use a single pathway to gain cell entry, but appears able to interact with various surface molecules for entry. As such, HAdV-D49 can transduce a broad range of cell types in vitro, with variable engagement of blood coagulation FX. Interestingly, when comparing in vivo biodistribution to adenovirus type 5, HAdV-D49 vectors show reduced liver targeting, whilst maintaining transduction of lung and spleen. Overall, this presents HAdV-D49 as a robust viral vector platform for ex vivo manipulation of human cells, and for in vivo applications where the therapeutic goal is to target the lung or gain access to immune cells in the spleen, whilst avoiding liver interactions, such as intravascular vaccine applications.

Natural IgM antibodies inhibit microvesicle-driven coagulation and thrombosis

Blood ◽  
2020 ◽  
Author(s):  
Georg Obermayer ◽  
Taras Afonyushkin ◽  
Laura Goederle ◽  
Florian Puhm ◽  
Waltraud C. Schrottmaier ◽  
...  
Keyword(s):  
Therapeutic Potential ◽  
Coagulation Factor ◽  
Mechanistic Explanation ◽  
Factor X ◽  
Thrombotic Risk ◽  
Igm Antibodies ◽  
Plasmatic Coagulation ◽  
Natural Igm

Thrombosis and the complications associated with it are a major cause of morbidity and mortality worldwide. Microvesicles (MVs), a class of extracellular vesicles, are increasingly recognized as mediators of coagulation and biomarkers of thrombotic risk. Thus, identifying factors targeting MV-driven coagulation may help in the development of novel antithrombotic treatments. We have previously identified a subset of circulating MVs that is characterized by the presence of oxidation-specific epitopes and bound by natural IgM antibodies targeting these structures. Here, we investigated whether natural IgM antibodies, which are known to have important anti-inflammatory house-keeping functions, inhibit the procoagulatory properties of MVs. We found that the extent of plasma coagulation is inversely associated with the levels of both free and MV-bound endogenous IgM. Moreover, the oxidation epitope-specific natural IgM antibody LR04, which recognizes malondialdehyde adducts, reduced MV-dependent plasmatic coagulation and whole blood clotting without affecting thrombocyte aggregation. Intravenous injection of LR04 protected mice from MV-induced pulmonary thrombosis. Of note, LR04 competed the binding of coagulation factor X/Xa to MVs, providing a mechanistic explanation for its anticoagulatory effect. Thus, our data identify natural IgM antibodies as hitherto unknown modulators of MV-induced coagulation in vitro and in vivo and their prognostic and therapeutic potential in the management of thrombosis.

scholarly journals Mycobacterium avium Biofilm Attenuates Mononuclear Phagocyte Function by Triggering Hyperstimulation and Apoptosis during Early Infection

2013 ◽  
Vol 82 (1) ◽  
pp. 405-412 ◽  
Author(s):  
Sasha J. Rose ◽  
Luiz E. Bermudez
Keyword(s):  
Immune System ◽  
Mycobacterium Avium ◽  
Innate Immune System ◽  
Innate Immune ◽  
Mononuclear Phagocytes ◽  
Bacterial Killing ◽  
Content Type ◽  
Tnf Α

ABSTRACTMycobacterium aviumsubsp.hominissuisis an opportunistic human pathogen that has been shown to form biofilmin vitroandin vivo. Biofilm formationin vivoappears to be associated with infections in the respiratory tract of the host. The reasoning behind howM. aviumsubsp.hominissuisbiofilm is allowed to establish and persist without being cleared by the innate immune system is currently unknown. To identify the mechanism responsible for this, we developed anin vitromodel using THP-1 human mononuclear phagocytes cocultured with establishedM. aviumsubsp.hominissuisbiofilm and surveyed various aspects of the interaction, including phagocyte stimulation and response, bacterial killing, and apoptosis.M. aviumsubsp.hominissuisbiofilm triggered robust tumor necrosis factor alpha (TNF-α) release from THP-1 cells as well as superoxide and nitric oxide production. Surprisingly, the hyperstimulated phagocytes did not effectively eliminate the cells of the biofilm, even when prestimulated with gamma interferon (IFN-γ) or TNF-α or cocultured with natural killer cells (which have been shown to induce anti-M. aviumsubsp.hominissuisactivity when added to THP-1 cells infected with planktonicM. aviumsubsp.hominissuis). Time-lapse microscopy and the TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) assay determined that contact with theM. aviumsubsp.hominissuisbiofilm led to early, widespread onset of apoptosis, which is not seen until much later in planktonicM. aviumsubsp.hominissuisinfection. Blocking TNF-α or TNF-R1 during interaction with the biofilm significantly reduced THP-1 apoptosis but did not lead to elimination ofM. aviumsubsp.hominissuis. Our data collectively indicate thatM. aviumsubsp.hominissuisbiofilm induces TNF-α-driven hyperstimulation and apoptosis of surveilling phagocytes, which prevents clearance of the biofilm by cells of the innate immune system and allows the biofilm-associated infection to persist.

scholarly journals In Vitro and In Vivo Evaluation of Human Adenovirus Type 49 as a Vector for Therapeutic Applications

2021 ◽  
Author(s):  
Emily A. Bates ◽  
John R. Counsell ◽  
Sophie Alizert ◽  
Alexander T. Baker ◽  
Natalie Suff ◽  
...  
Keyword(s):  
Viral Vector ◽  
Ex Vivo ◽  
Human Adenovirus ◽  
Adenovirus Type ◽  
Cell Types ◽  
In Vivo Evaluation ◽  
In Vivo Biodistribution ◽  
Adenovirus Type 5

The human adenovirus phylogenetic tree is split across seven species (A-G). Species D adenoviruses offer potential advantages for gene therapy applications, with low rates of preexisting immunity detected across screened populations. However, many aspects of the basic virology of species D, such as their cellular tropism, receptor usage and in vivo biodistribution profile, remain unknown. Here, we have characterized human adenovirus type 49 (HAdV-D49), a relatively understudied species D member. We report that HAdV-D49 does not appear to use a single pathway to gain cell entry but appears able to interact with various surface molecules for entry. As such, HAdV-D49 can transduce a broad range of cell types in vitro, with variable engagement of blood coagulation FX. Interestingly, when comparing in vivo biodistribution to adenovirus type 5, HAdV-D49 vectors show reduced liver targeting whilst maintaining transduction of lung and spleen. Overall, this presents HAdV-D49 as a robust viral vector platform for ex vivo manipulation of human cells and for in vivo applications where the therapeutic goal is to target the lung or gain access to immune cells in the spleen whilst avoiding liver interactions, such as intravascular vaccine applications.

scholarly journals Development and Assessment of Human Adenovirus Type 11 as a Gene Transfer Vector

2005 ◽  
Vol 79 (8) ◽  
pp. 5090-5104 ◽  
Author(s):  
Daniel Stone ◽  
Shaoheng Ni ◽  
Zong-Yi Li ◽  
Anuj Gaggar ◽  
Nelson DiPaolo ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Neutralizing Antibodies ◽  
Human Adenovirus ◽  
Adenovirus Type ◽  
Cell Types ◽  
Widespread Application ◽  
Human Cd34 ◽  
Immature Dendritic Cells

ABSTRACT Adenovirus vectors based on human serotype 5 (Ad5) have successfully been used as gene transfer vectors in many gene therapy-based approaches to treat disease. Despite their widespread application, many potential therapeutic applications are limited by the widespread prevalence of vector-neutralizing antibodies within the human population and the inability of Ad5-based vectors to transduce important therapeutic target cell types. In an attempt to circumvent these problems, we have developed Ad vectors based on human Ad serotype 11 (Ad11), since the prevalence of neutralizing antibodies to Ad11 in humans is low. E1-deleted Ad11 vector genomes were generated by homologous recombination in 293 cells expressing the Ad11-E1B55K protein or by recombination in Escherichia coli. E1-deleted Ad11 genomes did not display transforming activity in rodent cells. Transduction of primary human CD34+ hematopoietic progenitor cells and immature dendritic cells was more efficient with Ad11 vectors than with Ad5 vectors. Thirty minutes after intravenous injection into mice that express one of the Ad11 receptors (CD46), we found, in a pattern and at a level comparable to what is found in humans, Ad11 vector genomes in all analyzed organs, with the highest amounts in liver, lung, kidney, and spleen. Neither Ad11 genomes nor Ad11 vector-mediated transgene expression were, however, detected at 72 h postinfusion. A large number of Ad11 particles were also found to be associated with circulating blood cells. We also discovered differences in in vitro transduction efficiencies and in vivo biodistributions between Ad11 vectors and chimeric Ad5 vectors possessing Ad11 fibers, indicating that Ad11 capsid proteins other than fibers influence viral infectivity and tropism. Overall, our study provides a basis for the application of Ad11 vectors for in vitro and in vivo gene transfer and for gaining an understanding of the factors that determine Ad tropism.

scholarly journals A mutated factor X activatable by thrombin corrects bleedings in vivo in a rabbit model of antibody-induced hemophilia A

Haematologica ◽  
2019 ◽  
Vol 105 (9) ◽  
pp. 2335-2340
Author(s):  
Toufik Abache ◽  
Alexandre Fontayne ◽  
Dominique Grenier ◽  
Emilie Jacque ◽  
Alain Longue ◽  
...  
Keyword(s):  
Factor Viii ◽  
Hemophilia A ◽  
Factor Viia ◽  
Rabbit Model ◽  
Coagulation Factor ◽  
Factor Ix ◽  
Factor X ◽  
Factor Viiia

Rendering coagulation factor X sensitive to thrombin was proposed as a strategy that can bypass the need for factor VIII. In this paper, this non-replacement strategy was evaluated in vitro and in vivo in its ability to correct factor VIII but also factor IX, X and XI deficiencies. A novel modified factor X, named Actiten, was generated and produced in the HEK293F cell line. The molecule possesses the required post-translational modifications, partially keeps its ability to be activated by RVV-X, factor VIIa/tissue factor, factor VIIIa/factor IXa and acquires the ability to be activated by thrombin. The potency of the molecule was evaluated in respective deficient plasmas or hemophilia A plasmas, for some with inhibitors. Actiten corrects dose dependently all the assayed deficient plasmas. It is able to normalize the thrombin generation at 20 μg/mL showing however an increased lagtime. It was then assayed in a rabbit antibody-induced model of hemophilia A where, in contrast to recombinant factor X wild-type, it normalized the bleeding time and the loss of hemoglobin. No sign of thrombogenicity was observed and the generation of activated factor X was controlled by the anticoagulation pathway in all performed coagulation assays. This data indicates that Actiten may be considered as a possible non replacement factor to treat hemophilia's with the advantage of being a zymogen correcting bleedings only when needed.

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