scholarly journals Generation and Characterization of Typhoid Toxin-Neutralizing Human Monoclonal Antibodies

2020 ◽  
Vol 88 (10) ◽  
Author(s):  
Xuyao Jiao ◽  
Sarah Smith ◽  
Gabrielle Stack ◽  
Qi Liang ◽  
Allan Bradley ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Typhoid Fever ◽  
Severe Disease ◽  
Genetically Engineered ◽  
Human Monoclonal Antibodies ◽  
Genetically Engineered Mice ◽  
Typhoid Toxin

ABSTRACT Typhoid toxin is a virulence factor of Salmonella enterica serovar Typhi, the causative agent of typhoid fever, and is thought to be responsible for the symptoms of severe disease. This toxin has a unique A2B5 architecture with two active subunits, the ADP ribosyl transferase PltA and the DNase CdtB, linked to a pentameric B subunit, which is alternatively made of PltB or PltC. Here, we describe the generation and characterization of typhoid toxin-neutralizing human monoclonal antibodies by immunizing genetically engineered mice that have a full set of human immunoglobulin variable region genes. We identified several monoclonal antibodies with strong in vitro and in vivo toxin-neutralizing activity and different mechanisms of toxin neutralization. These antibodies could serve as the basis for the development of novel therapeutic strategies against typhoid fever.

Characterization of the In Vitro and In Vivo Activity of Monoclonal Antibodies to Human IL-18

Hybridoma ◽  
2000 ◽  
Vol 19 (5) ◽  
pp. 363-367 ◽  
Author(s):  
Steve Holmes ◽  
Julie A. Abrahamson ◽  
Niam Al-Mahdi ◽  
Sherin S. Abdel-Meguid ◽  
Yen Sen Ho
Keyword(s):  
Monoclonal Antibodies

scholarly journals Discrete Role for Cytosolic Phospholipase A2α in Platelets

2002 ◽  
Vol 196 (3) ◽  
pp. 349-357 ◽  
Author(s):  
Dennis A. Wong ◽  
Yoshihiro Kita ◽  
Naonori Uozumi ◽  
Takao Shimizu
Keyword(s):  
Therapeutic Potential ◽  
Genetically Engineered ◽  
Collagen Receptors ◽  
Cytosolic Phospholipase ◽  
Genetically Engineered Mice ◽  
Cytosolic Pla2 ◽  
In Vivo Effects ◽  
Deficient Mice

Among several different types of phospholipase A2 (PLA2), cytosolic PLA2 (cPLA2)α and group IIA (IIA) secretory PLA2 (sPLA2) have been studied intensively. To determine the discrete roles of cPLA2α in platelets, we generated two sets of genetically engineered mice (cPLA2α−/−/sPLA2-IIA−/− and cPLA2α−/−/sPLA2-IIA+/+) and compared their platelet function with their respective wild-type C57BL/6J mice (cPLA2α+/+/sPLA2-IIA−/−) and C3H/HeN (cPLA2α+/+/sPLA2-IIA+/+). We found that cPLA2α is needed for the production of the vast majority of thromboxane (TX)A2 with collagen stimulation of platelets. In cPLA2α-deficient mice, however, platelet aggregation in vitro is only fractionally decreased because small amounts of TX produced by redundant phospholipase enzymes sufficiently preserve aggregation. In comparison, adenosine triphosphate activation of platelets appears wholly independent of cPLA2α and sPLA2-IIA for aggregation or the production of TX, indicating that these phospholipases are specifically linked to collagen receptors. However, the lack of high levels of TX limiting vasoconstriction explains the in vivo effects seen: increased bleeding times and protection from thromboembolism. Thus, cPLA2α plays a discrete role in the collagen-stimulated production of TX and its inhibition has a therapeutic potential against thromboembolism, with potentially limited bleeding expected.

scholarly journals Monoclonal antibodies targeting the influenza virus N6 neuraminidase

2021 ◽  
Author(s):  
Shirin Strohmeier ◽  
Fatima Amanat ◽  
Juan Manuel Carreño ◽  
Florian Krammer
Keyword(s):  
Monoclonal Antibodies ◽  
Influenza A ◽  
Neutralization Assay ◽  
Escape Mutant ◽  
Influenza A Viruses ◽  
Highly Pathogenic ◽  
Lateral Ridge

AbstractInfluenza A viruses are a diverse species that include 16 hemagglutinin (HA) subtypes and 9 neuraminidase (NA) subtypes. While the antigenicity of many HA subtypes is reasonably well studied, less is known about NA antigenicity, especially when it comes to non-human subtypes that only circulate in animal reservoirs. The N6 NA subtypes are mostly found in viruses infecting birds. However, they have also been identified in viruses that infect mammals, such as swine and seals. More recently, highly pathogenic H5N6 subtype viruses have caused rare infections and mortality in humans. Here, we generated murine mAbs to the N6 NA, characterized their breadth and antiviral properties in vitro and in vivo and mapped their epitopes by generating escape mutant viruses. We found that the antibodies had broad reactivity across the American and Eurasian N6 lineages, but relatively little binding and inhibition of the H5N6 NA. Several of the antibodies exhibited strong NA inhibition activity and some also showed activity in the antibody dependent cellular cytotoxicity reporter assay and neutralization assay. In addition, we generated escape mutant viruses for six monoclonal antibodies and found mutations on the lateral ridge of the NA. Lastly, we observed variable protection in H4N6 and H5N6 mouse challenge models when the antibodies were given prophylactically.ImportanceThe N6 NA has recently gained prominence due to the emergence of highly pathogenic H5N6 viruses. Currently, there is limited characterization of the antigenicity of avian N6 neuraminidase. Our data is an important first step towards a better understanding of the N6 NA antigenicity.

scholarly journals Human Monoclonal Antibodies Directed against Toxins A and B Prevent Clostridium difficile-Induced Mortality in Hamsters

2006 ◽  
Vol 74 (11) ◽  
pp. 6339-6347 ◽  
Author(s):  
Gregory J. Babcock ◽  
Teresa J. Broering ◽  
Hector J. Hernandez ◽  
Robert B. Mandell ◽  
Katherine Donahue ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Clostridium Difficile ◽  
Human Monoclonal Antibodies ◽  
Toxin A ◽  
Hamster Model ◽  
Cytotoxic Effects ◽  
Toxin B ◽  
Reduction Of Mortality

ABSTRACT Clostridium difficile is the leading cause of nosocomial antibiotic-associated diarrhea, and recent outbreaks of strains with increased virulence underscore the importance of identifying novel approaches to treat and prevent relapse of Clostridium difficile-associated diarrhea (CDAD). CDAD pathology is induced by two exotoxins, toxin A and toxin B, which have been shown to be cytotoxic and, in the case of toxin A, enterotoxic. In this report we describe fully human monoclonal antibodies (HuMAbs) that neutralize these toxins and prevent disease in hamsters. Transgenic mice carrying human immunoglobulin genes were used to isolate HuMAbs that neutralize the cytotoxic effects of either toxin A or toxin B in cell-based in vitro neutralization assays. Three anti-toxin A HuMAbs (3H2, CDA1, and 1B11) could all inhibit the enterotoxicity of toxin A in mouse intestinal loops and the in vivo toxicity in a systemic mouse model. Four anti-toxin B HuMAbs (MDX-1388, 103-174, 1G10, and 2A11) could neutralize cytotoxicity in vitro, although systemic toxicity in the mouse could not be neutralized. Anti-toxin A HuMAb CDA1 and anti-toxin B HuMAb MDX-1388 were tested in the well-established hamster model of C. difficile disease. CDA1 alone resulted in a statistically significant reduction of mortality in hamsters; however, the combination treatment offered enhanced protection. Compared to controls, combination therapy reduced mortality from 100% to 45% (P < 0.0001) in the primary disease hamster model and from 78% to 32% (P < 0.0001) in the less stringent relapse model.

999 In Vitro and In Vivo Characterization of Neutralizing Monoclonal Antibodies Against Clostridium difficile Toxins a and B

2013 ◽  
Vol 144 (5) ◽  
pp. S-185
Author(s):  
Dominiek Staelens ◽  
Marlies Van de Wouwer ◽  
Els Brouwers ◽  
Silvia Caluwaerts ◽  
Borden Lacy ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Clostridium Difficile ◽  
In Vivo Characterization

Immunolocalization and characterization of the low molecular weight antigen (4–5 kDa) ofToxoplasma gondiithat elicits an early IgM response upon primary infection

Parasitology ◽  
1994 ◽  
Vol 108 (2) ◽  
pp. 139-145 ◽  
Author(s):  
S. Tomavo ◽  
G. Couvreur ◽  
M. A. Leriche ◽  
A. Sadak ◽  
A. Achbarou ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Molecular Weight ◽  
Monoclonal Antibodies ◽  
Primary Infection ◽  
Low Molecular Weight ◽  
Immuno Electron Microscopy ◽  
Surface Localization

SUMMARYA striking feature of toxoplasmic seroconversion is the prominent and early IgM response to a low molecular weight antigen of 4–5 kDa. Two different monoclonal antibodies directed against the 4–5 kDa antigen have been generated and used to characterize this molecule. Using these monoclonal antibodies, we could demonstrate the surface localization of the lowMrantigen by immunofluorescence and immuno-electron microscopy assays. By immunoblotting, we observed that one of the monoclonal antibodies was unable to recognize the 4–5 kDa antigen in tachyzoites propagated in cell culture, indicating an epitope variability betweenToxoplasma gondiitachyzoites grownin vivoandin vitro. We discuss the implications of this latter finding in the design of diagnostic reagents.

scholarly journals Novel epitopes of human monoclonal antibodies targeting the influenza virus N1 neuraminidase

2021 ◽  
Author(s):  
Ericka Kirkpatrick Roubidoux ◽  
Meagan McMahon ◽  
Juan Manuel Carreno ◽  
Christina Capuano ◽  
Kaijun Jiang ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Monoclonal Antibodies ◽  
Serial Passage ◽  
Viral Fitness ◽  
Escape Mutant ◽  
Human Antibody ◽  
Human Monoclonal Antibodies ◽  
Influenza Virus Neuraminidase

Influenza virus neuraminidase (NA) targeting antibodies are an independent correlate of protection against infection. Antibodies against the NA act by blocking enzymatic activity, preventing virus release and transmission. As we advance the development of improved influenza virus vaccines that incorporate standard amounts of NA antigen, it is important to identify the antigenic targets of human monoclonal antibodies (mAbs). Additionally, it is important to understand how escape from mAbs changes viral fitness. Here, we describe escape mutants generated by serial passage of A/Netherlands/602/2009 (H1N1) in the presence of human anti-N1 mAbs. We observed escape mutations on the N1 protein around the enzymatic site (S364N, N369T and R430Q) and also detected escape mutations located on the sides and bottom of the NA (N88D, N270D and Q313K/R). We found that a majority of escape mutant viruses had increased fitness in vitro but not in vivo. This work increases our understanding of how human antibody responses target the N1 protein.

scholarly journals Blocking gephyrin phosphorylation or microglia BDNF signaling prevents synapse loss and reduces infarct volume after ischemia

2020 ◽  
Author(s):  
Teresa Cramer ◽  
Raminder Gill ◽  
Zahra S Thirouin ◽  
Markus Vaas ◽  
Suchita Sampath ◽  
...  
Keyword(s):  
Hippocampal Formation ◽  
Infarct Volume ◽  
Point Mutations ◽  
Null Point ◽  
Genetically Engineered ◽  
Infarct Area ◽  
Synapse Plasticity ◽  
Genetically Engineered Mice

AbstractMicroglia interact with neurons to facilitate synapse plasticity; however, signal transducers between microglia and neuron remain unknown. Here, using in vitro organotypic hippocampal slice cultures and transient MCAO in genetically-engineered mice in vivo, we report that at 24 h post-ischemia microglia release BDNF to downregulate glutamatergic and GABAergic synapses within the peri-infarct area. Analysis of the CA1 hippocampal formation in vitro shows that proBDNF and mBDNF downregulate glutamatergic dendritic spines and gephyrin scaffold stability through p75NTR and TrkB receptors respectively. Post-MCAO, we report that in the peri- infarct area and in the corresponding contralateral hemisphere similar neuroplasticity occur through microglia activation and gephyrin phosphorylation at Ser268, Ser270 in vivo. Targeted deletion of the Bdnf gene in microglia or GphnS268A/S270A (phospho-null) point-mutations protect against ischemic brain damage, neuroinflamation and synapse downregulation normally seen post-MCAO. Collectively, we report that gephyrin phosphorylation and microglia derived BDNF faciliate synapse plasticity after transient ischemia.

scholarly journals Inhibition of Type 1 Cytokine–mediated Inflammation by a Soluble CD30 Homologue Encoded by Ectromelia (Mousepox) Virus

2002 ◽  
Vol 196 (6) ◽  
pp. 829-839 ◽  
Author(s):  
Margarida Saraiva ◽  
Philip Smith ◽  
Padraic G. Fallon ◽  
Antonio Alcami
Keyword(s):  
Viral Infections ◽  
Inflammatory Responses ◽  
Severe Disease ◽  
Interferon Γ ◽  
Reverse Signaling ◽  
Soluble Cd30

CD30 is up-regulated in several human diseases and viral infections but its role in immune regulation is poorly understood. Here, we report the expression of a functional soluble CD30 homologue, viral CD30 (vCD30), encoded by ectromelia (mousepox) virus, a poxvirus that causes a severe disease related to human smallpox. We show that vCD30 is a 12-kD secreted protein that not only binds CD30L with high affinity and prevents its interaction with CD30, but it also induces reverse signaling in cells expressing CD30L. vCD30 blocked the generation of interferon γ–producing cells in vitro and was a potent inhibitor of T helper cell (Th)1- but not Th2-mediated inflammation in vivo. The finding of a CD30 homologue encoded by ectromelia virus suggests a role for CD30 in antiviral defense. Characterization of the immunological properties of vCD30 has uncovered a role of CD30–CD30L interactions in the generation of inflammatory responses.

scholarly journals Preclinical Evaluation of Two Neutralizing Human Monoclonal Antibodies against Hepatitis C Virus (HCV): a Potential Treatment To Prevent HCV Reinfection in Liver Transplant Patients

2006 ◽  
Vol 80 (6) ◽  
pp. 2654-2664 ◽  
Author(s):  
Rachel Eren ◽  
Dorit Landstein ◽  
Dov Terkieltaub ◽  
Ofer Nussbaum ◽  
Arie Zauberman ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Liver Transplant ◽  
Immune Complexes ◽  
Human Liver ◽  
Human Monoclonal Antibodies ◽  
Transplant Patients

ABSTRACT Passive immunotherapy is potentially effective in preventing reinfection of liver grafts in hepatitis C virus (HCV)-associated liver transplant patients. A combination of monoclonal antibodies directed against different epitopes may be advantageous against a highly mutating virus such as HCV. Two human monoclonal antibodies (HumAbs) against the E2 envelope protein of HCV were developed and tested for the ability to neutralize the virus and prevent human liver infection. These antibodies, designated HCV-AB 68 and HCV-AB 65, recognize different conformational epitopes on E2. They were characterized in vitro biochemically and functionally. Both HumAbs are immunoglobulin G1 and have affinity constants to recombinant E2 constructs in the range of 10−10 M. They are able to immunoprecipitate HCV particles from infected patients' sera from diverse genotypes and to stain HCV-infected human liver tissue. Both antibodies can fix complement and form immune complexes, but they do not activate complement-dependent or antibody-dependent cytotoxicity. Upon complement fixation, the monoclonal antibodies induce phagocytosis of the immune complexes by neutrophils, suggesting that the mechanism of viral clearance includes endocytosis. In vivo, in the HCV-Trimera model, both HumAbs were capable of inhibiting HCV infection of human liver fragments and of reducing the mean viral load in HCV-positive animals. The demonstrated neutralizing activities of HCV-AB 68 and HCV-AB 65 suggest that they have the potential to prevent reinfection in liver transplant patients and to serve as prophylactic treatment in postexposure events.

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