Cryo-EM Studies of Virus-Antibody Immune Complexes

AbstractAntibodies play critical roles in neutralizing viral infections and are increasingly used as therapeutic drugs and diagnostic tools. Structural studies on virus-antibody immune complexes are important for better understanding the molecular mechanisms of antibody-mediated neutralization and also provide valuable information for structure-based vaccine design. Cryo-electron microscopy (cryo-EM) has recently matured as a powerful structural technique for studying bio-macromolecular complexes. When combined with X-ray crystallography, cryo-EM provides a routine approach for structurally characterizing the immune complexes formed between icosahedral viruses and their antibodies. In this review, recent advances in the structural understanding of virus-antibody interactions are outlined for whole virions with icosahedral T = pseudo 3 (picornaviruses) and T = 3 (flaviviruses) architectures, focusing on the dynamic nature of viral shells in different functional states. Glycoprotein complexes from pleomorphic enveloped viruses are also discussed as immune complex antigens. Improving our understanding of viral epitope structures using virus-based platforms would provide a fundamental road map for future vaccine development.

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Metal Oxide Nanoparticles in Therapeutic Regulation of Macrophage Functions

Nanomaterials ◽

10.3390/nano9111631 ◽

2019 ◽

Vol 9 (11) ◽

pp. 1631 ◽

Cited By ~ 11

Author(s):

Marina S. Dukhinova ◽

Artur. Y. Prilepskii ◽

Alexander A. Shtil ◽

Vladimir V. Vinogradov

Keyword(s):

Metal Oxide ◽

Molecular Mechanisms ◽

Viral Infections ◽

Metal Oxide Nanoparticles ◽

Cell Polarization ◽

Innate Immune ◽

Diagnostic Tools ◽

Oxide Nanoparticles ◽

Inflammatory Microenvironment ◽

Anti Inflammatory

Macrophages are components of the innate immune system that control a plethora of biological processes. Macrophages can be activated towards pro-inflammatory (M1) or anti-inflammatory (M2) phenotypes depending on the cue; however, polarization may be altered in bacterial and viral infections, cancer, or autoimmune diseases. Metal (zinc, iron, titanium, copper, etc.) oxide nanoparticles are widely used in therapeutic applications as drugs, nanocarriers, and diagnostic tools. Macrophages can recognize and engulf nanoparticles, while the influence of macrophage-nanoparticle interaction on cell polarization remains unclear. In this review, we summarize the molecular mechanisms that drive macrophage activation phenotypes and functions upon interaction with nanoparticles in an inflammatory microenvironment. The manifold effects of metal oxide nanoparticles on macrophages depend on the type of metal and the route of synthesis. While largely considered as drug transporters, metal oxide nanoparticles nevertheless have an immunotherapeutic potential, as they can evoke pro- or anti-inflammatory effects on macrophages and become essential for macrophage profiling in cancer, wound healing, infections, and autoimmunity.

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Structurally related but genetically unrelated antibody lineages converge on an immunodominant HIV-1 Env neutralizing determinant following trimer immunization

PLoS Pathogens ◽

10.1371/journal.ppat.1009543 ◽

2021 ◽

Vol 17 (9) ◽

pp. e1009543

Author(s):

Safia S. Aljedani ◽

Tyler J. Liban ◽

Karen Tran ◽

Ganesh Phad ◽

Suruchi Singh ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Molecular Mechanisms ◽

Vaccine Development ◽

Hypervariable Region ◽

Variable Region ◽

Tier 2 ◽

X Ray Crystallography ◽

Neutralizing Capacity ◽

Clade C ◽

Hiv 1

Understanding the molecular mechanisms by which antibodies target and neutralize the HIV-1 envelope glycoprotein (Env) is critical in guiding immunogen design and vaccine development aimed at eliciting cross-reactive neutralizing antibodies (NAbs). Here, we analyzed monoclonal antibodies (mAbs) isolated from non-human primates (NHPs) immunized with variants of a native flexibly linked (NFL) HIV-1 Env stabilized trimer derived from the tier 2 clade C 16055 strain. The antibodies displayed neutralizing activity against the autologous virus with potencies ranging from 0.005 to 3.68 μg/ml (IC50). Structural characterization using negative-stain EM and X-ray crystallography identified the variable region 2 (V2) of the 16055 NFL trimer to be the common epitope for these antibodies. The crystal structures revealed that the V2 segment adopts a β-hairpin motif identical to that observed in the 16055 NFL crystal structure. These results depict how vaccine-induced antibodies derived from different clonal lineages penetrate through the glycan shield to recognize a hypervariable region within V2 (residues 184–186) that is unique to the 16055 strain. They also provide potential explanations for the potent autologous neutralization of these antibodies, confirming the immunodominance of this site and revealing that multiple angles of approach are permissible for affinity/avidity that results in potent neutralizing capacity. The structural analysis reveals that the most negatively charged paratope correlated with the potency of the mAbs. The atomic level information is of interest to both define the means of autologous neutralization elicited by different tier 2-based immunogens and facilitate trimer redesign to better target more conserved regions of V2 to potentially elicit cross-neutralizing HIV-1 antibodies.

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Structurally related but genetically unrelated antibody lineages converge on an immunodominant HIV-1 Env neutralizing determinant following trimer immunization

10.1101/2021.04.09.439148 ◽

2021 ◽

Author(s):

Safia Aljedani ◽

Tyler J Liban ◽

Karen Tran ◽

Ganesh Phad ◽

Suruchi Singh ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Molecular Mechanisms ◽

Vaccine Development ◽

Hypervariable Region ◽

Variable Region ◽

Tier 2 ◽

X Ray Crystallography ◽

Neutralizing Capacity ◽

Clade C ◽

Hiv 1

Understanding the molecular mechanisms by which antibodies target and neutralize the HIV-1 envelope glycoprotein (Env) is critical in guiding immunogen design and vaccine development aimed at eliciting cross-reactive neutralizing antibodies (NAbs). Here, we analyzed monoclonal antibodies (mAbs) isolated from non-human primates (NHPs) immunized with variants of a native flexibly linked (NFL) HIV-1 Env stabilized trimer derived from the tier 2 clade C 16055 strain. The antibodies displayed neutralizing activity against the autologous virus with potencies ranging from 0.005 to 3.68 ug/ml (IC 50 ). Structural characterization using negative-stain EM and X-ray crystallography identified the variable region 2 (V2) of the 16055 NFL trimer to be the common epitope for these antibodies. The crystal structures revealed that the V2 segment adopts a β-hairpin motif identical to that observed in the 16055 NFL crystal structure. These results depict how vaccine-induced antibodies derived from different clonal lineages penetrate through the glycan shield to recognize a hypervariable region within V2 (residues 184-186) that is unique to the 16055 strain. They also provide an explanation for the potent autologous neutralization of these antibodies, confirming the immunodominance of this site and revealing that multiple angles of approach are permissible for affinity/avidity that results in potent neutralizing capacity. The structural analysis reveals that the most negatively charged paratope correlated with the potency of the mAbs. The atomic level information is of interest to both define the means of autologous neutralization elicited by different tier 2-based immunogens and facilitate trimer redesign to better target more conserved regions of V2 to potentially elicit cross-neutralizing HIV-1 antibodies.

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Challenges and Solutions to Viral Diseases of Finfish in Marine Aquaculture

Pathogens ◽

10.3390/pathogens10060673 ◽

2021 ◽

Vol 10 (6) ◽

pp. 673

Author(s):

Kizito K. Mugimba ◽

Denis K. Byarugaba ◽

Stephen Mutoloki ◽

Øystein Evensen ◽

Hetron M. Munang’andu

Keyword(s):

Marine Fish ◽

Management Practices ◽

Viral Infections ◽

Vaccine Development ◽

Fish Farming ◽

Economic Losses ◽

Diagnostic Tools ◽

Viral Diseases ◽

Marine Aquaculture ◽

Marine Fish Farming

Aquaculture is the fastest food-producing sector in the world, accounting for one-third of global food production. As is the case with all intensive farming systems, increase in infectious diseases has adversely impacted the growth of marine fish farming worldwide. Viral diseases cause high economic losses in marine aquaculture. We provide an overview of the major challenges limiting the control and prevention of viral diseases in marine fish farming, as well as highlight potential solutions. The major challenges include increase in the number of emerging viral diseases, wild reservoirs, migratory species, anthropogenic activities, limitations in diagnostic tools and expertise, transportation of virus contaminated ballast water, and international trade. The proposed solutions to these problems include developing biosecurity policies at global and national levels, implementation of biosecurity measures, vaccine development, use of antiviral drugs and probiotics to combat viral infections, selective breeding of disease-resistant fish, use of improved diagnostic tools, disease surveillance, as well as promoting the use of good husbandry and management practices. A multifaceted approach combining several control strategies would provide more effective long-lasting solutions to reduction in viral infections in marine aquaculture than using a single disease control approach like vaccination alone.

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Viral Myocarditis—From Pathophysiology to Treatment

Journal of Clinical Medicine ◽

10.3390/jcm10225240 ◽

2021 ◽

Vol 10 (22) ◽

pp. 5240

Author(s):

Heinz-Peter Schultheiss ◽

Christian Baumeier ◽

Ganna Aleshcheva ◽

C.-Thomas Bock ◽

Felicitas Escher

Keyword(s):

Molecular Mechanisms ◽

Viral Infections ◽

Treatment Options ◽

Viral Myocarditis ◽

Severe Heart Failure ◽

Diagnostic Methods ◽

Diagnostic Tools ◽

Public Health Issue ◽

Chronic Myocarditis ◽

Significant Public Health

The diagnosis of acute and chronic myocarditis remains a challenge for clinicians. Characterization of this disease has been hampered by its diverse etiologies and heterogeneous clinical presentations. Most cases of myocarditis are caused by infectious agents. Despite successful research in the last few years, the pathophysiology of viral myocarditis and its sequelae leading to severe heart failure with a poor prognosis is not fully understood and represents a significant public health issue globally. Most likely, at a certain point, besides viral persistence, several etiological types merge into a common pathogenic autoimmune process leading to chronic inflammation and tissue remodeling, ultimately resulting in the clinical phenotype of dilated cardiomyopathy. Understanding the underlying molecular mechanisms is necessary to assess the prognosis of patients and is fundamental to appropriate specific and personalized therapeutic strategies. To reach this clinical prerequisite, there is the need for advanced diagnostic tools, including an endomyocardial biopsy and guidelines to optimize the management of this disease. The severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) has currently led to the worst pandemic in a century and has awakened a special sensitivity throughout the world to viral infections. This work aims to summarize the pathophysiology of viral myocarditis, advanced diagnostic methods and the current state of treatment options.

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PERSPECTIVES OF APPLICATION OF ANTIBODIES AGAINST ENDOGLIN (CD105) FOR VISUALIZATION AND ANTI-ANGIOGENE THERAPY OF TUMORS

Problems in oncology ◽

10.37469/0507-3758-2018-64-4-504-507 ◽

2018 ◽

Vol 64 (4) ◽

pp. 504-507

Author(s):

Vladimir Klimovich ◽

Natalya Vartanyan ◽

Anastasiya Stolbovaya ◽

Lidiya Terekhina ◽

Olga Shashkova ◽

...

Keyword(s):

Endothelial Cells ◽

Monoclonal Antibodies ◽

Human Plasma ◽

Vascular Endothelium ◽

Immune Complexes ◽

Targeted Delivery ◽

Biological Fluids ◽

Therapeutic Drugs ◽

Cultured Endothelial Cells ◽

Hybridoma Technology

During last years monoclonal antibodies (MAB) directed against vascular endothelium markers demonstrated their efficiency for visualization and targeted delivery of therapeutic drugs to tumors. Endoglin (CD105) which serves as a key element that determines endothelial cells quiescence or activation is one of such markers. Endoglin is highly expressed on the vascular endothelium of growing tumors. A first panel of MAB against endoglin in our country was produced at the hybridoma technology laboratory of RRC RST named after A.M. Granov. On the basis of these MAB ELISA was created allowing detection of endoglin in human plasma and other biological fluids. Several MAB had been shown to bind endoglin on the membrane of the cultured endothelial cells and to persist there for several hours. During the first 30 min after binding some of the immune complexes “endoglin-MAB” were internalized into the cytoplasm and were found included in the endosomes. In future these MAB can be used to create the reagents for the addressed delivery of isotope tags both on the membrane and into the cytoplasm of endothelial cells.

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Circulating miRNAs Related to Epithelial–Mesenchymal Transitions (EMT) as the New Molecular Markers in Endometriosis

Current Issues in Molecular Biology ◽

10.3390/cimb43020064 ◽

2021 ◽

Vol 43 (2) ◽

pp. 900-916

Author(s):

Anna Zubrzycka ◽

Monika Migdalska-Sęk ◽

Sławomir Jędrzejczyk ◽

Ewa Brzeziańska-Lasota

Keyword(s):

Molecular Mechanisms ◽

Immune Cell ◽

Epithelial Mesenchymal Transition ◽

Clinical Symptoms ◽

Diagnostic Tools ◽

Endometrial Stromal Cells ◽

Disease Etiology ◽

Diagnostic Biomarkers ◽

Mesenchymal Transition ◽

Non Invasive

Endometriosis is a chronic gynecological disease defined by the presence of endometrial-like tissue found outside the uterus, most commonly in the peritoneal cavity. Endometriosis lesions are heterogenous but usually contain endometrial stromal cells and epithelial glands, immune cell infiltrates and are vascularized and innervated by nerves. The complex etiopathogenesis and heterogenity of the clinical symptoms, as well as the lack of a specific non-invasive diagnostic biomarkers, underline the need for more advanced diagnostic tools. Unfortunately, the contribution of environmental, hormonal and immunological factors in the disease etiology is insufficient, and the contribution of genetic/epigenetic factors is still fragmentary. Therefore, there is a need for more focused study on the molecular mechanisms of endometriosis and non-invasive diagnostic monitoring systems. MicroRNAs (miRNAs) demonstrate high stability and tissue specificity and play a significant role in modulating a range of molecular pathways, and hence may be suitable diagnostic biomarkers for the origin and development of endometriosis. Of these, the most frequently studied are those related to endometriosis, including those involved in epithelial–mesenchymal transition (EMT), whose expression is altered in plasma or endometriotic lesion biopsies; however, the results are ambiguous. Specific miRNAs expressed in endometriosis may serve as diagnostics markers with prognostic value, and they have been proposed as molecular targets for treatment. The aim of this review is to present selected miRNAs associated with EMT known to have experimentally confirmed significance, and discuss their utility as biomarkers in endometriosis.

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Role of Circulating Immune Complexes in the Pathogenesis of Canine Leishmaniasis: New Players in Vaccine Development

Microorganisms ◽

10.3390/microorganisms9040712 ◽

2021 ◽

Vol 9 (4) ◽

pp. 712

Author(s):

Cristina Cacheiro-Llaguno ◽

Nuria Parody ◽

Marta R. Escutia ◽

Jerónimo Carnés

Keyword(s):

Immune Complexes ◽

Vaccine Development ◽

Correct Diagnosis ◽

Circulating Immune Complexes ◽

Response To Treatment ◽

Canine Leishmaniasis ◽

Leishmania Infection ◽

Serum Samples ◽

Humoral Immune

During canine visceral leishmaniasis (CanL), due to Leishmania infantum (L. infantum), uncontrolled infection leads to a strong humoral immune response. As a consequence of the production of high antibody levels and the prolonged presence of parasite antigens, circulating immune complexes (CIC) are formed, which can be deposited in certain organs and tissues, inducing vasculitis, uveitis, dermatitis and especially glomerulonephritis and renal failure. A method to detect CIC and quantify their levels in serum samples from dogs infected with L. infantum has been recently described. It allowed demonstration of a correlation between CIC levels and disease severity. Thus, CIC measurement may be useful for diagnosis, assessment of disease progression and monitoring response to treatment. This is an interesting finding, considering that there remains an urgent need for identification of novel biomarkers to achieve a correct diagnosis and for optimal disease staging of dogs suffering from Leishmania infection. The objective of the present review is to shed light on the role of CIC in CanL, as well as to highlight their potential use not only as diagnostic and prognostic biomarkers but also as a valuable tool in vaccine development and new immunotherapy strategies to prevent or control disease outcome.

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Molecular mechanisms of metabotropic GABAB receptor function

Science Advances ◽

10.1126/sciadv.abg3362 ◽

2021 ◽

Vol 7 (22) ◽

pp. eabg3362

Author(s):

Hamidreza Shaye ◽

Benjamin Stauch ◽

Cornelius Gati ◽

Vadim Cherezov

Keyword(s):

G Protein ◽

Molecular Mechanisms ◽

Gabab Receptor ◽

Neurological Diseases ◽

Activation Mechanism ◽

Allosteric Modulators ◽

Inhibitory Neurotransmitter ◽

Therapeutic Drugs ◽

G Protein Coupled ◽

The Brain

Metabotropic γ-aminobutyric acid G protein–coupled receptors (GABAB) represent one of the two main types of inhibitory neurotransmitter receptors in the brain. These receptors act both pre- and postsynaptically by modulating the transmission of neuronal signals and are involved in a range of neurological diseases, from alcohol addiction to epilepsy. A series of recent cryo-EM studies revealed critical details of the activation mechanism of GABAB. Structures are now available for the receptor bound to ligands with different modes of action, including antagonists, agonists, and positive allosteric modulators, and captured in different conformational states from the inactive apo to the fully active state bound to a G protein. These discoveries provide comprehensive insights into the activation of the GABAB receptor, which not only broaden our understanding of its structure, pharmacology, and physiological effects but also will ultimately facilitate the discovery of new therapeutic drugs and neuromodulators.

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630. A 5-mRNA host response whole-blood classifier trained using patients with non-COVID-19 viral infections accurately predicts severity of COVID-19

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa439.824 ◽

2020 ◽

Vol 7 (Supplement_1) ◽

pp. S375-S376

Author(s):

ljubomir Buturovic ◽

Purvesh Khatri ◽

Benjamin Tang ◽

Kevin Lai ◽

Win Sen Kuan ◽

...

Keyword(s):

Logistic Regression ◽

Respiratory Failure ◽

Viral Infections ◽

Lamp Assay ◽

Training Data ◽

Diagnostic Tools ◽

Severe Respiratory Failure ◽

Proof Of Concept ◽

Risk Of Death ◽

Qpcr Platform

Abstract Background While major progress has been made to establish diagnostic tools for the diagnosis of SARS-CoV-2 infection, determining the severity of COVID-19 remains an unmet medical need. With limited hospital resources, gauging severity would allow for some patients to safely recover in home quarantine while ensuring sicker patients get needed care. We discovered a 5 host mRNA-based classifier for the severity of influenza and other acute viral infections and validated the classifier in COVID-19 patients from Greece. Methods We used training data (N=705) from 21 retrospective clinical studies of influenza and other viral illnesses. Five host mRNAs from a preselected panel were applied to train a logistic regression classifier for predicting 30-day mortality in influenza and other viral illnesses. We then applied this classifier, with fixed weights, to an independent cohort of subjects with confirmed COVID-19 from Athens, Greece (N=71) using NanoString nCounter. Finally, we developed a proof-of-concept rapid, isothermal qRT-LAMP assay for the 5-mRNA host signature using the QuantStudio 6 qPCR platform. Results In 71 patients with COVID-19, the 5 mRNA classifier had an AUROC of 0.88 (95% CI 0.80-0.97) for identifying patients with severe respiratory failure and/or 30-day mortality (Figure 1). Applying a preset cutoff based on training data, the 5-mRNA classifier had 100% sensitivity and 46% specificity for identifying mortality, and 88% sensitivity and 68% specificity for identifying severe respiratory failure. Finally, our proof-of-concept qRT-LAMP assay showed high correlation with the reference NanoString 5-mRNA classifier (r=0.95). Figure 1. Validation of the 5-mRNA classifier in the COVID-19 cohort. (A) Expression of the 5 genes used in the logistic regression model in patients with (red) and without (blue) mortality. (B) The 5-mRNA classifier accurately distinguishes non-severe and severe patients with COVID-19 as well as those at risk of death. Conclusion Our 5-mRNA classifier demonstrated very high accuracy for the prediction of COVID-19 severity and could assist in the rapid, point-of-impact assessment of patients with confirmed COVID-19 to determine level of care thereby improving patient management and healthcare burden. Disclosures ljubomir Buturovic, PhD, Inflammatix Inc. (Employee, Shareholder) Purvesh Khatri, PhD, Inflammatix Inc. (Shareholder) Oliver Liesenfeld, MD, Inflammatix Inc. (Employee, Shareholder) James Wacker, n/a, Inflammatix Inc. (Employee, Shareholder) Uros Midic, PhD, Inflammatix Inc. (Employee, Shareholder) Roland Luethy, PhD, Inflammatix Inc. (Employee, Shareholder) David C. Rawling, PhD, Inflammatix Inc. (Employee, Shareholder) Timothy Sweeney, MD, Inflammatix, Inc. (Employee)

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