scholarly journals The Integrin Binding Peptide, ATN-161, as a Novel Therapy for SARS-CoV-2 Infection

2020 ◽  
Author(s):  
Brandon Beddingfield ◽  
Naoki Iwanaga ◽  
Prem Chapagain ◽  
Wenshu Zheng ◽  
Chad J. Roy ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Protein Interaction ◽  
Cell Entry ◽  
Spike Protein ◽  
Novel Therapy ◽  
Binding Peptide ◽  
Viral Cell Entry ◽  
Α5β1 Integrin

SUMMARYMany efforts to design and screen therapeutics for severe acute respiratory syndrome coronavirus (SARS-CoV-2) have focused on inhibiting viral cell entry by disrupting ACE2 binding with the SARS-CoV-2 spike protein. This work focuses on inhibiting SARS-CoV-2 entry through a hypothesized α5β1 integrin-based mechanism, and indicates that inhibiting the spike protein interaction with α5β1 integrin (+/− ACE2), and the interaction between α5β1 integrin and ACE2 using a molecule ATN-161 represents a promising approach to treat COVID-19.

scholarly journals A hybrid soluble gp130/spike-nanobody fusion protein simultaneously blocks IL-6 trans-signaling and cellular infection with SARS-CoV2

2021 ◽  
Author(s):  
Julia Ettich ◽  
Julia Werner ◽  
Hendrik T. Weitz ◽  
Eva Mueller ◽  
Roland Schwarzer ◽  
...  
Keyword(s):  
Specific Inhibitor ◽  
Vero Cells ◽  
Organ Damage ◽  
Cell Entry ◽  
Spike Protein ◽  
Antibody Treatment ◽  
Glucocorticoid Treatment ◽  
Stat3 Phosphorylation ◽  
Dual Action ◽  
Viral Cell Entry

SARS-CoV2 infection can induce mild to life threatening symptoms. Especially individuals over 60 years of age or with underlying co-morbidities including heart or lung disease, and diabetes or immune compromised patients are at higher risk. Fatal multi-organ damage in COVID19 patients can be attributed to Interleukin (IL-)6 dominated cytokine storm. Consequently, IL-6R monoclonal antibody treatment for severe COVID19 cases has been approved for therapy. High concentrations of soluble IL-6R were found in COVID19 intensive care unit patients suggesting the involvement of IL-6 trans-signaling in disease pathology. Here, in analogy to bispecific antibodies (bsAbs), we developed the first bispecific IL-6 trans-signaling inhibitor c19s130Fc which blocks viral infection and IL-6 trans-signaling. c19s130Fc is a designer protein of the IL-6 trans-signaling inhibitor cs130 fused to a single domain nanobody directed against the receptor binding domain (RBD) of the SARS-CoV2 spike protein. c19s130Fc binds with high affinity to IL-6:sIL6R complexes as well as the spike protein of SARS-CoV2 as shown by surface plasmon resonance. Using cell-based assays, we demonstrate that c19s130Fc blocks IL-6 trans-signaling-induced proliferation and STAT3 phosphorylation of Ba/F3-gp130 cells as well as SARS-CoV2 infection and STAT3 phosphorylation in Vero cells. Taken together, c19s130Fc represents a new class of bispecific inhibitors consisting of a soluble cytokine receptor fused to anti-viral nanobodies and principally demonstrates the multi-functionalization of trans-signaling inhibitors. Importance The availability of effective SARS-CoV2 vaccines is a big step forward in managing the pandemic situation. In addition, therapeutic options e.g. monoclonal antibodies to prevent viral cell entry and anti-inflammatory therapies including glucocorticoid treatment are currently developed or in clinical use utilized to treat already infected patients. Here we report a novel dual-specific inhibitor to simultaneously target SARS-Cov2 infection and virus induced hyper-inflammation. This was achieved by fusing an inhibitor of viral cell entry with a molecule blocking IL-6, a key mediator of SARS-CoV2 induced hyper-inflammation. Through this dual action, this molecule may have the potential to efficiently ameliorate symptoms of COVID19 in infected individuals.

scholarly journals Differential Downregulation of ACE2 by the Spike Proteins of Severe Acute Respiratory Syndrome Coronavirus and Human Coronavirus NL63

2009 ◽  
Vol 84 (2) ◽  
pp. 1198-1205 ◽  
Author(s):  
Ilona Glowacka ◽  
Stephanie Bertram ◽  
Petra Herzog ◽  
Susanne Pfefferle ◽  
Imke Steffen ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Lung Injury ◽  
Vero Cells ◽  
Cell Entry ◽  
Spike Protein ◽  
Converting Enzyme ◽  
Human Coronavirus ◽  
Angiotensin Converting Enzyme 2 ◽  
Similar Activity ◽  
Human Coronaviruses

ABSTRACT The human coronaviruses (CoVs) severe acute respiratory syndrome (SARS)-CoV and NL63 employ angiotensin-converting enzyme 2 (ACE2) for cell entry. It was shown that recombinant SARS-CoV spike protein (SARS-S) downregulates ACE2 expression and thereby promotes lung injury. Whether NL63-S exerts a similar activity is yet unknown. We found that recombinant SARS-S bound to ACE2 and induced ACE2 shedding with higher efficiency than NL63-S. Shedding most likely accounted for the previously observed ACE2 downregulation but was dispensable for viral replication. Finally, SARS-CoV but not NL63 replicated efficiently in ACE2-positive Vero cells and reduced ACE2 expression, indicating robust receptor interference in the context of SARS-CoV but not NL63 infection.

scholarly journals A Furin Cleavage Site Inserted into the Spike Protein of SARS-CoV-2: A Structural Implication?

2020 ◽  
Author(s):  
Wei Li
Keyword(s):  
Cleavage Site ◽  
Structural Data ◽  
Cell Entry ◽  
Spike Protein ◽  
Spike Glycoprotein ◽  
Furin Cleavage ◽  
Furin Cleavage Site ◽  
Structural Consequence ◽  
Viral Cell Entry

One notable features of the SARS-CoV-2 genome is that the spike protein of SARS-CoV-2 has a functional polybasic (furin) cleavage site (RRAR) at the S1–S2 boundary through the insertion of 12 nucleotides encoding PRRA. To date, the furin cleavage site (FCS) remains an experimentally uncharted territory both structurally and functionally. For instance, whether or not FCS is actually cleaved, before or after viral cell entry or exit, still remains to be experimentally investigated. With currently available structural data, this article presents a computational structural characterization of the FCS inserted into SARS-CoV-2 spike glycoprotein, and puts forward a set of structural hypothesis against the hypothesis of SARS-CoV-2 from purposeful manipulation: (1), the inserted FCS does not alter, neither stabilize nor de-stabilize, the overall structure of SARS-CoV-2 spike glycoprotein; (2), the net structural consequence of FCS is the insertion of a furin cleavage site into SARS-CoV-2 spike glycoprotein, whose S1 and S2 subunits will still be bonded together even if the FCS is actually cleaved by furin protease.

scholarly journals SARS-CoV-2 variants of concern have acquired mutations associated with an increased spike cleavage

2021 ◽  
Author(s):  
Alba Escalera ◽  
Ana S. Gonzalez-Reiche ◽  
Sadaf Aslam ◽  
Ignacio Mena ◽  
Rebecca L. Pearl ◽  
...  
Keyword(s):  
Cell Entry ◽  
Host Susceptibility ◽  
Spike Protein ◽  
Protein Cleavage ◽  
S Protein ◽  
The Impact ◽  
Viral Cell Entry

For efficient cell entry and membrane fusion, SARS-CoV-2 spike (S) protein needs to be cleaved at two different sites, S1/S2 and S2 by different cellular proteases such as furin and TMPRSS2. Polymorphisms in the S protein can affect cleavage, viral transmission, and pathogenesis. Here, we investigated the role of arising S polymorphisms in vitro and in vivo to understand the emergence of SARS-CoV-2 variants. First, we showed that the S:655Y is selected after in vivo replication in the mink model. This mutation is present in the Gamma Variant Of Concern (VOC) but it also occurred sporadically in early SARS-CoV-2 human isolates. To better understand the impact of this polymorphism, we analyzed the in vitro properties of a panel of SARS-CoV-2 isolates containing S:655Y in different lineage backgrounds. Results demonstrated that this mutation enhances viral replication and spike protein cleavage. Viral competition experiments using hamsters infected with WA1 and WA1-655Y isolates showed that the variant with 655Y became dominant in both direct infected and direct contact animals. Finally, we investigated the cleavage efficiency and fusogenic properties of the spike protein of selected VOCs containing different mutations in their spike proteins. Results showed that all VOCs have evolved to acquire an increased spike cleavage and fusogenic capacity despite having different sets of mutations in the S protein. Our study demonstrates that the S:655Y is an important adaptative mutation that increases viral cell entry, transmission, and host susceptibility. Moreover, SARS-COV-2 VOCs showed a convergent evolution that promotes the S protein processing.

scholarly journals Peptides derived from the SARS-CoV-2 receptor binding motif bind to ACE2 but do not block ACE2-mediated host cell entry or pro-inflammatory cytokine induction

PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0260283
Author(s):  
Amit Mahindra ◽  
Gonzalo Tejeda ◽  
Mario Rossi ◽  
Omar Janha ◽  
Imogen Herbert ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Direct Interaction ◽  
Human Cell Line ◽  
Cell Entry ◽  
Host Cells ◽  
Spike Protein ◽  
Binding Motif ◽  
Viral Attachment ◽  
Contact Residues ◽  
Viral Cell Entry

SARS-CoV-2 viral attachment and entry into host cells is mediated by a direct interaction between viral spike glycoproteins and membrane bound angiotensin-converting enzyme 2 (ACE2). The receptor binding motif (RBM), located within the S1 subunit of the spike protein, incorporates the majority of known ACE2 contact residues responsible for high affinity binding and associated virulence. Observation of existing crystal structures of the SARS-CoV-2 receptor binding domain (SRBD)–ACE2 interface, combined with peptide array screening, allowed us to define a series of linear native RBM-derived peptides that were selected as potential antiviral decoy sequences with the aim of directly binding ACE2 and attenuating viral cell entry. RBM1 (16mer): S443KVGGNYNYLYRLFRK458, RBM2A (25mer): E484GFNCYFPLQSYGFQPTNGVGYQPY508, RBM2B (20mer): F456NCYFPLQSYGFQPTNGVGY505 and RBM2A-Sc (25mer): NYGLQGSPFGYQETPYPFCNFVQYG. Data from fluorescence polarisation experiments suggested direct binding between RBM peptides and ACE2, with binding affinities ranging from the high nM to low μM range (Kd = 0.207–1.206 μM). However, the RBM peptides demonstrated only modest effects in preventing SRBD internalisation and showed no antiviral activity in a spike protein trimer neutralisation assay. The RBM peptides also failed to suppress S1-protein mediated inflammation in an endogenously expressing ACE2 human cell line. We conclude that linear native RBM-derived peptides are unable to outcompete viral spike protein for binding to ACE2 and therefore represent a suboptimal approach to inhibiting SARS-CoV-2 viral cell entry. These findings reinforce the notion that larger biologics (such as soluble ACE2, ‘miniproteins’, nanobodies and antibodies) are likely better suited as SARS-CoV-2 cell-entry inhibitors than short-sequence linear peptides.

scholarly journals Immunological imprinting of the antibody response in COVID-19 patients

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Teresa Aydillo ◽  
Alexander Rombauts ◽  
Daniel Stadlbauer ◽  
Sadaf Aslam ◽  
Gabriela Abelenda-Alonso ◽  
...  
Keyword(s):  
Immune Response ◽  
Severe Acute Respiratory Syndrome ◽  
Antibody Response ◽  
Humoral Immune Response ◽  
Nucleocapsid Protein ◽  
Spike Protein ◽  
Ongoing Research ◽  
Variable Regions ◽  
Humoral Immune ◽  
Human Coronaviruses

AbstractIn addition to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), humans are also susceptible to six other coronaviruses, for which consecutive exposures to antigenically related and divergent seasonal coronaviruses are frequent. Despite the prevalence of COVID-19 pandemic and ongoing research, the nature of the antibody response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is unclear. Here we longitudinally profile the early humoral immune response against SARS-CoV-2 in hospitalized coronavirus disease 2019 (COVID-19) patients and quantify levels of pre-existing immunity to OC43, HKU1 and 229E seasonal coronaviruses, and find a strong back-boosting effect to conserved but not variable regions of OC43 and HKU1 betacoronaviruses spike protein. However, such antibody memory boost to human coronaviruses negatively correlates with the induction of IgG and IgM against SARS-CoV-2 spike and nucleocapsid protein. Our findings thus provide evidence of immunological imprinting by previous seasonal coronavirus infections that can potentially modulate the antibody profile to SARS-CoV-2 infection.

scholarly journals Replication, pathogenicity, and transmission of SARS-CoV-2 in minks

2020 ◽  
Author(s):  
Lei Shuai ◽  
Gongxun Zhong ◽  
Quan Yuan ◽  
Zhiyuan Wen ◽  
Chong Wang ◽  
...  
Keyword(s):  
Animal Model ◽  
Severe Acute Respiratory Syndrome ◽  
Subunit Vaccine ◽  
Lung Damage ◽  
Spike Protein ◽  
Pulmonary Lesions ◽  
Potential Strategy ◽  
Respiratory Droplets

Abstract Minks are raised in many countries and have transmitted severe acute respiratory syndrome–coronavirus 2 (SARS-CoV-2) to humans. However, the biologic properties of SARS-CoV-2 in minks are largely unknown. Here, we investigated and found that SARS-CoV-2 replicates efficiently in both the upper and lower respiratory tracts, and transmits efficiently in minks via respiratory droplets; pulmonary lesions caused by SARS-CoV-2 in minks are similar to those seen in humans with COVID-19. We further found that a spike protein-based subunit vaccine largely prevented SARS-CoV-2 replication and lung damage caused by SARS-CoV-2 infection in minks. Our study indicates that minks are a useful animal model for evaluating the efficacy of drugs or vaccines against COVID-19 and that vaccination is a potential strategy to prevent minks from transmitting SARS-CoV-2.

scholarly journals Long-term persistence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein-specific and neutralizing antibodies in recovered COVID-19 patients

2021 ◽  
Author(s):  
Jira Chansaenroj ◽  
Ritthideach Yorsaeng ◽  
Nasamon Wanlapakorn ◽  
Chintana Chirathaworn ◽  
Natthinee Sudhinaraset ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Symptom Onset ◽  
Neutralizing Antibodies ◽  
Serum Antibody ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Spike Protein ◽  
Immunological Study ◽  
Igg Antibody ◽  
Vaccine Schedule

Abstract Understanding antibody responses after natural severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can guide the coronavirus disease 2019 (COVID-19) vaccine schedule. This study aimed to assess the dynamics of SARS-CoV-2 antibodies, including anti-spike protein 1 (S1) immunoglobulin (Ig)G, anti-receptor-binding domain (RBD) total Ig, anti-S1 IgA, and neutralizing antibody against wild-type SARS-CoV-2 in a cohort of patients who were previously infected with SARS-CoV-2. Between March and May 2020, 531 individuals with virologically confirmed cases of SARS-CoV-2 infection were enrolled in our immunological study. The neutralizing titers against SARS-CoV-2 were detected in 95.2%, 86.7%, 85.0%, and 85.4% of recovered COVID-19 patients at 3, 6, 9, and 12 months after symptom onset, respectively. The seropositivity rate of anti-S1 IgG, anti-RBD total Ig, anti-S1 IgA, and neutralizing titers remained at 68.6%, 89.6%, 77.1%, and 85.4%, respectively, at 12 months after symptom onset. The half-life of neutralizing titers was estimated at 100.7 days (95% confidence interval = 44.5 – 327.4 days, R2 = 0.106). These results support that the decline in serum antibody levels over time depends on the symptom severity, and the individuals with high IgG antibody titers experienced a significantly longer persistence of SARS-CoV-2-specific antibody responses than those with lower titers.

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