A phenoxy-bridged trinuclear Ni(ii) complex: synthesis, structural elucidation and molecular docking with viral proteins

A novel phenoxy-bridged trinuclear nickel(ii) complex [Ni3(mu-L)2(bipy)3](1) (where H3L= (E)-2-hydroxy-N-(2-hydroxy-3,5-diiodophenyl)-3,5-diiodobenzohydrazonic acid, bipy = 2,2'-bipyridyl) has been designed and synthesized as a potential antivirus drug candidate. The trinuclear Ni(ii) complex [Ni3(mu-L)2(bipy)3](1) was fully characterized via single crystal X-ray crystallography. The unique structure of the trinuclear nickel(ii) complex crystallized in a trigonal crystal system with P3221 space group and revealed distorted octahedral coordination geometry around each Ni(ii) ion. The X-ray diffraction analysis established the existence of a new kind of trinuclear metal system containing nickel(ii)-nickel(ii) interactions with an overall octahedral-like geometry about the nickel(ii) atoms. The non-bonded Ni-Ni distance seems to be 3.067 and 4.455 A from the nearest nickel atoms. The detailed structural analysis and non-covalent supramolecular interactions are also investigated by single crystal structure analysis and computational approaches. Hirshfeld surfaces (HSs) and 2D fingerprint plots (FPs) have been explored in the crystal structure to investigate the intermolecular interactions. The preliminary analysis of redox and magnetic characterization was conducted using cyclic voltammetry measurements and a vibrating sample magnetometer (VSM), respectively. This unique structure shows good inhibition performance for SARS-CoV-2, Omicron and HIV viruses. For insight into the potential application of the Ni(ii) coordination complex as an effective antivirus drug, we have examined the molecular docking of the trinuclear Ni(ii) complex [Ni3(mu-L)2(bipy)3](1) with the receptor binding domain (RBD) from SARS-CoV-2 (PDB ID: 7MZF), Omicron BA.3 variant spike (PDB ID: 7XIZ), and HIV protease (PDB ID: 7WCQ) viruses. This structure shows good inhibition performance for SARS-CoV-2, Omicron S protein and HIV protease viruses;the binding energies (DELTAG) and the respective Ki/Kd (inhibition/dissociation constants) correlation values are -8.9 (2.373 muM or 2373 nM), -8.1 (1.218 muM or 1218 nM) and -7.9 (0.874 muM or 874 nM), respectively. The results could be used for rational drug design against SARS-CoV-2 Omicron variant and HIV protease viruses.Copyright © 2023 The Royal Society of Chemistry.

Omicron BQ.1.1 and XBB.1 unprecedentedly escape broadly neutralizing antibodies elicited by prototype vaccination.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariants have seriously attacked the antibody barrier established by natural infection and/or vaccination, especially the recently emerged BQ.1.1 and XBB.1. However, crucial mechanisms underlying the virus escape and the broad neutralization remain elusive. Here, we present a panoramic analysis of broadly neutralizing activity and binding epitopes of 75 monoclonal antibodies isolated from prototype inactivated vaccinees. Nearly all neutralizing antibodies (nAbs) partly or totally lose their neutralization against BQ.1.1 and XBB.1. We report a broad nAb, VacBB-551, that effectively neutralizes all tested subvariants including BA.2.75, BQ.1.1, and XBB.1. We determine the cryoelectron microscopy (cryo-EM) structure of VacBB-551 complexed with the BA.2 spike and perform detailed functional verification to reveal the molecular basis of N460K and F486V/S mutations mediating the partial escape of BA.2.75, BQ.1.1, and XBB.1 from the neutralization of VacBB-551. Overall, BQ.1.1 and XBB.1 raised the alarm over SARS-CoV-2 evolution with unprecedented antibody evasion from broad nAbs elicited by prototype vaccination.

REMDESIVIR RETAINS POTENT ACTIVITY AGAINST SARS-CoV-2 VARIANTS OF CONCERN

Background: Recent SARS-CoV-2 variants of concern (VOCs) have shown a progressive loss of sensitivity to monoclonal antibody therapeutics. Remdesivir (RDV) is a nucleotide analog prodrug that targets the viral RNA-dependent RNA polymerase (RdRp) Nsp12 and is approved to treat COVID-19 in hospitalized and non-hospitalized patients. Nsp12 is highly conserved across VOCs to date and RDV antiviral activity against previous VOCs (Alpha to Omicron BA.1) has been maintained. Here, we conduct a structural analysis of Nsp12 substitutions observed in recent Omicron subvariants (BA.2, BA.2.12.1, BA.4, BA.5 and BA.2.75) and assess RDV antiviral activity against clinical isolates and sitedirected mutants (SDMs) in a replicon system. Method(s): The prevalence of Nsp12 substitutions in Omicron subvariants was evaluated by analysis of sequences from the Global Initiative on Sharing Avian Influenza Data (GISAID) EpiCoV database. Structural analysis of identified substitutions was conducted on a prior cryo-electron microscopy-based model of the replication-transcription complex. Antiviral activity against subvariant clinical isolates was assessed by nucleoprotein ELISA in A549-hACE2-TMPRSS2 cells and by SDMs in the replicon system. Result(s): Genomic analysis of >1.4 million Omicron subvariant sequences revealed unique substitutions in Nsp12 compared to the ancestral WA1 strain. Besides P323L, present in all subvariants, G671S was observed in 95.9% of BA.2.75 sequences, F694Y was observed in <=1.9% of BA.4, BA.5 and BA.2.75 sequences, and Y521C was observed in 1.7% of BA.5 sequences. As anticipated, structural analysis of these substitutions showed no direct interaction with the incoming RDV nucleotide triphosphate or the viral RNA. Phenotyping of clinical isolates of Omicron subvariants BA.2, BA.2.12.1, BA.4, BA.5, and BA.2.75 consistently resulted in mean RDV EC50 values of 24.5 nM (BA.2) to 106.0 nM (BA.5). This represented 0.15-to 0.66-fold changes compared to WA1, indicating no loss of in vitro RDV antiviral activity against these VOCs. P323L, G671S, and F694Y were shown previously to have no impact on RDV antiviral activity. Similarly, the individual substitution Y521C showed no change in RDV susceptibility in the SARS-CoV-2 replicon system. Conclusion(s): RDV retained potent in vitro antiviral activity against all tested Omicron VOCs with potencies comparable to the WA1 isolate. These data support the continued use of RDV in patients infected with Omicron subvariants.

BROAD ANTIBODIES TO FUNCTIONALLY CONSTRAINED REGIONS OF SARS-CoV-2 SPIKE

Background: While remarkable and rapid progress was made in fighting the SARS-CoV-2 pandemic with vaccines and therapeutic antibodies, these approaches were quickly compromised by viral evolution. Therapeutic monoclonal antibodies (mAbs) that were once authorized for clinical use, which all target the receptor binding domain (RBD), are no longer effective against current variants of concern (VOCs) due to mutations in this region of Spike. Thus, to achieve durable protection against SARS-CoV-2, novel mAbs need to show breadth and potency across VOCs and target epitopes that are more constrained. Method(s): mAbs from an individual who had a breakthrough Delta VOC infection after vaccination were isolated from Spike-specific memory B cells. mAbs were assessed for binding affinity and neutralization potency using Spike-pseudotyped lentivirus (PSV) and live SARS-CoV-2 virus neutralization assays. Epitopes were mapped using deep mutational sequencing (DMS) and structural-based methods. Result(s): Three novel mAbs (C68.3, C68.13, C68.59) demonstrated binding breadth to Spikes from various VOCs including Omicron VOCs despite that C68 had not yet been exposed to Omicron. These mAbs potently neutralized the Wuhan-Hu-1 vaccine and Delta strains (IC50 = 9-61ng/mL), and early Omicron strains BA.1, BA.2, BA.5 (IC50 = 12-149 ng/mL). C68.3 and C68.59 retained potency against recent VOCs BQ.1.1 and XBB (IC50 = 121-122 ng/mL and 56-82 ng/mL, respectively) in the PSV assay. Similar neutralization activity was observed in the live virus assay. The potency of these mAbs was greater against Omicron VOCs than all but one of the mAbs previously authorized for treatment and they showed greater breadth. The mAbs target distinct epitopes on the Spike glycoprotein, two in the RBD (C68.3, C68.13) and one in an invariant region downstream of RBD in subdomain 1 (SD1) (C68.59). Structural analysis of C68.59 Fab binding to Spike trimer revealed significant allosteric changes to regions of Spike outside of the epitope in the S2 unit. Finally, DMS escape pathways showed these mAbs target regions highly conserved across VOCs that are also functionally constrained, suggesting escape could incur a fitness cost. Conclusion(s): Overall, these mAbs are novel in their breadth across VOCs and include a potent mAb targeting a rare epitope outside of the RBD in SD1. These mAbs focus on diverse, functionally constrained regions in Spike making them candidates for development as combination therapeutics with good durability against future VOCs.

The Impact of Life Stressors Associated With the COVID-19 Pandemic on Essential Workers of Color

Objective: Although essential workers of color in the United States have been notably at risk during the COVID-19 pandemic, less is known about the impacts of associated life stressors within this population. Thus, the purpose of this study is to understand (a) the impact of life stressors among a sample of essential workers of color during the COVID-19 pandemic and (b) associations between life stressors and sociodemographic characteristics, work, mental health, substance use, and eating behaviors. Method: Essential workers of color (N=319) completed the Holms-Rahe Life Stress Inventory to examine whether any set of 43 life stress experiences occurred because of COVID-19. We used latent class analysis to examine patterns of clustering. Results: We identified three latent classes: (a) minimally impacted, (b) moderately impacted, and (c) widely impacted. Most of our sample reported being minimally impacted (53%) or moderately impacted (35%) by life events during the pandemic. However, participants clustered within the widely impacted pattern (11%) reported significantly more concerns with substance use, binge eating, and perceived stress. Conclusions: Findings highlight distinct patterns of life-stress impacts caused by COVID-19 among essential workers of color. Social workers must consider how to implement policies to reduce the systematic challenges faced by those who are most marginalized.

Amyloidogenesis: What Do We Know So Far?

The study of protein aggregation, and amyloidosis in particular, has gained considerable interest in recent times. Several neurodegenerative diseases, such as Alzheimer's (AD) and Parkinson's (PD) show a characteristic buildup of proteinaceous aggregates in several organs, especially the brain. Despite the enormous upsurge in research articles in this arena, it would not be incorrect to say that we still lack a crystal-clear idea surrounding these notorious aggregates. In this review, we attempt to present a holistic picture on protein aggregation and amyloids in particular. Using a chronological order of discoveries, we present the case of amyloids right from the onset of their discovery, various biophysical techniques, including analysis of the structure, the mechanisms and kinetics of the formation of amyloids. We have discussed important questions on whether aggregation and amyloidosis are restricted to a subset of specific proteins or more broadly influenced by the biophysiochemical and cellular environment. The therapeutic strategies and the significant failure rate of drugs in clinical trials pertaining to these neurodegenerative diseases have been also discussed at length. At a time when the COVID-19 pandemic has hit the globe hard, the review also discusses the plausibility of the far-reaching consequences posed by the virus, such as triggering early onset of amyloidosis. Finally, the application(s) of amyloids as useful biomaterials has also been discussed briefly in this review.

Synthesis, crystal structure, DFT calculations, and Hirshfeld surface analysis of an NNN pincer type compound

The novel benzamide derivative NNN pincer type, N,N'-(azanediylbis(2,1-phenylene))bis(3-chlorobenzamide) (H3L), was synthesized from bis(2-nitrophenyl)amine starting material. The pincer ligand was characterized by 1H NMR, 13C NMR, COSY, HMQC, and FT-IR techniques. The geometry of pincer ligand was also confirmed by a single-crystal X-ray diffraction analysis. Structural analysis demonstrate that H3L is monoclinic and space group P21/n with Z = 4. It was find out the molecular conformation of the structure is promoted by intramolecular (N[sbnd]H⋅⋅⋅O, N[sbnd]H⋅⋅⋅N, and C[sbnd]H⋅⋅⋅O) and intermolecular (N(2)-H(2)⋅⋅⋅O(2)i, symmetry code (i) = 1/2 + x, 3/2-y, 1/2 + z) hydrogen bonds. The theoretical study of H3L was performed in the gaseous phase by B3LYP/6-311G(d,p) method to determine the structural properties of the title molecule, as a consequence the obtained data showed that the considerable agreement between the experimental and theoretical results. The reactivity and stability of the molecule were evaluated by calculating the HOMO–LUMO energy gap which was found as 6.5163 eV. In addition, FMO, NBO, NLO, DOS, RDG, MEP surface, and Mulliken atomic charge analyses were carried out. Hirshfeld surface analysis and two-dimensional fingerprint plots were investigated and the obtained data exposed that the most significant contributions to the crystal packing are from C···H/H···C (33.2%), H···H (31.5%), and H···Cl/Cl··H (18.9%) contacts. Furthermore, the molecular docking studies were performed to reveal the binding affinity between the title compound and the main protease (6LU7) of COVID-19 coronavirus. © 2023 Elsevier B.V.

Is there a Common Digital Market in the European Union? Implications for the European Digitalization Strategy

Due to the impact of the COVID‐19 pandemic, people has changed the way they work, learn and socialize. As result, it is important to identify the pre‐existing digital gaps to implement the European Union digitalization strategy. This study aims to identify typologies of internet use in the 28 European Union (EU) countries (at the time of the survey), based on the characteristics of the users and their internet usage patterns. A two‐level latent class analysis was applied. At the first level, individuals within each country were grouped according to their characteristics of internet use;and, simultaneously, at the second level, countries were grouped based on the similar structure of individual segments. Using data from Eurobarometer, results show that internet use in the EU digital market is not homogeneous. The European Commission should take these pre‐existing gaps into account in the EU digitalization strategy.

Posttraumatic Stress Disorder Checklist for Dsm-5 (Pcl-5). Latent Structure Analysis and Measurement Invariance in a Sample of Italian Health Workers during the Covid-19 Pandemic

The COVID-19 pandemic poses a significant threat to health workers (HW) in terms of posttraumatic stress symptoms (PTSS) and disorder (PTSD). Over the years, alternative PTSD structures have been proposed (DSM-5, Dysphoria, Dysphoric Arousal, Anhedonia, Externalizing Behaviors, Hybrid) and tested. To date, no studies have addressed this issue focusing on HW during the COVID-19 pandemic. This study investigated the fit of alternative PTSD structures in two Italian samples: HW during the COVID-19 pandemic, and university students in a pre-pandemic context. A total of 580 HW and 451 students completed the Posttraumatic Stress Disorder Checklist for DSM-5 (PCL-5) assessing PTSS. Confirmatory factor analysis investigated the best PTSD structure in each sample;measurement invariance was also inspected. The Anhedonia structure performed best in both samples;this model showed configural, metric, variances and covariances invariance. Results pave the way to the use of the PCL-5 to tailor intervention supporting HW during the pandemic.

POSTTRAUMATIC STRESS DISORDER CHECKLIST FOR DSM-5 (PCL-5). LATENT STRUCTURE ANALYSIS AND MEASUREMENT INVARIANCE IN A SAMPLE OF ITALIAN HEALTH WORKERS DURING THE COVID-19 PANDEMIC

The COVID-19 pandemic poses a significant threat to health workers (HW) in terms of posttraumatic stress symptoms (PTSS) and disorder (PTSD). Over the years, alternative PTSD structures have been proposed (DSM-5, Dysphoria, Dysphoric Arousal, Anhedonia, Externalizing Behaviors, Hybrid) and tested. To date, no studies have addressed this issue focusing on HW during the COVID-19 pandemic. This study investigated the fit of alternative PTSD structures in two Italian samples: HW during the COVID-19 pandemic, and university students in a pre-pandemic context. A total of 580 HW and 451 students completed the Posttraumatic Stress Disorder Checklist for DSM-5 (PCL-5) assessing PTSS. Confirmatory factor analysis investigated the best PTSD structure in each sample;measurement invariance was also inspected. The Anhedonia structure performed best in both samples;this model showed configural, metric, variances and covariances invariance. Results pave the way to the use of the PCL-5 to tailor intervention supporting HW during the pandemic. [ FROM AUTHOR]

SPR analysis on the interactions between heparin and SARS-CoV-2 spike protein RBDs from different variants

The COVID-19 pandemic has become a major human health concern. The pathogen responsible for COVID-19, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), invades its host through the interaction of its spike (S) protein with a host cell receptor, angiotensin-converting enzyme 2 (ACE2). In addition to ACE2, heparan sulfate (HS) on the surface of host cells also plays a significant role as a co-receptor. In this study, SARS-CoV-2 SP receptor binding domain (RBD) wild type (WT), Delta and Omicron variants were expressed in Expi293F cells and used in the kinetic and structural analysis on their interactions with heparin by Surface plasminogen resonance (SPR). Detailed kinetic and structural analysis of the interactions of SARS-CoV-2 SP RBDs with heparin provides important information for designing anti-SARS-CoV-2 molecules.

A Systematic Approach for Contextualizing Focal Structure Analysis in Social Networks

Focal Structures are key sets of individuals who may be responsible for coordinating events, protests, or leading citizen engagement efforts on social media networks. Discovering focal structures that can promote online social campaigns is important but complex. Unlike influential individuals, focal structures can effect large-scale complex social processes. In our prior work, we applied a greedy algorithm and bi-level decomposition optimization solution to identify focal structures in social media networks. However, the outcomes lacked a contextual representation of the focal structures that affected interpretability. In this research, we present a novel Contextual Focal Structure Analysis (CFSA) model to enhance the discovery and the interpretability of the focal structures to provide the context in terms of the content shared by individuals in the focal structures through their communication network. The CFSA model utilizes multiplex networks, where the first layer is the users-users network based on mentions, replies, friends, and followers, and the second layer is the hashtag co-occurrence network. The two layers have interconnections based on the user hashtag relations. The model's performance was evaluated on real-world datasets from Twitter related to domestic extremist groups spreading information about COVID-19 and the Black Lives Matter (BLM) social movement during the 2020–2021 time. The model identified Contextual Focal Structure (CFS) sets revealing the context regarding individuals’ interests. We then evaluated the model's efficacy by measuring the influence of the CFS sets in the network using various network structural measures such as the modularity method, network stability, and average clustering coefficient values. The ranking Correlation Coefficient (RCC) was used to conduct a comparative evaluation with real-world scenarios. © 2022, The Author(s), under exclusive license to Springer Nature Switzerland AG.

Causal Network Structure Analysis of Pharmaceutical and Biological Industry under the COVID-19

Nowadays, countries in the world are increasingly connected, and major emergencies affect the development of various industries, which makes particularly important to measure industry association.In this paper, we extract ordinary period structure before the outbreak of the COVID-19 and explode pharmaceutical biological industry network, then apply convergence cross mapping causal inference to describe the industry network, further establish the network of industry network topology to measure node and industry system risk. Empirical results show that the network structure of the pharmaceutical and biological industry is similar in the normal period and outbreak period before the epidemic, and the association within the industry was relatively stable. When the epidemic hit the network, the linkage of the pharmaceutical and biological industry is significantly enhanced, and the systemic risks and network efficiency are higher than usual. The network of pharmaceutical and biological industry is of strong robustness and strong ability to deal with emergencies, which provides some reference for grasping the stability of industry network structure and industry risk management under sudden shocks in pharmaceutical and biological industry. © 2021 IEEE.

Initial Adaptation Study of COVID-19 Peritraumatic Distress Index (CPDI) to Turkish Sample

Peritraumatic distress, which is regarded as a predictor of post-traumatic stress disorder (PTSD), expresses the feeling of fear and helplessness experienced in the face of a traumatic event. It has been evaluated that the COVID-19 pandemic can lead to peritraumatic reactions and PTSD symptoms. The study aimed to adapt the "COVID -19 Peritraumatic Distress Index (CPDI)", which was developed to determine peritraumatic distress levels during the COVID-19 pandemic, to Turkish culture. The study was conducted with 248 participants. The CPDI scale items were translated into the Turkish language. Confirmatory factor analysis (CFA) was performed to determine the construct validity of the scale. And the Cronbach alpha (a) reliability coefficient was calculated for reliability. As a result of the analyses, the original structure of the scale was confirmed and construct validity was ensured. This measurement tool can be used to assess the peritraumatic distress levels of the Turkish population.

Identification and functional-structural characterization of the enzymes linked with stable genome size increase of (+) RNA viruses

Nidovirales is an extraordinary order of complex positive-stranded RNA viruses including some of the largest RNA genomes (12-41 kb) among which notable human health burdens: SARS-CoV-1, SARS-CoV-2, MERS-CoV, etc. Recent advance in genome sequencing is slowly filling the gaps between and beyond the classified nidoviral families. Still, the research is lagging behind to understand the evolution of RNA genomes. For example, how are these large genome RNA viruses able to bypass the length and stability constraints of an RNA molecule? Is there any link between increasing length and gaining a functional domain or a special structural feature? To answer these questions, we started with database mining to extract novel nidoviral genomes and annotated different domains in polyproteins of classified and unclassified nidoviruses using HHpred and HHblits tools (Zimmermann L, et al. 2018). We observed a significant variation across the order regarding presence/absence, fold/structure type, co-factor (or enhancer) presence/absence, presence of one motif or the other and genome location of enzymes: Exonuclease (ExoN), N-7 Methyltransferase (MTase), 2'-O-MTase and RNA dependent RNA polymerase (RdRp). A trend seen with this bioinformatic analysis directly implies that stable RNA genome increase as well as maintenance is driven by the synergy of modifying enzymes: MTases, RNA proofreading by ExoNs and fast & processive RdRps (Ferron F, et al. 2021). Next, after their identification, we are trying to characterize these large RNA genome genetic markers: MTase(s) & ExoN, to have a comprehensive understanding of nidoviruses evolution. We have identified, expressed and purified a new nidoviral MTase from a Tobaniviridae family member, White Bream Virus (WBV). This enzyme is unique in terms of its location in ORF1a and not in ORF1b (Ferron F, et al. 2019). Functional and mutational studies show this new MTase to contain N-7 guanine specific, S-adenosyl-methionine (SAM) dependent capping activity (cap-0). Aligning with our predictions, structural characterization confirms that it has a Rossmann fold (RF) SAMdependent RNA-cap N7-guanine MTase. This study answers the missing link of capping activity in these members, which is somewhat only established for coronaviruses in this large genome order. Evaluating such enzymes is a step forward in the direction of fundamental understanding of how these RNA viruses are successfully expanding and maintaining their large genomes as well as coping up to fight against the host innate immunity.

STRUCTURAL CHARACTERIZATION of DNA-ENCODED HIV VACCINES INDUCED NEUTRALIZING ANTIBODY

Background: Rapid and large-scale deployment of COVID-19 mRNA vaccines highlights the potential utility of developing nucleic acid vaccines (such as RNA and DNA vaccines) against infectious diseases, including HIV-1. However, as compared to SARS-CoV-2, HIV-1 pose some unique challenges-induction of neutralizing antibodies (NAbs) against HIV-1 (frequently a correlate of protection) requires presentation of trimeric and highly conformational epitopes to the immune system, and whether nucleic acid vaccines can enable direct in vivo production of antigens that retain critical antigenic profile has not yet been elucidated. Additionally, it was previously reported that Tier 2 NAbs cannot be induced in mice due to a lack of antibody repertoire, and vaccine studies were suggested to be performed in larger mammals such as rabbits/NHPs, inadvertently slowing down and increasing the costs of preclinical HIV-1 vaccine studies. Methods: In our study, we used the Antigen Conformation Tracing In Vivo by ELISA (ACTIVE) assay developed in house to characterize antigenic profiles of vaccines produced in vivo (from transfected muscle tissues). We analyzed induced cellular responses, using stimulation with overlapping peptides followed by intracellular cytokine staining and IFN-g ELIspot assays. We analyzed induced humoral responses by using both binding ELISA assays and TZM-BL based neutralizing assays, and attempted to map induced NAb epitopes by engineering selectively mutated pseudovirus. We performed antigen-specific B-cell sorting, and used the 10x genomics pipeline to characterize antibody sequences of proliferating B-cell clones. Results: We confirmed that in vivo produced vaccines retained key trimeric conformational epitopes and glycan profiles. Compared to protein vaccination, DNA vaccination uniquely and strongly induced both TFH, CD4+, CD8+ T-cell responses, and Tier 2 NAbs mapped to a previously unreported Env C3/V5 epitope. 5 unique NAbs were isolated, and confirmed to bind to the epitope using a Cryo-EM structure of NAb-MD39 complex at 3.8Å resolution. Conclusion: Our study confirmed that with appropriate vaccine delivery technology, murine models can be appropriately used for HIV-1 vaccine studies aimed at generating NAb responses. In addition, beyond potential functional immunity gains, DNA vaccines permit in vivo folding of structured antigens and provide significant cost and speed advantages for enabling rapid evaluation of new HIV vaccines.

A NOVEL CLASS of ANTIVIRAL COMPOUNDS with POTENT ACTIVITY AGAINST SARS-CoV-2

Background: The SARS-CoV-2 pandemic has sickened over 245 million people, and has killed more than 5 million worldwide. Recent data proves that vaccinations are highly effective in preventing Covid-19 disease, however antigenic drift and other functional mutations in the virus genome reduce the efficacy of vaccines, indicating that the development of antiviral treatments remain a crucial priority. We report potent antiviral activity against SARS-CoV-2 for a promising, novel class of nitrogen-based heterocyclic compounds. Methods: 232 compounds based on the same class of nitrogen-based hetereocyclic molecules were synthesized to final purity of greater than 99%. This library was screened for antiviral phenotypes in a cytopathic effect (CPE) assay using VeroE6 cells and the SARS-CoV-2 WA1 isolate. Based on the results of the WA1 CPE screen, 47 lead candidates were structurally analyzed, and this information was utilized to design 56 additional compounds. A second antiviral CPE-based screen was performed using these 103 candidates in VeroE6 cells with the SARS-CoV-2 delta variant. Antiviral assays studying SARS-CoV-1 (Urbani) and MERS-CoV were performed in Vero 76 cells utilizing a Neutral Red cytopathic effect assay. Results: Within the same class of structurally related small molecules, we tested an initial set of 232 compounds using a CPE-based assay with VeroE6 cells and the USA/WA1 SARS-CoV-2 isolate. Of the compounds tested, 124 demonstrated potency 10 to 540-times higher than a Remdesivir control tested in parallel. Importantly, we observed no detectable toxicity for the vast majority of these compounds when tested up to a concentration of 30 μ M. The lead candidate in this screen displayed an IC50 of 0.02 μ M and a selectivity index of >1,500. Based on structural analysis of an initial 47 lead candidates, we synthesized 56 new molecules, and tested all 103 in a CPE-based assay using the delta variant, also observing efficacy against this variant of concern. Examples of this same class of compounds also display antiviral activity against SARS-CoV-1 (Urbani) and MERS-CoV in cell-based assays. Conclusion: We have identified a novel class of antiviral compounds with potent activity against SARS-CoV-2. High potency against both the early WA1 isolate and the more recent delta variant, as well as efficacy against SARS-CoV-1 and MERS-CoV, suggest that this class of antiviral compounds has pan-Coronavirus antiviral activity.

A FC-ENHANCED NON-NEUTRALIZING ANTIBODY DELAYS SARS-CoV-2-INDUCED DEATH in MICE

Background: Both neutralizing activity and Fc-mediated effector functions of antibodies are believed to contribute to protection against SARS-CoV-2. However, it is unclear if antibody effector functions alone could protect against SARS-CoV-2 infection. Methods: We isolated CV3-13 from a convalescent individual with potent Fc-mediated effector functions. Neutralization capacity of this antibody was measured by both a pseudovirus neutralization assay and an authentic virus microneutralization assay. We mutated the Fc-portion of CV3-13 to enhance (GASDALIE) or reduce (LALA) its capacity to mediate antibody dependant cellular cytotoxicity (ADCC). Structural analysis of CV3-13 was done by cryo-EM to characterize its epitope and its angle of approach. Finally, CV3-13 and CV3-13 GASDALIE were used in vivo in a K18-hACE2 transgenic mouse model challenged with SARS-CoV-2-nLuc to see if they altered viral replication and/or contributed to protection against SARS-CoV-2. Results: While CV3-13 did not neutralize SARS-CoV-2, it demonstrated nanomolar affinity towards the SARS-CoV-2 Spike and mediated strong ADCC. The cryo-EM structure of CV3-13 in complex with the SARS-CoV-2 Spike revealed that the antibody bound to a novel NTD epitope that partially overlapped with a frequently mutated NTD supersite in SARS-CoV-2 variants. Interestingly, this angle of approach was not observed for previously described NTD-directed antibodies. While CV3-13 did not alter the replication dynamics of SARS-CoV-2 in a K18-hACE2 transgenic mouse model, a Fc-enhanced CV3-13 significantly delayed neuroinvasion and death in prophylactic settings. Conclusion: CV3-13 represents a new class of non-neutralizing NTD-directed mAbs that can mediate Fc-effector functions both in vitro and in vivo. While effector functions alone did not protect K18-hACE2 mice from SARS-CoV-2-nLuc challenge, our data indicate that along with neutralization, additional antibody properties including Fc-mediated effector functions contribute to limiting viral spread and aid in fighting SARS-CoV-2 infection.

BIOFABRICATION OF NANOSTRUCTURED GELATIN FIBER VIA CENTRIFUGAL SPINNING METHOD

In this study, type-I bovine collagen solved in diluted acidic solutions (acidic acid and formic acid) and fed into the centrifugal spinning device to obtain nanofiber formation. The centrifugal spinning device is providing submicron fibers thanks to its rotational movement during the solution feeding. Different feeding ratios and rotational speeds are studied to optimize the process. Obtained na nofiber webs were observed via scanning electron microscopy (SEM). Structural characterization of samples was investigated via FTIR. Fabricated collagen-based nanofibers filtration performance was evaluated in terms of filtration efficiency and air permeability tests. Tests results are pointed that fabricated gelatin nanofibers can be an efficient alternative biomaterial design against Covid-19.