Mutations within clones were identified to be critical to the mAb epitope if they did not support reactivity of the test MAbs, but supported reactivity of other ZIKV antibodies. The amount of human mAbs in serum was detected using a capture ELISA with a standard curve of recombinant anti-ZIKV mAb ZIKV or an IgG1 isotype-matched control anti-influenza mAb, FLU-5J8.

Plates were developed using TMB substrate Thermo Fisher Scientific , and the reaction was stopped with H 2 SO 4. ELISA plates were read using a TriBar LB plate reader Berthold Technologies. The optical density values from the known quantity of ZIKV were fitted to a standard curve and compared with the optical density values of serum to determine the concentration of ZIKV The antibodies ZIKV, ZIKV and ZIKV failed to perform in this detection assay, even in cases in which they were used as purified IgG.

Serum concentration measurements for these mAbs are not available. Blood was collected from ZIKV-infected mice at various time points, allowed to clot at ambient temperature and serum was separated using centrifugation.

Viral RNA was isolated using the well Viral RNA kit Epigenetics , as described by the manufacturer. ZIKV RNA levels were determined using TaqMan one-step RT—qPCR as described previously Alternatively, ZIKV Dakar MA and Brazil strain titres were determined using a focus-forming assay on Vero cell monolayer cultures, as previously described For the prophylaxis experiments, mice were treated i.

For the therapeutic protection experiments, mice were treated i. For ZIKV infections, mice were inoculated by a s. The antibody ZIKV was used as a positive control, and the mAb FLU-5J8 or PBS was used as a negative control. The rhesus macaques were aged 5—7 years and were mixed male and female.

All of the animals were inoculated by the s. route with a target dose of 10 6 viral particles ~10 3 p. of ZIKV Brazil before mAb infusions. All of the animals were given physical exams, and blood was collected at the time of ZIKV inoculation and at indicated times after ZIKV inoculation.

Furthermore, all of the animals were monitored daily with an internal scoring protocol approved by the Institutional Animal Care and Use Committee. These studies were not performed blinded. Titration of virus in the indicated specimens was performed using RT—qPCR analysis as previously described 47 , Viral RNA was isolated from plasma and other tested specimens using a QIAcube HT QIAGEN system.

A QIAcube 96 Cador Pathogen kit or RNeasy 96 QIAcube HT kit was used for RNA extraction. cDNA of the wild-type BeH Cap gene was cloned into the pcDNA3. RNA quality was assessed by the Beth Israel Deaconess Medical Center molecular core facility.

Tenfold dilutions of the RNA were prepared for standards and reverse transcribed to cDNA. Primers were synthesized by Integrated DNA Technologies Coralville , and probes were obtained from Biosearch Technologies Petaluma.

Viral loads were calculated as virus particles per ml, and the assay sensitivity was copies per ml. A modified protocol using the commercially available human anti-ZIKV-Env IgG kit Alpha Diagnostics International was used to quantify ZIKV mAb levels in NHP serum samples. A four-parameter logistic standard curve was generated using Prism v.

were determined for continuous variables as noted. Survival curves were estimated using the Kaplan—Meier method, and an overall difference between groups was estimated using the two-sided log-rank Mantel—Cox test.

In the neutralization assays using focus-reduction neutralization tests, IC 50 values were calculated after log transformation of antibody concentrations using a three-parameter nonlinear fit analysis. In the RTCA neutralization assays, IC 50 values were estimated as cellular index change over time using nonlinear fit with variable slope analysis determined in the RTCA v.

Technical and biological replicates are described in the figure legends. Statistical analyses were performed using Prism v. Further information on research design is available in the Nature Research Reporting Summary linked to this article.

The main data supporting the results in this study are available within the paper and its Supplementary Information. The raw and analysed datasets generated during the study are too large to be publicly shared, yet they are available for research purposes from the corresponding authors on reasonable request.

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Cell Host Microbe 19 , — Download references. We thank M. Mayo for assistance with acquisition of the human survivor samples and coordination across study sites; J.

Govero for assistance with RNA protection experiments; A. Jones and K. Beeri for assistance and coordination of NGS sequencing timelines; J. Slaughter, M. Goff and R. Troseth for assistance with data analysis; and STEMCELL Technologies and ACEA Biosciences for providing resources.

This study was supported by Defense Advanced Research Projects Agency DARPA grant HR and HHS contract HHSNC to J. and B. The content is solely the responsibility of the authors and does not necessarily represent the official views of the DARPA.

These authors contributed equally: Pavlo Gilchuk, Robin G. Bombardi, Jesse H. Erasmus, Qing Tan, Rachel Nargi, Cinque Soto, Peter Abbink, Todd J. Vanderbilt Vaccine Center, Vanderbilt University Medical Center, Nashville, TN, USA.

Pavlo Gilchuk, Robin G. Bombardi, Rachel Nargi, Cinque Soto, Taylor Jones, James E. Pre-Clinical Vaccine Development, Infectious Disease Research Institute, Seattle, WA, USA.

Jesse H. Erasmus, Amit Khandhar, Jacob Archer, Elise Larson, Stacey Ertel, Brian Granger, Jasmine Fuerte-Stone, Steven G. Department of Medicine, Washington University School of Medicine, St Louis, MO, USA. Qing Tan, Lorellin A. Durnell, Michael S. Center for Virology and Vaccine Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA. Todd J. Suscovich, Vicky Roy, Thomas Broge, Thomas C. Linnekin, Caitlyn H. Linde, Matthew J.

Jenny Liang, Mallorie E. Department of Molecular Microbiology, Washington University School of Medicine, St Louis, MO, USA.

Andrew M. and Jane M. Sawdon and Peng used acyclovir directly to initiate the polymerization of ε-caprolactone to form the hydrophobic core of acyclovir—polycaprolactone. This was further grafted with methoxy polyethylene glycol to form the amphiphilic block copolymer which could self-assemble to form the nanosized polymeric micelles in the aqueous medium [ 78 ].

These polymeric micelles were found to enhance drug-loading capacities and eliminate drug-loading steps. In another study, a multifunctional poly l -lactic acid -b-poly ethylene glycol PLLA-b-PEG copolymer modified with a sialic acid derivative methyl-b-neuraminic acid, mNA was prepared and further used to form polymeric micelles.

Amantidine was loaded in these micelles and sialic acid is known to bind hemagglutinin of influenza viruses to inhibit the virus-mediated hemagglutination and infection [ ]. The advantages of polymeric nanoparticles as carrier system include controlled release, better cellular uptake, protection of drug molecule from degradation site-specific delivery, low toxicity, and their ability to be used as theranostics [ , ].

Polymeric nanoparticles have been widely explored for enhancing the efficacy of antiretroviral drugs due to their high applicability in targeting monocytes and macrophages in the brain and lymphatic system which are the principal reservoirs for viral dissemination during HIV infections [ ].

Various surface modifications and ligand conjugation approaches have been found to exhibit added advantages of promoting receptor-mediated endocytosis and hence increased uptake by the target cells.

Polyethylene oxide, polyethylene glycol, poloxamers, mannose, thiamine, and Tat are among the various surface modifiers tried for targeted HIV therapy [ ]. Polymer drug conjugates are comprised of a polymer and a therapeutic agent covalently bound, the therapeutic agent being a small molecule or a large molecule like protein [ ].

The purpose of conjugation is to achieve better efficacy through extended plasma stability and safety through targeted delivery [ , ]. Besides cancer, such conjugates have found tremendous potential in antiviral therapy. Some polymers are known to possess their inherent antiviral activity and their conjugation with antiviral drugs may act in a synergistic manner.

Conjugation of interferon α2A and α2A with polyethylene glycol PEG has been found effective against HCV [ ]. Polymers containing sialic acid functionality were designed and found to be potent inhibitors of viral entry through competitive multivalent binding to hyaluronic acid HA and NA on the viral envelope [ ].

Various negatively charged polymers like poly methacrylic acid PMAA and different glycosaminoglycans such as heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, and keratan sulfate have exhibited efficient binding to the HIV envelope resulting in shielding of the viral particles and their entry into the host cells [ , ].

AZT conjugates with naturally occurring polymers like chitosan, dextrin, and κ-carrageenan through succinic ester linker have resulted in longer plasma half-lives and high loading of AZT.

Phosphoamide-based prodrugs have been explored in which the antiviral drug stavudine was conjugated with chitosan through phosphoamide linkage. This resulted in greater antiviral potency and reduced toxicity.

Synthetic polymers like vinyl esters and methacrylates have been explored for conjugation with AZT and ribavirine showing beneficial results [ ]. In , Andersen et al. designed albumin—polymer—drug conjugates to ensure longer residential time and lymphatic accumulation, delivering the right ratio of individual drugs of ART to primary human T cells exhibiting strong protection from HIV infection in comparison with individual ART drugs.

The polymer used was N- 2-hydroxypropyl methacrylamide PHPMA [ ]. Dendrimers are highly branched, well-defined, monodisperse three-dimensional, synthetic nano-architectures, 2—10 nm in diameter, consisting of a central core, an inner shell made up of repeating units of the building blocks, and an outer shell containing numerous functionalities [ ].

The voids and flexible spaces within dendrimers facilitate entrapment of the guest molecules, and the surface functional groups allow interaction with the desired biological targets or can carry the drug by electrostatic interaction or chemical conjugation [ ].

They are synthesized by sequential addition of the building block molecules to the initiator through convergent or divergent approach, by click or lego chemistry [ ]. Dendrimers have the advantages being used as theranostics and show excellent uptake by cells, longer circulation times, and enhanced solubility and stability and targeted delivery [ ].

Among the commercially available dendrimers are polyamidoamine PAMAM , poly-propylene imine PPI , and poly- l -lysine dendrimers PLL [ ]. Dendrimers are functionalized mainly with three types of molecules: carbohydrates, peptides, and anionic groups. Interestingly, these dendrimers possess inherent antimicrobial or antiviral activities.

Dendrimers decorated with anionic groups interact with V3 loop of gp protein and interfere with early stages of viral replication. PPI dendrimers with randomly sulfated galactose residues and PLL dendrimers with sulfated galactose and cellobiose residues have demonstrated good antiviral activity [ ].

In recent times, glycodendropeptide with up to 16 peptides and 9 mannose units emerged as a promising tool for optimizing immune response against viral infections [ ].

Polyanionic carbosilane dendrimers PCDs have been explored against HIV and herpes simplex virus HSV -2 as topical microbicides [ ].

They are shown to inhibit viral entry fusion process and prevent these sexually transmitted infections. Some of them bind directly to HSV-2 virus, inactivating it, and some adhere to the host cell-surface proteins. They exhibit synergistic activity with antiviral drugs like acyclovir and tenofovir against HSV-2 in vitro.

The combination of PCDs with tenofovir and maraviroc as monotherapy has shown to enhance efficacy, reduce doses and side effects, and minimize emergence of multidrug-resistant mutants of HIV mutants resistant to nucleoside reverse transcriptase inhibitors [ ].

In another study, PCDs have shown to prevent HCV infection in cell culture [ ]. In a study carried out by Landers et al. In another study carried out by García-Gallego et al. In addition, these metallodendrimers also prevented the entry of virus in peripheral blood mononuclear cells taken as a model for second barrier against HIV infection [ ].

Ammonium terminated amphiphilic Janus dendrimers were shown to self-assemble in water to form micelles capable of carrying the antiviral drug camptothecin. These drug-loaded dendrimers were found to be highly efficacious against replicating HCV at lower working concentrations and hence displayed low toxicity and better therapeutic index than the free drug [ ].

Biodegradable poly phosphor-hydrazone dendrimers with end phosphoric acid functionalities and alkyl chains have been proposed for anti-HIV activity [ ].

Thiolated dendrimers loaded with acyclovir were developed by Yandrapu et al. which exhibited sustained release and mucoadhesion [ ]. Recently, Martínez-Gualda et al. synthesized a new class of dendrimers which are pentaerythritol derivatives containing multiple aromatic and nonaromatic amino acids on the periphery using the convergent approach.

These dendrimers were found to exhibit dual activity against HIV and enterovirus 71 EV71 responsible for hand-foot-and-mouth disease prevalent among children below 6 years of age.

They found dendrimer with peripheral N-methyl tryptophan to be most potent against HIV-1 and that with tyrosine to be most active against EV71 [ ]. A nanocapsule consists of nanosized structure 50— nm having a core and a shell. The drug is confined to the inner core, surrounded by the polymeric shell.

Nanocapsules exhibit advantages of high drug loading, controlled release, and targeted drug delivery. They are usually prepared by polymer coating, layer-by-layer method, nanoprecipitation, emulsion—diffusion, emulsion coacervation, emulsion evaporation, and double emulsification [ ]. In a study, nanocapsule consisting of poly iso -butylcyanoacrylate core, entrapping azidothymidine-triphosphate AZT-TP and polyethyleneimine which forms the shell, was designed to directly deliver AZT-TP into the cytoplasm [ ].

Nanospheres are smaller spherical structures of 10— nm diameter, where the drug is uniformly dispersed in the matrix system. This type of nanocarrier shows enhanced size-dependent characteristics, having the capability to prevent the drug from undergoing degradation. In addition, rapid drug clearance is observed due to smaller size.

It offers site-specific delivery and the required drug release profiles. Recommended preparation techniques used are solvent evaporation, polymerization solvent displacement techniques, and phase inversion temperature methods [ ]. Chitosan nanospheres loaded with acyclovir were synthesized using modified nanoemulsion template method for topical treatment of herpes.

The spheres were of the average size of nm and showed better permeation in in vitro skin permeation studies and higher potency against HSV-1 and HSV-2 than the free acyclovir itself [ ].

Cyclodextrins CDs are cyclic oligosaccharides made of six to twelve α- d -glucopyranose monomers linked by α1—4 linkages having exceptional hydrophobic interior surfaces and hydrophilic rims with primary and secondary —OH groups.

This type of molecular construct entraps the drug in a bucket-shaped cavity, thereby increasing the solubility of the drug and protecting the drug from degradation. Due to this, CDs become preferred delivery system for drugs [ ].

The common native or parent cyclodextrins are α-CD, β-CD, and γ-CD comprising of 6, 7, and 8 glycopyranose units and molecular weights of , , and Da, respectively [ ].

These CDs have a homogeneous crystalline structure offering numerous advantages like their unique ability to interact with a range of organic and inorganic lipophilic molecules and form inclusion complexes. But their use is restricted due to low solubility of the CDs.

Thus, alteration in CDs is being made to make them more suitable for their application in the pharmaceutical industry [ ].

β-CD and its derivatives are more widely used than α-CD and γ-CD because of their safety and ease of production. The structural framework of β-CD is attractive with a height of — pm, external diameter of pm, internal diameter of — pm, and cavity volume of — Å 3 [ ].

These dimensions make the most ideal hosts for the formation of inclusion complexes. Chemical and enzymatic modifications of macrocycle l in CD derivatives that self-assemble in aqueous solutions provide different shapes of supramolecular nano-assemblies vesicles, micelles, nanorods, nanospheres, and other kinds of nanoparticles and liquid crystalline structures of 30— nm in size depending on the concentration which are very useful for different types of nanodelivery systems [ ].

One of the common problems encountered with antiviral drugs is their poor bioavailability. A similar problem was found with the drug saquanavir. Couvreur and Vauthier formulated CDs loaded with saquanavir using poly alkylcyanoacrylate to deal with the issue. This improved solubility in water by fold.

Also, it was speculated that saquanavir could now bypass the efflux mechanism of P-gp, preventing its resistance [ ]. Similarly, in another study, acyclovir was loaded with CDs using the copolymer such as Eudragit RLPO®, and on evaluation, it was found that intracellular uptake of the drug increased and it had sustained drug release over a period of 24 h [ ].

Thus, by utilizing other drugs, CDs with different polymers can be formulated to increase the intracellular concentration of the drug. Several peptides from natural includes plants, arthropod venoms, amphibian skin, mammalian tissues and microbial sources includes bacteria, algae, and fungi have been known to possess broad-spectrum antiviral activities.

Various mechanisms, either targeting the virus or the host cells have been proposed for their antiviral activities. Some examples include magainin 1 and 2, dermaseptin S4 and temporin B from frog skin , clavanin from marine source , latarcin, protegrin from swine WBCs , cyclotides from plants , cecropin from moth , defensins and cathelicidins from mammals , and poly-γ-glutamic acid from bacteria exhibiting potent activity against various viruses like HIV, H1N1, DENV, and HCV [ , ].

In addition, certain peptides, rationally designed and synthesized depending on the structure of the viral protein and its interaction with the host cell protein, have shown great potential as antiviral agents. Enfuvirtide is the first peptide antiviral drug approved against HIV [ ].

Despite various advantages, various hurdles in their production, shorter half-life, and poor bioavailability have limited their use as antiviral agents.

Nanotechnology-based solutions have been explored for the delivery of antiviral peptides. Peptide—nanoparticle conjugate systems have been extensively studied. Emileh et al. have reported gold nanoparticle—peptide triazole conjugates to be active against HIV-1 by disrupting the interactions between host receptor proteins and trimeric envelope spike glycoprotein of virus [ ].

Recently, Alghrair et al. functionalized silver and gold nanoparticles with an antiviral peptide FluPep and reported increased antiviral potency against influenza A virus [ ]. Table 3 compiles a list of polymer-based nanoformulations for antiviral treatment. Carbon-based nanoformulations are comprise of carbon nanotubes, graphene oxide nanoparticles, and fullerenes.

Carbon nanotubes CNTs are cylindrical-shaped hollow nanomaterials, viewed as tubes made by rolling up of planar graphene sheets. They can be viewed as coming from the rolling up of a graphene sheet, named as single-walled carbon nanotubes SWCNTs , or a series of concentric rolled-up graphene sheets termed as multi-walled carbon nanotubes MWCNTs [ ].

The cylindrical structure is capped with fullerene sheets on one end or both ends. The sp 2 -hybridized carbon atoms in graphene sheets impart a unique strength to CNTs. In addition, they display other unique characteristics like high aspect ratio, high surface area, cell penetration capacity, and ultralight weight [ ].

The chemical vapor deposition CVD technique, laser-ablation technique, and electric arc-discharge techniques are commonly employed for the preparation of carbon nanotubes [ ]. Though CNTs are widely explored for delivering chemotherapeutic agents at the target site, their overall application in the biomedical field is limited due to pulmonary toxicity and high hydrophobicity [ ].

The proposed mechanisms for toxicity are uptaken by macrophages with subsequent generation of ROS and inflammatory mediators. However, functionalized CNTs have shown decreased toxicity and increased biodegradability. CNTs can be decorated with peptides, carbohydrates, and polymers and can be used for targeted therapy, when needed [ ].

In one study, Kumar et al. stated about protoporphyrin IX PPIX -conjugated multi-walled nanotubes MWNTs and its ability to treat influenza using photodynamic therapy. It was found that in the presence of visible light, PPIX-MWNT may indulge in mechanisms like RNA strand breakage, protein oxidation, or protein—RNA cross-linking caused by reactive oxygen species singlet oxygen and superoxide anion leading to inactivation of the influenza viral strain.

Probing into the inactivation mechanism of carbon nanotubes, they concluded that PPIX-MWNTs can be used for treating any viral infection as it displays nonspecificity in treating viral diseases. Also, PPIX-MWNT can be easily recovered through filtration and reused. Due to its multitarget mechanisms of antiviral action, it was proposed that PPIX-MWNTs have less chances of development of drug resistance [ ].

Nanostructures have shown antiviral effect in respiratory syncytial virus, a virus causing severe bronchitis and asthma. The treatment is generally done by combining nanoparticles and gene-silencing technologies. In a novel approach, MWCNTs were functionalized with recombinant dengue virus 3 envelop proteins.

This induced significant immune responses in mice [ ]. Similarly, conjugation of functionalized CNTs with B and T cell peptide epitopes could generate a multivalent system that was able to induce a strong immune response; thus, CNTs were considered to be good candidates for vaccine delivery [ ].

Further, functionalized CNTs were used for the transport of peptides such as foot-and-mouth virus peptide for vaccination [ ]. One of the most promising carbon-based nanomaterials with great potential for antiviral application is graphene.

Graphene G is a two-dimensional 2-D planar sheet of hexagonally arranged sp 2 -hybridized carbon atoms obtained from its three-dimensional 3-D material of graphite [ ]. It is chemically oxidized to graphene oxide GO to acquire oxygen bearing functional groups like hydroxyl, epoxide, and carboxylic acids [ ].

Graphene-based nanomaterials GBNs have high surface area, high loading capacity, and superior mechanical strength which make them attractive candidates for carrying antiviral agents [ ].

The oxygen-containing functional groups allow surface functionalization and conjugation strategies and show biocompatibility, reduced toxicity, and good dispersibility [ ]. The amphiphilicity of GO makes the incorporation of hydrophilic as well as hydrophobic moieties possible [ ].

In addition, these functional groups also provide attachment sites for various biological molecules like proteins, DNA, and RNA [ ]. Recently, Pokhrel et al. studied the interactions between graphene and VP40 viral matrix protein of Ebola virus using molecular dynamics simulations and graphene pelleting assay.

Graphene was found to interact strongly with VP at various interfaces crucial for the formation of the viral matrix.

They proposed the use of graphene-based nanoparticle solutions as disinfectant to prevent the Ebola epidemic [ ]. In another study, 18 sulfonated magnetic nanoparticles were anchored onto reduced graphene oxide SMRGO sheets and used to trap and destroy HSV-1 photothermally, upon their irradiation with near-infrared light.

It was found to be effective against 28 viral infections including HSV. It was found that SMRGO has higher entrapment efficiencies in comparison to magnetic nanoparticles due to increased entrapping efficiency, larger surface area, unique sheet-like structure, and outstanding photothermal characteristics shown by graphene [ ].

Fullerenes are among the first discoveries in symmetric carbon nanostructures and have received considerable attention in the case of antiviral research. Fullerenes are comprised fully of carbon atoms forming a nanosized caged hollow sphere. Buckminster fullerene C60 , also known as buckyball, is the most common form of fullerenes with 60 carbon atoms arranged in a spherical structure showing high symmetry [ ].

Due to their unique architecture, immense scope of derivatization, free radical scavenging activity, and low toxicity, they are widely studied for drug delivery and antimicrobial and antiviral activities [ ].

Fullerenes were found to fit inside the hydrophobic cavity of HIV proteases and inhibit HIV replication [ ].

Structure—activity relationship studies revealed trans-position of substitutions and positive charge near the cage to be important for antiviral activity. Fulleropyrolidines with two ammonium groups have been shown to be active against HIV-1 and HIV-2 [ ].

Also, fullerene C60 derivatized with two or more solubilizing side chains has been active when tested in CEM culture cells infected with HIV-1 and HIV-2 [ ].

Additionally, amino-acid derivatives of fullerene C60 are found to inhibit HIV and HCV replication [ , ]. Few studies aimed at screening fullerene derivatives for anti-influenza activity. Shoji et al. screened 12 fullerene derivatives for in vitro PA endonuclease inhibition.

PA represents the subunit of influenza A RNA polymerase which demonstrates endonuclease activity. It was found that 8 fullerene derivatives demonstrated endonuclease inhibiting potential. In the MDCK cell culture system, these fullerene derivatives inhibited influenza A virus infection and expression of viral nucleoprotein [ ].

Few of the studies also aimed at synthesizing anti-influenza fullerenes and evaluating their effectiveness against the influenza virus. Tollas and colleagues also prepared a fullerene conjugate having a thiosialosyl-a 2,6 -galactose disaccharide and evaluated to understand the multimeric interaction of a sialocluster with influenza virus NA and HA.

Results revealed that these fullerene derivatives did not target HA but was able to target influenza NA slightly [ ]. Table 4 gives an account of various carbon-based nanoformulations for antiviral treatment.

Quantum dots QDs 2—10 nm are semiconductor nanocrystals having the shape of dots. They are comprised of a semiconductor core, overcoated by a shell, and a cap leading to improved solubility in aqueous buffers [ ]. Fundamental semiconducting character and unique optical and electronic properties are attributed to the presence of the inorganic core consisting of semiconducting materials like silicon, cadmium selenide, cadmium sulfide, or indium arsenide [ ].

Quantum dots find interesting applications in biomedical imaging due to its limited light scattering, narrow emission bands, and low tissue penetration.

Quantum dots have been widely explored as theranostic platforms for simultaneous sensing, imaging, and therapy [ ]. The advantages of quantum dots as a drug carrier system include improved bioavailability and stability of drugs, increased circulation times, active targeting, and localized therapy.

In addition to this, QDs can be surface modified with targeting ligands [ , ]. Yong et al. utilized saquinavir and transferrin Tf -conjugated quantum dots for the treatment of HIV. In vitro studies demonstrated that higher concentrations of saquinavir were able to cross the BBB by this method [ ].

Metal and metal oxide nanoparticles have been widely explored for their antiviral activity. Among the various metal nanoparticles showing high efficacy are silver and gold, and among the various metal oxides are CuO, SiO 2 , TiO 2 , and CeO 2.

These nanoparticles have shown great efficacy against a broad spectrum of viruses like influenza H3N2 and H1N1 , HBV, HSV, HIV-1, HSV, dengue virus type-2, foot-and-mouth disease virus, and vesicular stomatitis virus [ ].

Metal nanoparticles by virtue of their unique shape, size, structure, and local-field enhancement action can interact with viral surface proteins through Kazimir interaction and van der Waals forces causing its inactivation [ ].

A variety of surface functionalizations with silane or thiol groups have shown to enhance interaction with biomolecules, affecting viral internalization in cells as well as the release of the drug molecule. Few researchers have also explored further grafting on functionalized metal nanoparticles in order to enhance their efficacy and selectivity [ ].

Gold nanoparticles AuNPs are colloids of nanosized particles of gold. AuNPs show special optical properties in the presence of light. When AuNP comes in contact with light, the oscillating electromagnetic field of light triggers coherent oscillation of the free gold electrons [ ]. This electron oscillation about the particle surface is responsible for a charge separation with regard to the ionic lattice, leading to a dipole oscillation in the path of the electric field of light.

However, when the amplitude of the oscillation becomes maximum at a particular frequency, photons get confined to a small particle size and leads to a special phenomenon known as surface plasmon resonance SPR [ ]. This SPR boosts all the radiative and nonradiative characteristics of the nanoparticles and, thus, has extensive application in areas of biological imaging, electronics, and materials science [ ].

In , Bayo et al. demonstrated that AuNP could enter the cells of various types like lymphocytes, macrophages, and brain microendothelial cells where HIV is known to replicate.

Further, they modified raltegravir RAL by introducing a thiol group that served as a linker between RAL and AuNP. Surprisingly, when the concentration of RAL was increased with the expectation of displaying high antiviral activity, it was found that anti-HIV activity was impaired.

The experiment showed positive results when a low concentration of RAL was loaded into AuNP. The authors also performed an experiment using free AuNP and the results proved that it does not have any antiviral activity.

So it was just a low concentration of RAL-loaded AuNP which turns it into an active compound having inhibitory action against HIV [ ]. Small interfering RNAs siRNAs which can target particular viral gene can be employed in the treatment of dengue. However, siRNAs are prone to degradation by serum nucleases and to rapid elimination on account of its small size and anionic character.

Paul et al. conjugated siRNAs with AuNPs and found that the complex had enhanced stability and could reduce dengue virus replication and the release of infectious virion in both pre- and post-infection conditions [ ].

A breakthrough in the arena of gold nanoparticles was the design and synthesis of long and flexible linkers which mimicked heparan sulfate proteoglycans HSPG , a target for viral attachment ligand.

This allowed effective attachment of the virus to HSPG, generating strong forces that eventually lead to viral deformation.

The mechanism was proposed by researchers on the basis of molecular dynamics simulations, electron microscopy images, and virucidal assays. These nanoparticles were nontoxic and effective against a broad range of viruses like HSV, human papilloma virus, respiratory syncytial virus RSV , dengue, and lentivirus.

Additionally, they were found to be active ex vivo in human cervicovaginal histocultures infected by HSV-2 and in vivo in mice infected with RSV [ ].

Recently, Halder et al. synthesized highly monodispersed gold nanoparticles, stabilized by gallic acid. Reduction of AuCl 4 using gallic acid was carried out using ultrasound-induced sonication which produced spherical nanoparticles in the range of 7—8 nm.

These nanoparticles selectively inhibited HSV with EC50 of Prevention of viral cell attachment and penetration was proposed as a mechanism of action of these nanoparticles [ ]. Silver metal possesses intrinsic antimicrobial activity due to its ability to interact with respiratory chain and electron transport chain enzymes and bacterial DNA.

It has been extensively studied since ancient times for fighting infections [ ]. Silver nanoparticles, on account of its small size and enormous surface area, facilitate rapid dissolution and have shown promising activity against a wide spectrum of viruses [ ].

They demonstrate less chances of development of resistance on account of multiplicity of targets they act upon [ ]. Nanoparticles of silver possess their own unique properties with regard to chemical stability, catalytic activity, high conductivity, and localized surface plasma resonance enabling researchers to envisage the implication of using AgNPs in disease therapeutics [ ].

Galdiero et al. were the first authors to describe the antiviral activity of silver nanoparticles against HIV They studied AgNP with three different surface functionalities: foamy carbon—coated AgNPs, poly N-vinylpyrrolidone PVP —coated AgNPs synthesized using glycerol as a reducing agent, and BSA-conjugated AgNPs.

They proposed that the interaction between these NPs and the HIV viral surface glycoprotein gp was size dependent, as only the NPs in the size range of 1—10 nm were able to bind to the virus. BSA- and PVP-coated nanoparticles demonstrated lower inhibitory activity than silver nanoparticles released from the carbon matrix in in vitro assays on laboratory-adapted HIV-1 strain.

AgNPs were shown to block the gp—CD4 interaction. Additionally, they also were proved to inhibit the post-entry stages of infection by complexing with S and O of thiols and phosphates on amino acids and nucleic acid or directly bind to RNA or DNA, thus reducing the rate of reverse transcription [ ].

Baram Pinto et al. synthesized mercaptoethane sulfonate MES —functionalized Au and AgNPs against HSV with a strategy to mimic heparan sulfate, present on the cell surface so that they compete for binding of the virus onto the cell. They inhibited HSV-1 infections by blocking the attachment and entry of the virus into the cell [ ].

AgNPs have been investigated for their activity against H1N1 influenza virus. Xiang et al. prepared and studied the activity of AgNP on H1N1 influenza A virus—induced apoptosis in MDCK cells [ ].

With the help of hemagglutination inhibition test, they found that AgNPs reduced or completely inhibited agglutination of RBCs.

During the MTT assay, they found that the antiviral activity of AgNPs continued for a prolonged period. Surface-decorated AgNPs were explored by Li et al. They proposed interference with ROS-mediated signaling pathway as a mechanism of anti-H1N1 activity [ ]. Similar studies were carried out by the same research group on AgNPs decorated with amantadine [ ].

Moreover, it was seen that reactive oxygen species ROS are formed either on the surface of AgNPs or via release of free silver ions under specific conditions. This ROS induces the cell death of either microbial cells or mammalian cells, depicting a unique antibacterial and antifungal attribute of AgNPs [ ].

Similarly, another study highlighted the use of gold and silver nanoparticles for effective delivery of antiviral peptide FluPep. Its conjugation with the noble metal nanoparticles enhanced the solubility as well as its antiviral activity.

The authors proposed Ag and AuNPs as an effective strategy for delivering the therapeutic peptide [ ]. In another study, modification of silver nanoparticles with tannic acid was able to reduce HSV-2 infections and inflammation in vitro and in vivo. The authors proposed binding of tannic acid with glycoproteins present on the surface of infectious virions as a mechanism for prevention of entry and spread to host cells.

The anti-HSV-2 activity of tannic acid—modified AgNP TAAgNP was more profound than tannic acid alone, and the direct interaction between tannic acid—modified AgNP with virions was essential for the activity.

Pretreatment of host cells with TAAgNPs did not inhibit the entry of the virus into the host cells. Antione et al. designed and synthesized zinc oxide tetrapod nanoparticles ZOTEN with engineered oxygen vacancies using the flame transport synthesis approach. This was achieved by burning mixture of zinc particles, polyvinyl butyrol particles, and ethanol in a furnace at °C, where Zn in vapor form combines with available oxygen resulting in uniform nucleation and growth of the same and forming tetrapod-like structures.

Nano-immunotherapy is one more way in which HSV-2 can be treated. The activity of ZOTEN was tested in female mice. It was seen that clinical signs of vaginal infection was greatly improved with use of this therapy. ZOTEN has high ability to trap HSV-2 virus and then it acts by increasing the presentation of the virus to mucosal APCs, which boosts T cell—mediated and Ab-mediated responses to the HSV-2 infection and, thus, suppresses viral activity [ ].

Despite potential advantages, the use of inorganic nanoparticles has become limited due to its potential toxicity.

Numerous studies have proven to cause toxicity as a result of administration of inorganic nanoparticles [ , ]. Examination of toxicity-related aspects with regard to inorganic nanoparticles is being done by researchers. Table 5 presents various inorganic material—based nanodelivery systems used for the treatment of viral infections.

The inability of a single drug to completely cure viral infection necessitates a multidrug therapy approach. In order to deliver two or more chemotherapeutic drugs with different physicochemical properties in a single delivery vehicle, lipid—polymer hybrid nanoparticles are developed [ ].

They offer benefits like high drug-loading capacity, stealth characteristics, high stability, biocompatibility, prolonged circulation time, and controlled drug release properties. The use of polymers allowed the systems too be surface functionalized and achieve targeted delivery [ ].

On account of several benefits offered by the use of lipid and polymer, several nanocarrier systems have been developed.

Polymer core—lipid shell nanoparticle consists of an inner polymeric core enclosed in one or more outer layers of lipid membranes lipid—PEG and lipoidal shell. Lipid bilayer-coated polymeric nanoparticles consist of a lipid bilayer coated with polymer. Polymer-caged nanoparticles are actually liposomes with their surface modified by cross-linked polymers for better functionality as shown in Fig.

Various methods like emulsification solvent evaporation, nanoprecipitation, high pressure homogenization, and self-assembly nanoprecipitation have been used for the preparation of these systems [ ]. Results illustrated positive outcomes with maximum drug-loading and encapsulation efficiencies were observed [ ].

Table 6 represents various lipid—polymer hybrid systems for antiviral therapy. Biomimetic lipid—polymer hybrid LPH -NPs are formed by modifying the surface of NPs with ligands that mimic cell surface proteins.

These LPH-NPs offer distinct advantages like long circulation time and cell-specific targeting, biocompatibility, increased efficacy, and attenuation of drug resistance [ ]. The approach is becoming increasingly popular in nanotherapeutics as well as in nanovaccines [ ].

Two strategies under this category are virus-like particles VLPs and virosomes. VLPs are self-assembled particles that are formed by incorporating the virus-derived capsid or envelope proteins into various naturally occurring proteins like ferritin, lumazine synthase, and encapsulin, displaying an advantage of precise structure and defined surface functionalities [ ].

Virosomes are virus-like particles having a modified phospholipid bilayer to incorporate the viral envelope glycoproteins like HA or NA. This vesicle is comprised of a reconstituted virus envelope lacking nucleocapsid includes genetic content of the source virus [ ].

The unique properties of these vesicles make them suitable to carry diverse payloads including drugs, antibodies, proteins, and contrast agents.

These systems hold considerable promise as they enable cellular entry, escape endolysosomal entrapment, and achieve targeted delivery [ ]. It is proposed that the risk of potential immunogenicity of such systems can be addressed by incorporating PEG and other targeting moieties in a bilipid layer of virosomes [ ].

Kanekiyo et al. genetically fused influenza HA protein with naturally occurring protein ferritin. This fusion glycoprotein could spontaneously self-assemble to create nanoparticles exposing HA trimeric spikes on their surfaces.

This designed vaccine elicited more potent and broader response than the conventional influenza vaccine [ ].

In another study, a self-assembling protein was rationally designed in silico to present the antigenic prefusion stabilized F-protein from RSV.

This was further synthesized to form self-assembled NP which had the advantages of having optimized stability and immunogenicity [ ]. Stimuli-responsive LPH-NPs SRNPs have the capacity to increase the therapeutic efficacy and to reduce the side effects of drugs by controlling the release of the encapsulated drug exactly at the target site in response to stimuli.

Various stimuli could be pH, temperature, and magnetic field [ ]. For instance, Clawson et al. synthesized LPH-NP using poly lactic-co-glycolic acid core and a lipid—PEG monolayer shell which could get disrupted at low acidic pH and release the drug.

The tunable pH sensitivity was achieved by synthesizing lipid— succinate —mPEG conjugate and using different molar concentrations of this [ ]. The advantage of such system included the stability of NP at neutral pH of blood but specific delivery at low pH sites in the body which includes the tumor microenvironment and deep bronchioles [ ].

Similarly, LPH-NPs containing magnetic beads have been synthesized for the controlled release of drug under the external stimulus of radiofrequency RF magnetic field [ ]. Thermosensitive nanomaterials have been recently utilized for delivering drugs at a specific target site.

You are currently viewing the international English version of our site. Would you like to consult the North America English version? A major innovation serving the fight against Coronaviruses As part of the fight against Coronaviruses, the Serge Ferrari group has developed a coating technology for its membranes which makes its surfaces virucidal.

VirHealth laboratory results The results obtained by comparing two membranes one treated, one untreated demonstrate a reduction in the number of viruses present on surfaces of: Source: independent VirHealth laboratory based on the Coronavirus E ISO standard, representative of human Coronaviruses.

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