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WIREs Nanomed Nanobiotechnol
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Plant viral nanoparticles for packaging and in vivo delivery of bioactive cargos

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Abstract Nanoparticles have unique capabilities and considerable promise for many different biological uses. One capability is delivering bioactive cargos to specific cells, tissues, or organisms. Depending on the task, there are multiple variables to consider including nanoparticle selection, targeting strategies, and incorporating cargo so it can be delivered in a biologically active form. One nanoparticle option, genetically controlled plant viral nanoparticles (PVNPs), is highly uniform within a given virus but quite variable between viruses with a broad range of useful properties. PVNPs are flexible and versatile tools for incorporating and delivering a wide range of small or large molecule cargos. Furthermore, PVNPs can be modified to create nanostructures that can solve problems in medical, environmental, and basic research. This review discusses the currently available techniques for delivering bioactive cargos with PVNPs and potential cargos that can be delivered with these strategies. This article is categorized under: Implantable Materials and Surgical Technologies > Nanomaterials and Implants Biology‐Inspired Nanomaterials > Protein and Virus‐Based Structures
Nanoscaffold and nanocarrier based applications of PVNPs. (a) Cargo loading and targeting PVNPs for vaccine and tissue engineering. (b) Native PVNPs for regulation of tumor microenvironment. (c) Cargo loading and targeting PVNPs with imaging and therapy options for tumors
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Expression and co‐expression for cargo loading during in vivo assembly. (a) Plasmids containing cargo‐CP fusion gene induced in expression systems. Expression of the cargo‐CP gene leads to the production of a chimera CP. The self‐assembly of the chimera CP subunits results in the production of the chimera capsid. (b) The assembly of the VNP‐cargo by expression of the CP gene and the cargo fusion gene. Capsid formation by the interactions of the cargo and CP subunits leading to the encapsulation of the cargo
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Chemical and enzymatic techniques for cargo loading. (a) Addressable bio‐conjugation: employing the reactive amino acid side chains in combination with coupling reagents like EDC and NHS. (b) Enzymatic approaches: synthesis of antigen‐VLP conjugates by enzyme‐mediated coupling of PVNP to cargo
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Cargo loading by the assembly, constrained synthesis and mineralization methods (a) Reassembly, technique and architectures. (a1) Statistical encapsulation process, the capsid assembly is triggered in the presence of cargo and some of the cargo ends up in the capsid as a function of the cargo and coat protein concentrations. (a2) This method, termed “polyacid association,” involves the simultaneous loading of cargo with polyacid (such as nucleic acid polymers or polystyrene sulfonic acid [PSA]) during the reassembly. (a3) Cargo is functionalized with a specific tag or an artificial origin‐of‐assembly site (OAS). The cargo associates with the coat protein prior to assembly and, ideally, triggers or directs the encapsulation process. (a4) The cargo is coated with an anionic surface charge to mimic RNA that interact with positively charged capsid proteins. Then, solvation and steric constraints organize the coat proteins into a symmetric capsid. (a5) Expression of cargo, purification and mixing with CP to enable assembly of capsid with cargo. (b) Constrained synthesis: precursor is added to PVNP, whereby a coupled deprotonation/electron transfer mechanism occurs with amino acids resulting in reduction of precursor to cargo, then nucleation of cargo leads to loading cargo at the interior and/or exterior capsid surfaces, (c) Mineralization: the reactive amino acids arrayed in the interior and exterior of PVNP are specifically bound to the cargos, which initializes the controlled growth of cargos and leads to nucleation and crystallization event
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Scheme of PVNPs as molecular containers, incubation, and infusion methods (a) the exterior surface, the interior surface, and the interface between subunits are all available for chemical and genetic manipulation in an assembled PVNP. (b) Incubation method: surface load PVNPs by incubating cargo with PVNP using electrostatic interactions and hydrogen bonding. (c) Simple infusion methods: PVNP is exposed to a solution of the cargo of interest (in yellow), washing and dialysis is used to remove excess cargo yielding intact PVNP with infused cargo. (d) Gating infusion method: the divalent ions associated with PVNP capsid are removed by dialyzing against divalent cation removal solutions (e.g., EDTA) at high pH, followed by incubation with cargo. The formation of pores within the capsid facilitates the infusion of molecules into the interior cavity of the capsid
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Typical steps in generating plant viruses and VLPs. The viral genome sequence encodes the sequences of proteins that mediate capsid production. These genes are inserted into an expression vector and used to generate capsids in appropriate expression systems
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Biology-Inspired Nanomaterials > Protein and Virus-Based Structures
Implantable Materials and Surgical Technologies > Nanomaterials and Implants

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