Induced Disassembly of a Virus-like Particle under Physiological Conditions for Venom Peptide Delivery

Abstract

Virus-like nanoparticles (VLPs) show considerable promise for the in vivo delivery of therapeutic compounds such as bioactive venom peptides. While loading and targeting protocols have been developed for numerous VLP prototypes, induced disassembly under physiological conditions of neutral pH, moderate temperature, and aqueous medium, remain a challenge. Here, we implement and evaluate a ring-opening metathesis polymerization (ROMP) general mechanism for controllable VLP disassembly that is independent of cell-specific factors or the manipulation environmental conditions such as pH and temperature that cannot be readily controlled in vivo. The ROMP substrate norbornene is covalently conjugated to surface-exposed lysine residues of a P22 bacteriophage-derived VLP, and ROMP is induced by treatment of water-soluble ruthenium catalyst AquaMet. Disruption of the P22 shell and release of a GFP reporter is confirmed via native agarose electrophoresis and quantitative microscopy and light scattering analyses. Our ROMP disassembly strategy does not depend on the particular structure or morphology of the P22 nanocontainer and is adaptable to other VLP prototypes for the potential delivery of venom peptides for pharmacological applications.

Bacteriophage P22 Capsid as a Pluripotent Nanotechnology Tool

“…Therefore, the retention of SP variants on the VLP interior is dependent on the particle morphology, which determines whether the putative binding sites are accessible and whether the capsid pore size is large enough to allow cargo passage through the capsid wall [ 13 ]. Other studies have developed different strategies for the release of cargo, such as exploiting the environmental pH to induce disassociation from the VLP structure and exit [ 9 , 12 ] or the programmed particle disassembly [ 76 , 77 ]. The vast array of methods for the internalization of cargo makes the varied types of molecules possible, which in turn makes the equally astounding number of applications possible, some of which are summarized in Table 1 .…”

“…Modifications on the structure can also be used to develop new strategies to bypass issues commonly encountered in development, such as the delivery of cargo. The incorporation of activated 5-Norbornene-2-carboxylic acid into the exterior of the P22 VLP for the programmed release of cargo under physiological pH has already been reported [ 76 , 77 ].…”

“…One of the most interesting methods for the release of cargo proposed the triggering of particle disassembly for the release of cargo in physiological pH under a defined and controllable stimulus, which did not rely on environmental conditions [ 76 , 77 ]. The strategy consisted of the chemical modification of the P22 VLP structure by conjugating Norbornene (5-Norbenen-2-Carboxylic acid) to exposed lysine residues on the particle’s exterior.…”

“…This initial study confirmed the potential of using Norbornene decoration and ROMP reaction for the triggered disassembly of P22 VLP, without being restricted to environmental conditions. However, the effective release of cargo would be confirmed only in a subsequent study, where the highly water-soluble AquaMet catalyst was also tested as a trigger [ 77 ]. In this new research, the release of the SP-GFP cargo was evaluated through fluorescence assay, which evaluated the changes in fluorescence in the aliquots of particles treated with AquaMet catalyst.…”

Rip It, Stitch It, Click It: A Chemist’s Guide to VLP Manipulation

“…Kelly et al demonstrated the use of ring-opening metathesis polymerization (ROMP) for capsid disassembly. (Kelly et al, 2021) First, the P22His6GFP was functionalized on the surface-exposed lysine residues with 5-norbornene-2-carboxylic acid using EDC and sulfo-NHS coupling. Aqua-Met-a ruthenium-based ROMP catalyst-was added for ROMP initiation and subsequent capsid disassembly.…”