Cell-Specific Delivery of Diverse Cargos by Bacteriophage MS2 Virus-like Particles

Abstract

Virus-like particles (VLPs) of bacteriophage MS2 possess numerous features that make them well-suited for use in targeted delivery of therapeutic and imaging agents. MS2 VLPs can be rapidly produced in large quantities using in vivo or in vitro synthesis techniques. Their capsids can be modified in precise locations via genetic insertion or chemical conjugation, facilitating the multivalent display of targeting ligands. MS2 VLPs also self-assemble in the presence of nucleic acids to specifically encapsidate siRNA and RNA-modified cargos. Here we report the use of MS2 VLPs to selectively deliver nanoparticles, chemotherapeutic drugs, siRNA cocktails, and protein toxins to human hepatocellular carcinoma (HCC). MS2 VLPs modified with a peptide (SP94) that binds HCC exhibit a 104-fold higher avidity for HCC than for hepatocytes, endothelial cells, monocytes, or lymphocytes and can deliver high concentrations of encapsidated cargo to the cytosol of HCC cells. SP94-targeted VLPs loaded with doxorubicin, cisplatin, and 5-fluorouracil selectively kill the HCC cell line, Hep3B, at drug concentrations < 1 nM, while SP94-targeted VLPs that encapsidate a siRNA cocktail, which silences expression of cyclin family members, induce growth arrest and apoptosis of Hep3B at siRNA concentrations < 150 pM. Impressively, MS2 VLPs, when loaded with ricin toxin A-chain (RTA) and modified to co-display the SP94 targeting peptide and a histidine-rich fusogenic peptide (H5WYG) that promotes endosomal escape, kill nearly 100% of Hep3B cells (1 × 106 cells/mL population) at an RTA concentration of 100 fM without affecting the viability of control cells. Our results demonstrate that MS2 VLPs, due to their tolerance of multivalent peptide display and their ability to specifically encapsidate a variety of disparate cargos, induce selective cytotoxicity of cancer in vitro and represent a significant improvement in the characteristics of VLP-based delivery systems.

Enhancing and initiating phage-based therapies

“…Typically, about 80 drug molecules are currently loaded in a shell 27.5 nm in diameter. 60 The molar drug concentration is over 10£ lower than for Doxil. (3) Drugs are covalently joined to RNA molecules, thereby generating complexity, a potential source of irreproducibility and problems in unloading.…”

“…In one study, these peptides produce uptake by hepatocarcinoma cells, but not normal liver cells, in culture. 60 A RNA phage-based DDV has the advantage of being genomically coded. Mature phage MS2 has been converted to a display vector.…”

Virus-like nanoparticles: Emerging Tools for Targeted Cancer Diagnostics and Therapeutics

“…Taking advantage of the natural cargo of viruses, a number of VLNPs can be loaded with various nucleic acid based therapeutics, such as plasmids, mRNAs and siRNAs. Requiring more extensive bioengineering, the encapsidation of guest proteins is an exciting area with applications in advanced vaccine design [13], cytotoxic protein delivery [14] and genome editing [15]. Encapsidation of inorganic cargos such as quantum dots [16] and metallic cores offer opportunities in molecular imaging.…”

“…A range of MRI and positron emission tomography agents have also been delivered by VLNPs [17]. Finally, VLNPs have been used to encapsidate high payloads of small molecule therapeutics such as doxorubicin [14], as well as photodynamic agents for light-triggered reactive oxygen generation [10,11].…”

Nature versus nurture in cellular behavior and disease.

“…The enormous capacity of the high-surface-area nanoporous core combined with the enhanced targeting efficacy enabled by the fluid supported lipid bilayer enable a single protocell loaded with a drug cocktail to kill a drug-resistant human hepatocellular carcinoma cell, representing a 106-fold improvement over comparable liposomes. [1] bind to HCC with high affinity due to recruitment of SP94 targeting peptides (magenta) to the cell surface, [2] become internalized via receptor-mediated endocytosis, and [3] release their cargo into the cytosol upon endosome acidification and protonation of the H5WYG fusogenic peptide (blue). Cargos modified with a NLS are transported through the nuclear pore complex and become concentrated in the nucleus [4].…”