Liver-specific Gene Delivery Using Engineered Virus-Like Particles of Hepatitis E Virus

Abstract

Virus-like particles (VLPs) possess great potential for organ-specific transport of therapeutic agents due to their central cavity surrounded by viral capsid proteins and similar tropism to their original viruses. The N-terminal truncated second open reading frame (Nt-ORF2) of the hepatotropic hepatitis E virus (HEV) forms VLPs via self-assembly. In the present study, we investigated whether HEV-LPs could deliver foreign genes specifically to the liver. HEV-LPs were obtained from Nt-ORF2 expression in Huh7 cells that were transduced with recombinant baculoviruses and purified by continuous density gradient centrifugation. The purified HEV-LPs efficiently penetrated liver-derived cell lines and the liver tissues. To evaluate HEV-LPs as gene delivery tools, we encapsulated foreign plasmids in HEV-LPs with disassembly/reassembly systems. Green fluorescence was detected at higher frequency in liver-derived Huh7 cells treated with HEV-LPs bearing GFP-encoding plasmids than in control cells. Additionally, HEV-LPs bearing Bax-encoding plasmids induced apoptotic signatures in Huh7 cells. In conclusion, HEV-LPs produced in mammalian cells can encapsulate foreign genes in their central cavity and specifically transport these genes to liver-derived cells, where they are expressed. The present study could contribute to advances in liver-targeted gene therapy.

Hepatitis E Infection in Nigeria: A Systematic Review

“…Generally, the risk factors of the disease include lack of access to cheap, safe and potable drinking water, unhygienic living conditions and indiscriminate disposal of human wastes, poverty, the insecurity that results to a strain on access to basic life amenities and limited access to quality health services [2]. Symptoms of the disease include mild fever, anorexia, nausea, jaundice, dark urine, pale stools and slight liver inflammation, although, rare cases have been reported to result to fulminant hepatitis in pregnant women [3], [4]. Diagnosis of HEV infection includes detection of the HEV in stool or serum samples and serological tests for identification of anti-HEV Ig G and Ig M [1].…”

In Vitro Assembly of Virus-Like Particles and Their Applications

“…Bacteriophage Qβ VLPs were disassembled in 6 M urea [34], whereas 2.5 M urea were sufficient to denature hepatitis B core protein (HBc) VLPs [35]. Reducing and chelating agents can also be used in VLP disassembly [30,[36][37][38][39][40][41].…”

“…Standard methods are sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) to determine size and purity, Western blot to confirm the identity, and ultraviolet (UV) spectroscopy with light scattering compensation, which monitors amino acids (phenylalanine, tyrosine and tryptophan) and nucleotides with an absorbance spectrum of about 240-300 nm, to determine concentrations [36,72,73]. Other protein quantitation methods, such as the bicinchoninic acid (BCA) assay and Bradford assay have been also used [18,36,74]. Size exclusion chromatography and dynamic light scattering (DLS) are widely used to characterize the size of VLPs [19,[75][76][77].…”

Microfluidic technologies for the synthesis and manipulation of biomimetic membranous nano-assemblies

“…103 Encephalitis, Yellow Fever and Zika Virus. 126 Beyond vaccinology, they are beginning to find use in point-of-care diagnostic research and biosensing, 127,128 as novel delivery vehicles for small molecule drugs and gene therapies, [129][130][131] and as biomimetic particles that could deepen our understanding of virus-cell interactions. 132,133 Despite this range, research incorporating microfluidic techniques into the production and manipulation of VLPs is limited.…”