RESUMO
The Middle East respiratory syndrome coronavirus (MERS-CoV), belonging to the family Coronaviridae and genus Betacoronavirus, has been recognized as a highly pathogenic virus. Due to the lack of therapeutic or preventive agents against MERS-CoV, developing an effective vaccine is essential for preventing a viral outbreak. To address this, we developed a recombinant S1 subunit of MERS-CoV spike protein fused with the human IgG4 Fc fragment (LV-MS1-Fc) in Chinese hamster ovary (CHO) cells. Thereafter, we identified the baculovirus gp64 signal peptide-directed secretion of LV-MS1-Fc protein in the extracellular fluid. To demonstrate the immunogenicity of the recombinant LV-MS1-Fc proteins, BALB/c mice were inoculated with 2.5 µg of LV-MS1-Fc. The inoculated mice demonstrated a significant humoral immune response, measured via total IgG and neutralizing antibodies. In addition, human dipeptidyl peptidase-4 (DPP4) transgenic mice vaccinated with LV-MS1-Fc showed the protective capacity of LV-MS1-Fc against MERS-CoV with no inflammatory cell infiltration. These data showed that the S1 and Fc fusion protein induced potent humoral immunity and antigen-specific neutralizing antibodies in mice, and conferred protection against coronavirus viral challenge, indicating that LV-MS1-Fc is an effective vaccine candidate against MERS-CoV infection.
Assuntos
Infecções por Coronavirus , Coronavírus da Síndrome Respiratória do Oriente Médio , Vacinas Virais , Animais , Anticorpos Neutralizantes , Anticorpos Antivirais , Células CHO , Infecções por Coronavirus/prevenção & controle , Cricetinae , Cricetulus , Dipeptidil Peptidase 4 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Glicoproteína da Espícula de Coronavírus/genética , Vacinas Sintéticas/genética , Vacinas Virais/genéticaRESUMO
The development of a synchronized delivery and imaging system for small interfering RNA (siRNA) is required for the clinical application of RNA interference (RNAi) in cancer treatment. Herein, we report a pH-responsive, magnetic nanoparticle-based siRNA delivery system that can facilitate the safe and efficient delivery and visualization of therapeutic siRNA by high-resolution magnetic resonance (MR) imaging. Cationic poly-l-lysine-graft-imidazole (PLI) with a reactive silane moiety was stably immobilized onto the surface of the assembled manganese ferrite nanoparticles (MFs) through an emulsion process, ensuring high water solubility, enhanced MR contrast effect, and endosome-disrupting functionality. The synthesized nanovectors were then complexed with siRNA targeting the CD44 gene via electrostatic interactions to verify the specific gene-silencing effect. The imidazolized magnetic nanovector (ImMNV) architectures developed here facilitated improved cellular internalization and exhibited a high level in vitro downregulation compared to non-imidazolized MNVs in metastatic breast cancer cells.
RESUMO
Combined cancer treatment via co-delivery of siRNAs and an anticancer drug can be a promising strategy due to the synergistic effect of simultaneously minimizing gene/drug administration. In this study, Bcl-xL siRNA and doxorubicin (DOX) are encapsulated into designed methoxy-poly(ethylene glycol)-block-poly(D,L-lactic acid) (mPEG-b-PLA) block copolymer polymersomes (PSomes). A study of the cytotoxicity of Bcl-xL siRNA and DOX co-encapsulated PSomes (CPSomes) shows more inhibited proliferation of MKN-45 and MKN-28 human gastric cancer cell lines than only gene- and drug-loaded ones. Consequently, these results demonstrate that co-delivery of genes and drugs using PSomes results in a synergistic efficacy and indicates the potential of PSomes as efficient nanocarriers for combined cancer therapy.
Assuntos
Antineoplásicos/farmacologia , Materiais Biocompatíveis/química , Doxorrubicina/farmacologia , Sistemas de Liberação de Medicamentos , Poliésteres/química , Polietilenoglicóis/química , RNA Interferente Pequeno/metabolismo , Proteína bcl-X/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Microscopia Crioeletrônica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Humanos , Concentração de Íons de Hidrogênio/efeitos dos fármacos , Hidrólise/efeitos dos fármacos , Concentração Inibidora 50 , Cinética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Tempo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo , Proteína bcl-X/genéticaRESUMO
We have developed a nanovector consisting of hyaluronic acid (HA) and poly-L-lysine-graft-imidazole (PLI)-based polyplexes containing Bcl-xL-specific shRNA-encoding plasmid DNA (HA/PLI/pDNA) for CD44 targeted gastric cancer therapy. The prepared ternary polyplexes have a negative surface charge of -24 mV and a size of approximately 100 nm at an N/P ratio of 5 with HA/PLI molar ratio of 0.03. Gel electrophoresis and cell viability experiments demonstrated that the ternary polyplexes showed high stability and no cytotoxicity due to the anchored HA molecules on the surface of PLI/pDNA binary polyplexes. Selective cancer cell death was achieved by CD44-mediated gene delivery and the internalized gene was effectively escaped from endosomes due to the buffering capacity of imidazole groups in an acidic environment. These nanovectors may be highly efficient gene delivery tools that allow the selective destruction of metastatic gastric cancer cells.
