Yuhua Wang1, Lu Zhang2, Shutao Guo2, Arash Hatefi3, Leaf Huang2. 1. Division of Molecular Pharmaceutics and Center for Nanotechnology in Drug Delivery, Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, USA. Electronic address: alwang@email.unc.edu. 2. Division of Molecular Pharmaceutics and Center for Nanotechnology in Drug Delivery, Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, USA. 3. Department of Pharmaceutics, Ernest Mario School of Pharmacy, University of Rutgers, Piscataway, USA.
Abstract
A novel recombinant protein tetra-H2A (TH) derived from histone H2A has been developed to replace protamine as a conditionally reversible, nucleic acid condensing agent. The novel protein will address the insufficient release of nucleic acid therapeutics, which is captured by protamine for siRNA delivery. TH is composed of 4 tandem repeats of the histone H2A N-terminal sequence, intervened by the cathepsin D cleavage site. The repeating H2A sequence enhances the binding affinity to anionic nucleic acids, forming more stable condensates, as demonstrated by the binding affinity assay. The TH/siRNA condensates are formulated into a core-membrane structured liposomal nanoparticle (NP). The endosomes of cancer cells are rich in cathepsin D, allowing on-site degradation of TH and facilitating the intracellular release of siRNA. The NPs assembled with TH produced a higher silencing efficiency of target genes in vitro and in vivo than the NPs assembled with protamine as the nucleic acid condensing agent. The exploitation of TH in the NP formulation exhibited a biocompatibility profile similar to that of protamine, with minimal immunostimulating and systemic toxicity observed after repeated administration.
A novel recombinant proteinn class="Species">tetra-n class="Gene">H2A (TH) derived from histone H2Ahas been developed to replace protamine as a conditionally reversible, nucleic acid condensing agent. The novel protein will address the insufficient release of nucleic acid therapeutics, which is captured by protamine for siRNA delivery. TH is composed of 4 tandem repeats of the histone H2A N-terminal sequence, intervened by the cathepsin D cleavage site. The repeating H2A sequence enhances the binding affinity to anionic nucleic acids, forming more stable condensates, as demonstrated by the binding affinity assay. The TH/siRNA condensates are formulated into a core-membrane structured liposomal nanoparticle (NP). The endosomes of cancer cells are rich in cathepsin D, allowing on-site degradation of TH and facilitating the intracellular release of siRNA. The NPs assembled withTH produced a higher silencing efficiency of target genes in vitro and in vivo thanthe NPs assembled with protamine as the nucleic acid condensing agent. The exploitation of TH in the NP formulation exhibited a biocompatibility profile similar to that of protamine, with minimal immunostimulating and systemic toxicity observed after repeated administration.
Keywords:
1,2-dioctadecanoyl-sn-glycero-3-phosphoethanolamine; 1,2-dioleoyl-3-trimethylammonium-propane; DOTAP; DSPE; Liposome; Nanoparticle; RNAi; Recombinant protein; TH; TH/ΔCathD; TH/ΔGALA; TH/ΔGALA/ΔCathD; Tumor targeted delivery; tetra-H2A polypeptide; tetra-H2A polypeptide without GALA; tetra-H2A polypeptide without GALA or cathepsin D cleavage site; tetra-H2A polypeptide without cathepsin D cleavage site
Authors: Charles P Lollo; Mariusz G Banaszczyk; Patricia M Mullen; Christopher C Coffin; Dongpei Wu; Alison T Carlo; Donna L Bassett; Erin K Gouveia; Dennis J Carlo Journal: Methods Mol Med Date: 2002
Authors: Danuta Balicki; Christopher D Putnam; Puthupparampil V Scaria; Ernest Beutler Journal: Proc Natl Acad Sci U S A Date: 2002-05-28 Impact factor: 11.205
Authors: Kai Shi; Yi Zhao; Lei Miao; Andrew Satterlee; Matthew Haynes; Cong Luo; Sara Musetti; Leaf Huang Journal: Mol Ther Date: 2017-03-06 Impact factor: 11.454