title: Corneal gene delivery: chitosan oligomer as a carrier of CpG rich, CpG free or S/MAR plasmid DNA.
abstract: BACKGROUND
  Corneal gene therapy can potentially treat acquired and inherited corneal disorders that otherwise lead to blindness. In a previous study on the development of effective vectors for corneal gene delivery, we showed that a particular formulation of chitosan-DNA nanoparticles, based on ultrapure chitosan oligomers injected into rat corneas, led to transgene expression that was 5.4-fold higher than that obtained using polyethylenimine-DNA nanoparticles.
  
  
  METHODS
  In the present study, we investigate the same formulation of chitosan-DNA nanoparticles as carriers of six different plasmids for corneal gene delivery. Size, zeta potential, the ability to condense plasmid DNA, and transfection efficiency in cell cultures and in rat corneas, were all investigated.
  
  
  RESULTS
  Size, zeta potential, the ability to condense plasmid DNA, and transfection efficiency in cell cultures did not substantially vary for nanoparticles based on different plasmids. One day post-injection of nanoparticles into rat corneas, we found that a CpG-free plasmid DNA, pCpG-Luc, which has an EF1α promoter, led to transgene expression that was 7.1-fold higher than that for gWiz-Luc, a commercially available plasmid DNA with a cytomegalovirus (CMV) promoter used in our previous study; 116.8-fold higher than that for pEPI-CMV, a commercially available plasmid that has a scaffold/matrix attachment region (S/MAR) sequence and a CMV promoter; and 76.8-fold higher than that for pEPI-UbC, an experimental plasmid that has an S/MAR sequence and a ubiquitin C promoter.
  
  
  CONCLUSIONS
  The present study reveals the potential of comparing various plasmids as an approach for enhancing transgene expression. The delivery system designed in the present study represents the next step in the development of effective vectors for corneal gene therapy.
authors: Department of Pharmaceutical Sciences, South College School of Pharmacy, Knoxville, TN 37922, USA. eklausner@southcollegetn.edu
authors: Eytan A
authors: EA
authors: Klausner
authors: Zhong
authors: Z
authors: Zhang
authors: Suet P
authors: SP
authors: Wong
authors: Robert L
authors: RL
authors: Chapman
authors: Michael V
authors: MV
authors: Volin
authors: Richard P
authors: RP
authors: Harbottle
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D048271:Chitosan / Q000008:administration & dosage* / Q000378:metabolism; D003316:Corneal Diseases / Q000235:genetics / Q000628:therapy*; D004247:DNA / Q000008:administration & dosage* / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008156:Luciferases; D053758:Nanoparticles / Q000008:administration & dosage*; D010957:Plasmids / Q000235:genetics; D051381:Rats; D014162:Transfection / Q000379:methods*; D019076:Transgenes / Q000235:genetics
chemical_list: D049452:Green Fluorescent Proteins; D004247:DNA; D048271:Chitosan; D008156:Luciferases