title: Chitosan-based therapeutic nanoparticles for combination gene therapy and gene silencing of in vitro cell lines relevant to type 2 diabetes.
abstract: Glucagon like peptide 1 (GLP-1), a blood glucose homeostasis modulating incretin, has been proposed for the treatment of type 2 diabetes mellitus (T2DM). However, native GLP-1 pharmacokinetics reveals low bioavailability due to degradation by the ubiquitous dipeptydil peptidase IV (DPP-IV) endoprotease. In this study, the glucosamine-based polymer chitosan was used as a cationic polymer-based in vitro delivery system for GLP-1, DPP-IV resistant GLP-1 analogues and siRNA targeting DPP-IV mRNA. We found chitosans to form spherical nanocomplexes with these nucleic acids, generating two distinct non-overlapping size ranges of 141-283 nm and 68-129 nm for plasmid and siRNA, respectively. The low molecular weight high DDA chitosan 92-10-5 (degree of deacetylation, molecular weight and N:P ratio (DDA-Mn-N:P)) showed the highest plasmid DNA transfection efficiency in HepG2 and Caco-2 cell lines when compared to 80-10-10 and 80-80-5 chitosans. Recombinant native GLP-1 protein levels in media of transfected cells reached 23 ng/L while our DPP-IV resistant analogues resulted in a fivefold increase of GLP-1 protein levels (115 ng/L) relative to native GLP-1, and equivalent to the Lipofectamine positive control. We also found that all chitosan-DPP-IV siRNA nanocomplexes were capable of DPP-IV silencing, with 92-10-5 being significantly more effective in abrogating enzymatic activity of DPP-IV in media of silenced cells, and with no apparent cytotoxicity. These results indicate that specific chitosan formulations may be effectively used for the delivery of plasmid DNA and siRNA in a combination therapy of type 2 diabetes.
authors.affiliation: Institute of Biomedical Engineering, Department of Chemical Engineering, École Polytechnique, P.O. Box 6079, Station Centre-ville, Montréal, Québec, Canada.
authors.forename: Myriam
authors.initials: M
authors.lastname: Jean
authors.forename: Mohamad
authors.initials: M
authors.lastname: Alameh
authors.forename: Diogo
authors.initials: D
authors.lastname: De Jesus
authors.forename: Marc
authors.initials: M
authors.lastname: Thibault
authors.forename: Marc
authors.initials: M
authors.lastname: Lavertu
authors.forename: Vincent
authors.initials: V
authors.lastname: Darras
authors.forename: Monica
authors.initials: M
authors.lastname: Nelea
authors.forename: Michael D
authors.initials: MD
authors.lastname: Buschmann
authors.forename: Abderrazzak
authors.initials: A
authors.lastname: Merzouki
mesh_terms: D000107:Acetylation; D018938:Caco-2 Cells; D055598:Chemical Phenomena; D048271:Chitosan / Q000009:adverse effects / Q000737:chemistry*; D003924:Diabetes Mellitus, Type 2 / Q000235:genetics / Q000628:therapy; D018819:Dipeptidyl Peptidase 4 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054873:Dipeptidyl-Peptidase IV Inhibitors / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism*; D020868:Gene Silencing; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods; D052216:Glucagon-Like Peptide 1 / Q000031:analogs & derivatives* / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use; D019073:HT29 Cells; D056945:Hep G2 Cells; D006801:Humans; D008970:Molecular Weight; D053758:Nanoparticles / Q000009:adverse effects / Q000737:chemistry* / Q000648:ultrastructure; D010316:Particle Size; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use
chemical_list: D054873:Dipeptidyl-Peptidase IV Inhibitors; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D052216:Glucagon-Like Peptide 1; D048271:Chitosan; C042807:DPP4 protein, human; D018819:Dipeptidyl Peptidase 4