title: Spacer-arm modulated gene delivery efficacy of novel cationic glycolipids: design, synthesis, and in vitro transfection biology.
abstract: Design, syntheses and relative in vitro gene delivery efficacies of six novel cationic glycolipids 1-6 containing open-form galactosyl units in CHO, COS-1, MCF-7 and A549 cells are described. The results of the present structure-activity investigation convincingly demonstrate that the in vitro gene delivery efficacies of galactosylated cationic glycolipids are strikingly dependent on the absence of a spacer-arm between the open-form galactose and the positively charged nitrogen atom in their headgroup region. While the cationic glycolipids 1-3 with no headgroup spacer unit between the positively charged nitrogen and galactose showed high in vitro gene transfer efficacies in all four cells (lipids 1 and 2 with myristyl and palmityl tails, respectively, being the most efficacious), lipids 4-6 with five-carbon spacer units between the quaternized nitrogen and galactose heads were essentially transfection incompetent. The transfection inhibiting role of the five-carbon spacer unit in the headgroup region of the present novel class of cationic lipids was demonstrated by both beta-galactosidase reporter gene expression and histochemical X-gal staining assays. Results of MTT assay-based cell viability measurements in representative MCF7 cells show that cell viabilities of lipoplexes (lipid:DNA complexes) prepared from all the lipids 1-6 are remarkably high. Thus, possibilities of differential cellular cytotoxicities playing any key role behind the strikingly contrasting transfection properties of lipids 1-3 with no spacer and lipids 4-6 with a spacer unit in the headgroup regions was ruled out. Electrophoresis gel patterns in DNase I sensitivity assays are consistent with more free DNA (accessible to DNase I) being present in lipoplexes of lipids 4-6 than in lipoplexes of lipids 1-3. Thus, the results of our DNase I protection experiments support the notion that enhanced degradation of DNA associated with lipoplexes of lipids 4-6 may play an important role in abolishing their in vitro gene transfer efficacies.
authors.affiliation: Division of Lipid Science and Technology, Indian Institute of Chemical Technology, Hyderabad-500 007, India.
authors.forename: Yenugonda Venkata
authors.initials: YV
authors.lastname: Mahidhar
authors.forename: Mukthavaram
authors.initials: M
authors.lastname: Rajesh
authors.forename: Arabinda
authors.initials: A
authors.lastname: Chaudhuri
mesh_terms: D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004247:DNA / Q000008:administration & dosage*; D003850:Deoxyribonuclease I / Q000737:chemistry; D005696:Galactosidases / Q000235:genetics; D017930:Genes, Reporter; D006017:Glycolipids / Q000138:chemical synthesis* / Q000737:chemistry / Q000633:toxicity; D006801:Humans; D008081:Liposomes / Q000138:chemical synthesis* / Q000737:chemistry; D010316:Particle Size; D055672:Static Electricity; D013329:Structure-Activity Relationship; D014162:Transfection
chemical_list: D006017:Glycolipids; D008081:Liposomes; D004247:DNA; D003850:Deoxyribonuclease I; D005696:Galactosidases