GTMB 7 - Gene Therapy & Molecular Biology

Martin et al: Advances in cationic lipid-mediated gene deliveryFigure 4: Modification to the headgroup moiety: inclusion of a hydroxyethyl group.Aminoglycosides have the further advantage of a knowninteraction with rRNA sequences (Moazed and Noller,1987; Hendrix et al, 1997). Thus we have recentlyreported acylation of the 6’ amine of kanamycin A,resulting in a series of transfection vectors typified byKanaChol (3β–[6’-kanamycin-carbamoyl]cholesterol)whose structure is shown in Figure 5 (Sainlos et al, 2003).KanaChol may be triply charged at physiological pH andwas found to be highly effective for in vitro genetransfection into a variety of mammalian cell lines,especially when formulated with DOPE (Belmont et al,2002). Gene transfer into the airway epithelium is anactive area of research as it could offer treatment for lungdiseases both inherited, such as cystic fibrosis, andacquired, such as asthma. Investigation into the usefulnessof KanaChol/DOPE liposomes to deliver reporter genesinto the mouse airways, confirmed its significant ability totransfect the respiratory epithelium. The applicability oflipid-derivatives of other members of the aminoglycosidefamily to gene delivery is currently under investigation.Guanidinium groups are also able to bind DNAphosphate anions, the interaction in this case beingcharacterised by a pair of parallel hydrogen bonds whichprovide binding strength by their structural organization.In addition, guanidinium groups are highly basic andevidence also exists for hydrogen-bonding to nucleic bases(in particular guanine). Their DNA binding ability shouldtherefore be relatively insensitive to environmentalmodifications (pH, proximity of other positively chargedgroups). Further, the group is found naturally in arginineamino acid residues which play a key role in DNA-bindingproteins such as histones and protamines. Two bisguanidiniumcholesterol derivatives have been synthesisedand tested within our group: BGSC (bis-guanidiniumspermidine-cholesterol)and BGTC (bis-guanidinium-trencholesterol)(Vigneron et al, 1996). The structure ofBGTC is shown in Figure 5. Both vectors were found tobe highly efficient for gene transfection in vitro into avariety of mammalian cell lines when formulated ascationic liposomes with DOPE. In addition, as BGTC wasfound to be soluble in aqueous medium, it could besuccessfully used for transfection when formulatedwithout DOPE. Further studies confirmed the ability ofBGTC/DOPE liposomes to efficiently transfect airwayepithelial cells in vivo (Oudrhiri et al, 1997). Interestinglyhowever, per-guanylation of the amino groups ofKanaChol caused a significant drop in transfection activity(Belmont et al, 2002).278