Non-viral Gene Therapy - Intracellular Delivery: Fundamentals and Applications - page 614

using NHS esters of fatty acids, in order to establish a structure-function relation-
ship (Abbasi et al. 2008 ). Both lipid type and substitution degree was thought to
influence the transfection efficiency. PLL substituted with lipids that are longer
than caprylic acid were effective DNA carriers for skin fibroblasts. Higher substitu-
tion of lipids also offered more efficient PLL-based vectors. Myristic, palmitic,
and stearic acid conjugated PLLs were suggested to be the most effective vehicles
(Incani et al. 2009 ).
In addition to modifications using small molecules, polymers have been
employed for tailoring the characteristics of PLL vectors. Grafting PLL using
Pluronic was shown to give rise to polymers with enhanced efficiency and lower
cytotoxicity (Jeon et al. 2003 ). On the other hand, conjugating PLL with polyeth-
ylene glycol (PEG) led to diblock copolymers (PEG- b -PLLs) that can form water-
soluble polyion complex (PIC) assemblies with DNA. The introduction of PEG
endowed the resulting PIC micelles with great colloidal stability, and can protect
the payload DNA from DNase I attacking (Katayose and Kataoka 1997 ). Further
studies revealed that pDNA-loaded PIC micelles exhibited practical gene transfection
efficacy, particularly under serum-containing medium. In the presence of 100 mM
chloroquine, PIC micellar formulations showed a gene expression comparable to
the lipofection toward cultured 293 cells (Itaka et al. 2003 ).

3.1.3 Polyaspartamides and Their Derivatives

Through a facile and quantitative aminolysis of poly(b-benzyl L-aspartate) (PBLA)
in the presence of oligoamines, Kataoka's group have developed a new type of
polycations, i.e. polyaspartamides (Nakanishi et al. 2007 ). Among these polymers,
polyaspartamide derivative bearing 1,2-diaminoethane side chains [PAsp(DET)]
(Table 1 ) was demonstrated to be the most efficient one, with transfection activity
comparable or even higher than that of PEI25 (Masago et al. 2007 ). Importantly,
this polymer exhibited negligible cytotoxicity. Further mechanism exploration indi-
cated that the pH-selective membrane destabilization profile of PAsp(DET) mainly
contribute to the amplified transfection and minimized toxicity (Miyata et al.
2008b ). PEGylation of PAsp(DET) could endow the resultant polyplexes with
improved stability for systemic administration (Kanayama et al. 2006 ). Moreover,
the efficiency of PEGylated PAsp(DET) could be further optimized by introducing
lysine unit (Miyata et al. 2007, 2008a ) or incorporating disulfide linkage (Takae
et al. 2008 ). Through systemic administration, the lysine-optimized polycation
could achieve clear in vivo transfection of the EGFP gene in fibrotic pancreatic
adenocarcinoma of BALB/c nude mice (Miyata et al. 2008a ). The polyplex micelles
PEGylated PAsp(DET) with disulfide linkage showed both a 1~3 orders of magni-
tude higher gene transfection efficiency and a more rapid onset of gene expression
than those based on PEGylated PAsp(DET) without disulfide linkages, due to much
more effective endosomal escape based on the PEG detachment in endosome
(Takae et al. 2008 ).