The engineering of BYL719 solubility dmso plasmid DNA encoding the therapeutic transgene under the influence of brain cell-specific promoters eliminates ectopic transgene expression and enables transgene expression in targeted regions of the CNS [2, 19–23, 27, 28]. Figure 1 Engineering of Trojan horse liposomes (THL). (a) A supercoiled plasmid DNA is encapsulated in the interior of the THL. The Inhibitors,research,lifescience,medical plasmid encodes for a coding sequence (cds), the expression of which is under the

influence of a promoter (pro), that is, SV40, … Table 1 Targeting MAbs for THL and target tissue. 2. Trojan Horse Liposome (THL) Technology THLs are pegylated liposomes containing a supercoiled plasmid DNA molecule in the interior of the liposome (Figure 1(a)). THLs are engineered with a mixture of naturally occurring lipids

that has been optimized for the encapsulation of plasmid DNA [4, 19]. The liposomes are comprised of 93% 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphocholine (POPC), 3% didodecyldimethylammonium Inhibitors,research,lifescience,medical bromide (DDAB), 3% distearoylphosphatidylethanolamine (DSPE)-PEG2000, and 1% DSPE-PEG2000-maleimide. The maleimide functional group allows Inhibitors,research,lifescience,medical for covalent conjugation of a thiolated MAb via a stable thioether linkage (Figure 1(a) and Table 1). A panel of species-specific peptidomimetic MAbs has been developed (Table 1), and their efficacy in delivering THLs to brain has been demonstrated in experimental animal models in vivo [1, 3, 4, 19–23, 27]. The 83-14 murine MAb to the HIR and the OX26 murine MAb to the rat TfR are used to target human and rat tissues, respectively (Table 1). The OX26 TfRMAb is active only in rats, so the rat 8D3 MAb against the mouse TfR is used in mice (Table 1) [20, 21, 27–33]. The 83-14 HIRMAb does not cross-react with Inhibitors,research,lifescience,medical the insulin receptor in rodents or even New World primates such as the squirrel monkey. However, this HIRMAb does cross-react with the insulin receptor of Old World primates such as the Rhesus monkey. Since the plasmid DNA must Inhibitors,research,lifescience,medical be delivered

to the nuclear compartment of brain cells, the THL must traverse both the BBB and the brain cell plasma membrane (BCM) behind the BBB (Figure 1(c)). Owing to high expression of the TfR or IR on both the BBB and BCM barriers, the targeting MAb enables the sequential receptor-mediated transcytosis of the THL across the BBB followed by Rutecarpine the receptor-mediated endocytosis of the THL into the brain cell (Figure 1(c)). THLs have also been successfully constructed to target human tumor cells in a scid mouse model wherein dual targeting MAbs were directed to the mouse TfR and HIR (Table 1 and Figure 1(a), i.e., MAb1 and MAb2) [23]. 3. Brain Expression of Reporter Genes In vivo applications of THLs were initially investigated with luciferase and lacZ reporter genes in vivo [3, 4, 20, 21, 34]. THLs were constructed with the expression plasmid of interest (i.e.