Prostate cancer is associated with high mortality and new therapeutic
strategies are necessary for improved patient outcome. The utilisation
of potent, sequence-specific small interfering RNA (siRNA) to facilitate
down-regulation of complementary mRNA sequences in vitro and in vivo
has stimulated the development of siRNA-based cancer therapies. However,
the lack of an effective siRNA delivery system significantly retards
clinical application. Amphiphilic polycations with 'stealth' capacity
have previously been synthesised by PEGylation of poly-l-lysine-cholic
acid (PLL-CA). The benzoic imine linker between PEG and PLL-CA was
designed to be stable at physiological pH but cleavable at lower pHs,
consistent with the extracellular environment of tumours and the
interior of endosomes/lysosomes. The selective hydrolysis of the PEG
linker at these targeted sites should provide enhanced cellular uptake
and endosomal escape while simultaneously ensuring prolonged blood
circulation times. In this study, physicochemical profiling demonstrated
nano-complex formation between the PLL derivatives and siRNA (200-280nm
in diameter). At physiological pH only a slight cationic surface charge
(<20mV) was detected, due to the masking effect of the PEG. In
contrast, significantly higher positive charges (∼20 to 30mV and
>40mV) were detected upon hydrolysis of the PEG linker at acidic pHs
(pH=6.8 and 5.5, respectively). The PEGylated complexes were stable in
serum without significant aggregation or decomplexation of siRNA for up
to 48h. At the cellular level, PEG-PLLs were comparable with the
commercial carrier INTERFRin™, in terms of cellular uptake, endosomal
escape and in vitro reporter gene knockdown. In vivo, utilising a mouse
model grafted with prostate carcinoma, significant tumour suppression
was achieved using PEGylated complexes without marked toxicity or
undesirable immunological response, this was accompanied by a
simultaneous reduction in target mRNA levels. In summary, the advantages
of these vectors include: the in vitro and in vivo silencing
efficiency, and the low toxicity and immunogenicity.