Approaches to Achieving Sub-cellular Targeting of Bioactives Using Pharmaceutical Nanocarriers - Intracellular Delivery: Fundamentals and Applications - page 61

2.2 Transport Through the Cytoplasm

The cytoplasm is a highly viscous space (Seksek et al. 1997 ; Goins et al. 2008 ),
which has a high concentration of dissolved solutes (Ellis and Minton 2003 ; Minton
2006 ) and as such can present a significant barrier to the movement of many poten-
tial bioactive molecules especially those with high molecular weight. However, in
the eukaryotic cells, transport of biomolecules such as proteins and lipids is gov-
erned by the actin and microtubule cytoskeletons (Vale 1987 ). The extent to which
this transport system can be utilized for nanocarrier mediated intracellular transport
remains to be explored. Interestingly it has recently been reported that PEGylation
of nanoparticles improves their cytoplasmic transport presumably by allowing
nanoparticles to evade non specific interactions in the highly crowded environment
of the cytoplasm (Suh et al. 2007 ). It is also interesting to consider in this context
the fact that endosomes are routinely trafficked through the cytosol in their normal
progression to lysosomes. If the endosomal vesicle can somehow be rerouted to
afford association with other membrane bound organelles like mitochondria or the
nucleus, there may well be no need to have the biologically active molecule enter
the cytosol. If for example, the nanocarrier components were to undergo a redistri-
bution to become part of the endosome and the targeting ligand was able to redis-
tribute to the surface of the endosomal vesicle, it might be possible then that the
vesicle would have an altered sub-cellular fate that could involve transport to and
association with a target compartment other than the lysosome. While there is
emerging evidence to suggest that in fact cells actively traffic nanocarriers in cell
membrane-derived vesicles (Ruan et al. 2007 ), the concept remains speculative
until more is understood about the mechanisms of intracellular molecular versus
vesicular transport. For now, the trend towards mediating endosomal release of
internalized nanocarrier and associated bioactive is arguably based on insights
gleaned from intracellular dynamics of viral particles that are in essence naturally
occurring nanocarriers. Viral particles are endocytosed and then are able to mediate
endosomal escape and subsequent nucleus specific delivery of their DNA. Based on
the premise that to efficiently deliver DNA to the nucleus, a delivery system must
penetrate through the plasma membrane and the nuclear envelope, prior to DNA
release in the nucleus, a strategy that involved step-wise membrane fusion was
devised. Using a multi-layered nanoparticle called a Tetra-lamellar Multi-functional
Envelope-type Nano Device (T-MEND) consisting of a DNA-polycation condensed
core coated with two nuclear membrane-fusogenic inner envelopes and two endo-
some-fusogenic outer envelopes, which are shed in stepwise fashion transgene
expression in non-dividing cells was reported to be dramatically increased (Akita
et al. 2009 ). A similar approach in designing a mitochondria specific delivery sytem
has been reported as well. Liposomal carriers called MITO-Porters which carry
octaarginine surface modifications to stimulate their entry into cells as intact vesi-
cles (via macropinocytosis) were prepared with lipid compositions that were identi-
fied in various experiments to promote both fusion with the mitochondrial
membrane and the release of liposomal cargo to the intra-mitochondrial compartment