ENGINEERING OF CELL-PENETRATING PEPTIDE-CONJUGATED INTRACELLULAR DELIVERY SYSTEMS - Biologically-Responsive Hybrid Biomaterials - page 216

Another example is gene delivery into immunocompetent cells to modulate
immune response. Antigen presenting cells (APC) are among the most important
cells of the immune system since they link the innate and the adaptative immune
responses, directing the type of immune response to be elicited. However, APC
are very resistant to transfection. To increase the efficiency of APC transfection,
we have used liposome-based lipoplexes additionally modified with cell-
penetrating TATp for better intracellular delivery of a model plasmid encoding
for the enhanced-green fluorescent protein (pEGFP). pEGFP-bearing lipoplexes
made of a mixture of PC:Chol:DOTAP (60:30:10 molar ratio) with the addition
of 2% mol of PEG-PE conjugate (plain-L) or TATp-PEG-PE (TATp-L) were
shown to effectively protect the incorporated DNA from degradation. Uptake
assays of rhodamine-labeled lipoplexes and transfections with the EGFP reporter
gene were performed with APC derived from the mouse spleen. TATp-L-based
lipoplexes allowed for significantly enhanced both, the uptake and transfection in
APC [127].
We have recently reported a double -targeted delivery system
simultaneously capable of extracellular accumulation and intracellular
penetration for gene therapy for the treatment of myocardial ischemia. We have
used low cationic liposomes-plasmid DNA complexes (lipoplexes) modified
with TATp and/or with monoclonal anti-myosin monoclonal antibody 2G4 (mAb
2G4) specific toward cardiac myosin, for targeted gene delivery to ischemic
myocardium. In vitro transfection of both normoxic and hypoxic cardiomyocytes
was enhanced by the presence of TATp as determined by fluorescence
microscopy and ELISA. The in vitro transfection was further enhanced by the
additional modification with mAb 2G4 antibody in the case of hypoxic, but not
normoxic cardiomyocytes. However, we did not observe a synergism between
TATp and mAb 2G4 ligands under our experimental condition. In in vivo
experiments, we have clearly demonstrated an increased accumulation of mAb
2G4-modified TATp lipoplexes in the ischemic rat myocardium and significantly
enhanced transfection of cardiomyocytes in the ischemic zone. Thus, the genetic
transformation of normoxic and hypoxic cardiomyocytes can be enhanced by
using lipoplexes modified with TATp and/or mAb 2G4 [128].
Another delivery system reported in literature is the 'multifunctional
envelope-type nano device' (MEND) [129]. This method involves three steps: (i)
DNA condensation with a polycation such as poly- -lysine (PLL), (ii) lipid film
hydration for the electrostatic binding of the condensed DNA, and (iii) sonication
to package the condensed DNA with lipids. Further MEND was surface-
modified with octaarginine on the envelope as a device for membrane
penetration to enhance cellular uptake, showed a 1000-fold higher transfection