Targeted Drug Delivery

Advances in the development of novel therapeutic molecules have exceeded advances in the development of delivery technologies to enable the therapeutic use of those molecules. An excellent case in point may be found in antisense oligonucleotides. These compounds, active only in the nucleus, have enormous potential; however their very poor trafficking into the nuclear compartment has rendered them of very little value. Other examples could be found, for example in plasmid DNA, transcription factors and antibodies against intracellular targets. In this sense, targeting of therapeutics to particular subcellular locations presents an important academic challenge. Even the challenge of delivery of macromolecular drugs to specific locations in the body, such as cancer metastases or sites of inflammation, represents an unmet and substantial challenge.
Self assembly of amphiphilic components has been explored for many years in drug delivery, the most notable example being development of liposome carriers. This challenge has been rather frustrating, in that the inherent instability of these systems has rendered them also of relatively little value. Rapid removal of the liposomes from the systemic circulation by the reticuloendothelial system of the liver has been addressed by PEGylation of the phospholipid components, however this has created a quandary: non-PEGylated lipid formulations are rapidly cleared from the circulation, and PEGylation to adequate extents dramatically decreases the stability of the supermolecular assembly.
We have sought to develop functional block copolymeric amphiphiles that are capable of assembly into lamellar phases and thus vesicles in water and that further demonstrate additional functionality useful in targeting. These vesicles are referred to as polymersomes, analogous to liposomes but formed from self-assembles of amphiphilic polymers. The structure PEG-polyproplyene sulfide-PEG offers unique features, in that oxidation of the thioether backbone under physiological conditions to the polysulfone provides for a transition from a lamellar-phase former to a soluble polymer, thus providing a defined route of elimination from the body. A number of polymer architectures can be formed, including those containing terminal functionalization with biological recognition moieties, e.g. to target the polymersomes to particular sites within the body.

The functionality of the PEG-PPS-PEG block copolymers may be exploited in targeting of particular anatomical and intracellular locations. The polymeric amphiphile may be formed into unilamellar polymersomes of diameter 100 - 200 nm, with wall thickness 5 - 10 nm. Bioactive compounds may be loaded within these polymersomes during fabrication, providing both entrapment and protection from biological destruction. Given that each polymeric amphiphile is PEGylated, very effective stealth properties should be obtainable, without sacrifice in stability. During the process of endosomal-lysosomal trafficking, the polymersomes degrade under the oxidative conditions found in the lysosome, releasing their contents; a destabilizing agent may also be contained within. Current work focuses on a preliminary reaction under the reductive conditions and moderately low pH found in the endosome, prior to lysosomal fusion, to provide for release and endosomal disruption at this early stage.

These examples demonstrate the level of conceptual design in these self-assembling systems for targeting drugs to specific intracellular and anatomical locations. If the cytoplasmic compartment can be reliably targeted, this opens the door to more effective use of antisense oligonucleotides, plasmid DNA in gene therapy, transcription factors, and antibodies directed against intracellular targets. Other examples of self-assembly are also being explored in current work. For example, in Nature, one finds protein structures that exploit sulfation, but not multi-sulfation; this is curious, since polysulfation in glycosaminoglycans is common. Why are these structures not employed in protein-protein recognition? To explore this, we have selected the example of peptide-analogs of heparin, a polysulfated glycosaminoglycan. Heparin binds to a number of growth factors and modulates their biological recognition. There is reason to expect that if one could develop a super-heparin, i.e. a heparin analog that binds to the protein very tightly, that it would serve as an inhibitor to receptor binding. Combinatorial libraries of sulfated amino acids, both naturally occurring and not, were formed alongside hydrophobic amino acid residues. These structures were demonstrated to bind to heparin-binding domains on proteins, using VEGF165 and antithrombin III as examples, and bind so strongly as to out-compete heparin by approximately 1000-fold. Thus, it appears that Nature did not employ multi-sulfated proteins in biological recognition because those recognitions are extremely strong, essentially irreversible without desulfation. As candidate therapeutics, this provides an interesting route toward drug discovery; current targets are peptide-analogs of heparin as anticoagulants and inhibitors of angiogenesis.