## 4 Drug Delivery
### Polyplex Delivery  
Polymeric systems have provided a viable avenue for the delivery of genes and proteins as polyionic complexes, or polyplexes [148]. Owing to the negatively charged backbone of nucleic acids and some peptides, cationic polymer segments are often used as complexing agents for stabilization and drug delivery. As miktoarm polymers contain multiple branched segments, they can be tuned such that disparate arms can be used for complexation within the same drug delivery system. In such an example, self-assembled micelles from ABC (A = PEG, B = PLL, C = PCL) miktoarm polymers, with PEG and PLL comprising the hydrophilic corona, and PCL forming the core, were prepared [45]. Whereas PEG and PCL were used to maintain the integrity of micelles by acting as corona-stabilizing hydrophilic and core-stabilizing hydrophobic segments, respectively, PLL was used for pDNA complexation. Upon self-assembly, the 31 nm micelles were capable of loading the hydrophobic drug paclitaxel, while being complexed with pDNA, enabling their codelivery. _In vitro_ studies showed a burst release of paclitaxel initially, but reached 65% cumulative release within 60 h. Unfortunately, blank micelles led to significant HeLa cell death, in which 61% of cells survived 6 \(\upmu\)g/mL polymer concentrations. Paclitaxel loading led to a further 20% reduction in cell survival. It was found that PEG-PLL-PCL miktoarm stars formed polyplexes with pDNA through electrostatic interactions at N/P (polymer amino (N)/nucleic acid phosphate (P)) ratios higher than 2. Binding affinities were found to be similar for paclitaxel loaded and unloaded micelles. Zeta potentials were measured for N/P ratios 0.5-32, and were found to change to positive values at N/P = 2. However, these continued to steadily rise afterward indicating further structural change/binding. _In vitro_ cell transfection experiments on HeLa cells revealed improved transfection in paclitaxel loaded micelles, likely due to their anti-mitotic effects. The first example of the use of core-crosslinked miktoarm stars micelles for drug/nucleic acid delivery was for micelles that contained PEG and poly(\(\beta\)-hydroxyethylenediamine-L-glutamate) (PHLG) arms in their coronae [61]. Core-crosslinked polymers prepared by radical polymerization are often symmetrical [149]. The inclusion of mixed arms, as in miktoarm stars, can yield more efficient drug delivery systems, as in this case, where PEG confers solubility and PHLG was used for polyplex formation. The pKa of these miktoarm stars was determined to be 8.6, thus leading to protonation of PHLG terminal amino groups at physiological pH. Due to its cationic arms, the miktoarm polymer formed a complex with negatively charged siRNA. Gel electrophoresis showed that complexes were formed at N/P ratios higher than 16, at which point, bands representing unbound siRNA were no longer seen. Flow cytometry of Cy3-labeled siRNA and Alq\({}_{3}\)-labeled miktoarm star cores showed that most siRNA was taken up into A549 cancer cells as a complex with miktoarm stars. Linear (AB) and AB\({}_{3}\) miktoarm stars (A = PEG, B = PGA) were synthesized based on the NCA ROP of PGA from a PEG macroinitiator, in order to study the effect of polymer topology on lysosome protein complex [67]. Having an isoelectric point at pH 11.3, lysozyme has a net positive charge at physiological pH, which allows for its complexation with polyanions, such as PGA, which is negatively charged at physiological pH. No protein complexation was seen at pH 2, where PGA miktoarm blocks contained completely protonated carboxylic acid groups. Complexation at pH 7.4-depended heavily on polymer topology, as well as the polymer/protein ratio. At ratios of \(>\)1, complete complexation was seen for the linear AB polymers, whereas a ratio higher than 2.5 was needed for AB\({}_{3}\) miktoarm stars, with the same hydrophilic/hydrophobic ratio.