Multivalency of targeting ligands on anti-cancer drug nano-carriers

Abstract

Targeted drug delivery is being focused in cancer research due to its advantages of limiting the side effects during treatment. Multivalence of targeting ligands decorated on the surface of nano-carriers plays an important role in the efficiency of selective cellular uptake and anti-cancer activities. Among many kinds of nano-carriers, oil-in-water nanoemulsions stabilized by the self-assembly of amphiphilic copolymers are the ones of the best candidates by far for encapsulation of hydrophobic drugs which occupy the majority of cancer chemotherapy. The purpose of this thesis is to develop an advanced delivery system for hydrophobic anti-cancer drugs with high avidity of targeting ligands, incorporating with diagnostic hydrophobic oil which also act as delivery phase for hydrophobic drug.

To achieve the goal, we first performed the investigation on increasing the stability of oil-in-water nanoemulsions using the FDA-proof biocompatible amphiphilic polymers mPEG-b-PCL, by two means of modifications on hydrophobic block and on hydrophilic block. The modifications on hydrophobic block were the decrease of crystallinity by adding certain percentages of lactic units among Caprolactone units, in diblock copolymer mPEG-b-PCLL and in triblock copolymer PCLL-b-PEG-b-PCLL. The decreased crystallinity was expected to increase the wettability of hydrophobic block into the core oil, thus decrease the dynamic size of nano-droplets and increase the stability of nanoemulsions. The results showed that incorporation of lactide in hydrophobic blocks indeed increased the stability of nanoemulsions, especially in diblock copolymer mPEG-b-PCLL with the non-polar hydrophobic oil (liquid paraffin). However, highly reduced crystallinity in triblock copolymer PCLL-b-PEG-b-PCLL was unsuccessful in stabilization of nanoemulsions under harsh physical stress such as freeze-thaw cycles, suggested that a certain degree of crystallinity is critically important for the structure robustness of nanoemulsions.

The modification on hydrophilic block were the substitution of PEG block by the linear polyglycerol (PG) block with highly increased hydrophilicity and modification sites. With the chemical structure of backbone chain similar to PEG, PG were the excellent candidate for stabilization of the drug loaded nanoemulsions. Moreover, with the terminated adjacent hydroxyls (vicinal diols), PG allows the facile and stable formation of covalent bonding with boronic ester, thus open new method for conjugation with targeting ligands to a high concentration. By far the concentration of targeting ligands which connected to PEG chains on the surface of nano-carriers reach 12%. In this project, we achieved the nanoemulsions with 21% of targeting ligands on each nano-droplets. In this project, lipiodol, a diagnostic oil for enhance the contrast of computed tomography image, was used in the core as drug delivery phase. Drug loaded lipiodol nanoemulsions stabilized by PG-b-PCL had size between 150-175 nm, encapsulated 3 wt-% of paclitaxel, and was stable in solutions containing salt and high concentration of proteins for over a week. The nanoemulsions incorporated with targeting ligands on the surface show significant increase in cellular uptake in vitro and excellent anti-cancer activity in vitro with the toxic concentration LD50 at smaller than 3 ng/mL. In vivo study showed that our highly multivalent targeting system effectively inhibit the growth of tumor at a treatment dose 4 times lower than that of available commercial drug (5 mg/kg compared to 20 mg/kg of Taxol). In vivo bio-distribution study showed that the drug loaded nanoemulsions accumulated mainly in the tumor and had no toxic to other internal organs. This study suggested that PG is a very promising corona polymer for the effective stabilization and multivalent functionalization of nano-carriers for tumor-targeted delivery of therapeutic and imaging agents.