PH Sensitive PEGylated Glucolipid Conjugate Micelles For Tumor Targeting Therapy

In recent years, environment-responsive conjugates have gained a growing interest in pharmaceutical research as a potential vehicle for antitumor drug delivery. Those smart drug delivery systems could mimic biological signals in a crude way, where an environmental stimulus results in a change in properties, following the improved antitumor efficiency and reduced side effects. According to the specificity of the tumor microenvironment, we design a pH sensitive PEGylated glucolipid conjugate for targeting delivery of the anticancer drug. The main objective of this study was aimed to obtain tumor microenvironment-responsive drug delivery system for efficient tumor therapy, with the characteristic of pH triggered PEG-cleavage resulting in the prolongation of circulation time and the increase of tumor cellular uptake.A glucolipid-like conjugate (stearic acid grafted chitosan, CS) was synthesized by the chemical reaction between chitosan (Mw:18.0 KDa) and stearic acid in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). The chemical structure of CS was confirmed by 1H-NMR spectrum. The degree of amino-substitution for CS was about 5.9%. CS could self-assemble to form micelles in aqueous medium, the critical micelle concentration (CMC) of which was 129.6μg/mL. The diameter and zeta potential of CS micelles with the concentration of 1 mg/mL were measured by Zetasizer analyzer, which were 27.6±1.8 nm and 39.7±1.6 mV, respectively.PH-sensitive PEGylated CS conjugates (PCCS) was synthesized by conjugating polyethylene glycol (PEG) to CS via a pH-responsive cis-aconityl linkage, using EDC and N-hydroxy-succinimide (NHS) as catalyst. At the same time, PEG was grafted to CS directly by the Schiff reaction to obtain pH-insensitive PEGylated CS conjugates (PCS), which was utilized for comparative study. The physicochemical characteristics of PCS and PCCS were determined. An increase in micellar size and CMC value as well as a decrease in zeta potential could be observed after PEGylation. Moreover, the PCCS micelles showed acid-triggered PEG cleavage manner. The cleavage rate of PEG form PCCS micelles increased significantly with the reductions of the pH for outer medium. For normal liver cells BRL-3A, the quantitative cellular percentage significantly reduced after PEG modification. As to liver tumor cells BEL-7402, a slightly increase in uptake percentage of PCCS was observed under weakly acid environment compared to normal physiological condition, indicating a potential advantage for PCCS to be drug delivery vehicle for antitumor therapy.Doxorubicin (DOX) was chosen as the model drug and loaded into the polymeric micelles by the ultrasound and dialysis method to produce DOX-loaded micelles (CS/DOX, PCS/DOX, PCCS/DOX). All the DOX-loaded micelles presented excellent DOX loading capacity with a drug encapsulation efficiency up to 86% when the DOX feeding amount was 5%. It was found a decrease in micellar size after DOX loading. After PEG modification, the in vitro release rate of DOX was demonstrated to be much slower. In particular, the drug release rate of PCCS/DOX was slightly accelerated by decreasing pH. In addition, in vitro cellular uptake and antitumor activitity tests showed that both the internalization ability and cytotoxicity of PCCS/DOX were enhanced as compared to pH-insensitive PCS/DOX.Taking PCS micelles as a comparative group, PCCS drug delivery system demonstrated to show a more accumulation in tumor tissue, followed by much more pronounced antitumor activity together with a security benefit in model mice.