Reversible Cross-Linked Mixed Micelles for pH Triggered Swelling and Redox Triggered Degradation for Enhanced and Controlled Drug Release

Abstract

Good stability and controlled drug release are important properties of polymeric micelles for drug delivery. A good candidate for drug delivery must have outstanding stability in a normal physiological environment, followed with low drug leakage and side effects. Moreover, the chemotherapeutic drug in the micellar core should also be quickly and "on-demand" released in the intracellular microenvironment at the tumor site, which is in favor of overcoming multidrug resistance (MDR) effects of tumor cells. In this work, a mixed micelle was prepared by the simple mix of two amphiphilic copolymers, namely PCL-SS-P(PEGMA-co-MAEBA) and PCL-SS-PDMAEMA, in aqueous solution. In the mixed micelle's core-shell structure, PCL blocks were used as the hydrophobic core, while the micellar hydrophilic shell consisted of two blocks, namely P(PEGMA-co-MAEBA) and PDMAEMA. In the micellar shell, PEGMA provided hydrophilicity and stability, while MAEBA introduced the aldehyde sites for reversible crosslinking. Meanwhile, the PDMAEMA blocks were also introduced in the micellar shell for pH-responding protonation and swelling of the micelle. The disulfide bonds between the hydrophobic core and hydrophilic shell had redox sensitive properties. Reversible cross-linked micelles (RCLMs) were obtained by crosslinking the micellar shell with an imine structure. RCLMs showed good stability and excellent ability against extensive dilution by aqueous solution. In addition, the stability in different conditions with various pH values and glutathione (GSH) concentrations was studied. Then, the anticancer drug doxorubicin (DOX) was selected as the model drug to evaluate drug entrapment and release capacity of mixed micelles. The in vitro release profiles indicated that this RCLM had controlled drug release. In the simulated normal physiological environment (pH 7.4), the drug release of the RCLMs was restrained obviously, and the cumulative drug release content was only 25.7 during 72 h. When it came to acidic conditions (pH 5.0), de-crosslinking of the micelles occurred, as well as protonation of PDMAEMA blocks and micellar swelling at the same time, which enhanced the drug release to a large extent (81.4%, 72 h). Moreover, the drug release content was promoted further in the presence of the reductant GSH. In the condition of pH 5.0 with 10 mM GSH, disulfide bonds broke-up between the micelle core and shell, followed by shedding of the shell from the inner core. Then, the micellar disassembly (degradation) happened based on the de-crosslinking and swelling, and the drug release was as high as 95.3%. The MTT assay indicated that the CLSMs showed low cytotoxicity and good biocompatibility against the HepG2 cells. In contrast, the DOX-loaded CLSMs could efficiently restrain the proliferation of tumor cells, and the cell viability after 48 h incubation was just 13.2%, which was close to that of free DOX. This reversible cross-linked mixed micelle with pH/redox responsive behaviors is a potential nanocarrier for chemotherapy.

Keywords: drug delivery; mixed micelles; reversible cross-link; stimulus-responsive.

Scheme 1

Schematic illustration for reversible cross-linked mixed micelles and the pH/redox-triggered controlled drug release.

Figure 1

The synthetic route of the copolymer PCL-SS-PDMAEMA.

Figure 2

¹H NMR spectra of PCL-SS-PDMAEMA.

Figure 3

GPC (Gel Permeation Chromatography) traces of PCL-SS-P(OEGMA-co-MAEBA) and PCL-SS-PDMAEMA.

Figure 4

FT–IR spectra of PCL-SS-iBuBr (A) and PCL-SS-PDMAEMA (B).

Figure 5

Graphs of intensity ratios (I_338.6/I_335.6) as functions of logarithm of copolymer mixture concentrations in aqueous solutions with different copolymeric mass ratios (PCL-SS-PDMAEMA to PCL-SS-P(PEGMA-co-MAEBA)) (The unit of C in the figures X-scale is mg/mL, and LgC is the corresponding logarithm of concentration C).

Figure 6

DLS results and TEM image of the cross-linked mixed micelles with DOX loading.

Figure 7

Size distribution of non-crosslinked micelles (NCMs) and reversible crosslinked micelles (SCMs) based on copolymer mixture before and after the 1000-fold water dilution.

Figure 8

Molecular structures and coarse-grained models of the polymers PCL-SS-PDMAEMA and PCL-SS-P(PEGMA-co-MAEBA).

Figure 9

The micelles self-assembled from different copolymers and their mixtures, PCL-SS-PDMAEMA micelle, PCL-SS-P(PEGMA-co-MAEBA) micelles and the mixed micelles.

Figure 10

In vitro drug release profiles of PCL-SS-PDMAEMA micelles (A), PCL-SS-P(PEGMA-co-MAEBA) cross-linked micelles (B) and reversible cross-linked mixed micelles (C).

Figure 11

The in vitro cytotoxicity of blank micelles against HepG2 cells for 48 h.

Figure 12

The in vitro cytotoxicity of free DOX and DOX-loaded mixed micelles against HepG2 cells for 48 h.