Purpose: Targeted delivery and accumulation of chemotherapeutics to tumor sites is in high demand and extremely challenging for anticancer therapy. Studies show that albumin is actively recruited into tumor tissue overexpressing albumin-binding proteins including secreted protein acidic and rich in cysteine (SPARC) and gp60. An albumin binding domain variant ABD035 was reported to have super strong affinity to serum albumin and the binding won't affect the in vivo process of endogenous albumin. Herein, we aimed to develop a novel ABD035-modified redox-responsive micelle loaded with paclitaxel (PTX) to enhance targeting ability and increase drug accumulation in tumor tissue and eventually achieve better anti-tumor effect.
Methods: Firstly, ABD035-PEG-SS-PTX4 was synthesized and ABD035-PEG-SS-PTX4 micelle was prepared through dialysis method. Then size distribution of the micelle was measured by dynamic light scattering (DLS) and the morphology was characterized by transmission electron microscope (TEM). The redox-responsive drug release of the ABD035-PEG-SS-PTX4 micelle was performed in PBS 7.4 containing different concentrations of glutathione (GSH). Binding kinetics of ABD035 peptide and ABD035-PEG-SS-PTX4 micelle to HSA was performed by surface plasmon resonance (SPR). BODIPY-labeled micelle was used for fluorescent studies. In vivo imaging was employed to investigate the targeting ability of the micelle. Immunofluorescence staining and confocal microscopy were used to observe the in vivo co-localization of the micelle and SPARC. Biodistribution study was conducted on MDA-MB-231-bearing nude mice and drug concentration in various tissues was detected using LC/MS/MS. Cellular viability was measured by MTT assay and cellular uptake were performed on MDA-MB-231 cells and photographed by confocal laser scanning microscope. In vivo antitumor effect and TUNEL assay were conducted on MDA-MB-231-bearing nude mice.
Results: The average size of PEG-SS-PTX4 and ABD035-PEG-SS-PTX4 micelles measured by DLS was 32 1 nm (PDI=0.136 0.036) and 29 2 nm (PDI=0.189 0.019), respectively. The release profile showed that prototype PTX was rapidly released from the micelle under 10 mM GSH (intracellular concentration) in PBS 7.4 while less than 6% of the total drug was released within 24 h under 0 or 2 μM GSH (extracellular concentration) in PBS 7.4. TEM images showed that the micelles were uniform and near spherical while the structure could be disintegrated under 10 mM GSH (Fig. 1). The binding kinetics of ABD035 peptide and ABD035-PEG-SS-PTX4 to HSA was similar and the ABD035-modified micelle has a strong binding response to albumin (KD=1.996×10-10) (Fig. 2).
The in vivo fluorescent images demonstrated that ABD035-modified micelles have enhanced tumor-targeting ability and co-localization of BODIPY-labeled ABD035-modified micelle and SPARC was observed in immunofluorescent staining images of ex vivo tumor slices. Bioditribution results further showed that ABD035-modified micelle could significantly increase drug accumulation in tumor tissues through binding with albumin and gp60-SPARC-mediated pathway (Fig. 3). Furthermore, the ABD035-modified micelle showed increased cellular uptake and significantly improved antitumor effect both in vitro and in vivo (Fig. 3).
Conclusion: The ABD035-modified micelle significantly improves the therapeutic effect in the animal model bearing triple negative breast cancer by increasing drug accumulation in tumor tissue, enhancing cell uptake efficiency, rapidly releasing prototype drug under intracellular reductive conditions and increasing tumor cell apoptosis. These results together vote the biomimetic delivery of ABD035-modified micelle as a promising strategy for cancer therapy.