Thermal-triggered loading and GSH-responsive releasing property of HBc particles for drug delivery

Hepatitis B core protein virus-like particles (HBc VLPs) have attracted wide attentions using as drug delivery vehicles, due to its excellent stability and easy in large scale production. Here in the present work, we report unique thermal-triggered loading and glutathione-responsive releasing property of the HBc particles for anticancer drug delivery. Through reversible temperature-dependent hole gating of the HBc particle capsid, about 4248 doxorubicin (DOX) were successfully encapsulated inside nanocage of a single nanoparticle at high HBc recovery of 83.2%, by simply incubating the DOX with HBc at 70 °C for 90 min. The new strategy was significantly superior to the disassembly-reassembly methods, which can only yield 3556 DOX loading at 52.3% HBc recovery. The thermal-sensitive drug entry channel in HBc was analyzed by molecular dynamic simulations, and the G113, G117 and R127 were identified as the key amino acid residues that are not conducive to the entrance of DOX but sensitive to temperature. Especially, the ΔGbind of R127 become even higher at high temperature, mutation of the R127 would be the first choice to make the drug entry thermodynamically easier. Due to plenty of disulfide bonds linking the HBc subunits, the HBc particles loaded with DOX exhibited intrinsic glutathione (GSH) responsivity for efficient controlled release in tumor sites. To further increase the tumor-targeting effect of the drug, Cyclo(Arg-Gly-Asp-d-Tyr-Lys) peptide was conjugated to the surface of HBc through a PEG linker. The prepared HBc-based anticancer drug showed significantly improved stability, tumor specificity, and in vivo anticancer activity on MCF7-bearing Balb/c-nu mice. Overall, our work demonstrated that the HBc VLPs can be an ideal drug carrier to fulfill requirement of the intelligent loading and "on demand" release of the therapeutic agents for efficient cancer therapy with minimal adverse effects.