Abstract
Tumors normally possess irregular vasculature, and tend to outstrip blood supplement and become hypoxia or ischemia, which results in the resistances of the tumor to chemotherapy treatment. New therapies are required to target hypoxic or ischemic areas in tumors. Recently, many studies have shown that monocytes and derived macrophages can be engineered to actively migrate toward tumors and infiltrate avascular and hypoxic areas. The properties of macrophages prompted us to propose the drug or genes delivery by macrophages to otherwise inaccessible areas within tumors. The aim of this study is to investigate the feasibility of using macrophages to infiltrate hypoxic or ischemic areas in tumors, as the carriers of drug-loaded phase-change droplets. We develope a drug-loaded droplet formulation (DOX-Droplet) with a high loading capacity of doxorubicin (DOX) drug, which was complexed to the lipid shell by both hydrophobic and electrostatic interactions. Ultrasound induced the transition from liquid droplets to gas bubbles was referred as the effect of acoustic droplet vaporization (ADV), and ADV can trigger encapsulated drug release from the droplet-loaded macrophages. DOX-Droplets were fabricated via the thin-film hydration method. The DOX drug encapsulation efficiency was 76.55 ± 5.97% estimated by a Fluorescence spectrophotometer. RAW 264.7 cells (mouse leukaemic monocyte macrophage cell line) were used to ingest the drug-loaded phase-change droplets. For evaluating DOX-Droplets uptake efficiency, cell viability and migration mobility of DOX-Droplet loaded macrophages, flow cytometric analysis, alarmarBlueTM assay, and transmembrane cell migration assay were measured, respectively. In vitro ultrasound triggering DOX release, a 3.5-MHz high-intensity focused ultrasound transducer was used. The DOX-Droplets loaded RAW 264.7 cells were insonated by a three-cycle, 200 Hz PRF and 12-MPa HIFU pulses for 10 min. After exposed to ultrasound the supernatant was collected and incubated with Tramp-C1 cells (mouse prostate cancer cell line) for 48 hr. The cytotoxicity of DOX released from DOX-Droplets loaded RAW 264.7 cells was evaluated by Tramp-C1 cell viability. The results demonstrate the feasibility of using macrophages to deliver DOX-loaded phase-change droplets. Cell viability assays reveals that the loading of droplets did not affect cell viability for 24 hr. But transmembrane cell migration assay reveals that the loading of droplets hampered the migration mobility of RAW 264.7 cells. In addition, Tramp-C1 cell viability analysis performed that ultrasound can trigger DOX release from DOX-droplet loaded RAW 264.7 cells effectively and the release DOX killed the Tramp-C1 cell. Future works include the drug payload and retention improvement of DOX-Droplet, the assessments of mechanical effect of droplet vaporization on therapy and the liberated drug payload via ultrasound-triggered vaporization in vivo.