Ultrasound enhanced siRNA delivery using cationic liposome-microbubble complexes for the treatment of squamous cell carcinoma.

Microbubble () contrast agents combined with ultrasound targeted microbubble cavitation () are a promising platform for site-specific therapeutic oligonucleotide delivery. We investigated UTMC-mediated delivery of siRNA directed against epidermal growth factor receptor (), to squamous cell carcinoma () via a novel MB-liposome complex (). were constructed by conjugation of cationic liposomes to the surface of CF gas-filled lipid MBs using biotin/avidin chemistry, then loaded with siRNA via electrostatic interaction. Luciferase-expressing SCC-VII cells () were cultured in Petri dishes. The Petri dishes were filled with media in which LPXs loaded with siRNA against firefly luciferase () were suspended. Ultrasound () (1 MHz, 100-µs pulse, 10% duty cycle) was delivered to the dishes for 10 sec at varying acoustic pressures and luciferase assay was performed 24 hr later. siRNA delivery was studied in SCC-VII tumor-bearing mice intravenously infused with a 0.5 mL saline suspension of EGFR siRNA LPX (7×10 LPX, 30 µg siRNA) for 20 min during concurrent US (1 MHz, 0.5 MPa spatial peak temporal peak negative pressure, five 100-µs pulses every 1 ms; each pulse train repeated every 2 sec to allow reperfusion of LPX into the tumor). Mice were sacrificed 2 days post treatment and tumor EGFR expression was measured (Western blot). Other mice (=23) received either EGFR siRNA-loaded LPX + UTMC or negative control () siRNA-loaded LPX + UTMC on days 0 and 3, or no treatment ("sham"). Tumor volume was serially measured by high-resolution 3D US imaging. Luc siRNA LPX + UTMC caused significant luciferase knockdown vs. no treatment control, <0.05) in SCC-VII-Luc cells at acoustic pressures 0.25 MPa to 0.9 MPa, while no significant silencing effect was seen at lower pressure (0.125 MPa). EGFR siRNA LPX + UTMC reduced tumor EGFR expression by ~30% and significantly inhibited tumor growth by day 9 (40% decrease in tumor volume vs. NC siRNA LPX + UTMC, <0.05). Luc siRNA LPXs + UTMC achieved functional delivery of Luc siRNA to SCC-VII-Luc cells . EGFR siRNA LPX + UTMC inhibited tumor growth and suppressed EGFR expression , suggesting that this platform holds promise for non-invasive, image-guided targeted delivery of therapeutic siRNA for cancer treatment.