In vitro potentiation of doxorubicin by unseeded controlled non-inertial ultrasound cavitation

Ultrasound-generated non-inertial cavitation has the ability to potentiate the therapeutic effects of cytotoxic drugs. We report a novel strategy to induce and regulate unseeded (without nucleation agents) non-inertial cavitation, where cavitation is initiated, monitored and regulated using a confocal ultrasound setup controlled by an instrumentation platform and a PC programmed feedback control loop. We demonstrate, using 4T1 murine mammary carcinoma as model cell line, that unseeded non-inertial cavitation potentiates the cytotoxicity of doxorubicin, one of the most potent drugs used in the treatment of solid tumors including breast cancer. Combined treatment with doxorubicin and unseeded non-inertial cavitation significantly reduced cell viability and proliferation at 72 h. A mechanistic study of the potential mechanisms of action of the combined treatment identified the presence of cavitation as required to enhance doxorubicin efficacy, but ruled out the influence of changes in doxorubicin uptake, temperature increase, hydroxyl radical production and nuclear membrane modifications on the treatment outcome. The developed strategy for the reproducible generation and maintenance of unseeded cavitation makes it an attractive method as potential preclinical and clinical treatment modality to locally potentiate doxorubicin.