Synergistic Enhancement of Breast Cancer Cell Death Using Ultrasound-Microbubbles in Combination with Cisplatin

摘要

Cisplatin (CDDP), an anti-cancer agent, can effectively treat several cancerous tumourstumors such as testicular, bladder, and ovarian cancers. CDDP binds to specific DNA bases causing 1,2-intrastrand cross-links, single strand and double strand breaks inducing apoptosis. However, the effectiveness of CDDP is limited in tumourtumors such as breast cancer due to drug resistance. In this study, the application of ultrasound-microbubble (USMB) in improving the therapeutic effect of CDDP in breast cancer cell line is investigated. Human breast cancer (MDA-MB-231) cells in suspension (2×10~6 cells/mL concentration and 0.6 mL volume) were treated with CDDP (3 μM, 30 μM and 300 μM) and USMB at 0.5 MHz pulse centered frequency, 60 s insonation time, 16 μs pulse duration, 1 kHz pulse repetition frequency, and 1.7% v/v (volume concentration) of Definity microbubble agent. Following USMB treatment, cells were plated in 96-well plates for 24 and 48-hour incubation, after which cell viability was measured using MTT assay (VMTT). Cell viability decreased significantly with the combined treatment of CDDP and USMB compared to CDDP alone (p<0.001) after both 24 and 48-hour incubation. After 24-hour incubation, the V_(MMT) was 40±2%, 32±1% and 18±1% with the combined treatment compared to 96±3%, 81±3% and 63±3% with CDDP alone at 3 μM, 30 μM and 300 μM, respectively. The combined treatment was additive at both concentrations (3 μM, p=0.9957) and (30 μM, p=0.6018) and synergistic at the highest concentration (300 μM, p=0.0169), based on Bliss Independence model with a 95% confidence interval of p<0.05. Furthermore, after 48-hour incubation, the V_(MTT) was 54±3%, 22±1% and 13±1% with the combined treatment compared to 94±9%, 68±3% and 44±2% with CDDP alone at 3 μM, 30 μM and 300 μM, respectively. The combined treatment was still additive at the lowest concentration (3 μM, p=0.6689) and synergistic at the higher concentrations (30 μM, p=0.0001) and (300 μM, p=0.0001) based on Bliss Independence model with a 95% confidence interval of p<0.05 after 48-hour incubation. The combination of ultrasound-microbubble and cisplatin synergistically enhances chemotherapeutic effectiveness in breast cancer cells. However, this enhanced effectiveness, in breast cancer cells (MDA-MB-231), is dependent on incubation time and cisplatin (CDDP) concentration.