Epigenetic therapy using targeted microbubbles as a potential treatment for breast cancer

Therapeutic efficacy of anti-cancer agents is a critical requirement however it is often limited due to poor delivery of the drugs to the tumour. The aim of this project was to use gas-filled, lipid-shelled microbubbles (MBs) to enhance therapeutic effects of an epigenetic drug, called decitabine (5-aza-2'-deoxycytidine or DAC), against triple-negative breast cancer (TNBC). MB-assisted drug delivery using ultrasound is proposed as a non-invasive approach for controllably triggered drug release, which may protect the drug from rapid degradation, enhance drug efficacy and reduce off-site side effects. In this study, treatment of human MDA-MB-231 breast cancer cells with low doses of DAC demonstrated up-regulation of epigenetically dysregulated tumour suppressor genes and immune response genes, as well as global DNA demethylation. These assays, alongside inhibition of tumour growth, were used as biomarkers for the assessment of DAC delivery in a mouse MDA-MB-231 TNBC xenograft model. To increase the accumulation of therapeutic MBs to the tumour site, vascular endothelial growth factor receptor 2 (VEGFR2) was evaluated as a suitable molecule for DAC delivery. Smaller tumours showed higher VEGFR2 expression in vivo, suggesting that VEGFR2-targeted MB drug delivery would be most effective during that stage of tumour growth. A flow assay verified that VEGFR2-targeted MBs were bound specifically to VEGFR2-expressing mouse endothelial SVR cells in vitro, indicating that enhanced binding to the mouse tumour vasculature is possible. Results from in vitro studies revealed that when DAC was delivered in combination with ultrasound destruction of MBs or encapsulated in liposomes, epigenetic effects were detected, suggesting that drug delivery was successful. The main in vivo experiment demonstrated that administration of DAC combined with VEGFR2- targeted MBs and ultrasound showed activation of tumour suppressor genes previously silenced by DNA methylation, as well as inhibition of tumour growth. These results suggest that delivery of DAC in this way could improve drug therapeutic efficacy, prime tumours for secondary anti-cancer agents and potentially reduce the required dose resulting in decreased off-site toxicities.