Uptake and cellular recovery mechanisms in microbubble-enhanced ultrasound delivery of nanoparticles for cancer therapy

摘要

Functionalised nanoparticles (NPs) have been shown to enable delivery of small silencing RNA molecules (siRNA) to cancer cells, disabling proliferation and providing a route to therapy. However the approach can be restricted to specific cell lines, limiting translation potential to in vivo treatment. Here, we investigate delivery of NPs to human breast cancer cell lines (MCF7), previously proven problematic in this regard, using ultrasound (US) and microbubbles (MB). In particular we examine potential mechanisms of NP uptake into cells, opportunities for optimisation and offer insight to cellular recovery post treatment. Treatment did not affect cell viability. TEM and EDX confirmed uptake of NPs into MCF7 cells via two potential routes; (i) at low MI vesicular confinement (Fig 1a) of NPs dominated, indicative of endocytosis, (ii) at higher MI both confined and free NPs were observed in the cytoplasm (Fig 1b) suggesting pore entry. In addition the presence of larger NP clusters at high MI suggests pinocytosis. Membrane pores observed via SEM support our hypothesis. Again two populations existed, large (approx. 1-2 μm) and more prevalent small pores (approx. 20-100 nm). All small pores appeared close to membrane `blebs' indicating that the recently reported sonoporation recovery mechanism may also underpin cellular preservation following minor insult at lower MI. No relationship between delivery route and frequency was observed. Since the therapeutic effect of NPs is strongly influenced by their delivery, MI could be a critical efficiency parameter. Work continues to determine influence of delivery route on breast cancer treatment.