MSc Defence: ENHANCEMENT OF ULTRASOUND ACOUSTIC ACTIVITY IN PROSTATE CANCER CELL LINES USING UNTARGETED NANOBUBBLES

Student: Sana Suboh

Supervisors: Dr. Michael Kolios and Dr. Agata Exner

Abstract

Nanobubbles (NBs) are nanoscale ultrasound contrast agents with potential applications in molecular imaging and targeted drug delivery. This study investigated how cell type, prostate- specific membrane antigen (PSMA) expression, incubation temperature, and incubation duration influence the acoustic activity and persistence of cell-associated untargeted nanobubbles.

PSMA expression was assessed in LNCaP, PIP, and Flu prostate cancer cell populations using immunofluorescence microscopy and flow cytometry. LNCaP showed high PSMA expression, unsorted PIP contained a heterogeneous PSMA-positive subpopulation, and Flu showed minimal detectable PSMA expression. For ultrasound experiments, LNCaP, PIP +Ve, PIP −Ve, PC3, and Flu cells were incubated with untargeted nanobubbles at a ratio of 20,000 NBs per cell for 30 or 60 min at either 4 °C or 37 °C. After removal of unbound nanobubbles, cell suspensions were imaged at room temperature using a Vevo 2100 ultrasound system with an MS250 transducer in a 1.5% agarose phantom. Nanobubble longevity was evaluated at post-incubation times of 0, 15, 30, and 60 min.

Acoustic activity was numerically higher after incubation at 4 °C than at 37 °C for all cell lines at both incubation times; however, the temperature difference reached statistical significance only for LNCaP after 30 min and for LNCaP, PIP +Ve, and Flu after 60 min. At 37 °C, LNCaP exhibited the highest acoustic activity, followed by PIP +Ve, whereas PIP −Ve, PC3, and Flu showed lower responses. Following a 60-min incubation and washing, measurable acoustic activity persisted over the post-incubation period but generally declined with time. These results are consistent with, but do not directly establish, an influence of nanobubble localization and cellular uptake on retained acoustic activity.

These findings demonstrate that nanobubble acoustic activity depends on incubation conditions and cell-line-specific interactions. Because untargeted nanobubbles were used, the observed differences likely reflect variations in nonspecific membrane association, cellular uptake, intracellular trafficking, retention, and nanobubble stability rather than PSMA expression alone.