In solid tumors, the limited diffusion of therapeutic molecules in the perivascular space is a known limitation impacting treatment efficacy. Ultrasound Targeted Microbubble Cavitation (UTMC) has been shown to increase vascular permeability and improve the delivery of therapeutic compounds including small molecules, antibodies (mAb), nanoparticles and even cells, notably across the blood-brain-barrier (BBB). In this study, we hypothesized that UTMC could improve the accumulation and biodistribution of mAb targeting the adenosinergic pathway (i.e. CD73) in mice bearing bilateral subcutaneous 4T1 mammary carcinoma. METHODS: A bolus of fluorescently labeled mAb was given intravenously, followed by a slow infusion of microbubbles. UTMC therapy (1 MHz, 850 kPa) was given under ultrasound image guidance for 5 minutes to the right side tumor only, using three different pulse lengths with identical ultrasound energy (5000cyc "long", 125x40cyc "mid" and 500x10cyc "short"), and leaving the left tumor as a paired control. Longitudinal accumulation at 0 h, 4 h and 24 h was measured using whole-body biofluorescence and confocal microscopy. RESULTS: Our data support an increase in antibody accumulation and extravasation (# extravasated vessels and extravasated signal intensity) at 0 h for all pulses and at 4 h for the mid and short pulses when compared to the control non treated side. However, this difference was not found at 24 h post UTMC, indicative of the transient nature of UTMC. Interestingly, confocal data supported that the highest extravasation range was obtained at 0 h with the long pulse and that the short pulse caused no increase in the extravasation range. Overall, the mid pulse was the only pulse to increase all our metrics (biofluorescence, fraction of extravasated vessels, amount of extravasated Ab, and extravasation range) at 0 h and 4 h time points. CONCLUSIONS: Our results support that UTMC can enhance antibody accumulation in solid tumors at the macroscopic and microscopic levels. This preferential accumulation was evident at early time points (0 h and 4 h) but had started to fade by 24 h, a time dependence that is consistent with the ultrasound blood brain barrier opening literature. Further development and optimization of this theranostic platform, such as repeated UTMC, could help improve antibody based therapies against solid cancer.

The lytic release of ATP due to cell and tissue injury constitutes an important source of extracellular nucleotides and may have physiological and pathophysiological roles by triggering purinergic signalling pathways. In the lungs, extracellular ATP can have protective effects by stimulating surfactant and mucus secretion. However, excessive extracellular ATP levels, such as observed in ventilator-induced lung injury, act as a danger-associated signal that activates NLRP3 inflammasome contributing to lung damage. Here, we discuss examples of lytic release that we have identified in our studies using real-time luciferin-luciferase luminescence imaging of extracellular ATP. In alveolar A549 cells, hypotonic shock-induced ATP release shows rapid lytic and slow-rising non-lytic components. Lytic release originates from the lysis of single fragile cells that could be seen as distinct spikes of ATP-dependent luminescence, but under physiological conditions, its contribution is minimal <1% of total release. By contrast, ATP release from red blood cells results primarily from hemolysis, a physiological mechanism contributing to the regulation of local blood flow in response to tissue hypoxia, mechanical stimulation and temperature changes. Lytic release of cellular ATP may have therapeutic applications, as exemplified by the use of ultrasound and microbubble-stimulated release for enhancing cancer immunotherapy in vivo.

In solid tumors, hypoxia is a well-known resistance mechanism to radiation therapy. It was previously shown that microbubbles (MBs), when exposed to an ultrasound pulse (US) can cause vasodilation in muscle tissue. Conceptually, the therapeutic pulse can be localized on the tumor by steering the US beam. This approach is therefore proposed as a targeted image-guided provascular therapy in tumors to reduce hypoxia before radiotherapy. However, the effects of US and MB conditions on the relative increase in tumor perfusion remain largely unknown.I would like to thank Dr. Francois Yu for teaching, guiding, and helping me grow as a person and a scientist. Without his work and constructive feedback, this project would not have been possible. I would like to thank:Ju Jing Tan and the rest of the 8 th -floor team for the help and knowledge provided for imaging ATP.Rodin and Amelie for the help with cell culture, chip manufacturing, and overall support for my project when creating, designing, and manufacturing the microfluidic chip. Boris, Dr. Loise Allard, Dr. Guy Cloutier, and the rest of the LBUM team for the constant support and feedback during the weekly presentations.The members of MTL YU Lab for your contributions feedback and overall support during these last few years.Nick, Cory, and all the rest of my friends for the encouragement and help all along.Dani, for reading, listening and supporting me throughout the project.Finally, my parents, my sister, and the rest of my family for their unconditional love and support.