Integrated development of a production procedure and characterization methodology of α-Bi2O3, BiONO3, Bi5O7NO3 ultra-fine particles as complementary contrasting agents for micro-bubbles in tumor imaging

Nowadays, molecular imaging is one of the most powerful technique in the monitoring of pathological processes of tumors, performed in a dynamically, real time, non-invasive level. To extend the diagnostic capability and imaging resolution, Ultrasounds Contrasting Agents (UCAs) have been largely used in clinical trials being promising for hospital applications. Despite their large potential, UCAs present limitation due their size range firstly, the impossibility to pass through vessel walls into the tumor tissue. This limitation can be overcome using nanosized UCAs because vascular endothelial gap in tumors is approximately 380–780 nm. Among nano-sized UCAs, many studies have been recently focused on preparation and activation of nanobubbles (NBs), composed of phospholipid (DPPE-PEG, DPPA, PEG-DPSE, etc…) shell and PFB gas core that are able to show optimal contrast enhancement abilities and some thermal therapeutic effect due to their ultrasound activated cavitation. Recently, research has been focused on the use of innovative nano-sized (or nano-structured) as UCAs. These materials are well-noticed to have unique optical, magnetics and chemical properties that allow a better contrast enhancement, increased sensitivity, controlled biodistribution, better spatial and temporal information, multi-functionality and multi-modal imaging across MRI, Positron Emission Tomography (PET), and ultrasounds (US). Additionally, nanomaterials can be functionalized in numerous ways due to their highly engineerability to exhibit significant properties as increased contrast sensitivity, binding avidity and targeting specificity. The aim of the project, made in collaboration with Medical Imaging (M. I.) Laboratory of University of Toronto, is to investigate possible solutions for innovative nano-sized UCAs optimized by the use of Bi-based (α-Bi2O3, BiONO3, Bi5O7NO3) micro- and nano-particles (MPs and NPs respectively) integrated with ultrasound responsive NBs. Particles studied were produced following two opposite ways: a solid-state reaction and a glycol assisted synthesis. The work is divided into 3 main parts: firstly, particles’ size reduction procedure was deeply investigated to be, at the same time, able to effectively reduce particles’ diameter in the nano-scale range and to be compatible with MBs production procedure. To overcome this last limitation, reduction procedure was conducted in MilliQ® water and in cyclooctane emulsions and stabilized by addition of DPPE-PEG2K as steric hindrance. After, the procedure was implemented in the production of ultrafine particles charged acoustic responsive MBs. For both steps Coulter Multisizer 4E SOP Counter, FESEM (Field Emission Scanning Electron Microscope), UV-VIS spectrophotometry and optical microscopy analysis were performed. In the third part of the project samples of Bi-based ultrafine particles charged MBs were tested with an MTT assay to determine CT26 mouse colon carcinoma cells’ viability when exposed. In conclusion the defined protocol was able to effectively produce stable charged MBs and the presence of nano sized Bi-based particles seems to have visible effects on cells’ proliferation. In the future of this work there will be tests in vivo and in vitro (agar phantom) of particles’ contrasting abilities after MBs activation under HIFU (High Intensity Focused Ultrasounds). Issues and future applications for this innovative category of nano particles have been rapidly explained at the end.