Intracellular Water Lifetime as a Tumor Biomarker to Monitor Doxorubicin Treatment via FFC-Relaxometry in a Breast Cancer Model

This study aims to explore whether the water exchange rate constants in tumor cells can act as a hallmark of pathology status and a reporter of therapeutic outcomes. It has been shown, using 4T1 cell cultures and murine allografts, that an early assessment of the therapeutic effect of doxorubicin can be detected through changes in the cellular water efflux rate constant k(io.) The latter has been estimated by analyzing the magnetization recovery curve in standard NMR T(1) measurements when there is a marked difference in the proton relaxation rate constants (R(1)) between the intra- and the extra-cellular compartments. In cellular studies, T(1) measurements were carried out on a relaxometer working at 0.5 T, and the required difference in R(1) between the two compartments was achieved via the addition of a paramagnetic agent into the extracellular compartment. For in-vivo experiments, the large difference in the R(1) values of the two-compartments was achieved when the T(1) measurements were carried out at low magnetic field strengths. This task was accomplished using a Fast Field Cycling (FFC) relaxometer that was properly modified to host a mouse in its probe head. The decrease in k(io) upon the administration of doxorubicin is the result of the decreased activity of Na(+)/K(+)-ATPase, as shown in an independent test on the cellular uptake of Rb ions. The results reported herein suggest that k(io) can be considered a non-invasive, early and predictive biomarker for the identification of responsive patients immediately from the first doxorubicin treatment

A Novel PSMA-Targeted Probe for NIRF-Guided Surgery and Photodynamic Therapy: Synthesis and Preclinical Validation

A total of 20% to 50% of prostate cancer (PCa) patients leave the surgery room with positive tumour margins. The intraoperative combination of fluorescence guided surgery (FGS) and photodynamic therapy (PDT) may be very helpful for improving tumour margin delineation and cancer therapy. PSMA is a transmembrane protein overexpressed in 90–100% of PCa cells. The goal of this work is the development of a PSMA-targeted Near InfraRed Fluorescent probe to offer the surgeon a valuable intraoperative tool for allowing a complete tumour removal, implemented with the possibility of using PDT to kill the eventual not resected cancer cells. PSMA-617 binding motif was conjugated to IRDye700DX-NHS and the conjugation did not affect the photophysical characteristics of the fluorophore. The affinity of IRDye700DX-PSMA-617 towards PCa cells followed the order of their PSMA expression, i.e., PC3-PIP > LNCaP > PC3, PC3-FLU. NIRF imaging showed a significant PC3-PIP tumour uptake after the injection of 1 or 5 nmol with a maximum tumour-to-muscle ratio (ca. 60) observed for both doses 24 h post-injection. Importantly, urine, healthy prostate, and the bladder were not fluorescent at 24 h post-injection. Flow cytometry and confocal images highlighted a co-localization of PSMA+ cells with IRDye700DX-PSMA uptake. Very interestingly, ex vivo analysis on a tumour specimen highlighted a significant PSMA expression by tumour-associated macrophages, likely attributable to extracellular vesicles secreted by the PSMA(+) tumour cells. FGS proved that IRDye700DX-PSMA was able to easily delineate tumour margins. PDT experiments showed a concentration-dependent decrease in cell viability (from 75% at 10 nM to 12% at 500 nM), whereas controls did not show any cytotoxicity. PC3-PIP tumour-bearing mice subjected to photodynamic therapy showed a delayed tumour growth. In conclusion, a novel PSMA-targeted NIRF dye with dual imaging-PDT capabilities was synthesized and displayed superior specificity compared to other small PSMA targeted molecules

In vivo assessment of tumour associated macrophages in murine melanoma obtained by low-field relaxometry in the presence of iron oxide particles

Acknowledgements The authors would like to acknowledge Dr Dana Dawson, University of Aberdeen, UK, for the supply of ferumoxytol. This project has received funding from the European Union Horizon 2020 research and innovation programme under grant agreement No 668119 (project “IDentIFY”) and it was performed in the frame of the COST Action AC15209 (EURELAX). Maria Rosaria Ruggiero was supported by a “FIRC-AIRC fellowship for Italy”. The Italian Ministry for Education and Research (MIUR) is gratefully acknowledged for yearly FOE funding to the Euro-BioImaging Multi-Modal Molecular Imaging Italian Node (MMMI). Data availability All data analysed during this study are included in this published article (and its supplementary information file). Other raw data required to reproduce these findings are available from the corresponding author on reasonable request.Peer reviewedPublisher PD

Imaging of Inflammation in Spinal Cord Injury: Novel Insights on the Usage of PFC-Based Contrast Agents

Labeling of macrophages with perfluorocarbon (PFC)-based compounds allows the visualization of inflammatory processes by 19F-magnetic resonance imaging (19F-MRI), due to the absence of endogenous background. Even if PFC-labeling of monocytes/macrophages has been largely investigated and used, information is lacking about the impact of these agents over the polarization towards one of their cell subsets and on the best way to image them. In the present work, a PFC-based nanoemulsion was developed to monitor the course of inflammation in a model of spinal cord injury (SCI), a pathology in which the understanding of immunological events is of utmost importance to select the optimal therapeutic strategies. The effects of PFC over macrophage polarization were studied in vitro, on cultured macrophages, and in vivo, in a mouse SCI model, by testing and comparing various cell tracking protocols, including single and multiple administrations, the use of MRI or Point Resolved Spectroscopy (PRESS), and application of pre-saturation of Kupffer cells. The blood half-life of nanoemulsion was also investigated by 19F Magnetic Resonance Spectroscopy (MRS). In vitro and in vivo results indicate the occurrence of a switch towards the M2 (anti-inflammatory) phenotype, suggesting a possible theranostic function of these nanoparticles. The comparative work presented here allows the reader to select the most appropriate protocol according to the research objectives (quantitative data acquisition, visual monitoring of macrophage recruitment, theranostic purpose, rapid MRI acquisition, etc.). Finally, the method developed here to determine the blood half-life of the PFC nanoemulsion can be extended to other fluorinated compounds

Monitoring tissue implants by field-cycling H-1-MRI via the detection of changes in the N-14-quadrupolar-peak from imidazole moieties incorporated in a "smart" scaffold material

Acknowledgements This work was performed in the frame of the COST Action AC15209 (EURELAX). The authors acknowledge the Italian Ministry of Research for FOE contribution to the EuroBioImaging MultiModal Molecular Imaging Italian Node (www.mmmi.unito.it). This project has received funding from the European Union Horizon 2020 research and innovation program under grant agreement No 668119 (project “IDentIFY”) and from the ATTRACT project funded by the EC under Grant Agreement No. 777222.Peer reviewedPostprin

Low-Field NMR Relaxometry for Intraoperative Tumour Margin Assessment in Breast-conserving Surgery

Funding: This project has received funding from the European Union Horizon 2020 research and innovation programme under grant agreement no. 668119 (project “IDentIFY”) and from AIRC under IG 2019, ID 23267 project (PI Geninatti Crich Simonetta). The authors acknowledge the Italian Ministry of Research for FOE contribution to the Euro-BioImaging MultiModal Molecular Imaging Italian Node (www.mmmi.unito.it accessed on 1 October 2020). Maria Rosaria Ruggiero was supported by a “FIRC-AIRC fellowship for Italy”. Acknowledgments: This work was performed in the frame of the COST Action CA15209 (EURELAX).Peer reviewedPublisher PD

Cobaltabis(dicarbollide) ([o-COSAN]−) as Multifunctional Chemotherapeutics: A Prospective Application in Boron Neutron Capture Therapy (BNCT) for Glioblastoma

Purpose: The aim of our study was to assess if the sodium salt of cobaltabis(dicarbollide) and its di-iodinated derivative (Na[o-COSAN] and Na[8,8′-I2-o-COSAN]) could be promising agents for dual anti-cancer treatment (chemotherapy + BNCT) for GBM. Methods: The biological activities of the small molecules were evaluated in vitro with glioblastoma cells lines U87 and T98G in 2D and 3D cell models and in vivo in the small model animal Caenorhabditis elegans (C. elegans) at the L4-stage and using the eggs. Results: Our studies indicated that only spheroids from the U87 cell line have impaired growth after treatment with both compounds, suggesting an increased resistance from T98G spheroids, contrary to what was observed in the monolayer culture, which highlights the need to employ 3D models for future GBM studies. In vitro tests in U87 and T98G cells conclude that the amount of 10B inside the cells is enough for BNCT irradiation. BNCT becomes more effective on T98G after their incubation with Na[8,8′-I2-o-COSAN], whereas no apparent cell-killing effect was observed for untreated cells. Conclusions: These small molecules, particularly [8,8′-I2-o-COSAN]−, are serious candidates for BNCT now that the facilities of accelerator-based neutron sources are more accessible, providing an alternative treatment for resistant glioblastoma