Cinzia Boffa

Sonosensitive theranostic liposomes for preclinical in vivo MRI-guided visualization of doxorubicin release stimulated by pulsed low intensity non-focused ultrasound

The main goal of this study was to assess the theranostic performance of a nanomedicine able to generate MRI contrast as a response to the release from liposomes of the antitumor drug Doxorubicin triggered by the local exposure to pulsed low intensity non focused ultrasounds (pLINFU). In vitro experiments showed that Gadoteridol was an excellent imaging agent for probing the release of Doxorubicin following pLINFU stimulation. On this basis, the theranostic system was investigated in vivo on a syngeneic murine model of TS/A breast cancer. MRI offered an excellent guidance for monitoring the pLINFU-stimulated release of the drug. Moreover, it provided: i) an in vivo proof of the effective release of the liposomal content, and ii) a confirmation of the therapeutic benefits of the overall protocol. Ex vivo fluorescence microscopy indicated that the good therapeutic outcome was originated from a better diffusion of the drug in the tumor following the pLINFU stimulus. Very interestingly, the broad diffusion of the drug in the tumor stroma appeared to be mediated by the presence of the liposomes themselves. The results of this study highlighted either the great potential of US-based stimuli to safely trigger the release of a drug from its nanocarrier or the associated significant therapeutic improvement. Finally, MRI demonstrated to be a valuable technique to support chemotherapy and monitoring the outcome. Furthermore, in this specific case, the theranostic agent developed has a high clinical translatability because the MRI agent utilized is already approved for human use.

A R2p /R1p ratiometric procedure to assess matrix metalloproteinase-2 activity by magnetic resonance imaging.

The approach to molecular imaging of enzymes by MRI typically relies upon imaging probes composed of an enzymecleavable moiety conjugated with a paramagnetic imaging reporter, such as a Gd chelate. Upon enzymatic processing, the probe is transformed into a fragment with an altered relaxivity, leading to a different capability to enhance contrast in MR images with respect to the parent species. Ideally, the unprocessed (intact) form of the probe should be completely silent while the processed (cleaved) form should have a high relaxivity (that is, high contrast enhancement). In such a way the appearance of contrast within images can be unambiguously attributed to the result of enzymatic activity and not to dynamic changes of tissue probe concentration. However, gadolinium-based agents as enzyme responsive agents are never completely silent and both forms (unprocessed and processed) contribute to the overall contrast enhancement as a function of their respective relaxivities and tissue local concentrations. Exact knowledge of the total concentration of Gd is essential to translate image contrast enhancement into the molar ratio of unprocessed versus processed forms, and thus into true enzyme activity maps. A viable solution to the concentration problem can be provided by the R2p/R1p ratiometric approach, which is based on the measurement of the ratio between the transverse and longitudinal paramagnetic contributions to the water proton relaxation rate; that is, R2p and R1p (with Ri = 1/Ti, i = 1,2). Although ratiometric approaches are well-established for CEST-MRI contrast agents, there are only a couple of examples of application to Gd based relaxation agents, namely for pH and temperature imaging. 5] Let us consider two Gd complexes, GdL and GdF, each characterized by its own transverse and longitudinal millimolar relaxivity (r2 and r1 terms respectively, in units of mm 1 s ). These species form the ratiometric pair, and the total paramagnetic relaxation enhancement of a mixture of the two can be expressed as:

In vivo MRI visualization of release from liposomes triggered by local application of pulsed low-intensity non-focused ultrasound.

UNLABELLED The work aimed at developing a MRI-guided protocol for the visualization of the release of material entrapped in liposomes stimulated by the local application of pulsed low-intensity non-focused ultrasound (pLINFU). The task was achieved by formulating liposomes filled up with the clinically approved paramagnetic agent gadoteridol, because the release of the agent from the nanovesicles is accompanied by a significant MRI signal enhancement. The protocol was validated in vivo on mice-bearing subcutaneous syngeneic B16 melanoma and i.v. injected with the paramagnetic liposomes. Upon exposing tumor to pLINFU (3MHz, insonation time 2min, duty cycle 50%) few minutes after liposomes injection, a signal enhancement of ca. 35% was detected. The effective release of the agent was confirmed by the strong enhancement measured in kidneys calyx and bladder due to the rapid renal excretion of the agent released in the tumor. FROM THE CLINICAL EDITOR In this paper, a pulsed low-intensity non-focused ultrasound-based technique was used to release a paramagnetic MRI contrast agent from liposomes, demonstrating the feasibility of this triggered release system in a mouse melanoma model for future research applications.

Design and testing of paramagnetic liposome-based CEST agents for MRI visualization of payload release on pH-induced and ultrasound stimulation

Abstract The development of nanomedicines in cancer therapy is constantly growing because of the advantages associated with the use of nanosized drug delivery systems. Among them, the possibility of accurate spatiotemporal control of the release of the chemotherapeutic from the carrier is one of the most interesting and clinically relevant. To further improve the therapy outcome, the clinical translation of imaging protocols for the in vivo visualization of the release step is of paramount importance. In this work, the combination of the great chemical versatility of liposomes and the outstanding potential of MRI chemical exchange saturation transfer agents has been successfully harnessed to image the selective release of the liposomal content stimulated by endogenous (variation of pH) and externally applied (nonfocused ultrasound) stimuli. The use of clinically safe components (both liposomes and MRI agents) and the good results obtained in vitro hold promise for a successful future in vivo translation.Graphical abstract

Low Foxp3 expression in negative sentinel lymph nodes is associated with node metastases in colorectal cancer

In a recent commentary, Sobhanu and Le Gouvello,1 take advantage of the account of Chaput et al 2 of a new population of T regulatory (Treg) lymphocytes (CD8+) to address the more general question of whether accumulation of Tregs (both CD8+ and conventional CD4+) must be considered a prognostic factor in colorectal cancer (CRC). Tregs (Foxp3+) play a pivotal role in maintaining immune system homeostasis through their ability to suppress immunological responses, including tumour immunity against tumour-associated antigens. In their interesting commentary, Sobhanu and Le Gouvello1 argue that the in vivo immunosuppressive effect of these cells in CRC still remains controversial. Actually, accordingly to the available data, we believe it reasonable to state that CD4+ Tregs do not contribute to CRC escape from host immunity. While earlier studies showed a higher density of tumour-infiltrating Tregs in advanced …

Successful in vivo MRI tracking of MSCs labeled with Gadoteridol in a Spinal Cord Injury experimental model

In this study, murine Mesenchymal Stem Cells (MSCs) labeled with the clinically approved MRI agent Gadoteridol through a procedure based on the hypo-osmotic shock were successfully tracked in vivo in a murine model of Spinal Cord Injury (SCI). With respect to iso-osmotic incubations, the hypo-osmotic labeling significantly increased the Gd(3+) cellular uptake, and enhanced both the longitudinal relaxivity (r1) of the intracellular Gadoteridol and the Signal to Noise Ratio (SNR) measured on cell pellets, without altering the biological and functional profile of cells. A substantial T1 Contrast Enhancement after local transplantation of 3.0×10(5) labeled cells in SCI mice enabled to follow their migratory dynamics in vivo for about 10days, and treated animals recovered from the motor impairment caused by the injury, indicating unaltered therapeutic efficacy. Finally, analytical and histological data corroborated the imaging results, highlighting the opportunity to perform a precise and reliable monitoring of the cell-based therapy.

A relaxometric method for the assessment of intestinal permeability based on the oral administration of gadolinium-based MRI contrast agents.

Herein, a new relaxometric method for the assessment of intestinal permeability based on the oral administration of clinically approved gadolinium (Gd)‐based MRI contrast agents (CAs) is proposed. The fast, easily performed and cheap measurement of the longitudinal water proton relaxation rate (R1) in urine reports the amount of paramagnetic probe that has escaped the gastrointestinal tract. The proposed method appears to be a compelling alternative to the available methods for the assessment of intestinal permeability. The method was tested on the murine model of dextran sulfate sodium (DSS)‐induced colitis in comparison with healthy mice. Three CAs were tested, namely ProHance®, MultiHance® and Magnevist®. Urine was collected for 24 h after the oral ingestion of the Gd‐containing CA at day 3–4 (severe damage stage) and day 8–9 (recovery stage) after treatment with DSS. The Gd content in urine measured by 1H relaxometry was confirmed by inductively coupled plasma‐mass spectrometry (ICP‐MS). The extent of urinary excretion was given as a percentage of excreted Gd over the total ingested dose. The method was validated by comparing the results obtained with the established methodology based on the lactulose/mannitol and sucralose tests. For ProHance and Magnevist, the excreted amounts in the severe stage of damage were 2.5–3 times higher than in control mice. At the recovery stage, no significant differences were observed with respect to healthy mice. Overall, a very good correlation with the lactulose/mannitol and sucralose results was obtained. In the case of MultiHance, the percentage of excreted Gd complex was not significantly different from that of control mice in either the severe or recovery stages. The difference from ProHance and Magnevist was explained on the basis of the (known) partial biliary excretion of MultiHance in mice. Copyright