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Cancer is one of the most devastating diseases that the world is currently facing, accounting for 10 million deaths in 2020 (WHO). In the last two decades, advanced medical imaging has played an ever more important role in the early detection of the disease, as it increases the chances of survival and the potential for full recovery. To date, dynamic glucose-enhanced (DGE) MRI using glucose-based chemical exchange saturation transfer (glucoCEST) has demonstrated the sensitivity to detect both d-glucose and glucose analogs, such as 3-oxy-methyl-d-glucose (3OMG) uptake in tumors. As one of the recent international efforts aiming at pushing the boundaries of translation of the DGE MRI technique into clinical practice, a multidisciplinary team of eight partners came together to form the “glucoCEST Imaging of Neoplastic Tumors (GLINT)” consortium, funded by the Horizon 2020 European Commission. This paper summarizes the progress made to date both by these groups and others in increasing our knowledge of the underlying mechanisms related to this technique as well as translating it into clinical practice.

The search for alternatives to Gd-containing magnetic resonance imaging (MRI) contrast agents addresses the field of Fe(III)-bearing species with the expectation that the use of an essential metal ion may avoid the issues raised by the exogenous Gd. Attention is currently devoted to highly stable Fe(III) complexes with hexacoordinating ligands, although they may lack any coordinated water molecule. We found that the hexacoordinated Fe(III) complex with two units of deferasirox, a largely used iron sequestering agent, owns properties that can make it a viable alternative to Gd-based agents. Fe(deferasirox)2 displays an outstanding thermodynamic stability, a high binding affinity to human serum albumin (three molecules of complex are simultaneously bound to the protein), and a good relaxivity that increases in the range 20-80 MHz. The relaxation enhancement is due to second sphere water molecules likely forming H-bonds with the coordinating phenoxide oxygens. A further enhancement was observed upon the formation of the supramolecular adduct with albumin. The binding sites of Fe(deferasirox)2 on albumin were characterized by relaxometric competitive assays. Preliminary in vivo imaging studies on a tumor-bearing mouse model indicate that, on a 3 T MRI scanner, the contrast ability of Fe(deferasirox)2 is comparable to the one shown by the commercial Gd(DTPA) agent. ICP-MS analyses on blood samples withdrawn from healthy mice administered with a dose of 0.1 mmol/kg of Fe(deferasirox)2 showed that the complex is completely removed in 24 h.

Gadolinium(III) complexes have been employed for more than 30 years as contrast agents in magnetic resonance imaging (MRI). In order to further improve the diagnostic accuracy of enhanced magnetic resonance images or to provide comparable enhancement at a reduced administered dose, current research is focusing on the development of GdIII-complexes characterized by higher relaxivity. In this study we describe the synthesis and the equilibrium, kinetic, relaxation and structural properties of two new GdIII-complexes based on modified 10-(2-hydroxypropyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (HP-DO3A) structure which, due to an intramolecular prototropic exchange, display more than two-fold higher relaxivity compared to currently available GdIII-based MRI contrast agents.

This study reports the development of a completely new class of MRI contrast agents, displaying remarkable relaxation effects in the absence of paramagnetic metal ions. Their detection requires the acquisition of images at variable magnetic field strength as provided by fast field cycling imaging scanners. They contain poly-histidine chains (poly-His), whose imidazole groups generate 14 N-quadrupolar-peaks that cause a relaxation enhancement of water protons at a frequency (1.38±0.3 MHz) that is readily detectable from the frequencies associated with endogenous proteins. The poly-His quadrupolar peaks are detectable only when the polymer is in a solid-like form, that is, at pH>6.6. Above this value, their intensity is pH dependent and can be used to report on the occurring pH changes. On this basis, the poly-His moieties were conjugated to biocompatible polymers, such as polylactic and glycolic acid, in order to form stable nanoparticles able to encapsulate structured water in their core. FFC images were acquired to assess their contrast-generating ability.

Objectives The targeting of tumor cells and their visualization with magnetic resonance imaging (MRI) is an important task in biomedicine. The low sensitivity of this technique is a significant drawback and one that may hamper the detection of the imaging reporters used.To overcome this sensitivity issue, this work explores the synergy between 2 strategies: (1) arginine, glycine, aspartic acid peptide (RGD)-functionalized giant unilamellar vesicles (GUVs) loaded with Gd complexes to accumulate large amounts of MRI contrast agent at the targeting site; and (2) the use of magnetization transfer contrast (MTC), which is a sensitive MRI technique for the detection of Gd complexes in the tumor region. Materials and methods Giant unilamellar vesicles were prepared using the gentle swelling method, and the cyclic RGD targeting moiety was introduced onto the external membrane. Paramagnetic Gd-containing complexes and the fluorescent probe rhodamine were both part of the vesicle membranes and Gd-complexes were also the payload within the inner aqueous cavity. Giant unilamellar vesicles that were loaded with the imaging reporters, but devoid of the RGD targeting moiety, were used as controls. U-87 MG human glioblastoma cells, which are known to overexpress the targets for RGD moieties, were used. In the in vivo experiments, U-87 MG cells were subcutaneously injected into nu/nu mice, and the generated tumors were imaged using MRI, 15 days after cell administration. Magnetic resonance imaging was carried out at 7 T, and T2W, T1W, and MTC/Z-spectra were acquired. Confocal microscopy images and Inductively Coupled Plasma Mass Spectrometry (ICP-MS) were used for result validation. Results In vitro results show that RGD GUVs specifically bind to U-87 MG cells. Microscopy demonstrates that (1) RGD GUVs were anchored onto the external surface of the tumor cells without any internalization; (2) a low number of GUVs per cell were clustered at specific regions; and (3) there is no evidence for macrophage uptake or cell toxicity. The MRI of cell pellets after incubation with RGD GUVs and untargeted ctrl-GUVs was performed. No difference in T1 signal was detected, whereas a 15% difference in MT contrast is present between the RGD GUV-treated cells and the ctrl-GUV-treated cells.Magnetic resonance imaging scans of tumor-bearing mice were acquired before and after (t = 0, 4 hours and 24 hours) the administration of RGD GUVs and ctrl-GUVs. A roughly 16% MTC difference between the 2 groups was observed after 4 hours. Immunofluorescence analyses and ICP-MS analyses (for Gd-detection) of the explanted tumors confirmed the specific accumulation of RGD GUVs in the tumor region. Conclusions RGD GUVs seem to be interesting carriers that can facilitate the specific accumulation of MRI contrast agents at the tumor region. However, the concentration achieved is still below the threshold needed for T1w-MRI visualization. Conversely, MTC proved to be sufficiently sensitive for the visualization of detectable contrast between pretargeting and posttargeting images.

The altered regulation of iron uptake and metabolism in cancerous cells, along with the potential of this metal to cause oxidative stress and cell death, makes iron overload an attractive therapeutic strategy for cancer treatment. In this study, the selective uptake of native HoS-ferritin (Horse-Spleen Ferritin) was assessed in TS/A breast cancer cells and compared with benign cystadenoma NMuMG. The higher expression of L-ferritin receptor SCARA5 led to an enhanced uptake in TS/A that is detected by the generation of a negative contrast in the corresponding MR images. The toxicity of HoS-ferritin toward TS/A cells has been investigated in detail in vitro, showing that cellular vitality is inversely related to the amount of internalized iron content. Finally, biodistribution and therapeutic efficacy of HoS-ferritin have been shown for the first time in vivo on a orthotopic breast cancer mice model, suggesting that iron overdose delivered by the HoS-ferritin can trigger selective mechanisms of regulated cell death.

8-Hydroxypyrene-1,3,6-trisulfonate (HPTS) is a small, hydrophilic fluorescent molecule. Since the pKa of the hydroxyl group is close to neutrality and quickly responds to pH changes, it is widely used as a pH-reporter in cell biology for measurements of intracellular pH. HPTS fluorescence (both excitation and emission spectra) at variable pH was measured in pure water in the presence of NaCl solution or in the presence of different buffers (PBS or hepes in the presence or not of NaCl) and in a solution containing BSA. pKa values have been obtained from the sigmoidal curves. Herein, we investigated the effect of mono-, di-, and trivalent cations (Na

Amide proton transfer-weighted (APTw) MR imaging shows promise as a biomarker of brain tumor status. Currently used APTw MRI pulse sequences and protocols vary substantially among different institutes, and there are no agreed-on standards in the imaging community. Therefore, the results acquired from different research centers are difficult to compare, which hampers uniform clinical application and interpretation. This paper reviews current clinical APTw imaging approaches and provides a rationale for optimized APTw brain tumor imaging at 3 T, including specific recommendations for pulse sequences, acquisition protocols, and data processing methods. We expect that these consensus recommendations will become the first broadly accepted guidelines for APTw imaging of brain tumors on 3 T MRI systems from different vendors. This will allow more medical centers to use the same or comparable APTw MRI techniques for the detection, characterization, and monitoring of brain tumors, enabling multi-center trials in larger patient cohorts and, ultimately, routine clinical use.

Changes in metabolism is an hallmark that characterizes tumour cells from healthy ones. Their detection can be highly relevant for staging the tumor and for monitoring the response to therapeutic treatments. Herein it is shown the readout of these changes can be achieved either by assessing the pH of the extracellular space in the tumour region and by monitoring real time transformations of hyperpolarized C-13 labelled substrates. Mapping pH in a MR image is possible by measuring the CEST response of an administered contrast agent such as Iopamidol that can provide accurate measurements of the heterogeneity of tumour acidosis. Direct detection of relevant enzymatic activities have been acquired by using Pyruvate and Fumarate hyperpolarized by the incorporation of a molecule of para-H

This work aims to investigate the supramolecular binding interactions that occur between iodinated X-ray contrast agents (CAs) and macrocyclic gadolinium (Gd)-based MRI contrast agents (GBCAs). This study provides some new insights in the renal excretion pathways of the two types of imaging probes.The water-proton relaxivities (rA significant increase in rThe reported results show that it is possible to detect the co-presence of iodinated agents and macrocyclic GBCAs in contrast-enhanced MR images. The new information may be useful in the design of novel experiments toward improved diagnostic outcomes.

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 kio. The latter has been estimated by analyzing the magnetization recovery curve in standard NMR T1 measurements when there is a marked difference in the proton relaxation rate constants (R1) between the intra- and the extra-cellular compartments. In cellular studies, T1 measurements were carried out on a relaxometer working at 0.5 T, and the required difference in R1 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 R1 values of the two-compartments was achieved when the T1 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 kio 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 kio can be considered a non-invasive, early and predictive biomarker for the identification of responsive patients immediately from the first doxorubicin treatment.

International audience; Dysfunctional elastin turnover plays a major role in the progression of atherosclerotic plaques. Failure of tropoelastin cross-linking into mature elastin leads to the accumulation of tropoelastin within the growing plaque, increasing its instability. Here we present Gd4-TESMA, an MRI contrast agent specifically designed for molecular imaging of tropoelastin within plaques. Gd4-TESMA is a tetrameric probe composed of a tropoelastin-binding peptide (the VVGS-peptide) conjugated with four Gd(III)-DOTA-monoamide chelates. It shows a relaxivity per molecule of 34.0 ± 0.8 mM-1 s-1 (20 MHz, 298 K, pH 7.2), a good binding affinity to tropoelastin (KD = 41 ± 12 μM), and a serum half-life longer than 2 h. Gd4-TESMA accumulates specifically in atherosclerotic plaques in the ApoE-/- murine model of plaque progression, with 2 h persistence of contrast enhancement. As compared to the monomeric counterpart (Gd-TESMA), the tetrameric Gd4-TESMA probe shows a clear advantage regarding both sensitivity and imaging time window, allowing for a better characterization of atherosclerotic plaques.

The search for alternatives to Gd-containing magnetic resonance imaging (MRI) contrast agents addresses the field of Fe(III)-bearing species with the expectation that the use of an essential metal ion may avoid the issues raised by the exogenous Gd. Attention is currently devoted to highly stable Fe(III) complexes with hexacoordinating ligands, although they may lack any coordinated water molecule. We found that the hexacoordinated Fe(III) complex with two units of deferasirox, a largely used iron sequestering agent, owns properties that can make it a viable alternative to Gd-based agents. Fe(deferasirox)

D-Glucose and 3-O-Methyl-D-glucose (3OMG) have been shown to provide contrast in magnetic resonance imaging-chemical exchange saturation transfer (MRI-CEST) images. However, a systematic comparison between these two molecules has yet to be performed. The current study deals with the assessment of the effect of pH, saturation power level (B1 ) and magnetic field strength (B0 ) on the MRI-CEST contrast with the aim of comparing the in vivo CEST contrast detectability of these two agents in the glucoCEST procedure. Phosphate-buffered solutions of D-Glucose or 3OMG (20 mM) were prepared at different pH values and Z-spectra were acquired at several B1 levels at 37°C. In vivo glucoCEST images were obtained at 3 and 7 T over a period of 30 min after injection of D-Glucose or 3OMG (at doses of 1.5 or 3 g/kg) in a murine melanoma tumor model (n = 3-5 mice for each molecule, dose and B0 field). A markedly different pH dependence of CEST response was observed in vitro for D-Glucose and 3OMG. The glucoCEST contrast enhancement in the tumor region following intravenous administration (at the 3 g/kg dose) was comparable for both molecules: 1%-2% at 3 T and 2%-3% at 7 T. The percentage change in saturation transfer that resulted was almost constant for 3OMG over the 30-min period, whereas a significant increase was detected for D-Glucose. Our results show similar CEST contrast efficiency but different temporal kinetics for the metabolizable and the nonmetabolizable glucose derivatives in a tumor murine model when administered at the same doses.

T1-weighted magnetic resonance images of water in the surroundings of a Nafion surface allowed the identification of the presence of a low-mobility zone (LMZ), 60 μm thick, consisting of water mole...

This study is focused on the development of innovative sensors to non-invasively monitor the tissue implant status by Fast-Field-Cycling Magnetic Resonance Imaging (FFC-MRI). These sensors are based on oligo-histidine moieties that are conjugated to PLGA polymers representing the structural matrix for cells hosting scaffolds. The presence of 14N atoms of histidine causes a quadrupolar relaxation enhancement (also called Quadrupolar Peak, QP) at 1.39 MHz. This QP falls at a frequency well distinct from the QPs generated by endogenous semisolid proteins. The relaxation enhancement is pH dependent in the range 6.5-7.5, thus it acts as a reporter of the scaffold integrity as it progressively degrades upon lowering the microenvironmental pH. The ability of this new sensors to generate contrast in an image obtained at 1.39 MHz on a FFC-MRI scanner is assessed. A good biocompatibility of the histidine-containing scaffolds is observed after its surgical implantation in healthy mice. Over time the scaffold is colonized by endogenous fibroblasts and this process is accompanied by a progressive decrease of the intensity of the relaxation peak. In respect to the clinically used contrast agents this material has the advantage of generating contrast without the use of potentially toxic paramagnetic metal ions.

Triple-negative breast cancer (TNBC) patients have usually poor outcome after chemotherapy and early prediction of therapeutic response would be helpful. [Breast tumor (4T1)-bearing mice were treated with doxorubicin or dichloroacetate for 1 week. PET/CT with [Doxorubicin-treated mice showed a significant decrease in tumor growth when compared to the control group. GlucoCEST imaging provided metabolic response after three cycles of treatment. Conversely, no variations were detected in [Metabolic changes during doxorubicin treatment can be predicted by glucoCEST imaging that appears more sensitive than [

Hyperpolarization‐enhanced magnetic resonance imaging can be used to study biomolecular processes in the body, but typically requires nuclei such as 13C, 15N, or 129Xe due to their long spin‐polarization lifetimes and the absence of a proton‐background signal from water and fat in the images. Here we present a novel type of 1H imaging, in which hyperpolarized spin order is locked in a nonmagnetic long‐lived correlated (singlet) state, and is only liberated for imaging by a specific biochemical reaction. In this work we produce hyperpolarized fumarate via chemical reaction of a precursor molecule with para‐enriched hydrogen gas, and the proton singlet order in fumarate is released as antiphase NMR signals by enzymatic conversion to malate in D2O. Using this model system we show two pulse sequences to rephase the NMR signals for imaging and suppress the background signals from water. The hyperpolarization‐enhanced 1H‐imaging modality presented here can allow for hyperpolarized imaging without the need for low‐abundance, low‐sensitivity heteronuclei., A proton singlet state on fumarate derived from parahydrogen can be broken by enzymatic conversion to malate, which releases hyperpolarized NMR signals. We demonstrate two “out‐of‐phase echo” pulse sequences to convert the I 1z I 2z state into net proton magnetization, and show how this can be used for hyperpolarized 1H imaging.

Herein, the synthesis and an extensive characterization of two novel Gd(AAZTA) (AAZTA=6-amino-6-methylperhydro-1,4-diazepine tetra acetic acid) derivatives functionalized with short (C

ProceduresTo preliminary assess the relationship between Manganese Enhanced Magnetic Resonance Imaging (MEMRI) and the expression of calcium receptors in human prostate and breast cancer animal models.MethodsNOD/SCID mice were inoculated with MDA-MB-231 breast cancer cells and prostate PC3 cancer cells to develop orthotopic or pseudometastatic cancer animal models. Mice were studied on a clinical 3T scanner by using a prototype birdcage coil before and after intravenous injection of MnCl2. Assessment of receptor's status was carried out after the MR images acquisition by immunohistochemistry on excised tumours.ResultsManganese contrast enhancement in breast or prostate cancer animal models well correlated with CaSR expression (pConclusionOur preliminary results suggest that MEMRI appears an efficient tool to characterize human breast and prostate cancer animal models in the presence of different expression level of calcium receptors.

Hyperpolarization-enhanced magnetic resonance imaging can be used to study biomolecular processes in the body, but typically requires nuclei such as 13C, 15N, or 129Xe due to their long spin‑polarization lifetimes and the absence of a proton‑background signal from water and fat in the images. Here we present a novel type of 1H imaging, in which hyperpolarized spin order is locked in a nonmagnetic long-lived correlated (singlet) state, and is only liberated for imaging by a specific biochemical reaction. In this work we produce hyperpolarized fumarate via chemical reaction of a precursor molecule with para-enriched hydrogen gas, and the proton singlet order in fumarate is released as antiphase NMR signals by enzymatic conversion to malate in D2O. Using this model system we show two pulse sequences to rephase the NMR signals for imaging and suppress the background signals from water. The hyperpolarization-enhanced 1H‑imaging modality presented here can allow for hyperpolarized imaging without the need for low‑abundance, low‑sensitivity heteronuclei.

Nowadays, magnetic resonance imaging (MRI) is one of the key, noninvasive modalities to detect and stage cancer which benefits from contrast agents (CA) to differentiate healthy from tumor tissue. An innovative class of MRI CAs is represented by Gd-loaded gold nanoparticles. The size, shape and chemical functionalization of Gd-loaded gold nanoparticles appear to affect the observed relaxation enhancement of water protons in their suspensions. The herein reported results shed more light on the determinants of the relaxation enhancement brought by Gd-loaded concave cube gold nanoparticles (CCGNPs). It has been found that, in the case of nanoparticles endowed with concave surfaces, the relaxivity is remarkably higher compared to the corresponding spherical (i.e., convex) gold nanoparticles (SPhGNPs). The main determinant for the observed relaxation enhancement is represented by the occurrence of a large contribution from second sphere water molecules which can be exploited in the design of high-efficiency MRI CA.

Nanotheranostic reagents that integrate magnetic resonance imaging (MRI) and photothermal therapy (PTT) offer a promising strategy for the treatment of human disease. However, classic gadolinium (Gd)-based T(1)-MRI contrast agents are limited by their low relaxivity. To address this, we produced Gd-loaded Tobacco mosaic virus (TMV) particles coated with the mussel-inspired biopolymer polydopamine (PDA). Such biocompatible nanotheranostic reagents can be used to facilitate PTT, guided by multimodal magnetic resonance/photoacoustic imaging. The r(1)-relaxivity of the Gd-TMV-PDA particles at 60 MHz was ~80 mM(−1)s(−1), compared to 13.63 mM(−1)s(−1) for the uncoated Gd-TMV particles. The Gd-TMV-PDA particles also promoted strong near-infrared absorption with high photothermal conversion efficiency (28.9%) and demonstrated excellent photoacoustic contrast. Multimodal imaging and PTT resulted in the effective killing of PC-3 prostate cancer cells. Gd-TMV-PDA nanoparticles therefore offer a promising theranostic approach that can now be tested in vivo in cancer models.

This contribution describes the use of Fast Field-cycling relaxometry (FFC-NMR) for the characterisation of Gd(III)- and Mn(II)-based contrast agents for MRI. Through a series of selected examples,...

The relaxivity of Gd(HP-DO3A) was studied as a function of pH and buffer composition in order to identify the main factors of the observed relaxation enhancement due to the exchange of the coordinated hydroxyl proton. It was established that the paramagnetic relaxation time, T1M, of the coordinated hydroxyl proton is about 50% shorter than that of the protons in the coordinated water molecule. The control of the p K of the coordinated alcoholic -OH moiety in the ligand is fundamental to utilize the proton exchange enhanced relaxivity under physio/pathologic conditions. A new derivative of Gd(HP-DO3A) was synthesized by replacing the -CH3 group with a -CF3 moiety. In this complex, the -OH group becomes more acidic. Consequently, the maximum contribution of the proton exchange to the relaxivity is shifted to a lower pH region with the fluorinated ligand.

Magnetic Resonance Imaging (MRI) enables to provide anatomical, functional and molecular information of pathological angiogenesis when used with properly tailored imaging probes. Functional studies have been the domain of Dynamic Contrast Enhancement (DCE) -MRI protocols from which it is possible to extract quantitative estimations on key parameters such as the volumes of vascular and extracellular compartments and the rates of the bidirectional exchange of the imaging reporters across the endothelial barrier. Whereas paramagnetic Gd-complexes able to reversibly bind to serum albumin act better than the clinically used small-sized, hydrophilic species, new findings suggest that an accurate assessment of the vascular volume is possible by analyzing images acquired upon the i.v. administration of Gd-labelled Red Blood Cells (RBCs). As far as it concerns molecular MRI, among the many available biomarkers, αvβ3 integrins are the most investigated ones. The low expression of these targets makes mandatory the use of nano-sized systems endowed with the proper signal enhancing capabilities. A number of targeted nano-particles have been investigated including micelles, liposomes, iron oxides and perfluorocarbon containing systems. Finally, a growing attention is devoted to the design and testing of "theranostic" agents based on the exploitation of MRI to monitor drug delivery processes and therapeutic outcome.

Mobile proton-containing solutes can be detected by MRI by the chemical exchange saturation transfer (CEST) method. CEST sensitivity is dramatically enhanced by using, as exchanging protons, the water molecules confined inside liposomes, shifted by a paramagnetic shift reagent. The chemical shift of the intraliposomal water resonance (δIL ) is affected by the overall shape of the supramolecular system. δIL of a spherical LipoCEST acts as a sensitive reporter of the distribution of streptavidin proteins anchored at the liposome surface by biotinylated phospholipids. This finding prompted the design of a MMP-2 responsive LipoCEST agent as the streptavidin moieties can be released from the liposome surfaces when a properly tailored enzyme-cleavable peptide is inserted on the phospholipids before the terminal biotin residues. δIL reports on the overall changes in the supramolecular architecture associated to the cleavage carried out by MMP-2.

PURPOSE To extend the pH detection range of iopamidol-based ratiometric chemical exchange saturation transfer (CEST) MRI at sub-high magnetic field and establish quantitative renal pH MRI. METHODS Chemical exchange saturation transfer imaging was performed on iopamidol phantoms with pH of 5.5 to 8.0 and in vivo on rat kidneys (n = 5) during iopamidol administration at a 4.7 T. Iopamidol CEST effects were described using a multipool Lorentzian model. A generalized ratiometric analysis was conducted by ratioing resolved iopamidol CEST effects at 4.3 and 5.5 ppm obtained under 1.0 and 2.0 µT, respectively. The pH detection range was established for both the standard ratiometric analysis and the proposed resolved approach. Renal pH was mapped in vivo with regional pH assessed by one-way analysis of variance. RESULTS Good-fitting performance was observed in multipool Lorentzian resolving of CEST effects (R2 s > 0.99). The proposed approach extends the in vitro pH detection range to 5.5 to 7.5 at 4.7 T. In vivo renal pH was measured to be 7.0 ± 0.1, 6.8 ± 0.1, and 6.5 ± 0.2 for cortex, medulla and calyx, respectively (P

Chemical Exchange Saturation Transfer (CEST) approach is a novel tool within magnetic resonance imaging (MRI) that allows visualization of molecules possessing exchangeable protons with water. Many molecules, employed as excipients for the formulation of finished drug products, are endowed with hydroxyl, amine or amide protons, thus can be exploitable as MRI-CEST contrast agents. Their high safety profiles allow them to be injected at very high doses. Here we investigated the MRI-CEST properties of several excipients (ascorbic acid, sucrose, N-acetyl-D-glucosamine, meglumine and 2-pyrrolidone) and tested them as tumor-detecting agents in two different murine tumor models (breast and melanoma cancers). All the investigated molecules showed remarkable CEST contrast upon i.v. administration in the range 1-3 ppm according to the type of mobile proton groups. A marked increase of CEST contrast was observed in tumor regions up to 30 min post injection. The combination of marked tumor contrast enhancement and lack of toxicity make these molecules potential candidates for the diagnosis of tumors within the MRI-CEST approach. (C) 2017 Elsevier B.V. All rights reserved.

The properties of LnIII -HPDO3A complexes as relaxation enhancers and paraCEST agents are essentially related to the hydroxylpropyl moiety. A series of three HPDO3A derivatives, with small modifications to the hydroxyl arm, were herein investigated to understand how heightened control can be gained over the parameters involved in the design of these agents. A full 1 H and 17 O-NMR relaxometric analysis was conducted and demonstrated that increasing the length of the OH group from the lanthanide centre significantly enhanced the water exchange rate of the gadolinium complex, but with a subsequent reduction in kinetic stability. Alternatively, the introduction of an additional methyl group, which increased the steric bulk around the OH moiety, resulted in the formation of almost exclusively the TSAP isomer (95 %) as identified by 1 H-NMR of the europium complex. The gadolinium analogue of this complex also exhibited a very fast water exchange rate, but with no detectable loss of kinetic stability. This complex therefore demonstrates a notable improvement over Gd-HPDO3A.

Objectives Being administered intravenously, the tissue that gadolinium-based contrast agents (GBCAs) for magnetic resonance imaging mostly encounter is blood. Herein, it has been investigated how much Gd is internalized by cellular blood components upon the in vitro incubation of GBCAs in human blood or upon intravenous administration of GBCAs to healthy mice. We report results that show how the superb sensitivity of inductively coupled plasma-mass spectrometry (ICP-MS) allows the detection of very tiny amounts of GBCAs entering red blood cells (RBCs) and white blood cells (WBCs). This finding may introduce new insights in the complex matter relative to excretion and retention pathway of administered GBCAs. Materials and methods The study was tackled by 2 independent approaches. First, human blood was incubated in vitro with 5 mM of GBCAs (gadoteridol, gadobenate dimeglumine, gadodiamide, and gadopentetate dimeglumine) for variable times (30 minutes, 1 hour, 2 hours, and 3 hours) at 37°C. Then, blood cell components were isolated by using the Ficoll Histopaque method, washed 3 times, mineralized, and analyzed by ICP-MS for total Gd quantification. Furthermore, blood components derived from human blood incubated with gadodiamide or gadoteridol underwent UPLC-MS (ultra performance liquid chromatography-mass spectrometry) analysis for determination of the amount of intact Gd-DTPA-BMA and Gd-HPDO3A. Second, the distribution of Gd in the blood components of healthy CD-1 mice was administered intravenously with a single dose (1.2 mmol/kg) of gadodiamide or gadoteridol. Blood samples were separated and processed at different time points (24 hours, 48 hours, 96 hours, and 10 days after GBCA administration). As for human blood, ICP-MS quantification of total Gd and UPLC-MS determination of the amount of intact GBCAs were carried out. Results The amount of Gd taken up by RBCs and WBCs was well detectable by ICP-MS. The GBCAs seem to be able to cross the membrane by diffusion (RBCs) or, possibly, by macropinocytosis (WBCs). Ex vivo studies allowed it to be established that the structure of the different GBCAs were not relevant to determine the amount of Gd internalized in the cells. Although the amount of Gd steadily decreases over time in gadoteridol-labeled cells, in the case of gadodiamide, the amount of Gd in the cells does not decrease (even 10 days after the administration of the GBCA). Moreover, while gadoteridol maintains its structural integrity upon cellular uptake, in the case of gadodiamide, the amount of intact complex markedly decreases over time. Conclusions The detection of significant amounts of Gd in RBCs and WBCs indicates that GBCAs can cross blood cell membranes. This finding may play a role in our understanding of the processes that are at the basis of Gd retention in the tissues of patients who have received the administration of GBCAs.

ProceduresTo assess the relationship between Manganese Enhanced Magnetic Resonance Imaging (MEMRI) and the expression of calcium receptors in human prostate and breast cancer animal models.MethodsNOD/SCID mice were inoculated with MDA-MB-231 breast cancer cells and prostate PC3 cancer cells to develop orthotopic or pseudometastatic cancer animal models. Mice were studied on a clinical 3T scanner by using a prototype birdcage coil before and after intravenous injection of MnCl2. Assessment of receptor’s status was carried out after the MR images acquisition by immunohistochemistry on excised tumours.ResultsManganese contrast enhancement in breast or prostate cancer animal models well correlated with CaSR expression (pConclusionMEMRI appears an efficient tool to characterize human breast and prostate cancer animal models in the presence of different expression level of CaSR.

Aims To determine individual glucose hydroxyl exchange rates at physiological conditions and use this information for numerical optimization of glucoCEST/CESL preparation. To give guidelines for in vivo glucoCEST/CESL measurement parameters at clinical and ultra-high field strengths. Methods Five glucose solution samples at different pH values were measured at 14.1 T at various B-1 power levels. Multi-B-1-Z-spectra Bloch-McConnell fits at physiological pH were further improved by the fitting of Z-spectra of five pH values simultaneously. The obtained exchange rates were used in a six-pool Bloch-McConnell simulation including a tissue-like water pool and semi-solid MT pool with different CEST and CESL presaturation pulse trains. In vivo glucose injection experiments were performed in a tumor mouse model at 7 T. Results and discussion Glucose Z-spectra could be fitted with four exchanging pools at 0.66, 1.28, 2.08 and 2.88 ppm. Corresponding hydroxyl exchange rates could be determined at pH = 7.2, T = 37 degrees C and 1X PBS. Simulation of saturation transfer for this glucose system in a gray matter-like and a tumor-like system revealed optimal pulses at different field strengths of 9.4, 7 and 3 T. Different existing sequences and approaches are simulated and discussed. The optima found could be experimentally verified in an animal model at 7 T. Conclusion For the determined fast exchange regime, presaturation pulses in the spin-lock regime (long recover time, short yet strong saturation) were found to be optimal. This study gives an estimation for optimization of the glucoCEST signal in vivo on the basis of glucose exchange rate at physiological conditions.

Breast Cancer is the most diffuse cancer among women and the treatment outcome is largely determined by its early detection. MRI at fixed magnetic field is already widely used for cancer detection. Herein it is shown that the acquisition of proton T1 at different magnetic fields adds further advantages. In fact, Fast Field Cycling Nuclear Magnetic Resonance Dispersion (FFC-NMRD) profiles have been shown to act as a high –sensitivity tool for cancer detection and staging in ex vivo murine breast tissues collected from Balb/NeuT mice. From NMRD profiles it was possible to extract two new cancer biomarkers, namely: (i) the appearance of 14N-quadrupolar peaks (QPs) reporting on tumor onset and (ii) the slope of the NMRD profile reporting on the progression of the tumor. By this approach it was possible to detect the presence of tumor in transgenic NeuT mice at a very early stage (5–7 weeks), when the disease is not yet detectable by using conventional high field (7 T) MRI and only minimal abnormalities are present in histological assays. These results show that, NMRD profiles may represent a useful tool for early breast cancer detection and for getting more insight into an accurate tumor phenotyping, highlighting changes in composition of the mammary gland tissue (lipids/proteins/water) occurring during the development of the neoplasia.

The work aimed at developing a novel MRI-based theranostic protocol for improving the anticancer efficacy of a Doxil-like liposomal formulation. The goal was achieved stimulating the intratumor release of the drug from the nanocarrier and favoring its diffusion in the lesion by the sequential application of low-intensity pulsed ultrasound. The protocol was tested on mice bearing a syngeneic breast cancer model. The combination of acoustic waves with different characteristics allowed for: i) the release of the drug and the co-encapsulated MRI agent (Gadoteridol) from the liposomes in the vessels of the tumor region, and ii) the extravasation of the released material, as well as intact liposomes, in the tumor stroma. The MR-T1 contrast enhancement measured in the tumor reported on the delivery and US-triggered release of Doxorubicin. The developed protocol resulted in a marked increase in the intratumor drug concentration that, in turn, led to the complete regression of the lesion. The protocol has a good clinical translatability because all the components of the theranostic agent (Doxorubicin, liposomes, Gadoteridol) are approved for human use.

Tumour progression depends on several sequential events that include the microenvironment remodelling processes and the switch to the angiogenic phenotype, leading to new blood vessels recruitment. Non-invasive imaging techniques allow the monitoring of functional alterations in tumour vascularity and cellularity. The aim of this work was to detect functional changes in vascularisation and cellularity through Dynamic Contrast Enhanced (DCE) and Diffusion Weighted (DW) Magnetic Resonance Imaging (MRI) modalities during breast cancer initiation and progression of a transgenic mouse model (BALB-neuT mice). Histological examination showed that BALB-neuT mammary glands undergo a slow neoplastic progression from simple hyperplasia to invasive carcinoma, still preserving normal parts of mammary glands. DCE-MRI results highlighted marked functional changes in terms of vessel permeability (K(trans) , volume transfer constant) and vascularisation (vp , vascular volume fraction) in BALB-neuT hyperplastic mammary glands if compared to BALB/c ones. When breast tissue progressed from simple to atypical hyperplasia, a strong increase in DCE-MRI biomarkers was observed in BALB-neuT in comparison to BALB/c mice (K(trans) = 5.3 ± 0.7E-4 and 3.1 ± 0.5E-4; vp = 7.4 ± 0.8E-2 and 4.7 ± 0.6E-2 for BALB-neuT and BALB/c, respectively) that remained constant during the successive steps of the neoplastic transformation. Consistent with DCE-MRI observations, microvessel counting revealed a significant increase in tumour vessels. Our study showed that DCE-MRI estimates can accurately detect the angiogenic switch at early step of breast cancer carcinogenesis. These results support the view that this imaging approach is an excellent tool to characterize microvasculature changes, despite only small portions of the mammary glands developed neoplastic lesions in a transgenic mouse model.

Objective: The early embryo implantation is characterized by enhanced uterine vascular permeability at the site of blastocyst attachment, followed by extracellular-matrix remodeling and angiogenesis. Two TG (transglutaminase) isoenzymes, TG2 (tissue TG) and FXIII (factor XIII), catalyze covalent cross-linking of the extracellular-matrix. However, their specific role during embryo implantation is not fully understood. Approach and Results: For mapping the distribution as well as the enzymatic activities of TG2 and FXIII towards blood-borne and resident extracellular-matrix substrates, we synthetized selective and specific low molecular weight substrate analogs for each of the isoenzymes. The implantation sites were challenged by genetically modifying the trophoblast cells in the outer layer of blastocysts, to either overexpress or deplete TG2 or FXIII, and the angiogenic response was studied by dynamic contrast–enhanced-magnetic resonance imaging. Dynamic contrast–enhanced-magnetic resonance imaging revealed a decrease in the permeability of decidual vasculature surrounding embryos in which FXIII were overexpressed in trophoblast cell. Reduction in decidual blood volume fraction was demonstrated when either FXIII or TG2 were overexpressed in embryonic trophoblast cell and was elevated when trophoblast cell was depleted of FXIII. These results were corroborated by histological analysis. Conclusions: In this study, we report on the isoenzyme-specific roles of TG2 and FXIII during the early days of mouse pregnancy and further reveal their involvement in decidual angiogenesis. Our results reveal an important magnetic resonance imaging-detectable function of embryo-derived TG2 and FXIII on regulating maternal angiogenesis during embryo implantation in mice.Visual Overview: An online visual overview is available for this article.

Summary MRI contrast agents (CAs) feature coordinated water molecule(s) (aqua ligands), which renders them unsuitable for magnetic resonance (MR) temperature mapping because of their resulting sensitivity to metabolic and physiological changes and/or their tendency to release toxic Gd3+ cations. Herein, we introduce an approach to temperature mapping based upon a coordinatively saturated gadolinium (Gd)-based metal-organic framework (MOF) that exhibits enhanced proton relaxation and high temperature sensitivity. The stable, non-toxic Gd zeolite-like MOF Gd-ZMOF was observed to generate a large enhancement in contrast as a result of a large (70%) contribution from second-sphere water relaxivity. Temperature mapping by clinical CAs and Gd-ZMOF by means of longitudinal (T1) relaxivity was investigated. Gd-ZMOF enabled the visualization of small temperature changes, especially in the thermal therapy region (41°C–45°C). In vivo thermal imaging demonstrates the feasibility of Gd-ZMOF as an MR thermometer and as a potential theranostic.

The discussed topic about gadolinium-based contrast agents (GBCA) safety has recently been revived due to the evidence of hyperintensities observed in the dentate nucleus (DN) and globus pallidus (GP) in the brain of patients with normal kidney function. Several preclinical studies have been conducted to understanding how the use of GBCAs can promote the gadolinium deposition in the brain. Here, we evaluate the impact of chronic cerebral hypoperfusion on gadolinium presence. T1 hyperintensities and BBB integrity were evaluated by MRI in chronically hypoperfused and healthy rats injected with either gadodiamide or hypertonic saline. Additionally, the assessment of glucose metabolism by PET imaging and the gadolinium content by ICP-MS was performed after the last MR scan. Chronically hypoperfused rats displayed a greater MRI T1w signal in the DCN and hippocampus compared to Sham-operated animals, suggesting gadolinium accumulation. Dynamic contrast-enhanced (DCE) MRI assessment of BBB permeability revealed loss of integrity (high Ktrans) after rat injury in the dentate nuclei and hippocampus. Ex vivo tissue analysis showed greater gadolinium retention in the cerebellum and subcortical regions, supporting the imaging finding. FDG-PET imaging of the cerebellum did not reveal abnormal uptake in the DCN after chronic cerebral hypoperfusion. Higher signal intensity followed by higher Gd concentration observed in DCN and hippocampus of animals subjected to cerebral injury can be associated with an increase in BBB permeability due to the applied vascular dementia animal model. Nonetheless, no glucose metabolism abnormalities were detected in chronically hypoperfused cerebellum.

The synthesis of poly[N,N-bis(3-aminopropyl)glycine] (PAPGly) dendrons Gd-based contrast agents (GdCAs) via an orthogonal protection of the different functional groups and an activation/coupling strategy wherein a specific number of synthetic steps add a generation to the existing dendron has been described. The aim of this protocol is to build up two different generations of dendrons (G-0 or dendron's core, and G-1) with peripheral NH2 groups to conjugate a 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (DO3A) derivative and afterwards to chelate with Gd3+ paramagnetic ions. These complexes, which have a well-defined molecular weight, are of relevance to MRI as an attempt to gain higher H-1 relaxivity by slowing down the rotation of molecule compared to monomeric Gd(III) complexes used as contrast agents and to increase the number of paramagnetic centers present in one molecular structure. From the study of their water H-1 longitudinal relaxation rate at different magnetic fields (NMRD, Nuclear Magnetic Relaxation Dispersion) and by evaluating the variable temperature O-17-NMR data we determined the parameters characterizing the water exchange rate and the rotational correlation time of each complex, both affecting H-1 relaxivity. Furthermore, these two novel PAPGly GdCAs were objects of i) an in vivo study to determine their biodistributions in healthy C57 mice at several time points, and ii) the Dynamic Contrast-Enhanced MRI (DCE-MRI) approach to assess their contrast efficiency measured in the tumor region of C57BL/6 mice transplanted subcutaneously with B16-F10 melanoma cells. The aim of the comparison of these two dendrons GdCAs, having different molecular weights (MW), is to understand how MW and relaxivity may influence the contrast enhancement capabilities in vivo at low magnetic field (1 T). Significant contrast enhancement was observed in several organs (vessel, spleen and liver), already at 5 min post-injection, for the investigated CAs. Moreover, these CAs induced a marked contrast enhancement in the tumor region, thanks to the enhanced permeability retention effect of those macromolecular structures.

Due to the implication of altered metabolism in a large spectrum of tissue function and disease, assessment of metabolic processes becomes essential in managing health. In this regard, imaging can play a critical role in allowing observation of biochemical and physiological processes. Nuclear imaging methods, in particular positron emission tomography, have been widely employed for imaging metabolism but are mainly limited by the use of ionizing radiation and the sensing of only one parameter at each scanning session. Observations in healthy individuals or longitudinal studies of disease could markedly benefit from non-ionizing, multi-parameter imaging methods. We therefore focus this review on progress with the non-ionizing radiation methods of MRI, hyperpolarized magnetic resonance and magnetic resonance spectroscopy, chemical exchange saturation transfer, and emerging optoacoustic (photoacoustic) imaging. We also briefly discuss the role of nuclear and optical imaging methods for research and clinical protocols.

The dentate nuclei of the cerebellum are the areas where gadolinium predominantly accumulates. It is not yet known whether gadolinium deposition affects brain functions.To assess whether gadolinium-dependent high signal intensity of the cerebellum on TObservational, cross-sectional.Fifteen patients affected by Crohn's disease were compared with 16 healthy age- and gender-matched control subjects. All participants underwent neurological, neurocognitive-psychological assessment, and blood sampling.1.5-T magnet blood oxygenation level-dependent (BOLD) functional MRI.High signal intensity on TAn unpaired two-sample t-test (age and sex were nuisance variables) was used to investigate between-group differences in cerebellar and dentate nucleus functional connectivity. Z-statistical images were set using clusters determined by Z 2.3 and a familywise error (FWE)-corrected cluster significance threshold of P = 0.05.Dentate nuclei RSFC was not different (P = n.s.) between patients with gadolinium-dependent high signal intensity on TCrohn's disease patients with gadolinium-dependent hyperintense dentate nuclei on unenhanced T2 Technical Efficacy Stage: 5 J. Magn. Reson. Imaging 2019;50:445-455.

The perturbation on the relaxation of solvent water protons induced by the presence of paramagnetic metal-containing systems has been exploited for the design of contrast agents in magnetic resonance imaging applications. Complexes of Gd3+ ions were identified early as the candidates of choice and several, highly stable, Gd-containing contrast agents entered into clinical practice two to three decades ago. Overall, the relationships between the structure and dynamics of paramagnetic metal complexes and the determinants of the proton relaxation enhancement have been well defined. On this basis a number of Gd-containing complexes have been synthetized and tested. Besides the currently used procedures devoted to visualizing tumor perfusion or abnormalities in organ function, Gd-containing contrast agents have found application in quantitative assessments such as DCE-MRI (dynamic contrast enhanced-MRI). At preclinical level, a number of paramagnetic systems have already been proposed as targeting agents of overregulated receptors/transporters or as responsive agents to tissue microenvironment parameters. These studies pave the way to molecular imaging studies with MRI modality.

The aim of this work was to investigate, by five different administration protocols, the impact of the dosage, the time passed after the last injection and the frequency of injections, on accumulation and distribution of Gd-containing species in the body tissues of healthy mice upon repeated injections of Gadolinium Based Contrast Agents (GBCAs). Gadodiamide and Gadoteridol have been compared. The amount of Gd retained in several tissues/organs (cerebrum, cerebellum, spleen, liver, kidneys, eyes, skin, bone and muscle) has been assessed by ICP-MS upon administration of the GBCAs i) at three weeks or three months after the last administration, ii) when one, three or twelve doses of GBCA were administered and iii) when administrations were made every two weeks. Gd was found in all tissues after the administration of Gadodiamide. Conversely, in the case of Gadoteridol, Gd was detected only in spleen, kidneys, liver and bone. The amounts of Gd found in spleen, liver and kidneys markedly decrease upon increasing the time that has passed after the last administration, whereas, in the case of Gadodiamide, the decrease of Gd found in bone, cerebrum and cerebellum appears to occur at a much slower rate. Overall, areas of long term deposition appear to be bone and spleen for both GBCAs. In conclusion, our findings demonstrate that intravenous multiple administrations of GBCAs is associated with extensive multiorgan retention which is reduced but not eliminated by the use of the macrocyclic Gadoteridol as well as by adopting reduced and/or less frequent dosing.

In glioma, the acidification of the extracellular tumor microenvironment drives proliferation, angiogenesis, immunosuppression, invasion and chemoresistance. Therefore, quantification of glioma extracellular pH (pHe) is of crucial importance. This study is focused on the application of the YbHPDO3A (ytterbium 1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane) probe for in vivo glioma pHe quantification using chemical exchange saturation transfer (CEST)-MRI and its correlation with tumor metabolism assessed by immunohistochemistry. The U87 glioma mouse model was used (n = 18) and MRI performed at 4.7 T. CEST-MRI of YbHPDO3A solutions at different pH values showed two resolved CEST spectra at 71 ppm and 99 ppm, both sensitive to pH variations, allowing therefore calculation of the ratiometric curve for in vivo pH quantification. In vivo MRI sequences consisted of T2w for tumor localization, T2w * to assess YbHPDO3A biodistribution by exploiting its magnetic susceptibility effect and CEST for glioma pHe mapping. T2w * images show that YbHPDO3A extravasates in tumor in regions with damaged blood-brain barrier. The pHe is calculated only in these regions. Hematoxylin/eosin histology and Ki-67, CA-IX (carbonic anhydrase 9) and NHE-1 immunohistochemical staining were performed; their expression rates were compared with the in vivo pHe values. On the basis of the cell proliferation marker Ki-67, two groups were defined: one group with a lower mitotic index (MI% 20% and an acidic pHe of 6.6 (high-proliferation/low-pH group). CA-IX and NHE-1 were over-expressed in the high-proliferation/low-pH group (CA-IX, 92 ± 7% versus 30 ± 13%; NHE-1, 84 ± 8% versus 35 ± 11%), indicating an acidic/hypoxic microenvironment. These immunohistochemical results are consistent with our pHe mapping (Pearson correlation coefficient > 0.70) and provide evidence for the feasibility of the CEST-MRI method with the YbHPDO3A probe for glioma pHe quantification at 4.7 T. Importantly, the YbHPDO3A probe has similar chemical and biological properties to the clinically approved MRI contrast agent GdHPDO3A. This makes the method promising for a clinical translation.

The aim of this study is to develop an innovative imaging guided approach based on Boron Neutron Capture Therapy, for the treatment of mesothelioma, assisted by the quantification of the in vivo boron distribution by MRI. The herein reported results demonstrate that overexpressed Low Density Lipoproteins receptors can be successfully exploited to deliver to mesothelioma cells a therapeutic dose of boron (26 μg/g), significantly higher than in the surrounding tissue (3.5 μg/g). Boron and Gd cells uptake was assessed by ICP-MS and MRI on two mesothelioma (ZL34, AE17) and two healthy (MRC-5 and NMuMg) cell lines. An in vivo model was prepared by subcutaneous injection of ZL34 cells in Nu/Nu mice. After irradiation with thermal neutrons, tumor growth was evaluated for 40 days by MRI. Tumor masses of boron treated mice showed a drastic reduction of about 80-85%. The obtained results appear very promising providing patients affected by this rare disease with an improved therapeutic option, exploiting LDL transporters.

From the early days of CEST agents' disclosure, it was evident that their potential for in vivo applications was strongly hampered by the intrinsic low sensitivity. Therefore, much work has been devoted to seek out suitable routes to achieve strong CEST contrast enhancement. The use of nanosized systems turned out to be a strategic choice, because a very large amount of CEST agents can be delivered at the site of interest. However, the breakthrough innovation in term of increase of sensitivity was found by designing the lipoCEST agents. The naturally inspired, liposomes vesicles, when loaded with paramagnetic lanthanide-based shift reagents, can be transformed into CEST probes. The large number of water molecules entrapped inside the inner cavity of the nanovesicles represents an enormous pool of exchanging protons for the generation of CEST contrast, whereas the presence of the shift reagent increases the separation in chemical shift of their nuclear magnetic resonance signal from that of the bulk water, thus allowing for a proper exchange regime for the activation of CEST contrast. From lipoCEST, it has been rather straightforward to evolve to cellCEST in order to exploit the cytoplasmatic water molecules as source of the CEST effect, once cells have been loaded with the proper shift reagent. The red blood cells were found to be particularly suitable for the development of the cellCEST concept. Finally, an understanding of the main determinants of the CEST effects in nanosized and cellular-sized agents has allowed the design of innovative lipoCEST/RBC aggregates for potential theranostic applications. WIREs Nanomed Nanobiotechnol 2016, 8:602-618. doi: 10.1002/wnan.1385 For further resources related to this article, please visit the WIREs website.