Concentrations of Gd–BOPTA in cholestatic fatty rat livers: role of transport functions through membrane proteins

Gd–BOPTA (gadobenate dimeglumine) is a magnetic resonance (MR) contrast agent that, after i.v. administration, distributes within the extracellular space, enters rat hepatocytes through the sinusoidal transporters organic anion transporting peptides (Oatps) and is excreted unchanged into bile through the multidrug resistance‐associated protein 2 (Mrp2). It is unclear how the hepatobiliary contrast agent would accumulate in cholestatic fatty livers from obese rats with bile flow impairment. Indeed, the expression of both Oatps and Mrp2 transporters is decreased in cholestatic hepatocytes. To assess this question, we measured on‐line the hepatic concentrations of ¹⁵³Gd–BOPTA with a gamma probe placed over perfused rat livers. During the perfusion of ¹⁵³Gd–BOPTA, we obtained a similar maximal hepatic concentration in normal and fatty livers despite the decreased expression and function of membrane transporters in fatty livers. By pharmacokinetic modeling and mathematical simulations, we show how changes of transport into and out of hepatocytes modify the concentrations of ¹⁵³Gd–BOPTA within hepatocytes. Mathematical simulations help to understand how each parameter (entry into hepatocytes, bile excretion, or efflux back to sinusoids) interferes with the hepatic concentrations. The hepatic concentrations of ¹⁵³Gd–BOPTA within hepatocytes rely on the entry into hepatocytes through the sinusoidal membrane and on two paths of exit, the efflux back to sinusoids and the elimination into bile. Understanding how ¹⁵³Gd–BOPTA accumulates in hepatocytes is then complex. However, such understanding is important to analyze liver imaging with hepatobiliary contrast agents in cholestatic fatty livers. Copyright © 2012 John Wiley & Sons, Ltd.     

Cellular uptake and imaging studies of gadolinium‐loaded single‐walled carbon nanotubes as MRI contrast agents

We quantify here, for the first time, the intracellular uptake (J774A.1 murine macrophage cells) of gadolinium‐loaded ultra‐short single‐walled carbon nanotubes (gadonanotubes or GNTs) in a 3 T MRI scanner using R₂ and R₂* mapping in vitro. GNT‐labeled cells exhibited high and linear changes in net transverse relaxations (ΔR₂ and ΔR) with increasing cell concentration. The measured ΔR₂* were about three to four times greater than the respective ΔR₂ for each cell concentration. The intracellular uptake of GNTs was validated with inductively coupled plasma optical emission spectrometry (ICP‐OES), indicating an average cellular uptake of 0.44 ± 0.09 pg Gd per cell or 1.69 × 10⁹ Gd³⁺ ions per cell. Cell proliferation MTS assays demonstrated that the cells were effectively labeled, without cytotoxicity, for GNTs concentrations ≤28 µM Gd. In vivo relaxometry of a subcutaneously‐injected GNT‐labeled cell pellet in a mouse was also demonstrated at 3 T. Finally, the pronounced R₂* effect of GNT‐labeled cells enabled successful in vitro visualization of labeled cells at 9.4 T. Copyright © 2010 John Wiley & Sons, Ltd.     

Nano assembly and encapsulation; a versatile platform for slowing the rotation of polyanionic Gd3+‐based MRI contrast agents

Encapsulating discrete Gd³⁺ chelates in nano‐assembled capsules (NACs) is a simple and effective method of preparing an MRI contrast agent capable of delivering a large payload of high relaxivity imaging agent. The preparation of contrast agent containing NACs had previously focussed on preparations incorporating GdDOTP^5‐ into the internal aggregate. In this report we demonstrate that other Gd³⁺ chelates bearing overall charges as low as 2‐ can also be used to prepare NACs. This discovery opens up the possibility of using Gd³⁺ chelates that have inner‐sphere water molecules that could further increase the relaxivity enhancement associated with the long τ_R that arises from encapsulation. However, encapsulation of the q = 1 chelate GdDTPA^2‐ did not give rise to a significant increase in relaxivity relative to encapsulation of the outer‐sphere chelate GdTTHA^3‐. This leads us to the conclusion that in the NAC interior proton transport is not mediated by movement of whole water molecules and the enhanced relaxivity of Gd³⁺ chelate encapsulated within NACs arises primarily from second sphere effects. Copyright © 2015 John Wiley & Sons, Ltd.     

Main applications of hybrid PET‐MRI contrast agents: a review

In medical imaging, the continuous quest to improve diagnostic performance and optimize treatment strategies has led to the use of combined imaging modalities. Positron emission tomography (PET) and computed tomography (CT) is a hybrid imaging existing already for many years. The high spatial and contrast resolution of magnetic resonance imaging (MRI) and the high sensitivity and molecular information from PET imaging are leading to the development of this new hybrid imaging along with hybrid contrast agents. To create a hybrid contrast agent for PET‐MRI device, a PET radiotracer needs to be combined with an MRI contrast agent. The most common approach is to add a radioactive isotope to the surface of a small superparamagnetic iron oxide (SPIO) particle. The resulting agents offer a wide range of applications, such as pH variation monitoring, non‐invasive angiography and early imaging diagnosis of atherosclerosis. Oncology is the most promising field with the detection of sentinel lymph nodes and the targeting of tumor neoangiogenesis. Oncology and cardiovascular imaging are thus major areas of development for hybrid PET‐MRI imaging systems and hybrid contrast agents. The aim is to combine high spatial resolution, high sensitivity, morphological and functional information. Future prospects include the use of specific antibodies and hybrid multimodal PET‐MRI‐ultrasound‐fluorescence imaging with the potential to provide overall pre‐, intra‐ and postoperative patient care. Copyright © 2015 John Wiley & Sons, Ltd.     

How transfer rates generate Gd‐BOPTA concentrations in rat liver compartments: implications for clinical liver imaging with hepatobiliary contrast agents

Following the injection of hepatobiliary contrast agents, MRI detects all molecules included in a region of interest but cannot estimate true concentrations in sinusoids, interstitium, hepatocytes or bile canaliculi. The aim of the study was to measure true concentrations in hepatocytes and to show how transfer rates across sinusoidal and canalicular membranes generate these concentrations. We perfused livers isolated from normal rats with 200 μM Gd‐DTPA and Gd‐BOPTA and measured clearances from sinusoids to liver and from hepatocytes to bile canaliculi or back to interstitium. We detected Gd‐BOPTA with a gamma probe and determined true concentrations in each liver compartment knowing their liver volumes. No pharmacokinetic modelling was applied. Gd‐BOPTA clearance from sinusoids to liver (2.5 ± 0.4 mL/min) was 50 times higher than that of Gd‐DTPA (0.05 ± 0.02 mL/min) when portal flow rate was 30 mL/min (p < 0.0001). Gd‐BOPTA clearance from sinusoids to liver was always superior to hepatocyte clearance, and hepatocyte Gd‐BOPTA concentrations measured by the probe increased over time. Gd‐BOPTA concentrations reached 439 ± 83 μM in hepatocytes and 15 × 700 ± 3100 μM in bile canaliculi, while concentrations in sinusoids were 200 μM. Gd‐BOPTA true concentrations in hepatocytes depend on the simultaneous clearances from sinusoids to hepatocytes and from hepatocytes to bile canaliculi and back to sinusoids. The study better defines how signal intensities are generated when hepatobiliary contrast agents are injected in clinical imaging. Copyright © 2016 John Wiley & Sons, Ltd.     

Real‐time 3D MRI of contrast agents in whole living mice

A specific mouse whole body coil and a dedicated gradient system at 4.7 T were coupled with an ultra‐fast 3D gradient echo MRI and keyhole reconstruction technique to obtain 3D whole‐body dynamic T₁‐weighted or T₂*‐weighted imaging. The technique was used to visualize the real‐time distribution of non‐targeting T₁ and T₂* contrast agent (CA) in a glioma‐bearing mouse model. T₁ dynamic contrast‐enhancement imaging was performed with a fast imaging with steady‐state precession sequence [echo time/repetition time (TE/TR), 1.32/3.7 ms] before and after CA injection (Gd–DOTA and BSA–Gd–DOTA) for 21 min. The temporal resolution was 1 image/6.5 s. T₂* imaging (TE/TR, 4/8 ms) was performed before and after iron‐based (small and ultra‐small particles of iron oxide) CA injection for 45 min. The temporal resolution was 1 image/14 s. Signal‐to‐noise ratio curves were determined in various mouse organs. The whole‐body coil and gradient systems made it possible to acquire data with sufficient and homogeneous signal‐to‐noise ratio on the whole animal. The spatial resolution allowed adequate depiction of the major organs, blood vessels and brain glioma. The distribution and the time‐course of T₁ and T₂* contrasts upon contrast agent injection were also assessed. 3D whole‐body mouse MRI is feasible at high spatial resolution in movie mode and can be applied successfully to visualize real‐time contrast agent distribution. This method should be effective in future preclinical molecular imaging studies. Copyright © 2011 John Wiley & Sons, Ltd.     

A review of responsive MRI contrast agents: 2005–2014

This review focuses on MRI contrast agents that are responsive to a change in a physiological biomarker. The response mechanisms are dependent on six physicochemical characteristics, including the accessibility of water to the agent, tumbling time, proton exchange rate, electron spin state, MR frequency or superparamagnetism of the agent. These characteristics can be affected by changes in concentrations or activities of enzymes, proteins, nucleic acids, metabolites, or metal ions, or changes in redox state, pH, temperature, or light. A total of 117 examples are presented, including ones that employ nuclei other than ¹H, which attests to the creativity of multidisciplinary research efforts to develop responsive MRI contrast agents. Copyright © 2014 John Wiley & Sons, Ltd.     

锰苄基酪氨酸乙二胺四乙酸的肝靶向性及其在小鼠肝癌模型中的运用

目的  评价锰苄基酪氨酸乙二胺四乙酸（Mn-BnO-TyrEDTA）MRI对比剂的肝靶向性及其在小鼠肝癌模型上的初步运用。方法  经尾静脉注射相同剂量的Mn-BnO-TyrEDTA和钆塞酸二钠（Gd-EOB-DTPA）,并对小鼠进行MRI动态增强扫描,持续1 h,以监测对比剂的分布和清除情况。同时在小鼠肝脏上注射肝癌细胞H22（1×10⁵/30 μL）和基质胶（25 μL）的混合液以建立肝癌模型,并对上述模型进行MRI。将钆喷葡胺作为对照组。结果  注射Mn-BnO-TyrEDTA 1 min后肝脏开始强化,信号强度在3 min时达到峰值并持续至30 min;1 min后血液和肾脏信号强度开始明显降低,3 min后可见对比剂进入膀胱。通过比较3 min归一化信噪比和对比噪声比,判断两种对比剂肝靶向的强弱：Mn-BnO-TyrEDTA的归一化信噪比和对比噪声比分别为3.07±0.38和11±4.0,Gd-EOB-DTPA的归一化信噪比和对比噪声比分别为2.06±0.11和4.4±0.4,差异有统计学意义（P<0.05）。与正常肝实质比较,小鼠肝癌模型的肝内病灶呈明显低信号。结论  Mn-BnO-TyrEDTA具有肝靶向性,并且经肝脏和肾脏双重途径清除,其在1~5 min的肝靶向性明显强于Gd-EOB-DTPA。小鼠肝癌模型的肝内病灶与周围正常肝实质具有较好的信号差异。     

Synthesis and characterization of a porphyrazine–Gd(III) MRI contrast agent and in vivo imaging of a breast cancer xenograft model

Porphyrazines (Pz), or tetraazaporphyrins, are being studied for their potential use in detection and treatment of cancer. Here, an amphiphilic Cu–Pz–Gd(III) conjugate has been prepared via azide–alkyne Huisgen cycloaddition or ‘click’ chemistry between an azide functionalized Pz and alkyne functionalized DOTA–Gd(III) analog for use as an MRI contrast agent. This agent, Cu–Pz–Gd(III), is synthesized in good yield and exhibits solution‐phase ionic relaxivity (r₁ = 11.5 mm ^?1 s^?1) that is approximately four times higher than that of a clinically used monomeric Gd(III) contrast agent, DOTA–Gd(III). Breast tumor cells (MDA‐MB‐231) associate with Cu–Pz–Gd(III) in vitro, where significant contrast enhancement (9.336 ± 0.335 contrast‐to‐noise ratio) is observed in phantom cell pellet MR images. This novel contrast agent was administered in vivo to an orthotopic breast tumor model in athymic nude mice and MR images were collected. The average T₁ of tumor regions in mice treated with 50 mg kg^?1 Cu–Pz–Gd(III) decreased relative to saline‐treated controls. Furthermore, the decrease in T₁ was persistent relative to mice treated with the monomeric Gd(III) contrast agent. An ex vivo biodistribution study confirmed that Cu–Pz–Gd(III) accumulates in the tumors and is rapidly cleared, primarily through the kidneys. Differential accumulation and T₁ enhancement by Cu–Pz–Gd(III) in the tumor's core relative to the periphery offer preliminary evidence that this agent would find application in the imaging of necrotic tissue. Copyright © 2014 John Wiley & Sons, Ltd.     

Effect of a mesitylene‐based ligand cap on the relaxometric properties of Gd(III) hydroxypyridonate MRI contrast agents

A series of new Gd(III) hydroxypyridonate complexes featuring a mesitylene (ME)‐derived ligand cap has been prepared. Relaxometric characterization reveals that the complexes tend to form large aggregates in solution with slow tumbling rates, as estimated from NMRD analysis, and unique pH‐dependent relaxivities. The solution behavior and relaxometric properties are compared with those observed for analogous TREN‐capped compounds, and the potential for use of these new ME‐capped complexes as pH‐responsive MRI contrast agents is explored. Copyright © 2009 John Wiley & Sons, Ltd.     

Serine‐ and mannose‐modified liposomal contrast agent for computed tomography: evaluation of the enhancement in rabbit liver VX‐2 tumor model

Kupffer cell imaging is a powerful tool for the detection of liver cancer. This diagnostic procedure depends on the faculty of the reticuloendothelial system (RES) which takes up foreign bodies, including small particles. The current study aimed to develop a novel RES targeting liposomal contrast agent that functionalized with serine or mannose, the moiety specifically binding to a corresponding receptor on phagocytic cells. Liposomes loaded with non‐ionic X‐ray contrast media, Iohexol, were prepared by supercritical carbon dioxide reverse‐phase evaporation method and were intravenously injected to healthy rabbits in order to evaluate the liver parenchymal enhancement in X‐ray computed tomography (CT). From 10 to 40 min after injection, the mean enhancement value of the liver parenchyma approached 45 and 34 Hounsfield units (HU) when serine‐modified iodinated liposomal contrast agent (ILCA) and mannose‐modified ILCA were applied, respectively. The tumor‐to‐liver contrast values were also evaluated after the administration of the prepared ILCA to rabbits with VX‐2 carcinoma. For serine‐modified ILCA, tumor‐to‐liver contrast was 82 HU at 1 min and >24 HU at 10–40 min; for mannose‐modified ILCA, the values were 58 HU at 0.5 min and >21 HU at 10–40 min. These vales estimated from the region of intrest and the imaging figures of liver indicate the potential of ILCA for clinical use. Copyright © 2010 John Wiley & Sons, Ltd.     

Assessment of tumor angiogenesis: dynamic contrast‐enhanced MRI with paramagnetic nanoparticles compared with Gd‐DTPA in a rabbit Vx‐2 tumor model

The purpose of this study was to evaluate the suitability of a macromolecular MRI contrast agent (paramagnetic nanoparticles, PNs) for the characterization of tumor angiogenesis. Our aim was to estimate the permeability of PNs in developing tumor vasculature and compare it with that of a low molecular weight contrast agent (Gd‐DTPA) using dynamic contrast‐enhanced MRI (DCE). Male New Zealand white rabbits (n = 5) underwent DCE MRI 12–14 days after Vx‐2 tumor fragments were implanted into the left hind limb. Each contrast agent (PNs followed by Gd‐DTPA) was evaluated using a DCE protocol and transendothelial transfer coefficient (K_i) maps were calculated using a two‐compartment model. Two regions of interest (ROIs) were located within the tumor core and hindlimb muscle and five ROIs were placed within the tumor rim. Comparisons were performed using repeated measures analysis of variance (ANOVA). The K_i values estimated using PNs were significantly lower than those obtained for Gd‐DTPA (p = 0.018). When PNs and Gd‐DTPA data were analyzed separately, significant differences were identified among tumor rim ROIs for PNs (p < 0.0001), but not for Gd‐DTPA data (p = 0.34). The mean K_i for the tumor rim was significantly greater than that of either the core or the hindlimb muscle for both contrast agents (p < 0.05 for each comparison). In summary, the extravasation of Gd‐DTPA was far greater than that of PNs, suggesting that PNs can reveal regional differences in tumor vascular permeability that are not otherwise apparent with clinical contrast agents such as Gd‐DTPA. These results suggest that PNs show potential for the noninvasive delineation of tumor angiogenesis. Copyright © 2010 John Wiley & Sons, Ltd.     

How to assess cytotoxicity of (iron oxide‐based) nanoparticles. A technical note using cationic magnetoliposomes

The range of different types of nanoparticles and their biomedical applications is rapidly growing, creating a need to thoroughly examine the effects these particles have on biological entities. One of the most commonly used nanoparticle types is iron oxide nanoparticles, which can be used as MRI contrast agents. The main research topic is the in vitro labeling of cells with iron oxide nanoparticles to render the cells detectable for MRI upon in vivo transplantation. For the correct evaluation of cell function and behavior in vivo, any effects of the nanoparticles on the cells must be completely ruled out. The present work provides a technical note where a detailed overview is given of several assays that could be useful to determine nanoparticle toxicity. The assays described focus on (i) nanoparticle internalization, (ii) immediate cell toxicity, (iii) cell proliferation, (iv) cell morphology, (v) cell functionality and (vi) cell physiology. Potential pitfalls, appropriate controls and advantages/disadvantages of the different assays are given. The main focus of this work is to provide a detailed guide to help other researchers in the field interested in setting up nanoparticle‐toxicity studies. Copyright © 2010 John Wiley & Sons, Ltd.     

Towards MRI contrast agents responsive to Ca(II) and Mg(II) ions: metal‐induced oligomerization of dota–bisphosphonate conjugates

In magnetic resonance imaging (MRI), paramagnetic complexes are utilized as contrast agents. Much attention has been paid to the development of new contrast agents responsive to pH, temperature or concentration of various components of body liquids. We report a new type of MRI probe sensing the concentrations of calcium and magnesium in biological media. The ligand do3ap^BP combines a dota‐like chelator with a bisphosphonate group. In the complex, the Gd(III ) ion is entrapped in the macrocyclic cavity whereas the bisphosphonate group is not coordinated and therefore is available for coordination with endogenous metal ions. In the presence of metal ions, Gd–do3ap^BP appears to show formation of coordination oligomers leading to an unprecedented increase in r₁ up to 200–500%. The extremely high relaxivity response makes this type of compound interesting for further studies as MRI ion‐responsive probes for biomedical research. Copyright © 2010 John Wiley & Sons, Ltd.     

Entrapment of a neutral Tm(III)‐based complex with two inner‐sphere coordinated water molecules into PEG‐stabilized vesicles: towards an alternative strategy to develop high‐performance LipoCEST contrast agents for MR imaging

Chemical exchange saturation transfer (CEST) probes issued from the encapsulation of a water proton paramagnetic shift reagent into the inner aqueous volume of lipid vesicles provide an emerging class of frequency‐selective contrast agents with huge potential in the field of molecular magnetic resonance imaging (MRI). This work deals with the generation of such LipoCEST agents properly designed to optimize, under isotonic conditions, the chemical shift offset of the intra‐liposomal water protons as well as the number of exchangeable protons under reasonably low radiofrequency (RF) fields of saturation. The strategy lies in the loading of poly(ethylene glycol)‐stabilized nanosized liposomes with uncharged lanthanide chelates, binding more than one water molecule in the first hydration sphere, exemplified here by [Tm(III)–DO3A (H₂O)₂] complex. The key properties of the probes are demonstrated by complementary NMR investigations. The residence lifetime of the water molecules coordinated to the lanthanide center was outstandingly short (9.5 ± 0.2 ns from ¹⁷O NMR), and indeed relevant for effective LipoCEST responsiveness. The ¹H NMR CEST spectra (7.01 T magnetic field) prove that the theoretically expected optimal sensitivity can be approximated in the nanomolar concentration range, at reasonably low RF presaturation pulses (6.7–12 μT) and saturation frequency offsets of the intra‐liposomal water protons beyond 10 ppm, making possible selective irradiation in biological environment. CEST‐MRI images (7.01 T magnetic field and 10–12 μT RF pulse) explicitly confirm the interest of these newly conceived LipoCEST agents, indeed among the most efficient ones developed so far under isosmotic conditions. Copyright © 2014 John Wiley & Sons, Ltd.     

Improving T1 and T2 magnetic resonance imaging contrast agents through the conjugation of an esteramide dendrimer to high‐water‐coordination Gd(III) hydroxypyridinone complexes

Commercial gadolinium magnetic resonance imaging (MRI) contrast agents are limited by low relaxivity (r₁) and coordination to only a single water molecule (q = 1). Consequently, gram quantities of these agents must be injected to obtain sufficient diagnostic contrast. In this study, MRI contrast agents for T₁ and T₂ relaxivity were synthesized using hydroxypyridinone and terephthalamide chelators with mesityl and 1,4,7‐triazacyclononane capping moieties. When covalently conjugated to a highly biocompatible esteramide dendrimer, T₂ relaxation rates up to 52 m m ^?1 s^?1 and T₁ relaxation rates up to 31 m m ^?1 s^?1 per gadolinium were observed under clinically relevant conditions. These values are believed to be brought about by using a dendritic macromolecule to decrease the molecular tumbling time of the small molecule complexes. These agents also show high aqueous solubility and low toxicity in vitro. In this study we report six new compounds: three discrete complexes and three dendrimer conjugates. Copyright © 2012 John Wiley & Sons, Ltd.