Use of Real-Life Safety Data From International Pharmacovigilance Databases to Assess the Importance of Symptoms Associated With Gadolinium Exposure.

OBJECTIVE: Recent scientific publications have reported cases of patients who complained from a variety of symptoms after they received a gadolinium-based contrast agent (GBCA). The aim of this study was to appreciate the importance of these clinical manifestations in the overall population by assessing the weight of "symptoms associated with gadolinium exposure" (SAGE) among the bulk of safety experiences reported to major health authorities. MATERIALS AND METHODS: Symptoms associated with gadolinium exposure were identified from a review of the scientific literature, and the corresponding preferred terms were searched in each system organ class (SOC) category recorded in the European and North American pharmacovigilance databases EudraVigilance (EV) and FDA Adverse Event Reporting System (FAERS), respectively. The numbers of SAGE per preferred term, and cumulatively per SOC, were recorded and their weights in the overall spectrum of adverse events (AEs) were determined for each GBCA. RESULTS: The analysis of the selected AEs revealed a significantly higher SAGE weight for gadobenate dimeglumine (EV: 25.83%, FAERS: 32.24%) than for gadoteridol (EV: 15.51%; FAERS: 21.13%) and significantly lower SAGE weights for gadobutrol (EV: 7.75%; FAERS: 13.31%) and gadoterate meglumine (EV: 8.66%; FAERS: 12.99%). A similar ranking was found for most of the SOCs except for "nervous system disorders," probably owing to a limitation in the methods of data selection. Furthermore, this analysis showed a greater percentage of reports mentioning a decrease in the quality of life of the patients when they were exposed to gadobenate dimeglumine or gadoteridol than to gadobutrol or gadoterate meglumine. CONCLUSION: This study showed that SAGE represent a significant percentage of the bulk of AEs reported to the health authorities for each GBCA. It provided real-life arguments suggesting that SAGE may be more prevalent with linear than macrocyclic GBCAs and that gadoteridol may present a higher SAGE risk than the other macrocyclic contrast agents.

Current and Future MR Contrast Agents: Seeking a Better Chemical Stability and Relaxivity for Optimal Safety and Efficacy.

This review summarizes 30 years of experience in the development and clinical use of magnetic resonance (MR) contrast agents. Despite their undisputable usefulness for disease diagnosis, gadolinium (Gd)-based contrast agents (GBCAs) have gone through 2 major safety crises. Approximately 10 years ago, the regulatory agencies decided to restrict the use of GBCAs to minimize the risk of nephrogenic systemic fibrosis in patients with severe renal insufficiency. Yet, following the recent discovery of Gd retention in brain, the same agencies adopted different positions ranging from suspension of marketing authorizations, changes in GBCA safety labeling, and performing preclinical and clinical studies to assess the potential long-term consequences of Gd accumulation on motor and cognitive functions. Besides, magnetic resonance imaging (MRI) has benefited from MR technological advances, which provide alternative solutions to increase the MR signal, generate new contrasts on MRI scans, and accelerate their acquisition and analysis. Altogether, GBCAs in combination with new MR techniques have found their place in the diagnostic pathway of various diseases. Despite the large research efforts to identify and develop alternative Gd-free MR agents, manganese- and iron-based contrast agents have failed to reach market approval. In this context, the development of next-generation MR contrast agents should focus on high-stability and high-relaxivity GBCAs, such as gadopiclenol, which offer the possibility to adapt the administered Gd dose to each indication while ensuring an optimal patient safety.

Future of Diagnostic Computed Tomography: An Update on Physicochemical Properties, Safety, and Development of X-ray Contrast Media.

Iodinated contrast media (CM) are utilized in approximately 40% of the 300 million computed tomography (CT) scans undertaken annually. This review focuses on the physicochemical properties and safety of iodinated CM, and the development of new x-ray CM, and it explores methods to optimize CT scanning parameters. It concludes that good x-ray CM should have high structural stability, hydrophilicity, and CT attenuation; low viscosity, osmolality, and protein binding; no metabolism and tissue accumulation; and a complete elimination.

Gadolinium Retention as a Safety Signal: Experience of a Manufacturer.

OBJECTIVES: The purpose of this manuscript is to review the successive regulatory actions and decisions following the initial publication by Kanda and colleagues in 2014 regarding gadolinium retention in the human brain after multiple gadolinium-based contrast agents (GBCAs) administrations. MATERIALS AND METHODS: Starting from 2014, the actions and decisions made by all regulatory authorities were collected and summarized region by region. Volumes of GBCA sales in 2018 per region and main countries are also presented as an indicator of patients' exposure to those products. RESULTS: All regulatory authorities agreed on the absence of evidence of any harmful effect of gadolinium retention in humans. However, based on the same amount of preclinical and clinical evidence available in adults and children, regulatory authorities used different approaches resulting in different actions and decisions regarding the labeling and market authorizations of GBCAs, as well as the specific actions requested to the manufacturers. CONCLUSIONS: The manufacturers of GBCAs had to face different situations according to the countries, due to the different positions and expectations from regulatory agencies. They have adapted their responses to the different positions of the regulatory agencies and conducted specific preclinical and clinical investigations to provide the expected evidence. It is also their responsibility to continuously monitor the benefit-risk balance of the products and to propose risk minimization measures to the regulatory agencies.

Revisiting the Pharmacokinetic Profiles of Gadolinium-Based Contrast Agents: Differences in Long-Term Biodistribution and Excretion.

OBJECTIVES: Gadolinium-based contrast agents (GBCAs) have been used for years for magnetic resonance imaging examinations. Because of their rapid blood clearance, they were considered as very safe products until some of them were shown to induce nephrogenic systemic fibrosis in patients with renal failure and hypersignals on T1-weighted unenhanced brain scans of patients with normal renal function. To date, these adverse effects have been related almost exclusively to the use of low-stability linear agents, which are more prone to release free gadolinium. The aim of the present meta-analysis was to ascertain the existence of a deep compartment for gadolinium storage in the body and to assess whether all the GBCAs present the same toxicokinetic profile. MATERIALS AND METHODS: Applying a systematic literature search methodology, all clinical and preclinical studies reporting time-dependent plasma concentrations and renal excretion data of gadolinium were identified and analyzed. Since the individual data were not available, the analysis focused on the average values per groups of subjects or animals, which had received a given GBCA at a given dose. The rate constants of the distribution phase (α), rapid elimination phase (ß), and residual excretion phase (γ) of gadolinium were determined in each group from the plasma concentration (Cp) time curves and the relative urinary excretion rate (rER) time curves, taking the 2-hour time point as a reference. Moreover, as bone may represent a reservoir for long-term gadolinium accumulation and slow release into the blood stream, the time curves of the relative concentration in the bone (rCB) of Gd-labeled GBCAs in mice or rats were analyzed taking day 1 concentrations as a reference. The ratio of gadolinium concentrations in the bone marrow (CBM) as compared with the bone (CB) was also calculated. RESULTS: The relative urinary excretion rate (rER) plots revealed a prolonged residual excretion phase of gadolinium in healthy volunteers, consistent with the existence of a deep compartment of distribution for the GBCAs. The rate constant γ of gadoterate meglumine (0.107 hour) is 5 times higher than that of the linear agents (0.020 ± 0.008 hour), indicating a much faster blood clearance for the macrocyclic GBCA. Similar results were obtained in the preclinical studies. A strong correlation was shown between the γ values of the different products and their respective thermodynamic stability constants (Ktherm). Greater clearance rates of Gd from murine bone were also found after gadoterate meglumine or gadoteridol injection (0.131-0.184 day) than after administration of the linear agents (0.004-0.067 day). The concentrations of Gd in the bone marrow (CBM) from animals exposed to either gadoterate meglumine or gadodiamide are higher than those in the bone (CB) for at least 24 hours. Moreover, the ratio of concentrations (CBM/CB) at 4 hours is significantly lower with the former agent than the latter (1.9 vs 6.5, respectively). CONCLUSIONS: Using a nonconventional pharmacokinetic approach, we showed that gadoterate meglumine undergoes a much faster residual excretion from the body than the linear GBCAs, a process that seems related to the thermodynamic stability of the different chelates. Gadolinium dissociation occurs in vivo for some linear chelates, a mechanism that may explain their long-term retention and slow release from bone. Potential consequences in terms of bone toxicity warrant further investigations.

Effects of gadolinium-based contrast agent concentrations (0.5 M or 1.0 M) on the diagnostic performance of magnetic resonance imaging examinations: systematic review of the literature.

Background To date there is no agreement as to what is the optimal concentration for gadolinium-based contrast agents (GBCAs). Purpose To assess whether diagnostic performance differences exist between 0.5 M and 1.0 M GBCAs used for magnetic resonance imaging (MRI). Material and Methods A PubMed literature search identified 21 clinical studies published between 2005 and 2013 which evaluated the diagnostic efficacy of both types of GBCAs. Study design, type of procedure, GBCA administration mode, imaging performances, impact on patient management, study limitations, and biases were analyzed. No statistical test was performed on pooled data. Results Sixteen comparative and five non-comparative studies were analyzed, involving 2183 patients who underwent MRI procedures for various indications. In 67% of the studies, 0.5 M and 1.0 M GBCAs were injected at equimolar gadolinium amounts per kg body weight. Only 33% applied the same molar flow rate for delivery of the GBCAs. No significant differences between GBCAs were reported for 23 out of 27 qualitative endpoints (mainly image quality, lesion, and vessel visualization) and 29 out of 40 quantitative endpoints. Three out of four studies with non-equimolar delivery rates showed better contrast-to-noise and signal-to-noise ratios for 1.0 M gadobutrol, without showing an impact on diagnostic performance. Methodological biases were identified in several studies impairing the interpretation of comparisons. Conclusion Imaging differences between 0.5 M and 1.0 M GBCAs were essentially observed under non-equimolar delivery rates. However, they did not result into greater diagnostic efficacy when performed under equimolar conditions.

Anti-inflammatory drug evaluation in ApoE-/- mice by ultrasmall superparamagnetic iron oxide-enhanced magnetic resonance imaging.

OBJECTIVES: The renin-angiotensin system and local phagocytic activity play a major role in atherosclerotic plaque development. Treatment with irbesartan, an antagonist of angiotensin II receptor, can decrease atherosclerotic lesion formation. Iron oxide-enhanced magnetic resonance imaging (MRI) can be successfully used to evaluate the phagocytic activity in the atherosclerotic plaque in mice. In this study, we used 2 iron oxide-enhanced MRI strategies, in vivo labeling by injection of iron oxide particles and injection of in vitro labeled macrophages, to investigate the effect of irbesartan on both atherosclerotic plaque size and macrophage content in apolipoprotein (Apo) E-deficient mice. MATERIALS AND METHODS: ApoE-/- female mice (C57BL/6 background; Charles-River, France) were divided into 2 groups (irbesartan treated [TG] or not treated [NTG]) and started on a high-fat diet (Harlan TD88137 Western Diet, 21% fat, 0.2% cholesterol). Animals underwent magnetic resonance examinations on a 7-T scanner at baseline and at 14 and 28 weeks of treatment. At each time point, 2 MRI sessions were performed, before and 48 hours after administration of an iron oxide agent (P904; Guerbet, France) or magnetically labeled macrophages (MФΦ). At the end of the follow-up, blood samples were taken for plasma lipid dosing and aorta samples for histology. The study was approved by the animal experimentation ethic committee of our institution.Vessel wall area measurements were performed on high-resolution spin echo transverse images. Multiecho gradient echo images acquired with the same geometry were used to calculate T2* maps of the vessel wall using a pixel-by-pixel monoexponential fit. Irbesartan effect on vessel wall area over time was assessed using a factorial analysis of variance test. T2* values of the vessel wall at pre- and post-ultrasmall superparamagnetic iron oxide (USPIO) administration were analyzed with a 1-way analysis of variance test with Bonferroni post hoc. RESULTS: Irbesartan treatment resulted in significantly smaller vessel wall areas at 28 weeks of treatment (P = 0.04). Postinjection values varied significantly over time for both the NTG-P904 (P = 0.02) and the TG-P904 (P = 0.01) groups. Furthermore, when comparing the TG-P904 with the NTG-P904 group at 28 weeks of treatment, a significant difference was obtained for both pre- and post-USPIO administration values (P = 0.01). In the labeled-macrophage group, postinjection T2* values were smaller than the preinjection ones for the NTG animals at 14 weeks of treatment. No T2* changes were observed in the TG-MΦ group.The difference between pre- and post-USPIO administration T2* values (ΔT2*) was significantly smaller in the TG-P904 group compared with the NTG-P904 group at 28 weeks of treatment. At this point, a good correlation (R = 0.7, P = 0.03) was found between the ΔT2* values in the P904 imaging group and the macrophage-covered area by immunohistological analysis. CONCLUSIONS: The present study illustrates an MRI follow-up of intraplaque macrophages using in vivo labeling by iron oxide particle injection and macrophage injection after in vitro USPIO labeling in the assessment of a therapeutic effect in a mouse model of atherosclerosis. Even though in vivo labeling is not fully specific of macrophage uptake, it enabled the detection of a treatment-related reduction in the macrophage content of atherosclerotic plaques in ApoE-/- mice.

Molecular and cellular targets of the MRI contrast agent P947 for atherosclerosis imaging.

P947 (DOTA-Gd-peptide) was recently identified as an MRI contrast agent for the detection and characterization of the matrix metalloproteinases (MMP)-rich atherosclerotic plaques. Because this product displays a broad spectrum affinity for the MMP family, we hypothesized that it may also recognize other metalloproteinases overactivated in vulnerable atherosclerotic plaques. Therefore, this study aimed at describing, at the molecular and cellular level, the interactions between P947 and proteases of atherosclerotic plaques. Fluorimetric assays were used to measure the in vitro affinity of P947 toward recombinant and purified MMPs, angiotensin-converting enzyme (ACE), endothelin-converting enzyme (ECE-1), neutral endopeptidase (NEP), and both aminopeptidases A and N (APA and APN). Using similar fluorimetric assays associated with specific substrates, enzymatic activities were measured in vulnerable and stable plaques collected from human atherosclerotic carotid arteries. Ex vivo affinity of P947 for metalloproteinases in vulnerable lesions was subsequently determined. Interaction between P947 and major cell types present in atherosclerotic plaques was also investigated in different cell lines: PMA-1-differentiated THP-1 (macrophage), Ox-LDL-treated THP-1 (foam cell), Jurkat cell line (lymphocyte), and human umbilical vein endothelial cell (HUVEC, endothelial cell). Molecular targeting of P947 was confirmed by fluorimetry, ICP-MS, and in vitro MRI approaches. Potential application of P947 for detecting atherosclerotic plaques by in vivo MRI was tested in a rabbit model of atherosclerosis. In vitro, P947 displayed affinities for purified MMPs, ACE, ECE-1, NEP, APA, and APN in the micromolar range. Interestingly, MMPs, ACE, and APN exhibited higher activities in vulnerable plaques from human atherosclerotic carotid samples, as compared to stable plaques. ECE-1, NEP, and APA had either no activity or the same low activity in both vulnerable and stable plaques. P947 showed micromolar affinities for MMPs, ACE, and APN secreted by plaque samples. Moreover, P947 bound to THP-1 macrophages and THP-1 foam cells in a concentration-dependent manner and with a higher intensity than the control contrast agents DOTA-Gd or P1135 (DOTA-Gd coupled to a scrambled peptide). In THP-1 macrophages, P947 inhibited largely (70%) and almost completely (95%) MMP and APN activities, respectively, which strongly suggested an MMP- and APN-dependent binding of P947 to these cells. This enzyme-specific binding was confirmed with in vitro MRI. Indeed, the T1 value of THP-1 cells decreased from 2.094 s (macrophages w/o P947) to 2.004 s (macrophages with 1 mM of P947). In addition, the Gd content measured by ICP-MS was 11.01 ± 1.05 fg Gd/macrophage when cells were incubated in the presence of P947 and only 5.18 ± 0.43 fg Gd/macrophage with the control product P1135. The difference of Gd concentration between both contrast agents corresponded to a specific accumulation of 5.83 fg Gd/cell, which may be detected by MRI. MR imaging in the atherosclerosis rabbit model showed enhancement of the aortic wall after P947 injection with a significant increase of CNR values from 0.21 ± 0.02 (before injection) to 0.37 ± 0.07 (after injection), demonstrating the efficacy of the contrast agent to detect the atherosclerotic plaques in vivo. Taken together, these data suggest that P947 may be an interesting contrast agent for in vivo molecular MR imaging of MMPs, ACE, and APN activities present in vulnerable atherosclerotic plaques.

Monitoring of arterial wall remodelling in atherosclerotic rabbits with a magnetic resonance imaging contrast agent binding to matrix metalloproteinases.

AIMS: P947 is a gadolinium-based magnetic resonance imaging (MRI) contrast agent with high affinity for several matrix metalloproteinases (MMPs) involved in arterial wall remodelling. We tested whether the intensity of enhancement detected in vivo in the arterial wall with P947 and MRI correlates with actual tissue MMP-related enzymatic activity measured in a rabbit atherosclerotic model subjected to dietary manipulations. METHODS AND RESULTS: Aortas of 15 rabbits in which atherosclerotic lesions were induced by balloon angioplasty and 4 months of hypercholesterolaemic diet were imaged at 'baseline' with P947-enhanced MRI. Atherosclerotic rabbits were divided into three groups: five rabbits were sacrificed ('baseline' group); five rabbits continued to be fed a lipid-supplemented diet ('high-fat' group); and five rabbits were switched from atherogenic to a purified chow diet ('low-fat' group). Four months later, a second P947-enhanced MRI was acquired in the 10 remaining rabbits. A significantly lower signal was detected in the aortic wall of rabbits from the 'low-fat' group as compared with rabbits from the 'high-fat' group (21 ± 6 vs. 46 ± 3%, respectively; P = 0.04). Such differences were not detected with the contrast agent P1135, which lacks the MMP-specific peptide sequence. In addition, the intensity of aortic wall enhancement detected with MRI after injection of P947 strongly correlated with actual MMP-2 gelatinolytic activity measured in corresponding aortic segments using zymography (r = 0.87). CONCLUSION: P947-enhanced MRI can distinguish dietary-induced variations in MMP-related enzymatic activity within plaques in an experimental atherosclerotic model, supporting its utility as a clinical imaging tool for in vivo detection of arterial wall remodelling.

Assessment of age modulated vascular inflammation in ApoE-/- mice by USPIO-enhanced magnetic resonance imaging.

OBJECTIVE: Inflammation within atherosclerotic lesions increases the risk for plaque rupture and thrombosis. A functional approach to plaque analysis is the intravenous administration of ultrasmall superparamagnetic particles of iron oxide (USPIO) that enables visualization of macrophages residing in the plaques. In this study, we sought to characterize the age-related inflammatory status associated with atherosclerosis lesion progression in ApoE mice using USPIO-enhanced magnetic resonance imaging (MRI). MATERIALS AND METHODS: A total of 24 ApoE mice were divided in 4 groups (N = 6) and were given a high cholesterol diet from 6 weeks of age to the end of the protocol. One group per MR time point was investigated at 10, 16, 24, and 34 weeks of age. Each MR examination was performed on a 4.7 T scanner and consisted of baseline and 48 hours post-USPIO administration imaging sessions. P904, a USPIO contrast agent (Guerbet, Paris, France) with a potential for plaque macrophage targeting, was used.Vessel wall area measurements were performed on high resolution spin echo transverse images. Multi-echo gradient-echo images acquired with the same geometry were used to calculate T2* maps of the vessel wall using a pixel-by-pixel monoexponential fit. A one-way analysis of variance was performed to characterize the temporal variation of vessel wall area, susceptibility artifact area, baseline, and post-USPIO T2* values. MR measurements were correlated with the histologic findings. RESULTS: A significant increase was found in the aortic wall area from 1.4 ± 0.2 at 10 weeks to 2.0 ± 0.3 mm at 34 weeks of age (P < 0.05). Concerning the post-USPIO MRI, signal loss regions, with patterns spanning from focal to the complete disappearance of the vessel wall, were observed on all postcontrast images. A significant increase in the size of the susceptibility artifact was observed from 0.5 ± 0.2 to 2.4 ± 1.0 at 24 weeks (P < 0.05) and to 2.0 ± 0.9 mm at 34 weeks (P < 0.05).The T2* values calculated on the 48 hours post-USPIO images were shorter compared with baseline. The decrease was 34% ± 16% at 10 weeks, 57% ± 11% at 16 weeks, 57% ± 16% at 24 weeks, and 48% ± 13% at 34 weeks.The Pearson's correlation test between measurement of aortic wall area performed on both MR images and histologic analysis showed a statistically significant correlation (r = 0.695 and P < 0.05). A correlation was also obtained between the signal loss area and the macrophages covered area (r = 0.68 and P < 0.05). CONCLUSIONS: This study demonstrated the feasibility of USPIO-enhanced MRI in assessing the inflammatory status related to the temporal progression of the atherosclerosis plaque in ApoE transgenic mice model of atherosclerosis. In our experimental conditions, the vascular inflammation peak, for the ApoE mice feeding high-fat/high-cholesterol diet is measured between 16 and 24 weeks of age.

In vivo evaluation of a new magnetic resonance imaging contrast agent (P947) to target matrix metalloproteinases in expanding experimental abdominal aortic aneurysms.

OBJECTIVE: Abdominal aortic aneurysm (AAA) rupture is a devastating event, and development of noninvasive methods to detect AAA at risk is needed. Matrix metalloproteinases (MMPs) play a major role in AAA growth and their subsequent rupture. This study was aimed to evaluate the ability of P947, a recently developed magnetic resonance imaging (MRI) contrast agent, to target MMPs in vivo in expanding experimental AAAs. MATERIALS AND METHODS: AAAs were induced in Wistar rats (n = 18) by perfusion of a segment of the abdominal aorta with porcine elastase. After 5 or 6 days of elastase perfusion, when the aortic segment was expanding and showed inflammation with high MMP levels, rats were injected either with P947 (n = 6), P1135, a scramble form of P947 (n = 6), or with the reference contrast agent Gadolinium-DOTA (Gd-DOTA) (n = 3). Sham-operated rats (n = 3) were injected with P947 as controls. Imaging was performed on the animals using a 1.5T MRI scanner before and at different times after injection of contrast agents (100 µmol/kg). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gelatin zymography of culture media conditioned by incubation with perfused aortic segment or control TA from elastase-perfused rats (n = 3) was performed to determine levels of MMP2 and MMP9. In addition, in situ gelatin zymography was used to localize these active MMPs on frozen histologic sections. RESULTS: The normalized signal enhancement determined on MRI images was higher in the perfused aortic segment of rats injected with P947 (162%) than in rats injected with P1135 (100%) or Gd-DOTA (117%) (P < 0.01 using the Friedman test) from 5 to 125 minutes after injection. The area of contrast enhancement on MRI images colocalized with the fluorescence generated by MMPs in the AAA inflammatory area, as detected by in situ zymography on histologic sections. CONCLUSION: Our data showed that MRI using P947 allows detection of MMP activity within the inflammatory wall of experimental AAAs, thus representing a potential noninvasive method to detect AAAs with a high risk of rupture.

Magnetic resonance molecular imaging of thrombosis in an arachidonic acid mouse model using an activated platelet targeted probe.

OBJECTIVE: Atherosclerotic plaque rupture leads to acute thrombus formation and may trigger serious clinical events such as myocardial infarction or stroke. Therefore, it would be valuable to identify atherothrombosis and vulnerable plaques before the onset of such clinical events. We sought to determine whether the noninvasive in vivo visualization of activated platelets was effective when using a target-specific MRI contrast agent to identify thrombi, hallmarks of vulnerable or high-risk atherosclerotic plaques. METHODS AND RESULTS: Inflammatory thrombi were induced in mice via topical application of arachidonic acid on the carotid. Thrombus formation was imaged with intravital fluorescence microscopy and molecular MRI. To accomplish the latter, a paramagnetic contrast agent (P975) that targets the glycoprotein alpha(IIb)beta(3), expressed on activated platelets, was investigated. The specificity of P975 for activated platelets was studied in vitro. In vivo, high spatial-resolution MRI was performed at baseline and longitudinally over 2 hours after injecting P975 or a nonspecific agent. The contralateral carotid, a sham surgery group, and a competitive inhibition experiment served as controls. P975 showed a good affinity for activated platelets, with an IC(50) (concentration of dose that produces 50% inhibition) value of 2.6 micromol/L. In thrombosed animals, P975 produced an immediate and sustained increase in MRI signal, whereas none of the control groups revealed any significant increase in MRI signal 2 hours after injection. More important, the competitive inhibition experiment with an alpha(IIb)beta(3) antagonist suppressed the MRI signal enhancement, which is indicative for the specificity of P975 for the activated platelets. CONCLUSIONS: P975 allowed in vivo target-specific noninvasive MRI of activated platelets.

Characterization of gadolinium-based dynamic susceptibility contrast perfusion measurements in permanent and transient MCAO models with volumetric based validation by CASL.

Perfusion imaging is crucial in imaging of ischemic stroke to determine 'tissue at risk' for infarction. In this study we compared the volumetric quantification of the perfusion deficit in two rat middle-cerebral-artery occlusion (MCAO) models using two gadolinium-based contrast agents (P1152 (Guerbet) and Magnevist (Bayer-Schering, Pittsburgh, PA, USA)) as compared with our well established continuous arterial spin labeling (CASL) perfusion imaging technique. Animals underwent either permanent MCAO or transient MCAO with 80-min reperfusion. Imaging was performed at four different time points after MCAO. A region-of-interest (ROI) analysis of the subregions of the ischemic zone (core, penumbra, transient reversal (TR), and sustained reversal (SR)) using P1152 showed significant reduction in blood flow in the core and TR subregions relative to the penumbral and SR subregions while occluded. After reperfusion, a significant increase in blood flow was recorded at all time points after reperfusion in all regions except TR. From the ROI analysis the threshold for the penumbra was determined to be -62+/-11% and this value was subsequently used for quantification of the volumetric deficit. The ischemic volume as defined by dynamic susceptibility contrast (DSC), was only statistically different from the CASL-derived ischemic volume when using Magnevist at post-reperfusion time points.

Peptidic targeting of phosphatidylserine for the MRI detection of apoptosis in atherosclerotic plaques.

Molecular and cellular imaging of atherosclerosis has garnered more interest at the beginning of the 21st century, with aims to image in vivo biological properties of plaque lesions. Apoptosis seems an attractive target for the diagnosis of vulnerable atherosclerotic plaques prone to a thrombotic event. The aim of the present work was to screen for apoptosis peptide binders by phage display with the final purpose to detect apoptotic cells in atherosclerotic plaques by magnetic resonance imaging (MRI). A phosphatidylserine-specific peptide identified by phage display was thus used to design an MRI contrast agent (CA), which was evaluated as a potential in vivo reporter of apoptotic cells. A library of linear 6-mer random peptides was screened in vitro against immobilized phosphatidylserine. Phage DNA was isolated and sequenced, and the affinity of peptides for phosphatidylserine was evaluated by enzyme-linked immunosorbent assay. The phosphatidylserine-specific peptide and its scrambled homologue were attached to a linker and conjugated to DTPA-isothiocyanate. The products were purified by dialysis and by column chromatography and complexed with gadolinium chloride. After their evaluation using apoptotic cells and a mouse model of liver apoptosis, the phosphatidylserine-targeted CA was used to image atherosclerotic lesions on ApoE(-/-) transgenic mice. Apoptotic cells were detected on liver and aorta specimens by the immunostaining of phosphatidylserine and of active caspase-3. Sequencing of the phage genome highlighted nine different peptides. Their alignment with amino acid sequences of relevant proteins revealed a frequent homology with Ca2+ channels, reminiscent of the function of annexins. Alignment with molecules involved in apoptosis provides a direct correlation between peptide selection and utility. The in vivo MRI studies performed at 4.7 T provide proof of concept that apoptosis-related pathologies could be diagnosed by MRI with a low molecular weight paramagnetic agent. The new CA could have real potential in the diagnosis and therapy monitoring of atherosclerotic disease and of other apoptosis-associated pathologies, such as cancer, ischemia, chronic inflammation, autoimmune disorders, transplant rejection, neurodegenerative disorders, and diabetes mellitus. The phage display-derived peptide could also play a potential therapeutic role through anticoagulant activity by mimicking the role of annexin V, the endogenous ligand of phosphatidylserine.