Comprehensive Speciation Analysis of Residual Gadolinium in Deep Cerebellar Nuclei in Rats Repeatedly Administered With Gadoterate Meglumine or Gadodiamide.

PURPOSE: Several preclinical studies have reported the presence of gadolinium (Gd) in different chemical forms in the brain, depending on the class (macrocyclic versus linear) of Gd-based contrast agent (GBCA) administered. The aim of this study was to identify, with a special focus on insoluble species, the speciation of Gd retained in the deep cerebellar nuclei (DCN) of rats administered repeatedly with gadoterate or gadodiamide 4 months after the last injection. METHODS: Three groups (N = 6/group) of healthy female Sprague-Dawley rats (SPF/OFA rats; Charles River, L'Arbresle, France) received a cumulated dose of 50 mmol/kg (4 daily intravenous administrations of 2.5 mmol/kg, for 5 weeks, corresponding to 80-fold the usual clinical dose if adjusted for man) of gadoterate meglumine (macrocyclic) or gadodiamide (linear) or isotonic saline for the control group (4 daily intravenous administrations of 5 mL/kg, for 5 weeks). The animals were sacrificed 4 months after the last injection. Deep cerebellar nuclei were dissected and stored at -80°C before sample preparation. To provide enough tissue for sample preparation and further analysis using multiple techniques, DCN from each group of 6 rats were pooled. Gadolinium species were extracted in 2 consecutive steps with water and urea solution. The total Gd concentrations were determined by inductively coupled plasma mass spectrometry (ICP-MS). Soluble Gd species were analyzed by size-exclusion chromatography coupled to ICP-MS. The insoluble Gd species were analyzed by single-particle (SP) ICP-MS, nanoscale secondary ion mass spectroscopy (NanoSIMS), and scanning transmission electron microscopy with energy-dispersive X-ray spectroscopy (STEM-EDX) for elemental detection. RESULTS: The Gd concentrations in pooled DCN from animals treated with gadoterate or gadodiamide were 0.25 and 24.3 nmol/g, respectively. For gadoterate, the highest amount of Gd was found in the water-soluble fractions. It was present exclusively as low-molecular-weight compounds, most likely as the intact GBCA form. In the case of gadodiamide, the water-soluble fraction of DCN was composed of high-molecular-weight Gd species of approximately 440 kDa and contained only a tiny amount (less than 1%) of intact gadodiamide. Furthermore, the column recovery calculated for this fraction was incomplete, which suggested presence of labile complexes of dissociated Gd3+ with endogenous molecules. The highest amount of Gd was detected in the insoluble residue, which was demonstrated, by SP-ICP-MS, to be a particulate form of Gd. Two imaging techniques (NanoSIMS and STEM-EDX) allowed further characterization of these insoluble Gd species. Amorphous, spheroid structures of approximately 100-200 nm of sea urchin-like shape were detected. Furthermore, Gd was consistently colocalized with calcium, oxygen, and phosphorous, strongly suggesting the presence of structures composed of mixed Gd/Ca phosphates. No or occasional colocalization with iron and sulfur was observed. CONCLUSION: A dedicated analytical workflow produced original data on the speciation of Gd in DCN of rats repeatedly injected with GBCAs. The addition, in comparison with previous studies of Gd speciation in brain, of SP element detection and imaging techniques allowed a comprehensive speciation analysis approach. Whereas for gadoterate the main fraction of retained Gd was present as intact GBCA form in the soluble fractions, for linear gadodiamide, less than 10% of Gd could be solubilized and characterized using size-exclusion chromatography coupled to ICP-MS. The main Gd species detected in the soluble fractions were macromolecules of 440 kDa. One of them was speculated to be a Gd complex with iron-binding protein (ferritin). However, the major fraction of residual Gd was present as insoluble particulate species, very likely composed of mixed Gd/Ca phosphates. This comprehensive Gd speciation study provided important evidence for the dechelation of linear GBCAs and offered a deeper insight into the mechanisms of Gd deposition in the brain.

Small Brain Lesion Enhancement and Gadolinium Deposition in the Rat Brain: Comparison Between Gadopiclenol and Gadobenate Dimeglumine.

OBJECTIVES: The aim of the set of studies was to compare gadopiclenol, a new high relaxivity gadolinium (Gd)-based contrast agent (GBCA) to gadobenate dimeglumine in terms of small brain lesion enhancement and Gd retention, including T1 enhancement in the cerebellum. MATERIALS AND METHODS: In a first study, T1 enhancement at 0.1 mmol/kg body weight (bw) of gadopiclenol or gadobenate dimeglumine was evaluated in a small brain lesions rat model at 2.35 T. The 2 GBCAs were injected in an alternated and cross-over manner separated by an interval of 4.4 ± 1.0 hours (minimum, 3.5 hours; maximum, 6.1 hours; n = 6). In a second study, the passage of the GBCAs into cerebrospinal fluid (CSF) was evaluated by measuring the fourth ventricle T1 enhancement in healthy rats at 4.7 T over 23 minutes after a single intravenous (IV) injection of 1.2 mmol/kg bw of gadopiclenol or gadobenate dimeglumine (n = 6/group). In a third study, Gd retention at 1 month was evaluated in healthy rats who had received 20 IV injections of 1 of the 2 GBCAs (0.6 mmol/kg bw) or a similar volume of saline (n = 10/group) over 5 weeks. T1 enhancement of the deep cerebellar nuclei (DCN) was assessed by T1-weighted magnetic resonance imaging at 2.35 T, performed before the injection and thereafter once a week up to 1 month after the last injection. Elemental Gd levels in central nervous system structures, in muscle and in plasma were determined by inductively coupled plasma mass spectrometry (ICP-MS) 1 month after the last injection. RESULTS: The first study in a small brain lesion rat model showed a ≈2-fold higher number of enhanced voxels in lesions with gadopiclenol compared with gadobenate dimeglumine. T1 enhancement of the fourth ventricle was observed in the first minutes after a single IV injection of gadopiclenol or gadobenate dimeglumine (study 2), resulting, in the case of gadopiclenol, in transient enhancement during the injection period of the repeated administrations study (study 3). In terms of Gd retention, T1 enhancement of the DCN was noted in the gadobenate dimeglumine group during the month after the injection period. No such enhancement of the DCN was observed in the gadopiclenol group. Gadolinium concentrations 1 month after the injection period in the gadopiclenol group were slightly increased in plasma and lower by a factor of 2 to 3 in the CNS structures and muscles, compared with gadobenate dimeglumine. CONCLUSIONS: In the small brain lesion rat model, gadopiclenol provides significantly higher enhancement of brain lesions compared with gadobentate dimeglumine at the same dose. After repeated IV injections, as expected for a macrocyclic GBCA, Gd retention is minimalized in the case of gadopiclenol compared with gadobenate dimeglumine, resulting in no T1 hypersignal in the DCN.

Reduced nontarget embolization and increased targeted delivery with a reflux-control microcatheter in a swine model.

PURPOSE: To evaluate the potential differences in non-target embolization and vessel microsphere filling of a reflux-control microcatheter (RCM) compared to a standard end-hole microcatheter (SEHM) in a swine model. MATERIALS AND METHODS: Radiopaque microspheres were injected with both RCM and SEHM (2.4-Fr and 2.7-Fr) in the kidneys of a preclinical swine model. Transarterial renal embolization procedures with RCM or SEHM were performed in both kidneys of 14 pigs. Renal arteries were selectively embolized with an automated injection protocol of radio-opaque microspheres. Ex-vivo X-ray microtomography images of the kidneys were utilized to evaluate the embolization by quantification of the deposition of injected microspheres in the target vs. the non-target area of injection. X-ray microtomography images were blindly analyzed by five interventional radiologists. The degree of vessel filling and the non-target embolization were quantified using a scale from 1 to 5 for each parameter. An analysis of variance was used to compare the paired scores. RESULTS: Total volumes of radio-opaque microspheres injected were similar for RCM (11.5±3.6 [SD] mL; range: 6-17mL) and SEHM (10.6±5.2 [SD] mL; range: 4-19mL) (P=0.38). The voxels enhanced ratio in the target (T) vs. non-target (NT) areas was greater with RCM (T=98.3% vs. NT=1.7%) than with SEHM (T=89% vs. NT=11%) but the difference was not significant (P=0.30). The total score blindly given by the five interventional radiologists was significantly different between RCM (12.3±2.1 [SD]; range: 6-15) and the standard catheter (11.3±2.5 [SD]; range: 4-15) (P=0.0073), with a significant decrease of non-target embolization for RCM (3.8±1.3 [SD]; range: 3.5-4.2) compared to SEHM (3.2±1.5 [SD]; range: 2.9-3.5) (P=0.014). CONCLUSION: In an animal model, RCM microcatheters reduce the risk of non-target embolization from 11% to 1.7%, increasing the delivery of microspheres of 98% to the target vessels, compared to SEHM microcatheters.

Safety and Gadolinium Distribution of the New High-Relaxivity Gadolinium Chelate Gadopiclenol in a Rat Model of Severe Renal Failure.

OBJECTIVE: The aim of this study was to investigate the toxicological profile of gadopiclenol, a new high-relaxivity macrocyclic gadolinium-based contrast agent (GBCA), in renally impaired rats, in comparison with 2 other macrocyclic GBCAs, gadoterate meglumine and gadobutrol, and 1 linear and nonionic GBCA, gadodiamide. METHODS: Renal failure was induced by adding 0.75% wt/wt adenine to the diet for 3 weeks. During the second week of adenine-enriched diet, the animals (n = 8/group × 5 groups) received 5 consecutive intravenous injections of GBCA at 2.5 mmol/kg per injection, resulting in a cumulative dose of 12.5 mmol/kg or saline followed by a 3-week treatment-free period after the last injection. The total (elemental) gadolinium (Gd) concentration in different tissues (brain, cerebellum, femoral epiphysis, liver, skin, heart, kidney, spleen, plasma, urine, and feces) was measured by inductively coupled plasma mass spectrometry. Transmission electron microscopy (and electron energy loss spectroscopy analysis of metallic deposits) was used to investigate the presence and localization of Gd deposits in the skin. Relaxometry was used to evaluate the presence of dissociated Gd in the skin, liver, and bone. Skin histopathology was performed to investigate the presence of nephrogenic systemic fibrosis-like lesions. RESULTS: Gadodiamide administrations were associated with high morbidity-mortality but also with macroscopic and microscopic skin lesions in renally impaired rats. No such effects were observed with gadopiclenol, gadoterate, or gadobutrol. Overall, elemental Gd concentrations were significantly higher in gadodiamide-treated rats than in rats treated with the other GBCAs for all tissues except the liver (where no significant difference was found with gadopiclenol) and the kidney and the heart (where statistically similar Gd concentrations were observed for all GBCAs). No plasma biochemical abnormalities were observed with gadopiclenol or the control GBCAs. Histopathology revealed a normal skin structure in the rats treated with gadopiclenol, gadoterate, and gadobutrol, contrary to those treated with gadodiamide. No evidence of Gd deposits on collagen fibers and inclusions in fibroblasts was found with gadopiclenol and its macrocyclic controls, unlike with gadodiamide. Animals of all test groups had Gd-positive lysosomal inclusions in the dermal macrophages. However, the textures differed for the different products (speckled texture for gadodiamide and rough-textured appearance for the 2 tested macrocyclic GBCAs). CONCLUSIONS: No evidence of biochemical toxicity or pathological abnormalities of the skin was observed, and similar to other macrocyclic GBCAs, gadoterate and gadobutrol, tissue retention of Gd was found to be low (except in the liver) in renally impaired rats treated with the new high-relaxivity GBCA gadopiclenol.

Speciation Analysis of Gadolinium in the Water-Insoluble Rat Brain Fraction After Administration of Gadolinium-Based Contrast Agents.

PURPOSE: To date, the analysis of gadolinium (Gd) speciation in the brain of animals administered with macrocyclic and linear Gd-based contrast agents (GBCAs) has been limited to Gd soluble in mild buffers. Under such conditions, less than 30% of the brain tissue was solubilized and the extraction recoveries of GBCAs into the aqueous phase were poor, especially in the case of the linear GBCAs. The aim of this study was to find the conditions to solubilize the brain tissue (quasi-)completely while preserving the Gd species present. The subsequent analysis using size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) was intended to shed the light on the speciation of the additionally recovered Gd. METHODS: Four groups of healthy female Sprague Dawley rats (SPF/OFA rats; Charles River, L'Arbresle, France) received randomly 5 intravenous injections (1 injection per week during 5 consecutive weeks) of either gadoterate meglumine, gadobenate dimeglumine, gadodiamide (cumulated dose of 12 mmol/kg), or no injection (control group). The animals were sacrifice 1 week (W1) after the last injection. Brain tissues were solubilized with urea solution, whereas tissues extracted with water served as controls. Total Gd concentrations were determined in the original brain tissue and its soluble and insoluble fractions by inductively coupled plasma-mass spectrometry (ICP-MS) to calculate the Gd accumulation and extraction efficiency. Size exclusion chromatography coupled to ICP-MS was used to monitor the speciation of Gd in the soluble fractions. The stability of GBCAs in the optimum conditions was monitored by spiking the brain samples from the untreated animals. The column recoveries were precisely determined in the purpose of the discrimination of weakly and strongly bound Gd complexes. The identity of the eluted species was explored by the evaluation of the molecular size and retention time matching with Gd chelates and ferritin standard. The speciation analyses were carried out for 2 different brain structures, cortex and cerebellum. RESULTS: The combination of water and urea extractions (sequential extraction) managed to solubilize efficiently the brain tissue (97% ± 1%) while preserving the stability of the initially injected form of GBCA. For macrocyclic gadoterate, 97% ± 1% and 102% ± 3% of Gd initially present in the cortex and cerebellum were extracted to the soluble fraction. For gadobenate, similar amounts of Gd (49% ± 1% and 46% ± 4%) were recovered from cortex and cerebellum. For gadodiamide, 48% ± 2% of Gd was extracted from cortex and 34% ± 1% from cerebellum. These extraction efficiencies were higher than reported elsewhere. The SEC-ICP-MS and the column recovery determination proved that Gd present at low nmol/g levels in brain tissue was exclusively in the intact GBCA form in all the fractions of brain from the animals treated with gadoterate. For the linear GBCAs (gadobenate and gadodiamide), 3 Gd species of different hydrodynamic volumes were detected in the urea-soluble fraction: (1) larger than 660 kDa, (2) approximately 440 kDa, and (3) intact GBCAs. The species of 440 kDa corresponded, on the basis of the elution volume, to a Gd3+ complex with ferritin. Gd3+ was also demonstrated by SEC-ICP-MS to react with the ferritin standard in 100 mM ammonium acetate (pH 7.4). In contrast to macrocyclic gadoterate, for linear GBCAs, the column recovery was largely incomplete, suggesting the presence of free, hydrolyzed, or weakly bound Gd3+ with endogenous ligands. CONCLUSIONS: The sequential extraction of rat brain tissue with water and urea solution resulted in quasi-complete solubilization of the tissue and a considerable increase in the recoveries of Gd species in comparison with previous reports. The macrocyclic gadoterate was demonstrated to remain intact in the brain 1 week after administration to rats. The linear GBCAs gadobenate and gadodiamide underwent ligand exchange reactions resulting in the presence of a series of Gd3+ complexes of different strength with endogenous ligands. Ferritin was identified as one of the macromolecules reacting with Gd3+. For the linear GBCAs, 3% of the insoluble brain tissue was found to contain more than 50% of Gd in unidentified form(s).

Long-Term Evaluation of Gadolinium Retention in Rat Brain After Single Injection of a Clinically Relevant Dose of Gadolinium-Based Contrast Agents.

PURPOSE: The aim of this study was to investigate the presence and chemical forms of residual gadolinium (Gd) in rat brain after a single dose of Gd-based contrast agent. METHODS: Four groups of healthy rats (2 sacrifice time-points, n = 10/group, 80 rats in total) were randomized to receive a single intravenous injection of 1 of the 3 Gd-based contrast agents (GBCAs) (gadoterate meglumine, gadobenate dimeglumine, or gadodiamide) or the same volume of 0.9% saline solution. The injected concentration was 0.6 mmol/kg, corresponding to a concentration of 0.1 mmol/kg in humans after body surface normalization between rats and humans (according to the US Food and Drug Administration recommendations). Animals were sacrificed at 2 washout times: 1 (M1) and 5 (M5) months after the injection. Total Gd concentrations were determined in cerebellum by inductively coupled plasma mass spectrometry. Gadolinium speciation was analyzed by size-exclusion chromatography coupled to inductively coupled plasma mass spectrometry after extraction from cerebellum. RESULTS: A single injection of a clinically relevant dose of GBCA resulted in the detectable presence of Gd in the cerebellum 1 and 5 months after injection. The cerebellar total Gd concentrations after administration of the least stable GBCA (gadodiamide) were significantly higher at both time-points (M1: 0.280 ± 0.060 nmol/g; M5: 0.193 ± 0.023 nmol/g) than those observed for macrocyclic gadoterate (M1: 0.019 ± 0.004 nmol/g, M5: 0.004 ± 0.002 nmol/g; P < 0.0001). Gadolinium concentrations after injection of gadobenate were significantly lower at both time-points (M1: 0.093 ± 0.020 nmol/g; M5: 0.067 ± 0.013 nmol/g; P < 0.05) than the Gd concentration measured after injection of gadodiamide. At the 5-month time-point, the Gd concentration in the gadoterate group was also significantly lower than the Gd concentration in the gadobenate group (P < 0.05). Gadolinium speciation analysis of the water-soluble fraction showed that, after injection of the macrocyclic gadoterate, Gd was still detected only in its intact, chelated form 5 months after injection. In contrast, after a single dose of linear GBCAs (gadobenate and gadodiamide), 2 different forms were detected: intact GBCA and Gd bound to soluble macromolecules (above 80 kDa). Elimination of the intact GBCA form was also observed between the first and fifth month, whereas the amount of Gd present in the macromolecular fraction remained constant 5 months after injection. CONCLUSIONS: A single injection of a clinically relevant dose of GBCA is sufficient to investigate long-term Gd retention in the cerebellar parenchyma. Administration of linear GBCAs (gadodiamide and gadobenate) resulted in higher residual Gd concentrations than administration of the macrocyclic gadoterate. Speciation analysis of the water-soluble fraction of cerebellum confirmed washout of intact GBCA over time. The quantity of Gd bound to macromolecules, observed only with linear GBCAs, remained constant 5 months after injection and is likely to represent a permanent deposition.

Contrast-to-Dose Relationship of Gadopiclenol, an MRI Macrocyclic Gadolinium-based Contrast Agent, Compared with Gadoterate, Gadobenate, and Gadobutrol in a Rat Brain Tumor Model.

Background Detection of cerebral lesions at MRI may benefit from a chemically stable and more sensitively detected gadolinium-based contrast agent (GBCA). Gadopiclenol, a macrocyclic GBCA with at least twofold higher relaxivity, is currently undergoing clinical trials in humans. Purpose To determine the relationship between MRI contrast enhancement and the injected dose of gadopiclenol in a glioma rat model compared with those of conventional GBCA at label dose. Materials and Methods Between April and July 2012, 32 rats implanted with C6 glioma received two intravenous injections at a 24-hour interval. The injections were randomly selected among five doses of gadopiclenol (0.025, 0.05, 0.075, 0.1, and 0.2 mmol/kg) and three reference GBCAs (gadoterate meglumine, gadobutrol, and gadobenate dimeglumine) at 0.1 mmol/kg. MRI tumor enhancement was assessed on T1-weighted images before and up to 30 minutes after injection. Two blinded radiologists visually and qualitatively scored contrast enhancement, border delineation, and visualization of tumor morphology. Quantitatively, variations in contrast-to-noise ratio (ΔCNR) between tumor and contralateral parenchyma were calculated at each time point and were compared for each treatment at 5 minutes by using a mixed model after normality test. Results A total of 24 rats underwent the complete protocol (n = 5-7 per group). A linear dose-dependent ΔCNR relationship was observed between 0.025 and 0.1 mmol/kg for gadopiclenol (R 2 = 0.99). No difference in ΔCNR was observed between the three reference GBCAs (P ≥ .55). Gadopiclenol resulted in twofold higher ΔCNR at 0.1 mmol/kg (P < .001 vs gadobutrol and gadoterate, P = .002 vs gadobenate) and similar ΔCNR at 0.05 mmol/kg (P = .56, P > .99, and P = .44 compared with gadobutrol, gadobenate, and gadoterate, respectively). For both readers, 0.05 mmol/kg of gadopiclenol improved contrast enhancement, border delineation, and visualization of tumor morphology (scores > 3 compared with scores between 2 and 3 for the marketed GBCA). Conclusion Gadopiclenol at 0.05 mmol/kg yielded comparable change in contrast-to-noise ratio and morphologic characterization of brain tumors compared with gadobenate, gadoterate, or gadobutrol at 0.1 mmol/kg. Published under a CC BY 4.0 license. Online supplemental material is available for this article. See also the editorial by Tweedle in this issue.

Physicochemical and Pharmacokinetic Profiles of Gadopiclenol: A New Macrocyclic Gadolinium Chelate With High T1 Relaxivity.

OBJECTIVES: We aimed to evaluate gadopiclenol, a newly developed extracellular nonspecific macrocyclic gadolinium-based contrast agent (GBCA) having high relaxivity properties, which was designed to increase lesion detection and characterization by magnetic resonance imaging. METHODS: We described the molecular structure of gadopiclenol and measured the r1 and r2 relaxivity properties at fields of 0.47 and 1.41 T in water and human serum. Nuclear magnetic relaxation dispersion profile measurements were performed from 0.24 mT to 7 T. Protonation and complexation constants were determined using pH-metric measurements, and we investigated the acid-assisted dissociation of gadopiclenol, gadodiamide, gadobutrol, and gadoterate at 37°C and pH 1.2. Applying the relaxometry technique (37°C, 0.47 T), we investigated the risk of dechelation of gadopiclenol, gadoterate, and gadodiamide in the presence of ZnCl2 (2.5 mM) and a phosphate buffer (335 mM). Pharmacokinetics studies of radiolabeled Gd-gadopiclenol were performed in Beagle dogs, and protein binding was measured in rats, dogs, and humans plasma and red blood cells. RESULTS: Gadopiclenol [gadolinium chelate of 2,2',2″-(3,6,9-triaza-1(2,6)-pyridinacyclodecaphane-3,6,9-triyl)tris(5-((2,3-dihydroxypropyl)amino)-5-oxopentanoic acid); registry number 933983-75-6] is based on a pyclen macrocyclic structure. Gadopiclenol exhibited a very high relaxivity in water (r1 = 12.2 mM·s at 1.41 T), and the r1 value in human serum at 37°C did not markedly change with increasing field (r1 = 12.8 mM·s at 1.41 T and 11.6 mM·s at 3 T). The relaxivity data in human serum did not indicate protein binding. The nuclear magnetic relaxation dispersion profile of gadopiclenol exhibited a high and stable relaxivity in a strong magnetic field. Gadopiclenol showed high kinetic inertness under acidic conditions, with a dissociation half-life of 20 ± 3 days compared with 4 ± 0.5 days for gadoterate, 18 hours for gadobutrol, and less than 5 seconds for gadodiamide and gadopentetate. The pharmacokinetic profile in dogs was typical of extracellular nonspecific GBCAs, showing distribution in the extracellular compartment and no metabolism. No protein binding was found in rats, dogs, and humans. CONCLUSIONS: Gadopiclenol is a new extracellular and macrocyclic Gd chelate that exhibited high relaxivity, no protein binding, and high kinetic inertness. Its pharmacokinetic profile in dogs was similar to that of other extracellular nonspecific GBCAs.

Evaluation of Gadopiclenol and P846, 2 High-Relaxivity Macrocyclic Magnetic Resonance Contrast Agents Without Protein Binding, in a Rodent Model of Hepatic Metastases: Potential Solutions for Improved Enhancement at Ultrahigh Field Strength.

PURPOSE: The aim of this study was to evaluate in vitro and in vivo the enhancement properties of experimental gadolinium (Gd)-based contrast agents (GBCAs) with different molecular weights and hydration numbers (P846 and gadopiclenol) compared with clinically approved low-molecular, extracellular agents (gadopentetate and gadoterate) at 9.4 T and to discuss influencing factors on r1 relaxivities. METHODS AND MATERIALS: All experiments were performed with a 9.4 T animal scanner (Bruker, Germany). We performed relaxometry measurements for all contrast agents in human plasma at 37°C using an IR-RARE sequence. In addition, we compared P846 with gadopentetate and gadopiclenol with gadoterate intraindividually in rats with hepatic colorectal cancer metastases (n = 10 each) acquiring T1-weighted FLASH sequences before and at 10 consecutive time points during 20 minutes. After intravenous contrast agent application, signal-to-noise ratios (SNRs), contrast-to-noise ratios (CNRs), and lesion enhancement (LE) for liver parenchyma and tumors were calculated based on region of interest measurements. RESULTS: Longitudinal relaxivities (r1) of the low-molecular agents were lower as compared with the experimental compounds. However, r1 of gadopentetate and gadoterate demonstrated only a moderate decrease of r1 at 9.4 T as compared with known data at lower field strengths (gadopentetate: r1 [at 9.4 T], 3.4 mM s/r1 [at 1.5 T], 4.1 mM s/gadoterate: r1 [at 9.4 T], 3.1 mM s/r1 [at 1.5 T], 3.6 mM s). In contrast, r1 of P846 showed a marked reduction at 9.4 T compared with 1.5 T (P846: r1 [at 9.4 T], 6.4 mM s/r1 [at 1.5 T], 32 mM s). Gadopiclenol provided the highest r1 in this study at 9.4 T and the drop of r1 as compared with lower field strength is less apparent (gadopiclenol: r1 [at 9.4 T], 8.7 mM s/r1 [at 1.5 T], 12.7 mM s).In vivo, P846 and gadopiclenol showed significantly higher SNR, CNR, and LE as compared with the low-molecular control agents (mean ± SD; SNRliver [gadopentetate, 18.1 ± 1.2; P846, 27.2 ± 1.5; P < 0.001]; SNRtumor [gadopentetate, 22.6 ± 1.9; P846, 40.1 ± 1.9; P < 0.001]; CNR [gadopentetate, 4.6 ± 1.0; P846, 12.9 ± 0.9; P < 0.001]; LE [gadopentetate, 7.2 ± 1.9; P846, 14.9 ± 1.9; P < 0.001]/SNRliver [gadoterate, 8.8 ± 0.5; gadopiclenol, 12.6 ± 1.3; P < 0.001]; SNRtumor [gadoterate, 11.3 ± 1.2; gadopiclenol, 20.9 ± 2.9; P < 0.001]; CNR [gadoterate, 2.5 ± 0.7; gadopiclenol, 8.3 ± 1.7; P < 0.001]; LE [gadoterate, 4.4 ± 1.2; gadopiclenol, 13.0 ± 2.9; P < 0.001]). Thus, for equal Gd doses, gadopiclenol and P846 increase the CNR of liver metastases by a factor of 2.5 to 3 at 9.4 T compared with gadoterate and gadopentetate. CONCLUSIONS: P846 and gadopiclenol provide superior enhancement at 9.4 T as compared with gadopentetate and gadoterate. However, the macromolecular agent P846 shows a marked decrease of r1 from 1.5 T to 9.4 T. This effect is less apparent for the low-molecular agents gadopiclenol, gadopentetate, and gadoterate. Yet, based on the higher hydration number, r1 of P846 and gadopiclenol are markedly higher as compared with the reference contrast agents. Thus, building compounds with moderately increased molecular size and hydration number, as implemented in gadopiclenol, seems to be a promising way to develop highly effective GBCAs.Advantages for gadopiclenol include a strong enhancement regardless of the external magnetic field strength, pharmacokinetics comparable to those of clinically approved extracellular GBCAs, and the potential to either improve sensitivity in diagnostic magnetic resonance imaging by improving lesion conspicuity or to perform studies with significantly reduced Gd-dose while at the same time providing comparable diagnostic accuracy. However, all this needs to be proven in clinical studies.

Does Age Interfere With Gadolinium Toxicity and Presence in Brain and Bone Tissues?: A Comparative Gadoterate Versus Gadodiamide Study in Juvenile and Adult Rats.

OBJECTIVES: The main objective of the study was to assess the effect of age on target tissue total gadolinium (Gd) retention after repeated administration of gadodiamide (linear) or gadoterate (macrocyclic) Gd-based contrast agent (GBCA) in rats. The secondary objective was to assess the potential developmental and long-term consequences of GBCA administration during neonatal and juvenile periods. MATERIALS AND METHODS: A total of 20 equivalent human clinical doses (cumulated dose, 12 mmol Gd/kg) of either gadoterate or gadodiamide were administered concurrently by the intravenous route to healthy adult and juvenile rats. Saline was administered to juvenile rats forming the control group. In juvenile rats, the doses were administered from postnatal day 12, that is, once the blood-brain barrier is functional as in humans after birth. The tests were conducted on 5 juvenile rats per sex and per group and on 3 adult animals per sex and per group. T1-weighted magnetic resonance imaging of the cerebellum was performed at 4.7 T during both the treatment and treatment-free periods. Behavioral tests were performed in juvenile rats. Rats were euthanatized at 11 to 12 weeks (ie, approximately 3 months) after the last administration. Total Gd concentrations were measured in plasma, skin, bone, and brain by inductively coupled plasma mass spectrometry. Cerebellum samples from the juvenile rats were characterized by histopathological examination (including immunohistochemistry for glial fibrillary acidic protein or GFAP, and CD68). Lipofuscin pigments were also studied by fluorescence microscopy. All tests were performed blindly on randomized animals. RESULTS: Transient skin lesions were observed in juvenile rats (5/5 females and 2/4 males) and not in adult rats having received gadodiamide. Persisting (up to completion of the study) T1 hyperintensity in the deep cerebellar nuclei (DCNs) was observed only in gadodiamide-treated rats. Quantitatively, a slightly higher progressive increase in the DCN/brain stem ratio was observed in adult rats compared with juvenile rats, whereas no difference was noted visually. In all tissues, total Gd concentrations were higher (10- to 30-fold higher) in the gadodiamide-treated groups than in the gadoterate groups. No age-related differences were observed except in bone marrow where total Gd concentrations in gadodiamide-treated juvenile rats were higher than those measured in adults and similar to those measured in cortical bone tissue. No significant treatment-related effects were observed in histopathological findings or in development, behavior, and biochemistry parameters. However, in the elevated plus maze test, a trend toward an anxiogenic effect was observed in the gadodiamide group compared with other groups (nonsignificant). Moreover, in the balance beam test, a high number of trials were excluded in the gadodiamide group because rats (mainly males) did not completely cross the beam, which may also reflect an anxiogenic effect. CONCLUSIONS: No T1 hyperintensity was observed in the DCN after administration of the macrocyclic GBCA gadoterate regardless of age as opposed to administration of the linear GBCA gadodiamide. Repeated administration of gadodiamide in neonatal and juvenile rats resulted in similar total Gd retention in the skin, brain, and bone to that in adult rats with sex having no effect, whereas Gd distribution in bone marrow was influenced by age. Further studies are required to assess the form of the retained Gd and to investigate the potential risks associated with Gd retention in bone marrow in juvenile animals treated with gadodiamide. Regardless of age, total Gd concentration in the brain and bone was 10- to 30-fold higher after administration of gadodiamide compared with gadoterate.

Multimodal Imaging Study of Gadolinium Presence in Rat Cerebellum: Differences Between Gd Chelates, Presence in the Virchow-Robin Space, Association With Lipofuscin, and Hypotheses About Distribution Pathway.

PURPOSE: The aim of this study was to investigate, based on in-depth multimodal imaging, the presence of Gd deposits, their ultrastructure, location, and co-location with endogenous elements, in the cerebellum, after repeated administrations of gadolinium-based contrast agents (GBCAs). METHODS: Rats sensitized by subtotal nephrectomy received 20 daily intravenous injections of 0.6 mmol Gd/kg for 5 weeks of commercial forms of either gadoterate, gadobenate or gadodiamide, or saline (n = 2/group). The study was randomized and blinded. Magnetic resonance imaging examination was performed weekly. One month after the last injection, electron microscopy analysis of the deep cerebellar nuclei, the granular layer of cerebellar cortex, and the choroid plexus was performed. Elemental analysis of deposits was carried out by electron energy loss spectroscopy. Secondary ion mass spectroscopy was used for complementary chemical mapping. RESULTS: A T1 hypersignal was evidenced in the deep cerebellar nuclei of rats treated with linear GBCAs, and Gd deposits were identified in all the studied cerebellar structures with gadobenate and gadodiamide (except in the granular layer in gadobenate-treated rats). No such effect was found with the macrocyclic GBCA gadoterate. Most of the Gd deposits revealed a characteristic spheroid "sea urchin-like" morphology, rich in phosphorus, and were localized in the basal lamina of microvessels, in the perivascular Virchow-Robin space, and in the interstitium. Gd was also identified in the glial cells, associated with lipofuscin pigments, for these same groups. CONCLUSIONS: Transmission electron microscopy analysis of cerebellums of renally impaired rats repeatedly injected with gadobenate and gadodiamide revealed the presence of Gd. Spheroid Gd depositions consisting of a filamentous meshwork were observed in the wall of microvessels, in perivascular Virchow-Robin space, and in the interstitium. Gd was also found in choroid plexus and was associated with pigments (likely lipofuscin) in glial cells. This is consistent with the involvement of the glymphatic distribution pathway for GBCAs. No insoluble Gd deposits were detected in rats injected with the macrocyclic GBCA gadoterate and controls.

One-year Retention of Gadolinium in the Brain: Comparison of Gadodiamide and Gadoterate Meglumine in a Rodent Model.

Purpose To compare the long-term brain elimination kinetics and gadolinium species in healthy rats after repeated injections of the contrast agents gadodiamide (a linear contrast agent) or gadoterate (a macrocyclic contrast agent). Materials and Methods Nine-week-old rats received five doses of 2.4 mmol gadolinium per kilogram of body weight over 5 weeks and were followed for 12 months with T1-weighted MRI (n = 140 rats, corresponding to seven time points, two contrast agents, and 10 rats per group). Animals were sacrificed at 1 week, 1 month, and 2, 3, 4, 5, and 12 months after the last injection. Brain and plasma were sampled to determine the total gadolinium concentration by using inductively coupled plasma mass spectrometry (ICP-MS). For the cerebellum, gadolinium speciation analysis was performed after mild extraction at four time points (1 month and 3, 5, and 12 months after the last injection) by using size exclusion chromatography and hydrophilic interaction liquid chromatography, both coupled to ICP-MS. Tissue gadolinium kinetics were fitted to estimate the area under the curves and tissue elimination half-lives over the 12-month injection-free period. Results T1 hyperintensity of the deep cerebellar nuclei was observed only in gadodiamide-treated rats and remained stable from the 1st month after the last injection (the ratio of the signal intensity of the deep cerebellar nuclei to the signal intensity of the brain stem at 1 year: 1.101 ± 0.023 vs 1.037 ± 0.022 before injection, P < .001). Seventy-five percent of the total gadolinium detected after the last injection of gadodiamide (3.25 nmol/g ± 0.30) was retained in the cerebellum at 1 year (2.45 nmol/g ± 0.35), with binding of soluble gadolinium to macromolecules. No T1 hyperintensity was observed with gadoterate, consistent with a rapid, time-dependent washout of the intact gadolinium chelate down to background levels (0.07 nmol/g ± 0.03). Conclusion After repeated administration of gadodiamide, a large portion of gadolinium was retained in the brain, with binding of soluble gadolinium to macromolecules. After repeated injection of gadoterate, only traces of the intact chelated gadolinium were observed with time-dependent clearance. Online supplemental material is available for this article.

Gadolinium Retention, Brain T1 Hyperintensity, and Endogenous Metals: A Comparative Study of Macrocyclic Versus Linear Gadolinium Chelates in Renally Sensitized Rats.

OBJECTIVES: This preclinical study was designed to compare gadolinium (Gd) brain uptake after repeated injections of a macrocyclic Gd-based contrast agent (GBCA) (gadoterate meglumine) or 2 linear GBCAs (L-GBCAs) (gadobenate dimeglumine or gadodiamide) on a translational model of moderate renal impairment in rats. METHODS: The study was carried out in subtotally nephrectomized rats. Animals received 4 intravenous injections per week of GBCA (gadoterate meglumine, gadobenate dimeglumine, or gadodiamide) for 5 weeks, resulting in a cumulative dose of 12 mmol/kg, followed by a 1-month injection-free period. T1 hyperintensity in the deep cerebellar nuclei (DCNs) was investigated, and brain structures were carefully dissected to determine elemental Gd, iron (Fe), copper (Cu), and zinc (Zn) distribution by mass spectrometry. Urinary excretion of endogenous metals was also investigated soon after GBCA administration and several days later in order to assess a potential transmetalation phenomenon. RESULTS: Unlike gadoterate, repeated injections of L-GBCAs gadobenate and gadodiamide both induced T1 hyperintensity in the DCNs. Fine dissection of cerebral and cerebellar structures demonstrated very low levels or absence of Gd after repeated injections of gadoterate, in contrast to the two L-GBCAs, for which the highest total Gd concentration was demonstrated in the DCNs (Gd concentration in DCNs after 4.5 weeks of injection-free period: 27.1 ± 6.5 nmol/g for gadodiamide [P < 0.01 vs saline and P < 0.05 vs gadoterate]; 12.0 ± 2.6 nmol/g for gadobenate [P < 0.09 vs saline]; compared with 1.4 ± 0.2 nmol/g for gadoterate [ns vs saline]). The distribution of Gd concentration among the various brain structures dissected was also well correlated with the Fe distribution in these structures. No difference in endogenous metal levels in brain structures was observed. However, injection of gadobenate or gadodiamide resulted in an increase in urinary Zn excretion (urinary Zn concentrations: 57.9 ± 20.5 nmol/mL with gadobenate [P < 0.01 vs gadoterate and saline] and 221.6 ± 83.3 nmol/L with gadodiamide [P < 0.0001 vs all other treatments] vs 8.1 ± 2.3 nmol/L with saline and 10.6 ± 4.8 nmol/L with gadoterate]). CONCLUSIONS: In a model of renally impaired rats, only traces of gadoterate meglumine were detected in the brain with no T1 hyperintensity of the DCNs, whereas marked Gd retention was observed in almost all brain areas after injections of the L-GBCAs, gadobenate dimeglumine and gadodiamide. Brain structures with higher Gd uptake corresponded to those structures containing more Fe. Urinary Zn excretion was significantly increased after a single injection of L-GBCAs.

Moderate Renal Failure Accentuates T1 Signal Enhancement in the Deep Cerebellar Nuclei of Gadodiamide-Treated Rats.

OBJECTIVES: The purpose of this preclinical study was to investigate whether moderate chronic kidney disease is a factor in potentiating gadolinium (Gd) uptake in the brain. MATERIALS AND METHODS: A comparative study was performed on renally impaired (subtotal nephrectomy) rats versus rats with normal renal function. The animals received 4 daily injections of 0.6 mmol Gd/kg a week for 5 weeks (cumulative dose of 12 mmol Gd/kg) of gadodiamide or saline solution. The MR signal enhancement in the deep cerebellar nuclei was monitored by weekly magnetic resonance imaging examinations. One week after the final injection, the total Gd concentration was determined by inductively coupled plasma mass spectrometry in different regions of the brain including the cerebellum, plasma, cerebrospinal fluid, parietal bone, and femur. RESULTS: After the administration of gadodiamide, the subtotal nephrectomy group presented a significantly higher T1 signal enhancement in the deep cerebellar nuclei and a major increase in the total Gd concentration in all the studied structures, compared with the normal renal function group receiving the same linear Gd-based contrast agent. Those potentiated animals also showed a pronounced hypersignal in the choroid plexus, still persistent 6 days after the last injection, whereas low concentration of Gd was found in the cerebrospinal fluid (<0.05 µmol/L) at this time point. Plasma Gd concentration was then around 1 µmol/L. Interestingly, plasma Gd was predominantly in a dissociated and soluble form (around 90% of total Gd). Total Gd concentrations in the brain, cerebellum, plasma, and bones correlated with creatinine clearance in both the gadodiamide-treated groups. CONCLUSIONS: Renal insufficiency in rats potentiates Gd uptake in the cerebellum, brain, and bones.

Linear Gadolinium-Based Contrast Agents Are Associated With Brain Gadolinium Retention in Healthy Rats.

OBJECTIVES: The aim of this study was to evaluate Gd retention in the deep cerebellar nuclei (DCN) of linear gadolinium-based contrast agents (GBCAs) compared with a macrocyclic contrast agent. MATERIALS AND METHODS: The brain tissue retention of Gd of 3 linear GBCAs (gadobenate dimeglumine, gadopentetate dimeglumine, and gadodiamide) and a macrocyclic GBCA (gadoterate meglumine) was compared in healthy rats (n = 8 per group) that received 20 intravenous injections of 0.6 mmol Gd/kg (4 injections per week for 5 weeks). An additional control group with saline was included. T1-weighted magnetic resonance imaging was performed before injection and once a week during the 5 weeks of injections and for another 4 additional weeks after contrast period. Total gadolinium concentration was measured with inductively coupled plasma mass spectrometry. Blinded qualitative and quantitative evaluations of the T1 signal intensity in DCN were performed, as well as a statistical analysis on quantitative data. RESULTS: At completion of the injection period, all the linear contrast agents (gadobenate dimeglumine, gadopentetate dimeglumine, and gadodiamide) induced a significant increase in signal intensity in DCN, unlike the macrocyclic GBCA (gadoterate meglumine) or saline. The T1 hypersignal enhancement kinetic was fast for gadodiamide. Total Gd concentrations for the 3 linear GBCAs groups at week 10 were significantly higher in the cerebellum (1.21 ± 0.48, 1.67 ± 0.17, and 3.75 ± 0.18 nmol/g for gadobenate dimeglumine, gadopentetate dimeglumine, and gadodiamide, respectively) than with the gadoterate meglumine (0.27 ± 0.16 nmol/g, P < 0.05) and saline (0.09 ± 0.12 nmol/g, P < 0.05). No significant difference was observed between the macrocyclic agent and saline. CONCLUSIONS: Repeated administrations of the linear GBCAs gadodiamide, gadobenate dimeglumine, and gadopentetate dimeglumine to healthy rats were associated with progressive and significant T1 signal hyperintensity in the DCN, along with Gd deposition in the cerebellum. This is in contrast with the macrocyclic GBCA gadoterate meglumine for which no effect was observed.

Non-clinical safety assessment of gadoterate meglumine (Dotarem(®)) in neonatal and juvenile rats.

The purpose of this study was to assess the safety of gadoterate meglumine, a gadolinium-based contrast agent used in magnetic resonance imaging, in neonatal and juvenile rats. Rats received a single intravenous administration on postnatal day (PND) 10 or 6 administrations (from PND 10 to 30), at doses of 0, 0.6, 1.25, and 2.5 mmol/kg/administration, i.e. equivalent to approximately 1, 2 and 4-times the usual human dose. The animals were sacrificed at the end of the treatment period or after a 60-day treatment-free period. No mortality and no significant treatment-related effect on clinical signs, macroscopic and histopathological findings, development, behavior, sexual maturation and hematology parameters were observed. Minor non-adverse changes were observed in clinical biochemistry and urinary parameters. Based on AUC0-t, gadoterate meglumine was more rapidly eliminated at PND 30 vs. PND 10, reflecting maturation of kidney function. At the end of the treatment period, Gd was measurable in all organs sampled after single or repeated dosing and levels were dose-dependent as expected, the highest ones being found in kidneys. The total Gd concentrations were similar in all the organs following a single or repeated dosing. At the end of the treatment-free period, only traces of gadolinium were quantifiable, almost exclusively in kidneys, reflecting the excretory function of this organ. In conclusion, single or repeated administration of gadoterate meglumine to juvenile rats was well tolerated.

Scientific and industrial challenges of developing nanoparticle-based theranostics and multiple-modality contrast agents for clinical application.

Designing of theranostics and dual or multi-modality contrast agents are currently two of the hottest topics in biotechnology and biomaterials science. However, for single entity theranostics, a right ratio of their diagnostic component and their therapeutic component may not always be realized in a composite suitable for clinical application. For dual/multiple modality molecular imaging agents, after in vivo administration, there is an optimal time window for imaging, when an agent is imaged by one modality, the pharmacokinetics of this agent may not allow imaging by another modality. Due to reticuloendothelial system clearance, efficient in vivo delivery of nanoparticles to the lesion site is sometimes difficult. The toxicity of these entities also remains poorly understood. While the medical need of theranostics is admitted, the business model remains to be established. There is an urgent need for a global and internationally harmonized re-evaluation of the approval and marketing processes of theranostics. However, a reasonable expectation exists that, in the near future, the current obstacles will be removed, thus allowing the wide use of these very promising agents.

T1-Weighted Hypersignal in the Deep Cerebellar Nuclei After Repeated Administrations of Gadolinium-Based Contrast Agents in Healthy Rats: Difference Between Linear and Macrocyclic Agents.

OBJECTIVES: To prospectively compare in healthy rats the effect of multiple injections of macrocyclic (gadoterate meglumine) and linear (gadodiamide) gadolinium-based contrast agents (GBCAs) on T1-weighted signal intensity in the deep cerebellar nuclei (DCN), including the dentate nucleus. MATERIALS AND METHODS: Healthy rats (n = 7/group) received 20 intravenous injections of 0.6 mmol of gadolinium (Gd) per kilogram (4 injections per week during 5 weeks) of gadodiamide, gadoterate meglumine, or hyperosmolar saline (control group). Brain T1-weighted magnetic resonance imaging was performed before and once a week during the 5 weeks of injections and during 5 additional weeks (treatment-free period). Gadolinium concentrations were measured with inductively coupled plasma mass spectrometry in plasma and brain. Blinded qualitative and quantitative evaluations of the T1 signal intensity in DCN were performed, as well as a statistical analysis on quantitative data. RESULTS: A significant and persistent T1 signal hyperintensity in DCN was observed only in gadodiamide-treated rats. The DCN-to-cerebellar cortex signal ratio was significantly increased from the 12th injection of gadodiamide (1.070 ± 0.024) compared to the gadoterate meglumine group (1.000 ± 0.033; P < 0.001) and control group (1.019 ± 0.022; P < 0.001) and did not significantly decrease during the treatment-free period. Total Gd concentrations in the gadodiamide group were significantly higher in the cerebellum (3.66 ± 0.91 nmol/g) compared with the gadoterate meglumine (0.26 ± 0.12 nmol/g; P < 0.05) and control (0.06 ± 0.10 nmol/g; P < 0.05) groups. CONCLUSIONS: Repeated administrations of the linear GBCA gadodiamide to healthy rats are associated with progressive and persistent T1 signal hyperintensity in the DCN, with Gd deposition in the cerebellum in contrast with the macrocyclic GBCA gadoterate meglumine for which no effect was observed.

Distribution profile of gadolinium in gadolinium chelate-treated renally-impaired rats: role of pharmaceutical formulation.

While not acutely toxic, chronic hepatic effect of certain gadolinium chelates (GC), used as contrast agent for magnetic resonance imaging, might represent a risk in renally-impaired patients due to free gadolinium accumulation in the liver. To answer this question, this study investigated the consequences of the presence of small amounts of either a soluble gadolinium salt ("free" Gd) or low-stability chelating impurity in the pharmaceutical solution of gadoteric acid, a macrocyclic GC with high thermodynamic and kinetic stabilities, were investigated in renally-impaired rats. Renal failure was induced by adding 0.75% adenine in the diet for three weeks. The pharmaceutical and commercial solution of gadoteric acid was administered (5 daily intravenous injections of 2.5 mmol Gd/kg) either alone or after being spiked with either "free" gadolinium (i.e., 0.04% w/v) or low-stability impurity (i.e., 0.06 w/v). Another GC, gadodiamide (low thermodynamic and kinetic stabilities) was given as its commercial solution at a similar dose. Non-chelated gadolinium was tested at two doses (0.005 and 0.01 mmol Gd/kg) as acetate salt. Gadodiamide induced systemic toxicity (mortality, severe epidermal and dermal lesions) and substantial tissue Gd retention. The addition of very low amounts of "free", non-chelated gadolinium or low thermodynamic stability impurity to the pharmaceutical solution of the thermodynamically stable GC gadoteric acid resulted in substantial capture of metal by the liver, similar to what was observed in "free" gadolinium salt-treated rats. Relaxometry studies strongly suggested the presence of free and soluble gadolinium in the liver. Electron microscopy examinations revealed the presence of free and insoluble gadolinium deposits in hepatocytes and Kupffer cells of rats treated with gadoteric acid solution spiked with low-stability impurity, free gadolinium and gadodiamide, but not in rats treated with the pharmaceutical solution of gadoteric acid. The presence of impurities in the GC pharmaceutical solution may have long-term biological consequences.