T1 Agents Research Articles

Double agents and secret agents: the emerging fields of exogenous chemical exchange saturation transfer and T2-exchange magnetic resonance imaging contrast agents for molecular imaging.

Two relatively new types of exogenous magnetic resonance imaging contrast agents may provide greater impact for molecular imaging by providing greater specificity for detecting molecular imaging biomarkers. Exogenous chemical exchange saturation transfer (CEST) agents rely on the selective saturation of the magnetization of a proton on an agent, followed by chemical exchange of a proton from the agent to water. The selective detection of a biomarker-responsive CEST signal and an unresponsive CEST signal, followed by the ratiometric comparison of these signals, can improve biomarker specificity. We refer to this improvement as a “double-agent” approach to molecular imaging. Exogenous T2-exchange agents also rely on chemical exchange of protons between the agent and water, especially with an intermediate rate that lies between the slow exchange rates of CEST agents and the fast exchange rates of traditional T1 and T2 agents. Because of this intermediate exchange rate, these agents have been relatively unknown and have acted as “secret agents” in the contrast agent research field. This review exposes these secret agents and describes the merits of double agents through examples of exogenous agents that detect enzyme activity, nucleic acids and gene expression, metabolites, ions, redox state, temperature, and pH. Future directions are also provided for improving both types of contrast agents for improved molecular imaging and clinical translation. Therefore, this review provides an overview of two new types of exogenous contrast agents that are becoming useful tools within the armamentarium of molecular imaging.

Bacillus amyloliquefaciens T1 as a potential control agent for cyanobacteria

Harmful cyanobacterial blooms, which force us to develop economical and environmental friendly ways for eutrophication control, are recognized as a nuisance that cause negative impacts on natural resources and humans. In the current study, strain Bacillus amyloliquefaciens T1 with anti-cyanobacterial activity against bloom-forming cyanobacterium Microcystis aeruginosa 905 was isolated from an eutrophication pond in Wuhan, China, and was identified by 16S ribosomal DNA (rDNA) sequence analysis. B. amyloliquefaciens T1 cell-free filtrate at a concentration of 2 % (v/v) showed a strong anti-cyanobacterial effect against M. aeruginosa 905 (initial cell density was 1.0 × 106 cells mL−1), and a reduction of 99.4 % in cell number was observed after incubation for 6 days. The anti-cyanobacterial effect was in accordance with the spore number rather than the cell number of B. amyloliquefaciens T1, suggesting that M. aeruginosa 905 was indirectly affected by the secretion of anti-cyanobacterial active substances. In addition, the interactions of B. amyloliquefaciens T1 with other cyanobacteria and green algae indicated that M. aeruginosa 907, M. aeruginosa 908, M. aeruginosa 912, and M. aeruginosa 7806 were obviously suppressed, while Anabaena flosaquae 1092 and Chlorella pyrenoidosa 415 proved to be not susceptible. Effects of environmental factors showed that the best inhibition efficiency would be achieved at 30 °C and pH 9.0, and the solid B. amyloliquefaciens T1 agent of B. amyloliquefaciens T1 could selectively kill the wild cyanobacteria from a shallow eutrophic pond. Based on the results obtained from this study, the anti-cyanobacterial bacterium B. amyloliquefaciens T1 had a potential application for eutrophication control in the natural environment.

RGD-targeted MnO nanoparticles as T1 contrast agents for cancer imaging - the effect of PEG length in vivo.

As magnetic resonance imaging (MRI) contrast agents, T1 Gd3+ chelates are generally the preferred option for radiologists over T2 iron oxide nanoparticles. The main reason for the popularity of T1 agents is the easier interpretation of T1-weighted MR images. However, the chemical versatility of nanoparticulate platforms makes them ideal candidates for the next generation of targeted MRI contrast agents. In this context, we present herein the design and preparation of a nanoparticulate contrast agent based on MnO, which presents T1 contrast enhancement properties as well as nanoparticle formulation. Functionalization of MnO nanoparticles with the extensively studied RGD peptide was used to target tumours over-expressing the αvβ3 integrin. PEG (polyethylene glycol) molecules were used to increase the blood half-life of the nanoparticles in vivo, and the effect of different PEG lengths on the final contrast on MR images was investigated.

Preparation and characterisation of magnetic mesoporous silica with a large pore size

A new kind of magnetic mesoporous nanoparticle composed of a uniform Fe3O4 nanoscale core and a larger pore size of SiO2 mesoporous shell (Fe3O4@meso-SiO2) has been successfully synthesised in a water/oil phase. The obtained core-shell structure is less than 100 nm in diameter and the pore size of mesoporous SiO2 layer is about 7 to 12 nm, which can be controllable through variation of styrene content. The magnetic property, drug delivery, magnetic resonance, and their in vitro c to HeLa cells were evaluated. The results showed that the magnetic mesoporous silica maintained superparamagnetic behaviour at room temperature, and possessed good transverse relaxivity and good biocompatibility. Additionally, the drug loading and release test indicated that the loading efficiency of DOX was 92%, and the Fe3O4@meso-SiO2 system showed a sustained-release property. The materials have great potential application for simultaneous MRI contrast T2 agent and targeting drug-delivery technology.

Antiferromagnetic Iron Nanocolloids: A New Generation in Vivo T1 MRI Contrast Agent

A novel T1 agent, antiferromagnetic α-iron oxide-hydroxide (α-FeOOH) nanocolloids with a diameter of 2-3 nm, has been successfully prepared. These nanocolloids, together with a post synthetic strategy performed in mesoporous silica, are a great improvement over the low T1-weighted contrast common in traditional magnetic silica nanocomposites. The intrinsic antiferromagnetic goethite (α-FeOOH) shows very low magnetization (M(z)) of 0.05 emu g(-1) at H = 2 T at 300 K (0.0006 emu g(-1) for FeOOH/WMSN-PEG), which is 2 orders of magnitude smaller than any current ultrasmall iron oxide NPs (>5 emu g(-1)) reported to date, hence ensuring the low r2 (∝ Mz) (7.64 mM(-1) s(-1)) and r2/r1 ratio (2.03) at 4.7 T. These biodegradable α-FeOOH nanocolloids also demonstrate excellent in vitro cellular imaging and in vivo MR vascular and urinary trace imaging capability with outstanding biocompatibility, which is exceptionally well secreted by the kidney and not the liver as with most nanoparticles, opening up a new avenue for designing powerful antiferromagnetic iron T1 contrast agents.

Gadolinium‐free T1 contrast agents for MRI: Tunable pharmacokinetics of a new class of manganese porphyrins

To evaluate a new class of manganese porphyrins with tunable pharmacokinetics as potential gadolinium (Gd)-free T1 agents for contrast-enhanced magnetic resonance imaging (MRI). Two new contrast agents, MnTCP and MnP2, were evaluated in four female rats. MRI was performed daily up to 3 days postinjection (0.05 mmol/kg) on a 3 T clinical scanner. T1 relaxation times and dynamic contrast-enhanced MRI were performed to assess contrast enhancement and clearance in blood, heart, liver, kidney, and muscle. Relative T1 decreases were similar for MnTCP and Gd-DTPA in all tissues but were significantly larger (P < 0.05) for MnP2 in blood, heart, kidney, and liver (2-6-fold larger). Clearance of MnTCP was similar to Gd-DTPA, with T1 returning to baseline by 40 minutes and complete elimination in 1 day. MnP2 was cleared from blood after 2 days and sustained a lowered T1 in other tissues for at least 1 hour (P < 0.05). The maximum enhancement, slope, and time-to-peak were similar between contrast agents. Only the parameter AUC60 differed, with MnP2 yielding the largest AUC60 values primarily through longer retention in tissue. MnTCP and MnP2 offer distinct applications as Gd-free T1 contrast agents. MnTCP behaves like a Gd-DTPA analog, while MnP2 provides significantly greater and longer positive signal enhancement.

A noncovalent, fluoroalkyl coating monomer for phosphonate-covered nanoparticles

Gadolinium-containing phosphonate-coated gold nanoparticles were prepared and then non-covalently coated with an amphiphilic fluorous monomer. The monomer spontaneously self-assembles into a non-covalent monolayer shell around the particle. The binding of the shell utilizes a guanidinium–phosphonate interaction analogous to the one exploited by the Wender molecular transporter system. Particle–shell binding was characterized by a 27% decrease in 19F T1 of the fluorous shell upon exposure to the paramagnetic gadolinium in the particle and a corresponding increase in hydrodynamic diameter from 3 nm to 4 nm. Interestingly, a much smaller modulation of 19F T1 is observed when the shell monomer is treated with a phosphonate-free particle. By contrast, the phosphonate-free particle is a much more relaxive 1H T1 agent for water. Together, these observations show that the fluoroalkylguanidinium shell binds selectively to the phosphonate-covered particle. The system's relaxivity and selectivity give it potential for use in 19F based nanotheranostic agents.

Toxicity of Magnetic Resonance Imaging Agents: Small Molecule and Nanoparticle

Magnetic resonance imaging (MRI) contrast agents have been used routinely for more than 20 years in order to increase sensitivity and specificity of lesion detection. MRI contrast agents (CAs) are usually categorized according to their magnetic behavior, biodistribution, and effect on the MR image. Typically, small molecular-weight gadolinium based CAs are examples of T1 agents, while magnetic nanoparticle (MNP) based CAs are examples of T2 agents. In addition to differences in magnetic relaxation behavior, small molecular-weight gadolinium based CAs and MNP based CAs show significantly different toxicity profiles. In the case of small molecular-weight gadolinium based CAs, many previous toxicological studies have reported favorable safety profiles of gadolinium based CAs. However, recently, a delayed serious adverse reaction known as nephrogenic systemic fibrosis (NSF) has been reported in patients, with a marked reduction in renal function after administration of certain types of gadolinium based CAs. For MNP based CAs, in addition to a wide spectrum of nanotoxicity common in nanomaterials, the emerging unexpected cytotoxicity of MNPs has become a new concern. Specifically, the combination of MNPs and strong static magnetic field (SMF) within MRI may give rise to potential adverse effects of MNPs in clinical application.

MRI Tracking of Macrophages Labeled with Glucan Particles Entrapping a Water Insoluble Paramagnetic Gd-Based Agent

This study is aimed at demonstrating the in vivo potential of Gd(III)-loaded glucan particles (Gd-GPs) as magnetic resonance imaging (MRI)-positive agents for labeling and tracking phagocytic cells. GPs were obtained from Saccharomyces cerevisae and loaded with the water-insoluble complex Gd-DOTAMA(C18)2. The uptake kinetics of Gd-GPs by murine macrophages was studied in vitro and the internalization mechanism was assessed by competition assays. The in vivo performance of Gd-GPs was tested at 7.05T on a mouse model of acute liver inflammation. The minimum number of Gd-GPs-labeled J774.A1 macrophages detected in vitro by MRI was ca. 300 cells/μl of agar, which is the lowest number ever reported for cells labeled with a positive T1 agent. Intravenous injection of macrophages labeled with Gd-GPs in a mouse model of liver inflammation enabled the MRI visualization of the cellular infiltration in the diseased area. Gd-GPs represent a promising platform for tracking macrophages by MRI as a T1 alternative to the golden standard T2-based iron oxide particles.

Promising strategies for Gd-based responsive magnetic resonance imaging contrast agents

Magnetic resonance imaging is a powerful imaging modality that is often coupled with paramagnetic contrast agents based on gadolinium to enhance sensitivity and image quality. Responsive contrast agents are key to furthering the diagnostic potential of MRI, both to provide anatomical information and to discern biochemical activity. Recent design of responsive gadolinium-based T₁ agents has made interesting progress, with the development of novel complexes which sense their chemical environment through changes in the coordination of water molecules, the molecular tumbling time or the number of metal centres. Particular promising design strategies include the use of multimeric systems, and the development of dual imaging probes.

High-Throughput Preparation of Hexagonally Ordered Mesoporous Silica and Gadolinosilicate Nanoparticles for use as MRI Contrast Agents

The development of biomedical nanoparticulate materials for use in diagnostics is a delicate balance between performance, particle size, shape, and stability. To identify materials that satisfy all of the criteria it is useful to employ automated high-throughput (HT) techniques for the study of these materials. The structure and performance of surfactant templated mesoporous silica is very sensitive to a wide number of variables. Variables, such as the concentration of the structure-directing agent, the cosolvent and dopant ions and also the temperature and concentration of quenching all have an influence on the structure, surface chemistry, and therefore, the performance of the mesoporous silica nanoparticles generated. Using an automated robotic synthetic platform, a technique has been developed for the high-throughput preparation of mesoporous silica and gadolinium-doped silicate (gadoliniosilicate) nanoparticulate MRI contrast agents. Twelve identical repeats of both the mesoporous silica and gadolinosilicate were synthesized to investigate the reproducibility of the HT technique. Very good reproducibility in the production of the mesoporous silica and the gadolinosilcate materials was obtained using the developed method. The performance of the gadolinosilicate materials was comparable as a T(1) agent to the commercial MRI contrast agents. This HT methodology is highly reproducible and an effective tool that can be translated to the discovery of any sol-gel derived nanomaterial.

Human Aortic Endothelial Cell Labeling with Positive Contrast Gadolinium Oxide Nanoparticles for Cellular Magnetic Resonance Imaging at 7 Tesla

Positive T₁ contrast using gadolinium (Gd) contrast agents can potentially improve detection of labeled cells on magnetic resonance imaging (MRI). Recently, gadolinium oxide (Gd₂O₃) nanoparticles have shown promise as a sensitive T₁ agent for cell labeling at clinical field strengths compared to conventional Gd chelates. The objective of this study was to investigate Gado CELLTrack, a commercially available Gd₂O₃ nanoparticle, for cell labeling and MRI at 7 T. Relaxivity measurements yielded r1 = 4.7 s⁻¹ mM⁻¹ and r₂/r₁ = 6.2. Human aortic endothelial cells were labeled with Gd₂O₃ at various concentrations and underwent MRI from 1 to 7 days postlabeling. The magnetic resonance relaxation times T₁ and T₂ of labeled cell pellets were measured. Cellular contrast agent uptake was quantified by inductively coupled plasma-atomic emission spectroscopy, which showed very high uptake compared to conventional Gd compounds. MRI demonstrated significant positive T₁ contrast and stable labeling on cells. Enhancement was optimal at low Gd concentrations, attained in the 0.02 to 0.1 mM incubation concentration range (corresponding cell uptake was 7.26 to 34.1 pg Gd/cell). Cell viability and proliferation were unaffected at the concentrations tested and up to at least 3 days postlabeling. Gd₂O₃ is a promising sensitive and stable positive contrast agent for cellular MRI at 7 T.

Conjugation to Biocompatible Dendrimers Increases Lanthanide T2 Relaxivity of Hydroxypyridinone (HOPO) Complexes for Magnetic Resonance Imaging (MRI).

Magnetic resonance imaging (MRI) contrast agents represent a worldwide billion-dollar market annually. While T1 relaxivity enhancement contrast agents receive greater attention and a significantly larger market share, the commercial potential for T2 relaxivity enhancing contrast agents remains a viable diagnostic option due to their increased relaxivity at high field strengths. Improving the contrast and biocompatibility of T2 MRI probes may enable new diagnostic prospects for MRI. Paramagnetic lanthanides have the potential to decrease T1 and T2 proton relaxation times, but are not commercially used in MRI diagnostics as T2 agents. In this article, oxygen donor chelates (hydroxypyridinone, HOPO, and terephthalamide, TAM) of various lanthanides are demonstrated as biocompatible macromolecular dendrimer conjugates for the development of T2 MRI probes. These conjugates have relaxivities up to 374 mm-1s-1 per dendrimer, high bioavailability, and low in vitro toxicity.

Silica-coated Iron-oxide Nanoparticles Synthesized as a T2 Contrast Agent for Magnetic Resonance Imaging by Using the Reverse Micelle Method

We synthesized iron-oxide (Fe3O4) nanoparticles by using the reverse micelle method and coated them with biocompatible silica. The coated nanoparticles were found to be spherical in the TEM images and showed a uniform size distribution with an average diameter of 10 nm. The T1 and the T2 relaxation times of hydrogen protons in aqueous solutions with various concentrations of silicacoated nanoparticles were determined by using a magnetic resonance (MR) scanner. We found that the T2 relaxivity was much larger than the T1 relaxivity for the nanoparticle contrast agent, which reflected the fact that the T2 relaxation was mainly influenced by outer sphere processes. The T2 relaxivity was found to be 15 times larger than that for the commercial Gd-DTPA-BMA contrast agent. This result demonstrates that silica-coated iron oxide nanoparticles are applicable as a T2 agent in magnetic resonance imaging.

NMR relaxation and magnetic properties of superparamagnetic nanoworms

Maghemite particles are used as T₂ contrast agents for magnetic resonance imaging, especially for molecular and cellular imaging. Linear clusters of particles - called nanoworms - were recently developed to enhance the targeting efficiency. In this work, the magnetic and NMR relaxation properties of these nanoworms are studied at multiple magnetic fields. After the usual saturation at 0.5 T, the magnetization of the worms is still increasing, which results in an appreciable increase of the transverse relaxivity at high magnetic fields. The obtained relaxivities are typical of superparamagnetic particles of iron oxide (SPIOs). The transverse relaxation of the worms is clearly more efficient than for the isolated grains, which is confirmed by computer simulations. At high field, the longitudinal relaxation of the worms is less pronounced than for the grains, as expected for SPIOs. The nanoworms thus constitute a promising T₂ agent for cellular and molecular imaging.

Modulation of T2 Relaxation Time by Light-Induced, Reversible Aggregation of Magnetic Nanoparticles

A reversible T2 contrast agent consisting of cross-linked anionic dextran coated iron oxide nanoparticles covalently coupled to a light-sensitive spiropyran (SP)/merocyanine (MC) motif was synthesized and characterized. In aqueous solution, light induced isomerization of the molecular switches between the hydrophobic SP isomer and hydrophilic MC isomer directs the aggregation and dispersion of the nanoparticles, respectively. When in the dark, where the MC form dominates, the probe has a T2 relaxation time of 37.09 ms (60 MHz, 37 degrees C) and two size populations at 70 and 540 nm. After irradiation with visible light, the T2 relaxation time is shortened 33.7%, and the size correspondingly shifts to a single population at 520 nm upon aggregation. This "smart" T2 agent provides the advantage of reversibility which may enable dynamic monitoring with MRI. In addition, the light responsiveness of this agent suggests the potential to employ them as MRI gene reporters for the luciferase expression system.

Tumor-targeting, superparamagnetic polymeric vesicles as highly efficient MRI contrast probes

Tumor-targeting polymeric vesicles were prepared using a mixture of two amphiphilic block copolymers, namely, folate-poly(ethylene glycol)-poly(D,L-lactide) (folate-PEG-PDLLA) and NH2-PEG-PDLLA. The copolymers were synthesized through sequential anionic polymerization in a well-controlled manner. Hydrophilic superparamagnetic iron oxide nanoparticles (SPIONs) were encapsulated in the aqueous core of the polymeric vesicles. Similar to the hydrophilic SPIONs, the SPION-loaded polymeric vesicles exhibited the superparamagnetic property. Moreover, the SPION-loaded polymeric vesicles demonstrated a higher r2/r1 value than a commercially available T2 agent, Resovist®, which can be attributed to the high SPION loading level and the SPION clustering effect in the aqueous core of the vesicles. Thus, tumor-targeting, SPION-loaded polymer vesicles can serve as highly efficient T2 contrast probes for cancer diagnosis and treatment.

Ultrasmall iron oxide nanoparticles: Synthesis, physicochemical, and magnetic properties

A new synthetic method leading to the formation of polycaprolactone (PCL)/iron oxide nanoparticles using polyvinylpyrrolidone (PVP) is described. The unique feature of this method is that the conventional polymerization step can be avoided. A stable colloidal dispersion of the PCL-coated iron oxide nanoparticles (PCLNP) is thus formed in the presence of PVP as a stabilizer. The PCLNP has a mean diameter of 20–50 nm as measured by TEM. Magnetic measurement by SQUID shows that the PCLNP has a strong superparamagnetism to be used as a liver MR T2 agent.

The detection limit of a Gd3+-based T1 agent is substantially reduced when targeted to a protein microdomain

Simple low molecular weight (MW) chelates of Gd(3+) such as those currently used in clinical MRI are considered too insensitive for most molecular imaging applications. Here, we evaluated the detection limit (DL) of a molecularly targeted low MW Gd(3+)-based T(1) agent in a model where the receptor concentration was precisely known. The data demonstrate that receptors clustered together to form a microdomain of high local concentration can be imaged successfully even when the bulk concentration of the receptor is quite low. A GdDO3A-peptide identified by phage display to target the anti-FLAG antibody was synthesized, purified and characterized. T(1-)weighted MR images were compared with the agent bound to antibody in bulk solution and with the agent bound to the antibody localized on agarose beads. Fluorescence competition binding assays show that the agent has a high binding affinity (K(D)=150 nM) for the antibody, while the fully bound relaxivity of the GdDO3A-peptide/anti-FLAG antibody in solution was a relatively modest 17 mM(-1) s(-1). The agent/antibody complex was MR silent at concentrations below approximately 9 microM but was detectable down to 4 microM bulk concentrations when presented to antibody clustered together on the surface of agarose beads. These results provided an estimate of the DLs for other T(1)-based agents with higher fully bound relaxivities or multimeric structures bound to clustered receptor molecules. The results demonstrate that the sensitivity of molecularly targeted contrast agents depends on the local microdomain concentration of the target protein and the molecular relaxivity of the bound complex. A model is presented, which predicts that for a molecularly targeted agent consisting of a single Gd(3+) complex with bound relaxivity of 100 mM(-1) s(-1) or, more reasonably, four tethered Gd(3+) complexes each having a bound relaxivity of 25 mM(-1) s(-1), the DL of a protein microdomain is approximately 690 nM at 9.4 T. These experimental and extrapolated DLs are both well below current literature estimates and suggests that detection of low MW molecularly targeted T(1) agents is not an unrealistic goal.

Transcription MRI: A New View of the Living Brain

Altered gene activities are underlying causes of many neurological disorders. The ability to detect, image, and report endogenous gene transcription using magnetic resonance (MR) holds great potential for providing significant clinical benefits. In this review, we present the development of conjugates consisting of gene-targeting short nucleic acids (oligodeoxynucleotides, or sODN) and superparamagnetic iron oxide nanoparticles (SPION, an MR susceptibility T2 agent) for reporting gene activity using transcription MRI (tMRI). We will discuss 1) the target specificity of sODN, 2) selection of contrast agents for tMRI, 3) the distribution and uptake, 4) sequence specificity, 5) histology of SPION and sODN, 6) data acquisition and quantitative analysis for tMRI, and 7) application of gene transcript—targeting nanoparticles in biology and medicine. We will also discuss methods of validating the correlation between results from conventional assays (in situ hybridization, PCR, histology Prussian blue stain and immunohistochemistry) in postmortem samples and retention of SPION-sODN using tMRI. The application of our novel contrast probe to report and target gene transcripts in the mesolimbic pathways of living mouse brains after amphetamine exposure will be discussed. Because of the targeting ability in the nucleic acid sequence, the concept of tMRI probes with complementary nucleic acid (antisense DNA or short interfering RNA) allows not only tracking, targeting, binding to intracellular mRNA, and manipulating gene action but also tracing cells with specific gene action in living brains. Transcription MRI will lend itself to myriad applications in living organs. NEUROSCIENTIST 14(5):503—520, 2008. DOI: 10.1177/1073858407309746