Molecular MRI has shown great promise in imaging of cardiovascular diseases. But new avenues are now being explored. MR probes are being investigated as vehicles for delivering therapeutics, multi-modal probes that use multiple imaging reporters are being utilized for detection of disease and integrated imaging systems that simultaneously image using two different modalities are being constructed and employed in imaging. Large constructs such as liposomes that carry 10-1000s of Gd ions have the potential to carry therapeutic loads as well. For instance, gadolinium(III) loaded liposomes targeted to αV
integrin (involved in plaque angiogenesis) were used in detection of neovasculature in plaques [62
]. On incorporation of the anti-angiogenic agent fumagilin into the probe, reduction in angiogenesis was observed which in turn decreased the uptake of the probe [87
]. These constructs act both in diagnosis and therapy of the disease and are sometimes called theranostics. These theranostics may help in detecting the disease and administering and monitoring therapy.
Although molecular MRI is well suited to image many important processes, multimodal-imaging strategies are being actively pursued concurrently. Dual MR-fluorescence probes have been developed and allow direct detection of plaque components using fluorescence imaging techniques. The high spatial resolution of fluorescence microscopy enables the fluorescence reporters in the probe to confirm probe localization ex vivo, providing an important validation strategy. Recently a trimodal probe using PET, fluorescence and magnetic-detectable moieties was synthesized by conjugating 64
Cu to a magneto-fluorescent probe [13
]. In vivo imaging of the plaque macrophages in the aortic roots of ApoE-/-
mice was successfully performed using MR and PET after injection of the trimodal probe. Similar type of multi-modal probes will probably be very significant in the future of molecular imaging of cardiovascular diseases.
Integrated MR-PET systems have been constructed for human imaging and will allow MR and PET imaging to be performed simultaneously, rather than sequentially as done with PET-CT [88
]. These systems will likely play an important role in the clinical translation of molecular imaging.
Within MRI, newer strategies of molecular MR imaging are being developed using either heteronuclei such as 19
] and hyperpolarized 13
] or exchange of magnetization from one pool of protons to the other as is done in chemical exchange saturation transfer (CEST) [94
19F-MRI uses fluorinated compounds such as perfluorocarbon emulsions to produce contrast. 19F is a suitable nucleus for MR due to its high magnetogyric ratio and high natural abundancy. In addition, 19F is not present in the body; hence the background in 19F-MRI is very low. However this technique also has its limitations. 19F-MRI has low sensitivity with a detection limit in the millimolar range, which is not suitable for detection of most in vivo targets (recall that conventional 1H MRI detects water present at about 90 molar). So molecular imaging probes that can deliver very large numbers of 19F per molecule will be required.
C MRI uses a water-soluble 13
C-enriched compound with a long relaxation time for imaging. Typically the 13
C concentration of the injected compound is 0.3-0.5 M and it is hyperpolarized to 20-40%. A major benefit of hyperpolarized contrast media is the excellent sensitivity with no background (high signal-to-noise ratio). Challenges include the distribution and availability of the hyperpolarization equipment and imaging hardware compatibility for imaging nonhydrogen nuclei, which is not available on all clinical scanners. However, commercial polarizers are now available. Hyperpolarized 13
C contrast agents were used for magnetic resonance angiography in rats and swine [93
]. Hyperpolarized 13
C MRI also offers the potential for investigating metabolic pathways. For instance, Merritt and colleagues studied the metabolism of [1-13
C]-pyruvate in a perfused rat heart. This field is likely to expand in the coming years.
CEST agents are based on the transfer of magnetization from a pool of exchangeable protons on the probe (e.g. amide N-H) to the free water protons [95
]. CEST agents contain exchangeable protons with a resonance frequency well separated from that of the free water peak. Upon selective saturation of the exchangeable proton peak, a drop in the free water peak is observed due to transfer of saturated magnetization. Polypeptides and sugars are used as diamagnetic CEST agents. Paramagnetic lanthanide (Eu, Dy, Ho, Er, Tm, Yb) chelates are employed as paramagnetic CEST agents (PARACEST) because of their large chemical shifts and fast exchange rates [94
]. CEST agents are attractive from the perspective of being “turned on” by the appropriate pulse sequence and also because of the potential for detecting multiple probes at once (provided the different probes have exchangeable hydrogens with different resonance frequencies) [94
]. A major challenge for this class of compounds remains sensitivity; thus far it appears that detection limits are an order of magnitude or more higher than for gadolinium or iron oxide based probes.
Molecular MRI is already playing an important role in basic science investigation and development of novel pharmaceuticals and has the potential of becoming an important tool in the detection of cardiovascular diseases. Serial, noninvasive imaging is also contributing to preclinical development where repeated measurements can be made to assess the extent of disease and response to treatment. Molecular MRI provides complementary information to that provided by genomic and proteomic readouts. It will help to detect and follow the changes in atherosclerotic plaque composition in vivo and identify the most relevant markers of plaque instability, which is the major cause of heart diseases.
- There are 4 different categories of molecular MR agents; superparamagnetic iron-oxide particles, discrete targeted peptides conjugated with gadolinium chelate(s), self-assembled gadolinium nanoparticles, and activatable “smart” probes.
- The superparamagnetic iron oxide nanoparticles are T2-contrast agents and produce negative contrast. They are prepared in variable sizes from 2-4000 nm and differ in their clearance properties. USPIO have been used to image macrophages associated with atherosclerotic plaque, surface receptors such as vascular cell adhesion molecule-1 (VCAM-1) and E-selectin, thrombosis components such as fibrin and activated platelets, and apoptosis.
- The discrete targeted peptides conjugated to Gd chelates are T1-contrast agents and produce positive contrast. These compounds are smaller than 10 kDa. They show fast uptake at the target and fast blood and renal clearance resulting in high target:background signal. Peptides-Gd conjugates have been used to target fibrin, an important marker of thrombosis and collagen, a protein which is upregulated in fibrosis.
- Self-assembled Gd constructs such as micelles or liposomes can carry large Gd payloads, 10s to 1000s of Gd3+ ions. They show slow blood clearance, and optimal imaging times may be 24 or 48 hrs post injection to minimize vascular background signal. Self-assembled Gd nanoparticles have been used to target atherosclerotic plaque components such as ECM proteins, macrophages and integrins involved in plaque angiogenesis.
- Activatable “smart” agents are chemically engineered substrates that undergo a physiochemical change as a result of enzymatic cleavage, pH change, cell internalization or interaction with ions. The effect of this activation is either an increase in the number of inner sphere water molecules, a polymerization of the substrate, or initiation of protein binding, all resulting in higher relaxivities and high target:background signal. Activatable probes that are based on enzyme activity of myeloperoxidase, tyrosinase, blood coagulation factor XII, thrombin-activatable fibrinolysis inhibitor (TAFI) and galactosidase have been used in molecular MR imaging.
- There is no one ideal strategy for molecular imaging. Each has its benefits and limitations and the best approach to take will likely depend on the scientific or clinical question to be answered.
- The challenges in the field of molecular imaging include optimization of the relaxivity of the probe without compromising the binding affinity for a particular target, the toxicity concerns associated with the probes and the economic and regulatory hurdles for clinical translation.
- The future of molecular MR imaging will likely include the use of multi-modal probes, exploitation of integrated systems for imaging and combined diagnostic-therapeutic applications for the nanoparticle-based contrast agents. The contribution of other MRI strategies including MRI using nuclei such as 19F and hyperpolarized 13C, and chemical exchange saturation transfer (CEST) agents will likely grow in the coming years.