Carbon-13 Hyperpolarization of α-Ketocarboxylates with Parahydrogen in Reversible Exchange.

Signal Amplification by Reversible Exchange (SABRE) is a relatively simple and fast hyperpolarization technique that has been used to hyperpolarize the α-ketocarboxylate pyruvate, a central metabolite and the leading hyperpolarized MRI contrast agent. In this work, we show that SABRE can readily be extended to hyperpolarize 13C nuclei at natural abundance on many other α-ketocarboxylates. Hyperpolarization is observed and optimized on pyruvate (P13C=17%) and 2-oxobutyrate (P13C=25%) with alkyl chains in the R-group, oxaloacetate (P13C=11%) and alpha-ketoglutarate (P13C=13%) with carboxylate moieties in the R group, and phenylpyruvate (P13C=2%) and phenylglyoxylate (P13C=2%) with phenyl rings in the R-group. New catalytically active SABRE binding motifs of the substrates to the hyperpolarization transfer catalyst-particularly for oxaloacetate-are observed. We experimentally explore the connection between temperature and exchange rates for all of these SABRE systems and develop a theoretical kinetic model, which is used to fit the hyperpolarization build-up and decay during SABRE activity.

Modeling Ligand Exchange Kinetics in Iridium Complexes Catalyzing SABRE Nuclear Spin Hyperpolarization.

Large signal enhancements can be obtained for NMR analytes using the process of nuclear spin hyperpolarization. Organometallic complexes that bind parahydrogen can themselves become hyperpolarized. Moreover, if parahydrogen and a to-be-hyperpolarized analyte undergo chemical exchange with the organometallic complex it is possible to catalytically sensitize the detection of the analyte via hyperpolarization transfer through spin-spin coupling in this organometallic complex. This process is called Signal Amplification By Reversible Exchange (SABRE). Signal intensity gains of several orders of magnitude can thus be created for various compounds in seconds. The chemical exchange processes play a defining role in controlling the efficiency of SABRE because the lifetime of the complex must match the spin-spin couplings. Here, we show how analyte dissociation rates in the key model substrates pyridine (the simplest six-membered heterocycle), 4-aminopyridine (a drug), and nicotinamide (an essential vitamin biomolecule) can be examined. This is achieved for the most widely employed SABRE motif that is based on IrIMes-derived catalysts by 1H 1D and 2D exchange NMR spectroscopy techniques. Several kinetic models are evaluated for their accuracy and simplicity. By incorporating variable temperature analysis, the data yields key enthalpies and entropies of activation that are critical for understanding the underlying SABRE catalyst properties and subsequently optimizing behavior through rational chemical design. While several studies of chemical exchange in SABRE have been reported, this work also aims to establish a toolkit on how to quantify chemical exchange in SABRE and ensure that data can be compared reliably.

Toward Ultra-High-Quality-Factor Wireless Masing Magnetic Resonance Sensing.

It has recently been shown that a bolus of hyperpolarized nuclear spins can yield stimulated emission signals similar in nature to maser signals, potentially enabling new ways of sensing hyperpolarized contrast media, including most notably [1-13C]pyruvate that is under evaluation in over 50 clinical trials for metabolic imaging of cancer. The stimulated NMR signal emissions lasting for minutes do not require radio-frequency excitation, offering unprecedented advantages compared to conventional MR sensing. However, creating nuclear spin maser emission is challenging in practice due to stringent fundamental requirements, making practical in vivo applications hardly possible using conventional passive MR detectors. Here, we demonstrate the utility of a wireless NMR maser detector, the quality factor of which was enhanced 22-fold (to 1,670) via parametric pumping. This active-feedback technique breaks the intrinsic fundamental limit of NMR detector circuit quality factor. We show the use of parametric pumping to reduce the threshold requirement for inducing nuclear spin masing at 300 MHz resonance frequency in a preclinical MRI scanner. Indeed, stimulated emission from hyperpolarized protons was obtained under highly unfavorable conditions of low magnetic field homogeneity (T2* of 3 ms). Greater gains of the quality factor of the MR detector (up to 1 million) were also demonstrated.

In vivo Metabolic Sensing of Hyperpolarized [1-13C]Pyruvate in Mice Using a Recyclable Perfluorinated Iridium Signal Amplification by Reversible Exchange Catalyst.

Real-time visualization of metabolic processes in vivo provides crucial insights into conditions like cancer and metabolic disorders. Metabolic magnetic resonance imaging (MRI), by amplifying the signal of pyruvate molecules through hyperpolarization, enables non-invasive monitoring of metabolic fluxes, aiding in understanding disease progression and treatment response. Signal Amplification By Reversible Exchange (SABRE) presents a simpler, cost-effective alternative to dissolution dynamic nuclear polarization, eliminating the need for expensive equipment and complex procedures. We present the first in vivo demonstration of metabolic sensing in a human pancreatic cancer xenograft model compared to healthy mice. A novel perfluorinated Iridium SABRE catalyst in a fluorinated solvent and methanol blend facilitated this breakthrough with a 1.2-fold increase in [1-13C]pyruvate SABRE hyperpolarization. The perfluorinated moiety allowed easy separation of the heavy-metal-containing catalyst from the hyperpolarized [1-13C]pyruvate target. The perfluorinated catalyst exhibited recyclability, maintaining SABRE-SHEATH activity through subsequent hyperpolarization cycles with minimal activity loss after the initial two cycles. Remarkably, the catalyst retained activity for at least 10 cycles, with a 3.3-fold decrease in hyperpolarization potency. This proof-of-concept study encourages wider adoption of SABRE hyperpolarized [1-13C]pyruvate MR for studying in vivo metabolism, aiding in diagnosing stages and monitoring treatment responses in cancer and other diseases.

Toward Next-Generation Molecular Imaging with a Clinical Low-Field (0.064 T) Point-of-Care MRI Scanner.

Low-field (LF) MRI promises soft-tissue imaging without the expensive, immobile magnets of clinical scanners but generally suffers from limited detection sensitivity and contrast. The sensitivity boost provided by hyperpolarization can thus be highly synergistic with LF MRI. Initial efforts to integrate a continuous-bubbling SABRE (signal amplification by reversible exchange) hyperpolarization setup with a portable, point-of-care 64 mT clinical MRI scanner are reported. Results from 1H SABRE MRI of pyrazine and nicotinamide are compared with those of benchtop NMR spectroscopy. Comparison with MRI signals from samples with known H2O/D2O ratios allowed quantification of the SABRE enhancements of imaged samples with various substrate concentrations (down to 3 mM). Respective limits of detection and quantification of 3.3 and 10.1 mM were determined with pyrazine 1H polarization (PH) enhancements of ∼1900 (PH ∼0.04%), supporting ongoing and envisioned efforts to realize SABRE-enabled MRI-based molecular imaging.

15N Hyperpolarization of Metronidazole Antibiotic in Aqueous Media Using Phase-Separated Signal Amplification by Reversible Exchange with Parahydrogen.

Metronidazole is a prospective hyperpolarized MRI contrast agent with potential hypoxia sensing utility for applications in cancer, stroke, neurodegenerative diseases, etc. We demonstrate a pilot procedure for production of ∼30 mM hyperpolarized [15N3]metronidazole in aqueous media by using a phase-separated SABRE-SHEATH hyperpolarization method, with nitrogen-15 polarization exceeding 2.2% on all three 15N sites achieved in less than 2 min. The 15N polarization T1 of ∼12 min is reported for the 15NO2 group at the clinically relevant field of 1.4 T in the aqueous phase, demonstrating a remarkably long lifetime of the hyperpolarized state. The produced aqueous solution of [15N3]metronidazole that contained only ∼100 µM of residual Ir was deemed biocompatible via validation through the MTT colorimetric test for assessing cell metabolic activity using human embryotic kidney HEK293T cells. This low-cost and ultrafast hyperpolarization procedure represents a major advance for the production of a biocompatible HP [15N3]metronidazole (and potentially other hyperpolarized drugs) formulation for MRI sensing applications.

Toward Lung Ventilation Imaging Using Hyperpolarized Diethyl Ether Gas Contrast Agent.

Hyperpolarized 129Xe gas was FDA-approved as an inhalable contrast agent for magnetic resonance imaging of a wide range of pulmonary diseases in December 2022. Despite the remarkable success in clinical research settings, the widespread clinical translation of HP 129Xe gas faces two critical challenges: the high cost of the relatively low-throughput hyperpolarization equipment and the lack of 129Xe imaging capability on clinical MRI scanners, which have narrow-bandwidth electronics designed only for proton (1H) imaging. To solve this translational grand challenge of gaseous hyperpolarized MRI contrast agents, here we demonstrate the utility of batch-mode production of proton-hyperpolarized diethyl ether gas via heterogeneous pairwise addition of parahydrogen to ethyl vinyl ether. An approximately 0.1-liter bolus of hyperpolarized diethyl ether gas was produced in 1 second and injected in excised rabbit lungs. Lung ventilation imaging was performed using sub-second 2D MRI with up to 2×2 mm2 in-plane resolution using a clinical 0.35 T MRI scanner without any modifications. This feasibility demonstration paves the way for the use of inhalable diethyl ether as a gaseous contrast agent for pulmonary MRI applications using any clinical MRI scanner.

MATRESHCA: Microtesla Apparatus for Transfer of Resonance Enhancement of Spin Hyperpolarization via Chemical Exchange and Addition.

We present an integrated, open-source device for parahydrogen-based hyperpolarization processes in the microtesla field regime with a cost of components of less than $7000. The device is designed to produce a batch of 13C and 15N hyperpolarized (HP) compounds via hydrogenative or non-hydrogenative parahydrogen-induced polarization methods that employ microtesla magnetic fields for efficient polarization transfer of parahydrogen-derived spin order to X-nuclei (e.g., 13C and 15N). The apparatus employs a layered structure (reminiscent of a Russian doll "Matryoshka") that includes a nonmagnetic variable-temperature sample chamber, a microtesla magnetic field coil (operating in the range of 0.02-75 microtesla), a three-layered mu-metal shield (to attenuate the ambient magnetic field), and a magnetic shield degaussing coil placed in the overall device enclosure. The gas-handling manifold allows for parahydrogen-gas flow and pressure control (up to 9.2 bar of total parahydrogen pressure). The sample temperature can be varied either using a water bath or a PID-controlled heat exchanger in the range from -12 to 80 °C. This benchtop device measures 62 cm (length) × 47 cm (width) × 47 cm (height), weighs 30 kg, and requires only connections to a high-pressure parahydrogen gas supply and a single 110/220 VAC power source. The utility of the device has been demonstrated using an example of parahydrogen pairwise addition to form HP ethyl [1-13C]acetate (P13C = 7%, [c] = 1 M). Moreover, the Signal Amplification By Reversible Exchange in SHield Enables Alignment Transfer to Heteronuclei (SABRE-SHEATH) technique was employed to demonstrate efficient hyperpolarization of 13C and 15N spins in a wide range of biologically relevant molecules, including [1-13C]pyruvate (P13C = 14%, [c] = 27 mM), [1-13C]-α-ketoglutarate (P13C = 17%), [1-13C]ketoisocaproate (P13C = 18%), [15N3]metronidazole (P15N = 13%, [c] = 20 mM), and others. While the vast majority of the utility studies have been performed in standard 5 mm NMR tubes, the sample chamber of the device can accommodate a wide range of sample container sizes and geometries of up to 1 L sample volume. The device establishes an integrated, simple, inexpensive, and versatile equipment gateway needed to facilitate parahydrogen-based hyperpolarization experiments ranging from basic science to preclinical applications; indeed, detailed technical drawings and a bill of materials are provided to support the ready translation of this design to other laboratories.

Carbon-13 Radiofrequency Amplification by Stimulated Emission of Radiation of the Hyperpolarized Ketone and Hemiketal Forms of Allyl [1-13C]Pyruvate.

13C hyperpolarized pyruvate is an emerging MRI contrast agent for sensing molecular events in cancer and other diseases with aberrant metabolic pathways. This metabolic contrast agent can be produced via several hyperpolarization techniques. Despite remarkable success in research settings, widespread clinical adoption faces substantial roadblocks because the current sensing technology utilized to sense this contrast agent requires the excitation of 13C nuclear spins that also need to be synchronized with MRI field gradient pulses. Here, we demonstrate sensing of hyperpolarized allyl [1-13C]pyruvate via the stimulated emission of radiation that mitigates the requirements currently blocking broader adoption. Specifically, 13C Radiofrequency Amplification by Stimulated Emission of Radiation (13C RASER) was obtained after pairwise addition of parahydrogen to a pyruvate precursor, detected in a commercial inductive detector with a quality factor (Q) of 32 for sample concentrations as low as 0.125 M with 13C polarization of 4%. Moreover, parahydrogen-induced polarization allowed for the preparation of a mixture of ketone and hemiketal forms of hyperpolarized allyl [1-13C]pyruvate, which are separated by 10 ppm in 13C NMR spectra. This is a good model system to study the simultaneous 13C RASER signals of multiple 13C species. This system models the metabolic production of hyperpolarized [1-13C]lactate from hyperpolarized [1-13C]pyruvate, which has a similar chemical shift difference. Our results show that 13C RASER signals can be obtained from both species simultaneously when the emission threshold is exceeded for both species. On the other hand, when the emission threshold is exceeded only for one of the hyperpolarized species, 13C stimulated emission is confined to this species only, therefore enabling the background-free detection of individual hyperpolarized 13C signals. The reported results pave the way to novel sensing approaches of 13C hyperpolarized pyruvate, potentially unlocking hyperpolarized 13C MRI on virtually any MRI system─an attractive vision for the future molecular imaging and diagnostics.