Protein−ligand interactions are frequently screened using nuclear magnetic resonance (NMR) spectroscopy. The dissociation constant (KD) of a ligand of interest can be determined via a spin-spin relaxation measurement of a reporter ligand, in a single scan when using hyperpolarization by means of dissolution dynamic nuclear polarization (D-DNP). Despite nearly instantaneous signal acquisition, a limitation of D-DNP for the screening of protein−ligand interactions is the required polarization time on the order of tens of minutes. Here, we introduce a multiplexed NMR experiment, where a single hyperpolarized ligand sample is rapidly mixed with protein injected into two flow cells. NMR detection is achieved simultaneously on both channels, resulting in a chemical shift resolved spin relaxation measurement. Spectral resolution allows the use of reference compounds for accurate quantification of concentrations. Simultaneous use of two concentration ratios between protein and ligand broadens the range of KD that is accurately measurable in a single experiment to at least an order of magnitude. In a comparison of inhibitors for the protein trypsin, the average KD values of benzamidine and benzylamine were found to be 12.6±1.4 μM and 207±22.3 μM from three measurements, based on KD = 142 μM assumed known for the reporter ligand 4-(trifluoromethyl)benzene-1-carboximidamide. Typical confidence ranges at 95% evaluated for single experiments were (8.3 μM, 20 μM) and (151 μM, 328 μM). The multiplexed detection of two or more hyperpolarized samples increases throughput of D-DNP by the same factor, improving the applicability to most multi-point measurements that would traditionally be achieved using titrations.

Fluorine-19 NMR and hyperpolarization form a powerful combination for drug screening. Under a competitive equilibrium with a selected fluorinated reporter ligand, the dissociation constant (KD) of other ligands of interest is measurable using a single-scan Carr-Purcell-Meiboom-Gill (CPMG) experiment, without the need for a titration. This method is demonstrated by characterizing the binding of three ligands with different affinities for the serine protease trypsin. Monte Carlo simulations show that the highest accuracy is obtained when about one-half of the bound reporter ligand is displaced in the binding competition. Such conditions can be achieved over a wide range of affinities, allowing for rapid screening of non-fluorinated compounds when a single fluorinated ligand to the binding pocket of interest is known.

Purpose To improve hyperpolarized 13C (HP‐13C) MRI by image denoising with a new approach, patch‐based higher‐order singular value decomposition (HOSVD). Methods The benefit of using a patch‐based HOSVD method to denoise dynamic HP‐13C MR imaging data was investigated. Image quality and the accuracy of quantitative analyses following denoising were evaluated first using simulated data of [1‐13C]pyruvate and its metabolic product, [1‐13C]lactate, and compared the results to a global HOSVD method. The patch‐based HOSVD method was then applied to healthy volunteer HP [1‐13C]pyruvate EPI studies. Voxel‐wise kinetic modeling was performed on both non‐denoised and denoised data to compare the number of voxels quantifiable based on SNR criteria and fitting error. Results Simulation results demonstrated an 8‐fold increase in the calculated SNR of [1‐13C]pyruvate and [1‐13C]lactate with the patch‐based HOSVD denoising. The voxel‐wise quantification of kPL (pyruvate‐to‐lactate conversion rate) showed a 9‐fold decrease in standard errors for the fitted kPL after denoising. The patch‐based denoising performed superior to the global denoising in recovering kPL information. In volunteer data sets, [1‐13C]lactate and [13C]bicarbonate signals became distinguishable from noise across captured time points with over a 5‐fold apparent SNR gain. This resulted in >3‐fold increase in the number of voxels quantifiable for mapping kPB (pyruvate‐to‐bicarbonate conversion rate) and whole brain coverage for mapping kPL. Conclusions Sensitivity enhancement provided by this denoising significantly improved quantification of metabolite dynamics and could benefit future studies by improving image quality, enabling higher spatial resolution, and facilitating the extraction of metabolic information for clinical research.

Porous silicon nanoparticles have recently garnered attention as potentially-promising biomedical platforms for drug delivery and medical diagnostics. Here, we demonstrate porous silicon nanoparticles as contrast agents for Si magnetic resonance imaging. Size-controlled porous silicon nanoparticles were synthesized by magnesiothermic reduction of silica nanoparticles and were surface activated for further functionalization. Particles were hyperpolarized via dynamic nuclear polarization to enhance their Si MR signals; the particles demonstrated long Si spin-lattice relaxation (T ) times (∼25 mins), which suggests potential applicability for medical imaging. Furthermore, Si hyperpolarization levels were sufficient to allow Si MRI in phantoms. These results underscore the potential of porous silicon nanoparticles that, when combined with hyperpolarized magnetic resonance imaging, can be a powerful theragnostic deep tissue imaging platform to interrogate various biomolecular processes in vivo.

Background Hyperpolarized 13C MRI quantitatively measures enzyme‐catalyzed metabolism in cancer and metabolic diseases. Whole‐abdomen imaging will permit dynamic metabolic imaging of several abdominal organs simultaneously in healthy and diseased subjects. Purpose Image hyperpolarized [1‐13C]pyruvate and products in the abdomens of healthy volunteers, overcoming challenges of motion, magnetic field variations, and spatial coverage. Compare hyperpolarized [1‐13C]pyruvate metabolism across abdominal organs of healthy volunteers. Study Type Prospective technical development. Subjects A total of 13 healthy volunteers (8 male), 21–64 years (median 36). Field Strength/Sequence A 3 T. Proton: T1‐weighted spoiled gradient echo, T2‐weighted single‐shot fast spin echo, multiecho fat/water imaging. Carbon‐13: echo‐planar spectroscopic imaging, metabolite‐specific echo‐planar imaging. Assessment Transmit magnetic field was measured. Variations in main magnetic field (ΔB0) determined using multiecho proton acquisitions were compared to carbon‐13 acquisitions. Changes in ΔB0 were measured after localized shimming. Improvements in metabolite signal‐to‐noise ratio were calculated. Whole‐organ regions of interests were drawn over the liver, spleen, pancreas, and kidneys by a single investigator. Metabolite signals, time‐to‐peak, decay times, and mean first‐order rate constants for pyruvate‐to‐lactate (kPL) and alanine (kPA) conversion were measured in each organ. Statistical Tests Linear regression, one‐sample Kolmogorov–Smirnov tests, paired t‐tests, one‐way ANOVA, Tukey's multiple comparisons tests. P ≤ 0.05 considered statistically significant. Results Proton ΔB0 maps correlated with carbon‐13 ΔB0 maps (slope = 0.93, y‐intercept = −2.88, R2 = 0.73). Localized shimming resulted in mean frequency offset within ±25 Hz for all organs. Metabolite SNR significantly increased after denoising. Mean kPL and kPA were highest in liver, followed by pancreas, spleen, and kidneys (all comparisons with liver were significant). Data Conclusion Whole‐abdomen coverage with hyperpolarized carbon‐13 MRI was feasible despite technical challenges. Multiecho gradient echo 1H acquisitions accurately predicted chemical shifts observed using carbon‐13 spectroscopy. Carbon‐13 acquisitions benefited from local shimming. Metabolite energetics in the abdomen compiled for healthy volunteers can be used to design larger clinical trials in patients with metabolic diseases. Evidence Level 2 Technical Efficacy Stage 1