Hyperpolarized ¹³C MRI: State of the Art and Future Directions

Abstract

Hyperpolarized (HP) carbon 13 (¹³C) MRI is an emerging molecular imaging method that allows rapid, noninvasive, and pathway-specific investigation of dynamic metabolic and physiologic processes that were previously inaccessible to imaging. This technique has enabled real-time in vivo investigations of metabolism that are central to a variety of diseases, including cancer, cardiovascular disease, and metabolic diseases of the liver and kidney. This review provides an overview of the methods of hyperpolarization and ¹³C probes investigated to date in preclinical models of disease. The article then discusses the progress that has been made in translating this technology for clinical investigation. In particular, the potential roles and emerging clinical applications of HP [1-¹³C]pyruvate MRI will be highlighted. The future directions to enable the adoption of this technology to advance the basic understanding of metabolism, to improve disease diagnosis, and to accelerate treatment assessment are also detailed.

©RSNA, 2019

Online supplemental material is available for this article.

Summary

Hyperpolarized carbon 13 MRI is an emerging molecular imaging technique that is actively undergoing clinical translation at multiple institutions.

Essentials

• ■ Hyperpolarization, achieved by means of dynamic nuclear polarization, dramatically enhances the MRI signal of carbon 13 (¹³C) labeled molecules by more than 10 000 fold.

• ■ Hyperpolarized ¹³C MRI allows in vivo probing of enzyme-mediated metabolic processes relevant to human diseases.

• ■ Work is ongoing in the clinical translation of hyperpolarized ¹³C MRI, with numerous emerging applications in oncology, diabetes, and heart disease, as well as metabolic diseases of the liver and kidney.

Introduction

Altered metabolism is central to many human diseases, such as cancer, cardiovascular disease, diabetes, and a variety of inflammatory conditions. The most commonly used imaging strategy in the clinic for interrogating metabolism, particularly in cancer, is PET with the glucose analog ¹⁸F fluorodeoxyglucose (FDG). FDG PET provides information regarding tissue glucose uptake and has been highly clinically successful. However, it cannot help assess downstream metabolism, which may be useful in the diagnosis and treatment monitoring of a variety of diseases.

Carbon 13 (¹³C) MRI is particularly attractive for metabolic imaging because carbon serves as the backbone of nearly all organic molecules, thus allowing the investigation of a wide range of biochemical processes that are relevant to human diseases. However, the low natural abundance of the ¹³C isotope, at 1.1%, has made in vivo imaging extremely challenging. This limitation has been overcome by the recent development of the dynamic nuclear polarization technique, which can dramatically, albeit temporarily, increase the signal of ¹³C-labeled molecules by more than 10 000 fold (1). Hyperpolarized (HP) ¹³C MRI has emerged as a powerful molecular imaging strategy that allows safe, nonradioactive, real-time, and pathway-specific investigation of dynamic metabolic and physiologic processes that were previously inaccessible to imaging. In this review, we will provide an overview of the methods of hyperpolarization and the various biologic processes that can be interrogated by using HP ¹³C probes, with a focus on HP ¹³C pyruvate. We will also summarize the technical and regulatory requirements of human HP ¹³C studies and highlight the emerging clinical applications of this molecular imaging technology.

Hyperpolarization

At body temperature and field strengths used in clinical MRI, the MRI signal of ¹³C endogenous nuclei is very low because of its low natural abundance (only about 1.1% of carbon is ¹³C) and low nuclear spin polarization (the spins are not well aligned to the external magnetic field). The relative signal of ¹³C to hydrogen 1 (¹H) is 0.016 based on the gyromagnetic ratio of ¹³C being approximately one-fourth of that of ¹H. The relative signal of ¹³C is further reduced by the low natural abundance of ¹³C of 1.1%, compared with more than 99% for ¹H. To improve the MRI signal of ¹³C nuclei, probes can be synthetically enriched to increase the concentration of the ¹³C label in a molecule, commonly enriched to 99% of ¹³C. MRI signal can be further increased dramatically by the process of hyperpolarization (Fig 1). The principle of hyperpolarization is based on the fact that at low temperature and high magnetic field, electrons have a very high level of polarization (ie, nearly all the electrons are aligned in the same direction). This high level of polarization can be transferred to ¹³C-labeled probes, increasing their MRI signals. This transfer of polarization is accomplished by mixing radicals (a source of free electrons) with the ¹³C-labeled probe(s) to be hyperpolarized and placing the mixture in a polarizer at a magnetic field typically of 3.0–5.0 T and at a low temperature (approximately 1 K). Microwave irradiation is then applied to transfer the polarization from unpaired electrons in a trityl radical to the ¹³C-labeled probes (1). Depending on the molecule to be polarized and the operating field strength and temperature of the polarizer, the hyperpolarization process takes between 30 and 120 minutes. The frozen HP sample is then rapidly dissolved by a heated and pressurized bolus of a biologically compatible buffer solution. The solution retains a high level of polarization and can be formulated to be at physiologic pH, osmolarity, and temperature for in vivo injection and metabolic investigations. In a 3.0-T field and at room temperature, the ¹³C thermal equilibrium polarization is approximately 0.00025% aligned with the external magnetic field; with hyperpolarization, the polarization increases to approximately 30%–40%, an increase of over 100 000 fold (1), thereby dramatically increasing the MRI signal. The enhanced signal, however, is typically available only for a short period of time (ie, 1–2 minutes), as the polarization decays back to its thermal equilibrium level at a rate dependent on the spin-lattice relaxation time (T1) of the ¹³C labeled nucleus. Therefore, rapid imaging is needed to acquire high signal-to-noise ratio metabolic data with minimal polarization loss and to measure fast metabolic processes.

Figure 1:

Processes for increasing MRI signal of carbon 13 (¹³C) nuclei. DNP = dynamic nuclear polarization, NMR = nuclear magnetic resonance.

HP ¹³C Probes

An advantage of HP ¹³C technology is the diverse array of probes that can be polarized. The most commonly studied HP probes have been endogenous biomolecules modified only by the ¹³C enrichment, and they have been applied to interrogate metabolic and physiologic processes associated with a variety of neoplastic, inflammatory, and metabolic diseases (Table). The selection of the ¹³C enrichment site should take into account two important considerations. First, the labeled carbon atom should have a long longitudinal relaxation time (T1), as the T1 determines how quickly the polarization of the probe decays back to thermal equilibrium once it is removed from the polarizer. Longer T1 facilitates preservation of the enhanced MRI signal and therefore more accurate quantification of metabolism in vivo. Carbon atoms that do not have directly attached protons, such as those in carbonyl groups, usually have longer T1 relaxation times. Another consideration is the chemical shift difference between the probe and its metabolites at the labeled position. Larger differences in chemical shift enable differentiation between the probe and metabolites more readily and therefore enable more accurate metabolic quantification. Successful HP probes additionally must be water soluble at physiologic pH values and have cellular uptake that is sufficiently rapid to allow observation of metabolism during the time frame of the HP study.

Selected Hyperpolarized Carbon 13 Probes Studied to Date

Note.—AcCoA = acetyl-coenzyme A, ALCAR = acetylcarnitine, ALT = alanine transaminase, βOHB = β-hydroxybutyrate, GA3P = glyceraldehyde-3-phosphate, G3P = glycerol-3-phosphate, IDH = isocitrate dehydrogenase, LDH = lactate dehydrogenase, PEP = phosphoenolpyruvate, PDH = pyruvate dehydrogenase, TCA = tricarboxylic acid, 2-HG = 2-hydroxyglutarate.

The most widely studied HP probe to date is [1-¹³C]pyruvate. It polarizes well (up to 50% polarization level in current clinical polarizers) and has a long T1 (approximately 67 seconds in solution at 3.0 T), thereby permitting in vivo investigation with high signal-to-noise ratio. Importantly, [1-¹³C]pyruvate is a highly biologically relevant probe, as pyruvate lies at a critical branch point of multiple metabolic pathways, including glycolysis, the tricarboxylic acid (TCA) cycle, and amino acid biosynthesis (Fig 2). On injection into a living system, HP [1-¹³C]pyruvate is rapidly taken up into the cells and metabolized within the cytosol into [1-¹³C]lactate and [1-¹³C]alanine by the enzymes lactate dehydrogenase (LDH) and alanine transaminase, respectively. HP [1-¹³C]pyruvate is also transported into the mitochondria and is converted by the enzyme pyruvate dehydrogenase (PDH) into ¹³C CO₂ and acetyl-coenzyme A, thereby serving as a readout of PDH activity and flux toward the TCA cycle. [1-¹³C]pyruvate has been used extensively to interrogate metabolism in a variety of diseases such as cancer, ischemia, and inflammation in preclinical models (discussed in detail below). Importantly, it has also been translated for clinical metabolic investigations and has been shown to be safe and feasible (2).

Figure 2:

Schematic of the metabolic pathways of pyruvate. [1-¹³C]pyruvate is rapidly taken up into the cells and metabolized within the cytosol into [1-¹³C]lactate and [1-¹³C]alanine by the enzymes lactate dehydrogenase (LDH) and alanine transaminase (ALT), respectively. Hyperpolarized [1-¹³C]pyruvate is also transported into the mitochondria and is converted by the enzyme pyruvate dehydrogenase (PDH) into ¹³C CO₂ and acetyl Co-A, with CO₂ in rapid equilibrium with ¹³C bicarbonate. TCA = tricarboxylic acid. Red circle = position of ¹³C labeling.

There are numerous other HP ¹³C probes, mainly composed of endogenous biomolecules, that have shown great promise for investigating metabolism relevant to human diseases. Examples include [2-¹³C]pyruvate for probing mitochondrial metabolism (3,4), [1,4-¹³C₂]fumarate for assessing tissue necrosis (5–9), ¹³C bicarbonate for measuring pH (10–13), [1-¹³C]dehydroascorbate (DHA) for interrogating redox capacity (14–18), and ¹³C urea for imaging perfusion (19–22). While many of these probes have so far been investigated only in preclinical disease models, several of them, including [2-¹³C]pyruvate, [1,4-¹³C₂]fumarate, and ¹³C urea are actively being evaluated for clinical translation.

HP Probes in Preclinical Disease Models

Preclinical studies are critical in testing the feasibility and safety of new HP probes and in developing and refining the methods for their use in specific diseases. In addition, preclinical HP investigations have contributed to our understanding of metabolism and its modification in disease processes and following therapy, thereby yielding important information that is of clinical value. There are more than 50 preclinical polarizers installed worldwide, facilitating a large number of scientific investigations in a variety of diseases.

Cancer

An increase in aerobic glycolysis and lactate production, also known as the Warburg effect, is characteristic of many tumors. Specifically, pyruvate generated from glucose metabolism is preferentially converted to lactate by LDH rather than entering the TCA cycle for oxidative metabolism and adenosine triphosphate generation. This lactate production occurs despite the presence of adequate tissue oxygenation. Hence, HP [1-¹³C]pyruvate is an ideal probe to noninvasively interrogate such metabolic reprogramming and has been applied to numerous models of cancer (23–33). For example, increased HP [1-¹³C]pyruvate-to-lactate conversion has been observed in a transgenic adenocarcinoma of mouse prostate (TRAMP) model, with the level of ¹³C lactate correlating with tumor histologic grade (23) (Fig 3). In a Myc oncogene–driven liver cancer model, increased HP [1-¹³C]pyruvate conversion to lactate and alanine was observed to precede tumor formation, and there was a dramatic reversal of pyruvate-to-lactate conversion during early tumor regression before any size change (24). HP [1-¹³C]pyruvate–to-lactate conversion and lactate efflux were able to help differentiate benign renal tumors from renal cell carcinomas in an ex vivo model of patient-derived renal tumor tissues (26). These studies have also demonstrated a mechanistic link between HP lactate signal and cellular alterations such as elevated expression of LDH and monocarboxylate transporters (lactate transporters) that are essential to fully understand cancer metabolism.

Figure 3:

Hyperpolarized (HP) carbon 13 (¹³C) MRI in a transgenic adenocarcinoma mouse model of prostate cancer. A, Representative hematoxylin-eosin–stained pathology sections and HP ¹³C spectra after injection of HP [1-¹³C]pyruvate in a normal mouse prostate, an early-stage prostate tumor, a late-stage prostate tumor, and a lymph node metastasis. At histologic examination, normal murine prostate was glandular, with secretory epithelial cells lining the glands (arrowhead). In prostate tumors, there was gradual replacement of the secretory epithelial cells by less differentiated epithelial cells, until the glands were completely eliminated and only anaplastic sheets of pleomorphic cells with irregular nuclei remained in the late stage tumors (arrow). The ¹³C spectra show an increase in ¹³C lactate and ¹³C lactate/pyruvate ratio in late-stage tumor and nodal metastasis. B, Axial T2-weighted anatomic MR image and overlay of HP ¹³C lactate image on T2-weighted image show a qualitatively high level of lactate in a late-stage tumor. Units for color bar = arbitrary units of signal intensity. C, Boxplots show quantitative ¹³C lactate signal in the four histologically defined groups, with late-stage tumors having significantly higher lactate than early-stage tumors. SNR = signal-to-noise ratio. (Adapted from reference 23.)

HP [1-¹³C]pyruvate has also been shown to be a useful probe to monitor early anticancer therapies. Successful therapies are usually associated with a decrease in pyruvate-to-lactate conversion, mediated by different mechanisms from various treatments (34–45). For example, in a model of glioblastoma, decreased lactate production was observed with early treatment response to everolimus (a mammalian target of rapamycin, or mTOR, inhibitor) before any tumor size change and was associated with a decrease in the LDH enzyme that mediates the pyruvate-to-lactate conversion (37). Treatment with temozolomide (a DNA damaging drug) in a glioblastoma model also resulted in lower lactate production but was related to a decrease in pyruvate kinase PKM2, a glycolytic enzyme that indirectly controls pyruvate metabolism (39). Interestingly, in an ovarian cancer model, treatment with a tyrosine kinase inhibitor (pazopanib) led to significantly higher HP pyruvate-to-lactate conversion at 2 days after treatment initiation, while no change was observed at FDG PET (40). The increased pyruvate-to-lactate conversion was hypothesized to reflect increased hypoxia with elevated lactate production as a result of the antiangiogenic effects of the drug (40).

In addition to [1-¹³C]pyruvate, multiple other probes have been used to study cancer metabolism preclinically. For example, increased HP [1,4-¹³C₂]fumarate–to-malate conversion was seen early following cancer therapy prior to significant changes in tumor size in models of lymphoma (5), breast cancer (46), liver cancer (47), and colon cancer (48), consistent with increased tumor necrosis. Hence, HP fumarate may be useful in providing another measure of early treatment response. HP ¹³C urea has been used in combination with HP [1-¹³C]pyruvate to investigate metabolism/perfusion mismatch in a prostate cancer model, and such mismatch was associated with more aggressive tumors (21). A recent study (49) also utilized a combination of [1-¹³C]pyruvate and HP ¹³C urea to monitor metabolic and perfusion changes following high-intensity focused ultrasound treatment in a TRAMP model. HP ¹³C bicarbonate has shown promise for in vivo pH mapping of tumors (10–13,50), which is of great interest, as an acidic tumor microenvironment is implicated in tumor aggressiveness, metastatic potential, and therapeutic response (51).

These initial studies demonstrate the promise of HP ¹³C MRI in advancing our understanding of cancer metabolism and metabolic response to treatment and, importantly, show the potential of this strategy in detecting early response versus nonresponse, which is critical for enabling adaptive therapies.

Cardiovascular Disease

Alterations in perfusion and energy metabolism are central to many cardiovascular diseases, making them a particularly promising application for HP ¹³C MRI. Perfusion can be mapped quantitatively by studying the distribution of HP ¹³C urea (52). Unlike T1-weighted ¹H MRI using gadolinium-based perfusion agents, the signal arising from HP ¹³C urea is linearly dependent on concentration, thus simplifying quantification. Quantitative perfusion mapping with HP ¹³C urea MRI may be useful in identifying ischemia in patients with balanced hypoperfusion from three-vessel disease, where qualitative gadolinium-enhanced MRI perfusion assessment may be insufficient (53). HP MRI with [1-¹³C]pyruvate can depict acute changes in myocardial metabolism after ischemia and reperfusion, thereby potentially enabling in vivo monitoring of the metabolic effects of reperfusion strategies (54). Additionally, simultaneous polarization and administration of HP ¹³C urea and [1-¹³C]pyruvate permit simultaneous interrogation of cardiac metabolism and perfusion (22). Because HP ¹³C MRI acquisitions take only 1–2 minutes, this modality has a speed advantage over methods such as PET metabolism/perfusion mismatch studies, where the imaging time approaches 30 minutes. The ability to rapidly assess metabolism/perfusion mismatch and myocardial viability is of clinical relevance in guiding revascularization following acute infarction, in clarifying the functional significance of a perfusion defect, and in guiding treatment selection for patients with angina in the absence of obstructive coronary artery disease.

HP MRI can also help monitor the flux from [1-¹³C]pyruvate to bicarbonate through the mitochondrial enzyme PDH as a way to examine the relative contribution of glucose and fatty acid oxidation (substrate selection) to energy production in the heart (55–60). In diabetes, the hyperthyroid heart, and dilated cardiomyopathy, flux through PDH in the heart is reduced as measured by HP [1-¹³C]pyruvate MRI (56–58). In contrast, in hypertrophic cardiomyopathy from hypertension, flux through PDH is increased, indicating a preference for glucose metabolism rather than fatty acid oxidation (60). Other studies have also utilized HP [2-¹³C]pyruvate MRI to interrogate TCA cycle metabolism during reperfusion and cardiac remodeling following myocardial infarction (61,62). These data show the potential of metabolic profiling with HP ¹³C MRI as a noninvasive tool to improve understanding of the mechanism of various heart diseases, provide more specific diagnoses, and provide therapeutic guidance.

Liver Disease

A growing body of literature has indicated a strong potential for HP ¹³C MRI in the diagnosis and monitoring of liver injury (63–69). For example, increased [1-¹³C]pyruvate–to-alanine-and-lactate conversion was observed in a model of chemically induced inflammatory liver injury (65), and decreased [1-¹³C]DHA–to–vitamin C conversion was noted in a model of diet-induced steatohepatitis (18). These results suggest that such an imaging approach may address a currently unmet clinical need for noninvasive diagnosis and treatment monitoring of nonalcoholic steatohepatitis (NASH), a disease of growing concern in the United States and worldwide. HP ¹³C dihydroxyacetone, a gluconeogenesis precursor, has been used to probe hepatic energy metabolism (68,69), the alteration of which is implicated in both NASH and cirrhosis. To increase the specificity of detected ¹³C metabolism, a clinically used liver-targeted contrast agent (ie, gadoxetate) can be administered prior to ¹³C probe injection to selectively suppress the ¹³C signal from normal hepatocytes (70), as compared with other cell types such as inflammatory cells.

Kidney Disease

In the kidneys, HP ¹³C MRI has been used to investigate metabolic changes related to hypoxia and oxidative stress (17,71–74), two key factors implicated in progressive kidney injury. Increased renal [1-¹³C]pyruvate–to-lactate conversion was observed in models of diabetes and has been hypothesized to be related to intrarenal pseudohypoxia from hyperglycemia (71,72). In models of acute kidney injury (ischemia-reperfusion injury), decreased [1-¹³C]pyruvate–to-bicarbonate conversion has been noted, corresponding to decreased PDH activity and mitochondrial energy production (73,74). The [1-¹³C]pyruvate–to-lactate conversion in the injured kidney changes dynamically during the evolution of acute kidney injury (73,74) and may potentially yield information on renal tubular injury during the progression from acute kidney injury to chronic kidney disease. Noninvasive predictors of such progressive kidney injuries are urgently needed given the increasing incidence of chronic kidney disease. The redox probe HP [1-¹³C]DHA has been shown to sensitively monitor the level of glutathione, a major antioxidant, during progression of diabetes and following treatment targeted to oxidative stress (17). Additionally, the corticomedullary gradient of urea, which is a key osmolyte in the kidney’s ability to concentrate urine, can be monitored by using HP ¹³C urea (75,76), suggesting the potential of this probe to monitor kidney function. While concerns about toxicity have limited the use of gadolinium-based contrast agents in patients with renal insufficiency, HP MRI with endogenous probes such as pyruvate and urea is particularly promising for the noninvasive diagnosis and monitoring of kidney disease.

Inflammation

Chronic inflammation is increasingly recognized as a common factor in many diseases and as a target for therapy. Inflammatory tissues have Warburg-like metabolism and hence can be interrogated by using HP pyruvate MRI. In addition to liver inflammation (as discussed above), increased HP [1-¹³C]pyruvate–to-lactate conversion has been shown to reflect inflammation in models of radiation-induced lung injury (77), neuroinflammation (78), and inflammatory arthritis (79). Response to anti-inflammatory treatment can similarly be monitored with HP [1-¹³C]pyruvate MRI (80). Additionally, recent work has explored new HP probes, such as HP [6-¹³C]arginine (81), that specifically target inflammatory cell metabolism. This is of particular interest in cancer treatment monitoring, as HP [1-¹³C]pyruvate cannot help differentiate lactate production from cancer cells from that associated with inflammatory cells.

Technical Advances Enabling Clinical Translation of HP ¹³C MRI

The true potential of HP ¹³C MRI lies in its rapid progress in clinical translation for patient care. Several technical advances have been made in recent years to achieve this goal. Figure 4 shows the requisite components of human HP ¹³C studies. These components are explained further below.

Figure 4:

Schematic shows the required components for clinical hyperpolarized carbon 13 (¹³C) MRI studies. The pharmacy kits used for preparing sterile ¹³C probes, the clinical polarizers with built-in quality-control units, and specialized MRI detector hardware are now commercially available. Many of the fast ¹³C pulse sequences and postprocessing tools are available as open source. In the case of [1-¹³C]pyruvate, the Investigational New Drug resource for its use is available from the National Cancer Institute to assist sites in obtaining U.S. Food and Drug Administration (FDA) regulatory approval for clinical HP MRI studies. k_PL = apparent rate constant for pyruvate-to-lactate conversion.

Clinical Polarizer

Clinical polarizers are now commercially available (SPINlab; GE Research Circle Technology, Waukesha, Wis) for patient studies (82), with 23 sited around the world. These clinical polarizers allow up to four samples to be polarized at the same time, permitting the injection of multiple probes during a single examination. The preparation of the probes uses a “pharmacy kit,” which is filled either in a licensed pharmacy in a sterile compounding environment or by using Good Manufacturing Practice with terminal sterilization following the polarization. The clinical polarizer contains a quality-control module that measures pH, pyruvate and residual radical concentration, polarization, and sample temperature prior to injection to ensure the probe is safe for clinical use. Current clinical polarizers can achieve up to 50% polarization of [1-¹³C]pyruvate, more than a twofold increase compared with the initial prototype polarizer used for the phase I human study.

Fast Imaging Techniques

The rapid metabolism and relatively short T1 relaxation times of most biologically relevant HP ¹³C probes have required the development of novel multichannel coil array systems, as well as specialized rapid pulse sequences, and many of these are now commercially available. Multichannel ¹³C radiofrequency coil arrays allow increased sensitivity and the use of rapid imaging techniques including parallel imaging. For abdominal studies, a commercial 16-channel torso array is available. For other applications, a variety of both custom and commercial ¹³C MRI coils have been developed with both single-channel receive and multichannel arrays of up to 32 channels for brain tumor studies.

In terms of pulse sequences, two general approaches can be used: a spectroscopic technique and an imaging-based technique. Rapid spectroscopic techniques, such as echo-planar spectroscopic imaging (EPSI), generate a ¹³C spectrum for every voxel that is imaged (83). These sequences are useful when the exact metabolites are not known, when there are numerous metabolites, or when inhomogeneity in the magnetic field causes the spectral frequencies to be shifted across the sample. In an alternative image-based approach, each metabolite is excited in turn and then imaged by using a conventional imaging sequence such as echo-planar imaging (84), spiral MRI (85), or balanced steady-state free precession (86). The advantage of this approach is that it is faster than EPSI and directly generates images on the MRI unit, which facilitates clinical interpretation. The disadvantages of the imaging-based approaches are that they require the metabolite frequencies to be known beforehand and they are sensitive to nonuniform magnetic fields. To improve the efficient use of HP signal as well as the accuracy and robustness of HP MRI, bolus tracking and real-time B₁ calibration methods have also been implemented recently (87).

Because many metabolic processes are rapid, fast dynamic HP ¹³C imaging of the substrates and their metabolites is advantageous (88). Dynamic ¹³C acquisition enables obtaining quantitative metabolic parameters such as the apparent rate constant for pyruvate-to-lactate conversion (k_PL), which is less dependent on the precise acquisition and contrast bolus timing. Because the HP ¹³C signal is nonrenewable, specialized fast imaging strategies are used to conserve magnetization and to allow dynamic data acquisition at multiple time points. For example, compressed sensing reconstruction can be used to minimize the amount of data required. Additionally, special radiofrequency pulses can be used to apply different flip angles to the HP ¹³C substrates and their metabolites to maximize the signal from the metabolites while preserving the magnetization of the substrates (89).

The MRI unit field strength affects the T1 relaxation time of HP probes. For example, HP ¹³C pyruvate has slightly shorter T1 at 3.0 T than at 1.5 T. However, the chemical shift separation between pyruvate and its metabolites in hertz at 3.0 T is twice that at 1.5 T; this enables better metabolic quantification. Therefore, current clinical studies of HP ¹³C pyruvate MRI are commonly performed with 3.0-T MRI units.

Data Analysis

Because HP ¹³C MRI signal changes rapidly over the course of a study, new approaches for data analysis and quantification are required. One common approach is to express the total signal of downstream metabolites (ie, lactate, alanine) as a fraction of the signal from the injected probe (ie, pyruvate). Alternatively, pharmacokinetic models have been proposed to quantify the apparent rate constants for substrate conversions (2,90–92). For example, k_PL can be calculated in patients (92). These approaches need to be further tested and refined in clinical studies to establish the most accurate and reproducible quantitative analysis methods for HP MRI.

Clinical HP Investigations

The two probes [1-¹³C]pyruvate and [2-¹³C]pyruvate are currently the probes that have received regulatory approval in the United States (U.S. Food and Drug Administration Investigational New Drug) for clinical investigation. Around the world, 10 sites have performed HP pyruvate MRI in more than 200 human subjects, with several other sites planning clinical studies in the near future. Table E1 (online) lists the up-to-date clinical trials that have registered with ClinicalTrials.Gov. These trials are predominantly oncology focused but also include investigations in heart and liver disease and traumatic brain injury, suggesting the potential of this technology for metabolic imaging in a range of human diseases.

Prostate Cancer

The first human study of HP [1-¹³C]pyruvate MRI was performed in study subjects with localized prostate cancer at the University of California San Francisco and established the safety and feasibility of this new metabolic imaging technique (2). In that study, a dose of up to 0.43 mL [1-¹³C]pyruvate per kilogram of body weight was administered. Despite being a supra-physiologic level of pyruvate, this dose is rapidly metabolized, and no significant adverse events have been reported to date in clinical studies that used the same dose. In the initial human study, elevated [1-¹³C]pyruvate–to-lactate conversion was demonstrated in the tumor compared with adjacent normal prostate tissue. In one study subject, elevated lactate signal was seen in a focus of tumor (later confirmed at biopsy) that was not visible on either the T2-weighted or the diffusion-weighted images. These results highlight the potential of this technology to improve tumor detection and characterization. The first results demonstrating metabolic response to androgen deprivation therapy by using HP [1-¹³C]pyruvate MRI have also been described (93) (Fig 5). After 6 weeks of treatment, there was complete abrogation of the HP ¹³C lactate signal while there was no change on T2-weighted images and only a modest change on apparent diffusion coefficient maps, illustrating the potential of HP lactate as a biomarker of treatment response. Preliminary data have also demonstrated that HP [1-¹³C]pyruvate MRI provides reproducible measurements of metabolic changes in the prostates of patients who underwent repeated injections of [1-¹³C]pyruvate (94). Several other clinical investigations in prostate cancer are currently ongoing, including studies to predict tumor grade in patients before prostatectomy and to assess early treatment response in men with metastatic prostate cancer.

Figure 5:

Representative axial T2-weighted MR image, water apparent diffusion coefficient (ADC) image, T2-weighted MR image with overlaid apparent rate constant for pyruvate (Pyr)-to-lactate (Lac) conversion (k_PL), and corresponding hyperpolarized (HP) carbon 13 (13C) spectral array in a 52-year-old patient with extensive high-grade (Gleason 4 + 5) prostate cancer, A, before therapy and, B, 6 weeks after initiation of androgen ablation and chemotherapy. Before treatment, the region of prostate cancer can be seen (arrows) as low signal on the T2-weighted and ADC images, with high HP lactate seen in the spectral array (vertical axis = arbitrary signal intensity units; horizontal axis = frequency in parts per million) and high k_PL. At 6 weeks after initiation of androgen deprivation therapy, there was near complete abrogation of elevated HP lactate peaks on the spectral array and associated marked reduction in tumor k_PL (maximum k_PL, 0.025 sec²¹ at baseline and 0.007 sec²¹ at follow-up). Notably, there was negligible change in the size of tumor on T2-weighted MR images and only a modest change on ADC images, supporting the utility of HP [1-¹³C]pyruvate MRI in detecting early metabolic responses prior to changes on conventional images. Concordant with these findings, the patient subsequently achieved a marked clinical response, with an undetectable serum prostate-specific antigen level 6 months after treatment initiation. (Reprinted, with permission, from reference 91.)

Brain Tumors

In an initial study of eight subjects with a diagnosis of primary brain tumors, HP [1-¹³C]pyruvate was rapidly transported across the blood-brain barrier and converted to the metabolites lactate and bicarbonate (95) (Fig 6). The non-tumor brain showed high bicarbonate signal that was not detected in the tumors, suggesting that this technique may be valuable for probing mitochondrial oxidative metabolism and its alteration in brain tumors. Another recent study (96) similarly confirmed the feasibility of acquiring high-quality metabolite maps in patients with primary or metastatic brain tumors. These initial clinical studies demonstrated successful technical implementation of HP [1-¹³C]pyruvate MRI for brain metabolism evaluation. While additional technical optimization is required, the data support further investigation of the clinical utility of this technology in patients with brain tumors, as well as in patients with other neurologic diseases.

Figure 6:

Hyperpolarized (HP) [1-¹³C]pyruvate MR images in two patients with treated glioblastoma multiforme. Patient A (top row) had progressing tumor at the time of the HP study, while patient B (bottom row) had stable tumor at the time of the HP study. HP [1-¹³C]pyruvate was rapidly transported across the blood-brain barrier and converted to the metabolites lactate and bicarbonate. There was substantial pyruvate-to-lactate and pyruvate-to-bicarbonate conversion in the normal brain. The tumor (arrows) in patient A, who had progressive tumor, showed moderate pyruvate-to-lactate conversion when compared with normal brain. The tumor (arrow) in patient B, who had clinically stable tumor, showed no substantial pyruvate-to-lactate conversion. Both tumors showed a lack of pyruvate-to-bicarbonate conversion, suggesting reduced mitochondrial oxidative metabolism. Color bars = metabolite ratios. The differences in the scales of the color bars are due to the single-band constant flip angle excitation scheme used for patient A and the multiband variable flip angle excitation scheme used for patient B. These initial findings support the use of HP [1-¹³C]pyruvate MRI for investigating metabolic reprogramming in brain tumors. FLAIR = fluid-attenuated inversion recovery, Gad = gadolinium. (Reprinted from reference 93.)

Cardiac Metabolism

The first images of HP ¹³C MRI in the human heart were reported in a pilot study of healthy volunteers (97). The metabolism of [1-¹³C]pyruvate and the generation of bicarbonate signal in the left ventricular myocardium were visualized with high signal-to-noise ratio. More recently, the same investigators also demonstrated the feasibility of using HP [1-¹³C]pyruvate to detect metabolic alterations in study subjects with hypertrophic cardiomyopathy (98). Preliminary data showed significantly elevated bicarbonate signal near the cardiac apex, corresponding to the known location of disease. There was also a different spatial distribution of the bicarbonate signal in cardiomyopathy compared with the normal heart. These initial data suggest the exciting possibilities of imaging altered cardiac energetics in patients with heart disease and potentially improving diagnosis and treatment monitoring in these patients.

Other Emerging Applications

In addition to the prostate, brain, and cardiac applications described above, preliminary human results have been obtained in the liver (Fig 7), kidney, pancreas, breast, and bone. As has already been suggested in initial patient data (93), the ability to rapidly monitor response to treatment is likely one of the most promising applications of HP ¹³C MRI. Particularly in the context of cancer, early detection of nonresponse is essential for minimizing toxicities and redirecting to more effective therapies. Other potential oncologic applications include risk stratification of tumors after the initial diagnosis and determining the most metabolically active region of tumors to target tissue sampling. Furthermore, metabolic changes occurring with diabetes, kidney, and liver disease are also promising areas for exploration in humans.

Figure 7:

Hyperpolarized (HP) [1-¹³C]pyruvate MR images in a patient with breast cancer metastatic to the liver. The study was performed by using a 16-channel abdomen carbon 13 (¹³C) coil array permitting coverage of the whole upper abdomen and a dynamic ¹³C echo-planar spectroscopic imaging technique at a 2-second time resolution. T2-weighted single-shot fast spin-echo (SSFSE) image (left) shows multiple liver metastases. The k_PL (the apparent rate constant for pyruvate-to-lactate conversion in sec²¹) map overlaid on the T2-weighted SSFSE image (right) shows liver metastases (arrows) with elevated pyruvate-to-lactate conversion, consistent with metabolically active tumors.

Comparison with PET

Although FDG PET and HP pyruvate MRI are both sensitive to the metabolic changes that occur in tumors, there are important distinctions that make the two methods complementary. As already discussed, HP MRI with [1-¹³C]pyruvate provides information at a key branch point of metabolic pathways involved in downstream glucose metabolism—information that is not accessible to FDG PET. For example, prostate cancer often has similar FDG avidity to the background prostate (99). In contrast, a grade-dependent increase in HP pyruvate-to-lactate conversion has been observed in a TRAMP model, as well in patient-derived prostate tissues (23,100), making HP pyruvate MRI a promising tool for assessing tumor aggressiveness. Additionally, FDG PET is sensitive to both glucose uptake and the ability of the cell to trap FDG within cells. For example, some hepatocellular carcinomas have low expression of the glucose transporter GLUT1 but high expression of glucose 6 phosphatase, which phosphorylates FDG and traps it intracellularly, thereby altering the FDG signal (101). Furthermore, because all PET agents produce photons with the same energy, the administered PET probes cannot be distinguished from their products, and it is difficult to image several different PET probes simultaneously. In contrast, because HP ¹³C probes and their metabolites have different resonance frequencies, they can be imaged simultaneously to provide information on multiple metabolic processes (13). Finally, because PET and HP ¹³C MRI depict complementary aspects of tumor metabolism, it is attractive to explore the combined performance of both modalities. In that context, the advent of the integrated PET/MRI units facilitate such multiparametric examinations.

Challenges and Future Directions

For HP ¹³C MRI to be adopted as a clinical molecular imaging tool to advance patient care, further efforts are needed in several areas. Importantly, these efforts will require a close collaboration between basic scientists and clinicians, as well as a strong partnership between academia, industry, and funding agencies.

Commercially available dual-tuned (¹H and ¹³C) multichannel coils that allow high signal-to-noise ratio imaging for all body areas are essential. Continuing work to improve pulse sequences that permit the imaging of different organs and with different HP ¹³C probes is necessary to increase clinical applicability. Further refinements are also needed to efficiently and reliably achieve higher and faster polarizations and to simplify methods of sterile HP probe delivery. To facilitate clinical adoption, automated procedures for HP probe delivery, imaging acquisition, and data analysis are also critical.

As indicated in a recent whitepaper (102), Food and Drug Administration approval and clinical adoption of HP ¹³C MRI will require establishing its clinical utility in subsequent larger phase II and III trials. Critical to this is the demonstration of reproducibility, which will necessitate the establishment of uniform standards for probe production, image acquisition protocol, and data analysis that enable multicenter trials. Early multicenter phase II trials should validate the findings from single-center trials, and indications supported by small multicenter trials can then be carried into larger phase II and phase III studies. These larger trials should not only evaluate safety and efficacy end points but also collect information on clinical impact and cost effectiveness, which are key data for coverage decisions by the Centers for Medicare and Medicaid Services and other payers. In the case of [1-¹³C]pyruvate, to assist sites planning clinical HP studies, the Investigational New Drug (IND) for the use of [1-¹³C]pyruvate was transferred to the National Cancer Institute, and the IND resource is available at https://imaging.cancer.gov/programs_resources/cancer-tracer-synthesis-resources/hyperpolarized-C13-pyruvate-documentation.htm.

An important consideration in the clinical adoption of HP ¹³C MRI is cost. The cost of a clinical polarizer is similar to that of a PET cyclotron, and the cost of a Good Manufacturing Practice dose of ¹³C pyruvate is similar to or less than that of many emerging PET agents. Given the cost, the usage of HP ¹³C MRI will likely be restricted to large academic medical centers, as in the cases of other high-end imaging technologies. Work is ongoing at multiple institutions to further improve the robustness, reliability, and efficiency of HP ¹³C MRI (in particular with ¹³C pyruvate) as a 5-minute add-on to a standard-of-care multiparametric MRI examination. This technology has the potential to provide cost-effective molecular imaging examinations to improve diagnosis and treatment monitoring for various diseases where current imaging methods show limitations.

Conclusions

HP ¹³C MRI is an emerging molecular imaging technique that is actively undergoing clinical translation at multiple institutions. Preclinical results suggest applications in oncology, diabetes, and heart disease, as well as metabolic disease of the liver and kidney. The tools required for human studies with HP ¹³C MRI are now commercially available, with clinical trials underway. Further expansion into the clinic will require continued improvement in imaging unit hardware and pulse sequences, formulation of new probes, quantification and standardization of results, and multicenter clinical trials.