MALDI MSI Separation of Same Donor's Fingermarks Based on Time of Deposition-A Proof-of-Concept Study.

Despite the advent of DNA profiling, fingerprints still play an important role in suspect identification. However, if single crime scene marks may be challenging to identify, overlapping fingermarks, understandably, pose an even greater challenge. In the last decade, mass spectrometry-imaging methods have provided a possible solution to the separation of fingermarks from two or more donors, based on the differential chemical composition. However, there are no studies attempting to separate overlapping marks from the same donor. This is important in relation to fingermark deposition at different times, which could be critical, for example, to ascertain legitimate access to the scene. In the work presented here, we investigate whether Matrix-Assisted Laser Desorption Ionisation Mass Spectrometry Imaging can separate the same donor's fingermarks deposited at different times based on intra-donor fingermark composition variability. Additionally, the hypothesis that the different times of deposition could be also determined was investigated in the view of linking the suspect at the scene at different times; the dating window of MALDI MSI within the selected molecular range was explored. Results show that it is possible to separate overlapping fingermarks from the same donor in most cases, even from natural marks. Fresh marks (0 days) could be separated from those of fourteen days of age, though the latter could not be distinguished from the set aged for seven days. Due to the use of only one donor, these are to be considered preliminary data, though findings are interesting enough to warrant further investigation of the capabilities and limitations of this approach using a larger cohort of donors.

N-(4-[18F]fluorobenzyl)cholylglycine, a potential tracer for positron emission tomography of enterohepatic circulation and drug-induced inhibition of ileal bile acid transport. A proof-of-concept PET/CT study in pigs.

INTRODUCTION: Enterohepatic circulation (EHC) of conjugated bile acids is an important physiological process crucial for bile acids to function as detergents and signal carriers. Perturbation of the EHC by disease or drugs may lead to serious and life-threatening liver and gastrointestinal disorders. In this proof-of-concept study in pigs, we investigate the potential of N-(4-[18F]fluorobenzyl)cholylglycine ([18F]FBCGly) as tracer for quantitative positron emission tomography (PET) of the EHC of conjugated bile acids. METHODS: The biodistribution of [18F]FBCGly was investigated by PET/CT in domestic pigs following intravenous and intraileal administration of the tracer. Hepatic kinetics were estimated from PET and blood data using a 2-tissue compartmental model and dual-input of [18F]FBCGly. The ileal uptake of [18F]FBCGly was investigated with co-injection of nifedipine and endogenous cholyltaurine. Dosimetry was estimated from the PET data using the Olinda 2.0 software. Blood, bile and urine samples were analyzed for possible fluorine-18 labelled metabolites of [18F]FBCGly. RESULTS: [18F]FBCGly was rapidly taken up by the liver and excreted into bile, and underwent EHC without being metabolized. Both nifedipine and endogenous cholyltaurine inhibited the ileal uptake of [18F]FBCGly. The flow-dependent hepatic uptake clearance was estimated to median 1.2 mL blood/min/mL liver tissue. The mean residence time of [18F]FBCGly in hepatocytes was 4.0 ± 1.1 min. Critical organs for [18F]FBCGly were the gallbladder wall (0.94 mGy/MBq) and the small intestine (0.50 mGy/MBq). The effective dose for [18F]FBCGly was 36 µSv/MBq. CONCLUSION: We have shown that [18F]FBCGly undergoes EHC in pigs without being metabolized and that its ileal uptake is inhibited by nifedipine and endogenous bile acids. Combined with our previous findings in rats, we believe that [18F]FBCGly has potential as PET tracer for assessment of EHC of conjugated bile acids under physiological conditions as well as conditions with perturbed hepatic and ileal bile acid transport.

A Retrospective Metabolomics Analysis of Gamma-Hydroxybutyrate in Humans: New Potential Markers and Changes in Metabolism Related to GHB Consumption.

GHB is an endogenous short-chain organic acid presumably also widely applied as a rape and knock out drug in cases of drug-facilitated crimes or sexual assaults (DFSA). Due to the endogenous nature of GHB and its fast metabolism in vivo, the detection window of exogenous GHB is however narrow, making it challenging to prove use of GHB in DFSA cases. Alternative markers of GHB intake have recently appeared though none has hitherto been validated for forensic use. UHPLC-HRMS based screening of blood samples for drugs of abuse is routinely performed in several forensic laboratories which leaves an enormous amount of unexploited data. Recently we devised a novel metabolomics approach to use archived data from such routine screenings for elucidating both direct metabolites from exogenous compounds, but potentially also regulation of endogenous metabolism and metabolites. In this paper we used UHPLC-HRMS data acquired over a 6-year period from whole blood analysis of 51 drivers driving under the influence of GHB as well as a matched control group. The data were analyzed using a metabolomics approach applying a range of advanced analytical methods such as OPLS-DA, LASSO, random forest, and Pearson correlation to examine the data in depth and demonstrate the feasibility and potential power of the approach. This was done by initially detecting a range of potential biomarkers of GHB consumption, some that previously have been found in controlled GHB studies, as well as several new potential markers not hitherto known. Furthermore, we investigate the impact of GHB intake on human metabolism. In aggregate, we demonstrate the feasibility to extract meaningful information from archived data here exemplified using GHB cases. Hereby we hope to pave the way for more general use of the principle to elucidate human metabolites of e.g. new legal or illegal drugs as well as for applications in more global and large scale metabolomics studies in the future.

The pathophysiology of Wilson's disease visualized: A human 64 Cu PET study.

BACKGROUND AND AIMS: Wilson's disease (WD) is a genetic disease with systemic accumulation of copper that leads to symptoms from the liver and brain. However, the underlying defects in copper transport kinetics are only partly understood. We sought to quantify hepatic copper turnover in patients with WD compared with heterozygote and control subjects using PET with copper-64 (64 Cu) as a tracer. Furthermore, we assessed the diagnostic potential of the method. APPROACH AND RESULTS: Nine patients with WD, 5 healthy heterozygote subjects, and 8 healthy controls were injected with an i.v. bolus of 64 Cu followed by a 90-min dynamic PET scan of the liver and static whole-body PET/CT scans after 1.5, 6, and 20 h. Blood 64 Cu concentrations were measured in parallel. Hepatic copper retention and redistribution were evaluated by standardized uptake values (SUVs). At 90 min, hepatic SUVs were similar in the three groups. In contrast, at 20 h postinjection, the SUV in WD patients (mean ± SEM, 31 ± 4) was higher than in heterozygotes (24 ± 3) and controls (21 ± 4; p < 0.001). An SUV-ratio of hepatic 64 Cu concentration at 20 and 1.5 h completely discriminated between WD patients and control groups (p < 0.0001; ANOVA). By Patlak analysis of the initial 90 min of the PET scan, the steady-state hepatic clearance of 64 Cu was estimated to be slightly lower in patients with WD than in controls (p = 0.04). CONCLUSIONS: 64 Cu PET imaging enables visualization and quantification of the hepatic copper retention characteristic for WD patients. This method represents a valuable tool for future studies of WD pathophysiology, and may assist the development of therapies, and accurate diagnosis.

Obeticholic acid improves hepatic bile acid excretion in patients with primary biliary cholangitis.

BACKGROUND & AIMS: Obeticholic acid (OCA) is an agonist of the nuclear bile acid receptor farnesoid X receptor, which regulates hepatic bile acid metabolism. We tested whether OCA treatment would influence hepatic transport of conjugated bile acids in patients with primary biliary cholangitis (PBC) who responded inadequately to treatment with ursodeoxycholic acid (UDCA). METHODS: Eight UDCA-treated patients with PBC with alkaline phosphatase ≥1.5 times the upper limit of normal range participated in a double-blind, placebo-controlled study. While continuing on UDCA, the patients were randomised to two 3-month crossover treatment periods with placebo and OCA, in random order, separated by a 1-month washout period without study treatment. After each of the two treatment periods, we determined rate constants for transport of conjugated bile acids between blood, hepatocytes, biliary canaliculi, and bile ducts by positron emission tomography of the liver using the conjugated bile acid tracer [N-methyl-11C]cholylsarcosine (11C-CSar). The hepatic blood perfusion was measured using infusion of indocyanine green and Fick's principle. RESULTS: Compared with placebo, OCA increased hepatic blood perfusion by a median of 11% (p = 0.045), the unidirectional uptake clearance of 11C-CSar from blood into hepatocytes by a median of 11% (p = 0.01), and the rate constant for secretion of 11C-CSar from hepatocytes into biliary canaliculi by a median of 73% (p = 0.03). This resulted in an OCA-induced decrease in the hepatocyte residence time of 11C-CSar by a median of 30% (p = 0.01), from group median 11 min to 8 min. CONCLUSIONS: This study of UDCA-treated patients with PBC showed that, compared with placebo, OCA increased the hepatic transport of the conjugated bile acid tracer 11C-CSar, and thus endogenous conjugated bile acids, from hepatocytes into biliary canaliculi. As a result, OCA reduced the time hepatocytes are exposed to potentially cytotoxic bile acids. LAY SUMMARY: Primary biliary cholangitis is a chronic liver disease in which the small bile ducts are progressively destroyed. We tested whether the treatment with obeticholic acid (OCA) would improve liver excretion of bile acids compared with placebo in 8 patients with primary biliary cholangitis. A special scanning technique (PET scan) showed that OCA increased the transport of bile acids from blood to bile. OCA thereby reduced the time that potentially toxic bile acids reside in the liver by approximately one-third.

Intravenous and oral copper kinetics, biodistribution and dosimetry in healthy humans studied by [64Cu]copper PET/CT.

PURPOSE: Copper is essential for enzymatic processes throughout the body. [64Cu]copper (64Cu) positron emission tomography (PET) has been investigated as a diagnostic tool for certain malignancies, but has not yet been used to study copper homeostasis in humans. In this study, we determined the hepatic removal kinetics, biodistribution and radiation dosimetry of 64Cu in healthy humans by both intravenous and oral administration. METHODS: Six healthy participants underwent PET/CT studies with intravenous or oral administration of 64Cu. A 90 min dynamic PET/CT scan of the liver was followed by three whole-body PET/CT scans at 1.5, 6, and 20 h after tracer administration. PET data were used for estimation of hepatic kinetics, biodistribution, effective doses, and absorbed doses for critical organs. RESULTS: After intravenous administration, 64Cu uptake was highest in the liver, intestinal walls and pancreas; the gender-averaged effective dose was 62 ± 5 µSv/MBq (mean ± SD). After oral administration, 64Cu was almost exclusively taken up by the liver while leaving a significant amount of radiotracer in the gastrointestinal lumen, resulting in an effective dose of 113 ± 1 µSv/MBq. Excretion of 64Cu in urine and faeces after intravenous administration was negligible. Hepatic removal kinetics showed that the clearance of 64Cu from blood was 0.10 ± 0.02 mL blood/min/mL liver tissue, and the rate constant for excretion into bile or blood was 0.003 ± 0.002 min- 1. CONCLUSION: 64Cu biodistribution and radiation dosimetry are influenced by the manner of tracer administration with high uptake by the liver, intestinal walls, and pancreas after intravenous administration, while after oral administration, 64Cu is rapidly absorbed from the gastrointestinal tract and deposited primarily in the liver. Administration of 50 MBq 64Cu yielded images of high quality for both administration forms with radiation doses of approximately 3.1 and 5.7 mSv, respectively, allowing for sequential studies in humans. TRIAL REGISTRATION NUMBER: EudraCT no. 2016-001975-59. Registration date: 19/09/2016.

The role of dopaminergic and serotonergic transmission in the processing of primary and monetary reward.

Natural rewards such as erotic stimuli activate common neural pathways with monetary rewards. In human studies, the manipulation of dopamine and serotonin play an important role in the processing of monetary rewards with less understood on its role on erotic stimuli. In this study, we investigate the neuromodulatory effects of dopaminergic and serotonergic transmission in the processing of erotic versus monetary visual stimuli. We scanned one hundred and two (N = 102) healthy volunteers using functional magnetic resonance imaging while performing a modified version of the well-validated monetary incentive delay task consisting of erotic, monetary and neutral visual stimuli. We show a role for enhanced central dopamine and lowered central serotonin levels in increasing activity in the right caudate and left anterior insula during anticipation of erotic relative to monetary rewards in healthy controls. We further show differential activation in the anticipation of natural versus monetary rewards with the former associated with ventromesial and dorsomesial activity and the latter with dorsal cingulate, striatal and anterior insular activity. These findings are consistent with preclinical and clinical findings of a role for dopaminergic and serotonergic mechanisms in the processing of natural rewards. Our study provides further insights into the neural substrates underlying reward processing for natural primary erotic rewards and yields importance for the neurochemical systems of addictive disorders including gambling disorder.

Human biodistribution, dosimetry, radiosynthesis and quality control of the bile acid PET tracer [N-methyl-11C]cholylsarcosine.

INTRODUCTION: [N-methyl-11C]cholylsarcosine ([11C]CSar) is a tracer for imaging and quantitative assessment of intrahepatic cholestatic liver diseases and drug-induced cholestasis by positron emission tomography (PET). The purpose of this study is to determine whole-body biodistribution and dosimetry of [11C]CSar in healthy humans. The results are compared with findings in a patient with primary sclerosing cholangitis (PSC) and a patient with primary biliary cholangitis (PBC) as well as with preclinical findings in pigs. Radiosynthesis and quality control for preparation of [11C]CSar for clinical use are also presented. METHODS: Radiosynthesis and quality control of [11C]CSar were set up in compliance with Danish/European regulations. Both healthy participants (3 females, 3 males) and patients underwent whole-body PET/CT to determine the biodistribution of [11C]CSar. The two patients were under treatment with ursodeoxycholic acid at the time of the study. Dosimetry was estimated from the PET data using the Olinda 2.0 software. RESULTS: The radiosynthesis provided [11C]CSar in a solution ready for injection. The biodistribution studies revealed that gallbladder wall, small intestine, and liver were critical organs in both healthy participants and patients with the gallbladder wall receiving the highest dose (up to 0.5 mGy/MBq). The gender-averaged (±SD) effective dose for the healthy participants was 6.2 ±â€¯1.4 µSv/MBq. The effective dose for the PSC and the PBC patient was 5.2 and 7.0 µSv/MBq, respectively. CONCLUSION: A radiosynthesis for preparation of [11C]CSar for clinical use was developed and approved by the Danish Medicines Agency. The most critical organ was the gallbladder wall although the amount of [11C]CSar in the gallbladder was found to vary significantly between individuals. The estimated effective dose for humans was comparable to that estimated in anesthetized pigs although the absorbed dose estimates to some organs, such as the stomach, was different. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: [11C]CSar PET/CT enables detailed quantitative assessment of patients with cholestatic liver disease by tracing the separate hepatobiliary transport steps of endogenous bile acids. The present work offers a radiosynthetic method and dosimetry data suitable for clinical implementation of [11C]CSar.

N-(4-[18F]fluorobenzyl)cholylglycine, a novel tracer for PET of enterohepatic circulation of bile acids: Radiosynthesis and proof-of-concept studies in rats.

INTRODUCTION: Enterohepatic circulation (EHC) of conjugated bile acids is an important physiological process crucial for regulation of intracellular concentrations of bile acids and their function as detergents and signal carriers. Only few bile acid-derived imaging agents have been synthesized and hitherto none have been evaluated for studies of EHC. We hypothesized that N-(4-[18F]fluorobenzyl)cholylglycine ([18F]FBCGly), a novel fluorine-18 labeled derivative of endogenous cholylglycine, would be a suitable tracer for PET of the EHC of conjugated bile acids, and we report here a radiosynthesis of [18F]FBCGly and a proof-of-concept study by PET/MR in rats. METHODS: A radiosynthesis of [18F]FBCGly was developed based on reductive alkylation of glycine with 4-[18F]fluorobenzaldehyde followed by coupling to cholic acid. [18F]FBCGly was investigated in vivo by dynamic PET/MR in anesthetized rats; untreated or treated with cholyltaurine or rifampicin. Possible in vivo metabolites of [18F]FBCGly were investigated by analysis of blood and bile samples, and the stability of [18F]FBCGly towards enzymatic de-conjugation by Cholylglycine Hydrolase was tested in vitro. RESULTS: [18F]FBCGly was produced with a radiochemical purity of 96% ±â€¯1% and a non-decay corrected radiochemical yield of 1.0% ±â€¯0.3% (mean ±â€¯SD; n = 12). PET/MR studies showed that i.v.-administrated [18F]FBCGly underwent EHC within 40-60 min with a rapid transhepatic transport from blood to bile. In untreated rats, the radioactivity concentration of [18F]FBCGly was approximately 15 times higher in bile than in liver tissue. Cholyltaurine and rifampicin inhibited the biliary secretion of [18F]FBCGly. No fluorine-18 metabolites of [18F]FBCGly were observed. CONCLUSION: We have developed a radiosynthesis of a novel fluorine-18 labeled bile acid derivative, [18F]FBCGly, and shown by PET/MR that [18F]FBCGly undergoes continuous EHC in rats without metabolizing. This novel tracer may prove useful in PET studies on the effect of drugs or diseases on the EHC of conjugated bile acids.

Quantitative PET of liver functions.

Improved understanding of liver physiology and pathophysiology is urgently needed to assist the choice of new and upcoming therapeutic modalities for patients with liver diseases. In this review, we focus on functional PET of the liver: 1) Dynamic PET with 2-deoxy-2-[18F]fluoro-D-galactose (18F-FDGal) provides quantitative images of the hepatic metabolic clearance Kmet (mL blood/min/mL liver tissue) of regional and whole-liver hepatic metabolic function. Standard-uptake-value (SUV) from a static liver 18F-FDGal PET/CT scan can replace Kmet and is currently used clinically. 2) Dynamic liver PET/CT in humans with 11C-palmitate and with the conjugated bile acid tracer [N-methyl-11C]cholylsarcosine (11C-CSar) can distinguish between individual intrahepatic transport steps in hepatic lipid metabolism and in hepatic transport of bile acid from blood to bile, respectively, showing diagnostic potential for individual patients. 3) Standard compartment analysis of dynamic PET data can lead to physiological inconsistencies, such as a unidirectional hepatic clearance of tracer from blood (K1; mL blood/min/mL liver tissue) greater than the hepatic blood perfusion. We developed a new microvascular compartment model with more physiology, by including tracer uptake into the hepatocytes from the blood flowing through the sinusoids, backflux from hepatocytes into the sinusoidal blood, and re-uptake along the sinusoidal path. Dynamic PET data include information on liver physiology which cannot be extracted using a standard compartment model. In conclusion, SUV of non-invasive static PET with 18F-FDGal provides a clinically useful measurement of regional and whole-liver hepatic metabolic function. Secondly, assessment of individual intrahepatic transport steps is a notable feature of dynamic liver PET.

On the Evolution of Bile Salts and the Farnesoid X Receptor in Vertebrates.

In recent decades, our knowledge of bile salts has undergone a vast development, and bile salts are now known not only for their detergent properties that aid in the absorption of dietary lipids but also for their interaction with specific nuclear and membrane receptors. In particular, it has been realized that the response of the farnesoid X receptor (FXR) to bile acids provides a signal bridge between the liver and small intestine, controlling the intracellular levels, biosynthesis, and enterohepatic circulation of bile acids. Therefore, FXR and bile acid signaling has become an attractive target for treatment of, for example, cholestatic liver diseases, diabetes, and colorectal cancer. Previously, interest in the structure and chemistry of bile salts has focused on their cellular toxicity and involvement in digestion. However, insight into the extensive variation in the structure of bile salts in vertebrates and the concurrent evolution of the FXR has become increasingly important as their role as signal molecules has become clearer. In this review, we therefore focus on common structural features of bile salts as well as evolutionary aspects of bile salts and the FXR in vertebrates. Ultimately, a better understanding of the evolution of bile salts and the FXR may expand our knowledge of their function in health and disease, including their function outside the gastrointestinal tract, and aid in the development of new strategies for treatment.

Functional assessment of hepatobiliary secretion by 11C-cholylsarcosine positron emission tomography.

Positron emission tomography (PET) with 11C-cholylsarcosine (11C-CSar), a radiolabelled synthetic N-methylglycine (sarcosine) conjugate of cholic acid, is a novel molecular imaging technique that enables quantitative assessment of the individual transport steps involved in hepatic secretion of conjugated bile acids. Here, we present the method and discuss its potential clinical and scientific applications based on findings in the first human study of healthy subjects and patients with cholestasis. We also present a clinical example of a patient studied during and six months after an episode of drug-induced cholestatic liver injury.

Hepatobiliary transport kinetics of the conjugated bile acid tracer 11C-CSar quantified in healthy humans and patients by positron emission tomography.

BACKGROUND & AIMS: Hepatobiliary secretion of bile acids is an important liver function. Here, we quantified the hepatic transport kinetics of conjugated bile acids using the bile acid tracer [N-methyl-11C]cholylsarcosine (11C-CSar) and positron emission tomography (PET). METHODS: Nine healthy participants and eight patients with varying degrees of cholestasis were examined with 11C-CSar PET and measurement of arterial and hepatic venous blood concentrations of 11C-CSar. RESULTS: Results are presented as median (range). The hepatic intrinsic clearance was 1.50 (1.20-1.76) ml blood/min/ml liver tissue in healthy participants and 0.46 (0.13-0.91) in patients. In healthy participants, the rate constant for secretion of 11C-CSar from hepatocytes to bile was 0.36 (0.30-0.62)min-1, 20 times higher than the rate constant for backflux from hepatocytes to blood (0.02, 0.005-0.07min-1). In the patients, rate constant for transport from hepatocyte to bile was reduced to 0.12 (0.006-0.27)min-1, 2.3times higher than the rate constant for backflux to blood (0.05, 0.04-0.09). The increased backflux did not fully normalize exposure of the hepatocyte to bile acids as mean hepatocyte residence time of 11C-CSar was 2.5 (1.6-3.1)min in healthy participants and 6.4 (3.1-23.7)min in patients. The rate constant for transport of 11C-CSar from intrahepatic to extrahepatic bile was 0.057 (0.023-0.11)min-1 in healthy participants and only slightly reduced in patients 0.039 (0.017-0.066). CONCLUSIONS: This first in vivo quantification of individual steps involved in the hepatobiliary secretion of a conjugated bile acid in humans provided new insight into cholestatic disease. LAY SUMMARY: Positron emission tomography (PET) using the radiolabelled bile acid (11C-CSar) enabled quantification of the individual steps of the hepatic transport of bile acids from blood to bile in man. Cholestasis reduced uptake and secretion and increased backflux to blood. These findings improve our understanding of cholestatic liver diseases and may support therapeutic decisions. CLINICAL TRIAL REGISTRATION NUMBER: The trial is registered at ClinicalTrials.gov (NCT01879735).

Hepatobiliary Secretion Kinetics of Conjugated Bile Acids Measured in Pigs by 11C-Cholylsarcosine PET.

UNLABELLED: The aim of this study was to develop a method for the quantification of hepatobiliary uptake and secretion of conjugated bile acids with PET and the (11)C-labeled conjugated bile acid analog [N-methyl-(11)C]cholylsarcosine ((11)C-CSar). METHODS: Six pigs (13 experiments) underwent dynamic (11)C-CSar PET of the liver with simultaneous measurements of hepatic blood perfusion and (11)C-CSar concentrations in arterial, portal, and hepatic venous blood. In 3 pigs (7 experiments), bile was collected from a catheter in the common hepatic duct. PET data were analyzed with a 2-tissue compartmental model with calculation of rate constants for the transport of (11)C-CSar among blood, hepatocytes, and intra- and extrahepatic bile ducts. PET results were validated against invasive blood and bile measurements. RESULTS: The directly measured rate of secretion of (11)C-CSar into bile was equal to the rate of removal from blood at steady state. Accordingly, hepatocytes did not accumulate bile acids but simply facilitated the transport of bile acids from blood to bile against a measured concentration gradient of 4,000. The rate constant for the secretion of (11)C-CSar from hepatocytes into bile in experiments with a catheter in the common hepatic duct was 25% of that in experiments without a catheter (P < 0.05); we interpreted this result to be mild cholestasis caused by the catheter. The catheter caused an increased backflux of (11)C-CSar from hepatocytes to blood, and hepatic blood flow was 25% higher than in experiments without the catheter. The capacity for the overall transport of (11)C-CSar from blood to bile, as quantified by intrinsic clearance, was significantly lower in experiments with the catheter than in those without the catheter (P < 0.001). PET and blood measurements correlated significantly (P < 0.05). CONCLUSION: The in vivo kinetics of hepatobiliary secretion of conjugated bile acids can now be determined by dynamic (11)C-CSar PET.

Radiosynthesis of N-¹¹C-Methyl-Taurine-Conjugated Bile Acids and Biodistribution Studies in Pigs by PET/CT.

UNLABELLED: During cholestasis, accumulation of conjugated bile acids may occur in the liver and lead to hepatocellular damage. Inspired by our recent development of N-(11)C-methyl-glycocholic acid-that is, (11)C-cholylsarcosine-a tracer for PET of the endogenous glycine conjugate of cholic acid, we report here a radiosynthesis of N-(11)C-methyl-taurine-conjugated bile acids and biodistribution studies in pigs by PET/CT. METHODS: A radiosynthesis of N-(11)C-methyl-taurine-conjugated bile acids was developed and used to prepare N-(11)C-methyl-taurine conjugates derived from cholic, chenodeoxycholic, deoxycholic, ursodeoxycholic, and lithocholic acid. The lipophilicity of these new tracers was determined by reversed-phase thin-layer chromatography. The effect of lipophilicity and structure on the biodistribution was investigated in pigs by PET/CT using the tracers derived from cholic acid (3α-OH, 7α-OH, 12α-OH), ursodeoxycholic acid (3α-OH, 7ß-OH), and lithocholic acid (3α-OH). RESULTS: The radiosyntheses of the N-(11)C-methyl-taurine-conjugated bile acids proceeded with radiochemical yields of 61% (decay-corrected) or greater and radiochemical purities greater than 99%. PET/CT in pigs revealed that the tracers were rapidly taken up by the liver and secreted into bile. There was no detectable radioactivity in urine. Significant reflux of N-(11)C-methyl-taurolithocholic acid into the stomach was observed. CONCLUSION: We have successfully developed a radiosynthesis of N-(11)C-methyl-taurine-conjugated bile acids. These tracers behave in a manner similar to endogenous taurine-conjugated bile acids in vivo and are thus promising for functional PET of patients with cholestatic diseases.

The lumped constant for the galactose analog 2-18F-fluoro-2-deoxy-D-galactose is increased in patients with parenchymal liver disease.

UNLABELLED: The galactose analog 2-(18)F-fluoro-2-deoxy-d-galactose ((18)F-FDGal) is a suitable PET tracer for measuring hepatic galactokinase capacity in vivo, which provides estimates of hepatic metabolic function. As a result of a higher affinity of galactokinase toward galactose, the lumped constant (LC) for (18)F-FDGal was 0.13 in healthy subjects. The aim of the present study was to test the hypothesis of a significantly different LC for (18)F-FDGal in patients with parenchymal liver disease. METHODS: Nine patients with liver cirrhosis were studied in connection with a previous study with determination of hepatic intrinsic clearance of ¹8F-FDGal (V*(max/K*(m)). The present study determined the hepatic removal kinetics of galactose, including hepatic intrinsic clearance of galactose (V(max)/K(m)) from measurements of hepatic blood flow and arterial and liver vein blood galactose concentrations at increasing galactose infusions. LC for ¹8F-FDGal was calculated as (V*(max)/K*(m))/(V(max)/K(m)). On a second day, a dynamic ¹8-FDGal PET study with simultaneous infusion of galactose (mean arterial galactose concentration, 6.1 mmol/L of blood) and blood samples from a radial artery was performed, with determination of hepatic systemic clearance of ¹8F-FDGal (K*(+gal) from linear analysis of data (Gjedde-Patlak method). The maximum hepatic removal rate of galactose was estimated from ¹8F-FDGal PET data (V(max)(PET)) using the estimated LC. RESULTS: The mean hepatic V(max) of galactose was 1.18 mmol/min, the mean K(m) was 0.91 mmol/L of blood and the mean V(max)/K(m) was 1.18 L of blood/min. When compared with values of healthy subjects, K(m) did not differ (P = 0.77), whereas both V(max) and V(max)/K(m) were significantly lower in patients (both P < 0.01). Mean LC for ¹8LF-FDGal was 0.24, which was significantly higher than the mean LC of 0.13 in healthy subjects (P < 0.0001). Mean K*(+gal) determined from the PET study was 0.019 L of blood/min/L of liver tissue, which was not significantly different from that in healthy subjects (P = 0.85). Mean hepatic V(max)(PET) was 0.57 mmol/min/L of liver tissue, which was significantly lower than the value in healthy subjects (1.41 mmol/min/L of liver tissue (P < 0.0001). CONCLUSION: Disease may change the LC for a pet tracer, and this study demonstrated the importance of using the correct LC.

Regional metabolic liver function measured in patients with cirrhosis by 2-[¹8F]fluoro-2-deoxy-D-galactose PET/CT.

BACKGROUND & AIMS: There is a clinical need for methods that can quantify regional hepatic function non-invasively in patients with cirrhosis. Here we validate the use of 2-[(18)F]fluoro-2-deoxy-d-galactose (FDGal) PET/CT for measuring regional metabolic function to this purpose, and apply the method to test the hypothesis of increased intrahepatic metabolic heterogeneity in cirrhosis. METHODS: Nine cirrhotic patients underwent dynamic liver FDGal PET/CT with blood samples from a radial artery and a liver vein. Hepatic blood flow was measured by indocyanine green infusion/Fick's principle. From blood measurements, hepatic systemic clearance (Ksyst, Lblood/min) and hepatic intrinsic clearance (Vmax/Km, Lblood/min) of FDGal were calculated. From PET data, hepatic systemic clearance of FDGal in liver parenchyma (Kmet, mL blood/mL liver tissue/min) was calculated. Intrahepatic metabolic heterogeneity was evaluated in terms of coefficient-of-variation (CoV, %) using parametric images of Kmet. RESULTS: Mean approximation of Ksyst to Vmax/Km was 86% which validates the use of FDGal as PET tracer of hepatic metabolic function. Mean Kmet was 0.157 mL blood/mL liver tissue/min, which was lower than 0.274 mL blood/mL liver tissue/min, previously found in healthy subjects (p<0.001), in accordance with decreased metabolic function in cirrhotic livers. Mean CoV for Kmet in liver tissue was 24.4% in patients and 14.4% in healthy subjects (p<0.0001). The degree of intrahepatic metabolic heterogeneity correlated positively with HVPG (p<0.05). CONCLUSIONS: A 20-min dynamic FDGal PET/CT with arterial sampling provides an accurate measure of regional hepatic metabolic function in patients with cirrhosis. This is likely to have clinical implications for the assessment of patients with liver disease as well as treatment planning and monitoring.

[N-methyl-11C]cholylsarcosine, a novel bile acid tracer for PET/CT of hepatic excretory function: radiosynthesis and proof-of-concept studies in pigs.

UNLABELLED: Excretion of conjugated bile acids into bile is an essential function of the liver, and impairment of canalicular bile acid secretion leads to cholestatic liver injury. However, hepatic excretory function cannot be quantified in vivo because of the lack of suitable methods. Cholylsarcosine is an analog of the endogenous bile acid conjugate cholylglycine and exhibits characteristics in vivo that led us to hypothesize that the (11)C-labeled form, that is, [N-methyl-(11)C]cholylsarcosine ((11)C-cholylsarcosine), would be a suitable PET tracer for quantification of hepatic excretory function. METHODS: A method for radiosynthesis of (11)C-cholylsarcosine was developed involving (11)C-methylation of glycine followed by conjugation with cholic acid. Blood-to-liver uptake and liver-to-bile excretion were investigated in vivo by dynamic (11)C-cholylsarcosine PET/CT of 2 anesthetized pigs. In pig 1, a second dynamic (11)C-cholylsarcosine PET/CT examination was preceded by a high dose of the endogenous bile acid conjugate cholyltaurine to investigate possible inhibition of the transhepatocellular transport of (11)C-cholylsarcosine. In pig 2, a second (11)C-cholylsarcosine administration was given to determine the biodistribution of the tracer by means of 5 successive whole-body PET/CT recordings. Possible formation of (11)C-metabolites was investigated by analysis of blood and bile samples from a third pig. RESULTS: The radiochemical yield was 13% ± 3% (n = 7, decay-corrected) and up to 1.1 GBq of (11)C-cholylsarcosine was produced with a radiochemical purity greater than 99%. PET/CT studies showed rapid blood-to-liver uptake and liver-to-bile excretion of (11)C-cholylsarcosine, with radioactivity concentrations being more than 90 times higher in the bile ducts than in liver tissue. Cholyltaurine inhibited the transhepatocellular transport of (11)C-cholylsarcosine, indicating that the tracer is transported by one or more of the same hepatic transporters as cholyltaurine. (11)C-cholylsarcosine underwent an enterohepatic circulation and reappeared in liver tissue and bile ducts after approximately 70 min. There were no detectable (11)C-metabolites in the plasma or bile samples, indicating that the novel conjugated bile acid (11)C-cholylsarcosine was not metabolized in the liver or in the intestines. The effective absorbed dose of (11)C-cholylsarcosine was 4.4 µSv/MBq. CONCLUSION: We have synthesized a novel conjugated bile acid analog, (11)C-cholylsarcosine, and PET/CT studies on anesthetized pigs showed that the hepatic handling of tracer uptake from blood and excretion into the bile was comparable to that for the endogenous bile acid cholyltaurine. This tracer may be valuable for future studies of normal and pathologic hepatic excretory functions in humans.