Karl Ploessl

Glutamine-based PET imaging facilitates enhanced metabolic evaluation of gliomas in vivo

Glutamine-based PET imaging takes advantage of gliomas’ glutamine addiction and can be used to assess metabolic nutrient uptake in gliomas. A PET approach to brain tumors Positron emission tomography, or PET, is a common method of imaging tumors by detecting their uptake of a radioactively labeled tracer. Radiolabeled glucose, in particular, is often used for this type of imaging, because tumor cells are often highly dependent on glycolysis and require large amounts of glucose to maintain their metabolism. Unfortunately, this method cannot be used to image brain tumors, because regular brain cells are also highly dependent on glucose. Now, Venneti et al. have used mouse models and human patients to show that radiolabeled glutamine, which is also taken up by tumor cells, can be used to image brain tumors and distinguish them from normal brain and even from tumors that are no longer growing. Glucose and glutamine are the two principal nutrients that cancer cells use to proliferate and survive. Many cancers show altered glucose metabolism, which constitutes the basis for in vivo positron emission tomography (PET) imaging with 18F-fluorodeoxyglucose (18F-FDG). However, 18F-FDG is ineffective in evaluating gliomas because of high background uptake in the brain. Glutamine metabolism is also altered in many cancers, and we demonstrate that PET imaging in vivo with the glutamine analog 4-18F-(2S,4R)-fluoroglutamine (18F-FGln) shows high uptake in gliomas but low background brain uptake, facilitating clear tumor delineation. Chemo/radiation therapy reduced 18F-FGln tumor avidity, corresponding with decreased tumor burden. 18F-FGln uptake was not observed in animals with a permeable blood-brain barrier or neuroinflammation. We translated these findings to human subjects, where 18F-FGln showed high tumor/background ratios with minimal uptake in the surrounding brain in human glioma patients with progressive disease. These data suggest that 18F-FGln is avidly taken up by gliomas, can be used to assess metabolic nutrient uptake in gliomas in vivo, and may serve as a valuable tool in the clinical management of gliomas.

PET Imaging of Glutaminolysis in Tumors by 18F-(2S,4R)4-Fluoroglutamine

Changes in gene expression, metabolism, and energy requirements are hallmarks of cancer growth and self-sufficiency. Upregulation of the PI3K/Akt/mTor pathway in tumor cells has been shown to stimulate aerobic glycolysis, which has enabled 18F-FDG PET tumor imaging. However, of the millions of 18F-FDG PET scans conducted per year, a significant number of malignant tumors are 18F-FDG PET–negative. Recent studies suggest that several tumors may use glutamine as the key nutrient for survival. As an alternative metabolic tracer for tumors, 18F-(2S,4R)4-fluoroglutamine was developed as a PET tracer for mapping glutaminolytic tumors. Methods: A series of in vitro cell uptake and in vivo animal studies were performed to demonstrate tumor cell addiction to glutamine. Cell uptake studies of this tracer were performed in SF188 and 9L glioblastoma tumor cells. Dynamic small-animal PET studies of 18F-(2S,4R)4-fluoroglutamine were conducted in 2 animal models: xenografts produced in F344 rats by subcutaneous injection of 9L tumor cells and transgenic mice with M/tomND spontaneous mammary gland tumors. Results: In vitro studies showed that both transformed 9L and SF188 tumor cells displayed a high rate of glutamine uptake (maximum uptake, ≈16% dose/100 μg of protein). The cell uptake of 18F-(2S,4R)4-fluoroglutamine by SF188 cells is comparable to that of 3H-l-glutamine but higher than that of 18F-FDG. The tumor cell uptake can be selectively blocked. Biodistribution and PET studies showed that 18F-(2S,4R)4-fluoroglutamine localized in tumors with a higher uptake than in surrounding muscle and liver tissues. Data suggest that certain tumor cells may use glutamine for energy production. Conclusion: The results support that 18F-(2S,4R)4-fluoroglutamine is selectively taken up and trapped by tumor cells. It may be useful as a novel metabolic tracer for tumor imaging.

Preparation and Characterization of l-[5-11C]-Glutamine for Metabolic Imaging of Tumors

Recently, there has been a renewed interest in the study of tumor metabolism above and beyond the Warburg effect. Studies on cancer cell metabolism have provided evidence that tumor-specific activation of signaling pathways, such as the upregulation of the oncogene myc, can regulate glutamine uptake and its metabolism through glutaminolysis to provide the cancer cell with a replacement of energy source. Methods: We report a convenient procedure to prepare l-[5-11C]-glutamine. The tracer was evaluated in 9L and SF188 tumor cells (glioma and astrocytoma cell lines). The biodistribution of l-[5-11C]-glutamine in rodent tumor models was investigated by dissection and PET. Results: By reacting 11C-cyanide ion with protected 4-iodo-2-amino-butanoic ester, the key intermediate was obtained in good yield. After hydrolysis with trifluoroacetic and sulfonic acids, the desired optically pure l-[5-11C]-glutamine was obtained (radiochemical yield, 5% at the end of synthesis; radiochemical purity, >95%). Tumor cell uptake studies showed maximum uptake of l-[5-11C]-glutamine reached 17.9% and 22.5% per 100 μg of protein, respectively, at 60 min in 9L and SF188 tumor cells. At 30 min after incubation, more than 30% of the activity appeared to be incorporated into cellular protein. Biodistribution in normal mice showed that l-[5-11C]-glutamine had significant pancreas uptake (7.37 percentage injected dose per gram at 15 min), most likely due to the exocrine function and high protein turnover within the pancreas. Heart uptake was rapid, and there was 3.34 percentage injected dose per gram remaining at 60 min after injection. Dynamic small-animal PET studies in rats bearing xenografted 9L tumors and in transgenic mice bearing spontaneous mammary gland tumors showed a prominent tumor uptake and retention. Conclusion: The data demonstrated that this tracer was favorably taken up in the tumor models. The results suggest that l-[5-11C]-glutamine might be useful for probing in vivo tumor metabolism in glutaminolytic tumors.

Comparative Evaluation of 18F-Labeled Glutamic Acid and Glutamine as Tumor Metabolic Imaging Agents

18F-labeled (2S,4R)-4-fluoro-l-glutamine (4F-GLN) has demonstrated high uptake in tumor cells that undergo high growth and proliferation. Similar tumor targeting properties have also been observed for 18F-labeled (2S,4R)-4-fluoro-l-glutamate (4F-GLU), suggesting that both are useful imaging agents. A new labeling procedure facilitates the preparation of 18F-(2S,4R)4F-GLN and 18F-(2S,4R)4F-GLU with confirmed radiochemical and enantiomeric purity. Here, we report the preparation and comparative evaluation of 18F-(2S,4R)4F-GLN and 18F-(2S,4R)4F-GLU as tumor metabolic imaging agents. Methods: Uptake of enantiomerically pure 18F-(2S,4R)4F-GLN and 18F-(2S,4R)4F-GLU was determined in 3 tumor cell lines (9L, SF188, and PC-3) at selected time points. The in vitro cell uptake mechanism was evaluated by inhibition studies in 9L cells. In vivo biodistribution and PET studies were performed on male F344 rats bearing 9L tumor xenografts. Results: In vitro cell uptake studies showed that 18F-(2S,4R)4F-GLN displayed higher uptake than 18F-(2S,4R)4F-GLU. Amino acid transport system ASC (alanine-serine-cysteine–preferring; in particular, its subtype ASCT2 [SLC1A5 gene]) and system Xc− (SLC7A11 gene) played an important role in transporting 18F-(2S,4R)4F-GLN and 18F-(2S,4R)4F-GLU, respectively, across the membrane. After being transported into cells, a large percentage of 18F-(2S,4R)4F-GLN was incorporated into protein, whereas 18F-(2S,4R)4F-GLU mainly remained as the free amino acid in its original form. In vivo studies of 18F-(2S,4R)4F-GLN in the 9L tumor model showed a higher tumor uptake than 18F-(2S,4R)4F-GLU, whereas 18F-(2S,4R)4F-GLU had a slightly higher tumor-to-background ratio than 18F-(2S,4R)4F-GLN. Imaging studies showed that both tracers had fast accumulation in 9L tumors. Compared with 18F-(2S,4R)4F-GLU, 18F-(2S,4R)4F-GLN exhibited prolonged tumor retention reflecting its incorporation into intracellular macromolecules. Conclusion: Differences in uptake and metabolism in tumor cells were found between 18F-(2S,4R)4F-GLN and 18F-(2S,4R)4F-GLU. Both agents are potentially useful as metabolic tracers for tumor imaging.

Dopamine Transporter Binding in Smokers and Nonsmokers

Purpose: Regional brain concentrations of dopamine transporters have been examined to elucidate the neurochemical substrates of neurologic and psychiatric conditions. Many of these conditions are associated with increased (or decreased) cigarette smoking prevalence; therefore, current smoking may confound the results of these investigations. This study determined whether healthy current smokers and nonsmokers exhibit differences in dopamine transporter (DAT) binding measured by Tc-99m TRODAT SPECT. Materials and Methods: Tc-99m TRODAT SPECT brain scans were retrospectively evaluated in 46 nonsmokers and 8 current smokers, all of whom had been recruited and screened as healthy controls for previous imaging studies. The scans were acquired approximately 3 hours after the intravenous administration of 740 MBq (20 mCi) of Tc-99m TRODAT and were reconstructed with a simple bandpass filter. Regions of interest (ROIs) were placed manually on subregions of the right and left basal ganglia and distribution volume ratios (DVRs) were compared for the smoker and nonsmoker groups. Results: There were significant decreases in DAT binding in current smokers compared with nonsmokers in the caudate nuclei bilaterally, the right anterior putamen and the left posterior putamen. Conclusion: Reduced DAT binding in ROIs relevant to movement disorders as well as other neuropsychiatric conditions may have important implications for evaluating scans in these patient populations.

Enantioselective Radiosynthesis of Positron Emission Tomography (PET) Tracers Containing [18F]Fluorohydrins

Herein, we describe an operationally straightforward radiosynthesis of a chiral transition metal fluoride catalyst, [(18)F](salen)CoF, and its use for late-stage enantioselective aliphatic radiofluorination. We demonstrate the utility of the method by preparing single enantiomer experimental and clinically validated PET tracers that contain base-sensitive functional groups, epimerizable stereocenters, and nitrogen-rich motifs. Unlike the conventional radiosyntheses of these targets with [(18)F]KF, labeling with (salen)CoF is possible in the last step and under exceptionally mild conditions. These results constitute a rare example of a nucleophilic radiofluorination using a transition metal fluoride and highlight the potential of such reagents to enhance traditional methods for labeling aliphatic hydrocarbons.

123I-ADAM SPECT imaging of serotonin transporter binding in patients with night eating syndrome: a preliminary report

Night eating syndrome (NES) represents a delay in the circadian pattern of food intake, manifested by evening hyperphagia and/or nocturnal awakenings accompanied by ingestions of food. A neurobiological marker of NES has been implicated with the recently discovered therapeutic response to the selective serotonin reuptake inhibitor (SSRI) sertraline. This pilot SPECT (single photon emission computed tomography) study compared the serotonin transporter (SERT) uptake ratios of night eaters with those of healthy controls. Six night eaters underwent SPECT imaging using the radiopharmaceutical (123)I-ADAM. Uptake, compared with that of the cerebellum, was obtained for the midbrain, basal ganglia, and temporal lobes; uptake ratios in night eaters were compared with those of six healthy controls. Night eaters had significantly greater SERT uptake ratios in the midbrain than healthy controls. These findings, in conjunction with the therapeutic response of NES to sertraline, indicate that the serotonin system is involved in the pathophysiology of NES.

Dopamine transporter levels in cocaine dependent subjects

Cocaine use is a significant problem in the US and it is well established that cocaine binds to the dopamine transporter (DAT) in the brain. This study was designed to determine if the DAT levels measured by 99mTc TRODAT SPECT (single photon emission computed tomography) brain scans are altered in cocaine dependent subjects and to explore clinical correlates of such alterations. SPECT brain scans were acquired on 21 cocaine dependent subjects and 21 healthy matched controls. There were significantly higher DAT levels in cocaine dependent subjects compared to controls for the anterior putamen (p=0.003; Cohens d effect size=0.98), posterior putamen (p<0.001; effect size=1.32), and caudate (p=0.003; effect size=0.97). DAT levels in these regions were 10%, 17%, and 8% higher in the cocaine dependent subjects compared to controls. DAT levels were unrelated to craving, severity of cocaine use, or duration of cocaine use, but DAT levels in the caudate and anterior putamen were significantly (p<0.05) negatively correlated with days since last use of cocaine.

[18F](2S,4S)-4-(3-Fluoropropyl)glutamine as a Tumor Imaging Agent

Although the growth and proliferation of most tumors is fueled by glucose, some tumors are more likely to metabolize glutamine. In particular, tumor cells with the upregulated c-Myc gene are generally reprogrammed to utilize glutamine. We have developed new 3-fluoropropyl analogs of glutamine, namely [18F](2S,4R)- and [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 3 and 4, to be used as probes for studying glutamine metabolism in these tumor cells. Optically pure isomers labeled with 18F and 19F (2S,4S) and (2S,4R)-4-(3-fluoropropyl)glutamine were synthesized via different routes and isolated in high radiochemical purity (≥95%). Cell uptake studies of both isomers showed that they were taken up efficiently by 9L tumor cells with a steady increase over a time frame of 120 min. At 120 min, their uptake was approximately two times higher than that of l-[3H]glutamine ([3H]Gln). These in vitro cell uptake studies suggested that the new probes are potential tumor imaging agents. Yet, the lower chemical yield of the precursor for 3, as well as the low radiochemical yield for 3, limits the availability of [18F](2S,4R)-4-(3-fluoropropyl)glutamine, 3. We, therefore, focused on [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4. The in vitro cell uptake studies suggested that the new probe, [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4, is most sensitive to the LAT transport system, followed by System N and ASC transporters. A dual-isotope experiment using l-[3H]glutamine and the new probe showed that the uptake of [3H]Gln into 9L cells was highly associated with macromolecules (>90%), whereas the [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4, was not (<10%). This suggests a different mechanism of retention. In vivo PET imaging studies demonstrated tumor-specific uptake in rats bearing 9L xenographs with an excellent tumor to muscle ratio (maximum of ∼8 at 40 min). [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4, may be useful for testing tumors that may metabolize glutamine related amino acids.

Multidentate 18F-Polypegylated Styrylpyridines As Imaging Agents for Aβ Plaques in Cerebral Amyloid Angiopathy (CAA)

β-Amyloid plaques (Aβ plaques) in the brain are associated with cerebral amyloid angiopathy (CAA). Imaging agents that could target the Aβ plaques in the living human brain would be potentially valuable as biomarkers in patients with CAA. A new series of (18)F styrylpyridine derivatives with high molecular weights for selectively targeting Aβ plaques in the blood vessels of the brain but excluded from the brain parenchyma is reported. The styrylpyridine derivatives, 8a-c, display high binding affinities and specificity to Aβ plaques (K(i) = 2.87, 3.24, and 7.71 nM, respectively). In vitro autoradiography of [(18)F]8a shows labeling of β-amyloid plaques associated with blood vessel walls in human brain sections of subjects with CAA and also in the tissue of AD brain sections. The results suggest that [(18)F]8a may be a useful PET imaging agent for selectively detecting Aβ plaques associated with cerebral vessels in the living human brain.