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.
We report a convenient procedure to prepare l-5-(11)C-glutamine. The tracer was evaluated in 9L and SF188 tumor cells (glioma and astrocytoma cell lines). The biodistribution of l-5-(11)C-glutamine in rodent tumor models was investigated by dissection and PET.
By reacting (11)C-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-(11)C-glutamine was obtained (radiochemical yield, 5% at the end of synthesis; radiochemical purity, >95%). Tumor cell uptake studies showed maximum uptake of l-5-(11)C-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-(11)C-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.
The data demonstrated that this tracer was favorably taken up in the tumor models. The results suggest that l-5-(11)C-glutamine might be useful for probing in vivo tumor metabolism in glutaminolytic tumors.
Rationale
F17464, a dopamine D3 receptor antagonist with relatively high D3 selectivity (70 fold vs D2
in vitro
), exhibits an antipsychotic profile in preclinical studies, and therapeutic efficacy ...was demonstrated in a randomized placebo-controlled clinical trial in patients with schizophrenia (Bitter et al. Neuropsychopharmacology 44(11):1917–1924,
2019
).
Objective
This open-label study in healthy male subjects aimed at characterizing F17464 binding to D3/D2 receptors and the time course of receptor occupancy using positron emission tomography (PET) imaging with a D3-preferring tracer,
11
C-(+)-PHNO.
Methods
PET scans were performed at baseline and following a single 30 mg or 15 mg dose of F17464 (3 subjects/dose), and blood samples were collected for pharmacokinetic analysis. Receptor occupancy was calculated based upon reduction in binding potential of the tracer following F17464 administration. The relationship between plasma F17464 concentration and D3/D2 receptor occupancy was modeled and the plasma concentration corresponding to 50% receptor occupancy (EC
50
) calculated.
Results
Both doses of F17464 robustly blocked
11
C-(+)-PHNO D3 receptor binding, with substantial occupancy from 1 h post-administration, which increased at 6–9 h (89–98% and 79-87% for the 30 mg and 15 mg groups, respectively) and remained detectable at 22 h. In contrast, D2 binding was only modestly blocked at all time points (< 18%).
F17464 exhibited a combination of rapid peripheral kinetics and hysteresis (persistence of binding 22 h post-dose despite low plasma concentration). The best estimate of the EC
50
was 19 ng ml
−1
(~ 40 nM).
Conclusion
Overall, F17464 was strongly D3-selective in healthy volunteers, a unique profile for an antipsychotic candidate drug.
Imaging agents targeting β-amyloid (Aβ) may be useful for diagnosis and treatment of patients with Alzheimer's disease (AD). Compounds 3e and 4e are fluorinated stilbene derivatives displaying high ...binding affinities for Aβ plaques in AD brain homogenates (K i = 15 ± 6 and 5.0 ± 1.2 nM, respectively). In vivo biodistributions of 18F3e and 18F4e in normal mice exhibited excellent brain penetrations (5.55 and 9.75% dose/g at 2 min), and rapid brain washouts were observed, especially for 18F4e (0.72% dose/g at 60 min). They also showed in vivo plaque labeling in APP/PS1 or Tg2576 transgenic mice, animal models for AD. Autoradiography of postmortem AD brain sections and AD homogenate binding studies confirmed the selective and specific binding properties to Aβ plaques. In conclusion, the preliminary results strongly suggest that these fluorinated stilbene derivatives, 18F3e and 18F4e, are suitable candidates as Aβ plaque imaging agents for studying patients with AD.
Glioblastoma (GBM) is the most common primary adult brain malignancy with an extremely poor prognosis and a median survival of fewer than two years. A key reason for this high mortality is that the ...blood-brain barrier (BBB) significantly restricts systemically delivered therapeutics to brain tumors. High-intensity focused ultrasound (HIFU) with microbubbles is a methodology being used in clinical trials to noninvasively permeabilize the BBB for systemic therapeutic delivery to GBM. Topotecan is a topoisomerase inhibitor used as a chemotherapeutic agent to treat ovarian and small cell lung cancer. Studies have suggested that topotecan can cross the BBB and can be used to treat brain metastases. However, pharmacokinetic data demonstrated that topotecan peak concentration in the brain extracellular fluid after systemic injection was ten times lower than in the blood, suggesting less than optimal BBB penetration by topotecan. We hypothesize that HIFU with microbubbles treatment can open the BBB and significantly increase topotecan concentration in the brain. We radiolabeled topotecan with
C and acquired static and dynamic positron emission tomography (PET) scans to quantify
C topotecan uptake in the brains of normal mice and mice after HIFU treatment. We found that HIFU treatments significantly increased
C topotecan brain uptake. Moreover, kinetic analysis of the
C topotecan dynamic PET data demonstrated a substantial increase in
C topotecan volume of distribution in the brain. Furthermore, we found a decrease in
C topotecan brain clearance, confirming the potential of HIFU to aid in the delivery of topotecan through the BBB. This opens the potential clinical application of
C topotecan as a tool to predict topotecan loco-regional brain concentration in patients with GBMs undergoing experimental HIFU treatments.
A new F-18-labeled phenylthiophenyl derivative specific for imaging of serotonin transporters (SERT) in the brain by positron emission tomography (PET) is described. Fluorinated phenylthiophenyl ...derivative, ACF, 2-(2-amino-4-chloro-5-fluorophenyl)thio-N,N-dimethyl-benzenmethanamine, was prepared by first coupling 2,5-dichloro-4-nitroaniline with 2-mercapto-N,N-dimethylbenzamide. The amino group of the coupled adduct was converted to a fluoro group through a Schiemann reaction. Subsequently, a one pot reduction of both nitro and amide groups by BH3−tetrahydrofuran yielded the nonradioactive ACF (yield 25%). In vitro binding assays using cell membrane homogenates of LLC cells expressing SERT, dopamine transporters (DAT), or norepinephrine transporters (NET) showed excellent binding affinity and selectivity for SERT (K i = 0.05, 3020, and 650 nM for SERT, DAT, and NET, respectively). For preparation of the 18FACF, the NH2 group of the initially coupled adduct was converted to the trimethylammonium salt, which was replaced by 18Ffluoride in the presence of Kryptofix 222 and potassium carbonate. The final product, 18FACF, was obtained after a borane and stannous chloride reduction reaction. The combined two step reaction gave a radiochemical yield of 10−15% (EOB) and a radiochemical purity of >99%. Synthesis of the novel PET tracer, 18FACF, as a probe for binding to SERT in the brain was successfully achieved. The new tracer 18FACF showed excellent brain penetration and selective localization after an iv injection in rats (brain uptake at 2, 30, 60, 120, and 240 min was 3.27, 1.28, 0.69, 0.21, and 0.06% dose/organ, respectively). The hypothalamus/cerebellum ratio at 60 min post iv injection was 3.55. This specific localization in the hypothalamus was blocked by pretreatment of (+)McN5652. This novel ligand is a potential PET tracer for in vivo evaluation of SERT in the brain.
Objectives: Glutamine is a readily available nutrient used for energy in both normal and tumor cells 1. L-11Cglutamine takes advantage of various tumor metabolic pathways for when 18FFDG is limited. ...There is therefore a need in the PET community for an automated synthesis of L-11Cglutamine. We have developed a method for the routine automated production of L-11Cglutamine. Methods: L-11Cglutamine was prepared with the use of an in-house Glutamine Module attached to a Siemens H11CN(Cyanide)Box. 11CCO2, produced from a Siemens Eclipse HP Cyclotron, is converted into H11CN with the use of nickel/platinum catalysts. Radioactive cyanide gas is trapped in reaction vessel 1 with the use of 18-crown-6/cesium hydrogen carbonate base in N,N-dimethylformamide. The glutamine precursor, (s)-tert-butyl 2-((tert-butoxycarbonyl)amine)-4- iodobutanoat, is then added and heated to 90 oC for 8 min. Trapping on a pre-conditioned t-C18 plus cartridge is achieved with the use of HPLC grade water. The radiolabeled intermediate is then released into reaction vessel 2 by way of acetonitrile. This is then dried at 120 oC over a stream of argon. This process is repeated with the use of acetonitrile. The dried intermediate is then deprotected with the use of (4:1) trifluoroacetic acid:sulfuric acid and heated to 90 oC for 5 min. Quenching with sterile water for injection and slowly passing through 5.0 g of Ag11 A-8 resin provided L-11Cglutamine. Chemical identity, purity and L:D isomeric ratio of the labeled radioligand was determined by reversed-phase HPLC. Quality control (QC) conditions were tested and passed for each production of L-11Cglutamine; these tests include radionuclidic purity, pH, chemical identity, ratio of L/D-glutamine isomer, excipients, half-life and sterility. An in vitro cell uptake assay was performed using a glioblastoma (GBM) cell line to evaluate the reactivity and specificity of L-11Cglutamine. Excess L-glutamine was used to block control samples. SOPs and Batch Record were created prior to validation runs. Results: L-11Cglutamine was successfully produced operating our automated in-house module connected to a Siemens H11CN Box. Routine production provided 17.5 - 32.5 mCi (n = 6) at end-of-synthesis (EOS), starting with approximately 1,400 mCi of 11CCO2. The average radiochemical purity = 95 ± 1.6% with an average enantiomeric excess (e.e.) = 96 ± 1.8 % and a pH value ranging from 7.5 to 8.0. Total synthesis time ranges from 74 to 82 min. In in vitro assay, L-11Cglutamine was taken up by GBM cells and was inhibited by ~65% when blocked with excess cold L-glutamine, indicating binding specificity. Conclusions: A method for the automated production of L-11Cglutamine was achieved with the use of an in-house module attached to a Siemens H11CN module. Specificity and reactivity of the produced L-11Cglutamine was validated in vitro using a glioblastoma cell line. This provides a platform for consistent-routine production of L-11Cglutamine which can be used as a potential PET imaging agent.
Development of imaging agents for pancreatic beta cell mass may provide tools for studying insulin-secreting beta cells and their relationship with diabetes mellitus. In this paper, a new imaging ...agent,
18F(+)-2-oxiranyl-3-isobutyl-9-(3-fluoropropoxy)-10-methoxy-2,3,4,6,7,11b-hexahydro-1H-pyrido2,1-
aisoquinoline
18F(+)
4, which displays properties targeting vesicular monoamine transporter 2 (VMAT2) binding sites of beta cells in the pancreas, was evaluated as a positron emission tomography (PET) agent for estimating beta cell mass in vivo. The hydrolyzable epoxide group of (+)
4 may provide a mechanism for shifting biodistribution from liver to kidney, thus reducing the background signal.
Both
18F- and
19F-labeled (+) and (−) isomers of
4 were synthesized and evaluated. Organ distribution was carried out in normal rats. Uptake of
18F(+)
4 in pancreas of normal rats was measured and correlated with blocking studies using competing drugs, (+)dihydrotetrabenazine (+)-DTBZ or 9-fluoropropyl-(+)dihydro tetrabenazine FP-(+)-DTBZ, (+)
2.
In vitro binding study of VMAT2 using rat brain striatum showed a
K
i value of 0.08 and 0.15 nM for the (+)
4 and (±)
4, respectively. The in vivo biodistribution of
18F(+)
4 in rats showed the highest uptake in the pancreas (2.68 %ID/g at 60 min postinjection). In vivo competition experiments with cold FP-(+)-DTBZ, (+)
2, (3.5 mg/kg, 5 min iv pretreatment) led to a significant reduction of pancreas uptake (85% blockade at 60 min). The inactive isomer
18F(−)
4 showed significantly lower pancreas uptake (0.22 %ID/g at 30 min postinjection). Animal PET imaging studies of
18F(+)4 in normal rats demonstrated an avid pancreatic uptake in rats.
The preliminary results suggest that the epoxide,
18F(+)
4, is highly selective in binding to VMAT2 and it has an excellent uptake in the pancreas of rats. The liver uptake was significantly reduced through the use of the epoxide group. Therefore, it may be potentially useful for imaging beta cell mass in the pancreas.
Objectives: Chronic pain results in long-term hyper-excitability (LTH) associated with activation of protein kinase G‐1α (PKG) in the dorsal root ganglion (DRG) with the region of pain. The balanol ...derivative NOP46, N-((3R,4R)-4-(4-(2-fluoro-3-methoxy-6-propoxybenzoyl)benzamido)pyrrolidin-3-yl)-1H-indazole-5-carboxamide has a high affinity to activated PKG. The chemical structure of NOP46 has an anisole moiety where a Carbon-11 methyl group can be inserted. The short half-life of 11CNOP46 permits multiple administrations of tracers in short intervals to the same subject and may provide a novel method to visualize chronic pain in human. Methods: O-Desmethyl 11CNOP46 was prepared by way of methylation with 11C-methyl triflate in GE-FxC Pro synthesis module. Chemical identity of the labeled product 11CNOP46 was determined by reversed-phase HPLC. In vitro biodistribution and dosimetry studies of 11CNOP46 was carried out in normal rats to determine the route of excretion and to estimate human dosimetry. GLP Toxicology studies were performed using IV dosing of 22, 44, and 110 μg/kg in anticipation of a first-in-human clinical trial. Results: 11CNOP46 was successfully produced on the GE FxC Pro platform. The range of the production results (n = 3) are Radiochemical yield = 27.7-31.8 mCi at end-of-synthesis (EOS), radiochemical purity > 99 %, drug concentration = 1.02-1.93 µg/mL, total chemical impurity = 0.09-0.12 µg/mL, and specific activity = 870-1,828 mCi/µmol. HPLC confirmed the chemical identity. Three successful validations were carried out. Rodent biodistribution studies were conducted with all major organs at 5-time points (3 male, 3 female rats for each time points). Human dosimetry was estimated using OLINDA software. Biodistribution study demonstrated primary hepatobiliary excretion and a calculated TEDE of 0.00806 and 0.005 rem/mCi in males and females, respectively. Preclinical toxicology studies demonstrated that a single intravenous (bolus) administration of NOP46 to Sprague Dawley rats at dose level of 110 μg/kg was systemically well tolerated with no evidence of any treatment related immediate or any delayed adverse effects. Conclusions: The production method of 11CNOP46 has been successfully developed and validated. Human dosimetry has been estimated using the data obtained from our rodent studies. First-in-human trial of 11CNOP46 may provide a novel method to assess chronic pain in human individuals.
This paper describes a novel series of
18F-labeled polyethyleneglycol (PEG)-stilbene derivatives as potential β-amyloid (Aβ) plaque-specific imaging agents for positron emission tomography (PET). In ...these series of compounds,
18F is linked to the stilbene through a PEG chain, of which the number of ethoxy groups ranges from 2 to 5. The purpose of adding PEG groups is to lower the lipophilicity and improve bioavailability. The syntheses of the “cold” compounds and the
18F-labeled PEG stilbene derivatives are successfully achieved. All of the fluorinated stilbenes displayed high binding affinities in an assay using postmortem AD brain homogenates (
K
i=2.9–6.7 nM). Labeling was successfully performed by a substitution of the mesylate group of
10a–d by
18Ffluoride giving the target compounds
18F
12a–d (EOS, specific activity, 900–1500 Ci/mmol; radiochemical purity >99%). In vivo biodistribution of these novel
18F ligands in normal mice exhibited excellent brain penetrations and rapid washouts after an intravenous injection (6.6–8.1 and 1.2–2.6 %dose/g at 2 and 60 min, respectively). Autoradiography of postmortem AD brain sections of
18F
12a–d confirmed the specific binding related to the presence of Aβ plaques. In addition, in vivo plaque labeling can be clearly demonstrated with these
18F-labeled agents in transgenic mice (Tg2576), a useful animal model for Alzheimer's disease. In conclusion, the preliminary results strongly suggest these fluorinated PEG stilbene derivatives are suitable candidates as Aβ plaque imaging agents for studying patients with Alzheimer's disease.
Accumulation of β-amyloid aggregates (Aβ) in the brain is linked to the pathogenesis of Alzheimer's disease (AD). We report a novel approach for producing 1,4-diphenyltriazoles as probes for ...targeting Aβ aggregates in the brain. The imaging probes, a series of substituted tricyclic 1,4-diphenyltriazoles showing excellent binding affinities to Aβ aggregates (K i = 4−30 nM), were conveniently assembled by “click chemistry.” Two radioiodinated probes, 125I10a and 125I10b, and two radiofluorinated probes, 18F17a and 18F17b, exhibited moderate lipophilicities and showed excellent initial brain penetrations and fast washouts from the normal mouse brain. In vitro autoradiography of postmortem AD brain sections and homogenates showed that these triazoles were binding to Aβ plaques. Preliminary results strongly suggest that use of click chemistry, which led to a 1,4-diphenyltriazole-based core, is a highly convenient and flexible approach for assembling novel imaging agents for targeting Aβ aggregates in senile plaques in the living human brain.
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