[en] Complete text of publication follows: Introduction There is evidence that clearance of beta-amyloid from brain into blood is mediated by P-glycoprotein (ABCB1) at the blood brain barrier (BBB). Pharmacological induction of ABCB1 is a currently investigated approach for the treatment of Alzheimer's disease. For the development of ABCB1-inducing therapeutics, the availability of a PET tracer to measure ABCB1 function at the BBB would be very helpful. Currently, available avid ABCB1 substrates for PET (e.g., (R)-[¹¹C]verapamil and [¹¹C]N-desmethyl-loperamide) are not suitable to measure ABCB1 induction at the BBB due to their very low brain uptake and presence of brain-penetrant radiolabeled metabolites. We evaluated the ability of the recently introduced weak ABCB1 substrate [¹¹C]metoclopramide ¹¹C]MCP) [1 (to measure ABCB1 induction at the mouse BBB. Methods [¹¹C]MCP was synthesized in a Tracerlab FX C Pro synthesis module by O-methylation of O-desmethyl-metoclopramide with [¹¹C]methyl triflate. Groups of female C57BL/6J mice aged approximately 170 days (n = 4-6 per group) underwent a baseline [¹¹C]MCP PET scan followed by treatment with two different validated ABCB1 induction protocols (PCN: 25 mg/kg i.p. over 7 days or rifampicin: 10 mg/kg i.p. over 5 days) or vehicle solution (safflower oil) and a second [¹¹C]MCP PET scan at the end of treatment. At the end of each PET scan, a small blood sample was collected from each animal and counted for radioactivity in a gamma counter. In separate groups of animals, radiolabelled metabolites of [¹¹C]MCP were analysed by radio-TLC at 15 min after injection (n = 2-5 per group). As an outcome parameter of ABCB1 function, the elimination slope for radioactivity washout from the brain was determined from the log-transformed brain time-activity curves (kE,brain, h^-1). All values are reported as mean ± SD. Results [¹¹C]MCP was synthesized in a decay-corrected radiochemical yield of 8.1 ± 1.6% (n = 15, based on [¹¹C]methane) in a total synthesis time of 31 min with a radiochemical purity >99% and a molar activity at end of synthesis of 96.5 ± 45.3 GBq/μmol. To the radiotracer solution carrier was added (2 mg/kg) to reduce peripheral metabolism of [¹¹C]MCP [1]. KE,brain was not significantly different between baseline, rifampicin-and vehicle-treated groups (0.687 ± 0.056 vs. 0.701 ± 0.075 and 0.775 ± 0.071 h^-1, p ≤ 0.05). Significantly higher kE,brain was observed in the PCN-treated group (0.883 ± 0.195 h^-1, p ≤ 0.001, 1-way ANOVA with Tukey's test) compared to the baseline group. No significant differences between the four groups were observed in blood radioactivity concentrations measured at the end of the PET scan. In plasma and brain, the percentage of unchanged [¹¹C (MCP at 15 min after injection was comparable for all groups (plasma: 33.9 ± 6.2%, brain: 90.9 ± 2.8%). Conclusion We established the synthesis of [¹¹C]MCP and evaluated two validated ABCB1 induction protocols. [¹¹C]MCP was found to lack brain-penetrant radiolabelled metabolites. Treatment with PCN significantly enhanced washout of radioactivity from the brain, which indicated that [¹¹C]MCP is a suitable PET tracer to measure ABCB1 induction at the BBB

[en] Aim of this study was to investigate if cellular ["1"8F]FBPA uptake can be increased upon preloading with amino acids. ["1"8F]FBPA uptake was assessed in HuH-7, CaCo-2 and B16-F1 cells pretreated with different concentrations or incubation times of L-BPA, L-tyrosine or L-DOPA. Without preloading, highest uptake of ["1"8F]FBPA was observed in B16-F1 cells, followed by CaCo-2 cells and HuH-7 cells. In all cell lines higher ["1"8F]FBPA accumulation (up to 1.65-fold) was obtained with increasing L-BPA, L-DOPA and L-tyrosine concentrations. - Highlights: • Uptake of ["1"8F]FBPA was assessed in HuH-7, CaCo-2 and B16-F1 cells. • ["1"8F]FBPA uptake is highly dependent on the incubation medium. • ["1"8F]FBPA uptake was higher after preloading with L-BPA, L-DOPA and L-tyrosine.

[en] We modified a commercially available synthesis module for nucleophilic ["1"8F]fluorinations (TRACERlab"T"M FX_F_D_G, GE Healthcare) to enable the reliable synthesis of 2-["1"8F]fluoro-4-borono-L-phenylalanine (["1"8F]FBPA) via direct electrophilic substitution of 4-borono-L-phenylalanine with ["1"8F]F_2 gas. ["1"8F]FBPA was obtained with a RCY of 8.5±2.0% and a radiochemical purity of 98±1% in a total synthesis time of 72±7 min (n=22). The modified synthesis module might also be useful for the synthesis of other ["1"8F]radiopharmaceuticals via electrophilic substitution reactions while still being suitable for nucleophilic substitution reactions. - Highlights: • Automated synthesis of ["1"8F]FBPA was developed via reaction of BPA with ["1"8F]F_2. • ["1"8F]FBPA was obtained with a RCY of 8.5±2.0%. • Nucleophilic synthesis module was adapted for electrophilic ["1"8F]fluorinations. • Modified synthesis module may also enable other electrophilic ["1"8F]fluorinations

[en] We transferred the previously published manual synthesis of ["1"8F]ciprofloxacin (decay-corrected RCY: 5.5±1.0%) to an automated synthesis module (TRACERlab"T"M FX_F_D_G, GE Healthcare) and observed a strong decrease in RCY (0.4±0.4%). When replacing the standard 15-mL glassy carbon reactor of the synthesis module with a 3-mL V-shaped borosilicate glass reactor a considerable improvement in RCY was observed. ["1"8F]Ciprofloxacin was obtained in a RCY of 2.7±1.4% (n=23) with a specific activity at EOS of 1.4±0.5 GBq/µmol in a synthesis time of 160 min. - Highlights: • Automated synthesis of ["1"8F]ciprofloxacin in a TRACERlab"T"M FX_F_D_G (GE Healthcare) synthesis module was developed. • Dependence of radiochemical yield on reactor type was observed. • 3-mL V-shaped borosilicate glass reactor gave higher radiochemical yield as compared with standard 15-mL glassy carbon reactor. • V-shaped borosilicate glass reactor might also give higher radiochemical yield for other ["1"8F]radiotracers than ["1"8F]ciprofloxacin

[en] Overactivity of the multidrug efflux transporter P-glycoprotein (P-gp) at the blood-brain barrier (BBB) is believed to play an important role in resistance to central nervous system drug treatment. (R)-[¹¹C]verapamil (VPM) PET can be used to measure the function of P-gp at the BBB, but low brain uptake of VPM hampers the mapping of regional differences in cerebral P-gp function and expression. The aim of this study was to evaluate the dose-response relationship of two potent P-gp inhibitors and to investigate if increased brain uptake of VPM mediated by P-gp inhibition can be used to assess regional differences in P-gp activity. Two groups of Sprague-Dawley rats (n = 12) underwent single VPM PET scans at 120 min after administration of different doses of the P-gp inhibitors tariquidar and elacridar. In an additional six rats, paired VPM PET scans were performed before and after administration of 3 mg/kg tariquidar. Inhibitor administration resulted in an up to 11-fold increase in VPM brain distribution volumes (DV) with half-maximum effective dose (ED₅₀) values of 3.0 ± 0.2 and 1.2 ± 0.1 mg/kg for tariquidar and elacridar, respectively. In paired PET scans, 3 mg/kg tariquidar resulted in regionally different enhancement of brain activity distribution, with lowest DV in cerebellum and highest DV in thalamus. Our data show that tariquidar and elacridar are able to increase VPM brain distribution in rat brain up to 11-fold over baseline at maximum effective doses, with elacridar being about three times more potent than tariquidar. Regional differences in tariquidar-induced modulation of VPM brain uptake point to regional differences in cerebral P-gp function and expression in rat brain. (orig.)

[en] Introduction: In recent years extra-corporal application of boron neutron capture therapy (BNCT) was evaluated for liver primary tumors or liver metastases. A prerequisite for such a high-risk procedure is proof of preferential delivery and high uptake of a ¹⁰B-pharmaceutical in liver malignancies. In this work we evaluated in a preclinical tumor model if [¹⁸F]FBPA tissue distribution measured with PET is able to predict the tissue distribution of [¹⁰B]L-BPA. Methods: Tumor bearing mice (hepatocellular carcinoma cell line, HuH-7) were either subject of a [¹⁸F]FBPA-PET scan with subsequent measurement of radioactivity content in extracted organs using a gamma counter or injected with [¹⁰B]L-BPA with tissue samples analyzed by prompt gamma activation analysis (PGAA) or quantitative neutron capture radiography (QNCR). The impact of L-tyrosine, L-DOPA and L-BPA preloading on the tissue distribution of [¹⁸F]FBPA and [¹⁰B]L-BPA was evaluated and the pharmacokinetics of [¹⁸F]FBPA investigated by compartment modeling. Results: We found a significant correlation between [¹⁸F]FBPA and [¹⁰B]L-BPA uptake in tumors and various organs as well as high accumulation levels in pancreas and kidneys as reported in previous studies. Tumor-to-liver ratios of [¹⁸F]FBPA ranged from 1.2 to 1.5. Preloading did not increase the uptake of [¹⁸F]FBPA or [¹⁰B]L-BPA in any organ and compartment modeling showed no statistically significant differences in [¹⁸F]FBPA tumor kinetics. Conclusions: [¹⁸F]FBPA-PET predicts [¹⁰B]L-BPA concentration after amino acid preloading in HuH-7 hepatocellular carcinoma models. Preloading had no effect on tumor uptake of [¹⁸F]FBPA. Advances in knowledge: Despite differences in chemical structure and administered dose [¹⁸F]FBPA and [¹⁰B]L-BPA demonstrate an equivalent biodistribution in a preclinical tumor model. Implications for patient care: [¹⁸F]FBPA-PET is suitable for treatment planning and dose calculations in BNCT applications for liver malignancies. However, alternative tracers with more favorable tumor-to-liver ratios should be investigated.

[en] One important mechanism for chemoresistance of tumours is overexpression of the adenosine triphosphate-binding cassette transporter P-glycoprotein (Pgp). Pgp reduces intracellular concentrations of chemotherapeutic drugs. The aim of this study was to compare the suitability of the radiolabelled Pgp inhibitors [¹¹C]tariquidar and [¹¹C]elacridar with the Pgp substrate radiotracer (R)-[¹¹C]verapamil for discriminating tumours expressing low and high levels of Pgp using small-animal PET imaging in a murine breast cancer model. Murine mammary carcinoma cells (EMT6) were continuously exposed to doxorubicin to generate a Pgp-overexpressing, doxorubicin-resistant cell line (EMT6AR1.0 cells). Both cell lines were subcutaneously injected into female athymic nude mice. One week after implantation, animals underwent PET scans with [¹¹C]tariquidar (n = 7), [¹¹C]elacridar (n = 6) and (R)-[¹¹C]verapamil (n = 7), before and after administration of unlabelled tariquidar (15 mg/kg). Pgp expression in tumour grafts was evaluated by Western blotting. [¹¹C]Tariquidar showed significantly higher retention in Pgp-overexpressing EMT6AR1.0 compared with EMT6 tumours: the mean ± SD areas under the time-activity curves in scan 1 from time 0 to 60 min (AUC_0-60) were 38.8 ± 2.2 min and 25.0 ± 5.3 min (p = 0.016, Wilcoxon matched pairs test). [¹¹C]Elacridar and (R)-[¹¹C]verapamil were not able to discriminate Pgp expression in tumour models. Following administration of unlabelled tariquidar, both EMT6Ar1.0 and EMT6 tumours showed increases in uptake of [¹¹C]tariquidar, [¹¹C]elacridar and (R)-[¹¹C]verapamil. Among the tested radiotracers, [¹¹C]tariquidar performed best in discriminating tumours expressing high and low levels of Pgp. Therefore [¹¹C]tariquidar merits further investigation as a PET tracer to assess Pgp expression levels in solid tumours. (orig.)

[en] 

Background

[¹¹C]erlotinib has been proposed as a PET tracer to visualize the mutational status of the epidermal growth factor receptor (EGFR) in cancer patients. For clinical use, a stable, reproducible and high-yielding radiosynthesis method is a prerequisite. In this work, two production schemes for [¹¹C]erlotinib applied in a set of preclinical and clinical studies, starting from either [¹¹C]CH₄ or [¹¹C]CO₂, are presented and compared in terms of radiochemical yields, molar activities and overall synthesis time. In addition, a time-efficient RP-HPLC method for quality control is presented, which requires not more than 1 min.

Results

[¹¹C]erlotinib was reliably produced applying both methods with decay-corrected radiochemical yields of 13.4 ± 6.2% and 16.1 ± 4.9% starting from in-target produced [¹¹C]CO₂ and [¹¹C]CH₄, respectively. Irradiation time for the production of [¹¹C]CO₂ was higher in order to afford final product amounts sufficient for patient application. Overall synthesis time was comparable, mostly attributable to adaptions in the semi-preparative HPLC protocol. Molar activities were 1.8-fold higher for the method starting from [¹¹C]CH₄ (157 ± 68 versus 88 ± 57 GBq/μmol at the end of synthesis).

Conclusions

This study compared two synthetic protocols for the production of [¹¹C]erlotinib with in-target produced [¹¹C]CO₂ or [¹¹C]CH₄. Both methods reliably yielded sufficiently high product amounts for preclinical and clinical use.

[en] Introduction: The adenosine triphosphate-binding cassette (ABC) transporter P-glycoprotein (Pgp) protects the brain from accumulation of lipophilic compounds by active efflux transport across the blood–brain barrier. Changes in Pgp function/expression may occur in neurological disorders, such as epilepsy, Alzheimer’s or Parkinson’s disease. In this work we investigated the suitability of the radiolabeled Pgp inhibitors [¹¹C]elacridar and [¹¹C]tariquidar to visualize Pgp density in rat brain with PET. Methods: Rats underwent a first PET scan with [¹¹C]elacridar (n = 5) or [¹¹C]tariquidar (n = 6) followed by a second scan with the Pgp substrate (R)-[¹¹C]verapamil after administration of unlabeled tariquidar at a dose which half-maximally inhibits cerebral Pgp (3 mg/kg). Compartmental modeling using an arterial input function and Logan graphical analysis were used to estimate rate constants and volumes of distribution (V_T) of radiotracers in different brain regions. Results: Brain PET signals of [¹¹C]elacridar and [¹¹C]tariquidar were very low (∼ 0.5 standardized uptake value, SUV). There was a significant negative correlation between V_T and K₁ (i.e. influx rate constant from plasma into brain) values of [¹¹C]elacridar or [¹¹C]tariquidar and V_T and K₁ values of (R)-[¹¹C]verapamil in different brain regions which was consistent with binding of [¹¹C]inhibitors to Pgp and efflux of (R)-[¹¹C]verapamil by Pgp. Conclusion: The small Pgp binding signals obtained with [¹¹C]elacridar and [¹¹C]tariquidar limit the applicability of these tracers to measure cerebral Pgp density. PET tracers with higher (i.e. subnanomolar) binding affinities will be needed to visualize the low density of Pgp in brain

[en] Objectives: With the aim to develop a PET tracer to visualize P-glycoprotein (Pgp) expression levels in different organs, the Pgp inhibitor MC113 was labeled with ¹¹C and evaluated using small-animal PET. Methods: [¹¹C]MC113 was synthesized by reaction of O-desmethyl MC113 with [¹¹C]methyl triflate. Small-animal PET was performed with [¹¹C]MC113 in FVB wild-type and Mdr1a/b^(-/-) mice (n = 3 per group) and in a mouse model of high (EMT6Ar1.0) and low (EMT6) Pgp expressing tumor grafts (n = 5). In the tumor model, PET scans were performed before and after administration of the reference Pgp inhibitor tariquidar (15 mg/kg). Results: Brain uptake of [¹¹C]MC113, expressed as area under the time-activity curve from time 0 to 60 min (AUC_0-60), was moderately but not significantly increased in Mdr1a/b^(-/-) compared with wild-type mice (mean ± SD AUC_0-60, Mdr1a/b^(-/-): 88 ± 7 min, wild-type: 62 ± 6 min, P = 0.100, Mann Whitney test). In the tumor model, AUC_0-60 values were not significantly different between EMT6Ar1.0 and EMT6 tumors. Neither in brain nor in tumors was activity concentration significantly changed in response to tariquidar administration. Half-maximum effect concentrations (IC₅₀) for inhibition of Pgp-mediated rhodamine 123 efflux from CCRFvcr1000 cells were 375 ± 60 nM for MC113 versus 8.5 ± 2.5 nM for tariquidar. Conclusion: [¹¹C]MC113 showed higher brain uptake in mice than previously described Pgp PET tracers, suggesting that [¹¹C]MC113 was only to a low extent effluxed by Pgp. However, [¹¹C]MC113 was found unsuitable to visualize Pgp expression levels presumably due to insufficiently high Pgp binding affinity of MC113 in relation to Pgp densities in brain and tumors.