High-dose chemotherapy with stem cell rescue to treat stage III homologous deficient breast cancer: factors influencing clinical implementation.

BACKGROUND: High-dose chemotherapy with autologous stem cell rescue (HDCT) is a promising treatment for patients with stage III, HER2-negative, homologous recombination deficient (HRD) breast cancer. Clinical effectiveness and cost-effectiveness are currently under investigation in an international multicenter randomized controlled trial. To increase the chance of successful introduction of HDCT into daily clinical practice, we aimed to identify relevant factors for smooth implementation using an early comprehensive assessment framework. METHODS: This is a qualitative, multi-stakeholder, exploratory research using semi-structured interviews guided by the Constructive Technology Assessment model, which evaluates the quality of a novel health technology by clinical, economic, patient-related, and organizational factors. Stakeholders were recruited by purposeful stratified sampling and interviewed until sufficient content saturation was reached. Two researchers independently created themes, categories, and subcategories by following inductive coding steps, these were verified by a third researcher. RESULTS: We interviewed 28 stakeholders between June 2019 and April 2021. In total, five overarching themes and seventeen categories were identified. Important findings for optimal implementation included the structural identification and referral of all eligible patients, early integration of supportive care, multidisciplinary collaboration between- and within hospitals, (de)centralization of treatment aspects, the provision of information for patients and healthcare professionals, and compliance to new regulation for the BRCA1-like test. CONCLUSIONS: In anticipation of a positive reimbursement decision, we recommend to take the highlighted implementation factors into consideration. This might expedite and guide high-quality equitable access to HDCT for patients with stage III, HER2-negative, HRD breast cancer in the Netherlands.

Cost-utility, cost-effectiveness, and budget impact of Internet-based cognitive behavioral therapy for breast cancer survivors with treatment-induced menopausal symptoms.

PURPOSE: Internet-based cognitive behavioral therapy (iCBT), with and without therapist support, is effective in reducing treatment-induced menopausal symptoms and perceived impact of hot flushes and night sweats (HF/NS) in breast cancer survivors. The aim of the current study was to evaluate the cost-utility, cost-effectiveness, and budget impact of both iCBT formats compared to a waiting list control group from the Dutch healthcare perspective. METHODS: A Markov model was constructed with a 5-year time horizon. Costs and health outcomes were measured alongside a randomized controlled clinical trial and included quality-adjusted life years (QALYs), overall levels of menopausal symptoms, and perceived impact of HF/NS. Uncertainty was examined using probabilistic and deterministic sensitivity analyses, together with a scenario analysis incorporating a different perspective. RESULTS: iCBT was slightly more expensive than the waiting list control, but also more effective, resulting in incremental cost-utility ratios of €23,331/QALY and €11,277/QALY for the guided and self-managed formats, respectively. A significant reduction in overall levels of menopausal symptoms or perceived impact of HF/NS resulted in incremental costs between €1460 and €1525 for the guided and €500-€753 for the self-managed format. The estimated annual budget impact for the Netherlands was €192,990 for the guided and €74,592 for the self-managed format. CONCLUSION: Based on the current trial data, the results indicate that both guided and self-managed iCBT are cost-effective with a willingness-to-pay threshold of well below €30,000/QALY. Additionally, self-managed iCBT is the most cost-effective strategy and has a lower impact on healthcare budgets.

SHARING AND COLLECTING HOSPITAL-BASED HEALTH TECHNOLOGY ASSESSMENT REPORTS INTERNATIONALLY: IS AN EXTENSIVE PARTICIPATION OF STAKEHOLDERS REALISTIC?

OBJECTIVES: The emergence of new medical technologies and budget restrictions has led to a substantial increase in the use of hospital-based health technology assessment (HB-HTA). This qualitative study explores whether there is a possibility and interest to realize the collection and dissemination of HB-HTA reports on an international scale by exploring the opinions from HB-HTA experts. METHODS: A survey was designed and sent to an international group of experts knowledgeable in HB-HTA from eighteen different countries. The survey contained questions about their opinions on the collection and distribution of HB-HTA information, and about the meaningful dimensions, barriers and values about a database. The data obtained were analyzed through the method of content analysis. RESULTS: A total of thirty-six experts (response rate of 18.3 percent) responded to the survey. The obtained data shows that all respondents agree that the collection of HB-HTA reports is useful. Moreover, 41.7 percent of respondents that are in the position of sharing HB-HTA reports (n = 24) mentioned that full HB-HTA reports can be shared. Many other respondents reported that confidentiality (45.7 percent) and investment into a database (40.0 percent) are important barriers for the dissemination of HB-HTA reports. CONCLUSIONS: There seem to be enough demand and willingness to share all or most reports by a large community of HB-HTA producers. Therefore, there is a need for a well-designed database with regular maintenance and complete, comparable, and qualitative HB-HTA reports. The database of the AdHopHTA project could potentially facilitate this process.

Early Cost-Effectiveness Analysis of Cytoreductive Surgery and Hyperthermic Intraperitoneal Chemotherapy for Gastric Cancer Patients with Limited Peritoneal Carcinomatosis.

BACKGROUND: Gastric cancer patients with peritoneal carcinomatosis (PC) have a poor prognosis, with a median overall survival of 10 months when treated with systemic chemotherapy only. Cohort studies showed that cytoreductive surgery with hyperthermic intraperitoneal chemotherapy (CRS/HIPEC) might improve the prognosis for gastric cancer patients with limited PC. Besides generating trial data on clinical effectiveness, it is crucial to timely collect information on economic aspects to guide the reimbursement decision-making process. No previous data have been published on the cost(-effectiveness) of CRS/HIPEC in this group of patients. Therefore, we performed an early model-based cost-effectiveness analysis of CRS/HIPEC for gastric cancer patients with limited PC in the Dutch setting. METHODS: We constructed a two-state (alive-dead) Markov transition model to evaluate costs and clinical outcomes from a Dutch healthcare perspective. Clinical outcomes, transition probabilities and utilities were derived from literature and verified by clinical experts in the field. Costs were measured using two available representative cohorts (2010-2017): one 'systemic chemotherapy only' cohort and one 'CRS/HIPEC' cohort (n = 10 each). Incremental cost-utility ratios (ICURs) were expressed as Euros per quality-adjusted life-year (QALY). We performed probabilistic and deterministic sensitivity, scenario, and value-of-information analyses using a willingness-to-pay (WTP) threshold of €80,000/QALY, which reflects the Dutch norm for severe diseases. RESULTS: In the base-case analysis, CRS/HIPEC yielded more QALYs (increment of 0.68) and more costs (increment of €34,706) compared with systemic chemotherapy only, resulting in an ICUR of €50,990/QALY. The probability that CRS/HIPEC was cost effective compared with systemic chemotherapy alone was 64%. To reduce uncertainty, the expected value of perfect information amounted to €4,021,468. The scenario analyses did not alter the results and showed that treatment costs, lifetime health-related quality of life and overall survival had the largest influence on the model. CONCLUSIONS: The presented early cost-effectiveness analysis suggests that adding CRS/HIPEC to systemic chemotherapy for gastric cancer patients with limited PC has a good chance of being cost-effectiveness compared with systemic chemotherapy alone when using a WTP of €80,000/QALY. However, there is substantial uncertainty in view of the current available data on effectiveness. Results from the ongoing phase III PERISCOPE II trial are therefore crucial for further decisions on treatment policy and its cost-effectiveness.

Binding characterization of N-(2-chloro-5-thiomethylphenyl)-N'-(3-[3 H]3 methoxy phenyl)-N'-methylguanidine ([3 H]GMOM), a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist.

Labeled with carbon-11, N-(2-chloro-5-thiomethylphenyl)-N'-(3-methoxyphenyl)-N'-methylguanidine ([11 C]GMOM) is currently the only positron emission tomography (PET) tracer that has shown selectivity for the ion-channel site of N-methyl-D-aspartate (NMDA) receptors in human imaging studies. The present study reports on the selectivity profile and in vitro binding properties of GMOM. The compound was screened on a panel of 80 targets, and labeled with tritium ([3 H]GMOM). The binding properties of [3 H]GMOM were compared to those of the reference ion-channel ligand [3 H](+)-dizocilpine maleate ([3 H]MK-801), in a set of concentration-response, homologous and heterologous inhibition, and association kinetics assays, performed with repeatedly washed rat forebrain preparations. GMOM was at least 70-fold more selective for NMDA receptors compared to all other targets examined. In homologous inhibition and concentration-response assays, the binding of [3 H]GMOM was regulated by NMDA receptor agonists, albeit in a less prominent manner compared to [3 H]MK-801. Scatchard transformation of homologous inhibition data produced concave upward curves for [3 H]GMOM and [3 H]MK-801. The radioligands showed bi-exponential association kinetics in the presence of 100 µmol L-1 l-glutamate/30 µmol L-1 glycine. [3 H]GMOM (3 nmol L-1 and 10 nmol L-1 ) was inhibited with dual affinity by (+)-MK-801, (R,S)-ketamine and memantine, in both presence and absence of agonists. [3 H]MK-801 (2 nmol L-1 ) was inhibited in a monophasic manner by GMOM under baseline and combined agonist conditions, with an IC50 value of ~19 nmol L-1 . The non-linear Scatchard plots, biphasic inhibition by open channel blockers, and bi-exponential kinetics of [3 H]GMOM indicate a complex mechanism of interaction with the NMDA receptor ionophore. The implications for quantifying the PET signal of [11 C]GMOM are discussed.

Synthesis and preliminary preclinical evaluation of fluorine-18 labelled isatin-4-(4-methoxyphenyl)-3-thiosemicarbazone ([18F]4FIMPTC) as a novel PET tracer of P-glycoprotein expression.

BACKGROUND: Several P-glycoprotein (P-gp) substrate tracers are available to assess P-gp function in vivo, but attempts to develop a tracer for measuring expression levels of P-gp have not been successful. Recently, (Z)-2-(5-fluoro-2-oxoindolin-3-ylidene)-N-(4-methoxyphenyl)hydrazine-carbothioamide was described as a potential selective P-gp inhibitor that is not transported by P-gp. Therefore, the purpose of this study was to radiolabel two of its analogues and to assess their potential for imaging P-gp expression using PET. RESULTS: [18F]2-(4-fluoro-2-oxoindolin-3-ylidene)-N-(4-methoxyphenyl)hydrazine-carbothioamide ([18F]5) and [18F]2-(6-fluoro-2-oxoindolin-3-ylidene)-N-(4-methoxyphenyl)hydrazine-carbothioamide ([18F]6) were synthesized and both their biodistribution and metabolism were evaluated in rats. In addition, PET scans were acquired in rats before and after tariquidar (P-gp inhibitor) administration as well as in P-gp knockout (KO) mice.Both [18F]5 and [18F]6 were synthesized in 2-3% overall yield, and showed high brain uptake in ex vivo biodistribution studies. [18F]6 appeared to be metabolically unstable in vivo, while [18F]5 showed moderate stability with limited uptake of radiolabelled metabolites in the brain. PET studies showed that transport of [18F]5 across the blood-brain barrier was not altered by pre-treatment with the P-gp inhibitor tariquidar, and uptake was significantly lower in P-gp KO than in wild-type animals and indeed transported across the BBB or bound to P-gp in endothelial cells. CONCLUSION: In conclusion, [18F]5 and [18F]6 were successfully and reproducibly synthesized, albeit with low radiochemical yields. [18F]5 appears to be a radiotracer that binds to P-gp, as showed in P-gp knock-out animals, but is not a substrate for P-gp.

Parametric Methods for Dynamic 11C-Phenytoin PET Studies.

In this study, the performance of various methods for generating quantitative parametric images of dynamic 11C-phenytoin PET studies was evaluated. Methods: Double-baseline 60-min dynamic 11C-phenytoin PET studies, including online arterial sampling, were acquired for 6 healthy subjects. Parametric images were generated using Logan plot analysis, a basis function method, and spectral analysis. Parametric distribution volume (VT) and influx rate (K1) were compared with those obtained from nonlinear regression analysis of time-activity curves. In addition, global and regional test-retest (TRT) variability was determined for parametric K1 and VT values. Results: Biases in VT observed with all parametric methods were less than 5%. For K1, spectral analysis showed a negative bias of 16%. The mean TRT variabilities of VT and K1 were less than 10% for all methods. Shortening the scan duration to 45 min provided similar VT and K1 with comparable TRT performance compared with 60-min data. Conclusion: Among the various parametric methods tested, the basis function method provided parametric VT and K1 values with the least bias compared with nonlinear regression data and showed TRT variabilities lower than 5%, also for smaller volume-of-interest sizes (i.e., higher noise levels) and shorter scan duration.

Radiosynthesis and biodistribution of a histamine H3 receptor antagonist 4-[3-(4-piperidin-1-yl-but-1-ynyl)-[11C]benzyl]-morpholine: evaluation of a potential PET ligand.

The potent histamine H(3) receptor antagonist JNJ-10181457 (1) was successfully labeled with (11)C in a novel one-pot reaction sequence, with high chemical yield (decay-corrected yield, 28+/-8%) and high specific radioactivity (56+/-26 GBq/mumol). The binding of [(11)C]1 to H(3) receptors was studied in vitro in rat brain and in vivo in rats and mice. The in vitro binding of [(11)C]1 in rat coronal brain slices showed high binding in the striatum, and this binding was blocked by histamine and by two known H(3) antagonists, JNJ-5207852 (2) and unlabeled Compound (1), in a concentration-dependent manner. The biodistribution of [(11)C]1 in rats was measured at 5, 10, 30 and 60 min. The uptake of [(11)C]1 in regions rich in H(3) receptors was highest at 30 min, giving 0.98%, 1.41%, 1.28% and 1.72% dose/g for the olfactory bulb, hippocampus, striatum and cerebral cortex, respectively. However, the binding of [(11)C]1 in the rat brain could not be blocked by pretreatment with either Compound (2) (30 min or 24 h pretreatment) or cold Compound (1) (30-min pretreatment). The biodistribution of [(11)C]1 in a second species (Balb/c mice) showed a higher overall uptake of the radioligand with an average brain uptake of 8.9% dose/g. In C57BL/6-H(3)(-/-) knockout mice, a higher brain uptake was also observed. Analyses of metabolites and plasma protein binding were also undertaken. It appeared that [(11)C]1 could not specifically label H(3) receptors in rodent brain in vivo. Possible causes are discussed.

Molecular imaging of aurora kinase A (AURKA) expression: Synthesis and preclinical evaluation of radiolabeled alisertib (MLN8237).

INTRODUCTION: Survival of patients after resection of colorectal cancer liver metastasis (CRCLM) is 36%-58%. Positron emission tomography (PET) tracers, imaging the expression of prognostic biomarkers, may contribute to assign appropriate management to individual patients. Aurora kinase A (AURKA) expression is associated with survival of patients after CRCLM resection. METHODS: We synthesized [(3)H]alisertib and [(11)C]alisertib, starting from [(3)H]methyl nosylate and [(11)C]methyl iodide, respectively. We measured in vitro uptake of [(3)H]alisertib in cancer cells with high (Caco2), moderate (A431, HCT116, SW480) and low (MKN45) AURKA expression, before and after siRNA-mediated AURKA downmodulation, as well as after inhibition of P-glycoprotein (P-gp) activity. We measured in vivo uptake and biodistribution of [(11)C]alisertib in nude mice, xenografted with A431, HCT116 or MKN45 cells, or P-gp knockout mice. RESULTS: [(3)H]Alisertib was synthesized with an overall yield of 42% and [(11)C]alisertib with an overall yield of 23%±9% (radiochemical purity ≥99%). Uptake of [(3)H]alisertib in Caco2 cells was higher than in A431 cells (P=.02) and higher than in SW480, HCT116 and MKN45 cells (P<.01). Uptake in A431 cells was higher than in SW480, HCT116 and MKN45 cells (P<.01). Downmodulation of AURKA expression reduced [(3)H]alisertib uptake in Caco2 cells (P<.01). P-gp inhibition increased [(3)H]alisertib uptake in Caco2 (P<.01) and MKN45 (P<.01) cells. In vivo stability of [(11)C]alisertib 90min post-injection was 94.7%±1.3% and tumor-to-background ratios were 2.3±0.8 (A431), 1.6±0.5 (HCT116) and 1.9±0.5 (MKN45). In brains of P-gp knockout mice [(11)C]alisertib uptake was increased compared to uptake in wild-type mice (P<.01) CONCLUSIONS: Radiolabeled alisertib can be synthesized and may have potential for the imaging of AURKA, particularly when AURKA expression is high. However, the exact mechanisms underlying alisertib accumulation need further investigation. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: Radiolabeled alisertib may be used for non-invasively measuring AURKA protein expression and to stratify patients for treatment accordingly.

Quantification of Dynamic 11C-Phenytoin PET Studies.

UNLABELLED: The overexpression of P-glycoprotein (Pgp) is thought to be an important mechanism of pharmacoresistance in epilepsy. Recently, (11)C-phenytoin has been evaluated preclinically as a tracer for Pgp. The aim of the present study was to assess the optimal plasma kinetic model for quantification of (11)C-phenytoin studies in humans. METHODS: Dynamic (11)C-phenytoin PET scans of 6 healthy volunteers with arterial sampling were acquired twice on the same day and analyzed using single- and 2-tissue-compartment models with and without a blood volume parameter. Global and regional test-retest (TRT) variability was determined for both plasma to tissue rate constant (K1) and volume of distribution (VT). RESULTS: According to the Akaike information criterion, the reversible single-tissue-compartment model with blood volume parameter was the preferred plasma input model. Mean TRT variability ranged from 1.5% to 16.9% for K1 and from 0.5% to 5.8% for VT. Larger volumes of interest showed better repeatabilities than smaller regions. A 45-min scan provided essentially the same K1 and VT values as a 60-min scan. CONCLUSION: A reversible single-tissue-compartment model with blood volume seems to be a good candidate model for quantification of dynamic (11)C-phenytoin studies. Scan duration may be reduced to 45 min without notable loss of accuracy and precision of both K1 and VT, although this still needs to be confirmed under pathologic conditions.

Synthesis and evaluation of radiolabeled analogs of the antidepressant drug zimelidine as potential SPECT-ligands for the serotonin transporter.

Z-3-(4-bromophenyl)-N,N-dimethyl-3-(3-pyridinyl)-2-propen-1-amine or zimelidine (ZIM) and its first metabolite nor-zimelidine, were radioiodinated via a nonisotopic exchange, using the Cu(I)-assisted nucleophilic labeling method. To evaluate their potential as SPECT ligands for the serotonin transporter (SERT), the biodistribution of both ligands was determined and pretreatment "blocking" studies performed. Both radioligands demonstrated a good brain penetration of 0.8-1% ID/g, stable after 60 min., p.i., and a brain/blood ratio of up to 3. In vivo brain distribution did not reveal specific binding. Blocking studies by pretreatment with a known SERT ligand, had minor influence on the uptake of [(123)I]I-ZIM, between the several isolated brain regions. It may therefore be concluded that [(123)I]I-ZIM and [(123)I]I-nor-ZIM do not appear to be promising SPECT ligands for the SERT.

The binding characteristics and orientation of a novel radioligand with distinct properties at 5-HT3A and 5-HT3AB receptors.

VUF10166 (2-chloro-3-(4-methyl piperazin-1-yl)quinoxaline) is a ligand that binds with high affinity to 5-HT3 receptors. Here we synthesise [(3)H]VUF10166 and characterise its binding properties at 5-HT3A and 5-HT3AB receptors. At 5-HT3A receptors [(3)H]VUF10166 displayed saturable binding with a Kd of 0.18 nM. Kinetic measurements gave monophasic association (6.25 × 10(7) M(-1) min(-1)) and dissociation (0.01 min(-1)) rates that yielded a similar Kd value (0.16 nM). At 5-HT3AB receptors two association (6.15 × 10(-7), 7.23 M(-1) min(-1)) and dissociation (0.024, 0.162 min(-1)) rates were seen, yielding Kd values (0.38 nM and 22 nM) that were consistent with values obtained in saturation (Kd = 0.74 nM) and competition (Ki = 37 nM) binding experiments respectively. At both receptor types, specific binding was inhibited by classical 5-HT3 receptor-selective orthosteric ligands (5-HT, allosetron, d-tubocurarine, granisetron, mCPBG, MDL72222, quipazine), but not by non-competitive antagonists (bilobalide, ginkgolide B, picrotoxin) or competitive ligands of other Cys-loop receptors (ACh, bicuculline, glycine, gabazine). To explore VUF10166 ligand-receptor interactions we used in silico modelling and docking, and tested the predictions using site directed mutagenesis. The data suggest that VUF10166 adopts a similar orientation to 5-HT3 receptor agonists bound in AChBP (varenicline) and 5HTBP (5-HT) crystal structures.

Development and characterization of [123I]iodotiagabine for in-vivo GABA-transporter imaging.

The increased knowledge of molecular changes associated with different neurological disorders calls for the development of novel radioligands. Tiagabine (Gabitril) is an anticonvulsive drug that binds selectively to GABA transporter-1 and thereby inhibits GABA uptake. As radioligands for in-vivo imaging of the GABA transporter are not yet available, we radiolabelled tiagabine and assessed its efficacy for in-vivo imaging of these transporters. Tiagabine was first brominated at its vinylic part, which was then exchanged with I. Next, anaesthetized rats received a bolus injection of [I]iodotiagabine in their tail vein, which was immediately followed by acquisition of planar and high-resolution micro-single-photon emission computed tomography (SPECT) images of the total body with special focus on the brain. Uptake in anatomical regions was assessed by coregistration of micro-SPECT with micro-CT images. Tiagabine labelling with I resulted in 50% yield and 99.7% radiochemical purity. Within 3 h after injection, SPECT demonstrated an increased signal-to-background ratio in the nasal mucosa and/or the Harderian glands but not in the brain. In addition we observed an increased signal-to-background ratio in organs such as the thyroid, heart, liver, kidney and bladder. More than 99% pure I-labelled tiagabine can be obtained and applied in animal micro-SPECT studies. However, this new radioligand is not taken up sufficiently by the brain and therefore cannot be used to successfully detect cerebral GABA transporters.

[11C]quinidine and [11C]laniquidar PET imaging in a chronic rodent epilepsy model: impact of epilepsy and drug-responsiveness.

INTRODUCTION: To analyse the impact of both epilepsy and pharmacological modulation of P-glycoprotein on brain uptake and kinetics of positron emission tomography (PET) radiotracers [(11)C]quinidine and [(11)C]laniquidar. METHODS: Metabolism and brain kinetics of both [(11)C]quinidine and [(11)C]laniquidar were assessed in naive rats, electrode-implanted control rats, and rats with spontaneous recurrent seizures. The latter group was further classified according to their response to the antiepileptic drug phenobarbital into "responders" and "non-responders". Additional experiments were performed following pre-treatment with the P-glycoprotein modulator tariquidar. RESULTS: [(11)C]quinidine was metabolized rapidly, whereas [(11)C]laniquidar was more stable. Brain concentrations of both radiotracers remained at relatively low levels at baseline conditions. Tariquidar pre-treatment resulted in significant increases of [(11)C]quinidine and [(11)C]laniquidar brain concentrations. In the epileptic subgroup "non-responders", brain uptake of [(11)C]quinidine in selected brain regions reached higher levels than in electrode-implanted control rats. However, the relative response to tariquidar did not differ between groups with full blockade of P-glycoprotein by 15 mg/kg of tariquidar. For [(11)C]laniquidar differences between epileptic and control animals were only evident at baseline conditions but not after tariquidar pretreatment. CONCLUSIONS: We confirmed that both [(11)C]quinidine and [(11)C]laniquidar are P-glycoprotein substrates. At full P-gp blockade, tariquidar pre-treatment only demonstrated slight differences for [(11)C]quinidine between drug-resistant and drug-sensitive animals.

Synthesis and preclinical evaluation of [11C]D617, a metabolite of (R)-[11C]verapamil.

OBJECTIVES: (R)-[(11)C]verapamil is widely used as a positron emission tomography (PET) tracer to evaluate P-glycoprotein (P-gp) functionality at the blood-brain barrier in man. A disadvantage of (R)-[(11)C]verapamil is the fact that its main metabolite, [(11)C]D617, also enters the brain. For quantitative analysis of (R)-[(11)C]verapamil data, it has been assumed that the cerebral kinetics of (R)-[(11)C]verapamil and [(11)C]D617 are the same. The aim of the present study was to investigate whether the cerebral kinetics of (R)-[(11)C]verapamil and [(11)C]D617 are indeed similar and, if so, whether [(11)C]D617 itself could serve as an alternative PET tracer for P-gp. METHODS: [(11)C]D617 was synthesized and its ex vivo biodistribution was investigated in male rats at four time points following intravenous administration of [(11)C]D617 (50 MBq) without (n=4) or with (n=4) pretreatment with the P-gp inhibitor tariquidar (15 mg·kg(-1), intraperitoneally). Brain distribution was further assessed using consecutive PET scans (n=8) before and after pretreatment with tariquidar (15 mg·kg(-1), intravenously), as well as metabolite analysis (n=4). RESULTS: The precursor for the radiosynthesis of [(11)C]D617, 5-amino-2-(3,4-dimethoxy-phenyl)-2-isopropyl-pentanitrile (desmethyl D617), was synthesized in 41% overall yield. [(11)C]D617 was synthesized in 58%-77% decay-corrected yield with a radiochemical purity of ≥99%. The homogeneously distributed cerebral volume of distribution (V(T)) of [(11)C]D617 was 1.1, and this increased 2.4-fold after tariquidar pretreatment. CONCLUSION: V(T) of [(11)C]D617 was comparable to that of (R)-[(11)C]verapamil, but its increase after tariquidar pretreatment was substantially lower. Hence, (R)-[(11)C]verapamil and [(11)C]D617 do not show similar brain kinetics after inhibition of P-gp with tariquidar.

Synthesis, biodistribution and PET studies in rats of (18)F-Labeled bridgehead fluoromethyl analogues of WAY-100635.

INTRODUCTION: In vitro screening of fluoromethyl bridge-fused ring (BFR) analogues of WAY-100635 (5a, 5b and 5c) has shown a high binding affinity and a good selectivity for the 5-HT(1A) receptor. As these compounds were designed to provide PET ligands with high metabolic stability, they are now radiolabeled with fluorine-18 and investigated in vivo. METHODS: BFR precursors were synthesized and reacted with fluorine-18 in dry MeCN in the presence of 2,2,2-kryptofix and K(2)CO(3). In rats, biodistribution and PET studies were performed using [(18)F]5a, [(18)F]5b and [(18)F]5c. The binding specificity was determined by administration of non-labeled WAY-100635 prior to the radiolabeled ligands. RESULTS: [(18)F]5 ligands were synthesized in overall radiochemical yields of 24%-45%, respectively with a radiochemical purity of >98%. Relatively good hippocampus to cerebellum ratios of 5.55, 4.79 and 5.45, respectively were reached at 45 min pi. However, PET studies indicated defluorination of the radioligands by showing high accumulation of radioactivity in the bones in the order of [(18)F]5a≈[(18)F]5b>[(18)F]5c. CONCLUSION: Also in vivo, the radioligands bind preferentially to the 5-HT(1A) receptor. Unfortunately, no metabolic stability with regard to defluorination was observed in rats.

[11C]phenytoin revisited: synthesis by [11C]CO carbonylation and first evaluation as a P-gp tracer in rats.

BACKGROUND: At present, several positron emission tomography (PET) tracers are in use for imaging P-glycoprotein (P-gp) function in man. At baseline, substrate tracers such as R-[11C]verapamil display low brain concentrations with a distribution volume of around 1. [11C]phenytoin is supposed to be a weaker P-gp substrate, which may lead to higher brain concentrations at baseline. This could facilitate assessment of P-gp function when P-gp is upregulated. The purpose of this study was to synthesize [11C]phenytoin and to characterize its properties as a P-gp tracer. METHODS: [11C]CO was used to synthesize [11C]phenytoin by rhodium-mediated carbonylation. Metabolism and, using PET, brain pharmacokinetics of [11C]phenytoin were studied in rats. Effects of P-gp function on [11C]phenytoin uptake were assessed using predosing with tariquidar. RESULTS: [11C]phenytoin was synthesized via [11C]CO in an overall decay-corrected yield of 22 ± 4%. At 45 min after administration, 19% and 83% of radioactivity represented intact [11C]phenytoin in the plasma and brain, respectively. Compared with baseline, tariquidar predosing resulted in a 45% increase in the cerebral distribution volume of [11C]phenytoin. CONCLUSIONS: Using [11C]CO, the radiosynthesis of [11C]phenytoin could be improved. [11C]phenytoin appeared to be a rather weak P-gp substrate.

Design, synthesis and in vitro evaluation of bridgehead fluoromethyl analogs of N-{2-[4-(2-methoxyphenyl)piperazin-1-yl]ethyl}-N-(pyridin-2-yl)cyclohexanecarboxamide (WAY-100635) for the 5-HT(1A) receptor.

Fluorinated analogs that are related to the 5-hydroxytryptamine (5-HT(1A)) antagonist, N-{2-[4-(2-methoxyphenyl)piperazin-1-yl]ethyl}-N-(pyridin-2-yl)cyclohexanecarboxamide (WAY-100635), have been synthesized and their binding affinity for the 5-HT(1A) receptor and other neurotransmitter receptors (adrenoceptors, sigma receptors, and dopamine receptors), and serotonin transporters was examined in vitro. These ligands were designed to provide a possible potential positron emission tomography (PET) ligand with high metabolic stability. To this end, the cyclohexyl moiety in WAY-100635 and in O-desmethyl WAY-100635 was replaced by a bridge-fused ring (BFR) such as adamantyl, cubyl, bicyclo[2.2.2]octyl and bicyclo[2.2.1]heptyl to reduce the metabolic rate of the amide bond hydrolysis, while a fluoromethyl group was introduced on the other bridgehead of the BFR to prevent defluorination by HF elimination. All synthesized analogs displayed high affinity in the (sub)nanomolar range for the 5-HT(1A) receptor, comparable to WAY-100635. In addition, 6b, 6c and 6d were reasonably selective to the 5-HT(1A) receptor over the above mentioned receptors. In human hepatocytes, 6b showed a suitable metabolic stability. In conclusion, the obtained data provides a promising starting point for the synthesis of the corresponding (18)F-labeled PET analogs.

Design, synthesis, radiolabeling, and in vitro and in vivo evaluation of bridgehead iodinated analogues of N-{2-[4-(2-methoxyphenyl)piperazin-1-yl]ethyl}-N-(pyridin-2-yl)cyclohexanecarboxamide (WAY-100635) as potential SPECT ligands for the 5-HT1A receptor.

Here we describe the design, synthesis, and pharmacological profile of 5-HT(1A) receptor ligands related to 1 (WAY-100635). The cyclohexyl moiety in 1 and its O-desmethylated analogue 3 were replaced by the bridgehead iodinated bridge-fused rings: adamantyl, cubyl, bicyclo[2.2.2]octyl, or bicyclo[2.2.1]heptyl. All analogues displayed a (sub)nanomolar affinity for the 5-HT(1A) receptor in vitro. Compounds 6b and 7b appeared to be selective for this receptor over other relevant receptors and could easily be iodinated with radioactive iodine-123. In humane hepatocytes, [(123)I]6b showed a low propensity for amide hydrolysis and a stable carbon-iodine bond. The biodistribution of [(123)I]6b and [(123)I]7b in rats revealed that the carbon-iodine bond was also stable in vivo. Unfortunately, the brain uptake and the specificity for both radioligands were significantly lower than those of the parent molecule 1. In conclusion, the designed tracers are not suitable for SPECT imaging.

Evaluation of [11C]laniquidar as a tracer of P-glycoprotein: radiosynthesis and biodistribution in rats.

At present, P-glycoprotein (P-gp) function can be studied using positron emission tomography (PET) together with a labelled P-gp substrate such as R-[11C]verapamil. Such a tracer is, however, less suitable for investigating P-gp (over)expression. Laniquidar is a third-generation P-gp inhibitor, which has been used in clinic trials for modulating multidrug resistance transporters. The purpose of the present study was to develop the radiosynthesis of [11C]laniquidar and to assess its suitability as a tracer of P-gp expression. The radiosynthesis of [11C]laniquidar was performed by methylation of the carboxylic acid precursor with [11C]CH3I. The product was purified by HPLC and reformulated over a tC18 Seppak, yielding a sterile solution of [11C]laniquidar in saline. For evaluating [11C]laniquidar, rats were injected with 20 MBq [11C]laniquidar via a tail vein and sacrificed at 5, 15, 30 and 60 min after injection. Several tissues and distinct brain regions were dissected and counted for radioactivity. In addition, uptake of [11C]laniquidar in rats pretreated with cyclosporine A and valspodar (PSC 833) was determined at 30 min after injection. Finally, the metabolic profile of [11C]laniquidar in plasma was determined. [11C]Laniquidar could be synthesized in moderate yields with high specific activity. Uptake in brain was low, but significantly increased after administration of cyclosporine A. Valspodar did not have any effect on cerebral uptake of [11C]laniquidar. In vivo rate of metabolism was relatively low. Further kinetic studies are needed to investigate the antagonistic behaviour of [11C]laniquidar at tracer level.