Clinical translation of 18F-fluoropivalate - a PET tracer for imaging short-chain fatty acid metabolism: safety, biodistribution, and dosimetry in fed and fasted healthy volunteers.

BACKGROUND: Fatty acids derived de novo or taken up from the extracellular space are an essential source of nutrient for cell growth and proliferation. Radiopharmaceuticals including 11C-acetate, and 18F-FAC (2-18F-fluoroacetate), have previously been used to study short-chain fatty acid (SCFA) metabolism. We developed 18F-fluoropivalate (18F-FPIA; 3-18F-fluoro-2,2-dimethylpropionic acid) bearing a gem-dimethyl substituent to assert metabolic stability for studying SCFA metabolism. We report the safety, biodistribution, and internal radiation dosimetry profile of 18F-FPIA in 24 healthy volunteers and the effect of dietary conditions. MATERIALS AND METHODS: Healthy volunteer male and female subjects were enrolled (n = 24), and grouped into 12 fed and 12 fasted. Non-esterified fatty acids (NEFA) and carnitine blood measurements were assessed. Subjects received 159.48 MBq (range, 47.31-164.66 MBq) of 18F-FPIA. Radiochemical purity was > 99%. Safety data were obtained during and 24 h after radiotracer administration. Subjects underwent detailed multiple whole-body PET/CT scanning with sampling of venous bloods for radioactivity and radioactive metabolite quantification. Regions of interest were defined to derive individual and mean organ residence times; effective dose was calculated using OLINDA 1.1. RESULTS: All subjects tolerated 18F-FPIA with no adverse events. Over 90% of radiotracer was present in plasma at 60 min post-injection. The organs receiving highest absorbed dose (in mGy/MBq) were the liver (0.070 ± 0.023), kidneys (0.043 ± 0.013), gallbladder wall (0.026 ± 0.003), and urinary bladder (0.021 ± 0.004); otherwise there was low tissue uptake. The calculated effective dose using mean organ residence times over all 24 subjects was 0.0154 mSv/MBq (SD ± 0.0010). No differences in biodistribution or dosimetry were seen in fed and fasted subjects, though systemic NEFA and carnitine levels reflected fasted and fed states. CONCLUSION: The favourable safety, imaging, and dosimetric profile makes 18F-FPIA a promising candidate radiotracer for tracing SCFA metabolism.

Use of carbon-11 labelled tool compounds in support of drug development.

The pharmaceutical industry is facing key challenges to improve return on R&D investment. Positron emission tomography (PET), by itself or in combination with complementary technologies such as magnetic resonance imaging (MRI), provides a unique opportunity to confirm a candidate's ability to meet the so-called 'three pillars' of drug development. Positive confirmation provides confidence for go/no-go decision making at an early stage of the development process and enables informed clinical progression. Whereas fluorine-18 has probably gained wider use in the community, there are benefits to using carbon-11 given the greater flexibility the use of this isotope permits in adaptive clinical study design. This review explores the scope of available carbon-11 chemistries and provides clinical examples to highlight its value in PET studies in support of drug development.

A general copper-mediated nucleophilic 18F†fluorination of arenes.

Molecules labeled with fluorine-18 are used as radiotracers for positron emission tomography. An important challenge is the labeling of arenes not amenable to aromatic nucleophilic substitution (SNAr) with [(18)F]F(-). In the ideal case, the (18)F fluorination of these substrates would be performed through reaction of [(18)F]KF with shelf-stable readily available precursors using a broadly applicable method suitable for automation. Herein, we describe the realization of these requirements with the production of (18)F arenes from pinacol-derived aryl boronic esters (arylBPin) upon treatment with [(18)F]KF/K222 and [Cu(OTf)2(py)4] (OTf = trifluoromethanesulfonate, py = pyridine). This method tolerates electron-poor and electron-rich arenes and various functional groups, and allows access to 6-[(18)F]fluoro-L-DOPA, 6-[(18)F]fluoro-m-tyrosine, and the translocator protein (TSPO) PET ligand [(18)F]DAA1106.

Evaluation and Application of a PET Tracer in Preclinical and Phase 1 Studies to Determine the Brain Biodistribution of Minzasolmin (UCB0599).

PURPOSE: Minzasolmin (UCB0599) is an orally administered, small molecule inhibitor of ASYN misfolding in development as a potential disease-modifying therapy for Parkinson's disease. Here we describe the preclinical development of a radiolabeled tracer and results from a phase 1 study using the tracer to investigate the brain distribution of minzasolmin. PROCEDURES: In the preclinical study, two radiolabeling positions were investigated on the S-enantiomer of minzasolmin (UCB2713): [11C]methylamine UCB2713 ([11C-N-CH3]UCB2713) and [11C]carbonyl UCB2713 ([11C-CO]UCB2713). Male C57 black 6 mice (N = 10) received intravenous [11C-N-CH3]UCB2713; brain homogenates were assessed for radioactivity and plasma samples analyzed by high-performance liquid chromatography. Positron emission tomography-computed tomography (PET-CT) was used to image brains in a subset of mice (n = 3). In the open-label, phase 1 study, healthy volunteers were scanned twice with PET-CT following injection with [11C]minzasolmin radiotracer (≤ 10 µg), first without, then with oral dosing with non-radiolabeled minzasolmin 360 mg. PRIMARY OBJECTIVE: to determine biodistribution of minzasolmin in the human brain; secondary objectives included minzasolmin safety/tolerability. RESULTS: Preclinical data supported the use of [11C]minzasolmin in clinical studies. In the phase 1 study, PET data showed substantial drug signal in the brain of healthy volunteers (N = 4). The mean estimated whole brain total distribution volume (VT) at equilibrium across all regions of interest was 0.512 mL/cm3, no difference in VT was observed following administration of minzasolmin 360 mg. Treatment-emergent adverse events (TEAEs) were reported by 75% (n = 3) of participants. No drug-related TEAEs, deaths, serious adverse events, or discontinuations were reported. CONCLUSION: Following positive preclinical results with the N-methyl labeled PET tracer, [11C]minzasolmin was used in the phase 1 study, which demonstrated that minzasolmin readily crossed the blood-brain barrier and was well distributed throughout the brain. Safety and pharmacokinetic findings were consistent with previous early-phase studies (such as UP0077, NCT04875962).

Use of LC-MS for the quality control of radiopharmaceuticals: example of [(18)F]ML10.

[(18)F]ML10 is a promising novel low molecular weight positron emission tomography probe for apoptosis. As part of the quality control to support clinical studies for cancer therapy monitoring in the GSK Clinical Imaging Centre, a simple and sensitive liquid chromatography mass spectrometry method has been developed and validated for the quantification of total ML10 and impurity content in the final product. Chromatographic separation of ML10 and its radiolabelling precursor and impurities was achieved. Mass curves were constructed from a concentration range of ML10 and known impurities and were linear. Quantification was achieved by comparison of the area under the curve for ML10 content (m/z = 205) and the mass curve. The method was validated over a concentration range of 0.1-1 µg/ml.

Widespread cell stress and mitochondrial dysfunction occur in patients with early Alzheimer's disease.

Cell stress and impaired oxidative phosphorylation are central to mechanisms of synaptic loss and neurodegeneration in the cellular pathology of Alzheimer's disease (AD). In this study, we quantified the in vivo expression of the endoplasmic reticulum stress marker, sigma 1 receptor (S1R), using [11C]SA4503 positron emission tomography (PET), the mitochondrial complex I (MC1) with [18F]BCPP-EF, and the presynaptic vesicular protein SV2A with [11C]UCB-J in 12 patients with early AD and in 16 cognitively normal controls. We integrated these molecular measures with assessments of regional brain volumes and cerebral blood flow (CBF) measured with magnetic resonance imaging arterial spin labeling. Eight patients with AD were followed longitudinally to estimate the rate of change of the physiological and structural pathology markers with disease progression. The patients showed widespread increases in S1R (≤ 27%) and regional reduction in MC1 (≥ -28%) and SV2A (≥ -25%) radioligand binding, brain volume (≥ -23%), and CBF (≥ -26%). [18F]BCPP-EF PET MC1 binding (≥ -12%) and brain volumes (≥ -5%) showed progressive reductions over 12 to 18 months, suggesting that they both could be used as pharmacodynamic indicators in early-stage therapeutics trials. Associations of reduced MC1 and SV2A and increased S1R radioligand binding with reduced cognitive performance in AD, although exploratory, suggested a loss of metabolic functional reserve with disease. Our study thus provides in vivo evidence for widespread, clinically relevant cellular stress and bioenergetic abnormalities in early AD.

Test-retest variability and reference region-based quantification of 18F-BCPP-EF for imaging mitochondrial complex I in the human brain.

Mitochondrial complex I (MC-I) is an essential regulator of brain bioenergetics and can be quantified in the brain using PET radioligand 18F-BCPP-EF. Here we evaluate the test-retest reproducibility of 18F-BCPP-EF in humans, and assess the use of a non-invasive quantification method (standardised uptake value ratio - SUVR). Thirty healthy volunteers had a 90-min dynamic 18F-BCPP-EF scan with arterial blood sampling, five of which received a second scan to be included in the test-retest analysis. Time-activity curves (TAC) were analysed using multilinear analysis 1 (MA1) and the two-tissue compartment model (2TC) to estimate volumes of distribution (VT). Regional SUVR-1 values were calculated from the 70 to 90-min TAC data using the centrum semiovale as a pseudo reference region, and compared to kinetic analysis-derived outcome measures. The mean absolute test-retest variability of VT ranged from 12% to 18% across regions. Both DVR-1and SUVR-1 had improved test-retest variability in the range 2%-7%. SUVR-1 was highly correlated with DVR-1 (r2 = 0.97, n = 30). In conclusion, 18F-BCPP-EF has suitable test-retest reproducibility and can be used to quantify MC-I in clinical studies.

Characterization of 3 PET Tracers for Quantification of Mitochondrial and Synaptic Function in Healthy Human Brain: 18F-BCPP-EF, 11C-SA-4503, and 11C-UCB-J.

Mitochondrial complex 1 is involved in maintaining brain bioenergetics; σ-1 receptor responds to neuronal stress; and synaptic vesicle protein 2A reflects synaptic integrity. Expression of each of these proteins is altered in neurodegenerative diseases. Here, we characterize the kinetic behavior of 3 PET radioligands-18F-BCPP-EF, 11C-SA-4503, and 11C-UCB-J-for the measurement of mitochondrial complex 1, σ-1 receptor, and synaptic vesicle protein 2A, respectively, and determine appropriate analysis workflows for their application in future studies of the in vivo molecular pathology of these diseases. Methods: Twelve human subjects underwent dynamic PET scans with each radioligand, including associated arterial blood sampling. A range of kinetic models was investigated to identify an optimal kinetic analysis method for each radioligand and a suitable acquisition duration. Results: All 3 radioligands readily entered the brain and yielded heterogeneous uptake consistent with the known distribution of the targets. The optimal models determined for the regional estimates of volume of distribution were multilinear analysis 1 (MA1) and the 2-tissue-compartment model for 18F-BCPP-EF, MA1 for 11C-SA-4503, and both MA1 and the 1-tissue-compartment model for 11C-UCB-J. Acquisition times of 70, 80, and 60 min for 18F-BCPP-EF, 11C-SA-4503, 11C-UCB-J, respectively, provided good estimates of regional volume of distribution values. An effect of age was observed on 18F-BCPP-EF and 11C-UCB-J signal in the caudate. Conclusion: These ligands can be assessed for their potential to stratify patients or monitor the progression of molecular neuropathology in neurodegenerative diseases.

Serotonin release measured in the human brain: a PET study with [11C]CIMBI-36 and d-amphetamine challenge.

Positron emission tomography (PET) enables non-invasive estimation of neurotransmitter fluctuations in the living human brain. While these methods have been applied to dopamine and some other transmitters, estimation of 5-hydroxytryptamine (5-HT; Serotonin) release has proved to be challenging. Here we demonstrate the utility of the novel 5-HT2A receptor agonist radioligand, [11C]CIMBI-36, and a d-amphetamine challenge to evaluate synaptic 5-HT changes in the living human brain. Seventeen healthy male volunteers received [11C]CIMBI-36 PET scans before and 3 h after an oral dose of d-amphetamine (0.5 mg/kg). Dynamic PET data were acquired over 90 min, and the total volume of distribution (VT) in the frontal cortex and the cerebellum derived from a kinetic analysis using MA1. The frontal cortex binding potential (BPNDfrontal) was calculated as (VTfrontal/VTcerebellum) - 1. ∆BPNDfrontal = 1 - (BPNDfrontal post-dose/BPNDfrontal baseline) was used as an index of 5-HT release. Statistical inference was tested by means of a paired Students t-test evaluating a reduction in post-amphetamine [11C]CIMBI-36 BPNDfrontal. Following d-amphetamine administration, [11C]CIMBI-36 BPNDfrontal was reduced by 14 ± 13% (p = 0.002). Similar effects were observed in other cortical regions examined in an exploratory analysis. [11C]CIMBI-36 binding is sensitive to synaptic serotonin release in the human brain, and when combined with a d-amphetamine challenge, the evaluation of the human brain serotonin system in neuropsychiatric disorders, such as major depression and Parkinson's disease is enabled.

In vivo quantification of regional dopamine-D3 receptor binding potential of (+)-PHNO: Studies in non-human primates and transgenic mice.

Examination of dopamine-D3 (D3) receptors with positron emission tomography (PET) have been hampered in the past by the lack of a PET ligand with sufficient selectivity for D3 over dopamine-D2 (D2) receptors. The two types co-localize in the brain, with D2 density significantly higher than D3, hence nonselective PET ligands inform on D2, rather than D3 status. [(11)C]-(+)-PHNO is a novel PET ligand with a preferential affinity for D3 over D2. We used the selective D3 antagonist, SB-277011 to dissect regional fractions of the [(11)C]-(+)-PHNO signal attributable to D3 and D2 in primate brain. The results were compared with quantitative autoradiography with (3)H-(+)-PHNO in wild-type, D2-knock-out, and D3-knock-out mice examined at baseline and following administration of SB-277011. Both sets of results converged to indicate a predominant D3-related component to (+)-PHNO binding in extra-striatal regions, with binding in the midbrain being entirely attributable to D3. The midbrain is thus an excellent target region to examine D3 receptor occupancy with [(11)C]-(+)-PHNO PET in vivo.

Investigation of the Brain Biodistribution of the Lipoprotein-Associated Phospholipase A2 (Lp-PLA2) Inhibitor [18F]GSK2647544 in Healthy Male Subjects.

PURPOSE: GSK2647544 is a potent and specific inhibitor of lipoprotein-associated phospholipase A2 (Lp-PLA2), which was in development as a potential treatment for Alzheimer's disease (AD). In order to refine therapeutic dose predictions and confirm brain penetration, a radiolabelled form of the inhibitor, [18F]GSK2647544, was manufactured for use in a positron emission tomography (PET) biodistribution study. PROCEDURES: [18F]GSK2647544 was produced using a novel, copper iodide (Cu(I)) mediated, [18F]trifluoromethylation methodology. Healthy male subjects (n = 4, age range 34-42) received an oral dose of unlabelled GSK2647544 (100 mg) and after 2 h an intravenous (iv) injection of [18F]GSK2647544 (average injected activity and mass were 106 ± 47 MBq and 179 ± 55 µg, respectively) followed by dynamic PET scans for 120 min. Defined regions of interest (ROI) throughout the brain were used to obtain regional time-activity curves (TACs) and compartmental modelling analysis used to estimate the primary outcome measure, whole brain volume of distribution (VT). Secondary PK and safety endpoints were also recorded. RESULTS: PET dynamic data were successfully obtained from all four subjects and there were no clinically significant variations of the safety endpoints. Inspection of the TACs indicated a relatively homogenous uptake of [18F]GSK2647544 across all the ROIs examined. The mean whole brain VT was 0.56 (95 % CI, 0.41-0.72). Secondary PK parameters, Cmax (geometric mean) and Tmax (median), were 354 ng/ml and 1.4 h, respectively. Metabolism of GSK2647544 was relatively consistent across subjects, with 20-40 % of the parent compound [18F]GSK2647544 present after 120 min. CONCLUSIONS: The study provides evidence that GSK2647544 is able to cross the blood brain barrier in healthy male subjects leading to a measurable brain exposure. The administered doses of GSK2647544 were well tolerated. Exploratory modelling suggested that a twice-daily dose of 102 mg, at steady state, would provide ~80 % trough inhibition of brain Lp-PLA2 activity. TRIAL REGISTRATION: Clintrials.gov: NCT01924858.

Methyl transfer reaction from monomethyltin reagent under palladium(0) catalysis: a versatile method for labelling with carbon-11.

The 11C-monomethylstannate prepared from [11C]-methyl iodide and Lappert's stannylene, was subject to a palladium-mediated cross-coupling reaction with an aryl halide under ligand-free conditions, to afford easily purified 11C-methyl(hetero)arenes in high radiochemical yields.

Boron-mediated polymerization of ylides derived from allylic arsonium salts: influence of the double bond substitution on the outcome.

New oligomers or polymers are prepared by polymerization of arsonium ylides in the presence of boron compounds.

Clinical Translation of a Click-Labeled 18F-Octreotate Radioligand for Imaging Neuroendocrine Tumors.

UNLABELLED: We conducted the first-in-human study of (18)F-fluoroethyl triazole [Tyr(3)] octreotate ((18)F-FET-ßAG-TOCA) in patients with neuroendocrine tumors (NETs) to evaluate biodistribution, dosimetry, and safety. Despite advances in clinical imaging, detection and quantification of NET activity remains a challenge, with no universally accepted imaging standard. METHODS: Nine patients were enrolled. Eight patients had sporadic NETs, and 1 had multiple endocrine neoplasia type 1 syndrome. Patients received 137-163 MBq (mean ± SD, 155.7 ± 8 MBq) of (18)F-FET-ßAG-TOCA. Safety data were obtained during and 24 h after radioligand administration. Patients underwent detailed whole-body PET/CT multibed scanning over 4 h with sampling of venous bloods for radioactivity and radioactive metabolite quantification. Regions of interest were defined to derive individual and mean organ residence times; effective dose was calculated with OLINDA 1.1. RESULTS: All patients tolerated (18)F-FET-ßAG-TOCA with no adverse events. Over 60% parent radioligand was present in plasma at 60 min. High tumor (primary and metastases)-to-background contrast images were observed. Physiologic distribution was seen in the pituitary, salivary glands, thyroid, and spleen, with low background distribution in the liver, an organ in which metastases commonly occur. The organs receiving highest absorbed dose were the gallbladder, spleen, stomach, liver, kidneys, and bladder. The calculated effective dose over all subjects (mean ± SD) was 0.029 ± 0.004 mSv/MBq. CONCLUSION: The favorable safety, imaging, and dosimetric profile makes (18)F-FET-ßAG-TOCA a promising candidate radioligand for staging and management of NETs. Clinical studies in an expanded cohort are ongoing to clinically qualify this agent.

Enhanced copper-mediated (18)F-fluorination of aryl boronic esters provides eight radiotracers for PET applications.

[(18)F]FMTEB, [(18)F]FPEB, [(18)F]flumazenil, [(18)F]DAA1106, [(18)F]MFBG, [(18)F]FDOPA, [(18)F]FMT and [(18)F]FDA are prepared from the corresponding arylboronic esters and [(18)F]KF/K222 in the presence of Cu(OTf)2py4. The method was successfully applied using three radiosynthetic platforms, and up to 26 GBq of non-carrier added starting activity of (18)F-fluoride.

Lapatinib access into normal brain and brain metastases in patients with Her-2 overexpressing breast cancer.

BACKGROUND: Brain metastases are common in human epidermal growth factor receptor (Her)-2-positive breast cancer. Drug access to brain metastases and normal brain is key to management of cranial disease. In this study, positron emission tomography (PET) scanning after administration of radiolabelled lapatinib was used to obtain direct evidence of cranial drug access. METHODS: Patients with Her-2+ metastatic breast cancer either with at least one 1-cm diameter brain metastasis or without brain metastases underwent dynamic carbon-11 radiolabelled lapatinib ([(11)C]lapatinib)-PET. Less than 20 µg of [(11)C]lapatinib was administered before and after 8 days of oral lapatinib (1,500 mg once daily). Radial arterial blood sampling was performed throughout the 90-min scan. The contribution of blood volume activity to the tissue signal was excluded to calculate lapatinib uptake in normal brain and metastases. Partitioning of radioactivity between plasma and tissue (V T) was calculated and the tissue concentration of lapatinib derived. Plasma lapatinib levels were measured and adverse events noted. RESULTS: Six patients (three with brain metastases) were recruited. About 80% plasma radioactivity corresponded to intact [(11)C]lapatinib after 60 min. PET signal in the brain corresponded to circulating radioactivity levels, with no [(11)C]lapatinib uptake observed in normal brain tissue. In contrast, radioactivity uptake in cranial metastases was significantly higher (p = 0.002) than that could be accounted by circulating radioactivity levels, consistent with [(11)C]lapatinib uptake in brain metastases. There was no difference in lapatinib uptake between the baseline and day 8 scans, suggesting no effect of increased drug access by inhibition of the drug efflux proteins by therapeutic doses of lapatinib. CONCLUSIONS: Increased lapatinib uptake was observed in brain metastases but not in normal brain. TRIAL REGISTRATION: ClinicalTrials.gov: NCT01290354.

An evaluation of the brain distribution of [(11)C]GSK1034702, a muscarinic-1 (M 1) positive allosteric modulator in the living human brain using positron emission tomography.

BACKGROUND: The ability to quantify the capacity of a central nervous system (CNS) drug to cross the human blood-brain barrier (BBB) provides valuable information for de-risking drug development of new molecules. Here, we present a study, where a suitable positron emission tomography (PET) ligand was not available for the evaluation of a potent muscarinic acetylcholine receptor type-1 (M1) allosteric agonist (GSK1034702) in the primate and human brain. Hence, direct radiolabelling of the novel molecule was performed and PET measurements were obtained and combined with in vitro equilibrium dialysis assays to enable assessment of BBB transport and estimation of the free brain concentration of GSK1034702 in vivo. METHODS: GSK1034702 was radiolabelled with (11)C, and the brain distribution of [(11)C]GSK1034702 was investigated in two anaesthetised baboons and four healthy male humans. In humans, PET scans were performed (following intravenous injection of [(11)C]GSK1034702) at baseline and after a single oral 5-mg dose of GSK1034702. The in vitro brain and plasma protein binding of GSK1034702 was determined across a range of species using equilibrium dialysis. RESULTS: The distribution of [(11)C]GSK1034702 in the primate brain was homogenous and the whole brain partition coefficient (V T) was 3.97. In contrast, there was mild regional heterogeneity for GSK1034702 in the human brain. Human whole brain V T estimates (4.9) were in broad agreement with primate V T and the f P/f ND ratio (3.97 and 2.63, respectively), consistent with transport by passive diffusion across the BBB. CONCLUSION: In primate and human PET studies designed to evaluate the transport of a novel M1 allosteric agonist (GSK1034702) across the BBB, we have demonstrated good brain uptake and BBB passage consistent with passive diffusion or active influx. These studies discharged some of the perceived development risks for GSK1034702 and provided information to progress the molecule into the next stage of clinical development. TRIAL REGISTRATION: Clinical trial details: 'Brain Uptake of GSK1034702: a Positron Emission Tomography (PET) Scan Study.'; clinicaltrial.gov identifier: NCT00937846 .

A broadly applicable [18F]trifluoromethylation of aryl and heteroaryl iodides for PET imaging.

Molecules labelled with the unnatural isotope fluorine-18 are used for positron emission tomography. Currently, this molecular imaging technology is not exploited at its full potential because many (18)F-labelled probes are inaccessible or notoriously difficult to produce. Typical challenges associated with (18)F radiochemistry are the short half-life of (18)F (<2 h), the use of sub-stoichiometric amounts of (18)F, relative to the precursor and other reagents, as well as the limited availability of parent (18)F sources of suitable reactivity ([(18)F]F(-) and [(18)F]F2). There is a high-priority demand for general methods allowing access to [(18)F]CF3-substituted molecules for application in pharmaceutical discovery programmes. We report the development of a process for the late-stage [(18)F]trifluoromethylation of (hetero)arenes from [(18)F]fluoride using commercially available reagents and (hetero)aryl iodides. This [(18)F]CuCF3-based protocol benefits from a large substrate scope and is characterized by its operational simplicity.