[11C]UCB2713 in vivo/ex vivo EvaluationRadiosynthesis of UCB2713

The objective of the preclinical study in mice was to evaluate the feasibility of radiolabeling UCB2713 at two different positions, to assess which compound would be most suitable for a future phase 1 study to evaluate the brain biodistribution of minzasolmin. To assess the brain biodistribution of minzasolmin in humans, the radiolabeling position should ideally be metabolically stable, for example, the [11C]carbonyl position.

In this preclinical study, two radiolabeling positions were investigated, [11C-N-CH3]UCB2713 and [11C]carbonyl UCB2713 ([11C-CO]UCB2713). The radiolabeling of [11C-N-CH3]UCB2713 was performed by N-methylation of the corresponding desmethyl precursor with [11C]methyl iodide and radiolabeling of [11C-CO]UCB2713 was performed by [11C]aminocarbonylation of the iodo precursor and the amino precursor with [11C]carbon monoxide in the presence of Pd(π-cinnamyl) chloride dimer and Xantphos (4,5-bis(diphenylphosphino)-9,9-dimethylxanthene) (Fig. 1b). For additional information on the precursor molecules and radiolabeling methods, see the electronic supplement. A high-performance liquid chromatography (HPLC) metabolite analysis method was developed to quantify the ratio of [11C-N-CH3]UCB2713 and other unidentified metabolites in plasma and brain homogenate samples. Relative quantification was used to provide the ratio of parent to metabolite in these samples.

Experimental Procedures

Preclinical experiments were conducted on male C57 black 6 mice (N = 10). In vivo experiments were performed on 3 mice and included a PET scan of the brain; ex vivo experiments were performed on all 10 mice and included samples of brain homogenate.

All experiments were performed in accordance with the United Kingdom (UK) Animals (Scientific Procedures) Act 1986 and the transposed European Union (EU) directive 2010/63/EU. Procedures used in this study were approved by the Animal Ethical Review Committee of Imperial College London.

For more information on the experimental procedures (administration of radioligand, collection of samples, brain sample processing, preparation of sample for analysis, plasma sample analysis, HPLC procedure and data analysis, PET-computed tomography (CT) scanning and image reconstruction, and PET-CT image data analysis) see the electronic supplement, including Supplemental Figs. 1 and 2.

Phase 1 PET Study in Healthy Male Participants (TM0017)Study Design and Eligibility

A phase 1, single-center, open-label study was designed to determine the brain biodistribution of tracer [11C]minzasolmin with PET imaging in healthy male participants; safety and tolerability of minzasolmin was also assessed. Participants were scanned twice with PET-CT, radiotracer injections of [11C]minzasolmin (≤ 10 µg) were administered ahead of each scan and following the first scan a 360 mg oral dose of minzasolmin was administered 2–4 h before the second scan (Fig. 2). A sample size of 2–10 participants was considered sufficient to meet the objectives, whilst limiting radiation exposure.

Fig. 2

Study design figure for TM0017. *PK samples were taken at 0 h (up to 5 min before dosing), and 0.25 h, 0.5 h, 1 h, 1.5 h, 2 h, 3 h, 4 h, 6 h, 8 h, 10 h, 12 h, 15 h, 24 h, 36 h, and 48 h after 360 mg oral minzasolmin. h hours, min minutes, PET-CT positron emission tomography-computed tomography, PK pharmacokinetics.

All participants in the study were healthy, non-smokers, male, ≥ 25 and ≤ 55 years of age, with a body weight of ≥ 50 kg, body mass index of ≥ 18.0 or ≤ 30.0 kg/m2, with adequate collateral flow to the radial and ulnar arteries in both hands, as determined by an Allen’s test. Key exclusion criteria for this study included a history of neurological disease or an abnormal brain structure, as assessed by magnetic resonance (MR) imaging.

This study was conducted under the auspices of an Independent Ethics Committee and in accordance with the current version of the applicable regulatory and International Council for Harmonization Good Clinical Practice requirements, the ethical principles that have their origin from the Declaration of Helsinki, and the local laws of the countries involved.

Objectives and Variables

The primary objective was to determine the biodistribution of minzasolmin in the human brain. Secondary objectives included evaluating the safety and tolerability of minzasolmin, determining the ratio between the concentration of minzasolmin in the brain and that in plasma, and determining the relationship between brain biodistribution and administered dose of minzasolmin.

The Generation of Radiolabeled [11C]minzasolmin

[11C]minzasolmin was prepared as a sterile solution and was infused via IV bolus at a maximum dose of 300 MBq (≤ 10 µg) during each PET-CT scan. [11C]minzasolmin was prepared under Good Manufacturing Practice conditions, in accordance with local Chemistry, Manufacturing, and Control procedures and quality specifications described in the local radiopharmacy manual.

PET-CT Imaging Procedures and Scanning

The preparation of radiopharmaceuticals and execution of PET-CT scans was performed by radiochemists, physicians, and technologists of Invicro (London, UK). Following local anesthesia, a catheter was inserted into the radial artery by an experienced physician; two venous catheters were also inserted. A maximum of 300 MBq of [11C]minzasolmin was administered by qualified personnel after participants had been positioned in the PET scanner. Upon administration, dynamic scanning started immediately and continued for 90 min. Each participant underwent two PET-CT scans; approximately 2 h before the second scan the participants took a single oral dose of minzasolmin 360 mg.

During PET-CT scanning, continuous and discrete blood samples were collected with sufficient frequency to allow accurate determination of the time course of arterial whole blood, plasma, and radio-metabolites of [11C]minzasolmin. Arterial input function data were acquired and processed for all PET scans.

PET-CT Image Analysis

The outcome parameter for the assessment of minzasolmin availability was the estimated whole brain total distribution volume (VT) at equilibrium, unless significant heterogeneity between brain regions was observed. Anatomical regions of interest (ROI) were defined via nonlinear deformation of a template MR image and associated neuroanatomical atlas, to match individual participants’ MR images. These ROI included, but were not limited to: parietal lobe, occipital lobe, frontal lobe, temporal lobe, striatum, cerebellum, thalamus, and the brain stem. Participants’ PET images were registered to their MR images, enabling the individualized ROI to be applied to the dynamic emission data to generate regional time activity curves (TACs). For additional information on the generation of regional TACs, see the electronic supplement.

Safety Measurements

Adverse events (AEs) experienced by participants during this study and their medical history were coded using Version 20.1 of the Medical Dictionary for Regulatory Activities [14]. Medications were coded according to the September 2017 version of the World Health Organization Drug Dictionary [15]. A treatment-emergent adverse event (TEAE) was defined as any AE with a start date at the time of or after the first administration of [11C]minzasolmin. Where dates were missing or partially missing, AEs were assumed to be treatment-emergent, unless there was clear evidence to suggest otherwise.

Other safety variables recorded include changes in 12-lead electrocardiogram (ECG) assessment and changes in vital signs (heart rate, systolic blood pressure, diastolic blood pressure, respiratory rate, and body temperature). Vital signs were measured in supine position following 5 min of rest and clinically significant abnormalities (according to the investigator) were recorded as an AE. All ECG recordings were performed following 5 min of rest in the supine position, in triplicate, with ≥ 1 min between each recording. If any clinically significant abnormalities occurred, an ECG data specialist was to review the recordings.

Statistical Methods

All statistical analyses were performed using SAS® version 9.4 (SAS Institute, Cary, NC, USA). Descriptive statistics were produced if there were three or more data points for a given dose level at a given time.