GABAA-α5 subunit-containing receptors have been shown to play a key modulatory role in cognition and represent a promising drug target for cognitive dysfunction, as well as other disorders. Here we report on the preclinical and early clinical profile of a novel GABAA-α5 selective negative allosteric modulator (NAM), basmisanil, which progressed into Phase II trials for intellectual disability in Down syndrome and cognitive impairment associated with schizophrenia. Preclinical pharmacology studies showed that basmisanil is the most selective GABAA-α5 receptor NAM described so far. Basmisanil bound to recombinant human GABAA-α5 receptors with 5 nM affinity and more than 90-fold selectivity versus α1, α2, and α3 subunit-containing receptors. Moreover, basmisanil inhibited GABA-induced currents at GABAA-α5 yet had little or no effect at the other receptor subtypes. An in vivo occupancy study in rats showed dose-dependent target engagement and was utilized to establish the plasma exposure to receptor occupancy relationship. At estimated receptor occupancies between 30 and 65% basmisanil attenuated diazepam-induced spatial learning impairment in rats (Morris water maze), improved executive function in non-human primates (object retrieval), without showing anxiogenic or proconvulsant effects in rats. During the Phase I open-label studies, basmisanil showed good safety and tolerability in healthy volunteers at maximum GABAA-α5 receptor occupancy as confirmed by PET analysis with the tracer [11C]-Ro 15-4513. An exploratory EEG study provided evidence for functional activity of basmisanil in human brain. Therefore, these preclinical and early clinical studies show that basmisanil has an ideal profile to investigate potential clinical benefits of GABAA-α5 receptor negative modulation.

Background and Purpose GABAA-α5 subunit-containing receptors have been shown to play a key modulatory role in cognition and represent a promising drug target for cognitive dysfunction, as well as other disorders. We describe the preclinical and clinical profile of basmisanil, a potent and highly selective negative allosteric modulator (NAM) of GABAA α5 receptors. Experimental Approach In vitro assays assessed binding and functional selectivity. In vivo occupancy studies measured target engagement. Effects on cognition were tested in rats (Morris water maze) and non-human primates (NHP; object retrieval) and potential side effects (anxiety and proconvulsant) were tested in rats. In healthy volunteers, target engagement and modulation of neuronal network activity were assessed using PET and EEG. Key Results Basmisanil bound to recombinant human GABAA-α5 receptors with 5 nM affinity and more than 90-fold selectivity versus α1, α2, and α3 subunit-containing receptors. Basmisanil inhibited GABA-induced currents at GABAA-α5 yet had little or no effect at the other receptor subtypes. In vivo, basmisanil demonstrated dose-dependent target engagement in rats. Basmisanil attenuated diazepam-induced spatial learning impairment in rats and improved executive function in NHPs. At these efficacious plasma concentrations, basmisanil had no anxiogenic and proconvulsant effects. In healthy volunteers, PET showed target engagement and established the plasma exposure to receptor occupancy relationship. Basmisanil modulated brain function reflected in characteristic changes of EEG spectral power. There were no serious adverse events. Conclusion and Implications Basmisanil is a highly potent and selective GABAA α5 receptor NAM with good safety and tolerability allowing for clinical testing in multiple brain disorders.

Tau aggregates and amyloid-β (Aβ) plaques are key histopathologic features in Alzheimer disease (AD) and are considered targets for therapeutic intervention as well as biomarkers for diagnostic in vivo imaging agents. This article describes the preclinical in vitro and in vivo characterization of 3 novel compounds—RO6958948, RO6931643, and RO6924963—that bind specifically to tau aggregates and have the potential to become PET tracers for future human use. Methods: RO6958948, RO6931643, and RO6924963 were identified as high-affinity competitors at the 3H-T808 binding site on native tau aggregates in human late-stage AD brain tissue. Binding of tritiated compounds to brain tissue sections of AD patients and healthy controls was analyzed by macro- and microautoradiography and by costaining of tau aggregates and Aβ plaques on the same tissue section using specific antibodies. All 3 tracer candidates were radiolabeled with a PET nuclide and tested in vivo in tau-naïve baboons to assess brain uptake, distribution, clearance, and metabolism. Results:3H-RO6958948, 3H-RO6931643, and 3H-RO6924963 bound with high affinity and specificity to tau aggregates, clearly lacking affinity for concomitant Aβ plaques in human AD Braak V tissue sections. The specificity of all 3 radioligands for tau aggregates was supported, first, by binding patterns in AD sections comparable to the tau-specific radioligand 3H-T808; second, by very low nonspecific binding in brain tissue devoid of tau pathology, excluding significant radioligand binding to any other central nervous system target; and third, by macroscopic and microscopic colocalization and quantitative correlation of radioligand binding and tau antibody staining on the same tissue section. RO6958948, RO6931643, and RO6924963 were successfully radiolabeled with a PET nuclide at high specific activity, radiochemical purity, and yield. After intravenous administration of 18F-RO6958948, 11C-RO6931643, and 11C-RO6924963 to baboons, PET scans indicated good brain entry, rapid washout, and a favorable metabolism pattern. Conclusion:18F-RO6958948, 11C-RO6931643, and 11C-RO6924963 are promising PET tracers for visualization of tau aggregates in AD. Head-to-head comparison and validation of these tracer candidates in AD patients and healthy controls will be reported in due course.

Tau aggregates and amyloid-beta (Aβ) plaques are key histopathological features in Alzheimer‘s disease (AD) and are considered as targets for therapeutic intervention as well as biomarkers for diagnostic in vivo imaging agents. This study describes the preclinical in vitro and in vivo characterization of the 3 novel compounds RO6958948, RO6931643 and RO6924963 that bind specifically to tau aggregates and have the potential to become positron emission tomography (PET) tracers for future human use. Methods: RO6958948, RO6931643 and RO6924963 were identified as high affinity competitors at the 3H-T808 binding site on native tau aggregates in human late stage AD brain tissue. Binding of tritiated compounds to brain tissue sections of AD patients and healthy controls (HC) was analyzed by macroand microautoradiography, and co-staining of tau aggregates and Aβ plaques on the same tissue section using specific antibodies. All 3 tracer candidates were radiolabeled with a PET nuclide and tested in vivo in tau-naïve baboons to assess brain uptake, distribution, clearance and metabolism. Results: 3HRO6958948, 3H-RO6931643 and 3H-RO6924963 bound with high affinity and specificity to tau aggregates, clearly lacking affinity for concomitant Aβ plaques in human AD Braak V tissue sections. The specificity of all 3 radioligands for tau aggregates was supported by (1) binding patterns in AD sections comparable to the tau-specific radioligand 3H-T808, (2) very low non-specific binding (NSB) in brain tissue devoid of tau pathology excluding significant radioligand binding to any other central nervous system (CNS) target and (3) macroscopic and microscopic colocalization and quantitative correlation of radioligand binding and tau antibody staining on the same tissue section. RO6958948, RO6931643 and RO6924963 were successfully radiolabeled with a PET nuclide at high specific activity, radiochemical purity and yield. Following intravenous administration in baboons, 18F-RO6958948, 11C-RO6931643 and 11C-RO6924963 PET scans indicated good brain entry, rapid washout and a favorable metabolism pattern. Conclusions: 18F-RO6958948, 11C-RO6931643 and 11C-RO6924963 are promising PET tracers for visualization of tau aggregates in AD. Head-to-head comparison and validation of these tracer candidates in AD patients and HCs will be reported in due course.

Aggregates of tau and beta amyloid (Aβ) plaques constitute the histopathological hallmarks of Alzheimer's disease and are prominent targets for novel therapeutics as well as for biomarkers for diagnostic in vivo imaging. In recent years much attention has been devoted to the discovery and development of new PET tracers to image tau aggregates in the living human brain. Access to a selective PET tracer to image and quantify tau aggregates represents a unique tool to support the development of any novel therapeutic agent targeting pathological forms of tau. The objective of the study described herein was to identify such a novel radiotracer. As a result of this work, we discovered three novel PET tracers (2-(4-[11C]methoxyphenyl)imidazo[1,2-a]pyridin-7-amine 7 ([11C]RO6924963), N-[11C]methyl-2-(3-methylphenyl)imidazo[1,2-a]pyrimidin-7-amine 8 ([11C]RO6931643), and [18F]2-(6-fluoropyridin-3-yl)pyrrolo[2,3-b:4,5-c']dipyridine 9 ([18F]RO6958948)) with high affinity for tau neurofibrillary tangles, excellent selectivity against Aβ plaques, and appropriate pharmacokinetic and metabolic properties in mice and non-human primates.

[11C]RO6924963, [11C]RO6931643, and [18F]RO6958948 have been reported as promising PET tracers for tau imaging based on in vitro studies and preclinical PET data (Honer, HAI 2015). Here we describe the first evaluation of these novel radiotracers in humans (ClinicalTrials.gov: NCT02187627). Seven amyloid PET positive Alzheimer's disease (AD) patients (4M:3F; 64-86 y; MMSE: 16-24), and 7 young healthy subjects (HS) (5M:2F; 25-38 y) each received 2 different tau tracers (1 additional HS was withdrawn). Dynamic 90 min scans were obtained after bolus injection of [11C]RO6924963 (4HS:2AD), [11C]RO6931643 (5HS:7AD), or [18F] RO6958948 (5HS:5AD). Arterial blood sampling was performed in 13/14 HS and 8/14 AD. Regions were defined on MRI, and PET data was quantified by plasma-reference graphical analysis (for VT) and target-cerebellum ratio (SUVR60-90). SUVRimages were also analyzed voxel-wise. Peak SUV values were approximately 3, 1.5 and 3.5 for [11C]RO6924963, [11C]RO6931643, and [18F]RO6958948, respectively. The retention of [11C]RO6931643 and [18F]RO6958948 in HS was much lower than that of [11C]RO6924963. For [11C]RO6931643 and [18F]RO6958948 (in AD), highest uptake was seen in the frontal (Fr), temporal (Tp), parietal (Pa), occipital (Oc), and fusiform (Fs) cortices, and the entorhinal area (ER). SUVR60-90 values ranged from 1.22 (L. Fr) to 1.55 (L. Oc) for [11C]RO6931643, and 1.39 (R.Fr) to 2.25 (L. Fs) for [18F]RO6958948, and VT ranged from 2.28 (R. ER) to 3.14 (L. Tp), and 3.98 (R. ER) to 5.62 (L. Tp), respectively. Regional analysis of SUVR and VT for [11C]RO6931643 and [18F]RO6958948 clearly allowed AD and HS to be distinguished. When compared to HS, the two tracers showed robust group effects (F>90; p=<10−6; two-way ANOVA) on SUVR and VT, and significant group differences (p<0.05 with Bonferroni correction; without overlap) in 6 of 12 regions for [11C]RO6931643 and 4 of 12 for [18F]RO6958948 (using SUVR). Voxel-wise analysis of SUVR revealed clusters of significantly higher uptake in AD compared to HS in Fs for [11C]RO6931643 (7 AD vs. 5 HS), and [18F]RO6958948 (5 AD vs. 5 HS). There were no radiolabelled metabolites nor defluorination of [18F]RO6958948.

The late-stage fluorination of common synthetic building blocks and drug leads is an appealing reaction for medicinal chemistry. In particular, fluorination of benzylic C-H bonds provides a means to attenuate drug metabolism at this metabolically labile position. Here we report two complimentary strategies for the direct fluorination of benzylic C-H bonds using N-fluorobenzenesulfonimide and either a decatungstate photocatalyst or AIBN-initiation.

The NK3 receptor is a GPCR that is prominently expressed in limbic areas of the brain, many of which have been implicated in schizophrenia. Phase II clinical trials in schizophrenia with two selective NK3 antagonists (osanetant and talnetant) have demonstrated significant improvement in positive symptoms. The objective of this study was to characterize the properties of a novel dual NK2/NK3 antagonist, RO5328673. [(3)H]RO5328673 bound to a single saturable site on hNK2, hNK3 and gpNK3 with high-affinity. RO5328673 acted as an insurmountable antagonist at both human and guinea-pig NK3 receptors in the [(3)H]IP accumulation assay. In binding kinetic analyses, [(3)H]RO5328673 had fast association and dissociation rates at hNK2 while it had a fast association rate and a remarkably slow dissociation rate at gp and hNK3. In electrophysiological recordings of gp SNpc, RO5328673 inhibited the senktide-induced potentiation of spontaneous activity of dopaminergic neurons with an insurmountable mechanism of action. RO5328673 exhibited in-vivo activity in gerbils, robustly reversing the senktide-induced locomotor activity. The TM2 residue gpNK3-A114(2.58) (threonine in all other species) was identified as the critical residue for the RO5328673's slower dissociation kinetics and stronger insurmountable mode of antagonism in the guinea-pig as compared to hNK3-T139(2.58). Using site-directed mutagenesis, [(3)H]RO5328673 binding and rhodopsin-based modeling, the important molecular determinants of the RO5328673-binding pocket of hNK3 were determined. A comparison of the RO5328673-binding pocket with that of osanetant showed that two antagonists have similar contact sides on hNK3 binding crevice except for three mutations V95L(1.42), Y247W(5.38), V255I(5.46), which behaved differently between interacting modes of two antagonists in hNK3.

Recently we have documented research efforts aimed at new classes of oxetanes as well as spiro-heteroalicyclic ring systems (which we have termed 'Compact Modules') designed to expand the palette of tailored module scaffolds available to medicinal chemists, which constitute an important role for synthetic chemistry in the drug discovery process. An essential component for this process is to provide access to specific molecular topologies with functional group diversity, essential for generating leads that discriminate among biological targets, therefore promoting selectivity and enhancing the safety profile of the final clinical candidates.

With N-Boc-protected 4-(allylaminomethyl)-2-(5H)furanones as starting materials, a photochemical approach is presented to give 3,9-diazatricyclo[5.3.0.0(1,5)]decanes as conformationally restricted bis-pyrrolidines. The products are orthogonally protected at the two nitrogen atoms and exhibit, depending on the substitution pattern at positions C5, C6, and C7, latent C-2 symmetry. When the furanones had a phenyl group at the 3-position (X-3), alternative photochemical pathways were observed.