Discovery and Preclinical Characterization of BIIB129, a Covalent, Selective, and Brain-Penetrant BTK Inhibitor for the Treatment of Multiple Sclerosis.

Multiple sclerosis (MS) is a chronic disease with an underlying pathology characterized by inflammation-driven neuronal loss, axonal injury, and demyelination. Bruton's tyrosine kinase (BTK), a nonreceptor tyrosine kinase and member of the TEC family of kinases, is involved in the regulation, migration, and functional activation of B cells and myeloid cells in the periphery and the central nervous system (CNS), cell types which are deemed central to the pathology contributing to disease progression in MS patients. Herein, we describe the discovery of BIIB129 (25), a structurally distinct and brain-penetrant targeted covalent inhibitor (TCI) of BTK with an unprecedented binding mode responsible for its high kinome selectivity. BIIB129 (25) demonstrated efficacy in disease-relevant preclinical in vivo models of B cell proliferation in the CNS, exhibits a favorable safety profile suitable for clinical development as an immunomodulating therapy for MS, and has a low projected total human daily dose.

Utilization of Cyclic Amides as Masked Aldehyde Equivalents in Reductive Amination Reactions.

An operationally simple protocol has been discovered that couples primary or secondary amines with N-aryl-substituted lactams to deliver differentiated diamines in moderate to high yields. The process allows for the partial reduction of a lactam in the presence of Cp2ZrHCl (Schwartz's reagent), followed by a reductive amination between the resulting hemiaminal and primary or secondary amine. These reactions can be telescoped in a one-pot fashion to significantly simplify the operation. The scope of amines and substituted lactams of various ring sizes was demonstrated through the formation of a range of differentiated diamine products. Furthermore, this methodology was expanded to include N-aryl pyrrolidinone substrates with an enantiopure ester group at the 5-position, and α-amino piperidinones were prepared with complete retention of stereochemical information. The development of this chemistry has enabled the consideration of lactams as useful synthons.

Highly hindered 2-(aryl-di-tert-butylsilyl)-N-methyl-imidazoles: a new tool for the aqueous 19F- and 18F-fluorination of biomolecule-based structures.

A new class of silicon-based fluoride acceptors with a C-linked heterocycle as the leaving group was synthesized in one step from commercial chemicals, and linked to biomolecules. The resulting conjugates were efficiently 19F-fluorinated in aqueous mixtures, and switching to 18F-labelling provided nucleoside- and peptide-based bioconjugates with excellent molar activities suitable for biological applications.

What about αvß3 integrins in molecular imaging in oncology?

Non-invasive investigation of integrin expression is an interesting approach in nuclear medicine department. Indeed, integrins are overexpressed in a wide array of diseases, including tumor neoangiogenesis, cardiovascular pathologies, immune dysfunction, etc. Different targets have been identified in order to be detected and quantified for angiogenesis and vascular remodeling, among them VEGF, matrix metalloproteases, and integrins (αvß3, but also α5ß1 and αvß6). Their targeting appears of great interest either for early diagnosis, aggressiveness staging of the disease or for selection of responders to new-targeted therapies. Thus, αvß3 is a biomarker of angiogenesis that specifically binds to RGD containing peptides. Many different strategies were attempted to develop RGD peptides for single photon emission tomography (SPECT) and positron emission tomography (PET) imaging. This review is mainly focused on αvß3-targeting in oncology. We will present an overview of the tracers mostly used on nuclear imaging techniques, those in clinical trials, the recent development concerning the 18F-labeling strategies, the 68Ga-complex chemistry and different approaches of therapy.

Design and Synthesis of Piperazine Sulfonamide Cores Leading to Highly Potent HIV-1 Protease Inhibitors.

Using the HIV-1 protease binding mode of MK-8718 and PL-100 as inspiration, a novel aspartate binding bicyclic piperazine sulfonamide core was designed and synthesized. The resulting HIV-1 protease inhibitor containing this core showed an 60-fold increase in enzyme binding affinity and a 10-fold increase in antiviral activity relative to MK-8718.

Ex Vivo and In Vivo Imaging and Biodistribution of Aptamers Targeting the Human Matrix MetalloProtease-9 in Melanomas.

The human Matrix MetalloProtease-9 (hMMP-9) is overexpressed in tumors where it promotes the release of cancer cells thus contributing to tumor metastasis. We raised aptamers against hMMP-9, which constitutes a validated marker of malignant tumors, in order to design probes for imaging tumors in human beings. A chemically modified RNA aptamer (F3B), fully resistant to nucleases was previously described. This compound was subsequently used for the preparation of F3B-Cy5, F3B-S-acetylmercaptoacetyltriglycine (MAG) and F3B-DOTA. The binding properties of these derivatives were determined by surface plasmon resonance and electrophoretic mobility shift assay. Optical fluorescence imaging confirmed the binding to hMMP-9 in A375 melanoma bearing mice. Quantitative biodistribution studies were performed at 30 min, 1h and 2 h post injection of 99mTc-MAG-aptamer and 111In-DOTA-F3B. 99mTc radiolabeled aptamer specifically detected hMMP-9 in A375 melanoma tumors but accumulation in digestive tract was very high. Following i.v. injection of 111In-DOTA-F3B, high level of radioactivity was observed in kidneys and bladder but digestive tract uptake was very limited. Tumor uptake was significantly (student t test, p<0.05) higher for 111In-DOTA-F3B with 2.0%ID/g than for the 111In-DOTA-control oligonucleotide (0.7%ID/g) with tumor to muscle ratio of 4.0. Such difference in tumor accumulation has been confirmed by ex vivo scintigraphic images performed at 1h post injection and by autoradiography, which revealed the overexpression of hMMP-9 in sections of human melanomas. These results demonstrate that F3B aptamer is of interest for detecting hMMP-9 in melanoma tumor.

Evaluation of (68)Ga-DOTA-TOC PET/CT for the detection of duodenopancreatic neuroendocrine tumors in patients with MEN1.

CONTEXT: Somatostatin receptor scintigraphy with (111)In-pentetreotide (SRS) is used to detect duodenopancreatic neuroendocrine tumors (dpNETs) in multiple endocrine neoplasia type 1 (MEN1). However, SRS has limited sensitivity for this purpose. Positron emission tomography/computed tomography (PET/CT) with (68)Ga-DOTA-TOC has a higher rate of sporadic dpNETs detection than SRS but there is little data for dpNETs detection in MEN1. PURPOSE: To compare the performances of (68)Ga-DOTA-TOC PET/CT, SRS and contrast-enhanced computed tomography (CE-CT) to diagnose dpNETs in MEN1. DESIGN AND SETTING: Single-institution prospective comparative study PATIENTS AND METHODS: Nineteen consecutive MEN1 patients (aged 47 ± 13 years) underwent (68)Ga-DOTA-TOC PET/CT, SRS, and CE-CT within 2 months in random order. Blinded readings of images were performed separately by experienced physicians. Unblinded analysis of CE-CT, combined with additional magnetic resonance imaging, endoscopic-ultrasound, (18)F-2-fluoro-deoxy-D-glucose ((18)F-FDG) PET/CT or histopathology results served as reference standard for dpNETs diagnosis. RESULTS: The sensitivity of (68)Ga-DOTA-TOC PET/CT, SRS, and CE-CT was 76, 20, and 60 %, respectively (p < 0.0001). All the true-positive lesions detected by SRS were also depicted on (68)Ga-DOTA-TOC PET/CT. (68)Ga-DOTA-TOC PET/CT detected lesions of smaller size than SRS (10.7 ± 7.6 and 15.2 ± 5.9 mm, respectively, p < 0.03). False negatives of (68)Ga-DOTA-TOC PET/CT included small dpNETs (<10 mm) and (18)F-FDG PET/CT positive aggressive dpNETs. No false positives were recorded. In addition, whole-body mapping with (68)Ga-DOTA-TOC PET/CT identified extra-abdominal MEN1-related tumors including one neuroendocrine thymic carcinoma identified by the three imaging procedures, one bronchial carcinoid undetected by CE-CT and three meningiomas undetected by SRS. CONCLUSIONS: Owing to higher diagnostic performance, (68)Ga-DOTA-TOC PET/CT (or alternative (68)Ga-labeled somatostatin analogues) should replace (111)In-pentetreotide in the investigation of MEN1 patients.

Simplified Quantification Method for In Vivo SPECT Imaging of the Vesicular Acetylcholine Transporter with 123I-Iodobenzovesamicol.

UNLABELLED: (123)I-iodobenzovesamicol is a SPECT radioligand selective for the vesicular acetylcholine transporter (VAChT) and used to assess the integrity of cholinergic pathways in various neurologic disorders. The current noninvasive method for quantitative analysis of (123)I-iodobenzovesamicol, based on multilinear reference tissue model 2 (MRTM2), requires repeated scans for several hours, limiting its application in clinical trials. Our objective was to validate a simplified acquisition method based on a single (123)I-iodobenzovesamicol static scan preserving the quantification accuracy. Three acquisition times were tested comparatively to a kinetic analysis using MRTM2. METHODS: Six healthy volunteers underwent a dynamic SPECT acquisition comprising 14 frames over 28 h and an MR imaging scan. MR images were automatically segmented, providing the volumes of 19 regions of interest (ROIs). SPECT datasets were coregistered with MR images, and regional time-activity curves were derived. For each ROI, a complete MRTM2 pharmacokinetic analysis, using the cerebellar hemispheres as the reference region, led to the calculation of a (123)I-iodobenzovesamicol-to-VAChT binding parameter, the nondisplaceable binding potential (BP(ND-MRTM2)). A simplified analysis was also performed at 5, 8, and 28 h after injection, providing a simplified BP(ND), given as BP(ND-t) = C(ROI) - C(cerebellar hemispheres)/C(cerebellar hemispheres), with C being the averaged radioactive concentration. RESULTS: No significant difference was found among BP(ND-5 h), BP(ND-8 h), and BP(ND-MRTM2) in any of the extrastriatal regions explored. BP(ND-28 h) was significantly higher than BP(ND-5 h), BP(ND-8 h), and BP(ND-MRTM2) in 9 of the 17 regions explored (P < 0.05). BP(ND-5 h), BP(ND-8 h), and BP(ND-28 h) correlated significantly with BP(ND-MRTM2) (P < 0.05; ρ = 0.99, 0.98, and 0.92, respectively). In the striatum, BP(ND-28 h) was significantly higher than BP(ND-5 h) and BP(ND-8 h). BP(ND-5 h) differed significantly from BP(ND-MRTM2) (P < 0.05), with BP(ND-5 h) being 43.6% lower. CONCLUSION: In the extrastriatal regions, a single acquisition at 5 or 8 h after injection provides quantitative results similar to a pharmacokinetic analysis. However, with the highest correlation and accuracy, 5 h is the most suitable time to perform an accurate (123)I-iodobenzovesamicol quantification. In the striatum, none of the 3 times has led to an accurate quantification.

A phantom-based method to standardize dose-calibrators for new ß+-emitters.

Quantitative imaging with PET requires accurate measurements of the amount of radioactivity injected into the patient and the concentration of radioactivity in a given region. Recently, new positron emitters, such as (124)I, (89)Zr, (82)Rb, (68)Ga, and (64)Cu, have emerged to promote PET development, but standards are still largely lacking. Therefore, we propose to validate a simple, robust, and replicable methodology, not requiring the use of any standards, to accurately calibrate a dose-calibrator for any ß(+)-emitter. On the basis of (18)F cross-calibration, routinely performed with fluorine-18-fluorodeoxyglucose (F-FDG) in nuclear medicine departments, a methodology was developed using ß(+)-emitting' phantoms to cross-calibrate the dose-calibrator for measuring the activity of positron emitters and quantifying the standardized uptake value (SUV). Ga phantoms filled with activities measured with various dose-calibrator settings were imaged to establish calibration curves (SUV values as a function of the dose-calibrator settings) and to identify the setting value, yielding an SUV value of 1.00 g/ml, reflecting an accurate measurement of (68)Ga activity. Activities measured with the identified setting were finally checked with a γ-counter. The setting of 772±1 was identified as ensuring that the studied dose-calibrator is correctly calibrated for (68)Ga to ensure an SUV value of 1.00±0.01 g/ml. γ-Ray spectrometry confirmed the accurate measurement of Ga activities by the dose-calibrator (relative error of 2.9±1.5%). We have developed a phantom-based method to accurately standardize dose-calibrators for any ß(+)-emitter, without any standards.

Associations between infants' crying, sleep and cortisol secretion and mother's sleep and well-being.

BACKGROUND: Infants' continuous crying is a challenge both for the child and the principal caregiver. However, the links between infants' sleep, crying and cortisol secretion and mothers' well-being and sleep have been scarcely investigated. The aim of the present study was therefore to examine the link between cortisol secretion, crying and sleep of infants characterized by infantile colic (IC) and mothers' psychological well-being and own sleep. METHODS: Mothers of 24 infants characterized by IC (mean age = 8 weeks, SD = 1.5 weeks) completed a series of questionnaires regarding the infant's crying and sleeping patterns. Infants' sleep was objectively assessed with actigraphs. Cortisol secretion was measured by means of saliva samples in the mornings after waking. After 4 weeks, infants were assessed once again. Mothers completed questionnaires assessing their psychological well-being (depressive symptoms, family strain) and sleep. RESULTS: Mothers' psychological well-being and sleep was greatly predicted by infants' morning saliva cortisol levels, sleep disruptions and crying intensity, whereas infants' crying duration and volume had low predictive value. CONCLUSIONS: Mothers with infants characterized by IC are at increased risk for reporting impaired sleep, developing depressive symptoms and reporting higher family strain. Most importantly, this risk seems to be greater if their infants' sleep is fragmented.

A new SiF-Dipropargyl glycerol scaffold as a versatile prosthetic group to design dimeric radioligands: synthesis of the [(18) F]BMPPSiF tracer to image serotonin receptors.

A novel SiX-dipropargyl glycerol scaffold (X: H, F, or (18) F) was developed as a versatile prosthetic group that provides technical advantages for the preparation of dimeric radioligands based on silicon fluoride acceptor pre- or post-labeling with fluorine-18. Rapid conjugation with the prosthetic group takes place in microwave-assisted click conjugation under mild conditions. Thus, a bivalent homodimeric SiX-dipropargyl glycerol derivatized radioligand, [(18) F]BMPPSiF, with enhanced affinity was developed by using click conjugation. High uptake of the radioligand was demonstrated in 5-HT1A receptor-rich regions in the brain with positron emission tomography. Molecular docking studies (rigid protein-flexible ligand) of BMPPSiF and known antagonists (WAY-100635, MPPF, and MefWAY) with monomeric, dimeric, and multimeric 5-HT1A receptor models were performed, with the highest G score obtained for docked BMPPSiF: -6.766 as compared with all three antagonists on the monomeric model. Multimeric induced-fit docking was also performed to visualize the comparable mode of binding under in vivo conditions, and a notably improved G score of -8.455 was observed for BMPPSiF. These data directly correlate the high binding potential of BMPPSiF with the bivalent binding mode obtained in the biological studies. The present study warrants wide application of the SiX-dipropargyl glycerol prosthetic group in the development of ligands for imaging with enhanced affinity markers for specific targeting based on peptides, nucleosides, and lipids.

Low brain penetrant CB1 receptor agonists for the treatment of neuropathic pain.

Novel, low brain penetrant, orally bioavailable CB1 receptor agonists were designed starting from a mature lead series of potent brain penetrant CB1 receptor agonists. Increasing the calculated polar surface area was found to be a good strategy for reducing brain penetration whilst retaining drug-like properties. This in silico approach led to the discovery of LBP1, an orally bioavailable, low brain penetrant CB1 receptor agonist with robust activity in rodent models of neuropathic pain and a good preclinical therapeutic profile, which was selected for clinical development.

Design, synthesis and structure-activity relationships of (indo-3-yl) heterocyclic derivatives as agonists of the CB1 receptor. Discovery of a clinical candidate.

We report an expansion of the structure-activity relationship (SAR) of a novel series of indole-3-heterocyclic CB1 receptor agonists. Starting from the potent but poorly soluble lead, 1, a rational approach was taken in order to balance solubility, hERG activity and potency while retaining the desired long duration of action within the mouse tail flick test. This led to the discovery of compound 38 which successfully progressed into clinical development.

Discovery of potent and orally bioavailable heterocycle-based cannabinoid CB1 receptor agonists.

Novel 3-(1H-indol-3-yl)-1,2,4-oxadiazoles and -thiadiazoles were synthesized and found to be potent CB1 cannabinoid receptor agonists. The oral bioavailability of these compounds could be dramatically improved by optimization studies of the side chains attached to the indole and oxadiazole cores, leading to identification of a CB1 receptor agonist with good oral activity in a range of preclinical models of antinociception and antihyperalgesia.

Silicon-based chemistry: an original and efficient one-step approach to [18F]-nucleosides and [18F]-oligonucleotides for PET imaging.

Take it eaSi! Nucleosides, dinucleotides, and one oligonucleotide, all modified by click chemistry, have for the first time been directly and very efficiently labeled with (18)F by using a silicon-based, one-step approach that opens the way for the development of a new class of positron emission tomography (PET) tracers (see graphic).

Design, synthesis, and structure-activity relationships of indole-3-heterocycles as agonists of the CB1 receptor.

Novel indole-3-heterocycles were designed and synthesized and found to be potent CB1 receptor agonists. Starting from a microsomally unstable lead 1, a bioisostere approach replacing a piperazine amide was undertaken. This was found to be a good strategy for improving stability both in vitro and in vivo. This led to the discovery of 24, which had an increased duration of action in the mouse tail flick test in comparison to the lead 1.

Tandem copper-catalysed aryl and alkenyl amination reactions: the synthesis of N-functionalised indoles.

A Cu-diamine complex effectively catalyses tandem C-N bond formation on 2-(2-haloalkenyl)-aryl halide substrates, to deliver a series of N-functionalised indoles. Anilines, amides and carbamates are all effective coupling partners under the developed conditions.

Identification of substituted 6-amino-4-phenyltetrahydroquinoline derivatives: potent antagonists for the follicle-stimulating hormone receptor.

Substituted 6-amino-4-phenyl-tetrahydroquinoline derivatives are described that are antagonists for the G(s)-protein-coupled human follicle-stimulating hormone (FSH) receptor. These compounds show high antagonistic efficacy in vitro using a CHO cell line expressing the human FSH receptor. Antagonist 10 also showed a submicromolar IC(50) in a more physiologically relevant rat granulosa cell assay and was found to significantly inhibit follicle growth and ovulation in an ex vivo mouse model. This compound class may open the way toward a novel, nonsteroidal approach for contraception.