Efficient Screening of Target-Specific Selected Compounds in Mixtures by 19 F NMR Binding Assay with Predicted 19 F NMR Chemical Shifts.

Ligand-based 19 F NMR screening is a highly effective and well-established hit-finding approach. The high sensitivity to protein binding makes it particularly suitable for fragment screening. Different criteria can be considered for generating fluorinated fragment libraries. One common strategy is to assemble a large, diverse, well-designed and characterized fragment library which is screened in mixtures, generated based on experimental 19 F NMR chemical shifts. Here, we introduce a complementary knowledge-based 19 F NMR screening approach, named 19 Focused screening, enabling the efficient screening of putative active molecules selected by computational hit finding methodologies, in mixtures assembled and on-the-fly deconvoluted based on predicted 19 F NMR chemical shifts. In this study, we developed a novel approach, named LEFshift, for 19 F NMR chemical shift prediction using rooted topological fluorine torsion fingerprints in combination with a random forest machine learning method. A demonstration of this approach to a real test case is reported.

Comprehensive and High-Throughput Exploration of Chemical Space Using Broadband 19 F†NMR-Based Screening.

Fragment-based lead discovery has become a fundamental approach to identify ligands that efficiently interact with disease-relevant targets. Among the numerous screening techniques, fluorine-detected NMR has gained popularity owing to its high sensitivity, robustness, and ease of use. To effectively explore chemical space, a universal NMR experiment, a rationally designed fragment library, and a sample composition optimized for a maximal number of compounds and minimal measurement time are required. Here, we introduce a comprehensive method that enabled the efficient assembly of a high-quality and diverse library containing nearly 4000 fragments and screening for target-specific binders within days. At the core of the approach is a novel broadband relaxation-edited NMR experiment that covers the entire chemical shift range of drug-like 19 F motifs in a single measurement. Our approach facilitates the identification of diverse binders and the fast ligandability assessment of new targets.

Cerebrospinal Fluid Aß42/Aß40 as a Means to Limiting Tube- and Storage-Dependent Pre-Analytical Variability in Clinical Setting.

BACKGROUND: Alzheimer's disease (AD) cerebrospinal fluid (CSF) biomarkers have recently been included in the criteria for AD diagnosis. Unfortunately, their wider use in routine and interpretation require more standardization, particularly for the pre-analytical steps. In particular, amyloid-ß (Aß)42 peptide measurement is strongly influenced by the type of collection tube and by repeated freeze/thaw cycles. OBJECTIVE: The objectives of this study were to compare, in clinical setting, the impact of collection tubes and the repetition of freeze/thaw cycles on Aß42 and Aß40 concentrations and consequently determine if the Aß42/Aß40 ratio could resolve the diagnosis difficulties related to these pre-analytical parameters. METHODS: CSF from 35 patients was collected in different polypropylene (PP) and stored at - 80°C after sampling. For CSF collected in the reference tube, three successive freeze-thaw cycles were done. Aß42 and Aß40 concentrations were determined in each condition in order to calculate the Aß42/Aß40 ratio. RESULTS: Our results showed that CSF Aß42 and Aß40 values were significantly different according to the type of collection tube and the number of freeze/thaw cycles. Although the calculation of the Aß42/Aß40 ratio eliminated the effect of PP tube-dependent variation, this was not the case for freeze-thaw cycle-associated variation. CONCLUSION: The use of Aß42/Aß40 ratio rather than Aß42 alone could contribute toward pre-analytical standardization, thus allowing the general use of CSF AD biomarkers in routine practice.

Gastric Wall Fatty Infiltration in Patients Without Overt Gastrointestinal Disease.

OBJECTIVE: The purpose of this study was to evaluate the frequency of gastric wall fatty infiltration in patients without overt gastrointestinal disease. MATERIALS AND METHODS: A retrospective study included patients who underwent unenhanced MDCT for renal colic. Two radiologists reviewed all of the images and evaluated for the presence of gastric wall fatty infiltration. The following patient characteristics were also recorded: sex, age, body mass index, total and visceral fat area, and presence of colic or ileal fat halo sign, or hepatic steatosis. A t test and Fisher test were used to compare the results between patients with and patients without gastric wall fatty infiltration. RESULTS: Gastric wall fatty infiltration was present in 25 of 120 (21%) patients in the study. Mean age, weight, body mass index, visceral and total fat areas, proportion of hepatic steatosis, number of men, and frequency of the presence of colic and ileal fat halo signs were significantly higher among patients with gastric wall fatty infiltration than in those without infiltration. CONCLUSION: Gastric wall fatty infiltration was significantly more frequent in men older than 45 years and patients with a body mass index greater than 25. It may represent a normal finding, but its relation to other pathologic conditions related to obesity remains to be explored.

Discovery of Novel Allosteric Non-Bisphosphonate Inhibitors of Farnesyl Pyrophosphate Synthase by Integrated Lead Finding.

Farnesyl pyrophosphate synthase (FPPS) is an established target for the treatment of bone diseases, but also shows promise as an anticancer and anti-infective drug target. Currently available anti-FPPS drugs are active-site-directed bisphosphonate inhibitors, the peculiar pharmacological profile of which is inadequate for therapeutic indications beyond bone diseases. The recent discovery of an allosteric binding site has paved the way toward the development of novel non-bisphosphonate FPPS inhibitors with broader therapeutic potential, notably as immunomodulators in oncology. Herein we report the discovery, by an integrated lead finding approach, of two new chemical classes of allosteric FPPS inhibitors that belong to the salicylic acid and quinoline chemotypes. We present their synthesis, biochemical and cellular activities, structure-activity relationships, and provide X-ray structures of several representative FPPS complexes. These novel allosteric FPPS inhibitors are devoid of any affinity for bone mineral and could serve as leads to evaluate their potential in none-bone diseases.

Low-dose CT with automatic tube current modulation, adaptive statistical iterative reconstruction, and low tube voltage for the diagnosis of renal colic: impact of body mass index.

OBJECTIVE: The objective of our study was to evaluate the impact of body mass index (BMI) on dose, diagnostic performance, and image quality of a low-dose CT examination for renal colic. MATERIALS AND METHODS: This retrospective study included all patients who underwent a low-dose CT examination for renal colic performed during the year 2012 with automatic tube current modulation, adaptive statistical iterative reconstruction, and a low tube voltage (kV). Three readers independently reviewed all images for the presence of renal colic and evaluated diagnostic confidence and image quality. The results and doses were compared among patients grouped by body mass index (BMI) and between patients with a BMI<25 and those with a BMI≥25. RESULTS: Eighty-six patients were included in the study: 39 patients had a BMI<25 and 47 had a BMI≥25. No statistically significant difference was found between the accuracy rates for the diagnosis of renal colic when the rates of the three independent readers were averaged for both BMI groups (95.7% vs 96.4%, respectively; p=0.83). Image quality and diagnostic confidence scores were significantly better for patients with a BMI≥25 than for patients with a BMI<25 (mean image quality score, 3.7 vs 3.4, p<0.001; mean diagnostic confidence score, 2.8 vs 2.5, p<0.001). The mean radiation dose for patients with a BMI<25 was 2.4 mGy compared with 3.7 mGy for patients with a BMI≥25 (p<0.001). CONCLUSION: The diagnostic performance of our low-dose CT protocol for renal colic was excellent for all patients, and image quality and diagnostic confidence were significantly better for patients with a BMI≥25. However, our protocol also required exposure to a greater dose of radiation for these overweight and obese patients.

Time efficient detection of protein-ligand interactions with the polarization optimized PO-WaterLOGSY NMR experiment.

The identification of compounds that bind to a protein of interest is of central importance in contemporary drug research. For screening of compound libraries, NMR techniques are widely used, in particular the Water-Ligand Observed via Gradient SpectroscopY (WaterLOGSY) experiment. Here we present an optimized experiment, the polarization optimized WaterLOGSY (PO-WaterLOGSY). Based on a water flip-back strategy in conjunction with model calculations and numerical simulations, the PO-WaterLOGSY is optimized for water polarization recovery. Compared to a standard setup with the conventional WaterLOGSY, time consuming relaxation delays have been considerably shortened and can even be omitted through this approach. Furthermore, the robustness of the pulse sequence in an industrial setup was increased by the use of hard pulse trains for selective water excitation and water suppression. The PO-WaterLOGSY thus yields increased time efficiency by factor of 3-5 when compared with previously published schemes. These time savings have a substantial impact in drug discovery, since significantly larger compound libraries can be tested in screening campaigns.