Discovery of Affinity-Based Probes for Btk Occupancy Assays

Bruton's tyrosine kinase (Btk) is an attractive target for the treatment of a wide array of B-cell malignancies and autoimmune diseases. Small-molecule covalent irreversible Btk inhibitors targeting Cys481 have been developed for the treatment of such diseases. In clinical trials, probe molecules are required in occupancy studies to measure the level of engagement of the protein by these covalent irreversible inhibitors. The result of this pharmacodynamic (PD) activity provides guidance for appropriate dosage selection to optimize inhibition of the drug target and correlation of target inhibition with disease treatment efficacy. This information is crucial for successful evaluation of drug candidates in clinical trials. Based on the pyridine carboxamide scaffold of a novel solvent-accessible pocket (SAP) series of covalent irreversible Btk inhibitors, we successfully developed a potent and selective affinity-based biotinylated probe 12 (2-[(4-{4-[5-(1-{5-[(3aS,4S,6aR)-2-oxo-hexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamido}-3,6,9,12-tetraoxapentadecan-15-amido)pentanoyl]piperazine-1-carbonyl}phenyl)amino]-6-[1-(prop-2-enoyl)piperidin-4-yl]pyridine-3-carboxamide). Compound 12 has been used in Btk occupancy assays for preclinical studies to determine the therapeutic efficacy of Btk inhibition in two mouse lupus models driven by TLR7 activation and type I interferon.     

Intersecting Xenobiology and Neometabolism To Bring Novel Chemistries to Life

The diversity of life relies on a handful of chemical elements (carbon, oxygen, hydrogen, nitrogen, sulfur and phosphorus) as part of essential building blocks; some other atoms are needed to a lesser extent, but most of the remaining elements are excluded from biology. This circumstance limits the scope of biochemical reactions in extant metabolism – yet it offers a phenomenal playground for synthetic biology. Xenobiology aims to bring novel bricks to life that could be exploited for (xeno)metabolite synthesis. In particular, the assembly of novel pathways engineered to handle nonbiological elements (neometabolism) will broaden chemical space beyond the reach of natural evolution. In this review, xeno-elements that could be blended into nature's biosynthetic portfolio are discussed together with their physicochemical properties and tools and strategies to incorporate them into biochemistry. We argue that current bioproduction methods can be revolutionized by bridging xenobiology and neometabolism for the synthesis of new-to-nature molecules, such as organohalides.     

Proton-Detected Solid-State NMR of the Cell-Free Synthesized α-Helical Transmembrane Protein NS4B from Hepatitis C Virus

Proton-detected 100 kHz magic-angle-spinning (MAS) solid-state NMR is an emerging analysis method for proteins with only hundreds of microgram quantities, and thus allows structural investigation of eukaryotic membrane proteins. This is the case for the cell-free synthesized hepatitis C virus (HCV) nonstructural membrane protein 4B (NS4B). We demonstrate NS4B sample optimization using fast reconstitution schemes that enable lipid-environment screening directly by NMR. 2D spectra and relaxation properties guide the choice of the best sample preparation to record 2D ¹H-detected ¹H,¹⁵N and 3D ¹H,¹³C,¹⁵N correlation experiments with linewidths and sensitivity suitable to initiate sequential assignments. Amino-acid-selectively labeled NS4B can be readily obtained using cell-free synthesis, opening the door to combinatorial labeling approaches which should enable structural studies.     

Carbon Chain Length Modulates MDA-MB-231 Breast Cancer Cell Killing Mechanisms by Mitochondrially Targeted Aryl−Urea Fatty Acids

Targeting the tumor cell mitochondrion could produce novel anticancer agents. We designed an aryl−urea fatty acid ( 1 g ; 16({[4-chloro-3-(trifluoromethyl)phenyl]carbamoyl}amino)hexadecanoic acid) that disrupted the mitochondrion and decreased MDA-MB-231 breast cancer cell viability. To optimize the aryl−ureas the present study evaluated mitochondrial targeting by 1 g analogues containing alkyl chains between 10–17 carbons. Using the dye JC-1, the C12−C17 analogues efficiently disrupted the mitochondrial membrane potential (IC₅₀s 3.5±1.2 to 7.6±1.1 μM) and impaired ATP production; shorter analogues were less active. 7-Aminoactinomycin D/annexin V staining and flow cytometry showed that these agents activated the killing mechanisms of necrosis and apoptosis to varying extents (7-aminoactinomycin D/annexin V staining ratios 4.3–6.0). Indeed, 1 g and its C17 analogue preferentially activated necrosis and apoptosis, respectively (ratios 2.1 and 16). Taken together, alkyl chain length is a determinant of mitochondrial targeting by aryl−ureas and can be varied to develop analogues that activate apoptosis or necrosis in a regulated fashion.     

Interview mit Hans-Christian Boos zum Thema „Unternehmertum in der IT-Branche“

Ohne Zusammenfassung     

Modellierungs- und Simulationsverfahren für die optische Verbindungstechnik auf elektrischen Leiterplatten

Optical interconnection technology on the printed circuit board level is a key technology for future microelectronic equipment. The consideration of functional, technological, and economical requirements results in a hybrid solution, where electrical and optical interconnects are integrated into one substrate called electrical optical printed circuit board. The significant part of the entire design process for electrical optical printed circuit boards is marked by the design supporting modelling and simulation of optical interconnects. Based on an abstract model for an entire optical interconnect a simulation model for optical multimode-waveguides is presented, taking into account all significant waveguide properties. Apart from that, the modeling of active components (laser- and photo-diodes) is addressed.     

E-Government-Initiative BundOnline 2005 — Gesellschaftlicher Mehrwert oder unwirtschaftliche Geldverschwendung?

Ohne Zusammenfassung     

Automatic fog detection and estimation of visibility distance through use of an onboard camera

In this paper, we will present a technique for measuring visibility distances under foggy weather conditions using a camera mounted onboard a moving vehicle. Our research has focused in particular on the problem of detecting daytime fog and estimating visibility distances; thanks to these efforts, an original method has been developed, tested and patented. The approach consists of dynamically implementing Koschmieder's law. Our method enables computing the meteorological visibility distance, a measure defined by the International Commission on Illumination (CIE) as the distance beyond which a black object of an appropriate dimension is perceived with a contrast of less than 5%. Our proposed solution is an original one, featuring the advantage of utilizing a single camera and necessitating the presence of just the road and sky in the scene. As opposed to other methods that require the explicit extraction of the road, this method offers fewer constraints by virtue of being applicable with no more than the extraction of a homogeneous surface containing a portion of the road and sky within the image. This image preprocessing also serves to identify the level of compatibility of the processed image with the set of Koschmieder's model hypotheses. Nicolas Hautiére graduated from the École Nationale des Travaux Publics de l'État, France (2002). He received his M.S. and Ph.D. degree in computer vision, respectively, in 2002 and 2005 from Saint-Étienne University (France). From 2002, he is a researcher in the Laboratoire Central des Ponts et Chaussées (LCPC), Paris, France. His research interests include trafic engineering, computer vision, and pattern recognition. Jean-Philippe Tarel graduated from the École Nationale des Ponts et Chaussées, Paris, France (1991). He received his Ph.D. degree in Applied Mathematics from Paris IX-Dauphine University in 1996 and he was with the Institut National de Recherche en Informatique et Automatique (INRIA) from 1991 to 1996. From 1997 to 1998, he was a research associate at Brown University, USA. From 1999, he is a researcher in the Laboratoire Central des Ponts et Chaussées (LCPC), Paris, France, and from 2001 to 2003 in the INRIA. His research interests include computer vision, pattern recognition, and shape modeling. Jean Lavenant graduated from the École Nationale des Travaux Publics de l'État, Lyon, France (2001). He received the M.S. degree in Computer Vision from Jean Monnet university of Saint-Étienne in 2001. In 2001, he was a researcher in the Laboratoire Central des Ponts et Chaussées (LCPC). In 2002, he was a system engineer in Chicago (USA). He is currently an engineer for the french ministry of transports. Didier Aubert received the M.S. and Ph.D. degree, respectively, in 1985 and 1989 from the National Polytechnical Institut of Grenoble (INPG). From 1989--1990, he worked as a research scientist on the development of an automatic road following system for the NAVLAB at Carnegie Mellon University. From 1990–1994, he worked in the research department of a private company (ITMI). During this period he was the project leader of several projects dealing with computer vision. He is currently a researcher at INRETS since 1995 and works on Road traffic measurements, crowd monitoring, automated highway systems, and driving assistance systems for vehicles. He is an image processing expert for several companies, teaches at Universities (Paris VI, Paris XI, ENPC, ENST) and is at the editorial board of RTS (Research - Transport - Safety).