Iterative model-building, structure refinement, and density modification with the PHENIX AutoBuild Wizard Thomas C. Terwilliger a* , Ralf W. Grosse-Kunstleve b , Pavel V. Afonine b , Nigel W. Moriarty b , Peter Zwart b , Li-Wei Hung a , Randy J. Read c , Paul D. Adams b* a b Los Alamos National Laboratory, Mailstop M888, Los Alamos, NM 87545, USA Lawrence Berkeley National Laboratory, One Cyclotron Road, Bldg 64R0121, Berkeley, CA 94720, USA. c Department of Haematology, University of Cambridge, Cambridge CB2 0XY, UK. * Email: terwill@lanl.gov or PDAdams@lbl.gov Running title: The PHENIX AutoBuild Wizard Abstract The PHENIX AutoBuild Wizard is a highly automated tool for iterative model- building, structure refinement and density modification using RESOLVE or TEXTAL model- building, RESOLVE statistical density modification, and phenix.refine structure refinement. Recent advances in the AutoBuild Wizard and phenix.refine include automated detection and application of NCS from models as they are built, extensive model completion algorithms, and automated solvent molecule picking. Model completion algorithms in the AutoBuild Wizard include loop-building, crossovers between chains in different models of a structure, and side-chain optimization. The AutoBuild Wizard has been applied to a set of 48 structures at resolutions ranging from 1.1 A to 3.2 A, resulting in a mean R-factor of 0.24 and a mean free R factor of 0.29. The R-factor of the final model is dependent on the quality of the starting electron density, and relatively independent of resolution. Keywords: Model building; model completion; macromolecular models; Protein Data Bank; structure refinement; PHENIX Introduction Iterative model-building and refinement is a powerful approach to obtaining a complete and accurate macromolecular model. The approach consists of cycles of building an atomic model based on an electron density map for a macromolecular structure, refining the structure, using the refined structure as a basis for improving the map, and building a new model. This type of approach has been carried out in a semi-automated fashion for many years, with manual model-building iterating with automated refinement (Jensen, 1997). More recently, with the development first of ARP/wARP (Perrakis et al., 1999), and later other procedures including RESOLVE iterative model-building and refinement (Terwilliger,

Iterative model-building, structure refinement, and density modification with the PHENIX AutoBuild Wizard

Tumours display considerable variation in the patterning and properties of angiogenic blood vessels, as well as in their responses to anti-angiogenic therapy. Angiogenic programming of neoplastic tissue is a multidimensional process regulated by cancer cells in concert with a variety of tumour-associated stromal cells and their bioactive products, which encompass cytokines and growth factors, the extracellular matrix and secreted microvesicles. In this Review, we discuss the extrinsic regulation of angiogenesis by the tumour microenvironment, highlighting potential vulnerabilities that could be targeted to improve the applicability and reach of anti-angiogenic cancer therapies.

Microenvironmental regulation of tumour angiogenesis

During the past decade, proof of the principle that peptide receptors can be used successfully for in vivo targeting of human cancers has been provided. The molecular basis for targeting rests on the in vitro observation that peptide receptors can be expressed in large quantities in certain tumors. The clinical impact is at the diagnostic level: in vivo receptor scintigraphy uses radiolabeled peptides for the localization of tumors and their metastases. It is also at the therapeutic level: peptide receptor radiotherapy of tumors emerges as a serious treatment option. Peptides linked to cytotoxic agents are also considered for therapeutic applications. The use of nonradiolabeled, noncytotoxic peptide analogs for long-term antiproliferative treatment of tumors appears promising for only a few tumor types, whereas the symptomatic treatment of neuroendocrine tumors by somatostatin analogs is clearly successful. The present review summarizes and critically evaluates the in vitro data on peptide and peptide receptor expression in human cancers. These data are considered to be the molecular basis for peptide receptor targeting of tumors. The paradigmatic peptide somatostatin and its receptors are extensively reviewed in the light of in vivo targeting of neuroendocrine tumors. The role of the more recently described targeting peptides vasoactive intestinal peptide, gastrin-releasing peptide, and cholecystokinin/gastrin is discussed. Other emerging and promising peptides and their respective receptors, including neurotensin, substance P, and neuropeptide Y, are introduced. This information relates to established and potential clinical applications in oncology.

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Peptide receptors as molecular targets for cancer diagnosis and therapy.

Natural products to drugs: natural product derived compounds in clinical trials

Lipid metabolism, in particular the synthesis of fatty acids (FAs), is an essential cellular process that converts nutrients into metabolic intermediates for membrane biosynthesis, energy storage and the generation of signalling molecules. This Review explores how different aspects of FA synthesis promote tumorigenesis and tumour progression. FA synthesis has received substantial attention as a potential target for cancer therapy, but strategies to target this process have not yet translated into clinical practice. Furthermore, efforts to target this pathway must consider the influence of the tumour microenvironment.

The multifaceted roles of fatty acid synthesis in cancer

Pharmacomodulation of torasemide, a diuretic sulfonylurea, led to the discovery of two novel diuretics, a sulfonylthiourea (BM 20) and a sulfonynylcyanoguanidine (BM 106). BM 27, a lipophilic sulfonylurea, exhibited neuroprotective properties associated to an anticonvulsant activity. As BM 27, two lipophilic sulfonythioureas (BM 11 and BM 34) revealed an anticonvulsant profile similar to that of phenytoin. Finally the synthesis of torasemide derivatives led to the development of a sulfonylcyanoguanidine (BM 144) with a thromboxane A2 antagonist potency.

Pharmacomodulation of torasemide led to original diuretic, neuroprotective, anticonvulsant and antithrombotic drugs

Knowing the potential therapeutic interest of AMPA potentiation, patents have been published for 15 years relating the discovery of novel compounds able to allosterically bind to the AMPA receptors and leading to a potentiation of the AMPA signals. Besides this classical approach, a new target for the medicinal chemist has been defined due to the recent findings describing the modulation of AMPA receptors by neuronal proteins, and has led to the design of dipyrazole compounds by Aventis. These new dipyrazole derivatives may be considered as ‘indirect AMPApams’, constituting a new pharmacological class. Based on their in vitro activity, it is probable that these compounds could be of therapeutic value for the treatment of cognitive disorders, schizophrenia or depression.

Novel dipyrazoles acting as glutamate modulators

Novel thiadiazine derivatives useful as AMPA receptor modulators, preparation method, and pharmaceutical compositions containing them

3-Methyl-4H-pyrido[3,2-e][1,2,4]thiadiazine 1,1-dioxide

AIMS
The present work describes the synthesis and the biological evaluation of novel compounds acting as pyruvate dehydrogenase kinase (PDK) inhibitors. These drugs should become a new therapeutic approach for the treatment of pathologies improved by the control of the blood lactate level.


METHODS
Four series of compounds belonging to N-(4-(N-alkyl/aralkylsulfamoyl)phenyl)-2-methylpropanamides and to 1,2,4-benzothiadiazine 1,1-dioxides were prepared and evaluated as PDK inhibitors.


RESULTS
The newly synthesized N-(4-(N-alkyl/aralkylsulfamoyl)phenyl)-2-methylpropanamides structurally related to previously reported reference compounds 4 and 5 were found to be potent PDK inhibitors (i.e. 10d: IC50 = 41 nM). 1,2,4-Benzothiadiazine 1,1-dioxides carrying a (methyl/trifluoromethyl)-propanamide moiety at the 6-position were also designed as conformationally restricted ring-closed analogues of N-(4-(N-alkyl/aralkylsulfamoyl)phenyl)-2-hydroxy-2-methylpropanamides. Most of them were found to be less potent than their ring-opened analogues. Interestingly, the best choice of hydrocarbon side chain at the 4-position was the benzyl chain, providing 11c (IC50 = 3.6 µM) belonging to "unsaturated" 1,2,4-benzothiadiazine 1,1-dioxides, and 12c (IC50 = 0.5 µM) belonging to "saturated' 1,2,4-benzothiadiazine 1,1-dioxides.


CONCLUSION
This work showed that ring-closed analogues of N-(4-(N-alkyl/aralkylsulfamoyl)phenyl)-2-hydroxy-2-methylpropanamides were less active as PDK inhibitors than their corresponding ring-opened analogues. However, the introduction of a bulkier substituent at the 4-position of the 1,2,4-benzothiadiazine 1,1-dioxide core structure, such as a benzyl or a phenethyl side chain, was allowed, opening the way to the design of new inhibitors with improved PDK inhibitory activity.

/pdf/design-synthesis-and-evaluation-of-novel-pyruvate-2i302vnz.pdf

Pascal De Tullio

Papers

Pharmacomodulation of torasemide led to original diuretic, neuroprotective, anticonvulsant and antithrombotic drugs

Novel dipyrazoles acting as glutamate modulators

Novel thiadiazine derivatives useful as AMPA receptor modulators, preparation method, and pharmaceutical compositions containing them

3-Methyl-4H-pyrido[3,2-e][1,2,4]thiadiazine 1,1-dioxide

Design, Synthesis, and Evaluation of Novel Pyruvate Dehydrogenase Kinase Inhibitors.