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34548568306
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Siu, L. L; Carducci, M.; Pearce, L.; Maclean, M.; Sullivan, R.; Li, Z.; Kalita, A.; Chen, E. X.; Pili, R.; Longstreth, J.; Martell, R. E.; Reid, G. K. Abstracts of Posters, C77: Phase I Study of Isotype Selective Histone Deactylase (HDAC) Inhibitor MGCD0103 Given as Three-Times Weekly Oral Dose in Patients (pts) with Advanced Solid Tumors, AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics: Discovery, Biology, and Clinical Applications, Philadelphia, Pennsylvania, November 14-18, 2005.
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note
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2, and 0.1 mg/mL BSA. HDAC enzymatic activities were determined by the following procedure: 3× serial dilutions of a 10 mM solution of inhibitor were performed in DMSO followed by a 20× dilution into Assay buffer. 20 μl HDAC was preincubated with 5 μL diluted compound at RT for 10 min. The reaction was initiated by the addition of 25 μL of the appropriate substrate (HDACs 1, 2, 3, and 6: fluor-de-Lys substrate KI-104; HDAC8: fluor-de-Lys HDAC8 substrate KI-178), incubated 15 (HDAC8) or 60 (HDACs 1, 2, 3, and 6) minutes at 37 °C, before adding 50 μL of the appropriate development solution. The development solution for HDACs 1, 2, 3, and 6 was 167×-diluted 20× Developer Concentrate (BIOMOL: KI-105) plus 10 μM SAHA. For HDAC8, the development solution was 100×-diluted 5× Developer Concentrate (BIOMOL: KI-176) plus 10 μM SAHA. The assay was read in a VictorV plate reader (Perkin-Elmer, Wellesley, MA) at Ex 360 nm/Em 460 nm. The final substrate concentration was 30 μM and final HDAC concentrations in the reaction were 1 nM for HDACs 1, 3, and 6), 2 nM for HDAC2, and 15 nM for HDAC8. Carboxy-terminal FLAG-tagged human HDACs 1, 2, 3 (co-expressed with the domain of SMRT), and 6 were overexpressed in mammalian cells and affinity purified using an anti-Flag antibody matrix, eluted from the matrix with 100 μg/mL of a competing FLAG peptide in 20 mM Tris-Cl, pH 8.0, 150 mM NaCl, 10% glycerol, and protease inhibitor cocktail (Roche cat. # 1836153).
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34548594616
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note
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2, viable cells were quantitated using Vialight Plus (Cambrex) according to manufacturer's instructions.
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35
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34548584422
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note
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+ 446.2, exp. 446.2.
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36
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4744370522
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Heltweg B., Dequiedt F., Marshall B.L., Brauch C., Yoshida M., Nishino N., Verdin E., and Jung M. J. Med. Chem. 47 (2004) 5235
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Nishino, N.6
Verdin, E.7
Jung, M.8
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38
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3142562372
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HDAC8 crystal structure (1T64)
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HDAC8 crystal structure (1T64). Somoza J.R., Skene R.J., Katz B.A., Mol C., Ho J.D., Jennings A.J., Luong C., Arvai A., Buggy J.J., Chi E., Tang J., Sang B.-C., Verner E., Wynands R., Leahy E.M., Dougan D.R., Snell G., Navre M., Knuth M.W., Swanson R.V., McRee DE., and Tari L.W. Structure 12 (2004) 1325
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Structure
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Somoza, J.R.1
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Jennings, A.J.6
Luong, C.7
Arvai, A.8
Buggy, J.J.9
Chi, E.10
Tang, J.11
Sang, B.-C.12
Verner, E.13
Wynands, R.14
Leahy, E.M.15
Dougan, D.R.16
Snell, G.17
Navre, M.18
Knuth, M.W.19
Swanson, R.V.20
McRee, DE.21
Tari, L.W.22
more..
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39
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3142562372
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HDAC8 crystal structure (1T64)
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HDAC8 crystal structure (1T64). Somoza J.R., Skene R.J., Katz B.A., Mol C., Ho J.D., Jennings A.J., Luong C., Arvai A., Buggy J.J., Chi E., Tang J., Sang B.-C., Verner E., Wynands R., Leahy E.M., Dougan D.R., Snell G., Navre M., Knuth M.W., Swanson R.V., McRee DE., and Tari L.W. Structure 12 (2004) 1325
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Structure
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Somoza, J.R.1
Skene, R.J.2
Katz, B.A.3
Mol, C.4
Ho, J.D.5
Jennings, A.J.6
Luong, C.7
Arvai, A.8
Buggy, J.J.9
Chi, E.10
Tang, J.11
Sang, B.-C.12
Verner, E.13
Wynands, R.14
Leahy, E.M.15
Dougan, D.R.16
Snell, G.17
Navre, M.18
Knuth, M.W.19
Swanson, R.V.20
McRee, DE.21
Tari, L.W.22
more..
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40
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0033539092
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HDLP crystal structure (1C3S)
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HDLP crystal structure (1C3S). Finnin M.S., Donigian J.R., Cohen A., Richon V.M., Rifkind R.A., Marks P.A., Breslow R., and Pavletich N.P. Nature 401 (1999) 188
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Finnin, M.S.1
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Marks, P.A.6
Breslow, R.7
Pavletich, N.P.8
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41
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0034567210
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Comparative protein structure modeling
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An alignment of the two crystal structures and the amino acid sequence for hHDAC1 (retrieved from SwissProt, accession ID: Q13547) was accomplished using the sequence/structure alignment tools in MOE (Chemical Computing Group, Montreal, Canada). This alignment was then used in Quanta/Modeler5 (Accelrys, San Diego, CA, USA) to generate a set of homology models. That with the least number of violations was further energy minimized using CHARMm 200 steps steepest descents without the catalytic Zn or any waters present:. Webster D.M. (Ed), Humana Press, New Jersey
-
An alignment of the two crystal structures and the amino acid sequence for hHDAC1 (retrieved from SwissProt, accession ID: Q13547) was accomplished using the sequence/structure alignment tools in MOE (Chemical Computing Group, Montreal, Canada). This alignment was then used in Quanta/Modeler5 (Accelrys, San Diego, CA, USA) to generate a set of homology models. That with the least number of violations was further energy minimized using CHARMm 200 steps steepest descents without the catalytic Zn or any waters present:. Sánchez R., and Sali A. Comparative protein structure modeling. In: Webster D.M. (Ed). Protein Structure Prediction: Methods and Protocols Vol. 143 (2000), Humana Press, New Jersey 97-129
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Protein Structure Prediction: Methods and Protocols
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Sali, A.2
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84986512474
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44
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10444230239
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Distance Geometry and Molecular Conformation
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The 150 conformers were generated and energy minimized using MMFFs with a distance-dependent dielectric of 2r:. Bawden D. (Ed), Research Studies Press, Wiley, New York
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The 150 conformers were generated and energy minimized using MMFFs with a distance-dependent dielectric of 2r:. Crippen C.M., and Havel T.F. Distance Geometry and Molecular Conformation. In: Bawden D. (Ed). Chemometrics Research Studies Series (1988), Research Studies Press, Wiley, New York
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(1988)
Chemometrics Research Studies Series
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Crippen, C.M.1
Havel, T.F.2
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46
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34548596012
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note
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During the course of energy minimization within the context of the enzyme model, only slight differences in the presentation of the co-minimizing residues (those within 5 Å of the inhibitor) were noted between the starting and final enzyme structure. The catalytic Zn and chelating residues were virtually unchanged during the minimization process.
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34548589609
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2 for 24 hours with either 0.2% DMSO or increasing concentrations of 7i dissolved in DMSO (final DMSO concentration = 0.2%). Whole cell lysates were made by disrupting cell membranes with 2% deoxycholate followed by sonication. Using these HCT116 lysates as antigen, H2BK5 acetylation was quantified using an indirect ELISA and normalized to total H2B levels obtained using a second indirect ELISA. Primary antibodies used were rabbit anti-acetylated H2BK5 (Cell Signaling) and sheep anti-histone H2B (Abcam). Goat anti-rabbit IgG HRP (Bio-Rad) and rabbit anti-sheep-HRP (Jackson ImmunoResearch) were used as secondary antibodies, respectively. TMB substrate (Pierce) was used for detection with absorbance read at 450 nm.
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