1. Volume 17, Issue 6, Pages 659-664 (June 2010)
Chemical Proteomics Identifies Nampt as the Target of CB30865, An Orphan Cytotoxic Compound 
Tracey C. Fleischer, Brett R. Murphy, Jeffrey S. Flick, Ryan T. Terry-Lorenzo, Zhong-Hua Gao, Thaylon Davis, Rena McKinnon, Kirill Ostanin, J. Adam Willardsen, J. Jay Boniface 
Chemistry & Biology 
Volume 17, Issue 6, Pages (June 2010)
DOI: /j.chembiol
Copyright © 2010 Elsevier Ltd Terms and Conditions

2. Figure 1 CB30865 Analogs Interact with Nampt
(A) Quinazolinone structures.
(B) Viability assay. Mean ATP levels of two replicates normalized to a vehicle control are plotted with the standard deviation.
Chemistry & Biology  , DOI: ( /j.chembiol )
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3. Figure 2 MPI-0479883 Interacts with Nampt
(A) MPI affinity purification. SDS-PAGE gel showing pull-downs with EA- or MPI beads in the presence (+) or absence (−) of 10 μM uncoupled MPI (Block).
(B) LC-MS/MS analysis of pull-downs. The heat map indicates the total number of spectra corresponding to the listed proteins.
(C) α-Nampt immunoblot of pull-downs. Lanes are the same as in (A).
(D) Ruby-stained gel showing MPI pull-downs blocked with vehicle (−), free MPI (3p), or MPI (2p).
(E) Ruby-stained gel and α-Nampt immunoblot of pull-downs done with MPI or MPI in the presence (+) or absence (−) of 10 μM of their respective free compound (Block). Density of compound was varied by coupling at the concentrations indicated.
Chemistry & Biology  , DOI: ( /j.chembiol )
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4. Figure 3 MPI-0479883 Inhibits Nampt Activity In Vitro and In Vivo
(A) Schematic of the in vitro Nampt assay.
(B) Dose response of MPI in the Nampt assay. Graphs indicate average Nampt activity (resorufin fluorescence) normalized to a vehicle control. The standard deviation between duplicate samples is shown.
(C) PAR immunofluorescence assay. Cells treated with vehicle or MPI and H2O2 where indicated were stained with α-PAR antibody and Hoechst to mark nuclei.
(D) The PAR intensity for cells treated with MPI in the presence or absence of NA was quantified and the mean from duplicate experiments plotted with the standard deviation.
(E) NAD levels in cells treated with indicated amounts of MPI in the presence (NA) or absence (vehicle) of 10 μM nicotinic acid. The mean and the standard error are shown.
(F) Cytotoxicity of MPI in the absence (vehicle) or presence (NA) of 10 μM nicotinic acid. Mean ATP levels of two replicates normalized to a vehicle control are plotted with the standard deviation.
Chemistry & Biology  , DOI: ( /j.chembiol )
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