1. Volume 12, Issue 12, Pages 1337-1347 (December 2005)
Chemistry and Biology of Moverastins, Inhibitors of Cancer Cell Migration, Produced by Aspergillus 
Yasushi Takemoto, Hidenori Watanabe, Kenji Uchida, Koji Matsumura, Koichi Nakae, Etsu Tashiro, Kazutoshi Shindo, Takeshi Kitahara, Masaya Imoto 
Chemistry & Biology 
Volume 12, Issue 12, Pages (December 2005)
DOI: /j.chembiol
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2. Figure 1 Structural Determination of Moverastin
(A) Structure of moverastin.
(B) HMBC and DQF COSY correlations of moverastin.
Chemistry & Biology  , DOI: ( /j.chembiol )
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3. Figure 2 Synthesis of Diastereomeric Mixture of Moverastin
Reagents and conditions: (a) TsOH, HO(CH2)2OH, toluene reflux (at 70°C), quant; (b) O3, CH2Cl2-pyridine, then Me2S, 99%; (c) H2, Pd-C, EtOAc, quant; (d) KHMDS, 5-chloro-2-[N,N-bis(trifluoromethanesulfonyl)amino]pyridine, THF, −78°C, 64%; (e) n-BuLi, CH2=CHCH2Br, TMEDA, THF, −20°C∼room temperature, 60%; (f) n-BuLi, DMF, TMEDA, ether, −20°C∼room temperature, 54%; (g) O3, CH2Cl2, then Ph3P, 74%; (h) CrCl2, NiCl2, DMF, room temperature, 62%; (i) concentrated HCl, THF, room temperature, 82%.
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4. Figure 3 Separation of the Two Diastereomers of Moverastin
Reagents and conditions: (a) (R)-MTPACl, DMAP, CH2Cl2, room temperature, 81%, then HPLC separation; (b) NaOMe, MeOH, 0°C, then concentrated HCl, THF, 0°C, 69% for (10R)-moverastin (1), 61% for (10S)-moverastin (2).
Chemistry & Biology  , DOI: ( /j.chembiol )
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5. Figure 4 Effect of Moverastins A and B on Migration in EC17 Cells
(A) Wounding was introduced into confluent cultures of EC17 cells, as described in Experimental Procedures. The cells were incubated with various concentrations of moverastin A or moverastin B.
(B) EC17 cells were incubated with various concentrations of moverastin A (a) or moverastin B (b) in the top chamber for 24 hr. Then, the number of cells that migrated through the filter to the lower surface was counted. The results are the mean ± SD of five different fields.
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6. Figure 5 Effect of Moverastins A and B on FTase In Vitro
(A) Partially purified enzymes from EC17 cells were incubated with [3H]-FPP plus recombinant GST-H-Ras for FTase assay or [3H]-GGPP plus recombinant GST-Rho A for GGTase assay in the presence or absence of moverastin A or moverastin B. The reaction was terminated by the addition of TCA. The radioactivity of the TCA-insoluble fraction was measured. The results are the mean ± SD of four independent experiments.
(B) Partially purified enzymes from EC17 cells were incubated with [3H]-FPP plus recombinant GST-H-Ras for FTase assay or [3H]-GGPP plus recombinant GST-Rho A for GGTase assay in the presence or absence of moverastin A. The reaction was terminated by adding SDS sample buffer, and the product solution was subjected to SDS-PAGE and analyzed by autoradiography.
(C) Lineweaver-Burk plot of initial velocity versus GST-H-Ras concentration. Moverastin A concentrations are 0 (rhombus), 1 (square), 3 (triangle), and 10 (circle) μg/ml, respectively.
(D) Dixon plot of initial velocity versus moverastin A concentration. GST-H-Ras concentrations are 65 (rhombus), 217 (square), and 650 (triangle) μg/ml, respectively.
Chemistry & Biology  , DOI: ( /j.chembiol )
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7. Figure 6 Effect of Moverastin A on H-Ras Translocation in EC17 Cells
(A) EC17 cells were incubated with moverastin A for 24 hr. Proteins were extracted and separated into cytosolic and membrane fractions, and then H- or N-Ras were detected by immunoblotting as described in Experimental Procedures. Total cell lysate was immunoblotted with anti-H-Ras or anti-N-Ras antibody.
(B) EC17 cells were incubated with 10 μg/ml of moverastin A for the periods indicated. H-Ras in cytosolic and membrane fractions was detected by immunoblotting as described in Experimental Procedures. Total cell lysate was immunoblotted with anti-H-Ras, anti-phospho-Akt, anti-Akt, anti-phospho-Erk, or anti-Erk antibody. The data shown are representative of three independent experiments.
(C) EC17 cells were incubated with the indicated concentrations of LY or U0126 for 24 hr. Then, cells were collected, total cell lysate was prepared, and equal amounts of lysate were examined by Western blotting using anti-phospho-Akt, anti-Akt, anti-phospho-Erk, or anti-Erk antibody.
(D) EC17 cells were incubated with the indicated concentrations of LY or U0126 in the top chamber for 24 hr. The cells that migrated through the filter to the lower surface were counted. The results are the mean ± SD of five different fields.
Chemistry & Biology  , DOI: ( /j.chembiol )
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