nf- κ b-inhibiting naphthopyrones from the fijian echinoderm comanthus parvicirrus

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NF- B-Inhibiting Naphthopyrones from the Fijian Echinoderm Comanthus parvicirrus. Florence Folmer, William T. A. Harrison, Jioji N. Tabudravu, Marcel Jaspars,*, William Aalbersberg, Klaus Feussner, Anthony D. Wright, Mario Dicato, and Marc Diederich - PowerPoint PPT Presentation

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  • NF-B-Inhibiting Naphthopyrones from the Fijian Echinoderm Comanthus parvicirrusFlorence Folmer, William T. A. Harrison, Jioji N. Tabudravu, Marcel Jaspars,*, William Aalbersberg, Klaus Feussner,Anthony D. Wright, Mario Dicato, and Marc Diederich

    Department of Chemistry, UniVersity of Aberdeen, Old Aberdeen, AB24 3UE, U.K., Institute of Applied Sciences, Faculty of Science and Technology, UniVersity of the South Pacific, P.O. Box 1168, SuVa, Fiji Islands, Australian Institute of Marine Science, TownsVille 4810, Queensland, Australia, and Laboratoire de Biologie Molculaire et Cellulaire du Cancer, Hpital Kirchberg, 9, Rue Edward Steichen, L-2540 Luxembourg, Luxembourg

    Received June 18, 2007 97.06.10

  • naphthopyrones 6-methoxycomaparvin (1)brown needle-shaped crystalsUV (MeOH) max (log) 245 (4.57), 290 (4.37), 380 (3.77) nmIR (KBr) max 3341, 2950, 1667, 1615, 1571, 1537, 1448 cm-1LRESIMS m/z 331 [M + H]+ (C18H18O6) HRESIMS m/z 331.1176 [M + H]+, (calc for C18H19O6)1H NMR (400 MHz, CDCl3) :6.23 (1H, s, H-3)7.01 (1H, d, J=1.6 Hz, H-7)6.43 (1H, d, J=1.6 Hz, H-9)6.23 (2H, t J=7.7 Hz, H-11)1.8 (2H, m, H-12)Methine (CH)Methylene (CH2)1.01 (3H, t, H-13)3.94 (3H, s, H-16)3.95 (3H, s, H-15)Methyl (CH3)12.90 (s, OH-5)

  • 13C NMR :170.1 (C-2)183.4 (C-4)147.1 (C-4a)109.3 (C-5)134.7 (C-6)158.3 (C-6a)104.9 (C-8)160.2 (C-10)136.1 (C-10a)152.4 (C-10b)109.5 (C-3)96.3 (C-7)96.7 (C-9)36.5 (C-11)20.0 (C-12)13.7 (C-13)60.4 (C-15)56.2 (C-16)

  • HMBC

  • COSY

  • single-crystal X-ray diffraction analysis

  • 6-methoxycomaparvin 5-methyl ether (2)brown needle-shaped crystalsUV (MeOH) max (log) 240 (4.58), 280 (4.51), 365 (4.05) nmIR (KBr) max 3229, 2960, 1642, 1584, 1415 cm-1LRESIMS m/z 345 [M + H]+ (C19H20O6) HRESIMS m/z 345.1337 [M + H]+, (calc for C19H21O6)1H NMR (400 MHz, CDCl3) :naphthopyrones 6-methoxycomaparvin (1)6-methoxycomaparvin 5-methyl ether (2)12.90 (s, OH-5)3.93 (s, H-14)

  • naphthopyrones 6-methoxycomaparvin (1)6-methoxycomaparvin 5-methyl ether (2)13C NMR :62.0 (C-14)

  • HMBC

  • Reporter Gene AssaysLuciferase Reporter Gene AssayFor rapid, sensitive determination of luciferase enzyme activity in transfected cells or in tissues isolated from transgenic animalsTransient transfections of K562 cells- pNF-BLuc K562 cells

  • Figure 1. Luciferase activity of pNF-BLuc K562 cells pretreatedfor 2 h with different concentrations (in g/mL) of 6-methoxycomaparvin(1) (A) or 6-methoxycomaparvin 5-methyl ether (2) (B),and treated for 2 h with 20 ng/mL TNF-.

  • Electrophoretic mobility shift assay (EMSA)Oligonucleotide (contain NF-B binding site )The probe was hybridized and labeled with[-32P]ATP(Contain p50/p65)

  • Figure 2. Inhibition of TNF-R-induced NF-B-DNA bindingactivity by 6-methoxycomaparvin (1) (A) and 6-methoxycomaparvin5-methyl ether (2) (B). For supershift/immunodepletion experiments, incubationwith 2 g of anti-p50, anti-p52, anti-p65, anti c-Rel, and anti-RelBantibodies (C).

  • Figure 3. Western blot analysis showing that 6-methoxycomaparvin(1) and 6-methoxycomaparvin 5-methyl ether (2) inhibit TNF- induceddegradation of IB and the consequent translocation ofp50 and p65 into the nucleus.

  • Figure 4. Effects of 6-methoxycomaparvin (1) and 6-methoxycomaparvin 5-methyl ether (2) on the kinase activity of IKK. No enzyme refers to a control in the absence of IKK.The negative control was performed in the presence of IKK, but in the absence of any test compound. Calbio IV ,which is an IKK inhibitor.

  • Figure 5. Effects of 6-methoxycomaparvin (1) and 6-methoxycomaparvin5-methyl ether (2) on the proteolytic activity of the 26S proteasome in K562 cells, at a concentration of 100 g/mL (300M). Control refers to a negative control without any test compound. Different concentrations of the known proteasome inhibitor MG132 (Z-Leu-Leu-Leu-CHO) were used as positive controls.

  • Both compounds inhibit the activation of the transcription factor NF-B, which plays an important role in cancer development and inflammation, and the mechanism of action of the two compounds was investigated.Both naphthopyrones 1 and 2 completely inhibit TNF--induced NF-B activation by inhibiting the enzymatic activity of the kinase IKK.