Abstract
The goal of this study is to identify a novel inhibitor with anti-inflammatory and antiproliferative properties for the treatment of psoriasis. Compound f152A1 [(3S,5Z,8S,11E)-8,9,16-trihydroxy-14-methoxy-3-methyl-3,4,9,10-tetrahydro-1H-benzo[c][1]oxacyclotetradecine1,7(8H)-dione] was identified as the main active metabolite with strong inhibitory activity against tumor necrosis factor-α (TNFα) transcription in a fraction originated from the fermentation broth of the fungus Curvularia verruculosa. Although active in cell-based assays, f152A1 was unstable in plasma and liver microsome preparations, thus limiting its pharmaceutical utilization. To improve the metabolic properties of f152A1, a medicinal chemistry program was undertaken, resulting in the generation of over 400 analogs of f152A1. Eventually, E6201 [(3S,4R,5Z,8S,9S,11E)-14-(ethylamino)-8,9,16-trihydroxy-3,4-dimethyl-3,4,9,19-tetrahydro-1H-2-benzoxacyclotetradecine-1,7(8H)-dione] was identified as a promising analog in this series. In the present study, we characterized the in vitro activities of E6201 and discovered that the compound inhibits lipopolysaccharide-activated TNFα reporter activity in THP-1-33 cells with an IC50 value of 50 nM and selectively inhibits mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK)-1 and MEK kinase-1 in cell-free biochemical assays. In addition, E6201 showed inhibitory activity in several other cell-based systems: 1) phosphorylation of c-jun N-terminal kinase and p38 MAPKs; 2) nuclear factor-κB and activated protein-1 activation in various cell types; 3) interleukin (IL)-2 production from human lymphocytes; 4) hyperproliferation of human keratinocytes; 5) IL-8 production from human keratinocytes; and 6) proinflammatory cytokine production from human peripheral blood mononuclear cells. Based on the data presented here, E6201 may be beneficial for treatment of inflammatory and hyperproliferative diseases such as psoriasis through its anti-inflammatory activities on immune cells and antihyperproliferative activities on keratinocytes.
- TNFα, tumor necrosis factor-α
- MEK1, mitogen-activated protein kinase/extracellular signal-regulated kinase kinase-1
- MEKK1, MEK kinase-1
- BSA, bovine serum albumin
- MAPK, mitogen-activated protein kinase
- MKK, MAPK kinase
- ERK, extracellular signal-regulated kinase
- NF-κB, nuclear factor-κB
- IκBα, inhibitory protein of NF-κB
- IKK, IκB kinase
- AP-1, activator protein-1
- E6201, (3S,4R,5Z,8S,9S,11E)-14-(ethylamino)-8,9,16-trihydroxy-3,4-dimethyl-3,4,9,19-tetrahydro-1H-2-benzoxacyclotetradecine-1,7(8H)-dione
- f152A1, LL-Z1640-2, (3S,5Z,8S,9S,11E)-8,9,16-trihydroxy-14-methoxy-3-methyl-3,4,9,10-tetrahydro-1H-benzo[c][1]oxacyclotetradecine-1,7(8H)-dione
- f152D2 (hypothemycin), (1aR,3S,4S,9S,15bR,Z)-3,4,12-trihydroxy-14-methoxy-9-methyl-3,4,8,9-tetrahydro-1aH-benzo[c]oxireno[2,3-e][1]oxacyclotetradecine-5,11(2H,15bH)-dione
- f152B1, (3S,5E,8S,9S,11E)-8,9,16-trihydroxy-14-methoxy-3-methyl-3,4,9,10-tetrahydro-1H-benzo[c][1]oxacyclotetradecine-1,7(8H)dione
- f152C, (E)-8,9,16-trihydroxy-14-methoxy-3-methyl-3,4,5,6,9,10-hexahydro-1H-benzo[c][1]oxacyclotetradecine-1,7(8H)-dione
- ER-803064, (3S,4R,5Z,8S,9S,11E)-8,9,16-trihydroxy-14-methoxy-3,4-dimethyl-3,4,9,10-tetrahydro-1H-benzo[c][1]oxacyclotetradecine-1,7(8H)-dione
- IL, interleukin
- EGF, epidermal growth factor
- EGFR, EGF receptor
- FGFR1, fibroblast growth factor receptor 1
- VEGFR2, vascular endothelial growth factor receptor-2
- JNK, c-jun N-terminal kinase
- MBP, myelin basic protein
- PDGFRβ, platelet-derived growth factor receptor-β
- PKC, protein kinase C
- TAK1, tissue growth factor β-activated kinase-1
- FBS, fetal bovine serum
- PBMCs, peripheral blood mononuclear cells
- PLAP, placental alkaline phosphatase
- LPS, lipopolysaccharide
- PHA-P, phytohemagglutinin-P
- PMA, phorbol 12-myristate 13-acetate
- HRP, horseradish peroxidase
- GSH, glutathione
- GST, glutathione S-transferase
- ECL, enhanced chemiluminescence
- ELISA, enzyme-linked immunosorbent assay
- HTRF, homogeneous time-resolved fluorescence
- UTR, untranslated region
- 2-ME, 2-mercaptoethanol
- THP-1, a human acute monocytic leukemia cell line
- Eu (K)-PY20, europium cryptate-conjugated antiphosphotyrosine antibody, PY20
- Eu (K)-PT66, europium cryptate-conjugated antiphosphotyrosine antibody, PT66
- XL665-SA, streptavidin-conjugated-XL665
- Syk, spleen tyrosine kinase
- PGK, phosphoglycerine kinase
- GA-1000, gentamicin/amphotericin-B
- ZAP-70, 70-kDa ζ-associated protein.
Footnotes
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This work was supported by Eisai Co., Ltd.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.109.156554
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ABBREVIATIONS:
- Received May 23, 2009.
- Accepted August 13, 2009.
- © 2009 by The American Society for Pharmacology and Experimental Therapeutics
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