Kabiramides, The Actin-Binding Trisoxazole Macrolides from the Thai marine sponge Pachastrissa Nux, and Semisynthetic Cluorescent Derivatives of Kabiramide C

Abstract

Three trisoxazole macrolides, including two known kabiramides C and D, and a new kabiramide F were isolated from the marine sponge Pachastrissa nux, collected from Sichang Island in the Gulf of Thailand. Their identification and structure elucidation were achieved by analyses of MS, UV, IR, 1D-NMR, and 2D-NMR spectral data as well as comparison with the literatures. The major compound, kabiramide C (KabC), was structurally modified to give the key intermediate 7-(4-aminomethyl-1H-1,2,3-triazol-1-yl) kabiramide C (AMT-KabC) by using Mitsunobu reaction and 1,3-dipolar cycloaddition reaction. Furthermore, five fluorescent conjugates of kabiramide C, including derivatives of tetramethylrhodamine (TMR-KabC), rhodol green (RG-KabC), IC5 (IC5-KabC), dapoxyl (DAP-KabC), and fluorescein diester (FDE-KabC) were synthesized by coupling AMT-KabC with N-succinimidyl esters of the fluorescence dyes. The steady state fluorescence techniques, including fluorescence emission spectral analysis, fluorescence resonance energy transfer (FRET), fluorescence anisotroupy (FA), and iodide quenching were used to determine the actin binding properties of these kabiramide derivatives. All of the isolated and semisynthetic fluorescent kabiramide derivatives bound stoichiometrically to G-actin in 1:1 complex. Five fluorescent kabiramides were further separately introduced to living NIH 3T3 cells. Only TMR-KabC and FDE-KabC showed good permeability through plasma membrane of the cells and expressed intense fluorescence at the protrusion sites. Additionally, kabiramides C, D, and F exhibited equal antifungal activity against Candida albicans with inhibition zone of 22 mm at the concentration of 100 µg/disc.