Novel Antileukemic Sterol Glycosides from Ajuga salicifolia

Two novel and three new sterol glycosides were isolated from the MeOH extract of the aerial parts of Ajuga salicifolia (L.) <SC>Schreber</SC>. The structures of the compounds were elucidated as (3R,16S,17S,20R,22S,23S, 24S,25S)-16,23 : 16,27 : 22,25-triepoxy-3-(β-<SC>D</SC>-glucopyranosyloxy)coprostigmast-7-en-17-ol (1), (3R,16S,17S, 20R,22S,23S,24S,25S)-16,23 : 16,27 : 22,25-triepoxy-3-{[β-<SC>D</SC>-glucopyranosyl-(1→2)-β-<SC>D</SC>-glucopyranosyl]oxy}coprostigmast-7-en-17-ol (2), (3R,16S,17R,20S,22R,24S,25S)-22,25-epoxy-3,27-bis(β-<SC>D</SC>-glucopyranosyloxy)coprostigmast-7-en-16-ol (3), (3R,16S,17R,20S,22R,24S,25S)-22,25-epoxy-3-{[β-<SC>D</SC>-glucopyranosyl-(1→2)-β-<SC>D</SC>-glucopyranosyl]oxy}-27-(β-<SC>D</SC>-glucopyranosyloxy)coprostigmast-7-en-16-ol (4), and (3R,16R,17S,20R,22S,23S, 24S,25S)-22,25-epoxy-3-(β-<SC>D</SC>-glucopyranosyloxy)coprostigmast-7-ene-16,17,23,27-tetrol 27-acetate (5) by means of 1D and 2D NMR spectroscopy and HR-MALDI mass spectrometry. The novel compounds, which consist of three additional ring systems at the coprostigmastane skeleton, were named ajugasalicioside A (1) and B (2), and the new compounds C (3), D (4) and E (5). In our cytotoxicity assays (HeLa cells, Jurkat T cells, and peripheral mononuclear blood cells), ajugasaliciosides A–D specifically inhibited the viability and growth of Jurkat T-leukemia cells at concentrations below 10 μ<SC>M</SC>. Ajugasalicioside A (1; (IC<INF>50</INF>=6 μ<SC>M</SC>) and C (3; IC<INF>50</INF>=3 μ<SC>M</SC>) were the most active compounds. Ajugasalicioside A (1) induced cell-cell contact, inhibited Jurkat T cell proliferation, and up-regulated mRNA levels of the cell-cycle regulator cyclin D1, which might be an indication for cell differentiation. Furthermore, 1 down-regulated the mRNA levels of the NF-κB subunit p65 in a concentration-dependent manner. These effects were not found for ajugasalicioside B (2), which has an additional glucose unit, and the onset of cytotoxicity of 2 (IC<INF>50</INF>=10 μ<SC>M</SC>) was delayed by 24 h.