Salirasib (Farnesyl Thiosalicylic Acid)(Ki=2.6 μM), an effective competitive prenylated protein methyltransferase (PPMTase) inhibitor, inhibits Ras methylation with potential antineoplastic activity.

PPMTase:2.6 μM(Ki)

Salirasib 抑制人类 Ha-ras 转化的 Rat1 细胞生长，这与其对 PPMTase 的抑制作用密切相关。[1] Salirasib 在 Rat-1 成纤维细胞、Ras 转化的 Rat-1 细胞和 B16 黑色素瘤细胞中抑制 Ras 的甲基化。Salirasib 还降低细胞膜中 Ras 的水平，并独立于甲基化抑制 Ras 依赖的细胞生长，但通过调节 Ras-Raf 通讯达到这一点。[2] 在 Ras 转化的 EJ 细胞中，Salirasib 干扰 Raf-1 和 MAPK 的激活，并抑制 DNA 合成。[3]

在Panc-1移植的裸鼠中，Salirasib (5 mg/kg i.p.) 显著抑制肿瘤生长且无系统性毒性。[4] 在雄性Wistar大鼠中，Salirasib (5 mg/kg i.p.) 显著抑制硫代乙酰胺引起的肝硬化。[5] 在先天性肌肉营养不良症的dy(2J)/dy(2J)小鼠模型中，Salirasib (5 mg/kg i.p.) 减轻纤维化并改善肌肉力量。[6]

PPMTase Assays : Synaptosomal membranes of rat brain cerebellum or total membranes of cultured cell lines (100,000 × g pellet) are used for methyltransferase assays in the cell-free systems. Methyltransferase assays are performed at 37°C in 50 mM Tris-HCl buffer, pH 7.4, using 100 μg of protein, 25 μM [methyl-3H]AdoMet (300,000 cpm/nmol), and 50 μM AFC (prepared as a stock solution in DMSO) in a total volume of 50 μL. DMSO concentration in all assays is 8%. Various AFC concentrations are used in several experiments as indicated in the text. Reactions are terminated after 10 min by addition of 500 μL of chloroform:methanol (1:1) with subsequent addition of 250 μL of Water, mixing, and phase separation. A 125-μL portion of the chloroform phase is dried at 40°C, and 200 μl of 1 N NaOH, 1% SDS solution is added. The methanol thus formed is counted by the vapor phase equilibrium method. Typical background counts (no AFC added) are 50-100 cpm, while typical reactions with AFC yield 500-6,000 cpm. Assays are performed in triplicate, and background counts are subtracted. Methylation of endogenous substrates and gel electrophoresis are performed. Protein carboxyl methylation in intact cells is determined using 100 μCi/mL [methyl-3H]methionine. Cells are assayed in near confluent cultures grown in 10-cm plates with 5 mL of labeling medium.

Cells are grown in 24-well plates. 2 h after plating, the cells receive either solvent or FTS freshly prepared from a stock solution to yield the final indicated concentrations in 0.1% DMSO. Media are replaced every 4 days with fresh medium containing the solvent or the drug. Separate experiments indicate that the solvent itself has no effect on cell growth. On the indicated days, the cells are detached from plates by trypsin/EDTA and counted under the light microscope. All assays are performed in quadruplicate. In parallel experiments, cells are stained either with trypan blue or with MTT, and the stained cells are examined under the light microscope. In some MTT-stained cultures, the cells are dissolved in 0.2 mL of 100% DMSO, and the extent of staining is determined spectrophotometrically using an enzyme-linked immunosorbent assay reader.(Only for Reference)