EPZ015666_PRMT5,EPZ015666是一种有效的PRMT5酶活性抑制剂，IC50为22 nM

体外

用EPZ015666处理MCL细胞系导致SmD3甲基化和细胞死亡的抑制，IC50值在纳摩尔范围内。EPZ015666，一种有效的肽竞争性和SAM合作抑制剂，相对于其他甲基转移酶，对PRMT5具有>10,000倍的特异性。

体内

EPZ015666是口服生物可利用的并且适合于体内研究。我们对携带皮下Z-138和Maver-1异种移植物的严重联合免疫缺陷（SCID）小鼠进行了21天功效研究，每日两次（BID）口服给药4个剂量组：每公斤25,50,100和200毫克体重（mg/kg）。在连续给药21天后，对动物实施，并分析血液和组织以确定甲基标记药效学和肿瘤生长抑制（TGI）之间的关系。在两种MCL模型中，EPZ015666在21天后显示剂量依赖性暴露和TGI。与载体治疗的肿瘤相比，在第21天测量的所有EPZ015666剂量组中的肿瘤显示出重量，体积和肿瘤生长的统计学显着差异。以200mg/kg BID给药诱导Z-138细胞中的肿瘤停滞，21天后具有>93％TGI，而Maver-1细胞显示>70％TGI。另外，使用Granta-519细胞系测试第三次MCL异种移植物，该细胞系是在第18天达到终点的快速生长模型，并且在200mg/kg组中显示出剂量依赖性功效，45％TGI。EPZ015666在所有三种模型中都具有良好的耐受性，在200 mg/kg剂量组中体重损失*小，没有其他临床观察

Kinase Assay

EPZ015666 is serially diluted threefold from 1,000 to 0.051 nM ａnd spotted into a 384-well polypropylene V-bottom microplate.3H-SAM is serially diluted twofold in assay buffer for a seven-point dilution series with a top concentration of 700 nM(final assay concentration).Reactions are initiated by the addition of 4 nM enzyme ａnd 40 nM peptide(final assay concentrations for both).Reactions are incubated for 60 min ａnd quenched by the addition of 10μL per well of 600μM unlabeled SAM in assay buffer(final assay concentration).For the peptide competition,EPZ015666 is serially diluted threefold from 1,000 to 0.051 nM ａnd spotted into a 384-well polypropylene V-bottom microplate.Peptide is serially diluted twofold in assay buffer for a seven-point dilution series with a top concentration of 480 nM(final assay concentration).Reactions are initiated by the addition of 4 nM enzyme ａnd 75 nM 3H-SAM(final assay concentrations for both).Reactions are incubated for 60 min,and the reactions are quenched by the addition of 10μL per well of 600μM unlabeled SAM in assay buffer(final assay concentration).MCE has not independently confirmed the accuracy of these methods.They are for reference only

Cell Assay

EPZ015666 is dissolved in DMSO ａnd stored,and then diluted with appropriate medium(final DMSO 0.2%)before use[1].

Cultured cells in linear/log-phase growth are split to a seeding density of 2×105 cells/mL in 2-20 mL of media,depending on the yield required at the end of the growth period.Compound is diluted in DMSO ａnd added to each culture vessel with a final DMSO concentration of 0.2%.Cells are allowed to grow for 96 h undisturbed.At the conclusion of each treatment period,cells are harvested by centrifugation(5 min,1,200 rpm),and cell pellets are rinsed once with PBS before being frozen on dry ice pending further processing.Long-term proliferation assays are performed on all MCL lines,with slight adjustments to initial seeding densities,depending on growth characteristics for each cell line.All assays are carried out for 12 d[1].MCE has not independently confirmed the accuracy of these methods.They are for reference only

更新时间：2024/1/2 10:10:55