图1所示。发现命中化合物CPD1和化学先导HB007。(A) LN229细胞为基础的NCI文库药物筛选工作流程，通过Western blots和细胞活力测定，鉴定了11种活性化合物，其中D5为命中化合物(红色突出部分)。(B)命中化合物CPD1和先导化合物HB007的化学结构和药理特性。cLogP，分割系数log P;PSA，极表面积。(C) LN229细胞用指定剂量的CPD1处理72小时，并通过Western blots分析偶联和非偶联/游离SUMO1(如图所示)。分别以SUMO2/3和β-actin作为选择性和负载对照。(D) LN229细胞用CDP1处理72小时，用点状斑点法检测总SUMO1浓度，并显示负载蛋白的数量(右)。使用ImageJ(下)评估网点强度。(E和F) LN229细胞共转染myc - ubc9和YFP-SUMO1或YFP-SUMO3或空载体作为对照，并对UBC9-SUMO1 (E)和UBC9-SUMO3偶联物(F)(图右)进行myc IP和Western blotting，全细胞裂解液(WCL)作为加载对照。 (G) LN229 cells were treated with a series of dilutions of CPD1 or HB007 for 5 days and examined by cell viability for cell growth inhibition with the IC50 values indicated (points: n = 6). (H) HCT116 cells were treated with HB007 for the time indicated and analyzed by Western (left) and dot blots (right) for conjugated and total SUMO1 concentrations. (I) HCT116 cells were treated with DMSO (control), CPD1, or HB007 for 72 hours with the indicated doses (micromolar) and analyzed by Western blotting using the indicate antibodies (left). (J) LN229 cells were cotransfected with Flag-CDK6 and YFP-SUMO1, treated with HB007 for 24 hours, and subjected to Flag IP and Western blotting using CDK6 and green fluorescent protein (GFP) antibody (that recognizes YFP) for SUMO1-CDK6 conjugates as indicated (right). (K) Myc IP and Western blotting for SUMO1-UBC9 conjugates as indicated (right) in myc-UBC9– and YFP-SUMO1–transfected LN229 cells after CPD1 and HB007 treatment for 24 hours. Credit: DOI: 10.1126/scitranslmed.abh1486
