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目的:评价以耻垢分枝杆菌(Msm)进行抗结核药物筛选的可行性与局限性,并应用该菌快速筛选新型抗结核药物先导物。方法:测定抗结核药物对Msm的最小抑菌浓度(MIC);用基于微孔板的高通量方法筛选抑制Msm和结核分枝杆菌(MTB)的化合物;用MTT法测定抗结核化合物的细胞毒性。结果:靶向于分枝菌酸和蛋白质合成、能量和核酸代谢的药物,在抑制Msm和MTB方面具有平行性;从7万余样品中筛选到5个对MTB(H37Rv)的MIC小于10μg·mL-1的化合物;其中的IMBYH2232为喹啉胺类化合物,对MTB(H37Rv)的MIC为0.125μg·mL-1,该化合物对巨噬细胞J774A.1、人肝细胞L02和人肾上皮细胞293T的ID50分别为12.5,15.2和10.5μg·mL-1。结论:本研究确定了Msm高效快速表型筛选模型的可行性与局限性,发现了具有极强的抗结核活性的化合物。
OBJECTIVE: To evaluate the feasibility and limitations of anti-TB drug screening with M. smegmatis (Msm) and to rapidly screen novel anti-TB drug precursors using this bacterium. Methods: The minimum inhibitory concentration (MIC) of anti-tuberculosis drugs against Msm was determined. The compounds that inhibit Msm and MTB were screened by microplate-based high-throughput method. The anti-tuberculosis compounds were determined by MTT assay toxicity. RESULTS: The drugs targeted to mycolic acid and protein synthesis, energy and nucleic acid metabolism had parallel inhibition of Msm and MTB. Five MICs out of 70k samples were screened for MTB (H37Rv) less than 10 μg · mL -1; IMBYH2232 is a quinolinamine compound, and the MIC for the MTB (H37Rv) is 0.125 μg · mL-1, and the compound has no effect on macrophage J774A.1, human hepatocyte L02 and human renal epithelial cells The ID50 of 293T was 12.5, 15.2 and 10.5μg · mL-1, respectively. Conclusion: This study identified the feasibility and limitations of a rapid and efficient screening phenotype for Msm and found a compound with strong anti-TB activity.