ML-323 is a highly reversible and highly effective selective inhibitor of USP1-UAF1 with an IC50 value of 76nM. The Ki value of ML-323 for free enzymes is 68nM. USP1-UAF1 is closely related to the DNA damage response and can be used for the study of anti-tumor drug resistance [1]. ML-323 has anti-cancer activity against various cancers [2-3].
In vitro, in the Ub-Rho experiment, ML-323 (0.08-114μM; 1-2h) exhibited dose-dependent inhibition of USP1-UAF1 with an IC50 value of 76nM [1]. ML-323 (0-200μM; 48h) inhibited the growth of HCCLM3 and SMMC-7721 HCC cells and the formation of colonies in a dose-dependent manner [4]. ML-323 (0-400nM; 0, 4, 8 and 16h) could down-regulate the expression of USP1 in colorectal cancer cells in a dose-dependent and time-dependent manner [5].
In vivo, the injection treatment of ML-323 (10mg/kg; 7 days) significantly increased the body weight of mice with colitis models and decreased the MPO activity in colonic tissues and ulcer areas, and had a significant inhibitory effect on the levels of inflammatory markers (TNF-α, IL-6, IL-17 and IL-1β) in the colonic tissues of colitis mice [6]. ML-323 (25 and 50mg/kg/day; oral; twice a week for 8 weeks) treatment significantly reduced the body weight of mice fed a high-fat diet (HFD), improved insulin and glucose sensitivity, and significantly decreased the fat content and size of adipocytes in white adipose tissue, reduced lipid accumulation, triglycerides, free fatty acids and macrophage infiltration in the liver of the mice [7].
References:
[1] Liang Q, Dexheimer T S, Zhang P, et al. A selective USP1–UAF1 inhibitor links deubiquitination to DNA damage responses[J]. Nature chemical biology, 2014, 10(4): 298-304.
[2] Sun Y, Sha B, Huang W, et al. ML323, a USP1 inhibitor triggers cell cycle arrest, apoptosis and autophagy in esophageal squamous cell carcinoma cells[J]. Apoptosis, 2022, 27(7): 545-560.
[3] Song B, Jiang Y, Jiang Y, et al. ML323 suppresses the progression of ovarian cancer via regulating USP1-mediated cell cycle[J]. Frontiers in Genetics, 2022, 13: 917481.
[4] Wang L, Hu T, Shen Z, et al. Inhibition of USP1 activates ER stress through Ubi-protein aggregation to induce autophagy and apoptosis in HCC[J]. Cell Death & Disease, 2022, 13(11): 951.
[5] Xu X, Mei X, Han K, et al. The deubiquitinating enzyme USP1 is auto-ubiquitinated and destabilized by ML323 in colorectal cancer cells[J]. Eurasian J Med Oncol, 2023, 7: 174-179.
[6] Lai Y, Liu J, Hu X, et al. N6-methyladenosine (m6A)-forming enzyme METTL3 controls UAF1 stability to promote inflammation in a model of colitis by stimulating NLRP3[J]. Scientific Reports, 2025, 15(1): 5876.
[7] Kim M S, Baek J H, Lee J A, et al. Deubiquitinase USP1 enhances CCAAT/enhancer-binding protein beta (C/EBPβ) stability and accelerates adipogenesis and lipid accumulation[J]. Cell Death & Disease, 2023, 14(11): 776.
ML-323是一种可逆性强、高效的USP1-UAF1的选择性抑制剂，IC50值为76nM。ML-323对游离酶的Ki值为68nM。USP1-UAF1与DNA损伤反应密切相关，可用于抗肿瘤耐药性的研究 [1]。ML-323对多种癌症具有抗癌活性 [2-3]。
在体外，在Ub-Rho实验中，ML-323（0.08-114μM; 1-2h）对USP1-UAF1剂量依赖性抑制，其IC50值为76nM [1]。ML-323（0-200μM; 48h）以剂量依赖性方式抑制HCCLM3和SMMC-7721 HCC细胞的生长和菌落形成 [4]。ML-323（0-400nM; 0, 4, 8和16h）能够以剂量依赖性和时间依赖性方式下调结直肠癌细胞中USP1的表达 [5]。
在体内，ML-323（10mg/kg; 7d）的注射治疗显著增加了结肠炎模型小鼠的体重并降低结肠组织和溃疡区域的MPO活性，对结肠炎小鼠结肠组织中炎症标志物（TNF-α、IL-6、IL-17和IL-1β）水平有显著抑制作用[6]。ML-323（25 and 50mg/kg/day; oral; twice a week, 8 weeks）治疗显著减轻高脂饮食（HFD）小鼠体重以及改善胰岛素和葡萄糖敏感性，并且显著降低白色脂肪组织中的脂肪量和脂肪细胞大小，减少了小鼠肝脏中的脂质积累、甘油三酯、游离脂肪酸和巨噬细胞浸润 [7]。