背景:蛋白质精氨酸甲基转移酶(PRMT)通过调节癌症中的精氨酸甲基化来调节蛋白质生物学活性,并且越来越被认为是潜在的药物靶标。针对PRMT的抑制剂目前处于临床试验的早期阶段,需要更多的候选药物。FlavokawainA(FKA),从卡瓦植物中提取,已被认为是膀胱癌(BC)的潜在化疗药物,但其作用机制尚不清楚。
方法:我们首先确定了II型PRMT的作用,PRMT5,在BC组织样本中进行细胞学实验。然后我们利用生物信息学工具,包括计算模拟,虚拟筛选,分子对接,和能源分析,确定PRMT5抑制剂用于BC治疗的潜在用途。进行体外和体内共IP和突变测定以阐明PRMT5抑制剂的分子机制。药理学实验,如生物层干涉法,CETSA,和下拉测定进一步用于提供复杂结合过程的直接证据。
结果:在PRMT中,PRMT5被鉴定为BC的治疗靶标。PRMT5在BC中的表达与不良预后相关,操纵其表达可能会影响癌细胞的生长。通过筛选和广泛的实验验证,我们认识到一种天然产物,FKA,是一种新的小的PRMT5抑制剂分子。我们注意到该产品可以抑制BC的作用,在体外和体内,通过抑制PRMT5。我们进一步证明了FKA通过与PRMT5的Y304和F580结合来阻断组蛋白H2A和H4的对称精氨酸二甲基化。
结论:总之,我们的研究强烈表明,PRMT5是膀胱癌潜在的表观遗传治疗靶点,FKA可作为PRMT5的靶向抑制剂用于膀胱癌的治疗。
BACKGROUND: Protein arginine methyltransferases (PRMTs) regulate protein biological activity by modulating arginine methylation in cancer and are increasingly recognized as potential drug targets. Inhibitors targeting PRMTs are currently in the early phases of clinical trials and more candidate drugs are needed. Flavokawain A (FKA), extracted from kava plant, has been recognized as a potential chemotherapy drug in bladder cancer (BC), but its action mechanism remains unclear.
METHODS: We first determined the role of a type II PRMT, PRMT5, in BC tissue samples and performed cytological experiments. We then utilized bioinformatics tools, including computational simulation, virtual screening, molecular docking, and energy analysis, to identify the potential use of PRMT5 inhibitors for BC treatment. In vitro and in vivo co-IP and mutation assays were performed to elucidate the molecular mechanism of PRMT5 inhibitor. Pharmacology experiments like bio-layer interferometry, CETSA, and pull-down assays were further used to provide direct evidence of the complex binding process.
RESULTS: Among PRMTs, PRMT5 was identified as a therapeutic target for BC. PRMT5 expression in BC was correlated with poor prognosis and manipulating its expression could affect cancer cell growth. Through screening and extensive experimental validation, we recognized that a natural product, FKA, was a small new inhibitor molecule for PRMT5. We noticed that the product could inhibit the action of BC, in vitro and in vivo, by inhibiting PRMT5. We further demonstrated that FKA blocks the symmetric arginine dimethylation of histone H2A and H4 by binding to Y304 and F580 of PRMT5.
CONCLUSIONS: In summary, our research strongly suggests that PRMT5 is a potential epigenetic therapeutic target in bladder cancer, and that FKA can be used as a targeted inhibitor of PRMT5 for the treatment of bladder cancer.