c-Kit蛋白是涉及多种信号通路的信号转导蛋白，对细胞增殖、凋亡和分化等多种细胞事件起重要作用。c-Kit基因启动子上的特殊DNA二级结构，包括G四链体和i-结构，可以作为基因转录调控的“分子开关”，是发展新型抗癌药物的潜在重要靶点。我们通过SPR、FRET、CD、MST、NMR、双荧光素酶报告试验、Western blot、qPCR、免疫荧光、MTT试验、克隆形成、细胞刮擦、细胞凋亡、细胞周期分析和Transwell试验等多项实验筛选并评估了化合物对c-Kit的影响。经过广泛的筛选，我们发现比撒喜啶衍生物B05对c-Kit基因启动子上的双i-结构具有选择性结合和稳定作用，可以下调c-Kit基因的转录和翻译，从而抑制细胞增殖和转移。B05在HGC-27细胞上表现出强效的抗肿瘤活性，并在HGC-27移植瘤小鼠模型中明显抑制肿瘤生长。B05可以与c-Kit启动子上的双i-结构发生卓越的选择性相互作用，从而使选择性调节基因的转录和翻译成为可能。B05可进一步开发成为靶向c-Kit启动子i-结构的选择性抗癌剂。不同原癌基因启动子上的i-结构具有多样性，在同一启动子上同时结合双i-结构能够显著下调基因转录，并降低剂量，从而提高选择性。这一新策略为开发选择性DNA靶向配体照亮了道路。版权所有 © 2023 Elsevier B.V.

c-Kit protein is a signal transduction protein involved in multiple signal pathways, which play an important role in a variety of cellular events such as cell proliferation, apoptosis and differentiation. Special DNA secondary structures on the promoter of c-Kit gene, including G-quadruplex and i-motif structures, could act as "molecular switch" for gene transcriptional regulation, which are potentially important target for development of new anti-cancer drugs.We screened and evaluated the effect of compounds on c-Kit through several experiments, including SPR, FRET, CD, MST, NMR, dual-luciferase reporter assay, Western blot, qPCR, immunofluorescence, MTT assay, colony formation, cell scrape, cell apoptosis, cell cycle analysis, and transwell assay.After extensive screening, we found that bisacridine derivative B05 had selective binding and stabilization to dual i-motif structures on c-Kit gene promoter, which could down-regulate c-Kit gene transcription and translation, resulting in inhibition of cell proliferation and metastasis. B05 exhibited potent anti-tumor activity on HGC-27 cells, and strongly suppressed tumor growth in HGC-27 xenograft mice model.B05 could interact with c-Kit promoter dual i-motif structures with excellent selectivity, which make it possible for selective regulation of gene transcription and translation. B05 could be further developed for selective anti-cancer agent targeting c-Kit promoter i-motifs.i-Motifs on different proto-oncogene promoters are diversified, and especially binding of dual i-motifs on the same promoter simultaneously could significantly down-regulate gene transcription with decreased dosage, and therefore increasing the selectivity. This new strategy shed bight light on development of selective DNA-targeting ligands.Copyright © 2023 Elsevier B.V. All rights reserved.