Acetylation of NF-κB's RelA subunit at lysine-310 (AcLys310) helps to maintain constitutive NF-κB activity in cancers such as triple-negative breast cancer (TNBC). Bromodomain-containing factor BRD4 binds to acetylated RelA to promote the activity of NF-κB. Hence, interfering with the acetylated RelA-BRD4 interaction is a potential strategy for treating NF-κB-driven TNBC. Here, a new compound 13a was obtained by structural optimization and modification of our previously reported compound. In comparison with the well-known BRD4 inhibitor (+)-JQ1, 13a showed more potent anticancer activity in NF-κB-active MDA-MB-231 cells. Mechanistically, 13a antagonized the protein–protein interaction (PPI) between BRD4 and acetylated RelA, decreased levels of IL-6, IL-8, Snail, Vimentin, and ZEB1, induced cell senescence and DNA damage, and weakened the adhesion, metastasis, and invasion ability of TNBC cells. Our results provide insights into avenues for the further development of potent BRD4-acetylated RelA PPI inhibitors. Moreover, our findings highlight the effectiveness and feasibility of blocking the interaction between BRD4 and acetylated RelA against NF-κB-active cancers, and of screening antagonists of this PPI.