This study reports the synthesis, characterization, computational analysis, and in vitro anticancer activity of novel heteroleptic copper(II) coordination compounds, [Cu(phen)(ipbi)(H2O)](NO3) (1) and [Cu2(phen)2(bisipbi)(H2O)2](NO3)2 (2). Here, phen: represents 1,10-phenanthroline, ipbi refers to 2-(imidazo[1,2-a]pyridin-8yl)benzimidazole (L1), and bis-ipbi (L2) denotes its bis-derivative. Compounds 1 and 2 are five-coordinate copper(II) complexes, where the metal centers bind to the deprotonated nitrogen atom of the benzimidazole ring, the tertiary nitrogen atom of the imidazopyridine ring, and the nitrogen atoms of the phenanthroline ligands. A water molecule occupies the fifth coordination site in each complex. The biological studies revealed that both compounds exhibit DNA binding through groove interactions, DNA cleavage activity, transcription inhibition, and significant cytotoxic activity. Compounds 1 and 2 demonstrated superior inhibitory activity against various cancer cell lines compared to the positive control (Cisplatin) and the parent complex [Cu(phen)2(H2O)]2+. Specifically, compound 1 showed the most potent IC50 value of 1.23 mu M against the triple-negative breast cancer cell line MDA-MB-231. The IC50 values for MDA-MB-231 were 1.23 mu M (1) and 1.28 mu M (2), compared to 2.77 mu M for the reference compound. Similar enhancements were observed against MCF-7 (10.59 and 12.56 mu M vs. 9.72 mu M), HepG2 (2.57 and 2.77 mu M vs. 30.38 mu M), DLD-1 (5.25 and 5.57 mu M vs. 60.79 mu M), and Beas-2B (6.10 and 12.99 mu M vs. 11.16 mu M). Computational and experimental studies, including UV-vis absorption and FTIR spectroscopy, revealed good agreement. Notably, complex 2 exhibited the smallest chemical hardness and highest electrophilicity, indicating a greater likelihood of electronic transitions. These findings suggest that complexes 1 and 2 are promising candidates for further exploration as anticancer agents.