A series of novel naphthalimide derivatives with 4-[4-(3,3-diphenylallyl)piperazin-1-yl]benzoic acid as side chain were designed and synthesized. Their antitumor activities were evaluated against a variety of cancer cell lines in vitro. Preliminary results showed that most of the derivatives had cytotoxic activity comparable with that of amonafide, with IC50 values of 10−6–10−5 m. Interestingly, compound 12e had the unique antitumor activity against MCF-7 among the cancer cell lines tested. More importantly, flow cytometric analysis indicated that compared with amonafide, the target compounds could effectively induce G2/M arrest and progress to apoptosis in HL-60 cells after double staining with annexin V–FITC and propidium iodide. The present work provided a novel class of naphthalimide-based derivatives with potential apoptosis-inducing and improved antitumor activity for further optimization.