Clofazimine, a riminophenazine antimicrobial agent, and its analogue B669 were investigated for their effects on FaDu cells, a human squamous carcinoma cell line. These agents, at concentrations within the therapeutic range (0.25–2 µg/ml), caused a dose-dependent tumor cell cytotoxicosis which was greatly enhanced in the presence of human neutrophils. The neutrophil-mediated increment in tumoricidal activity, but not the direct antitumor effects of the drugs per se, was inhibited by catalase. The effects of these drugs on three more cell carcinoma lines as well as on two primary cultures and a noncarcinoma cell line were also investigated and compared with the activity of the standard antitumor chemotherapeutic agents bleomycin, cisplatin, and methotrexate. All seven cultures were sensitive to clofazimine and B669 compared to six that were sensitive to cisplatin, three that were sensitive to bleomycin, and one that was sensitive to methotrexate. The treatment of FaDu cells with clofazimine and B669 was associated with enhanced activity of phospholipase A₂, as evidenced by increased release of radiolabeled arachidonate and lysophosphatidylcholine from membrane phospholipids. Inhibitors of arachidonic acid metabolism, protein kinase C inhibitors, as well as water and lipid soluble antioxidants failed to protect the cells against the cytotoxic activity of clofazimine and B669. However, α-tocopherol, a lysophospholipid-complexing agent, completely blocked the antiproliferative effects of the riminophenazines and also protected the cells against the direct cytotoxic effect of lysophosphatidylcholine, while the lysophospholipid-neutralizing enzyme lysophospholipase protected against the riminophenazines. These observations demonstrate that the tumoricidal properties of clofazimine and B669 are probably due to increases in the lysophospholipid content of cell membranes.