The present study evaluates whether the in vitro activity of immunotoxins can be enhanced by verapamil or by various antagonists of calmodulin (dansylcadaverine, trifluoperazine, chlorpromazine). The following immunotoxins made with either Pseudomonas exotoxin (PE), recombinant ricin A chain (rRTA), or ricin A chain (RTA) were used: HB21-PE and 454A12-rRTA that both recognize the human transferrin receptor; and 260F9-rRTA and 454C11-RTA that both react with human ovarian and breast cancer cells. The cytotoxicity of these immunotoxins was determined in human ovarian carcinoma cell lines and KB cells. Verapamil, that was demonstrated previously to enhance the cell-killing activity of PE immunotoxins, enhanced the activity of several ricin A chain immunotoxins, including 454A12-rRTA, 260F9-rRTA, and 454C11-RTA. Comparing 50% inhibitory dose values for inhibition of protein synthesis by 454A12-rRTA, enhancement ranged from 2- to greater than 25-fold, was dependent on the concentration of verapamil, and was greatest at short incubation times. In addition, the cytotoxicity of HB21-PE and of selected RTA immunotoxins was increased up to 30-fold by the addition of various calmodulin antagonists. The enhancing drugs did not decrease the specificity of the immunotoxins.