IDENTIFICATION OF DIPHTHERIA-TOXIN VIA SCREENING AS A POTENT CELL-CYCLE AND P53-INDEPENDENT CYTOTOXIN FOR HUMAN PROSTATE-CANCER THERAPEUTICS

BACKGROUND. Metastatic human prostate cancer requires novel therapeuti c strategies in order to overcome its low proliferative rate and its r esistance to conventional chemotherapeutic agents. To identify potenti al cytotoxin gene products for use in experimental therapeutics such a s in vivo gene therapy, an in vitro screen was designed. METHODS. Eigh t recombinant cellular toxins were tested for activity against a spect rum of metastatic human prostate cancer cell phenotypes. Pseudomonas e xotoxin A, ricin, tumor necrosis factor alpha (TNF-alpha), diphtheria toxin (DT), Crotalus durissus terrificus toxin, crotalus adamenteus to xin, Naja naja toxin, and Naja mocambique toxin were evaluated. Compar ative survival distinguished the relative potencies of these cytotoxin s for irreparable prostate cancer cell death. RESULTS. Of the phosphol ipase A2 toxins, Crotalus durissus terrificus and Naja mocambique are active against the PSA secreting LNCaP cell line; however, the effect is reversible, and no other hormone refractory prostate cell line test ed is sensitive. Screening identified toxin-specific differences: dose -dependent cytotoxic activity against all human prostate cancer cell l ines tested was only identified for ricin and diphtheria toxin (DT) as highly potent. DT has an IC50 in the range of 20-00 pM by clonogenic survival and kills irreversibly by both apoptosis as well as nonapotot ic pathways. Acquisition of p53 mutant status conferred no reduction i n sensitivity to DT cytotoxicity. Cell cycle arrest by aphidicolin did not protect human prostate cells from irreversible DT-induced cell de ath. TNF-alpha had modest cytostatic activity in the screen; however, the combination of TNF-alpha and DT resulted in marked acceleration of the time to prostate cancer cell death. CONCLUSIONS. The rational scr eening of cytotoxins allows the identification of cell cycle-independe nt agents of variable potency against human prostate cancer. DT-mediat ed cell death is cell cycle independent, and p53 independent, making i t particularly attractive for application to cytoreductive gene therap y, targeted monoclonal antibodies, and prodrug delivery of toxins appl ied to human prostate cancer therapeutics. (C) 1998 Wiley-Liss, Inc.