SELECTIVE SENSITIZATION TO DNA-DAMAGING AGENTS IN A HUMAN RHABDOMYOSARCOMA CELL-LINE WITH INDUCIBLE WILD-TYPE P53 OVEREXPRESSION

Drug-induced cytotoxicity or apoptosis may be influenced by the expres sion of the p53 tumor suppressor gene and by the specific oncogene exp ressed, which may dictate the threshold at which a cytotoxic response may be induced. The objective of the study was to elucidate how DNA-da maging agents with different mechanisms of action were sensitized in t he context of expression of the Pax3/FKHR fusion protein, a transforma tion event unique to alveolar rhabdomyosarcomas (ARMSs), and wild-type p53 (wtp53). A wtp53 cDNA was subcloned into the pGRE5-2/EBV vector w ith dexamethasone-inducible overexpression and transfected into Rh30 A RMS cells that express Pax3/FKHR and a mutant p53 phenotype. Following dexamethasone induction of wtp53 overexpression in a derived clone (C l.#27), growth was slowed, and cells accumulated in G(1). Functional w tp53 activity was demonstrated by selective transactivation of p50-2, a wtp53 chloramphenicol acetyltransferase reporter construct, and by u p-regulated expression of endogenous p21(Waf1). Data demonstrated p53- dependent sensitization (greater than or equal to 4-fold) to bleomycin , actinomycin D, and 5-fluorouracil and considerably less p53-dependen ce (less than or equal to 2-fold) for doxorubicin, topotecan, etoposid e, and cisplatin in Cl.#27 compared to an equivalent clone containing the pGRE5-EBV vector alone (VC#3). Data demonstrate that ARMS cells sh ow a selective sensitization to DNA-damaging agents when wtp53 is over expressed. The cytotoxic activity of agents that are not potentiated s ubstantially must, therefore, depend upon p53-independent factors that relate to the mechanism of drug action.