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Response of Cultured Human Cell Lines from Rhabdomyosarcoma Xenografts to Treatment with Chloroethylnitrosoureas¹

Department of Biochemical and Clinical Pharmacology, St. Jude Children's Research Hospital, Memphis, Tennessee 38101

DNA interestrand cross-links are thought to be the cytotoxic lesion resulting from treatment of cells with the chloroethylnitrosoureas (CENUs). We showed in an earlier study that the resistance of xenografts of pediatric rhabdomyosarcoma to therapy with CENUs correlates with levels of O⁶-alkylguanine-DNA alkyltransferase. We now demonstrate a relationship between levels of the alkyltransferase and CENU-induced cytotoxicity and DNA-interstrand cross-link formation in two cell lines recently established from such rhabdomyosarcoma xenografts. Rh18 cells were derived from the HxRh18 xenograft line, which contains the alkyltransferase and is relatively resistant to CENUs, and Rh28 cells were derived from the HxRh28 xenograft line, which lacks detectable alkyltransferase activity and is sensitive to treatment with the CENUs. In vitro, Rh28 cells were 5- to 6-fold more sensitive to growth inhibition by 1-(2-chloroethyl)-1-nitrosourea, chlorozotocin, and 1,3-bis(2-chloroethyl)nitrosourea than Rh18 cells. Extracts of Rh18 cells contained 3.8 units of the alkyltransferase per mg of protein, whereas such activity was undetectable in Rh28 cells, a unit of the alkyltransferase being defined as 1 pmol of [³H]methyl transferred from [³H]methyl-labeled DNA to protein. DNA interstrand cross-links, measured by alkaline elution 6 h after a 1-h pulse treatment with CENU, could not be detected in Rh18 cells but were found in the Rh28 line. The phenotypes of the parental xenograft lines defined by their alkyltransferase levels and by responses to CENU therapy of the mice have clearly been retained in the cultured cell lines, and as predicted, cross-link formation was inhibited in the alkyltransferase-containing Rh18 cells. These two new cell lines thus provide a useful model for studying the role of DNA repair in rhabdomyosarcoma resistance to these alkylating agents.

¹ Supported by NIH Awards CA 36888, CA 21765, and CA 23099 and by American Lebanese Syrian Associated Charities.

² Present address: Department of Chemistry, University of Texas at Austin, Austin, TX 78712-1096.

³ To whom requests for reprints should be addressed.

Received 8/30/88. Revised 11/ 7/88. Accepted 11/15/88.