Summary
Cyclophosphamide (CPA) is widely used against leukemic and lymphoproliferative diseases, but in vitro studies on response to this agent so far have been limited to instable derivatives with poor galenic properties. ASTA Z 7557 is a newly synthesized “activated cyclophosphamide” that circumvents the need for hepatic activation and has good stability. The critical cytotoxic lesions after exposure to bifunctional alkylating agents presumably are DNA interstrand crosslinks (ISC). We have, therefore, examined the formation and apparent removal of ISC after in vitro treatment with ASTA Z 7557 by use of the highly sensitive alkaline elution technique. Survival of murine L1210 cells was determined after 1 hour in vitro exposure with a D 37 value of 5.7 μg/ml (from the initial shoulder part of the survival curve) and a Do value of 1.5 μg/ml (from the exponential part of the curve). Previous labelling of L1210 cells by 125IUdR simplified the alkaline elution procedure but there was some cytotoxicity of the radiochemical itself with a reduction of cloning efficiency from 77% to 61 %. The maximum of ISC was observed at 6 h after initiation of treatment with much of the damage apparently removed at 24 h. The simultaneous presence of DNA single strand breaks (SSB), however, confounds the analysis of DNA damage at 24 h and early cytolysis and unaided death of human lymphocytes often preclude the analysis of macromolecular damage at this time. Human peripheral blood cells isolated from patients with leukemic or lymphoproliferative diseases showed a remarkable heterogeneity with regard to the formation of ISC at 3 h. Thus, analysis of macromolecular damage may become an additional prognostic factor for response to CPA beyond the morphologic classification of these diseases.
Similar content being viewed by others
References
Brock N: Comparative pharmacologic study in vitro and in vivo with cyclophosphamide (NSC-26271), cyclophosphamide metabolites, and plain nitrogen mustard compounds. Cancer Treat Rep 60:301–307, 1976
Connors TA: Alkylating drugs, nitrosoureas and alkyltriazenes. In HM Pinedo (ed): Cancer Chemotherapy 1982. The EORTC Cancer Chemotherapy Annual 4. Excerpta Medica, Amsterdam, Oxford, 1982, pp 29–50
Bedford P, Fox BW: Repair of DNA interstrand crosslinks after busulphan. A possible mode of resistance. Cancer Chemother Pharmacol8: 3–7, 1982
Ducore JM, Erickson LC, Zwelling LA, Laurent G, Kohn KW: Comparative studies of DNA cross-linking and cytotoxicity in Burkitt's lymphoma cell lines treated with cisdiamminedichloroplatinum (II) and L-phenylalanine mustard. Cancer Res 42:897–902, 1982
Ramonas LM, Erickson LC, Klesse W, Kohn KW, Zaharko DS: Differential cytotoxicity and DNA cross-linking produced by polymeric and monomeric activated analogues of cyclophosphamide in mouse L1210 leukemia cells. Molec Pharmacol 19:331–336, 1981
Ramonas LM, Erickson LC, Ringsdorf H, Zaharko DS: Effect of dose, schedule, and route of administration on the in vivo toxicity and antitumor activity of two activated sulfhydryl derivatives of cyclophosphamide. Cancer Res 40:3704–3708, 1980
Surya YA, Rosenfeld JM, Hillcoat BL: Cross-linking of DNA in L1210 cells and nuclei treated with cyclophosphamide and phosphoramide mustard. Cancer Treat Rep 62:23–29, 1978
Erickson LC, Osieka R, Sharkey NA, Kohn KW: Measurement of DNA damage in unlabeled mammalian cells analyzed by alkaline elution and a fluorometric DNA assay. Anal Biochem 106:169–174, 1980
Chu M-Y, Fischer GA: The incorporation of 3H-cytosine arabinoside and its effect on murine leukemic cells (L5178Y). Biochem Pharmacol 17:753–767, 1968
Bloomer WD, McLaughlin WH, Adelstein SJ: Therapeutic implications of iodine-125 cytotoxicity. Int J Rad Oncol Biol Phys 8:1903–1908, 1982
Erickson LC, Bradley MO, Kohn KW: Strand breaks in DNA from normal and transformed human cells treated with 1,3-bis(2-chloroethyl)-1-nitrosourea. Cancer Res 37:3744–3750, 1977
Wetherley-Mein G, Thomson AER, O'Connor TWE, Peel WE, Singh AK: Colchicine ultrasensitivity of lymphocytes in chronic lymphocytic leukaemia. Br J Haematol 54:111–120, 1983
Draeger U, Hohorst H-J: Permeation of cyclophosphamide (NSC-26271) metabolites into tumor cells. Cancer Treat Rep 60:423–427, 1976
Goldenberg GJ, Land HB, Cormack DV: Mechanism of cyclophosphamide transport by L5178Y lymphoblasts in vitro. Cancer Res 34:3274–3282, 1974
Huang AT, Kremer WB, Laszlo J, Setlow RB: DNA repair in human leukaemic lymphocytes. Nature New Biol 240:114–116, 1972
Waldstein EA, Cao E-H, Miller ME, Cronkite EP, Setlow RB: Extracts of chronic lymphocytic leukemia lymphocytes have a high level of DNA repair activity for 06-methylguanine. Proc Natl Acad Sci USA 79:4786–4790, 1982
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Osieka, R., Pannenbäcker, R. & Schmidt, C.G. Macromolecular DNA-damage in murine and human leukemic and lymphoid cells after in vitro exposure to ASTA Z 7557 (INN mafosfamide). Invest New Drugs 2, 161–168 (1984). https://doi.org/10.1007/BF00232346
Issue Date:
DOI: https://doi.org/10.1007/BF00232346