This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Cavanaugh, P. F. Articles by Bergeron, R. J. Search for Related Content PubMed PubMed Citation Articles by Cavanaugh, P. F., Jr. Articles by Bergeron, R. J.

Characterization of L1210 Cell Growth Inhibition by the Bacterial Iron Chelators Parabactin and Compound II¹

Grace Cancer Drug Center, Roswell Park Memorial Institute, Buffalo, New York 14263 [P. F. C., O. S. F., C. W. P., Z. P. P.], and Department of Medicinal Chemistry, J. Hillis Miller Health Center, University of Florida, Gainesville, Florida 36210 [R. J. B., D. T.]

Microbial siderophores represent a class of iron chelators characterized by their high affinity (i.e., formation constants, >10⁴⁰ M) for ferric iron. Previously, we demonstrated that the bacterial siderophores, N-[3-(2,3-dihydroxybenzamido)propyl]-N-[4-(2,3-dihydroxybenzamino)butryl]-2-(2-hydroxyphenyl)trans-5-methyloxazoline-4-carboxamide (Parabactin) and N¹,N⁸-bis(2,3-dihydroxybenzoyl)spermidine (Compound II), inhibit the growth of L1210 cells and the replication of DNA (but not RNA) viruses at low micromolar concentrations (Biochem. Biophys. Res. Commun., 121: 848–854, 1984). The basis for this antiproliferative effect on L1210 cells has now been investigated further. Onset of growth inhibition induced by 5 µM Parabactin occurs much earlier than with an equimolar concentration of Compound II but, once established by either chelator, inhibition appears to be irreversible. Growth inhibition was fully preventable with exogenous FeCl₃ when given at the same time as the chelators. Flow cytometric analysis revealed a G₁-S cycle block following treatment for 4 h with either 5 µM Parabactin or 30 µM Compound II. The block was readily reversed with exogenous FeCl₃, allowing cells to progress to mid-S phase by 3 h and to G₁ again by 9 h. Thereafter, cells accumulated at a second block located at S phase. The treatment conditions required for the initial cell cycle block (at 4 h) were adapted for subsequent studies. Clonogenicity of L1210 cells in soft agar following a 4-h exposure was reduced to 22% of control by 5 µM Parabactin and to 16% by 30 µM Compound II. Neither growth inhibition in suspension culture nor decreased clonogenicity in soft agar could be reversed with exogenous iron, following treatment with the chelators. Both chelators caused an early and significant decrease in [¹⁴C]thymidine incorporation over the 4-h period (50% inhibitory concentration at 4 h, 0.4 µM for Parabactin and 6.0 µM for Compound II). [³H]Uridine incorporation was inhibited later than [¹⁴C]thymidine and to a much lesser extent, while [³H]leucine incorporation was not significantly affected. Treatment of cells with 5 µM Parabactin or Compound II for 4 h decreased deoxyadenosine triphosphate pools by 38 and 70%, respectively, and increased deoxythymidine triphosphate pools by 67 and 36%, respectively, suggesting interference with ribonucleotide reductase. Indeed, extracts of cells treated for 4 h with either 5 µM Parabactin or 30 µM Compound II exhibit a 97 to 98% decrease in cytidine-5'-diphosphate reductase activity compared to control, whereas DNA polymerase was elevated slightly. These data suggest that interference with DNA biosynthesis by inhibition of the non-heme iron-containing enzyme, ribonucleotide reductase, represents one site of drug action involved in the early antiproliferative effects of the bacterial siderophores, Compound II and Parabactin.

¹ This investigation was supported by American Cancer Society Postdoctoral Fellowship Grant PF-2425 (P. F. C.); National Institute of Arthritis and Metabolic Diseases Grant AM-29936; and Grants CA-33321, CA-37606, and CA-24538 from the National Cancer Institute, Department of Health, Education, and Welfare, and the Veterans Administration.

² To whom requests for reprints should be addressed.

Received 11/16/84. Revised 7/ 3/85. Accepted 7/ 8/85.