aphidicolin and cordycepin

Adozelesin is a highly potent alkylating agent that undergoes binding in the minor groove of double-stranded DNA (ds-DNA) at A-T-rich sequences followed by covalent bonding with N-3 of adenine in preferred sequences. On the basis of its high-potency, broad-spectrum in vivo antitumor activity and its unique mechanism of action, adozelesin has entered clinical trial. We report herein the cytotoxicity for Chinese hamster ovary (CHO) cells of several agents, including antitumor drugs, combined with adozelesin. The additive, synergistic, or antagonistic nature of the combined drug effect was determined for most combinations using the median-effect principle. The results show that in experiments using DNA- and RNA-synthesis inhibitors, prior treatment with the DNA inhibitor aphidicolin did not affect the lethality of adozelesin. Therefore, ongoing DNA synthesis is not needed for adozelesin cytotoxicity. Combination with the RNA inhibitor cordycepin also did not affect adozelesin cytotoxicity. In experiments with alkylating agents, combinations of adozelesin with melphalan or cisplatin were usually additive or slightly synergistic. Adozelesin-tetraplatin combinations were synergistic at several different ratios of the two drugs, and depending on the schedule of exposure to drug. In experiments using methylxanthines, adozelesin combined synergistically with noncytotoxic doses of caffeine or pentoxifylline and resulted in several logs of increase in adozelesin cytotoxicity. In experiments with hypomethylating agents, adozelesin combined synergistically with 5-azacytidine (5-aza-CR) and 5-aza-2'-deoxycytidine (5-aza-2'-CdR). Combinations of adozelesin with tetraplatin or 5-aza-2'-CdR were also tested against B16 melanoma cells in vitro and were found to be additive and synergistic, respectively. The synergistic cytotoxicity to CHO cells of adozelesin combinations with tetraplatin, 5-aza-CR, or pentoxifylline was not due to increased adozelesin uptake or increased alkylation of DNA by adozelesin.

Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Aphidicolin; Azacitidine; Benzofurans; Caffeine; Cell Survival; CHO Cells; Cisplatin; Cricetinae; Cricetulus; Cyclohexanecarboxylic Acids; Cyclohexenes; Decitabine; Deoxyadenosines; DNA; Dose-Response Relationship, Drug; Drug Synergism; Duocarmycins; Indoles; Melanoma, Experimental; Melphalan; Mutagens; Organoplatinum Compounds; Pentoxifylline; Tumor Cells, Cultured

Treatment of human K-562-J leukemia cells for 1 h with the topoisomerase II-reactive drugs VP-16, VM-26, or mAMSA resulted in a dose-dependent inhibition of proliferation and in an increase in the percentage of cells staining positive for hemoglobin, a marker of erythroid differentiation. Staining for hemoglobin of up to about 60% of the cells was observed at 20 microM VP-16, 1 microM VM-26, and 8 microM mAMSA. Such treatment also caused a G2/M arrest in the cell cycle. Incubation of the cells with radiolabeled VP-16 indicated that the induced erythroid differentiation was not due to continuous cell exposure to a residual amount of the drug. VP-16-induced erythroid differentiation was also not affected by DNA, RNA, or protein synthesis inhibitors. Differentiation induction and the G2/M arrest evoked by VP-16, VM-26, and mAMSA were, however, reduced in the presence of novobiocin. Our results indicate that topo-reactive drugs that cause G2/M arrest in the K-562-J cell cycle can induce in these cells erythroid differentiation after a short and irreversible interaction with their target molecule(s).

Topics: Amsacrine; Aphidicolin; Cell Cycle; Cell Differentiation; Cycloheximide; Deoxyadenosines; Dose-Response Relationship, Drug; Etoposide; G2 Phase; Hemoglobins; Humans; Kinetics; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Melanins; Mitosis; Novobiocin; Teniposide; Topoisomerase II Inhibitors; Tumor Cells, Cultured

To search for possible synergy between topoisomerase (topo) II-directed chemotherapeutic agents and topo I-directed agents, IL-60 human progranulocytic leukemia cells were incubated with etoposide in the absence or presence of camptothecin (CPT). Treatment of HL-60 cells for 1 h with 15-20 microM etoposide resulted in the death of 99-99.9% of the cells as assessed by colony formation in soft agar. Unexpectedly, simultaneous incubation with 1 microM CPT increased the survival of etoposide-treated cells as much as 30-fold. Inhibition of etoposide cytotoxicity was observed at CPT concentrations as low as 0.01 microM and was one-half maximal at 0.1 microM. CPT also antagonized the cytotoxicity of 4'-(9-acridinylamino)methanesulfon-M-anisidide and daunorubicin, two structurally unrelated topo II-directed agents. Topotecan, a CPT analogue currently undergoing Phase I clinical trials, had a similar effect. Studies using an alkaline unwinding assay (to measure DNA strand breaks) and Western blotting (to assess formation of covalent adducts involving topo II) revealed that CPT did not alter the ability of etoposide to stabilize topo II-DNA adducts. CPT is a potent inhibitor of both DNA and RNA synthesis. To further assess the mechanism by which CPT diminished the cytotoxicity of topo II-directed agents, inhibitors of DNA synthesis or RNA synthesis were substituted for CPT. Aphidicolin, an inhibitor of replicative DNA polymerases, enhanced the survival of etoposide-treated HL-60 cells less than 3-fold. In contrast, inhibitors of RNA synthesis (cordycepin or 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole) enhanced the survival of etoposide-treated HL-60 cells as much as 20-fold. The potential biological and therapeutic implications of these results are discussed.

Topics: Amsacrine; Aphidicolin; Blotting, Western; Camptothecin; Colony-Forming Units Assay; Cycloheximide; Daunorubicin; Deoxyadenosines; Dichlororibofuranosylbenzimidazole; Diterpenes; DNA Polymerase II; DNA Replication; Drug Synergism; Etoposide; Female; Humans; Leukemia, Promyelocytic, Acute; Mutagens; RNA; Topoisomerase II Inhibitors; Topotecan; Tumor Cells, Cultured

The primary cytotoxic mechanism of camptothecin has been proposed to involve an interaction between the replication machinery and the camptothecin-mediated topoisomerase I-DNA cleavable complex (Y. H. Hsiang, M.G. Lihou, and L.F. Liu, Cancer Res., 49:5077-5082, 1989). In the present study, we show that killing of V79 cells by the topoisomerase II poisons 4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA) and etoposide may involve ongoing RNA synthesis in addition to ongoing DNA synthesis. V79 cells synchronized by mitotic shake-off were treated with topoisomerase poisons in the presence of inhibitors of nucleic acid synthesis. S-Phase V79 cells were more sensitive to the topoisomerase I poison camptothecin and the topoisomerase II poison m-AMSA than G1-phase cells. The greater sensitivity of S-phase cells to killing by m-AMSA and camptothecin was abolished during cotreatment, but not posttreatment, with aphidicolin, suggesting that ongoing DNA synthesis in involved in cell killing by both topoisomerase I and II poisons. Cotreatment with transcription inhibitors, such as 5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole or cordycepin, partially protected cells from the cytotoxic effects of m-AMSA but had no effect on camptothecin-mediated cytotoxicity. These results suggest that ongoing RNA transcription may be involved in cell killing by topoisomerase II poisons but not topoisomerase I poisons. Cotreatment with camptothecin reduced m-AMSA-mediated cytotoxicity in G1-phase V79 cells, suggesting a possible antagonism between topoisomerase I and II poisons. This antagonistic effect between topoisomerase I and II poisons could be explained by the strong inhibitory effect of camptothecin on RNA transcription.

Topics: Amsacrine; Animals; Antibiotics, Antineoplastic; Aphidicolin; Camptothecin; Cell Survival; Cricetinae; Cricetulus; Cycloheximide; Deoxyadenosines; Dichlororibofuranosylbenzimidazole; Diterpenes; DNA; Etoposide; Mutagens; Nucleic Acids; RNA; Time Factors; Topoisomerase I Inhibitors; Topoisomerase II Inhibitors