novobiocin and Fibrosarcoma

Topoisomerase I and topoisomerase II allow a metabolically active cell to mobilize its supercoiled chromosomal DNA and undergo replication, transcription, recombination, and repair. Several topoisomerase inhibitors have recently been shown to be active in preclinical systems. Topotecan (SK&F 104,864), a water-soluble camptothecin analog, is an inhibitor of topoisomerase I. Novobiocin is an inhibitor of topoisomerase II. Lonidamine depletes cellular adenosine 5'-triphosphate (ATP) and may impede energy-dependent DNA repair, MCF-7 human breast-cancer cells were treated in vitro with topotecan, novobiocin, and lonidamine alone, in paired combinations, and in combination with CDDP and melphalan. The three enzyme inhibitors alone and in combination did not increase tumor cell sensitivity to CDDP. However, the combinations of topotecan/novobiocin and lonidamine/novobiocin did enhance the cytotoxicity of melphalan. Mice bearing the FSaII fibrosarcoma were treated in vivo with topotecan, novobiocin, and lonidamine alone, in paired combinations, and in combination with CDDP, melphalan, BCNU, and cyclophosphamide. The combination of topotecan/novobiocin had the greatest impact on tumor cell sensitivity to each cytotoxic agent tested in both tumor cell-survival and tumor growth-delay assays. This sensitization was greatest at the highest concentrations of the cytotoxic agent tested. Combinations of topoisomerase I and topoisomerase II inhibitors may be useful as modulators of antitumor alkylating agents.

Topics: Adenocarcinoma; Alkylating Agents; Animals; Antineoplastic Combined Chemotherapy Protocols; Breast Neoplasms; Camptothecin; Drug Synergism; Female; Fibrosarcoma; Humans; Indazoles; Male; Mice; Mice, Inbred C3H; Novobiocin; Topotecan; Tumor Cells, Cultured

Our previous in vitro studies demonstrated marked synergy with alkylating agents when novobiocin was present during and after alkylating agent exposure. To determine whether this effect is observed in vivo, novobiocin was administered daily for 3 days prior to alkylating agent treatment, during alkylating agent treatment, and for 2 days after completion of alkylating agent treatment. When combined with cis-diamminedichloroplatinum(II), 1,3-bis(2-chloroethyl)-1-nitrosourea, or cyclophosphamide, there was significant enhancement of the growth delay of the FSaIIC fibrosarcoma implanted s.c. in C3H mice when compared with alkylating agents alone. In a second assay using ex vivo studies of tumor cells exposed in vivo, single doses of 100 mg/kg of novobiocin followed by cis-diamminedichloroplatinum(II) resulted in a 3- to 4-fold increase in tumor cell killing by cis-diamminedichloroplatinum(II). At a dose of 100 mg/kg of 1,3-bis(2-chloroethyl)-1-nitrosourea there was about a 7-fold increase in tumor cell kill upon addition of novobiocin. Cyclophosphamide showed a dose response effect with novobiocin, reaching 13-fold at a dose of 300 mg/kg of cyclophosphamide. In all cases bone marrow elements were affected less than were neoplastic cells, suggesting that the combination of novobiocin and alkylating agents may be a clinically useful strategy.

Topics: Alkylating Agents; Animals; Antineoplastic Combined Chemotherapy Protocols; Bone Marrow; Carmustine; Cell Survival; Cyclophosphamide; Fibrosarcoma; Male; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Novobiocin; Organoplatinum Compounds

DNA-DNA crosslinks are the lethal cellular mechanism of bifunctional alkylating agent cytotoxicity. Novobiocin, an inhibitor of DNA topoisomerase II, impairs eukaryotic DNA repair of alkylating agent adducts and may increase the number of adducts and their resultant cytotoxicity in malignant cells. The effect of novobiocin on clonogenic survival and DNA crosslinking due to cisplatin (cDDP) and carmustine (BCNU) was studied. Novobiocin caused synergistic cytotoxicity in Chinese hamster ovary cells exposed to cDDP or BCNU. Novobiocin and cDDP increased the formation of DNA-DNA interstrand crosslinks six-fold greater than cDDP alone. The effect was schedule dependent. Novobiocin and cDDP or BCNU markedly reduced in vivo growth of a murine fibrosarcoma without increased host toxicity. As a modulating agent of cytotoxicity due to DNA-DNA crosslinking, novobiocin may enhance the clinical effectiveness of the alkylating agents in human cancer and offer insight into new therapeutic strategies.

Topics: Alkylating Agents; Animals; Antineoplastic Combined Chemotherapy Protocols; Carmustine; Cell Survival; Cisplatin; Cricetinae; Cricetulus; DNA; Drug Synergism; Female; Fibrosarcoma; Leukemia L1210; Mice; Novobiocin; Ovary; Topoisomerase II Inhibitors

Post-synthetic enzymatic hypermethylation of DNA was induced in hamster fibrosarcoma cells by the DNA synthesis inhibitors cytosine arabinoside, hydroxyurea and aphidicolin. This effect required direct inhibition of DNA polymerase alpha or reduction in deoxynucleotide pools and was not specific to a single cell type. At equivalently reduced levels of DNA synthesis, neither cycloheximide, actinomycin D nor serum deprivation affected DNA methylation in this way. The topoisomerase inhibitors nalidixic acid and novobiocin caused significant hypomethylation indicating that increased 5-mCyt content was not a necessary consequence of DNA synthesis inhibition. The induced hypermethylation occurred predominantly in that fraction of the DNA synthesized in the presence of inhibitor; was stable in the absence of drug; was most prominent in low molecular weight DNA representing sites of initiated but incomplete DNA synthesis; and occurred primarily within CpG dinucleotides, although other dinucleotides were overmethylated as well. Drug-induced CpG hypermethylation may be capable of silencing genes, an effect which may be relevant to the aberrantly expressed genes characteristic of neoplastic cells.

Topics: Animals; Aphidicolin; Base Sequence; Cell Line; Chromatography, High Pressure Liquid; Cricetinae; Cycloheximide; Cytarabine; Dactinomycin; Diterpenes; DNA; DNA Polymerase II; DNA Replication; Fibrosarcoma; Hydroxyurea; Methylation; Nalidixic Acid; Novobiocin