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PubMedDate not supplied

Phase I and pharmacokinetic study of novobiocin in combination with VP-16 in patients with refractory malignancies.

PURPOSE: The coumarin antibiotic novobiocin potentiates the activity of etoposide (VP-16) in vitro by increasing intracellular accumulation of VP-16. The drug efflux pump inhibited by novobiocin appears to be distinct from both of the major proteins associated with the multidrug resistance phenotype in human cancers, the 170-kDa P-glycoprotein and the 190-kDa multidrug resistance protein. In a recent study, we found that novobiocin augmented VP-16 accumulation ex vivo in 16 of 24 fresh tumor samples at concentrations that could be achieved in vivo. Therefore, we conducted a clinical trial to determine the maximum tolerated dose and the pharmacokinetics of novobiocin when given in combination with VP-16. PATIENTS AND METHODS: Patients with refractory cancer were treated with VP-16 on days 1, 3, and 5. Antiemetics, consisting of ondansetron and dexamethasone, were given 60 minutes before the VP-16 was administered. Novobiocin was given orally 30 minutes before the VP-16, and the dose was escalated in successive groups of patients according to a standard dose escalation design. Treatment cycles were repeated every 4 weeks. Plasma concentrations of novobiocin were determined during the first treatment cycle by high-performance liquid chromatography. RESULTS: Thirty-three patients were treated for a total of 69 cycles. Eleven patients were treated with a starting dose of VP-16 of 120 mg/m2, and three of these patients experienced neutropenic fever. The dose of VP-16 was reduced to 100 mg/m2, and an additional 22 patients were enrolled. The dose of novobiocin ranged from 3 to 9 g. At a novobiocin dose of at least 5.5 g, plasma concentrations of at least 150 microM were sustained for 24 hours. Dose-limiting toxicities consisted of neutropenic fever and reversible hyperbilirubinemia. Nausea, which was a limiting toxicity in other trials of novobiocin, was well controlled with the use of serotonergic antiemetics. Diarrhea was common but mild in most patients. DISCUSSION: In previously treated patients, the recommended dose of novobiocin in this schedule is 7 g/m2/day. Novobiocin does not appear to augment the toxicity of VP-16 to the bone marrow or the gastrointestinal mucosa. Plasma concentrations of novobiocin equivalent to the levels required to modulate VP-16 in vitro are readily achievable for total but not unbound free drug.

Adult↗
PubMed1992

Potentiation by novobiocin of the cytotoxic activity of etoposide (VP-16) and teniposide (VM-26).

The coumermycin antibiotic novobiocin, which interacts with the nuclear enzyme topoisomerase II, produced supra-additive toxicity to WEHI-3B D+ leukemia cells at clinically achievable concentrations, when combined with teniposide (VM-26) or etoposide (VP-16). Simultaneous exposure of cells to both agents was required for maximum efficacy of the combination. Novobiocin also produced supra-additive toxicity to A549 human lung carcinoma cells when combined with VM-26 or VP-16. At concentrations above the peak plasma levels achievable in patients, novobiocin lost its potentiating activity. Exposure of WEHI-3B D+ cells to novobiocin did not modify the cytotoxicity produced by the topoisomerase II inhibitor m-AMSA, whereas, in contrast, novobiocin antagonized the cytotoxicity of m-AMSA in A549 cells. Although it has been suggested that inhibitors of the syntheses of DNA and RNA interfere with the cytotoxic activity of the epipodophyllotoxins, maximum potentiation of the cytotoxicities of VP-16 and VM-26 occurred at novobiocin concentrations that decreased the rates of synthesis of both DNA and RNA in WEHI-3B D+ cells by about 50%. The number of DNA-topoisomerase-II covalent complexes stabilized by VM-26 in WEHI-3B D+ cells was greatly increased when cells were exposed simultaneously to VM-26 and novobiocin for 1 hr, but not when cells were treated with m-AMSA and novobiocin for the same period of time. Novobiocin did not affect the amount of covalent complexes produced by VM-26 in isolated nuclei, suggesting that the potentiating activity of novobiocin was not due to its direct interaction with the nuclear topoisomerase II enzyme. Our findings suggest that therapeutic levels of novobiocin may be capable of enhancing the clinical activities of VP-16 and VM-26.

Adenocarcinoma↗
PubMed2004

Novobiocin induces a distinct conformation of Hsp90 and alters Hsp90-cochaperone-client interactions.

Hsp90 functions to facilitate the folding of newly synthesized and denatured proteins. Hsp90 function is modulated through its interactions with cochaperones and the binding and hydrolysis of ATP. Recently, novobiocin has been shown to bind to a second nucleotide binding site located within the C-terminal domain of Hsp90. In this report, we have examined the effect of novobiocin on Hsp90 function in reticulocyte lysate. Novobiocin specifically inhibited the maturation of the heme-regulated eIF2alpha kinase (HRI) in a concentration-dependent manner. Novobiocin induced the dissociation of Hsp90 and Cdc37 from immature HRI, while the Hsp90 cochaperones p23, FKBP52, and protein phosphatase 5 remained associated with immature HRI. Proteolytic fingerprinting of Hsp90 indicated that novobiocin had a distinct effect on the conformation of Hsp90, and molybdate lowered the concentration of novobiocin required to alter Hsp90's conformation by 10-fold. The recombinant C-terminal domain of Hsp90 adopted a proteolytic resistant conformation in the presence of novobiocin, indicating that alteration of Hsp90/cochaperone interactions was not the cause of the novobiocin-induced protease resistance within Hsp90's C-terminal domain. The concentration dependence of this novobiocin-induced conformation change correlated with the dissociation of Hsp90 and Cdc37 from immature HRI and novobiocin-induced inhibition of Hsp90/Cdc37-dependent activation of HRI's autokinase activity. The data suggest that binding of novobiocin to the C-terminal nucleotide binding site of Hsp90 induces a change in Hsp90's conformation leading to the dissociation of bound kinase. The unique structure and properties of novobocin-bound Hsp90 suggest that it may represent the "client-release" conformation of the Hsp90 machine.

Animals↗
PubMed1995

Phase I and pharmacologic study of the alkylating agent modulator novobiocin in combination with high-dose chemotherapy for the treatment of metastatic breast cancer.

PURPOSE: Resistance to alkylators may potentially be overcome by drugs that inhibits DNA repair, thus improving the efficacy of high-dose chemotherapy. This trial was performed to determine if novobiocin, an agent that inhibits DNA repair, could be given with high-dose alkylators. Study aims were to define the toxicities and maximal-tolerated dose (MTD) of novobiocin and the pharmacokinetics of novobiocin and high-dose cyclophosphamide and thiotepa. PATIENTS AND METHODS: Thirty-eight women with responsive metastatic breast cancer received high-dose cyclophosphamide (3 to 6 g/m2 over 4 days), thiotepa (400 to 800 mg/m2), and novobiocin (0.5 to 5.0 g/d x 7, orally) with autologous marrow support. Toxicity was monitored. The pharmacology of novobiocin, cyclophosphamide, and thiotepa was evaluated. RESULTS: There were no toxic deaths. The MTD of novobiocin was 4 g/d. All seven patients treated at 5 g/d developed grade III/IV mucositis and vomiting. The severity of mucositis correlated with the plasma levels of novobiocin. Other severe toxicities were not observed. Plasma novobiocin levels > or = 100 micrograms/mL, which are associated with reversal of drug resistance in animal models, were consistently seen at dose levels greater than 2 g. The dispositions of cyclophosphamide and thiotepa were not altered by novobiocin. CONCLUSION: Novobiocin may be given with high-dose alkylators in doses that produce plasma levels that augment the activity of these cytotoxics in experimental models. The pharmacology of high-dose cyclophosphamide and thiotepa is unaffected. Novobiocin 4 g/d orally for 7 days is recommended for future study.

Adult↗
PubMed1992

Novobiocin-induced accumulation of etoposide (VP-16) in WEHI-3B D+ leukemia cells.

A previous report from this laboratory demonstrated that novobiocin produced supra-additive cytotoxicity when combined with etoposide (VP-16) or teniposide (VM-26) in WEHI-3B D+ and A549 cells. The increase in cytotoxicity was accompanied by an increase in the formation of drug-stabilized protein-DNA covalent complexes. We now report that novobiocin increased the amount of VP-16-induced covalent complexes between the 170 kDa form of topoisomerase II and DNA in WEHI-3B D+ cells, as measured by the band-depletion immunoblotting assay, while it did not affect the extractable topoisomerase II activity, measured by the unknotting of P4 phage DNA and by a DNA cleavage assay. Novobiocin progressively increased the steady-state concentration of intracellular VP-16. Removal of novobiocin resulted in a rapid return of VP-16 to levels comparable to those seen with VP-16 alone. The increased accumulation of VP-16 was accounted for by an increase in the exchangeable fraction only. The novobiocin-mediated increase in the steady-state concentration of VP-16 occurred whether novobiocin was added simultaneously with VP-16 or was added after a steady-state level of VP-16 had been achieved. Novobiocin did not affect the initial rate of uptake of VP-16; however, it inhibited the efflux of the epipodophyllotoxin. In fact, when cells were loaded with the same level of VP-16 in the presence or absence of novobiocin, the efflux curves in the presence or absence of novobiocin were significantly different. We conclude that the inhibition of VP-16 efflux by novobiocin is responsible for the increase in VP-16 accumulation, leading to increased formation of VP-16-stabilized topoisomerase-II-DNA covalent complexes and increased cytotoxicity.

Animals↗
PubMed1981

Dependence of mammalian DNA synthesis on DNA supercoiling. III. Characterization of the inhibition of replicative and repair-type DNA synthesis by novobiocin and nalidixic acid.

Novobiocin and nalidixic acid, inhibitors of the bacterial enzyme DNA gyrase, inhibit DNA, RNA and protein synthesis in several human and rodent cell lines. The sensitivity of DNA synthesis (both replicative and repair) to inhibition by novobiocin and nalidixic acid is greater than that of protein synthesis. Novobiocin inhibits RNA synthesis about half as effectively as it does DNA synthesis, whereas nalidixic acid inhibits both equally well. Replicative DNA synthesis, as measured by incorporation of [3H]thymidine, is blocked by novobiocin in a number of cell strains; the inhibition is reversible with respect to both DNA synthesis and cell killing, and continues for as long as 20--30 h if the cells are kept in novobiocin-containing growth medium. Both novobiocin and nalidixic acid inhibit repair DNA synthesis (measured by BND-cellulose chromatography) induced by ultraviolet light or N-methyl-N'-nitro-N-nitrosoguanidine (but not that induced by methyl methanesulfonate) at lower concentration (as low as 5 micrograms/ml) than those required to inhibit replicative DNA synthesis (50 micrograms/ml or greater). Neither novobiocin nor nalidixic acid alone induces DNA repair synthesis. Incubation of ultraviolet-irradiated cells with 10--100 micrograms/ml novobiocin results in little, if any, further reduction of colony-forming ability (beyond that caused by the ultraviolet irradiation). Novobiocin at sufficiently low concentrations (200 micrograms/ml) apparently generates a quiescent state (in terms of cellular DNA metabolism) from which recovery is possible. Under more drastic conditions of time in contact with cells and concentration, however, novobiocin itself induces mammalian cell killing.

Cell Line↗
PubMed1985

Novobiocin inhibition of DNA excision repair may occur through effects on mitochondrial structure and ATP metabolism, not on repair topoisomerases.

Novobiocin inhibits DNA topoisomerases. It also inhibits excision repair of DNA photodamage, blocking both repair synthesis and the earlier step of incision at u.v. damage sites (as measured by the accumulation of DNA strand breaks in u.v.-irradiated interphase cells treated with DNA synthesis inhibitors such as hydroxyurea or cytosine arabinoside). It has been supposed, therefore, that novobiocin affects repair by blocking a putative topoisomerase step prior to incision. But we find that novobiocin also has a marked dose- and time-dependent effect on mitochondria: in cells exposed to novobiocin, mitochondria swell and their cristae become disrupted, and the intracellular ATP:ADP ratio is lowered, though the membrane potential is maintained as judged by rhodamine 123 fluorescence. Mitotic cells are more resistant to mitochondrial disruption by novobiocin than are interphase cells. This correlates with a relative resistance of u.v.-irradiated mitotic cells to the inhibition of incision by novobiocin. The chromosomal decondensation that results from the accumulation of DNA breaks due to incision when u.v.-irradiated mitotic cells are treated with hydroxyurea and cytosine arabinoside is largely suppressed by novobiocin. Furthermore, the suppression of induced strand break accumulation is partly due to a suppression by novobiocin of the uptake and phosphorylation of cytosine arabinoside; breaks accumulated in u.v.-irradiated cells in the presence of aphidicolin, an inhibitor of DNA polymerase alpha that does not require phosphorylation, are less novobiocin-sensitive. We conclude that the effects of novobiocin on excision repair are more likely to be due to a non-specific effect on ATP metabolism than to a specific effect on a repair-related topoisomerase.

Adenosine Triphosphate↗
PubMed1977

Morphological changes associated with novobiocin resistance in Bacillus licheniformis.

Spontaneously occurring novobiocin-resistant (Nov) mutants of Bacillus licheniformis ATCC 9945, resistant to low levels of novobiocin (15 mug/ml), were isolated with a frequency of 3 in 106 organisms. Such isolates grew well, but nearly all exhibited consistent plleiotropic alterations in colonial and cell morphologies. One mutant, nov-12, grew as chains of unseparated but clearly distinct daughter cells in the absence of novobiocin in liquid culture. When novobiocin was present, nov-12 grew as very long "filaments" which were, however, septate. Septa formed in the presence of the antibiotic were normal, except that no annular clevage of the septal wall was observed. Septa were also irregularly positioned along the filament. These observations were compared with previous findings on the effects of novobiocin and novobiocin resistance described for other organisms. It was concluded that the primary action of novobiocin might differ in gram-positive and gram-negative organisms. However, when the low-level novobiocin sensitivity, normally associated with gram-positive organisms, was genetically abolished in Nov strains of B. licheniformis they became susceptible to an action of novobiocin more analogous to that found for gram-negative organisms. The morphological alterations associated with the Nov phenotype in this organism, together with observations in other organisms, indicate that novobiocin resistance might be generally useful in the search for mutants of gram-positive organisms with altered cell walls.

Bacillus↗
PubMed1992

Development and characterization of a WEHI-3B D+ monomyelocytic leukemia cell line resistant to novobiocin and cross-resistant to other topoisomerase II-targeted drugs.

A novobiocin-resistant subline of WEHI-3B D+ murine monomyelocytic leukemia cells was developed by the continuous exposure of cells to this agent in vitro. Sensitive (WEHI-3B/S) and novobiocin-resistant (WEHI-3B/NOVO) sublines were cloned in vitro. WEHI-3B/NOVO cells were stable in the absence of novobiocin for more than 3 months, and the sensitive and resistant clones displayed the same growth rate, cell cycle distribution, cell size, DNA and protein content, and cloning efficiency. Novobiocin has been shown to compete with ATP for the ATP-binding site of topoisomerase II; therefore, intracellular ATP levels can influence the cellular sensitivity to novobiocin. High-performance liquid chromatographic analysis of total cell extracts demonstrated that no difference exists between WEHI-3B/S and WEHI-3B/NOVO cells in the content of ATP. Furthermore, exposure of both cell lines to novobiocin did not affect intracellular ATP levels. In addition to an approximately 2-fold level of resistance to novobiocin, the WEHI-3B/NOVO subline was also 7- and 11-fold cross-resistant to the topoisomerase II-targeted drugs, teniposide and etoposide (VP-16), respectively. A lower level of cross-resistance, comparable to that of novobiocin, was observed in WEHI-3B/NOVO cells for the intercalating topoisomerase II-reactive drugs, doxorubicin, 4'-(9-acridinylamino)methanesulfon-m-anisidide and aclacinomycin A, while the sensitivity to the cytotoxic action of the non-topoisomerase II-acting agents, camptothecin and vincristine, was not altered. After 3-6 h of exposure to 1 microM VP-16, WEHI-3B/S cells accumulated in the S and G2 + M phases of the cell cycle. Similar changes were detected in WEHI-3B/NOVO cells only after exposure to a 10-fold higher concentration of VP-16. Exposure to 150 microM novobiocin caused an accumulation of WEHI-3B/S cells in the G0-G1 phase of the cell cycle but did not affect the cell cycle distribution of WEHI-3B/NOVO cells, while camptothecin induced the same type and extent of changes in the cell cycle distribution of both cell lines. Although the WEHI-3B/NOVO subline appeared to be less responsive to the differentiation-inducing activity of novobiocin and teniposide, the capacity of WEHI-3B/NOVO cells to respond to the differentiation-inducing agent 13-cis-retinoic acid was not significantly different from that of WEHI-3B/S cells. A slight decrease in the accumulation of VP-16 occurred in the resistant cell line, which did not appear to be of sufficient magnitude to account for the 11-fold increase in the degree of resistance to this agent.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Triphosphate↗
PubMed1988

Na transport stimulation by novobiocin: transepithelial parameters and evaluation of ENa.

The action of the antibiotic novobiocin on transepithelial Na transport was studied in isolated skins obtained from two different frog species. In Rana esculenta addition of novobiocin to the outer bath (1 mM) resulted in a sustained and reversible stimulation of the short-circuit current, transepithelial potential, and transepithelial conductance. Similar, though more variable and much less pronounced changes were observed in Rana temporaria. In the presence of amiloride (0.1 mM) novobiocin had no effect on any of the investigated transport parameters and all novobiocin induced changes were fully reversed when amiloride was given subsequently. At reduced external Na concentration or low pH the action of novobiocin was found to be greatly attenuated. In the presence of novobiocin an increased affinity to amiloride and a linearization of the transepithelial current-voltage relationship was observed. The results are consistent with the view that novobiocin increases the Na permeability of the outer membrane, possibly by an attenuation of an Na self-inhibition mechanism. In addition, the driving force of transepithelial Na transport was estimated by means of novobiocin. Several different methods were employed, providing varying results. As shown in an Appendix, for the most part the discrepancies can be explained by changes in the intracellular Na and K concentration. In some cases, novobiocin induced large secondary increases in the skin conductance which can be referred to an increased Cl permeability.

Amiloride↗
PubMed1992

Novobiocin-induced anti-proliferative and differentiating effects in melanoma B16.

The antibiotic drug novobiocin was evaluated for its anti-tumour properties in B16 melanoma cells. Novobiocin is shown to inhibit melanoma B16 cell proliferation. The anti-proliferative effect was gradually reversible upon removal of novobiocin from the culture medium. Growth inhibition by novobiocin was accompanied by phenotypic alterations, that included morphological changes, lipid accumulation and marked increases in the activities of NADPH cytochrome c reductase and gamma glutamyl transpeptidase. In vivo administration of repeated i.p. doses of novobiocin, to mice implanted with B16 melanoma cells resulted in growth retardation. The combined treatment of the B16 melanoma cells with novobiocin and other chemical inducers of differentiation was examined in a cell growth assay. Novobiocin and sodium butyrate inhibited cell growth in a near additive manner, while combination of novobiocin with the GTP-depleting agents, tiazofurin or mycophenolic acid resulted in a synergistic decrease in cell growth. Our results support the contention further that novobiocin and other differentiating agents might be of potential value in melanoma therapy.

Animals↗
PubMed2000

Novobiocin and related coumarins and depletion of heat shock protein 90-dependent signaling proteins.

BACKGROUND: Heat shock protein 90 (Hsp90) interacts with and stabilizes several oncogenic protein kinases (e.g., p185(erbB2), p60(v-src), and Raf-1) and is required for the stability and dominant-negative function of mutated p53 protein. Two unrelated antibiotics, geldanamycin and radicicol, bind specifically to an atypical nucleotide-binding pocket of Hsp90, a site that shares homology with the adenosine triphosphate (ATP)-binding domain of bacterial DNA gyrase B. This interaction leads to destabilization of proteins that interact with Hsp90. Since the nucleotide-binding site of gyrase B is targeted by coumarin antibiotics (e.g., novobiocin), we investigated whether these drugs can also interact with Hsp90 and affect its activity. METHODS: We used immobilized novobiocin, geldanamycin, or radicicol to isolate either endogenous Hsp90 from cell lysates or Hsp90 deletion fragments translated in vitro. Effects of the coumarin antibiotics novobiocin, chlorobiocin, and coumermycin A1 on several proteins interacting with Hsp90 were assessed in vitro and in vivo. RESULTS: Hsp90 binding to immobilized novobiocin was competed by soluble coumarins and ATP but not by geldanamycin or radicicol. A carboxy-terminal Hsp90 fragment bound immobilized novobiocin but not immobilized geldanamycin, while a geldanamycin-binding amino-terminal fragment did not bind novobiocin. All three coumarins markedly reduced cellular levels of p185(erbB2), p60(v-src), Raf-1, and mutated p53. Furthermore, novobiocin reduced Raf-1 levels in the spleens of mice treated with the drug. CONCLUSIONS: These coumarin antibiotics, particularly novobiocin, represent a first-generation alternative to other Hsp90-targeting drugs that are not as well tolerated. Novobiocin's unique interaction with Hsp90 identifies an additional site on this protein amenable to pharmacologic interference with small molecules.

Aminocoumarins↗
PubMed1982

Novobiocin resistance marker in Haemophilus influenzae that is not expressed on a plasmid.

The plasmid pNov2, carrying a cloned chromosomal marker conferring resistance to at least 2.5 micrograms of novobiocin per ml, was constructed with a new Haemophilus influenzae cloning vehicle, pDM2. The novobiocin marker of pNov2 was not normally expressed, but in Rec+ cells approximately one in 10(4) cells in a culture of a transformant became novobiocin resistant, a frequency about four orders of magnitude higher than the spontaneous mutation frequency. Variants of such cells that had lost the plasmid were also novobiocin resistant. Since Rec- cultures bearing pNov2 showed novobiocin resistance only at the normal mutation frequency, we concluded that the Rec+ novobiocin-resistant transformants arose because of a rare recombination between plasmid and chromosome in which the chromosome acquired the novobiocin marker from the plasmid. Evidence is presented that novobiocin sensitivity is dominant over this particular novobiocin resistance marker.

Chloramphenicol↗
PubMed2000

Structure-activity studies of novobiocin analogs as modulators of the cytotoxicity of etoposide (VP-16).

We have previously reported that the antibiotic novobiocin enhanced the toxicity of the anticancer agent etoposide (VP-16) to several drug-sensitive and -resistant tumor cell lines. The increase in VP-16 cytotoxicity produced by novobiocin was not due to the combined effects of these agents on topoisomerase II, but to inhibition by novobiocin of VP-16 efflux, which in turn led to increased accumulation of VP-16 and increased formation of potentially lethal VP-16-stabilized topoisomerase II-DNA covalent complexes. We have now identified novobiocin analogs that are essentially equivalent to novobiocin as inhibitors of the activity of topoisomerase II, but that are more potent than novobiocin (a) as modulators of the cytotoxicity of VP-16 to WEHI-3B leukemia and A549 lung carcinoma cells and (b) in increasing VP-16 accumulation in these cell lines. Thus, removal of the sugar moiety of novobiocin to form novobiocic acid enhanced the potency of the antibiotic as a modulator of VP-16, whereas the substituted coumarin ring alone (U-7587) was devoid of VP-16 modulatory activity. Modifications of the side chain of novobiocin significantly influenced modulatory activity, with cyclonovobiocic acid, which was formed from novobiocic acid by acid-catalyzed cycloaddition, being the most active in enhancing the cytotoxicity of VP-16. The increased potency of novobiocic acid and cyclonovobiocic acid as modulators of VP-16 activity was achieved with no change from novobiocin in the capacity of these analogs to inhibit the catalytic activity of mammalian topoisomerase II, indicating a change in the specificity of these analogs.

ATP-Binding Cassette Transporters↗
PubMed2003

[Novobiocin inhibits angiogenesis and shows synergistic effect with vincristine].

AIM: To study the anti-angiogenic activity of novobiocin and its mechanism of action. METHODS: The anti-angiogenic activity of novobiocin was determined using chick embryo chorioallantoic membrane(CAM) assay. MTT assay, zymography and related assays were used to observe the effects of drugs on bovine aorta endothelial cells and human pulmonary carcinoma PG cells. RESULTS: Novobiocin at the doses of 100 and 200 micrograms/egg inhibited angiogenesis by 31.6% and 68.7% in CAM, respectively. The combination of novobiocin and vincristine enhanced the anti-angiogenic effect. Novobiocin inhibited the proliferation of bovine aortic endothelial cells in a concentration-dependent manner. In addition, novobiocin suppressed MMP-2 secretion, migration, and tube formation of endothelial cells. As determined by MTT assay, novobiocin in combination with vincristine displayed synergistic effect on the proliferation of PG cells, CONCLUSION: This study demonstrates that novobiocin is active in suppressing angiogenesis and the anti-angiogenic activity may be enhanced by combination with vincristine. The anti-angiogenic activity of novobiocin may be related, at least in part, to its inhibition of cell proliferation, cell migration, tube formation and secretion of matrix metalloproteinases.

Angiogenesis Inhibitors↗
PubMed1989

Novobiocin- and phorbol-12-myristate-13-acetate-induced differentiation of human leukemia cells associated with a reduction in topoisomerase II activity.

Studies were conducted to determine the possible involvement of DNA topoisomerase II (Topo II) in the induction of differentiation in two human promyelocytic HL-60 leukemia cell variants that are either susceptible or resistant to differentiation induced by phorbol-12-myristate-13-acetate (PMA), a protein kinase C activator. The acquisition of maturation markers and changes in the activity, level, and phosphorylation of Topo II were determined after treatment with either novobiocin, a Topo II inhibitor, or PMA. Novobiocin at 50-150 microM induced differentiation in the HL-205 cells but not in the HL-525 cells, although both cell types were equally susceptible to novobiocin-evoked cytotoxicity. In both cell types, novobiocin induced similar reductions in topoisomerase I activity but different reductions in Topo II activity. Treatment with novobiocin at 150 microM for 6 h or at 2 mM for 30 min resulted in a 4-fold or higher reduction in Topo II activity in the differentiation-susceptible HL-205 cells but not in the differentiation-resistant HL-525 cells. A differential response in Topo II activity was also observed after treatment with PMA. The novobiocin-evoked decrease in Topo II activity seems to be due to an enhanced enzyme proteolysis, whereas the PMA-elicited decrease in Topo II activity is associated with an increase in Topo II phosphorylation. 1-(5-Isoquinolinesulfonyl)-2-methylpiperazine, which is an inhibitor of protein kinases, including protein kinase C, diminished the novobiocin-elicited proteolysis of Topo II and the PMA-induced Topo II phosphorylation, as well as the decrease in Topo II activity and the acquisition of differentiation markers induced by either novobiocin or PMA. These results suggest that induction of differentiation in HL-60 cells by novobiocin or PMA is associated with a reduction in Topo II activity, mediated directly or indirectly by a protein kinase(s), perhaps protein kinase C.

Amsacrine↗
PubMed1993

Reversal of etoposide resistance in non-P-glycoprotein expressing multidrug resistant tumor cell lines by novobiocin.

Previous reports from this laboratory have demonstrated that novobiocin produces supraadditive cytotoxicity and increases the formation of drug-stabilized topoisomerase II-DNA covalent complexes in WEHI-3B myelomonocytic leukemia and A549 lung carcinoma cells when combined with etoposide (VP-16). Inhibition of the efflux of VP-16 by novobiocin is responsible for the increase in VP-16 accumulation, which in turn leads to increased formation of VP-16-stabilized topoisomerase II-DNA covalent complexes and increased cytotoxicity. We now report that novobiocin synergistically enhanced the sensitivity of the multidrug resistant variants, WEHI-3B/NOVO and A549(VP)28, to VP-16, causing almost complete reversal of the resistance to the epipodophyllotoxin. These two tumor cell variants are resistant to several topoisomerase II-targeted drugs, particularly VP-16, but not to Vinca alkaloids; this finding corresponds to the fact that they do not overexpress the P-glycoprotein. The effects of novobiocin in these resistant sublines are mediated through the intracellular accumulation of VP-16, resulting in an increase in the formation of lethal VP-16-induced topoisomerase II-DNA covalent complexes. In the P-glycoprotein expressing multidrug resistant HCT116(VM)34 colon carcinoma and L1210/VMDRC0.06 leukemia cell lines, the latter being transfected with the human mdr-1 gene, novobiocin did not potentiate the cytotoxic activity of VP-16 nor increase the intracellular accumulation of VP-16 and the formation of covalent complexes, whereas their normal counterparts were sensitive to the potentiating activity of novobiocin when used in combination with VP-16. These results indicate that the action of novobiocin on the intracellular transport of VP-16 is not directed at the level of the P-glycoprotein, but that the action of novobiocin is antagonized by the presence of the P-glycoprotein. Since novobiocin is a clinically available antibiotic, has numerous structural analogues available for comparative studies, and has a relatively low toxicity profile, this drug, as well as structurally related agents, would appear to have significant clinical potential in combination with an epipodophyllotoxin for the treatment of non-P-glycoprotein expressing multidrug resistant tumors.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

The properties of ion channels formed by the coumarin antibiotic, novobiocin, in lipid bilayers.

The coumarin antibiotic novobiocin forms ion channels of varying conductances in lipid bilayers. The conductances (about 20, 22, 14, 7 and 2 pS for 100 mM NH4Cl, CsCl, KCl, NaCl and LiCl, respectively) and selectivities (cation transference numbers in the range of 0.97-0.98) of one type of novobiocin-induced channel are similar to those found for channels formed by gramicidin A, an antibiotic of very different structure. The conductance of novobiocin channels of this type was independent of the species of the membrane lipid. This observation suggests that novobiocin molecules directly form these channels, and that channels are not formed through defects in lipid structure. The similarity in conductance and ion selectivity between channels induced by novobiocin and those formed by gramicidin A suggests that these structurally different molecules form channels with comparable internal diameter and internal surface charge distribution. Using HPLC purification we argue that the channel-forming activity of novobiocin is related to the activity of the novobiocin molecule itself, and not to a contaminant of the commercially available novobiocin sodium salt preparation.

Cations, Monovalent↗