Discriminant analysis. 2. A new index for antitumor activity

The antitumor activities of 17 nitrogen analogs of 1, 4-benzoquinone, expressed in eight different ways, were subjected to discriminant analysis. Different methods of measuring the effectiveness of antitumor agents give different results. Methods based on dose levels such as the optimum dose or the lethal dose appear to be more useful in classifying compounds as antitumor agents than methods based on the maximum antitumor effect. A new index for antitumor activity involving the maximum antitumor effect, the optimum dose, and the therapeutic index, examined by discriminant analysis with substitutive parameters as discriminatory variables, can give valuable information for the design of new antitumor drugs.     

Lethal and mutagenic action of N-nitroso-N-methylbiuret on the producers of levorin, amphotericin and mycoheptin]

The lethal and mutagenic effect of N-nitrozo-N-methylbiuret (NMB) on the organisms producing levorin, amphotericin B and mycoheptin was studied. The mutagen effect depended on the dose, culture and physiological state of the spores. NMB had a low mutagenic effect on the levorin-producing organism characterized by high activity and genetic homogenicity with respect to the colony morphology and antibiotic production. As for the organisms producing amphotericin B and mycoheptin characterized by high genetic heterogenicity, significant variation of all the features studied was observed on their exposure to the mutagen. Inspite of diverse reaction of the organisms producing levorin, amphotericin B and mycoheptin to the effect of NMB mutants with increased antibiotic production were obtained from the three cultures. The lethal and mutagenic effect of NMB on the mycoheptin-producing organism depended on the process of the spore DNA replication. The spores during the DNA replication period were least sensitive to the lethal effect of the mutagen and most mutable with the respect to the colony morphology. For selection of highly active and stable strains exposure to NMB of the spores of the mycoheptin-producing organism during replication of DNA proved to be more effective than that of the spores during the lag-phase.     

Differences in the consequences of the decay of radioactive64Cu atoms incorporated in cells under either in vitro or in vivo conditions

Summary To study the lethal effect of⁶⁴Cu under in vitro conditions, asynchronous mammalian cells were used. A lethal effect does exist as a consequence of the decay itself of a few⁶⁴Cu atoms incorporated in cellular DNA. This lethal effect is characterized by an exponential survival curve with no shoulder and no tail; it exists even for non-dividing cells. The lethal efficiency per decay is very high. To test the⁶⁴Cu lethal effect under in vivo conditions, experiments were performed with ascitic cells developing in mice. In this case, the lethal effect also exists, but it is not a function of the⁶⁴Cu doses injected in the mice. Faced with this puzzling result, a systemic approach was necessary to understand and counteract ascitic cells developing in mice.Dedicated to Prof. L.E. Feinendegen on the occasion of his 60th birthday     

Calmodulin antagonist W 13 prevents DNA repair after bleomycin treatment of human urological tumor cells growing on extracellular matrix

1. The combined application of DNA strand-scission agents (bleomycin) and inhibitors of recovery from lethal damage (calmodulin antagonist W-13) could be a novel and potentially important approach to cancer therapy. 2. As determined by alkaline elution, both DNA-DNA and DNA-protein cross-links in bleomycin-treated cells were revealed by the presence of the proteinase K assay. 3. This lethal effect could be potentiated by the addition of calmodulin antagonist W-13 that prevents the repair of DNA strand breaks and DNA cross-links caused by bleomycin. 4. The results indicated that combinations of bleomycin and W-13 were more effective than treatment with either single agent. Isobologram analysis suggests synergistic effect of these drugs. 5. Therefore, the rational use of combinations of DNA-strand-scission agents and inhibitors of recovery from lethal damage based on mechanistic considerations should result with improved therapeutic regimens for the treatment of cancer.     

A study of deleterious gene structure in plants using Markov chain Monte Carlo

The characteristics of deleterious genes have been of great interest in both theory and practice in genetics. Because of the complex genetic mechanism of these deleterious genes, most current studies try to estimate the overall magnitude of mortality effects on a population, which is characterized classically by the number of lethal equivalents. This number is a combination of several parameters, each of which has a distinct biological effect on genetic mortality. In conservation and breeding programs, it is important to be able to distinguish among different combinations of these parameters that lead to the same number of lethal equivalents, such as a large number of mildly deleterious genes or a few lethal genes, The ability to distinguish such parameter combinations requires more than one generation of mating. We propose a model for survival data from a two-generation mating experiment on the plant species Brassica rapa, and we enable inference with Markov chain Monte Carlo. This computational strategy is effective because a vast amount of missing genotype information must be accounted for. In addition to the lethal equivalents, the two-generation data provide separate information on the average intensity of mortality and the average number of deleterious genes carried by an individual. In our Markov chain Monte Carlo algorithm, we use a vector proposal distribution to overcome inefficiency of a single-site Gibbs sampler. Information about environmental effects is obtained from an outcrossing experiment conducted in parallel with the two-generation mating experiments.     

Modality of the antimitotic acitivity of the diamine cis-dichloroplatinum (II) studied on the plant cell]

In the order of decreasing doses, cis-platinum (II) diamine dichloride acting on root meristem of Allium sativum produces first an irreversible metostatic effect quickly followed by lethal effect, then a variable mitodepression accompanied by important chromosomal alterations: numerous disruptions and stickiness. This chromatoclasic effect persists at very low concentrations but stathmodieretic effect is not observed. The chromatoclasic activity of the compound is compared to that of other antitumoral drugs.     

Characterization of conditionally expressed mutants affecting age-specific survival in inbred lines of Drosophila melanogaster: lethal conditions and temperature-sensitive periods

The specific genetic basis of inbreeding depression is poorly understood. To address this question, two conditionally expressed lethal effects that were found to cause line-specific life span reductions in two separate inbred lines of Drosophila melanogaster were characterized phenotypically and genetically in terms of whether the accelerated mortality effects are dominant or recessive. The mortality effect in one line (I4) is potentially a temperature-sensitive semilethal that expresses in adult males only and is partially dominant. The other line (I10) responds as one would expect for a recessive lethal. It requires a cold shock for expression and is cold sensitive. Flies exhibiting this lethal condition responded as pupae and freshly eclosed imagoes. The effect is recessive in both males and females. The expression of the lethal effects in both lines is highly dependent upon environmental conditions. These results will serve as a basis for more detailed and mechanistic genetic research on inbreeding depression and are relevant to sex- and environment-specific effects on life span observed in quantitative trait loci studies using inbred lines.     

Schedule dependence of the lethal effects of 6-azauridine and cytosine arabinoside on murine leukemia cels (L5178Y) in culture

The antineoplastic drugs 6-azauridine and cytosine arabinoside exhibit a supra-additive lethal effect on murine L5178Y lymphoblasts if exposure to 6-azauridine precedes exposure to cytosine arabinoside; an additive effect is seen if cytosine arabinoside precedes 6-azauridine, while a sub-additive effect is obtained when the two drugs are present simultaneously. The potentiation of the effect of cytosine arabinoside by 6-azauridine increases for $\text{2}\text{1}\text{2}\text{hours}$ following the removal of the 6-azauridine from the culture.     

Use of 8-methoxypsoralen and light (lambda--365 nm) for studying repair in yeasts]

The lethal effect of 8-metoxypsoralen (8/MOP) plus light (lambda = 365 nm) on the haploid radioresistant and UV-sensitive strains of Saccharomyces cerevisiae was studied. The mutation uvs1 increased the sensitivity to the lethal effect of 8-MOP more than 2.8 times as compared to radioresistant strain. The method of repeated irradiation allowed to study kinetics of excision of monoadducts induced by 8-MOP. The mutant uvs1 was characterized by the absence of excision of monoadducts. The radioresistant strain removed monoadducts very efficiently (80%) after the incubation in complete liquid medium for 2.5 hours at 28 degrees before repeated irradiation. After the incubation of this strain in buffer (pH 7.0) monoadducts were removed considerably less efficiently (30%).     

Electrolytic reduction of nitroheterocyclic drugs leads to biologically important damage in DNA

The effect of electrolytic reduction of nitroimidazole drugs on biologically active DNA was studied. The results show that reduction of the drugs in the presence of DNA affects inactivation for both double-stranded (RF) and single-stranded phi X174 DNA. However, stable reduction products did not make a significant contribution to the lethal damage in DNA. This suggests that probably a short-lived intermediate of reduction of nitro-compounds is responsible for damage to DNA.     

The genetic structure of Balkan populations of Drosophila subobscura

Although Drosophila subobscura has been a model organism for European and American population geneticists, little information is available on the genetic structure of its natural populations. In this paper we report the estimates of some population parameters. We have used data from lethal allelism tests in four Balkan populations (Kamariste, Djerdap and Petnica in Serbia and Zanjic in Montenegro). In all populations, lethal genes were found to have a deleterious effect on heterozygotes. The N(e) values varied greatly from 370 (Petnica) to 19413 (Kamariste), depending on the habitat conditions and some environmental factors. Finally, gene flow between the Balkan populations was detected by the estimates of N(m) (from 4.68 in Petnica to 106.2 in Kamariste) and m (from 0.0041 in Djerdap to 0.0126 in Petnica). These results agree with those obtained in a previous study where the frequencies of allelism between populations were greater than predicted by independently arising lethal genes.     

Protective effect of chlorpromazine against the lethality of interleukin 1 in adrenalectomized or actinomycin D-sensitized mice

Interleukin 1 (IL-1) and Tumor Necrosis Factor (TNF) are thought to play a key role in septic shock and inflammation. We have tested the effect of dexamethasone (DEX) and chlorpromazine (CPZ) on the lethal effect of IL-1, TNF and endotoxin. Two different experimental models were used to sensitize mice to the lethal effect of IL-1: adrenalectomy and pretreatment with actinomycin D. CPZ (4 mg/kg) was found to protect mice against IL-1 and endotoxin toxicity in all cases, while DEX had a protective effect only in adrenalectomized mice. In contrast to its protective effect against IL-1 and endotoxin, CPZ did not protect mice against TNF. These findings might be useful in the analysis of the differences in the actions of IL-1 and TNF in vivo, and in the development of new drugs preventing their toxicity.     

Molecular nature of the pathogenic effect induced by B. cereus

The paper shows that biological properties of the DL-toxin are determined by structural particularities of this substance. It is supposed that the component B with mol. mass of 42,000 performs the ligand function enabling fixation of the toxin on a target cell. The component A with mol. mass of 37,000 has been characterized as an activator in reproducing oedematous and diarrheagenic effect. The component C activates the induction of the lethal effect. The optimal ratios of these components needed for reproduction of the respective biological effects are: B + A-6:1; B + C-5:1. In terms of specific activity, cereolysine is more active than the DL-toxin in inducing of the lethal effect. However, the later has bigger capability to break vascular permeability and it also reveals enteropathogenic properties. It is shown that among the exoproteins, produced by the strain 96 of B. cereus, the contents of the DL-toxin considerably exceeds the one of cereolysine. Thus, the DL-toxin plays the dominating role in reproducing oedematous, diarrheagenic and lethal effects.     

Correlation between In Vitro Cytotoxicity and In Vivo Lethal Activity in Mice of Epsilon Toxin Mutants from Clostridium perfringens

Epsilon toxin (Etx) from Clostridium perfringens is a pore-forming protein with a lethal effect on livestock, producing severe enterotoxemia characterized by general edema and neurological alterations. Site-specific mutations of the toxin are valuable tools to study the cellular and molecular mechanism of the toxin activity. In particular, mutants with paired cysteine substitutions that affect the membrane insertion domain behaved as dominant-negative inhibitors of toxin activity in MDCK cells. We produced similar mutants, together with a well-known non-toxic mutant (Etx-H106P), as green fluorescent protein (GFP) fusion proteins to perform in vivo studies in an acutely intoxicated mouse model. The mutant (GFP-Etx-I51C/A114C) had a lethal effect with generalized edema, and accumulated in the brain parenchyma due to its ability to cross the blood-brain barrier (BBB). In the renal system, this mutant had a cytotoxic effect on distal tubule epithelial cells. The other mutants studied (GFP-Etx-V56C/F118C and GFP-Etx-H106P) did not have a lethal effect or cross the BBB, and failed to induce a cytotoxic effect on renal epithelial cells. These data suggest a direct correlation between the lethal effect of the toxin, with its cytotoxic effect on the kidney distal tubule cells, and the ability to cross the BBB.     

Intracellular calcium antagonist protects cultured peritoneal macrophages against anthrax lethal toxin-induced cytotoxicity

The lethal toxin ofBacillus anthracis is central to the pathogenesis of anthrax. Using primary cultures of mouse peritoneal macrophages, we have demonstrated that intracellular calcium release inhibitors protect against anthrax lethal toxin-induced cytotoxicity. The cytolytic effect of anthrax lethal toxin was markedly reduced by dantrolene, an inhibitor of calcium release from intracellular calcium stores. Pretreatment of macrophages with cyclosporin A, which has been shown to be a potent inhibitor of calcium release from mitochondria, also protected cells against cytotoxicity. These results indicate that calcium release from intracellular store may be an essential step for the propagation of anthrax lethal toxin-induced cell damage in macrophages. Thus our findings suggest that dantrolene, cyclosporin A, and possibly other drugs affecting intracellular calcium pools might be effectively preventing the toxicity from anthrax lethal toxin. This revised version was published online in July 2006 with corrections to the Cover Date.     

Action of carminomycin on leukemic cell proliferation and some problems of its use in the combined therapy of leukemias

The effect of carminomycin on the mitotic cycle of the cells of the transplantable leukemia L-1210 and the therapeutic activity of other antitumor drugs, such as phopurine and cyclophosphane was studied on mice BDF1. It was found that the cells in phases S and G2 of the mitotic cycle were most sensitive to carminomycin. Transfer G1--S and phase G1 were characterized by resistance to the antibiotic effect. When carminomycin was used in combination with phopurine or cyclophosphane, clear dependence of the therapeutic efficacy on the treatment scheme was noted. Simultaneous administration of all the three drugs resulted in potentiation of the antileukemic effect. An analogous effect was observed when carminomycin was administered prior to phopurine or cyclophosphane. When the order of the drugs use was reverse, the efficacy of the combined therapy was significantly lower than the summation antileukemic effect of the drugs.     

Dominant lethal effect of 60Co gamma radiation in Biomphalaria glabrata (SAY, 1818)

The dominant lethal effects of gamma radiation of 60Co in the snail Biomphalaria glabrata were studied. Three groups of 13 wild-type snails were irradiated with single doses of 2.5; 10 and 20 Gy. Crossings were carried out at intervals of 7, 17, 23, 30 and 36 days after irradiation. The dominant lethal effect was observed only at the first crossing occurring 7 days after irradiation with 2.5 Gy. With 10 and 20 Gy, the induction of lethal mutations was detected at 7, 17 and 23 days after irradiation; a dose-response effect was observed. The effect was stronger 7 days after irradiation, decreasing in the succeeding crossings up to 30 days. Cell-killing effects on germ cells were detected in the crossings at 23 days and 30 days after irradiation with 20 Gy. After 36 days, frequencies of malformations resumed background levels; crossing rates partially recovered. These results show that gamma radiation affected all the stages of spermatogenesis. Germ cells at later phases were more sensitive to the mutagenic effect of radiation and the cell killing effects were observed on the youngest cells. This response was similar to the highly homogeneous pattern observed in widely different species and allowed us to estimate some parameters of spermatogenesis in B. glabrata.     

Effect of the inducers of cellular differentiation and of ionizing radiation on thymus lymphocytes: chromatin degradation and programmed cell death

The cytotoxic response of thymocytes to chemical inducers of differentiation does not represent a non-specific toxic action of these drugs. The death of thymocytes treated with the inducers or with gamma-rays is associated with internucleosomal chromatin fragmentation. All treatments are more effective on the most radiosensitive sub-population of these cells. This subpopulation is characterized by the maximal level of spontaneous DNA lesions. Incubation of thymocytes with the inducers of differentiation raises the level of these lesions. It is suggested that the processes of thymocyte death after the inducer treatment or irradiation and of cellular differentiation have features in common, and the capacity of thymocytes to limit or reverse the potentially lethal effects of these treatments is determined by the level of pre-existing spontaneous DNA lesions.     

Role of the activation gate in determining the extracellular potassium dependency of block of HERG by trapped drugs

Drug induced long QT syndrome (diLQTS) results primarily from block of the cardiac potassium channel HERG (human-ether-a-go-go related gene). In some cases long QT syndrome can result in the lethal arrhythmia torsade de pointes, an arrhythmia characterized by a rapid heart rate and severely compromised cardiac output. Many patients requiring medication present with serum potassium abnormalities due to a variety of conditions including gastrointestinal dysfunction, renal and endocrine disorders, diuretic use, and aging. Extracellular potassium influences HERG channel inactivation and can alter block of HERG by some drugs. However, block of HERG by a number of drugs is not sensitive to extracellular potassium. In this study, we show that block of WT HERG by bepridil and terfenadine, two drugs previously shown to be trapped inside the HERG channel after the channel closes, is insensitive to extracellular potassium over the range of 0 mM to 20 mM. We also show that bepridil block of the HERG mutant D540K, a mutant channel that is unable to trap drugs, is dependent on extracellular potassium, correlates with the permeant ion, and is independent of HERG inactivation. These results suggest that the lack of extracellular potassium dependency of block of HERG by some drugs may in part be related to the ability of these drugs to be trapped inside the channel after the channel closes.     

Mutagenic activity of dioxydine]

A previous evaluation of mutagenic activity of some drugs and perspective substances is carried out using indicator microorganisms. The mutagenicity of dioxydine, a drag with discovered antibacterial activity, is investigated. Dioxydine is shown to induce reversions in mutant of Salmonella typhimurium TA-1950, the indicator strain which demonstrates mutagenic activity of agents, producing mutations of base pair substitution type. Dioxydine proved to affect logariphmiically growing bacterial culture with great activity. Mutageni effect of dioxydine is not modified itself in microsomal oxidation system in vitro. Some data concerning participation of excision reparation enzyme (uvr-B+ gene product) in repair of lethal damages induced by dioxydine, have been obtained. The dioxydine ability to cause bacterial gene mutations in host mediated assay as well as dominant and recessive sex-linked lethal mutations in Drosophila is demonstrated. Dioxydine is capable of inducing chromosome aberrations in bone marrow cells and dominant lethal mutations in mouse germ cells.