The effectiveness of low-dose of eCG in timed AI Nelore (Bos indicus) heifers.

This study evaluated the efficacy of the administration of different doses of equine chorionic gonadotropin (eCG; 0 IU, 200 IU, or 300 IU) at the time of the progesterone device removal in 2-year-old Nelore (Bos indicus) heifers synchronized for fixed-timed artificial insemination (FTAI). On day 0 (D0), a total of 398 heifers received 2 mg of oestradiol benzoate i.m., 0.53 mg of cloprostenol i.m., and an eight-day previously used (second use) intravaginal device containing 1 g of progesterone (P4). Eight days later (D8), simultaneous with the P4 device removal, 0.5 mg of oestradiol cypionate i.m. and 0.53 mg of cloprostenol i.m. were administered. At the same time, heifers were randomly assigned to receive one of the following treatments: G-0 IU (n = 141; no eCG treatment), G-200 IU (n = 132; treated with 200 IU of eCG), and G-300 IU (n = 125; treated with 300 IU of eCG). FTAI was performed 48 h after the P4 device removal (D10). Ultrasonographic evaluations were performed at D0, D10, and D17. Heifers were scanned to measure the size of the largest follicle (LF), the presence, number, and size of the corpus luteum (CL), and the ovulation rate. Subsequently, at D40, the heifers underwent scanning to determine the pregnancy rate and identify any twin pregnancies. Additionally, at D70, scans were performed to assess pregnancy loss (PG). Data were analysed by orthogonal contrasts [C1 (eCG effect): control x (200 IU + 300 IU) and C2 (eCG dose effect): 200 IU × 300 IU]. On D0, CL presence was similar between the groups [G-0 IU = 65.2% (92/141), G-200 IU = 55.3% (73/132), and G-300 IU = 63.2% (79/125); p = .16]. No interactions between the presence of CL on D0 and eCG treatment were found for any of the variables (p > .05). The diameter of the LF at FTAI (D10) was not influenced by eCG treatment (p = .22) or eCG dose (p = .18). However, treatment with eCG increased the diameter of the CL at D17 (G-0 IU = 15.7 ± 0.3 mmb , G-200 IU = 16.6 ± 0.2 mma , and G-300 IU = 16.6 ± 0.3 mma ; p = .001), regardless of the dose used (p = .94). The ovulation rate was higher in heifers treated with eCG [G-0 IU = 79.4%b (112/141), G-200 IU = 90.2%a (119/132), and G-300 IU = 93.6%a (117/125); p = .002], but there was no effect of eCG dose (p = .36). Pregnancy per AI (P/AI) on D40 [G-0 IU = 32.6%b (46/141), G-200 IU = 42.4%a (56/132), and G-300 IU = 42.4%a (53/125); P = 0.05] and D70 [G-0 IU = 29.1%b (41/141), G-200 IU = 40.9%a (54/132), and G-300 IU = 40.8%a (51/125); p = .02] were higher on heifers that received eCG; however, no dose effect was observed for P/AI on D40 (p = .89) nor D70 (p = .98). Pregnancy loss between D40 and D70 tended to reduce (p = .07) in eCG-treated heifers without dose effect (p = .91). Heifers with CL at D0 presented a greater follicle diameter (LF) on D10 (With CL = 11.2 ± 0.2 mm and Without CL = 10.2 ± 0.2 mm; p = .05), CL diameter on D17 (With CL = 15.8 ± 0.03 mm and Without CL = 11.8 ± 0.6 mm; p = .01), and ovulation rate [With CL = 95.5% (233/244) and Without CL = 74.7% (115/154); p = .01]. However, no difference in pregnancy rate at D40 (p = .52) and D70 (p = .84) was found. In conclusion, eCG treatment increases ovulation and pregnancy rates of heifers submitted to a FTAI protocol. Furthermore, eCG treatment increases the diameter of the CL after FTAI and reduces pregnancy losses. No dose effect was observed, suggesting Nelore (Bos indicus) heifers respond to 200 IU of eCG treatment for FTAI.

An intense source for cold cluster ions of a specific composition.

The demand for nanoscale materials of ultra-high purity and narrow size distribution is addressed. Clusters of Au, C60, H2O, and serine are produced inside helium nanodroplets using a combination of ionization, mass filtering, collisions with atomic or molecular vapor, and electrostatic extraction, in a specific and novel sequence. The helium droplets are produced in an expansion of cold helium gas through a nozzle into vacuum. The droplets are ionized by electron bombardment and subjected to a mass filter. The ionic and mass-selected helium droplets are then guided through a vacuum chamber filled with atomic or molecular vapor where they collide and "pick up" the vapor. The dopants then agglomerate inside the helium droplets around charge centers to singly charged clusters. Evaporation of the helium droplets is induced by collisions in a helium-filled radio frequency (RF)-hexapole, which liberates the cluster ions from the host droplets. The clusters are analyzed with a time-of-flight mass spectrometer. It is demonstrated that using this sequence, the size distribution of the dopant cluster ions is distinctly narrower compared to ionization after pickup. Likewise, the ion cluster beam is more intense. The mass spectra show, as well, that ion clusters of the dopants can be produced with only few helium atoms attached, which will be important for messenger spectroscopy. All these findings are important for the scientific research of clusters and nanoscale materials in general.

[Responsibility due to medication errors in France: a study based on SHAM insurance data]. / Responsabilité liée aux erreurs médicamenteuses : données SHAM.

PURPOSE: The safe medication practices at the hospital constitute a major public health problem. Drug supply chain is a complex process, potentially source of errors and damages for the patient. SHAM insurances are the biggest French provider of medical liability insurances and a relevant source of data on the health care complications. METHODS: The main objective of the study was to analyze the type and cause of medication errors declared to SHAM and having led to a conviction by a court. We did a retrospective study on insurance claims provided by SHAM insurances with a medication error and leading to a condemnation over a 6-year period (between 2005 and 2010). RESULTS: Thirty-one cases were analysed, 21 for scheduled activity and 10 for emergency activity. Consequences of claims were mostly serious (12 deaths, 14 serious complications, 5 simple complications). The types of medication errors were a drug monitoring error (11 cases), an administration error (5 cases), an overdose (6 cases), an allergy (4 cases), a contraindication (3 cases) and an omission (2 cases). Intravenous route of administration was involved in 19 of 31 cases (61%). The causes identified by the court expert were an error related to service organization (11), an error related to medical practice (11) or nursing practice (13). Only one claim was due to the hospital pharmacy. CONCLUSION: The claim related to drug supply chain is infrequent but potentially serious. These data should help strengthen quality approach in risk management.

Longitudinal trends in utilization of endocrine therapies for breast cancer: an international comparison.

WHAT IS KNOWN AND OBJECTIVE: Endocrine therapy is an effective treatment for post-menopausal women with 'oestrogen receptor-positive' invasive breast cancers. There are two main types of endocrine therapies: selective oestrogen receptor modulators (tamoxifen) and aromatase inhibitors (anastrozole, letrozole and exemestane). The aim of this study was to compare the patterns of use of endocrine therapies for breast cancer in women between nine developed countries. METHODS: A longitudinal, cross-national drug utilization study was conducted. The endocrine therapies included were tamoxifen and the aromatase inhibitors: anastrozole, letrozole and exemestane. Annual drug utilization data were collected from Australia, Denmark, England, Finland, France, Iceland, the Netherlands, Norway and Sweden over the period 2001-2012. Utilization was measured in DDD/1000 inhabitants/day and was also adjusted for breast cancer incidence and female population statistics. RESULTS AND DISCUSSION: Total use of endocrine therapies either increased or remained steady in all countries. Total endocrine therapy usage was consistently highest in England and France. Norway showed the lowest usage of endocrine therapies overall, using only 1.80 DDD/1000 inhabitants/day in 2012. Downward trends in tamoxifen use and upward trends in aromatase inhibitors were seen across all countries over the study period. By 2012, aromatase inhibitors represented over half of total endocrine therapy use in all countries, and as high as 74% and 80% in France and Denmark, respectively. WHAT IS NEW AND CONCLUSION: Our analysis found a shift in use of endocrine therapy from tamoxifen to aromatase inhibitors. This trend is consistent with major clinical guidelines endorsing preferential use of aromatase inhibitors in post-menopausal women. Stabilization or small increase in tamoxifen use in the recent years may reflect the recognition of tamoxifen as still an appropriate first-line treatment. The similarity in utilization patterns may be due to the relatively comparable healthcare systems in the countries, namely universal health insurance and pharmaceutical coverage. Differences in utilization observed could be due to differences in breast cancer incidence, prescribing behaviours, interpretation of new trial evidence, and timing of drug marketing approval and reimbursement between countries.

Comparison of different methods for an accurate assessment of cytotoxicity in the in vitro micronucleus test. I. Theoretical aspects.

A decrease in the cytokinesis-block proliferation index (CBPI) or replication index (RI) is routinely used to determine cytotoxicity of a test compound and therefore the choice of its appropriate test concentration for the in vitro micronucleus (MN) test conducted in the presence of cytochalasin B. As a number of laboratories prefer to conduct the in vitro MN test in the absence of cytochalasin B, it is important that selected test concentrations, based on cytotoxicity, should be similar to what they would have been if cytochalasin B had been used, and should be relevant of a true cytotoxicity. By using models to analyse the dynamics of the cell cultures with and without cytochalasin B we have compared different methods for evaluation of cytotoxicity, and demonstrate that relative decrease in population doubling or relative increase in cell counts are the most appropriate measures of cytotoxicity to compare with reduction in CBPI or RI.

An in vitro study on 5-HT6 receptor antagonist induced hepatotoxicity based on biochemical assays and toxicogenomics.

We investigated the effects of two 5-HT(6) receptor antagonists on rat primary hepatocytes using a combined biochemical and toxicogenomics approach. Both compounds share the same pharmacological target, but displayed strikingly different toxicity profiles in pre-clinical animal studies: While R7199 caused hepatic steatosis in rats, no hepatotoxicity was observed with R0074. Here, we partially reproduced the steatosis findings seen in vivo using primary rat hepatocytes. Biochemical analyses and gene expression results generally supported the findings observed in the animal model and also allowed the differentiation of both compounds with regards to hepatotoxic potential. In particular, the induction of Cyp 2B and Cyp 3A1 directly correlates to the findings in the livers of treated animals. The effects on genes of the steroideogenic pathway relate to the deregulation of cholesterol homeostasis. We also observed the inhibition of beta-oxidation, indicating impaired lipid metabolism. Hence, gene expression analysis in combination with biochemical parameters can provide additional insight into the possible mechanisms underlying adverse events.

Absence of interference of serum IgGs from patients with breast cancer and thyroid autoimmunity on the function of human iodide symporter gene stably transfected in CHO cells.

The cause of the association between breast cancer (BC) and thyroid autoimmunity is still unknown. Na+/I- symporter (NIS) is highly expressed in BC cells, and previous studies demonstrated that iodine content in BC is lower than in remote normal breast tissue, suggesting a disorder of iodide uptake in BC. In this study, we evaluated the presence of putative serum autoantibodies able to block the function of NIS in BC patients with thyroid autoimmunity. IgGs were obtained from: a) 11 patients with BC and high antithyroglobulin (TgAb) and antithyroperoxidase (TPOAb) autoantibodies serum concentration; b) 34 patients with Hashimoto's thyroiditis (HT) (1 was euthyroid, 4 had subclinical hypothyroidism and 29 were overtly hypothyroid); c) 15 control subjects. The biological activity of NIS was studied using a chinese hamster ovary (CHO) cell line stably expressing NIS (NIS-CHO). The course of iodide accumulation in NIS-CHO was studied after addition of Na125 I in culture medium. The accumulation of iodide linearly increased between 2 and 10 min, reaching a plateau at 45 min. The preincubation of NIS-CHO with IgGs purified from sera of BC with the highest levels of TPOAb and TgAb caused an inhibition of iodine uptake of no more than 5%. Similar results were obtained using IgGs purified from patients with HT and control subjects. Our data showed no interference of autoantibodies on iodine uptake in patients with BC and thyroid autoimmunity and the very low percentage of inhibition of iodine uptake cannot explain the lower content of iodine in BC tissue.

Effect of two 5-HT6 receptor antagonists on the rat liver: a molecular approach.

Serotonin is involved in disorders of the central nervous system; thus, specific 5-HT(6) receptor antagonists have therapeutic potential. Nevertheless, preclinical tests showed that Ro 65-7199 caused hepatic steatosis. Here, we investigated the hepatic effects of Ro 65-7199 and Ro 66-0074 using toxicogenomics. The profiles obtained after exposure of rats to both compounds clearly show that two pharmacologically closely related compounds with different toxicological profiles can be distinguished based on gene expression profiles. Moreover, side effects can be detected earlier with toxicogenomics than with conventional end points. A possible link between the sterol metabolic pathway, the induction of CYP2B, and the hepatic fat accumulation was also established. Summarizing, gene expression profiles allow both compounds to be distinguished according to their toxicity and provide mechanistic insights. The results clearly show the power of toxicogenomics as a tool for obtaining characteristic fingerprints at early time-points and for generating mechanistic hypotheses.

In vitro micronucleus assay with Chinese hamster V79 cells - results of a collaborative study with in situ exposure to 26 chemical substances.

A collaborative study with 10 participating laboratories was conducted to evaluate a test protocol for the performance of the in vitro micronucleus (MN) test using the V79 cell line with one treatment and one sampling time only. A total of 26 coded substances were tested in this study for MN-inducing properties. Three substances were tested by all 10 laboratories and 23 substances were tested by three or four laboratories in parallel. Six aneugenic, 7 clastogenic and 6 non-genotoxic chemicals were uniformly recognised as such by all laboratories. Three chemicals were tested uniformly negative by three laboratories although also clastogenic properties have been reported for these substances. Another set of three clastogenic substances showed inconsistent results and one non-clastogenic substance was found to be positive by one out of three laboratories. Within the study, the applicability of the determination of a proliferation index (PI) as an internal cytotoxicity parameter in comparison with the determination of the mitotic index (MI) was also evaluated. Both parameters were found to be useful for the interpretation of the MN test result with regard to the control of cell cycle kinetics and the mode of action for MN induction. The MN test in vitro was found to be easy to perform and its results were mainly in accordance with results from chromosomal aberration tests in vitro.

Two-dimensional database of mouse liver proteins: changes in hepatic protein levels following treatment with acetaminophen or its nontoxic regioisomer 3-acetamidophenol.

Overdose of acetaminophen (APAP) causes acute hepatotoxicity in rodents and man. The mechanism underlying APAP-induced liver injury remains unclear, but experimental evidence strongly suggests that activation of APAP and subsequent formation of protein adducts are involved in hepatotoxicity. Using proteomics technologies, we constructed a two-dimensional protein database for mouse liver, comprising 256 different gene products and investigated the proteins affected after APAP-induced hepatotoxicity. Adult male mice received a single dose of APAP (100 or 300 mg/kg) or its nontoxic regioisomer 3-acetamidophenol (AMAP, 300 mg/kg). The extent of liver damage was assessed 8 h after administration by increased liver enzyme release and histopathology. Changes in the protein level were studied by comparison of the intensities of the corresponding spots on two-dimensional (2-D) gels. The expression level of about 35 of the identified proteins was modified due to treatment with APAP or AMAP. The observed changes were usually in the order of 10-50% of the control value and were more marked in the high- than in the low-dose of APAP-treated animals. Most of the changes caused by AMAP occurred in a subset of the proteins modified by APAP. Many of the proteins showing changed expression levels are either known targets for covalent modification by N-acetyl-p-benzoquinoneimine (NAPQI) or involved in the regulation of mechanisms that are believed to drive APAP-induced hepatotoxicity.

Report from the In Vitro Micronucleus Assay Working Group.

At the Washington International Workshop on Genotoxicity Test Procedures (March 25-26, 1999), the current methodologies and data for the in vitro micronucleus test were reviewed. From this, guidelines for the conduct of specific aspects of the protocol were developed. Because there are a number of important in vitro micronucleus validation studies in progress, it was not possible to design a definitive, internationally harmonized protocol at this time. Agreement was achieved on the following topics: Cells. The choice of cells is flexible, yet the choice of cell type should be justified and take into consideration doubling time, spontaneous frequency of micronuclei, and genetic background. Slide preparation. A fixation method that preserves the cytoplasm and cytoplasmic boundaries, and minimizes clumping should be used. Use of fluorescent DNA-specific dyes is encouraged for better detection of small micronuclei. Analysis. Micronuclei should have a diameter less than one-third of the main nucleus, and should be clearly distinguishable from the main nucleus. In the cytokinesis-block method, binucleated cells selected for analysis should have two clearly distinguishable main nuclei. Cells where the main nucleus(ei) is undergoing apoptosis should not be scored for micronuclei because the assumed micronuclei may have been the result of nuclear fragmentation during the apoptotic process. Toxicity. Cytotoxicity can be measured by various methods including cell growth, cell counts, nucleation (i.e., percent binucleated), division/proliferation index, confluence. A majority of the group recommended that the highest concentration should induce at least 50% cytotoxicity (by whatever measure is selected). Cytochalasin B. There is much debate regarding the use of cytochalasin B. For human lymphocytes, the use of cytochalasin B (6 microg/ml [lymphocytes cultured from whole blood cells] and 3-6 microg/ml [isolated lymphocyte cultures]) is recommended. For cell lines, because there were no definitive data showing a clear advantage or disadvantage of the use of cytochalasin B for a variety of chemicals, the majority opinion of the group was that at this time, the use of cytochalasin B for cell lines is considered optional. Further studies (many chemicals of a variety of potencies, tested both with and without cytochalasin B) are clearly needed to resolve this issue. Number of doses. At least three concentrations should be scored for micronuclei. Treatment/harvest times. At this time, there are not enough data to define the most appropriate treatment/harvest times. Following the principles of the in vitro metaphase assay (with or without metabolic activation), it was agreed that there was a need for a short treatment followed by a recovery time in the absence of test chemical, there was a need for a long treatment (maybe with and without recovery time), and ideally, treatment should cover cells in different cell cycle stages.

Thresholds in genotoxicity responses.

It has been commonly accepted that risk assessments of genotoxic chemicals are based on linear extrapolation methods. However, there is substantial evidence that some chemicals may be genotoxic only at high doses by mechanisms that do not occur at low doses, or only under specific conditions in genotoxicity assays, but are inactive at concentrations within the range of human exposure levels. There are a variety of possible mechanisms of thresholded genotoxicity, including disruption of cell division and chromosome segregation, inhibition of DNA synthesis, overloading of oxidative defence mechanisms, metabolism or plasma binding capacity, disturbances of metal homeostasis, cytotoxicity and physiological perturbations in in vivo assays. The degrees of evidence supporting the proposed mechanisms are variable and not all are sufficiently robust to be universally accepted as yet by the scientific community. However, a survey of industrial companies indicated that data have been accepted by some regulatory authorities indicating thresholds contributing to genotoxicity responses.

Genotoxicity testing of biotechnology-derived products. Report of a GUM task force. Gesellschaft für Umweltmutationsforschung.

Various aspects of genotoxicity testing of biotechnology-derived products are discussed based on information gathered from a questionnaire which was sent to about 30 predominantly European companies. Feedback was received from 13 companies on 78 compounds, mostly recombinant proteins but also on a number of nonrecombinant proteins, which had been assessed for genotoxicity in a total of 177 tests. Four of the 78 compounds appeared to elicit reproducible genotoxic effects. For one of these compounds, the activity could be related to a nonpeptidic linker molecule. No scientifically convincing rationale for the other three compounds could be established, although, at least for two compounds, their activity may be connected with the enzymatic/hormonal activity. In addition to the survey, published reports on genotoxicity testing of biotechnology products were reviewed. The data are discussed relative to whether genotoxicity testing is a valuable exercise when assessing potentially toxic liabilities of biotechnology-derived compounds. It is concluded that genotoxicity testing is generally inappropriate and unnecessary, a position which is in accordance with the available guidelines addressing this area. For the 'average' protein, electrophilic reactions are difficult to envision. Indirect reactions via DNA metabolism and growth regulation seem possible for only very specific proteins such as nucleases, growth factors, cytokines. No information on testing of different types of biotechnology-derived products (e.g., ribozymes, antisense-oligonucleotides, DNA vaccines) has been received in the questionnaires. Discussion of their potential to cause genotoxic changes was based on literature reports. Even for those products for which concerns of genotoxic/tumourigenic potential cannot be completely ruled out, e.g., because of their interaction with DNA metabolism or proliferation control, the performance of standard genotoxicity assays generally appears to be of little value. All information, including also information on the occurrence of genotoxic impurities, has been utilized to formulate a decision tree approach for the genotoxicity testing of biotechnology-derived products.

Suppression of mutagenic activity of a series of 5HT2c receptor agonists by the incorporation of a gem-dimethyl group: SAR using the Ames test and a DNA unwinding assay.

A positive result in the Ames test is generally taken as a strong indication for a genotoxic (i.e. DNA damaging) property of the test compound, often sufficient to cause termination of its development as a new therapeutic agent. A number of serotonin receptor ligands have been tested for their mutagenic potential in the Ames assay at an early stage of development. For several compounds increases in the number of revertant colonies were observed in strain TA1537. Consequently, structure-activity relationship investigations were undertaken to search for compounds without mutagenic liability. All compounds are three ringed heterocyclic structures consisting of a benzene ring, a central (generally non-aromatic) 5- or 6-membered ring and a pyrrole or pyrazole ring. Using a gel shift assay we provide evidence that the observed genotoxic effects are strongly influenced by the intercalating properties of the compounds. The highest mutagenic response was seen with a compound possessing a central aromatic ring. The mutagenic activity of the naphthaleno derivatives appears to be stronger when compared with the indeno compounds, probably because of the less curved structure. Dimethyl substitution of the indeno substructure reduces the intercalating ability of the compounds and leads to loss of mutagenic activity. Pyrazole analogues of both indeno and naphthaleno structures appear to produce stronger mutagenic responses than the pyrrole derivatives.

Aneuploidy: a report of an ECETOC task force.

Aneuploidy plays a significant role in adverse human health conditions including birth defects, pregnancy wastage and cancer. Although there is clear evidence of chemically induced aneuploidy in experimental systems, to date there are insufficient data to determine with certainty if chemically induced aneuploidy contributes to human disease. However, since there is no reason to assume that chemically induced aneuploidy will not occur in human beings, it is prudent to address the aneugenic potential of chemicals in the safety assessment process. A wide range of methods has been described for the detection of chemically induced aneuploidy including subcellular systems, tests with fungi, plants and Drosophila as well as in vitro mammalian systems and in vivo mammalian somatic and germ cell assays. However, none of these methods is sufficiently validated or widely used in routine screening. Underlying the efforts to develop aneuploidy-specific assays is the presumption that current genetic toxicology tests do not detected chemicals that have aneuploidy-inducing potential. To address this, we have critically evaluated data from standard genetic toxicology assays for 16 known or suspected aneugens. The conclusions from the review are listed below. 1. At present there are only nine chemicals that can be classified as definitive aneugens, as determined by positive results in in vivo rodent assays. 2. As expected, the majority of definitive and suspected aneugens are negative in the bacterial mutation assay. 3. The majority of definitive aneugens evaluated induce polyploidy in vitro. With few exception, they also induced structural chromosome aberrations in vitro. 4. All of the definitive aneugens that have been sufficiently tested induce micronuclei in rodent bone marrow cells in vivo. A number of these chemicals also induced structural chromosome aberrations in vivo. 5. There is no evidence for a unique germ cell aneugen, that is a chemical that induces aneuploidy in germ cells and not in somatic cells. Furthermore, an analysis of several databases indicates the proportion of chemicals which induce polyploidy and not chromosome aberrations in vitro is low. Based on these conclusions, the following recommendations are made: for screening purposes, a standard genotoxicity test battery (including an in vitro cytogenetic assay with an assessment of polyploidy and clastogenicity at the same harvest time) should be performed; in the absence of polyploidy induction in vitro no further evaluation of aneuploidy-inducing potential is needed; if polyploidy is observed, in vitro follow-up testing to investigate further the aneuploidy-inducing potential should be conducted; such follow-up testing will generally start with the conduct of a standard in vivo somatic cell micronucleus assay; if the in vivo somatic cell micronucleus assay is negative, with adequate evidence of exposure of the bone marrow to the test compound, no further testing of aneuploidy-inducing potential is needed; if the in vivo somatic cell micronucleus assay is positive, further information on mechanisms of micronucleus induction can be obtained by using kinetochore/centromeric staining in vitro and/or in vivo; an assessment of potential germ cell aneuploidy activity may then be considered; aneuploidy induction which does not involve the direct interaction of a chemical or its metabolite(s) with DNA is expected to have a threshold. This must be considered in the risk assessment of such chemicals; this is not addressed by current risk assessment guidelines.

The photomutagenicity of fluoroquinolones and other drugs.

Induction of DNA damage as a consequence of exposure to UV light has been established as the major and still increasing cause of skin cancer. Absorption of the photon energy may be either directly by the DNA molecules (for wavelengths < 320 nm) or may be by endogenous or exogenous chemicals (sensitizers) with the potential of energy or electron transfer to DNA. Oxygen-mediated reactions (often called type II reactions) appear to be the most important mechanism since molecular oxygen is a good and abundant substrate for triplet excited sensitizers. Energy transfer to molecular oxygen is possible for wavelengths in the near UV and in the visible part of the solar spectrum since the energy of the excited oxygen molecule ((1)O2*) is comparatively low. A few light-absorbing pharmaceuticals have long been known to cause photo(geno)toxic effects. Notably psoralene and chlorpromazine derivatives have been established as photomutagens and the reaction mechanisms have been identified. The fluoroquinolone antibiotics have more recently been recognized as being photomutagenic. The type of DNA damage and the modulation by antioxidants indicate the involvement of reactive oxygen species (ROS) but other mechanisms are also reported at least for some derivatives. In routine genotoxicity studies we observed a photomutagenic activity of a compound under development as an anxiolytic agent in the Ames tester strain TA102 at 'normal laboratory illumination' conditions. Further investigations showed strong photogenotoxic activity in tests for gene mutations and chromosomal aberrations in mammalian cells. The compound proved to be a potent (1)O2-producer. The finding led to termination of development but in the course of studies several structural analogues have been tested for which structure activity relationships will be described. The relevance of photogenotoxic properties of drugs for predicting adverse effects in man will be discussed.

Comparative evaluation of the in vitro micronucleus test and the in vitro chromosome aberration test: industrial experience.

Because of its rapidness, simplicity and potential for automation, the measurement of micronucleated cells in vivo is not only equivalent to the analysis of chromosome aberrations, but often even preferred within routine genotoxicity testing. In order to evaluate the correlation between the in vitro micronucleus assay (MNT) and the in vitro chromosome aberration test (CA), we collected data from four pharmaceutical companies obtained either in Chinese hamster cell lines (CHO-K5, CHO-K1, V79) or in human peripheral blood lymphocytes. Among the 57 compounds included in this comparison, 45 compounds gave rise to concordant results in both assays (26 compounds negative in both assays; 19 compounds positive in both assays). The high percentage of concordance, i.e. about 79% is very promising and can be even increased to about 88% by omitting the 3 aneugenic compounds and 2 compounds inducing endoreduplicated chromosomes which were found positive only in the in vitro MNT. The results are remarkable in particular considering that most of the compounds evaluated are 'standard' pharmaceutical compounds and thus are at most weak inducers of chromosome damage. Our comparison strongly supports that the in vitro micronucleus test is a suitable alternative to the in vitro chromosome aberration assay. Moreover, the MNT has the potential of not only detecting clastogens but additionally aneuploidy inducing chemicals.

Mutation induction and mutation spectra of S. typhimurium TA100 after exposure to isohistidines.

L-Isohistidine and D,L-isohistidine, but not D-isohistidine, caused an increase of the number of mutant colonies in S. typhimurium strain TA100. Spontaneous and also sodium azide or 2-aminoanthracene induced mutant numbers were enhanced by L-isohistidine and by an isomeric mixture of D,L- and L-isohistidine. These effects could not be attributed to a growth-enhancing property. The colony probe hybridization procedure was used to investigate the effects of the histidines on the spontaneous and azide-induced spectra of the hisG46 allele in strain TA100. D,L-Isohistidine, but not the D-isomer, caused and increase of transitions (CCC-->CTC) and transversions (CCC-->CAC) in the spontaneous spectrum. Sodium azide alone induced a strong increase of CCC-->CTC transitions; combination with the D,L-isohistidine led to a further enhancement of this type of base substitutions, whereas with the L-isomer, no such effect was observed. This supports the hypothesis that the activity of D,L-isohistidine is probably not due to DNA-damaging properties, but rather to indirect mechanisms, such as enhancement of the infidelity of DNA replication and/or interference with DNA-repair or proofreading functions.