Teratogenicity Assay Research Articles

Evaluation of genotoxicity and teratogenicity of phillyrin

Phillyrin is extracted from Forsythia suspensa (Thunb.) Vahl, is significantly higher in (unripe Forsythiae Fructus) Qing qiao than in (ripe Forsythiae Fructus) Lao qiao fruits of the plant. However, the toxicity of phillyrin has not been adequately investigated. The study investigates the genetic and teratogenic effects of phillyrin to determine its safety profile. Assessing the genotoxicity and teratogenicity of phillyrin involved various tests, such as the bacterial reverse mutation assay, mammalian erythrocyte micronucleus assay, spermatocyte chromosome aberration assay, and teratogenicity assay. The results demonstrated that phillyrin exhibited no discernible impact on the following: number of colonies that spontaneously revert for Salmonella typhimurium TA 97, TA98, TA100, TA102, and TA1535, frequency of bone marrow polychromatic erythrocytes, and the rate of chromosomal aberrations. In the teratogenicity test, the pregnant rats exhibited no signs of toxicity or abnormal changes, and the growth, embryonic development, and visual anatomy of each pup were normal. In comparison with the negative control group, there were no significant differences in fetal body weight, mortality, deformity rate, malformed nest rate, gravid uterus weight, average number of fetuses per litter, fetal body length, or visceral and skeletal development in each dose group. In conclusion, these findings provide evidence that phillyrin does not exhibit genotoxic or teratogenic effects, supporting its potential safety for pharmacological applications.

Evaluation of temperature- and ethanol-related developmental degree variations by a new scoring system (FETAX-score) applicable to Frog Embryo Teratogenicity Assay: Xenopus

The aim of the present work is to propose a new quantitative assessment method (FETAX-score) for determining the degree of Xenopus laevis embryo development intended for use in embryotoxicity studies. Inspired by a similar scoring system used to evaluate developmental delays (young-for-age phenotypes) in rat embryos cultured in vitro, the FETAX-score was established by considering seven morphological features (head, naris, mouth, lower jaw, tentacles, intestine, anus) that are easily evaluable in tadpoles during the late stages of development at the conclusion of the test. Given that X. laevis development is temperature-dependent and that temperatures below 14°C and above 26°C are teratogenic, the FETAX-score was tested in embryos maintained at 17, 20, 23 and 26°C. No abnormalities were observed in any group, while the total score was temperature-related, suggesting that the FETAX-score is sensitive to moderate distress that does not influence general morphology. Intestine and anus were the least sensitive structures to temperature variations. To assess the applicability of the FETAX-score in developmental toxicological studies, we evaluated FETAX-score in tadpoles exposed during the morphogenetic period to Ethanol (Eth) at concentrations of 0, 0.25, 0.5, 1, 1.5, and 2 % v/v. Gross malformations were observed only in tadpoles from the Eth 2 % group. By contrast, data analysis of the other Eth groups showed dose-related reductions in the FETAX-score. Tentacles were the most sensitive structures to Eth-related delays. These results support the use of the FETAX-score to quantitatively assess developmental deviations in FETAX embryotoxicity studies.

Ethnomycological Expedition: Mycophagy, Species Listing and Toxicity Assessment of Macrofungi in Bambang Nueva Vizcaya, Philippines

In the Philippines, studies have been conducted on the taxonomic identification of wild macrofungi. However, data about the utilization, diversity, and toxicity of wild macrofungi are still insufficient especially from the mountainous hard to reach areas of the country. Therefore, this study was conducted to document the macrofungal species utilization and diversity of wild macrofungi in Barangay Buag, Bambang, Nueva Vizcaya to come up with a species listing of macrofungi in the area and screening of selected macrofungi for its teratogenic and cytotoxic activity using Danio rerio embryo and Artemia salina nauplii. Purposive sampling method was used in the collection of the macrofungi species. Results showed that 15 macrofungi were declared as utilized either as food, medicine, or as an additional source of income by the community. During the collection, 53 macrofungal samples were collected; of these eight were utilized and identified by the community with their local names. However, only seven of the eight species were identified scientifically using their morphological and molecular characteristics namely: Auricularia cornea, Schizophyllum commune, Coprinus disseminatus, Mycena sp., Ganoderma lucidum, Polyporous tenuiculus, and Volvariella volvacea. Out of the 53 macrofungal samples collected 44 (39 morphologically and 5 molecularly) were identified, 24 were identified to species level, and 20 were identified to genus level, all belonging to 20 families 32 genera and 24 species. The molecularly identified macrofungi are Ganoderma neo-japonicum, Ganoderma applanatum, Polyporus tenuiculus, Agaricus erectosquamosus and Helotiales sp. Lyophilized hot water extract (LHWE) of Ganoderma applanatum and Thelepora sp. generally exhibited teratogenic effects against developing D. rerio embryos during the teratogenicity assay. The varying treatment concentration of the LHWE of G. applanatum and Thelephora sp. significantly revealed toxic effects such as coagulation and no heartbeat, for teratogenic effects of the extract, growth retardation, scoliosis, malformation of head, and malformation of tail were the noted effects on the developing D. rerio. Furthermore, the other abnormalities that were not mentioned as teratogenic endpoints were observed such as unpigmented tail, and curved embryo body. The LHWE of G. applanatum and Thelephora sp. projected an LC50 of 979.33 ppm (mildly toxic) and 10.763 ppm (highly toxic) respectively which was based on the brine shrimp lethality assay.

Polymethylmethacrylate nanoplastics can cause developmental malformations in early life stages of Xenopus laevis

Polymethylmethacrylate (PMMA) production has increased almost 20% over the last years. With its release into the aquatic environment, its breakdown or degradation to nano dimensions (nanoplastics-NPLs) due to biological and physical/mechanical action is, theoretically, anticipated. The occurrence of PMMA-NPLs in aquatic ecosystems may thus cause adverse effects particularly to early life stages of amphibians, which may be in contact with PMMA-NPLs suspended in the water column or deposited in upper layers of the sediments. Accordingly, this work aimed at assessing the effects of PMMA-NPLs to aquatic early life stages of the model anuran species Xenopus laevis. To attain this objective, two types of toxicity assays were carried out by exposing embryos [Nieuwkoop and Faber (NF) stage 8-11] or tadpoles (NF 45) to three concentrations of PMMA-NPLs (1, 100 and 1000 μg/L): i) 96-h embryo teratogenicity assay, where survival, malformation, and total body length (BL) of embryos were assessed; and ii) 48-h feeding rate assay, where survival, feeding (FR), malformations and growth rates (body weight-BW and BL) of tadpoles were evaluated. PMMA-NPLs exposure had no significant effects on mortality, malformations of X. laevis embryos but BL was lower at 1000 μg PMMA-NPLs/L. In tadpoles, no effects on survival or FR were observed after exposure to PMMA-NPLs, but significant changes occured in BW and BL. Moreover, anatomical changes in the abdominal region (externalization of the gut) were observed in 62.5% of the tadpoles exposed to 1000 μg PMMA-NPLs/L. Despite the lack of knowledge regarding the environmental levels of NPLs, it is expected that sediments constitute a sink for these contaminants, where they can become available for organisms that, like tadpoles, feed on the organic matter at the surface of sediments. Considering the continuous release and subsequent accumulation of PMMA, the malformations obtained in the feeding assays suggest that, in the future, these nano-polymers may constitute a risk for aquatic life stages of amphibians.

Concordance of 3 alternative teratogenicity assays with results from corresponding in vivo embryo-fetal development studies: Final report from the International Consortium for Innovation and Quality in Pharmaceutical Development (IQ) DruSafe working group 2

An IQ DruSafe working group evaluated the concordance of 3 alternative teratogenicity assays (rat whole embryo culture, rWEC; zebrafish embryo culture, ZEC; and murine embryonic stem cells, mESC) with findings from rat or rabbit embryo-fetal development (EFD) studies. Data for 90 individual compounds from 9 companies were entered into a database. In vivo findings were deemed positive if malformations or embryo-fetal lethality were reported in either species. Each company used their own criteria for deciding whether the alternative assay predicted the in vivo findings. Standard concordance parameters were calculated, positive and negative predictive values (PPV and NPV) were adjusted for the aggregate portfolio prevalence of positive compounds (established by a survey of participating companies), and positive and negative likelihood ratios (LR+ and iLR-) were calculated. Of the 3 assays, only rWEC data were robustly predictive, particularly for negative predictions (NPVadj = 92%). However, both LR+ (4.92) and iLR- (4.72) were statistically significant for the rWEC assay. When analyzed separately for rats, the NPVadj and iLR-values for the rWEC assay increased to 96% and 9.75, respectively. These data suggest that a negative rWEC outcome could defer or replace a rat EFD study in certain regulatory settings.

Design, Synthesis, Molecular Docking Studies and Antitubercular Evaluation of Hexahydroquinolin-2-yl Benzamide Derivatives

A new series of hexahydroquinolin-2-yl benzamide derivatives (BZ1-10) were designed and synthesized. The synthesized compounds were characterized by 1H NMR, IR and ESI-MS spectra and also subjected for molecular docking studies with the target DNA gyrase enzyme (PBD ID: 4B6C). The molecular docking results of synthesized derivatives indicated the best docking score of -5.105 and -5.02 for BZ9 and BZ4, respectively. All the synthesized compounds were screened for antitubercular activity against H37RV strain, among all, two compounds exhibited significant activity at 12.5 μg/mL and 25 μg/mL concentrations. Thus, the MIC values are in between range of 12.5 and 6.25 μg/mL concentrations. The teratogenicity assay of synthesized compounds was performed in zebrafish larvae, out of the ten compounds, BZ4, BZ6 and BZ8 compounds were found to safer at 0.5 μM concentration without any abnormalities.

AEO-7 surfactant is \u201csuper toxic\u201d and induces severe cardiac, liver and locomotion damage in zebrafish embryos

BackgroundFatty alcohol polyoxyethylene ether-7 (AEO-7), a non-ionic surfactant, has recently been receiving extensive attention from the ocean pipeline industry for its ability to inhibit corrosion. However, the present lack of information concerning the potential environmental toxicity of AEO-7, especially towards aquatic organisms, is a major impediment to its wider application. Here, we assess potential adverse effects of AEO-7 on zebrafish embryos employing a variety of assays, including (i) a mortality/survival assay which allowed the median lethal concentration (LC50) to be calculated; (ii) a teratogenicity assay on the basis of which the no observed effect concentration (NOEC) was determined; and (iii) specific assays of cardiotoxicity, neurotoxicity (based on locomotion), hematopoietic toxicity (the level of hemoglobin as revealed by o-dianisidine staining) and hepatotoxicity (liver steatosis and yolk retention examined by staining with Oil Red O).ResultsAEO-7 caused mortality with a calculated LC50 of 15.35 μg/L, which, according to the U.S. Fish and Wildlife Service (USFWS) Acute Toxicity Rating scale, should be considered “super toxic”. Although at its NOEC (0.8 μg/L), there were no signs of significant teratogenicity, cardiotoxicity, or hemopoiesis toxicity, 3.2 µg/L AEO-7 exerted dramatic detrimental effects on organ development.ConclusionOn the basis of these findings, we recommend that the industrial usage and environmental impact of AEO-7 be re-evaluated and strictly monitored by environmental and public health organizations.

Organ-specific toxicity evaluation of stearamidopropyl dimethylamine (SAPDMA) surfactant using zebrafish embryos

Surfactants are widely used in the industry of detergents, household products, and cosmetics. SAPDMA is a cationic surfactant that is used mostly in cosmetics, conditioning agents and has recently gained attention as a corrosion inhibitor in the sea pipelines industry. In this regard, literature concerning the ecotoxicological classification of SAPDMA on aquatic animals is lacking. This study aims to evaluate the potential ecotoxicity of SAPDMA using the aquatic zebrafish embryo model. The potential toxic effects of SAPDMA were assessed by different assays. This includes (i) mortality/survival assay to assess the median lethal concentration (LC50); (ii) teratogenicity assay to assess the no observed effect concentration (NOEC); (iii) organ-specific toxicity assays including cardiotoxicity, neurotoxicity (using locomotion assay), hematopoietic toxicity (hemoglobin synthesis using o-dianisidine staining), hepatotoxicity (liver steatosis and yolk retention using Oil Red O (ORO) stain); (iv) cellular cytotoxicity (mitochondrial membrane potential) by measuring the accumulation of JC-1 dye into mitochondria. Exposure of embryos to SAPDMA caused mortality in a dose-dependent manner with a calculated LC50 of 2.3 mg/L. Thus, based on the LC50 value and according to the Fish and Wildlife Service (FWS) Acute Toxicity Rating Scale, SAPDMA is classified as “moderately toxic”. The No Observed Effect Concentration (NOEC) concerning a set of parameters including scoliosis, changes in body length, yolk, and eye sizes was 0.1 mg/L. At the same NOEC concentration (0.1 mg/L), no organ-specific toxicity was detected in fish treated with SAPDMA, except hepatomegaly with no associated liver dysfunctions. However, higher SAPDMA concentrations (0.8 mg/L) have dramatic effects on zebrafish organ development (eye, heart, and liver development). Our data recommend a re-evaluation of the SAPDMA employment in the industry setting and its strictly monitoring by environmental and public health agencies.

DVL mutations identified from human neural tube defects and Dandy-Walker malformation obstruct the Wnt signaling pathway

Wnt signaling pathways, including the canonical Wnt/β-catenin pathway, planar cell polarity pathway, and Wnt/Ca2+ signaling pathway, play important roles in neural development during embryonic stages. The DVL genes encode the hub proteins for Wnt signaling pathways. The mutations in DVL2 and DVL3 were identified from patients with neural tube defects (NTDs), but their functions in the pathogenesis of human neural diseases remain elusive. Here, we sequenced the coding regions of three DVL genes in 176 stillborn or miscarried fetuses with NTDs or Dandy-Walker malformation (DWM) and 480 adult controls from a Han Chinese population. Four rare mutations were identified: DVL1 p.R558H, DVL1 p.R606C, DVL2 p.R633W, and DVL3 p.R222Q. To assess the effect of these mutations on NTDs and DWM, various functional analyses such as luciferase reporter assay, stress fiber formation, and in vivo teratogenic assay were performed. The results showed that the DVL2 p.R633W mutation destabilized DVL2 protein and upregulated activities for all three Wnt signalings (Wnt/β-catenin signaling, Wnt/planar cell polarity signaling, and Wnt/Ca2+ signaling) in mammalian cells. In contrast, DVL1 mutants (DVL1 p.R558H and DVL1 p.R606C) decreased canonical Wnt/β-catenin signaling but increased the activity of Wnt/Ca2+ signaling, and DVL3 p.R222Q only decreased the activity of Wnt/Ca2+ signaling. We also found that only the DVL2 p.R633W mutant displayed more severe teratogenicity in zebrafish embryos than wild-type DVL2. Our study demonstrates that these four rare DVL mutations, especially DVL2 p.R633W, may contribute to human neural diseases such as NTDs and DWM by obstructing Wnt signaling pathways.

From mRNA Expression of Drug Disposition Genes to In Vivo Assessment of CYP-Mediated Biotransformation during Zebrafish Embryonic and Larval Development.

The zebrafish (Danio rerio) embryo is currently explored as an alternative for developmental toxicity testing. As maternal metabolism is lacking in this model, knowledge of the disposition of xenobiotics during zebrafish organogenesis is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this study was to assess cytochrome P450 (CYP) activity in zebrafish embryos and larvae until 14 d post-fertilization (dpf) by using a non-specific CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR) and a CYP1-specific substrate, i.e., 7-ethoxyresorufin (ER). Moreover, the constitutive mRNA expression of CYP1A, CYP1B1, CYP1C1, CYP1C2, CYP2K6, CYP3A65, CYP3C1, phase II enzymes uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1) and sulfotransferase 1st1 (SULT1ST1), and an ATP-binding cassette (ABC) drug transporter, i.e., abcb4, was assessed during zebrafish development until 32 dpf by means of quantitative PCR (qPCR). The present study showed that trancripts and/or the activity of these proteins involved in disposition of xenobiotics are generally low to undetectable before 72 h post-fertilization (hpf), which has to be taken into account in teratogenicity testing. Full capacity appears to be reached by the end of organogenesis (i.e., 120 hpf), although CYP1—except CYP1A—and SULT1ST1 were shown to be already mature in early embryonic development.

Analysis of Lethality and Malformations During Zebrafish (Danio rerio) Development.

The versatility offered by zebrafish (Danio rerio) makes it a powerful and an attractive vertebrate model in developmental toxicity and teratogenicity assays. Apart from the newly introduced chemicals as drugs, xenobiotics also induce abnormal developmental abnormalities and congenital malformations in living organisms. Over the recent decades, zebrafish embryo/larva has emerged as a potential tool to test teratogenicity potential of these chemicals. Zebrafish responds to compounds as mammals do as they share similarities in their development, metabolism, physiology, and signaling pathways with that of mammals. The methodology used by the different scientists varies enormously in the zebrafish embryotoxicity test. In this chapter, we present methods to assess lethality and malformations during zebrafish development. We propose two major malformations scoring systems: binomial and relative morphological scoring systems to assess the malformations in zebrafish embryos/larvae. Based on the scoring of the malformations, the test compound can be classified as a teratogen or a nonteratogen and its teratogenic potential is evaluated.

The Ascidian Embryo Teratogenicity assay in Ciona intestinalis as a new teratological screening to test the mixture effect of the co-exposure to ethanol and fluconazole

The aim of this work was to evaluate the Ascidian Embryo Teratogenicity assay (AET) as new alternative invertebrate model to test the developmental effects of the co-exposure to ethanol and fluconazole. Ciona intestinalis embryos were exposed to the azolic fungicide fluconazole, (FLUCO, 7.8–250μM), to ethanol (Eth, 0.01–0.5%) and to their mixture (0.01% Eth+FLUCO 7.8–250μM) from neurula to larval stage. At the end of the exposure period, larvae were morphologically evaluated and benchmark analysis performed by using the PROAST modelling software. Both compounds were teratogenic in a concentration-related manner, particularly affecting the pigmented organs. The co-exposure to Eth enhanced the effects of FLUCO, the additive hypothesis was not rejected by the modelling. The results demonstrated that AET could be considered a good vertebrate-free alternative model for toxicological investigation in embryos.

In Vitro Biotransformation of Two Human CYP3A Probe Substrates and Their Inhibition during Early Zebrafish Development.

At present, the zebrafish embryo is increasingly used as an alternative animal model to screen for developmental toxicity after exposure to xenobiotics. Since zebrafish embryos depend on their own drug-metabolizing capacity, knowledge of their intrinsic biotransformation is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this in vitro study was to assess the activity of cytochrome P450 (CYP)—a group of drug-metabolizing enzymes—in microsomes from whole zebrafish embryos (ZEM) of 5, 24, 48, 72, 96 and 120 h post-fertilization (hpf) by means of a mammalian CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR). The same CYP activity assays were performed in adult zebrafish liver microsomes (ZLM) to serve as a reference for the embryos. In addition, activity assays with the human CYP3A4-specific Luciferin isopropyl acetal (Luciferin-IPA) as well as inhibition studies with ketoconazole and CYP3cide were carried out to identify CYP activity in ZLM. In the present study, biotransformation of BOMR was detected at 72 and 96 hpf; however, metabolite formation was low compared with ZLM. Furthermore, Luciferin-IPA was not metabolized by the zebrafish. In conclusion, the capacity of intrinsic biotransformation in zebrafish embryos appears to be lacking during a major part of organogenesis.

Recent Advances in Disease Modeling and Drug Discovery for Diabetes Mellitus Using Induced Pluripotent Stem Cells.

Diabetes mellitus (DM) is a widespread metabolic disease with a progressive incidence of morbidity and mortality worldwide. Despite extensive research, treatment options for diabetic patients remains limited. Although significant challenges remain, induced pluripotent stem cells (iPSCs) have the capacity to differentiate into any cell type, including insulin-secreting pancreatic β cells, highlighting its potential as a treatment option for DM. Several iPSC lines have recently been derived from both diabetic and healthy donors. Using different reprogramming techniques, iPSCs were differentiated into insulin-secreting pancreatic βcells. Furthermore, diabetes patient-derived iPSCs (DiPSCs) are increasingly being used as a platform to perform cell-based drug screening in order to develop DiPSC-based cell therapies against DM. Toxicity and teratogenicity assays based on iPSC-derived cells can also provide additional information on safety before advancing drugs to clinical trials. In this review, we summarize recent advances in the development of techniques for differentiation of iPSCs or DiPSCs into insulin-secreting pancreatic β cells, their applications in drug screening, and their role in complementing and replacing animal testing in clinical use. Advances in iPSC technologies will provide new knowledge needed to develop patient-specific iPSC-based diabetic therapies.

Abstract 1793: Novel radiation mitigators and anticancer drugs

Abstract The possibility of a radiation disaster from a nuclear detonation or accident has existed for over 50 years and spawned much of the basic research in radiobiology in the 1950-60s. The recent Fukushima accident was yet another reminder that there remains a dire need to develop novel therapies against radiation-induced toxicities. Here we report on the development of two novel radiation countermeasure therapies: Yel001 and Yel002. These small, biologically active, drug-like molecules were uncovered in the DEL high throughput assay reducing radiation-induced cyto- and geno-toxicity in yeast. Radiation-modulating activity was further confirmed in yeast plate-based DEL Assay: addition of either Yel001 or Yel002 to irradiated cultures reduced cell death and genomic instability. Further, Yel compounds increases survival to 75% in vivo following an LD100/30 dose of ionizing radiation (IR) with the first therapeutic injection administered 24 hours post exposure followed by injections at 48,72,96, and 120 hours. Additionally, treatment with Yel001 and Yel002 compounds reduces radiation-induced leukemia from 90% to to 50% and 40% respectively. Of note, treatment with either Yel001 or Yel002 reduced spontaneous leukemia rate from 10% to 0%. Furthermore, Yel002 significantly prolonged the life of Atm deficient mice. Treatment with Yel002 following IR accelerates the recovery of the hematopoietic cells and protects stem cells after sub-lethal exposures. In addition, treatment with Yel002 reduces I131, EMS, MMS, UV, cigarette smoke extract as well as nitrogen mustard induced toxicity as well as genotoxicity showing a broad application spectrum. Proteomics shows that Yel002 induces DNA repair mediated by Atm signaling, homologous recombination, nonhomologous end joining, base excision repair, DNA damage binding proteins and chromosome segregation functions. Toxicity has not been observed in neither in vitro or in vivo administrations, even in a two generation teratogenicity assay. Overall, Yel compounds have much potential as stockpile therapies for radiation-induced lethality and cancer as well as spontaneous cancer: they are highly effective when administered up to 24hours post exposure, they reduce radiation-induced sequelae such as leukemia, and appear to have an acceptable toxicity profile. Citation Format: Robert H. Schiestl, Michael Davoren, Yelena Rivina. Novel radiation mitigators and anticancer drugs. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1793. doi:10.1158/1538-7445.AM2015-1793

Optimization and Performance Assessment of the Chorion-Off [Dechorinated] Zebrafish Developmental Toxicity Assay.

The Dechorinated Zebrafish Embryo Developmental toxicity assay was originally developed from a training set of 31 compounds and reported to be 87% concordant with in vivo teratogenicity data (Brannen, K. C., Panzica-Kelly, J. M., Danberry, T. L., and Augustine-Rauch, K. A. (2010). Development of a zebrafish embryo teratogenicity assay and quantitative prediction model. Birth Defects Res. 89, 66-77.). The assay includes scoring larva treated in a concentration range for malformations of specific morphological structures/organ systems. The model includes identifying a no-adverse-effect-level (NOAEL) and the concentration resulting in 25% lethality (LC25) at 5 days postfertilization. An LC25/NOAEL ratio ≥10 classifies a compound positive for teratogenic potential. A consortium effort evaluated a modified version of this assay which involved enzymatic chorion treatment instead of manual dissection and used experimental replicates for final classification. The modified assay achieved an 85% overall predictivity (Gustafson, A. L., Stedman, D. B., Ball, J., Hillegass, J. M., Flood, A., Zhang, C. X., Panzica-Kelly, J., Cao, J., Coburn, A., Enright, B. P., et al. (2012). Inter-laboratory assessment of a harmonized zebrafish developmental toxicology assay - progress report on phase I. Reprod. Toxicol. 33, 155-164.). The objective of this study was to perform a thorough performance evaluation of the dechorinated assay by repeating the original training set and testing additional compounds in experimental replicates. When the initial training set was repeated with inclusion of experimental replicates, the overall predictivity was 83%. Model performance was tested with an additional 34 compounds and achieved overall predictivity of 74%. When the training and test sets were combined (63 compounds) the assay's final sensitivity was 83% and the specificity was 71%. Total predictivity was 78% with relatively balanced predictivity for nonteratogens (77%) and teratogens (78%). The chorion-off assay achieved superior sensitivity (83%) compared with sensitivity (63-74%) reported by consortium efforts testing a similar assay with chorion-intact embryos (Ball, J. S., Stedman, D. B., Hillegass, J. M., Zhang, C. X., Panzica-Kelly, J., Coburn, A., Enright, B. P., Tornesi, B., Amouzadeh, H. R., Hetheridge, M., et al. (2014). Fishing for teratogens: a consortium effort for a harmonized zebrafish developmental toxicology assay. Toxicol. Sci. 139, 210-219.). Additional protocol modifications were made to increase assay throughput.

Abstract 4909: Novel radiation mitigators and cancer chemoprevention drugs

Abstract The possibility of a radiation disaster from a nuclear detonation or accident has existed for over 50 years and spawned much of the basic research in radiobiology in the 1950-60s. The recent Fukushima accident was yet another reminder that there remains a dire need to develop novel therapies against radiation-induced toxicities. Here we report on the development of two novel radiation countermeasure therapies: Yel001 and Yel002. These small, biologically active, drug-like molecules were uncovered in the DEL high throughput assay reducing radiation-induced cyto- and geno-toxicity in yeast. Radiation-modulating activity was further confirmed in yeast plate-based DEL Assay: addition of either Yel001 or Yel002 to irradiated cultures reduced cell death and genomic instability. Further, Yel compounds increases survival to 75% in vivo following an LD100/30 dose of ionizing radiation (IR) with the first therapeutic injection administered 24 hours post exposure followed by injections at 48,72,96, and 120 hours. Additionally, treatment with Yel001 and Yel002 compounds reduces radiation-induced leukemia from 90% to to 50% and 40% respectively. Of note, treatment with either Yel001 or Yel002 reduced spontaneous leukemia rate from 10% to 0%. Furthermore, Yel002 significantly prolonged the life of Atm deficient mice. Treatment with Yel002 following IR accelerates the recovery of the hematopoietic cells and protects stem cells after sub-lethal exposures. In addition, treatment with Yel002 reduces I131, EMS, MMS, UV, cigarette smoke extract as well as nitrogen mustard induced toxicity as well as genotoxicity showing a broad application spectrum. Proteomics shows that Yel002 induces DNA repair mediated by Atm signaling, homologous recombination, nonhomologous end joining, base excision repair, DNA damage binding proteins and chromosome segregation functions. Toxicity has not been observed in neither in vitro or in vivo administrations, even in a two generation teratogenicity assay. Overall, Yel compounds have much potential as stockpile therapies for radiation-induced lethality and cancer as well as spontaneous cancer: they are highly effective when administered up to 24hours post exposure, they reduce radiation-induced sequelae such as leukemia, and appear to have an acceptable toxicity profile. Citation Format: Robert H. Schiestl, Yelena Rivina, Michael Davoren. Novel radiation mitigators and cancer chemoprevention drugs. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4909. doi:10.1158/1538-7445.AM2014-4909

Concise Review: Drug Discovery in the Age of the Induced Pluripotent Stem Cell

For decades, the paradigm of drug discovery and development has relied on immortalized cell lines, animal models of human disease, and clinical trials. With the discovery of induced pluripotent stem cell (iPSC) technology in 2007, a new human in vitro drug testing platform has potentially augmented this set of tools by providing additional ways to screen compounds for safety and efficacy. The growing number of human disease models made with patient-specific iPSCs has made it possible to conduct research on a wide range of disorders, including rare diseases and those with multifactorial origin, as well as to simulate drug effects on difficult-to-obtain tissues such as brain and cardiac muscle. Toxicity and teratogenicity assays developed with iPSC-derived cells can also provide an additional layer of safety before advancing drugs to clinical trials. The incorporation of iPSC technology into drug therapy development holds promise as a more powerful and nuanced approach to personalized medicine.

Evaluation on genotoxicity and teratogenicity of aqueous extract from Cyclocarya paliurus leaves.

Tremendous attentions have been attracted to the foods labeled with natural, green, organic, and nuisanceless conception of healthy diet. Therefore, it is of great significance to establish relative defining guidance for safe assessment of botanicals. Cyclocarya paliurus (Batal.) Iljinsk (family Cyclocaryaceae), called sweet tea tree, is a well-known edible and medicinal plant, which has been widely used in China as drug formulation for the treatment of hypertension and diabetes. Despite its benefits, no reports have been described on the safe assessment of C. paliurus leaves aqueous extract. In this study, we have conducted the genotoxicity assay (including Ames test, bone marrow polychromatic erythrocyte micronucleus test, and sperm abnormality test in mice) and traditional teratogenicity assay in rats (maternal toxicity, embryo toxicity, and teratogenicity test) to assess the genetic and teratogenic safety of aqueous extracts from C. paliurus leaves. Results of each assay show that the highest dose of C. paliurus leaves aqueous extract is considered relatively nonmutagenic and nonteratogenic, revealing that C. paliurus leaves possess safety and quality as a functional additional ingredient in food.

Organic Nanovesicular Cargoes for Sustained Drug Delivery: Synthesis, Vesicle Formation, Controlling “Pearling” States, and Terfenadine Loading/Release Studies

“Sustained drug delivery systems” which are designed to accomplish long-lasting therapeutic effect are one of the challenging topics in the area of nanomedicine. We developed an innovative strategy to prepare nontoxic and polymer stabilized organic nanovesicles (diameter: 200 nm) from a novel bolaamphiphile, where two hydrogen bonding acetyl cytosine molecules connected to 4,4′′-positions of the 2,6-bispyrazolylpyridine through two flexible octyne chains. The nanovesicles behave like biological membrane by spontaneously self-assembling into “pearl-like” chains and subsequently forming long nanotubes (diameter: 150 nm), which further develop into various types of network-junctions through self-organization. For drug loading and delivery applications, the nanovesicles were externally protected with biocompatible poly(ethyleneglycol)-2000 to prevent them from fusion and ensuing tube formation. Nontoxic nature of the nanovesicles was demonstrated by zebrafish teratogenicity assay. Biocompatible nanovesicles were loaded with “terfenadine” drug and successfully utilized to transport and release drug in sustained manner (up to 72 h) in zebrafish larvae, which is recognized as an emerging in vivo model system.