Development, Use, and Validation of (Q)SARs for Predicting Genotoxicity and Carcinogenicity: Experiences from Italian National Institute of Health Activities.

In silico approaches are now increasingly accepted in several areas of toxicology to rapidly assess chemical hazard without the need for animal testing. Among in silico tools, quantitative and qualitative structure-activity approaches ((Q)SARs) are the most typically applied methods to predict hazard in the absence of experimental data. This paper provides an overview of different protocols that can be applied while dealing with (Q)SARs in different scenarios, namely, (Q)SAR development, use, and validation. Examples of protocols adopted in the three scenarios are reported, derived from the authors' experience in working at the Predictive Toxicology unit of the Italian National Institute of Health, focusing on the endpoints of carcinogenicity and genotoxicity.The illustrated activities are in line with the Institute's mission, the main center of research, control, and technical-scientific advice on public health in Italy.

Evaluation of arsenic induced genotoxicity and its impact on life processes of Daphnia magna.

Heavy metals like arsenic is ubiquitously present in the environment. Geologic and anthropogenic activities are the root cause behind high concentration of arsenic in natural water bodies demanding strict monitoring of water quality prior to human consumption and utilization. In the present study, we have employed Daphnia magna for studying the biological effects of environmentally relevant high concentration of arsenic in water. In acute toxicity study, the LC50 value for 24hr exposure was found to be 2.504 mg/L, which gradually decreased with increase in time period (24hr- 96hr) to 2.011 mg/ L at 96hr. Sub-chronic toxicity was evaluated over 12 days using sub-lethal concentrations (5 %, 10 %, 15 %, and 20 % of the 24-hr LC50). Survivability in Daphnia showed a decreasing trend from 96 % to 91 % with increase in arsenic concentrations from 5 % of LC50 24 hr value to 20 % of LC 50 24hr value respectively. Alongside decreased survivability, there was a significant reduction in body size, with organisms exposed to the highest concentration of arsenic measuring 0.87±0.01 mm compared to 1.51±0.10 mm in the control group. Reproductive potential declined concentration dependently with exposure, with the highest reduction observed at 20 % of LC50 24hr value, where offspring numbers decreased to 7±1 from 23±5 in the control. Heart rate decreased in concentration and time-dependent manners, with the lowest rates observed at the highest arsenic concentration (279±16 bpm after 24hr and 277±27 bpm after 48hr). Comet assay and micronucleus assay conducted after 48 hrs of exposure revealed concentration-dependent genotoxic effects in Daphnia magna. Our results indicate negative impact on physiology and reproduction of Daphnia magna at environmentally existent concentration of arsenic. Also Daphnia magna could serve as a sensitive test system for investigating arsenic contamination in water bodies.

Development of a genotoxicity/carcinogenicity assessment method by DNA adductome analysis.

Safety evaluation is essential for the development of chemical substances. Since in vivo safety evaluation tests, such as carcinogenesis tests, require long-term observation using large numbers of experimental animals, it is necessary to develop alternative methods that can predict genotoxicity/carcinogenicity in the short term, taking into account the 3Rs (replacement, reduction, and refinement). We established a prediction model of the hepatotoxicity of chemicals using a DNA adductome, which is a comprehensive analysis of DNA adducts that may be used as an indicator of DNA damage in the liver. An adductome was generated with LC-high-resolution accurate mass spectrometer (HRAM) on liver of rats exposed to various chemicals for 24 h, based on two independent experimental protocols. The resulting adductome dataset obtained from each independent experiment (experiments 1 and 2) and integrated dataset were analyzed by linear discriminant analysis (LDA) and found to correctly classify the chemicals into the following four categories: non-genotoxic/non-hepatocarcinogens (-/-), genotoxic/non-hepatocarcinogens (+/-), non-genotoxic/hepatocarcinogens (-/+), and genotoxic/hepatocarcinogens (+/+), based on their genotoxicity/carcinogenicity properties. A prototype model for predicting the genotoxicity/carcinogenicity of the chemicals was established using machine learning methods (using random forest algorithm). When the prototype genotoxicity/carcinogenicity prediction model was used to make predictions for experiments 1 and 2 as well as the integrated dataset, the correct response rates were 89 % (genotoxicity), 94 % (carcinogenicity) and 87 % (genotoxicity/carcinogenicity) for experiment 1, 47 % (genotoxicity), 62 % (carcinogenicity) and 42 % (genotoxicity/carcinogenicity) for experiment 2, and 52 % (genotoxicity), 62 % (carcinogenicity), and 48 % (genotoxicity/carcinogenicity) for the integrated dataset. To improve the accuracy of the toxicity prediction model, the toxicity label was reconstructed as follows; Pattern 1: when +/+ and -/- chemicals were used from the toxicity labels +/+, +/-, -/+ and -/-; and Pattern 2: when +/+, +/-, and -/+ other than -/- were replaced with the label "Others". As a result, chemicals with only +/+ and -/- toxicity labels were used and the correct response rates were approximately 100 % for the measured data in experiment 1, 53 %-66 % for the data in experiment 2, and 59-73 % for the integrated data, all of which were 10 %-30 % higher compared with the data before the label change. In contrast, when the toxicity labels were replaced with -/- and "Others", they reached nearly 100 % in the measured data from experiment 1, 65 %-75 % in the data from experiment 2, and 70 %-78 % in the integrated data, all of which were 10 %-50 % higher compared with the data before the label change.

Evaluation of the in vivo acute toxicity and in vitro genotoxicity and mutagenicity of synthetic ß-carboline alkaloids with selective cytotoxic activity against ovarian and breast cancer cell lines.

The aim of this study was to evaluate the in vitro cytotoxic, genotoxic, and mutagenic potential and to determine the in silico ADME parameters of two synthetic ß-carboline alkaloids developed as prototypes of antitumor agents (NQBio-06 and NQBio-21). Additionally, acute toxicity of the compounds was evaluated in mice. The results from the MTT assay showed that NQBio-06 presented higher cytotoxicity in the ovarian cancer cell line TOV-21 G (IC50 = 2.5 µM, selectivity index = 23.7). NQBio-21 presented an IC50 of 6.9 µM and a selectivity index of 14.5 against MDA-MB-231 breast cancer cells. Comet assay results showed that NQBio-06 did not induce chromosomal breaks in vitro, but NQBio-21 was genotoxic with and without metabolic activation (S9 fraction). Micronucleus assay showed that both compounds were mutagenic. In addition, metabolic activation enhanced this effect in vitro. The in silico predictions showed that the compounds met the criteria set by Lipinski's rules, had strong prediction for intestinal absorption, and were possible substrates for P-glycoprotein. The in vivo results demonstrated that both the compounds exhibited low acute toxicity. These results suggest that the mechanisms underlying the cytotoxicity of NQBio-06 and NQBio-21 are related to DNA damage induction and that the use of S9 enhanced these effects. In vivo analysis showed signs of toxicity after a single administration of the compounds in mice. These findings highlight the potential of ß-carboline compounds as sources for the development of new anticancer chemotherapeutic agents.

Differential genotoxicity of Polygoni Multiflori in rat and human: insights from Ames test and S9 metabolic activation system.

The Ames test is used worldwide to initially screen the mutagenic potential of new chemicals. In the standard Ames test, S. typhimurium strains (TA100, TA98, TA1535, and TA1537) and Escherichia coli (WP2uvrA) are treated with substances with/without cytochrome P450s (CYPs)-induced rat S9 fractions for identifying mutagens and pro-mutagens. However, many substances show completely different toxicity patterns depending on whether the liver S9 fraction belongs to rats or humans. The natural product Polygoni Multiflori Radix (PMR) can also show bacterial reverse mutation, followed by the rat or human liver S9 fraction. While PMR elicits reverse mutations in the TA1537 strain in rat liver S9 but not in human liver S9, this mechanism has not been verified yet. To explain this, the differences in metabolic enzymes compositions commonly observed between rats and humans have been implicated. This study aimed to explore the key factors that cause differences in the genotoxicity of PMR between rat and human liver S9 metabolic enzymes. The results of next-generation sequencing (NGS) analysis showed that both rat and human metabolic enzymes caused similar mutations in TA1537. However, when the metabolic enzymes in each S9 fraction were analyzed using ion mobility tandem mass spectrometry (IM-MS), rat- and human-specific enzymes were identified among the cytochrome (CYP) family, especially aryl hydrocarbon receptor (AHR)-related CYPs. These findings suggest that CYP1A1 isoforms contribute to the mechanism of PMR in the Ames test. Therefore, an in vitro Ames test might be more reliable in predicting genotoxicity for both rodents and humans. This will also help overcome the limitations of laboratory animal-based toxicity evaluations, which provide unreliable results due to interspecies differences between humans and rodents.

Benefits and concerns of probiotics: an overview of the potential genotoxicity of the colibactin-producing Escherichia coli Nissle 1917 strain.

Recently, the mounting integration of probiotics into human health strategies has gathered considerable attention. Although the benefits of probiotics have been widely recognized in patients with gastrointestinal disorders, immune system modulation, and chronic-degenerative diseases, there is a growing need to evaluate their potential risks. In this context, new concerns have arisen regarding the safety of probiotics as some strains may have adverse effects in humans. Among these strains, Escherichia coli Nissle 1917 (EcN) exhibited traits of concern due to a pathogenic locus in its genome that produces potentially genotoxic metabolites. As the use of probiotics for therapeutic purposes is increasing, the effects of potentially harmful probiotics must be carefully evaluated. To this end, in this narrative review article, we reported the findings of the most relevant in vitro and in vivo studies investigating the expanding applications of probiotics and their impact on human well-being addressing concerns arising from the presence of antibiotic resistance and pathogenic elements, with a focus on the polyketide synthase (pks) pathogenic island of EcN. In this context, the literature data here discussed encourages a thorough profiling of probiotics to identify potential harmful elements as done for EcN where potential genotoxic effects of colibactin, a secondary metabolite, were observed. Specifically, while some studies suggest EcN is safe for gastrointestinal health, conflicting findings highlight the need for further research to clarify its safety and optimize its use in therapy. Overall, the data here presented suggest that a comprehensive assessment of the evolving landscape of probiotics is essential to make evidence-based decisions and ensure their correct use in humans.

Relative genotoxicity of polycyclic aromatic hydrocarbons inferred from free energy perturbation approaches.

Utilizing molecular dynamics and free energy perturbation, we examine the relative binding affinity of several covalent polycyclic aromatic hydrocarbon - DNA (PAH-DNA) adducts at the central adenine of NRAS codon-61, a mutational hotspot implicated in cancer risk. Several PAHs classified by the International Agency for Research on Cancer as probable, possible, or unclassifiable as to carcinogenicity are found to have greater binding affinity than the known carcinogen, benzo[a]pyrene (B[a]P). van der Waals interactions between the intercalated PAH and neighboring nucleobases, and minimal disruption of the DNA duplex drive increases in binding affinity. PAH-DNA adducts may be repaired by global genomic nucleotide excision repair (GG-NER), hence we also compute relative free energies of complexation of PAH-DNA adducts with RAD4-RAD23 (the yeast ortholog of human XPC-RAD23) which constitutes the recognition step in GG-NER. PAH-DNA adducts exhibiting the greatest DNA binding affinity also exhibit the least RAD4-RAD23 complexation affinity and are thus predicted to resist the GG-NER machinery, contributing to their genotoxic potential. In particular, the fjord region PAHs dibenzo[a,l]pyrene, benzo[g]chrysene, and benzo[c]phenanthrene are found to have greater binding affinity while having weaker RAD4-RAD23 complexation affinity than their respective bay region analogs B[a]P, chrysene, and phenanthrene. We also find that the bay region PAHs dibenzo[a,j]anthracene, dibenzo[a,c]anthracene, and dibenzo[a,h]anthracene exhibit greater binding affinity and weaker RAD4-RAD23 complexation affinity than B[a]P. Thus, the study of PAH genotoxicity likely needs to be substantially broadened, with implications for public policy and the health sciences. This approach can be broadly applied to assess factors contributing to the genotoxicity of other unclassified compounds.

Assessment of receptor-mediated activity (AhR and ERα), mutagenicity, and teratogenicity of metal shredder wastes in Wallonia, Belgium.

In this study, hazardous wastes including fluff, dust, and scrubbing sludge were sampled in 2019 from two metal shredding facilities located in Wallonia, Belgium. To assess the extent of the contamination, a global approach combining chemical and biological techniques was used, to better reflect the risks to health and the environment. The samples investigated induced significant in vitro aryl hydrocarbon receptor (AhR) agonistic bioactivities and estrogenic receptor (ERα) (ant)agonistic bioactivities in the respective CALUX (chemical activated luciferase gene expression) bioassays. The mutagenicity of the samples was investigated with the bacterial reverse gene mutation test using the Salmonella typhimurium TA98 and TA100 strains. Except for the sludge sample (site 3), all samples induced a mutagenic response in the TA98 strain (± S9 metabolic fraction) whereas in the TA100 strain (+ S9 metabolic fraction), only the sludge sample (site 2) showed a clear mutagenic effect. The in vivo toxicity/teratogenicity of the shredder wastes was further evaluated with zebrafish embryos. Except for the dust sample (site 2), all samples were found to be teratogenic as they returned teratogenic indexes (TIs) > 1. The high levels of contamination, the mutagenicity, and the teratogenicity of these shredder wastes raise significant concerns about their potential negative impacts on both human health and environment.

Microbial genotoxin-elicited host DNA mutations related to mitochondrial dysfunction, a momentous contributor for colorectal carcinogenesis.

Gut microbe dysbiosis increases repetitive inflammatory responses, leading to an increase in the incidence of colorectal cancer. Recent studies have revealed that specific microbial species directly instigate mutations in the host nucleus DNA, thereby accelerating the progression of colorectal cancer. Given the well-established role of mitochondrial dysfunction in promoting colorectal cancer, it is reasonable to postulate that gut microbes may induce mitochondrial gene mutations, thereby inducing mitochondrial dysfunction. In this review, we focus on gut microbial genotoxins and their known and potential targets in mitochondrial genes. Consequently, we propose that targeted disruption of genotoxin transport pathways may effectively reduce the rate of mitochondrial gene mutations and yield substantial benefits for the prevention of colorectal carcinogenesis.

Evaluation of levamlodipine benzenesulfonate compound I for embryo-fetal developmental toxicity in SD rats and genotoxicity.

In the present study, the effects of levamlodipine benzenesulfonate on the development of fertile Sprague-Dawley (SD) rats, their embryos, and littermates were assessed using an embryo-fetal developmental toxicity test. Maternal body weight reduction was observed at a dose of 20 mg/kg, but it recovered after treatment cessation. The 20 mg/kg dose group showed a skewed sex ratio in fetal rats, with a higher proportion of males. While some effects on fetal sternum development were observed at 20 mg/kg, no skeletal malformations were observed. No significant gross morphological abnormalities were detected in the dams (mothers), no significant embryotoxicity or foetotoxicity in fetal rats and no significant effects on fetal length and weight development at doses of 5 and 10 mg/kg. Genotoxicity was evaluated using a combination of the Ames test, the Chinese hamster ovary (CHO) cell chromosome aberration assay, and the ICR mouse bone marrow micronucleus test. The Ames test results indicated substantial bacteriostatic effects at doses of 500 and 5000 mg/dish, with no mutagenicity observed at doses of 0.5, 5, and 50 mg/dish. No significant effect on the aberration rate of CHO cell chromosomes was found at doses of 2.8, 5.6, and 11.2 mg/mL. In the ICR mouse micronucleus test, no micronucleus-inducing effect was observed at doses of 3.125, 6.25, and 12.5 mg/kg in each treatment group. In conclusion, under the conditions of this experiment, the no-observed-adverse-effect level (NOAEL) for developmental toxicity of levamlodipine benzenesulfonate in fertile SD rats, their embryos, and littermates was established to be 10 mg/kg/day. Levamlodipine benzenesulfonate did not exhibit significant genotoxicity.

Photomutagenicity of N-nitrosoproline dissolved in non-aqueous solvent, oleic acid.

In the present study, we investigated the genotoxicity of the active products formed from N-nitrosoproline (NPRO) dissolved in oleic acid following ultraviolet A (UVA) irradiation, bypassing the need for metabolic activation. We previously demonstrated the photomutagenicity of NPRO dissolved in a phosphate-buffered solution. It has been suggested that the association of the nitrosamine group with acid ions facilitates rapid photodissociation and photoactivation. We hypothesized that NPRO's inherent carboxyl group may mimic an acid, inducing photodissociation and photomutagenicity, even in a non-aqueous solvent lacking acidic ions. Following UVA irradiation, NPRO dissolved in oleic acid exhibited a dose-dependent mutagenic activity. Similar results were obtained when NPRO was dissolved in linoleic acid and triolein. Nitric oxide formation, which is dependent on NPRO concentration, is accompanied by mutagenic activity. The mutagenicity spectrum obtained in response to NPRO irradiation followed the absorption curve of NPRO dissolved in oleic acid. Irradiated NPRO in oleic acid displayed relative stability, retaining approximately 18, 36, and 63 % of initial mutagenicity after 10 days of storage at 25, 4, and -20 °C, respectively. Thus NPRO stored in a fatty environment undergoes photoactivation upon irradiation, leading to genotoxicity.

Levothyroxine synthesis impurities are neither mutagenic nor genotoxic: Insilico, Ames test and micronucleus test studies.

In vitro and in silico tests were used to assess the possible genotoxicity and mutagenicity of five impurities that may be present in levothyroxine, a drug used for thyroid hormone replacement therapy. Neither ToxTree nor VEGA (Virtual Models for evaluating the properties of chemicals within a global architecture) identified cause for concern for any of the impurities. Ames test results (doses up to 1 mg per plate), with or without metabolic activation, were negative. The micronucleus test with TK6 (human lymphoblastoid) cells, at doses up to 500 µg/mL, with or without metabolic activation, also gave negative results.

Ecotoxicological studies of direct and indirect genotoxicity with Artemia: a integrative review.

Artemia is a brine shrimp genus adapted to extreme habitats like ranges salinity from 5-25 g/L and in temperatures from 9 to 35 °C. It is widely distributed and used as an environmental quality biomarker. Artemia franciscana and Artemia salina species are commonly used in ecotoxicological studies and genotoxicity assays due to their short life cycle, high fecundity rate, easy culture, and availability. Thus, considering the importance of these tests in ecotoxicological studies, the present study aimed to present Artemia genus as a biological model in genotoxicity research. To this end, we reviewed the literature, analyzing data published until July 2023 in the Web of Science, SCOPUS, Embase, and PubMed databases. After screening, we selected 34 studies in which the genotoxicity of Artemia for various substances. This review presents the variability of the experimental planning of assays and biomarkers in genotoxicity using Artemia genus as a biological model for ecotoxicological studies and show the possibility of monitoring biochemical alterations and genetic damage effects. Also highlight innovative technologies such as transcriptomic and metabolomic analysis, as well as studies over successive generations to identify changes in DNA and consequently in gene expression.

Deep active learning with high structural discriminability for molecular mutagenicity prediction.

The assessment of mutagenicity is essential in drug discovery, as it may lead to cancer and germ cells damage. Although in silico methods have been proposed for mutagenicity prediction, their performance is hindered by the scarcity of labeled molecules. However, experimental mutagenicity testing can be time-consuming and costly. One solution to reduce the annotation cost is active learning, where the algorithm actively selects the most valuable molecules from a vast chemical space and presents them to the oracle (e.g., a human expert) for annotation, thereby rapidly improving the model's predictive performance with a smaller annotation cost. In this paper, we propose muTOX-AL, a deep active learning framework, which can actively explore the chemical space and identify the most valuable molecules, resulting in competitive performance with a small number of labeled samples. The experimental results show that, compared to the random sampling strategy, muTOX-AL can reduce the number of training molecules by about 57%. Additionally, muTOX-AL exhibits outstanding molecular structural discriminability, allowing it to pick molecules with high structural similarity but opposite properties.

Olive Mill Wastewater Extract: In Vitro Genotoxicity/Antigenotoxicity Assessment on HepaRG Cells.

Olive mill wastewater (OMWW), with its high level of phenolic compounds, simultaneously represents a serious environmental challenge and a great resource with potential nutraceutical activities. To increase the knowledge of OMWW's biological effects, with an aim to developing a food supplement, we performed a chemical characterisation of the extract using the Liquid Chromatography-Quadrupole Time-of-flight spectrometry (LC-QTOF) and an in vitro genotoxicity/antigenotoxicity assessment on HepaRG ™ cells. Chemical analysis revealed that the most abundant phenolic compound was hydroxytyrosol. Biological tests showed that the extract was not cytotoxic at the lowest tested concentrations (from 0.25 to 2.5 mg/mL), unlike the highest concentrations (from 5 to 20 mg/mL). Regarding genotoxic activity, when tested at non-cytotoxic concentrations, the extract did not display any effect. Additionally, the lowest tested OMWW concentrations showed antigenotoxic activity (J-shaped dose-response effect) against a known mutagenic substance, reducing the extent of DNA damage in the co-exposure treatment. The antigenotoxic effect was also obtained in the post-exposure procedure, although only at the extract concentrations of 0.015625 and 0.03125 mg/mL. This behaviour was not confirmed in the pre-exposure protocol. In conclusion, the present study established a maximum non-toxic OMWW extract dose for the HepaRG cell model, smoothing the path for future research.

Genotoxicity of selected cannabinoids in human lymphoblastoid TK6 cells.

Natural non-psychoactive cannabinoids such as cannabigerol (CBG), cannabidiol (CBD), cannabichromene (CBC), cannabidivarin (CBDV), and cannabinol (CBN) are increasingly consumed as constituents of dietary products because of the health benefits claims. Cannabinoids may reduce certain types of pain, nausea, and anxiety. Anti-inflammatory and even anti-carcinogenic properties have been discussed. However, there are insufficient data available regarding their potential (geno-)toxic effects. Therefore, we tested CBG, CBD, CBC, CBDV, and CBN for their genotoxic potential and effects on mitosis and cell cycle in human lymphoblastoid TK6 cells. The selected cannabinoids (except CBDV) induced increased micronuclei formation, which was reduced with the addition of a metabolic activation system (S9 mix). CBDV induced micronuclei only after metabolic activation. Mitotic disturbances were observed with all tested cannabinoids, while G1 phase accumulation of cells was observed for CBG, CBD and CBDV. The genotoxic effects occurred at about 1000-fold higher concentrations than are reported as blood levels from human consumption. However, the results clearly indicate a need for further research into the genotoxic effects of cannabinoids. The mechanism of the mitotic disturbance, the shape of the dose-response curves and the possible effects of mixtures of cannabinoids are aspects which need clarification.

Analysis of transcriptional response in haploid and diploid Schizosaccharomyces pombe under genotoxic stress.

Whole genome duplications are implicated in genome instability and tumorigenesis. Human and yeast polyploids exhibit increased replication stress and chromosomal instability, both hallmarks of cancer. In this study, we investigate the transcriptional response of Schizosaccharomyces pombe to increased ploidy generally, and in response to treatment with the genotoxin methyl methanesulfonate (MMS). We find that treatment of MMS induces upregulation of genes involved in general response to genotoxins, in addition to cell cycle regulatory genes. Downregulated genes are enriched in transport and sexual reproductive pathways. We find that the diploid response to MMS is muted compared to the haploid response, although the enriched pathways remain largely the same. Overall, our data suggests that the global S. pombe transcriptome doubles in response to increased ploidy but undergoes modest transcriptional changes in both unperturbed and genotoxic stress conditions.

Designing and applying a methodology to assess sperm cell viability and DNA damage in a model amphipod.

Sperm quality is defined as the sperm cell ability to successfully fertilize eggs and allow normal embryo development⁠. Few studies explore sperm quality using aquatic invertebrates. Parhyale hawaiensis is a marine amphipod with a circumtropical distribution and considered a model for evolution, development, and ecotoxicological studies. We aimed to develop a methodology to collect sperm cells of P. hawaiensis and evaluate their viability and DNA damage (comet assay). We directly exposed the sperm cells to different mutagenic agents to optimize/develop the protocols. Then, as a proof of concept, we exposed the males to mutagenic compounds (EMS, benzo[a]pyrene (BaP), azo and anthraquinone dyes) at non-lethal concentrations verified by the proposed viability test and analyzed their sperm cells for DNA damage (comet assay). Organisms exposed to EMS presented a clear concentration response in the DNA damage response. We also showed that BaP was able to induce a statistically significant increase in DNA damage of the sperm cells. For the two dyes, although DNA damage increased, statistically differences were not observed. We believe we successfully developed a test to detect genotoxicity of chemicals in sperm cells using an invertebrate model. The protocol for sperm cell viability needs to be further explored with different chemicals to verify its utility as a toxicity endpoint. The developed genotoxicity test has the advantages to employ organisms that are easily cultivated in reduced space, use simple laboratory resources and reduced amount of material and reagents. Positive responses with this model could be used to disclose new germ cell mutagen candidates which could be further confirmed in vertebrates' systems.

A novel procedure for selection of molecular descriptors: QSAR model for mutagenicity of nitroaromatic compounds.

Nitroaromatic compounds (NACs) stand out as pervasive organic pollutants, prompting an imperative need to investigate their hazardous effects. Computational chemistry methods play a crucial role in this exploration, offering a safer and more time-efficient approach, mandated by various legislations. In this study, our focus lay on the development of transparent, interpretable, reproducible, and publicly available methodologies aimed at deriving quantitative structure-activity relationship models and testing them by modelling the mutagenicity of NACs against the Salmonella typhimurium TA100 strain. Descriptors were selected from Mordred and RDKit molecular descriptors, along with several quantum chemistry descriptors. For that purpose, the genetic algorithm (GA), as the most widely used method in the literature, and three alternative algorithms (Boruta, Featurewiz, and ForwardSelector) combined with the forward stepwise selection technique were used. The construction of models utilized the multiple linear regression method, with subsequent scrutiny of fitting and predictive performance, reliability, and robustness through various statistical validation criteria. The models were ranked by the Multi-Criteria Decision Making procedure. Findings have revealed that the proposed methodology for descriptor selection outperforms GA, with Featurewiz showing a slight advantage over Boruta and ForwardSelector. These constructed models can serve as valuable tools for the quick and reliable prediction of NACs mutagenicity.

The Allium cepa comet assay for environmental sample assessment: a scoping review.

The purposes of this review were to investigate the application of the comet assay in Allium cepa root cells to assess the genotoxicity of environmental samples and to analyse the experimental procedures employed. A literature search was performed selecting articles published between January 2000 and October 2023 from online databases using the combined search terms 'comet assay' and 'A. cepa'. Only 18 papers met the inclusion criteria. None of these were published in the first eight years (2000-2007), highlighting the increasing interest in using the comet assay on A. cepa to analyse environmental samples over the last decade. The majority of the selected studies (15/18, 83%) were performed on samples belonging to the water compartment on onion bulbs. Half of the selected studies (9/18) were conducted to demonstrate the DNA damaging effect of the sample, while the other half of the studies not only recognized the presence of genotoxic agents but also addressed possible remediation measures. Detailed analysis of the experimental procedures revealed heterogeneity in many key steps, such as exposure time, test controls, nuclei isolation solutions, duration of electrophoresis, and number of nuclei scored. This literature review has shown that the comet assay on A. cepa, although recognized as an appropriate tool, is underutilized in environmental toxicology. Greater standardization could lead to its more widespread use, providing valuable information on the genotoxicity of environmental samples and the ability of different processes to mitigate their negative effects on plants.