Neonicotinoids are systemic, broad-spectrum insecticides that are largely applied on crops for human consumption. Recently added to the EU watch list as a drug to be monitored (2018/840/ EC), acetamiprid (ACE) is a chlorinated nicotine pesticide primarily used to prevent Lepidoptera and Hemiptera pests. Environmental transformation of acetamiprid produces metabolites that can exhibit very high persistency in soil and high mobility in groundwaters. This combined with high water solubility, facilitates the transport and contamination of nearby freshwater compartments. Additionally to the active ingredients (a.i.), insecticides used in agricultural production contain several additives, solvents and excipients that are listed as inert but can exhibit relevant toxicological properties. The main objective of this work was to assess the toxicity of acetamiprid as an active ingredient (96% purity) and its commercial formulation EPIK SG® (20% p/p a.i.) to freshwater non-target organisms of different trophic levels. Worst-case scenarios of toxic exposure were evaluated, by carrying short-term toxicity assays with primary producers (Raphidocelis subcapitata, 72h-growth inhibition assay), primary consumers (Daphnia magna, 48h immobilization assay) and secondary consumers (Hydra viridissima, 96h mortality assay; Xenopus laevis and Pelophylax perezi, 96h FETAX assay, 96h tadpole acute toxicity assay). Behaviour assays were also performed with Xenopus laevis tadpoles. Mortality and sublethal endpoints were evaluated and concentrations causing 20 and 50% of lethal (LCx) and sublethal effects (ECx) were estimated. Primary producers were the less sensitive species to EPIK SG^® and acetamiprid (EC_{50, EPIK SG®} = 517.8 mg/L of a.i., EC_{50, ACE} = 747.9 mg/L for R. subcapitata growth rate). H. viridissima revealed the highest lethality to EPIK SG® (LC₅₀ = 22.04 mg/L of a.i.) and P. perezi to acetamiprid (LC₅₀ = 134.4 mg/L). Behaviour testing proved to be much more sensitive than traditional toxicity tests. EPIK SG® was the most toxic compound to all freshwater species tested, showing that formulation additives can exhibit a higher toxicity by themselves and/or enhance the toxicity of active ingredients. These findings highlight the necessity to invest in alternative models of conventional formulas, minimizing the toxicity and persistence in the environment, contributing towards more sustainable practices.

The naturally occurring resveratrol (3,5,4′-trihydroxy-trans-stilbene) is a phytoalexin produced by plants in response to various stresses, promoting disease resistance. It received a great attention for its ability to impact on multiple key processes in cancer cell biology, although limitations in terms of poor solubility in water and extensive phase II metabolism reduce the bioavailability to less than 1%. Chemically, resveratrol contains a stibene scaffold that represents a suitable tool for chemical modifications with the aim to obtain derivatives with enhanced bioavailability and pharmacological activity. Bearing in mind that the 4′-OH was considered essential for the antioxidant/antitumor activity, we synthesized analogs of the resveratrol in which the 4′-OH was preserved, but the 3,5-OH moiety was replaced with 4-substituted phenyls. Some of the synthesized compounds have proven to be active in inhibiting cell viability across three distinct PC cell lines, as compared to the parent compound resveratrol. In this presentation, an insight into the more active compound will be described. In particular, we will refer to the ability of the most active analogue in modulating cancer-relevant molecular pathways, in a panel of PC cell lines with distinct genetic profiles. The results will provide to gain insights into the mechanisms involved in antiproliferative action, supporting the potential value in the search for effective and safe agents in PC treatment.

Cancer remains a paramount global health challenge with multifaceted mechanisms driving its proliferation. One pivotal target in this complex landscape is the Mitotic kinesin Eg5, a key player during cell division. Inhibiting this protein holds immense potential to combat cancer effectively. Building on a recent breakthrough, our study aims to enhance the inhibitory efficacy against Eg5 by optimizing a promising novel compound identified in our previous research, which has shown superior inhibitory properties compared to a standard treatment. As we progress through the drug discovery funnel, our focus shifts to lead optimization. We seek to elevate the binding affinity of the lead compound while simultaneously enhancing its Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) properties. This optimization phase is critical for ensuring the compound's therapeutic efficacy, safety, and potential for clinical translation. Our proposed research aligns with the urgent need for innovative cancer treatments and addresses a vital aspect of drug development. By refining the lead compound's molecular interactions and ADMET profile, we endeavor to contribute to the advancement of targeted cancer therapy, potentially bringing us closer to a transformative solution in the battle against cancer.

Prostate cancer is a prevalent malignancy that ranks second in both incidence and mortality rates among males. Although different treatment options are available, they have drawbacks (e.g., limited specificity, high cytotoxicity, and development of resistance). To address these shortcomings, researchers are focusing on more efficient alternative strategies. One of such approach involves the development of more efficient drug delivery systems. Polymeric nanoparticles (NPs), particularly those composed of bovine serum albumin (BSA), are emerging as a promising, environmental-friendly strategy for targeted drug delivery. These NPs offer several advantages, including biocompatibility, stability, and biodegradability. The main objective of this study was to synthesize BSA nanoparticles, using a genipin-based crosslinking method, to be used as carriers for two compounds: an antimetabolite (5-fluorouracil) and an antidepressant (sertraline), which have shown potential efficacy in the treatment of prostate cancer. The effects of the synthesized particles (with and without loaded drugs) on the PNT-2 cell line (normal prostate cells) and the 22Rv1 cell line (prostate cancer cells) viability to validate its potential utility. Of the synthesized particles, BSA NPs with a size range of 200–400 nm with low size variability (polydispersity index < 0.2) were selected for encapsulation procedures. BSA NPs and genipin exhibited non-toxicity in both cell lines, while sertraline-loaded BSA NPs showed a nullifying effect. These results highlight the promising potential of BSA NPs for biological applications.

Millions of deaths are caused every year by cancer, which is one of the most widespread diseases in the world. New innovative therapies have been developed by nanomedicine in recent years to address this issue, specifically using drug delivery methods and approaches. Among the many materials used to create nanocarriers, biopolymers like chitosan are widely utilized. The aim of this work is to describe the synthesis of chitosan nanocapsules (NCs) and the encapsulation of anticancer compounds. Two substances have been chosen: doxorubicin (DOX), a drug largely employed in the treatment of cancer for decades, and aloe-emodin (AE), an anthraquinone isolated from the plant of Aloe vera L., that has been shown to have a strong anti-carcinogenic effect. The commercially available A.vera latex was used to extract AE. The obtained Dox-loaded and AE-loaded chitosan NCs were characterized by Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM), and Dynamic Light Scattering (DLS). Moreover, UV-vis spectroscopy was used to analyze the encapsulation efficiency of these nanomaterials. Finally, cytotoxicity and in vitro release studies of active agents have been conducted on the cell line of human adenocarcinoma breast MDA-MB-231. The results obtained showed that doxorubicin and aloe-emodin were released gradually from NCs over time and with a dose-dependent effect, resulting in a significant reduction in cellular viability. Moreover, immunofluorescence observations have revealed the uptake of DOX-NCs and AE-NCs in MDA-MB-231 cells and their localization both in the cytoplasm and in the nuclear region.

Nanoparticles are widely studied in nanomedicine for their potential use as drug delivery systems due to their possibility of engineering them for multiple purposes. In the context of nanocarriers, liposomes can be a powerful tool for the transport of bioactive molecules, thanks to their unique characteristics, such as low toxicity and ability to encapsulate a wide range of drugs, and, above all, the ability to finely tune their features by modulating the formulation.

In this scenario, we studied how the presence of different lipid components can play a central role by influencing the physico-chemical characteristics of the final nanosystems. In particular, we mainly focused on how the inclusion of cationc diasteromeric amphiphiles within the formulation can define the characteristics of the liposomes as a whole, in terms of charge, size, fluidity, ability to encapsulate and retain different types of probes, and how these features can then dictate different biological outcome and final fate.

In this work, we design and investigated liposomes composed of natural phospholipids and cholesterol in mixture with synthetic diastereomeric cationic gemini amphiphiles. All formulations were characterized in terms of mean diameter, polydispersity index (PDI) and stability over time by dynamic laser light scattering measurements (DLS). The most promising nanosystems were then evaluated from a biological point of view, in particular uptake experiments were carried out on a monolayer of human iPSC derived brain microvascular endothelial cells (iBMECs) and transport experiment were performed across an in vitro human BBB model (iBMECs in coculture with human astrocyets), in order to evaluate the different biological response.

Acknowledgments: This research was supported by Regione Lazio through the projects “Torno Subito” Ed. 2018 (PROT.N. 1179), and “Lipobarr”(PROT.N. 85-2017-15057), by Ministero dell' Università e della Ricerca through the project "CNCCS-Centre for rare, neglected and poverty related diseases", by Sapienza University of Rome through “Fondi Avvio alla ricerca 2021” PROT.N. 36805), and by HORIZON-EIC-2022-PATHFINDEROPEN-01 through the project “Bioaction” PROT.N. 101098972).

Anacardium occidentale (cashew) stem bark is used in traditional medicine to treat various kinds of diseases such as diabetes, fever, liver disorder and bronchitis in different parts of the world, including Nigeria. This study was aimed at phytochemical profiling of methanol extract of Anarcardium occidentale (cashew) stem bark (MEAOSB) and evaluation of its effect on antioxidant and liver function markers. In the experimental design of this study, a total of 25 Wistar albino rats divided into five groups, of five rats each, were used. Hepatotoxicity was induced with 2 ml/kg of carbon tetrachloride (CCl₄) in all the groups except group one (1) which served as normal control. Group 2 was induced but not treated and served as the positive control while group 3-5 were induced and treated with standard (silymarin), 200 and 400 mg/kg of the extract respectively. Treatment lasted a week. Phytochemical profiling was done with GCMS while biochemical analyses of the parameters were determined by standard protocol. As observed from the results, induction with CCl₄ significantly decreased the activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione transferase and concentrations of glutathione and malondialdehyde. In contrast, induction resulted in an increase in liver markers such as alkaline phosphatase, aspartate transaminase, alanine transaminase and total bilirubin. However treatment with MEAOSB significantly (P < 0.05) elicited antioxidant and hepatoprotective effect by increasing the levels of antioxidant markers and lowering the levels serum liver markers which were in turn comparable with that of the reference drug silymarin. Findings from this research indicate that high doses of CCl₄-induction could result in Oxidative and hepatic damage and MEAOSB has the potential of scavenging radicals and protecting the liver against CCl₄-induced oxidation and hepatotoxicity. The observed effect of MEAOSB could be due to the presence of Aspidospermidin-17-ol, hexadecanoic acid,methyl ester and Aziridinone in the extract shown by the GCMS analysis.

The combretazet beta-lactams are cis-restricted analogues of combretastatin A-4, chemically manipulated to enhance their in vitro and in vivo stability. They are extremely potent anti-proliferative agents targeting the colchicine-binding site of tubulin, as microtubule destabilisers leading to G₂/M phase arrest, karyorrhexis and ultimately, apoptosis and cell death. 3-Hydroxyl combretazets demonstrate excellent IC₅₀ values in subnanomolar range across a panel of cancer cell lines. We have previously reported chiral diastereomeric resolution of b-lactam racemates using chiral derivatising reagent N-Boc-L-proline to afford enantiopure b-lactams, albeit with large process mass intensities (PMI) of approximately 150,000 kg/kg and modest yields (5-10%), insufficient for progression toward in vivo pre-clinical toxicology studies. The 3S,4S enantiomer was isolated in >90% enantiomeric excess (ee). However, poor diastereomeric excess was obtained for the second eluting 3R,4R diastereomer yielding equally poor ee (50-60%) for its corresponding enantiomer. The process was labour and solvent intensive. Committed to the design of a sustainable, green and safer resolution process, biocatalysis using lipase chemoenzymatic mediated kinetic resolution (KR) toward combretazet enantiomers was investigated.Candida antarctica lipase B (CAL-B) catalysed KR of 3-acetoxy beta-lactams toward their corresponding enantioenriched 3-hydroxyl 3S,4S beta-lactams in 70-80% ee, comparable to values achieved via diastereomeric resolution. PMI was reduced by 1,300-30,000 fold compared to diastereomeric resolution. Candida antarctica KR of combretazet b-lactams is an accessible enantioseparation strategy, reducing environmental impact while overcoming issues associated with poor yield, with KR yielding sufficient quantities of beta-lactam enantiomers for pre-clinical in vivo toxicological evaluation.

Tire wear particles (TWP), also known as tire dust, are formed when friction occurs between tires and the road surface. As the particles continue to come into contact with the road surface, they can reach increasingly smaller sizes, in the micro and nanoscale, making them more susceptible to transportation by wind and/or water. Physical damage to organisms as well as parallel toxicity exerted by compounds on the TWP composition that are released over time have already been reported. However, to induce quantifiable effects, the concentrations used are amplified and therefore fall outside the sphere of environmental relevance, while studies evaluating the ageing of the particles and the parallel release of compounds from their composition over time have yet to be reported. The study aimed to evaluate the toxicity of a TWP suspension at environmentally relevant concentrations ranging between 0.5 and 512 mg TWP/L and at three ageing stages: immediately after preparation (0dA), 7 days after preparation (7dA), and 30 days after preparation (30dA). The zebrafish model species Danio rerio was chosen to perform these assays due to its utility as a homolog for potential effects that TWP may cause on humans, and due to the multiplicity of sublethal endpoints that D. rerio allows for analysis. The 96h assays followed the OECD guideline 236 (OECD, 2013) using both embryos and post-hatching larvae to determine potential different routes of uptake. Mortality and malformations were registered every 24h, in addition to the determination of the heartbeat rate (HBR), total and interocular length (TL and IL, respectively), swim bladder area (SBA), and behaviour (total swim distance and time, thigmotaxis).

No significant mortality and malformations were registered in all assays of embryos and larvae (>10%). In embryos, HBR presented a pattern of tachycardia between concentrations of 0.5 and 8 mg TWP/L, whilst a pattern of bradycardia at higher concentrations (<32 mg TWP/L), for all ageing stages. Also in embryos, TL decreased significantly (p<0.05) at the highest concentration (512 mg TWP/L) compared to the control for all ageing stages, whilst IL was significantly (p<0.05) higher at concentrations <8 mg TWP/L compared with the control for 0dA. The SBA was significantly lower at 128 and 512 mg TWP/L compared to the control in organisms exposed to 30dA (p<0.05). In the post-hatching larvae, it is to highlight the significant increment in IL at concentrations <8 mg TWP/L for 0dA and 7dA suspensions. Larvae SBA decreased significantly (p<0.05) at all tested concentrations (except 0.5 mg TWP/L) compared to the control of 0dA. About behavioural assessment carried out at the end of each exposure, it is worth noting that activity time or distance travelled were zero or almost zero during periods of exposure to light at concentrations between 32 and 512 mg TWP/L for all stages of ageing of the suspensions, compared to the control organisms. The results obtained highlight that, in addition to the lack of ecotoxicological studies on the effects of TWP, experimental designs must be carefully planned to increase the level of realism of the experiments and to increase knowledge and understanding of the impact that this type of particulate may have on ecosystems and, ultimately, human health.

Proliferating cell nuclear antigen (PCNA) is a key regulatory protein in human DNA metabolism. PCNA forms a ring-like structure around the DNA that serves as a binding platform for a multitude of other proteins involved in DNA metabolism. Ataxia-telangiectasia-like disorder type 2 (ATLD2) is a neurodegenerative disease associated with impaired PCNA function. Recently, a mutation in the PCNA gene of ATLD2 patients was identified that encodes for the NM_002592.2(PCNA): c.443G > C(p.C148S) variant. However, the molecular effect of this mutation was unclear. Here we used non-equilibrium free (NEQ) energy calculations to predict the effect of this single amino acid mutation on PCNA stability and on the interaction of PCNA with one of its binding partner’s, p15. No change in binding affinity was predicted, while a significant decrease in the folding free energy was predicted for the PCNA^C148S variant. These results were validated experimentally by differential scanning fluorimetry and a FRET-based PCNA-p15 interaction assay. The experiments confirmed a reduced folding free energy of the variant, while no direct influence of the mutation on the PCNA-p15 interaction is detected. However, the lower folding free energy of the variant caused a time-dependent denaturation of PCNA^C148S. This provides a possible molecular explanation for the association of the PCNA^C148S variant with ATLD2. This study therefore underlines the effectiveness of NEQ alchemical free energy calculations in the analysis of disease conferring mutations.