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Peripheral nervous system (PNS) toxicity assessment in non-clinical safety studies is challenging and relies mostly on histopathological assessment. The present work aims to identify blood-based biomarkers that could detect peripheral neuropathy in rats upon exposure to neurotoxic compounds. Three anticancer agents (oxaliplatin, cisplatin, paclitaxel) and a developmental compound (NVS-1) were assessed in male rats (Wistar Han). Clinical and/or functional endpoints (i.e., electronic Von Frey, Cold Plate, and Paw Pressure tests) and blood biomarkers (i.e., neurofilament light chain (NfL), neurofilament heavy chain (NF-H), microtubule-associated protein Tau (Tau), neuron specific enolase (NSE), vascular endothelial growth factor A (VEGFA), and glial fibrillary acidic protein (GFAP)) were assessed. Drug exposure and histopathological evaluations were conducted on selected nervous tissues. Oxaliplatin, cisplatin and paclitaxel treatment resulted in a significant decrease of nociceptive thresholds. Clinical signs suggestive of PNS toxicity were observed with NVS-1. NfL was consistently increased in the NVS-1 study and correlated with moderate microscopic findings in dorsal root ganglia (DRG). Only minimal microscopic findings were observed in oxaliplatin-treated animals, whereas no treatment-related microscopic findings were observed in animals treated with cisplatin and paclitaxel. For all compounds, exposure was confirmed in the PNS tissues. Clinical and functional changes were observed with all the compounds evaluated. NfL levels in plasma proved to be the most sensitive indicator of PNS toxicities, capturing moderate nervous degeneration in DRG. A combined approach that includes both functional assessments and biomarker measurements offers a more comprehensive evaluation than histopathological analysis alone when monitoring drug-induced neurotoxicity in rat models.

Cell Painting is an established community-based microscopy-assay platform that provides high-throughput, high-content data for biological readouts. In November 2022, the JUMP-Cell Painting Consortium released the largest publicly available Cell Painting dataset with CellProfiler features, comprising more than 2 billion cell images. This dataset is designed for predicting the activity and toxicity of 115k drug compounds, with the aim to make cell images as computable as genomes and transcriptomes. In this context, our paper introduces a scalable and computationally efficient data analytics workflow created to meet the needs of researchers. This data-driven workflow facilitates the comparison of drug treatment effects through significant and biologically relevant insights. The workflow consists of two parts: first, the Equivalence score (Eq. score), a straightforward yet sophisticated metric highlighting relevant deviations from negative controls based on cell image morphology; second, the scalability of the workflow, by utilizing the Eq. scores on a large scale to predict and classify the subtle morphological changes in cell image profiles. By doing so, we show classification improvements compared to using the raw CellProfiler features on the CPJUMP1-pilot dataset on three types of perturbations. We hope that our workflow's contributions will enhance drug screening efficiency and streamline the drug development process. As this process is resource-intensive, every incremental improvement is valuable. Through our collective efforts in advancing the understanding of high-throughput image-based data, we aim to reduce both the time and cost of developing new, life-saving treatments.

Chemical risk assessments typically focus on single substances, often overlooking real-world co-exposures to chemical mixtures. Mixture toxicology studies using representative mixtures can reveal potential chemical interactions, but these do not account for the unique chemical profiles that occur in the blood of diverse individuals. Here we used the H295R steroidogenesis assay to screen personalized mixtures of 24 persistent organic pollutants (POPs) for cytotoxicity and endocrine disruption. Each mixture was reconstructed at a human exposure relevant concentration (1×), as well as at 10- and 100-fold higher concentration (10×, 100×) by acoustic liquid handling based on measured blood concentrations in a Swedish cohort. Among the twelve mixtures tested, nine mixtures decreased the cell viability by 4–18%, primarily at the highest concentration. While the median and maximum mixtures based on the whole study population induced no measurable effects on steroidogenesis at any concentration, the personalized mixture from an individual with the lowest total POPs concentration was the only mixture that affected estradiol synthesis (35% increase at the 100× concentration). Mixtures reconstructed from blood levels of three different individuals stimulated testosterone synthesis at the 1× (11–15%) and 10× concentrations (12–16%), but not at the 100× concentration. This proof-of-principle personalized toxicity study illustrates that population-based representative chemical mixtures may not adequately account for the toxicological risks posed to individuals. It highlights the importance of testing a range of real-world mixtures at relevant concentrations to explore potential interactions and non-monotonic effects. Further toxicological studies of personalized contaminant mixtures could improve chemical risk assessment and advance the understanding of human health, as chemical exposome data become increasingly available.

Developmental neurotoxicity (DNT) is a branch of toxicology that examines the effects of chemicals on the developing nervous system. Traditional methods for assessing DNT mainly rely on animal testing, which raises ethical concerns, is time-consuming, and expensive. Consequently, there is a shift towards alternative methods, such as in vitro and in silico approaches, which offer faster and more efficient testing. The overall aim of this thesis was to contribute to the development and integration of alternative methods for DNT assessment, employing both in vitro and in silico techniques. In this work, the human neuroblastoma SH-SY5Y cell line was utilized as a robust, cost-effective, and easy-to-use model for DNT evaluation. Through RNA sequencing and morphological observation, it was determined that the SH-SY5Y cell line can differentiate into a more neuron-like phenotype (Paper I). Additionally, neurite outgrowth and the mRNA expression of genes important for neuronal development were studied by exposing the cells to chemicals known to induce DNT (Paper II). The thesis has also focused on acrylamide, a neurotoxic compound that may also cause DNT. In Paper I, it was found that acrylamide inhibited neuronal differentiation by suppressing neurite outgrowth at non-cytotoxic concentrations. Moreover, acrylamide altered the expression of several genes involved in the retinoic acid and CREB signaling pathways. The hypothesis that acrylamide impairs neuronal differentiation by depleting glutathione, leading to oxidative stress, was tested but not supported in the SH-SY5Y cells (Paper III). In Paper IV, we performed an in vitro to in vivo extrapolation by using a novel physiologically based toxicokinetic (PBTK) model for pregnant women, to assess the biological relevance of the acrylamide concentrations that affected neuronal differentiation of SH-SY5Y cells. The results revealed that doses that humans may be exposed to through food intake, resulted in fetal plasma acrylami

The present study aims to characterise the pharmacokinetics of rifampicin (RIF) in tuberculosis (TB) patients with and without HIV co-infection, considering the formation of 25-O-desacetyl-rifampicin (desRIF). It is hypothesised that the metabolite formation, HIV co-infection and drug formulation may further explain the interindividual variation in the exposure to RIF. Pharmacokinetic, clinical, and demographic data from TB patients with (TB-HIV+ group; n = 18) or without HIV (TB-HIV- group; n = 15) who were receiving RIF as part of a four-drug fixed-dose combination (FDC) regimen (RIF, isoniazid, pyrazinamide, and ethambutol) were analysed, along with the published literature data on the relative bioavailability of different formulations. A population pharmacokinetic model, including the formation of desRIF, was developed and compared to a model based solely on the parent drug. HIV co-infection does not alter the plasma exposure to RIF and the desRIF formation does not contribute to the observed variability in the RIF disposition. The relative bioavailability and RIF plasma exposure were significantly lower than previously reported for the standard regimen with FDC tablets. Furthermore, participants weighting less than 50 kg do not reach the same RIF plasma exposure as compared to those weighting >50 kg. In conclusion, as no covariate was identified other than body weight on CL/F and Vd/F, low systemic exposure to RIF is likely to be caused by the low bioavailability of the formulation.

Bioluminescence- and fluorescence-based resonance energy transfer assays have gained considerable attention in pharmacological research as high-throughput scalable tools applicable to drug discovery. To this end, G proteincoupled receptors represent the biggest target class for marketed drugs, and among them, orphan G proteincoupled receptors have the biggest untapped therapeutic potential. In this review, the cases where biophysical methods, BRET and FRET, were employed for deorphanization and ligand discovery studies on orphan G proteincoupled receptors are listed and discussed.

Background and main body Pharmacokinetics (PK) and pharmacodynamics (PD) are central concepts to guide the dosage and administration of drug therapies and are essential to consider for both healthcare professionals and researchers in therapeutic planning and drug discovery. PK/PD properties of a drug significantly influence variability in response to treatment, including therapeutic failure or excessive medication-related harm. Furthermore, suboptimal PK properties constitute a significant barrier to further development for some candidate treatments in drug discovery. This article describes how extracellular vesicles (EVs) affect different aspects of PK and PD of medications and their potential to modulate PK and PD properties to address problematic PK/PD profiles of drugs. We reviewed EVs' intrinsic effects on cell behaviours and medication responses. We also described how surface and cargo modifications can enhance EV functionalities and enable them as adjuvants to optimise the PK/PD profile of conventional medications. Furthermore, we demonstrated that various bioengineering strategies can be used to modify the properties of EVs, hence enhancing their potential to modulate PK and PD profile of medications. Conclusion This review uncovers the critical role of EVs in PK and PD modulation and motivates further research and the development of assays to unfold EVs' full potential in solving PK and PD-related problems. However, while we have shown that EVs play a vital role in modulating PK and PD properties of medications, we postulated that it is essential to define the context of use when designing and utilising EVs in pharmaceutical and medical applications.

Background Vincristine-induced peripheral neuropathy (VIPN) is a common adverse effect of vincristine, a drug often used in pediatric oncology. Previous studies demonstrated large inter- and intrapatient variability in vincristine pharmacokinetics (PK). Model-informed precision dosing (MIPD) can be applied to calculate patient exposure and individualize dosing using therapeutic drug monitoring (TDM) measurements. This study set out to investigate the PK/pharmacodynamic (PKPD) relationship of VIPN and determine the utility of MIPD to support clinical decisions regarding dose selection and individualization. Methods Data from 35 pediatric patients were utilized to quantify the relationship between vincristine dose, exposure and the development of VIPN. Measurements of vincristine exposure and VIPN (Common Terminology Criteria for Adverse Events [CTCAE]) were available at baseline and for each subsequent dosing occasions (1–5). A PK and PKPD analysis was performed to assess the inter- and intraindividual variability in vincristine exposure and VIPN over time. In silico trials were performed to portray the utility of vincristine MIPD in pediatric subpopulations with a certain age, weight and cytochrome P450 (CYP) 3A5 genotype distribution. Results A two-compartmental model with linear PK provided a good description of the vincristine exposure data. Clearance and distribution parameters were related to bodyweight through allometric scaling. A proportional odds model with Markovian elements described the incidence of Grades 0, 1 and ≥ 2 VIPN overdosing occasions. Vincristine area under the curve (AUC) was the most significant exposure metric related to the development of VIPN, where an AUC of 50 ng⋅h/mL was estimated to be related to an average VIPN probability of 40% over five dosing occasions. The incidence of Grade ≥ 2 VIPN reduced from 62.1 to 53.9% for MIPD-based dosing compared with body surface area (BSA)-based dosing in patients. Dose decreases occurred in 81.4% o, De två första författarna delar förstaförfattarskapet

Background: Standard dosages of analgesic and sedative drugs are given to intensive care patients. The resulting range of blood concentrations and corresponding clinical responses need to be better examined. The purpose of this study was to describe daily dosages, measured blood concentrations, and clinical responses in critically ill patients. The purpose was also to contribute to establishing whole blood concentration reference values of the drugs investigated. Methods: A descriptive study of prospectively collected data from 302 admissions to a general intensive care unit (ICU) at a university hospital. Ten drugs (clonidine, fentanyl, morphine, dexmedetomidine, ketamine, ketobemidone, midazolam, paracetamol, propofol, and thiopental) were investigated, and daily dosages recorded. Blood samples were collected twice daily, and drug concentrations were measured. Clinical responses were registered using Richmond agitation-sedation scale (RASS) and Numeric rating scale (NRS). Results: Drug dosages were within recommended dose ranges. Blood concentrations for all 10 drugs showed a wide variation within the cohort, but only 3% were above therapeutic interval where clonidine (57 of 122) and midazolam (38 of 122) dominated. RASS and NRS were not correlated to drug concentrations. Conclusion: Using recommended dose intervals for analgesic and sedative drugs in the ICU setting combined with regular monitoring of clinical responses such as RASS and NRS leads to 97% of concentrations being below the upper limit in the therapeutic interval. This study contributes to whole blood drug concentration reference values regarding these 10 drugs.

Cancer continues to pose a significant global health challenge, claiming numerous lives prematurely and necessitating the use of potent cytotoxic chemotherapeutics such as 5-fluorouracil (5-FU). Chemotherapeutics are efficiently killing cancer cells but the treat-ments frequently cause off-target effects such as chemotherapy-induced mucositis (CIM), characterized by debilitating symptoms like diarrhoea, that require a medical intervention. In this study, we elucidated the efficacy of melatonin and misoprostol in 5-FU-induced small intestinal mucositis. Morphological and cellular changes in the je-junum, along with colonic faecal water content were quantified in rats as markers for CIM. Additionally, the effects of melatonin were investigated in vitro on 5-FU treated murine intestinal organoids. The results showed that melatonin prevented villus atrophy in the rat jejunal mucosa and upheld cell viability in murine intestinal organoids. In contrast, misoprostol alone or in combination with melatonin did not significantly affect CIM caused by 5-FU. These in vivo and in vitro experiments provide promising insights that melatonin may be used as a preventive and/or adjuvant combination therapy to reduce CIM, holding the potential to enhance cancer treatment outcomes and improve patient quality-of-life., Contributed equally to this work: Karsten Peters, Ada Lerma Clavero

The nucleic acid topoisomerases (TOP) are an evolutionary conserved mechanism to solve topological problems within DNA and RNA that have been historically well-established as a chemotherapeutic target. During investigation of trends within clinical trials, we have identified a very high number of clinical trials involving TOP inhibitors, prompting us to further evaluate the current status of this class of therapeutic agents. In total, we have identified 233 unique molecules with TOP-inhibiting activity. In this review, we provide an overview of the clinical drug development highlighting advances in current clinical uses and discussing novel drugs and indications under development. A wide range of bacterial infections, along with solid and hematologic neoplasms, represent the bulk of clinically approved indications. Negative ADR profile and drug resistance among the antibacterial TOP inhibitors and anthracycline-mediated cardiotoxicity in the antineoplastic TOP inhibitors are major points of concern, subject to continuous research efforts. Ongoing development continues to focus on bacterial infections and cancer; however, there is a degree of diversification in terms of novel drug classes and previously uncovered indications, such as glioblastoma multiforme or Clostridium difficile infections. Preclinical studies show potential in viral, protozoal, parasitic and fungal infections as well and suggest the emergence of a novel target, TOP III beta. We predict further growth and diversification of the field thanks to the large number of experimental TOP inhibitors emerging.

The basic principle for the treatment of idiopathic diarrhoea (functional diarrhoea K59.1) is to delay transit through the gut in order to promote the absorption of electrolytes and water. Under mild conditions, bulking agents may suffice. With increasing severity, antidiarrhoeal pharmaceuticals may be added in a stepwise manner. In diarrhoea of unknown aetiology, peripherally-acting opioid receptor agonists, such as loperamide, are first-line treatment and forms the pharmaceutical basis of antidiarrheal treatment. As second-line treatment opium drops have an approved indication for severe diarrhoea when other treatment options fail. Beyond this, various treatment options are built on experience with more advanced treatments using clonidine, octreotide, as well as GLP-1 and GLP-2 analogs which require specialist knowledge the field.

Mycotoxins are secondary metabolites of filamentous fungi and ubiquitous dietary contaminants. Aflatoxins, a group of mycotoxins with high prevalence and toxicity, have raised a high level of public health concern, the most prevalent and toxic being aflatoxin B1 (AFB1). Many aspects appertaining to AFB1 poisoning are not well understood. Yet this information is necessary to devise appropriate surveillance and mitigation strategies against human and animal aflatoxicosis. This review provides an in-depth update of work carried out on mycotoxin poisoning, particularly aflatoxicosis in humans and animals, to identify gaps in knowledge. Hypotheses explaining the functional significance of mycotoxins in fungal biology and their dietary epidemiological data are presented and briefly discussed. The toxicology of aflatoxins and the challenges of their mitigation are discussed in depth. It was concluded that the identification of potential mycotoxin-hazard-prone food items and quantification of the associated risk of cancer ailments in humans is a prime priority. There is a dearth of reliable sampling methodologies for estimating AFB1 in animal feed. Data update on AFB1 in animal feed and its implication in animal production, mitigation strategies, and elucidation of risk factors to this hazard is required. To reduce the burden of aflatoxins, surveillance employing predictive technology, and biocontrol strategies seem promising approaches.

Objectives: Clofazimine is a promising drug for the treatment of nontuberculous mycobacterial (NTM) diseases. Accumulation of clofazimine to reach steady-state plasma concentrations takes months. A loading dose may reduce the time to steady-state-like concentrations. We evaluated the pharmacokinetics (PK), safety and tolerability of a loading dose regimen in patients with NTM disease. Methods: Adult participants received a 4-week loading dose regimen of 300 mg clofazimine once daily, followed by a maintenance dose of 100 mg once daily (combined with other antimycobacterial drugs). Blood samples for PK analysis were collected on three occasions. A population PK model for clofazimine was developed and simulations were performed to assess the time to reach steady-state-like (target) concentrations for different dosing regimens. Results: Twelve participants were included. The geometric mean peak and trough clofazimine concentrations after the 4-week loading phase were 0.87 and 0.50 mg/L, respectively. Adverse events were common, but mostly mild and none led to discontinuation of clofazimine. Our loading dose regimen reduced the predicted median time to target concentrations by 1.5 months compared to no loading dose (3.8 versus 5.3 months). Further time benefit was predicted with a 6-week loading dose regimen (1.4 versus 5.3 months). Conclusion: A 4-week loading dose regimen of 300 mg once daily reduced the time to target clofazimine concentrations and was safe and well-tolerated. Extending the loading phase to 6 weeks could further decrease the time to target concentrations. Using a loading dose of clofazimine is a feasible strategy to optimize treatment of NTM disease. Clinical trials registration: NCT05294146.

Background: Cost-utility analyses (CUAs) increasingly use models to predict long-term outcomes and translate trial data to real-world settings. Model structure uncertainty affects these predictions. This study compares pharmacometric against traditional pharmacoeconomic model evaluations for CUAs of sunitinib in gastrointestinal stromal tumors (GIST). Methods: A two-arm trial comparing sunitinib 37.5 mg daily with no treatment was simulated using a pharmacometric-based pharmacoeconomic model framework. Overall, four existing models [time-to-event (TTE) and Markov models] were re-estimated to the survival data and linked to logistic regression models describing the toxicity data [neutropenia, thrombocytopenia, hypertension, fatigue, and hand-foot syndrome (HFS)] to create traditional pharmacoeconomic model frameworks. All five frameworks were used to simulate clinical outcomes and sunitinib treatment costs, including a therapeutic drug monitoring (TDM) scenario. Results: The pharmacometric model framework predicted that sunitinib treatment costs an additional 142,756 euros per quality adjusted life year (QALY) compared with no treatment, with deviations - 21.2% (discrete Markov), - 15.1% (continuous Markov), + 7.2% (TTE Weibull), and + 39.6% (TTE exponential) from the traditional model frameworks. The pharmacometric framework captured the change in toxicity over treatment cycles (e.g., increased HFS incidence until cycle 4 with a decrease thereafter), a pattern not observed in the pharmacoeconomic frameworks (e.g., stable HFS incidence over all treatment cycles). Furthermore, the pharmacoeconomic frameworks excessively forecasted the percentage of patients encountering subtherapeutic concentrations of sunitinib over the course of time (pharmacoeconomic: 24.6% at cycle 2 to 98.7% at cycle 16, versus pharmacometric: 13.7% at cycle 2 to 34.1% at cycle 16). Conclusions: Model structure significantly influences CUA predictions. The pharmacometric-based model framework mor

Endocrine Disrupting Chemicals (EDCs) are compounds that interfere with normal hormone balances. Reproductive functions rely on precise endocrine regulation involving hormones produced in the brain and ovaries. Several epidemiological studies have shown associations with EDC exposure and adverse fertility outcomes. However, effects of EDCs on the endometrium are still not well investigated. Therefore, this PhD thesis aimed to unravel the effects of EDCs, especially phthalates, on endometrial cell function. Study I showed the effects of several EDCs on decidualisation, a key process for a successful pregnancy. Protein kinases involved in the decidualisation process, as well as PRL and IGFBP1 excretion, were reduced by different EDCs. To continue our search for candidate genes and pathways affected by EDCs, we exposed different endometrial primary cells and cell lines to phthalate mixtures in study II. Phthalate mixtures were based on urine samples from the Midlife Women’s Health study cohort. All mixtures affected endometrial cells, with MEHHP being the main actor in the phthalate mixtures. Many genes and pathways found with RNA sequencing were involved with the cytoskeleton and mitochondria. Therefore, we conducted study III, where we investigated the effect of MEHHP on mitochondrial metabolism and structure in the endometrial stromal cell line T-HESC. While mitochondrial metabolism was not significantly affected overall, a significant concentration-response effect was observed in spare respiratory capacity. Mitochondrial structure was significantly altered by MEHHP exposure, resulting in decreased branching and volume, but increased sphericity. In conclusion, EDCs affected endometrial cells by disrupting decidualisation; phthalates specifically affected genes related to mitochondria and the cytoskeleton. MEHHP had the most substantial effect on gene expression, significantly affecting spare respiratory capacity and mitochondrial structure. Future studies should

Drug-induced liver injury (DILI) has been a significant challenge in drug discovery, often leading to clinical trial failures and necessitating drug withdrawals. Over the last decade, the existing suite of in vitro proxy-DILI assays has generally improved at identifying compounds with hepatotoxicity. However, there is considerable interest in enhancing the in silico prediction of DILI because it allows for evaluating large sets of compounds more quickly and cost-effectively, particularly in the early stages of projects. In this study, we aim to study ML models for DILI prediction that first predict nine proxy-DILI labels and then use them as features in addition to chemical structural features to predict DILI. The features include in vitro (e.g., mitochondrial toxicity, bile salt export pump inhibition) data, in vivo (e.g., preclinical rat hepatotoxicity studies) data, pharmacokinetic parameters of maximum concentration, structural fingerprints, and physicochemical parameters. We trained DILI-prediction models on 888 compounds from the DILI data set (composed of DILIst and DILIrank) and tested them on a held-out external test set of 223 compounds from the DILI data set. The best model, DILIPredictor, attained an AUC-PR of 0.79. This model enabled the detection of the top 25 toxic compounds (2.68 LR+, positive likelihood ratio) compared to models using only structural features (1.65 LR+ score). Using feature interpretation from DILIPredictor, we identified the chemical substructures causing DILI and differentiated cases of DILI caused by compounds in animals but not in humans. For example, DILIPredictor correctly recognized 2-butoxyethanol as nontoxic in humans despite its hepatotoxicity in mice models. Overall, the DILIPredictor model improves the detection of compounds causing DILI with an improved differentiation between animal and human sensitivity and the potential for mechanism evaluation. DILIPredictor required only chemical structures as input for prediction

Background/Objectives: Many pregnant women globally suffer from depression and are routinely prescribed selective serotonin reuptake inhibitors (SSRIs). These drugs function by blocking the re-uptake of serotonin by the serotonin transporter (SERT) into neurons, resulting in its accumulation in the presynaptic cleft. Despite a large amount of research suggesting a potential link to neurodevelopmental disorders in children whose mothers took these drugs during pregnancy, their possible adverse effects are still debated, and results are contradictory. On the other hand, there is an immediate need for improved cell-based models for developmental neurotoxicity studies (DNT) to minimize the use of animals in research. Methods: In this study, we aimed to assess the effects of clinically relevant concentrations of paroxetine (PAR), fluoxetine (FLX), and citalopram (CIT)-on maturing neurons derived from human neural stem cells using multiple endpoints. Results: Although none of the tested concentrations of FLX, CIT, or PAR significantly affected cell viability, FLX (10 mu M) exhibited the highest reduction in viability compared to the other drugs. Regarding neurite outgrowth, CIT did not have a significant effect. However, FLX (10 mu M) significantly reduced both mean neurite outgrowth and mean processes, PAR significantly reduced mean processes, and showed a trend of dysregulation of multiple genes associated with neuronal development at therapeutic-relevant serum concentrations. Conclusions: Transcriptomic data and uptake experiments found no SERT activity in the system, suggesting that the adverse effects of FLX and PAR are independent of SERT.

In this work we present the synthesis of benzimidazole-quinoline hybrids series (9a-c and 10a-f), characterized using spectroscopy studies (FT-IR, 1H NMR, and mass spectroscopy). The precursor for the hybrid compounds consists of two schemes: i. synthesis of substituted quinoline-4-carboxylic acids (3a-b) with various acetophenones. ii. The key intermediates (8a-c) were obtained initially from the benzimidazole-5-carboxylates (7a-c), were efficiently synthesized by 'one pot' nitro reductive cyclization reaction between ethyl 3-nitro-4-(substituted amino) benzoates 6a-c and 5-bromothiophene-2-carbaldehyde. iii. Further, the benzimidazole esters (7a-c) were converted into the corresponding hydrazides (8a-c) and then finally obtained the benzimidazole-quinoline hybrids series (9a-c and 10a-f). Compounds 7a and 7b were crystallized and their molecular structures were determined using a single crystal X-ray diffraction method. The resultant compounds from the synthesis were screened (in-silico and in-vitro) for their anti-cancer activities (human melanoma cell line (A375) and human breast cancer cell line (MDA-MB-231)). The p53 receptor protein was used for the molecular docking analysis and compound (name) 10b binds the target site with four hydrogen bonds (-6.25 Kcal/mol). The antioxidant activity revealed compounds 9a (IC50 = 604.8 mu g/mL) and 9b (IC50 = 604.8 mu g/mL 683.7 mu g/mL) to exhibit the highest percentage of inhibition and lowest IC50 value. In addition, compounds 10a and 10b showed high scavenging activity. The compounds 9a (A375: IC50 = 34.7 f 0.9 mu g/mL and MDA-MB-231: IC50 = 20.4 f 1.1 mu g/mL), 10a (A375: IC50 = 19.6 f 1.3 mu g/mL and MDA-MB-231: IC50 = 37.0 f 1.3 mu g/mL) and 10b (A375: IC50 = 16.5 f 1.5 mu g/mL and MDA-MB-231: IC50 = 13.4 f 1.5 mu g/mL) showed the significant cytotoxicity against these human cancer cell lines (melanoma and breast cancer) and can be potential anti-cancer molecules.

Mycobacterium avium complex pulmonary disease is treated with an azithromycin, ethambutol, and rifampicin regimen, with limited efficacy. The role of rifampicin is controversial due to inactivity, adverse effects, and drug interactions. Here, we evaluated the efficacy of clofazimine as a substitute for rifampicin in an intracellular hollow-fiber infection model. THP-1 cells, which are monocytes isolated from peripheral blood from an acute monocytic leukemia patient, were infected with M. avium ATCC 700898 and exposed to a regimen of azithromycin and ethambutol with either rifampicin or clofazimine. Intrapulmonary pharmacokinetic profiles of azithromycin, ethambutol, and rifampicin were simulated. For clofazimine, a steady-state average concentration was targeted. Drug concentrations and bacterial densities were monitored over 21 days. Exposures to azithromycin and ethambutol were 20%-40% lower than targeted but within clinically observed ranges. Clofazimine exposures were 1.7 times higher than targeted. Until day 7, both regimens were able to maintain stasis. Thereafter, regrowth was observed for the rifampicin-containing regimen, while the clofazimine-containing regimen yielded a 2 Log10 colony forming unit (CFU) per mL decrease in bacterial load. The clofazimine regimen also successfully suppressed the emergence of macrolide tolerance. In summary, substitution of rifampicin with clofazimine in the hollow-fiber model improved the antimycobacterial activity of the regimen. Clofazimine-containing regimens merit investigation in clinical trials.

Model-informed precision dosing (MIPD) stands as a significant development in personalized medicine to tailor drug dosing to individual patient characteristics. MIPD moves beyond traditional therapeutic drug monitoring (TDM) by integrating mathematical predictions of dosing, and considering patient-specific factors (patient characteristics, drug measurements) as well as different sources of variability. For this purpose, rigorous model qualification is required for the application of MIPD in patients. This review delves into new methods in model selection and validation, also highlighting the role of machine learning in improving MIPD, the utilization of biosensors for real-time monitoring, as well as the potential of models integrating biomarkers for efficacy or toxicity for precision dosing. The clinical evidence of TDM and MIPD is discussed for various medical fields including infection medicine, oncology, transplant medicine, and inflammatory bowel diseases, thereby underscoring the role of pharmacokinetics/pharmacodynamics and specific biomarkers. Further research, particularly randomized clinical trials, is warranted to corroborate the value of MIPD in enhancing patient outcomes and advancing personalized medicine.

The overall aims of the projects included in this thesis were to discover, functionally characterise, explore biological safety and evaluate potential utility of cysteine-rich peptides, named alpha-nemertides, from marine worms (Nemertea sp.). One of the main outcomes of this thesis is the discovery of a new subfamily of nemertean peptide toxins, from the epidermal mucus of an unexplored marine worm (Paper I). Structure-activity relationships of alpha-nemertides was investigated via alanine scanning on the archetype nemertide alpha-1 (Paper II). Finally, a preliminary investigation of nemertide alpha-1 feeding repellence and acute toxicity was perform in vivo on pest insects (Paper III-IV) alongside with the analysis of its human and biological safety (Paper III). In Manuscript I, the discovery and initial characterisation of a new subfamily of nemertean peptide toxins from Lineus viridis is presented. The new subfamily was named delta-nemertide and the archetype, nemertide delta-1, was successfully isolated and described. Furthermore, transcriptomic analysis led to the addition of seven delta-nemertide putative sequences. Nemertide delta-1 share many similarities with the previously described nemertide alpha-1 except for a 30-fold higher potency on measured insect voltage-gated sodium channels. In Manuscript II, the study focused on determining the importance for selectivity and acute toxicity of each amino acid from nemertide alpha-1 sequence. The mutants’ toxicity was assessed in vitro on a selection of voltage-gated sodium channels and in vivo on Artemia salina. Five positions crucial for overall selectivity were highlighted and one mutant named S12A demonstrated specific selectivity to insect voltage-gated sodium channels. In Manuscript III, nemertide alpha-1 displayed feeding deterrence and feeding behaviour changes at the low molecular level on green peach aphids (Myzus persicae), a known pest insect. In parallel, the peptide demonstrated attractive envir

Abuse of the highly toxic compound fentanyl and its analogues is increasing, raising serious public health concerns due to their potency and availability. Therefore, there is a need for decontamination methodologies to safely remove fentanyl to avoid harmful exposure. In this study, the efficacy of commercial and in-house synthesized decontamination agents (Dahlgren Decon, RSDL (Reactive Skin Decontamination Lotion), FAST-ACT (First applied sorbent treatment against chemical threats), GDS2000, alldecont MED, bleach, Domestos Spray Bleach, Effekt Klor, MgO, TiO2-nanodiamond, and CeO2) were evaluated for the degradation of fentanyl and carfentanil under controlled laboratory conditions and on wooden floor surfaces. Liquid chromatography/mass spectrometry analysis showed that oxidative decontamination agents were the most effective, with N-oxides identified as major degradation products. The physiological effects of these N-oxides were also investigated regarding their ability to activate the µ-opioid receptor and their metabolism in human liver microsomes. The results provide empirical evidence that complements prior research findings on the degradation of fentanyl and carfentanil using a variety of decontamination agents.

Objectives: Antihypertensives reduce the risk of myocardial infarction and stroke. Restrictions during the COVID-19 pandemic limited access to healthcare, which may have had a negative impact on drug prescribing. This study aimed to assess the effect of the COVID-19 pandemic on the initiation of antihypertensive drugs. Design: Interrupted time series study using a segmented linear regression model. Setting: Swedish population assessed through linked national healthcare registers. Participants: 720 300 new users of antihypertensives. Intervention: March 2020, COVID-19 pandemic onset. Main outcomes: measures The change in the initiation of antihypertensives expressed as monthly cumulative incidence, stratified by age and sex. Data on dispensed prescriptions of diuretics, beta-blockers, calcium channel blockers, ACE inhibitors (ACEi) and angiotensin receptor blockers were extracted from the Swedish Prescribed Drug Register, from March 2018 to November 2021. Initiation (new use) was defined as having no previous dispensations before March 2019. Monthly cumulative incidence in March 2019-November 2021 was calculated as the number of patients initiating each drug class in each month divided by the population. Results: The start of the pandemic was associated with an immediate drop in the initiation of any antihypertensive, but no sustained effects were observed, as the incidence continued to increase in the postinterruption period by +0.02% each month in both sexes. The immediate drop was statistically significant for ACEi in both sexes and all antihypertensive classes except diuretics in patients >65 years. & Acy; significant postintervention trend change was observed for initiation of diuretics (+0.013% overall), driven mainly by a significant increase in patients >65 years. Similar findings were also observed for diuretics in females (+0.02%) and ACEi (+0.03%) in patients >65 years. Conclusions: The pandemic had an immediate negative short-term effect, but

Early detection of drug-drug interactions (DDIs) can facilitate timely drug development decisions, prevent unnecessary restrictions on patient enrollment, resulting in clinical study populations that are not representative of the indicated study population, and allow for appropriate dose adjustments to ensure safety in clinical trials. All of these factors contribute to a streamlined drug approval process and enhanced patient safety. Here we describe a new approach for early prediction of the magnitude of change in exposure for cytochrome P450 (P450) CYP3A4-related DDIs of small- molecule anticancer drugs based on the model-based extrapolation of human-CYP3A4-transgenic mice pharmacokinetics to humans. Victim drugs brigatinib and lorlatinib were evaluated with the new approach in combination with the perpetrator drugs itraconazole and rifampicin. Predictions of the magnitude of change in exposure deviated at most 0.99- to 1.31-fold from clinical trial results for inhibition with itraconazole, whereas exposure predictions for the induction with rifampicin were less accurate, with deviations of 0.22- to 0.48-fold. Results for the early prediction of DDIs and their clinical impact appear promising for CYP3A4 inhibition, but validation with more victim and perpetrator drugs is essential to evaluate the performance of the new method. SIGNIFICANCE STATEMENT The described method offers an alternative for the early detection and assessment of potential clinical impact of CYP3A4-related drug-drug interactions. The model was able to adequately describe the inhibition of CYP3A4 metabolism and the subsequent magnitude of change in exposure. However, it was unable to accurately predict the magnitude of change in exposure of victim drugs in combination with an inducer.

Objectives Applying physiologically-based pharmacokinetic (PBPK) modelling in sepsis could help to better understand how PK changes are influenced by drug- and patient-related factors. We aimed to elucidate the influence of sepsis pathophysiology on the PK of meropenem by applying PBPK modelling. Methods A whole-body meropenem PBPK model was developed and evaluated in healthy individuals, and renally impaired non-septic patients. Sepsis-induced physiological changes in body composition, organ blood flow, kidney function, albumin, and haematocrit were implemented according to a previously proposed PBPK sepsis model. Model performance was evaluated, and a local sensitivity analysis was conducted. Results The model-predicted PK metrics (AUC, Cmax, CL, Vss) were within 1.33-fold-error margin of published data for 87.5% of the simulated profiles in healthy individuals. In sepsis, the model provided good predictions for literature-digitised average plasma and tissue exposure data, where the model-predicted AUC was within 1.33-fold-error margin for 9 out 11 simulated study profiles. Furthermore, the model was applied to individual plasma concentration data from 52 septic patients, where the model-predicted AUC, Cmax, and CL had a fold-error ratio range of 0.98–1.12, with alignment of the predicted and observed variability. For Vss, the fold-error ratio was 0.81, and the model underpredicted the population variability. CL was sensitive to renal plasma clearance, and kidney volume, whereas Vss was sensitive to the unbound fraction, organ volume fraction of the interstitial compartment, and the organ volume. Conclusions These findings may be extended to more diverse drug types and support a more mechanistic understanding of the effect of sepsis on drug exposure.

Peptide toxins from marine invertebrates have found use as drugs and in biotechnological applications. Many marine habitats, however, remain underexplored for natural products, and the Southern Ocean is among them. Here, we report toxins from one of the top predators in Antarctic waters: the nemertean worm Parborlasia corrugatus (McIntosh, 1876). Transcriptome mining revealed a total of ten putative toxins with a cysteine pattern similar to that of alpha nemertides, four nemertide-beta-type sequences, and two novel full-length parborlysins. Nemertean worms express toxins in the epidermal mucus. Here, the expression was determined by liquid chromatography combined with mass spectrometry. The findings include a new type of nemertide, 8750 Da, containing eight cysteines. In addition, we report the presence of six cysteine-containing peptides. The toxicity of tissue extracts and mucus fractions was tested in an Artemia assay. Notably, significant activity was observed both in tissue and the high-molecular-weight mucus fraction, as well as in a parborlysin fraction. Membrane permeabilization experiments display the membranolytic activity of some peptides, most prominently the parborlysin fraction, with an estimated EC50 of 70 nM.

Cognitive functions like learning and memory, are natural abilities that humans are highly dependent on to function properly. Neurological disorders like dementia, ischemia, and substance-use disorder may all severely impair cognitive functions. The complexity of the brain and its neurobiology make it challenging to develop potent new therapies that can prevent the onset of disease or relieve symptoms. Few new drugs have been successful in the last decades, thus new targets are wanted. In 1995, a new aminopeptidase member was discovered in fat and muscle cells, the insulin-regulated aminopeptidase (IRAP). IRAP is expressed in many different tissues, including several regions of the brain associated with cognition. The endogenous hexapeptide angiotensin IV (AngIV) and similar ligands binds to IRAP, and causes inhibition of its enzymatic activity. This inhibitory intervention is suggested to improve cognitive functions. The overall aim of the present thesis was to investigate the effects of inhibiting IRAP in the brain. The project has focused on studying restorative effects, cognitive markers, and effects on memory performance using the synthetic IRAP inhibitors HA08 and compound 9 (C9). These two IRAP inhibitors improved cellular function in damaged primary neuronal cultures, an effect that was more prominent in hippocampal cell cultures. The difference in effect between hippocampus and cortical cell cultures may be linked to the composition of cell types and the expression pattern of IRAP in these regions. C9 was also demonstrated to have positive effects on drebrin expression, a protein highly concentrated in dendritic spines. Furthermore, C9 regulated the distribution of microtubule-associated protein 2 (MAP2) positive neurons and glial fibrillary acidic protein (GFAP) positive astrocytes, key markers for improved synaptic activity and cognitive functions, in primary neuronal cultures. Although many positive results were obtained from the in vitro studies, behavi

Metabolic dysfunction-associated steatohepatitis (MASH) is the progressive form of liver steatosis, the most common liver disease, and substantially increases the mortality rate. However, limited therapies are currently available to prevent MASH development. Identifying potential pharmacological treatments for the condition has been hampered by its heterogeneous and complex nature. Here, we identified a hepatic nonneuronal cholinergic signaling pathway required for metabolic adaptation to caloric overload. We found that cholinergic receptor nicotinic alpha 2 subunit (CHRNA2) is highly expressed in hepatocytes of mice and humans. Further, CHRNA2 is activated by a subpopulation of local acetylcholine-producing macrophages during MASH development. The activation of CHRNA2 coordinates defensive programs against a broad spectrum of MASH-related pathogenesis, including steatosis, inflammation, and fibrosis. Hepatocyte-specific loss of CHRNA2 signaling accelerates the disease onset in different MASH mouse models. Activation of this pathway via pharmacological inhibition of acetylcholine degradation protects against MASH development. Our study uncovers a hepatic nicotinic cholinergic receptor pathway that constitutes a cell-autonomous self-defense route against prolonged metabolic stress and holds therapeutic potential for combatting human MASH. The complexity of metabolic dysfunction-associated steatohepatitis (MASH) has impaired the development of pharmacological treatments for the condition. This study shows that nicotinic acetylcholine receptor signaling serves as an intrinsic protective mechanism against extended metabolic stress in the liver.

Background: Pharmacogenetic profiling and therapeutic drug monitoring (TDM) have both been proposed to manage inter-individual variability (IIV) in drug exposure. However, determining the most effective approach for estimating exposure for a particular drug remains a challenge. This study aimed to quantitatively assess the circumstances in which pharmacogenetic profiling may outperform TDM in estimating drug exposure, under three sources of variability (IIV, inter-occasion variability (IOV) and residual unexplained variability (RUV)). Methods: Pharmacokinetic models were selected from the literature corresponding to drugs for which pharmacogenetic profiling and TDM are both clinically considered approaches for dose-individualization. The models were used to simulate relevant drug exposures (Ctrough or AUC) under varying degrees of IIV, IOV and RUV. Results: Six drug cases were selected from the literature. Model-based simulations demonstrated that the percentage of patients for whom pharmacogenetic exposure predictions is superior to TDM differs for each drug case: tacrolimus (11.0%), tamoxifen (12.7%), efavirenz (49.2%), vincristine (49.6%), risperidone (48.1%) and 5-FU (100%). Generally, in the presence of higher unexplained IIV in combination with lower RUV and IOV, exposure was best estimated by TDM, whereas under lower unexplained IIV in combination with higher IOV or RUV, pharmacogenetic profiling was preferred. Conclusions: For the drugs with relatively low RUV and IOV (e.g., tamoxifen and tacrolimus), TDM estimated true exposure the best. Conversely, for drugs with similar or lower unexplained IIV (e.g., efavirenz or 5-FU, respectively) combined with relatively high RUV, pharmacogenetic profiling provided the most accurate estimate for most patients. However, genotype prevalence and the relative influence of genotypes on the PK, as well as the ability of TDM to accurately estimate AUC with a limited number of samples, had an impact. The results could be use

Artificial intelligence is revolutionizing protein structure prediction, providing unprecedented opportunities for drug design. To assess the potential impact on ligand discovery, we compared virtual screens using protein structures generated by the AlphaFold machine learning method and traditional homology modeling. More than 16 million compounds were docked to models of the trace amine-associated receptor 1 (TAAR1), a G protein-coupled receptor of unknown structure and target for treating neuropsychiatric disorders. Sets of 30 and 32 highly ranked compounds from the AlphaFold and homology model screens, respectively, were experimentally evaluated. Of these, 25 were TAAR1 agonists with potencies ranging from 12 to 0.03 mu M. The AlphaFold screen yielded a more than twofold higher hit rate (60%) than the homology model and discovered the most potent agonists. A TAAR1 agonist with a promising selectivity profile and drug-like properties showed physiological and antipsychotic-like effects in wild-type but not in TAAR1 knockout mice. These results demonstrate that AlphaFold structures can accelerate drug discovery.

Purpose Model-based methods can predict pediatric exposure and support initial dose selection. The aim of this study was to evaluate the performance of allometric scaling of population pharmacokinetic (popPK) versus physiologically based pharmacokinetic (PBPK) models in predicting the exposure of tyrosine kinase inhibitors (TKIs) for pediatric patients (>= 2 years), based on adult data. The drugs imatinib, sunitinib and pazopanib were selected as case studies due to their complex PK profiles including high inter-patient variability, active metabolites, time-varying clearances and non-linear absorption.Methods Pediatric concentration measurements and adult popPK models were derived from the literature. Adult PBPK models were generated in PK-Sim (R) using available physicochemical properties, calibrated to adult data when needed. PBPK and popPK models for the pediatric populations were translated from the models for adults and were used to simulate concentration-time profiles that were compared to the observed values.Results Ten pediatric datasets were collected from the literature. While both types of models captured the concentration-time profiles of imatinib, its active metabolite, sunitinib and pazopanib, the PBPK models underestimated sunitinib metabolite concentrations. In contrast, allometrically scaled popPK simulations accurately predicted all concentration-time profiles. Trough concentration (Ctrough) predictions from the popPK model fell within a 2-fold range for all compounds, while 3 out of 5 PBPK predictions exceeded this range for the imatinib and sunitinib metabolite concentrations.Conclusion Based on the identified case studies it appears that allometric scaling of popPK models is better suited to predict exposure of TKIs in pediatric patients >= 2 years. This advantage may be attributed to the stable enzyme expression patterns from 2 years old onwards, which can be easily related to adult levels through allometric scaling. In some instances, bo

Per- and polyfluoroalkyl substances (PFAS) are endocrine disruptors with unambiguous neurotoxic effects. However, due to variability in experimental models, population characteristics, and molecular endpoints, the elucidation of mechanisms underlying PFAS-induced neurotoxicity remains incomplete. In this review, we utilized the adverse outcome pathway (AOP) framework, a comprehensive tool for evaluating toxicity across multiple biological levels (molecular, cellular, tissue and organ, individual, and population), to elucidate the mechanisms of neurotoxicity induced by PFAS. Based on 271 studies, the reactive oxygen species (ROS) generation emerged as the molecular initiating event 1 (MIE1). Subsequent key events (KEs) at the cellular level include oxidative stress, neuroinflammation, apoptosis, altered Ca2+ signal transduction, glutamate and dopamine signaling dyshomeostasis, and reduction of cholinergic and serotonin. These KEs culminate in synaptic dysfunction at organ and tissue levels. Further insights were offered into MIE2 and upstream KEs associated with altered thyroid hormone levels, contributing to synaptic dysfunction and hypomyelination at the organ and tissue levels. The inhibition of Na+/I− symporter (NIS) was identified as the MIE2, initiating a cascade of KEs at the cellular level, including altered thyroid hormone synthesis, thyroid hormone transporters, thyroid hormone metabolism, and binding with thyroid hormone receptors. All KEs ultimately result in adverse outcomes (AOs), including cognition and memory impairment, autism spectrum disorders, attention deficit hyperactivity disorders, and neuromotor development impairment. To our knowledge, this review represents the first comprehensive and systematic AOP analysis delineating the intricate mechanisms responsible for PFAS-induced neurotoxic effects, providing valuable insights for risk assessments and mitigation strategies against PFAS-related health hazards.

The circadian rhythm plays a crucial role in regulating biological processes with a 24-hour period, including gene expression. However, in vitro systems often lack synchronization of cellular clocks, leading to inaccuracies in toxicity testing and failing to mimic human physiologically relevant conditions. To address this limitation, three experiments were conducted to investigate the synchronization of SH-SY5Y cells and the subsequent effects of neurotoxic agents. The first experiment aimed to identify the most effective entrainment factor for synchronizing the circadian rhythm in SH-SY5Y cells. Among the factors tested, dexamethasone was found to be the most effective, generating high-amplitude oscillations of BMAL1 and PER2 with a 24-hour periodicity. The second experiment evaluated the lethal effects of different concentrations of the pesticides cypermethrin and chlorpyrifos on the synchronized SH-SY5Y cells. The results demonstrated that the concentrations of these pesticides could lethally affect the cells, emphasizing the critical role of dose in toxicity assessment. The third experiment investigated the potential differences in toxicity from cypermethrin and chlorpyrifos when cells were exposed at different circadian time points. The findings indicate that the toxicity levels of these pesticides vary depending on the time of exposure relative to the circadian rhythm, suggesting that the timing of exposure is a crucial factor in assessing neurotoxic effects. Overall, this study underscores the importance of considering circadian timing in toxicity testing to better replicate human physiological conditions and improve the accuracy of in vitro models.

The superfamily of voltage-gated potassium (KV) channels is crucial for the normal function of several tissues and represents an attractive pharmacological target for treating disorders such as epilepsy and cardiac arrhythmias. However, any drug designed to target a KV channel must be capable of discriminating between different members within the superfamily, lest they plague the user with deleterious side effects. Such rational design requires structural and functional insight into how the selectivity of a molecule can be tailored to suit the intended target. This thesis combines the use of electrophysiological and computational techniques to investigate the molecular basis for how the function of hKV7 and hERG channels can be modulated by different lipophilic compounds with known or suspected effects on ion channels. These include polyunsaturated fatty acids (PUFAs), cannabidiol (CBD), and synthetic cannabinoid receptor agonists (SCRAs). Using the two-electrode voltage clamp technique on Xenopus oocytes, we find that both PUFAs and CBD modulate the function of hKV7 channels in subtype-specific manners. PUFAs facilitated the activation of hKV7 channels, except for hKV7.4 channels which were instead inhibited. Molecular dynamics simulations revealed that structural differences in the voltage-sensing domain of hKV7.4 conferred a unique, inhibitory PUFA interaction site absent in the other hKV7 subtypes. Once this site was neutralised by mutagenesis, PUFAs facilitated hKV7.4 activation. In the case of CBD, we observed three different responses: inhibition of channels with hKV7.1 subunits, potentiated voltage-sensitivity of channels with hKV7.2 or hKV7.3 subunits and enhanced maximum conductance of channels with hKV7.4 or hKV7.5 subunits. However, these responses were evoked from the same interaction site in the pore domain, indicating a more complex subtype-specific mechanism of action. Finally, using an automated patch-clamp system we screened 36 different SCRAs, Updates:2024-10-02 The thesis was first published online. The online published version reflects the printed version. 2024-11-08 The thesis was updated with an errata list which is also downloadable from the DOI landing page. Before this date the PDF has been downloaded 94 times.

Fever elicited by bacterial lypopolyssacharide (LPS) is mediated by pro-inflammatory cytokines, which activate central mediators and regulate the hypothalamic temperature setpoint. This response is often accompanied by morphological changes involving the extracellular matrix, neurons and glial cells, with significant health impacts. The NK1 receptor is involved in the febrile response induced by LPS but its effects over the extracellular matrix in the context of neuroinflammation remain unknown. The present work aims to clarify the extracellular changes associated with NK1 signaling in LPS-induced fever. Male Wistar rats were exposed to LPS intraperitoneally. Experimental groups were pre-treated intracerebroventricularly with the NK1 selective inhibitor SR140333B or saline. Histological changes involving the brain extracellular matrix were evaluated using hematoxylin and eosin, Mason's trichrome, picrosirius, alcian blue, periodic acid Schiff's stains. The expression of matrix metalloproteinase 9 (MMP9) was studied using confocal microscopy. Fever was accompanied by edema, perivascular lymphoplamacytic and neutrophylic infiltration, spongiosis and MMP9 overexpression. SR140333B significantly reduced LPS-induced fever (p < 0.0001), MMP9 overexpression (p < 0.01) and associated histological changes. These results contribute to characterize cerebral extracellular matrix changes associated with LPS-induced fever. Overall, the present work supports a role for NK1 receptor in these neuroinflammatory changes, involving MMP9 overexpression, edema and leukocytic infiltration., Funding Agencies|Fundacao de Amparo a Pesquisa e ao Desenvolvimento Cientifico e Tecnologico do Maranhao (FAPEMA) [00431/12]; Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES/CAPES Sandwich Program) [007414/2014-05]; CAPES/PNPD; CAPES [001]; Research Center of the Portuguese Oncology Institute of Porto [PI86-CI-IPOP-66-2017]; FCT/MCTES (PIDDAC): LEPABE [UIDB/00511/2020, UIDP/00511/2020, LA/P/0045/2020]

Modern healthcare faces a significant challenge, namely that 25-70% of patients with common diseases do not benefit from standard treatments despite the availability of over 13,000 drugs registered in DrugBank. This discrepancy is likely due to these diseases' complex and heterogeneous molecular nature rather than a lack of therapeutic options. Emerging technologies have revealed the immense molecular complexity underlying common diseases. For instance, singlecell RNA sequencing (scRNA-seq) has demonstrated altered gene interactions in and across multiple cell types in numerous tissues. Furthermore, these technologies have revealed vast molecular differences between patients with the same diagnosis. There is a wide gap between this complexity and the current diagnostic and therapeutic approaches. Aim: To bring personalized medicine one step closer to the clinic; this thesis focuses on developing digital disease models that can capture the molecular biological complexity of disease in individual patients. We aim to harness these disease models to identify optimal treatments for each individual patient. Paper I: We started by exploring the usefulness of OMIC-based approaches for diagnostic and therapeutic predictions. Utilizing a single-cell RNA-sequenced mouse model of antigen-induced arthritis, we aimed to prioritize cell types and therapeutic targets. Initial pathway enrichment analyses did not yield relevant prioritization, prompting an investigation into network-based approaches. Multi-cellular disease models (MCDMs) for AIA and human rheumatoid arthritis were constructed, incorporating predicted cell type interactions. Centrality analysis indicated that these interactions could quantify a cell type’s relative importance in disease pathogenesis. We hypothesized that transcriptomic alterations in central cell types might reflect the MCDM, serving as potential diagnostic markers. An analysis of CD4+ T cells from patients with 13 different inflammatory diseases and

High relapse rate during abstinence is a defining characteristic of drug addiction. We previously found that opioid seeking progressively increases after voluntary abstinence induced by adverse consequences of oxycodone seeking (crossing an electric barrier). Functional MRI revealed that this effect is associated with changes in functional connectivity within medial orbitofrontal cortex (mOFC)- and dorsomedial striatum (DMS)-related circuits. Here, we used a pharmacological manipulation and fMRI to determine the causal role of mOFC and DMS in oxycodone seeking after electric barrier-induced abstinence. We trained rats to self-administer oxycodone (6 h/day, 14 days). Next, we induced voluntary abstinence by exposing them to an electric barrier for 2 weeks. We inactivated the mOFC and DMS with muscimol+baclofen (GABA(a) and GABA(b) receptor agonists) and then tested them for relapse to oxycodone seeking on abstinence days 1 or 15 without the electric barrier or oxycodone. Inactivation of DMS (p < 0.001) but not mOFC decreased oxycodone seeking before or after electric barrier-induced abstinence. Functional MRI data revealed that DMS inactivation decreased cerebral blood volume levels in DMS and several distant cortical and subcortical regions (corrected p < 0.05). Furthermore, functional connectivity of DMS with several frontal, sensorimotor, and auditory regions significantly increased after DMS inactivation (corrected p < 0.05). Finally, an exploratory analysis of an existing functional MRI dataset showed that DMS inactivation restored voluntary abstinence-induced longitudinal changes in DMS functional connectivity with these brain regions (p < 0.05). Results indicate a role of DMS and related brain circuits in oxycodone seeking after voluntary abstinence, suggesting potential targets for intervention., Funding Agencies|National Institutes of Health

White-faced sakis (Pithecia pithecia) are commonly considered as frugivores but are unusual among primates as they do not specialize on ripe fruits but rather include a high proportion of unripe fruits into their diet, even during seasons when ripe fruits are available. Using a two-bottle preference test of short duration we therefore assessed whether this dietary specialization affects the taste responsiveness and sour-taste tolerance of four adult white-faced sakis for five food-associated acids. We found taste preference thresholds of the sakis to be 1-10 mM for citric acid, 0.5-20 mM for ascorbic acid, 2-10 mM for malic acid, 0.1-1 mM for tannic acid, and 2-20 mM for acetic acid, respectively. When given the choice between a reference solution of 50 mM sucrose and mixtures containing varying concentrations of sucrose plus citric acid, the sakis displayed a high sour-taste tolerance and required only 100 mM of sucrose (when mixed with 10 mM citric acid) or 200 mM of sucrose (when mixed with 30 or 50 mM citric acid), respectively, to prefer the sweet-sour mixture over the purely sweet 50 mM sucrose reference solution. These results demonstrate that white-faced sakis have a well-developed taste sensitivity for food-associated acids which is not inferior to that of primates specializing on ripe fruits. Compared to other platyrrhine primates, the sakis displayed a markedly higher sour-taste tolerance. These results may therefore reflect an evolutionary adaptation to the dietary specialization of the white-faced sakis to sour-tasting unripe fruits.

Objectives: Linezolid treatment has a high risk of toxicity and adverse drug reactions (ADR) are frequent. Few studies have investigated risk factors of major ADRs separately, therefore, we aimed to evaluate major ADRs including peripheral neuropathy in relation to risk factors and drug concentration levels of linezolid in a high-resource setting for multidrug-resistant tuberculosis (MDR-TB). Methods: We conducted a retrospective cohort study including participants treated with a linezolid-containing MDR-TB regimen in Sweden 1992-2018. Data was collected from medical records. ADRs were classified according to Common Terminology Criteria for Adverse Events (version 5.0). Results: Of all participants (n = 132), 43.2% were female and the median age 28 y. The median linezolid treatment was 6.5 months (IQR 3.0-12.7) with a median daily dose of 9.6 mg/kg/d. Any ADR was seen in 58.3% (n = 77) of participants, with 35.6% having peripheral neuropathy (n = 47), 27.3% anaemia (n = 36), 22.0% leukopenia (n = 36) while 6.1% (n = 8) had optic neuritis. The median time for peripheral neuropathy was 3.6 months (IQR 2.1-5.9) and 8.3 months (6.2-10.7) for optic neuritis. A >2.0 mg/L trough concentration (n = 40) was associated with anaemia (P = 0.0038) and thrombocytopenia (P = 0.009) but not with peripheral neuropathy. In multivariable analysis, a dose >= 12 mg/kg/d was associated with time to peripheral neuropathy (HR 2.89, 95% CI 1.08-7.74, P = 0.035), anaemia (HR 6.62, 95% CI 2.22-19.8, P = 0.001) and leukopenia (HR 5.23, 95% CI 1.48-18.5, P = 0.010). Conclusions: Linezolid ADRs were frequent in a high-resource setting. Structured, regular follow-up for ADRs and adjusting dosing according to body weight followed-up by monitoring of drug concentrations early may reduce toxicity. (c) 2024 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ ), Funding Agencies|Swedish Heart-Lung Foundation [20190229, 20210407, 20220859, 20150237, 20190226]; Swedish research council [2016-02043, 2018-1256]; ALF in Region Ostergotland, Sweden [987589]

The biological response to nanomaterials exposure depends on their properties, route of exposure, or model organism. Titanium dioxide nanoparticles (TiO2 NPs) are among the most used nanomaterials; however, concerns related to oxidative stress and metabolic effects resulting from their ingestion are rising. Therefore, in the present work, we addressed the metabolic effects of citrate-coated 45 nm TiO2 NPs combining bioaccumulation, tissue ultrastructure, and proteomics approaches on gilthead seabream, Sparus aurata and Japanese carpet shell, Ruditapes philippinarum. Sparus aurata was exposed through artificially contaminated feeds, while R. philippinarum was exposed using TiO2 NPs-doped microalgae solutions. The accumulation of titanium and TiO2 NPs in fish liver is associated with alterations in hepatic tissue structure, and alteration to the expression of proteins related to lipid and fatty acid metabolism, lipid breakdown for energy, lipid transport, and homeostasis. While cellular structure alterations and the expression of proteins were less affected than in gilthead seabream, atypical gill cilia and microvilli and alterations in metabolic-related proteins were also observed in the bivalve. Overall, the effects of TiO2 NPs exposure through feeding appear to stem from various interactions with cells, involving alterations in key metabolic proteins, and changes in cell membranes, their structures, and organelles. The possible appearance of metabolic disorders and the environmental risks to aquatic organisms posed by TiO2 NPs deserve further study., Funding Agencies|Portuguese Foundation for the Sci-ence and Technology (FCT) [UIDB/04423/2020, UIDP/04423/2020]; Scientific Employment Stimulus Program [2023.06491, CEE-CIND/03767/2018, 2020.04021]; European Union - Interreg POCTEP [07-12-ACUINA-NO_1_E]

Agonists at mu opioid receptors relieve acute pain, however, their long-term use is limited by side effects, which may involve beta-arrestin2. Agonists biased against beta-arrestin2 recruitment may be advantageous. However, the classification of bias may be compromised by assays utilising overexpressed mu receptors which overestimate efficacy for G-protein activation. There is a need for re-evaluation with restricted receptor availability to determine accurate agonist efficacies. We depleted mu receptor availability in PathHunter CHO cells using the irreversible antagonist, beta-funaltrexamine (beta-FNA), and compared efficacies and apparent potencies of twelve agonists, including several previously reported as biased, in beta-arrestin2 recruitment and cAMP assays. With full receptor availability all agonists had partial efficacy for stimulating beta-arrestin2 recruitment relative to DAMGO, while only TRV130 and buprenorphine were partial agonists as inhibitors of cAMP accumulation. Limiting receptor availability by prior exposure to beta-FNA (100 nM) revealed morphine, oxycodone, PZM21, herkinorin, U47700, tianeptine and U47931e are also partial agonists in the cAMP assay. The efficacies of all agonists, except SR-17018, correlated between beta-arrestin2 recruitment and cAMP assays, with depleted receptor availability in the latter. Furthermore, naloxone and cyprodime exhibited non-competitive antagonism of SR-17018 in the beta-arrestin2 recruitment assay. Limited antagonism by naloxone was also non-competitive in the cAMP assay, while cyprodime was competitive. Furthermore, SR-17018 only negligibly diminished beta-arrestin2 recruitment stimulated by DAMGO (1 mu M), whereas fentanyl, morphine and TRV130 all exhibited the anticipated competitive inhibition. The data suggest that SR-17018 achieves bias against beta-arrestin2 recruitment through interactions with mu receptors outside the orthosteric agonist site. This article is part of the Special Issue on "Ligand Bi, Funding Agencies|NIAA (BJA/RCoA) [WKR0-2017-0066, WKR0-2019-0067]; Versus Arthritis Grant [MR/W002566/1]; UKRI

Voltage-gated ion channels are responsible for the electrical excitability of neurons and cardiomyocytes. Thus, they are obvious targets for pharmaceuticals aimed to modulate excitability. Compounds activating voltage-gated potassium (KV) channels are expected to reduce excitability. To search for new KV-channel activators, we performed a high-throughput screen of 10,000 compounds on a specially designed Shaker KV channel. Here, we report on a large family of channel-activating compounds with a carboxyl (COOH) group as the common motif. The most potent COOH activators are lipophilic (4 < LogP <7) and are suggested to bind at the interface between the lipid bilayer and the channel's positively charged voltage sensor. The negatively charged form of the COOH-group compounds is suggested to open the channel by electrostatically pulling the voltage sensor to an activated state. Several of the COOH-group compounds also activate the therapeutically important KV7.2/7.3 channel and can thus potentially be developed into antiseizure drugs. The COOH-group compounds identified in this study are suggested to act via the same site and mechanism of action as previously studied COOH-group compounds, such as polyunsaturated fatty acids and resin acids, but distinct from sites for several other types of potassium channel-activating compounds., Funding Agencies|Swedish Re-search Council [2020-01019]; Swedish Heart-Lung Foundation [20210596]; Swedish Brain Foundation [2022-0219]; County of Ostergotland

Drug use is prevalent in prisons with drugs associated with depressant effects found to be more prevalent than stimulants. Synthetic cathinones (SCats; often sold as "bath salts", "ecstasy", "molly", and "monkey dust") are the second largest category of new psychoactive substances (NPS) currently monitored by the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) and are commonly used as substitutes for regulated stimulants, such as amphetamine, cocaine, and MDMA. N,N-dimethylpentylone (also known as dimethylpentylone, dipentylone, and bk-DMBDP) was detected for the first time in the Scottish prisons in seven powder samples seized between January and July 2023. Samples were analyzed using gas chromatography-mass spectrometry (GC-MS), ultra-high performance liquid chromatography-quadrupole time of flight mass spectrometry (UPLC-QToF-MS), and nuclear magnetic resonance imaging (NMR). Dimethylpentylone was detected alongside other drugs in four samples, including the novel benzodiazepine desalkylgidazepam (bromonordiazepam) and the synthetic cannabinoid receptor agonists (SCRAs) MDMB-INACA and MDMB-4en-PINACA., Funding Agencies|Leverhulme Trust [01865]; Leverhulme Research Centre for Forensic Science; [RC-2015-011]