Toward a responsible future: recommendations for AI-enabled clinical decision support.

BACKGROUND: Integrating artificial intelligence (AI) in healthcare settings has the potential to benefit clinical decision-making. Addressing challenges such as ensuring trustworthiness, mitigating bias, and maintaining safety is paramount. The lack of established methodologies for pre- and post-deployment evaluation of AI tools regarding crucial attributes such as transparency, performance monitoring, and adverse event reporting makes this situation challenging. OBJECTIVES: This paper aims to make practical suggestions for creating methods, rules, and guidelines to ensure that the development, testing, supervision, and use of AI in clinical decision support (CDS) systems are done well and safely for patients. MATERIALS AND METHODS: In May 2023, the Division of Clinical Informatics at Beth Israel Deaconess Medical Center and the American Medical Informatics Association co-sponsored a working group on AI in healthcare. In August 2023, there were 4 webinars on AI topics and a 2-day workshop in September 2023 for consensus-building. The event included over 200 industry stakeholders, including clinicians, software developers, academics, ethicists, attorneys, government policy experts, scientists, and patients. The goal was to identify challenges associated with the trusted use of AI-enabled CDS in medical practice. Key issues were identified, and solutions were proposed through qualitative analysis and a 4-month iterative consensus process. RESULTS: Our work culminated in several key recommendations: (1) building safe and trustworthy systems; (2) developing validation, verification, and certification processes for AI-CDS systems; (3) providing a means of safety monitoring and reporting at the national level; and (4) ensuring that appropriate documentation and end-user training are provided. DISCUSSION: AI-enabled Clinical Decision Support (AI-CDS) systems promise to revolutionize healthcare decision-making, necessitating a comprehensive framework for their development, implementation, and regulation that emphasizes trustworthiness, transparency, and safety. This framework encompasses various aspects including model training, explainability, validation, certification, monitoring, and continuous evaluation, while also addressing challenges such as data privacy, fairness, and the need for regulatory oversight to ensure responsible integration of AI into clinical workflow. CONCLUSIONS: Achieving responsible AI-CDS systems requires a collective effort from many healthcare stakeholders. This involves implementing robust safety, monitoring, and transparency measures while fostering innovation. Future steps include testing and piloting proposed trust mechanisms, such as safety reporting protocols, and establishing best practice guidelines.

A Smart System for Remote Monitoring of Patients in Palliative Care (HumanITcare Platform): Mixed Methods Study.

BACKGROUND: Due to the complexities of advanced illnesses and their treatments, it can be difficult for patients in palliative care to maintain their quality of life. Telemedicine interventions in chronic disease management engage patients in their care, provide continuous follow-up by their health care providers, identify symptoms earlier, and allow a quick response to illness-related decline. OBJECTIVE: We aimed to detail and reflect on the design of an app and evaluate its feasibility to monitor the clinical situation of patients with advanced illnesses. METHODS: This study used a mixed methods design using qualitative methods to inform app development and design and quantitative methods for data collection and analysis of patient evaluations. Palliative care units in 2 Spanish university hospitals (Nuestra Señora de la Candelaria in Santa Cruz de Tenerife and University Hospital Complex of Ferrol in A Coruña) carried out a literature review, designed the study protocol, and obtained approval from the Ethics Committee from June to December 2020. In addition, focus group meetings were held, and the design and technical development of the app were elaborated on and subsequently presented in the participating palliative care units. From January to March 2021, the app was made public on the App Store and Play Store, and a pilot study with patients was carried out in April to September 2021. RESULTS: Six focus group meetings were held that included doctors, nurses, app developers, technology consultants, and sponsors. In addition, the technology consultants presented their results 3 times in the participating palliative care units to obtain feedback. After the app's final design, it was possible to publish it on the usual servers and begin its evaluation in patients (n=60, median age 72 years). Sixty percent (n=36) of the participants were women and 40% (n=24) were men. The most prevalent advanced pathology was cancer (n=46, 76%), followed by other diseases (n=7, 12%) and amyotrophic lateral sclerosis (n=5, 8%). Seventy percent (n=42) of the patients were already in follow-up prior to the start of the study, while 30% (n=18) were included at the start of their follow-up. The information in the app was collected and entered by relatives or caregivers in 60% (n=36) of the cases. The median follow-up was 52 (IQR 14-104) days. In all, 69% (n=41) had a follow-up >30 days (10 were deceased and 9 were missing data). The use of the different sections of the app ranged from 37% (n=22) for the glycemic record to 90% (n=54) for the constipation scale). Patients and caregivers were delighted with its ease of use and usefulness. CONCLUSIONS: Incorporating an intelligent remote patient monitoring system in clinical practice for patients in palliative care can improve access to health services and provide more information to professionals.

Cardio4Health Study, a Cardiac Telerehabilitation Pilot Program Aimed at Patients After an Ischemic Event: Cross-sectional Study.

BACKGROUND: Center-based cardiac rehabilitation programs (CRPs) reduce morbidity and mortality after an ischemic cardiac event; however, they are widely underused. Home-based CRP has emerged as an alternative to improve patient adherence; however, its safety and efficacy remain unclear, especially for older patients and female patients. OBJECTIVE: This study aimed to develop a holistic home-based CRP for patients with ischemic heart disease and evaluate its safety and impact on functional capacity, adherence to a healthy lifestyle, and quality of life. METHODS: The 8-week home-based CRP included patients of both sexes, with no age limit, who had overcome an acute myocardial infarction in the previous 3 months, had a left ventricular ejection fraction of ≥40%, and had access to a tablet or mobile device. The CRP was developed using a dedicated platform designed explicitly for this purpose and included 3 weekly exercise sessions combining tailored aerobic and strength training and 2 weekly educational session focused on lifestyle habits, therapeutic adherence, and patient empowerment. RESULTS: We initially included 62 patients, of whom 1 was excluded for presenting with ventricular arrhythmias during the initial stress test, 5 were excluded because of incompatibility, and 6 dropped out because of a technological barrier. Ultimately, 50 patients completed the program: 85% (42/50) were male, with a mean age of 58.9 (SD 10.3) years, a mean left ventricular ejection fraction of 52.1% (SD 6.72%), and 25 (50%) New York Heart Association functional class I and 25 (50%) New York Heart Association II-III. The CRP significantly improved functional capacity (+1.6 metabolic equivalent tasks), muscle strength (arm curl test +15.5% and sit-to-stand test +19.7%), weekly training volume (+803 metabolic equivalent tasks), adherence to the Mediterranean diet, emotional state (anxiety), and quality of life. No major complications occurred, and adherence was excellent (>80%) in both the exercise and educational sessions. In the subgroup analysis, CRP showed equivalent beneficial effects irrespective of sex and age. In addition, patient preferences for CRP approaches were equally distributed, with one-third (14/50, 29%) of the patients preferring a face-to-face CRP, one-third (17/50, 34%) preferring a telematic CRP, and one-third (18/50, 37%) preferring a hybrid approach. Regarding CRP duration, 63% (31/50) of the patients considered it adequate, whereas the remaining 37% (19/50) preferred a longer program. CONCLUSIONS: A holistic telematic CRP dedicated to patients after an ischemic cardiac event, irrespective of sex and age, is safe and, in our population, has achieved positive results in improving maximal aerobic capacity, weekly training volume, muscle strength, quality of life, compliance with diet, and anxiety symptoms. The preference for a center- or home-based CRP approach is diverse among the study population, emphasizing the need for a tailored CRP to improve adherence and completion rates.

Development of a web tool to calculate the cumulative dose of glucocorticoids.

INTRODUCTION: Glucocorticoids are associated with serious side effects related to dosing and time of use. Unfortunately, there is no standard method for determining glucocorticoid exposure, especially in patients undergoing long-term treatment. OBJECTIVE: The aim of this work was to create a free and easy-to-use web application to calculate, in a systematic way, the total cumulative dose of corticosteroids. METHODS: The total cumulative dose is calculated as the sum of all periods of treatment with different doses of corticosteroids, and is expressed as the equivalent dose of prednisone in mg. Glucocorticoid doses during periods in which the available information is missing or incomplete are estimated by systematic assumptions. RESULTS: A simulation exercise using standard patterns of steroid use in polymyalgia rheumatica, and giant cell arteritis showed that even when the period of no information reached 50% of the time, the accuracy of the calculator had a mean absolute percentage error (MAPE)<7%. CONCLUSION: This tool simplifies and standardizes the glucocorticoids cumulative dose calculation, thereby minimizing bias in the assessment of glucocorticoid cumulative dose.

Depressive symptom severity is a major risk factor for frailty in community-dwelling older adults with depression. A prospective study.

BACKGROUND: Depression and frailty are closely related, but the mechanisms by which depressed older adults are at an increased risk of becoming frail are still not well understood. AIM: To assess socioeconomic and depression-related risk factors for frailty in older adults with depression. METHODS: Observational and prospective cohort study, with 12-month follow-up, of nonfrail community-dwelling subjects aged ≥70 years old with depression. The main study factors were clinical characteristics of depression, including symptom severity (Hamilton Depression Rating Scale), accompanying anxiety and cognitive symptoms, pharmacological treatment, and social factors including educational level, income, housing conditions and living circumstances, and social network. Frailty status was established according to Fried criteria. RESULTS: We recruited and analysed 216 subjects (mean age 76.5 years; 74% women), 65 (30%) of whom were lost to follow-up. Annual incidence of frailty was 23.2 new cases/100 persons. Age, female gender, osteoarthritis, pain, number of medications, major depression, first-degree family history of depression, depressive symptom severity, low educational level, and low-income level were risk factors for frailty. The multivariate analysis showed that age (odds ratio [OR] = 1.16; 95% confidence interval [CI]: 1.04-1.29), visual analogue scale (VAS)-pain (OR = 1.25; 95% CI: 1.01-1.55), and severe or very severe depressive symptoms (OR = 37.36; 95% CI: 2.68-518.53) were significantly associated with incident frailty at 12 months of follow-up. CONCLUSIONS: Both clinical and social characteristics are risk factors for frailty, but severity of depressive symptoms had the highest independent effect on frailty in depressed aged subjects. Frailty requires a multidisciplinary approach that pays special attention to pain and depressed mood.

The Role of DNA Damage Response in Dysbiosis-Induced Colorectal Cancer.

The high incidence of colorectal cancer (CRC) in developed countries indicates a predominant role of the environment as a causative factor. Natural gut microbiota provides multiple benefits to humans. Dysbiosis is characterized by an unbalanced microbiota and causes intestinal damage and inflammation. The latter is a common denominator in many cancers including CRC. Indeed, in an inflammation scenario, cellular growth is promoted and immune cells release Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS), which cause DNA damage. Apart from that, many metabolites from the diet are converted into DNA damaging agents by microbiota and some bacteria deliver DNA damaging toxins in dysbiosis conditions as well. The interactions between diet, microbiota, inflammation, and CRC are not the result of a straightforward relationship, but rather a network of multifactorial interactions that deserve deep consideration, as their consequences are not yet fully elucidated. In this paper, we will review the influence of dysbiosis in the induction of DNA damage and CRC.

Acute arterial ischemia in COVID-19.

Since the coronavirus pandemic set in in Spain in March 2020, a noteworthy increase in the incidence of acute limb ischemia (ALI) has been observed. It has been recently discovered that SARS-CoV 2 may lead to ALI secondary to arterial thrombosis. Elevation of D-dimer (DD) in patients with coronavirus infection (COVID-19) indicates that a hypercoagulable state causes acute arterial thrombosis. A remarkably high DD elevation has been reported to be a poor prognosis factor in COVID-19. The ways in which SARS-CoV 2 results in arterial thrombosis may be multiple. On the other hand, surgical revascularization for ALI is associated with poor outcomes in COVID-19 patients, probably in relation to hypercoagulability. Here, we describe two ALI cases in patients who required urgent surgical treatment for limb salvage and were positive for the novel coronavirus infection (COVID 19).

Desde que a pandemia pelo novo coronavírus se estabeleceu na Espanha, em março de 2020, um aumento notável da incidência de isquemia aguda de membros foi observado. Recentemente, descobriu-se que o coronavírus 2 causador da síndrome respiratória aguda grave (SARS-CoV-2) pode ocasionar isquemia aguda de membros secundária à trombose arterial. A elevação do D-dímero em pacientes acometidos pela doença do novo coronavírus (COVID-19) indica o estado de hipercoagulabilidade como causa da trombose arterial aguda. Vale destacar que a alta elevação do D-dímero foi relatada como um fator de prognóstico reservado na COVID-19. Há diversas maneiras pelas quais o SARS-CoV-2 pode resultar em trombose arterial. Em pacientes com COVID-19, a revascularização cirúrgica para isquemia aguda de membros está associada a desfechos desfavoráveis, provavelmente relacionados a hipercoagulabilidade. Descrevemos dois casos de isquemia aguda de membros de pacientes que necessitaram de tratamento cirúrgico de urgência para salvamento de membro e que haviam testado positivo para COVID-19.

Biocompatible DNA/5-Fluorouracil-Gemini Surfactant-Functionalized Gold Nanoparticles as Promising Vectors in Lung Cancer Therapy.

The design and preparation of novel nanocarriers to transport cancer drugs for chemotherapy purposes is an important line of research in the medical field. A new 5-fluorouracil (5-Fu) transporter was designed based on the use of two new biocompatible gold nanosystems: (i) a gold nanoparticle precursor, Au@16-Ph-16, stabilized with the positively charged gemini surfactant 16-Ph-16, and (ii) the compacted nanocomplexes formed by the precursor and DNA/5-Fu complexes, Au@16-Ph-16/DNA-5-Fu. The physicochemical properties of the obtained nanosystems were studied by using UV-visible spectroscopy, TEM, dynamic light scattering, and zeta potential techniques. Method tuning also requires the use of circular dichroism, atomic force microscopy, and fluorescence spectroscopy techniques for the prior selection of the optimal relative Au@16-Ph-16 and DNA concentrations (R = CAu@16-Ph-16/CDNA), biopolymer compaction/decompaction, and 5-Fu release from the DNA/5-Fu complex. TEM experiments revealed the effective internalization of the both precursor and Au@16-Ph-16/DNA-5-Fu-compacted nanosystems into the cells. Moreover, cytotoxicity assays and internalization experiments using TEM and confocal microscopy showed that the new strategy for 5-Fu administration enhanced efficacy, biocompatibility and selectivity against lung cancer cells. The differential uptake among different formulations is discussed in terms of the physicochemical properties of the nanosystems.

Psychosocial factors associated with frailty in the community-dwelling aged population with depression. A cross-sectional study.

OBJECTIVE: There is a two-way relationship between frailty and depression, but the mechanisms by which one may influence the other are not well understood. The objective of this study was to evaluate the relationship between psychosocial factors and frailty in community-dwelling aged populations with depression. DESIGN: Observational cross-sectional study. SITE: 5 primary care centres. PARTICIPANTS: Community-dwelling subjects with depression aged ≥70 years. MAIN MEASUREMENTS: Frailty status was established according to Fried criteria, depression and depression severity were evaluated by DSM-IV criteria and the Hamilton Depression Rating Scale, respectively, and psychosocial factors were assessed using the Gijón Social-Familial Evaluation Scale and ad hoc questionnaires. RESULTS: Recruited were 338 subjects (mean age 77.2 years), 82% women and 36.1% rated as frail. A dose-response relationship was observed between depression severity and frailty risk. Widowhood was a risk factor for frailty, while a higher educational level, home internet, stairs in the home, and an active social life had a protective effect. A multivariate analysis showed that age, number of drugs, and depression severity were independent risk factors for frailty, while an active social life was a protective factor. The severity of depressive symptoms showed higher association with frailty than other clinical and socio-demographic characteristics. CONCLUSIONS: In depressed elderly subjects, frailty is associated with psychologiocal factors such as the intensity of depressive symptoms and with social factors such as education level, widowhood, loneliness, and limited social life. More research is required to better understand the modifiable psychological risk factors for frailty.

In Vitro Anticancer Activity and Mechanism of Action of an Aziridinyl Galactopyranoside.

We recently screened a series of new aziridines ß-D-galactopyranoside derivatives for selective anticancer activity and identified 2-methyl-2,3-[N-(4-methylbenzenesulfonyl)imino]propyl 2,3-di-O-benzyl-4,6-O-(S)-benzylidene-ß-D-galactopyranoside (AzGalp) as the most promising compound. In this article, we explore the possible mechanisms involved in the cytotoxicity of this aziridine and evaluate its selective anticancer activity using cancer cells and normal cells from a variety of tissues. Our data show that AzGalp induces DNA damage (comet assay). Cells deficient in the nucleotide excision repair (NER) pathway were hypersensitive to the cytotoxicity of this compound. These results suggest that AzGalp induces bulky DNA adducts, and that cancer cells lacking a functional NER pathway may be particularly vulnerable to the anticancer effects of this aziridine. Several experiments revealed that neither the generation of oxidative stress nor the inhibition of glycolysis played a significant role in the cytotoxicity of AzGalp. Combinations of AzGalp with oxaliplatin or 5-fluorouracil slightly improved the ability of both anticancer drugs to selectively kill cancer cells. AzGalp also showed selective cytotoxicity against a panel of malignant cells versus normal cells; the highest selectivity was observed for two acute promyelocytic leukemia cell lines. Additional preclinical studies are necessary to evaluate the anticancer potential of AzGalp.

Acute arterial ischemia in COVID-19 / Isquemia arterial aguda em pacientes com COVID-19

Abstract Since the coronavirus pandemic set in in Spain in March 2020, a noteworthy increase in the incidence of acute limb ischemia (ALI) has been observed. It has been recently discovered that SARS-CoV 2 may lead to ALI secondary to arterial thrombosis. Elevation of D-dimer (DD) in patients with coronavirus infection (COVID-19) indicates that a hypercoagulable state causes acute arterial thrombosis. A remarkably high DD elevation has been reported to be a poor prognosis factor in COVID-19. The ways in which SARS-CoV 2 results in arterial thrombosis may be multiple. On the other hand, surgical revascularization for ALI is associated with poor outcomes in COVID-19 patients, probably in relation to hypercoagulability. Here, we describe two ALI cases in patients who required urgent surgical treatment for limb salvage and were positive for the novel coronavirus infection (COVID 19).

Resumo Desde que a pandemia pelo novo coronavírus se estabeleceu na Espanha, em março de 2020, um aumento notável da incidência de isquemia aguda de membros foi observado. Recentemente, descobriu-se que o coronavírus 2 causador da síndrome respiratória aguda grave (SARS-CoV-2) pode ocasionar isquemia aguda de membros secundária à trombose arterial. A elevação do D-dímero em pacientes acometidos pela doença do novo coronavírus (COVID-19) indica o estado de hipercoagulabilidade como causa da trombose arterial aguda. Vale destacar que a alta elevação do D-dímero foi relatada como um fator de prognóstico reservado na COVID-19. Há diversas maneiras pelas quais o SARS-CoV-2 pode resultar em trombose arterial. Em pacientes com COVID-19, a revascularização cirúrgica para isquemia aguda de membros está associada a desfechos desfavoráveis, provavelmente relacionados a hipercoagulabilidade. Descrevemos dois casos de isquemia aguda de membros de pacientes que necessitaram de tratamento cirúrgico de urgência para salvamento de membro e que haviam testado positivo para COVID-19.

Remote Monitoring Telemedicine (REMOTE) Platform for Patients With Anxiety Symptoms and Alcohol Use Disorder: Protocol for a Case-Control Study.

BACKGROUND: Monitoring mental health outcomes has traditionally been based on heuristic decisions, often based on scarce, subjective evidence, making the clinical decisions made by professionals, as well as the monitoring of these diseases, subject to flaws. However, the digital phenotype, which refers to the analysis of data collected by measuring human behavior with mobile sensors and smart bracelets, is a promising tool for filling this gap in current clinical practice. OBJECTIVE: The objectives of this study are to develop the digital phenotyping of patients with alcohol use disorder and anxiety symptoms using data collected from a mobile device (ie, smartphone) and a wearable sensor (ie, Fitbit) and to analyze usability and patient satisfaction with the data collection service provided by the app. METHODS: We propose to conduct a study among a group of 60 participants split into two subgroups-experimental and control-of 30 participants each. The experimental group will be recruited by physicians from the Hospital Clínic de Barcelona, and the control group will be recruited on a volunteer basis through fliers and social media. All participants will go through pretraining to ensure technological capability and understanding of tasks, then each participant will download the HumanITcare app and will be given a wearable sensor (ie, Fitbit). Throughout the 4-month period, participants will be monitored on a range of factors, including sleep cycle, heart rate, movement patterns, and sociability. All data from the wearable sensors and the mobile devices will be saved and sent to the HumanITcare server. Participants will be asked to complete weekly questionnaires about anxiety, depression, and alcohol use disorder symptoms. Research assistants will ensure timely responses. The data from both sensors will then be compared to the questionnaire responses to determine how accurately the devices can predict the same symptoms. RESULTS: The recruitment phase was completed in November 2019 and all the data were collected by the end of December 2019. Data are being processed; this process is expected to be completed by October 2020. CONCLUSIONS: This study was created and conducted as a pilot study with the Hospital Clínic de Barcelona, with the purpose of exploring the feasibility of our approach. The study is focused on patients diagnosed with anxiety and alcohol use disorder, but participants were also monitored for depressive symptoms throughout the trial, although these were not part of the initial inclusion criteria. A limitation to our study was the exclusive use of Android smartphones over iOS devices; this could result in a potential selection bias, due to the accessibility and affordability of Android phones as opposed to iOS-based phones. Another limitation might be that reviews of usability and satisfaction could be confounded by factors such as age and familiarity. An additional function that we might add in future studies is the ability for patients to manage their own data. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/16964.

Challenges to rutile-based geoscientific tools: low-temperature polymorphic TiO2 transformations and corresponding reactive pathways.

Rutile, a common accessory mineral in a wide variety of rocks, is the most stable naturally occurring TiO2 polymorph. The relationship between its trace element composition and formation conditions has provided geoscientists with discriminant tools for fingerprinting geological processes, such as magmatic evolution and subduction zone metamorphism, alongside applications to the study of sediment provenance. In the present work, volcaniclastic rock samples belonging to Fara and Saiq Formations, outcropping in Jebel Akhdar mountains, Oman, are studied with Raman spectroscopy and Electron Microprobe (EMP) aiming: of (i) the identification of different naturally-occurring TiO2 polymorphs, (ii) the evaluation of their trace element contents in relation with hydrothermal alteration features, and (iii) the analysis of the mineral reactive pathways behind the observed textural relationships. Raman investigations demonstrated that interstitial, fine-grained TiO2 corresponds to anatase, whereas rutile occurs as isolated single grains. EMP determinations further revealed that an identified Nb-enrichment in anatase is coupled with a corresponding Nb-depletion in rutile. The combination of the obtained results with petrographic observations enabled unravelling the TiO2 reactive pathways affecting the studied samples. Thus, a coupled polymorphic dissolution-precipitation reaction assisting rutile-to-anatase conversion has been defined, together with the role of Nb in further stabilizing the structure of the lower temperature polymorph. Semi-quantitative thermometric considerations suggest that rutile substrates are likely of magmatic origin, whereas anatase formation is clearly associated with a lower temperature aqueous environment. The gathered results raise fundamental questions concerning the application of commonly used rutile-based geochemical and thermometric tools.

The Cockayne syndrome protein B is involved in the repair of 5-AZA-2'-deoxycytidine-induced DNA lesions.

The Cockayne Syndrome Protein B (CSB) plays an essential role in Transcription-Coupled Nucleotide Excision Repair (TC-NER) by recruiting repair proteins once transcription is blocked with a DNA lesion. In fact, CSB-deficient cells are unable to recover from transcription-blocking DNA lesions. 5-Aza-2'-deoxycytidine (5-azadC) is a nucleoside analogue that covalently traps DNA methyltransferases (DNMTs) onto DNA. This anticancer drug has a double mechanism of action: it reverts aberrant hypermethylation in tumour-suppressor genes, and it induces DNA damage. We have recently reported that Homologous Recombination and XRCC1/PARP play an important role in the repair of 5-azadC-induced DNA damage. However, the mechanisms involved in the repair of the DNMT adducts induced by azadC remain poorly understood. In this paper, we show for the first time the importance of CSB in the repair of azadC-induced DNA lesions. We propose a model in which CSB initiates a signalling pathway to repair transcription blocks induced by incorporated 5-azadC. Indeed, CSB-deficient cells treated with 5-azadC show a delay in the repair of trapped DNMT1, increased levels of DNA damage and reduced survival.

Enhanced Cytotoxic Activity of Mitochondrial Mechanical Effectors in Human Lung Carcinoma H520 Cells: Pharmaceutical Implications for Cancer Therapy.

Cancer cell mitochondria represent an attractive target for oncological treatment as they have unique hallmarks that differ from their healthy counterparts, as the presence of a stronger membrane potential that can be exploited to specifically accumulate cytotoxic cationic molecules. Here, we explore the selective cytotoxic effect of 10-N-nonyl acridine orange (NAO) on human lung carcinoma H520 cells and compare them with healthy human lung primary fibroblasts. NAO is a lipophilic and positively charged molecule that promotes mitochondrial membrane adhesion that eventually leads to apoptosis when incubated at high micromolar concentration. We found an enhanced cytotoxicity of NAO in H520 cancer cells. By means Fluorescence lifetime imaging microscopy (FLIM) we also confirmed the formation of H-dimeric aggregates originating from opposing adjacent membranes that interfere with the mitochondrial membrane structure. Based on our results, we suggest the mitochondrial membrane as a potential target in cancer therapy to mechanically control the cell proliferation of cancer cells.

Selective cytotoxic activity and DNA damage by an epoxyalkyl galactopyranoside.

Preclinical Research & Development Several clinically useful anticancer drugs selectively kill cancer cells by inducing DNA damage; the genomic instability and DNA repair defects of cancer cells make them more vulnerable than normal cells to the cytotoxicity of DNA-damaging agents. Because epoxide-containing compounds can induce DNA damage, we have used the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to evaluate the selective cytotoxicity of three epoxyalkyl galactopyranosides against A549 lung cancer cells and MRC-5 lung normal cells. Compound (2S,3S)-2,3-epoxydecyl 4,6-O-(S)-benzylidene-ß-d-galactopyranoside (EDBGP) showed the highest selective anticancer activity and was selected for mechanistic studies. After observing that EDBGP induced cellular DNA damage (comet assay), we found that cells deficient in nucleotide excision repair were hypersensitive to the cytotoxicity of this compound; this suggests that EDBGP may induce bulky DNA adducts. EDBGP did not inhibit glycolysis (glucose consumption and lactate production). Pretreatment of lung cancer cells with several antioxidants did not reduce the cytotoxicity of EDBGP, thereby indicating that reactive oxygen species do not participate in the anticancer activity of this compound. Finally, EDBGP was screened against a panel of cancer cells and normal cells from several tissues, including three genetically modified skin fibroblasts with increasing degree of malignancy. Our results suggest that epoxyalkyl galactopyranosides are promising lead compounds for the development of new anticancer agents.

Polymorphism genotyping based on loop-mediated isothermal amplification and smartphone detection.

The genotyping of a single-nucleotide polymorphism (SNP) is addressed through methods based on loop-mediated isothermal amplification (LAMP) combined with user-friendly optical read-outs to cover the current demand for point-of-care DNA biomarker detection. The modification of primer design and reaction composition improved the assay selectivity yielding allele-specific results and reducing false-positive frequency. Furthermore, the reduced cost, ease of use and effectiveness of colorimetric detection (solution and hybridisation chip formats) were availed for the image capture by a smartphone, reching high sensitivity. In order to evaluate their discriminating capacities, LAMP-based methods were applied to human samples to genotype a SNP biomarker (rs1954787) located in the GRIK4 gene and related to the treatment response to anti-depressants drugs. Sensitive (limit of detection: 100 genomic DNA copies), reproducible (< 15% error), fast (around 70 min) and low-cost assays were accomplished. Patient subgroups were correctly discriminated, agreeing with reference sequencing techniques. The achieved analytical performances using the developed amplification-detection principles confirmed the approach potential for point-of-care optical DNA testing.

Zebularine induces replication-dependent double-strand breaks which are preferentially repaired by homologous recombination.

Zebularine is a second-generation, highly stable hydrophilic inhibitor of DNA methylation with oral bioavailability that preferentially target cancer cells. It acts primarily as a trap for DNA methyl transferases (DNMTs) protein by forming covalent complexes between DNMT protein and zebularine-substrate DNA. It's well documented that replication-blocking DNA lesions can cause replication fork collapse and thereby to the formation of DNA double-strand breaks (DSB). DSB are dangerous lesions that can lead to potentially oncogenic genomic rearrangements or cell death. The two major pathways for repair of DSB are non-homologous end joining (NHEJ) and homologous recombination (HR). Recently, multiple functions for the HR machinery have been identified at arrested forks. Here we investigate in more detail the importance of the lesions induced by zebularine in terms of DNA damage and cytotoxicity as well as the role of HR in the repair of these lesions. When we examined the contribution of NHEJ and HR in the repair of DSB induced by zebularine we found that these breaks were preferentially repaired by HR. Also we show that the production of DSB is dependent on active replication. To test this, we determined chromosome damage by zebularine while transiently inhibiting DNA synthesis. Here we report that cells deficient in single-strand break (SSB) repair are hypersensitive to zebularine. We have observed more DSB induced by zebularine in XRCC1 deficient cells, likely to be the result of conversion of SSB into toxic DSB when encountered by a replication fork. Furthermore we demonstrate that HR is required for the repair of these breaks. Overall, our data suggest that zebularine induces replication-dependent DSB which are preferentially repaired by HR.

RENEB intercomparison exercises analyzing micronuclei (Cytokinesis-block Micronucleus Assay).

PURPOSE: In the framework of the 'Realizing the European Network of Biodosimetry' (RENEB) project, two intercomparison exercises were conducted to assess the suitability of an optimized version of the cytokinesis-block micronucleus assay, and to evaluate the capacity of a large laboratory network performing biodosimetry for radiation emergency triages. Twelve European institutions participated in the first exercise, and four non-RENEB labs were added in the second one. MATERIALS AND METHODS: Irradiated blood samples were shipped to participating labs, whose task was to culture these samples and provide a blind dose estimate. Micronucleus analysis was performed by automated, semi-automated and manual procedures. RESULTS: The dose estimates provided by network laboratories were in good agreement with true administered doses. The most accurate estimates were reported for low dose points (≤ 0.94 Gy). For higher dose points (≥ 2.7 Gy) a larger variation in estimates was observed, though in the second exercise the number of acceptable estimates increased satisfactorily. Higher accuracy was achieved with the semi-automated method. CONCLUSION: The results of the two exercises performed by our network demonstrate that the micronucleus assay is a useful tool for large-scale radiation emergencies, and can be successfully implemented within a large network of laboratories.

The PARP inhibitor Olaparib disrupts base excision repair of 5-aza-2'-deoxycytidine lesions.

Decitabine (5-aza-2'-deoxycytidine, 5-azadC) is used in the treatment of Myelodysplatic syndrome (MDS) and Acute Myeloid Leukemia (AML). Its mechanism of action is thought to involve reactivation of genes implicated in differentiation and transformation, as well as induction of DNA damage by trapping DNA methyltranferases (DNMT) to DNA. We demonstrate for the first time that base excision repair (BER) recognizes 5-azadC-induced lesions in DNA and mediates repair. We find that BER (XRCC1) deficient cells are sensitive to 5-azadC and display an increased amount of DNA single- and double-strand breaks. The XRCC1 protein co-localizes with DNMT1 foci after 5-azadC treatment, suggesting a novel and specific role of XRCC1 in the repair of trapped DNMT1. 5-azadC-induced DNMT foci persist in XRCC1 defective cells, demonstrating a role for XRCC1 in repair of 5-azadC-induced DNA lesions. Poly (ADP-ribose) polymerase (PARP) inhibition prevents XRCC1 relocation to DNA damage sites, disrupts XRCC1-DNMT1 co-localization and thereby efficient BER. In a panel of AML cell lines, combining 5-azadC and Olaparib cause synthetic lethality. These data suggest that PARP inhibitors can be used in combination with 5-azadC to improve treatment of MDS and AML.