Determinants of non-alcoholic fatty liver disease in young people: Maternal, neonatal, and adolescent factors.

AIM: To assess the impact of maternal, neonatal, and adolescent factors on the development of non-alcoholic fatty liver disease (NAFLD) in a cohort of 14- to 19-year-old adolescents. METHODS: This study is part of the Early Vascular Ageing in the YOUth study, a single-center cross-sectional study conducted in western Austria. Maternal and neonatal factors were extracted from the mother-child booklet, adolescent factors were evaluated by a face-to-face interview, physical examination, and fasting blood analyses. Liver fat content was assessed by controlled attenuation parameter (CAP) using signals acquired by FibroScan® (Echosense, Paris, France). The association of maternal, neonatal, and adolescent factors with CAP values was analyzed using linear regression models. RESULTS: In total, 595 adolescents (27.2% male) aged 17.0 ± 1.3 years were included. 4.9% (n = 29) showed manifest NAFLD with CAP values above the 90th percentile. Male sex (p < 0.001), adolescent triglyceride levels (p = 0.021), Homeostatic Model Assessment for Insulin Resistance index and BMI z-score (p < 0.001, each) showed a significant association with liver fat content in the multivariable analysis. Maternal pre-pregnancy BMI was associated with CAP values after adjustment for sex, age, and birth weight for gestational age (p < 0.001), but this association was predominantly mediated by adolescent BMI (indirect effect b = 1.18, 95% CI [0.69, 1.77]). CONCLUSION: Components of the metabolic syndrome were the most important predictors of adolescent liver fat content. Therefore, prevention of NAFLD should focus on lifestyle modification in childhood and adolescence.

Determinants of Diet Quality in Adolescents: Results from the Prospective Population-Based EVA-Tyrol and EVA4YOU Cohorts.

(1) Background: Unhealthy dietary behaviors are estimated to be one of the leading causes of death globally and are often shaped at a young age. Here, we investigated adolescent diet quality and its predictors, including nutrition knowledge, in two large Central European cohorts. (2) Methods: In 3056 participants of the EVA-Tyrol and EVA4YOU prospective population-based cohort studies aged 14 to 19 years, diet quality was assessed using the AHEI-2010 and DASH scores, and nutrition knowledge was assessed using the questionnaire from Turconi et al. Associations were examined utilizing multivariable linear regression. (3) Results: The mean overall AHEI-2010 score was 42%, and the DASH score was 45%. Female participants (60.6%) had a significantly higher diet quality according to the AHEI-2010 and DASH score. AHEI-2010 and DASH scores were significantly associated (p < 0.001) with sex, school type, smoking, and total daily energy intake. The DASH score was additionally significantly associated (p < 0.001) with age, socioeconomic status, and physical activity. Participants with better nutrition knowledge were more likely to be older, to attend a general high school, to live in a high-income household, to be non-smokers, and to have a higher diet quality according to the AHEI-2010 and DASH score. (4) Conclusions: Predictors of better diet quality included female sex, physical activity, educational level, and nutrition knowledge. These results may aid focused interventions to improve diet quality in adolescents.

Evaluation of two fully automated ADAMTS13 activity assays in comparison to manual FRET assay.

INTRODUCTION: The objective of the present study was to evaluate and compare the validity and utility of two fully automated ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) activity assays for clinical diagnostic decision-making and to compare their performance. METHODS: Two automated ADAMTS13 activity assays (Werfen HemosIL® AcuStar ADAMTS13 Activity, Technoclone Technofluor ADAMTS13 Activity) were compared with a manual FRET assay (BioMedica ACTIFLUOR ADAMTS13 Activity). The following samples were used: 13 acute phase TTP (thrombotic thrombocytopenic purpura) samples from 11 different patients, one sample from a patient with congenital ADAMTS13 deficiency, 16 samples from control patients, three follow-up samples from TTP patients in long-term remission and one sample from a patient with stem cell transplantation related thrombotic microangiopathy (TMA). The WHO 1st International Standard for ADAMTS13 and several dilutions of normal plasma with ADAMTS13-depleted normal plasma were also tested. Statistical analysis included descriptive statistics, sensitivity and specificity, Passing & Bablok regression and Bland-Altman plot. RESULTS: The quantitative comparison between the HemosIL® (x) and Technofluor (y) methods showed a strong correlation (Pearson r = 0.98, n = 49). When considering an ADAMTS13 activity of <10% as a hallmark for the diagnosis of TTP, two fully automated assays were both able to identify all TTP- and non-TTP-samples correctly, resulting in sensitivities and specificities of 100%. CONCLUSION: Both fully automated ADAMTS13 activity assays showed a good diagnostic performance and quantitative correlation among themselves, discriminating reliably between TTP- and non-TTP-patients.

One-Year Follow-Up of COVID-19 Patients Indicates Substantial Assay-Dependent Differences in the Kinetics of SARS-CoV-2 Antibodies.

Determination of antibody levels against the nucleocapsid (N) and spike (S) proteins of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are used to estimate the humoral immune response after SARS-CoV-2 infection or vaccination. Differences in the design and specification of antibody assays challenge the interpretation of test results, and comparative studies are often limited to single time points per patient. We determined the longitudinal kinetics of antibody levels of 145 unvaccinated coronavirus disease 2019 (COVID-19) patients at four visits over 1 year upon convalescence using 8 commercial SARS-CoV-2 antibody assays (from Abbott, DiaSorin, Roche, Siemens, and Technoclone), as well as a virus neutralization test (VNT). A linear regression model was used to investigate whether antibody results obtained in the first 6 months after disease onset could predict the VNT results at 12 months. Spike protein-specific antibody tests showed good correlation to the VNT at individual time points (rS, 0.74 to 0.92). While longitudinal assay comparison with the Roche Elecsys anti-SARS-CoV-2 S test showed almost constant antibody concentrations over 12 months, the VNT and all other tests indicated a decline in serum antibody levels (median decrease to 14% to 36% of baseline). The antibody level at 3 months was the best predictor of the VNT results at 12 months after disease onset. The current standardization to a WHO calibrator for normalization to binding antibody units (BAU) is not sufficient for the harmonization of SARS-CoV-2 antibody tests. Assay-specific differences in absolute values and trends over time need to be considered when interpreting the course of antibody levels in patients. IMPORTANCE Determination of antibodies against SARS-CoV-2 will play an important role in detecting a sufficient immune response. Although all the manufacturers expressed antibody levels in binding antibody units per milliliter, thus suggesting comparable results, we found discrepant behavior between the eight investigated assays when we followed the antibody levels in a cohort of 145 convalescent patients over 1 year. While one assay yielded constant antibody levels, the others showed decreasing antibody levels to a varying extent. Therefore, the comparability of the assays must be improved regarding the long-term kinetics of antibody levels. This is a prerequisite for establishing reliable antibody level cutoffs for sufficient individual protection against SARS-CoV-2.

Borderline and weakly positive antibody levels against the S-protein of SARS-CoV-2 exhibit limited agreement with virus neutralization titres.

The presence of neutralizing antibodies against SARS-CoV-2 in a large number of people is - besides cellular immunity - important to overcome the SARS-CoV-2 pandemic. While determination of neutralizing antibodies via virus neutralization tests are laborious, assays to determine the antibody levels serologically are fully automated and widely available. Correlations between these methodologies were recently given by the manufacturers, however performance in samples close to the cut off value have not yet been fully validated. Thus, we analysed 22 borderline and low positive (<100 BAU/ml) samples and 9 high positive (≥ 100 BAU/ml) from infected and/or vaccinated individuals and compared the SARS-CoV-2 IgG II Quant assay (Abbott), LIAISON SARS-CoV-2 TrimericS IgG (Diasorin), Elecsys Anti-SARS-CoV-2 S (Roche), and SARS-CoV-2 IgG (Siemens) with results obtained from a virus neutralization test. Based on the cut off values given by Abbott, Diasorin, Roche, and Siemens, the positive serologic results were concordant with the virus neutralization test in 100%, 76%, 88%, and 71%, respectively, while in turn, negative ones were in agreement in 29%, 79%, 93%, and 86%, respectively. In conclusion, weakly positive, serologic results are challenging to correctly predict the presence of neutralizing antibodies. Our study suggests, that different cut off values (for positivity vs. presence of neutralizing antibodies) could improve the test's performance, but determination thereof requires more samples to be analysed.

Diagnostic Yield of a Systematic Vascular Health Screening Approach in Adolescents at Schools.

PURPOSE: A significant proportion of noncommunicable diseases in adults has its roots in adolescence, and this is particularly true for cardiovascular disease and stroke. Detection of vascular and metabolic risk factors at young ages may aid disease prevention. METHODS: In 2,088 adolescents sampled from the general population of Tyrol, Austria, and South Tyrol, Italy, we systematically assessed the frequency of yet unknown vascular and metabolic risk conditions that require further diagnostic workup or intervention (lifestyle counselling or pharmacotherapy). The health screening included medical history taking, fasting blood analysis, and blood pressure and body measurements and was performed at schools. To recruit a representative sample of adolescents, equal proportions (about 67%) of schools were invited per school type and region. RESULTS: Adolescents were on average 16.4 (standard deviation 1.1) years old, and 56.4% were female. A proportion of 22.8% (95% confidence interval [CI], 19.6-26.3) had previous or current physician-confirmed diseases. The health screening newly detected relevant medical conditions in 45.4% [95% CI, 41.5-49.4] (55.8% [95% CI, 52.7-58.7] in boys and 37.4% [95% CI, 35.0-39.8] in girls, p < .001). The most prevalent were elevated blood pressure and hypertension, metabolic syndrome, hypercholesterolemia, hypertriglyceridemia, hyperuricemia, and subclinical hypothyroidism. Detection of risk conditions did not depend on socioeconomic status but increased with age and body mass index. CONCLUSIONS: Vascular health screening in adolescents at schools has a high diagnostic yield and may aid guideline-recommended prevention in the youth. Implementation should carefully consider national differences in healthcare systems, resources, and existing programs.

A comprehensive antigen production and characterisation study for easy-to-implement, specific and quantitative SARS-CoV-2 serotests.

BACKGROUND: Antibody tests are essential tools to investigate humoral immunity following SARS-CoV-2 infection or vaccination. While first-generation antibody tests have primarily provided qualitative results, accurate seroprevalence studies and tracking of antibody levels over time require highly specific, sensitive and quantitative test setups. METHODS: We have developed two quantitative, easy-to-implement SARS-CoV-2 antibody tests, based on the spike receptor binding domain and the nucleocapsid protein. Comprehensive evaluation of antigens from several biotechnological platforms enabled the identification of superior antigen designs for reliable serodiagnostic. Cut-off modelling based on unprecedented large and heterogeneous multicentric validation cohorts allowed us to define optimal thresholds for the tests' broad applications in different aspects of clinical use, such as seroprevalence studies and convalescent plasma donor qualification. FINDINGS: Both developed serotests individually performed similarly-well as fully-automated CE-marked test systems. Our described sensitivity-improved orthogonal test approach assures highest specificity (99.8%); thereby enabling robust serodiagnosis in low-prevalence settings with simple test formats. The inclusion of a calibrator permits accurate quantitative monitoring of antibody concentrations in samples collected at different time points during the acute and convalescent phase of COVID-19 and disclosed antibody level thresholds that correlate well with robust neutralization of authentic SARS-CoV-2 virus. INTERPRETATION: We demonstrate that antigen source and purity strongly impact serotest performance. Comprehensive biotechnology-assisted selection of antigens and in-depth characterisation of the assays allowed us to overcome limitations of simple ELISA-based antibody test formats based on chromometric reporters, to yield comparable assay performance as fully-automated platforms. FUNDING: WWTF, Project No. COV20-016; BOKU, LBI/LBG.

Clinical validation of the Siemens quantitative SARS-CoV-2 spike IgG assay (sCOVG) reveals improved sensitivity and a good correlation with virus neutralization titers.

OBJECTIVES: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections cause coronavirus disease 2019 (COVID-19) and induce a specific antibody response. Serological assays detecting IgG against the receptor binding domain (RBD) of the spike (S) protein are useful to monitor the immune response after infection or vaccination. The objective of our study was to evaluate the clinical performance of the Siemens SARS-CoV-2 IgG (sCOVG) assay. METHODS: Sensitivity and specificity of the Siemens sCOVG test were evaluated on 178 patients with SARS-CoV-2-infection and 160 pre-pandemic samples in comparison with its predecessor test COV2G. Furthermore, correlation with virus neutralization titers was investigated on 134 samples of convalescent COVID-19 patients. RESULTS: Specificity of the sCOVG test was 99.4% and sensitivity was 90.5% (COV2G assay 78.7%; p<0.0001). S1-RBD antibody levels showed a good correlation with virus neutralization titers (r=0.843; p<0.0001) and an overall qualitative agreement of 98.5%. Finally, median S1-RBD IgG levels increase with age and were significantly higher in hospitalized COVID-19 patients (median levels general ward: 25.7 U/mL; intensive care: 59.5 U/mL) than in outpatients (3.8 U/mL; p<0.0001). CONCLUSIONS: Performance characteristics of the sCOVG assay have been improved compared to the predecessor test COV2G. Quantitative SARS-CoV-2 S1-RBD IgG levels could be used as a surrogate for virus neutralization capacity. Further harmonization of antibody quantification might assist to monitor the humoral immune response after COVID-19 disease or vaccination.

Evaluation of four commercial, fully automated SARS-CoV-2 antibody tests suggests a revision of the Siemens SARS-CoV-2 IgG assay.

OBJECTIVES: Serological tests detect antibodies against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in the ongoing coronavirus disease-19 (COVID-19) pandemic. Independent external clinical validation of performance characteristics is of paramount importance. METHODS: Four fully automated assays, Roche Elecsys Anti-SARS-CoV-2, Abbott SARS-CoV-2 IgG, Siemens SARS-CoV-2 total (COV2T) and SARS-CoV-2 IgG (COV2G) were evaluated using 350 pre-pandemic samples and 700 samples from 245 COVID-19 patients (158 hospitalized, 87 outpatients). RESULTS: All tests showed very high diagnostic specificity. Sensitivities in samples collected at least 14 days after disease onset were slightly lower than manufacturers' claims for Roche (93.0%), Abbott (90.8%), and Siemens COV2T (90.3%), and distinctly lower for Siemens COV2G (78.8%). Concordantly negative results were enriched for immunocompromised patients. ROC curve analyses suggest a lowering of the cut-off index for the Siemens COV2G assay. Finally, the combination of two anti-SARS-CoV-2 antibody assays is feasible when considering borderline reactive results. CONCLUSIONS: Thorough on-site evaluation of commercially available serologic tests for detection of antibodies against SARS-CoV-2 remains imperative for laboratories. The potentially impaired sensitivity of the Siemens COV2G necessitates a switch to the company's newly filed SARS-CoV-2 IgG assay for follow-up studies. A combination of tests could be considered in clinical practice.

Chemical imaging and assessment of cadmium distribution in the human body.

The scientific interest in cadmium (Cd) as a human health damaging agent has significantly increased over the past decades. However, particularly the histological distribution of Cd in human tissues is still scarcely defined. Using inductively coupled plasma-mass spectrometry (ICP-MS), we determined the concentration of Cd in 40 different human tissues of four body donors and provided spatial information by elemental imaging on the microscopic distribution of Cd in 8 selected tissues by laser ablation (LA)-ICP-MS. ICP-MS results show that Cd concentrations differ by a factor of 20 000 between different tissues. Apart from the well know deposits in kidney, bone, and liver, our study provides evidence that muscle and adipose tissue are underestimated Cd pools. For the first time, we present spatially resolved Cd distributions in a broad panel of human soft tissues. The defined histological structures are mirrored by sharp cut differences in Cd concentrations between neighboring tissue types, particularly in the rectum, testis, and kidneys. The spatial resolution of the Cd distribution at microscopic level visualized intratissue hot spots of Cd accumulation and is suggested as a powerful tool to elucidate metal based toxicity at histological level.

Tumor microenvironment in focus: LA-ICP-MS bioimaging of a preclinical tumor model upon treatment with platinum(IV)-based anticancer agents.

The selection of drug candidates for entering clinical development relies on in vivo testing in (solid) tumor animal models. However, the heterogeneity of tumor tissue (e.g. in terms of drug uptake or tissue composition) is rarely considered when testing novel drug candidates. Therefore, we used the murine colon cancer CT-26 tumor model to study the spatially-resolved drug distribution in tumor tissue upon repetitive treatment of animals over two weeks with three investigational platinum(IV)-based anticancer agents, oxaliplatin or satraplatin. A quantitative laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) imaging method revealed a heterogeneous platinum distribution, which correlated well with the histologic features of the tumor and surrounding tissue at the microscopic level. In most of the cases, higher amounts of intratumoral platinum were found in the surrounding tissue than in the malignant parts of the sample. This indicates that determination of average platinum amounts (e.g. by microwave-assisted digestion of the sample followed by analysis with ICP-MS) might overestimate the drug uptake in tumor tissue causing misleading conclusions. In addition, we studied the platinum distribution in the kidneys of treated animals to probe if accumulation in the cortex and medulla predict potential nephrotoxicity. A 10-fold increase of platinum in the cortex of the kidney over the medulla was observed for oxaliplatin and satraplatin. Although these findings are similar to those in the platinum distribution of the nephrotoxic anticancer drug cisplatin, treatment with the compounds of our study did not show signs of nephrotoxicity in clinical use or clinical trials (oxaliplatin, satraplatin) and did not result in the alteration of renal structures. Thus, predicting the side effects based on bioimaging data by LA-ICP-MS should be considered with caution. To the best of our knowledge, this is the first LA-ICP-MS study on spatially-resolved platinum accumulation in tissues after repetitive platinum-based anticancer drug treatment of mice bearing a preclinical tumor model.

Extravasation of Pt-based chemotherapeutics - bioimaging of their distribution in resectates using laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS).

Platinum-based drugs (cisplatin, carboplatin and oxaliplatin) are widely used in cancer treatment. They are administered intravenously, thus accidental extravasations of infusions can occur. This may cause severe complications for the patient as the toxic platinum compounds likely persist in subcutaneous tissue. At high concentrations, platinum toxicity in combination with local thrombosis may result in tissue necrosis, eventually requiring surgical intervention. To describe tissue distribution at the anatomic level, we quantified drug extravasation in cryosections of various tissues (muscle, nerve tissue, connective tissue, fat tissue) by means of quantitative laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) and compared the resulting data with bulk analysis of microwave-assisted digestion of tissue samples followed by ICP-MS analysis. Samples of three patients receiving systemic chemotherapy either via peripheral venous access or central access via port-a-cath® were analyzed. Pt was enriched up to 50-times in connective tissue when compared with muscle tissue or drain samples collected over five days. The large areas of subcutaneous fat tissue showed areactive necrosis and average Pt concentrations (determined upon sample digestion) ranged from 0.2 µg g(-1) (therapy with 25 mg m(-2) cisplatin, four weeks after peripheral extravasation) to 10 µg g(-1) (therapy with 50 mg m(-2) oxaliplatin: four weeks after port-a-cath® extravasation). A peripheral nerve subjected to bioimaging by LA-ICP-MS showed a 5-times lower Pt concentration (0.2 µg g(-1)) than the surrounding connective tissue (1.0 µg g(-1)). This is in accordance with the patient showing no signs of neurotoxicity during recovery from extravasation side-effects. Thus, bioimaging of cutaneous nerve tissue may contribute to understand the risk of peripheral neurotoxic events.

Comparative in vitro and in vivo pharmacological investigation of platinum(IV) complexes as novel anticancer drug candidates for oral application.

Platinum(IV) complexes are promising candidates as prodrugs for oral application in anticancer chemotherapy. However, only a few Pt(IV) compounds entered (pre)clinical trials, e.g. satraplatin, while most of the others were only tested in vitro. Aim of the study was investigation of the in vivo pharmacological behavior as well as the anticancer activity of two novel platinum(IV) complexes vs. satraplatin. The drugs were selected due to significantly different in vitro cytotoxicity while sharing some physicochemical properties (e.g. lipophilicity). Initial experiments indicated that the highly in vitro cytotoxic compound 1 ((OC-6-33)-dichloridobis((4-ethoxy)-4-oxobutanoato)-bis(ethylamine)platinum(IV)) was also characterized by high drug absorption and tissue platinum levels after oral application. Interestingly, analysis of serum samples using SEC-ICP-MS revealed that the administered drugs have completely been metabolized and/or bound to proteins in serum within 2 h after treatment. With regard to the activity in vivo, the outcomes were rather unexpected: although potent anticancer effect of 1 was observed in cell culture, the effects in vivo were rather minor. Nevertheless, 1 was superior to 2 ((OC-6-33)-diammine(cyclobutane-1,1-dicarboxylato)-bis((4-cyclopentylamino)-4-oxobutanoato)platinum(IV)) after i.p. administration, which was, at least to some extent, in accordance to the cell culture experiments. After oral gavage, both compounds exhibited comparable activity. This is remarkable considering the distinctly lower activity of 2 in cell culture as well as the low platinum levels detected both in serum and tissues after oral application. Consequently, our data indicate that the prediction of in vivo anticancer activity by cell culture experiments is not trivial, especially for orally applied drugs.

Quantitative bioimaging by LA-ICP-MS: a methodological study on the distribution of Pt and Ru in viscera originating from cisplatin- and KP1339-treated mice.

Laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) was used to study the spatially-resolved distribution of ruthenium and platinum in viscera (liver, kidney, spleen, and muscle) originating from mice treated with the investigational ruthenium-based antitumor compound KP1339 or cisplatin, a potent, but nephrotoxic clinically-approved platinum-based anticancer drug. Method development was based on homogenized Ru- and Pt-containing samples (22.0 and 0.257 µg g(-1), respectively). Averaging yielded satisfactory precision and accuracy for both concentrations (3-15% and 93-120%, respectively), however when considering only single data points, the highly concentrated Ru sample maintained satisfactory precision and accuracy, while the low concentrated Pt sample yielded low recoveries and precision, which could not be improved by use of internal standards ((115)In, (185)Re or (13)C). Matrix-matched standards were used for quantification in LA-ICP-MS which yielded comparable metal distributions, i.e., enrichment in the cortex of the kidney in comparison with the medulla, a homogenous distribution in the liver and the muscle and areas of enrichment in the spleen. Elemental distributions were assigned to histological structures exceeding 100 µm in size. The accuracy of a quantitative LA-ICP-MS imaging experiment was validated by an independent method using microwave-assisted digestion (MW) followed by direct infusion ICP-MS analysis.

Metal-Arene Complexes with Indolo[3,2-c]-quinolines: Effects of Ruthenium vs Osmium and Modifications of the Lactam Unit on Intermolecular Interactions, Anticancer Activity, Cell Cycle, and Cellular Accumulation.

Six novel ruthenium(II)- and osmium(II)-arene complexes with three modified indolo[3,2-c]quinolines have been synthesized in situ starting from 2-aminoindoloquinolines and 2-pyridinecarboxaldehyde in the presence of [M(p-cymene)Cl(2)](2) (M = Ru, Os) in ethanol. All complexes have been characterized by elemental analysis, spectroscopic techniques ((1)H, (13)C NMR, IR, UV-vis), and ESI mass spectrometry, while four complexes were investigated by X-ray diffraction. The complexes have been tested for antiproliferative activity in vitro in A549 (non-small cell lung), SW480 (colon), and CH1 (ovarian) human cancer cell lines and showed IC(50) values between 1.3 and >80 µM. The effects of Ru vs Os and modifications of the lactam unit on intermolecular interactions, antiproliferative activity, and cell cycle are reported. One ruthenium complex and its osmium analogue have been studied for anticancer activity in vivo applied both intraperitoneally and orally against the murine colon carcinoma model CT-26. Interestingly, the osmium(II) complex displayed significant growth-inhibitory activity in contrast to its ruthenium counterpart, providing stimuli for further investigation of this class of compounds as potential antitumor drugs.

Influence of extracellular pH on the cytotoxicity, cellular accumulation, and DNA interaction of novel pH-sensitive 2-aminoalcoholatoplatinum(II) complexes.

Extracellular acidity is a frequent pathophysiological condition of solid tumors offering possibilities for improving the tumor selectivity of molecular therapy. This might be accomplished by prodrugs with low systemic toxicity, attaining their full antitumor potency only under acidic conditions, such as bis(2-aminoalcoholato-κ(2)N,O)platinum(II) complexes that are activated by protonation of alcoholato oxygen, resulting in cleavage of platinum-oxygen bonds. In this work, we examined whether the pH dependency of such compounds is reflected in differential biological activity in vitro. In particular, the pH dependence of cytotoxicity, cellular accumulation, DNA platination, GMP binding, effects on DNA secondary structure, cell cycle alterations, and induction of apoptosis was investigated. Enhanced cytotoxicity of five of these complexes in non-small-cell lung cancer (A549) and colon carcinoma (HT-29) cells at pH 6.0 in comparison with pH 7.4 was confirmed: 50 % growth inhibition concentrations ranged from 42 to 214 µM in A549 cells and from 35 to 87 µM in HT-29 cells at pH 7.4 and decreased at pH 6.0 to 11-50 and 7.3-25 µM, respectively. The effects induced by all five pH-sensitive compounds involve increased 5'-GMP binding, cellular accumulation, and DNA platination as well as stronger effects on DNA secondary structure at pH 6.0 than at pH 7.4. As exemplified by treatment of A549 cells with a 2-amino-4-methyl-1-pentanolato complex, induction of apoptosis is enhanced at pH 6.5. These results confirm the increased reactivity and in vitro activity of these compounds under slightly acidic conditions, encouraging further evaluation of ring-closed aminoalcoholatoplatinum(II) derivatives in solid tumors in vivo.

Nanoscalic silver possesses broad-spectrum antimicrobial activities and exhibits fewer toxicological side effects than silver sulfadiazine.

Silver has been used successfully for decades as an antibacterial agent and has become a standard treatment for burns and bacterial skin infections. Silver-containing creams, particularly silver sulfadiazine (SSD), possess effective activities against bacteria and fungi. However, there is serious concern that silver ions applied to denuded skin might be absorbed in significant amounts, thus introducing the risk of silver deposition, potentially leading to internal organ injury. In view of these facts we compared the percutaneous absorption and the antimicrobial potency of SSD with a new composition, nanoscalic silver (NSAg). In a murine model topical application of NSAg resulted in significantly lower percutaneous absorption and internal organ deposition compared to SSD. Strikingly, antimicrobial activity of NSAg used as a 0.1% formulation was comparable not only with 0.1% SSD against different bacterial strains including methicillin-resistant Staphylococcus aureus, but also against different yeast and dermatophyte species. FROM THE CLINICAL EDITOR: Nanoscale silver (NSAg) was demonstrated to have significantly lower percutaneous absorption and less accumulation in multiple organs when applied to denuded skin. Its antimicrobial activity against MRSA was not only comparable to silver sulfadiazine, but the formulation was also effective against different yeast and dermatophyte species.

Influence of ascorbic acid on the activity of the investigational anticancer drug KP1019.

Ascorbic acid has been previously discussed to have antitumor potential through its interaction with transition metal ions such as iron and copper. Furthermore, ascorbic acid may act as a reducing agent for Ru(III) compounds such as indazolium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] (KP1019), an investigational anticancer drug which is supposed to be activated by reduction, prior to binding to cellular target proteins. Therefore, we investigated the influence of ascorbic acid on the activity of this antitumor metal complex in cell culture studies. We show that co-incubation of equicytotoxic, constant amounts of KP1019 with high concentrations of ascorbic acid (50-700 µM) increases cytotoxicity of the ruthenium anticancer drug in the human colon carcinoma cell line SW480, human cervical carcinoma KB-3-1 cells, and the multidrug-resistant subline KBC-1, whereas addition of low concentrations (2.7-50 µM) has a strong chemoprotective effect in the human colon carcinoma cell line SW480, but not in multidrug-resistant KBC-1 cells. Although cellular uptake of KP1019 is not altered, ascorbic acid induce stronger interaction of the ruthenium compound with DNA both in SW480 cells and under cell-free conditions with plasmid DNA. Even if DNA interactions probably play a subordinate role in vivo given the extensive protein binding of the compound, our data exemplify that ascorbic acid enhances the reactivity of KP1019 with biomolecules. Moreover, we demonstrate that the levels of KP1019-generated reactive oxygen species are markedly decreased by co-incubation with ascorbic acid. Conclusively, our results indicate that application of high doses of ascorbic acid might increase the anticancer effects of KP1019.

Metabolization of [Ru(eta(6)-C (6)H (5)CF (3))(pta)Cl (2)]: a cytotoxic RAPTA-type complex with a strongly electron withdrawing arene ligand.

The anticancer ruthenium-arene compound [Ru(eta(6)-C(6)H(5)CF(3))(pta)Cl(2)] (where pta is 1,3,5-triaza-7-phosphatricyclo[3.3.1.1]decane), termed RAPTA-CF3, with the electron-withdrawing alpha,alpha,alpha-trifluorotoluene ligand, is one of the most cytotoxic RAPTA compounds known. To rationalize the high observed cytotoxicity, the hydrolysis of RAPTA-CF3 in water and brine (100 mM sodium chloride) and its reactions with the protein ubiquitin and a double-stranded oligonucleotide (5'-GTATTGGCACGTA-3') were studied using NMR spectroscopy, high-resolution Fourier transform ion cyclotron resonance mass spectrometry, and gel electrophoresis. The aquation of the ruthenium-chlorido complex was accompanied by a loss of the arene ligand, independent of the chloride concentration, which is a special property of the compound not observed for other ruthenium-arene complexes with relatively stable ruthenium-arene bonds. Accordingly, the mass spectra of the biomolecule reaction mixtures contained mostly [Ru(pta)]-biomolecule adducts, whereas [Ru(pta)(arene)] adducts typical of other RAPTA compounds were not observed in the protein or DNA binding studies. Gel electrophoresis experiments revealed a significant degree of decomposition of the oligonucleotide, which was more pronounced in the case of RAPTA-CF3 compared with RAPTA-C. Consequently, facile arene loss appears to be responsible for the increased cytotoxicity of RAPTA-CF3.

Transferring the concept of multinuclearity to ruthenium complexes for improvement of anticancer activity.

Multinuclear platinum anticancer complexes are a proven option to overcome resistance of established anticancer compounds. Transferring this concept to ruthenium complexes led to the synthesis of dinuclear Ru(II)-arene compounds containing a bis(pyridinone)alkane ligand linker. A pronounced influence of the spacer length on the in vitro anticancer activity was found, which is correlated to the lipophilicity of the complexes. IC(50) values in the same dimension as for established platinum drugs were found in human tumor cell lines. No cross-resistance to oxoplatin, a cisplatin prodrug, was observed for the most active complex in three resistant cell lines; in fact, a 10-fold reversal of sensitivity in two of the oxoplatin-resistant lines was found. (Bio)analytical characterization of the representative examples showed that the ruthenium complexes hydrolyze rapidly, forming predominantly diaqua species that exhibit affinity toward transferrin and DNA, indicating that both proteins and nucleobases are potential targets.