Validation, Deployment, and Real-World Implementation of a Modular Toolbox for Alzheimer's Disease Detection and Dementia Risk Reduction: The AD-RIDDLE Project.

The Real-World Implementation, Deployment, and Validation of Early Detection Tools and Lifestyle Enhancement (AD-RIDDLE) project, recently launched with the support of the EU Innovative Health Initiative (IHI) public-private partnership and UK Research and Innovation (UKRI), aims to develop, test, and deploy a modular toolbox platform that can reduce existing barriers to the timely detection, and therapeutic approaches in Alzheimer's disease (AD), thus accelerating AD innovation. By focusing on health system and health worker practices, AD-RIDDLE seeks to improve and smooth AD management at and between each key step of the clinical pathway and across the disease continuum, from at-risk asymptomatic stages to early symptomatic ones. This includes innovation and improvement in AD awareness, risk reduction and prevention, detection, diagnosis, and intervention. The 24 partners in the AD-RIDDLE interdisciplinary consortium will develop and test the AD-RIDDLE toolbox platform and its components individually and in combination in six European countries. Expected results from this cross-sectoral research collaboration include tools for earlier detection and accurate diagnosis; validated, novel digital cognitive and blood-based biomarkers; and improved access to individualized preventative interventions (including multimodal interventions and symptomatic/disease-modifying therapies) across diverse populations, within the framework of precision medicine. Overall, AD-RIDDLE toolbox platform will advance management of AD, improving outcomes for patients and their families, and reducing costs.

Metabolic profile modifications in milk after enrofloxacin administration studied by liquid chromatography coupled with high resolution mass spectrometry.

High resolution accurate mass spectrometry (HRMS) operating in full scan MS mode was used in the search and identification of metabolites in raw milk from cows medicated with enrofloxacin. Data consisting of m/z features were taken throughout the entire chromatogram of milk samples from medicated animals and were compared with blank samples. Twenty six different compounds were identified. Some of them were attributed to structures related to enrofloxacin while others were dipeptides or tripeptides. Additionally, enrofloxacin was administered in a controlled treatment for three days. Milk was collected daily from the first day of treatment and until four days after in the search for the identified compounds. The obtained data were chemometrically treated by Principal Component Analysis. Samples were classified by this method into three different groups corresponding to days 1-2, day 3 and days 4-7 considering the different concentration profile evolution of metabolites during the days studied. Tentative metabolic pathways were designed to rationalize the presence of the newly identified compounds.

Monolithic silica columns with covalently attached octaproline chiral selector. Dependence of performance on derivatization degree and comparison with a bead-based analogue.

A monolithic silica gel chromatographic matrix was derivatized repetitively with an octaproline-derived chiral selector (CS). The increasingly derivatized column was tested after each derivatization reaction. The enantioseparation ability, resolution and efficiency were found to depend on the content of CS attained after each reaction. Moreover, enantioselectivity and performance of the column with the highest CS coverage were compared to those of a bead-based chiral stationary phase (CSP) counterpart. The octaproline-derivatized monolithic column demonstrated increased enantioseparation factors, resolution and broader applicability than the particle-based column. Finally, the loading capacity of the CSPs was also examined. The monolithic octaproline-derived column permits the separation of 3-20 times higher molar amounts of the tested analytes (depending on the compound considered) than the particle-based counterpart. The enhanced capabilities of the derivatized monolithic column with respect to that of a bead-based counterpart cannot be explained only on the basis of an increased CS coverage. The involvement of an effect produced by the chromatographic silica support structure in the obtained results is discussed.

High resolution mass spectrometry in the identification of transformation products and metabolites from ß-lactam antibiotics in thermally treated milk.

Antibiotics such as ß-lactam derivatives (penicillins and cephalosporins) are frequently used in veterinary medicine. The presence of these antibiotics together with their metabolites and/or products produced in subsequent treatments at which milk is submitted (sterilization, pasteurization), may be responsible for bacterial resistance, allergy and/or toxicity on sensitive individuals. In this study, liquid chromatography coupled with high resolution mass spectrometry (LC-HRMS) is used to identify transformation products (TPs) from four ß-lactam antibiotics (amoxicillin (AMOX), cephapirin (PIR), ceftiofur (TIO) and penicillin G (PENG)) in thermally treated cow milk. In addition, milk from cows medicated with PENG has also been analogously treated and studied. The detected TPs come mainly from hydrolysis and decarboxylation reactions. Products more strongly degraded respect to parent compounds (of lower molecular weight) were obtained after treating milk at higher temperatures. Products identified in milk from cows medicated with PENG have been classified as TPs when coming from chemical/thermal degradation, and metabolites when resulting from the biological drug metabolism. While TPs are the result of hydrolysis and decarboxylation processes, as already indicated, an enzymatic conjugation with amino acids is suggested to be involved in the formation of metabolites.

Identification of a novel metabolite in the degradation of pyrene by Mycobacterium sp. strain AP1: actions of the isolate on two- and three-ring polycyclic aromatic hydrocarbons.

Mycobacterium sp. strain AP1 grew with pyrene as a sole source of carbon and energy. The identification of metabolites accumulating during growth suggests that this strain initiates its attack on pyrene by either monooxygenation or dioxygenation at its C-4, C-5 positions to give trans- or cis-4,5-dihydroxy-4,5-dihydropyrene, respectively. Dehydrogenation of the latter, ortho cleavage of the resulting diol to form phenanthrene 4,5-dicarboxylic acid, and subsequent decarboxylation to phenanthrene 4-carboxylic acid lead to degradation of the phenanthrene 4-carboxylic acid via phthalate. A novel metabolite identified as 6,6'-dihydroxy-2,2'-biphenyl dicarboxylic acid demonstrates a new branch in the pathway that involves the cleavage of both central rings of pyrene. In addition to pyrene, strain AP1 utilized hexadecane, phenanthrene, and fluoranthene for growth. Pyrene-grown cells oxidized the methylenic groups of fluorene and acenaphthene and catalyzed the dihydroxylation and ortho cleavage of one of the rings of naphthalene and phenanthrene to give 2-carboxycinnamic and diphenic acids, respectively. The catabolic versatility of strain AP1 and its use of ortho cleavage mechanisms during the degradation of polycyclic aromatic hydrocarbons (PAHs) give new insight into the role that pyrene-degrading bacterial strains may play in the environmental fate of PAH mixtures.

Correlation of p53 mutations with resistance to platinum-based chemotherapy and shortened survival in ovarian cancer.

PURPOSE: The p53 tumor suppressor gene plays a central role in cell cycle regulation and induction of apoptosis. We analyzed p53 alterations and their impact on response to chemotherapy and clinical outcome in ovarian cancer patients. EXPERIMENTAL DESIGN: One hundred seventy-eight ovarian carcinomas, snap frozen and stored at -80 degrees C, were analyzed for mutations of the p53 gene (exons 2-11) by single-strand conformation polymorphism and DNA sequencing and for p53 overexpression by immunohistochemistry (monoclonal antibody DO7). RESULTS: p53 mutations were found in 56% (99 of 178) of the tumors, and 62% of these were located in evolutionary highly conserved domains of the gene. Time to progression and overall survival were significantly shortened in patients with p53 mutations compared with wild-type p53 (P = 0.029 and P = 0.014) and patients with mutations in highly conserved domains as opposed to nonconserved domains or wild-type p53 (P = 0.010 and P = 0.007). p53 protein overexpression (>10% positively stained nuclei) was found in 62% (110 of 178). Time to progression and overall survival were shorter in cases with p53 overexpression (cutpoint, 10%: P = 0.071 and P = 0.056) but only marginally significant. Resistance to adjuvant cisplatin or carboplatin chemotherapy was significantly more frequent in patients with p53 overexpression (P = 0.001) or p53 missense mutations (P = 0.008) than patients with normal p53. CONCLUSIONS: p53 alterations correlate significantly with resistance to platinum-based chemotherapy, early relapse, and shortened overall survival in ovarian cancer patients in univariate analysis. In multivariable analysis though, p53 was not an independent prognostic factor.

Amphioxus Evx genes: implications for the evolution of the Midbrain-Hindbrain Boundary and the chordate tailbud.

Evx genes are widely used in animal development. In vertebrates they are crucial in gastrulation, neurogenesis, appendage development and tailbud formation, whilst in protostomes they are involved in gastrulation and neurogenesis, as well as segmentation at least in Drosophila. We have cloned the Evx genes of amphioxus (Branchiostoma floridae), and analysed their expression to understand how the functions of Evx have evolved between invertebrates and vertebrates, and in particular at the origin of chordates and during their subsequent evolution. Amphioxus has two Evx genes (AmphiEvxA and AmphiEvxB) which are genomically linked. AmphiEvxA is prototypical to the vertebrate Evx1 and Evx2 genes with respect to its sequence and expression, whilst AmphiEvxB is very divergent. Mapping the expression of AmphiEvxA onto a phylogeny shows that a role in gastrulation, dorsal-ventral patterning and neurogenesis is probably retained throughout bilaterian animals. AmphiEvxA expression during tailbud development implies a role for Evx throughout the chordates in this process, whilst lack of expression at the homologous region to the vertebrate Midbrain-Hindbrain Boundary (MHB) is consistent with the elaboration of the full organiser properties of this region being a vertebrate innovation.

Evaluation of the contribution to enantioselectivity of quinine and quinidine scaffolds in chemically and physically mixed chiral selectors.

Three "dimeric" C(9)-carbamates of quinine (QN) and quinidine (QD), that is, QN-QN, QD-QD, and QN-QD (chemically prepared mixture of the two cinchona-derived subunits), separated by an ethylene spacer were synthesized and used as chiral selectors for HPLC and capillary electrophoresis (CE) for the resolution of chiral acids. The chiral recognition abilities of these dimers and of several physically prepared mixtures thereof were compared in order to estimate the contribution of every cinchona scaffold to the overall enantioselectivity. The diverse phenomena observed in nonaqueous capillary electrophoresis (NACE), either using the selector added to the background electrolyte (BGE) in the total filling or partial filling mode, led us to rationalize, taking into account the relative mobilities of the chiral selectors in the capillary. The chromatographic and electrophoretic properties were compared with those of the corresponding "monomeric" QN and QD carbamates.

Covalently bonded polysaccharide derivatives as chiral stationary phases in high-performance liquid chromatography.

Polysaccharide derivatives have been extensively used as chromatographic chiral selectors in chiral stationary phases (CSPs) for the separation of enantiomers by HPLC. When coated onto a silica matrix, they represent nowadays one of the most popular type of CSPs. However, they are only compatible with a limited choice of solvents. The main drawback of these CSPs is related to the solubility of the chiral selector in a number of solvents, which limits their applicability. The different attempts which have been described up to now to overcome this problem by covalently fixing the chiral selector to a matrix are reviewed in this paper.

Characterization of doubly substituted polysaccharide derivatives.

Derivatives of cellulose, amylose and chitosan, bearing simultaneously 10-undecenoyl and arylaminocarbonyl or benzoyl groups were characterized by the combined use of 1H NMR and elemental analysis. The mathematical manipulation of elemental analysis data permits the calculation of the degree of substitution for each kind of substituent. The method was validated and is applicable to other derivatives.

Preparation of mixed 10-undecenoyl/phenylaminocarbonyl or benzoyl derivatives of chitosan.

The preparation of mixed 10-undecenoyl/phenylaminocarbonyl or benzoyl derivatives of chitosan is described. Several mixed derivatives were prepared using differently substituted aryl isocyanates or benzoyl chlorides. The reactivity of the starting polysaccharide was found to be influenced by the acetyl content.

The amphioxus Hox cluster: deuterostome posterior flexibility and Hox14.

The amphioxus (Branchiostoma floridae) Hox cluster is a model for the ancestral vertebrate cluster, prior to the hypothesized genome-wide duplications that may have facilitated the evolution of the vertebrate body plan. Here we describe the posterior (5') genes of the amphioxus cluster, and report the isolation of four new homeobox genes. Vertebrates possess 13 types of Hox gene (paralogy groups), but we show that amphioxus possesses more than 13 Hox genes. Amphioxus is now the first animal in which a Hox14 gene has been found. Our mapping and phylogenetic analysis of amphioxus "Posterior Class" Hox genes reveals that these genes are evolving at a faster rate in deuterostomes than in protostomes, a phenomenon we term Posterior Flexibility.

Environmental tobacco smoke interference in the assessment of the health impact of a municipal waste incinerator on children through urinary thioether assay.

The urinary elimination of thioethers urinary thioethers (UT) was studied in 83 schoolchildren living in two different areas of the city of Mataró, with special attention paid to the influence of a waste incinerator and of the smoking habits of their parents. The mean UT values were 8.79+/-3.23 and 7.52+/-3.23 mmol/mol creatinine in the area close to the incinerator (A1) and in the area far away from it (A2) respectively (statistically n.s.). Children exposed to environmental tobacco smoke (ETS) at home presented increased levels of UT (8.60+/-3.11 vs 5.93+/-3.22 mmol/mol creatinine; P=0.002). Considering the two exposures together (waste incinerator and ETS), no differences were found between the two areas studied (A1 and A2) in non-exposed (ETS) children, whereas slight differences were found when comparing highest ETS exposed children from the two areas (10. 18+/-2.70 vs 8.00+/-3.42 mmol/mol creatinine; P=0.04). Exposure to ETS could affect health more than pollutants from a waste incinerator and may interfere with non-selective assays such as urinary thioethers.

Enantioselective separation of several piperidine-2, 6-diones on a covalently bonded cellulose 3,5-dimethylphenyl carbamate/10-undecenoate chiral selector.

A series of piperidine-2, 6-dione drugs were enantiomerically resolved on a covalently bonded cellulose 3,5-dimethylphenyl carbamate/10-undecenoate chiral stationary phase (CSP), under normal- or reversed-phase conditions. A single column that can be applied in both modes may be advantageous when considering the shorter setup time required and the solubility of the compound to be analysed since many samples possess different solubilities. The covalently bonded CSP was compared to a commercially available chiral adsorbent, Chiralcel OD, which was previously used in our laboratory for the enantiomeric resolution of the above-mentioned drug series. Chiralcel OD is used in the normal-phase mode and is more fragile than the column used in this study. The user is restricted to the range of solvents available as eluents. Hence, it was of interest to look at the possible advantages of using a chemically bonded phase that can be applied in dual mode.

Mammary serum antigen (MSA), Ca 549, CA 15-3 and CEA in breast cancer preoperative sensitivity and correlation to prognostic factors.

Previously a higher preoperative sensitivity of the mucin like glycoprotein Mammary Serum Antigen (MSA) compared to established tumor markers was reported. The aim of our study was the comparison of MSA and CA 549, CA 15-3 and CEA in primary breast cancer and the correlation to prognostic factors. In 119 patients MSA and CA 549 serum levels were analysed by ELISA, CEA and CA 15-3 levels by LIA. We received the following sensitivities: 30.2% for MSA (cut off = 55 U/mL), 21.8% for CA 549 (cut off = 12.6 U/mL), 20.5% for CA 15-3 (cut off = 25 U/ml) and 10.7% for CEA (cut off = 6 ng/ml). Significant correlations were found between MSA concentrations and grading (p = 0.006), tumor size (p = 0.005) and metastases (p = 0.03). No correlations were found to tumor type, hormone receptors, lymph node status and cathepsin D. MSA is a valuable tumor marker with the highest preoperative sensitivity. Further studies on the value of MSA in the follow-up are necessary.

Boeck sarcoidosis of the breast: mammographic, ultrasound, and MR findings.

The rare case of sarcoidosis of the breast is presented. The mammographic, ultrasound, and MR appearances are described.

[Can the high-resolution 13-MHz ultrasonography facilitate the preoperative localization and marking of microcalcification clusters?]. / Kann die hochauflösende 13-MHz-Sonographie die präoperative Lokalisation und Markierung von Mikroverkalkungsgruppen erleichtern?

PURPOSE: The value of 13 MHz ultrasound regarding the preoperative localisation in combination with mammography were investigated into. MATERIAL AND METHOD: Out of 112 mammaographically detected calcifications, 30 (30 patients) were clusters of microcalcifications. Out of these, 23 were classified as suspicious of malignancy and were preoperatively localised with a 7.5 and 13 MHz probe. Upon marking the skin, a fine needle was inserted, and after due correction blue dye or coal solution was instilled. RESULTS: Following mammographically shown position, 23 clusters of microcalcifications could be localised using the 13 MHz probe. A localisation with the 7.5 MHz probe was impossible in all cases. A single correction of the needle's site was necessary with 35% (8/23) of patients. The maximum distance of the needle's tip in case of misposition measured 11 mm. CONCLUSION: Giving a known mammographic position and a sonographic perceptibility of clusters of microcalcifications in a maximal depth of penetration of 2 cm, 13 MHz high resolution ultrasound examination in combination with mammography prove to be a valuable means in facilitating the preoperative localisation.

[Significance of PCNA proliferating fraction for prognosis of ovarian carcinoma]. / Die Bedeutung der PCNA-Proliferationsfraktion für die Prognose des Ovarialkarzinoms.

The prognostic value of the PCNA-proliferative fraction as compared to conventional clinical and histomorphological factors (FIGO-stage, tumour type, histological grading, lymph node status, size of residual tumour) was investigated in 81 ovarian cancer patients. Categorisation of PCNA-expression into tumours with low and high proliferative activity (< 20%/ > or = 20% according to laboratory standards) had the highest prognostic value. Categorisation on the basis of the median value (< or = 34%/> 34%) or classification of PCNA as a continuous variable did not prove advantageous. PCNA-proliferative fraction was significantly directly correlated with histological grading (p = 0.006). Tumours with a high PCNA expression had a greater frequency of macroscopically detectable residual tumours. In univariate survival analyses patients with highly proliferating tumours had a worse outcome than patients with tumour of low proliferation (PCNA or = 20%/20%, p = 0.012; PCNA < or = 34%/ > 34%, p = 0.08). The result was consistent in subgroup of FIGO III-tumours (p = 0.031, PCNA < 20%/> or = 20%), of FIGO l-tumours (p = 0.036, PCNA < 20%/> or = 20%), of carcinomas without post-operative residual tumour (p = 0.03 PCNA < 20%/> or = 20%) and also of FIGO III-tumour without residual tumours (p = 0.041 PCNA < 20%/> or = 20%). Multivariate survival analysis comprising all the patients revealed PCNA expression (< 20%/> or = 20%) as an independent prognostic factor second to the size of the residual tumour. In patients with FIGO III-tumours PCNA proves significant as an independent factor after the size of the residual tumour was removed from the model. Thus, PCNA provides additional information which may prove beneficial in determining prognostic estimates for ovarian cancer.

[C-erbB-2, PCNA and histomorphologic factors in breast carcinoma after oral contraceptive use]. / C-erbB-2, PCNA und histomorphologische Faktoren beim Mammakarzinom nach Einnahme oraler Kontrazeptiva.

The influence of oral contraceptives (OC) on histomorphological and molecular biological prognostic factors was studied in 471 breast cancer patients. Differences in histological tumor type, histological grade, tumor size, lymph node status, hormonal receptor status, PCNA expression and c-erbB-2 protein overexpression were investigated in relation to the duration of OC use (< 5 years/ > or = 5 years) and the time since last use. A total of 297 (63%) patients had used oral contraceptives at some time in their life; 186 patients (39.5%) had used OC's for 5 years or more. There were no significant differences in the tumor characteristics investigated with respect to OC use in general. Neither long-term use at some time in their life nor long-term use until breast cancer diagnosis had an effect on histomorphological and molecular biological factors. Thus, steroid hormones contained in OC's had no direct effect on prognostic factors in breast cancer.