Improved HIV-1 RNA quantitation by COBAS AmpliPrep/COBAS TaqMan HIV-1 Test, v2.0 using a novel dual-target approach.

BACKGROUND: HIV-1 RNA viral load is a key parameter for reliable treatment monitoring of HIV-1 infection. Accurate HIV-1 RNA quantitation can be impaired by primer and probe sequence polymorphisms as a result of tremendous genetic diversity and ongoing evolution of HIV-1. A novel dual HIV-1 target amplification approach was realized in the quantitative COBAS AmpliPrep/COBAS TaqMan HIV-1 Test, v2.0 (HIV-1 TaqMan test v2.0) to cope with the high genetic diversity of the virus. OBJECTIVES AND STUDY DESIGN: The performance of the new assay was evaluated for sensitivity, dynamic range, precision, subtype inclusivity, diagnostic and analytical specificity, interfering substances, and correlation with the COBAS AmpliPrep/COBAS TaqMan HIV-1 (HIV-1 TaqMan test v1.0) predecessor test in patients specimens. RESULTS: The new assay demonstrated a sensitivity of 20 copies/mL, a linear measuring range of 20-10,000,000 copies/mL, with a lower limit of quantitation of 20 copies/mL. HIV-1 Group M subtypes and HIV-1 Group O were quantified within +/-0.3 log(10) of the assigned titers. Specificity was 100% in 660 tested specimens, no cross reactivity was found for 15 pathogens nor any interference for endogenous substances or 29 drugs. Good comparability with the predecessor assay was demonstrated in 82 positive patient samples. In selected clinical samples 35/66 specimens were found underquantitated in the predecessor assay; all were quantitated correctly in the new assay. CONCLUSIONS: The dual-target approach for the HIV-1 TaqMan test v2.0 enables superior HIV-1 Group M subtype coverage including HIV-1 Group O detection. Correct quantitation of specimens underquantitated in the HIV-1 TaqMan test v1.0 test was demonstrated.

Robust volume-targeted balanced steady-state free-precession coronary magnetic resonance angiography in a breathhold at 3.0 Tesla: a reproducibility study.

BACKGROUND: Transient balanced steady-state free-precession (bSSFP) has shown substantial promise for noninvasive assessment of coronary arteries but its utilization at 3.0 T and above has been hampered by susceptibility to field inhomogeneities that degrade image quality. The purpose of this work was to refine, implement, and test a robust, practical single-breathhold bSSFP coronary MRA sequence at 3.0 T and to test the reproducibility of the technique. METHODS: A 3D, volume-targeted, high-resolution bSSFP sequence was implemented. Localized image-based shimming was performed to minimize inhomogeneities of both the static magnetic field and the radio frequency excitation field. Fifteen healthy volunteers and three patients with coronary artery disease underwent examination with the bSSFP sequence (scan time = 20.5 ± 2.0 seconds), and acquisitions were repeated in nine subjects. The images were quantitatively analyzed using a semi-automated software tool, and the repeatability and reproducibility of measurements were determined using regression analysis and intra-class correlation coefficient (ICC), in a blinded manner. RESULTS: The 3D bSSFP sequence provided uniform, high-quality depiction of coronary arteries (n = 20). The average visible vessel length of 100.5 ± 6.3 mm and sharpness of 55 ± 2% compared favorably with earlier reported navigator-gated bSSFP and gradient echo sequences at 3.0 T. Length measurements demonstrated a highly statistically significant degree of inter-observer (r = 0.994, ICC = 0.993), intra-observer (r = 0.894, ICC = 0.896), and inter-scan concordance (r = 0.980, ICC = 0.974). Furthermore, ICC values demonstrated excellent intra-observer, inter-observer, and inter-scan agreement for vessel diameter measurements (ICC = 0.987, 0.976, and 0.961, respectively), and vessel sharpness values (ICC = 0.989, 0.938, and 0.904, respectively). CONCLUSIONS: The 3D bSSFP acquisition, using a state-of-the-art MR scanner equipped with recently available technologies such as multi-transmit, 32-channel cardiac coil, and localized B0 and B1+ shimming, allows accelerated and reproducible multi-segment assessment of the major coronary arteries at 3.0 T in a single breathhold. This rapid sequence may be especially useful for functional imaging of the coronaries where the acquisition time is limited by the stress duration and in cases where low navigator-gating efficiency prohibits acquisition of a free breathing scan in a reasonable time period.

Aortic vessel wall magnetic resonance imaging at 3.0 Tesla: a reproducibility study of respiratory navigator gated free-breathing 3D black blood magnetic resonance imaging.

The purpose of this study was to evaluate a free-breathing three-dimensional (3D) dual inversion-recovery (DIR) segmented k-space gradient-echo (turbo field echo [TFE]) imaging sequence at 3T for the quantification of aortic vessel wall dimensions. The effect of respiratory motion suppression on image quality was tested. Furthermore, the reproducibility of the aortic vessel wall measurements was investigated. Seven healthy subjects underwent 3D DIR TFE imaging of the aortic vessel wall with and without respiratory navigator. Subsequently, this sequence with respiratory navigator was performed twice in 10 healthy subjects to test its reproducibility. The signal-to-noise (SNR), contrast-to-noise ratio (CNR), vessel wall sharpness, and vessel wall volume (VWV) were assessed. Data were compared using the paired t-test, and the reproducibility of VWV measurements was evaluated using intraclass correlation coefficients (ICCs). SNR, CNR, and vessel wall sharpness were superior in scans performed with respiratory navigator compared to scans performed without. The ICCs concerning intraobserver, interobserver, and interscan reproducibility were excellent (0.99, 0.94, and 0.95, respectively). In conclusion, respiratory motion suppression substantially improves image quality of 3D DIR TFE imaging of the aortic vessel wall at 3T. Furthermore, this optimized technique with respiratory motion suppression enables assessment of aortic vessel wall dimensions with high reproducibility.

Automated scoring of AFLPs using RawGeno v 2.0, a free R CRAN library.

Amplified Fragment Length Polymorphisms (AFLPs) are a cheap and efficient protocol for generating large sets of genetic markers. This technique has become increasingly used during the last decade in various fields of biology, including population genomics, phylogeography, and genome mapping. Here, we present RawGeno, an R library dedicated to the automated scoring of AFLPs (i.e., the coding of electropherogram signals into ready-to-use datasets). Our program includes a complete suite of tools for binning, editing, visualizing, and exporting results obtained from AFLP experiments. RawGeno can either be used with command lines and program analysis routines or through a user-friendly graphical user interface. We describe the whole RawGeno pipeline along with recommendations for (a) setting the analysis of electropherograms in combination with PeakScanner, a program freely distributed by Applied Biosystems; (b) performing quality checks; (c) defining bins and proceeding to scoring; (d) filtering nonoptimal bins; and (e) exporting results in different formats.

Mico-cocktail validation for CYP3A and CYP2D6 assessment using a low dose of midazolam (0.075 mg) and dextromethorphan (2.5 mg)

Background/Aim: Cocktail approach is generally preferred to individual administration of probes in order to characterize the activity of multiple enzymes. However, cocktail strategy has several drawbacks such as drug-drug interactions, tolerability and toxicity. Hence, there is a need to develop cocktails using low doses of probes. Our aim was to investigate whether the simultaneous oral administration of microdoses of midazolam (MDZ) and dextromethorphan (DEM) can be used to assess the simultaneous activities of CYP3A and CYP2D6. Methods: As part of a 5 arm randomized cross-over control trial on the analgesic efficacy of oxycodone, ten healthy young non-smoking males received the following combinations of drugs: Quinidine (Q)+ ketoconazole (K) or Q+placebo (P) or K+P or P+P. In all cases MDZ (0.075 mg) and DEM (2.5 mg) were administrated 1 hour after Q, K or P. CYP2D6 and CYP3A activities were determined after urine collection during 8 hours (ratio DEM/DOR), and a blood sample (EDTA) after 30 min (ratio 1-OH-MDZ/MDZ). DEM and DOR analysis was performed using LC-fluorescence. MDZ and 1-OH-MDZ determination was performed using GC-MS. Allele's variants of CYP2D6 were detected using the AmpliChipTMCYP450 (Roche). Results: CYP2D6 genotype predicted 1 poor (PM), 1 intermediate (IM), 7 extensive (EM) and 2 ultra rapid (UM) metabolizers. A good correlation was obtained between the predicted and the measured phenotypes except for 1 EM phenotyped as UM. Two duplications for alleles *41/*41xN and *1/*2xN were detected and the two volunteers were phenotyped as UM. A potent inhibition of CYP2D6 or CYP3A4 was obtained when Q or K were used. Mean metabolic ratio DEM/DOR in P and K groups were 0.015 (±0.028) and 0.015 (±0.019). It significantly increased in Q and QK groups (0.668 (±0.676) and 0.743 (±1.038)). Mean 1-OH-MDZ/MDZ in P, Q were 2.73 (±1.05) and 2.55 (±1.40) while it significantly decreased in K and QK groups (0.11 (±0.05), 0.10 (±0.05)). Moreover, there were no statistically significant differences between QK and K sessions for CYP3A and between QK and Q for CYP2D6 which indicate that there is no interaction between the two metabolic pathways. Conclusion: Simultaneous assessment of CYP3A and CYP2D6 activities can be obtained by low oral doses (micro-cocktail) of MDZ and DEM. Specific inhibitors such as Q or K modulates selectively CYP2D6 or CYP3A activities.

The Brief Psychiatric Rating Scale (version 4.0) factorial structure and its sensitivity in the treatment of outpatients with unipolar depression.

The 24-item Brief Psychiatric Rating Scale (BPRS, version 4.0) enables the rater to measure psychopathology severity. Still, little is known about the BPRS's reliability and validity outside of the psychosis spectrum. The aim of this study was to examine the factorial structure and sensitivity to change of the BPRS in patients with unipolar depression. Two hundred and forty outpatients with unipolar depression were administered the 24-item BPRS. Assessments were conducted at intake and at post-treatment in a Crisis Intervention Centre. An exploratory factor analysis of the 24-item BPRS produced a six-factor solution labelled "Mood disturbance", "Reality distortion", "Activation", "Apathy", "Disorganization", and "Somatization". The reduction of the total BPRS score and dimensional scores, except for "Activation", indicates that the 24-item BPRS is sensitive to change as shown in patients that appeared to have benefited from crisis treatment. The findings suggest that the 24-item BPRS could be a useful instrument to measure symptom severity and change in symptom status in outpatients presenting with unipolar depression.

3. Formation continue: promotion du mouvement et de l'activité physique chez les 0-6 ans

Sedentary lifestyle in children is increasing at an alarming rate. Now, promotion of physical activity by health professionals is a promising way. To support childhood specialists in this role, a transdisciplinary training is being developped.

Blind docking of 260 protein-ligand complexes with EADock 2.0.

Molecular docking softwares are one of the important tools of modern drug development pipelines. The promising achievements of the last 10 years emphasize the need for further improvement, as reflected by several recent publications (Leach et al., J Med Chem 2006, 49, 5851; Warren et al., J Med Chem 2006, 49, 5912). Our initial approach, EADock, showed a good performance in reproducing the experimental binding modes for a set of 37 different ligand-protein complexes (Grosdidier et al., Proteins 2007, 67, 1010). This article presents recent improvements regarding the scoring and sampling aspects over the initial implementation, as well as a new seeding procedure based on the detection of cavities, opening the door to blind docking with EADock. These enhancements were validated on 260 complexes taken from the high quality Ligand Protein Database [LPDB, (Roche et al., J Med Chem 2001, 44, 3592)]. Two issues were identified: first, the quality of the initial structures cannot be assumed and a manual inspection and/or a search in the literature are likely to be required to achieve the best performance. Second the description of interactions involving metal ions still has to be improved. Nonetheless, a remarkable success rate of 65% was achieved for a large scale blind docking assay, when considering only the top ranked binding mode and a success threshold of 2 A RMSD to the crystal structure. When looking at the five-top ranked binding modes, the success rate increases up to 76%. In a standard local docking assay, success rates of 75 and 83% were obtained, considering only the top ranked binding mode, or the five top binding modes, respectively.