Selective killing of ATM- or p53-deficient cancer cells through inhibition of ATR.

Here we report a comprehensive biological characterization of a potent and selective small-molecule inhibitor of the DNA damage response (DDR) kinase ATR. We show a profound synthetic lethal interaction between ATR and the ATM-p53 tumor suppressor pathway in cells treated with DNA-damaging agents and establish ATR inhibition as a way to transform the outcome for patients with cancer treated with ionizing radiation or genotoxic drugs.

Colorectal cancer: texture analysis of portal phase hepatic CT images as a potential marker of survival.

PURPOSE: To assess the utility of texture analysis of liver computed tomographic (CT) images by determining the effect of acquisition parameters on texture and by comparing the abilities of texture analysis and hepatic perfusion CT to help predict survival for patients with colorectal cancer. MATERIALS AND METHODS: The study comprised a phantom test and a clinical evaluation of 48 patients with colorectal cancer who had consented to retrospective analysis of hepatic perfusion CT data acquired during a research study approved by the institutional review board. Both components involved texture analysis to quantify the relative contribution of CT features between 2 and 12 pixels wide to overall image brightness and uniformity. The effect of acquisition factors on texture was assessed on CT images of a cylindric phantom filled with water obtained by using tube currents between 100 and 250 mAs and voltages between 80 and 140 kVp. Texture on apparently normal portal phase CT images of the liver and hepatic perfusion parameters were related to patient survival by using Kaplan-Meier survival analysis. RESULTS: A texture parameter that compared the uniformity of distribution of CT image features 10 and 12 pixels wide exhibited the least variability with CT acquisition parameters (maximum coefficient of variation, 2.6%) and was the best predictor of patient survival (P < .005). There was no significant association between survival and hepatic perfusion parameters. CONCLUSION: The study provides preliminary evidence that analysis of liver texture on portal phase CT images is potentially a superior predictor of survival for patients with colorectal cancer than CT perfusion imaging. SUPPLEMENTAL MATERIAL: http://radiology.rsnajnls.org/cgi/content/full/2502071879/DC1.

Proteomic analysis of the mouse mammary gland is a powerful tool to identify novel proteins that are differentially expressed during mammary development.

After lactation, the mouse mammary gland undergoes apoptosis and tissue remodelling as the gland reverts to its prepregnant state. This complex change was investigated using 2-DE. An integrated database was produced from lactation and involution proteomes. Forty-four molecular cluster indexes (MCIs) that showed altered expression from lactation to involution were selected for MS analysis. Of these, 32 gave protein annotations, 18 of which were unequivocal proteins. Selected proteins were then studied across all of development, including pregnancy, using data integrated from another proteome database. Two proteins, the RNA polymerase B transcription factor 3 (BTF3) and the minichromosome maintenance protein 3 (MCM3), although initially selected on the basis of the lactation/involution criteria, had expression profiles that indicated an additional role in mammary development and were further analysed. BTF3, a transcription factor previously not described in the mammary gland, was up-regulated strongly in pregnancy, indicating an involvement in alveolar growth. MCM3's expression was greatest in pregnancy and late involution, decreasing through lactation. Immunohistochemistry localised MCM3 to the mammary epithelium, where a greater proportion of cells stained than for the proliferation marker Ki67. MCM3 expression during lactation may identify cells that are licensed to repopulate the gland during cell loss in lactation and following involution.

Proteomic analysis of mouse mammary terminal end buds identifies axonal growth cone proteins.

Ductal morphogenesis in the mouse mammary gland occurs mainly postnatally and is driven by specialized structures at the ends of the developing ducts, the terminal end buds (TEBs), which later regress once ductal growth is complete. To identify proteins that are specifically associated with migration of TEBs we developed a novel method of isolating TEBs, which eliminated the mammary stroma. The protein expression profile of the TEBs was then compared with that of isolates taken from the 4th inguinal mammary gland of adult virgin mice using two-dimensional (2-D) gel electrophoresis and mass spectrometry (MS) analysis (matrix-assisted laser desorption/ionization and quadrupole time of flight). Following construction of an integrated protein expression database, 44 protein features which showed differential expression levels between the two sets were chosen for MS analysis. Of these, 24 gave protein annotations whereas the other 20 produced unidentified peptides. Fourteen unequivocal proteins were identified from these 24, whereas the remaining 10 matched more than one protein within a single 2-D gel feature. Several of the identified proteins were associated with the cytoskeleton and have previously been reported in axonal growth cones, suggesting that they may influence cell shape and motility within the advancing TEBs, in a similar fashion to migrating axons.

Proteome analysis of adipogenesis.

Adipose tissue plays a crucial endocrine role in controlling whole body glucose homeostasis and insulin sensitivity. Given the substantial rise in obesity and obesity-related diseases such as diabetes, it is important to understand the molecular basis of adipocyte differentiation and its control. Many studies have successfully exploited gene array technology to monitor changes in the profile of expressed genes during adipocyte differentiation, although this method only measures changes at the level of individual mRNA species. Using two-dimensional polyacrylamide gel electrophoresis, high-throughput image analysis, and candidate picking coupled with sequencing mass spectrometry, we have followed the changes in protein expression profile that occur during the differentiation of 3T3-L1 fibroblasts into adipocytes in response to dexamethasone, isobutyl methyl xanthine and insulin, or to the PPARgamma agonist, ciglitazone. Using this technique we have found alterations in the profile of over 2000 protein species during adipogenesis. Our studies reveal previously unknown alterations during adipogenesis in the expression or mobility (on sodium dodecyl sulfate-polyacrylamide gel electrophoresis) of coactosin, which promotes actin filament destabilization, several signalling molecules, including RhoGDI-1, RhoGDI-2 and EHD1, and NEDD5 a protein involved in cytokinesis.

Precision of bone densitometry measurements: when is change true change and does it vary across bone density values?

The precision error of the bone densitometer is used to interpret significant change in bone mineral density (BMD) in serial studies. The precision error can be expressed as standard deviation (SD) or coefficient of variation (CV). The aims of this study are to determine the precision error over a range of BMD values and to demonstrate the application of the precision error in clinical practice. A bone phantom was used consisting of a perspex block with eight compartments containing varying amounts of hydroxyapatite powder to simulate a range of bone densities. The block was scanned 21 times and manual regions placed over each compartment to measure the BMD in each compartment. There were no significant differences in the variances or SD for all eight compartments, that is, over the range of BMD normally encountered in clinical practice. However, the calculated CV show a progressive fall in values as the BMD rises. Therefore, the SD should be used to calculate significant BMD change. In a practise with quality control procedures in place to detect calibration drift and with appropriately trained personnel, a change of approximately 0.05 g/cm2 is generally regarded as being a significant change at a 95% confidence level.

The diagnostic role of dual femur bone density measurement in low-impact fractures.

A high correlation has been documented between the left and right femoral bone mineral densities in the normal population. This suggests that dual femur measurements are not justified in clinical practice. This study evaluated whether this premise holds for subjects who have lost bone mass and have sustained fractures with minimal trauma. Seventy-eight women aged 31-83 years (mean=66 years) with previous low-impact fractures had both proximal femora measured using dual-energy X-ray absorptiometry. There were significant correlations between values in the left and right total hip (TH) (r=0.95; p<0.05) and in the left and right femoral neck (FN) (r=0.90; p<0.05). The mean differences between the left and right TH and FN densities were not significant. However, the range of the limits of agreement for the TH (-0.074 to 0.086 g/cm2) and FN (-0.115 to 0.105 g/cm2) were greater than the 95% confidence interval for true change for the TH (0.05 g/cm2) and FN (0.07 g/cm2). Any longitudinal BMD assessment therefore needs to measure the same proximal femur to get a reliable comparison. A one-tailed analysis showed that for the TH, 7.5% of subjects had a T-score discordance greater than or equal to 0.5 and 0.5% had a T-score discordance greater than or equal to 1. For the FN, 9% had a T-score discordance greater than or equal to 0.5 and 2.5% had a T-score discordance greater than or equal to 1. The use of dual femur measurements increases the diagnostic yield by about 10% in subjects with prior minimal trauma fractures.

Centralized assessment of dual-energy X-ray absorptiometry (DEXA) in multicenter studies of HIV-associated lipodystrophy.

BACKGROUND: The accurate measurement of total body and subcutaneous fat is essential if therapeutic interventions, aimed at preventing or reversing lipodystrophy syndrome, are to be adequately assessed. The aim of this study was to investigate the variability of dual-energy X-ray absorptiometry (DEXA) scans analysis performed at local sites compared to central analysis in a multicenter clinical trial. METHOD: The PIILR study was a multicenter randomized clinical trial in which 80 HIV-infected patients with physician-documented lipodystrophy had serial measurements of body composition performed with Lunar DEXA scans. Scans were analyzed at local sites and then were reanalyzed centrally. RESULTS: DEXA scans from 73 patients who completed 24 weeks study were compared. Greater variation in the locally analyzed results than in the centrally reanalyzed data was noted, with arm, leg, and combined limb fat being most divergent between the local and centralized assessments (ratio of local to central standard deviation was 1.28, 1.31, and 1.35, respectively). The magnitude of this variance was enough to alter statistically relevant differences between study populations. CONCLUSION: Quality assurance is an important issue in the use of DEXA scans to determine body fat composition in multicenter research studies. A central quality assurance site should be incorporated to reduce variability in results.

Comparative evaluation of the DPX and DPX-IQ lunar densitometer systems following software upgrade.

Bone densitometry research departments perform system and software upgrades infrequently in order to maintain high precision. This study compares the results obtained on a Lunar densitometer with DPX, and DPX-IQ installed to achieve year 2000 compliance. The DPX-IQ provides an improved femur edge detection algorithm with an expanded reference database. Two hundred data files for each measurement site acquired on DPX were reanalyzed on DPX-IQ. There was no change to the bone mineral density (BMD), bone mineral content (BMC), T-scores or Z-scores for the L2-L4 spine, radius (ultradistal and 33%), and total body. There was a significant high correlation for the femoral neck BMD (r = 0.98; p < 0.05). The mean differences in BMD, BMC, T-scores, and Z-scores at the femoral neck and Ward's and trochanteric regions were not significant (p > 0.05). The limits of agreement within the 95% confidence interval for the femoral neck BMD using the Bland and Altman method was between -0.057 and 0.063 g/cm(2). This order of magnitude magnifies the long-term precision error and alters the usual confidence limits for interpretation of true change in densitometry practice. Therefore, it is important for reanalysis of DPX data files with the DPX-IQ to be performed so that longitudinal changes in BMD can be accurately assessed.