High-throughput identification of combinatorial ligands for DNA delivery in cell culture

Finding the optimal combinations of ligands for tissue-specific delivery is tedious even if only a few well-established compounds are tested. The cargo affects the receptor-ligand interaction, especially when it is charged like DNA. The ligand should therefore be evaluated together with its cargo. Several viruses have been shown to interact with more than one receptor, for efficient internalization. We here present a DNA oligonucleotide-based method for inexpensive and rapid screening of biotin labeled ligands for combinatorial effects on cellular binding and uptake. The oligonucleotide complex was designed as a 44 bp double-stranded DNA oligonucleotide with one central streptavidin molecule and a second streptavidin at the terminus. The use of a highly advanced robotic platform ensured stringent processing and execution of the experiments. The oligonucleotides were fluorescently labeled and used for detection and analysis of cell-bound, internalized and intra-cellular compartmentalized constructs by an automated line-scanning confocal microscope, IN Cell Analyzer 3000. All possible combinations of 22 ligands were explored in sets of 2 and tested on 6 different human cell lines in triplicates. In total, 10 000 transfections were performed on the automation platform. Cell-specific combinations of ligands were identified and their relative position on the scaffold oligonucleotide was found to be of importance. The ligands were found to be cargo dependent, carbohydrates were more potent for DNA delivery whereas cell penetrating peptides were more potent for delivery of less charged particles. © 2008 American Institute of Physics

Digital breast tomosynthesis (DBT): recommendations from the Italian College of Breast Radiologists (ICBR) by the Italian Society of Medical Radiology (SIRM) and the Italian Group for Mammography Screening (GISMa)

This position paper, issued by ICBR/SIRM and GISMa, summarizes the evidence on DBT and provides recommendations for its use. In the screening setting, DBT in adjunct to digital mammography (DM) increased detection rate by 0.5\u20132.7\u2030 and decreased false positives by 0.8\u20133.6% compared to DM alone in observational and double-testing experimental studies. The reduction in recall rate could be less prominent in those screening programs which already have low recall rates with DM. The increase in radiation exposure associated with DM/DBT protocols has been solved by the introduction of synthetic mammograms (sDM) reconstructed from DBT datasets. Thus, whenever possible, sDM/DBT should be preferred to DM/DBT. However, before introducing DBT as a routine screening tool for average-risk women, we should wait for the results of randomized controlled trials and for a statistically significant and clinically relevant reduction in the interval cancer rate, hopefully associated with a reduction in the advanced cancer rate. Otherwise, a potential for overdiagnosis and overtreatment cannot be excluded. Studies exploring this issue are ongoing. Screening of women at intermediate risk should follow the same recommendations, with particular protocols for women with previous BC history. In high-risk women, if mammography is performed as an adjunct to MRI or in the case of MRI contraindications, sDM/DBT protocols are suggested. Evidence exists in favor of DBT usage in women with clinical symptoms/signs and asymptomatic women with screen-detected findings recalled for work-up. The possibility to perform needle biopsy or localization under DBT guidance should be offered when DBT-only findings need characterization or surger