6 research outputs found
In Silico Enhanced Restriction Enzyme Based Methylation Analysis of the Human Glioblastoma Genome Using Agilent 244K CpG Island Microarrays
Genome wide methylation profiling of gliomas is likely to provide important clues to improving treatment outcomes. Restriction enzyme based approaches have been widely utilized for methylation profiling of cancer genomes and will continue to have importance in combination with higher density microarrays. With the availability of the human genome sequence and microarray probe sequences, these approaches can be readily characterized and optimized via in silico modeling. We adapted the previously described HpaII/MspI based Methylation Sensitive Restriction Enzyme (MSRE) assay for use with two-color Agilent 244K CpG island microarrays. In this assay, fragmented genomic DNA is digested in separate reactions with isoschizomeric HpaII (methylation-sensitive) and MspI (methylation-insensitive) restriction enzymes. Using in silico hybridization, we found that genomic fragmentation with BfaI was superior to MseI, providing a maximum effective coverage of 22,362 CpG islands in the human genome. In addition, we confirmed the presence of an internal control group of fragments lacking HpaII/MspI sites which enable separation of methylated and unmethylated fragments. We used this method on genomic DNA isolated from normal brain, U87MG cells, and a glioblastoma patient tumor sample and confirmed selected differentially methylated CpG islands using bisulfite sequencing. Along with additional validation points, we performed a receiver operating characteristics (ROC) analysis to determine the optimal threshold (pāā¤ā0.001). Based on this threshold, we identified ā¼2,400 CpG islands common to all three samples and 145 CpG islands unique to glioblastoma. These data provide general guidance to individuals seeking to maximize effective coverage using restriction enzyme based methylation profiling approaches
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Noninterruptive Clinical Decision Support Decreases Ordering of Respiratory Viral Panels during Influenza Season
ObjectiveA growing body of evidence suggests that testing for influenza virus alone is more appropriate than multiplex respiratory viral panel (RVP) testing for general populations of patients with respiratory tract infections. We aimed to decrease the proportion of RVPs out of total respiratory viral testing ordered during influenza season.MethodsWe implemented two consecutive interventions: reflex testing for RVPs only after a negative influenza test, and noninterruptive clinical decision support (CDS) including modifications of the computerized physician order entry search behavior and cost display. We conducted an interrupted time series of RVPs and influenza polymerase chain reaction tests pre- and postintervention, and performed a mixed-effects logistic regression analysis with a primary outcome of proportion of RVPs out of total respiratory viral tests. The primary predictor was the intervention period, and covariates included the provider, clinical setting, associated diagnoses, and influenza incidence.ResultsFrom March 2013 to April 2019, there were 24,294 RVPs and 26,012 influenza tests (nā=ā50,306). Odds of ordering an RVP decreased during the reflex testing period (odds ratio: 0.432, 95% confidence interval: 0.397-0.469), and decreased more dramatically during the noninterruptive CDS period (odds ratio: 0.291, 95% confidence interval: 0.259-0.327).DiscussionThe odds of ordering an RVP were 71% less with the noninterruptive CDS intervention, which projected 4,773 fewer RVPs compared with baseline. Assuming a cost equal to Medicare reimbursement rates for RVPs and influenza tests, this would generate an estimated averted cost of $1,259,474 per year.ConclusionNoninterruptive CDS interventions are effective in reducing unnecessary and expensive testing, and avoid typical pitfalls such as alert fatigue