The pooled prevalence values provided from this study provide a basis for an evidence-based approach to the management of children with this frequently occurring condition.

A 15-gene prognostic signature for early-stage, completely resected, non-small-cell lung carcinoma, (which distinguishes between patients with good and poor prognoses) was clinically validated in prior studies. To achieve operational efficiencies, this study was designed to evaluate the assay's performance in RNA-stabilized tissue as an alternative to the fresh-frozen tissue format originally used to develop the assay. The percent concordance between matched tissue formats was 84% (95% Wilson CI, 70%-92%), a level of agreement comparable to the inherent reproducibility of the assay observed within biological replicates of fresh-frozen tissue. Furthermore, the analytical performance of the assay using the RNA-stabilized tissue format was evaluated. When compared to an accredited reference laboratory, the clinical laboratory achieved a concordance of 94% (95% Wilson CI, 81%-98%), and there was no evidence of bias between the laboratories. The lower limit of quantitation for the target RNA concentration was confirmed to be, at most, 12.5 ng/μL. The assay reportable range defined in terms of risk score units was determined to be -4.295 to 4.210. In a large-scale precision study, the assay showed high reproducibility and repeatability. When subjected to a maximal amount of genomic DNA, a potential contaminant, the assay still produced the expected results. The 15-gene signature was confirmed to produce reliable results and, thus, is suitable for its intended use.

A formalin-fixed paraffin-embedded tissue-based prognostic assay to assess the risk for recurrence in stage II colon cancer has recently been clinically validated. This study describes the analytical performance and quality control measures of the assay. The reportable range was determined to be [-1.129, 1.414] in risk score units. The accuracy was evaluated with a split sample comparison within the production lab and between the production lab and a reference lab. The concordance between the replicates within the production lab was 79% (95% confidence interval, 64%-91%). There was no evidence of bias, and the concordance was 78% (95% confidence interval, 61%-90%) between the labs. The lab-to-lab concordance was further evaluated by simulating risk scores from the full reportable range. The simulation suggested a higher concordance. The sensitivity study demonstrated that the percentage of tumor tissue did not impact the risk score and that RNA concentration of 9.5 ng/μL was a conservative determination of the analyte lower limit of quantification. From the precision study, the repeatability and reproducibility estimates were 0.1267 and 0.0548 in risk score units, respectively. Furthermore, multifaceted quality control measures were implemented, such as proper tissue processing steps, high-risk and low-risk controls, nontemplate control, and a gene expression-based classifier to evaluate the cDNA amplification kit, a key reagent in the assay. In conclusion, this study demonstrates the strong analytical performance of the assay and further supports its use as an objective standardized prognostic test for stage II colon cancer.

7532 Background: Recent clinical studies have demonstrated the benefit of adjuvant chemotherapy (ACT) in some early-stage non-small cell lung cancer (NSCLC) patients. A 15-gene signature, developed using fresh frozen (FF) tissue, has been shown to be an independent prognostic marker that identifies high risk patients who may benefit from ACT. Use of this signature in tissue preserved in an RNA stabilization reagent is desired for easier access to tumor tissue in the clinical setting. Methods: Matched FF and RNAlater-preserved (RNAL) tissues were obtained from 43 NSCLC patients. Patients provided written consent for the collection of tumor tissue at the time of surgery under an IRB approved protocol. Each tissue sample was split into 2 pieces, creating biological replicates for each tissue format. For each patient, RNA was extracted from 4 tissue pieces (2 FF, 2 RNAL), followed by microarray-based genomic profiling (Affymetrix U133 Plus 2.0). The 15-gene signature was applied to each profile, generating a numerical risk score and a risk category (high, low) using methods previously established (Zhu 2010 J Clin Oncol). The level of agreement was evaluated within biological replicates of each tissue format, as well as between the averaged biological replicates of matched FF and RNAL tissues. Results: The concordance in risk category between averaged biological replicates of matched FF and RNAL tissues was 84%, with a Pearson correlation of 0.74. This level of agreement is comparable to the inherent reproducibility of the assay observed within biological replicates of FF tissue, which demonstrated concordance of 79% and Pearson correlation of 0.83. In addition, a statistical in silico simulation was used to demonstrate that if the risk scores in this study had spanned the full dynamic range of the assay while maintaining the same level of inherent reproducibility observed in the current study, the level of concordance would be 89% with a Pearson correlation of 0.93. Conclusions: The level of agreement between matched FF and RNAL tissues is not inferior to that seen within FF biological replicates. Therefore, the 15-gene signature maintains its performance when used in RNAlater-preserved NSCLC tissues.