NGS and NanoString Platforms Play Complementary Role in Bedside Advanced Heart Failure Outcome Prediction Test Development

Purpose In order to create a clinical assay that preoperatively predicts 1-year survival status of AdHF patients following surgical/interventional therapies and to select the appropriate commercial platform, we compared the properties of Next Generation Sequencing (NGS) to NanoString (NS). Methods 8ml of blood was collected in CPT tubes from 25 AdHF patients (11 heart failure control, 4 mechanical circulatory support surgery, and 10 heart transplant surgery) at UCLA Medical Center 2015-2016. Purified RNA samples were aliquoted in two sets: one was processed using NGS transcriptome analysis (Illumina HiSeq 2500) and compared to 770 genes represented on NS pre-existing PanCancer IO 360 Gene Expression research panel. Prior to statistical analysis, expression values were log transformed. Pearson correlation coefficients were computed for each sample. Gene expression values were compared between NS and NGS across the set of matched genes and for each of the matched genes across the set of samples. Genes were grouped by average NGS expression, and the NS-NGS correlation for each group was computed. Results Out of 770 genes, 734 overlapped between both platforms and showed high intrasample correlation (R^2 = 0.82). Genes with higher expression demonstrated higher correlation across both platforms (Figure). NS demonstrated high robustness: Gene signature scores were similar across different RNA input concentrations, cartridges, and outcomes; sensitivity: Probe counts between 100 and 500 detected and quantified. The RNA input of 100 ng yielded the most precise result and were used for downstream analysis; and reproducibility: Technical replicates with equal concentration (100ng input) showed extremely high correlation (R^2 > 0.99), thus demonstrating high reproducibility. Conclusion Both platforms have meaningful, complementary roles, NGS for discovery and NanoString for commercialization and can be used for AdHF-biomarker test development.