Next-generation DNA sequencing (NGS) is rapidly becoming an indispensable tool for genome-directed cancer diagnostics, but next-generation RNA sequencing (RNA-seq) is currently not standardly used in clinical diagnostics for expression assessment. However, multi-gene RNA diagnostic assays are used increasingly in the routine diagnosis of early stage breast cancer. Two of the most widely used tests are currently available only as a central laboratory service, which limits their clinical use.
Agendia scientists evaluated the use of RNA-seq as a de-centralized methodology to perform such tests. The MammaPrint and BluePrint RNA-seq tests were found to be equivalent to the clinically validated microarray tests. The RNA-seq tests were highly reproducible when performed in different locations, stable over-time. The MammaPrint RNA-seq test was clinically validated. These data demonstrate that RNA-seq can be used as a de-centralized platform, yielding results substantially equivalent to results derived from the predicate diagnostic device.
A: Expression heatmaps of the MammaPrint 70 genes measured with NGS (left panel) and microarray (right panel) in 204 matched NGS and microarray patient samples. Samples are ordered based on their microarray MammaPrint Index indicated as scatterplot graph next to the heatmap. Genes are clustered based on their NGS expression. B: Expression of the BluePrint 80 genes measured with NGS (left panel) and with microarray (right panel) in 204 matched NGS and microarray patient samples. Samples are ordered based on their microarray BluePrint subtype indicated as color bar next to the heatmap (Luminal-type, blue; HER2-type yellow, Basal-type, red). Genes are clustered based on their NGS expression. Samples are showed in rows, genes are showed in columns. Genes are clustered using Hierarchical Clustering, Pearson correlation distance, and average linkage. Data are normalized, Log2 transformed and median centered prior visualization. The color bar underneath the heatmap reports the expression range (from -3 to +3: 70 genes; from -5 to +5: 80 genes). NGS=Next-Generation Sequencing.