GSVA methods outline. The input for the GSVA algorithm are a gene expression matrix in the form of log2 microarray expression values or RNA-seq counts and a database of gene sets. 1. Kernel estimation of the cumulative density function (kcdf). The two plots show two simulated expression profiles mimicking 6 samples from microarray and RNA-seq data. The x-axis corresponds to expression values where each gene is lowly expressed in the four samples with lower values and highly expressed in the other two. The scale of the kcdf is on the left y-axis and the scale of the Gaussian and Poisson kernels is on the right y-axis. 2. The expression-level statistic is rank ordered for each sample. 3. For every gene set, the Kolmogorov-Smirnov-like rank statistic is calculated. The plot illustrates a gene set consisting of 3 genes out of a total number of 10 with the sample-wise calculation of genes inside and outside of the gene set. 4. The GSVA enrichment score is either the maximum deviation from zero (top) or the difference between the two sums (bottom). The two plots show two simulations of the resulting scores under the null hypothesis of no gene expression change (see main text). The output of the algorithm is a matrix containing pathway enrichment scores for each gene set and sample.