Differential expression of IFN-γ and IFN-α in EOT compared to pre-treatment samples in all cohorts; hierarchical clustering based on genes differentially expressed at EOT compared to pre-treatment samples in each treatment cohort and dendrogram showing the degree of relatedness of samples based on imiquimod-induced genes in the blue group. The 2-ΔΔCT describes (a) IFN-γ and (b) IFN-α gene expression fold change at EOT relative to baseline after normalization according to the endogenous reference cyclophilin G. CT equals the mean cycle times of duplicate wells and ΔΔCT = (CT, Target-CT, cyclophilin) EOT - (CT, Target-CT, cyclophilin) baseline. The fold-change data were transformed using logarithm10. The box and whisker style box plot gives the median and interquartile range (box), 1.5 of the inter-quartile range (whiskers), points outside the whiskers (square symbols) and the mean (cross symbol). Statistics: p values refer to 2-sample t-tests between treatment and control groups. (c) Based on a paired t-test cut-off p2 value < 0.05, 1,311 genes were differentially expressed between the pre-treatment and EOT samples in the q12 × 2 (orange) cohort. Reclustering of these genes identified a node of 65 genes uniquely upregulated in the imiquimod-treated EOT samples (part i). Similar analyses were performed for the other imiquimod-treated cohorts; 1,578 genes were differentially expressed in the q12 × 4 (blue) cohort, including an imiquimod-specific node of 263 genes (part ii and the vertical blue bar in adjacent complete data set); 650 genes were differentially expressed in the q24 × 4 (green) cohort, including an imiquimod-specific node of 58 genes (part iii); and 495 genes were differentially expressed in the q24 × 8 (pink) cohort, including an imiquimod-specific node of 23 genes (part iv). A Venn diagram displays the extent of overlap among genes differentially expressed in the three most informative orange, blue and green groups (part v). (d) Reclustering of all BCC samples based on the imiquimod-specific 263-gene signature identified in the q12 × 4 (blue) cohort. Straight lines identify imiquimod-treated EOT samples color coded according to treatment regimen; dashed lines identify vehicle cream-treated EOT samples and unlabeled are the all pre-treatment samples. A diagram illustrating the strategy used to prepare Figure 1c,d is available as Additional data file 4.