Extended Data Fig. 5: Stratification of rectal adenocarcinomas using the consensus molecular subtypes (CMS) classification.

(a) Expression levels for selected genes stratified by CMS group. Genes were annotated using the signatures from Budinska et al.⁷⁰. (b) TMB stratified by CMS groups. Sample sizes are: CMS1 (n = 11), CMS2 (n = 26), CMS3 (n = 26), and CMS4 (n = 38). (c) FGA stratified by CMS groups. Sample sizes are: CMS1 (n = 11), CMS2 (n = 26), CMS3 (n = 26), and CMS4 (n = 38). (d) Percentage of KRAS mutated tumors by CMS group. (e) ssGSEA scores for selected pathways from the Hallmark dataset35. Sample sizes are: CMS1 (n = 11), CMS2 (n = 26), CMS3 (n = 26), and CMS4 (n = 38). (f) DFS for LARC patients treated with NAT, stratified by CMS group. (g) Levels of CA9 gene expression as a function of KRAS and PIK3CA mutational status. Double mutants and KRAS-mutant tumors had significantly higher expression of CA9 compared to wild-type tumors, p = 1.3e-07 and p = 4.65e-05, respectively. Sample sizes are: Double-mutant (n = 8), KRAS-mutant (n = 26), PIK3CA-mutant (n = 6), and wild-type (n = 5). Statistical significance was assessed using a two-sided Mann–Whitney U-test. (h) Expression of L1CAM stratified by CMS group. L1CAM expression was higher in CMS2 and CMS4 compared to CMS3, q = 0.0498 and q = 0.096, respectively. Sample sizes are: CMS1 (n = 11), CMS2 (n = 26), CMS3 (n = 26), and CMS4 (n = 38). (i) Validation of transcriptomic findings using an independent cohort of 15 LARC cases from Kamran et al.¹⁰ Differential gene expression was conducted using DESeq2 and the p-values attained by the Wald test were corrected using false discovery rates. In panels (B), (C), (E) and (H), statistical significance was assessed using a two-sided Mann–Whitney U-test. P values were corrected using the Bonferroni method and significant results are denoted as *q < 0.05, **q < 0.01, ***q < 0.005 and ****q < 0.001. In panels (B), (C), (E), (G), and (H), boxplots’ center lines indicate medians, edges indicate the interquartile range, and the whiskers extend to the highest and lowest values not considered outliers.