Flow diagram of exploring RPLS dichotomous classification

Baseline characteristics of training cohort and validation cohort

Cell cycle, DNA damage and repair, and metabolism are dysregulated in RPLS

Volcano plot of the DEGs in 8 normal vs 8 RPLS tissues (A). Venn diagram showed shared genes between DEGs and prognostic genes (B). GSEA analysis of RPLS tumors, including HALLMARK gene sets and REACTOME gene sets (C). Circular plots of the prognostic genes in GO, KEGG, and enrichWP (D).

RPLS subgroups (G1 and G2) based on cell cycle, DNA damage and repair, and metabolism

tSNE exhibited the subgroups (G1 and G2) of RPLS (A). Survival cures of OS (B) and DFS (C) in G1 and G2. The hierarchical clustering heatmap of dysregulated pathways in G1 and G2 (D). Histograms revealed the difference of pathological composition ratio (E), surgery times (F), and MDM2 (G) in G1 and G2. Violin plot of the microenvironmental scores in G1 and G2 (H).

RPLS classification strategy (C1 and C2) derived from RPLS subgroups

NMF for a re-classification of training cohort 1 (C1 and C2) (A). Survival cures of OS (B) and DFS (C) in C1 and C2. Histograms revealed the difference in pathological composition ratio (D), MDM2 (E), and surgery times (F) in C1 and C2.

RPLS dichotomous classification (Cluster_C1 and Cluster_C2) derived from RPLS clusters

Heatmap of biomarkers identified (LEP and PTTG1) in C1 and C2 (A). ROC curves of the machine learning models to identify C1 and C2 (B). Correlation between LEP and PTTG1 expression (C). Survival curves of OS (D) and DFS (E) in Cluster_C1 (low risk) and Cluster_C2 (high risk) groups. GSEA of HALLMARK gene sets in Cluster_C1 and Cluster_C2 (F). Histograms revealed the difference of pathological composition ratio (G), MDM2 level (H), and surgery times (I) in Cluster_C1 and Cluster_C2. Sankey diagram indicated the correlation among G1/G2, C1/C2, and Cluster_C1/C2 (J).

Validation of the RPLS dichotomous classification in another 241 RPLS cohort

Representative IHC staining images of LEP (A) and PTTG1 (B). Survival curves of OS (C) and DFS (D) in high-risk and low-risk groups.

Survival nomogram of LEP+PTTG1 model in validation cohort

The difference of surgery times (A), pathological composition (B), and surgery times (C) in high-risk and low-risk groups. Nomograms for OS was developed in REASR cohort with four factors: sex, age, risk score, and differentiation (D). ROC curves of 1-, 2-, and 3-year OS in validation cohort (E). Calibration curves of predicting 1-, 2-, and 3-year OS in validation cohort (F).