Extended Data Fig. 8: Effects of tamoxifen and E2 in mouse endometrial epithelial cells.

(a) Pathway enrichment analysis on the differently expressed genes identified by DEseq2 from comparing estradiol (E2) versus tamoxifen (Tam) in endometrial epithelial cells (Q < 0.01, Benjamini-Hochberg-corrected two-sided Wald test). Bar plot depicts the odds ratio of pathway enrichment (MSigDB oncogenic signatures); purple line indicates the Q-values from the Benjamini-Hochberg-corrected two-sided Fisher’s exact tests. (b) Venn diagram showing the genes upregulated after tamoxifen treatment compared to vehicle (veh) control, and genes upregulated after E2 compared to veh control (DEseq2, log₂FC > 1, Q < 0.01, Benjamini-Hochberg-corrected two-sided Wald test). (c–e) Pathway enrichment analysis (MSigDB oncogenic signatures) on: (c) genes upregulated with tamoxifen treatment versus vehicle that are not shared with the E2-upregulated genes (n = 962, as shown in the Venn diagram in b); (d) shared genes upregulated by tamoxifen treatment versus vehicle and E2 treatment versus vehicle; and (e) genes upregulated by E2 treatment versus vehicle but not upregulated by tamoxifen versus vehicle. Bar plots depict the odds ratio of pathway enrichment, Q-values from the Benjamini-Hochberg-corrected two-sided Fisher’s exact tests. (f) Comparison of differentially expressed genes (log₂ fold change [FC]) between tamoxifen (tam) over vehicle control (x-axis) and tamoxifen plus alpelisib (y-axis). Genes with FDR < 0.05 (DEseq2) are categorized and color-coded. (g) Representative immunohistochemistry images of ER expression (nuclear brown staining) in epithelial (black arrow) and stromal cells (red arrow) in the uteri from mice treated with vehicle control, estrogen (E2), and tamoxifen. Scale bars: 10μm, H&E counterstaining. Each treatment group represents independently repeated experiments with similar results: vehicle (n = 2 mice), E2 (n = 3 mice), and tamoxifen (n = 5 mice). (h) Heatmap of log₂ FPKM expression levels (scale 0-5) of genes related to Igf1r in mouse endometrial epithelial cells from vehicle (Veh), estradiol (E2), tamoxifen (Tam) and tamoxifen plus alpelisib (Tam+Alp) treatment cohorts.