(A) Forty-eight genes (outlined by thick black lines) from the Venn diagram shown in Figure 3A were upregulated in quiescent clusters across 3 of 4 samples and selected as the ancestor-like CSC signature genes. (B) UMAP visualization of scRNA-Seq data from PDX models shown in Figure 1B colored using GSVA score of the ancestor-like CSC signature. (C and D) UMAP visualization of single-nucleus RNA-Seq data from NAC-sensitive or NAC-resistant TNBC cancer tissues shown in Figure 1D colored using GSVA score for the ancestor-like CSC signature and compared between pre- and mid-/post-treatment subgroups (C). Wilcoxon’s rank sum test (D) was used to determine significant P value. (E) Kaplan-Meier survival analysis between high (>0) and low (<0) subgroups of the ancestor-like CSC signature score in METABRIC cohort of breast cancer patients who received chemotherapy (not hormone therapy; n = 213) or belonged to TNBC subtype (n = 299). OS, overall survival; PFS, progression-free survival. (F) Kaplan-Meier survival analysis between NRP1^hiFXYD3^lo and NRP1^hiFXYD3^hi groups in METABRIC cohort of TNBC breast cancer patients. n = 149. Medians were used for cutoff value. P value was obtained using log-rank test. (G) Representative images of H&E staining and immunofluorescence staining of paired tumor samples of pre- and post-NAC from patients with TNBC, using antibodies against FXYD3 and NRP1 or IGF1R. Nuclei were stained using DAPI. Yellow arrows indicate cells double-positive for NRP1 and FXYD3 (top) and cells double-positive for IGF1R and FXYD3 (bottom). Scale bars: 20 μm. (H) Quantification of the ratio (percent) of the cells double-positive for NRP1 and FXYD3 to total NRP1-positive cells (top) and the cells double-positive for IGF1R and FXYD3 to total IGF1R-positive cells (bottom). n = 5 random fields were collected for each condition. Statistical significance was determined by 2-tailed Mann-Whitney U tests. Results are shown as means ± SEM.