Fig. 1: MS analysis of mutp53-dependent secreted proteins in human PDAC cells.

A Differentially secreted proteins detected by MS analysis in the CM of PANC-1 cells transiently transfected with siTP53^R273H or siScramble. The 25 proteins significantly downsecreted are indicated by the light lavender-shaded rectangle underlaid on the plot. Vertical dashed lines indicate log2 fold change = ±0.5. Horizontal dashed line indicates the cut-off p-value p = 0.05. TPM1, CSTN1, HMGA1 and CP2R1 proteins are marked on the plot. B Differentially secreted proteins detected by MS analysis in the CM of AsPC-1 cells transiently transfected with TP53^R273H or MOCK plasmid. The 83 proteins significantly hypersecreted are indicated by the light teal-shaded rectangle underlaid on the plot. Among them, TPM1, CSTN1, HMGA1 and CP2R1 proteins are specifically highlighted. Vertical dashed lines indicate log2 fold change = ±0.5. Horizontal dashed line indicates the cut-off p-value p = 0.05. C Venn diagram indicating the overlaps of the differentially mut or wt p53-dependent secreted proteins detected by MS analysis. D Histogram showing 4 proteins (Tropomyosin 1 (TPM1), Calsyntenin 1 (CSTN1), High Mobility Group A1 (HMGA1), Cytochrome P450 Family 2 Subfamily R Member 1 (CP2R1)) hypersecreted by AsPC-1 (p53-null) cells overexpressing TP53^R273H and downsecreted by PANC-1 cells after KD of the same hot-spot mutp53 isoform. E UMAP visualization of all identified cell types present in the pancreatic microenvironment subset by disease state: Adjacent Normal (n = 3), Healthy (n = 6) and Tumor (n = 16). Data source: Pancreatic Tissue Single Cell Atlas. F UMAP visualizations showing HMGA1 expression across the major cell populations subset by disease state (Adjacent Normal, Healthy, Tumor). Data source: Pancreatic Tissue Single Cell Atlas. G HMGA1 gene expression level in tumor derived epithelial cells compared to adjacent normal or healthy epithelial cells (number of cells: 892 Adj.Normal; 14,380 Healthy; 9484 Tumor). Data source: Pancreatic Tissue Single Cell Atlas. H HMGA1 gene expression level in tumor compared to normal tissue. Data source: GEPIA database. * p < 0.01. I HMGA1 expression correlates with the mutational status of TP53 (n = 66 no-mutation, n = 62 missense mutations). Data source: cBioportal database, TCGA PanCancer Atlas. (Wilcoxon test). **** p < 0.0001. J HMGA1 expression level in PDAC patient with TP53^mut (n = 24 no-mutation, n = 43 missense mutations Data source: cBioportal database, QCMG Nature 2016. (Wilcoxon test). **** p < 0.0001. K Kaplan-Meier (KM) plot of survival probability (log-rank test) for PDAC patients only as obtained from KM-plotter database using the default parameters. L Kaplan-Meier survival plot after PDAC patients’ stratification for tumor stage (S2, S3, S4) in KM plotter database showing HMGA1 expression is a prognostic factor in advanced pancreatic cancer. M Volcano plot of differential gene expression (DGE) analysis for metastatic vs primary tumors using the microarray dataset GSE71729 showing HMGA1 is significantly highly expressed in metastatic patients (Log2FC = 2.383837; −log10(Adj. p-value) = 10.47756). Red highlights indicate significant regulated genes; black highlights indicate non-significant genes. Vertical dashed lines indicate log2 fold change = ±1. Horizontal dashed line indicates p = 0.01. N Immunoblot validation of HMGA1 KO in human PANC-1 cell line. KO denotes HMGA1 KO cells, while the minus sign (-) represents the parental cells. Vinculin was used as a loading control. O Representative images, with corresponding magnifications, of PANC-1 (top) and HMGA1-KO PANC-1 (bottom) cells invading through Matrigel-coated transwell inserts (8 μm pore size) after 24 h of incubation at 37 °C. Right panel: bar plot quantification of the percentage of invasive cells. Data are presented as mean ± SD (n = 4). (Unpaired t-test). **p < 0.01. P Tumor volume (mm³) from subcutaneous injection of either PANC-1 or HMGA1 KO PANC-1 cells in immunodeficient mice. Data plotted are mean tumor volumes + SEM (n = 6 for each cohort). (Two-way ANOVA). ****p < 0.0001. Q Individual tumor volumes + SEM at the endpoint (Unpaired t-test). *p < 0.05.