Fig 2: Targeting CD49b delays the tumor growth in vivo(A) Representative images of human PDAC cell lines cultured on COLI or COLIV-coated plates, with or without vatelizumab (10 μg/mL), following a 2-h incubation.(B) Representative images of mouse KPC cells treated with or without vatelizumab (10 μg/mL) compared to anti mouse-CD49b antibody (10 μg/mL) on COL I and COL IV coated substrates, following a 2-h incubation. Scale bars, 600 μm.(C) Histogram showing human CD49b expression in KPCwt and KPC49bKO.(D) Representative images of mouse KPC cells expressing human CD49b treated with or without vatelizumab (10 μg/mL) on COL I- and COL IV-coated substrates after 2-h incubation. Scale bars, 600 μm.(E) C57BL/6J mice were subcutaneously inoculated with KPC49bKOhCD4b cells (1 × 106 cells). Tumors were allowed to grow before mice were randomly assigned to PBS or vatelizumab (200 μg) intratumoral treatment groups. Tumor growth was measured three times per week.(F) Bar graph shows immune cell infiltration in tumors, (G) C57BL/6J mice were subcutaneously inoculated with KPCwt cells (1 × 16 cells) on the right flank. Mice were randomly grouped to IgG control or amCD49b (200 μg) intratumoral treatment groups. Tumor growth was measured three times per week.(H) Bar graph shows immune cell infiltration in tumors. Statistical significance: (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001). Data are shown as mean ± SEM.

Fig 3: The expression of collagen receptors in pancreatic cancer(A) Log2 expression levels of collagen receptors in human PDAC cell lines based on CCLE RNA-seq data (RMA-normalized).(B) The percentage of total human PDAC cell lines stained with CD49b, CD29, CD49a, DDR1, and LAIR-1 by FACS.(C) Representative FACS histogram of KPC cell lines stained with CD49b, CD29, and CD49a.(D) Representative western blot of CD49b, CD29, and DDR1.(E–G) Collagen receptors expression across cell clusters in mouse scRNA-seq (GEO: GSE166298).(F) ITGA2 expression within various cell clusters.(G) Volcano blot of ITGA2 vs. ITGA1 clusters ((logFC) cutoff ±1).(H) Heatmap of scRNA-seq showing the expression of collagen receptors among cellular clusters in normal pancreas (GSE159343) vs. KPC mouse model (GSE166298).(I) C57Bl6J mice were inoculated with KPC cell line (Hy15549) in the right flank. (I) Bar graph of CD49a+ CD29+ and CD49b+ CD29+ bar graph in tumor, pancreas, and spleen.(J) Representative dot plot comparing the CD49b+ CD29+ on EPCAM+ cells in tumor and pancreas.(K) Representative immunofluorescence (IF) image of tumor stained with CD49b (yellow), Col1 (red), and DAPI (blue). Data are presented as means ± SD of at least three replicates. Statistical significance (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001).

Fig 4: Colocalization of CD49b and CK19 in human PDAC Tissue(A) Collagen receptors expression among various cell clusters from scRNA-seq datasets of treatment naive human PDAC patients (GEO: GSE166298 ((n = 16 patients); with matched adjacent normal pancreas (n = 3 patients) and GSE155698 (n = 18 treatment naive patients).(B) tSNE FACS clustering shows the collagen receptors expression in resected human PDAC specimens (adjacent-tumor and tumor).(C) Bar graph comparing the collagen receptors expression among the immune cells (CD45+), epithelial cells (EPCAM+), endothelial cells (CD45− EPCAM− CD31+), and other cells including CAFs (CD45− EPCAM− CD31−).(D) Kaplan-Meier survival analysis (TCGA, n = 177) stratified into two groups based on the median CD49b expression level. Log rank (Gehan-Breslow-Wilcoxon) test was used.(E) Spatial transcriptomics in PDAC patients (GSE211895) showing clusters of epithelium and stroma, ITAG2 is localized in epithelium clusters.(F) Immunofluorescence staining of human PDAC tissue CD49b (green) and CK19 (red). Scale bars, 250 μm.(G) Pearson’s correlation test between CD49b, KRT19 and ACTA2 (TCGA dataset). Data are presented as means ± SD of at least three replicates. Statistical significance (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001).

Fig 5: CD8+ T cells control tumor growth in CD49b knockout cell line(A) Tumor growth following the subcutaneous injection of KPCwt or KPC49bKO cells (1 × 106 cells) into immunodeficient nude mice (12 mice for each group).(B) Tumor growth in C57BL/6 mice with prior treatment with anti-IgG or anti-CD8 antibody (n = 5 tumors per group).(C) Pearson correlation analysis between collagen receptors expression and CD8a+ levels in the TCGA PDAC dataset.(D) Spatial transcriptions spots of KRT18, CD49b and CD8a in PDAC patients (GSE235315).(E) Donut chart showing the spatial spots that express CD49b+ CD8a+, CD49b+ CD8a−, CD49b−CD8a+, and CD49b− CD8a−.(F) The spots that express either CD49b+ CD8a−, CD49b−CD8a+ are correlated and represented as percentage per total spots for each patient.(G) HE, trichrome and multiplex immunofluorescent staining of naive PDAC specimen stained with aCD49b, aCD8, and COL1. Bar graph represents the quantification of CD8+ T cells and CD49b expression levels in both the tumor core and margin regions.