Figure 3. Immune profiling of pancreatic ductal adenocarcinoma (PDAC) orthotopic tumours in response to therapy.

(A) Timeline of triple treatment regimen (maraviroc, αPD1, and radiotherapy) following orthotopic injection of KPC-F cells. A total of 12 days post-orthotopic injection of 500 KPC-F cells in the pancreas of wildtype C57BL/6 mice, mice were treated as follows: 7 consecutive days of 10 mg/kg intraperitoneal injection of maraviroc and 4 alternating days of 10 mg/kg intraperitoneal injection of αPD1. Mice were followed for up to 30 days following the start of the treatment regimen. Tumor volumes were measured by MRI and growth curves of individual treatment groups are plotted with or without radiotherapy as measured by MRI. Average growth curves ± SD are depicted in bold, individual mice are shaded (without IR; dashed, with IR; solid). Insert: expanded view of triple combination to show ‘responders’ display a significant benefit over RT alone. (B) Quantification of pancreatic tumors derived from (A) stained by IHC for p53. (C) Quantification of necrotic areas in pancreatic tumors derived from (A) based on H&E staining. (D) Quantification of infiltrating CD8 T-cells in pancreatic tumors derived from (A) by flow cytometry. (E) Profiling of infiltrating immune cells in pancreatic tumors derived from (A) by flow cytometry as in Figure 2B. Single, live cells were included for analysis and are represented as frequencies of Live/CD45⁺ cells or total CD3⁺ for FoxP3⁺ Tregs. Significance was tested using the Welch and Brown-Forsythe ANOVA for parametric data. ^#p<0.05, ^##p<0.01, or Kruskal-Wallis test for non-parametric data ^#p<0.01; pairwise comparisons (Student’s t-test). *p<0.05, **p<0.01, ***p<0.005, ****p<0.001.

Figure 3—figure supplement 1. Accessory data to support pancreatic ductal adenocarcinoma (PDAC) tumour model immune monitoring.

(A) Representative images of cross-sectional anatomy of a mouse abdomen, obtained through respiratory gated MRI (left), SARRP (center; labeled CT), and an overlay of the MRI and CT images for tumor-specific dosage of 4 Gy (right). (B) Gross tumor volume at day 30 post-treatment as measured by MRI of mice as treated in Figure 3A. Data are represented as mean tumor volume ± SD. (C) Quantitative measurement of DAPI⁺ cells based on H&E staining of mice treated as in Figure 3A. (D) Percentage of αSMA⁺ cells based on IHC. (E) Quantification of IHC slides showing the percentage of CD8⁺ T-cell infiltrating the tumor (left). Tumor borders are marked as dashed lines. T=tumor, NT=non-tumor (right). (F) Profiles of infiltrating immune cells in pancreatic tumors by all treatments including single control treatments as measured by Aurora Cytek. Single, live cells were included for analysis and represented as frequencies of CD45⁺ cells in percentage. *p<0.05, **p<0.01, ***p<0.005.