Murine- and Human-Derived Autologous Organoid/Immune Cell Co-Cultures as Pre-Clinical Models of Pancreatic Ductal Adenocarcinoma
Abstract
:Simple Summary
Abstract
1. Introduction
2. Results
2.1. Increased PMN-MDCS Infiltration Correlated with Tumor Growth in Orthotopically Transplanted Mice
2.2. Organoids Derived from Cabozantinib-Treated Mouse Tumors Exhibit a Decreased Stromal Cell Compartment That Correlates with Increased CD8+ Cells
2.3. PMN-MDCSs Disrupt the Efficacy of Checkpoint Inhibition in Mouse-Derived Organoid/Immune Cell Co-Cultures
2.4. Generation of PDAC Patient-Derived Organoids and Orthotopic Transplantation
2.5. DSP Reveals a Correlation between Infiltrating PMN-MDSCs, Arg1 Expression and Increased Stroma in Patient Tissue
2.6. Depletion of PMN-MDSCs from Patient-Derived Organoid/Immune Cell Co-Cultures Maximizes the Effect of Anti-PD-1/PD-L1 Interaction
3. Discussion
4. Methods
4.1. Mouse Orthotopic Transplants and Treatment
4.2. Generation of Mouse- and Human-Derived Pancreatic Cancer Organoids
4.3. Extraction and Culture of Murine and Human Immune Cells
4.4. Human- and Mouse-Derived Pancreatic Cancer/Immune Cell Co-Cultures
4.5. Human- and Mouse- Derived MDSC-CTL Titration Assay
4.6. Testing Efficacy of Different MDSC Inhibitors Using Human- and Mouse-Derived MDSC-CTL Co-Culture
4.7. Immunofluorescence and Immunohistochemistry
4.8. Flow Cytometry
4.9. Quantitative RT-PCR (qRT-PCR)
4.10. NanoString Technologies Digital Spatial Profiling (DSP)
4.11. Luminex Assay
4.12. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
PDAC | Pancreatic Ductal Adenocarcinoma |
PD-l1 | Program Death Ligand 1 |
PD-1 | Program Death 1 |
MDSC | Myeloid-Derived Suppressor Cell |
PMN-MDSC | Polymorphonuclear MDSC |
M-MDSC | Monocytic MDSC |
CTL | Cytotoxic T- Lymphocyte |
DC | Dendritic Cell |
PanIN | Pancreatic Intraepithelial Neoplasia |
Treg | Regulatory T cell |
GZMB | Granzyme B |
SMA | Smooth Muscle Actin |
VEGF | Vascular Endothelial Growth Factor |
GMCSF | Granulocyte-Macrophage Colony Stimulating Factor |
TGFβ | Transforming Growth Factor beta |
CFSE | Carboxyfluorescein Succinimidyl Ester |
NSG | NOD Scid Gamma |
ROS | Reactive Oxygen Species |
ROI | Region of Interest |
RET | Ret Proto-Oncogene |
DSP | Digital Spatial Profiling |
FLT3 | fms-Like Tyrosine Kinase 3 |
IDO-1 | Indoleamine 2,3-Dioxygenase 1 |
VISTA | V-domain Ig Suppressor of T Cell Activation |
SLFN | Schlafen |
DPBS | Dulbecco’s Phosphate-Buffered Saline |
DMEM/F12 | Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12 |
FCS | Fetal Bovine Serum |
HBSS | Hank’s Balanced Salt Solution |
FGF-10 | Fibroblast Growth Factor 10 |
FGF-2 | Basic Fibroblast Growth Factor |
ATRA | All-Trans Retinoic Acid |
TIL | Tumor Infiltrating Lymphocytes |
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Holokai, L.; Chakrabarti, J.; Lundy, J.; Croagh, D.; Adhikary, P.; Richards, S.S.; Woodson, C.; Steele, N.; Kuester, R.; Scott, A.; et al. Murine- and Human-Derived Autologous Organoid/Immune Cell Co-Cultures as Pre-Clinical Models of Pancreatic Ductal Adenocarcinoma. Cancers 2020, 12, 3816. https://doi.org/10.3390/cancers12123816
Holokai L, Chakrabarti J, Lundy J, Croagh D, Adhikary P, Richards SS, Woodson C, Steele N, Kuester R, Scott A, et al. Murine- and Human-Derived Autologous Organoid/Immune Cell Co-Cultures as Pre-Clinical Models of Pancreatic Ductal Adenocarcinoma. Cancers. 2020; 12(12):3816. https://doi.org/10.3390/cancers12123816
Chicago/Turabian StyleHolokai, Loryn, Jayati Chakrabarti, Joanne Lundy, Daniel Croagh, Pritha Adhikary, Scott S. Richards, Chantal Woodson, Nina Steele, Robert Kuester, Aaron Scott, and et al. 2020. "Murine- and Human-Derived Autologous Organoid/Immune Cell Co-Cultures as Pre-Clinical Models of Pancreatic Ductal Adenocarcinoma" Cancers 12, no. 12: 3816. https://doi.org/10.3390/cancers12123816