BET inhibition modulates the pancreatic stromal cell landscape by inducing shifts in the polarization of cancer-associated fibroblasts. Open Access
Minocha, Trisha (Spring 2024)
Abstract
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is the most malignant neoplasm of the pancreas, and is refractory to most therapeutic strategies, including immunotherapy. We have previously demonstrated that loss of CDKN2A in PDAC tumor cells promotes an immunologically cold tumor microenvironment, which can be counteracted through the use of bromodomain and extraterminal domain inhibitors (BETi). The immunosuppressive PDAC tumor microenvironment is also characterized by a dense desmoplastic stroma, rich in cancer associated fibroblasts (CAFs). These CAFs are a highly plastic population of cells, composed of myofibroblast, inflammatory, and antigen presenting CAFs (myCAFs, iCAFs, and apCAFs, respectively). Previous research has suggested BETi can alter the function of CAFs, suggesting this therapeutic strategy may be effective at targeting both the tumor and stromal compartments of PDAC. Here, we hypothesize that BETi enhances anti-tumor immunity in CDKN2A-deficient PDAC by modulating the phenotype and function of PDAC CAFs.
METHODS: KPCluc-derived murine CAF cells were cultured into spheroids and used to evaluate the effects of BETi on CAF phenotype. CAF populations were evaluated by flow cytometry and the gene expression profiles of sorted CAFs were obtained through NanoString analysis. Lastly, signaling within CAF cells was evaluated via Western blot.
RESULTS: Direct treatment of CAFs with BETi induced a dramatic dose- and time-dependent polarization of CAF spheroids towards a myCAF phenotype. However, while BETi induced p-NF-κB expression CAF cells, this signaling was not found to contribute towards myCAF polarization. Further analysis of CAF gene expression profiles revealed that BETi reduced the expression of genes involved in establishing a stiff, immunosuppressive environment within the tumor, suggesting BETi's potential to re-program tumor architecture and promote immune accessibility.
CONCLUSIONS: BET inhibition demonstrates the potential to modulate CAF cell phenotype and mitigate the immunosuppressive nature of PDAC tumors. Our data suggest BETi is a rational treatment approach to re-model the pancreatic stromal cell landscape and serves the potential to increase the efficacy of the immunotherapy in this genetically defined immunotherapy-resistant patient population.
Table of Contents
INTRODUCTION----------------------------------------------------------------------------------------1
METHODS---------------------------------------------------------------------------------------------- 8
Cell culture and drug treatments------------------------------------------------------------------------- 8
Western blotting ---------------------------------------------------------------------------------------- 8
Spheroid treatment, flow cytometry analysis and cell sorting----------------------------------------------8
Statistical Methods-------------------------------------------------------------------------------------- 9
RESULTS------------------------------------------------------------------------------------------------10
BETi promotes a myCAF phenotype among CAF cells-----------------------------------------------------10
BETi polarizes CAF cells towards the myCAF Phenotype in a time- and dose-dependent fashion-----------11
NF-κB and STING inhibition does not influence JQ1 driven CAF Cell Polarization------------------------ 13
NanoString analysis of sorted CAF cells------------------------------------------------------------------15
Differential gene expression upon JQ1 treatment in iCAFs------------------------------------------------ 16
Differential gene expression upon JQ1 treatment in myCAFs---------------------------------------------- 19
JQ1 treatment differentially impacts gene expression in iCAFs and myCAFs------------------------------- 21
DISCUSSION--------------------------------------------------------------------------------------------24
REFERENCES------------------------------------------------------------------------------------------- 30
FIGURES
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Table 1---------------------------------------------------------------------------------------------------18
Table 2--------------------------------------------------------------------------------------------------- 20
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BET inhibition modulates the pancreatic stromal cell landscape by inducing shifts in the polarization of cancer-associated fibroblasts. () | 2024-04-10 01:27:38 -0400 |
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