PhD Scientific Days 2025

Poster Session I. - A: Molecular Medicine

Exploring the Colorectal Cancer Immune Microenvironment with Patient-Derived Organoids

Name of the presenter

Hajdó Szabolcs

Institute/workplace of the presenter

Department of Genetics, Cell- and Immunobiology

Authors

Szabolcs Hajdó¹, Idan Carmi¹, Barbara Molnár-Érsek¹, Zoltán Wiener¹

1: Department of Genetics, Cell- and Immunobiology, Semmelweis University

Text of the abstract

Introduction: Colorectal cancer (CRC) is the third most frequent cancer type, and is a major cause of cancer-related death. Most CRC patients do not benefit from immunotherapy because of the reduced immune cell activity in the tumor microenvironment (TME). NK cells are one of the important cytotoxic cells with the capacity of direct tumor cell killing and IFN-gamma secretion. The most aggressive CRC subtype (CMS4) is characterised by fibroblast accumulation and an immunosuppressive milieu.

Aims: Our study aimed to establish a complex co-culture system with patient-derived CRC organoids (PDOs), fibroblasts, and NK cells to characterize the resulting tumor-immune cell interaction in this 3D in vitro model, and to identify mechanisms leading to immunosuppression.

Methods: NK cells from donor PBMCs were sorted. Isolated NK cells and/or cancer-associated fibroblasts (CAF) were incorporated into different co-culture systems, functional assays were performed and analyzed by flow cytometry, ELISA, and confocal microscopy. In the co-cultures, PDOs were plated inside extracellular matrix (ECM) containing gel droplets or on top of ECM coatings.

Results: NK cells were able to kill colorectal cancer cells, although the cytotoxic activity was modest. The killing activity was enhanced in the presence of IL-2. We demonstrated that NK cells invade the ECM droplets and the PDOs cultured inside them. Incorporation of CAFs in the co-cultures decreased both the cytotoxic activity and the IFN-gamma secretion of NK cells. Of note, we detected several immune checkpoint molecules such as PD-L1, HVEM, and CD155 both on PDO cells and CAFs.

Conclusions: Our findings confirm that NK cells can kill colorectal cancer cells in PDOs. Our model also highlights that fibroblast accumulation in the tumor stroma, a major feature of the most aggressive CRC subclass, is associated with a dampened NK cell killing and secretory activity. Further investigations are needed to elucidate cellular interactions in the CRC immune microenvironment that influence NK cell function.

Funding: K137554, 2024-1.2.3-HU-RIZONT-2024-00003, TKP2021-EGA-24 (NRDI Office, Hungary), SE250+ PhD Scholarship to Sz.H. Ethical permission: TUKEB 2015, 51323-4/2015/EKU.