Page 116 - SRPSKO DRUŠTVO ISTRAŽIVAČA RAKA

Serbian Association for Cancer Research SDIRSACR

induced tumor-derived exosomes (hiTDExs) from pharyngeal cancer cells.
Material and Methods: Protein profile changes in human umbilical vein endothelial cells (HUVECs) treated with hiTDExs
were analyzed using a human cytokine antibody array assay. The expression of the target proteins was examined in
HNSCC patient tissues with and without metastasis.
Results: hiTDExs significantly modulated cytokine expression in HUVEC-derived secretomes. Array analysis revealed
at least 10 cytokines with altered expression compared to normoxic exosome-treated controls. Notably, CCL26, EGFR,
IL-3, and IL-8 expression was significantly altered. The functional role of these factors in remodeling the pharyngeal
cancer microenvironment, particularly in promoting metastasis and hypoxia, requires further investigation.
Conclusions: Our findings offer novel insights into how dynamic interactions between endothelial cells and hiTDExs
drive metastasis. While chemokine-based therapies remain under development, CCL26 emerges as a promising target
for future HNSCC treatments.

Acknowledgments and funding: This work was supported by the Scientific and Technological Research Council of
Turkey (TUBITAK), Grant/Award Number: 123Z593.

P20

Exosomes as Key Mediators of Fibroblast Reprogramming into CAFs via Oncogenic miRNA Transfer and
Activation of Cancer-Related Pathways

Milica Jaksic Karisik, Milos Lazarevic, Dijana Mitic, Jelena Carkic, Maja Milosevic Markovic, Jelena Milasin

Department of Human Genetics, School of Dental Medicine, University of Belgrade, Belgrade, Serbia

Keywords: exosomes, fibroblasts, miR-21, oral cancer, tumor microenvironment

Background: The tumor microenvironment is shaped by complex interactions between tumor cells and
stromal cells, among which fibroblasts play a key role. Exosomes, as important mediators of intercellular
communication, transfer regulatory molecules—including microRNAs—that can modulate the behavior of
recipient cells. This study aimed to investigate the effects of exosomes derived from healthy, premalignant,
and malignant oral epithelial cells on the expression of key oncogenic signaling pathways in fibroblasts.
Methods: Exosomes were isolated from cell lines of healthy epithelium (HaCaT), dysplastic cells (DOK), and oral
squamous cell carcinoma (SCC15 and SCC25). Expression of miR-21 was analyzed in exosomes from each group,
followed by treatment of normal fibroblasts with these exosomes for 24 hours. Total RNA was extracted from fibroblasts
using the Trizol method, and expression of genes involved in carcinogenesis (PIK3CA, AKT, NOTCH1, HES1) was
analyzed by qPCR. Fibroblasts without treatment served as the reference control for gene expression normalization.
Results: MiR-21 expression was significantly higher in exosomes derived from malignant cells compared to
those from healthy and premalignant cells, while no significant difference was observed between healthy
and dysplastic cells. Fibroblasts treated with cancer-derived exosomes showed a significant upregulation of
PIK3CA(p=0.041), AKT (p<0.0001), NOTCH1 (p=0.0003), and HES1 (p<0.0001) compared to the healthy exosome-
treated group. Treatment with dysplastic cell-derived exosomes resulted in a significant increase in AKT (p=0.04)
and HES1 (p=0.01) expression. No changes were observed in fibroblasts treated with exosomes from healthy cells.
Conclusions: Exosomes from malignant and premalignant epithelial cells induce activation of oncogenic signaling
pathways in fibroblasts, with miR-21 potentially playing a key role in fibroblast reprogramming toward a pro-tumorigenic
phenotype.

Acknowledgments and funding: This research was funded by the Science Fund of the Republic of Serbia, #GRANTNO
7750038, ORAL CANCER—NEW APPROACHES IN PREVENTION, CONTROL, AND POSTOPERATIVE REGENERATION—AN
IN VITRO STUDY—ORCA–PCR.

101