Annexin A1 Released in Extracellular Vesicles by Pancreatic Cancer Cells Activates Components of the Tumor Microenvironment, through Interaction with the Formyl-Peptide Receptors

Abstract

Pancreatic cancer (PC) is one of the most aggressive cancers in the world. Several extracellular factors are involved in its development and metastasis to distant organs. In PC, the protein Annexin A1 (ANXA1) appears to be overexpressed and may be identified as an oncogenic factor, also because it is a component in tumor-deriving extracellular vesicles (EVs). Indeed, these microvesicles are known to nourish the tumor microenvironment. Once we evaluated the autocrine role of ANXA1-containing EVs on PC MIA PaCa-2 cells and their pro-angiogenic action, we investigated the ANXA1 paracrine effect on stromal cells like fibroblasts and endothelial ones. Concerning the analysis of fibroblasts, cell migration/invasion, cytoskeleton remodeling, and the different expression of specific protein markers, all features of the cell switching into myofibroblasts, were assessed after administration of wild type more than ANXA1 Knock-Out EVs. Interestingly, we demonstrated a mechanism by which the ANXA1-EVs complex can stimulate the activation of formyl peptide receptors (FPRs), triggering mesenchymal switches and cell motility on both fibroblasts and endothelial cells. Therefore, we highlighted the importance of ANXA1/EVs-FPR axes in PC progression as a vehicle of intercommunication tumor cells-stroma, suggesting a specific potential prognostic/diagnostic role of ANXA1, whether in soluble form or even if EVs are captured in PC.

ANXA1 Contained in EVs Regulates Macrophage Polarization in Tumor Microenvironment and Promotes Pancreatic Cancer Progression and Metastasis

“…In addition, in this case, we used as control the endothelial cells in the presence of their own growth medium (ctrl) and with HUVEC medium: macrophage growth medium 1:1 (ctrl sup). The experimental point with Ac2-26 was not explained because it was already shown in our previous study [34]. These results highlighted by the fluorescence images have been also proved by the histograms representing the densitometry analysis performed has reported in the Material and Methods section (Figure 4G).…”

“…MIA PaCa-2 cells (ATCC ® CRL-1420; Manassas, VA, USA) were grown as reported in [34]. ANXA1 knockout (KO) MIA PaCa-2 cells were generated as described in [15] and kept in selection by 700 µg/mL neomycin (Euroclone; Milan, Italy).…”

“…These latter (40-100 nm diameter) are small membranous extracellular vesicles (EVs) released from nonmalignant and malignant cells in the extracellular space and body fluids [21,[25][26][27][28][29][30]. EVs play a central role promoting cell-to-cell communication, cell proliferation, migration and invasion and are involved in organizing premetastatic niches as key modulators of the tumor microenvironment [24,[31][32][33][34][35]. Moreover, previous studies have shown that vesicles regulate the interactions between tumor and immune cells, such as TAMs [32,33].…”

Annexin A1 Expression Is Associated with Epithelial–Mesenchymal Transition (EMT), Cell Proliferation, Prognosis, and Drug Response in Pancreatic Cancer

“…In addition to its relationship with cancer cells, ANXA1 expression is also associated with multiple cells in the TME, such as fibroblasts, and, with angiogenesis, the generation of new vessels and metastasis [16,17]. Novizio et al reported that the ANXA1 extracellular vesicle (EV) complex participates in tumor cells-stroma intercommunication as a vehicle during PC progression, suggesting that ANXA1 may have potential prognostic and diagnostic roles [18].…”

The Cellular and Biological Impact of Extracellular Vesicles in Pancreatic Cancer

“…However, tumor cells were required to already be present at the future metastatic site in order for EVs to be able to enhance PSC migration and recruitment, as EVs are not a source of PDGFb [ 66 ]. Besides miRNAs and TGFb, Novizio et al reported that Annexin A1 released in PCC-EVs drives a mesenchymal switch and turns fibroblasts into myofibroblasts through Formyl Peptide Receptors (FPRs) activation [ 67 ]. Tumor cell-derived EVs isolated from PDA patient sera, as with other carcinoma cell lines, mediate the transfer of mRNA, such as that of telomerase—hTERT mRNA—to non-telomerase expressing somatic cells, fibroblasts, marking them with augmented proliferation and delayed senescence, as well as protection from DNA damage and subsequent apoptosis.…”