Hodgkin Lymphoma-Derived Extracellular Vesicles Change the Secretome of Fibroblasts Toward a CAF Phenotype

Abstract

Secretion of extracellular vesicles (EVs) is a ubiquitous mechanism of intercellular communication based on the exchange of effector molecules, such as growth factors, cytokines, and nucleic acids. Recent studies identified tumor-derived EVs as central players in tumor progression and the establishment of the tumor microenvironment (TME). However, studies on EVs from classical Hodgkin lymphoma (cHL) are limited. The growth of malignant Hodgkin and Reed–Sternberg (HRS) cells depends on the TME, which is actively shaped by a complex interaction of HRS cells and stromal cells, such as fibroblasts and immune cells. HRS cells secrete cytokines and angiogenic factors thus recruiting and inducing the proliferation of surrounding cells to finally deploy an immunosuppressive TME. In this study, we aimed to investigate the role of tumor cell-derived EVs within this complex scenario. We observed that EVs collected from Hodgkin lymphoma (HL) cells were internalized by fibroblasts and triggered their migration capacity. EV-treated fibroblasts were characterized by an inflammatory phenotype and an upregulation of alpha-smooth muscle actin (α-SMA), a marker of cancer-associated fibroblasts. Analysis of the secretome of EV-treated fibroblast revealed an enhanced release of pro-inflammatory cytokines (e.g., IL-1α, IL-6, and TNF-α), growth factors (G-CSF and GM-CSF), and pro-angiogenic factors such as VEGF. These soluble factors are known to promote HL progression. In line, ingenuity pathway analysis identified inflammatory pathways, including TNF-α/NF-κB-signaling, as key factors directing the EV-dependent phenotype changes in fibroblasts. Confirming the in vitro data, we demonstrated that EVs promote α-SMA expression in fibroblasts and the expression of proangiogenic factors using a xenograft HL model. Collectively, we demonstrate that HL EVs alter the phenotype of fibroblasts to support tumor growth, and thus shed light on the role of EVs for the establishment of the tumor-promoting TME in HL.

Extracellular Vesicles: New Players in Lymphomas

“…Lymphoma cells play an important role in directing tumor immune escape by creating a feasible tumor microenvironment. A number of strategies are adopted by the cells to obtain this goal: (1) the direct recruitment of immune cells [67]; (2) the indirect “education” of normal cells to release a specific pattern of cytokines or chemokines [68,69,70]; and (3) the reprogramming of resident fibroblasts to cancer-associated fibroblasts (CAF) [71]. It is known that Treg cells exert immune-suppressive activity on lymphoma-infiltrating cytotoxic T cells, resulting in ineffective immune clearance [72].…”

“…LCEV-dependent cell-to-cell communication between distant lymphoma cells also involves educational mechanisms that are driven by CD30+ LCEVs. This mainly includes bidirectional cross-talk between malignant cells and resident fibroblasts [71]. Recent data have demonstrated that HL cells release LCEVs that express the full-length CD30 receptor and can locally modify the LME via interactions between their ligands (CD30L+) and neighboring cells.…”

The Immunosuppressive Microenvironment of Classic Hodgkin Lymphoma and Therapeutic Approaches to Counter It

“…HRS cells can educate fibroblasts to become CAFs. Dorsam et al [107] found that HL extracellular vesicles can change the secretome of fibroblasts toward a CAF phenotype. These vesicles were internalized by fibroblasts, which increased their migratory capacity, showed an inflammatory phenotype and increased expression of alpha-smooth muscle actin, a marker of CAFs.…”

“…These vesicles were internalized by fibroblasts, which increased their migratory capacity, showed an inflammatory phenotype and increased expression of alpha-smooth muscle actin, a marker of CAFs. Extracellular vesicle-treated fibroblasts enhanced the release of pro-inflammatory cytokines (e.g., IL-1α, IL-6, and TNF-α), growth factors (G-CSF and GM-CSF), and the pro-angiogenic factor VEGF [107] ( Figure 1). These findings were confirmed using a cHL xenograft model [107].…”

“…Extracellular vesicle-treated fibroblasts enhanced the release of pro-inflammatory cytokines (e.g., IL-1α, IL-6, and TNF-α), growth factors (G-CSF and GM-CSF), and the pro-angiogenic factor VEGF [107] ( Figure 1). These findings were confirmed using a cHL xenograft model [107].…”