Alberto Monroy-García

TGF-β Induces the Secretion of Extracellular Vesicles Enriched with CD39 and CD73 from Cervical Cancer Cells

The presence of TGF-β in the tumor microenvironment of cervical cancer (CC) is important for tumor progression. In this study, we analyzed the effect of TGF-β on the expression of the ectonucleotidases CD39 and CD73, which are involved in the generation of adenosine (Ado), in CC cells and in extracellular vesicles (EVs) secreted by these cells. Treatment of HeLa and CaSki cells for 72 h with recombinant human TGF-β increased the expression of CD39 and CD73 by 20 and 30% and by 40 and 100%, respectively. The addition of SB505124, an inhibitor of the TGF-β1 receptor, or GW4869, an inhibitor of exosome formation and release, reduced the expression and release of both ectonucleotidases in CC cells. Furthermore, TGF-β promoted the secretion of medium-large EVs (>130 nm) in HeLa cells (HeLa + TGF-β/EVs) and CaSki cells (CaSki + TGF-β/EVs), which increased the expression of CD39 (>20%) and CD73 (>60%), and EVs obtained from cells treated with TGF-β had a greater capacity to generate Ado than did EVs obtained from cells cultured in the absence of this factor (HeLa/EVs and CaSki/EVs). These findings suggest that the production of TGF-β in the CC TME can promote neoplastic progression through the secretion of EVs enriched with CD39 and CD73. Therefore, the inhibition of CD39+ CD73+ EVs could be a strategy for the treatment of CC.

Evidence that cervical cancer cells cultured as tumorspheres maintain high CD73 expression and increase their protumor characteristics through TGF‐β production

Abstract Recently, a link between the biological activity of CD73 and tumorigenicity in solid tumors has been proposed. We previously reported that the generation of adenosine (Ado) by the activity of CD73 in cervical cancer (CC) cells induces transforming growth factor‐beta 1 (TGF‐β1) production to maintain CD73 expression. In the present study, we analyzed the participation of TGF‐β1 in CD73 expression and the development of protumoral characteristics in CaSki CC cells cultured as tumorspheres (CaSki‐T) and in monolayers (CaSki‐M). Compared with those in CaSki‐M cells, CD73 expression and Ado generation ability were significantly increased in CaSki‐T cells. CaSki‐T cells exhibited enrichment in the CSC‐like phenotype due to increases in the expression levels of stem cell markers (CD49f, CK17, and P63; OCT4 and SOX2), greater sphere formation efficiency (SFE), and an increase in the percentage of side population (SP) cells. Interestingly, compared with CaSki‐M cells, CaSki‐T cells produced a greater amount of TGF‐β1 and presented a marked protumor phenotype characterized by a significant decrease in the expression of major histocompatibility complex class‐I (MHC‐I) molecules, an increase in the expression of multidrug resistance protein‐I (MRP‐I) and vimentin, and an increase in the protein expression levels of Snail‐1 and Twist, which was strongly reversed with TGF‐β1 inhibition. These results suggest that the presence of TGF‐β1−CD73–Ado feedback loop can promote protumoral characteristics in the CC tumor microenvironment.

Adenosine increases PD‐L1 expression in mesenchymal stromal cells derived from cervical cancer through its interaction with A2AR/A2BR and the production of TGF‐β1

AbstractMesenchymal stromal cells (MSCs) together with malignant cells present in the tumor microenvironment (TME), participate in the suppression of the antitumor immune response through the production of immunosuppressive factors, such as transforming growth factor beta 1 (TGF‐β1). In previous studies, we reported that adenosine (Ado), generated by the adenosinergic activity of cervical cancer (CeCa) cells, induces the production of TGF‐β1 by interacting with A2AR/A2BR. In the present study, we provide evidence that Ado induces the production of TGF‐β1 in MSCs derived from CeCa tumors (CeCa‐MSCs) by interacting with both receptors and that TGF‐β1 acts in an autocrine manner to induce the expression of programmed death ligand 1 (PD‐L1) in CeCa‐MSCs, resulting in an increase in their immunosuppressive capacity on activated CD8+ T lymphocytes. The addition of the antagonists ZM241385 and MRS1754, specific for A2AR and A2BR, respectively, or SB‐505124, a selective TGF‐β1 receptor inhibitor, in CeCa‐MSC cultures significantly inhibited the expression of PD‐L1. Compared with CeCa‐MSCs, MSCs derived from normal cervical tissue (NCx‐MSCs), used as a control and induced with Ado to express PD‐L1, showed a lower response to TGF‐β1 to increase PD‐L1 expression. Those results strongly suggest the presence of a feedback mechanism among the adenosinergic pathway, the production of TGF‐β1, and the induction of PD‐L1 in CeCa‐MSCs to suppress the antitumor response of CD8+ T lymphocytes. The findings of this study suggest that this pathway may have clinical importance as a therapeutic target.