Insluin and epithelial growth factor (EGF) promote programmed death ligand 1(PD-L1) production and transport in colon cancer stem cells

Abstract

Background Programmed cell death ligand 1 (PD-L1) is an important immune-inhibitory protein expressed on cancer cells to mediate cancer escape through interaction with PD-1 expressed on activated T lymphocytes (T cells). Previously, we reported that colon and breast cancer stem cells (CSCs) expressed much higher levels of PD-L1 than their parental cells, suggesting they will be more resistant to immune attack. Methods We investigated the underlining mechanism of PD-L1 increase in colon CSCs, with a special focus on the effect of insulin and epithelial growth factor (EGF), the two fundamental components to sustain the metabolism and stemness in the culture of CSCs. Results We found that insulin increased the total and surface PD-L1 levels through PI3K/Akt/mTOR pathway as the increase could be inhibited by the dual inhibitor of the pathway, BEZ235. EGF didn’t affect the total PD-L1 levels of CSCs but increased the cell surface protein levels by flow cytometry analysis, indicating EGF promotes the transport of PD-L1 to the cell surface. Blocking cell surface PD-L1 with a specific antibody resulted in a significant reduction of tumour sphere formation but didn’t interfere with the sphere growth, suggesting that cell surface PD-L1 may act as an adhering molecule for CSCs. Conclusions Apart from the essential roles in metabolism and stemness, insulin and EGF involve in up-regulation of PD-L1 expression in colon CSCs, therefore the inhibition of insulin and EGF/EGFR pathways can be considered for cancer immunotherapy or combined with PD-1/PD-L1 antibody-based cancer immunotherapy to eliminate CSCs.

Transforming growth factor beta induces fibroblasts to express and release the immunomodulatory protein PD‐L1 into extracellular vesicles

“…TGFβ is considered as one of the most profibrotic cytokines and it is well established that TGFβ is a crucial modulator of ECM production in fibrogenic processes . As such, we first established that TGFβ upregulated PD‐L1 in both HuLFs and murine fibroblast cell lines (Figures and ); which extends previous work showing that PD‐L1 expression is upregulated by various cytokines, including IFN‐γ, GM‐CSF, IL‐17, TNF‐α, and EGF . To address whether PD‐L1 had a central role in profibrotic TGFβ signaling, PD‐L1 was knocked down with siRNA and the cellular response to TGFβ was examined.…”

“…65 As such, we first established that TGFβ upregulated PD-L1 in both HuLFs and murine fibroblast cell lines (Figures 1 and 2); which extends previous work showing that PD-L1 expression is upregulated by various cytokines, including IFN-γ, GM-CSF, IL-17, TNF-α, and EGF. [66][67][68][69][70] To address whether PD-L1 had a central role in profibrotic TGFβ signaling, PD-L1 was knocked down with siRNA and the cellular response to TGFβ was examined. Inhibition of PD-L1 expression significantly reduced TGFβ induced profibrotic target genes, cell migration, and soft agar colony formation (Figure 3).…”

Cancer stemness contributes to cluster formation of colon cancer cells and high metastatic potentials

“…For colon cancer HT-29 cells, we recently showed that after SP1 culturing, the clustered cells increased the expression of stem cell markers including CD133, CD24 and CD44. 26 Collectively, our data showed that after SP1 culturing, colon cancer cells have enriched cancer stemness. Therefore, upon subsequent culturing of the enriched cancer stemness of SP1 culture (ie SP2), spherical clusters with >20% population of single cells formed within 24 hours.…”

Cell stemness, epithelial-to-mesenchymal transition, and immunoevasion: Intertwined aspects in cancer metastasis

“…The synergy between EGF and insulin effects induced a remarkable increase in PD-L1 expression on the surfaces of CSCs. This elevated expression enhanced the ability of CSCs to form spheres [85]. The importance of proper glycosylation for maintaining PD-L1 stability was also emphasized by Cha et al [86].…”

“…Inhibition of EGF by gefitinib affected PD-L1 levels by increasing its degradation [84]. Similarly, in colon cancer, Chen and coworkers found that EGF facilitated PD-L1 transport to the surface of cancer cells, which stabilized the protein [85]. They also found that insulin had a stimulatory effect on PD-L1 synthesis, which was mediated by activating the PI3k/ Akt pathway.…”