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Immunosuppression in malignant melanoma induced by tumor-derived extracellular vesicles

Fleming, Viktor

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Abstract

The aim of this project was to investigate the role of tumor-derived extracellular vesicles (EV) in the induction of immunosuppression in malignant melanoma. Here, we focused on the effect of tumor-derived EV isolated from the Ret murine melanoma model (Ret) on bone marrow (BM)-derived immature myeloid cells (IMC). We demonstrated that IMC efficiently took up Ret-EV that resulted in the secretion of inflammatory molecules and upregulation of miRNA resembling an immunosuppressive phenotype. Furthermore, we found that Ret-EV upregulated the expression of PD-L1 on IMC. This PD-L1 expression was induced due the TLR signaling pathway, where TLR4 played a dominant role followed by TLR2 and TLR7. The TLR signaling led to the activation of NF-B, thereby inducing the transcription of PD-L1. Blocking the NF-B pathway diminished the Ret-EV-mediated PD-L1 upregulation on IMC. To test whether IMC becomes immunosuppressive upon the treatment with Ret-EV, we performed inhibition of T cell proliferation and IFN- secretion assays. Here, we demonstrated that IMC became immunosuppressive and converted into MDSC. The immunosuppressive activity of Ret-EV-treated IMC was mainly due to the induction of PD-L1. By blocking PD-L1 with neutralizing antibodies, we could almost completely abrogate the immunosuppressive properties of Ret-EV-treated IMC. Investigating the ligands for this TLR-dependent upregulation of PD-L1, we found that the inducible heat shock protein (HSP) 86 was the dominant ligand on Ret-EV, inducing TLR4 signaling in IMC upon Ret-EV treatment. Inhibition of all inducible HSP on EV by KNK-437 resulted in the reduction of the Ret-EV mediated conversion of IMC into MDSC. Furthermore, we stably knocked-down HSP86 on Ret cells. By co-culturing HSP86-deficient Ret cells with IMC, we could not observe a PD-L1 upregulation, whereas the scramble control showed a strong increase in PD-L1 expression. When using DMA to block EV secretion, the EV-mediated PD-L1 upregulation was strongly diminished. Finally, we observed an impaired tumor growth of HSP86-deficient Ret cells compared to wild type cells in vivo that was accompanied by reduced levels of MDSC expressing PD-L1 in the tumor microenvironment (TME). Summary VIII Taken together, our findings demonstrate a critical role in converting IMC into MDSC for HSP86 on EV that could be a promising target for immunotherapy.

Item Type: Dissertation
Supervisor: Umansky, Prof. Dr. Viktor
Date of thesis defense: 13 December 2018
Date Deposited: 15 Jan 2019 14:52
Date: 2019
Faculties / Institutes: The Faculty of Bio Sciences > Dean's Office of the Faculty of Bio Sciences
Subjects: 570 Life sciences
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