Unit:
Faculty of MedicineLibrary of the School of Health Sciences
Author:
Christofides Anthos
Dissertation committee:
Δημήτριος Τ. Μπούμπας, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Βασιλική Α. Μπουσιώτη, Καθηγήτρια, Ιατρική Σχολή, Πανεπιστήμιο Harvard και BIDMC Κέντρο Καρκίνου, Beth Israel Deaconess Medical Center
Παναγιώτης Βεργίνης, Αναπληρωτής Καθηγητής, Ιατρική Σχολή, Πανεπιστήμιο Κρήτης
Σωτήριος Τσιόδρας, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Δημήτριος Βασιλόπουλος, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Παναγιώτης Τσιριγώτης, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Αντώνιος Φανουριάκης, Επίκουρος Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Original Title:
Ο ρόλος του PD-1 στην ενεργοποίηση των μακροφάγων
Translated title:
The role of PD-1 in macrophage polarization and fate commitment
Summary:
Background
PD-1 is a checkpoint receptor that has been shown to inactivate T cells through SHP-2 recruitment. However, tumor bearing mice with conditional deletion of SHP-2 in T cells did not demonstrate any differences in tumor progression compared to wild type mice. Both PD-1 and SHP-2 are also expressed in myeloid cells, including the myeloid cells progenitors. PD-1 deletion in myeloid cells can induce differentiation of myeloid progenitors towards mature myeloid cells, while SHP-2 gain- of-function mutations are known to be implicated in Acute Myelogenous Leukemia, by preventing differentiation, through dephosphorylation of IRF-8 and HOXA10. Nevertheless, no PD-1/SHP- 2 interaction has been described in primary healthy myelocytes. The goal of this study, is to investigate the PD-1 / SHP-2 interaction in myeloid cells in the context of tumor and the role of this axis in myeloid cell fate commitment and anti-tumor immunity.
Methods
We generated mice with conditional deletion of Ptpn11 gene (encoding for Shp-2) in myeloid cells (Shp2f/fLysMCre), or T cells (Shp2f/fLckCre) and used Shp2f/f mice as control. We also used mice with myeloid-specific deletion of the Pdcd1 gene (encoding for PD-1). Tumor cells were injected subcutaneously and tumor growth was monitored. We used two different tumor cell lines: MC17- 51 fibrosarcoma and B16-F10 melanoma. At termination, tumors, spleens and bone marrows were collected, processed (details below), and immune populations were identified by flow cytometry. For cell isolation we used Fluorescence-Activated Cell Sorting (FACS) and magnetic beads and for signaling studies we used bone marrow cells cultured with GM-CSF and IL-3.
Results
Mice with myeloid-specific SHP-2 ablation had significantly decelerated tumor growth compared to mice with T cell-specific SHP-2 ablation or control mice, but did not gain extra benefit from PD-1 blockade. SHP-2 deletion in myeloid cells resulted in T cell activation and myeloid cell activation and differentiation, while MDSCs had diminished suppressive properties and monocytes acquired lasting anti-tumor properties. Furthermore, by using RNA sequencing and gene set enrichment analysis in TAMs and PMN MDSCs, we demonstrated that SHP-2 deletion enriched gene pathways related to activation, differentiation, phagocytosis and antigen presentation in both these myeloid cell types. Myeloid specific PD-1 ablation had very similar effects in T cells and myeloid cells, and RNAseq of PD-1 deficient TAMs showed greater than 50% overlap in gene expression with SHP-2 deficient TAMs. Furthermore, we determined that in myeloid cells, GM-CSF induced phosphorylation of PD-1, recruited SHP-2 to the GM-CSF receptor and facilitated PD-1-SHP-2 interaction. Finally, either PD-1 or SHP-2 deletion enhanced GM-CSF-depended myeloid differentiation by abrogating SHP-2 medicated dephosphorylation of IRF8 and HOXA10.
Conclusion
With the current study, we showed that PD-1-SHP-2 interaction can play a role in the differentiation of bone marrow myelocytes and myeloid cell progenitors and, subsequently, can affect their function and anti-tumor immunity.
Main subject category:
Health Sciences
Keywords:
PD-1, Tumor microenvironemnt, Myeloid cells, Emergency myelopoiesis, SHP-2 phosphatase
