Etude des voies de recrutement des cellules dendritiques dans une tumeur solide

2017 
The concept of immunosurveillance suggests that the innate and adaptative immune system eliminate developing tumors. However, tumor development is associated with important modifications of the stroma which, by multiple mechanisms, restrain the immune response notably by affecting dendritic cells (DC) recruitment and functions. My thesis project aims at deciphering how the tumor environment alters the mechanisms and pathways of DC recruitment and impairs their functions. First, we determine if the site of tumor transplantation affects tumor immunogenicity. We show that tumors transplanted in the dermis (i.d.), an environment containing multiple DC subsets, induce a protective anti-tumoral immune response and tumor rejection. By contrast, the same tumor implanted in the subcutaneous tissue (s.c), mainly containing monocytes, is not rejected. Rejection of i.d. tumor is associated with a rapid (within 2 days) recruitment of DC within the tumor and rapid migration of DC towards tumor draining lymph nodes (dLN) where they present the tumor antigens (TAA) to CD4 and CD8 T lymphocyte. These events also occur for s.c. tumors but with a delayed kinetic. Thus the kinetic of DC mobilisation is decisive for tumor immunogenicity. Analysis of the DC subpopulations (TIDC) infiltrating the i.d. or s.c. tumors at 4 days (D4) and 8 days (D8) post-tumor transplantation, revealed that the different DC subpopulations are present at similar frequencies. Based on these findings, we proposed that i.d. tumor are rapidly infiltrated by dermal DC (DDC), whereas in s.c. tumor, the absence of inflammatory signals would limit DDC recruitment. In this latter case, DC would mainly come from local differentiation of blood-born precursors of DC (pre-cDC). Local differentiation of pre-cDC within the immunosuppressor tumor environment may affect their differentiation program and functions. We found that pre-cDC infiltrate i.d. and s.c. tumors starting at D4 and their frequency increases at D8. To determine the DC origin in tumors, we use CD11c-DTR-GFP mice in which CD11c+ cells express a fusion protein constituted by the diphtheria toxin receptor (DTR) and GFP. To track DDC trafficking within tumors, we injected anti-MHCII antibody before tumor implantation and analysed MHCII+ CD11c+ DDC infiltration in tumor by biphotonic microscopy. At D3 post-tumor transplantation, the DDC infiltration was higher in i.d. than s.c. tumors. To analyse the impact of this early DDC recruitment on anti-tumor immunity, we inhibit early recruitment of DC by injection of pertussis toxin (PTX), a chemokine receptor protein G inhibitor, during the three first days of tumor development. For i.d. tumors, PTX treatment induced 60% reduction of DC recruitment starting at D4. In s.c. tumor, while this effect was observable at D3 (60% reduction) and increased to 80% at D4. These results suggest that early recruitment of DDC to i.d. tumors may be be chemokine independent. PTX treatment, which inhibits DDC migration from tumors to dLN inhibits the early TAA presentation to CD4 and CD8 T cells but did not impaired i.d. tumor rejection. Collectively, these results suggest that a first wave of DDC may infiltrate i.d. tumor. The initial wave of DDC may rapidly activate the adaptive immune system and induce protective anti-tumoral immune response. For s.c. tumor, this first wave in delayed or limited. Tumor neoangioenesis would permit an input of pre-cDC which would differentiate locally into cDC1 and cDC2. To consolidate this model we are developing new protocols to efficiently inhibit early recruitment of DDC in i.d. tumor. Moreover, to determine the DC origin and pathways of DC recruitment in tumors, we exploit several experimental approaches to directly analyse DDC migration toward i.d. and s.c. tumor.
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