SCIENCE-JOBS-DE

Regulation of tumor - associated macrophage and myeloid - derived suppressor cell activation and its neutralization in transgenic mouse melanoma model (Mannheim)

Melanoma  immunotherapy is  not  satisfactory  due  to  the  accumulation  of  chronic inflammatory factors and immunosuppressive myeloid cells such as tumor-associated macrophages (TAM) and myeloid-derived  suppressor  cells  (MDSC)  in  tumor  lesions.  The  goal  of  the  project  is  to  better understand  the  molecular  mechanisms  underlying  the  inhibition  of  anti-tumor  immune  responses mediated by TAM and  MDSC, and to design novel therapeutic strategies targeting these myeloid cells in melanoma. We will study the role of signaling molecules (p38 MAPK and S100A8/A9) and microRNA  in  the  capacity  of  TAM  and  MDSC  to  inhibit  anti-tumor  reactivity  of  T  cells.  Results  of the   project   will   help   to   develop   novel   efficient  human   melanoma   treatments   neutralizing immunosuppression in the tumor microenvironment.

Specific Aims
Aim 1:Investigating  signaling  molecules  and  miR  involved  in  the  recruitment,  expansion  and activation of TAM and MDSC during melanoma progression
Aim  2:Studying  the  neutralization  of  immunosuppression  induced  by TAM  and  MDSC  using  the modulation of their relevant signaling pathways and miR

Experimental program
1.We  will  test  the  expression  and  activation  (phosphorylation) of  such  signaling  molecules  as S100A8/A9 and p38 MAPK in F4/80high CD11b+Gr1-TAM and CD11b+Gr1+ MDSC in melanoma lesions  and  lymphoid  organs  using  FACS.  Differentially
up-and  down-regulated  miR  in  TAM and  MDSC  will  be  studied  by  AffymetrixMicroarray  analysis  and  real-time  quantitative  PCR.Functional  relevance  of  identified  miR  will  be  analyzed  using  lentiviral  constructs.TAM  and MDSC  will  be  characterized  by  the  expression  of  arginase-1  and  inducible  nitric  oxide synthase.  The  activity  of  these  myeloid  cells  will  be  determined  by  the  inhibition of  T  cell proliferation  upon  the  co-culture  with  TAM  and/or  MDSC.  Inflammatory  factors  (VEGF,  IL-1, IL-6, TNF-, TGF-, etc.) will be detected in melanoma lesions by multiplex technology.Tumor-infiltrating T cells will be validated measuring the -chain expression by FACS.
2.We  will  suppress  signaling  pathways  involved  in  the  expansion  and  activation  of  MDSC  and TAM  with  specific  inhibitors  p38  MAPK  (SB203580  and  RO3201195).  Activating  miR  will  be blocked by the sponge preventing the interaction of this miRNA with its targets. TAM or MDSC isolated from melanoma  lesions  will  be  treated with  these inhibitors followed  by  co-incubation with  activated  syngeneic  T  cells.  T-cell  proliferation  and -chain  expression  will  be  tested  by
Susanne Melchers
susanne.melchers@gmx.de
Klinik fĂĽr Dermatologie
Mannheim