title: A novel CXCL10-based GPI-anchored fusion protein as adjuvant in NK-based tumor therapy.
abstract: BACKGROUND
  Cellular therapy is a promising therapeutic strategy for malignant diseases. The efficacy of this therapy can be limited by poor infiltration of the tumor by immune effector cells. In particular, NK cell infiltration is often reduced relative to T cells. A novel class of fusion proteins was designed to enhance the recruitment of specific leukocyte subsets based on their expression of a given chemokine receptor. The proteins are composed of an N-terminal chemokine head, the mucin domain taken from the membrane-anchored chemokine CX3CL1, and a C-terminal glycosylphosphatidylinositol (GPI) membrane anchor replacing the normal transmembrane domain allowing integration of the proteins into cell membranes when injected into a solid tumor. The mucin domain in conjunction with the chemokine head acts to specifically recruit leukocytes expressing the corresponding chemokine receptor.
  
  
  RESULTS
  A fusion protein comprising a CXCL10 chemokine head (CXCL10-mucin-GPI) was used for proof of concept for this approach and expressed constitutively in Chinese Hamster Ovary cells. FPLC was used to purify proteins. The recombinant proteins efficiently integrated into cell membranes in a process dependent upon the GPI anchor and were able to activate the CXCR3 receptor on lymphocytes. Endothelial cells incubated with CXCL10-mucin-GPI efficiently recruited NK cells in vitro under conditions of physiologic flow, which was shown to be dependent on the presence of the mucin domain. Experiments conducted in vivo using established tumors in mice suggested a positive effect of CXCL10-mucin-GPI on the recruitment of NK cells.
  
  
  CONCLUSIONS
  The results suggest enhanced recruitment of NK cells by CXCL10-mucin-GPI. This class of fusion proteins represents a novel adjuvant in cellular immunotherapy. The underlying concept of a chemokine head fused to the mucin domain and a GPI anchor signal sequence may be expanded into a broader family of reagents that will allow targeted recruitment of cells in various settings.
authors.affiliation: AG Klinische Biochemie, Medizinische Klinik und Poliklinik IV, Klinikum der Universität München, Munich, Germany.
authors.forename: Niklas
authors.initials: N
authors.lastname: Muenchmeier
authors.forename: Sophia
authors.initials: S
authors.lastname: Boecker
authors.forename: Lorenz
authors.initials: L
authors.lastname: Bankel
authors.forename: Laura
authors.initials: L
authors.lastname: Hinz
authors.forename: Nicole
authors.initials: N
authors.lastname: Rieth
authors.forename: Constantin
authors.initials: C
authors.lastname: Lapa
authors.forename: Anna N
authors.initials: AN
authors.lastname: Mendler
authors.forename: Elfriede
authors.initials: E
authors.lastname: Noessner
authors.forename: Ralph
authors.initials: R
authors.lastname: Mocikat
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Nelson
mesh_terms: D000276:Adjuvants, Immunologic / Q000627:therapeutic use*; D000818:Animals; D016466:CHO Cells; D002448:Cell Adhesion; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D054357:Chemokine CXCL10 / Q000627:therapeutic use*; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D005260:Female; D017261:Glycosylphosphatidylinositols / Q000302:isolation & purification / Q000627:therapeutic use*; D006801:Humans; D007167:Immunotherapy; D007694:Killer Cells, Natural / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D009077:Mucins / Q000378:metabolism; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000627:therapeutic use*; D012212:Rheology
chemical_list: D000276:Adjuvants, Immunologic; D054357:Chemokine CXCL10; D017261:Glycosylphosphatidylinositols; D009077:Mucins; D011993:Recombinant Fusion Proteins
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