Generation of GPI-linked CCL5 based chemokine receptor antagonists for the suppression of acute vascular damage during allograft transplantation.
Limiting the acute vascular damage associated with leukocyte infiltration is a central issue in solid organ transplantation. The family of chemotactic cytokines (chemokines) helps to regulate leukocyte recruitment. Systemic treatment with the chemokine ligand-5 (CCL5) based antagonist Met-RANTES has previously shown to suppress acute damage to transplanted kidneys by blocking effector cell recruitment. To address problems associated with systemic long-term administration of chemokine receptor antagonists, a chemokine based reagent was designed to be integrated into endothelial surfaces of the organ just before transplantation. Proteins anchored by glycosylphosphatidylinositol (GPI), when purified and added to cells, are efficiently incorporated into their cell surface membranes. A series of modifications were introduced into the CCL5 protein to generate a functional antagonist. These included the addition of an N-terminal methionine group, a mutation to render the protein a dimer and a GPI signal sequence for surface expression. The resultant protein was stably expressed in CHO cells, GPI anchorage was confirmed and the protein purified by FPLC. Exogenously administered Met-CCL5(dimer)-GPI was efficiently inserted into the membrane of microvascular endothelial cells. The reagent is being tested in murine models of renal transplantation. The effect on subsequent immune induced damage will be assessed.
Notohamiprodjo
Mike
M
Medizinische Poliklinik, Schillerstrasse 42, 80336 Ludwig-Maximilians-University of Munich, Kölnerplatz 1, 80803 Munich, German Cancer Research Center, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.
Djafarzadeh
Roghieh
R
Mojaat
Anke
A
von Lüttichau
Irene
I
Gröne
Hermann-Josef
HJ
Nelson
Peter J
PJ
D000208:Acute Disease / Q000628:therapy; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002478:Cells, Cultured; D018946:Chemokine CCL5; D019742:Chemokines, CC / Q000737:chemistry*; D006224:Cricetinae; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000473:pathology; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D016030:Kidney Transplantation; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D019707:Receptors, Chemokine / Q000037:antagonists & inhibitors*; D014184:Transplantation, Homologous
C503460:Ccl5 protein, mouse; D018946:Chemokine CCL5; D019742:Chemokines, CC; D017261:Glycosylphosphatidylinositols; D019707:Receptors, Chemokine