A His19 to Ala mutant of melanoma growth-stimulating activity is a partial antagonist of the CXCR2 receptor.
Melanoma growth stimulating activity (MGSA) and IL-8 are related chemokines that are potent chemoattractants and activators of neutrophils both in vitro and in vivo. Increasing evidence suggests that these molecules play an important role in inflammation; thus, antagonists of their action could be useful therapeutically as antiinflammatory agents. We have generated an MGSA mutant, H19A, that shows a dissociation between receptor binding and biologic activity. The biologic activity of the H19A mutant is between 133-fold and 282-fold less potent than that of wild-type MGSA measured by three independent assays of neutrophil function, i.e., elastase release chemotaxis and the up-regulation of CD18. In addition, pretreatment of cells with the H19A mutant inhibited the ability of MGSA to induce elastase release and chemotaxis and to increase intracellular calcium. However, competition binding studies in cells transfected with the CXCR2 receptor and in neutrophils demonstrate that the receptor affinity of the H19A mutant is only 13-fold less than that of wild-type MGSA. These studies suggest that the mutant MGSA is defective in activating signaling through the receptor and indicate that binding to the receptor is not sufficient to activate a biologic response. The dissociation between receptor binding and activation for this mutant suggests that it should be possible to design antagonists of MGSA that may be of clinical utility.
Baly
D L
DL
Genentech, Inc., South San Francisco, CA 94080, USA. baly@gene.com
Horuk
R
R
Yansura
D G
DG
Simmons
L C
LC
Fairbrother
W J
WJ
Kotts
C
C
Wirth
C M
CM
Gillece-Castro
B L
BL
Toy
K
K
Hesselgesser
J
J
Allison
D E
DE
D019943:Amino Acid Substitution; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors / Q000235:genetics / Q000494:pharmacology*; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D003571:Cytochalasin B / Q000494:pharmacology; D006133:Growth Substances / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8 / Q000494:pharmacology; D019272:Leukocyte Elastase / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000502:physiology; D019707:Receptors, Chemokine / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D018123:Receptors, Interleukin / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D023062:Receptors, Interleukin-8A; D023063:Receptors, Interleukin-8B; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection
D015703:Antigens, CD; C475829:CXCL1 protein, human; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors; D006133:Growth Substances; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8; D019707:Receptors, Chemokine; D018123:Receptors, Interleukin; D023062:Receptors, Interleukin-8A; D023063:Receptors, Interleukin-8B; D011993:Recombinant Fusion Proteins; D003571:Cytochalasin B; D019272:Leukocyte Elastase