Effect of single-chain antibody targeting of the ligand-binding domain in the anaplastic lymphoma kinase receptor.
The tyrosine kinase receptor anaplastic lymphoma kinase (ALK) and its ligand, the growth factor pleiotrophin (PTN), are highly expressed during the development of the nervous system and have been implicated in the malignant progression of different tumor types. Here, we describe human single-chain variable fragment (scFv) antibodies that target the ligand-binding domain (LBD) in ALK and show the effect in vitro and in vivo. The ALK LBD was used as a bait in a yeast two-hybdrid system to select human scFv from a library with randomized complementarity-determining region 3 domains. Surface plasmon resonance showed high-affinity binding of the selected scFv. The anti-ALK scFv competed for binding of PTN to ALK in intact cells and inhibited PTN-dependent signal transduction through endogenous ALK. Invasion of an intact endothelial cell monolayer by U87MG human glioblastoma cells was inhibited by the anti-ALK scFv. In addition, the growth of established tumor xenografts in mice was reversed after the induction of the conditional expression of the anti-ALK scFv. In archival malignant brain tumors expression levels of ALK and PTN were found elevated and appear correlated with poor patient survival. This suggests a rate-limiting function of the PTN/ALK interaction that may be exploited therapeutically.
Stylianou
D C
DC
Lombardi Cancer Center, Georgetown University, Washington, DC 20057, USA.
Auf der Maur
A
A
Kodack
D P
DP
Henke
R T
RT
Hohn
S
S
Toretsky
J A
JA
Riegel
A T
AT
Wellstein
A
A
D000595:Amino Acid Sequence; D000077548:Anaplastic Lymphoma Kinase; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D001419:Bacteria / Q000166:cytology / Q000276:immunology; D001667:Binding, Competitive; D001932:Brain Neoplasms / Q000235:genetics / Q000276:immunology / Q000473:pathology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019295:Computational Biology; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D042783:Endothelial Cells / Q000473:pathology; D000939:Epitopes / Q000276:immunology; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000473:pathology; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology; D008024:Ligands; D051379:Mice; D000078792:Midkine; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000276:immunology; D017434:Protein Structure, Tertiary; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000737:chemistry* / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases; D015398:Signal Transduction / Q000276:immunology
D000906:Antibodies; D002352:Carrier Proteins; D016207:Cytokines; D000939:Epitopes; D007135:Immunoglobulin Variable Region; D008024:Ligands; D012333:RNA, Messenger; C066845:pleiotrophin; D000078792:Midkine; C000626173:ALK protein, human; C000626174:Alk protein, mouse; D000077548:Anaplastic Lymphoma Kinase; D011505:Protein-Tyrosine Kinases; D020794:Receptor Protein-Tyrosine Kinases; D017027:Protein Tyrosine Phosphatases