Antibody fragments directed against different portions of the human neural cell adhesion molecule L1 act as inhibitors or activators of L1 function

• Published in: PloS one 2012-12, Vol.7 (12), p.e52404
• Main Authors: Wang, Yan, Loers, Gabriele, Pan, Hong-Chao, Gouveia, Ricardo, Zhao, Wei-Jiang, Shen, Yan-Qin, Kleene, Ralf, Costa, Julia, Schachner, Melitta
• Format: Article
• Language: English
• Subjects: Adhesion; Amino acids; Angiogenesis; Antibodies; Axonogenesis; BAX protein; Bcl-2 protein; Binding; Biology; Cancer; Cell adhesion; Cell adhesion & migration; Cell adhesion molecules; Cell Line; Cell migration; Cell Movement - drug effects; Cell proliferation; Cell Proliferation - drug effects; Cell survival; Chromatography; Cloning; Enzyme-linked immunosorbent assay; Fibronectin; Fibronectins; Fragmentation; Fragments; Homology; Humans; Hydrogen peroxide; Hydrogen Peroxide - toxicity; Immunoglobulins; Kinases; L1 protein; Medicine; Metastases; Mutation; Nervous system; Neural cell adhesion molecule; Neural Cell Adhesion Molecule L1 - agonists; Neural Cell Adhesion Molecule L1 - antagonists & inhibitors; Neural Cell Adhesion Molecule L1 - metabolism; Neurites - drug effects; Neuroblastoma cells; Neuroblasts; Neurosciences; Phosphorylation; Phosphorylation - drug effects; Protein Binding; Protein Interaction Domains and Motifs; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Recombinant Proteins - pharmacology; Single-Chain Antibodies - isolation & purification; Single-Chain Antibodies - metabolism; Single-Chain Antibodies - pharmacology; src-Family Kinases - metabolism; Synaptogenesis; Toxicity; Tumors
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0052404

The neural cell adhesion molecule L1 plays important roles in neuronal migration and survival, neuritogenesis and synaptogenesis. L1 has also been found in tumors of different origins, with levels of L1 expression correlating positively with the metastatic potential of tumors. To select antibodies targeting the varied functions of L1, we screened the Tomlinson library of recombinant human antibody fragments to identify antibodies binding to recombinant human L1 protein comprising the entire extracellular domain of human L1. We obtained four L1 binding single-chain variable fragment antibodies (scFvs), named I4, I6, I13, and I27 and showed by enzyme-linked immunosorbent assay (ELISA) that scFvs I4 and I6 have high affinity to the immunoglobulin-like (Ig) domains 1-4 of L1, while scFvs I13 and I27 bind strongly to the fibronectin type III homologous (Fn) domains 1-3 of L1. Application of scFvs I4 and I6 to human SK-N-SH neuroblastoma cells reduced proliferation and transmigration of these cells. Treatment of SK-N-SH cells with scFvs I13 and I27 enhanced cell proliferation and migration, neurite outgrowth, and protected against the toxic effects of H(2)O(2) by increasing the ratio of Bcl-2/Bax. In addition, scFvs I4 and I6 inhibited and scFvs I13 and I27 promoted phosphorylation of src and Erk. Our findings indicate that scFvs reacting with the immunoglobulin-like domains 1-4 inhibit L1 functions, whereas scFvs interacting with the fibronectin type III domains 1-3 trigger L1 functions of cultured neuroblastoma cells.