title: Anti-Mesothelin Nanobodies for Both Conventional and Nanoparticle-Based Biomedical Applications.
abstract: Mesothelin, a cancer biomarker overexpressed in tumors of epithelial origin, is a target for nanotechnology-based diagnostic, therapeutic, and prognostic applications. The currently available anti-mesothelin antibodies present limitations, including low penetration due to large size and/or lack of in vivo stability. Single domain antibodies (sdAbs) or nanobodies (Nbs) provide powerful solutions to these specific problems. We generated a phage-display library of Nbs that were amplified from B cells of a llama that was immunized with human recombinant mesothelin. Two nanobodies (Nb A1 and Nb C6) were selected on the basis of affinity (K(D) = 15 and 30 nM, respectively). Nb A1 was further modified by adding either a cysteine to permit maleimide-based bioconjugations or a sequence for the site-specific metabolic addition of a biotin in vivo. Both systems of conjugation (thiol-maleimide and streptavidin/biotin) were used to characterize and validate Nb A1 and to functionalize nanoparticles. We showed that anti-mesothelin Nb A1 could detect native and denatured mesothelin in various diagnostic applications, including flow cytometry, western blotting, immunofluorescence, and optical imaging. In conclusion, anti-mesothelin Nbs are novel, cost-effective, small, and single domain reagents with high affinity and specificity for the tumor-associated antigen mesothelin, which can be simply bioengineered for attachment to nanoparticles or modified surfaces using multiple bioconjugation strategies. These anti-mesothelin Nbs can be useful in both conventional and nanotechnology-based diagnostic, therapeutic and prognostic biomedical applications.
authors: Andrew M
authors: AM
authors: Prantner
authors: Marc
authors: M
authors: Turini
authors: Brigitte
authors: B
authors: Kerfelec
authors: Shree
authors: S
authors: Joshi
authors: Daniel
authors: D
authors: Baty
authors: Patrick
authors: P
authors: Chames
authors: Nathalie
authors: N
authors: Scholler
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D058851:GPI-Linked Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D006367:HeLa Cells; D006801:Humans; D000090204:Mesothelin; D053758:Nanoparticles / Q000737:chemistry / Q000627:therapeutic use* / Q000648:ultrastructure; D015202:Protein Engineering / Q000379:methods; D013347:Subcellular Fractions / Q000276:immunology* / Q000473:pathology
chemical_list: D000911:Antibodies, Monoclonal; D058851:GPI-Linked Proteins; D000090204:Mesothelin