NaLi-H1: A universal synthetic library of humanized nanobodies providing highly functional antibodies and intrabodies

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Abstract

In vitro selection of antibodies allows to obtain highly functional binders, rapidly and at lower cost. Here, we describe the first fully synthetic phage display library of humanized llama single domain antibody (NaLi-H1: Nanobody Library Humanized 1). Based on a humanized synthetic single domain antibody (hs2dAb) scaffold optimized for intracellular stability, the highly diverse library provides high affinity binders without animal immunization. NaLi-H1 was screened following several selection schemes against various targets (Fluorescent proteins, actin, tubulin, p53, HP1). Conformation antibodies against active RHO GTPase were also obtained. Selected hs2dAb were used in various immunoassays and were often found to be functional intrabodies, enabling tracking or inhibition of endogenous targets. Functionalization of intrabodies allowed specific protein knockdown in living cells. Finally, direct selection against the surface of tumor cells produced hs2dAb directed against tumor-specific antigens further highlighting the potential use of this library for therapeutic applications.

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Sandrine Moutel

Institut Curie, Paris, France

Competing interests
Sandrine Moutel, Co-inventor on a patent application (filled under ref: WO/2015/063331) that covers the hs2dAb scaffold and the commercial use of the library. The library has been licensed to Hybrigenics Service SA, which will perform screens on a fee-for-service basis.A consultant for Hybrigenics Service SA.
Nicolas Bery

Inserm, UMR 1037-CRCT, Toulouse, France

Competing interests
No competing interests declared.
Virginie Bernard

Institut Curie, Paris, France

Competing interests
No competing interests declared.
Laura Keller

Inserm, UMR 1037-CRCT, Toulouse, France

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0002-1786-9760
Emilie Lemesre

Institut Curie, Paris, France

Competing interests
No competing interests declared.
Ario de Marco

Institut Curie, Paris, France

Competing interests
No competing interests declared.
Laetitia Ligat

plateau de protéomique, CRCT, Toulouse, France

Competing interests
No competing interests declared.
Jean-Christophe Rain

Hybrigenics Service SA, Paris, France

Competing interests
Jean-Christophe Rain, Employed by, and a stockholder in, Hybrigenics Service SA.
Gilles Favre

Inserm, UMR 1037-CRCT, Toulouse, France

Competing interests
No competing interests declared.
Aurélien Olichon

Inserm, UMR 1037-CRCT, Toulouse, France

For correspondence
aurelien.olichon@inserm.fr

Competing interests
Aurélien Olichon, Co-inventor on a patent application (filled under ref: WO/2015/063331) that covers the hs2dAb scaffold and the commercial use of the library. The library has been licensed to Hybrigenics Service SA, which will perform screens on a fee-for-service basis.
Franck Perez

Institut Curie, Paris, France

For correspondence
Franck.Perez@curie.fr

Competing interests
Franck Perez, Co-inventor on a patent application (filled under ref: WO/2015/063331) that covers the hs2dAb scaffold and the commercial use of the library. The library has been licensed to Hybrigenics Service SA, which will perform screens on a fee-for-service basis.

"This ORCID iD identifies the author of this article:" 0000-0002-9129-9401

Funding

Institut Curie

Sandrine Moutel
Virginie Bernard
Ario de Marco
Franck Perez

Centre National de la Recherche Scientifique

Sandrine Moutel
Franck Perez

Institut National de la Santé et de la Recherche Médicale

Nicolas Bery
Laura Keller
Laetitia Ligat
Gilles Favre
Aurélien Olichon

Agence Nationale de la Recherche (ANR-09-BIOT-05)

Sandrine Moutel
Jean-Christophe Rain
Franck Perez

Institut National de la Santé et de la Recherche Médicale (ITS-201103)

Ario de Marco
Franck Perez

Fondation pour la Recherche Médicale (DEQ20120323723)

Franck Perez

Groupe de Recherche of the Claudius Regaud Institute

Gilles Favre
Aurélien Olichon

LABEX CellTisPhyBio (11-LBX-0038)

Sandrine Moutel
Virginie Bernard
Ario de Marco
Franck Perez

IDEX Paris Sciences Lettres (ANR-10-IDEX-0001-02 PSL)

Sandrine Moutel
Virginie Bernard
Ario de Marco
Franck Perez

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.