Camelid Single-Domain Antibodies: Promises and Challenges as Lifesaving Treatments

Abstract

Since the discovery of camelid heavy-chain antibodies in 1993, there has been tremendous excitement for these antibody domains (VHHs/sdAbs/nanobodies) as research tools, diagnostics, and therapeutics. Commercially, several patents were granted to pioneering research groups in Belgium and the Netherlands between 1996-2001. Ablynx was established in 2001 with the aim of exploring the therapeutic applications and development of nanobody drugs. Extensive efforts over two decades at Ablynx led to the first approved nanobody drug, caplacizumab (Cablivi) by the EMA and FDA (2018-2019) for the treatment of rare blood clotting disorders in adults with acquired thrombotic thrombocytopenic purpura (TPP). The relatively long development time between camelid sdAb discovery and their entry into the market reflects the novelty of the approach, together with intellectual property restrictions and freedom-to-operate issues. The approval of the first sdAb drug, together with the expiration of key patents, may open a new horizon for the emergence of camelid sdAbs as mainstream biotherapeutics in the years to come. It remains to be seen if nanobody-based drugs will be cheaper than traditional antibodies. In this review, I provide critical perspectives on camelid sdAbs and present the promises and challenges to their widespread adoption as diagnostic and therapeutic agents.

Keywords: VHH; biotherapeutics; bispecific VHH; camelid heavy-chain antibody; camelid mice; caplacizumab; nanobody; single-domain antibody.

Figure 1

Schematic representation of camelid IgGs and llama serum IgG fractionation on protein A and protein G. (a) The comparative structures of each respective IgG isotype has been shown on top of lanes 2, 4, and 6. (b) Llama immunoglobulin serum was fractionated on protein G and A and ran on reducing and non-reducing SDS-PAGE. Lane 1: MW Marker; lane 2: IgG2 (Protein A) (non-reduced: NR); ); lane 3: IgG2 (reduced: R); lane 4: IgG1 (Protein A&G) (NR); lane 5: IgG1 (R); lane 6, IgG3 (Protein A&G)) (NR); lane 7: IgG3 (R); (c) the VHH folding structure of two β-sheets with five and four β-strands is shown on the right with CDR loops shown in dark green (CDR1), red (CDR2), and blue (CDR3).

Figure 2

Visualization of both CDR and non-CDR contacts in nanobodies. VHH A.20 (PDB 4NBX; approximately 2.5 × 4 × 3 nm in size) is shown as a (a) cartoon with an overlaid mesh and (b) surface representation. The molecule is orientated with a view looking down on the paratope, with CDR1/2/3 colored in dark green, red, and blue, respectively. The flexible non-CDR regions are highlighted in light colors, including the N-terminus (green), framework 2 (blue), C’’D loop (red), and DE loop (orange). The contact points in CDR and non-CDR hotspots for VHH A.20 with Toxin A are highlighted as yellow sticks, and β-strands are labeled A through G. Structural assignments are based on Zavrtanik et al. (2018).