Improved anti-IgG and HSA affinity ligands: Clinical application of VHH antibody technology

Abstract

Large scale, highly specific purification of valuable proteins from blood and removal of undesirable components promise to have wide therapeutic applications. Moreover, depletion of bulk proteins from blood is a prerequisite for clinical proteomics. Here we describe the development of specific, high affinity Camelid antibody fragments (VHH) derived from immune libraries for purification and depletion of the bulk protein HSA and IgG from human serum and plasma for therapeutic and research purposes. The anti-IgG VHH substantially improved depletion of IgGs from blood over the classical method based on protein A. To demonstrate the improved performance of VHH based IgG depletion, we analyzed the presence of auto-antibodies in human plasma before and after depletion from two groups of patients with auto-immune disease: Goodpasture syndrome (GP) and systemic lupus erythematosus (SLE). VHHs can be produced efficiently and cost effectively in Saccharomyces cerevisiae, a genetically regarded as safe (GRAS) microorganism. A good manufacturing process (GMP) for purification of these VHHs has also been developed. Moreover, as VHHs are single protein chains, they can be coupled relatively easily to solid matrices. These three factors are important for developing affinity purification medication.

Dual anti-idiotypic purification of a novel, native-format biparatopic anti-MET antibody with improved in vitro and in vivo efficacy

“…3). Altogether, these data demonstrated that the two anti-idiotype VHHs specifically recognized the CDRs of their targets with affinities allowing their capture as well as their elution21.…”

“…Thus, the physical and thermal stability, or better, the capacity of VHHs to refold completely after denaturation enables the apparent infinite reusability of the columns15. These characteristics explained that these binding domains have already been successfully applied as affinity ligands in chromatography processes covering a variety of biotherapeutics16171819, can serve as target-specific capture reagents20, and can replace the conventional and expensive protein A purification step2122.…”

Specific immuno capturing of the Staphylococcal superantigen toxic‐shock syndrome toxin‐1 in plasma

“…In addition to the immunodepletion of TSST-1 from human plasma, we believe that these columns can be equipped with other ligands in order to deplete other contaminants from plasma. Consistently, a VHH has been recently discovered which can be used to deplete the bulk plasma protein IgG (Klooster et al, 2007). This VHH display improved depletion over the classical method based on protein A depletion, and the authors concluded that the VHH can be used for antibody depletion in the auto-immune diseases Goodpasture's syndrome and systemic lupus erythematosus.…”

Nanobody-derived nanobiotechnology tool kits for diverse biomedical and biotechnology applications

“…[1][2][3] For example, antihuman IgG VHH has been developed for IgG purification and depletion from blood, outperforming canonical protein A-based method. 38 Apart from concave epitopes on properly folded proteins, nanobodies could recognize small linear peptide sequences, which has been confirmed by isolating anti-EPEA VHHs. EPEA is a C-terminal tetra-amino-acid Glu-Pro-Glu-Ala sequence that can be cloned as a tag behind any protein, 39 facilitating a rapid and robust affinity purification of proteins.…”