Human anti-CD30 recombinant antibodies by guided phage antibody selection using cell panning

Abstract

SummaryIn various clinical studies, Hodgkin's patients have been treated with anti-CD30 immunotherapeutic agents and have shown promising responses. One of the problems that appeared from these studies is the development of an immune response against the nonhuman therapeutics, which limits repeated administration and reduces efficacy. We have set out to make a recombinant, human anti-CD30 single-chain variable fragment (scFv) antibody, which may serve as a targeting moiety with reduced immunogenicity and more rapid tumour penetration in similar clinical applications. Rather than selecting a naive phage antibody library on recombinant CD30 antigen, we used guided selection of a murine antibody in combination with panning on the CD30-positive cell line L540. The murine monoclonal antibody Ki-4 was chosen as starting antibody, because it inhibits the shedding of the extracellular part of the CD30 antigen. This makes the antibody better suited for CD30-targeting than most other anti-CD30 antibodies. We have previously isolated the murine Ki-4 scFv by selecting a mini-library of hybridoma-derived phage scFv-antibodies via panning on L540 cells. Here, we report that phage display technology was successfully used to obtain a human Ki-4 scFv version by guided selection. The murine variable heavy (VH) and light (VL) chain genes of the Ki-4 scFv were sequentially replaced by human V gene repertoires, while retaining only the major determinant for epitope-specificity: the heavy-chain complementarity determining region 3 (CDR3) of murine Ki-4. After two rounds of chain shuffling and selection by panning on L540 cells, a fully human anti-CD30 scFv was selected. It competes with the parental monoclonal antibody Ki-4 for binding to CD30, inhibits the shedding of the extracellular part of the CD30 receptor from L540 cells and is thus a promising candidate for the generation of anti-CD30 immunotherapeutics.

Conversion of a murine monoclonal antibody A13 targeting epidermal growth factor receptor to a human monoclonal antibody by guided selection

“…There are several reports mentioning similar lower affinity of human clones from guided selection compared to the murine counterpart (Klimka et al, 2000;Wang et al, 2000;Osbourn et al, 2005). Human anti-CD30 mAb which was isolated by guided selection showed 10-fold lower affinity compared to the mouse mAb (Klimka et al, 2000). The lower affinity and as a result lower efficacy of human clones from guided selection require the improvement of affinity.…”

“…Others reported the retention of epitope specificity between human scFv from guided selection and murine mAb which is already characterized the epitope and efficacy (Jespers et al, 1994;Klimka et al, 2000;Figini et al, 2009). And it is the advantage of guided selection that the epitope of well characterized murine mAb is retained in the human counterpart mAb.…”

“…It is a serial transition from a single murine antibody, via murine-human chimeric forms, to a panel of human antibodies with similar characteristics to those of the starting murine antibody (Osbourn et al, 2005). Several murine mAbs were converted to human antibodies successfully with retention of specificity of original murine mAbs (Beiboer et al, 2000;Klimka et al, 2000;Wang et al, 2000;Schmidt et al, 2001;Kim and Hong, 2007).…”

A Highly Conserved Interspecies V H in the Human Genome

“…This corresponds to other examples of human V L chain selection where the guided selection did not yield the most similar human counterpart sequence. 22,26,27,29,36 For the antigen binding site, more important than sequence homology are the canonical structures defined by Chothia et al 38,39 Previous studies have shown the isolation of human antibodies by guided selection, which shared these structural elements with the parental mouse antibody. In our case the three light chains did not share similar L1 and L3 canonical structures with the mouse antibody BSW17.…”

Preparation of single chain variable fragment of MG7mAb by phage display technology

“…MG 7 antibody was confirmed to be of great value and good potency in the targeting gene therapy of gastric cancer due to the overexpression of its corresponding antigen in a large proportion of patients with gastric cancer. But owing to its murine origin, like many other similar antibodies, MG7 antibody can elicit human anti-mouse immunoreaction and thus its use in clinical practice is restricted [2,3] . One of the efficient solutions to this problem is to remove the constant region of antibody which makes main contribution to the immunogenicity of the murine antibody to human being.…”