| Status | 已發表Published |
| Discovery and Development of Therapeutic TACE (ADAM17) Antibodies | |
Kwok, H. F.  ; Botkjaer, K.; Tape, C.; Huang, Y.; McCafferty, J.; Murphy, G.
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| 2015-03-01 | |
| Source Publication | Frontiers In Cancer Biology & Drug Development
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| Abstract | TNF-Alpha Converting Enzyme (TACE) is a membrane-bound zinc metalloprotease (MP) that may play a significant role in tumour biology, notably by the conversion of many inactive cell-surface ligands into active soluble ligands; e.g. ErbB-ligands such as TGF-Alpha, HB-EGF, EGF, Amphiregulin and Epigen. Moreover, TACE can stimulate local inflammation by solubilising TNF-Alpha, and aid immunological evasion by removing tumour cell surface MICA. Due to the homology between MP active sites, the development of small molecule TACE inhibitors has been plagued with unwanted nonspecific MP activity. Recently, our group has successfully produced a specific inhibitory TACE antibody [D1(A12)] and showed the effective inhibition of TACE. However, D1(A12) only recognises human version of TACE and cannot be used for in vivo applications in mouse cancer models. Here we set out to further develop a ‘Human and Mouse Cross-Reactive’ anti-TACE antibody as part of an anti-cancer drug development programme. In this study, we developed clinically applicable inhibitory antibodies using contemporary phage-display technology. Therapeutic antibodies must be of human origin if they are to survive in vivo. We exposed human TACE ectodomain to a phagedisplay scFv library in the initial selection and further included the mouse TACE ectodomain in the affinity maturation selection. We totally screened over 2,000 individual antibody fragments; and a portfolio of TACE-binding ScFvs was identified, expressed and purified. These antibodies were screened for their ability to bind both recombinant human and mouse TACE and biochemically inhibit TACE-mediated proteolytic activity. The novel ‘Human and Mouse Cross-Reactive’ TACE inhibitory antibody A9 was identified. Initial screening identified ScFv clone A9 as an efficient inhibitor of both recombinant human & mouse TACE biochemical activity. However, the kinetic data suggested the half-life of A9 ScFv was inadequate for in vivo work. To further develop A9 as a prototype drug, the A9 ScFv was reformatted to a clinically applicable human IgG1 format. BIAcore kinetic analysis revealed that A9 IgG1 had a nanomolar affinity [KD = approx. 5-10nM] and retained its cross-reactivity with human and mouse TACE. The reformatted A9 has subsequently proved to be an inhibitor of cell surface TACE activity in mouse cancer cell shedding activities in vitro. Unfortunately, with the 250nM IC50 in the cell based assay, the affinity of A9 IgG1 will not adequate for in vivo applications. In order to improve the affinity of A9 to address this issue, we further perform affinity maturation using complementarity determining region (CDR) randomisation approach. The improved A9 derivatives with mature subnanomolar affinities are currently being screened and will be selected for specific inhibitory TACE antibody leads for in vivo applications in mouse cancer models. |
| Keyword | Cross-Reactive Anti-TACE Antibody Mouse Models |
| Language | 英語English |
| The Source to Article | PB_Publication |
| PUB ID | 13921 |
| Document Type | Conference paper |
| Collection | DEPARTMENT OF BIOMEDICAL SCIENCES Faculty of Health Sciences |
| Corresponding Author | Kwok, H. F. |
| Recommended Citation GB/T 7714 | Kwok, H. F.,Botkjaer, K.,Tape, C.,et al. Discovery and Development of Therapeutic TACE (ADAM17) Antibodies[C], 2015. |
| APA | Kwok, H. F.., Botkjaer, K.., Tape, C.., Huang, Y.., McCafferty, J.., & Murphy, G. (2015). Discovery and Development of Therapeutic TACE (ADAM17) Antibodies. Frontiers In Cancer Biology & Drug Development. |
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