Alternative non-antibody scaffolds for molecular recognition

Skerra, Arne

doi:10.1016/j.copbio.2007.04.010

Cited by 275 publications

(199 citation statements)

References 50 publications

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“…Thus, the VLR system of jawless vertebrates truly behaves like the Ig-based system of jawed vertebrates, with the potential to provide an effective humoral antipathogen shield. VLRs also hold considerable potential as natural non-Ig antibodies for various biotechnology applications (27,28). These highly stable modular single-chain polypeptides of relatively small size (15-25 kDa) can bind a broad range of antigenic determinants with high affinity and specificity, and can be readily engineered for improved binding properties.…”

Section: <-----Lrrnt -----><------Lrr1 ------><----Lrrve ------><---Cp ---><-----------------Lrrct ----------------mentioning

confidence: 99%

High-affinity lamprey VLRA and VLRB monoclonal antibodies

Tasumi

Velikovsky

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

104

138

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Section: <-----Lrrnt -----><------Lrr1 ------><----Lrrve ------><---Cp ---><-----------------Lrrct ----------------mentioning

confidence: 99%

High-affinity lamprey VLRA and VLRB monoclonal antibodies

Tasumi

Velikovsky

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

104

138

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“…Several types of proteins have been used as scaffolds to create collections ("libraries") of potential binders (4). A suitable scaffold protein contains a robust structurally defined framework and a spatially defined surface area that may be more or less randomly modified.…”

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confidence: 99%

Tumor Targeting Using Affibody Molecules: Interplay of Affinity, Target Expression Level, and Binding Site Composition

Tolmachev

Tran

Rosik³

et al. 2012

J Nucl Med

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Radionuclide imaging of cancer-associated molecular alterations may contribute to patient stratification for targeting therapy. Scaffold high-affinity proteins, such as Affibody molecules, are a new, promising class of probes for in vivo imaging. Methods. The effects of human epidermal growth factor receptor 2 (HER2) affinity and binding site composition of HER2-binding Affibody molecules, and of the HER2 density on the tumor targeting, were studied in vivo. The tumor uptake and tumor-to-organ ratios of Affibody molecules with moderate (dissociation constant [K D ] 5 10 29 M) or high (K D 5 10 210 M) affinity were compared between tumor xenografts with a high (SKOV-3) and low (LS174T) HER2 expression level in BALB/C nu/nu mice. Two Affibody molecules with similar affinity (K D 5 10 210 M) but having alternative amino acids in the binding site were compared. Results. In SKOV-3 xenografts, uptake was independent of affinity at 4 h after injection, but high-affinity binders provided 2-fold-higher tumor radioactivity retention at 24 h. In LS174T xenografts, uptake of high-affinity probes was already severalfold higher at 4 h after injection, and the difference was increased at 24 h. The clearance rate and tumor-to-organ ratios were influenced by the amino acid composition of the binding surface of the tracer protein. Conclusion. The optimal affinity of HER2-binding Affibody molecules depends on the expression of a molecular target. At a high expression level (.10 6 receptors per cell), an affinity in the low-nanomolar range is sufficient. At moderate expression, subnanomolar affinity is desirable. The binding site composition can influence the imaging contrast. This information may be useful for development of imaging agents based on scaffold affinity proteins.

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“…S8). Thus, KD scaffold with two potential binding surfaces for target recognition can be developed into bivalent or bispecific, but single domain variants, such as agonists inducing receptor clustering, providing a distinct advantage over the reported non-Ab scaffolds with a single binding surface (2)(3)(4)(5)(6)(7)(8)(9). Further, the small and modular structural features of the KD scaffold might offer favorable properties of non-Ab domain scaffolds (2, 3), such as cheap production in a microbial system and easy modifications for fusion with effector molecules (e.g., Fc fusion) and multivalent or multispecific constructs with enhanced functionality by tandem linkages of the same or different KD variants using flexible linkers.…”

Section: Discussionmentioning

confidence: 99%

“…Target-specific binding proteins with high affinity can be isolated from a synthetic library of alternative protein scaffolds, in which particular regions on a rigid scaffold core have been randomized (4)(5)(6)(7)(8)(9)(10). These scaffolds often have favorable characteristics, such as small size, high stability, ease of modification, and/or costefficient production, compared to Abs (2,3).…”

mentioning

confidence: 99%

Engineering of a human kringle domain into agonistic and antagonistic binding proteins functioning in vitro and in vivo

Lee

Park

Sung

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Here, we report the development of target-specific binding proteins based on the kringle domain (KD) (∼80 residues), a ubiquitous modular structural unit occurring across eukaryotic species. By exploiting the highly conserved backbone folding by core residues, but using extensive sequence variations in the seven loop regions of naturally occurring human KDs, we generated a synthetic KD library on the yeast cell surface by randomizing 45 residues in the loops of a human KD template. We isolated KD variants that specifically bind to anticancer target proteins, such as human death receptor 4 (DR4) and/or DR5, and that function as agonists to induce apoptotic cell death in several cancer cell lines in vitro and inhibit tumor progression in mouse models. Combined treatments with KD variants possessing different recognition sites on the same target protein exerted synergisitic tumoricidal activities, compared to treatment with individual variants. In addition to the agonists, we isolated an antagonistic KD variant that binds human tumor necrosis factor-α (TNFα) and efficiently neutralizes TNFα-induced cytotoxicity in vitro and in vivo. The KD scaffold with seven flexible loops protruding from the central core was strongly sequence-tolerant to mutations in the loop regions, offering a potential advantage of distinct binding sites for target recognition on the single domain. Our results suggest that the KD scaffold can be used to develop target-specific binding proteins that function as agonists or antagonists toward given target molecules, indicative of their potential use as biotherapeutics.

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Alternative non-antibody scaffolds for molecular recognition

Cited by 275 publications

References 50 publications

High-affinity lamprey VLRA and VLRB monoclonal antibodies

High-affinity lamprey VLRA and VLRB monoclonal antibodies

Tumor Targeting Using Affibody Molecules: Interplay of Affinity, Target Expression Level, and Binding Site Composition

Engineering of a human kringle domain into agonistic and antagonistic binding proteins functioning in vitro and in vivo

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