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Dual-display of small molecules enables the discovery of ligand pairs and facilitates affinity maturation

Nature Chemistry volume 7, pages 241–249 (2015)Cite this article

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Abstract

In contrast to standard fragment-based drug discovery approaches, dual-display DNA-encoded chemical libraries have the potential to identify fragment pairs that bind simultaneously and benefit from the chelate effect. However, the technology has been limited by the difficulty in unambiguously decoding the ligand pairs from large combinatorial libraries. Here we report a strategy that overcomes this limitation and enables the efficient identification of ligand pairs that bind to a target protein. Small organic molecules were conjugated to the 5′ and 3′ ends of complementary DNA strands that contain a unique identifying code. DNA hybridization followed by an inter-strand code-transfer created a stable dual-display DNA-encoded chemical library of 111,100 members. Using this approach we report the discovery of a low micromolar binder to alpha-1-acid glycoprotein and the affinity maturation of a ligand to carbonic anhydrase IX, an established marker of renal cell carcinoma. The newly discovered subnanomolar carbonic anhydrase IX binder dramatically improved tumour targeting performance in vivo.

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Figure 1: Design, synthesis and encoding of the dual-display DNA-encoded chemical library.
Figure 2: Affinity-based selection procedure for the isolation of simultaneously binding fragment pairs from a dual-display chemical library.
Figure 3: Selection results from the 111,100-member dual-display library.
Figure 4: Hit validation of selected pharmacophore pair A-117/B-113 against AGP.
Figure 5: Hit validation of selected pharmacophore pair A-493/B-202 binding to CAIX.
Figure 6: Long-lasting residence of selected library compound–dye conjugate at the tumour site.

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Acknowledgements

This work was supported by ETH Zürich, the Swiss National Science Foundation (SNSF), Philochem AG and Krebsliga Schweiz/Krebsforschung Schweiz (KFS-2839-08-2011). R.M.F. acknowledges a VPFW-ETH postdoctoral fellowship endowed by ETH Zurich and Marie-Curie actions. The authors thank M. Jaggi, I. Mafli and A. Nauer for help with library and ligand synthesis, C. Aquino and L. Opitz (Functional Genomics Center Zurich) for help with high-throughput DNA sequencing, Y. Zhang, F. Buller, H. Röst, M. Stravs, G. Jackson and A. Rabenseifner for help with sequencing data analysis and software implementation, and L. Urner for help with NMR spectra analysis. The authors also thank C. Hess, G. Hausammann, T. Hemmerle, M. Weber, E. Perrino, A. Baumann, M. Bühler and A. Zemann for help with experimental work. The authors are grateful to I. Jelezarov for critically reviewing the ITC data and to F. Samain for discussions. The authors thank J. Kunze and D. Reker for technical support with protein graphics implementation. Instant JChem (ChemAxon) was used for structure and data management (http://www.chemaxon.com).

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Authors and Affiliations

  1. Department of Chemistry and Applied Biosciences, Institute of Pharmaceutical Sciences, ETH Zurich, Zurich, 8093, Switzerland

    Moreno Wichert, Nikolaus Krall, Willy Decurtins, Raphael M. Franzini, Petra Schneider, Dario Neri & Jörg Scheuermann

  2. Philochem AG, Otelfingen, 8112, Switzerland

    Francesca Pretto

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Contributions

M.W., D.N. and J.S. designed the project. M.W. and J.S. constructed the dual-display library. W.D. and R.F. provided target proteins. W.D. designed and performed the selections. M.W., W.D. and J.S. analysed high-throughput DNA screening data. M.W. and P.S. performed ITC experiments. M.W. performed hit validation experiments. M.W., N.K. and F.P. performed in vivo experiments. M.W., D.N. and J.S. wrote the manuscript.

Corresponding authors

Correspondence to Dario Neri or Jörg Scheuermann.

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Competing interests

D.N. is a co-founder and shareholder of Philochem AG (Otelfingen, Switzerland) and J.S. is a board member of Philochem AG.

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Wichert, M., Krall, N., Decurtins, W. et al. Dual-display of small molecules enables the discovery of ligand pairs and facilitates affinity maturation. Nature Chem 7, 241–249 (2015). https://doi.org/10.1038/nchem.2158

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