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Second-generation DNA-templated macrocycle libraries for the discovery of bioactive small molecules

An Author Correction to this article was published on 01 October 2019

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Abstract

DNA-encoded libraries have emerged as a widely used resource for the discovery of bioactive small molecules, and offer substantial advantages compared with conventional small-molecule libraries. Here, we have developed and streamlined multiple fundamental aspects of DNA-encoded and DNA-templated library synthesis methodology, including computational identification and experimental validation of a 20 × 20 × 20 × 80 set of orthogonal codons, chemical and computational tools for enhancing the structural diversity and drug-likeness of library members, a highly efficient polymerase-mediated template library assembly strategy, and library isolation and purification methods. We have integrated these improved methods to produce a second-generation DNA-templated library of 256,000 small-molecule macrocycles with improved drug-like physical properties. In vitro selection of this library for insulin-degrading enzyme affinity resulted in novel insulin-degrading enzyme inhibitors, including one of unusual potency and novel macrocycle stereochemistry (IC50 = 40 nM). Collectively, these developments enable DNA-templated small-molecule libraries to serve as more powerful, accessible, streamlined and cost-effective tools for bioactive small-molecule discovery.

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Fig. 1: DNA-templated macrocycle library synthesis scheme.
Fig. 2: Identification of an orthogonal codon set for second-generation DNA-templated libraries.
Fig. 3: Building blocks for the second-generation DNA-templated macrocycle library.
Fig. 4: Distribution of physical parameters among library members from the second-generation macrocycle library and first-generation library.
Fig. 5: Approaches to the assembly of DNA template libraries.
Fig. 6: In vitro selection of the 256,000-membered DNA-templated macrocycle library for binding to IDE.

Change history

  • 01 October 2019

    An amendment to this paper has been published and can be accessed via a link at the top of the paper.

References

  1. Hüser, J., Mannhold, R., Kubinyi, H. & Folkers, G. High-Throughput Screening in Drug Discovery (Wiley, Weinheim, 2006).

  2. Macarron, R. et al. Impact of high-throughput screening in biomedical research. Nat. Rev. Drug Discov. 10, 188–195 (2011).

    CAS  Article  Google Scholar 

  3. Dandapani, S. & Marcaurelle, L. A. Grand Challenge commentary: accessing new chemical space for ‘undruggable’ targets. Nat. Chem. Biol. 6, 861–863 (2010).

    CAS  Article  Google Scholar 

  4. Brenner, S. & Lerner, R. A. Encoded combinatorial chemistry. Proc. Natl Acad. Sci. USA 89, 5381–5383 (1992).

    CAS  Article  Google Scholar 

  5. Gartner, Z. J. & Liu, D. R. The generality of DNA-templated synthesis as a basis for evolving non-natural small molecules. J. Am. Chem. Soc. 123, 6961–6963 (2001).

    CAS  Article  Google Scholar 

  6. Gartner, Z. J. et al. DNA-templated organic synthesis and selection of a library of macrocycles. Science 305, 1601–1605 (2004).

    CAS  Article  Google Scholar 

  7. Zimmermann, G. & Neri, D. DNA-encoded chemical libraries: foundations and applications in lead discovery. Drug Discov. Today 21, 1828–1834 (2016).

    CAS  Article  Google Scholar 

  8. Goodnow, R. A. A Handbook for DNA-encoded Chemistry: Theory and Applications for Exploring Chemical Space and Drug Discovery (Wiley, Hoboken, NJ, 2014).

  9. Franzini, R. M., Neri, D. & Scheuermann, J. DNA-encoded chemical libraries: advancing beyond conventional small-molecule libraries. Acc. Chem. Res. 47, 1247–1255 (2014).

    CAS  Article  Google Scholar 

  10. Krall, N., Scheuermann, J. & Neri, D. Small targeted cytotoxics: current state and promises from DNA-encoded chemical libraries. Angew. Chem. Int. Ed. 52, 1384–1402 (2013).

    CAS  Article  Google Scholar 

  11. Mannocci, L., Leimbacher, M., Wichert, M., Scheuermann, J. & Neri, D. 20 Years of DNA-encoded chemical libraries. Chem. Commun. 47, 12747–12753 (2011).

    CAS  Article  Google Scholar 

  12. Kleiner, R. E., Dumelin, C. E. & Liu, D. R. Small-molecule discovery from DNA-encoded chemical libraries. Chem. Soc. Rev. 40, 5707–5717 (2011).

    CAS  Article  Google Scholar 

  13. Scheuermann, J. & Neri, D. DNA-encoded chemical libraries: a tool for drug discovery and for chemical biology. ChemBioChem 11, 931–937 (2010).

    CAS  Article  Google Scholar 

  14. Clark, M. A. Selecting chemicals: the emerging utility of DNA-encoded libraries. Curr. Opin. Chem. Biol. 14, 396–403 (2010).

    CAS  Article  Google Scholar 

  15. Buller, F., Mannocci, L., Scheuermann, J. & Neri, D. Drug discovery with DNA-encoded chemical libraries. Bioconjug. Chem. 21, 1571–1580 (2010).

    CAS  Article  Google Scholar 

  16. Clark, M. A. et al. Design, synthesis and selection of DNA-encoded small-molecule libraries. Nat. Chem. Biol. 5, 647–654 (2009).

    CAS  Article  Google Scholar 

  17. Doyon, J. B., Snyder, T. M. & Liu, D. R. Highly sensitive in vitro selections for DNA-linked synthetic small molecules with protein binding affinity and specificity. J. Am. Chem. Soc. 125, 12372–12373 (2003).

    CAS  Article  Google Scholar 

  18. Scheuermann, J. & Neri, D. Dual-pharmacophore DNA-encoded chemical libraries. Curr. Opin. Chem. Biol. 26, 99–103 (2015).

    CAS  Article  Google Scholar 

  19. Wrenn, S. J., Weisinger, R. M., Halpin, D. R. & Harbury, P. B. Synthetic ligands discovered by in vitro selection. J. Am. Chem. Soc. 129, 13137–13143 (2007).

    CAS  Article  Google Scholar 

  20. Li, Y., Zhao, P., Zhang, M., Zhao, X. & Li, X. Multistep DNA-templated synthesis using a universal template. J. Am. Chem. Soc. 135, 17727–17730 (2013).

    CAS  Article  Google Scholar 

  21. Hansen, M. H. et al. A yoctoliter-scale DNA reactor for small-molecule evolution. J. Am. Chem. Soc. 131, 1322–1327 (2009).

    CAS  Article  Google Scholar 

  22. Chan, A. I., McGregor, L. M. & Liu, D. R. Novel selection methods for DNA-encoded chemical libraries. Curr. Opin. Chem. Biol. 26, 55–61 (2015).

    CAS  Article  Google Scholar 

  23. Satz, A. L. DNA encoded library selections and insights provided by computational simulations. ACS Chem. Biol. 10, 2237–2245 (2015).

    CAS  Article  Google Scholar 

  24. Satz, A. L. Simulated screens of DNA encoded libraries: the potential influence of chemical synthesis fidelity on interpretation of structure–activity relationships. ACS Comb. Sci. 18, 415–424 (2016).

    CAS  Article  Google Scholar 

  25. Connors, W. H., Hale, S. P. & Terrett, N. K. DNA-encoded chemical libraries of macrocycles. Curr. Opin. Chem. Biol. 26, 42–47 (2015).

    CAS  Article  Google Scholar 

  26. Levin, J. I. Macrocycles in Drug Discovery (Royal Society of Chemistry, London, 2014).

  27. Driggers, E. M., Hale, S. P., Lee, J. & Terrett, N. K. The exploration of macrocycles for drug discovery—an underexploited structural class. Nat. Rev. Drug Discov. 7, 608–624 (2008).

    CAS  Article  Google Scholar 

  28. Marsault, E. & Peterson, M. L. Macrocycles are great cycles: applications, opportunities, and challenges of synthetic macrocycles in drug discovery. J. Med. Chem. 54, 1961–2004 (2011).

    CAS  Article  Google Scholar 

  29. White, C. J. & Yudin, A. K. Contemporary strategies for peptide macrocyclization. Nat. Chem. 3, 509–524 (2011).

    CAS  Article  Google Scholar 

  30. Yudin, A. K. Macrocycles: lessons from the distant past, recent developments, and future directions. Chem. Sci. 6, 30–49 (2015).

    CAS  Article  Google Scholar 

  31. Villar, E. A. et al. How proteins bind macrocycles. Nat. Chem. Biol. 10, 723–731 (2014).

    CAS  Article  Google Scholar 

  32. Dougherty, P. G., Qian, Z. & Pei, D. Macrocycles as protein–protein interaction inhibitors. Biochem. J. 474, 1109–1125 (2017).

    CAS  Article  Google Scholar 

  33. Giordanetto, F. & Kihlberg, J. Macrocyclic drugs and clinical candidates: what can medicinal chemists learn from their properties? J. Med. Chem. 57, 278–295 (2014).

    CAS  Article  Google Scholar 

  34. Gartner, Z. J., Kanan, M. W. & Liu, D. R. Expanding the reaction scope of DNA-templated synthesis. Angew. Chem. Int. Ed. 41, 1796–1800 (2002).

    CAS  Article  Google Scholar 

  35. Gartner, Z. J., Kanan, M. W. & Liu, D. R. Multistep small-molecule synthesis programmed by DNA templates. J. Am. Chem. Soc. 124, 10304–10306 (2002).

    CAS  Article  Google Scholar 

  36. Li, X. & Liu, D. R. DNA-templated organic synthesis: nature’s strategy for controlling chemical reactivity applied to synthetic molecules. Angew. Chem. Int. Ed. 43, 4848–4870 (2004).

    CAS  Article  Google Scholar 

  37. Calderone, C. T., Puckett, J. W., Gartner, Z. J. & Liu, D. R. Directing otherwise incompatible reactions in a single solution by using DNA-templated organic synthesis. Angew. Chem. Int. Ed. 41, 4104–4108 (2002).

    CAS  Article  Google Scholar 

  38. O’Reilly, R. K., Turberfield, A. J. & Wilks, T. R. The evolution of DNA-templated synthesis as a tool for materials discovery. Acc. Chem. Res. 50, 2496–2509 (2017).

    Article  Google Scholar 

  39. Malone, M. L. & Paegel, B. M. What is a ‘DNA-compatible’ reaction? ACS Comb. Sci. 18, 182–187 (2016).

    CAS  Article  Google Scholar 

  40. Satz, A. L. et al. DNA compatible multistep synthesis and applications to DNA encoded libraries. Bioconjug. Chem. 26, 1623–1632 (2015).

    CAS  Article  Google Scholar 

  41. Tse, B. N., Snyder, T. M., Shen, Y. & Liu, D. R. Translation of DNA into a library of 13000 synthetic small-molecule macrocycles suitable for in vitro selection. J. Am. Chem. Soc. 130, 15611–15626 (2008).

    CAS  Article  Google Scholar 

  42. Mullard, A. DNA tags help the hunt for drugs. Nature 530, 367–369 (2016).

    CAS  Article  Google Scholar 

  43. Kleiner, R. E., Dumelin, C. E., Tiu, G. C., Sakurai, K. & Liu, D. R. In vitro selection of a DNA-templated small-molecule library reveals a class of macrocyclic kinase inhibitors. J. Am. Chem. Soc. 132, 11779–11791 (2010).

    CAS  Article  Google Scholar 

  44. Georghiou, G., Kleiner, R. E., Pulkoski-Gross, M., Liu, D. R. & Seeliger, M. A. Highly specific, bisubstrate-competitive Src inhibitors from DNA-templated macrocycles. Nat. Chem. Biol. 8, 366–374 (2012).

    CAS  Article  Google Scholar 

  45. Maianti, J. P. et al. Anti-diabetic activity of insulin-degrading enzyme inhibitors mediated by multiple hormones. Nature 511, 94–98 (2014).

    CAS  Article  Google Scholar 

  46. Aleem, S. U. et al. Structural and biochemical basis for intracellular kinase inhibition by Src-specific peptidic macrocycles. Cell Chem. Biol. 23, 1103–1112 (2016).

    CAS  Article  Google Scholar 

  47. Snyder, T. M., Tse, B. N. & Liu, D. R. Effects of template sequence and secondary structure on DNA-templated reactivity. J. Am. Chem. Soc. 130, 1392–1401 (2008).

    CAS  Article  Google Scholar 

  48. Lipinski, C. A., Lombardo, F., Dominy, B. W. & Feeney, P. J. Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv. Drug Deliv. Rev. 23, 3–25 (1997).

    CAS  Article  Google Scholar 

  49. Veber, D. F. et al. Molecular properties that influence the oral bioavailability of drug candidates. J. Med. Chem. 45, 2615–2623 (2002).

    CAS  Article  Google Scholar 

  50. Pye, C. R. et al. Nonclassical size dependence of permeation defines bounds for passive adsorption of large drug molecules. J. Med. Chem. 60, 1665–1672 (2017).

    CAS  Article  Google Scholar 

  51. Bockus, A. T. et al. Probing the physicochemical boundaries of cell permeability and oral bioavailability in lipophilic macrocycles inspired by natural products. J. Med. Chem. 58, 4581–4589 (2015).

    CAS  Article  Google Scholar 

  52. Hewitt, W. M. et al. Cell-permeable cyclic peptides from synthetic libraries inspired by natural products. J. Am. Chem. Soc. 137, 715–721 (2015).

    CAS  Article  Google Scholar 

  53. Matsson, P. & Kihlberg, J. How big is too big for cell permeability? J. Med. Chem. 60, 1662–1664 (2017).

    CAS  Article  Google Scholar 

  54. Over, B. et al. Structural and conformational determinants of macrocycle cell permeability. Nat. Chem. Biol. 12, 1065–1074 (2016).

    CAS  Article  Google Scholar 

  55. Doak, B. C., Over, B., Giordanetto, F. & Kihlberg, J. Oral druggable space beyond the rule of 5: insights from drugs and clinical candidates. Chem. Biol. 21, 1115–1142 (2014).

    CAS  Article  Google Scholar 

  56. Doak, B. C., Zheng, J., Dobritzsch, D. & Kihlberg, J. How beyond rule of 5 drugs and clinical candidates bind to their targets. J. Med. Chem. 59, 2312–2327 (2016).

    CAS  Article  Google Scholar 

  57. Matsson, P., Doak, B. C., Over, B. & Kihlberg, J. Cell permeability beyond the rule of 5. Adv. Drug Deliv. Rev. 101, 42–61 (2016).

    CAS  Article  Google Scholar 

  58. Watkins, N. E. & SantaLucia, J. Nearest-neighbor thermodynamics of deoxyinosine pairs in DNA duplexes. Nucleic Acids Res. 33, 6258–6267 (2005).

    CAS  Article  Google Scholar 

  59. Irwin, J. J. et al. An aggregation advisor for ligand discovery. J. Med. Chem. 58, 7076–7087 (2015).

    CAS  Article  Google Scholar 

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Acknowledgements

This paper is dedicated to Hisashi Yamamoto on the occasion of his 75th birthday. This work was supported by US National Institutes of Health (NIH) R35 GM118062, DARPA HR0011-17-2-0049, the Howard Hughes Medical Institute, and the F-Prime Biomedical Research Initiative.

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D.L.U. and D.R.L. designed the research and wrote the manuscript. D.L.U. conducted all the experimental, analytical and computational work for the development and synthesis of the library. Selections and library regeneration were optimized and conducted by D.L.U. and A.I.C. Macrocyclic hits were synthesized and purified by A.I.C. and IDE inhibition assays were conducted by J.P.M. All authors edited the manuscript.

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Correspondence to David R. Liu.

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Usanov, D.L., Chan, A.I., Maianti, J.P. et al. Second-generation DNA-templated macrocycle libraries for the discovery of bioactive small molecules. Nature Chem 10, 704–714 (2018). https://doi.org/10.1038/s41557-018-0033-8

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