Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Design of phosphodiesterase 4D (PDE4D) allosteric modulators for enhancing cognition with improved safety

Abstract

Phosphodiesterase 4 (PDE4), the primary cAMP-hydrolyzing enzyme in cells, is a promising drug target for a wide range of conditions. Here we present seven co-crystal structures of PDE4 and bound inhibitors that show the regulatory domain closed across the active site, thereby revealing the structural basis of PDE4 regulation. This structural insight, together with supporting mutagenesis and kinetic studies, allowed us to design small-molecule allosteric modulators of PDE4D that do not completely inhibit enzymatic activity (Imax 80–90%). These allosteric modulators have reduced potential to cause emesis, a dose-limiting side effect of existing active site–directed PDE4 inhibitors, while maintaining biological activity in cellular and in vivo models. Our results may facilitate the design of CNS therapeutics modulating cAMP signaling for the treatment of Alzheimer's disease, Huntington's disease, schizophrenia and depression, where brain distribution is desired for therapeutic benefit.

Your institute does not have access to this article

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: PDE4D Constructs.
Figure 2: Capping the active site by UCR2.
Figure 3: A common pharmacophore for PDE4 inhibitors accessing the UCR2 binding pose.
Figure 4: Critical pharmacophore elements that determine partial kinetic behavior of PDE4 allosteric modulators.
Figure 5: Effects of compounds on mouse models of cognition and a behavioral correlate of emesis.
Figure 6: Effects of compounds on emesis.
Figure 7: A model to explain how PDE4 regulatory domains control PDE4 activity.

Accession codes

Accessions

Protein Data Bank

References

  1. Omori, K. & Kotera, J. Overview of PDEs and their regulation. Circ. Res. 100, 309–327 (2007).

    CAS  PubMed  Google Scholar 

  2. Wang, H., Robinson, H. & Ke, H. The molecular basis for different recognition of substrates by phosphodiesterase families 4 and 10. J. Mol. Biol. 371, 302–307 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Bolger, G. et al. A family of human phosphodiesterases homologous to the dunce learning and memory gene product of Drosophila melanogaster are potential targets for antidepressant drugs. Mol. Cell. Biol. 13, 6558–6571 (1993).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Jacobitz, S., McLaughlin, M.M., Livi, G.P., Burman, M. & Torphy, T.J. Mapping the functional domains of human recombinant phosphodiesterase 4A: structural requirements for catalytic activity and rolipram binding. Mol. Pharmacol. 50, 891–899 (1996).

    CAS  PubMed  Google Scholar 

  5. Rocque, W.J. et al. Human recombinant phosphodiesterase 4B2B binds (R)-rolipram at a single site with two affinities. Biochemistry 36, 14250–14261 (1997).

    CAS  PubMed  Google Scholar 

  6. Beard, M.B. et al. UCR1 and UCR2 domains unique to the cAMP-specific phosphodiesterase family form a discrete module via electrostatic interactions. J. Biol. Chem. 275, 10349–10358 (2000).

    CAS  PubMed  Google Scholar 

  7. MacKenzie, S.J. et al. Long PDE4 cAMP specific phosphodiesterases are activated by protein kinase A-mediated phosphorylation of a single serine residue in Upstream Conserved Region 1 (UCR1). Br. J. Pharmacol. 136, 421–433 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Sette, C. & Conti, M. Phosphorylation and activation of a cAMP-specific phosphodiesterase by the cAMP-dependent protein kinase. Involvement of serine 54 in the enzyme activation. J. Biol. Chem. 271, 16526–16534 (1996).

    CAS  PubMed  Google Scholar 

  9. Shakur, Y., Pryde, J.G. & Houslay, M.D. Engineered deletion of the unique N-terminal domain of the cyclic AMP-specific phosphodiesterase RD1 prevents plasma membrane association and the attainment of enhanced thermostability without altering its sensitivity to inhibition by rolipram. Biochem. J. 292, 677–686 (1993).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Bolger, G.B. et al. The unique amino-terminal region of the PDE4D5 cAMP phosphodiesterase isoform confers preferential interaction with beta-arrestins. J. Biol. Chem. 278, 49230–49238 (2003).

    CAS  PubMed  Google Scholar 

  11. Bolger, G.B. et al. Scanning peptide array analyses identify overlapping binding sites for the signalling scaffold proteins, beta-arrestin and RACK1, in cAMP-specific phosphodiesterase PDE4D5. Biochem. J. 398, 23–36 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Lehnart, S.E. et al. Phosphodiesterase 4D deficiency in the ryanodine-receptor complex promotes heart failure and arrhythmias. Cell 123, 25–35 (2005).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Blokland, A., Schreiber, R. & Prickaerts, J. Improving memory: a role for phosphodiesterases. Curr. Pharm. Des. 12, 2511–2523 (2006).

    CAS  PubMed  Google Scholar 

  14. Houslay, M.D., Schafer, P. & Zhang, K.Y. Keynote review: phosphodiesterase-4 as a therapeutic target. Drug Discov. Today 10, 1503–1519 (2005).

    CAS  PubMed  Google Scholar 

  15. DeMarch, Z., Giampa, C., Patassini, S., Bernardi, G. & Fusco, F.R. Beneficial effects of rolipram in the R6/2 mouse model of Huntington's disease. Neurobiol. Dis. 30, 375–387 (2008).

    CAS  PubMed  Google Scholar 

  16. Zhang, H.T. Cyclic AMP-specific phosphodiesterase-4 as a target for the development of antidepressant drugs. Curr. Pharm. Des. 15, 1688–1698 (2009).

    CAS  PubMed  Google Scholar 

  17. Giembycz, M.A. Life after PDE4: overcoming adverse events with dual-specificity phosphodiesterase inhibitors. Curr. Opin. Pharmacol. 5, 238–244 (2005).

    CAS  PubMed  Google Scholar 

  18. Spina, D. PDE4 inhibitors: current status. Br. J. Pharmacol. 155, 308–315 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Robichaud, A. et al. Deletion of phosphodiesterase 4D in mice shortens alpha(2)-adrenoceptor-mediated anesthesia, a behavioral correlate of emesis. J. Clin. Invest. 110, 1045–1052 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Robichaud, A., Savoie, C., Stamatiou, P.B., Tattersall, F.D. & Chan, C.C. PDE4 inhibitors induce emesis in ferrets via a noradrenergic pathway. Neuropharmacology 40, 262–269 (2001).

    CAS  PubMed  Google Scholar 

  21. Aoki, M. et al. Studies on mechanisms of low emetogenicity of YM976, a novel phosphodiesterase type 4 inhibitor. J. Pharmacol. Exp. Ther. 298, 1142–1149 (2001).

    CAS  PubMed  Google Scholar 

  22. Giembycz, M.A. Development status of second generation PDE4 inhibitors for asthma and COPD: the story so far. Monaldi Arch. Chest Dis. 57, 48–64 (2002).

    CAS  PubMed  Google Scholar 

  23. Conn, P.J., Christopoulos, A. & Lindsley, C.W. Allosteric modulators of GPCRs: a novel approach for the treatment of CNS disorders. Nat. Rev. Drug Discov. 8, 41–54 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Altucci, L., Leibowitz, M.D., Ogilvie, K.M., de Lera, A.R. & Gronemeyer, H. RAR and RXR modulation in cancer and metabolic disease. Nat. Rev. Drug Discov. 6, 793–810 (2007).

    CAS  PubMed  Google Scholar 

  25. Hoffmann, R., Wilkinson, I.R., McCallum, J.F., Engels, P. & Houslay, M.D. cAMP-specific phosphodiesterase HSPDE4D3 mutants which mimic activation and changes in rolipram inhibition triggered by protein kinase A phosphorylation of Ser-54: generation of a molecular model. Biochem. J. 333, 139–149 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Houslay, M.D. & Adams, D.R. PDE4 cAMP phosphodiesterases: modular enzymes that orchestrate signalling cross-talk, desensitization and compartmentalization. Biochem. J. 370, 1–18 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Saldou, N. et al. Comparison of recombinant human PDE4 isoforms: interaction with substrate and inhibitors. Cell. Signal. 10, 427–440 (1998).

    CAS  PubMed  Google Scholar 

  28. Souness, J.E. & Rao, S. Proposal for pharmacologically distinct conformers of PDE4 cyclic AMP phosphodiesterases. Cell. Signal. 9, 227–236 (1997).

    CAS  PubMed  Google Scholar 

  29. Brideau, C., Van Staden, C., Styhler, A., Rodger, I.W. & Chan, C.C. The effects of phosphodiesterase type 4 inhibitors on tumour necrosis factor-alpha and leukotriene B4 in a novel human whole blood assay. Br. J. Pharmacol. 126, 979–988 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Reid, P. Roflumilast Altana Pharma. Curr. Opin. Investig. Drugs 3, 1165–1170 (2002).

    CAS  PubMed  Google Scholar 

  31. Lorimer, D. et al. Gene Composer: database software for protein construct design, codon engineering, and gene synthesis. BMC Biotechnol. 9, 36 (2009).

    PubMed  PubMed Central  Google Scholar 

  32. Raymond, A. et al. Combined protein construct and synthetic gene engineering for heterologous protein expression and crystallization using Gene Composer. BMC Biotechnol. 9, 37 (2009).

    PubMed  PubMed Central  Google Scholar 

  33. Lim, J., Pahlke, G. & Conti, M. Activation of the cAMP-specific phosphodiesterase PDE4D3 by phosphorylation. Identification and function of an inhibitory domain. J. Biol. Chem. 274, 19677–19685 (1999).

    CAS  PubMed  Google Scholar 

  34. Wang, P. et al. Expression, purification, and characterization of human cAMP-specific phosphodiesterase (PDE4) subtypes A, B, C, and D. Biochem. Biophys. Res. Commun. 234, 320–324 (1997).

    CAS  PubMed  Google Scholar 

  35. Xu, R.X. et al. Crystal structures of the catalytic domain of phosphodiesterase 4B complexed with AMP, 8-Br-AMP, and rolipram. J. Mol. Biol. 337, 355–365 (2004).

    CAS  PubMed  Google Scholar 

  36. Robichaud, A., Tattersall, F.D., Choudhury, I. & Rodger, I.W. Emesis induced by inhibitors of type IV cyclic nucleotide phosphodiesterase (PDE IV) in the ferret. Neuropharmacology 38, 289–297 (1999).

    CAS  PubMed  Google Scholar 

  37. Card, G.L. et al. Structural basis for the activity of drugs that inhibit phosphodiesterases. Structure 12, 2233–2247 (2004).

    CAS  PubMed  Google Scholar 

  38. McCahill, A. et al. In resting COS1 cells a dominant negative approach shows that specific, anchored PDE4 cAMP phosphodiesterase isoforms gate the activation, by basal cyclic AMP production, of AKAP-tethered protein kinase A type II located in the centrosomal region. Cell. Signal. 17, 1158–1173 (2005).

    CAS  PubMed  Google Scholar 

  39. Chambers, R.J. et al. A new chemical tool for exploring the role of the PDE4D isozyme in leukocyte function. Bioorg. Med. Chem. Lett. 16, 718–721 (2006).

    CAS  PubMed  Google Scholar 

  40. Souness, J.E. et al. Suppression of eosinophil function by RP 73401, a potent and selective inhibitor of cyclic AMP-specific phosphodiesterase: comparison with rolipram. Br. J. Pharmacol. 115, 39–46 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Mihara, T. et al. Pharmacological characterization of a novel, potent adenosine A1 and A2A receptor dual antagonist, 5-[5-amino-3-(4-fluorophenyl)pyrazin-2-yl]-1-isopropylpyridine-2(1H)-one (ASP5854), in models of Parkinson's disease and cognition. J. Pharmacol. Exp. Ther. 323, 708–719 (2007).

    CAS  PubMed  Google Scholar 

  42. Bailey, C.H., Bartsch, D. & Kandel, E.R. Toward a molecular definition of long-term memory storage. Proc. Natl. Acad. Sci. USA 93, 13445–13452 (1996).

    CAS  PubMed  Google Scholar 

  43. Robichaud, A. et al. Assessing the emetic potential of PDE4 inhibitors in rats. Br. J. Pharmacol. 135, 113–118 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Hirose, R. et al. Correlation between emetic effect of phosphodiesterase 4 inhibitors and their occupation of the high-affinity rolipram binding site in Suncus murinus brain. Eur. J. Pharmacol. 573, 93–99 (2007).

    CAS  PubMed  Google Scholar 

  45. Ueno, S., Matsuki, N. & Saito, H. Suncus murinus: a new experimental model in emesis research. Life Sci. 41, 513–518 (1987).

    CAS  PubMed  Google Scholar 

  46. MacKenzie, S.J., Baillie, G.S., McPhee, I., Bolger, G.B. & Houslay, M.D. ERK2 mitogen-activated protein kinase binding, phosphorylation, and regulation of the PDE4D cAMP-specific phosphodiesterases. The involvement of COOH-terminal docking sites and NH2-terminal UCR regions. J. Biol. Chem. 275, 16609–16617 (2000).

    CAS  PubMed  Google Scholar 

  47. Millar, J.K. et al. DISC1 and PDE4B are interacting genetic factors in schizophrenia that regulate cAMP signaling. Science 310, 1187–1191 (2005).

    CAS  PubMed  Google Scholar 

  48. Verde, I. et al. Myomegalin is a novel protein of the golgi/centrosome that interacts with a cyclic nucleotide phosphodiesterase. J. Biol. Chem. 276, 11189–11198 (2001).

    CAS  PubMed  Google Scholar 

  49. Bolger, G.B. et al. Attenuation of the activity of the cAMP-specific phosphodiesterase PDE4A5 by interaction with the immunophilin XAP2. J. Biol. Chem. 278, 33351–33363 (2003).

    CAS  PubMed  Google Scholar 

  50. Millar, J.K. et al. Genomic structure and localisation within a linkage hotspot of Disrupted In Schizophrenia 1, a gene disrupted by a translocation segregating with schizophrenia. Mol. Psychiatry 6, 173–178 (2001).

    CAS  PubMed  Google Scholar 

  51. Murdoch, H. et al. Isoform-selective susceptibility of DISC1/phosphodiesterase-4 complexes to dissociation by elevated intracellular cAMP levels. J. Neurosci. 27, 9513–9524 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Zhang, J. et al. Histopathology of vascular injury in Sprague-Dawley rats treated with phosphodiesterase IV inhibitor SCH 351591 or SCH 534385. Toxicol. Pathol. 36, 827–839 (2008).

    CAS  PubMed  Google Scholar 

  53. Naganuma, K. et al. Discovery of selective PDE4B inhibitors. Bioorg. Med. Chem. Lett. 19, 3174–3176 (2009).

    CAS  PubMed  Google Scholar 

  54. Hersperger, R., Bray-French, K., Mazzoni, L. & Muller, T. Palladium-catalyzed cross-coupling reactions for the synthesis of 6, 8-disubstituted 1,7-naphthyridines: a novel class of potent and selective phosphodiesterase type 4D inhibitors. J. Med. Chem. 43, 675–682 (2000).

    CAS  PubMed  Google Scholar 

  55. Singh, J. et al. Biaryl inhibitors for treating pulmonary and cardiovascular disorders. PCT/US2008/084193 (2008).

  56. Deschenes, D. et al. Substituted 8-arylquinoline phosphodiesterase-4 inhibitors. WO 94/22852 (2000).

  57. Wilhelm, R. et al. Optionally substituted pyrido[2,3-D]pyridine-2,4(1H,3H)-diones and pyrido[2,3-D]pyrimidine-2(1H,3H)-ones. US 5,264,437 (1993).

Download references

Acknowledgements

The development of Gene Composer software used to design protein constructs was supported in part by the National Institute of General Medical Sciences–National Center for Research Resources, co-sponsored PSI-2 Specialized Center Grant U54 GM074961 for the Accelerated Technologies Center for Gene to 3D Structure. The authors would like to thank M. Smith, M.H. Haraldsson, G.V. Halldorsdottir, B.B. Sigurdsson, G. Bragason, I. Saemundsdottir, B. Gudmundsdottir, T.J. Dagbjartsdottir, K. Astradsdottir, S. Gunnarsdottir, B. Eiriksdottir, N. Zhou, D. Sullins, P. Rauen, A. Motta, W. Zeller, J. Christensen and M. O'Connell for contributions to the research. We also thank Dr. Akira Ito and colleagues at Dainippon Sumitomo and Dr. Klaus Mendla and colleagues at Boerhinger Ingelheim for contributions to the animal studies.

Author information

Authors and Affiliations

Authors

Contributions

A.B.B., P.W., B.L.S., L.J.S. and M.E.G. contributed to the structural and molecular biology experiments. O.T.M., J.M.B, M.T., S.H. and M.E.G. contributed to kinetic, safety and efficacy studies. J.S., T.H., A.S.K. and M.E.G. contributed to medicinal chemistry experiments. A.B.B. and M.E.G. wrote the manuscript.

Corresponding authors

Correspondence to Alex B Burgin or Mark E Gurney.

Ethics declarations

Competing interests

The authors are employees of deCODE genetics, Inc. or subsidiaries thereof.

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–16, Supplementary Tables 1–4 and Supplementary Notes (PDF 1076 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Burgin, A., Magnusson, O., Singh, J. et al. Design of phosphodiesterase 4D (PDE4D) allosteric modulators for enhancing cognition with improved safety. Nat Biotechnol 28, 63–70 (2010). https://doi.org/10.1038/nbt.1598

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt.1598

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing