Drug binding and subtype selectivity in G-protein-coupled receptors

Transcription

1 Drug binding and subtype selectivity in G-protein-coupled receptors Albert C. Pan The Aspect of Time in Drug Design Schloss Rauischholzhausen Marburg, Germany Thursday, March 27 th, 2014

2 D. E. Shaw Research Independent research lab founded in 2002, in New York CIty, US 100+ biologists, chemists, computer scientists and engineers Impact on human health Major tool: long timescale molecular dynamics

3 BINDING KINETICS, LEAD OPTIMIZATION AND DRUG DISCOVERY

4 Free energy Drug binding kinetics D + R DR K d = exp( G d /RT) kon G on k off G off G d D + R (unbound) DR (bound) Binding reaction coordinate

5 Residence time and efficacy A2A receptor agonists 1/k off Residence time correlates better with efficacy than affinity Data from Guo et al., British Journal of Pharmacology 166, 1846 (2012)

6 Free energy Elucidating the molecular determinants of binding kinetics has been difficult kon G on k off G off G d D + R (unbound) DR (bound) Binding reaction coordinate A. C. Pan, D. W. Borhani, R. O. Dror, D. E. Shaw, Drug Discovery Today (2013), 18, 667

7 PATHWAY AND MECHANISM OF DRUG BINDING TO G-PROTEIN-COUPLED RECEPTORS R. O. Dror, A. C. Pan, D. W. Arlow et al., PNAS (2011), 108, 13118

8 G-protein G-protein-coupled receptors Binding Membrane Activation G-protein coupling GPCR

9

10 A b-blocker binding to b 2 AR

11 Bound pose matches crystal structure Alprenolol/b 2 AR Propranolol/b 2 AR Simulation Crystal structure (Cherezov et al., 2010) Alprenolol/b 1 AR Simulation Crystal structure (Wacker et al., 2010) Simulation Crystal structure (Warne et al., 2008)

12 Binding simulations in accord with experiment Binding affinity (kcal/mol) Simulation * Caron and Lefkowitz, 1976; Limbird and Lefkowitz, 1976 Experiment ± * K on (M -1 s -1 ) 3.1 x x 10 7

13 Open questions How does the drug enter and exit the protein? Are there metastable conformations along that pathway? What controls binding and unbinding rates? What is the nature of the energetic barriers that control these rates? Are there alternative binding sites?

14 Drugs bind along a dominant pathway 11/12 trajectories 1/12 trajectories Drug does not bind via partitioning into the lipid

15 Metastable intermediates along the dominant binding pathway Alternative metastable binding poses observed in other trajectories

16 Two barriers to drug entry

17 Physical origin of the first barrier? Probably not due to electrostatics or protein/ligand degrees of freedom

18 Waters around ligand Dehydration may contribute to the initial barrier Unbound Extracellular Vestibule 10 Bound Time (µs) Ligand loses ~50% of associated water as it enters vestibule

19 Dehydration may contribute to the initial barrier Ligand entering the vestibule

20 Drug associates with a variety of positions on the receptor surface Density of nitrogen positions Density of ring center-of-mass positions

21 A recent crystal structure of carvedilol bound to b 1 AR is consistent with an alternative pose observed in simulation Warne et al., Structure (2012)

22 Conclusions (beta-2) Beta blockers bind along a dominant pathway Drugs tend to pause in the extracellular vestibule as they bind Drugs encounter two barriers to drug entry, one 15 Angstroms from the binding pocket Molecular dynamics are a powerful way for probing drug binding pathways in atomic detail

23 SUBTYPE SELECTIVITY OF DRUG INTERACTIONS WITH MUSCARINIC RECEPTORS A. C. Kruse, J. Hu, A. C. Pan et al., Nature (2012), 482, 552

24 Extracellular The orthosteric sites of muscarinic receptors are highly conserved M2 M3 Intracellular Orthosteric binding site Haga et al., Nature (2012) 482:547 Kruse et al., Nature (2012) 482:552

25 A metastable intermediate in the extracellular vestibule Unbinding simulation Binding simulation

26 A metastable intermediate in the extracellular vestibule Unbinding simulation Binding simulation As predicted from beta-blocker binding simulations

27 Non-conserved residues interact with the ligand in the metastable state H5 H2 F221 Y177 K523 H5 L225 F181 H7 N419 H6 M2 M3

28 Kinetic selectivity in muscarinic receptors M2 M3 Tiotropium (Spiriva) is kinetically selective for M3 Casarosa et al., JPET (2009)

29 Kinetic selectivity Lu and Tonge, Current Opinion in Chemical Biology, 14, 467 (2010)

30 Different loop dynamics may be responsible for slower M3 dissociation Extracellular Loop 2 (ECL2) M3 (Crystal-like) H5 M2 M3 H6 H7 M2 (Dynamic) Kruse et al., Nature (2012) 482:552

31 Extracellular loop dynamics in M2 M2 crystal structure M2 simulation

32 Conclusions (muscarinic) Identified and characterized a potential allosteric site in the EV of M2 and M3 Faster ligand dissociation in M2 vs. M3 may be due to different ECL2 loop dynamics

33 Acknowledgments Daniel Arlow David Borhani Ron Dror Hillary Green Paul Maragakis Ansgar Philippsen Yibing Shan Huafeng Xu and David Shaw Brian Kobilka, Andrew Kruse, Daniel Rosenbaum, and Bill Weis (Stanford) Jurgen Wess (NIH) Woody Sherman (Schrödinger)