Structure-based discovery of opioid analgesics with reduced side effects

Morphine is an alkaloid from the opium poppy used to treat pain. The potentially lethal side effects of morphine and related opioids—which include fatal respiratory depression—are thought to be mediated by μ-opioid-receptor (μOR) signalling through the β-arrestin pathway or by actions at other receptors. Conversely, G-protein μOR signalling is thought to confer analgesia. Here we computationally dock over 3 million molecules against the μOR structure and identify new scaffolds unrelated to known opioids. Structure-based optimization yields PZM21—a potent Gi activator with exceptional selectivity for μOR and minimal β-arrestin-2 recruitment. Unlike morphine, PZM21 is more efficacious for the affective component of analgesia versus the reflexive component and is devoid of both respiratory depression and morphine-like reinforcing activity in mice at equi-analgesic doses. PZM21 thus serves as both a probe to disentangle μOR signalling and a therapeutic lead that is devoid of many of the side effects of current opioids

Toward Male Individualization with Rapidly Mutating Y-Chromosomal Short Tandem Repeats

Peer reviewe

Comparative MD Simulations Indicate a Dual Role for Arg1323.50 in Dopamine-Dependent D2R Activation.

Residue Arg3.50 belongs to the highly conserved DRY-motif of class A GPCRs, which is located at the bottom of TM3. On the one hand, Arg3.50 has been reported to help stabilize the inactive state of GPCRs, but on the other hand has also been shown to be crucial for stabilizing active receptor conformations and mediating receptor-G protein coupling. The combined results of these studies suggest that the exact function of Arg3.50 is likely to be receptor-dependent and must be characterized independently for every GPCR. Consequently, we now present comparative molecular-dynamics simulations that use our recently described inactive-state and Gα-bound active-state homology models of the dopamine D2 receptor (D2R), which are either bound to dopamine or ligand-free, performed to identify the function of Arg1323.50 in D2R. Our results are consistent with a dynamic model of D2R activation in which Arg1323.50 adopts a dual role, both by stabilizing the inactive-state receptor conformation and enhancing dopamine-dependent D2R-G protein coupling

Active-State Models of Ternary GPCR Complexes: Determinants of Selective Receptor-G-Protein Coupling

Based on the recently described crystal structure of the β2 adrenergic receptor - Gs-protein complex, we report the first molecular-dynamics simulations of ternary GPCR complexes designed to identify the selectivity determinants for receptor-G-protein binding. Long-term molecular dynamics simulations of agonist-bound β2AR-Gαs and D2R-Gαi complexes embedded in a hydrated bilayer environment and computational alanine-scanning mutagenesis identified distinct residues of the N-terminal region of intracellular loop 3 to be crucial for coupling selectivity. Within the G-protein, specific amino acids of the α5-helix, the C-terminus of the Gα-subunit and the regions around αN-β1 and α4-β6 were found to determine receptor recognition. Knowledge of these determinants of receptor-G-protein binding selectivity is essential for designing drugs that target specific receptor/G-protein combinations

Active-State Model of a Dopamine D2 Receptor - Gai Complex Stabilized by Aripiprazole-Type Partial Agonists

Partial agonists exhibit a submaximal capacity to enhance the coupling of one receptor to an intracellular binding partner. Although a multitude of studies have reported different ligand-specific conformations for a given receptor, little is known about the mechanism by which different receptor conformations are connected to the capacity to activate the coupling to G-proteins. We have now performed molecular-dynamics simulations employing our recently described active-state homology model of the dopamine D2 receptor-Gai protein-complex coupled to the partial agonists aripiprazole and FAUC350, in order to understand the structural determinants of partial agonism better. We have compared our findings with our model of the D2R-Gai-complex in the presence of the full agonist dopamine. The two partial agonists are capable of inducing different conformations of important structural motifs, including the extracellular loop regions, the binding pocket and, in particular, intracellular G-protein-binding domains. As G-protein-coupling to certain intracellular epitopes of the receptor is considered the key step of allosterically triggered nucleotide-exchange, it is tempting to assume that impaired coupling between the receptor and the G-protein caused by distinct ligand-specific conformations is a major determinant of partial agonist efficacy

Concluding model of the predicted impact of Arg132^3.50 on dopamine-dependent activation of D2R.

<p>At the inactive-state system (left row), an equilibrium between a formed and a broken ionic lock was observed, whereas the presence of dopamine was found to reduce the stability of the ionic lock. At active-state D2R (right row), dopamine increased receptor-G protein interactions via the formation of an ionic interaction between Arg132^3.50 of D2R and Asp350 of Gα_i. For clarity, TM3 and TM6 are colored in light-blue and dark-blue, respectively.</p

Distances of ionic lock residues at the inactive-state systems A and B.

<p>(A) Close view on representative conformations of an intact (= closed, grey) and a broken (= open, orange) ionic lock between residues Arg132^3.50 of TM3 (light-blue) and Glu368^6.30 of TM6 (dark-blue). In addition, Arg132^3.50 is stabilized by Asp131^3.49 of TM3. (B, C) Distances between the side chains of residues Arg132^3.50 (Cζ) and Glu368^6.30 (Cδ) in the course of the simulations A and B are shown. Cumulative occurrences of certain distances for system A (B) predominantly show distances, which are consistent with an intact ionic lock (green boxes). In contrast, the latter distances are less frequently populated at the dopamine-bound system B (C), when higher occurrences were observed for larger distances, consistent with an open ionic lock.</p

Schematic overview of the main simulation systems and their simulation times.

<p>To visually help distinguish the apo- (in which dopamine is absent, A and C) from the dopamine-bound complexes (dopamine in orange, B and D), TM 1, 2 and 7 are colored in light-grey and dark-grey, respectively. For clarity, TM3 and TM6 are colored in light-blue and dark-blue, respectively. The active-state systems (C and D) are represented by the characteristic outward movement of TM6 and the presence of Gα_i (in green). The asterisk refers to a previously published simulation. The simulation times of each system are given in bold.</p

Total occurrences of distances larger than 9.5 Å between Arg132^3.50 and Glu368^6.30 at the simulation systems A and B.

<p>The fractions of simulation time within the systems A and B, in which the distances between the Cα-atoms of Arg132^3.50 and Glu368^6.30 were found to be larger than 9.5 Å. The values above the bars represent mean ± standard error of the mean of the simulation systems A and B and indicate a higher frequency of distances larger than 9.5 Å in the presence of dopamine (unpaired t-test, two-tailed P value = 0.0960).</p

Conformational classification of the simulation systems A-D.

<p>(A, B) Intracellular view on cytoplasmic receptor domains of the simulations systems A-D indicating unchanged global conformational states throughout the MD simulations: An overlay of average structures of representative simulation systems (each derived from the final 25ns simulation times) is shown for (A) inactive-state systems A (light-grey) and B (light-orange) and (B) the active-state systems C (dark-grey) and D (brown). For comparison, the X-ray structures of inactive-state D3R (blue) and active-state β2AR (green), which were used for homology modeling, are depicted. (C, D) The distances (and occurrences of these distances) between the intracellular ends of TM3 and TM6, measured as the distances between the Cα-atoms of Arg132^3.50 and Glu368^6.30, are shown for (C) the inactive-state systems A and B and (D) the active-state systems C and D. For comparison, these distances at crystal structures of D3R and β2AR are highlighted with dashed lines. Colors are used as described above.</p