Recent patenting activities in the discovery and development of vasopressin V2 receptor agonists

Introduction: Vasopressin V₂ agonists are well known as effective therapies in the treatment of central diabetes insipidus and nocturnal enuresis. Furthermore, given their mode of action, these particular agonists have more recently been considered, in both the pharmaceutical industry and in academia, as viable therapies for urological conditions such as nocturia.

Areas covered: For the past 10 years, significant progress has been made in the discovery and development of vasopressin V₂ agonists. This article provides the reader with information on the recent progress in the discovery and development of these compounds based on patents published from 2002 onward. Specifically, the article looks at the discovery of new non-peptide agonists as well as well as novel formulations of the vasopressin V₂ agonist desmopressin.

Expert opinion: The V₂ receptor is currently one of the hottest therapeutic targets investigated for the treatment of urinary disorders such as nocturia and central diabetes. Over the past 10 years, significant progress has been made in the discovery and development of vasopressin V₂ receptor agonists, for the treatment of these disorders. The author anticipates that these agonists will be launched to market in the not-too-distant future.     

Recent discovery and development of non-peptide vasopressin V2 receptor agonists

Desmopressin, 1-desamino-8-D-arginine vasopressin (DDAVP), is a peptide analogue of arginine vasopressin (AVP), which is an agonist of the AVP V₂ receptor. It is used for the treatment of central diabetes insipidus, the control of nocturnal enuresis and may also be of use for controlling nocturia. However, DDAVP is a peptide analogue with variable bioavailability, even via an intranasal route or in oral formulation. Therefore, the discovery and the development of an orally effective non-peptide V₂ agonist may be particularly useful in all respects. The continuous filing of patents since 1995 evidences the importance of this field.     

Molecular modeling of vasopressin receptor and in silico screening of V1b receptor antagonists

Introduction: G protein-coupled receptors (GPCRs) are integral membrane proteins which contain seven-transmembrane-spanning alpha-helices. GPCR-mediated signaling has been associated with various human diseases, positioning GPCRs as attractive targets in the drug discovery field. Recently, through advances in protein engineering and crystallography, the number of resolved GPCR structures has increased dramatically. This growing availability of GPCR structures has greatly accelerated structure-based drug design (SBDD) and in silico screening for GPCR-targeted drug discovery.

Areas covered: The authors introduce the current status of X-ray crystallography of GPCRs and what has been revealed from the resolved crystal structures. They also review the recent advances in SBDD and in silico screening for GPCR-targeted drug discovery and discuss a docking study, using homology modeling, with the discovery of potent antagonists of the vasopressin 1b receptor.

Expert opinion: Several innovative protein engineering techniques and crystallographic methods have greatly accelerated SBDD, not only for already-resolved GPCRs but also for those structures which remain unclear. These technological advances are expected to enable the determination of GPCR-fragment complexes, making it practical to perform fragment-based drug discovery. This paves the way for a new era of GPCR-targeted drug discovery.     

Highly potent antidiuretic antagonists: conformational studies of vasopressin analogues modified with 1-naphthylalanine enantiomers at position 2

In this paper, we investigated the structure-activity relationship of two vasopressin analogues, [Cpa(1),(L-1-Nal)(2) ]AVP (I) and [Cpa(1),(D-Nal)(2) ]AVP (II) by NMR spectroscopy and molecular modeling. Both peptides exhibit antioxytocic and antipressor potency. Inversion of configuration of the residue at position 2 converted a weak antidiuretic agonist (peptide I) into a highly potent antidiuretic antagonist (peptide II). For this reason, the purpose of our study was to explain the causes of different interactions of the analogues with V(2) receptors. The results have shown that both analogues display the tendency to adopt β-turns in the 1-4 and 2-5 fragments, which is characteristic of OT and V(1a) receptors antagonists. In addition, the [Cpa(1),(L-1-Nal)(2) ]AVP (I) shows the propensity to assume β-turn at position 7,8, which is believed to enhance antidiuretic activity, although not being crucial for its appearance. Moreover, the C-terminal amide group seems to be crucial for signal transduction. Its high accessibility in [Cpa(1),(L-1-Nal)(2) ]AVP (I) in contrast to [Cpa(1),(D-1-Nal)(2) ]AVP (II), probably results in V(2) receptor activation.     

QSAR analysis and molecular modeling of ABCG2-specific inhibitors

In addition to its critical role is controlling drug availability and protecting sensitive organs and stem cells through cellular detoxification, breast cancer resistance protein (BCRP/ABCG2) plays an important role in cancer cell resistance to chemotherapy, together with P-glycoprotein/ABCB1. A main approach to abolish multidrug resistance is to find out specific inhibitors of the drug-efflux activity, able to chemosensitize cancer cell proliferation. Many efforts have been primarily focused on ABCB1, discovered thirty years ago, whereas very few studies have concerned ABCG2, identified much more recently. This review describes the main types of inhibitors presently known for ABCG2, and how quantitative structure-activity relationship analysis among series of compounds may lead to build up molecular models and pharmacophores allowing to design lead inhibitors as future candidates for clinical trials. A special attention is drawn on flavonoids which constitute a structurally-diverse class of compounds, well suited to identify potent ABCG2-specific inhibitors.     

Characterization of RWJ-351647, a novel nonpeptide vasopressin V2 receptor antagonist

1. Antagonists of the V(2) vasopressin (AVP) receptor are aquaretic agents, inhibiting water resorption without stimulating electrolyte excretion. In this set of experiments, a novel V(2) receptor antagonist, RWJ-351647, was characterized in vitro and in vivo. 2. RWJ-351647 displaced (3)H-AVP binding from cloned human V(2) and V(1A) receptors with Ki values of 1 nmol/L and 24 nmol/L. In assays using transfected HEK293 cells expressing either human or rat V(2) receptors, RWJ-351647 inhibited AVP-induced cAMP accumulation with Ki values of 3 nmol/L and 6 nmol/L, respectively. 3. RWJ-351647 was very selective in binding assays and showed only weak functional antagonist activity at either the cloned human V(1B) and oxytocin receptors or the human platelet V(1A) receptor. No agonist activity was seen with the compound at any receptor. 4. Pharmacokinetic studies in rats showed RWJ-351647 to be 41.9% bioavailable after a single oral administration. After repeated daily dosing over 5 days, the oral bioavailability remained at 43.9% with no change in the compound peak plasma levels or clearance rate. 5. In efficacy studies, RWJ-351647 increased urine output and decreased urine osmolality with oral doses as low as 0.1 mg/kg and 1.0 mg/kg in rats and cynomolgus monkeys, respectively. In a multiple dose study in primates, RWJ-351647 maintained a consistent aquaretic effect over 10 days without increasing sodium or potassium excretion. 6. In summary, RWJ-351647 was shown to be a selective and potent V(2) receptor antagonist with sustainable aquaretic activity in both rats and primates. The preclinical data suggest that RWJ-351647 is a potent and effective aquaretic agent with potential for use in diseases characterized by water retention.     

非肽类精氨酸加压素受体拮抗剂的研究进展

精氨酸加压素(AVP)可以通过与不同的AVP受体亚型结合产生抗利尿、收缩血管、加强记忆、参与体温和免疫调节、参与社会行为调节等不同的生理作用。AVP在体内参与雷诺氏综合征、痛经、焦虑症、抑郁症、慢性充血性心力衰竭、肝硬化、抗利尿激素分泌紊乱综合征等疾病的过程。新型高效的非肽类AVP受体拮抗剂一直是世界范围内的研究热点,本文从结构及构效关系等方面介绍了非肽类AVP受体拮抗剂的研究进展。     

Antidepressant and anxiolytic profiles of newly synthesized arginine vasopressin V1B receptor antagonists: TASP0233278 and TASP0390325

BACKGROUND AND PURPOSE

Vasopressin V_1B receptor antagonists may be effective for the treatment of depression and anxiety and the objective of this study was to characterize the pharmacological profiles of two newly synthesized arginine vasopressin receptor 1B (V_1B receptor) antagonists, TASP0233278 and TASP0390325.

EXPERIMENTAL APPROACH

We investigated the in vitro profiles of TASP0233278 and TASP0390325. In addition, the effect of TASP0390325 on the increase in plasma adrenocorticotropic hormone (ACTH) levels induced by corticotropin-releasing factor (CRF)/desmopressin (dDAVP) was investigated. We also investigated the antidepressant and anxiolytic profiles of TASP0233278 and TASP0390325 in animal models.

KEY RESULTS

Both TASP0233278 and TASP0390325 showed a high affinity and potent antagonist activity for V_1B receptors. Oral administration of TASP0390325 antagonized the increase in plasma ACTH levels induced by CRF/dDAVP in rats, indicating that TASP0390325 blocks the anterior pituitary V_1B receptor in vivo. Oral administration of TASP0233278 or TASP0390325 also exerted antidepressant effects in two models of depression (a forced swimming test and an olfactory bulbectomy model). Moreover, TASP0233278 improved depressive-like behaviour induced by repeated treatment with corticosterone, a model that has been shown to be resistant to treatment with currently prescribed antidepressants. In addition to depression models, TASP0233278 or TASP0390325 exerted anxiolytic effects in several anxiety models (social interaction, elevated plus-maze, stress-induced hyperthermia, separation-induced ultrasonic vocalization and sodium lactate-induced panic-like responses in panic-prone rats).

CONCLUSION

TASP0233278 and TASP0390325 are potent and orally active V_1B receptor antagonists with antidepressant and anxiolytic activities in rodents.     

Characterizing the Free‐Energy Landscape of MDM2 Protein–Ligand Interactions by Steered Molecular Dynamics Simulations

Inhibition of p53–MDM2 interaction by small molecules is considered to be a promising approach to re‐activate wild‐type p53 for tumor suppression. Several inhibitors of the MDM2–p53 interaction were designed and studied by the experimental methods and the molecular dynamics simulation. However, the unbinding mechanism was still unclear. The steered molecular dynamics simulations combined with Brownian dynamics fluctuation–dissipation theorem were employed to obtain the free‐energy landscape of unbinding between MDM2 and their four ligands. It was shown that compounds 4 and 8 dissociate faster than compounds 5 and 7 . The absolute binding free energies for these four ligands are in close agreement with experimental results. The open movement of helix II and helix IV in the MDM2 protein‐binding pocket upon unbinding is also consistent with experimental MDM2‐unbound conformation. We further found that different binding mechanisms among different ligands are associated with H‐bond with Lys51 and Glu25. These mechanistic results may be useful for improving ligand design.     

QSAR modeling on dopamine D2 receptor binding affinity of 6-methoxy benzamides

QSAR modeling was performed on 58 (S) N-[(1-ethyl-2-pyrrolidinyl) methyl]-6-methoxy benzamides as dopamine (DA) D2 receptor antagonists to identify the structural requirements for DA D2 receptor binding affinity. The study pointed out that the presence of hydrophobic substituents at R3 position and electron-donating groups at R5 position increased the biological activity. Substitutions at phenyl ring favored the binding affinity of these benzamides. Ethyl group and iodine at R3 position were advantageous to the activity whereas nitro group at phenyl ring hindered the antagonistic activity.     

Pharmacological characterization of RWJ-676070, a dual vasopressin V(1A)/V(2) receptor antagonist

The dysregulation of arginine vasopressin (AVP) release and activation of vasopressin V(1A) and V(2) receptors may play a role in disease. The in vitro and in vivo pharmacology of RWJ-676070, a potent, balanced antagonist of both the V(1A) and V(2) receptors is described. RWJ-676070 binding and intracellular functional antagonist activity was characterized using cells expressing V(1A), V(1B) or V(2) receptors. Its inhibition of V(1A) receptor-mediated contraction of vascular rings and platelet aggregation was determined. V(2) receptor-medated aquaresis was determined in rats, dogs and monkeys. V(1A) receptor-mediated inhibitory activity was assessed in vivo in a vasopressin-induced hypertension model and in normotensive rats and in two hypertensive rat models. RWJ-676070 inhibited AVP binding to human V(1A) and V(2) receptors (Ki=1 and 14 nM, respectively). RWJ-676070 inhibited V(1A) receptor-induced intracellular calcium mobilization and V(2) receptor-induced cAMP accumulation with Ki values of 14 nM and 13 nM, respectively. The compound was slightly less potent against rat V(1A) receptors. RWJ-676070 inhibited V(1A) receptor-mediated vasoconstriction in rat and dog vascular rings and AVP-induced human platelet aggregation. Dose dependent aquaresis was demonstrated in rats, dogs and monkeys following oral administration. RWJ-676070 inhibited AVP-induced hypertension in rats but had no effect on arterial pressure in normotensive and spontaneously hypertensive rats but did decrease arterial pressure in Dahl, salt-sensitive hypertensive rats. RWJ-676070 is a new, potent antagonist of V(1A) and V(2) receptors that may be useful for treatment of diseases benefiting from balanced inhibition of both V(1A) and V(2) receptors.     

Novel linear antagonists of the antidiuretic (V2) and vasopressor (V1) responses to vasopressin

We report the solid phase synthesis of a series of 16 linear analogues of the cyclic antagonist of the antidiuretic (V₂) and the vasopressor (V₁) responses to arginine vasopressin (AVP), d(CH₂)₅[d -Tyr(Et)²,Val⁴]AVP (A). Peptide 1, the linear precursor of (A), (CH₂)₅(SH)-CH₂-CO-d -Tyr(Et)-Phe-Val-Asn-Cys-Pro-Arg-Gly-NH₂ was modified at position six with α-L-aminobutyric acid (Abu) to give peptide 2. Further modifications of the Abu⁶ analogue (No. 2) at position one by substituting cyclohexylacetic acid (Caa), cyclohexylpropionic acid (Cpa), 1-adamantaneacetic acid (Aaa), phenylacetic acid (Phaa), tert.-butylacetic acid (t-Baa), isovaleric acid (Iva), propionic acid (Pa), l -penicillamine (P), tert.-butoxycarbonyl (Boc) or omitting any substituent at this position, and/or in combination with Arg-NH₂⁹, Ala-NH₂, ⁹d -Arg⁸-Arg-NH₂⁹, and desGly⁹ modifications yielded the remaining 14 peptides. All 16 peptides were examined for agonistic and antagonistic potencies in AVP V₂ and V₁ assays in rats. Apart from the Cpa¹ analogue and the analogue lacking any substituent in the 1-position, all exhibit substantial V₂ and V₁ antagonism. A number are as potent as (A) as V₂ antagonists. With an anti-V₂ pA₂= 8.11 ± 0.07, Aaa-d -Tyr(Et)-Phe-Val-Asn-Abu-Pro-Arg-Arg-NH₂ (No. 6) is as potent as any cyclic AVP V₂ antagonist reported to date. The Pa¹ analogue of No. 6 exhibits promising anti-V₂/anti-V₁ selectivity. These findings prove conclusively that a ring structure is not a requirement for recognition of or for binding to AVP V₂ or V₁ receptors. This discovery thus offers a promising new approach to the design of peptide and non-peptide antagonists of AVP and perhaps also to other cyclic peptides such as somatostatin, atrial-natriuretic factor, insulin, and the recently discovered endothelin. Some of these linear antagonists may be of value as pharmacological tools and as therapeutic agents.     

Conformational properties of a cyclic peptide bradykinin B2 receptor antagonist using experimental and theoretical methods

Abstract: The solution conformation of the cyclic peptide J324 (cyclo^0,6‐[Lys⁰,Glu⁶,d ‐Phe⁷]BK), an antagonist targeted at the bradykinin (BK) B₂ receptor, has been investigated using experimental and theoretical methods. In order to gain insight into the structural requirements essential for BK antagonism, we carried out molecular dynamics (MD) simulations using simulated annealing as the sampling protocol. Following a free MD simulation we performed simulations using nuclear Overhauser enhancement (NOE) distance constraints determined by NMR experiments. The low‐energy structures obtained were compared with each other, grouped into families and analyzed with respect to the presence of secondary structural elements in their backbone. We also introduced new ways of plotting structural data for a more comprehensive analysis of large conformational sets. Finally, the relationship between characteristic backbone conformations and the spatial arrangement of specific pharmacophore centers was investigated.     

Conformational preferences of proline derivatives incorporated into vasopressin analogues: NMR and molecular modelling studies

In this study, arginine vasopressin analogues modified with proline derivatives - indoline-2-carboxylic acid (Ica), (2S,4R)-4-(naphthalene-2-ylmethyl)pyrrolidine-2-carboxylic acid (Nmp), (2S,4S)-4-aminopyroglutamic acid (APy) and (2R,4S)-4-aminopyroglutamic acid, (Apy) - were examined using NMR spectroscopy and molecular modelling methods. The results have shown that Ica is involved in the formation of the cis peptide bond. Moreover, it reduces to a great extent the conformational flexibility of the peptide. In turn, incorporation of (2S,4R)-Nmp stabilizes the backbone conformation, which is heavily influenced by the pyrrolidine ring. However, the aromatic part of the Nmp side chain exhibits a high degree of conformational freedom. With analogues IV and V, introduction of the 4-aminopyroglumatic acid reduces locally conformational space of the peptides, but it also results in weaker interactions with the dodecylphosphocholine/sodium dodecyl sulphate micelle. Admittedly, both analogues are adsorbed on the micelle's surface but they do not penetrate into its core. With analogue V, the interactions between the peptide and the micelle seem to be so weak that conformational equilibrium is established between different bound states.     

Synthesis of [2H, 13C] and [14C] labeled vasopressin V1a/V2 receptor antagonist RWJ‐676070 and its stable labeled N‐des‐benzoyl metabolite

Stable isotope‐labeled ([¹³C₂, D₃]) and carbon 14‐labeled vasopressin receptor antagonist RWJ‐676070, a spirobenzazepine amide, were prepared in separate syntheses for use in drug metabolism studies. The stable isotopically labeled sample was prepared starting from [¹³C₂, D₆]dimethyl sulfate and [¹³C]copper (I) cyanide in nine steps with a 14% overall isotopic yield. The ¹⁴C label was introduced in five steps starting from [¹⁴C]potassium cyanide to provide material having a specific activity of 58 mCi/mmol (2.15 GBq/mmol). Selective hydrolysis of the metabolically labile amide bond in [¹³C₂, D₃]RWJ‐676070 gave the corresponding labeled metabolite in one step. Copyright © 2008 John Wiley & Sons, Ltd.     

NMR and molecular modeling characterization of RGD containing peptides

The tripeptide sequence arginine-glycine-aspartic acid (RGD) has been shown to be the key recognition segment in numerous cell adhesion proteins. The solution conformation and dynamics in DMSO-d₆ of the cyclic pentapeptides, Ac-Cys-Arg-Gly-Asp-Cys-OH (CRGDC), a potent fibrinogen receptor antagonist, and Ac-Cys-Arg-Gly-D-Asp-Cys-OH (CRGdC), a weak fibrinogen receptor antagonist, have been characterized by nuclear magnetic resonance (NMR) spectroscopy and molecular modeling. ¹H-¹H distance constraints derived from two-dimensional NOE spectroscopy and torsional angle constraints obtained from ³JNH-h α coupling constants, combined with computer-assisted modeling using conformational searching algorithms and energy minimization have allowed several low energy conformations of the peptides to be determined. Low temperature studies in combination with molecular dynamics simulations suggest that each peptide does not exist in a single, well-defined conformation, but as an equilibrating mixture of conformers in fast exchange on the NMR timescale. The experimental results can be fit by considering pairs of low energy conformers. Despite this inherent flexibility, distinct conformational preferences were found which may be related to the biological activity of the peptides.