Expedited discovery of second generation NK-1 antagonists: identification of a nonbasic aryloxy substituent.

Solution-phase, parallel-synthesis techniques were used to optimize a series of nonbasic NK-1 antagonists, resulting in the identification of (R)-26, an orally bioavailable compound with subnanomolar potency.

Substance P regulates PTH secretion through the neurokinin-1 receptor.

The primary regulator of PTH secretion is serum ionized Ca(2+); however, neuropeptide-containing nerve fibers have been localized to the parathyroid gland. The purpose of this study was to determine whether or not substance P (SP) regulates PTH secretion. In dispersed porcine parathyroid cells, SP reversibly inhibited 0.5 mM CaCl(2)-induced PTH secretion (IC(50) = 0.29 nM) and had no effect at CaCl(2) concentrations of 1.5 mM and greater. At 0.5 mM CaCl(2), treatment with a NK-1 selective receptor agonist resulted in a concentration-dependent decrease in PTH secretion (IC(50) = 0.21 nM). In contrast, NK-2 and NK-3 receptor agonists were approximately 100-fold less active than SP or the NK-1 receptor selective agonist. An enantiospecific reversal of the effects of SP on PTH secretion was observed with LY306740, a potent selective NK-1 receptor antagonist (K(i) = 0.125 nM). In porcine parathyroid cells, expression of mRNA for the NK-1 receptor was observed using RT-PCR. In summary, a novel neuroendocrine pathway is described whereby the neuropeptide, SP, regulates PTH secretion through NK-1 receptors.

Structure-activity relationship of a series of diaminoalkyl substituted benzimidazole as neuropeptide Y Y1 receptor antagonists.

A series of benzimidazoles (4) was synthesized and evaluated in vitro as potent and selective NPY Y1 receptor antagonists. Substitution of the piperidine nitrogen of 4 with appropriate R groups resulted in compounds with more than 80-fold higher affinity at the Y receptor compared to the parent compound 5 (R = H). The most potent benzimidazole in this series was 21 (Ki = 0.052 nM).

Structure-activity relationships of a series of benzothiophene-derived NPY Y1 antagonists: optimization of the C-2 side chain.

A series of benzo[b]thiophene-derived NPY-1 receptor antagonists is described. Systematic modification of the C-2 substituent afforded a 1000-fold range in Y1 receptor affinity. Appropriate substitution at the ortho and para positions of the C-2 phenyl ether produced a synergistic effect on Y1 binding affinity, which led to the discovery of the most active ligands, 12t (K(i) = 15 nM), 12u (K(i) = 11 nM), and 12v (K(i) = 13 nM).

Synthesis and evaluation of a series of novel 2-[(4-chlorophenoxy)methyl]benzimidazoles as selective neuropeptide Y Y1 receptor antagonists.

A series of novel benzimidazoles (BI) derived from the indole 2 was synthesized and evaluated as selective neuropeptide Y (NPY) Y1 receptor antagonists with the aim of developing antiobesity drugs. In our SAR approach, the (4-chlorophenoxy)methyl group at C-2 was kept constant and a series of BIs substituted with various piperidinylalkyl groups at N-1 was synthesized to identify the optimal spacing and orientation of the piperidine ring nitrogen relative to the benzimidazole. The 3-(3-piperidinyl)propyl in 33 was found to maximize affinity for the Y1 receptor. Because of the critical importance of Arg33 and Arg35 of NPY binding to the Y1 receptor, the incorporation of an additional aminoalkyl functionality to the structure of 33 was explored. Methyl substitution was used to probe where substitution on the aromatic ring was best tolerated. In this fashion, the C-4 was chosen for the substitution of the second aminoalkyl functionality. Synthesis of such compounds with a phenoxy tether using the 4-hydroxybenzimidazole 11 was pursued because of their relative ease of synthesis. Functionalization of the hydroxy group of 45 with a series of piperidinylalkyl groups provided the dibasic benzimidazoles 55-62. Among them, BI 56 demonstrated a Ki of 0.0017 microM, which was 400-fold more potent than 33. To evaluate if there was a stereoselective effect on affinity for these BIs, the four constituent stereoisomers (69-72) of the BI 60 were prepared using the S- and R-isomers of bromide 17. Antagonist activity of these BIs was confirmed by measuring the ability of selected compounds to reverse NPY-induced forskolin-stimulated cyclic AMP. The high selectivity of several BI antagonists for the Y1 versus Y2, Y4, and Y5 receptors was also shown.

Structure-activity relationships of a series of 1-substituted-4-methylbenzimidazole neuropeptide Y-1 receptor antagonists.

The characterization of a novel series of NPY-1 receptor antagonists derived from the 4-methylbenzimidazole 4 is described. Appropriate substitution on the piperidyl nitrogen of 4 led to systematic increases in Y-1 receptor affinity, to approximately 50-fold, and to the discovery of the importance of a second basic substituent.

LY303870, a centrally active neurokinin-1 antagonist with a long duration of action.

The selective neurokinin (NK)-1 antagonist LY303870 has high affinity and specificity for human and guinea pig brain NK-1 receptors labeled with 125I-substance P. It has approximately 15- to 30-fold lower affinity for rat and mouse brain NK-1 receptors, consistent with previously reported species differences in the affinities of nonpeptide antagonists for NK-1 receptors. In vivo, LY303870 blocked the characteristic, caudally directed, biting and scratching response elicited by intrathecal administration of the selective NK-1 agonist Ac-[Arg6,Sar9,Met(O2)11]substance P6-11 in conscious mice. The potentiation of the tail-flick response elicited by intrathecal administration of the NK-1 agonist [Sar9,Met(O2)11]substance P in rats was also selectively blocked by LY303870. When tested in a model of persistent nociceptive activation induced by tissue injury (the formalin test), LY303870 blocked licking behavior in the late phase of the formalin test, in a dose-dependent manner. After oral administration of 10 mg/kg, the blockade of the late-phase licking behavior was evident for at least 24 hr. Ex vivo binding studies in guinea pigs showed that orally administered LY303870 potently inhibited binding to central and peripheral NK-1 receptors labeled with 125I-substance P. This inhibition was long-lasting, consistent with other in vivo activities. LY306155, the opposite enantiomer of LY303870, was less active in all of the functional assays. In rodents, LY303870 did not exhibit any neurological, motor, cardiovascular, gastrointestinal or autonomic side effects at doses of < or = 50 mg/kg p.o. Thus, LY303870 is a potent, centrally active, NK-1 antagonist in vivo, with long-lasting oral activity.

The non-peptide NK-1 receptor antagonist LY303870 inhibits neurogenic dural inflammation in guinea pigs.

LY303870 is a competitive, high affinity NK-1 receptor antagonist. It was tested in the trigeminal stimulation-induced neurogenic dural inflammation model of migraine. The neurogenic inflammation theory of migraine pain proposes that substance P, acting through NK-1 receptors, causes dural inflammation which enhances migraine pain. LY303870 administration potently inhibited neurogenic dural inflammation as measured by plasma protein extravasation caused by electrical stimulation of the trigeminal ganglion in guinea pigs. It was active in this model when administered via intravenous, oral or inhalation routes. LY306155, the enantiomer of LY303870 with lower affinity for the NK-1 receptor, was much less potent than LY303870 in this model. LY303870, at oral doses of 1, 10 and 100 microg/kg, produced a long, dose-dependent inhibition of dural inflammation, demonstrating a suitable duration of action for a potential use in acute migraine and migraine prophylaxis.

Neuropeptide Y receptor antagonists in obesity.

Neuropeptide Y (NPY) is a 36 amino acid amidated peptide with high sequence homology to the endocrine peptides, peptide YY (PYY) and pancreatic polypeptide (PP). They appear to interact with a family of receptors that possess high affinity for one or more of these peptides. Five members of the receptor family have been cloned, with several additional members postulated through pharmacological evidence. All are members of the seven transmembrane domain-G-protein coupled receptor family. The Y1 receptor is the best characterised, with several nonpeptide antagonists available. This receptor appears to mediate a constriction of the peripheral vasculature and the 'anxiolytic' effects of centrally administered NPY. Less is known about the other receptors in the family. The Y2 receptor is believed to be presynaptic and mediates a reduction in neurotransmitter release. The Y4 receptor appears to be the receptor for pancreatic polypeptide, with high amounts of mRNA for this receptor found in the periphery, but lower levels in the brain. The Y5 receptor is expressed in the hypothalamus and has been postulated to be the receptor which mediates the increased food consumption seen following centrally administered NPY. Finally, the Y6 receptor has been cloned in the mouse and other species, but does not appear to encode a functional gene product in humans. Several types of nonpeptide Y1 and a series of Y5 antagonists have been described in the patent literature, though these compounds have limitations that will confine their use to preclinical studies. Nevertheless, considerable progress has been made in understanding the role of NPY and its receptors in experimental obesity. The next step will be the discovery of potent and selective nonpeptide antagonists, to add further credence to the therapeutic potential.

Peripheral effects of three novel non-peptide tachykinin NK1 receptor antagonists in the anaesthetized rat.

Three novel non-peptide tachykinin NK1 receptor antagonists were assessed on the transient fall in mean arterial blood pressure and the salivation induced by i.v. substance P (0.65 nmol/kg) in the urethane-anaesthetized rat. LY303241 ((R)-1-[N-(2-methoxybenzyl)acetylamino]-3-(1H-indol-3-yl)-2-[N-(2-(4- phenylpiperazin-1-yl)acetyl)amino]propane), LY303870 ((R)-1-[N-(2-methoxybenzyl)acetylamino]-3-(1H-indol-3-yl)-2-[N-(2-(4-(++ +piperidin-1 -yl)piperidin-1-yl)acetyl)amino]propane) and LY306740 ((R)-1-[N-(2-methoxybenzyl)acetylamino]-3-(1H-indol-3-yl)-2-[N-(2-(4 -cyclohexylpiperazin-1-yl)acetyl)amino]propane) (65 nmol-9 micromol/kg i.v.; 5 min earlier) inhibited both the vasodepressor and salivary responses to substance P in a dose-dependent manner. LY303241 and LY306740 were more potent in inhibiting the vascular response to substance P while LY303870 was more potent in inhibiting the salivary response. LY303870 and LY306740 were devoid of direct effects while LY303241 decreased blood pressure and heart rate for 1 and 10 min, respectively. The antagonists act in a stereoselective and specific manner since the opposite (S) enantiomers of LY303870 (LY306155) and LY306740 (LY307679) failed to block the effects of substance P. In addition, LY303241, LY303870 and LY306740 neither affected the hypotension and the salivation induced by carbachol nor the increases in mean arterial pressure and heart rate induced by the tachykinin NK2 receptor agonist [beta-Ala8]neurokinin A-(4-10). Only LY303241 attenuated the decreases in mean arterial pressure and heart rate evoked by the tachykinin NK3 receptor agonist senktide. LY303870 and LY306740 appear to be the most interesting antagonists since they act in a specific and selective manner at the tachykinin NK1 receptor. The difference in the order of potency of the three antagonists to inhibit the hypotension and salivation elicited by substance P could be ascribed to their pharmacodynamic features or to the existence of different signal transduction mechanisms or receptor subtypes.

[3H]LY303870, a novel nonpeptide radioligand for the NK-1 receptor.

We synthesized a potent and selective antagonist radioligand for the neurokinin (NK)-1 receptor and characterized its binding to guinea pig striatal membranes. (R)-N-[2-[Acetyl[3H3][(2-methoxyphenyl)-methyl]amino]- 1-(1H-indol-3-ylmethyl) ethyl][1,4'-bipiperidine]- 1'-acetamide ([3H]LY303870) binds to a single class of sites with an equilibrium KD of 0.22 nM and a Bmax of 723 fmol/mg of protein. Unlabeled LY303870 potently inhibited the binding with an IC50 of 0.56 nM, whereas the less active (S)-enantiomer (LY306155) was substantially less potent. The nonpeptide NK-1 antagonists (+/-)-CP96,345 and (+/-)-RP 67580 had IC50 values of 0.74 and 49 nM, respectively. Substance P (SP) was also a potent inhibitor with with an IC50 of 3.1 nM. The inhibition by SP could be separated into two components: a high-affinity component with a Ki of 0.53 nM and a lower-affinity component with a Ki of 155 nM. Addition of 100 microM guanylyl 5'-imidodiphosphate [Gpp(NH)p] in the incubation increased the relative amount of the low-affinity agonist state of the receptor. Consistent with the antagonist properties of LY303870, the dissociation rate of [3H]-LY303870 was not changed by the presence of 100 microM Gpp(NH)p. The distribution of [3H]LY303870 binding sites in the guinea pig brain closely matched the distribution of NK-1 receptors labeled by [3H]SP. Therefore, [3H]LY303870 is a potent and selective antagonist radioligand for NK-1 receptors in guinea pig brain. In addition, regulation of NK-1 agonist affinity by guanine nucleotides is similar to that seen for monoaminergic receptors.

3-Aryl-1,2-diacetamidopropane derivatives as novel and potent NK-1 receptor antagonists.

Early structure-activity studies on racemic tryptophan ester and amide NK-1 antagonists 5-7 led to the discovery that the potency of the series could be markedly increased by moving the carbonyl function in these molecules to an off-chain position as in the 3-aryl-1,2-diacetamidopropane 9. Further medicinal chemistry incorporating this change resulted in the discovery of a novel series of highly potent aryl amino acid derived NK-1 antagonists of the R stereoisomeric series (IC50's = 100 pM to > 5 microM). Compounds in this series were shown to be competitive antagonists using an in vitro NK-1 smooth muscle assay, and this data correlated well with observed human NK-1 binding affinities. Two of these agents, (R)-25 and (R)-32, blocked intrathecal NK-1 agonist-driven [Ac-[Arg6,Sar9,Met(O2)11]- substance P 6-11 (Ac-Sar9)] nociceptive behavior in mice. Both compounds potently blocked the neurogenic dural inflammation following trigeminal ganglion stimulation in the guinea pig after intravenous administration. Further, upon oral administration in this model, (R)-32 was observed to be very potent (ID50 = 91 ng/kg) and have a long duration of action (> 8 h at 1 micrograms/kg). Compound (R)-32, designated LY303870, is currently under clinical development as an NK-1 antagonist with a long duration of action.