Joseph Frey

Behavioural effects of receptor-specific substance P agonists

&NA; Septide and senktide are synthetic substance P (SP) agonists with extremely high selectivity for 1 of the 3 known SP receptor subtypes. When injected intrathecally, they produced dramatically different behavioural effects. Septide, the selective SP‐P receptor agonist, evoked intense, compulsive scratching, biting and licking of the hind limb, with no sign of motor flaccidity, and without measurable effect on responses to noxious thermal or mechanical stimulation of the foot or tail. In contrast, senktide, the selective SP‐N receptor agonist, produced profound, but transient, motor flaccidity, reduced response to noxious stimuli and, at low doses, ‘wet‐dog shakes.’ These various symptoms, all previously associated with SP and/or synthetic SP analogues, appear therefore to derive from activation of distinct SP receptor subtypes.

The effective ‘size’ of the tris(trimethylsilyl)silyl group in several molecular environments

The effective size of the tris(trimethylsilyl)silyl group in several molecular environments has been estimated. 2,2-Dimesityl-1-tris(trimethylsilyl)silylethanol 1g has been prepared and its structure determined by X-ray crystallography. The Mes–CC torsional angles are 59.6 (φ2) and 63.3°(φ2) and the CC–Si bond angle α4 is 133.8°. The two-ring flip barrier for the correlated rotation of the two mesityl rings around the Mes–C bonds is ΔGc‡= 10.2 kcal mol–1. The structures of enols Mes2CC(OH)R, R = H, Me, Et, Pri, But(1a–1e), Me3Si (1f), (Me3Si)3Si (1g) and (Me3Si)3C (1h) and the two-ring flip barriers have been calculated by the MM2* force-field. The calculated and the experimental values are in good agreement, except for somewhat lower calculated α4 for 1b–1e and a shorter C–Si distance in 1g. From the linear correlations between the observed cos φ2 or ΔGc‡ values and Es values for the enols 1a–1e, and the values observed for 1g an average Es value of –1.46 has been calculated for (Me3Si)3Si. MM2* calculations gave an A value for (Me3Si)3Si of 4.89 kcal mol–1. These steric parameters resemble those for the But group (Es=–1.54; A= 4.9 kcal mol–1) indicating a similar effective size for the But and (Me3Si)3Si groups in these specific environments. (Me3Si)3C is significantly larger (A= 13.3 kcal mol–1; estimated Es=–3.7).

Synthesis and biological activity of peptide hydroxamate inhibitors of degradation of substance P analogues

A series of hydroxamic acid derivatives of peptides related to fragments of substance P (SP) were synthesized. Methyl, ethyl or N-hydroxy-succinimide ester precursors of the desired peptides were prepared by using classical peptide synthesis methodology and these were reacted with excess hydroxylamine in either ethanol or N,N-dimethylformamide. The products were characterized by chromatographic methods, amino acid analysis and fast atom bombardment mass spectrometry. The inhibition of the degradation of the radiolabelled substrate desamino-[3-125I-tyrosyl5]SP(5–11) ([(125I)BH5]SP(5–11)) by these compounds in rat hypothalamus preparations was determined. The most potent inhibitors found were Boc-Phe-Phe-Phe-NHOH (12d, IC50 = 4 μM), Boc-Phe-Phe-Trp-NHOH (9, IC50 = 5 μM) and desamino-Tyr-Phe-Phe-Gly-NHOH (22, IC50 = 1.8 μM). A model describing the interaction of these compounds with the active site is proposed.

17O and13C NMR spectra of stable simple enols

17 O NMR spectra of twelve stable simple enois 1[Ar2CC(R)OH, Ar = crowded aryl group, R = H (1c, Ar = Mes), But, m- and p-substituted aryl, mesityl] were measured in CDCl3 and of six of them in [2H6] DMSO. 13C spectra of all the enois were measured in both solvents. These are the first values measured for enois not strongly intramolecularly hydrogen bonded. The δ(17O) values in CDCl3 are divided into three groups which parallel the steric bulk of R: (i) 70.5–74.9 when R = H; (ii) 84.1–87.4 when R =m- and p-substituted aryl; (iii) 93.0–98.5 ppm for R = But, mesityl (Mes). ΔδO =δ(17O)[[2H6]DMSO]–δ(17O)[CDCl3]= 7–10 for group (i), ca. 3 for group (ii) and –17 for trimesitylethenol 1l. IGLO calculations on CH2CHOH (4) give a shallow parabola for the δ(17O)vs. the CC–O–H torsional angle θ plot, with maxima at θ= 0 and 180°(syn and anti planar conformations) and a minimum at θ= 90°. δ(13C-2) values show an approximate mirror-image change while δ(13C-1) changes slightly with θ. Calculation on the solvates 4·OSH2 with syn, anti and perpendicular conformations show trends similar to those for uncomplexed 4 with upfield shifts of δ(17O, 13C-2) and downfield shift of δ(13C-1, 1H) in all conformations. When the calculations also included the effect of the reaction field, the δ(17O) values of the perpendicular and anti conformations differ little from those of uncomplexed 4. PM3 calculations for four enois including 1c and 1l also show a minimum at θ= 90° and the plot of ΔfH for 1c and 1lvs. θ resemble the calculated energy (by MP2/6-311 G**)vs. θ plot for 4. No correlation exists between the Ar–CC torsion angles or λmax of the enols with δ(17O), but the polar effect of the α-substituent affects the δ(17O) values. The solvent effect is ascribed to OH–[2H6]DMSO hydrogen bonding, accompanied by a conformational change from syn-periplanar in CDCl3 to mainly anti-clinal in [2H6]DMSO. A lower association with DMSO for bulkier Rs and an accompanying change of θ towards 90° qualitatively account for the ΔδO values. Several correlations between the δ(17O), δ(1H) and δ(13C) shifts or of δ(13C) and Hammetts σ values were found.

Crystal structure of the solid state photoreactive 2,2′,4,4′,6,6′-hexaisopropylbenzil

2,2′,4,4′,6,6′-Hexaisopropylbenzil undergoes an intramolecular solid state photochemical cyclization initiated by hydrogen abstraction by a carbonyl oxygen. The crystal structure of the diketone has been determined by X-ray diffraction methods and the molecular geometry was found to possess an inversion centre. There are three hydrogen atoms (and another three are related by the inversion centre) in close contact (2.68, 2.86 and 2.99 A) with an oxygen atom, which suggests the feasibility of both γ- and δ-hydrogen abstraction. The molecular conformation and geometry at the central diketone moiety are compared with those of other diaryl diketones.

2,2',4,4',6-PENTA-TERT-BUTYLBENZIL : AN UNEXPECTED PRODUCT IN THE ATTEMPTED COUPLING OF TRI-TERT-BUTYLBENZOYL CHLORIDE WITH MAGNESIUM-MAGNESIUM IODIDE

Reduction of 2,4,6-tri-tert-butylbenzoyl chloride with Mg–MgI2 in 1∶1 diethyl ether–benzene under ultrasonic irradiation gives the hexa-tert-butyl-benzil and -benzoin and also 2,2′,4,4′,6-penta-tert-butylbenzil. Possible mechanisms for the loss of the But group are discussed.

Receptor-specific substance-p agonists: Behavioral effects upon intrathecal injection

Intrathecal (IT) vasopressin (VAS) produces prolonged analgesia on the tail flick and formalin tests and a transient analgesia on the hot plate and tail shock vocalization tests. This study further characterizes the tail flick suppression as to 1) whether analgesia depends only on spinal circuitry, 2) whether ITVAS acts via pressor-type VAS receptors, and 3) whether a metabolite (VAS-4-g) has similar IT effects. Ninety-two male Sprague-Dawley rats were implanted with lumbosacral IT catheters. Thirty-six of these rats had their spinal cords transected at the second thoracic vertebra and 21 had sham spinalizations. These 57 rats were injected IT with either 0, 0.25, 2.5, or 25 ng VAS. The remaining 35 rats were injected IT with either (a) VAS (25 ng), (b) VAS (25 ng) and a VAS antagonist, dET2AVP (25 or 50 ng), or (c) VAS-4-g (2.5, 25, or 50 ng). Tail flick latencies were measured prior to drug injection, and then every 5 min for 40 min after injection. IT VAS significantly suppressed the tail flick reflex for the 40 min test period in both spinalized and intact rats. However, analgesia was delayed in spinalized rats. In intact rats, dET2AVP produced a dosedependent block of analgesia. The antagonist is currently being tested alone. VAS-4-g had no effect on tail flick latency. These data indicate that: (a) IT VAS exerts its effects at least partially at the spinal level, but the delay in onset of analgesia in spinalized rats suggests that there may also be a supraspinal component; (b) the effects of VAS may be via Vl-like receptors since the pressorspecific antagonist appears to block the tail flick suppression; and Cc) the fact that VAS-4-g has no effect on TF latency suggests that VAS, rather than a metabolite, produces analgesia. Grant WNS20037

Synthesis and Biological Activity of Agonists for the Neuronal Tachykinin Receptor in Guinea Pig Ileum

The classification of tachykinin receptors into two subclasses, the SP-P and the SP-E receptors has been well established (1). Recently Laufer et. al. (2) characterized, in the guinea pig ileum, a third tachykinin receptor subclass, designated as SP-N receptor. The SP-N receptor is located on the enteric cholinergic neurons, and madiates the release of acetylcholine. We found that the SP analog (pGlu6,(N-Me)Phe8) SP6-11 acts as a selective potent agonist for the SP-N receptor (EC50=0.5 nM) while its potency for the SP-P receptor is much lower (EC50=500 nM). Conceivably N-methylation of Phe8 in the hexapeptide sequence of substance P (SP), induces selectivity toward the SP-N receptor. Therefore we set to probe the constraints at the nitrogen of Phe7-Phe8 amide bond by changing the alkyl group at this site.