Novel sst2-selective somatostatin agonists. Three-dimensional consensus structure by NMR

The 3D NMR structures of six octapeptide agonist analogues of somatostatin (SRIF) in the free form are described. These analogues, with the basic sequence H-DPhe/Phe2-c[Cys3-Xxx7-DTrp8-Lys9-Thr10-Cys14]-Thr-NH2 (the numbering refers to the position in native SRIF), with Xxx7 being Ala/Aph, exhibit potent and highly selective binding to human SRIF type 2 (sst2) receptors. The backbone of these sst2-selective analogues have the usual type-II' beta-turn reported in the literature for sst2/3/5-subtype-selective analogues. Correlating the biological results and NMR studies led to the identification of the side chains of DPhe2, DTrp8, and Lys9 as the necessary components of the sst2 pharmacophore. This is the first study to show that the aromatic ring at position 7 (Phe7) is not critical for sst2 binding and that it plays an important role in sst3 and sst5 binding. This pharmacophore is, therefore, different from that proposed by others for sst2/3/5 analogues.

Conformational analysis of a potent SSTR3-selective somatostatin analogue by NMR in water solution

The three-dimensional structure of a potent SSTR3-selective analogue of somatostatin, cyclo(3-14)H-Cys(3)-Phe(6)-Tyr(7)-D-Agl(8)(N(beta) Me, 2-naphthoyl)-Lys(9)-Thr(10)-Phe(11)-Cys(14)-OH (des-AA(1, 2, 4, 5, 12, 13)[Tyr(7), D-Agl(8)(N(beta) Me, 2-naphthoyl)]-SRIF) (peptide 1) has been determined by (1)H NMR in water and molecular dynamics (MD) simulations. The peptide exists in two conformational isomers differing mainly by the cis/trans isomerization of the side chain in residue 8. The structure of 1 is compared with the consensus structural motifs of other somatostatin analogues that bind predominantly to SSTR1, SSTR2/SSTR5 and SSTR4 receptors, and to the 3D structure of a non-selective SRIF analogue, cyclo(3-14)H-Cys(3)-Phe(6)-Tyr(7)-D-2Nal(8)-Lys(9)-Thr(10)-Phe(11)-Cys(14)-OH (des-AA(1, 2, 4, 5, 12, 13)[Tyr(7), D-2Nal(8)]-SRIF) (peptide 2). The structural determinant factors that could explain selectivity of peptide 1 for SSTR3 receptors are discussed.

Three-dimensional consensus structure of sst2-selective somatostatin (SRIF) antagonists by NMR

The three-dimensional NMR structures of seven octapeptide analogs of somatostatin (SRIF), based on octreotide, with the basic sequence H-Cpa/Phe2-c[DCys3-Xxx7-DTrp/DAph(Cbm)8-Lys9-Thr10-Cys14]-Yyy-NH2 (the numbering refers to the position in native SRIF), with Xxx7 being Aph(Cbm)/Tyr/Agl(NMe,benzoyl) and Yyy being Nal/DTyr/Thr, are presented here. Most of these analogs exhibit potent and highly selective binding to sst2 receptors, and all of the analogs are antagonists inhibiting receptor signaling. Based on their consensus 3D structure, the pharmacophore of the sst2-selective antagonist has been defined. The pharmacophore involves the side chains of Cpa2, DTrp/DAph(Cbm)8, and Lys9, with the backbone for most of the sst2-selective antagonists comprised a Type-II' beta-turn. Hence, the sst2-selective antagonist pharmacophore is very similar to the sst2-selective agonist pharmacophore previously described.

Titration and implementation of neurally adjusted ventilatory assist in critically ill patients

BACKGROUND: Neurally adjusted ventilatory assist (NAVA) delivers assist in proportion to the patient's respiratory drive as reflected by the diaphragm electrical activity (EAdi). We examined to what extent NAVA can unload inspiratory muscles, and whether unloading is sustainable when implementing a NAVA level identified as adequate (NAVAal) during a titration procedure. METHODS: Fifteen adult, critically ill patients with a Pao(2)/fraction of inspired oxygen (Fio(2)) ratio < 300 mm Hg were studied. NAVAal was identified based on the change from a steep increase to a less steep increase in airway pressure (Paw) and tidal volume (Vt) in response to systematically increasing the NAVA level from low (NAVAlow) to high (NAVAhigh). NAVAal was implemented for 3 h. RESULTS: At NAVAal, the median esophageal pressure time product (PTPes) and EAdi values were reduced by 47% of NAVAlow (quartiles, 16 to 69% of NAVAlow) and 18% of NAVAlow (quartiles, 15 to 26% of NAVAlow), respectively. At NAVAhigh, PTPes and EAdi values were reduced by 74% of NAVAlow (quartiles, 56 to 86% of NAVAlow) and 36% of NAVAlow (quartiles, 21 to 51% of NAVAlow; p < or = 0.005 for all). Parameters during 3 h on NAVAal were not different from parameters during titration at NAVAal, and were as follows: Vt, 5.9 mL/kg predicted body weight (PBW) [quartiles, 5.4 to 7.2 mL/kg PBW]; respiratory rate (RR), 29 breaths/min (quartiles, 22 to 33 breaths/min); mean inspiratory Paw, 16 cm H(2)O (quartiles, 13 to 20 cm H(2)O); PTPes, 45% of NAVAlow (quartiles, 28 to 57% of NAVAlow); and EAdi, 76% of NAVAlow (quartiles, 63 to 89% of NAVAlow). Pao(2)/Fio(2) ratio, Paco(2), and cardiac performance during NAVAal were unchanged, while Paw and Vt were lower, and RR was higher when compared to conventional ventilation before implementing NAVAal. CONCLUSIONS: Systematically increasing the NAVA level reduces respiratory drive, unloads respiratory muscles, and offers a method to determine an assist level that results in sustained unloading, low Vt, and stable cardiopulmonary function when implemented for 3 h.

Improved impurity fingerprinting of heparin by high resolution (1)H NMR spectroscopy

For improving the identification of potential heparin impurities such as oversulfated chondroitin sulfate (OSCS) the standard 2D (1)H-(1)H NMR NOESY was applied. Taking advantage of spin diffusion and adjusting the experimental parameters accordingly additional contaminant-specific signals of the corresponding sugar ring protons can easily be detected. These are usually hidden by the more intense heparin signals. Compared to the current 1D (1)H procedure proposed for screening commercial unfractionated heparin samples and focusing on the contaminants acetyl signals more informative and unique fingerprints may be obtained. Correspondingly measured (1)H fingerprints of a few potential impurities are given and their identification in two contaminated commercial heparin samples is demonstrated. The proposed 2D NOESY method is not intended to replace the current 1D method for detecting and quantifying heparin impurities but may be regarded as a valuable supplement for an improved and more reliable identification of these contaminants.

[Optimised glycaemic control and dose titration with insulin glargine (Lantus) by general practitioners in Switzerland: results of the OPTI-LAN Postmarketing Study]

National and international societies have published guidelines regarding glycaemic control in type-2 diabetes mellitus. Clinical studies have shown that glycaemic control of type-2 diabetes mellitus can be improved using simple algorithms for titration of insulin Glargine (Lantus). It is unclear, to what degree published guidelines are adopted in daily practice in Switzerland.

Use of a computerized closed-loop sodium nitroprusside titration system for antihypertensive treatment after open heart surgery

This study evaluates the clinical applicability of administering sodium nitroprusside by a closed-loop titration system compared with a manually adjusted system. The mean arterial pressure (MAP) was registered every 10 and 30 sec during the first 150 min after open heart surgery in 20 patients (group 1: computer regulation) and in ten patients (group 2: manual regulation). The results (16,343 and 2,912 data points in groups 1 and 2, respectively), were then analyzed in four time frames and five pressure ranges to indicate clinical efficacy. Sixty percent of the measured MAP in both groups was within the desired +/- 10% during the first 10 min. Thereafter until the end of observation, the MAP was maintained within +/- 10% of the desired set-point 90% of the time in group 1 vs. 60% of the time in group 2. One percent and 11% of data points were +/- 20% from the set-point in groups 1 and 2, respectively (p less than .05, chi-square test). The computer-assisted therapy provided better control of MAP, was safe to use, and helped to reduce nursing demands.

Time-dependent interactions of the two porphyrinic compounds chlorin e6 and mono-L-aspartyl-chlorin e6 with phospholipid vesicles probed by NMR spectroscopy

The distribution processes of chlorin e6 (CE) and monoaspartyl-chlorin e6 (MACE) between the outer and inner phospholipid monolayers of 1,2-dioleoyl-phosphatidylcholine (DOPC) vesicles were monitored by 1H NMR spectroscopy through analysis of chemical shifts and line widths of the DOPC vesicle resonances. Chlorin adsorption to the outer vesicle monolayer induced changes in the DOPC 1H NMR spectrum. Most pronounced was a split of the N-methyl choline resonance, allowing for separate analysis of inner and outer vesicle layers. Transbilayer distribution of the chlorin compounds was indicated by time-dependent characteristic spectral changes of the DOPC resonances. Kinetic parameters for the flip-flop processes, that is, half-lives and rate constants, were obtained from the experimental data points. In comparison to CE, MACE transbilayer movement was significantly reduced, with MACE remaining more or less attached to the outer membrane layer. The distribution coefficients for CE and MACE between the vesicular and aqueous phase were determined. Both CE and MACE exhibited a high affinity for the vesicular phase. For CE, a positive correlation was found between transfer rate and increasing molar ratio CE/DOPC. Enhanced membrane rigidity induced by increasing amounts of cholesterol into the model membrane was accompanied by a decrease of CE flip-flop rates across the membrane. The present study shows that the movement of porphyrins across membranes can efficiently be investigated by 1H NMR spectroscopy and that small changes in porphyrin structure can have large effects on membrane kinetics.

High-field 1H T1 and T2 NMR Relaxation Time Measurements of H2O in Homeopathic Preparations of Quartz, Sulfur and Copper Sulfate

Quantitative meta-analyses of randomized clinical trials investigating the specific therapeutic efficacy of homeopathic remedies yielded statistically significant differences compared to placebo. Since the remedies used contained mostly only very low concentrations of pharmacologically active compounds, these effects cannot be accounted for within the framework of current pharmacology. Theories to explain clinical effects of homeopathic remedies are partially based upon changes in diluent structure. To investigate the latter, we measured for the first time high-field (600/500 MHz) 1H T1 and T2 nuclear magnetic resonance relaxation times of H2O in homeopathic preparations with concurrent contamination control by inductively coupled plasma mass spectrometry (ICP-MS). Homeopathic preparations of quartz (10c–30c, n = 21, corresponding to iterative dilutions of 100−10–100−30), sulfur (13x–30x, n = 18, 10−13–10−30), and copper sulfate (11c–30c, n = 20, 100−11–100−30) were compared to n = 10 independent controls each (analogously agitated dilution medium) in randomized and blinded experiments. In none of the samples, the concentration of any element analyzed by ICP-MS exceeded 10 ppb. In the first measurement series (600 MHz), there was a significant increase in T1 for all samples as a function of time, and there were no significant differences between homeopathic potencies and controls. In the second measurement series (500 MHz) 1 year after preparation, we observed statistically significant increased T1 relaxation times for homeopathic sulfur preparations compared to controls. Fifteen out of 18 correlations between sample triplicates were higher for controls than for homeopathic preparations. No conclusive explanation for these phenomena can be given at present. Possible hypotheses involve differential leaching from the measurement vessel walls or a change in water molecule dynamics, i.e., in rotational correlation time and/or diffusion. Homeopathic preparations thus may exhibit specific physicochemical properties that need to be determined in detail in future investigations.