Bradykinin antagonists with dehydrophenylalanine analogues at position 5

Continuing the studies on structural requirements of bradykinin antagonists, it has been found that analogues with dehydrophenylalanine (ΔPhe) or its ring‒substituted analogues (ΔPhe(X)) at position 5 act as antagonists on guinea pig pulmonary artery, and on guinea pig ileum. Because both organs are considered to be bradykinin B₂receptor tissues, the analogues with ΔPhe or ΔPhe(X) at position 5, but without any replacement at position 7, seem to represent a new structural type of B₂receptor antagonist. All the analogues investigated act as partial antagonists; they inhibit the bradykinin‒induced contraction at low concentrations and act as agonists at higher concentrations. Ring substitutions by methyl groups or iodine reduce both the agonistic and antagonistic activity. Only substitution by fluorine gives a high potency. Incorporation of ΔPhe into different representative antagonists with key modifications at position 7 does not enhance the antagonist activity of the basic structures, with one exception. Only the combination of ΔPhe at position 5 with D Phe at position 7 increases the antagonistic potency on guinea pig ileum by about one order of magnitude. Radio‒ligand binding studies indicate the importance of position 5 for the discrimination of B₂receptor subtypes. The binding affinity to the low‒affinity binding site (K_L) was not significantly changed by replacement of Phe by ΔPhe. In contrast, ring‒methylation of ΔPhe results in clearly reduced binding to K_L. The affinity to the high‒affinity binding site (K_H) was almost unchanged by the replacement of Phe in position 5 by ΔPhe, whereas the analogue with 2‒methyl‒dehydrophenylalanine completely failed to detect the K_H‒site. The peptides were synthesized on the Wang‒resin according to the Fmoc/Bu^tstrategy using Mtr protection for the side chain of Arg. The dehydrophenylalanine analogues were prepared by a strategy involving PyBop couplings of the dipeptide unit Fmoc‒Gly‒ΔPhe(X)‒OH to resin‒bound fragments. © 1998 European Peptide Society and John Wiley & Sons, Ltd.     

Synthesis and biological activity of nociceptin/orphanin FQ analogues substituted in position 7 or 11 with Calpha,alpha-dialkylated amino acids

Previous structure-activity and NMR studies on nociceptin/orphanin FQ (N/OFQ) demonstrated that Aib substitution of Ala(7) and/or Ala(11) increases the peptide potency through an alpha helix structure induction mechanism. On these bases we synthesised and evaluated pharmacologically in the mouse vas deferens assay a series of N/OFQ-NH(2) analogues substituted in position 7 and 11 with Calpha,alpha-disubstituted cyclic, linear and branched amino acids. None of the 20 novel N/OFQ analogues produced better results than [Aib(7)]N/OFQ-NH(2). Thus, this substitution was combined with other chemical modifications known to modulate peptide potency and/or efficacy generating compound 21 [Nphe(1)Aib(7)Arg(14)Lys(15)]N/OFQ-NH(2) (coded as UFP-111), compound 22 [(pF)Phe(4)Aib(7)Arg(14)Lys(15)]N/OFQ-NH(2) (UFP-112) and compound 23 [Phe(1)Psi(CH(2)-NH)Gly(2)(pF)Phe(4)Aib(7)Arg(14)Lys(15)]N/OFQ-NH(2) (UFP-113). These novel peptides behaved as highly potent NOP receptor ligands showing full (UFP-112) and partial (UFP-113) agonist and pure antagonist (UFP-111) activities in a series of in vitro functional assays performed on pharmacological preparations expressing native as well as recombinant NOP receptors.     

Synthesis and antiviral activity of C-5 substituted analogues of d4T bearing methylamino- or methyldiamino-linker arms

A general strategy is reported for the preparation of C-5-methylamino- or methyldiamino-d4T analogues of "different sizes". Reactions of the 2',3'-didehydro-2',3'-dideoxy-C-5 hydroxymethyl precursor (7) with either polymethylene diamines (n = 6, 8, 10 and 12) or propargylamine proceed regioselectively via substitution reactions at the C-5 position of uracil. The compounds were evaluated for antiviral activity and cytotoxicity. No significant activity was observed for compounds 9, 11, and 13, but 10 and 12 exhibited a weak activity against HIV-1.     

Enkephalin analogues with 2',6'-dimethylphenylalanine replacing phenylalanine in position 4

Four Leu-enkephalin (Enk) analogues containing 2',6'-dimethyphenylalanine (Dmp) in position 4 were prepared and tested for their receptor binding and biological activities. Among the analogues prepared, [2', 6'-dimethyltyrosine, D-Dmp4]Enk was found to be an antagonist toward mu and delta opioid receptors with pA2 values of 6.90 and 5.57, respectively.     

Structure-activity relationships in a series of NPY Y5 antagonists: 3-amido-9-ethylcarbazoles,core-modified analogues and amide isosteres

Beginning with carbazole 1a, the amide and alkyl substituents were optimized to maintain potency while adding solubilizing groups. Efforts to replace the 3-amino-9-ethylcarbazole core, a known carcinogen, used the SAR generated in the carbazole series for guidance and led to the synthesis of a number of core-modified analogues. In addition, an isosteric series, in which the amide was replaced with an imidazole, was prepared. Two potent new series lacking the putative toxicophore were identified from these endeavors.     

Synthesis and antiviral activity of C-5 substituted beta-D- and beta-L-D4T analogues

A series of beta-D-2',3'-didehydro-2',3'-dideoxy-nucleosides bearing a tether attached at the C-5 position and their beta-L-counterparts was synthesized. Their inhibitory activities against human immunodeficiency virus (HIV) were investigated and compared to establish relationship(s) between compound structure and their antiviral activity. No significant activity was observed for beta-D- and beta-L-modified nucleosides respectively 7a-c and 14a-c, but 7d and 14d exhibited a weak activity against HIV-1.     

Oxytocin analogs with oxygen-containing side chains in position 3

We have synthesized three oxytocin analogs containing an oxygen atom in the amino acid side chain in position 3 to determine the influences of increased side chain length and of hydrophilicity on the potencies and specificities of the resulting analogs. These three analogs: [3-O-methylhomoserine] oxytocin, [3-O-ethylserine] oxytocin, and [3-O-methylthreonine] oxytocin - have the following activities in U/mg: 490, 208, 265, milk ejection; 125, 129, 63, uterus in vivo; 0.2, 16, 0.03, antidiuretic; and 0.1, 0.5, 0.1, pressor. The results show that a longer side chain, [3-O-methylhomoserine] and [3-O-ethylserine] vs. [3-O-methylthreonine], tends to increase all activities. Moving the hydrophilic oxygen farther away from the peptide backbone, on the other hand, decreases vasopressin-like activities but increases or has no effect on oxytocin-like activities.     

Acyclic nucleoside phosphonates with 5-azacytosine base moiety substituted in C-6 position

Two methods for preparation of 6-substituted derivatives of anti DNA-viral agent 1-(S)-[3-hydroxy-2-(phosphonomethoxy)propyl]-5-azacytosine (HPMP-5-azaC) were developed: (1) ammonia mediated ring-opening reaction of diisopropyl esters of HPMP-5-azaC (4) to carbamoylguanidine derivatives followed by ring-closure reaction with orthoesters and (2) condensation reaction of 6-substituted 5-azacytosines with diisopropyl (1S)-[2-hydroxy-1-tosyloxymethyl)ethoxy]methylphosphonate (15). Deprotection of diisopropyl esters to free phosphonic acids was performed with bromotrimethylsilane in acetonitrile followed by hydrolysis. In contrast to parent compound HPMP-5-azaC, a substantial decrease of antiviral activity in case of 6-substituted analogues occurred. Surprisingly, N-3 isomer of 6-methyl-HPMP-5-azaC in the form of isopropyl ester revealed activity against RNA viruses (Sindbis virus).     

Oxytocin antagonists with changes in the Asn5 position shed light on hormone-oxytocin receptor interactions.

Since oxytocin agonists and antagonists have different structure-activity relationships, we have investigated the stereostructural and stereoelectronic requirements of the Asn5 residue in oxytocin antagonists by the synthesis of four analogues of the potent, prolonged acting oxytocin antagonist [Pen1,D-Phe2,Thr4,Orn8]-oxytocin (I) in which Asn5 was replaced respectively with Thr (II), Leu5 (III), Asp5 (IV) and Tyr5 (V). These analogues had pA2 values in the antioxytocic in vitro rat uterine assay of 7.23 (I), 7.16 (II), 6.67 (III), 7.21 (IV), and 6.76 (IV), respectively. All were also found to be weakly potent in the in vivo anti-vasopressor assay in the rat. These studies demonstrate very different structural and stereoelectronic requirements for oxytocin agonists and antagonists when they interact with the oxytocin uterine receptor.     

Design and SAR of new substituted purines bearing aryl groups at N9 position as HIV-1 Tat-TAR interaction inhibitors

Twenty-four purine derivatives bearing aryl groups at N9 position were designed and synthesized as HIV-1 Tat-TAR interaction inhibitors. All the compounds showed high antiviral activities in inhibiting the formation of SIV-induced syncytium in CEM174 cells. Ten of them with low cytotoxicities were evaluated by Tat dependent HIV-1 LTR-driven CAT gene expression colorimetric enzyme assay in human 293T cells at a concentration of 30 microM, indicating effective inhibitory activities of blocking the Tat-TAR interaction. The aryl groups at N9 position affected the binding affinities between compounds and TAR RNA, showing some specificities of aryl groups to TAR RNA.     

Conformational studies of cyclic enkephalin analogues with L- or D-proline in position 3.

The conformation of a series of cyclic enkephalin analogues of a general formula X(1)-cyclo[Y(2)-Z(3)-Nal(4)-Leu(5)] (Nal: beta-(2-naphthyl)alanine), where X = Tyr, Phe, or Phe(NO(2)), Y = D-Dab or L-Dab (Dab: 2,4-diaminobutyric acid), and Z = D-Pro or L-Pro, was studied by means of NMR spectroscopy and theoretical conformational analysis with the Empirical Conformational Energy Program for Peptides and Proteins force field plus solvation. The NMR measurements were performed in dimethyl sulfoxide solution. The nuclear Overhauser effect intensities and coupling constants were used to compute the statistical weights of the conformations of the ensemble generated in global conformational searches. The purpose of this study was to determine whether introducing the D- or L-proline residue in position 3 can produce peptides with both rigid backbone and significant separation of the pharmacophore groups in position 1 and 4 (as required for high affinity for the mu-type opioid receptors). It was found that the analogues with D-Dab in position 2 and D-Pro in position 3 possess a stable type II' beta-turn at positions 3 and 4, which rigidifies the cyclic backbone; this finding was confirmed by independent measurements of the temperature coefficients of the amide protons, which indicated very significant screening of the Leu(5) amide proton from the solvent. However, these analogues were found to possess a short interchromophore distance. The analogues containing both Dab and Pro in the L-configuration are characterized by a larger interchromophore distance; however, they do not possess a stable beta-turn and have therefore a higher conformational flexibility. The modifications proposed in this work are therefore not likely to lead to enkephalin analogues with a high affinity for the mu-receptors.     

Synthesis and potent antimicrobial activity of some novel methyl or ethyl 1H-benzimidazole-5-carboxylates derivatives carrying amide or amidine groups

A series of benzimidazole-5-carboxylic acid alkyl ester derivatives carrying amide or amidine substituted methyl or phenyl groups at the position C-2 were synthesised and evaluated for antibacterial and antifungal activities against S. aureus, methicillin resistant S. aureus (MRSA), S. faecalis, methicillin resistant S. epidermidis (MRSE), E. coli and C. albicans. The results showed that while all simple acetamides are essentially inactive, aromatic amides and amidines have potent antibacterial activities. Aromatic amidine derivatives 13 f-h exhibited the best inhibitory activity with 1.56-0.39 microg/mL MIC values against MRSA and MRSE.     

Berberine derivatives, with substituted amino groups linked at the 9-position, as inhibitors of acetylcholinesterase/butyrylcholinesterase

Berberine derivatives with substituted amino groups linked at the 9-position using different carbon spacers were designed, synthesized, and biologically evaluated as inhibitors of acetylcholinesterase. Compound 10b, with a cyclohexylamino group linked to berberine by a three carbon spacer, gave the most potent inhibitor activity with an IC(50) of 0.020 μM for AChE. Kinetic studies revealed mixed inhibition of AChE, and molecular modeling simulations of the AChE-inhibitor complex confirmed that compounds bound to both the catalytic active site and the peripheral anionic site.     

Synthesis of certain 2'-deoxyuridine derivatives containing substituted phenoxy groups attached to C-5'; evaluation as potential dUTP analogues

Derivatives of 2'-deoxyuridine in which the 5'-OH group is replaced by a 2,3,6-trifluoro-5-hydroxy-4-nitrophenoxy or a 4-carboxy-2,3,6-trifluoro-5-hydroxyphenoxy group have been prepared for evaluation as possible dUTP analogues. They showed a weak ability to displace radiolabelled dUTP from a dUTP-binding antiserum. The corresponding compounds lacking the three fluorine substituents were prepared for comparison.     

Synthesis of novel C-5 substituted d4T analogues bearing linker arms as potential anti-HIV agents.

This work reports the synthesis of 2',3'-didehydro-2',3'-dideoxy-thymidine analogues bearing several kinds of amino-linker arms at the C-5 position of the pyrimidine moiety. C-5 is an attractive position since a flexible chain may permit the triphosphates to be generated. The beta-D- and beta-L-d4T analogues were synthesized following a multi-step reaction from D-ribose and D-xylose, from D- and L-arabinose (towards an oxazoline ring) or from uridine and then were reacted with alkylene diamines.     

Protective effects of GLP-1 analogues exendin-4 and GLP-1(9-36) amide against ischemia-reperfusion injury in rat heart

Glucagon-Like Peptide-1 (GLP-1) is an incretin peptide secreted from intestinal L-cells, whose potent plasma glucose-lowering action has prompted intense efforts to develop GLP-1 receptor-targeting drugs for treatment of diabetic hyperglycemia. More recently, GLP-1 and its analogues have been shown to exert cardiovascular effects in a number of experimental models. Here we tested exendin-4 (Exe-4), a peptide agonist at GLP-1 receptors, and GLP-1(9-36) amide, the primary endogenous metabolite of GLP-1 (both in the concentration range 0.03-3.0 nM), for their protective effects against ischemia-reperfusion injury (IRI) in an isolated rat heart preparation. When administered, the agents were only present for the first 15 min of a 120 min reperfusion period (postconditioning protocol). Exe-4, but not GLP-1(9-36) amide, showed a strong infarct-limiting action (from 33.2% +/-2.7% to 14.5% +/-2.2% of the ischemic area, p<0.05). This infarct size-limiting effect of Exe-4 was abolished by exendin(9-39) (Exe(9-39)), a GLP-1 receptor antagonist. In contrast, both Exe-4 and GLP-1(9-36) amide were able to augment left ventricular performance (left ventricular developed pressure and rate-pressure product) during the last 60 min of reperfusion. These effects were only partially antagonized by Exe(9-39). We suggest that Exe-4, in addition to being currently exploited in treatment of diabetes, may present a suitable candidate for postconditioning trials in clinical settings of IRI. The divergent agonist effects of Exe-4 and GLP-1(9-36), along with correspondingly divergent antagonistic efficacy of Exe(9-39), seem consistent with the presence of more than one type of GLP-1 receptor in this system.     

Synthesis and charge-transfer properties of two acth analogues containing pentamethylphenylalanine in position 9.

In order to investigate the possible role of the Trp residue in ACTH as a change-transfer donor in the activation of ACTH receptors, two ACTH analogues (beta-corticotrophins (I-24) with L-Ser1 and D-Ser1, respectively) containing pentamethylphenylalanine instead of Trp have been synthesized. In these syntheses a new, alkaline-labile, amino-protecting group, the methylsulphonylethyloxycarbonyl group, was employed. The association constants of ACTH (I-24) and (Pmp9)-ACTH (I-24) with the water-soluble acceptor paraquat, were nearly equal.     

Opioid receptor binding and antinociceptive activity of the analogues of endomorphin-2 and morphiceptin with phenylalanine mimics in the position 3 or 4

Endomorphin-2 (EM-2) and morphiceptin are the same class of putative mu-opioid receptor ligands. To investigate the effectiveness of phenylglycine (Phg, L or D) and homophenylalanine (Hfe) as the surrogates of phenylalanine in the position 3 and/or 4 of them, a series of their analogues were synthesized. Opioid receptor binding affinities were determined. Two analogues, [Hfe3]EM-2 and [Phg4] (EM-2/morphiceptin), showed different but potent antinociceptive activity in mouse hot-plate test, the results combined with their half-lives of degradation by mouse brain homogenate could also present some evidence to the in vivo degradative mechanism of EM-2.