Exchange protein activated by cAMP (Epac) mediates cAMP activation of p38 MAPK and modulation of Ca2+-dependent K+ channels in cerebellar neurons.

The exchange factor directly activated by cAMP (Epac) is a newly discovered direct target for cAMP and a guanine-nucleotide exchange factor for the small GTPase Rap. Little is known about the neuronal functions of Epac. Here we show that activation of Epac by specific cAMP analogs or by the pituitary adenylate cyclase-activating polypeptide induces a potent activation of the Ca2+-sensitive big K+ channel, slight membrane hyperpolarization, and increased after-hyperpolarization in cultured cerebellar granule cells.

The epimerization of peptide aldehydes--a systematic study.

Peptide aldehydes are interesting targets as enzyme inhibitors, and can be used for pseudopeptide chemistry or ligation. However, they are known to be subjected to epimerization during synthesis or purification. By (1)H NMR, a model dipeptide aldehyde can be used to check the possible epimerization occurring during synthesis.

Solid-phase synthesis of arginine-containing peptides and fluorogenic substrates using a side-chain anchoring approach.

Attachment of an amino acid to a solid support by its side chain is sometimes necessary to take advantage of an alpha-carboxylic group available for diverse modifications, including the incorporation of a fluorophore for the preparation of fluorogenic substrates. In contrast to most other amino acids, anchoring the guanidinium group of an arginine to a resin requires the use of a supplementary linker. To avoid the usually multistep synthesis of such a linker as well as its difficult attachment to the guanidine group, we developed a simple method where the guanidine group is built on a Rink amide resin.

Biological activity of silylated amino acid containing substance P analogues.

The need to replace natural amino acids in peptides with nonproteinogenic counterparts to obtain new medicinal agents has stimulated a great deal of innovation on synthetic methods. Here, we report the incorporation of non-natural silylated amino acids in substance P (SP), the binding affinity for the two hNK-1 binding sites and, the potency to stimulate phospholipase C (PLC) and adenylate cyclase of the resulting peptide. We also assess the improvement of their stability towards enzyme degradation.

Microtubule-associated protein 2 (MAP2) expression during synaptic plasticity in the guinea pig cochlea.

The expression of different isoforms of microtubule-associated proteins 2 (MAP2), including the low molecular weight form MAP2c present mainly in developing neurons, was investigated in the primary auditory neurons after alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) perfusion in the guinea pig cochlea. MAP2 expression appeared to be tightly regulated in the repairing neurons. Neurite regrowth seems to involve the MAP2c isoform.

From beta-N-tert-butyloxycarbonyl N-carboxyanhydrides to beta-aminoacid derivatives.

beta-N-tert-butyloxycarbonyl-N-carboxyanhydrides are very reactive beta-amino acid derivatives. They react cleanly and smoothly with different nucleophiles like aminoesters, enolates, N-methyl-d-glucamine, amidoximes to afford in good to excellent yields peptides, beta-amino ketocompounds, beta-aminosugars and functionalized disubstituted 1,2,4-oxadiazoles.

Solution and solid-supported synthesis of 3,4,5-trisubstituted 1,2,4-triazole-based peptidomimetics.

[reaction: see text] 3,4,5-Trisubstituted 1,2,4-triazoles were synthesized in solution from various thioamides and hydrazides in smooth experimental conditions leading to peptidomimetic scaffolds. This strategy was found to be compatible with the usual peptide synthesis protecting groups. This methodology was then applied on solid support by anchoring alpha-amino acids through their amino function to an activated carbonate resin.

Synthesis of various 3-substituted 1,2,4-oxadiazole-containing chiral beta 3- and alpha-amino acids from Fmoc-protected aspartic acid.

Various 3-substituted chiral 1,2,4-oxadiazole-containing Fmoc-beta(3)- and -alpha-amino acids were synthesized from Fmoc-(l or d)-Asp(OtBu)-OH and Fmoc-l-Asp-OtBu, respectively, in three steps (i.e., condensation of an aspartyl derivative with differentially substituted amidoximes, formation of the 1,2,4-oxadiazole, and cleavage of the tert-butyl ester).

Optically pure beta-substituted beta-hydroxy aspartates as glutamate transporter blockers.

A short asymmetric synthesis of optically pure beta-substituted beta-hydroxy aspartates is described. The key step is an aldol reaction between a glycine enolate derived from an oxazinone intermediate used as chiral auxiliary and various alpha-keto esters. Excellent diastereomeric excesses are obtained.

Imaging combinatorial libraries by mass spectrometry: from peptide to organic-supported syntheses.

Supported peptide and drug-like organic molecule libraries were profiled in single nondestructive imaging static secondary ion mass spectrometric experiments. The selective rupture of the bond linking the compound and the insoluble polymeric support (resin) produced ions that were characteristic of the anchored molecules, thus allowing unambiguous resin bead assignment. Very high sensitivity and specificity were obtained with such a direct analytical method, which avoids the chemical release of the molecules from the support.

Post-synthesis incorporation of a lipidic side chain into a peptide on solid support.

A new strategy for the synthesis of lipopeptides has been developed. Using Weinreb (N-methoxy, N-methyl) amide as an aldehyde function precursor on the side chains of Asp or Glu residues, this new strategy avoids the synthesis of a lipidic amino acid residue before its incorporation in the peptide sequence. The aldehyde generated on the solid support can react with ylides leading to unsaturated or saturated side chains or with various nucleophiles to yield non-coded amino acid residues incorporated into the sequence.

Gastrin induces over-expression of genes involved in human U373 glioblastoma cell migration.

Astrocytic tumors are the most common and the most malignant primary tumors of the central nervous system. We had previously observed that gastrin could significantly modulate both cell proliferation and migration of astrocytoma cells. We have investigated in the present study which genes could be targeted by gastrin in tumor astrocyte migration.

Urethane N-carboxyanhydrides from beta-amino acids.

A general method has been developed for the synthesis of N-tert-butyloxycarbonyl N-carboxyanhydrides from beta-amino acids using Vilsmeier complex. These beta-UNCA are stable, and the reactivity with different nucleophiles (alcohol, amine, lithium enolate) was studied.

Synthesis of a new insoluble polymer-supported chiral alcohol auxiliary and its first application to nucleophilic addition to ketenes.

The preparation of a new optically active alcohol with a carboxylic function that allowed its attachment to an amine-functionalized insoluble polymer is described. Its first use as a polymer supported chiral auxiliary is demonstrated by asymmetric transformation of two racemic aryl propionic acids via ketene formation (95-96% ee).

A silaproline-containing dipeptide.

The silaproline-containing dipeptide N-(3, 3-dimethyl-1-pivaloyl-1-aza-3-sila-5-cyclopentylcarbonyl)-L- alanine isopropylamide, C(17)H(33)N(3)O(3)Si, has two independent molecules in the asymmetric unit and each adopts a beta-II folded conformation, where the amide on the terminal C interacts intramolecularly with the pivaloyl O atom. The five-membered silaproline ring is C(beta)-puckered, an infrequent conformation for the homologous proline ring.

A synthetic glycine-extended bombesin analogue interacts with the GRP/bombesin receptor.

alpha-amidation of a peptide (which takes place from a glycine-extended precursor) is required to produce biologically active amidated hormones, such as gastrin-releasing peptide (GRP)/Pyr-Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH(2) (bombesin). It was shown that glycine-extended gastrin mediates mitogenic effects on various cell lines by interacting with a specific receptor, different from the classical CCK(1) or CCK(2) receptors. On the basis of this observation, we have extended the concept of obtaining active glycine-extended forms of others amidated peptides to produce new active analogues.

Mass spectrometry in combinatorial chemistry.

In the fast expanding field of combinatorial chemistry, profiling libraries has always been a matter of concern--as illustrated by the buoyant literature over the past seven years. Spectroscopic methods, including especially mass spectrometry and to a lesser extent IR and NMR, have been applied at different levels of combinatorial library synthesis: in the rehearsal phase to optimize the chemistry prior to library generation, to confirm library composition, and to characterize after screening each structure that exhibits positive response. Most of the efforts have been concentrated on library composition assessment.

A rational approach to the design and synthesis of a new bradykinin B(1) receptor antagonist.

We have previously synthesized a potent and selective B(1) bradykinin receptor antagonist, JMV1645 (H-Lys-Arg-Pro-Hyp-Gly-Igl-Ser-D-BT-OH), containing a dipeptide mimetic ((3S)-amino-5-carbonylmethyl-2,3-dihydro-1, 5-benzothiazepin-4(5H)-one (D-BT) moiety) at the C-terminal. Analogues of this potent B(1) bradykinin receptor antagonist in which the central Pro(2)-Hyp(3)-Gly(4)-Igl(5) tetrapeptide has been replaced by constrained N-1-substituted-1,3,8-triazaspiro¿4. 5decan-4-one ring system were synthesized.

Synthesis and biological evaluation of bradykinin B(1)/B(2) and selective B(1) receptor antagonists.

We recently described a potent bradykinin B(2) receptor agonist (JMV1116) obtained by replacing the D-Tic-Oic dipeptide moiety of HOE140 by a (3S)-amino-5-(carbonylmethyl)-2,3-dihydro-1, 5-benzothiazepin-4(5H)-one (D-BT) moiety. This compound inhibited the specific binding of [(3)H]BK on membranes of CHO cells expressing the human cloned B(2) receptor with nanomolar affinity and contracted both isolated rat uterus and human umbilical vein. These data demonstrated that D-BT could be a good mimic of the Pro-Phe dipeptide.

Identification of synthetic by-products in combinatorial libraries using high performance liquid chromatography-electrospray ionization mass spectrometry.

High performance liquid chromatography (HPLC), electrospray ionization mass spectrometry (ESI) and high performance liquid chromatography coupled to mass spectrometry (LC-MS) were used to analyze randomly chosen samples from parallel syntheses carried out on derivatized polypropylene crowns compatible with a Multipin solid support system. Side-reactions and by-products were clearly identified, and the yields of the expected molecules were unexpectedly low for most samples. LC-MS was superior to HPLC with absorbance detection or electrospray mass spectrometry alone for determining the identity and purity of each desired combinatorial compounds.

A side-reaction in the SPPS of Trp-containing peptides.

Syntheses of several Trp-containing peptides on a Wang solid support afforded significant amounts of a side-product. 1H-NMR and MS data showed that an unexpected alkylation by the linker has occurred on the indole nucleus. This was observed whatever the scavenger used, and whatever the position of the Trp residue in the sequence, unless it was in the C-terminal position.

Synthesis and characterization of bradykinin B(2) receptor agonists containing constrained dipeptide mimics.

We have previously shown that substitution of the D-Tic-Oic dipeptide by a (3S)-[amino]-5-(carbonylmethyl)-2,3-dihydro-1, 5-benzothiazepin-4(5H)-one (D-BT) moiety in the bradykinin B(2) receptor antagonist HOE 140 resulted in a full potent and selective bradykinin B(2) receptor agonist (H-DArg-Arg-Pro-Hyp-Gly-Thi-Ser-D-BT-Arg-OH, JMV1116) exhibiting a high affinity for the human receptor (K(i) 0.7 nM). In the present study, we have investigated the effects of replacement of the D-Tic-Oic moiety by various constrained dipeptide mimetics.

Design and synthesis of potent bradykinin agonists containing a benzothiazepine moiety.

A bradykinin analogue (H-Arg-Pro-Pro-Gly-Phe-Ser-D-BT-Arg-OH, 3) in which the Pro-Phe dipeptide was replaced by the (3S)[amino]-5-(carbonylmethyl)-2,3-dihydro-1, 5-benzothiazepin-4(5H)-one (D-BT) moiety has been synthesized. The same modification was performed on the potent bradykinin B(2) receptor antagonist HOE 140 (H-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-Tic-Oic-Arg-OH), in which the -D-Tic-Oic- moiety was replaced by D-BT to yield H-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-BT-Arg-OH, 1 (JMV1116). These compounds were examined in vitro for their binding affinity toward bradykinin B(1) and B(2) receptors as well as for their ability to interfere with bradykinin-induced contraction of both human umbilical vein and rat uterus.

A delayed-early response nuclear gene encoding MRPL12, the mitochondrial homologue to the bacterial translational regulator L7/L12 protein.

We have characterized a new delayed-early response mRNA encoding a 21-kDa product (MRPL12) that accumulates during the G1 phase of growth-stimulated cells. MRPL12 is the mammalian homologue to chloroplastic and bacterial L12 ribosomal proteins. Immunofluorescence microscopy and cell fractionation indicate a predominant mitochondrial localization in various mammalian cell lines.