Dehydroaminobutyric Acid Research Articles

Toxicity and structure-activity relationship (SAR) of α,β-dehydroamino acids against human cancer cell lines

A library of N-protected dehydroamino acids, namely dehydroalanine, dehydroaminobutyric acid and dehydrophenylalanine derivatives, was screened in three human cancer cell lines [(lung (A549), gastric (AGS) and neuroblastoma (SH-SY5Y)] in order to characterize their toxicological profile and identify new molecules with potential anticancer activity. Results showed N-protected dehydrophenylalanine and dehydroaminobutyric acid derivatives have no or low toxicity for all tested cell lines. The N-protected dehydroalanines exhibit significant toxic effects and the AGS and SH-SY5Y cells were significantly more vulnerable than A549 cells. Four α,β-dehydroalanine derivatives, with IC50<62.5μM, were selected to investigate the pathways by which these compounds promote cell death. All compounds, at their IC50 concentrations, were able to induce apoptosis in both AGS and SH-SY5Y cell lines. In both cell lines, loss of mitochondrial membrane potential (ΔΨm) was found and caspase activity was increased, namely endoplasmic reticulum-resident caspase-4 in AGS cells and caspase-3/7 in SH-SY5Y cells. When evaluated in a non-cancer cell line, the molecules displayed no to low toxicity, thus suggesting some degree of selectivity for cancer cells. The results indicate that α,β-dehydroalanine derivatives can be considered a future resource of compounds able to work as anticancer drugs.

Facile access to α,β-dehydroalanine and α,β-dehydroamino butyric acid derivatives from DL-serines and threonines

ABSTRACTNot only α,β-dehydroamino acids are important constituents for a number of bioactive peptides in nature, but also they are important building blocks for a variety of synthetic amino acids in organic synthesis. Methods to prepare dehydroamino acids have been reported extensively in the literature; however, efficient and convenient protocols are still required. Here we have developed a convenient method to prepare dehydroalanine (ΔAla) and dehydroamino butyric acid (ΔAbu) derivatives derived from DL-serines and DL-threonines, respectively. 4-Toluenesulfonyl chloride (TsCl) and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) were employed in this procedure, which carried out activation of hydroxyl group and β-elimination in one pot. Because it is convenient and easy to handle, this method will attract the attention of synthetic chemists.

Dehydrodipeptide Hydrogelators Containing Naproxen N-Capped Tryptophan: Self-Assembly, Hydrogel Characterization, and Evaluation as Potential Drug Nanocarriers.

In this work, we introduce dipeptides containing tryptophan N-capped with the nonsteroidal anti-inflammatory drug naproxen and C-terminal dehydroamino acids, dehydrophenylalanine (ΔPhe), dehydroaminobutyric acid (ΔAbu), and dehydroalanine (ΔAla) as efficacious protease resistant hydrogelators. Optimized conditions for gel formation are reported. Transmission electron microscopy experiments revealed that the hydrogels consist of networks of micro/nanosized fibers formed by peptide self-assembly. Fluorescence and circular dichroism spectroscopy indicate that the self-assembly process is driven by stacking interactions of the aromatic groups. The naphthalene groups of the naproxen moieties are highly organized in the fibers through chiral stacking. Rheological experiments demonstrated that the most hydrophobic peptide (containing C-terminal ΔPhe) formed more elastic gels at lower critical gelation concentrations. This gel revealed irreversible breakup, while the C-terminal ΔAbu and ΔAla gels, although less elastic, exhibited structural recovery and partial healing of the elastic properties. A potential antitumor thieno[3,2-b]pyridine derivative was incorporated (noncovalently) into the gel formed by the hydrogelator containing C-terminal ΔPhe residue. Fluorescence and Förster resonance energy transfer measurements indicate that the drug is located in a hydrophobic environment, near/associated with the peptide fibers, establishing this type of hydrogel as a good drug-nanocarrier candidate.

Balticidins A-D, antifungal hassallidin-like lipopeptides from the Baltic Sea cyanobacterium Anabaena cylindrica Bio33.

Balticidins A-D (1-4), four new antifungal lipopeptides, were isolated from the laboratory-cultivated cyanobacterium Anabaena cylindrica strain Bio33 isolated from a water sample collected from the Baltic Sea, Rügen Island, Germany. Fractionation of the 50% aqueous MeOH extract was performed by bioassay-guided silica gel column chromatography followed by SPE and repeated reversed-phase HPLC. The main fraction containing the compounds exhibited a strong and specific antifungal activity with inhibition zones in an agar-diffusion assay from 21 to 32 mm against Candida albicans, Candida krusei, Candida maltosa, Aspergillus fumigatus, Microsporum gypseum, Mucor sp., and Microsporum canis. The structures were elucidated by multidimensional (1)H and (13)C NMR spectroscopy, HRESIMS, amino acid analysis, and sugar analysis. Spectroscopic data analysis afforded an unambiguous sequence of R.CHO(S1).CHOH.CONH-Thr(1)-Thr(2)-Thr(3)-HOTyr(4)-Dhb(5)-D-Gln(6)-Gly(7)-NMeThr(8)(S2)-L-Gln COOH(9), in which Dhb is dehydroaminobutyric acid, S1 is d(-)-arabinose-(3-1)-D-(+)-galacturonic acid, S2 is D-(+)-mannose, and R is the aliphatic residue -C13H26Cl or -C13H27. Besides NMeThr, D-allo-Thr, D-Thr, and L-Thr were identified, but the position of the enantiomers in the sequence is not clear. The four balticidins differ in their cyclic (2, 4)/linear (1, 3) core and the presence (1, 2)/absence (3, 4) of chlorine in the aliphatic unit.

Asymmetric Synthesis of Enantiomerically Enriched a-Amino Acids Containing 2-Furyl- and 2-Thienyl-1,2,4-triazoles in the Side-Chain

An efficient method for the asymmetric synthesis of a-amino acids, containing furyl- and thiophenyl-substituted triazoles in their side-chain, is reported. The strategy relies on Michael addition of 3,4,5-substituted 1,2,4-triazoles to the C=C bond of chiral NiII complexes containing the Schiff base formed from dehydroamino acids (dehydroalanine and (E + Z)-dehydroaminobutyric acid) and from chiral auxiliaries, i. e. (S)-2-N-(N0-benzylprolyl)aminobenzophenone and (S)-2-N- (N0-2-chlorobenzylprolyl) aminobenzophenone. The reactions proceeded with good to very good diastereoselectivity. Hydrolysis of the diastereomeric mixtures of metal complexes afforded the enantiomerically pure a-amino acids with high enantiomeric excess (ee&gt; 98%).

Synthesis of 2,6-Bis(oxazolyl)pyridine Ligands for Luminescent LnIII Complexes

New bis(oxazolyl)pyridine ligands for LnIII ions were prepared by using an expeditious methodology from threonine and dipicolinic chloride. The synthetic strategy includes a dehydration step to give a bis(dehydroaminobutyric acid) derivative followed by bromination and cyclization with DBU. Photophysical studies of EuIII and TbIII complexes of these ligands showed that the 2,6-bis(oxazolyl)pyridine moiety acts as an effective sensitizer for lanthanide luminescence and indicate the formation of 3:1 complexes [Ln{bis(oxazole)pyridine}3]3+.

Selective Reduction of C=C Double Bonds in Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry of Microcystins

Cyanobacteria are photosynthetic bacteria encountered in various aquatic environments. Some of them are able to produce powerful toxins called cyanotoxins. Among cyanotoxins, microcystins (MCs) constitute a group of closely related cyclic heptapeptides. Their sequences are made up of classical amino acids as well as post- translational modified ones. Interestingly, in vivo metabolism of microcystins seems to be greatly dependent on various minor structural differences and particularly those of the seventh amino acid, which can be either dehydroalanine (or a derivative), dehydroaminobutyric acid (or a derivative), serine or alanine. As a consequence, microcystins have been classified on the basis of the nature of this singular amino acid. A major difficulty in the classification of such toxins is that some of them share the same molecular masses and the same molecular formulas. Consequently, a simple mass measurement is not sufficient to determine the structure and the class of a toxin of interest. Heavy and expensive techniques are used to classify them, such as multi-dimensional nuclear magnetic resonance and amino acid analysis. In this work, a new matrix-assisted laser desorption/ionization time-of-flight method leading to an easy classification of MCs is proposed. The methodology relies on the reductive properties of the matrix 1,5-diaminonaphtalene (1,5-DAN) which appears to be able to selectively reduce the double carbon-carbon bond belonging to the seventh amino acid. Moreover, the yield of reduction seems to be influenced by the degree of substitution of this double bond, allowing a discrimination between dehydroalanine and dehydroaminobutyric acid. This selective reduction was confirmed by the study of three synthetic peptides by mass spectrometry and tandem mass spectrometry. According to these results, the use of reductive matrices seems to be promising in the study of microcystins and in their classification. More generally, 1,5-DAN allows the selective reduction of double carbon-carbon bonds. This property could also be employed in the characterization of others types of compound displaying double bonds (petrochemistry, metabolomics....).

An improved procedure for the synthesis of dehydroamino acids and dehydropeptides from the carbonate derivatives of serine and threonine using tetrabutylammonium fluoride

Dehydroamino acids are important precursors for the synthesis of a number of unnatural amino acids and are structural components in many biologically active peptide derivatives. However, efficient synthetic procedures for their production in large amounts and without side reactions are limited. We report here an improved procedure for the synthesis of dehydroalanine and dehydroamino butyric acid from the carbonate derivatives of serine and threonine using TBAF. The antiselective E(2) elimination of the carbonate derivatives of serine and threonine using TBAF is milder and more efficient than other available procedures. The elimination reaction is completed in less than 10 min with various carbonate derivatives studied and the methodology is very efficient for the synthesis of dehydroamino acids and dehydropeptides. The procedure thus provides an easy access to key synthetic precursors and can be used to introduce interesting structural elements to designed peptides.

Pyrenylamino Acids: Synthesis, Photophysical and Electrochemical Studies

AbstractSeveral β‐pyrenyldehydroamino acids and a pyrenylalanine derivative have been synthesized in good‐to‐high yields from dehydroamino acids by several types of reactions. A β‐[(pyren‐1‐yl)methylamino]alanine was prepared by treating the methyl ester of N,N‐(di‐tert‐butoxycarbonyl)dehydroalanine with (pyren‐1‐yl)methylamine hydrochloride in the presence of an excess of potassium carbonate. The E isomer of the methyl esters of N‐(tert‐butoxycarbonyl)‐β‐(1,2,4‐triazol‐1‐yl)dehydroalanine and dehydroaminobutyric acid were treated with (pyren‐1‐yl)methylamine hydrochloride in the presence of triethylamine to give the E isomers of the β‐aminomethylpyrene dehydroalanine and dehydroaminobutyric acid derivatives. β‐(Pyren‐1‐yl)dehydrophenylalanine and dehydroaminobutyric acid derivatives were obtained from the pure stereoisomer of the corresponding β‐bromodehydroamino acids and (pyren‐1‐yl)boronic acid by Suzuki cross‐coupling. This reaction was also applied successfully to β‐bromodehydrodipeptides. The electrochemical behaviour of some of the compounds prepared was studied by cyclic voltammetry. The peak potentials for the oxidation and reduction of pyrenylalanines were similar to those reported for pyrene. However, it was found that when the pyrene ring was conjugated with the dehydroamino acid moiety, the compounds had higher oxidation and reduction peak potentials than pyrene. The fluorescence properties of four of the pyrenylamino acids synthesized were evaluated in cyclohexane and alcohols. The methyl ester of N,N‐bis(tert‐butoxycarbonyl)‐β‐[(pyren‐1‐yl)methylamino]alanine presented high fluorescence quantum yields in cyclohexane and ethanol (ΦF = 0.45 and 0.35, respectively). (© Wiley‐VCH Verlag GmbH &amp; Co. KGaA, 69451 Weinheim, Germany, 2008)

Synthesis of Substituted Oxazoles from N‐Acyl‐β‐hydroxyamino Acid Derivatives

AbstractSeveral N‐acyl‐β‐hydroxyamino acids were prepared and treated with di‐tert‐butyl dicarbonate in the presence of 4‐(dimethylamino)pyridine, followed by treatment with N,N,N′,N′‐tetramethylguanidine to give the corresponding N‐acyldehydroamino acids in good to high yields. These were then treated with I2/K2CO3 followed by 1,8‐diazabicyclo[5.4.0]undec‐7‐ene. The methyl esters of N‐acyldehydroaminobutyric acid gave the corresponding substituted oxazoles in good to high yields. The N‐acyldehydrophenylalanines gave 5‐phenyloxazole derivatives in low to moderate yields together with β‐iododehydrophenylalanines. Under the same conditions, N‐acyldehydroalanines failed to give the corresponding oxazoles. However, when the reaction was carried out in the absence of DBU, it was possible to isolate the β,β‐diiododehydroalanine derivatives. Although the reason for the different reactivities of the N‐acyldehydroamino acids is not completely clear to us, cyclic voltammetry studies showed that the less‐reactive derivatives have higher reduction potentials. This suggests that the double bonds in dehydroaminobutyric acid derivatives are more susceptible to electrophilic attack by iodine.(© Wiley‐VCH Verlag GmbH &amp; Co. KGaA, 69451 Weinheim, Germany, 2008)

Reactivity of Dehydroamino Acids and Dehydrodipeptides TowardsN‐Bromosuccinimide: Synthesis of β‐Bromo‐ and β,β‐Dibromodehydroamino Acid Derivatives and of Substituted 4‐Imidazolidinones

AbstractWe have developed a modification of our previously reported high‐yielding method for the synthesis ofN,N‐diacyldehydroamino acid derivatives to prepareN‐monoprotected dehydroamino acids and dehydrodipeptides. Thus, several dehydroalanine, dehydroaminobutyric acid and dehydrophenylalanine derivatives have been prepared by treating the correspondingL‐serine,L‐threonine andD,L‐3‐phenylserine (threo‐type) derivatives with 1 equiv. of di‐tert‐butyl dicarbonate and 4‐(dimethylamino)pyridine. The reaction proceeded with the initial formation of anO‐tert‐butyl carbonate which, by treament withN,N,N′,N′‐tetramethylguanidine, underwent β elimination to give the corresponding dehydroamino acid derivative. This two‐step method can be carried out as a one‐pot procedure and is stereoselective, giving only theZisomer. TheN‐monoprotected dehydroamino acids were treated withN‐bromosuccinimide and thereafter with triethylamine to afford several β,β‐dibromodehydroalanines or β‐bromo‐, β‐alkyl‐ or β‐aryldehydroalanines. The latter were obtained as mixtures ofEandZisomers. An increased stereoselectivity towards the formation of theZisomer was observed with dehydrophenylalanine and when 4‐tolylsulfonyl was used as theN‐protecting group. In the case of dehydrodipeptides, the reaction with NBS and triethylamine afforded the corresponding brominated dehydrodipeptides when theN‐protecting group was other than 4‐tolylsulfonyl. However, when the reagent was a peptide with a dehydroamino acid as the second residue and anN‐(4‐tolylsulfonyl) group the corresponding 2,2‐disubstituted 1‐(4‐tolylsulfonyl)imidazolidin‐4‐ones were obtained in good‐to‐high yields. (© Wiley‐VCH Verlag GmbH &amp; Co. KGaA, 69451 Weinheim, Germany, 2007)

An efficient synthesis of dehydroamino acids and dehydropeptides from O-Cbz and O-Eoc derivatives of serine and threonine

A simple and efficient method for the synthesis of dehydroamino acids from serine and threonine is reported. Various O-Cbz and O-Eoc derivatives of serine and threonine are prepared using CbzCl and EocCl, respectively, and are subjected to an anti-selective elimination on treatment with K 2CO 3 in DMF at 65 °C to afford dehydroalanine and dehydroamino butyric acid derivatives, respectively, in excellent yields. The high stability of these carbonate derivatives of serine and threonine allows their use in normal peptide synthesis as protected serine and threonine residues. Peptides synthesized by incorporating O-Cbz or O-Eoc derivatives undergo ready elimination under the reported conditions, to give the corresponding dehydropeptides in excellent yields. The reaction conditions are mild enough not to cause the racemization of other stereogenic centers present in the peptide.

Hassallidin A, a Glycosylated Lipopeptide with Antifungal Activity from the Cyanobacterium Hassallia sp.

Hassallidin A (1), a new antifungal glycosylated lipopeptide, was isolated from an epilithic cyanobacterium collected in Bellano, Italy, identified as Tolypothrix (basionym Hassallia) species. Chemical, mass spectrometric, and spectroscopic analyses, including one- and two-dimensional NMR, were performed to determine an esterified eight-residue cyclic peptide linked with a carbohydrate and a fatty acid residue. Chiral GC-MS analysis revealed the occurrence of the nonproteinogenic amino acids D-allo-Thr, D-Thr, D-Tyr, D-Gln, and dehydroaminobutyric acid (Dhb) within the peptide moiety. The additional components of hassallidin A could be identified as alpha,beta-dihydroxytetradecanoic acid (Dht) and mannose. This is the first report on a cyclic peptide of cyanobacterial origin that contains both a fatty acid and a carbohydrate moiety. Compound 1 exhibits antifungal activity against Aspergillus fumigatus and Candida albicans with MIC values of 4.8 microg/mL for both test organisms.

Phase I Clinical and Pharmacokinetic Study of Kahalalide F in Patients with Advanced Androgen Refractory Prostate Cancer

The purpose is to determine the maximum tolerated dose, profile of adverse events, and dose-limiting toxicity of Kahalalide F (KF) in patients with androgen refractory prostate cancer. Furthermore, the pharmacokinetics after KF administration and preliminary antitumor activity were evaluated. KF is a dehydroaminobutyric acid-containing peptide isolated from the marine herbivorous mollusk, Elysia rufescens. Adult patients with advanced or metastatic androgen refractory prostate cancer received KF as an i.v. infusion over 1 hour, during five consecutive days every 3 weeks. The starting dose was 20 microg per m(2) per day. Clinical pharmacokinetics studies were done in all patients using noncompartmental analysis. Prostate-specific antigen levels were evaluated as a surrogate marker for activity against prostate cancer. Thirty-two patients were treated at nine dose levels (20-930 microg per m(2) per day). The maximum tolerated dose on this schedule was 930 microg per m(2) per day. The dose-limiting toxicity was reversible and asymptomatic Common Toxicity Criteria grade 3 and 4 increases in transaminases. The recommended dose for phase II studies is 560 microg per m(2) per day. Pharmacokinetics analysis revealed dose linearity up to the recommended dose. Thereafter, a more than proportional increase was observed. Elimination was rapid with a mean (SD) terminal half-life (t(1/2)) of 0.47 hour (0.11 hour). One patient at dose level 80 microg per m(2) per day had a partial response with a prostate-specific antigen decline by at least 50% for > or =4 weeks. Five patients showed stable disease. KF can be given safely as a 1-hour i.v. infusion during five consecutive days at a dose of 560 microg per m(2) per day once every 3 weeks.

Studies on the formation of lysinomethylalanine and histidinomethylalanine in milk products.

From reaction mixtures consisting of N-acetyldehydroaminobutyric acid methyl ester and N alpha-acetyl-L-lysine or N alpha-acetyl-L-histidine, respectively, distinct amounts of the cross-link amino acids N epsilon-(2-amino-2-carboxy-1-methyl-ethyl)-L-lysine (lysinomethylalanine, LMeAL) and N tau-(2'-amino-2'-carboxy-1'-methyl-ethyl)-L-histidine (histidinomethylalanine, HMeAL) were isolated via preparative ion-exchange chromatography and identified by 1H- and 13C-nuclear magnetic resonance. In the amino acid chromatogram, both compounds eluted clearly separated from other basic amino acids. However, neither LMeAL nor HMeAL could be detected in numerous acid hydrolysates of a range of milk products. In model studies, threonine showed a significantly lower tendency for an alkali-induced beta-elimination reaction compared to serine. The reactivity of the resulting dehydroaminobutyric acid towards nucleophiles was more than tenfold lower as compared to dehydroalanine. Thus, the formation of LMeAL as well as of HMeAL during food processing is negligible.

Conformation of dehydropeptides

Six model dipeptide methylamides containing dehydroaminobutyric acid (delta Abu) of the type Boc-X-delta z Abu-NHCH3 and Box-X-delta E Abu-NHCH3, X = Ala, Val, Phe (Boc = tert-butoxycarbonyl), have been synthesized and their solution conformations explored using 300 MHz 1H NMR and IR spectroscopy. Studies based on delineation of intramolecularly hydrogen bonded NH groups in CDCl3 and (CD3)2SO revealed that none of the NH groups is appreciably solvent shielded. Difference NOE (Nuclear Overhauser Effect) studies have also failed to detect the presence of any discernible turn structure in these peptides. These studies indicate that the conformational preferences of peptides containing, alpha, beta-dehydroaminobutyric acid are different from those of delta ZPhe and delta ZLeu. It appears that steric interactions due to the beta-substituent in the dehydroamino acid moiety play an important role. Unlike delta ZPhe and delta ZLeu, which have relatively large beta-substituents, phenyl and isopropyl, respectively, and stabilize a beta-turn, the beta-methyl group of delta ZAbu or delta EAbu is readily accommodated in extended conformation. Clearly, the size of beta-substituent in dehydroamino acid crucially influences the conformational preferences. Thus, it may be possible to use different dehydroamino acids to introduce variable but definite constraints in synthetic peptides.