SYNTHESIS AND SCREENING OF NEW ENANTIOMERICALLY ENRICHED NON-PROTEIN (S) AND (R)-á-AMINO ACIDS, PEPTIDES AND POLYMER
SYNTHESIS AND SCREENING OF A NEW GENERATION OF OPTICALLY ACTIVE NON-PROTEINOGENIC á-AMINO ACIDS, PEPTIDES AND POLYMERS CONTAINING UNSATURATED GROUPS IN THE SIDE CHAIN
Tech Area / Field
- CHE-SYN/Basic and Synthetic Chemistry/Chemistry
- MED-DRG/Drug Discovery/Medicine
8 Project completed
Senior Project Manager
Algiyev S K
Yerevan State University, Armenia, Yerevan
- Agricultural University of Georgia, Georgia, Tbilisi\nScientific and Production Center "Armbiotechnology" NAS RA, Armenia, Yerevan
- University of Texas at San Antonio, USA, TX, San Antonio\nUniversität Rostock, Germany, Rostock\nUniversitat Politècnica de Catalunya, Spain, Barcelona\nUniversite de Paris-Sud, France, Orsay
Project summaryThe goal of the project is development of methods for the synthesis of new enantiomerically enriched (S) and (R)-á-amino acids, containing acetylene groups in the side chain, peptides and polymers based thereon; study of antibacterial, antitumor properties to reveal potentially pharmacologically active preparations.
Amino acids playing a central role in all process of a live cell are widely used in biosynthesis of molecules that have specific biological activity. As a rule, in synthesis of peptides and other amino acid-based biopreparations only natural amino acids were previously used [1-3]. However, in the last 20 years in production of peptides and other medications enantiomerically enriched non-proteinogenic amino acids containing different functional groups (aliphatic, aromatic and heterocyclic) in the side chain have been used. Among similar compounds amino acids containing acetylene bond in the side chain radical occupy a special place .
Such amino acids were used as selective inhibitors of Endothelin-converting enzymes, inhibitors of thrombin and cathepsin B [4-5], inactivators of pyridoxalphosphate-dependent ã-cystathionase, growth inhibitors of B. Subtillis B-50, etc. In particular, propargylglycylchloroalanyl dipeptides have a strong antibacterial effect, 2-aminohex-5-ynoic acid inhibits growth of B. Subtillis B-50 [6-10], arylacetylene amino phosphonate dipeptide inhibits Endothelin-Converting Enzyme-1 (ECE 1) and Neutral Endopeptidase (NEP), etc . Unsaturated amino acids also were used as a protein bioconjugate. . They represent very interesting building blocks. Indeed, alkenyl- or alkynylside chains can be functionalized by many chemical functions and offer a wide range of possible transformations. Particularly, unsaturated á-amino acids give access to many synthetic applications in all fields of chemistry Among them, metal catalysed cross-coupling reactions and cross metathesis are commonly used to generate peptide modifications They can also be substrates of biological interest for fluorination for example as PET radiotracers. They are very interesting and useful tools for « click » chemistry in peptidomimetic drug design or covalent modification of proteins. They can also be incorporated in compounds as beta-turn inducer to promote secondary structures. Finally they can be used for the preparation of stapled peptides. Stapled peptides were first designed by Verdine as a hydrocarbon staple that ‘locked’ a flexible peptide into the shape of an alpha-helix by reaction of two á,á-disubstituted non-natural amino acids bearing an olefin side chain . In recent years a great number of papers have appeared on stapled pepides and it has been demonstrated that also monosubstituted alkenyl amino acids are suitable for their preparation .
Within the framework of the project it is also envisaged to synthesize non-proteinogenic amino acids containing acetylene bonds in the side chain radical as well as to study their antibacterial and antitumor properties. The proposed project is a continuation of the research fulfilled in the framework of two ISTC projects #A-1247 and #A-1677. In the course of these projects realization about 100 novel aliphatic and heterocycle-substituted non-proteinogenic alpha amino acids and peptides containing non-proteinogenic amino acids have been synthesized. Synthesized compounds were screened on microbiological, biochemical and cytological models to reveal their biological activity. As a result new inhibitors of serine proteases as well as compounds with antibacterial activity were revealed.
Non-proteinogenic á-amino acids containing acetylene bond belong to a class of substrates that irreversibly inhibit the enzyme (suicide substrate). There are three ways of “suicidal” effect of the acetylene substrate on the active site of the enzyme: 1) by carbanion formation at an adjacent carbon followed by propargylic rearrangement to an allene, 2) by oxidation of the side group to ketone and 3) by monoxygeneration to oxypropene . This fact is the crux in selection of biological models to reveal pharmacological activities of new non-proteinogenic á-amino acids and peptides that are scheduled to be synthesized within the frame of this project.
Antibacterial compounds are inhibitors of enzymes involved in the biosynthesis of peptidoglycan – component of the cell wall. Such enzymes are e.g. alanine racemase, DAla:Dala ligase and aminotransferase of D-amino acids. All these enzymes are inhibited by alanine analogs. It is especially important to take into account the microbiological specificity of the enzyme, i.e. search for inhibitors should be conducted considering the enzyme origin. Due to the discovery of the phenomenon of inactivation (irreversible inhibition) of the enzyme based on the operating mechanism of the enzyme itself (substrate interacting with the active site of the enzyme causes enzyme suicide), the search for new inhibitors among such compounds is especially attractive. Non-proteinogenic amino acids and peptides containing acetylene bond in the side-chain radical are expected to have high level of specificity since they are suicide substrates .
Another targeted study of biological activity of new non-proteinogenic á-amino acids containing acetylene bond in the side-chain radical and peptides are inhibitors of metalloproteases. Matrix metalloproteases (ММРs) belong to zinc-dependent metalloproteases. They play an important role in degradation of extracellular matrix in both norm and at various pathologies . The interaction of cells with extracellular matrix is a key moment for the human body normal development and functioning. However, matrix metalloproteases are also responsible for many proteolytic processes that lead to tumor development. They are involved in matrix hydrolysis thus promoting spread of metastases in oncological patients. Metalloprotease inhibitors are able to prevent this process. In this connection, metalloproteases are targets for creation of antitumor drugs. Unfortunately, clinical trials of gelatinize inhibitors on oncological patients have not revealed therapeutic effect so far; moreover undesirable side effects were registered. The search for new highly specific compounds able to inhibit metalloproteases is one of the directions in creation of antitumor drugs. At the preliminary stage our enantiomerically enriched (S)-2-amino-2-(benzyl)pent-4-ynoic and (S)-2-amino-2(3,4-dichlorobenzyl)pent-4-ynoic new á-amino acids were studied on biological targets and proved to have inhibiting activity in relation to metalloproteinases. It should be mentioned that non-proteinogenic á-amino acids containing acetylene bond in the side-chain radical are key units in the synthesis of compounds that are inhibitors of thrombin and cathepsin B [1,4].
In addition, the incorporation of non-proteinogenic amino acids (NPAAs) into the polymeric backbones can result in multiple positive effects such as: enhanced stability compared to NPAAs based peptides, better bioavailability (e.g. via the preparation of nanoparticles), increased membrane penetration (e.g. via imparting cationic nature to macromolecules), prolonged action and decreased immunogenicity. It is important that these polymers will be degraded after the fulfillment of their functions, and the degradation fragments can be either assimilated or cleared from the body. One of the most popular ways to render amino acid based on polymers biodegradable is the incorporation of easily hydrolysable ester bonds into their backbones . It is also important for the realization of biological activities that the polymers be water soluble. This can be achieved by incorporating ether bonds in the polymeric backbones as it has been demonstrated for arginine-based biodegradable ester polymers, or by attaching to the polymeric chains solubilizing (e.g. PEG) groups via lateral functional groups. In other words, we plan to construct NPAAs based ester and ether-ester polymers, and these polymers will be four classes – poly(ester amide)s & poly(ether ester amide)s, and poly(ester urethane)s & poly(ether ester urethane)s. Some cationic polymers to enhance membrane penetration and water insoluble polymers for making nanoparticle are included in the research plan as well [17-18].
Different synthetic approaches have been developed to obtain chiral unsaturated á-amino acids. They have been prepared by chiral induction with auxiliaries as Sulfinylimino acetate or bislactime ether alkaloids has also been employed. Furthermore they have been synthesized by thioclaisen rearrangement, which further led to â-substituted-ã-ä unsaturated amino acids. Synthetic routes starting from boron derivatives through Petasis reaction and phosphor derivatives through Wittig reactions also proved their efficiency. Kinetic resolution using cinchona .
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