Martina Hrast | University of Ljubljana (original) (raw)
Papers by Martina Hrast
Pharmaceuticals, Nov 3, 2020
Drug discovery programs against the antibacterial target UDP-N-acetylglucosamine enolpyruvyl tran... more Drug discovery programs against the antibacterial target UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) have already resulted in covalent inhibitors having small threeand five-membered heterocyclic rings. In the current study, the reactivity of four-membered rings was carefully modulated to obtain a novel family of covalent MurA inhibitors. Screening a small library of cyclobutenone derivatives led to the identification of bromo-cyclobutenaminones as new electrophilic warheads. The electrophilic reactivity and cysteine specificity have been determined in a glutathione (GSH) and an oligopeptide assay, respectively. Investigating the structure-activity relationship for MurA suggests a crucial role for the bromine atom in the ligand. In addition, MS/MS experiments have proven the covalent labelling of MurA at Cys115 and the observed loss of the bromine atom suggests a net nucleophilic substitution as the covalent reaction. This new set of compounds might be considered as a viable chemical starting point for the discovery of new MurA inhibitors.
Antibiotics, Apr 15, 2021
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
PubMed, 2013
MurF is an essential bacterial enzyme that is involved in the last intracellular stage of peptido... more MurF is an essential bacterial enzyme that is involved in the last intracellular stage of peptidoglycan biosynthesis, and therefore it has the potential to be exploited as a target for the development of new antibacterials. Here, we report on the expression, purification and biochemical characterization of MurF from an important pathogen, Streptococcus pneumoniae. Additionally, ligand-based virtual screening was successfully used and a new hit compound with micromolar inhibitory activities against MurF enzymes from S. pneumoniae and Escherichia coli was identified.
ChemMedChem, Dec 11, 2019
Supporting information for this article is given via a link at the end of the document.
Tetrahedron, Aug 1, 2013
ABSTRACT The synthesis of U-shaped conformationally constrained analogues of peptides based on th... more ABSTRACT The synthesis of U-shaped conformationally constrained analogues of peptides based on the 3-amino-2-oxo-1,5-diazabicyclo[3.3.0]octane-8-carboxylic acid scaffold was developed. [3+2] Cycloadditions of (1Z,4R*,5R*)-1-arylmethylidene-4-benzyloxycarbonylamino-3-oxo-5-phenylpyrazolidin-1-ium-2-ides to tert-butyl acrylate and tert-butyl methacrylate gave the corresponding racemic cycloadducts, in most cases as mixtures of isomers, which were separated by preparative chromatography. Selective deprotection of the C- and the N-terminal of these heterocyclic dipeptides followed by coupling with (S)-alanine derivatives and chromatographic separation furnished the non-racemic tripeptides as target compounds. The structures of racemic cycloadducts and non-racemic final products were determined by NMR spectroscopy and X-ray diffraction. The synthesized compounds were also evaluated for inhibition of MurD ligase and d-alanine:d-alanine ligase.
Bioorganic Chemistry, Aug 1, 2014
The widespread emergence of resistant bacterial strains is becoming a serious threat to public he... more The widespread emergence of resistant bacterial strains is becoming a serious threat to public health. This thus signifies the need for the development of new antibacterial agents with novel mechanisms of action. Continuous efforts in the design of novel antibacterials remain one of the biggest challenges in drug development. In this respect, the Mur enzymes, MurA-F, that are involved in the formation of UDP-N-acetylmuramyl-pentapeptide can be genuinely considered as promising antibacterial targets. This review provides an in-depth insight into the recent developments in the field of inhibitors of the MurA-F enzymes. Special attention is also given to compounds that act as multiple inhibitors of two, three or more of the Mur enzymes. Moreover, the reasons for the lack of preclinically successful inhibitors and the challenges to overcome these hurdles in the next years are also debated.
ACS Medicinal Chemistry Letters, Nov 3, 2022
Thiazoles exhibit a wide range of biological activities and therefore represent useful and attrac... more Thiazoles exhibit a wide range of biological activities and therefore represent useful and attractive building blocks. To evaluate their usefulness and pinpoint their liabilities in fragment screening campaigns, we assembled a focused library of 49 fragment-sized thiazoles and thiadiazoles with various substituents, namely amines, bromides, carboxylic acids, and nitriles. The library was profiled in a cascade of biochemical inhibition assays, redox activity, thiol reactivity, and stability assays. Our study indicates that when thiazole derivatives are identified as screening hits, their reactivity should be carefully addressed and correlated with specific on-target engagement. Importantly, nonspecific inhibition should be excluded using experimental approaches and in silico predictions. To help with validation of hits identified in fragment screening campaigns, we can apply our highthroughput profiling workflow to focus on the most tractable compounds with a clear mechanism of action.
Acta Chimica Slovenica, Jun 15, 2022
Optimized conditions for the synthesis of fully deprotected (2R)-aziridine containing dipeptides ... more Optimized conditions for the synthesis of fully deprotected (2R)-aziridine containing dipeptides are described. Preparation of fully protected N-and C-terminal aziridine containing dipeptides was found to be straightforward and high yielding for the majority of compounds, whereas their full deprotection was possible only for C-terminal analogs. Deprotection of N-terminal derivatives using standard procedures of peptide chemistry was found difficult providing only mixtures of unidentifiable products. The described molecules have potential as building blocks in synthetic chemistry, in the chemical biology arena, as covalent modifiers, and as biomarkers.
Bioorganic Chemistry, Feb 1, 2022
The therapeutic indications for monoamine oxidases A and B (MAO-A and MAO-B) inhibitors that have... more The therapeutic indications for monoamine oxidases A and B (MAO-A and MAO-B) inhibitors that have emerged from biological studies on animal and cellular models of neurological and oncological diseases have focused drug discovery projects upon identifying reversible MAO inhibitors. Screening of our in-house academic compound library identified two hit compounds that inhibit MAO-B with IC50 values in micromolar range. Two series of indole (23 analogues) and 3-(benzyloxy)benzyl)piperazine (16 analogues) MAO-B inhibitors were derived from hits, and screened for their structure-activity relationships. Both series yielded low micromolar selective inhibitors of human MAO-B, namely indole 2 (IC50 = 12.63 ± 1.21 µM) and piperazine 39 (IC50 = 19.25 ± 4.89 µM), which is comparable to selective MAO-B inhibitor isatin (IC50 = 6.10 ± 2.81 µM), yet less potent in comparison to safinamide (IC50 = 0.029 ± 0.002 µM). Selective MAO-B inhibitors 2, 14, 38 and 39 exhibited favourable permeation of the blood-brain barrier and low cytotoxicity in the human neuroblastoma cell line SH-SY5Y.
Bioorganic & Medicinal Chemistry, Sep 1, 2011
D-Alanine:D-alanine ligase (Ddl), an intracellular bacterial enzyme essential for cell wall biosy... more D-Alanine:D-alanine ligase (Ddl), an intracellular bacterial enzyme essential for cell wall biosynthesis, is an attractive target for development of novel antimicrobial drugs. This study focused on an extensive evaluation of two families of Ddl inhibitors encountered in our previous research. New members of both families were obtained through similarity search and synthesis. Ellipticines and 9-acridinylamines were both found to possess inhibitory activity against Ddl from Escherichia coli and antimicrobial activity against E. coli and Staphylococcus aureus. Ellipticines with a quaternary methylpyridinium moiety were the most potent among all studied compounds, with MIC values as low as 2 mg/L in strains with intact efflux mechanisms. Antimicrobial activity of the studied compounds was connected to membrane damage, making their development as antibacterial drug candidates unlikely unless analogues devoid of this nonspecific effect can be discovered.
Pharmaceuticals, Nov 29, 2022
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Molecules, Apr 1, 2019
Despite the great importance of β-lactam antibiotics, there is still a limited number of syntheti... more Despite the great importance of β-lactam antibiotics, there is still a limited number of synthetic approaches for the formation of β-lactam-containing dipeptides. In this study, we report upon the stereoselective preparation of β-lactam-containing pseudopeptides, where different reaction conditions and NH 2 protective groups were tested to obtain compounds that contain 3-amino-azetidin-2-one. We demonstrate that the protective group is essential for the outcome of the reaction. Successful implementation of dibenzyl-protected serine-containing dipeptides through the Mitsunobu reaction can provide the desired products at high yields and stereoselectivity.
Bioorganic Chemistry, Dec 1, 2009
Enzymes involved in the biosynthesis of bacterial peptidoglycan represent important targets for d... more Enzymes involved in the biosynthesis of bacterial peptidoglycan represent important targets for development of new antibacterial drugs. Among them, Mur ligases (MurC to MurF) catalyze the formation of the final cytoplasmic precursor UDP-N-acetylmuramyl-pentapeptide from UDP-N-acetylmuramic acid. We present the design, synthesis and biological evaluation of a series of phosphorylated hydroxyethylamines as new type of small-molecule inhibitors of Mur ligases. We show that the phosphate group attached to the hydroxyl moiety of the hydroxyethylamine core is essential for good inhibitory activity. The IC 50 values of these inhibitors were in the micromolar range, which makes them a promising starting point for the development of multiple inhibitors of Mur ligases as potential antibacterial agents. In addition, 1-(4methoxyphenylsulfonamido)-3-morpholinopropan-2-yl dihydrogen phosphate 7a was discovered as one of the best inhibitors of MurE described so far.
Research Square (Research Square), Jul 18, 2023
MurC, D, E, and F are ATP-dependent ligases involved in the stepwise assembly of the tetrapeptide... more MurC, D, E, and F are ATP-dependent ligases involved in the stepwise assembly of the tetrapeptide stem of forming peptidoglycan. As highly conserved targets found exclusively in bacterial cells, they are of signi cant interest for antibacterial drug discovery. In this study, we employed a computer-aided molecular design approach to identify potential inhibitors of MurF. A biochemical inhibition assay was conducted, screening twenty-four avonoids and related compounds against MurC-F, resulting in the identi cation of quercitrin, myricetin, and (-)-epicatechin as MurF inhibitors with IC 50 values of 143 µM, 139 µM, and 92 µM, respectively. Notably, (-)-epicatechin demonstrated mixed type inhibition with ATP and uncompetitive inhibition with D-Ala-D-Ala dipeptide and UM3DAP substrates. Furthermore, in silico analysis using Sitemap and subsequent docking analysis using Glide revealed two plausible binding sites for (-)-epicatechin. The study also investigated the crucial structural features required for activity, with a particular focus on the substitution pattern and hydroxyl group positions, which were found to be important for the activity. The study highlights the signi cance of computational approaches in targeting essential enzymes involved in bacterial peptidoglycan synthesis.
European journal of medicinal chemistry, Mar 1, 2023
European journal of medicinal chemistry, May 1, 2020
This is a PDF file of an article that has undergone enhancements after acceptance, such as the ad... more This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Molecules, Jul 13, 2022
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Pharmaceuticals, Nov 3, 2020
Drug discovery programs against the antibacterial target UDP-N-acetylglucosamine enolpyruvyl tran... more Drug discovery programs against the antibacterial target UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) have already resulted in covalent inhibitors having small threeand five-membered heterocyclic rings. In the current study, the reactivity of four-membered rings was carefully modulated to obtain a novel family of covalent MurA inhibitors. Screening a small library of cyclobutenone derivatives led to the identification of bromo-cyclobutenaminones as new electrophilic warheads. The electrophilic reactivity and cysteine specificity have been determined in a glutathione (GSH) and an oligopeptide assay, respectively. Investigating the structure-activity relationship for MurA suggests a crucial role for the bromine atom in the ligand. In addition, MS/MS experiments have proven the covalent labelling of MurA at Cys115 and the observed loss of the bromine atom suggests a net nucleophilic substitution as the covalent reaction. This new set of compounds might be considered as a viable chemical starting point for the discovery of new MurA inhibitors.
Antibiotics, Apr 15, 2021
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
PubMed, 2013
MurF is an essential bacterial enzyme that is involved in the last intracellular stage of peptido... more MurF is an essential bacterial enzyme that is involved in the last intracellular stage of peptidoglycan biosynthesis, and therefore it has the potential to be exploited as a target for the development of new antibacterials. Here, we report on the expression, purification and biochemical characterization of MurF from an important pathogen, Streptococcus pneumoniae. Additionally, ligand-based virtual screening was successfully used and a new hit compound with micromolar inhibitory activities against MurF enzymes from S. pneumoniae and Escherichia coli was identified.
ChemMedChem, Dec 11, 2019
Supporting information for this article is given via a link at the end of the document.
Tetrahedron, Aug 1, 2013
ABSTRACT The synthesis of U-shaped conformationally constrained analogues of peptides based on th... more ABSTRACT The synthesis of U-shaped conformationally constrained analogues of peptides based on the 3-amino-2-oxo-1,5-diazabicyclo[3.3.0]octane-8-carboxylic acid scaffold was developed. [3+2] Cycloadditions of (1Z,4R*,5R*)-1-arylmethylidene-4-benzyloxycarbonylamino-3-oxo-5-phenylpyrazolidin-1-ium-2-ides to tert-butyl acrylate and tert-butyl methacrylate gave the corresponding racemic cycloadducts, in most cases as mixtures of isomers, which were separated by preparative chromatography. Selective deprotection of the C- and the N-terminal of these heterocyclic dipeptides followed by coupling with (S)-alanine derivatives and chromatographic separation furnished the non-racemic tripeptides as target compounds. The structures of racemic cycloadducts and non-racemic final products were determined by NMR spectroscopy and X-ray diffraction. The synthesized compounds were also evaluated for inhibition of MurD ligase and d-alanine:d-alanine ligase.
Bioorganic Chemistry, Aug 1, 2014
The widespread emergence of resistant bacterial strains is becoming a serious threat to public he... more The widespread emergence of resistant bacterial strains is becoming a serious threat to public health. This thus signifies the need for the development of new antibacterial agents with novel mechanisms of action. Continuous efforts in the design of novel antibacterials remain one of the biggest challenges in drug development. In this respect, the Mur enzymes, MurA-F, that are involved in the formation of UDP-N-acetylmuramyl-pentapeptide can be genuinely considered as promising antibacterial targets. This review provides an in-depth insight into the recent developments in the field of inhibitors of the MurA-F enzymes. Special attention is also given to compounds that act as multiple inhibitors of two, three or more of the Mur enzymes. Moreover, the reasons for the lack of preclinically successful inhibitors and the challenges to overcome these hurdles in the next years are also debated.
ACS Medicinal Chemistry Letters, Nov 3, 2022
Thiazoles exhibit a wide range of biological activities and therefore represent useful and attrac... more Thiazoles exhibit a wide range of biological activities and therefore represent useful and attractive building blocks. To evaluate their usefulness and pinpoint their liabilities in fragment screening campaigns, we assembled a focused library of 49 fragment-sized thiazoles and thiadiazoles with various substituents, namely amines, bromides, carboxylic acids, and nitriles. The library was profiled in a cascade of biochemical inhibition assays, redox activity, thiol reactivity, and stability assays. Our study indicates that when thiazole derivatives are identified as screening hits, their reactivity should be carefully addressed and correlated with specific on-target engagement. Importantly, nonspecific inhibition should be excluded using experimental approaches and in silico predictions. To help with validation of hits identified in fragment screening campaigns, we can apply our highthroughput profiling workflow to focus on the most tractable compounds with a clear mechanism of action.
Acta Chimica Slovenica, Jun 15, 2022
Optimized conditions for the synthesis of fully deprotected (2R)-aziridine containing dipeptides ... more Optimized conditions for the synthesis of fully deprotected (2R)-aziridine containing dipeptides are described. Preparation of fully protected N-and C-terminal aziridine containing dipeptides was found to be straightforward and high yielding for the majority of compounds, whereas their full deprotection was possible only for C-terminal analogs. Deprotection of N-terminal derivatives using standard procedures of peptide chemistry was found difficult providing only mixtures of unidentifiable products. The described molecules have potential as building blocks in synthetic chemistry, in the chemical biology arena, as covalent modifiers, and as biomarkers.
Bioorganic Chemistry, Feb 1, 2022
The therapeutic indications for monoamine oxidases A and B (MAO-A and MAO-B) inhibitors that have... more The therapeutic indications for monoamine oxidases A and B (MAO-A and MAO-B) inhibitors that have emerged from biological studies on animal and cellular models of neurological and oncological diseases have focused drug discovery projects upon identifying reversible MAO inhibitors. Screening of our in-house academic compound library identified two hit compounds that inhibit MAO-B with IC50 values in micromolar range. Two series of indole (23 analogues) and 3-(benzyloxy)benzyl)piperazine (16 analogues) MAO-B inhibitors were derived from hits, and screened for their structure-activity relationships. Both series yielded low micromolar selective inhibitors of human MAO-B, namely indole 2 (IC50 = 12.63 ± 1.21 µM) and piperazine 39 (IC50 = 19.25 ± 4.89 µM), which is comparable to selective MAO-B inhibitor isatin (IC50 = 6.10 ± 2.81 µM), yet less potent in comparison to safinamide (IC50 = 0.029 ± 0.002 µM). Selective MAO-B inhibitors 2, 14, 38 and 39 exhibited favourable permeation of the blood-brain barrier and low cytotoxicity in the human neuroblastoma cell line SH-SY5Y.
Bioorganic & Medicinal Chemistry, Sep 1, 2011
D-Alanine:D-alanine ligase (Ddl), an intracellular bacterial enzyme essential for cell wall biosy... more D-Alanine:D-alanine ligase (Ddl), an intracellular bacterial enzyme essential for cell wall biosynthesis, is an attractive target for development of novel antimicrobial drugs. This study focused on an extensive evaluation of two families of Ddl inhibitors encountered in our previous research. New members of both families were obtained through similarity search and synthesis. Ellipticines and 9-acridinylamines were both found to possess inhibitory activity against Ddl from Escherichia coli and antimicrobial activity against E. coli and Staphylococcus aureus. Ellipticines with a quaternary methylpyridinium moiety were the most potent among all studied compounds, with MIC values as low as 2 mg/L in strains with intact efflux mechanisms. Antimicrobial activity of the studied compounds was connected to membrane damage, making their development as antibacterial drug candidates unlikely unless analogues devoid of this nonspecific effect can be discovered.
Pharmaceuticals, Nov 29, 2022
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Molecules, Apr 1, 2019
Despite the great importance of β-lactam antibiotics, there is still a limited number of syntheti... more Despite the great importance of β-lactam antibiotics, there is still a limited number of synthetic approaches for the formation of β-lactam-containing dipeptides. In this study, we report upon the stereoselective preparation of β-lactam-containing pseudopeptides, where different reaction conditions and NH 2 protective groups were tested to obtain compounds that contain 3-amino-azetidin-2-one. We demonstrate that the protective group is essential for the outcome of the reaction. Successful implementation of dibenzyl-protected serine-containing dipeptides through the Mitsunobu reaction can provide the desired products at high yields and stereoselectivity.
Bioorganic Chemistry, Dec 1, 2009
Enzymes involved in the biosynthesis of bacterial peptidoglycan represent important targets for d... more Enzymes involved in the biosynthesis of bacterial peptidoglycan represent important targets for development of new antibacterial drugs. Among them, Mur ligases (MurC to MurF) catalyze the formation of the final cytoplasmic precursor UDP-N-acetylmuramyl-pentapeptide from UDP-N-acetylmuramic acid. We present the design, synthesis and biological evaluation of a series of phosphorylated hydroxyethylamines as new type of small-molecule inhibitors of Mur ligases. We show that the phosphate group attached to the hydroxyl moiety of the hydroxyethylamine core is essential for good inhibitory activity. The IC 50 values of these inhibitors were in the micromolar range, which makes them a promising starting point for the development of multiple inhibitors of Mur ligases as potential antibacterial agents. In addition, 1-(4methoxyphenylsulfonamido)-3-morpholinopropan-2-yl dihydrogen phosphate 7a was discovered as one of the best inhibitors of MurE described so far.
Research Square (Research Square), Jul 18, 2023
MurC, D, E, and F are ATP-dependent ligases involved in the stepwise assembly of the tetrapeptide... more MurC, D, E, and F are ATP-dependent ligases involved in the stepwise assembly of the tetrapeptide stem of forming peptidoglycan. As highly conserved targets found exclusively in bacterial cells, they are of signi cant interest for antibacterial drug discovery. In this study, we employed a computer-aided molecular design approach to identify potential inhibitors of MurF. A biochemical inhibition assay was conducted, screening twenty-four avonoids and related compounds against MurC-F, resulting in the identi cation of quercitrin, myricetin, and (-)-epicatechin as MurF inhibitors with IC 50 values of 143 µM, 139 µM, and 92 µM, respectively. Notably, (-)-epicatechin demonstrated mixed type inhibition with ATP and uncompetitive inhibition with D-Ala-D-Ala dipeptide and UM3DAP substrates. Furthermore, in silico analysis using Sitemap and subsequent docking analysis using Glide revealed two plausible binding sites for (-)-epicatechin. The study also investigated the crucial structural features required for activity, with a particular focus on the substitution pattern and hydroxyl group positions, which were found to be important for the activity. The study highlights the signi cance of computational approaches in targeting essential enzymes involved in bacterial peptidoglycan synthesis.
European journal of medicinal chemistry, Mar 1, 2023
European journal of medicinal chemistry, May 1, 2020
This is a PDF file of an article that has undergone enhancements after acceptance, such as the ad... more This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Molecules, Jul 13, 2022
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY