I02-Macromolecular Crystallography
I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
I24-Microfocus Macromolecular Crystallography
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Sitthivut
Charoensutthivarakul
,
Sherine E.
Thomas
,
Amy
Curran
,
Karen P.
Brown
,
Juan M.
Belardinelli
,
Andrew J.
Whitehouse
,
Marta
Acebron-Garcia-De-Eulate
,
Jaspar
Sangan
,
Subramanian G.
Gramani
,
Mary
Jackson
,
Vitor
Mendes
,
R. Andres
Floto
,
Tom L.
Blundell
,
Anthony G.
Coyne
,
Chris
Abell
Diamond Proposal Number(s):
[9537, 14043, 18548]
Abstract: Mycobacterium abscessus (Mab) has emerged as a challenging threat to individuals with cystic fibrosis. Infections caused by this pathogen are often impossible to treat due to the intrinsic antibiotic resistance leading to lung malfunction and eventually death. Therefore, there is an urgent need to develop new drugs against novel targets in Mab to overcome drug resistance and subsequent treatment failure. In this study, SAICAR synthetase (PurC) from Mab was identified as a promising target for novel antibiotics. An in-house fragment library screen and a high-throughput X-ray crystallographic screen of diverse fragment libraries were explored to provide crucial starting points for fragment elaboration. A series of compounds developed from fragment growing and merging strategies, guided by crystallographic information and careful hit-to-lead optimization, have achieved potent nanomolar binding affinity against the enzyme. Some compounds also show a promising inhibitory effect against Mab and Mtb. This work utilizes a fragment-based design and demonstrates for the first time the potential to develop inhibitors against PurC from Mab.
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Jan 2022
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
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Philip
Hinchliffe
,
Diego M.
Moreno
,
Maria-Agustina
Rossi
,
Maria F.
Mojica
,
Veronica
Martinez
,
Valentina
Villamil
,
Brad
Spellberg
,
George L.
Drusano
,
Claudia
Banchio
,
Graciela
Mahler
,
Robert A.
Bonomo
,
Alejandro J.
Vila
,
James
Spencer
Diamond Proposal Number(s):
[12342, 17212]
Abstract: Metallo-β-lactamase (MBL) production in Gram-negative bacteria is an important contributor to β-lactam antibiotic resistance. Combining β-lactams with β-lactamase inhibitors (BLIs) is a validated route to overcoming resistance, but MBL inhibitors are not available in the clinic. On the basis of zinc utilization and sequence, MBLs are divided into three subclasses, B1, B2, and B3, whose differing active-site architectures hinder development of BLIs capable of “cross-class” MBL inhibition. We previously described 2-mercaptomethyl thiazolidines (MMTZs) as B1 MBL inhibitors (e.g., NDM-1) and here show that inhibition extends to the clinically relevant B2 (Sfh-I) and B3 (L1) enzymes. MMTZs inhibit purified MBLs in vitro (e.g., Sfh-I, Ki 0.16 μM) and potentiate β-lactam activity against producer strains. X-ray crystallography reveals that inhibition involves direct interaction of the MMTZ thiol with the mono- or dizinc centers of Sfh-I/L1, respectively. This is further enhanced by sulfur-π interactions with a conserved active site tryptophan. Computational studies reveal that the stereochemistry at chiral centers is critical, showing less potent MMTZ stereoisomers (up to 800-fold) as unable to replicate sulfur-π interactions in Sfh-I, largely through steric constraints in a compact active site. Furthermore, in silico replacement of the thiazolidine sulfur with oxygen (forming an oxazolidine) resulted in less favorable aromatic interactions with B2 MBLs, though the effect is less than that previously observed for the subclass B1 enzyme NDM-1. In the B3 enzyme L1, these effects are offset by additional MMTZ interactions with the protein main chain. MMTZs can therefore inhibit all MBL classes by maintaining conserved binding modes through different routes.
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Aug 2021
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I03-Macromolecular Crystallography
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Diamond Proposal Number(s):
[11088]
Abstract: Chagas disease, an infectious condition caused by Trypanosoma cruzi, lacks treatment with drugs with desired efficacy and safety profiles. To address this unmet medical need, a set of trypanocidal compounds were identified through a large multicenter phenotypic-screening initiative and assembled in the GSK Chagas Box. In the present work, we report the screening of the Chagas Box against T. cruzi malic enzymes (MEs) and the identification of three potent inhibitors of its cytosolic isoform (TcMEc). One of these compounds, TCMDC-143108 (1), came out as a nanomolar inhibitor of TcMEc, and 14 new derivatives were synthesized and tested for target inhibition and efficacy against the parasite. Moreover, we determined the crystallographic structures of TcMEc in complex with TCMDC-143108 (1) and six derivatives, revealing the allosteric inhibition site and the determinants of specificity. Our findings connect phenotypic hits from the Chagas Box to a relevant metabolic target in the parasite, providing data to foster new structure–activity guided hit optimization initiatives.
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Jul 2021
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I24-Microfocus Macromolecular Crystallography
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Diamond Proposal Number(s):
[12346]
Abstract: Metallo-β-lactamases (MBLs) can efficiently catalyze the hydrolysis of all classes of β-lactam antibiotics except monobactams. While serine-β-lactamase (SBL) inhibitors (e.g., clavulanic acid, avibactam) are established for clinical use, no such MBL inhibitors are available. We report on the synthesis and mechanism of inhibition of N-sulfamoylpyrrole-2-carboxylates (NSPCs) which are potent inhibitors of clinically relevant B1 subclass MBLs, including NDM-1. Crystallography reveals that the N-sulfamoyl NH2 group displaces the dizinc bridging hydroxide/water of the B1 MBLs. Comparison of crystal structures of an NSPC and taniborbactam (VRNX-5133), presently in Phase III clinical trials, shows similar binding modes for the NSPC and the cyclic boronate ring systems. The presence of an NSPC restores meropenem efficacy in clinically derived E. coli and K. pneumoniae blaNDM-1. The results support the potential of NSPCs and related compounds as efficient MBL inhibitors, though further optimization is required for their clinical development.
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May 2021
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I04-Macromolecular Crystallography
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Abstract: Here we have described a systematic structure activity relationship (SAR) of a set of compounds inspired from cladosporin, a tool compound that targets parasite (Plasmodium falciparum) lysyl tRNA synthetase (KRS). Four sets of analogues, synthesized based on point changes in the chemical scaffold of cladosporin and other logical modifications and hybridizations, were assessed using high throughput enzymatic and parasitic assays along with in vitro pharmacokinetics. Co-crystallization of the most potent compound in our series (CL-2) with PfKRS revealed its structural basis of enzymatic binding and potency. Further, we report that CL-2 has performed better than cladosporin in terms of metabolic stability. It thus represents a new lead for further optimization toward the development of antimalarial drugs. Collectively, along with a lead compound, the series offers insights on how even the slightest chemical modification might play an important role in enhancing or decreasing the potency of a chemical scaffold.
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Apr 2021
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I03-Macromolecular Crystallography
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Diamond Proposal Number(s):
[13775]
Open Access
Abstract: Integrase strand transfer inhibitors (INSTIs) block the integration step of the retroviral lifecycle and are first-line drugs used for the treatment of HIV-1/AIDS. INSTIs have a polycyclic core with heteroatom triads, chelate the metal ions at the active site, and have a halobenzyl group that interacts with viral DNA attached to the core by a flexible linker. The most broadly effective INSTIs inhibit both wild-type (WT) integrase (IN) and a variety of well-known mutants. However, because there are mutations that reduce the potency of all of the available INSTIs, new and better compounds are needed. Models based on recent structures of HIV-1 and red-capped mangabey SIV INs suggest modifications in the INSTI structures that could enhance interactions with the 3′-terminal adenosine of the viral DNA, which could improve performance against INSTI resistant mutants. We designed and tested a series of INSTIs having modifications to their naphthyridine scaffold. One of the new compounds retained good potency against an expanded panel of HIV-1 IN mutants that we tested. Our results suggest the possibility of designing inhibitors that combine the best features of the existing compounds, which could provide additional efficacy against known HIV-1 IN mutants.
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Mar 2021
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I04-Macromolecular Crystallography
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David T.
Davies
,
Simon
Leiris
,
Nicolas
Sprynski
,
Jérôme
Castandet
,
Clarisse
Lozano
,
Justine
Bousquet
,
Magdalena
Zalacain
,
Srinivas
Vasa
,
Praveen K.
Dasari
,
Ramesh
Pattipati
,
Naresh
Vempala
,
Swetha
Gujjewar
,
Syamkumar
Godi
,
Raju
Jallala
,
Rajashekar Reddy
Sathyap
,
Narasimha A.
Darshanoju
,
Vengala R.
Ravu
,
Ramakrishna R.
Juventhala
,
Narender
Pottabathini
,
Somesh
Sharma
,
Srinivasu
Pothukanuri
,
Kirsty
Holden
,
Peter
Warn
,
Francesca
Marcoccia
,
Manuela
Benvenuti
,
Cecilia
Pozzi
,
Stefano
Mangani
,
Jean-Denis
Docquier
,
Marc
Lemonnier
,
Martin
Everett
Diamond Proposal Number(s):
[21741]
Abstract: The clinical effectiveness of the important β-lactam class of antibiotics is under threat by the emergence of resistance, mostly due to the production of acquired serine- (SBL) and metallo-β-lactamase (MBL) enzymes. To address this resistance issue, multiple β-lactam/β-lactamase inhibitor combinations have been successfully introduced into the clinic over the past several decades. However, all of those combinations contain SBL inhibitors and, as yet, there are no MBL inhibitors in clinical use. Consequently, there exists an unaddressed yet growing healthcare problem due to the rise in recent years of highly resistant strains which produce New Delhi metallo (NDM)-type metallo-carbapenemases. Previously, we reported the characterization of an advanced MBL inhibitor lead compound, ANT431. Herein, we discuss the completion of a lead optimization campaign culminating in the discovery of the preclinical candidate ANT2681, a potent NDM inhibitor with strong potential for clinical development.
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Aug 2020
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I03-Macromolecular Crystallography
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João A.
Ribeiro
,
Alexander
Hammer
,
Gerardo A.
Libreros-Zúñiga
,
Sair M.
Chavez-Pacheco
,
Petros
Tyrakis
,
Gabriel S.
De Oliveira
,
Timothy
Kirkman
,
Jamal
El Bakali
,
Silvana A.
Rocco
,
Mauricio L.
Sforça
,
Roberto
Parise-Filho
,
Anthony G.
Coyne
,
Tom L.
Blundell
,
Chris
Abell
,
Marcio V. B.
Dias
Abstract: Dihydrofolate reductase (DHFR), a key enzyme involved in folate metabolism, is a widely explored target in the treatment of cancer, immune diseases, bacteria, and protozoa infections. Although several antifolates have proved successful in the treatment of infectious diseases, they have been underexplored to combat tuberculosis, despite the essentiality of M. tuberculosis DHFR (MtDHFR). Herein, we describe an integrated fragment-based drug discovery approach to target MtDHFR that has identified hits with scaffolds not yet explored in any previous drug design campaign for this enzyme. The application of a SAR by catalog strategy of an in house library for one of the identified fragments has led to a series of molecules that bind to MtDHFR with low micromolar affinities. Crystal structures of MtDHFR in complex with compounds of this series demonstrated a novel binding mode that considerably differs from other DHFR antifolates, thus opening perspectives for the development of relevant MtDHFR inhibitors.
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Jul 2020
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I03-Macromolecular Crystallography
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Leah S.
Torrie
,
David A.
Robinson
,
Michael G.
Thomas
,
Judith V.
Hobrath
,
Sharon M
Shepherd
,
John M.
Post
,
Eun-Jung
Ko
,
Rafael Augusto
Alves Ferreira
,
Claire J.
Mackenzie
,
Karolina
Wrobel
,
Darren
Edwards
,
Ian H.
Gilbert
,
David W.
Gray
,
Alan H.
Fairlamb
,
Manu
De Rycker
Diamond Proposal Number(s):
[14980]
Open Access
Abstract: Methionyl-tRNA synthetase (MetRS) is a chemically validated drug target in the kinetoplastid parasites Trypanosoma brucei and Leishmania donovani. To date, all kinetoplastid MetRS inhibitors described bind in a similar way to an expanded methionine pocket and an adjacent, auxiliary pocket. In the current study we have identified a structurally novel class of inhibitors containing a 4,6-diamino substituted pyrazolopyrimidine core (‘MetRS02’ series). Crystallographic studies revealed that ‘MetRS02’ compounds bind to an allosteric pocket in L. major MetRS not previously described and enzymatic studies demonstrated a non-competitive mode of inhibition. Homology modelling of the Trypanosoma cruzi MetRS enzyme revealed key differences in the allosteric pocket between the T. cruzi and Leishmania enzymes. These provide a likely explanation for the lower ‘MetRS02’ potencies that we observed for the T. cruzi enzyme compared to the Leishmania enzyme. The identification of a new series of MetRS inhibitors and the discovery of a new binding site in kinetoplastid MetRS enzymes provides a novel strategy in the search for new therapeutics for kinetoplastid diseases.
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Apr 2020
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I04-Macromolecular Crystallography
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Fraser
Cunningham
,
Jorge
Esquivias
,
Raquel
Fernández-Menéndez
,
Arancha
Pérez
,
Ana
Guardia
,
Jaime
Escribano
,
Cristina
Rivero
,
Mythily
Vimal
,
Mónica
Cacho
,
Paco
De Dios-Antón
,
María Santos
Martínez-Martínez
,
Elena
Jiménez
,
Leticia
Huertas Valentín
,
María José
Rebollo-López
,
Eva María
López-Román
,
Verónica
Sousa-Morcuende
,
Joaquín
Rullas
,
Margaret
Neu
,
Chun-Wa
Chung
,
Robert H.
Bates
Diamond Proposal Number(s):
[5799]
Abstract: In the course of optimizing a novel indazole sulfonamide series that inhibits β-ketoacyl-ACP synthase (KasA) of Mycobacterium tuberculosis, a mutagenic aniline metabolite was identified. Further lead optimization efforts were therefore dedicated to eliminating this critical liability by removing the embedded aniline moiety or modifying its steric or electronic environment. While the narrow SAR space against the target ultimately rendered this goal unsuccessful, key structural knowledge around the binding site of this underexplored target for TB was generated to inform future discovery efforts.
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Mar 2020
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