Potent β-Lactam Inhibitors of Human Cytomegalovirus Protease

Yoakim, Christiane; Ogilvie, William W.; Cameron, Dale R.; Chabot, Catherine; Grand‐Maître, Chantal; Guse, Ingrid; Haché, Bruno; Kawai, Stephen H.; Naud, Julie; O'Meara, Jeff A.; Plante, Raymond; Déziel, Robert

doi:10.1177/095632029800900502

Cited by 35 publications

(24 citation statements)

References 29 publications

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“…Since yeast encodes a pleiotrophic drug resistance network with a variety of ABC transporters that efficiently clear small molecules within the cell, we sensitized the strain by deleting the three major drug efflux pumps Pdr5p, Snq2p, and Yor1p. In such a setup, the yeast system shows a sensitivity towards small molecules similar to that shown in a mammalian cellular assay, at least for the ␤-lactam inhibitors tested, with a calculated EC 50 of 31 M in the yeast assay versus 78 M in a mammalian plaque reduction assay (42).…”

Section: Discussionmentioning

confidence: 99%

“…In order to assess sensitivity of the selection system towards small molecules, the two validated HCMV protease inhibitors BI31 and BI36 of Boehringer Ingelheim were applied (42) Figure 5A shows that increasing concentrations of both BI31 and BI36 in cells expressing the active protease correlate with increasing cell proliferation. The calculated EC 50 of 31 M for BI36 suggests that the yeast-based assay exhibits the same sensitivity as that of a mammalian cellular assay, at least for this class of compounds (42). At BI36 concentrations of Ͼ100 M, HCMV protease was strongly inhibited, as evidenced by proliferation rates similar to those of cells expressing the inactive protease.…”

Section: Modulation Of Hcmv Protease Expression Level Correlates Withmentioning

confidence: 99%

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Novel Yeast Cell-Based Assay To Screen for Inhibitors of Human Cytomegalovirus Protease in a High-Throughput Format

Cottier¹,

Barberis²,

Lüthi³

2006

Antimicrob Agents Chemother

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The protease encoded by the human cytomegalovirus (HCMV) is an attractive target for antiviral drug development because of its essential function in viral replication. We describe here a cellular assay in the yeast Saccharomyces cerevisiae for the identification of small molecule inhibitors of HCMV protease by conditional growth in selective medium. In this system, the protease cleavage sequence is inserted into the N-(5-phosphoribosyl)anthranilate isomerase (Trp1p), a yeast protein essential for cell proliferation in the absence of tryptophan. Coexpression of HCMV protease with the engineered Trp1p substrate in yeast cells results in site-specific cleavage and functional inactivation of the Trp1p enzyme, thereby leading to an arrest of cell proliferation. This growth arrest can be suppressed by the addition of validated HCMV protease inhibitors. The growth selection system presented here provides the basis for a high-throughput screen to identify HCMV protease inhibitors that are active in eukaryotic cells.Herpesviruses are widely present in nature and afflict many species throughout the animal kingdom (14). The most frequent human infection is caused by the human cytomegalovirus (HCMV), affecting up to 80% of the general population. This highly prevalent member of the herpesvirus family is responsible for opportunistic infections in immunocompromised individuals, notably AIDS patients and organ transplant recipients (for reviews, see references 9 and 29). Antiviral agents currently licensed for the treatment of HCMV infections include ganciclovir and its orally bioavailable prodrug valganciclovir as well as foscarnet, cidofovir, and fomivirsen (9). All these drugs are nucleoside analogues that ultimately target, either directly or indirectly, the viral DNA polymerase. The only exception is fomivirsen, an antisense oligonucleotide approved for HCMV retinitis that blocks translation of HCMV immediate-early mRNA. Unfortunately for the patients, the clinical usefulness of these drugs is limited: they exhibit toxic side effects, including bone marrow toxicity and nephrotoxicity, and they need to be injected either intravenously or intraocularly (9). Moreover, HCMV strains with reduced susceptibility resulting from chronic antiviral treatment are becoming more and more frequent. Thus, improved alternative drugs with novel mechanisms of action are needed for treating HCMV infections.Herpesviruses encode a serine protease that cleaves the assembly protein precursor, a major component of the intermediate capsid (reviewed in reference 23). Studies with herpes simplex virus type 1 protease, a close homologue of HCMV protease, showed that viral DNA cannot be packaged into virions without this cleavage, resulting in an empty nucleocapsid (17, 32). HCMV protease is expressed as a 708-amino-acid precursor encoded by the UL80 open reading frame. Autocatalytic cleavage is observed at two consensus sequences called maturational (M) and release (R) sites (23). M-site cleavage removes a 6-kDa C-terminal tail, which mediates inter...

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Section: Discussionmentioning

confidence: 99%

Section: Modulation Of Hcmv Protease Expression Level Correlates Withmentioning

confidence: 99%

Novel Yeast Cell-Based Assay To Screen for Inhibitors of Human Cytomegalovirus Protease in a High-Throughput Format

Cottier¹,

Barberis²,

Lüthi³

2006

Antimicrob Agents Chemother

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“…These compounds inhibit HCMV protease via the opening of the β-lactam ring followed by acyl enzyme complex formation. Among them, N-methylthiotetrazole-containing compounds were found to be most potent inhibitors in enzymatic assays [115]. Subsequently, Ogilvie et al reported only modest antiviral activity of monobactam inhibitors, despite the fact that many compounds were very effective in enzymatic assays [116].…”

Section: H Current Activity In Drug Discoverymentioning

confidence: 98%

Impact of Recombinant DNA Technology and Protein Engineering on Structure-Based Drug Design: Case Studies of HIV-1 and HCMV Proteases

Kan

2002

CTMC

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Structure-based drug design is an organized, multidisciplinary endeavor undertaken by scientists from many different scientific fields. The success of structure-based drug design was only made possible by advances in structure biology that provides the three-dimensional structure of the drug design target with which small molecular chemical ligands interact. Visualization of the conformation and interactions of a small molecule ligand bound to the protein target in the co-crystal structure of the protein:ligand complex enables the design of new chemical compounds with improved binding affinity and specificity. With the advances in molecular biology, lab automation, and computational science, genomic data have now become available for the human genome, as well as various other organisms. The pharmaceutical industry is currently putting forth tremendous effort in the area of functional genomics and structural genomics in attempts to decipher functions and structures of protein encoded by genes, with the ultimate goal of identifying novel targets for drug discovery and development. This chapter discusses the significant impact made by recombinant DNA technology and protein engineering on structural biology and, more specifically, on structure-based drug design.

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“…The penicillin with the β-lactam ring has been reported earlier as antiviral protease inhibitors 48,49 , but they were not explored further as better antiviral agents were existing. Also, like N3 inhibitor, there were several molecules reported being covalent inhibitors of M pro for SARS-CoV and MERS-CoV.…”

Section: Covalent Dockingmentioning

confidence: 99%

In Silico Drug Repurposing of Penicillins to Target Main Protease M^proof SARS-CoV-2

et al. 2020

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Background: SARS coronavirus-2 (SARS-CoV-2) infection causes Novel Coronavirus Disease . It is a respiratory tract infection and currently becoming pandemic worldwide affecting more than 50 lakh people. As of now, there is no treatment or vaccine developed for disease management. The main protease, M pro in SARS-CoV-2 is a druggable target explored by many scientists. We targeted this with the well-known approach of drug repurposing by using computational tools. Methods: Schrödinger software was used for the study. Ligands were prepared from US-FDA drug-bank by importing it to Maestro graphical user interphase, optimised using LigPrep, and molecular geometry minimized using OPLS3e force-field. M pro crystal structure 6LU7 was downloaded from PDB and optimised. Molecules were docked using CovDock module in Glide docking. Further, molecular dynamics simulations were carried out for 100 ns using Desmond module. Results: In docking and molecular interactions studies, penicillins emerged as hits with consistent binding pattern by forming hydrophilic, hydrophobic, electrostatic interactions. The molecular dynamics simulations confirmed the interactions. Phenoxymethylpenicillin and Carbenicillin were found to interact consistently and appeared to be the most promising. Conclusion: Usually, antibiotics are discouraged from using in the viral pandemic because of the development of resistance. Azithromycin was combined with hydroxychloroquine to treat COVID-19. Penicillins are less potent and first-line antibiotics for most of the bacterial infections. This study suggests Phenoxymethylpenicillin and Carbenicillin can be tried along with hydroxychloroquine. Further, this study shows the possible exploration by drug repurposing using computer-aided docking tools and the potential roles of beta-lactams in COVID-19.

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Potent β-Lactam Inhibitors of Human Cytomegalovirus Protease

Cited by 35 publications

References 29 publications

Novel Yeast Cell-Based Assay To Screen for Inhibitors of Human Cytomegalovirus Protease in a High-Throughput Format

Novel Yeast Cell-Based Assay To Screen for Inhibitors of Human Cytomegalovirus Protease in a High-Throughput Format

Impact of Recombinant DNA Technology and Protein Engineering on Structure-Based Drug Design: Case Studies of HIV-1 and HCMV Proteases

In Silico Drug Repurposing of Penicillins to Target Main Protease M^proof SARS-CoV-2

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Potent β-Lactam Inhibitors of Human Cytomegalovirus Protease

Cited by 35 publications

References 29 publications

Novel Yeast Cell-Based Assay To Screen for Inhibitors of Human Cytomegalovirus Protease in a High-Throughput Format

Novel Yeast Cell-Based Assay To Screen for Inhibitors of Human Cytomegalovirus Protease in a High-Throughput Format

Impact of Recombinant DNA Technology and Protein Engineering on Structure-Based Drug Design: Case Studies of HIV-1 and HCMV Proteases

In Silico Drug Repurposing of Penicillins to Target Main Protease Mpro of SARS-CoV-2

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In Silico Drug Repurposing of Penicillins to Target Main Protease M^proof SARS-CoV-2