Biology subjects

Lieske, J.

Publications and source records attributed to Lieske, J..

5 recordsLinked to original sources

bioRxiv2023

Structural elucidation and antiviral activity of cathepsin L inhibitors with carbonyl and epoxide warheads

Emerging RNA viruses including SARS-CoV-2 continue to be a major threat around the globe. The cell entry of SARS-CoV-2 particles via the endosomal pathway involves the cysteine protease cathepsin L (CatL) among other proteases. CatL is rendered as a promising drug target in the context of different viral and lysosome-related diseases. Hence, drug discovery and structure-based optimization of inhibitors is of high pharmaceutical interest. We herein verified and compared the anti-SARS-CoV-2 activity of a set of carbonyl and succinyl-epoxide-based inhibitors, which have previously been identified as cathepsin inhibitors. Calpain inhibitor XII (CI-XII), MG-101 and CatL inhibitor IV (CLI-IV) possess antiviral activity in the very low nanomolar IC50 range in Vero E6 cells. Experimental structural data on how these and related compounds bind to CatL are however notably lacking, despite their therapeutic potential. Consequently, we present and compare crystal structures of CatL in complex with 14 compounds, namely BOCA (N-BOC-2-aminoacetaldehyde), CLI-IV, CI-III, CI-VI, CI-XII, the main protease -ketoamide inhibitor 13b, MG-101, MG-132 as well as E-64d (aloxistatin), E-64, CLIK148, CAA0225, TC-I (CID 16725315) and TPCK at resolutions better than 2 [A]. Overall, the presented data comprise a broad and solid basis for structure-guided understanding and optimization of CatL inhibitors towards protease drug development.

biochemistry↗

bioRxiv2021

SARS-CoV-2 papain-like protease PLpro in complex with natural compounds reveal allosteric sites for antiviral drug design

SARS-CoV-2 papain-like protease (PLpro) covers multiple functions. Beside the cysteine-protease activity, PLpro has the additional and vital function of removing ubiquitin and ISG15 (Interferon-stimulated gene 15) from host-cell proteins to aid coronaviruses in evading the hosts innate immune responses. We established a high-throughput X-ray screening to identify inhibitors by elucidating the native PLpro structure refined to 1.42 [A] and performing co-crystallization utilizing a diverse library of selected natural compounds. We identified three phenolic compounds as potential inhibitors. Crystal structures of PLpro inhibitor complexes, obtained to resolutions between 1.7-1.9 [A], show that all three compounds bind at the ISG15/Ub-S2 allosteric binding site, preventing the essential ISG15-PLpro molecular interactions. All compounds demonstrate clear inhibition in a deISGylation assay, two exhibit distinct antiviral activity and one inhibited a cytopathic effect in a non-cytotoxic concentration range. These results highlight the druggability of the rarely explored ISG15/Ub-S2 PLpro allosteric binding site to identify new and effective antiviral compounds. Importantly, in the context of increasing PLpro mutations in the evolving new variants of SARS-CoV-2, the natural compounds we identified may also reinstate the antiviral immune response processes of the host that are down-regulated in COVID-19 infections.

biochemistry↗

bioRxiv2020

Massive X-ray screening reveals two allosteric drug binding sites of SARS-CoV-2 main protease

The coronavirus disease (COVID-19) caused by SARS-CoV-2 is creating tremendous health problems and economical challenges for mankind. To date, no effective drug is available to directly treat the disease and prevent virus spreading. In a search for a drug against COVID-19, we have performed a massive X-ray crystallographic screen of two repurposing drug libraries against the SARS-CoV-2 main protease (Mpro), which is essential for the virus replication and, thus, a potent drug target. In contrast to commonly applied X-ray fragment screening experiments with molecules of low complexity, our screen tested already approved drugs and drugs in clinical trials. From the three-dimensional protein structures, we identified 37 compounds binding to Mpro. In subsequent cell-based viral reduction assays, one peptidomimetic and five non-peptidic compounds showed antiviral activity at non-toxic concentrations. We identified two allosteric binding sites representing attractive targets for drug development against SARS-CoV-2.

biophysics↗

bioRxiv2020

Catalytic cleavage of HEAT and subsequent covalent binding of the tetralone moiety by the SARS-CoV-2 main protease

Here we present the crystal structure of SARS-CoV-2 main protease (Mpro) covalently bound to 2-methyl-1-tetralone. This complex was obtained by co-crystallization of Mpro with HEAT (2-(((4-hydroxyphenethyl)amino)methyl)-3,4-dihydronaphthalen-1(2H)-one) in the framework of a large X-ray crystallographic screening project of Mpro against a drug repurposing library, consisting of 5632 approved drugs or compounds in clinical phase trials. Further investigations showed that HEAT is cleaved by Mpro in an E1cB-like reaction mechanism into 2-methylene-1-tetralone and tyramine. The catalytic Cys145 subsequently binds covalently in a Michael addition to the methylene carbon atom of 2-methylene-1-tetralone. According to this postulated model HEAT is acting in a pro-drug-like fashion. It is metabolized by Mpro, followed by covalent binding of one metabolite to the active site. The structure of the covalent adduct elucidated in this study opens up a new path for developing non-peptidic inhibitors.

biophysics↗

bioRxiv2019

Deep phenotyping of a healthy human HAO1 knockout informs therapeutic development for primary hyperoxaluria type 1.

Primary Hyperoxaluria Type 1 (PH1) is a rare autosomal recessive metabolic disorder of oxalate metabolism leading to kidney failure as well as multi-organ damage. Overproduction of oxalate occurs in the liver due to an inherited genetic defect in the enzyme alanine-glyoxylate aminotransferase (AGXT), causing pathology due to the insolubility of calcium oxalate crystals in body fluids. The main current therapy is dual liver-kidney transplant, which incurs high morbidity and has poor availability in some health systems where PH1 is more prevalent. One approach currently in active clinical investigation targets HAO1 (hydroxyacid oxidase 1), encoding glycolate oxidase, to reduce substrate levels for oxalate production. To inform drug development, we sought individuals with reduced HAO1 function due to naturally occurring genetic variation.\n\nAnalysis of loss of function variants in 141,456 sequenced individuals suggested individuals with complete HAO1 knockout would only be observed in 1 in 30 million outbred people. However in a large sequencing and health records program (Genes & Health), in populations with substantial autozygosity, we identified a healthy adult individual predicted to have complete knockout of HAO1 due to an ultra rare homozygous frameshift variant (rs1186715161, ENSP00000368066.3:p.Leu333SerfsTer4). Primary care and hospital health records confirmed no apparently related clinical phenotype. At recall, urine and plasma oxalate levels were normal, however plasma glycolate levels (171 nmol/mL) were 12 times the upper limit of normal in healthy, reference individuals (mean+2sd=14 nmol/mL, n=67) while her urinary glycolate levels were 6 times the upper limit of normal. Comparison with preclinical and phase 1 clinical trial data of an RNAi therapeutic targeting HAO1 (lumasiran) suggests the individual likely retains <2% residual glycolate oxidase activity.\n\nThese results provide important data to support the safety of HAO1 inhibition as a potential chronic therapy for a devastating metabolic disease (PH1). We also suggest that the effect of glycolate oxidase suppression in any potential other roles in humans beyond glycolate oxidation do not lead to clinical phenotypes, at least in this specific individual. This demonstrates the value of studying the lifelong complete knockdown of a target protein in a living human to aid development of a potential therapeutic, both in de-risking the approach and providing potential hypotheses to optimize its development. Furthermore, therapy for PH1 is likely to be required lifelong, in contrast to data from chronicity studies in non-human species or relatively short-term therapeutic studies in people. Our approach demonstrates the potential for improved drug discovery through unlocking relevant evidence hiding in the diversity of human genetic variation.

genetics↗