A screen for modulators of large T antigen’s ATPase activity uncovers novel inhibitors of Simian Virus 40 and BK virus replication

Seguin, Sandlin P.; Ireland, Alex W.; Gupta, Tushar; Wright, Colin M.; Miyata, Yoshinari; Wipf, Peter; Pipas, James M.; Gestwicki, Jason E.; Brodsky, Jeffrey L.

doi:10.1016/j.antiviral.2012.07.012

Cited by 17 publications

(27 citation statements)

References 66 publications

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“…Four inhibitors were tested: MAL2-11B, hexachlorophene, bithionol and MAL2-11B tetrazole. Hexachlorophene and bithionol were identified as SV40 LT ATPase inhibitors in a high throughput screen that sought small molecule inhibitors of SV40 LT using a colorimetric reporter assay [36,37]. In contrast, MAL2-11B is a derivative of a chaperone modulator that similarly inhibited SV40 LT ATPase activity but also abrogated the propagation of SV40 as well as BKV in infected mammalian cells [38].…”

Section: Resultsmentioning

confidence: 99%

“…Several inhibitors, including MAL2-11B, bithionol, hexachlorophene and MAL2-11B tetrazole were identified by our ongoing analyses of structure–activity relationships and via a high throughput screen using SV40 LT [36,37,39,44]. MAL2-11B and the tetrazole inhibited the ATPase activity of SV40 LT and inhibited SV40 and BKV DNA replication in cell culture.…”

Section: Discussionmentioning

confidence: 99%

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The conserved core enzymatic activities and the distinct dynamics of polyomavirus large T antigens

Brodsky

Pipas

2015

Archives of Biochemistry and Biophysics

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Several human polyomaviruses including JCV, BKV and TSV are associated with diseases, particularly in immunosuppressed patients. While the large T antigen (LT) encoded by the monkey polyomavirus SV40 is well studied, and possesses intrinsic ATPase and DNA helicase activities, the LTs of the human polyomaviruses are relatively uncharacterized. In order to evaluate whether these enzymatic activities, which are required for viral DNA replication, are conserved between polyomaviruses, we performed a comparative study using the LTs from JCV, TSV and SV40. The ATPase and DNA helicase activities and the interaction with the cellular tumor suppressor p53 were assayed for the purified Zn-ATPase domains of the three LTs. We found that all Zn-ATPases were active ATPases. The Zn-ATPase domains also functioned as DNA helicases, although the measured kinetic constants differed among the three proteins. In addition, when tested against four small molecule ATPase inhibitors, the Zn-ATPase domains of TSV was more resistant than that of SV40 and JCV. Our results show that, while LTs from JCV and TSV share the core ATPase and DNA helicase activities, they possess important functional differences that might translate into their respective abilities to infect and replicate in hosts.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

The conserved core enzymatic activities and the distinct dynamics of polyomavirus large T antigens

Brodsky

Pipas

2015

Archives of Biochemistry and Biophysics

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“…Activity against both polyomavirus BK and SV40 was evaluated using recombinant SV40 protein. SV40 LTA ATPase inhibition >20–35% was observed in 46 compounds that included flavones, polyphenols and bisphenols, but only two compounds, bithionol and hexachlorophene, were evaluated in detail (Seguin et al, 2012b). Both compounds were more cytotoxic than our LDN series of compounds: the MTS assay generated IC 50 values of 68 and 55 μM respectively.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of large T antigen ATPase activity as a potential strategy to develop anti-polyomavirus JC drugs

et al. 2014

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Introduction This study evaluates polyomavirus JC (JCV) large T antigen (LTA) as a potential target for drug development. LTA is a hexameric protein with a helicase activity that is powered by ATP binding and hydrolysis. The helicase and ATPase function is critical for viral replication. Methods Recombinant JCV LTA was produced in an Escherichia coli based expression plasmid. ATPase activity was measured using the malachite green assay. A high throughput screen was completed using a brain-biased library of 75,000 drug-like compounds selected for physicochemical properties consistent with blood brain barrier permeability. Results Five compounds showed non-competitive inhibition of ATPase activity with an EC50 ≤ 15 μM. Modest antiviral activity was demonstrated in an immunofluorescence assay for JCV VP-1 expression in COS7 cells (EC50 15, 18, 20, 27, and 52 μM respectively). The compounds also inhibited viral replication in a real time PCR assay at comparable concentrations. LD50 in the MTS96 and Cell TiterGlo assays was >100 μM for all compounds in COS7 as well as HEK293 cells. However, two compounds inhibited cell proliferation in culture with IC50 values of 43 and 34 μM respectively. Despite substantial amino acid similarity between polyomavirus JC, BK and SV40 proteins, these compounds differ from those previously reported to inhibit SV40 LTA ATPase in chemical structure as well as a non-competitive mechanism of inhibition. Conclusion LTA ATPase is a valid target for discovery. Additional screening and chemical optimization is needed to develop clinically useful compounds with less toxicity, which should be measured by metabolic as well as cell proliferation assays.

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“…Importantly, both bithionol and hexachlorophene inhibit SV40 and BKV cell culture, and they are less toxic than BPA. 124 …”

Section: Inhibitors Of Helicase-catalyzed Atp Hydrolysismentioning

confidence: 99%

“…156 ( C ) Inhibitors of simian virus 40 (SV40) TAg-catalyzed ATP hydrolysis: bisphenol A (CID 6623), IC 50 = 41 μM 128 ; bithionol (CID 2406), IC 50 = 4 μM. 124 ( D ) Hepatitis C virus (HCV) helicase inhibitors: CID 42618092, IC 50 = 10 μM 157 ; aurintricarboxylic acid (CID 2259), IC 50 = 1.4 μM. 158 …”

Section: Figurementioning

confidence: 99%

Discovering New Medicines Targeting Helicases: Challenges and Recent Progress

et al. 2013

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Helicases are ubiquitous motor proteins that separate and/or rearrange nucleic acid duplexes in reactions fueled by adenosine triphosphate (ATP) hydrolysis. Helicases encoded by bacteria, viruses, and human cells are widely studied targets for new antiviral, antibiotic, and anticancer drugs. This review summarizes the biochemistry of frequently targeted helicases. These proteins include viral enzymes from herpes simplex virus, papillomaviruses, polyomaviruses, coronaviruses, the hepatitis C virus, and various flaviviruses. Bacterial targets examined include DnaB-like and RecBCD-like helicases. The human DEAD-box protein DDX3 is the cellular antiviral target discussed, and cellular anticancer drug targets discussed are the human RecQ-like helicases and eIF4A. We also review assays used for helicase inhibitor discovery and the most promising and common helicase inhibitor chemotypes, such as nucleotide analogues, polyphenyls, metal ion chelators, flavones, polycyclic aromatic polymers, coumarins, and various DNA binding pharmacophores. Also discussed are common complications encountered while searching for potent helicase inhibitors and possible solutions for these problems.

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A screen for modulators of large T antigen’s ATPase activity uncovers novel inhibitors of Simian Virus 40 and BK virus replication

Cited by 17 publications

References 66 publications

The conserved core enzymatic activities and the distinct dynamics of polyomavirus large T antigens

The conserved core enzymatic activities and the distinct dynamics of polyomavirus large T antigens

Inhibition of large T antigen ATPase activity as a potential strategy to develop anti-polyomavirus JC drugs

Discovering New Medicines Targeting Helicases: Challenges and Recent Progress

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