Protein kinase inhibitors of the quinazoline class exert anti-cytomegaloviral activity in vitro and in vivo

Schleiss, Mark R.; Eickhoff, Jan; Auerochs, Sabrina; Leis, Martina; Abele, S.; Rechter, Sabine; Choi, Yoonsuk; Anderson, Jodi; Scott, Gillian; Rawlinson, William D.; Michel, Detlef; Ensminger, Stephan; Klebl, Bert; Stamminger, Thomas; Marschall, Manfred

doi:10.1016/j.antiviral.2008.01.154

Cited by 74 publications

(62 citation statements)

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“…This finding prompted us to analyse additional protein kinase inhibitors, particularly those inhibiting the viral protein kinase pUL97. Two inhibitors of pUL97 kinase activity, Ax7396 and Gö6976, belonging to two different chemical classes, quinazoline and indolocarbazole, both of which inhibit pUL97 kinase activity as well as HCMV replication in vitro (Herget et al, 2004;Marschall et al, 2001Marschall et al, , 2002Schleiss et al, 2008), also induced intranuclear microspeckled accumulation of pUL69 similarly to roscovitine (Fig. 1a,v,vi and viii).…”

Section: Resultsmentioning

confidence: 99%

Cytomegaloviral protein kinase pUL97 interacts with the nuclear mRNA export factor pUL69 to modulate its intranuclear localization and activity

Thomas

Rechter

Milbradt

et al. 2009

Journal of General Virology

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Human cytomegalovirus encodes a number of phosphorylation-regulated proteins, including the autophosphorylating protein kinase pUL97 and the nuclear mRNA export factor pUL69. Recently, it was reported that the kinase inhibitor roscovitine induces an intranuclear aggregation of pUL69 in infected fibroblasts. Here, we demonstrate that pUL97-specific kinase inhibitors induce a similar pUL69 aggregation. Furthermore, a direct pUL69-pUL97 interaction was demonstrated by coimmunoprecipitation analyses. Deletion mapping identified the domains required for interaction in both proteins (1-140/478-532 in pUL69 and 231-336 in pUL97). Further analysis of the immunoprecipitates by in vitro kinase assays demonstrated the phosphorylation of pUL69 by pUL97. However, catalytically inactive mutants of pUL97 and interaction-negative fragments of pUL69 were phosphorylation-negative. Moreover, an analysis of the pUL69-mediated nuclear RNA export indicated a correlation of the export efficiency with the presence of active pUL97 kinase. These data suggest a specific pUL69-pUL97 interaction and pUL97-mediated phosphorylation which influences the regulatory activities of pUL69.

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Section: Resultsmentioning

confidence: 99%

Cytomegaloviral protein kinase pUL97 interacts with the nuclear mRNA export factor pUL69 to modulate its intranuclear localization and activity

Thomas

Rechter

Milbradt

et al. 2009

Journal of General Virology

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“…Anticancer agent Iressa (ZD1839) A is a clinically approved example of quinazoline-based inhibitors which is in phase III clinical trials for cancer used to inhibit the receptor tyrosine kinase of epidermal growth factor. also exhibit pronounced biological activities as antimicrobial, 7,8 anti-HIV (NNRTI), 9,10 anti-tumour, [11][12][13][14][15][16] potential analgesic and anti-inflammatory activity. 17 Non-proteinogenic amino acids are major component in a number of drugs including β-lactam antibiotics 18 and glutamate antagonists.…”

Section: Introductionmentioning

confidence: 99%

Synthesis of methyl [3-alkyl-2-(2,4-dioxo-3,4-dihydro-2H-quinazolin-1-yl)-acetamido] alkanoate

Ismail¹,

Ali²,

Fathalla³

et al. 2017

Arkivoc

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A series of methyl [3-alkyl-2-(2,4-dioxo-3,4-dihydro-2H-quinazolin-1-yl)-acetamido] alkanoate 10-13a-f has been developed on the basis of the N-chemoselective reaction of 3-substituted quinazoline-2,4-diones 3a-d with ethyl chloroacetate and azide coupling method with amino acid ester hydrochloride. The precursor quinazoline diones 3a-d chemoselective reactions were studied using DFT(B3LYP)/6-311G level of theory and were prepared by a new rearrangement method from the corresponding 2-(3-methyl-4-oxo-3,4-dihydroquinazolin-2-ylthio) acetohydrazide 6.

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“…Genetic deletion of UL97 results in a severe (10-to 1,000-fold) replication defect in vitro (8-10), as does mutation of the K355 lysine residue, which is critical for kinase activity (6). UL97 is therefore an attractive target for antiviral drug development (1,(11)(12)(13)(14) as an alternative to the UL54 DNA polymerase target of all currently licensed HCMV antivirals, including ganciclovir (GCV).Maribavir (MBV) is a benzimidazole riboside pUL97 inhibitor that inhibits HCMV replication potently (15) but variably depending on cell culture conditions (16). While MBV showed promise in early clinical trials, two recently published phase III trials demonstrated that MBV was no more effective than a placebo at preventing HCMV viremia after allogeneic stem cell transplantation or liver transplantation (17, 18).…”

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Differential Properties of Cytomegalovirus pUL97 Kinase Isoforms Affect Viral Replication and Maribavir Susceptibility

Webel

Hakki

Prichard

et al. 2014

J Virol

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The human cytomegalovirus (HCMV)-encoded kinase pUL97 is required for efficient viral replication. Previous studies described two isoforms of pUL97, the full-length isoform (M1) and a smaller isoform likely resulting from translation initiation at codon 74 (M74). Here, we report the detection of a third pUL97 isoform during viral infection resulting from translation initiation at codon 157 (isoform M157). The consistent expression of isoform M157 as a minor component of pUL97 during infection with clinical and laboratory-adapted HCMV strains was suppressed when codon 157 was mutagenized. Viral mutants expressing specific isoforms were generated to compare their growth and drug susceptibility phenotypes, as well as pUL97 intracellular localization patterns and kinase activities. The exclusive expression of isoform M157 resulted in substantially reduced viral growth and resistance to the pUL97 inhibitor maribavir while retaining susceptibility to ganciclovir. Confocal imaging demonstrated reduced nuclear import of amino-terminal deletion isoforms compared to isoform M1. Isoform M157 showed reduced efficiency of various substrate protein interactions and autophosphorylation, whereas Rb phosphorylation was preserved. These results reveal differential properties of pUL97 isoforms that affect viral replication, with implications for the antiviral efficacy of maribavir. IMPORTANCEThe HCMV UL97 kinase performs important functions in viral replication that are targeted by the antiviral drug maribavir. Here, we describe a naturally occurring short isoform of the kinase that when expressed by itself in a recombinant virus results in altered intracellular localization, impaired growth, and high-level resistance to maribavir compared to those of the predominant full-length counterpart. This is another factor to consider in explaining why maribavir appears to have variable antiviral activity in cell culture and in vivo.T he product of the human cytomegalovirus (HCMV) UL97 gene (pUL97) is a serine/threonine protein kinase that phosphorylates itself and multiple viral and host proteins (1). It is expressed early in infection (2, 3), localizes predominantly to the nucleus using amino-terminal nuclear localization signals (4, 5), and is also observed in the cytoplasm later in the infection cycle (2, 5, 6). It is also present in the HCMV virion (7). Genetic deletion of UL97 results in a severe (10-to 1,000-fold) replication defect in vitro (8-10), as does mutation of the K355 lysine residue, which is critical for kinase activity (6). UL97 is therefore an attractive target for antiviral drug development (1,(11)(12)(13)(14) as an alternative to the UL54 DNA polymerase target of all currently licensed HCMV antivirals, including ganciclovir (GCV).Maribavir (MBV) is a benzimidazole riboside pUL97 inhibitor that inhibits HCMV replication potently (15) but variably depending on cell culture conditions (16). While MBV showed promise in early clinical trials, two recently published phase III trials demonstrated that MBV was no more...

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Protein kinase inhibitors of the quinazoline class exert anti-cytomegaloviral activity in vitro and in vivo

Cited by 74 publications

References 39 publications

Cytomegaloviral protein kinase pUL97 interacts with the nuclear mRNA export factor pUL69 to modulate its intranuclear localization and activity

Cytomegaloviral protein kinase pUL97 interacts with the nuclear mRNA export factor pUL69 to modulate its intranuclear localization and activity

Synthesis of methyl [3-alkyl-2-(2,4-dioxo-3,4-dihydro-2H-quinazolin-1-yl)-acetamido] alkanoate

Differential Properties of Cytomegalovirus pUL97 Kinase Isoforms Affect Viral Replication and Maribavir Susceptibility

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