GCV resistance due to the mutation A594P in the cytomegalovirus protein UL97 is partially reconstituted by a second mutation at D605E

Ijichi, Osamu; Michel, Detlef; Mertens, Thomas; Miyata, Koichiro; Eizuru, Yoshito

doi:10.1016/s0166-3542(01)00202-9

Cited by 33 publications

(19 citation statements)

References 31 publications

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“…It has been described that combinations of mutations may act synergistically with regard to drug resistance or may act as compensatory mutations. Such a partial compensation of reduced drug susceptibility has previously been described for a pair of UL97 mutations (24). In our study, we investigated whether the D413E mutation was able to compensate at least partially for the reduced FOS susceptibility conferred by the E756K mutation (5, 6).…”

Section: Discussionmentioning

confidence: 81%

“…Information concerning the effects of combinations of mutations is even scarcer, although such combinations are increasingly found, especially in UL54 sequences from patients' specimens. The resulting phenotype of combined mutations is difficult to predict, since it has been shown in a few case reports that combined mutations may potentiate resistance (20,30,33) or, in contrast, may compensate for and reduce the loss of susceptibility conferred by the single mutations (24).…”

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confidence: 99%

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Differentiation between Polymorphisms and Resistance-Associated Mutations in Human Cytomegalovirus DNA Polymerase

Dittmann

Ersing

Mertens

et al. 2010

Antimicrob Agents Chemother

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Specific mutations in the human cytomegalovirus (HCMV) DNA polymerase (pUL54) are known to confer resistance against all currently licensed drugs for treatment of HCMV infection and disease. Following the widespread use of antivirals, the occurrence of HCMV drug resistance is constantly increasing. Recently, diagnostic laboratories have started to replace phenotypic drug resistance testing with genotypic resistance testing. However, the reliability and success of genotypic testing highly depend on the availability of high-quality phenotypic resistance data for each individual mutation and for combinations of mutations, with the latter being increasingly found in patients' HCMV isolates. We performed clonal marker transfer experiments to investigate the impacts of 7 different UL54 point mutations and also of combinations of these mutations on drug susceptibility and viral replicative fitness. We show that several mutations-S695T, A972V, K415R, S291P, and A692V-of suspected but uncertain drug susceptibility phenotype, either alone or in combination, were not relevant to antiviral drug resistance. In contrast, the combination of two mutations individually characterized previously-E756K and D413E-conferred high-grade loss of susceptibility to all three antivirals. Our results have been added to the newly available database of all published HCMV resistance mutations (http://www.informatik.uni-ulm .de/ni/mitarbeiter/HKestler/hcmv/index.html). These data will allow better interpretation of genotypic data and further improve the basis for drug resistance testing.

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Section: Discussionmentioning

confidence: 81%

mentioning

confidence: 99%

Differentiation between Polymorphisms and Resistance-Associated Mutations in Human Cytomegalovirus DNA Polymerase

Dittmann

Ersing

Mertens

et al. 2010

Antimicrob Agents Chemother

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“…For cell-associated clinical isolates, the number of plaque-forming cells (PFCs) present in an infected culture was estimated according to previously described methods (19). The following clinically isolated virus strains were used: strains 01, 03, and 05 were isolated from patients with congenital HCMV infection; strains 02 and 04 were from patients with bone marrow transplantation and kidney transplantation (21), respectively; strain 06 was from a patient with lung transplantation whose clinical course will be described elsewhere; strains 07 and 08 were isolated from a patient with CMV retinitis and encephalitis, as reported previously (13,32); and strain 09 was from a patient with AIDS (14).…”

Section: Methodsmentioning

confidence: 99%

Novel Real-Time Monitoring System for Human Cytomegalovirus- Infected Cells In Vitro That Uses a Green Fluorescent Protein-PML-Expressing Cell Line

Ueno

Eizuru

Katano

et al. 2006

Antimicrob Agents Chemother

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Promyelocytic leukemia (PML) bodies are discrete nuclear foci that are intimately associated with many DNA viruses. In human cytomegalovirus (HCMV) infection, the IE1 (for "immediate-early 1") protein has a marked effect on PML bodies via de-SUMOylation of PML protein. Here, we report a novel real-time monitoring system for HCMV-infected cells using a newly established cell line (SE/15) that stably expresses green fluorescent protein (GFP)-PML protein. In SE/15 cells, HCMV infection causes specific and efficient dispersion of GFP-PML bodies in an IE1-dependent manner, allowing the infected cells to be monitored by fluorescence microscopy without immunostaining. Since a specific change in the detergent solubility of GFP-PML occurs upon infection, the infected cells can be quantified by GFP fluorescence measurement after extraction. With this assay, the inhibitory effects of heparin and neutralizing antibodies were determined in small-scale cultures, indicating its usefulness for screening inhibitory reagents for laboratory virus strains. Furthermore, we established a sensitive imaging assay by counting the number of nuclei containing dispersed GFP-PML, which is applicable for titration of slow-growing clinical isolates. In all strains tested, the virus titers estimated by the GFP-PML imaging assay were well correlated with the plaque-forming cell numbers determined in human embryonic lung cells. Coculture of SE/15 cells and HCMV-infected fibroblasts permitted a rapid and reliable method for estimating the 50% inhibitory concentration values of drugs for clinical isolates in susceptibility testing. Taken together, these results demonstrate the development of a rapid, sensitive, quantitative, and specific detection system for HCMV-infected cells involving a simple procedure that can be used for titration of low-titer clinical isolates.

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“…Recently, in transplant patients and AIDS patients, 28 phenotypically GCV-sensitive strains with changes of amino acids in the UL97 gene have been encountered. The 605 mutation was observed in only one isolate (C325) but it was not related to a resistant phenotypic to the GCV (Gilbert et al, 2002;Ijichi et al, 2002;Lurain et al, 2001) showed in vitro, in one resistant strain (A594P), that the 605 mutation would partially reconstitute the GCV phosphorylation activity and that it would work as a "compensatory mutation". Mutations in the UL97 gene, which confer GCV resistance, cluster in two regions which are highly conserved in all protein kinases.…”

Section: Resultsmentioning

confidence: 97%

Cytomegalovirus UL97 mutations associated with ganciclovir resistance in immunocompromised patients from Argentina

Puch¹,

Ochoa²,

Carballal³

et al. 2004

Journal of Clinical Virology

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GCV resistance due to the mutation A594P in the cytomegalovirus protein UL97 is partially reconstituted by a second mutation at D605E

Cited by 33 publications

References 31 publications

Differentiation between Polymorphisms and Resistance-Associated Mutations in Human Cytomegalovirus DNA Polymerase

Differentiation between Polymorphisms and Resistance-Associated Mutations in Human Cytomegalovirus DNA Polymerase

Novel Real-Time Monitoring System for Human Cytomegalovirus- Infected Cells In Vitro That Uses a Green Fluorescent Protein-PML-Expressing Cell Line

Cytomegalovirus UL97 mutations associated with ganciclovir resistance in immunocompromised patients from Argentina

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