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Herpesviridae Dec 2010The UL97 protein kinase of human cytomegalovirus phosphorylates the antiviral drug ganciclovir and is the target of maribavir action. A detailed enzyme kinetic analysis...
BACKGROUND
The UL97 protein kinase of human cytomegalovirus phosphorylates the antiviral drug ganciclovir and is the target of maribavir action. A detailed enzyme kinetic analysis of maribavir on the various enzymatic functions of wild type and ganciclovir resistant forms of UL97 is required.
METHODS
Wild type and site directed mutant forms of the human cytomegalovirus UL97 gene product were expressed using recombinant baculoviruses and the purified products used to assess the effects of maribavir on the ganciclovir (GCV) kinase and protein kinase (PK) activities.
RESULTS
Maribavir was a potent inhibitor of the autophosporylation of the wild type and all the major GCV resistant UL97 mutants analysed (M460I, H520Q, A594V and L595F) with a mean IC50 of 35 nM. The M460I mutation resulted in hypersensitivity to maribavir with an IC50 of 4.8 nM. A maribavir resistant mutant of UL97 (L397R) was functionally compromised as both a GCV kinase and a protein kinase (~ 10% of wild type levels). Enzyme kinetic experiments demonstrated that maribavir was a competitive inhibitor of ATP with a Ki of 10 nM.
DISCUSSION
Maribavir is a potent competitive inhibitor of the UL97 protein kinase function and shows increased activity against the M460I GCV-resistant mutant which may impact on the management of GCV drug resistance in patients.
PubMed: 21429239
DOI: 10.1186/2042-4280-1-4 -
ChemistrySelect Dec 2022Given the rapid progression of the coronavirus disease 2019 (COVID-19) pandemic, an ultrafast response was urgently required to handle this major public crisis. To...
Given the rapid progression of the coronavirus disease 2019 (COVID-19) pandemic, an ultrafast response was urgently required to handle this major public crisis. To contain the pandemic, investments are required to develop diagnostic tests, prophylactic vaccines, and novel therapies. Lately, nucleoside analog (NA) antivirals topped the scene as top options for the treatment of COVID-19 caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections. Meanwhile, the continuous generation of new lineages of the SARS-CoV-2 Omicron variant caused a new challenge in the persistent COVID-19 battle. Hitting the two crucial SARS-CoV-2 enzymes RNA-dependent RNA polymerase (RdRp) and 3'-to-5' exoribonuclease (ExoN) collectively together using only one single ligand is a very successful new approach to stop SARS-CoV-2 multiplication and combat COVID-19 irrespective of the SARS-CoV-2 variant type because RdRps and ExoNs are broadly conserved among all SARS-CoV-2 strains. Herein, the current comprehensive study investigated most NAs libraries, searching for the most ideal drug candidates expectedly able to perfectly act through this double tactic. Gradual computational filtration gave rise to six different promising NAs, which are riboprine, forodesine, tecadenoson, nelarabine, vidarabine, and maribavir, respectively. Further biological assessment proved for the first time, using the anti-RdRp/ExoN and anti-SARS-CoV-2 bioassays, that riboprine and forodesine, among all the six tested NAs, are able to powerfully inhibit the replication of the new virulent strains of SARS-CoV-2 with extremely minute anti-RdRp and anti-SARS-CoV-2 EC values of about 0.22 and 0.49 μM for riboprine and about 0.25 and 0.73 μM for forodesine, respectively, surpassing both remdesivir and the new anti-COVID-19 drug molnupiravir. The prior data supported these biochemical findings, suggesting that riboprine and forodesine molecules strongly hit the key catalytic pockets of the SARS-CoV-2 (Omicron variant) RdRp's and ExoN's main active sites. Additionally, the ideal pharmacophoric features of riboprine and forodesine molecules render them typical dual-action inhibitors of SARS-CoV-2 replication and proofreading, with their relatively flexible structures open for diverse types of chemical derivatization. In Brief, the current important results of this comprehensive study revealed the interesting repurposing potentials of, mainly, the two nucleosides riboprine and forodesine to effectively shut down the polymerase/exoribonuclease-RNA nucleotides interactions of the SARS-CoV-2 Omicron variant and consequently treat COVID-19 infections, motivating us to rapidly begin the two drugs' broad preclinical/clinical anti-COVID-19 bioevaluations, hoping to combine both drugs soon in the COVID-19 treatment protocols.
PubMed: 36718467
DOI: 10.1002/slct.202201912 -
Antiviral Research Dec 2019In a Phase 2 clinical trial, 120 subjects with cytomegalovirus (CMV) infection refractory or resistant to standard therapy were randomized equally to 3 doses of oral... (Randomized Controlled Trial)
Randomized Controlled Trial
In a Phase 2 clinical trial, 120 subjects with cytomegalovirus (CMV) infection refractory or resistant to standard therapy were randomized equally to 3 doses of oral maribavir treatment, and 70% achieved undetectable plasma CMV DNA within 12 weeks. At study entry, standard diagnostic UL97 genotyping was available for 71 subjects, with 60 (85%) revealing well-characterized ganciclovir resistance mutations that did not preclude a therapeutic response to maribavir. Central laboratory testing of a range of UL97 codons (288-468) not fully covered by standard genotyping was done on 93 subjects at baseline. This detected no previously known maribavir resistance mutations, but identified atypical mutations in 3 subjects, including a P-loop substitution F342Y, and ATP-binding region substitutions K359E/Q. By recombinant phenotyping, K359E and K359Q each conferred a nearly 4-fold increased ganciclovir 50% inhibitory concentration (EC50) without maribavir resistance, whereas F342Y conferred a 6-fold increased ganciclovir EC50 and a 4.5-fold increased maribavir EC50. The subject with F342Y detected at baseline did not achieve plasma CMV DNA clearance after 12 weeks of maribavir therapy and later developed an additional UL97 substitution H411Y known to confer 12- to 20-fold increased MBV EC50 by itself. The combination of F342Y and H411Y was shown to increase the maribavir EC50 by 56-fold. Diagnostic genotyping of UL97 should be expanded to cover the ATP-binding region beginning at codon 335 to enable the detection of atypical resistance mutations and further correlation of their clinical significance.
Topics: Benzimidazoles; Cell Line; Cytomegalovirus; Cytomegalovirus Infections; Drug Resistance, Viral; Ganciclovir; Genes, Viral; Genotyping Techniques; Humans; Inhibitory Concentration 50; Mutation; Phosphotransferases (Alcohol Group Acceptor); Ribonucleosides
PubMed: 31568799
DOI: 10.1016/j.antiviral.2019.104616 -
Biology of Blood and Marrow... Jan 2015The 2015 Tandem American Society for Blood and Marrow Transplantation/Center for International Blood and Marrow Transplant Meetings provide an opportunity to review the... (Review)
Review
The 2015 Tandem American Society for Blood and Marrow Transplantation/Center for International Blood and Marrow Transplant Meetings provide an opportunity to review the current status and future perspectives on therapy for cytomegalovirus (CMV) infection in the setting of hematopoietic stem cell transplantation (HSCT). After many years during which we have seen few tangible advances in terms of new antiviral drugs, we are now experiencing an exciting period of late-stage drug development, characterized by a series of phase III trials incorporating a variety of novel agents. These trials have the potential to shift our current standard therapeutic strategies, which generally involve pre-emptive therapy based on sensitive molecular surveillance, towards the prophylactic approaches we see more generally with other herpes viruses such as herpes simplex and varicella zoster. This comes at a time when the promise of extensive preclinical research has been translated into encouraging clinical responses with several cellular immunotherapy strategies, which have also been moved towards definitive late-stage clinical trials. How these approaches will be integrated with the new wave of antiviral drugs remains open to conjecture. Although most of the focus of these cellular immunotherapy studies has been on adaptive immunity, and in particular T cells, an increasing awareness of the possible role of other cellular subsets in controlling CMV infection has developed. In particular, the role of natural killer (NK) cells is being revisited, along with that of γδ T cells. Depletion of NK cells in mice results in higher titers of murine CMV in tissues and increased mortality, whereas NK cell deficiency in humans has been linked to severe CMV disease. We will review recent progress in these areas.
Topics: Acetates; Adoptive Transfer; Animals; Antiviral Agents; Benzimidazoles; Cell- and Tissue-Based Therapy; Clinical Trials, Phase III as Topic; Cytomegalovirus; Cytomegalovirus Infections; Cytosine; Hematopoietic Stem Cell Transplantation; Humans; Isoxazoles; Killer Cells, Natural; Leflunomide; Mice; Organophosphonates; Quinazolines; Ribonucleosides; T-Lymphocytes
PubMed: 25452035
DOI: 10.1016/j.bbmt.2014.11.002 -
Intervirology 2016Drug-resistant isolates of human cytomegalovirus (HCMV) have led to the development of new anti-HCMV drugs. Maribavir (MBV) is a novel inhibitor of the HCMV viral...
OBJECTIVE
Drug-resistant isolates of human cytomegalovirus (HCMV) have led to the development of new anti-HCMV drugs. Maribavir (MBV) is a novel inhibitor of the HCMV viral kinase. Resistance to MBV is mapped to gene UL27, a viral nuclear protein. In this study, we investigated UL27 polymorphisms in MBV-naive HIV-positive and HCMV congenitally infected clinical samples.
METHODS
DNA was extracted from 20 CMV-positive HIV (9/20) and congenitally infected (11/20) patients and used for UL27 polymerase chain reaction amplification. Sanger sequencing and multiple sequence alignment of products was performed.
RESULTS
K90 was the most prevalent polymorphism in both HIV-positive and congenitally infected patients. Polymorphisms Q54, D123, and R107 (10%) were seen in more than one sample. There were significantly more polymorphisms in the HIV-positive samples (p = 0.038).
CONCLUSION
HCMV pUL27 is highly variable in adult immunocompromised HIV-positive patients.
PubMed: 28402975
DOI: 10.1159/000471484 -
Antiviral Research Aug 2012Resistance to the experimental human cytomegalovirus (CMV) UL97 kinase inhibitor maribavir has been mapped to UL97 mutations at codons 353, 397, 409 and 411, in the...
Resistance to the experimental human cytomegalovirus (CMV) UL97 kinase inhibitor maribavir has been mapped to UL97 mutations at codons 353, 397, 409 and 411, in the kinase ATP-binding region, and to mutations in the UL27 gene. We studied the maribavir susceptibility phenotypes of additional UL97 mutations observed in vitro and in clinical trials, and the effect of simultaneous mutation in both UL97 and UL27. In vitro selection under maribavir identified a new locus of UL97 mutation within the conserved kinase p-loop (L337M), which conferred low grade maribavir resistance (3.5-fold increased EC50) without ganciclovir cross-resistance. During maribavir Phase III CMV prevention clinical trials, three previously unknown UL97 sequence variants were detected in plasma samples after 27-98 days of drug exposure (I324V, S334G and S386L). These variants did not confer any drug resistance despite proximity to mutations that confer maribavir resistance. The UL27 resistance mutation R233S, when added to strains containing UL97 mutations L337M or V353A, doubled their maribavir EC50s. These results expand the range of UL97 maribavir-resistance mutations into another part of the kinase ATP-binding region, but offer no genotypic evidence that development of drug resistance affected the outcomes of Phase III maribavir clinical trials after drug exposure of up to 14 weeks. There is a potential for increased maribavir resistance in UL27-UL97 double mutants.
Topics: Antiviral Agents; Benzimidazoles; Clinical Trials, Phase III as Topic; Cytomegalovirus; Cytomegalovirus Infections; Drug Resistance, Viral; Humans; Mutation, Missense; Phosphotransferases (Alcohol Group Acceptor); Ribonucleosides
PubMed: 22664236
DOI: 10.1016/j.antiviral.2012.05.013 -
Journal of Virology Nov 20031-(beta-D-Ribofuranosyl)-2,5,6-trichlorobenzimidazole (TCRB) and its 2-bromo analog, BDCRB, are potent and selective inhibitors of human cytomegalovirus (HCMV) DNA...
1-(beta-D-Ribofuranosyl)-2,5,6-trichlorobenzimidazole (TCRB) and its 2-bromo analog, BDCRB, are potent and selective inhibitors of human cytomegalovirus (HCMV) DNA processing and packaging. Since they are readily metabolized in vivo, analogs were synthesized to improve biostability. One of these, 1-(beta-L-ribofuranosyl)-2-isopropylamino-5,6-dichlorobenzimidazole (1263W94; maribavir), inhibits viral DNA synthesis and nuclear egress. Resistance to maribavir was mapped to UL97, and this viral kinase was shown to be a direct target of maribavir. In the present study, an HCMV strain resistant to TCRB and BDCRB was passaged in increasing concentrations of maribavir, and resistant virus was isolated. This strain (G2) grew at the same rate as the wild-type virus and was resistant to both BDCRB and maribavir. Resistance to BDCRB was expected, because the parent strain from which G2 was isolated was resistant due to known mutations in UL56 and UL89. However, no mutations were found in UL97 or other relevant open reading frames that could explain resistance to maribavir. Because sequencing of selected HCMV genes did not identify the resistance mutation, a cosmid library was made from G2, and a series of recombinant G2 wild-type viruses were constructed. Testing the recombinants for sensitivity to maribavir narrowed the locus of resistance to genes UL26 to UL32. Sequencing identified a single coding mutation in ORF UL27 (Leu335Pro) as the one responsible for resistance to maribavir. These results establish that UL27 is either directly or indirectly involved in the mechanism of action of maribavir. They also suggest that UL27 could play a role in HCMV DNA synthesis or egress of HCMV particles from the nucleus.
Topics: Amino Acid Sequence; Antiviral Agents; Benzimidazoles; Cells, Cultured; Chromosome Mapping; Cytomegalovirus; Drug Resistance, Viral; Humans; Microbial Sensitivity Tests; Molecular Sequence Data; Ribonucleosides; Sequence Analysis, DNA; Viral Plaque Assay; Viral Proteins
PubMed: 14557635
DOI: 10.1128/jvi.77.21.11499-11506.2003 -
Blood Jun 2008The anti-cytomegalovirus (CMV) activity and safety of oral maribavir in CMV-seropositive allogeneic stem-cell transplant recipients were evaluated in a randomized,... (Randomized Controlled Trial)
Randomized Controlled Trial
Maribavir prophylaxis for prevention of cytomegalovirus infection in allogeneic stem cell transplant recipients: a multicenter, randomized, double-blind, placebo-controlled, dose-ranging study.
The anti-cytomegalovirus (CMV) activity and safety of oral maribavir in CMV-seropositive allogeneic stem-cell transplant recipients were evaluated in a randomized, double-blind, placebo-controlled, dose-ranging study. After engraftment, 111 patients were randomized to receive CMV prophylaxis with maribavir (100 mg twice daily, 400 mg once daily, or 400 mg twice daily) or placebo. Within the first 100 days after transplantation, the incidence of CMV infection based on CMV pp65 antigenemia was lower in each of the respective maribavir groups (15%, P = .046; 19%, P = .116; 15%, P = .053) compared with placebo (39%). Similarly, the incidence of CMV infection based on plasma CMV DNA was lower in each of the respective maribavir groups (7%, P = .001; 11%, P = .007; 19%, P = .038) compared with placebo (46%). Anti-CMV therapy was also used less often in patients receiving each respective dose of maribavir (15%, P = .001; 30%, P = .051; 15%, P = .002) compared with placebo (57%). There were 3 cases of CMV disease in placebo patients but none in the maribavir patients. Adverse events, mostly taste disturbance, nausea, and vomiting, were more frequent with maribavir. Maribavir had no adverse effect on neutrophil or platelet counts. These results show that maribavir can reduce the incidence of CMV infection and, unlike ganciclovir, does not cause myelosuppression.
Topics: Adult; Antiviral Agents; Benzimidazoles; Cytomegalovirus Infections; Dose-Response Relationship, Drug; Double-Blind Method; Female; Humans; Male; Middle Aged; Nausea; Ribonucleosides; Stem Cell Transplantation; Taste Disorders; Transplantation, Homologous; Vomiting
PubMed: 18285548
DOI: 10.1182/blood-2007-11-121558 -
Transplantation and Cellular Therapy Jul 2022Outcomes of refractory (Rf) cytomegalovirus (CMV) infection (CMVi) after hematopoietic cell transplantation (HCT) are poor owing to limited treatment options and...
Outcomes of refractory (Rf) cytomegalovirus (CMV) infection (CMVi) after hematopoietic cell transplantation (HCT) are poor owing to limited treatment options and treatment related toxicities. Maribavir, an orally bioavailable CMV antiviral, was recently approved for treatment of Rf-CMVi. Real-world studies quantifying the burden of Rf-CMVi prior to maribavir provide a benchmark for evaluating the net value of novel treatments. Here we report the incidence, clinical outcomes, and healthcare resource utilization (HRU) associated with Rf-CMVi in the first year post-HCT in a cohort of CMV-seropositive HCT recipients (R+) who underwent HCT between January 1, 2014, and December 31, 2017, at Memorial Sloan Kettering Cancer Center and were managed exclusively by preemptive therapy. CMVi was defined as CMV viremia treated preemptively. Rf-CMVi was defined as a <1 log decrease and CMV viral load >1000 U/mL after ≥14 days of appropriately dosed therapy. Welldays were defined as alive days not hospitalized and off CMV antivirals by 1 year post-HCT. The impact of Rf-CMVi on mortality and HRU was examined in multivariable models. Of the 286 R+ patients, 145 (50.7%) developed CMVi (99 no Rf-CMVi and 46 Rf-CMVi). Compared with the no Rf-CMVi group, the Rf-CMVi group had higher rates of CMV EOD (23.9% versus 10.1%; P = .030), CMV-related mortality (9.5% versus .0%; P = .002), and all-cause mortality (33.3% versus 15.6%; adjusted P = .049). Rf-CMVi was an independent predictor for readmission (adjusted odds ratio [aOR], 3.24; 95% confidence interval [CI], 2.19 to 4.87; P < .0001); CMV-related readmission (aOR, 9.48; 95% CI, 5.83 to 15.80; P < .0001), and decreased well days (adjusted arithmetic mean ratio, .72; 95% CI, .58 to .89; P = .001) in the first year post-HCT. Rf-CMVi is associated with increased mortality and increased HRU at 1 year after HCT. Improved therapies for Rf-CMVi have the potential of improving HCT outcomes and reducing HRU.
Topics: Antiviral Agents; Cytomegalovirus; Cytomegalovirus Infections; Hematopoietic Stem Cell Transplantation; Humans; Transplant Recipients; Viremia
PubMed: 35476955
DOI: 10.1016/j.jtct.2022.04.016 -
International Journal of Molecular... Jan 2021Human cytomegalovirus (HCMV) is a human pathogenic herpesvirus associated with a variety of clinical symptoms. Current antiviral therapy is not always effective, so that...
Human cytomegalovirus (HCMV) is a human pathogenic herpesvirus associated with a variety of clinical symptoms. Current antiviral therapy is not always effective, so that improved drug classes and drug-targeting strategies are needed. Particularly host-directed antivirals, including pharmaceutical kinase inhibitors (PKIs), may help to overcome problems of drug resistance. Here, we focused on utilizing a selection of clinically relevant PKIs and determined their anticytomegaloviral efficacies. Particularly, PKIs directed to host or viral cyclin-dependent kinases, i.e., abemaciclib, LDC4297 and maribavir, exerted promising profiles against human and murine cytomegaloviruses. The anti-HCMV in vitro activity of the approved anti-cancer drug abemaciclib was confirmed in vivo using our luciferase-based murine cytomegalovirus (MCMV) animal model in immunocompetent mice. To assess drug combinations, we applied the Bliss independence checkerboard and Loewe additivity fixed-dose assays in parallel. Results revealed that (i) both affirmative approaches provided valuable information on anti-CMV drug efficacies and interactions, (ii) the analyzed combinations comprised additive, synergistic or antagonistic drug interactions consistent with the drugs' antiviral mode-of-action, (iii) the selected PKIs, especially LDC4297, showed promising inhibitory profiles, not only against HCMV but also other α-, β- and γ-herpesviruses, and specifically, (iv) the combination treatment with LDC4297 and maribavir revealed a strong synergism against HCMV, which might open doors towards novel clinical options in the near future. Taken together, this study highlights the potential of therapeutic drug combinations of current developmental/preclinical PKIs.
Topics: Aminopyridines; Animals; Antiviral Agents; Benzimidazoles; Cell Line; Cytomegalovirus; Cytomegalovirus Infections; Drug Combinations; Drug Resistance, Viral; Ganciclovir; Humans; Mice; Protein Kinase Inhibitors; Pyrazoles; Ribonucleosides; Triazines; Virus Replication
PubMed: 33430060
DOI: 10.3390/ijms22020575