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Introduction: Human cytomegalovirus (Orthoherpesviridae: Herpesvirales: Cytomegalovirus: Cytomegalovirus humanbeta 5 ) (HCMV) is one of the most commonly detected viruses in recipients of allogeneic hematopoietic stem cell (allo-HSCT) transplants. However, the emergence of resistance to antiviral drugs such as ganciclovir (GCV) poses a challenge in managing these patients. This study aims to investigate the prevalence and impact of mutations in the HCMV UL97 gene associated with resistance to GCV on the course of infection among allo-HSCT patients., Materials and Methods: The study examined the association between UL97 mutations and the clinical course of HCMV infection in allo-HSCT patients. Genetic sequencing was performed to identify mutations, and their impact on viral replication and resistance to GCV was assessed., Results and Discussion: Six mutations were identified (D490A, T502A, C592G, C592F, E596G, C603W). C592G, C592F, E596G, and C603W are associated with resistance to antiviral drugs, while D490A and T502A described for the first time. When comparing patients with wild-type and those carrying the mutant variant, several parameters of peripheral blood were significantly lower in the former group. The median time to peak viral load following allo-HSCT, duration of viremia, and rate of virological response to high-dose therapy also differed significantly between the two groups., Conclusion: It was shown that approximately one third (4 out of 14) of allogeneic stem cell transplant recipients had mutations associated with resistance to GCV. Patients carrying the mutant variant of HCMV had longer viremia and took longer to achieve a negative virological test result after starting high-dose therapy. Performing genotyping may help make more evidence-based therapeutic decisions.

Efflux transporters, namely ATP-binding cassette (ABC), are one of the primary reasons for cancer chemoresistance and the clinical failure of chemotherapy. Ganciclovir (GCV) is an antiviral agent used in herpes simplex virus thymidine kinase (HSV-TK) gene therapy. In this therapy, HSV-TK gene is delivered together with GCV into cancer cells to activate the phosphorylation process of GCV to active GCV-triphosphate, a DNA polymerase inhibitor. However, GCV interacts with efflux transporters that are responsible for the resistance of HSV-TK/GCV therapy. In the present study, it was explored whether GCV and its more lipophilic derivative ( 1 ) could inhibit effluxing of another chemotherapeutic, methotrexate (MTX), out of the human breast cancer cells. Firstly, it was found that the combination of GCV and MTX was more hemocompatible than the corresponding combination with compound 1 . Secondly, both GCV and compound 1 enhanced the cellular accumulation of MTX in MCF-7 cells, the MTX exposure being 13-21 times greater compared to the MTX uptake alone. Subsequently, this also reduced the number of viable cells (41-56%) and increased the number of late apoptotic cells (46-55%). Moreover, both GCV and compound 1 were found to interact with breast cancer resistant protein (BCRP) more effectively than multidrug-resistant proteins (MRPs) in these cells. Since the expression of BCRP was higher in MCF-7 cells than in MDA-MB-231 cells, and the cellular uptake of GCV and compound 1 was smaller but increased in the presence of BCRP-selective inhibitor (Fumitremorgin C) in MCF-7 cells, we concluded that the improved apoptotic effects of higher MTX exposure were raised mainly from the inhibition of BCRP-mediated efflux of MTX. However, the effects of GCV and its derivatives on MTX metabolism and the quantitative expression of MTX metabolizing enzymes in various cancer cells need to be studied more thoroughly in the future.

Valganciclovir (VGCV) is the prodrug of ganciclovir (GCV). The objective of this study was to establish a population pharmacokinetic model (PPK) of GCV to investigate the PK characteristics of GCV after administration of VGCV in adult Chinese renal allograft recipients. Seventy Chinese renal allograft recipients were given 450 mg (n = 41) or 900 mg (n = 29) VGCV daily. Blood samples were drawn 0-24 hours after 5 days' therapy, and GCV plasma levels were determined. The PPK model was constructed using nonlinear mixed-effects modeling, and the Bayesian estimation of AUC 0-24h was constructed for an individual patient based on limited plasma samples. The PK of GCV was best described by a 2-compartment model with a first-order absorption process. The CL/F, V 2 /F, Q/F, V 3 /F, K a , and lag time of GCV were 15.8 ± 0.71 L/h, 10.9 ± 2.38 L, 3.98 ± 0.40 L/h, 167 ± 44.0 L, 0.23 ± 0.0078 1/h, and 0.93 ± 0.017 hours, respectively. Clearance of creatinine was found to have a significant impact on the CL/F of GCV (P < .01). Sampling strategies consisted of plasma concentrations 0 and 2 and 0, 2, and 4 hours after VGCV administration were shown to be suitable for the estimation of the GCV AUC 0-24h . The PPK model was acceptable and can describe the PK of GCV in Chinese renal transplant patients administered VGCV. The AUC 0-24h of GCV in Chinese renal transplant patients can be calculated by a limited sampling strategy method., (© 2020, The American College of Clinical Pharmacology.)

Objectives: Valganciclovir (VGCV) treatment is recommended for the prevention of cytomegalovirus (CMV) infection in renal allograft recipients. The aim of the present study is to investigate the pharmacokinetic characteristics of ganciclovir (GCV) after administration of VGCV in Chinese adult renal allograft recipients and estimate the exposure to GCV using limited sampling strategy (LSS)., Methods: Forty Chinese renal allograft recipients were given 450 mg or 900 mg VGCV daily. Blood samples were drawn before treatment and 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12, and 24 h after 5 days of VGCV therapy, and the plasma concentrations of VGCV and GCV were determined using a liquid chromatography-mass spectrometry assay. The major pharmacokinetic parameters for GCV and VGCV were determined using a noncompartmental assay. Multiple stepwise linear regression analysis was conducted to establish a model equation for the estimation of the GCV AUC 0-24 h in Chinese patients using LSS., Results: In the 450 and 900 mg groups, the C max for VGCV was 0.2 ± 0.10 and 0.4 ± 0.16 mg/L, respectively; the C max for GCV was 4.2 ± 1.1 and 8.6 ± 1.6 mg/L, respectively; and the AUC 0-24 h for GCV was 28.4 ± 8.4 and 60.7 ± 17.5 mg·h/L, respectively. For the establishment of LSS models, 40 patients were divided into the training group (n = 24) and validation group (n = 16). The model equations used for the calculation of AUC 0-24 h for GCV were established in the training group by using multiple linear regression assay. Equations including AUC = 8.1 + 29.7 × C 0 + 5.7 × C 4 (r 2 = 0.91) and AUC = - 0.4 + 11.0 × C 0 + 2.1 × C 2 + 13.7 × C 8 (r 2 = 0.98) were acceptable. The %MPE and %MAPE values obtained from the validation group for the two model equations were 5.89 ± 14.5% and 12.1 ± 9.53%, and - 1.30 ± 4.40% and 3.28 ± 3.11%, respectively., Conclusions: The LSS models that included C 0 and C 4 or C 0 , C 2 , and C 8 in the estimation of AUC 0-24 h for GCV had favorable performance and can be used for therapeutic drug monitoring in the prevention of CMV infection using VGCV in Chinese renal allograft recipients.

Abstract: The limitations of the ganciclovir (GCV)/herpes simplex virus thymidine kinase (HSV1 TK: EC 2.7.1.21) system as a suicide gene therapy approach have been extensively studied over the years. In our study, we focused on improving the cytotoxic profile of the GCV/equine herpes virus-4 thymidine kinase (EHV4 TK: EC 2.7.1.21) system. Our approach involved the structure-guided mutagenesis of EHV4 TK in order to switch its ability to preferentially phosphorylate the natural substrate deoxythymidine (dT) to that of GCV. We performed steady-state kinetic analysis, genetic complementation in a thymidine kinase-deficient Escherichia coli strain, isothermal titration calorimetry, and analysis of GCV-induced cell killing through generation of HEK 293 stable cell-lines expressing EHV4 TK mutants and wild-type EHV4 TK. We found that the EHV4 TK S144H-GFP mutant preferentially phosphorylates GCV and confers increased GCV-induced cytotoxicity compared to wild-type EHV4 TK. [Copyright &y& Elsevier]

Background: Ganciclovir/valganciclovir plays an important role in the treatment and prevention of cytomegalovirus disease after organ transplantation. Material and methods: We developed and validated a simple chromatographic method by ultra-performance liquid chromatography tandem mass spectrometry to measure plasma concentration of ganciclovir in human plasma. Chromatographic separation was achieved using an Acquity® UPLC® BEH™ (2.1×50mm id, 1.7μm) reverse-phase C18 column, with a water/methanol linear gradient containing ammonium acetate/formic acid at a 0.4mL/min flow rate. Ganciclovir and its internal standard (acyclovir) were detected by electrospray ionization mass spectrometry in positive ion multiple reaction monitoring mode. Results: The limit of detection and quantification were 0.03 and 0.06mg/L, respectively, and linearity was observed between 0.06 and 30.0mg/L. Intra-day and day-to-day coefficients of variation and relative biases ranged from 3.6 to 5.4%, 4.2 to 6.2%, −2.6 to −1.1% and −4.0 to −2.8%, respectively. Recovery values were greater than 81.9%. Evaluation of the matrix effect showed ion suppression for ganciclovir and acyclovir. No carry-over was observed. Conclusions: The validated method is useful for both therapeutic drug monitoring and pharmacokinetic studies. It could be applied to the daily clinical laboratory practice to measure the concentration of ganciclovir in human plasma. [ABSTRACT FROM AUTHOR]

Highlights: [•] Homologous recombination deficiency enhances ganciclovir cytotoxicity >14-fold. [•] Vorinostat inhibits homologous recombination repair events. [•] Vorinostat+ganciclovir is synergistic only in cells with homologous recombination. [•] Vorinostat does not alter ganciclovir metabolism or cell cycle distribution. [•] Ganciclovir synergy is due to vorinostat suppression of Rad51, Rad51 foci and CtIP. [Copyright &y& Elsevier]

Abstract: We characterised by pyrosequencing, the dynamics of cytomegalovirus populations harbouring mutations A594V in gene UL97 and A834P and Q578H in gene UL54 in a haematopoietic stem cell transplant recipient. Unexpected re-emergence of A594V and decrease of A834P under CMX001 were shown to depend on both the selection pressure exerted by the antiviral treatments and the immune response. [Copyright &y& Elsevier]

Abstract: Background: Human herpesvirus 6 (HHV-6) and human cytomegalovirus (HCMV) are major opportunistic pathogens in solid organ transplant (SOT) recipients. The use of antivirals for the treatment of HCMV disease can result in the development of drug resistance mutations in HCMV and also potentially in HHV-6. Objectives: The emergence of HHV-6 drug resistance mutations was evaluated in SOT recipients at the onset of HCMV disease and following treatment with ganciclovir (GCV) or valganciclovir (VGCV). Study design: Detection of HHV-6 was performed by real-time PCR from whole blood samples serially obtained from SOT recipients treated for HCMV disease with an induction dose of intravenous GCV or oral VGCV for 21 days followed by VGCV maintenance for 28 days in both arms. Baseline and last positive HHV-6 samples were tested for mutations in the genes encoding the protein kinase (U69) and the DNA polymerase (U38). Results: The rate of HHV-6 viraemia among SOT patients with HCMV disease at baseline was 3.2% (5/155). All isolates belonged to the HHV-6B species. Mutations L213I and Y479H were detected at baseline and at later times in the U69 kinase. Mutation L213I was previously reported as polymorphism whereas the role of mutation Y479H in drug resistance is unknown. Mutations D854E and E855Q found in the DNA polymerase were known as natural variants. Conclusions: The incidence of HHV-6 viraemia in SOT recipients with established HCMV disease before initiation of antiviral therapy was low. Treatment with GCV or VGCV did not induce the emergence of HHV-6 drug resistance mutations. [Copyright &y& Elsevier]

Abstract: This manuscript describes the determination of Ganciclovir (GCV), active component of the antiviral drug Valcyte®, and its ester prodrug Valganciclovir (VGC) in human and rat plasma, using liquid chromatography coupled to tandem mass spectrometry. Protein precipitation with acetonitrile was followed by hydrophilic interaction liquid chromatography on a silica column with 4min run time. After electrospray ionization, the compounds were detected in positive ion selected reaction monitoring (SRM) mode. The lower limits of quantification (LLOQ) were 16ng/mL for GCV and 4ng/mL for VGC in human and rat plasma. Inter-day and intra-day precisions and inaccuracies were below 15% and between 85 and 115%, respectively. Five-fold deuterated GCV and VGC were used as internal standards and compensated for any matrix effect. The method was successfully applied to samples from a rat pharmacokinetic study. The feasibility of blood analysis as dried blood spots (DBS) was investigated. [Copyright &y& Elsevier]

We evaluated the cytotoxic and apoptotic effects of two purine nucleoside analogues, acyclovir (ACV) and ganciclovir (GCV), on lymphoma cells stably harboring Kaposi's sarcoma-associated herpesvirus (KSHV). Colorimetric caspase assay, flow cytometry, and immunoblotting with antibodies against apoptosis-related molecules revealed that GCV has cytotoxic activity toward KSHV-infected primary effusion lymphoma cells, while ACV has weak or little activity. In addition to the GCV-induced cytotoxicity, apoptosis via caspase-7/8, cleavage of poly(ADP-ribose) polymerase, and accumulation of p53 and p21 were induced by GCV treatment. In contrast, neither ACV nor GCV have cytotoxicity- or apoptosis-inducing activities toward uninfected cells. [ABSTRACT FROM AUTHOR]

An improved total synthesis of [18F]FHPG starting from 1,3-dibenzyloxy-2-propanol and guanine has been developed. [18F]FHPG was prepared by nucleophilic substitution of the appropriate precursor with [18F]KF/Kryptofix 2.2.2 followed by a quick deprotection reaction and purification with a simplified Silica Sep-Pak solid-phase extraction (SPE) method in 10–15% radiochemical yield, and 70 min synthesis time from end of bombardment (EOB). [ABSTRACT FROM AUTHOR]