Ganciclovir

Optimization of Ganciclovir use in allogeneic hematopoietic cell transplant recipients – the role of therapeutic drug monitoring
d,e,f and Michelle Yonga,b,c
aDepartments of Infectious Diseases, The National Centre for Infections in Cancer, Peter MacCallum Cancer Centre Melbourne, Australia; bSir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Australia; cVictorian Infectious Diseases Services Department, Royal Melbourne Hospital, Parkville VIC, Australia; dUniversity of Queensland Centre for Clinical Research, Faculty of Medicine & Centre for Translational Anti-infective Pharmacodynamics, School of Pharmacy, The University of Queensland, Brisbane, Australia; eDepartments of Pharmacy and Intensive Care Medicine, Royal Brisbane and Women’s Hospital, Brisbane, Australia; fDivision of Anaesthesiology Critical Care Emergency and Pain Medicine, Nîmes University Hospital, University of Montpellier, Nîmes, France

ABSTRACT
Introduction: Cytomegalovirus (CMV) is an opportunistic infectious complication that can occur after allogeneic hematopoietic cell transplantation (HCT). The mainstay of treatment and prevention of this infection is ganciclovir and its ester prodrug valganciclovir. There is conflicting evidence on the clinical utility of routine ganciclovir therapeutic drug monitoring (TDM) as a means to optimize treatment. Areas covered: This review aims to describe the current knowledge of the pharmacokinetic and pharmacodynamic characteristics of ganciclovir and valganciclovir, and to explore the evidence and challenges surrounding ganciclovir TDM within the allogeneic HCT cohort.
Expert opinion: Ganciclovir TDM is important to optimize efficacy in selected patient groups where there are variable pharmacokinetic factors or inadequate response to treatment. However, defined pharmacokinetic exposures which correlate with treatment efficacy and toxicity remain elusive. Prospective clinical studies in specific patient groups are required to clarify this issue. Alternative TDM targets such as the intracellular ganciclovir triphosphate should be explored as they may prove to have better correlation with clinical outcomes and adverse effects. With recent advances in CMV immune monitoring, novel approaches integrat- ing TDM with specific CMV immune phenotyping in a predictive model will be advantageous in optimizing ganciclovir dosing by combining TDM with a risk stratification approach.
ARTICLE HISTORY Received 18 July 2020 Accepted 11 November 2020
KEYWORDS
Allogeneic; CMV; ganciclovir; TDM; valganciclovir

1.Introduction
Human cytomegalovirus (CMV) infection is ubiquitous globally with adult seroprevalence ranging from 45% to 100% in devel- oped and developing countries, respectively, [1,2]. Following primary infection, complex host immune response interplay controls and directs the virus into latency [1]. This immune control is disrupted in allogeneic hematopoietic stem cell transplantation (HCT), enabling the development of active infection which can range from asymptomatic viremia to tis- sue-invasive CMV disease caused by the direct cytotoxic effects of the virus [3]. Approximately 70% of CMV seropositive recipients will experience early CMV reactivation (within the first 100 days post-transplant) [4]. The antiviral drug ganciclo- vir (and orally administered pro-drug, valganciclovir) has been the cornerstone for the management of CMV prophylaxis, infection, and disease.
Since early clinical usage, there have been attempts to explore a relationship between ganciclovir exposure with clin- ical response and adverse effects. To date, there is no con- sensus on the clinical pharmacokinetic (PK) parameter to be measured, and target exposures which correlate with clinical outcomes or toxicity. However, the published work so far has highlighted significant deficits in the current weight- and

renally-adjusted dosing algorithms which raise questions of whether such issues could be overcome through the use of therapeutic drug monitoring (TDM). At present, the majority of studies describing the clinical pharmacokinetics (PK) and phar- macodynamics of ganciclovir are dated and have been pre- dominantly conducted in solid organ transplant recipients [5]
or involve small heterogeneous subsets of immunocompro- mised patients. Results from such studies may not be transla- table to allogeneic HCT recipients who have a unique pathology characteristics that can impact on systemic antiviral exposure such as the presence of graft-versus-host-disease of the gastrointestinal tract, steroid exposure, acute renal func- tion changes and host immune reconstitution post-transplant. TDM-guided dosing can bypass the prediction of the effect of these variables, optimizing therapeutic efficacy, and minimiz- ing potential adverse effects.

1.1.Historical discovery of ganciclovir
Ganciclovir was first discovered in 1980 and subsequently emerged as the first high potency anti-CMV agent in 1989 [6]. Prior to official approval by the U.S. Food and Drug Administration [7], it was first used in 1984 through compas-

CONTACT Su Ann Ho [email protected] Departments of Infectious Diseases, The National Centre for Infections in Cancer, Peter MacCallum Cancer Centre, Melbourne, Vic 3002, Australia
© 2020 Informa UK Limited, trading as Taylor & Francis Group

2.Ganciclovir and valganciclovir

Article highlights
● The activity of ganciclovir is determined by the susceptibility of the specific viral strain in the individual patient and intracellular concen- tration of the active compound.
● Ganciclovir dosing algorithms result in highly variable interindividual concentrations despite recommended adjustments according to body weight and renal function.
● There is little consensus on a therapeutic range which correlates with clinical outcomes and toxicity, with individual institutions extrapolat- ing results from either in vitro studies or earlier studies performed in AIDS patients and the renal transplant group.
● Ganciclovir TDM may be beneficial in select patient groups, namely, those with highly variable pharmacokinetic profiles such as patients with fluctuating renal function and the pediatric population, and to minimize the risk of drug-related hematological toxicity.
● Future studies should investigate novel approaches to ganciclovir TDM with more specific measures of antiviral activity such as intra- cellular ganciclovir concentrations, host CMV specific T-cell functional immune markers and pharmacogenomic screening to provide more accurate reflection of the anti-CMV activity.

sionate access, in a bone marrow transplant patient with CMV pneumonitis then subsequently in an AIDS cohort with CMV retinitis [6,8,9]. At that stage, there were only in vitro and preliminary animal data supporting its anti-CMV effects, and interestingly the recommended dose at that time remains until today [8].

1.2. Efficacy of ganciclovir and valganciclovir in CMV infection
These antiviral drugs have led to a significant reduction of early CMV disease from 10 to 40% historically [10,11] to, 3 to 6% in the allogeneic HCT cohort [4]. As a result, CMV disease- related mortality is significantly lower today (<2%) compared with the 70% mortality in previous reports of patients with CMV pneumonitis [11,12]. Current CMV management strate- gies in seropositive patients include antiviral chemoprophy- laxis or preemptive therapy [11,13]. A universal prophylaxis approach aims to administer antiviral drug therapy to all at- risk HCT where the donor or recipient is CMV seropositive patients based on pretransplant serology, prior to detection of CMV virus [11,14,15]. On the other hand, preemptive ther- apy requires close surveillance of CMV DNA in blood with initiation of treatment when CMV DNA or antigen thresholds are detected with increments beyond predetermined viral load criteria [11,14]. The use of universal prophylactic ganci- clovir increased the incidence of late CMV infection occurring beyond the 100-day post-transplant period [11,14] with increased incidence of CMV pneumonia [16] which has a high mortality and a high nonrelapse mortality.
Both ganciclovir and valganciclovir have similar efficacy in the clearance of CMV viremia and safety profile [3,17–19]. They can be used interchangeably as primary preemptive strategy in patients with no gastrointestinal (GIT) absorption issues [11]. In fact, there is evidence supporting valganciclovir use in allogeneic HCT recipients with stable mild to moderate graft versus host disease of the GIT [20].
2.1.Mechanism of action
Ganciclovir is a guanosine analog that is bioactivated in CMV infected cells [21]. It is phosphorylated by viral thymidine kinase to the monophosphate form in a rate-limiting step, after which cellular enzymes convert it to ganciclovir tripho- sphate, the active form which exerts the viral inhibitory effect [22–24]. The triphosphate form competitively inhibits viral DNA polymerase [24,25] and it is also incorporated into the growing viral DNA chain, markedly slowing down replication rather than terminating the chain [21,26]. Animal studies demonstrate ganciclovir does not eradicate CMV, but func- tions as a virustatic agent, with viral DNA synthesis resuming once ganciclovir is removed [22,24,27,28]. It is also highly specific for infected cells, with minimal levels of the tripho- sphate form found in uninfected cells [21,22,24,27,29].

2.2.Overview pharmacodynamic properties
The antiviral activity of ganciclovir has been examined in in vitro cell cultures and by convention it is expressed as the concentration required to inhibit CMV replication by 50% (IC50) [30]. There is a wide range in IC50 values depending on the type of isolate, with the reference laboratory strain AD169 mean being 0.9 mg/L while clinical isolates range from 0.2 to 3.48 mg/L [29,31–34].

2.3.Overview of pharmacokinetic properties
Valganciclovir is a L-valyl ester prodrug of ganciclovir, which was developed to overcome the poor oral bioavailability of the par- ent drug [35,36]. It has superseded oral ganciclovir in today’s practice due to significant improvement of bioavailability (up to 10-fold) and the ease of administration with the present formula- tion [35–37]. The moderately high bioavailability of valganciclovir (60%) is related to the recognition of the prodrug as a substrate by the intestinal peptide transporter PEPT1 [37]. It is subse- quently rapidly metabolized to ganciclovir by intestinal and hepatic esterases [20] with minimal concentration of valganci- clovir present in circulation present after absorption [37].
Key PK properties are depicted in (Table 1): briefly ganci- clovir is administered as a 1-h infusion with peak concentra- tions varying linearly, with no accumulation observed with conventional dosing provided renal function is normal [38]. The administration of intravenous ganciclovir 5 mg/kg results in a peak serum concentration (Cmax) of approximately 10.0 mg/L [21] compared to a Cmax of 6.6 mg/L after valganci- clovir 900 mg [37]. The lower Cmax for the oral formulation is expected due to PK limitations with absorption. However, numerous studies have demonstrated serum concentrations of these agents are at least equivalent at their respective dosages [20,39] due to higher systemic exposures of valganci- clovir in HCT recipients [40,41].
In vitro studies have demonstrated the intracellular active form persists in infected cells for extended periods of up to 18 hours [23,24] despite removal of the parent drug from the culture med- ium. The complex dynamic interplay between viral replication and

Table 1. Basic pharmacokinetic properties of ganciclovir and valganciclovir (24, 37, 39, 45, 69, 70).
Parameter Ganciclovir Valganciclovir

Formulation
500 mg/10 mL vial (Cymeve®)
450 mg tablets (Valcyte®)
50 mg/mL powder for oral solution (Valcyte®)

Treatment dose in normal renal function iv. 5 mg/kg twice daily p.o. 900 mg twice daily
Maintenance dose in normal renal function 5 mg/kg daily 900 mg daily
Absolute oral bioavailability of ganciclovir Not applicable 60 to 80%
High fat meal increases systemic exposure by 30%
Volume of distribution 30 to 70 L Rapidly hydrolyzed to ganciclovir Tissue concentrations

Vitreous fluid Lung
Liver Testes
Cerebrospinal fluid
44 to 65% of serum concentration Same as serum concentration Same as serum concentration Same as serum concentration
24 to 67% of serum concentration

Protein binding 1 to 2% Nil significant as rapidly hydrolyzed to ganciclovir

Half life
3 hours (plasma)
13 hours (vitreous fluid)
0.47 hours, rapidly hydrolyzed to ganciclovir

Elimination 90 to 100% unmetabolised in urine within 24 hours
Metabolism Nil significant Only 1 to 2% VGCV detected in plasma after oral administration

host/viral cellular enzymes in the formation of ganciclovir phos- phorylated metabolites [37] suggest, systemic exposure as repre- sented by area under the concentration-time curve [36] may be a more reliable surrogate measure of clinical activity.
There are PK differences between transplant patients com- pared to healthy and HIV subjects, peak concentrations for valganciclovir take twice as long to be achieved at 3 hours instead of 1.6 hours [37,42]. In addition, systemic exposure for both the intravenous and oral formulations are twice as high in the transplant group as well [37]. PK studies have shown ganciclovir is primarily eliminated through the kidneys via glomerular filtration and tubular secretion, with 85 to 100% recovered unchanged in urine when renal function is normal [22,25,37,43]. This probably explains the higher exposure of ganciclovir achieved in transplant patients where studies have demonstrated altered renal clearance of the drug [44,45].

2.4.Toxicity
Ganciclovir-related toxicity has been difficult to evaluate due to several confounding factors, namely the patient’s under- lying immunocompromised state likely to contribute to mea- surable physiological changes [38]. In addition, specific adverse effects attributable to the drug can be difficult to define. For example, concomitant medications such as myco- phenolate and trimethoprim-sulfamethoxazole and concurrent infections potentially contribute to myelotoxicity and nephro- toxicity and therefore be confounding factors [9,37].
Nevertheless, the most significant adverse effect of these agents is hematological toxicity with neutropenia being the most commonly reported (up to 90%) [22,43,46] followed by thrombocytopenia and anemia being less common [38]. Ganciclovir induces neutropenia by dose-dependent inhibition of DNA polymerase in hematopoietic precursors [47,48]. The incidence of myelosuppression occurred more frequently in HCT recipients than other transplant groups [12], with studies suggesting a reversible exposure-dependent effect [49–53].
Neurotoxicity has been reported in 5% of patients in early trials of ganciclovir and published case reports, with a broad
range of signs from headaches, confusional states, seizures to psychosis with hallucinations [22,43]. Central nervous system (CNS) adverse effects appear to be related to high plasma and CSF ganciclovir concentrations in the setting of intravenous administration and renal impairment, however there is no known specific toxic range [22,37]. Rare cases of hepatotoxicity have also been associated with intravenous ganciclovir [37].

3.Ganciclovir and therapeutic drug monitoring
TDM has a well-established role in dose optimization of certain antimicrobial agents such as aminoglycosides and vancomycin due to a known narrow therapeutic window and risk of nephrotoxicity. This is, however still a contentious issue in the prescribing of ganciclovir and valganciclovir. Currently, there are no data correlating in vitro susceptibilities with in vivo pharmacokinetic parameters to provide a defined tar- get PK/PD ratio for these antivirals [37]. PK studies have shown total body clearance of ganciclovir is correlated with creatinine clearance (CrCl) [32]. Some have argued, because of this pre- dictability, TDM is unnecessary [25,32,37]. The tolerability pro- file of ganciclovir has been difficult to assess, given recipients are immunocompromised with a degree of disease contribut- ing to myelosuppression, in addition to potential concurrent nephrotoxic agents. However, neutropenia has been consis- tently reported in most published studies and appears to be dose-related which resolves on cessation of the drug [53]. The rates of myelosuppression are approximately 20 to 50% during induction therapy [38,46,54] resulting in the need for cessation of the drug or concomitant administration of granulocyte colony-stimulating-factor.

3.1.Associations of ganciclovir concentration with clinical outcomes
3.1.1.Allogeneic HCT recipients
To date, the data on the clinical utility of ganciclovir TDM is mixed and is based primarily on a handful of small studies [12,55–57]. Early studies in AIDS patients on ganciclovir for

CMV retinitis had demonstrated the measured trough concen- trations of 0.6 mg/L predicted for disease relapse or progres- sion [58–60]. However, this has not been consistently reproduced in the transplant setting [56,61]. In addition, there is no consensus on the target concentration range or which PK exposure best correlates with clinical efficacy of ganciclovir. This issue remains unresolved in the allogeneic HCT cohort where many individual centers depend on the conventional renal-adjusted dosing schedule. Patients who do not respond to ganciclovir often have their treatment changed to alternative agents which are potentially more toxic, as clinicians have no alternative method of optimizing ganciclovir dosing [62,63].
Various older and small studies over the last three decades have sought correlation between ganciclovir concentrations and its systemic exposure (described as area under the curve) with clinical efficacy. One of the main aims in earlier studies was to examine whether conventional dosing correlated with in vitro CMV minimum inhibitory concentration (MIC) and IC50 and the associated clinical outcomes [32,64]. Early clinical studies on ganciclovir included HCT recipients within a heterogeneous group of immunocompromised patients (Table 2) [12] with severe CMV infection and end-organ dis- ease. There was no clear relationship demonstrated between peak and/or trough concentrations of ganciclovir and clinical response, although the level of exposure was mostly above the IC50 of known CMV strains.
In contrast, a prospective observational study in 39 patients, 15 of whom were bone marrow transplant recipients (Table 2) [65], showed ganciclovir was effective with 72% having clinical improvement and elimination of CMV from cultures. The reported mean trough concentration of 1.4 mg/L and peak of 11.5 mg/L were associated with an overall good response to ganciclovir. Patients with CMV pneumonitis had more treatment failures, only half clinically improved. Neutropenia occurred in one-third of patients; however, there was no relationship with levels in blood. This study had several limitations with only half of patients having serum concentrations collected and no corre- sponding details on specific outcomes, in addition there was a large range within the measured trough and peak levels of up to 10-fold.
A recent study looked at ganciclovir trough plasma concen- trations with clinical outcomes in 13 HCT recipients on preemp- tive treatment (Table 2) [57]. Trough plasma concentrations did not reliably predict clearance of viremia, as three responders had undetectable trough concentrations. The small sample size, lim- ited PK pharmacokinetic parameters measured and absence of viral load kinetics post-treatment, precludes a more systematic analysis of ganciclovir concentrations.
The largest clinical study of ganciclovir TDM in HCT was a retrospective analysis of 82 immunocompromised hosts where 33 were allogeneic HCT recipients (Table 2) [55]. In this heterogeneous group, there was no association between serum peak or trough concentrations with clearance of vire- mia. It is worth noting that, the institution reference range spanned a wide range with target peak concentration 3.0 to 12.5 mg/L and trough 1.0 to 3.0 mg/L. TDM was done in selected cases at the discretion of the treating clinician which introduces case selection bias. Dose adjustment due

to poor response and off-target serum concentration occurred in one-fifth of patients, however clinical outcomes of this group were not reported.

3.1.2.Ganciclovir TDM studies in other patient populations
A majority of more recent ganciclovir TDM studies have been conducted in SOT recipients, where ganciclovir prophylaxis starts immediately post-transplant. In a randomized, double- blind, multicenter PK study of 372 patients comparing oral GCV and VGCV for prophylaxis in SOT [36], ganciclovir sys- temic exposure, measured by area under the curve over a time period was analyzed. An AUC of more than 45 mg.h/L was associated with a better antiviral activity with a low incidence (3%) of breakthrough viremia during prophylaxis management as compared to a lower mean AUC of 28 µg.h/L where there was 10% breakthrough viremia. This higher systemic exposure was found to be superior in suppressing CMV viremia up to 4-months post-transplant.
A prospective study in SOT recipients assessed the utility of a renal-based dosing nomogram in 69 patients with 25 patients receiving treatment and the others prophylaxis [31]. This series found serum concentration did not correlate with clinical efficacy nor did it have a predictive value in determin- ing patients who relapse or develop recurrent infection [31]. This would correlate with ganciclovir’s function as a virustatic agent rather than eradicating the virus [24,27,64]. Another common observation is that infection recurs in the presence of continued immune incompetence [31,56,65]. Similarly, another prospective study measuring trough levels in renal and liver transplant recipients on valganciclovir as prophylaxis, showed no correlation between ganciclovir levels and level of viremia [66].
There are however, reports on the utility of ganciclovir TDM in select group of patients who have suboptimal response to treatment [61]. In a cohort of 51 lung transplant recipients, 11 patients had persistent viremia after 2 weeks of treatment, five of them were discovered to have ganciclovir-resistant infec- tion, all of whom had consistently low trough levels of less than 1.1 mg/L.

3.2.Associations of ganciclovir concentration with toxicity
In vitro toxicology studies on ganciclovir have demonstrated high concentrations up to 100-fold that of antiviral activity are required to inhibit proliferation of uninfected host cell lines [24,37]. The exception to this, is hematopoietic progenitor cells, where toxic effects are observed at concentrations simi- lar to those required to inhibit CMV. Granulocyte-macrophage progenitors appear more susceptible compared to the ery- throid cell lines [37]. Following intravenous administration of conventional ganciclovir doses, peak plasma concentrations are generally higher than the hematopoietic toxic thresholds, which suggests that a degree of bone marrow suppression is likely with this regimen [38].
Numerous studies investigating the relationships between sys- temic ganciclovir concentrations and myelosuppression have not

consistently found reliable correlations [12,31,55,57,65,67]. It is worth noting these studies had collected limited blood samples which corresponded to either serum ganciclovir peak and/or trough concentrations. Two large prospective pharmacodynamic studies in more than 300 SOT recipients found a trend of increased incidence of leukopenia and neutropenia in patients with higher systemic exposures of ganciclovir as measured by AUC [36,68]. There was a modest 5–10% increase in incidence of neutropenia and leukopenia when ganciclovir AUCs were increased by two- thirds [36]. Earlier studies indicated the development of neutro- penia is associated with the total dose of ganciclovir administered, usually occurring before a total cumulative dose of 200 mg/kg [38]. This adverse effect is reversible on cessation of the drug, with evidence of recovery within seven days of the treatment with- drawal [25].

4.Current practice for optimizing the efficacy of val/
ganciclovir allogeneic HCT recipients
4.1.Dosing algorithms
Currently, clinicians rely on renal- and weight-based dosing algorithms for ganciclovir and valganciclovir which are then combined with quantitative monitoring of CMV PCR levels, to assess for clinical effect. However, this approach may vary across different institutions depending on which reference nomogram is adopted (Table 3) [69–72]. The main rationale for this approach is to ensure there is adequate clearance of the drug, in order to maintain a similar systemic exposure to patients with normal renal function [37,44,73]. These differing dosing algorithms highlight the role of TDM in standardizing antiviral exposures in individuals to ensure optimal efficacy with minimal toxicity (see Table 3).
Another potential confounding factor, is the different methods of calculating renal function in clinical practice. Measured creatinine clearance (by 24-h urine collection) is challenging, instead surrogate measures such as serum crea- tinine levels and prediction formulas (based on a combination of patient age, sex, weight, ethnicity, serum creatinine level) are commonly used in routinely [74,75]. Some institutions use the readily available electronic estimates of glomerular filtra- tion rate (eGFR) which is based on the Modification of Diet in Renal Disease (MDRD) formula. A prospective study of SOT recipients [66] found eGFR underestimated measured creati- nine clearance (24-h urine collection) by more than 20% in two-thirds of patients leading to underdosing of valganciclovir in half of the cohort [66]. This study demonstrated the Cockroft-Gault equation was able to provide a 95% accuracy for estimating renal function. In addition, assessments of [36]

renal function utilizing ideal body weight will likely incorrectly predict ganciclovir dosing regimen [76,77].
Population PK studies have identified select groups of patients who will benefit from ganciclovir TDM [55,78–80]. Up to half of critically ill patients with renal impairment (eGFR<50 mL/min) were at risk of subtherapeutic ganciclovir exposure levels, when receiving ganciclovir based on standard dosing nomograms [79]. There was also a high degree of plasma concentration interpatient variability not fully accounted by differences in eGFR. Other potential covariates such as volume of distribution and the role of active tubular secretion of ganciclovir were proposed [79]. Another vulner- able group are patients on different modalities of renal repla- cement therapy, which contribute to changes in the antiviral PK parameters [81]. System-specific factors such as filter mate- rial, membrane type, in conjunction with haemofiltration and dialyzate rates are some of the confounders to changes in drug clearance [80].
Observational studies in the pediatric age group have also demonstrated wide variability of exposure levels, especially in neonates where there is rapid physiological changes in renal function [62,82,83]. Numerous dosing regimens have been used in clinical practice [84–86], however drug exposures have been inconsistent across the full pediatric age and renal function range. Some studies have demonstrated val- ganciclovir dosing based only on body surface area (BSA) resulted in low concentrations in infants and young children, similar findings were seen in ganciclovir, when standard adult dosing of 5 mg/kg was administered [62,83] groups. Recently, a model-based dosing algorithm for ganciclovir based on BSA and creatinine clearance according to the Schwarz formula has been implemented in the pediatric population [87]

4.2.PK parameters utilized for dosing titration
Traditional parameters commonly measured and referenced as a target range for ganciclovir TDM are serum peak and trough concentrations. PK studies have demonstrated that both of these levels achieved with ganciclovir exceeded the IC50 referenced from laboratory and clinical CMV viral strains which generally ranges from 0.1 to 2.8 mg/L [21,25,32,33]. A PK/PD study of oral ganciclovir in AIDS patients with CMV retinitis demonstrated that area under the concentration-time curve was superior in predict- ing clinical efficacy compared to Cmax and Cmin [88,89]. Such comparative studies between the different PK parameters have not been conducted in transplant patients.
Of course, plasma ganciclovir concentration remains a surrogate of the active intracellular triphosphorylated molecule

Table 3. Different induction phase dosing nomograms for ganciclovir: Dosing regimen A (70), dosing regimen B (72).

Ganciclovir dosing regimen according to renal function (A)
Creatinine clearance (mL/min)
(B)
Creatinine clearance (mL/min) [Serum creatinine (µmol/L)]

5 mg/kg 12 hourly >50 ≥70 [<125]
2.5 mg/kg 12 hourly 25 to 50 50 to 69 [125 to 175]
2.5 mg/kg 24 hourly 10 to 25 25 to 49 [176 to 350]
1.25 mg/kg 24 hourly <10 10 to 24 [>350]
1.25 mg/kg THREE times weekly post hemodialysis < 10 [>350 (and on hemodialysis)]

which is responsible for drug efficacy [21,60]. A pioneering study measuring intracellular ganciclovir and its metabolites in a group of kidney transplant recipients on prophylaxis valganciclovir, indi- cated a significant association between the accumulated levels of the triphosphate metabolite form with neutropenia [54]. Interestingly, this study also demonstrated no correlation between intracellular ganciclovir and the plasma exposure indices [54].

4.3.Dosing titration according to model-based predictions
A consistent finding from PK pharmacokinetic studies is the presence of large interindividual variation of ganciclovir expo- sures [55,62,76,90]. Apart from the contribution of differences in renal clearance, body weight and sex have been described to predict differences in volume of distribution in a population PK pharmacokinetic study [91] of valganciclovir and oral gan- ciclovir. However, despite correcting for these covariates, there remains a large unexplained interpatient variability of up to 28% [91]. Serum concentration variability is likely to impact on clinical efficacy, as evidenced by studies demonstrating sub- therapeutic levels leading to the development of resistant CMV disease [77,92].
Here, Bayesian forecasting is likely to demonstrate an improved capacity for achieving therapeutic targets compared to conven- tional dosing nomograms [93,94]. This strategy relies on use of population PK pharmacokinetic data for the specific target cohort, which is presently lacking in the allogeneic HCT group. In the pediatric population, model-based approaches have been com- monly used in drug dosing to bridge data between adult and pediatric groups [87]. A method of combining physiologically based pharmacokinetic modeling (PBPK) with population PK data, has provided more confidence in establishing these antiviral agents dosing regimens for neonates and infants [95]. This strat- egy has made some key changes in establishing a new recom- mended dosing schedule for ganciclovir and for valganciclovir, selecting the best-fit schedule across all ages groups [84,87].

5.Other considerations for optimization of ganciclovir in allogeneic HCT recipients
5.1.Preclinical data
The dosage recommendation for ganciclovir is based on in vitro studies of different strains of CMV resulting in a wide range of IC50 values [37,62]. It has been suggested that a more robust pharma- codynamic standard such as IC95 should be utilized as complete viral suppression is the ultimate goal of therapy [30]. Current guidelines recommend a CMV preemptive strategy where fre- quent monitoring of CMV using nucleic acid testing (NAT) in blood or plasma [11] triggers the commencement of antiviral treatment. The time-consuming process of viral culture and mea- suring IC50 of each isolate is not required [96]. Ganciclovir is utilized empirically, with genotypic antiviral resistance mutations (UL54, UL97) screened for when the individual does not respond to treatment after a period of appropriate treatment [96]. In contrast, pharmacokinetic-pharmacodynamic (PK/PD) denominators of bacterial infections are routinely confirmed by culture and suscept-

ibility, thereby providing predictions of efficacy of the selected antimicrobial.
To date, there are no clear data that demonstrate pharma- codynamic parameters such as in vitro susceptibility values, in vivo ganciclovir serum levels or time over IC50 predicts clinical efficacy [31,37]. Importantly, the host-CMV specific T-cell immunity and T-cell reconstitution [97] plays a role in the overall clinical efficacy of ganciclovir. A study looking at the CMV replication dynamics supported this concept whereby a high antiviral activity (IC95) is required to eliminate viral growth in a CMV naive transplant patient as opposed to an experienced CMV immune host [98].

5.2.Host immune status
Newer trends in allogeneic transplantation have increased the proportion of vulnerable patients to CMV infection. For example the development of new immune therapies such as ruxolitinib and the availability of reduced-intensity conditioning in stem cell transplant. The number of HCT recipients aged 60 and older has tripled over the last two decades [11], while recipients ≥70 year represented 6% of allogeneic transplants for acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) [99]. The effect of immunosenescence in this relatively new group of older transplant patients has not been quantified but needs to be considered as well. Wider accessibility to stem cell transplant in countries such as Latin America and Asia, where CMV seroprevalence is higher also increases the likelihood of CMV infection in transplanted patients. In addition, there is a greater availability of unmatched transplantation where donors are generally younger and are CMV seronegative. These factors in combination contribute to more HCT recipients who are at risk for CMV infection.

6.Conclusion
There is a paucity of large, well-conducted clinical data evaluating ganciclovir TDM in the allogeneic transplant group. International guidelines in SOT and hematological malignancies [11,100] have identified this issue as an area of need. Recently, there has been a large focus on newer novel anti-CMV drugs such as letermovir. However, case reports of [101,102] letermovir resistance highlight the importance of optimizing our available antivirals, like val/
ganciclovir which remain first-line treatment options.
There is much work to be done from redefining fundamen- tal issues of in vitro inhibitory concentration thresholds, clinical pharmacodynamic studies that correlate efficacy with drug exposure. As such, current evidence indicates ganciclovir TDM is highly likely to be useful in patients with fluctuating renal function and at high risk of bone marrow suppression and/or in patients not responding to treatment as expected, as well as the pediatric population. Future studies identifying these specific patient populations would ensure ganciclovir TDM is utilized effectively and efficiently. A much anticipated role of TDM in this area would be the minimization of myelo- toxicity risk associated with these agents, and therefore

improving accessibility to wider cohort of patients, reducing the premature cessation of treatment and preserving alterna- tive anti-CMV agents for refractory and resistant cases.

7. Expert opinion
To date, ganciclovir TDM has been a passive process, mainly focused on achieving drug exposures in patients that match in vitro inhibitory thresholds. Whilst most institutions and inter- national consensus guidelines do not recommend routine TDM, they also identify this as an important area for future research. There are three key research areas that will improve the clinical application of ganciclovir TDM in order to maximize antiviral activity and minimize the likelihood of drug toxicity by indivi- dualized dosing. These include exploring more precise measures of val/ganciclovir anti-CMV activity along with its correlation with outcomes and toxicity, and the incorporation drug concentra- tions with the interplay between CMV replication dynamics and host immunity (Figure 1).
Firstly, the measurement of the active intracellular form of ganciclovir will likely provide a more accurate prediction

of clinical efficacy. Conventional methods of measuring plasma concentrations of this agent have yet to provide strong guidance to clinicians for a TDM target for dose optimization. If studies can characterize the PK of intracel- lular ganciclovir triphosphate, it will enable adjustment of current val/ganciclovir dosing regimens in order to achieve more robust therapeutic ranges. Studies looking at CMV viral dynamics during ganciclovir treatment have revealed a unique interplay between drug activation, virus replica- tion, and subsequent control of replication (81). The distinc- tive patterns of viral load decays ranged from a transient rebound pattern, rapid decline to a slow rate of decline, viral kinetics could potentially be utilized to optimize gan- ciclovir dosage regimens. A novel approach of including CMV viral load kinetics in a PK/PD model can potentially provide an individualized dosing with tailored antiviral activity efficacy for each replicative phase.
Up to a third of allogeneic HCT recipients who develop detectable CMV viremia experience spontaneous clearance, a method to predict and risk stratify this group of patients will enable a more focused management for vulnerable patients which would include ganciclovir TDM. Given host

Figure 1. Host, CMV virus and val/ganciclovir interactions.

immune status plays an integral role in this opportunistic infection, methods to track cellular immune recovery by way of phenotypic and functional immunophenotyping uti- lizing flow cytometry and diagnostic assays such as Quantiferon-CMV® can be incorporated into a predictive model as means to identify patient groups for ganciclovir TDM. This method could also play a part in the interpreta- tion drug concentrations in individual patients whether higher end of therapeutic range should be targeted in those who are slow to respond to treatment.
TDM also aims to minimize toxicity, here, more specific markers such as intracellular ganciclovir triphosphate can be considered as means to predict neutropenia in patients on val/ganciclovir. Pharmacogenomics is becoming an integral partner to TDM as it can provide information on the impact of genetic variations of interindividual differences in drug disposition and toxicity allowing earlier prediction of dosing needs. Intracellular transportation of ganciclovir has not been fully defined; however, there is evidence that a membrane transporter – multidrug resistance-associated protein 4 (MRP4) is involved in the efflux transfer of ganci- clovir metabolites (82). Genetic polymorphism of genes encoding this transporter can contribute to the accumula- tion of the triphosphate form and risk of developing neu- tropenia (83). In addition, individuals who have an excess of MRP4 may be at risk of developing refractory CMV infection and subsequently resistance.
As we move forward in the era of precision medicine, novel approaches have to be considered in the effort to ensure applic- ability and transferability of ganciclovir TDM into clinical practice. Fundamental changes to conventional TDM practices by adopt- ing an integrative method to include CMV-host immunity factors, host pharmacogenomics influences, with ganciclovir PK/PD fac- tors, and enhancement of these agents in vitro inhibitory con- centration targets will contribute to the overall goal of optimizing these agents. Looking more broadly, these proposed principles of TDM could potentially be extended to the other anti-CMV agents as means of ensuring efficient use these finite agents.

Acknowledgments
J.A. Roberts would like to acknowledge funding from the Australian National Health and Medical Research Council for a Centre of Research Excellence (APP1099452) and a Practitioner Fellowship (APP1117065). The authors wish to thank Dr Nyein Chan Aung for his illustration.

Declaration of interest
The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

Reviewer disclosures
Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

ORCID
Monica Slavin http://orcid.org/0000-0002-8443-314X
Jason A. Roberts http://orcid.org/0000-0001-6218-435X

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