Currently, there are two common strategies used for cytomegalovirus (CMV) prevention in organ transplant recipients: prophylaxis and preemptive therapy. In a prophylaxis strategy, antivirals are commenced early posttransplant and continued for a defined period of time such as 3 months. In a preemptive strategy, patients are monitored at regular intervals (e.g., weekly for the first 12 weeks) using a molecular diagnostic assay. In patients with early evidence of viral replication, treatment is initiated to prevent the development of CMV disease. Both strategies have their advantages and disadvantages. However, in the highest risk group for CMV (donor seropositive and recipient seronegative [D+/R−]), current guidelines suggest that prophylaxis may have advantages over a preemptive strategy (1, 2).
One of the main limitations with a standard 3-month course of prophylaxis is the occurrence of late-onset CMV disease, defined as CMV disease occurring beyond 3 months posttransplant often shortly after the discontinuation of antiviral prophylaxis. The reported incidence of late-onset disease is variable but ranges from 17% to 37% in D+/R− patients receiving 3 months of prophylaxis (2). In this group, disease usually develops between 3 and 6 months posttransplant. Several strategies have been proposed to prevent late-onset CMV disease. These include prolongation of antiviral prophylaxis or use of serial molecular surveillance after the discontinuation of prophylaxis. The latter strategy involves use of regular viral load or antigenemia monitoring after discontinuing antiviral prophylaxis. Patients with viral replication above a certain threshold are then treated preemptively.
This latter strategy is also sometimes referred to as a hybrid prophylaxis/preemptive therapy approach. Its purpose is specifically to provide the potential benefits of prophylaxis and yet prevent the problem of late-onset CMV disease commonly observed with prophylaxis. Clear guidelines do not exist with respect to exactly how such a strategy should be carried out (target patient population, type of test, threshold, period of surveillance, or intensity of surveillance). In addition, there are limited data on the efficacy of this approach in D+/R− patients. Our center has been using this hybrid prevention in high-risk patients. We performed once-weekly viral load testing for 8 weeks after the completion of prophylaxis. Protocol-defined preemptive therapy is then initiated based on predefined thresholds of viremia. The purpose of this study was to evaluate the efficacy of this hybrid approach for the prevention of late onset CMV disease.
A total of 83 CMV D+/R− patients were identified; however, 12 patients were excluded from analysis (no antiviral prophylaxis [n=2] and no postprophylaxis molecular monitoring [n=10]). The demographic characteristics and immune suppression regimens of the patient population are listed in Table 1. Median age at the time of transplant was 51 years (range 3–72 years). Antiviral prophylaxis lasted a median of 98 days (interquartile range 96–117 days). It consisted of intravenous ganciclovir for 1 to 2 weeks followed by oral ganciclovir (during 2004) or valganciclovir (from 2005 onward) using standard doses adjusted for renal function.
CMV Viremia and Disease
CMV viremia (>500 copies/mL) occurred in 37 of 71 (52.1%) patients during the first-year posttransplant. Of the viremic patients, 19 of 37 patients had viremia first detected during the postprophylaxis surveillance period, and 18 of 37 patients had their first detectable viremia only after surveillance (measurement performed for assessment of symptoms). Symptomatic CMV disease occurred in 29 of 71 (40.8%) patients during the first-year posttransplant. CMV disease occurred during the surveillance period in 13 of 29 (44.8%; 2 tissue-invasive and 11 viral syndrome) patients and after the surveillance period in 16 of 29 (55.2%; 1 tissue-invasive and 15 viral syndrome) patients. One patient with tissue-invasive CMV disease was diagnosed at autopsy 97 days after completion of the postprophylaxis surveillance period and had no viral load investigation during the hospital admission. Median time to CMV disease was 214 days (range 93–356 days) posttransplant. All episodes of disease (except one diagnosed at autopsy) were treated with ganciclovir or valganciclovir.
Clinical Utility of Surveillance
Approximately half of the viremia episodes during the first-year posttransplant were detected during the 8-week surveillance period postprophylaxis (n=19/37, 51.3%). The first positive viral load was after a median of 35 days from the beginning of the surveillance period (range 11–53 days). The median first detectable viral load during surveillance was 10,250 copies/mL (range 1620–2,666,500 copies/mL). The median peak viral load observed within this group was 100,550 copies/mL (range 1985–2,666,500 copies/mL). On the basis of routine surveillance, preemptive therapy was successfully initiated before the development of symptoms in three patients (two liver transplant and one kidney transplant). An additional three patients (all liver recipients) had asymptomatic low-level viremia, which spontaneously cleared without reaching the threshold for initiation of preemptive therapy. In the remaining 13 patients with detectable viremia during the surveillance period, patients either had symptoms (CMV disease) at the time of first detectable viremia (n=7) or became symptomatic between the first positive sample and subsequent sample but before the initiation of preemptive therapy (n=6). Virologic and sampling characteristics of each patient subgroup are summarized in Table 2.
A significant proportion of patients developed symptomatic CMV disease only after the end of the 8-week postprophylaxis surveillance period (n=16/29 total disease cases). The median time to presentation of this late disease was 64 (range 9–180) days after the completion of the surveillance period. Viral load at disease onset was 39,830 copies/mL (range 3095–16,700,000 copies/mL).
Kinetic Parameters of Viral Doubling
In six patients, rapid increases in viral load during the surveillance period meant that preemptive therapy could not be started in time to prevent CMV disease. Plotting of viral loads over time before treatment initiation allowed calculation of the in vivo viral doubling time. All six patients had an exponential increase in viral load over time. An example of three of these patients is shown in Figure 1. The median doubling time before the start of therapy for these six patients was 1.10 days (range 0.91–1.46 days) reflecting rapid viral growth.
Compliance to Postprophylaxis Surveillance
We also evaluated the compliance to postprophylaxis surveillance. Overall, during the 8 weeks after the discontinuation of the antiviral prophylaxis, a median of six samples were collected at a median interval of 8 days (mean 8.64, range 3.5–34 days). We analyzed a best-case scenario of how many cases of disease could have been prevented if compliance with weekly monitoring for 8-week postprophylaxis was 100%. At best, an additional 3 of 29 (10.3%) cases of CMV disease may have been prevented through enhanced compliance.
Analysis by Organ Type
An analysis of the utility of postprophylaxis surveillance was carried out for each organ transplant type and is shown in Table 3. For example, in the 29 D+/R− kidney (or kidney/pancreas) recipients, CMV viremia occurred in 12 (41.4%) and CMV disease in 9 (31.0%). One patient benefited from preemptive antiviral therapy, whereas in most cases, viremia and disease occurred only after the 8-week surveillance period (Table 3). Similar results were observed in liver transplant recipients. Of note, the highest CMV disease rate was observed in lung (including heart-lung) transplant recipients (10/12 [83.3%]), and in none of these patients was preemptive therapy successfully deployed.
Current guidelines recommend antiviral prophylaxis as the preferred strategy for CMV prevention in D+/R− transplant patients. However, after the discontinuation of prophylaxis, the occurrence of late-onset CMV disease is a major problem. The use of a hybrid strategy where preemptive monitoring is used after prophylaxis has been proposed as a way to prevent late-onset CMV disease (1); however, data assessing the utility of hybrid prevention strategies are limited. In this study, we demonstrate in a moderately large cohort of D+/R− transplant recipients that a number of limitations exist to this hybrid approach. A significant portion of disease developed only after the 8-week surveillance period. Preemptive antiviral therapy was successfully administered in only three patients, and 13 of the 71 patients still developed late-onset CMV disease during the surveillance period. Therefore, the hybrid approach was not fully effective because of a combination of factors including rapid viral doubling time, occurrence of disease after the monitoring period, and use of a high threshold for preemptive therapy. An analysis by organ subtype demonstrates that these limitations exist despite the transplanted organ (Table 3).
There are limited data assessing this hybrid prophylaxis/ preemptive strategy in D+/R− transplant patients. Boillat Blanco et al. (3) assessed a similar strategy in 30 D+/R− kidney transplant patients and showed that CMV disease occurred simultaneous to the first positive viral load in five of seven patients. In an analysis of the PV16000 database, which included 364 non-lung D+/R− patients who received 3 months of antiviral prophylaxis, the utility of monitoring for predicting late-onset disease was assessed. Monitoring was performed at the time of discontinuation of prophylaxis and then every 2 to 4 weeks until month 6 and then at less frequent intervals (4). Overall, of 64 cases of late-onset disease, viral load monitoring was not able to predict CMV disease in 40 (62%) patients. One of the main criticisms of that study was that viral load monitoring was performed at infrequent intervals (every 2–4 weeks). In a study comparing a hybrid strategy (n=10) versus preemptive therapy alone (n=11) after pediatric liver transplantation, no case of CMV disease was observed in either arm (5). In a study of 122 pediatric liver transplant recipients (43 D+/R−), a minimum of 14 days of prophylaxis was followed by surveillance. CMV disease developed in 12 patients (9.8%) during the follow-up, the majority of whom were D+/R−, had detectable CMV viremia, and had rapid development of symptoms after initial detection of viremia (6). The incidence of disease in this study was low likely because no threshold value for treatment was used, and all viremia was treated.
Alternative strategies to prevent late-onset CMV disease include simply using a preemptive strategy as opposed to a prophylaxis approach in D+R− patients or prolonging antiviral prophylaxis beyond 3 months (7–10).
There are potential ways to improve on the efficacy of this hybrid strategy. The first would be to lower the viral load threshold at which therapy is initiated. A lower threshold would likely have prevented some other cases of CMV disease but may result in treatment of low-level viremia that would have spontaneously cleared. Indeed, optimal thresholds for use in preemptive therapy in general have not been established. The recent international CMV consensus stated that no specific threshold could be agreed upon because such thresholds were assay dependent and likely also dependent on the patient population being studied (1). To prevent cases of disease occurring after the surveillance period, an extension of the weekly surveillance period beyond 8-week postprophylaxis may be of benefit but may be difficult in patient compliance and cost. For example, to diagnose all 18 episodes of viremia that were detected after the surveillance period, an extra 26 weeks of testing would be required. In addition, based on rapid viral load doubling times observed, even more frequent monitoring (more than once weekly) may be necessary in D+/R− patients (6) consistent with previous reports assessing viral kinetics in transplant patients (11). The feasibility of intensive monitoring (i.e., sampling more than once weekly) is unclear and may be difficult because of lack of compliance and cost.
Another important issue is that the utility of this strategy may vary across different transplanted organ groups. Different transplant types may have higher or lower risk of CMV. Lung transplant patients in particular seem to be at high risk of late-onset disease (12). In our cohort, lung transplant patients did have a higher rate of viremia and disease. None of the six lung patients with viremia in the surveillance period had preemptive therapy instituted before the development of CMV disease. In patients with the highest risk of postprophylaxis disease, prolongation of prophylaxis may be a more viable option, whereas in specific organ types with lower rates of late-onset disease, postprophylaxis surveillance may be more suitable. The hybrid strategy used in our study seems to be more useful in kidney and liver recipients who tended to clear viremia spontaneously and in whom preemptive strategies were sometimes successful. Further studies in individual organ groups would need to be performed to better define this. Our study has a number of limitations. First, we only evaluated a single threshold for preemptive therapy and a single duration of surveillance. The optimal duration, frequency, and thresholds for use in a postprophylaxis preemptive strategy, therefore, cannot be well defined based on these data. However, the sample size is robust enough to provide valuable conclusions that have not been previously well shown in the literature; that is, for prevention of late-onset disease in D+/R− patients, a preemptive strategy would need longer than 8 weeks of surveillance with low thresholds for initiation of antivirals and possible twice weekly than once weekly monitoring. Another limitation of our study was the retrospective design. However, the viremia endpoints were objective, and the disease definitions based on standard ones recommended for use in clinical studies (2, 13).
In summary, the use of standard viral load surveillance after completing CMV prophylaxis in patients who were D+/R− was associated with only modest benefit, given that it did not prevent CMV disease in a number of patients. The reasons include the use of a high threshold for preemptive therapy, disease occurring beyond the surveillance period, and rapid viral load doubling times. Further studies are needed to evaluate and compare improved hybrid strategies to alternative approaches for prevention of late-onset CMV viremia and disease in D+/R− patients.
MATERIALS AND METHODS
We conducted a retrospective chart review of all CMV D+/R− solid organ transplant recipients at our institution. Patients were identified through a solid organ transplant registry maintained by a study investigator and viral load data from the Provincial Laboratory for Public Health's database. We reviewed transplants occurring at the University of Alberta Hospital over a 4-year period. This represented a time period when a uniform prophylaxis protocol was used in conjunction with a nucleic acid testing for CMV viral load monitoring approach. We included all organ recipients who fulfilled the following criteria: (i) aged older than 18 months; (ii) CMV-seronegative recipient of organ(s) from CMV-seropositive donors; (iii) received antiviral prophylaxis; and (iv) monitored with at least one viral load measurement during the expected surveillance postprophylaxis period. The study was approved by the institutional ethics board.
The institutional protocol was to administer universal antiviral prophylaxis to CMV D+/R− patients before the seventh day posttransplant and until 3 months posttransplant for kidney, liver, heart, and pancreas recipients (including combined organ). For D+/R− lung transplant, combined heart and lung transplant, and small bowel recipients, 6 months of antiviral prophylaxis was administered. Prophylaxis consisted of valganciclovir 900 mg/day or intravenous ganciclovir 5 mg/kg once daily followed by oral ganciclovir 3 g/day (during 2004) or valganciclovir 900 mg daily (2005 onward) upon recovery of oral intake with all doses adjusted for renal function. No molecular surveillance was performed during prophylaxis.
After prophylaxis was discontinued, weekly surveillance during the next 8 weeks was performed using a quantitative real-time plasma-based polymerase chain reaction assay. This time period was chosen based on data from a study comparing oral ganciclovir versus valganciclovir prophylaxis in 364 CMV D+/R− patients (4). In this study, after prophylaxis, a significant proportion of viremia occurred in the first 8 weeks. The lower limit of accurate quantitation was 500 copies/mL (14). Results were available to clinicians within 24 hr of sample receipt in the laboratory. The threshold for initiation of preemptive antiviral therapy was set at 25,000 copies/mL of plasma. This was based on the quantitative relationship between our center's in-house developed assay results and a commercially available quantitative polymerase chain reaction assay used in a natural history study of CMV viral loads that best predicted CMV disease in liver transplant patients seropositive for CMV before transplant (15). In asymptomatic patients with lower levels of viral of replication, viral load was repeated once weekly. If the viral load was between 10,000 and 25,000 copies/mL, it was repeated every 3 days. In patients with greater than 25,000 copies/mL antiviral therapy consisted of either intravenous ganciclovir 10 mg/kg/day or oral valganciclovir 900 mg twice daily, adjusted for renal function, for at least 14 days or greater until viremia was undetectable. Antiviral therapy was also instituted to symptomatic patients regardless of their viral load. There was no standard approach to reduction of immunosuppression, and this was at the discretion of the treating physician. Generally, for patients on triple immunosuppression, the mycophenolate mofetil dose was reduced by 25% to 50%. Further surveillance was also performed at any time posttransplant if lymphocyte-depleting antibodies were used for treatment of rejection.
Outcomes and Analysis
Definitions of CMV disease were comparable with those in the American Society of Transplantation Infectious Diseases Guidelines (2) and American Society of Transplantation Recommendations for Screening, Monitoring and Reporting of Infectious Complications in Immunosuppression Trials in Recipients of Organ Transplantation (13). Patients were followed for 1 year from the time of transplant. Allograft rejection was defined as biopsy-proven and treated, except in lung recipients, in whom rejection could be clinically diagnosed. Descriptive statistics were used for demographics and viral load calculations. Mann-Whitney U test was used for comparison of continuous variables and Fisher's exact test for categorical variables. Viral kinetics analysis (doubling time) for selected cases was performed as previously described (16). Briefly, doubling time calculations require at least three viral load measurements and are performed using a mathematical model to calculate the exponential increase of the viral load before the start of antiviral therapy.
The authors thank Ms. Jayne Fenton for compiling Provincial Laboratory of Public Health's database information for this study.
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