The success of increasingly potent immunosuppression in reducing the incidence and severity of allograft rejection after liver transplantation has been accompanied by an increasing incidence of Epstein–Barr virus (EBV)-associated posttransplant lymphoproliferative disease (PTLD). The cause of PTLD is multifactorial but is frequently associated with recent EBV infection. Most cases of PTLD occur during the first year after transplantation, with a smaller percentage occurring after the first posttransplantation year. Up to 90% of PTLD in the first posttransplant year is triggered by a primary EBV infection occurring in the first 3 to 4 months after transplantation. Pretransplant seronegativity and young age at the time of transplantation are risk factors for development of PTLD. The increased use of EBV-positive organs (split-livers, reduced-size liver segments, and living donors) in young, primarily EBV-negative patients has led to a greater prevalence of EBV donor–recipient mismatching and places more patients at risk of primary EBV infection.
Epstein–Barr virus enters the B cell by binding to the CD21 molecule on the surface of the B cell. After entering the B cell, the viral genome forms an episome and remains latent in B cells. Latent infection results in proliferation of B cells and clinical disease. Latent infection is characterized by the expression of nine of the genes encoded by the virus. These include six Epstein–Barr nuclear antigens (EBNA-1, -2, -3A, -3B, -3C, and -LP) and three integral latent membrane proteins (LMP-1, -2, and -2B) . Two nontranslated, small ribonucleic acids, EBER-1 and -2, are also encoded and are essential for maintenance of latent infection and for transformation and immortalization of B cells. In immunocompetent children, cellular immune responses control B-cell proliferation. Early control of B-cell proliferation results from non-major histocompatibility complex (MHC)–restricted natural killer cell activity. Subsequently, major histocompatibility complex–restricted cytotoxic T lymphocytes appear, directed against latent antigens (LMP-1 and EBNA-2) on the B cell. These cytotoxic T lymphocytes check uncontrolled B-cell production. However, in immunocompromised children, the number of cytotoxic T lymphocytes is reduced and B-cell proliferation is uncontrolled.
The association of EBV infection, impaired T-cell immunity and proliferation of B cells, resulting in a heterogeneous group of lymphoproliferative disorders, has been known for nearly 20 years. There are several convincing lines of evidence associating EBV infection with PTLD. Not only is the EBV genome detected in tumors from patients with PTLD, but EBV viremia is significantly increased in immunocompromised children with PTLD. The number of EBV-infected lymphocytes found during long-term latent infection in immunocompetent children is approximately 1 in 10 5 or 10 6 B cells and increases only 10-fold during an acute infection. In contrast, immunosuppressed transplant patients are less able to contain the virus, and, at the time of diagnosis of PTLD, the number of EBV-infected peripheral blood lymphocytes is 20 to 100,000 times greater than those in immunocompetent children or in children who underwent transplantation in whom PTLD did not develop (1). The technology of quantitative EBV polymerase chain reaction (EBV PCR), based on an end-point determination requiring serial dilution or the use of competitive internal standards, has permitted earlier identification of patients who seem to be at significant risk of development of PTLD and allowed early initiation of preemptive therapy to potentially prevent development of histopathologic PTLD.
In this issue of the Journal of Pediatric Gastroenterology and Nutrition, Kogan et al. (2) report their preliminary experience using the technique of quantitative competitive EBV PCR to detect asymptomatic EBV replication and to guide preemptive therapy to prevent progression to PTLD. They describe 13 patients who were EBV seronegative at the time of transplant and subsequently at high risk for development of PTLD. The donor liver was EBV seropositive in eight of these patients. All the patients were given intravenous ganciclovir in the immediate postoperative period. Epstein–Barr virus viral load increased in nine patients (69%), and immunosuppression was subsequently reduced in seven of the nine. After these interventions, EBV load decreased and PTLD developed in none of the seven patients. This is not a randomized, controlled investigation and treatment included two interventions: reduction of immunosuppression and administration of ganciclovir. However, this clinical experience adds to the existing body of knowledge regarding the effectiveness of preemptive therapy based on quantitative measurement of viral load; supports the need for a formal, multicenter, randomized controlled study; and highlights the need for good science and data regarding preemptive therapy.
Reports showing a correlation between EBV viral load, measured by EBV PCR in peripheral blood lymphocytes, and PTLD date back to at least 1994 (1). Studies have shown an 80% correlation between EBV viral load (detected by EBV PCR) and PTLD (3). Epstein–Barr virus viral load has also been shown to increase before clinical detection of PTLD and to decrease with effective therapy (4). In 1996, Green et al. (5) defined “threshold” EBV PCR levels of more than 40 genomes/10 5 peripheral blood lymphocytes for pretransplant EBV-seronegative patients and more than 200 genomes/10 5 lymphocytes for pretransplant EBV-seropositive patients. Patients whose EBV PCR levels exceeded these thresholds were preemptively treated to halt the progression of EBV infection to PTLD. Epstein–Barr virus PCR values more than 500 genomes/10 5 lymphocytes seemed to correlate with an increased risk of development of PTLD (6). After successful preemptive therapy, there may be intermittent or persistent rebound in EBV viral load in asymptomatic patients (7). This rebound may represent a balance between the reconstituted immune system (EBV-specific cytotoxic T lymphocytes) and latently infected B cells.
As the clinical significance of PTLD has increased, the focus of posttransplantation therapy has shifted to preemptive treatment of the disease. One strategy is to identify EBV infection as early as possible and to institute treatment to prevent asymptomatic EBV infection from progressing to symptomatic disease. Although the value of EBV PCR in identifying patients at risk of development of PTLD or diagnosing PTLD is becoming more accepted, the approach to treatment is controversial.
Recommendations for preemptive treatment include reducing immunosuppression and administering antiviral agents. This current approach to preemptive therapy is based on clinical reports and limited series, without the advantage of results from large, controlled studies. Although there is general agreement to reduce immunosuppression when the EBV viral load increases, there is no consensus as to how much to reduce or for how long to maintain reduced levels. However, it is not clear that all patients who reach threshold values of EBV PCR require therapeutic changes, including reducing immunosuppression, which may place the patient at risk of allograft rejection. The relative amount of immunosuppression at the time of the EBV primary infection or reactivation, the patient's history of rejection episodes, and other host variables must be taken into account before reducing immunosuppression based solely on PCR values.
The effectiveness of antiviral therapy has not been established. For antiviral prophylaxis to have any significant impact on EBV-induced PTLD, the antiviral therapy should be started at or before the time of transmission of EBV from the donor to the recipient. Although data from controlled randomized trials suggest little benefit of antiviral prophylaxis in decreasing the incidence of EBV PTLD, nonrandomized studies show potential benefit. McDiarmid et al (8) developed a protocol using preemptive intravenous ganciclovir for 100 days in high-risk liver transplant recipients, i.e., EBV-seropositive donor (D+) and EBV-seronegative recipient (R−) followed by 2 years of oral acyclovir administration. Children who underwent transplantation and who are at low risk (D+R+, D−R−, D−R+) received intravenous ganciclovir only during the initial hospital stay, followed by 2 years of treatment with oral acyclovir. Epstein–Barr virus PCR was monitored every 2 to 3 months in both groups. Tacrolimus immunosuppression was reduced in patients in both groups if the EBV PCR exceeded threshold levels. In no child in the high-risk group, preemptively treated with 100 days of ganciclovir, did PTLD develop. Since instituting this protocol, the overall incidence of PTLD at this center has decreased from 10% to 5%. These results imply that antiviral prophylaxis may be effective if administered at or before the time that the lytic or replicative form of EBV is transmitted from the donor to the recipient. However, the key element in the study by McDiarmid et al. (8) study may have been EBV PCR monitoring and therapeutic intervention based on the finding of increasing viral load.
The report by Kogan et al. (2) reinforces the value of monitoring quantitative competitive PCR as a guide to therapy and reaffirms the correlation between increased EBV viral load and PTLD. Although quantitative competitive PCR is helpful, the technique is expensive and time consuming. New assay techniques, which are less labor intensive and provide real-time PCR monitoring, have reduced turn-around time to less than 1 hour, permitting more frequent monitoring (9). It may be that no single value of EBV PCR is predictive of PTLD, and monitoring “trends” in serial quantitative EBV PCR values, coupled with monitoring of EBV-specific cytotoxic T cells, may be more valuable in guiding management of immunosuppressed patients with EBV and those at risk of development of PTLD.
The heterogeneity among reports from different transplant centers argues for a standardized multicenter approach to evaluate the outcome of therapy of PTLD. It will be important to determine whether the current approach will reduce the incidence, and morbidity and mortality rates of PTLD. It will also be important to evaluated the effectiveness of old and new therapies, including treatment with intravenous immunoglobulins, cytomegalovirus-specific immunoglobulin G, monoclonal antibodies, interferon, adaptive immunotherapy, and chemotherapy.
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