Management of post-transplant Epstein-Barr virus-related lymphoproliferative disease in solid organ and hematopoietic stem cell recipients

Epstein-Barr virus (EBV)-related post-transplant lymphoproliferative disease (PTLD) is one of the most serious complications associated with solid organ and hematopoietic stem cell transplantation. PTLD is most frequently seen with primary EBV infection post-transplant, a common scenario for pediatric solid organ recipients. Risk factors for infection or reactivation of EBV following solid organ transplant are stronger immunosuppressive therapy regimens, and being seronegative for receptor. For hematopoietic stem cell transplantation, the risk factors relate to the type of transplant, human leukocyte antigen disparity, the use of stronger immunosuppressants, T-cell depletion, and severe graft-versus-host disease. Mortality is high, and most frequent in patients who develop PTLD in the fi rst six months post-transplant. The primary goal of this article is to provide an overview of the clinical manifestations, diagnosis, accepted therapies, and management of EBV infection in transplant recipients, and to suggest that the adoption of monitoring protocols could contribute to a reduction in related complications.

Epstein-Barr virus (EBV) is a deoxyribonucleic acid (DNA) virus of the herpes family that infects B-cells and the epithelial cells of the oropharynx.Infection results in different degrees of activation and proliferation of B-cells, and a latent infection is established within these cells.An intact immune system can control this infection, through either B-cell apoptosis, or cytotoxic T-cell activity.However, in the presence of post-transplant immunosuppression, immune control ceases to be effective and there is a risk of lymphoproliferative disease development 1 .
Although all cases of post-transplant lymphoproliferative disease (PTLD) are not caused by EBV, EBV-related PTLD is one of the main complications associated with relevant morbidity and mortality in solid organ transplant recipients.Its incidence is higher in children than in adults, and varies depending on the transplanted organ: 33% for combined liverkidney transplantation, 20% for small intestine, 15% for heart, 10% for liver, 6.4% for lungs, and 1.2% for kidneys.The observed differences may be related to the amount of lymphoid tissue transplanted, and the level of immunosuppressive therapy required to prevent transplant rejection [2][3][4][5] .EBV-related PTLD occurs more frequently in children owing to primary infection, while its occurrence in adults is a result of either primary infection or reactivation 6,7 .Presentation can vary from polyclonal reactive hyperplasia of B-cells, to monoclonal or polyclonal lymphomas.
Analysis of records from a database of 200,000 solid organs transplantations, at 271 centers in 42 European countries, showed that the frequency of EBV-related PTLD is highest for heartlung transplantation (42.7%), followed by the liver (21.8%), heart (16%), and kidney (10.3%) 1 .Mortality due to EBV-related PTLD is also high, ranging from 25% to 48%, and is more frequent in patients who develop PTLD in the fi rst six months post-transplant 8 .The main risk factors are: I) having transplant receptors susceptible to EBV; II) being less than 5 years of age (risk decreases with age), III) being in the fi rst 12 months post-transplant; and IV) a high level of immunosuppressive
For patients undergoing hematopoietic stem cell transplantation (HSCT), immunosuppressive therapy can lead to a mono-or poly-clonal proliferation of B-cells infected with EBV, resulting in the development of PTLD.The highest incidence of disease is in the fi rst fi ve months post-transplant, with reported levels ranging from 0.5 to 22% based on different studies.Additional risk factors that are also signifi cant are the use of unrelated donor material and graft-versus-host disease (GVHD) [9][10][11] .

CLINICAL SPECTRUM
There are no pathognomonic signs for PTLD; thus, it is essential to maintain a high degree of clinical suspicion and surveillance in order to make an early diagnosis.EBV infection and/or PTLD should be considered if the following signs or symptoms are present: I) a persistent fever, alone, or in association with gastrointestinal manifestations (diarrhea, abdominal pain, intestinal bleeding, protein-losing enteropathy, and, more rarely, intestinal perforation/obstruction); II) lymphadenopathy; III) tonsil or adenoid hypertrophy; IV) mononucleosis-like syndrome; V) hepatomegaly or splenomegaly; VI) anemia, leukopenia, pancytopenia, or hemophagocytosis; VII) graft dysfunction; or VIII) other pulmonary or central nervous system-associated signs or symptoms 5,12 .

DIAGNOSIS OF EBV-RELATED PTLD IN TRANSPLANT RECIPIENTS
A number of analyses are recommended for effective diagnosis and monitoring of EBV infection.Virus capsid antigen immunoglobulin (IgG and IgM (anti-VCA), and Epstein Barr nuclear antigen (EBNA) serology of the transplant recipient are recommended pre-transplant to determine if they may be susceptible to primary infection, reactivation, or carry a latent infection 9 .In addition, polymerase chain reaction (PCR) monitoring of viral replication should be conducted immediately post-transplant and periodically throughout the following year, depending on the risk status of the case 13,14 ; there is evidence to support the effi cacy of systematic PCR evaluation of EBV viral load in high-risk patients 14 .
Patients showing persistent viral replication, or those with initially high EBV levels, have a higher risk of progressing to PTLD.However, there are no precise protocols to determine the frequency of follow-up.Measurement of EBV load at the time of transplant, monthly for the fi rst six months, and then every two months for the following six months has been suggested.In addition, weekly viral load measurements are recommended for the fi rst six months for high-risk HSCT patients.Patients with two or more of the following risk factors are considered highrisk: susceptible, having an unrelated donor or human leukocyte antigens (HLA) disparity, or undergoing haploidentical transplantation, ATG therapy, or T-cell depletion [9][10][11] .
It is also important to mention that international standards have yet to be defi ned for EBV viral load measurement, this will be necessary to enhance inter-laboratory reliability 5 .
The term PTLD represents a spectrum of lymphoproliferative states that include benign conditions such as lymphoid hyperplasia, infectious mononucleosis-like disease, and malignancies, however, when not specifi ed, PTLD generally refers to the neoplastic end of this spectrum.Neoplasia is defi ned as any two of the following three characteristics: I -destruction of the underlying lymph node architecture; II -monoclonality; and III -evidence of EBV infection in neoplastic cells 15 .
If PTLD is suspected a biopsy of the affected organ is recommended, with diagnosis based on World Health Organization (WHO) criteria.WHO PTLD classification includes four categories: I) early lesions; II) polymorphic PTLD; III) monomorphic PTLD, and IV) classical Hodgkin lymphoma-type PTLD 16,17 .Early lesions are characterized by a polyclonal lymphoid proliferation with B-, T-and plasma cells with phenotypic homogeneity.In polymorphic PTLD, lymphoid cells form destructive extranodal masses, tissue architecture is damaged, and there is a proliferation of monoclonal B-cells.Monomorphic PTLD often presents with RAS, Myc, and P53 mutations and, in some cases, an altered B-cell immunophenotype with low or minimal CD20 expression [17][18][19] .
Other examinations conducted to determine the extent of disease include bone marrow biopsy, lumbar puncture for cerebrospinal fl uid (CSF) collection, and analysis besides radiological assessment.Chest and abdomen tomography and central nervous system (CNS) magnetic nuclear resonance can contribute to the diagnosis of PTLD in organs or systems located in the corresponding areas 20 , and, if a patient cannot receive a contrast for investigation of disease in the abdomen, magnetic nuclear resonance is a good alternative.Endoscopy and colonoscopy should be performed in patients with unexplained abdominal complaints and diarrhea following the exclusion of infection with other pathogens.

PREVENTION OF PTLD
Strategies for the prevention of EBV-related PTLD under investigation include immunoprophylaxis, chemoprophylaxis, and preemptive therapy.PCR analysis of peripheral blood for EBV viral load has been proven to be effective for early diagnosis of EBV-related PTLD, particularly for high-risk patients 5 .The detection of EBV DNA through quantitative real-time PCR (RTq-PCR) is currently considered the gold standard 5,21 .The adoption of monitoring protocols by different services has reduced the incidence of this complication, as observed by Imadome K and others; it has also improved overall PTLD-related survival and reduced morbidity for both solid organ transplantation and HSCT 2,5,22,23 .The monitoring of EBV viral load could indicate when immunosuppressive therapy should be reduced and/or antiviral agents administered.However, further research is required in order to develop individualized monitoring programs specifi c to transplant type and risk level 24 .

TREATMENT OF PTLD
There is no consensus on PTLD treatment; however, the main therapeutic options are as follows:  A reduction in immunosuppressive therapy should be considered as the fi rst-line of treatment, and must be individualized to each patient.With this strategy response