Posttransplant Lymphoproliferative Disease (PTLD)

The vast majority of cases of posttransplant lymphoproliferative disease (PTLD) are caused by Epstein-Barr virus (EBV) infection.[4, 5] EBV is a herpes virus that is thought to infect up to 95% of the population by adulthood.[6]

Primary infection with EBV in childhood usually results in mild, self-limiting illness.[6] In immunocompetent older children and adults, EBV infection causes infectious mononucleosis.[7] Once EBV infection occurs, the virus persists in a person for life, latently in B-cell lymphocytes and chronically replicating in the cells of the oropharynx.[8]

EBV was discovered in the 1960s by electron microscopy of cells cultured from a Burkitt lymphoma.[9] Subsequently, EBV has also been associated with non-Hodgkin lymphoma and oral hairy leukoplakia in patients with HIV infection and with nasopharyngeal carcinoma, particularly in patients from Southeast Asia.[7, 10, 11]

Structurally, the EBV genome is enclosed in a nuclear capsid surrounded by a glycoprotein envelope.The EBV genome is a linear DNA molecule that encodes for approximately 100 viral proteins that are expressed during replication. The CD21 molecule on the surface of the B cell is the target receptor of the EBV glycoprotein envelope. Infection of B-cell lymphocytes with EBV results in either viral replication and B-cell lysis (ie, lytic replication) or a transformation of the cell with only partial EBV genome expression (ie, latency).

Cell transformation is associated with B-cell activation and continuous proliferation. In immunocompetent patients, proliferation of transformed B cells usually is controlled by cytotoxic T cells. This is not the case, however, with patients who are immunosuppressed.[6]

During latency, the viral genome adopts an episomal configuration and expresses only 9 proteins. This change in gene expression creates increased difficulty for T-cell recognition, facilitating persistent EBV infection. Latency occurs in resting memory B cells. The 9 EBV proteins expressed during latency are the latent membrane proteins LMP-1, LMP-2A, and LMP-2B and the nuclear antigens EBNA-1, EBNA-2, EBNA-3A, EBNA-3B, EBNA-3C, and EBNA-LP.

LMP-1 is considered an oncogene. Its expression results in increased levels of CD23, which is a B-cell activation antigen. LMP-1 also induces expression of bcl-2, an inhibitor of apoptosis in infected cells. LMP-2 prevents EBV reactivation in latently infected cells. EBNA-1 maintains the episomal configuration of the latent virus. EBNA-2 up-regulates the expression of LMP-1 and LMP-2, which are necessary for transformation of the B cell.[6]

The lymphoproliferative tissue from most cases of PTLD has detectable EBV DNA and expression of 3 antigens in particular: EBNA-1, EBNA-2, and LMP-1. Two of these 3 proteins usually are not expressed in other EBV-related malignancies and so may be distinguishing features. Of note, the classic t(8;14) or t(8;22) chromosomal translocations observed in Burkitt lymphoma are often not detected in the tumors of patients with PTLD, even in cases of monomorphic/monoclonal B cell lymphoproliferations.[8, 12, 13]

Vietzen et al reported that genetic features in the transplant recipient and the EBV strain affect the risk of PTLD. In hosts with the HLA-E∗0103/0103 genotype, EBV strains that encode for the LMP-1 peptide variants GGDPHLPTL and GGDPPLPTL, presented by HLA-E, elicit strong inhibitory NKG2A+ NK and CD8+ T-cell responses.[14]

EBV infection normally results in both a humoral and cellular immune response by the host. Cellular immunity—CD4 and CD8 T cells and natural killer cells—is thought to be more important in controlling proliferation of infected B lymphocytes. Antibodies produced to viral capsid and nuclear proteins facilitate the diagnosis of EBV infection. In individuals who are immunocompetent, these mechanisms prevent outgrowth of EBV-infected lymphocytes. The immunosuppression required to prevent graft rejection post transplantation impairs T-cell immunity, potentially allowing for uncontrolled proliferation of EBV-infected B-cells, which may result in a spectrum of B-cell proliferations that range from hyperplasia to true lymphoma.[15, 16, 17, 18]

In the initial stages of PTLD, proliferation is polyclonal. With mutation and selective growth, the lesion becomes oligoclonal and, later, monoclonal. Lymphocytes from patients treated with cyclosporine following transplantation do not exhibit an appropriate T-cell response to EBV-infected B cells in vitro. The activity of natural killer cells also is reduced for several months following transplantation, contributing to the impairment of the most important regulator of proliferation: cellular immune response.[5, 19, 20]

The mechanisms that cause EBV-negative PTLD are not well understood. This disorder often has a later onset, monomorphic histology, and a more aggressive course than EBV-positive PTLD, suggesting that it may have a different pathophysiology.[21, 22]

Frequency

The frequency of posttransplant lymphoproliferative disease (PTLD) depends largely on the type of transplant received and the immunosuppression that the particular transplant requires. Primary EBV infection, such as may develop in an EBV-seronegative recipient who receives an allograft from an EBV-seropositive donor, is recognized as probably the most significant risk factor for developing PTLD. It is therefore not surprising that in general, PTLD rates are reported to be higher in pediatric transplant recipients than in adult transplant recipients.[23, 24, 25, 26]

The incidence of PTLD varies with the type of transplanted allograft. Reported rates are higher in heart, heart-lung, and small bowel transplants than in kidney and liver transplants. This presumably reflects in part the need for more intense immunosuppression to maintain certain types of allografts. The site of lymphoproliferative disease also depends on the type of graft. The lungs are frequently a site of involvement in patients undergoing heart-lung or heart-alone transplantation. Similarly, in small bowel transplants, the grafted bowel is commonly a site of PTLD. In cardiac transplants, the heart itself seldom is involved.

Kidney transplantation

A French kidney transplant registry consisting of data from adult patients who were enrolled prospectively from 1998 through 2007 found a 1% incidence of PTLD after 5 years, which increased to 2.1% by 10 years post transplantation. The more recently the transplant was performed, however, the less likely the recipient was to develop PTLD.[22]

In a Canadian single-center cohort study that included 1642 patients who received kidney transplants from 2000 through 2012, the incidence rate of PTLD was 0.18 cases per 100 person-years. EBV mismatch between donor and recipient was a strong risk factor.[27]

A report from the Organ Procurement and Transplantation Network (OPTN) of the United States reported PTLD incidence following kidney transplantation ranging from 0.6-1.5%. This study found an association between Medicare status, white race, and receipt of T-cell–depleting antibodies with the development of PTLD. A November 2012 report from the US OPTN found an incidence rate of 1.58 cases of PTLD per 1000 patient years post transplantation.[28]

In pediatric kidney transplant recipients, the reported incidence of PTLD ranges from 4.4-6.9%, with a wide range of times until onset—from less than 6 months to greater than 3 years.[29, 30] The majority of cases reported were B-cell proliferations. Age younger than 5 years at the time of transplantation and EBV-positive allograft donor to an EBV-seronegative recipient both significantly increased the risk ratio for developing PTLD. Many of the cases occurred while patients were still receiving steroids for immunosuppression.

Heart transplantation

In adults, heart transplantation is associated with approximately a 5% cumulative incidence of PTLD. Some reports have suggested that it may be as high as 8.9%.[31] Many reports of PTLD in heart transplant patients are single-institution studies and thus there may be greater variation seen in these reports compared with registry or cooperative studies. The US OPTN reports a rate of 2.24 cases of PTLD per 1000 patient years post heart transplantation.[28]

In children, the rate of PTLD following heart transplantation is even higher. Manlhiot et al found a 13% rate of PTLD at 4 years after pediatric heart transplantation at the Hospital for Sick Children at the University of Toronto. In this study, higher EBV load and longer induction of immunosuppression were associated with greater risk for PTLD.

Liver transplantation

An analysis of the Scientific Registry of Transplant Recipients of the United States reported PTLD rates of approximately 1% following liver transplantation in adult and pediatric recipients. The US OPTN reports a rate of 2.44 cases of PTLD per 1000 patient years post liver transplantation.

A retrospective analysis from the Mayo Clinic of 1206 orthotopic liver transplants (OLT) reported a cumulative incidence of PTLD of 1.1% at 18 months and 4.7% at 15 years. The authors suggest that previous reports of a lower incidence of PTLD late after OLT may warrant further examination as survival of OLT recipients improves.[32]

A retrospective study of 431 adult liver transplant recipients by Zimmerman et al reported that 11 (2.6%) patients developed PTLD. Multivariate analysis in that study revealed that pretransplant steroid treatment and liver transplantation for autoimmune hepatitis were risk factors for developing PTLD.[33]

More recent reports of PTLD in pediatric liver transplant recipients show rates that range from 3.8-4.7%. This is improved from earlier studies that reported PTLD rates as high as 5-15% for pediatric liver transplant.[34] The higher incidence of PTLD that was initially observed with tacrolimus as compared with cyclosporine[35] has more recently been attributed to the earlier practice of targeting higher tacrolimus levels. With greater experience using this immunosuppressive agent and targeting of lower therapeutic levels, tacrolimus no longer appears to be associated with a higher incidence of PTLD in pediatric liver transplant recipients.

Lung transplantation

Lung transplantation is associated with a relatively high rate of PTLD, with an incidence of approximately 5%. The median time to development of PTLD has been reported to be between 12 and 40 months, depending on the study. A report of 705 lung transplant recipients identified 34 cases of PTLD, for an incidence of 4.8%.[36] Among the 17 patients diagnosed within 11 months of transplantation, 85% had polymorphic PTLD and the most common site involved was the allograft, in 71% of the patients. PTLD that developed late was typically monomorphic and was more likely to involve the GI tract (39%) than the lung allograft (28%). The US OPTN reports a rate of 5.72 cases of PTLD per 1000 post transplantation patient years, with most cases occurring less than 1 year post transplantation.

HSCT

In the setting of HSCT, PTLD rates vary greatly depending on the conditioning regimen and the amount of T-cell depletion involved in either the conditioning or the product provided. Rates as high as 71% have been reported in the setting of very extreme T-cell depletion. More recently, however, rates of PTLD in adult HSCT range from 0.2% with no T-cell depletion to 1.7% with antithymocyte globulin treatment.[21]

Similar results are seen in pediatric HSCT. PTLD occurs in less than 1% of non–T-cell–depleted grafts from matched siblings, compared with as high as 30% of patients with unrelated or HLA-mismatched donors when extensive T-cell depletion of the donor bone marrow is performed. Other risk factors for PTLD in HSCT recipients include treatment of graft versus host disease with antithymocyte globulin or anti–T-cell monoclonal antibodies.[37]

PTLD represents a heterogeneous group of tumors, ranging from B-cell hyperplasia to immunoblastic lymphoma, the latter portending a grim prognosis.[3, 38, 39]  All PTLD, however, irrespective of histology, is potentially life-threatening.

The presentation and clinical course for PTLD may be quite variable. At one end of the spectrum is fulminant disease with diffuse involvement, which may result in the rapid demise of the patient. At the other end of the spectrum are localized lesions that may be indolent and slow growing over months. The former tends to occur early post transplantation after primary EBV infection and often involves the allograft itself. Late-onset PTLD tends to be monoclonal, EBV-negative, and often more refractory to therapy.[19]

In a retrospective review of 32 adult and pediatric SOT patients with PTLD, the 5-year survival rate was 59%.[38]  Nearly half of the patients were diagnosed within the first year following transplantation. Characteristics associated with poorer survival were diagnosis within the first year post transplantation, monoclonal tumors, and presentation with an infectious mononucleosis–like syndrome.

While advances in the diagnosis and treatment of PTLD have improved outcomes for patients with PTLD, studies continue to report high rates of mortality from PTLD. A series of adult kidney and heart transplant recipients reported by Wasson et al in 2006 had a mortality rate of 26.6%. Similarly, a more recent report of pediatric kidney transplant patients also found a 25% mortality rate related to PTLD.[30]

Central nervous system (CNS) involvement in EBV-PTLDs portends a very poor prognosis. In a case series of CNS PTLDs after HSCT, 7 of the 9 patients died a median of 17 days from diagnosis.[40]

Several studies have attempted to identify prognostic factors for PTLD outcomes. For example, Evens et al identified hypoalbuminemia as a strong prognostic factor in their multicenter study of 80 adult SOT recipients with PTLD. Similarly, Khedmat and Taheri performed a literature review and found evidence to suggest that CD20-positive PTLD portends earlier disease appearance and inferior outcome.

LeBlond et al applied the non-Hodgkin lymphoma International Prognostic Index (IPI) to a series of 61 adult patients with PTLD following kidney, lung, liver, or heart transplantation to analyze factors that may be predictive for shorter survival. In univariate analysis, the following were found to be poor prognostic features:

• Performance status (PS) ≥2
• Increased number of sites involved (ie, >1 vs 1)
• Primary central nervous system (CNS) involvement
• T-cell origin
• Monoclonality
• Nondetection of EBV in the tumor
• Treatment based on chemotherapy (in addition to reduction in immunosuppression)

In multivariate analysis, PS less than 2 and decreased number of disease sites (ie, 1 vs >1) both were associated with improved survival. These determinants were used to identify 3 levels of risk stratification in terms of survival probability. For intermediate-risk patients (ie, PS ≥2 or 2 or more sites), median survival time with treatment was 34 months. For high-risk patients (ie, PS ≥2 and 2 or more sites), median survival time was 1 month. Survival time for low-risk patients (ie, PS < 2 and < 2 sites) was not defined. This risk stratification system was found to be more helpful in determining prognosis for PTLD patients than the IPI used for immunocompetent patients with non-Hodgkin lymphoma.

Although a number of prognostic scores have been proposed in different transplantation populations, none has been externally validated or widely adopted. The rarity and clinical diversity of the disease, as well as the nature of the reporting of PTLD cases, which is often retrospective and from single-institution studies, have contributed to the challenge of developing and validating a universally accepted system of risk classification.

Whether posttransplant lymphoproliferative disease (PTLD) presents as localized or disseminated disease, the tumors are often aggressive, rapidly progressive, and potentially life-threatening. Clinical presentation is variable and includes the following:

• Fever (57%)
• Lymphadenopathy (38%)
• Gastrointestinal symptoms (including obstruction [27%])
• Infectious mononucleosis–like syndrome, which can be fulminant (19%)
• Pulmonary signs and symptoms (15%)
• Central nervous system (CNS) symptoms (13%)
• Weight loss (9%)

Involvement of the allografted tissue can cause a decline in its function. This organ failure may be the presenting symptom.

Having a high index of suspicion and clinical vigilance is critical, because patients may present with nonspecific symptoms or systemic signs (eg, hepatosplenomegaly, fever, fatigue) that may otherwise not initially suggest a diagnosis of PTLD. A rising blood level of EBV viral load by quantitative polymerase chain reaction (PCR) measurement in this clinical setting should raise concern for PTLD.[41, 4, 25, 42, 43]

The most common sites of involvement are are as follows[44, 37] :

• Lymph nodes
• Liver
• Lung
• Kidney
• Bone marrow
• Small intestine
• Spleen
• CNS
• Large bowel
• Tonsils
• Salivary glands

Less common sites of involvement include the oral cavity, skin, and subcutaneous lesions. T-cell lymphoproliferative disorders not associated with EBV infection tend to occur at extranodal sites.

The initial evaluation for posttransplant lymphoproliferative disease (PTLD) should include a complete history and physical examination, complete blood cell count, comprehensive chemistry panel, and Epstein-Barr virus (EBV) evaluation. Attention should be given to the metabolic panel for signs of tumor lysis syndrome.

Elevated lactate dehydrogenase (LDH) concentrations may represent rapid cell turnover, but this has not been shown to be prognostic in PTLD. Additional laboratory tests specific for allograft function should also be obtained.

The EBV status of the recipient usually is established pretransplantation. Donor EBV status may not be routinely tested because the incidence of infection with EBV in the general adult population is so high.

In primary EBV infection, EBV viral capsid antigen immunoglobulin M (IgM) titers are elevated. Reactivation of EBV infection is characterized by more than a 4-fold rise in EBV viral capsid antigen immunoglobulin G (IgG) titers, compared with previously recorded EBV viral capsid antigen IgG titers. No change in titer suggests past infection. However, in immunocompromised patients, the antibody titer response may be a less reliable marker of acute infection or reactivation; thus, the absence of change in EBV antibody titers does not exclude a diagnosis of PTLD.

The EBV viral load in the peripheral blood, measured by quantitative PCR, is the most commonly used laboratory test to monitor patients who are at risk for developing PTLD after solid organ transplantation (SOT) or bone marrow transplantation. A single elevated EBV PCR value is less informative than a trend of rising (or falling) values over time. Because testing for EBV by blood PCR is performed using different techniques in different laboratories, it may not be valid to compare results from one laboratory to another. A negative EBV PCR does not rule out the presence of PTLD.

The diagnosis of PTLD can only be made by histologic confirmation of tumor tissue. PTLDs are categorized pathologically by the WHO classification system (see Background and Histologic Findings). Histopathologically, the lesion may demonstrate plasmacytic hyperplasia, B-cell hyperplasia, B-cell lymphoma, or immunoblastic lymphoma. Less commonly, T-cell lymphoma or Hodgkin lymphoma may manifest as PTLD.

Evaluation of tumor tissue for the presence of EBV is very important and is typically performed with immunohistologic staining of paraffin-embedded tissue. In situ hybridization with the EBV-encoded RNA (Epstein-Barr early region [EBER]-1) probe (which labels EBV-encoded RNA in infected cells) is a reliable means of detecting EBV in tissue. While the majority of PTLDs are EBV-positive, EBV-negative PTLDs are seen typically in tumors that present late post transplantation.

Establishing the clonality of the lesion is also important. Tumors can be monoclonal, oligoclonal, polyclonal, or mixed. Clonality evaluation is typically performed by immunohistochemical staining of surface immunoglobulin light chains or by molecular determination of immunoglobulin or T-cell receptor rearrangement.

Radiological evaluation includes computed tomography (CT) or magnetic resonance imaging (MRI) scan of the neck, chest, abdomen, pelvis, and head, looking for evidence of any abnormal nodal or extranodal masses.

The role of [18 F]2-fluoro-2-deoxyglucose (FDG) positron emission tomography (PET)/CT scanning is still under evaluation. However, PET/CT can be useful for staging disseminated disease.

Obtaining bone marrow aspirate and biopsy is appropriate to determine whether marrow is involved in the disease process.

If a suspicion of CNS or neurological involvement exists, lumbar puncture should be performed for routine cerebrospinal fluid (CSF) evaluation and examination of the CSF for malignant cells. In addition to the standard tests, the fluid may also be analyzed for EBV DNA, using polymerase chain reaction (PCR).

The pathological diagnosis of posttransplant lymphoproliferative disease (PTLD) is based on the World Health Organization (WHO) classification and includes the following 4 main categories[3] :

1. Early lesions
2. Polymorphic PTLD
3. Monomorphic PTLD
4. Classic Hodgkin lymphoma

The monomorphic subgroup includes both B-cell and T-cell neoplasms.[45] This classification system is now universally accepted for defining distinct subtypes of PTLD. In practice, however, a clear separation between the different subtypes is not always possible; early lesions, polymorphic PTLD, and monomorphic PTLD probably represent a spectrum of disease, and more than one subtype may sometimes be present in a given patient.

Prior to the adoption of the WHO system for classification of PTLD, an alternative classification system had been proposed by Knowles et al in 1995. This system classified PTLD into the following 3 distinct categories:

1. Plasmacytic hyperplasia, which arises most commonly in the oropharynx or lymph nodes and nearly always is polyclonal, with a lack of oncogene or tumor suppressor gene alteration
2. Polymorphic B-cell hyperplasia and polymorphic B-cell lymphoma, which can be nodal or extranodal, nearly always are monoclonal, usually contain a single form of EBV, and lack oncogene or tumor suppressor gene alteration
3. Immunoblastic lymphoma with widely disseminated disease, which is monoclonal, contains a single form of EBV, and contains alterations of one or more oncogene or tumor suppressor genes

This classification system is no longer routinely used. However, some earlier literature may reference it.

The images below show histologic findings consistent with PTLD.

Biopsy of gingival tissue (400 X) with hematoxylin and eosin stain demonstrates polymorphous infiltrate of atypical lymphoid cells, which is consistent with posttransplant lymphoproliferative disease (PTLD).

Biopsy of gingival tissue (400 X). Epstein-Barr virus encoded RNA (EBER) study shows numerous positive cells, which is consistent with posttransplant lymphoproliferative disease (PTLD).

The management of posttransplant lymphoproliferative disease (PTLD) remains a challenge and generally without a standardized therapeutic approach that can be applied to all patients. Despite this diversity, reduction of immunosuppression (RIS) remains the cornerstone for treatment of Epstein-Barr virus (EBV)–driven B-cell PTLD, independent of histology.

Starzl et al were the first to suggest reduction, or withdrawal, of immunosuppression as a treatment option for PTLD.[46] This strategy allows the patient's natural immunity to recover and gain control over proliferating EBV-infected cells. Benkerrou et al reported complete regression in about 40% of patients after reduction or discontinuation of immunosuppressive therapy.[47] Patients with less-aggressive or polyclonal PTLD tend to respond more favorably to this management approach, as compared with patients with clinically aggressive PTLD.

In patients who have an inadequate response to reduction of immunosuppression, the humanized anti-CD20 monoclonal antibody rituximab has become a standard of care. Rituximab can be given as monotherapy or in combination with chemotherapy, concurrently or sequentially.[48] Reports have demonstrated the safety and efficacy of single-agent rituximab in the treatment of CD20-expressing PTLD, generally with a response rate of approximately 50%.[49, 50, 51]

However, relapse of PTLD is not uncommon after rituximab monotherapy. In addition, the kinetics of disease response to rituximab may be slower than what is observed with chemotherapy, making it a less effective therapeutic option for patients with clinically aggressive or fulminant PTLD. These observations have led investigators to combine rituximab with chemotherapy.

The PTLD-1 trial, a prospective, international, multicenter, phase II study, found rituximab followed by CHOP (cyclophosphamide, doxorubicin [hydroxydaunorubicin], vincristine [Oncovin], and prednisone) chemotherapy to be a safe and effective treatment for adult solid organ transplant (SOT) patients with PTLD that fails to respond to reduction in immunosuppression. Patients received 4 weekly courses of rituximab followed by CHOP. However, the efficacy of chemotherapy was offset by its toxicity, with an 11% CHOP-associated mortality rate and a 9% rate of adverse events significant enough to halt CHOP treatment.[52]

Further analysis of PTLD-1, along with the subsequent PTLD-2 trial, suggested a protocol for the use of rituximab and chemotherapy in CD20-positive B-cell PTLD in adults that balances risks and benefits. The studies established that patients who enter complete remission with rituximab induction therapy, or who have less than three international prognostic index (IPI) risk factors at diagnosis and enter partial remission, are at low risk and can continue with rituximab monotherapy.[53]

In contrast, other patients are at high risk and can benefit from the addition of CHOP. This includes patients who have three or more IPI risk factors and attain partial remission, those with stable disease, and those with progressive disease who are non-thoracic SOT recipients. Thoracic SOT (ie, lung, heart) recipients with progressive PTLD are at very high risk, and even with intensive chemoimmunotherapy their prognosis remains poor.[14]

National Comprehensive Cancer Network (NCCN) guidelines recommend CHOP, given sequentially or concurrently with rituximab, as chemoimmunotherapy for monomorphic PTLD (B-cell type) and polymorphic PTLD. For frail patients who cannot tolerate anthracycline, no specific regimen has been identified but options may include CEOP ( cyclophosphamide, etoposide, vincristine, prednisone) or CVP (cyclophosphamide, vincristine, prednisone). For T cell–type monomorphic PTLD, the NCCN recommends brentuximab vedotin plus CHP (cyclophosphamide, doxorubicin, prednisone) for CD30+ cases, CHOP, or CHOP plus etoposide (CHEOP).[54]

SOT patients are often not able to tolerate full-dose chemotherapy owing to end-organ toxicity or risk of allograft dysfunction. For cardiac transplant patients, the dose of doxorubicin is often reduced over concerns of myocardial toxicity. Patients who develop PTLD after hematopoietic stem cell transplant (HSCT) are often not able to tolerate chemotherapy owing to myelotoxicity.

Despite these challenges, with diligent supportive care and careful monitoring for toxicity, chemotherapy can be safely administered to most SOT patients. Given the risk of treatment-related morbidity and mortality, this strategy is often reserved for patients in whom front-line therapy with reduction of immunosuppression and/or rituximab failed, for patients with CD20-negative PTLD, or for patients with clinically fulminant PTLD, including those with CNS involvement.

In an effort to decrease the chemotherapy-related toxicity observed in SOT recipients, a low-dose chemotherapy regimen consisting of cyclophosphamide and prednisone was piloted in 36 children with PTLD in whom initial reduction of immunosuppression failed.[55] The overall response rate was 83%, and 2 patients died of treatment-related toxicity. The PTLD relapse rate was 19%.

To further assess the efficacy of this regimen, a Children’s Oncology Group phase II trial of low-dose cyclophosphamide and prednisone together with rituximab was conducted.[56] Fifty-four pediatric patients with PTLD were enrolled in the study, and the majority had monomorphic disease. The complete response rate in this study was 69%, and the 2-year event-free survival rate was 71%; there was one death due to infection during therapy.

The choice of therapy ideally attempts to balance the risk of life-threatening PTLD with the risk of allograft failure and treatment-related morbidity. In patients who have undergone solid organ transplantation (SOT), reduction of immunosuppression may risk allograft rejection. In addition, SOT recipients are often at greater risk for organ toxicity and opportunistic infections that may complicate chemotherapy administration. In hematopoietic stem cell transplantation (HSCT) patients with PTLD, reduction of immunosuppression may increase the risk of graft versus host disease.

Additional therapeutic measures that have been used, each with varying degrees of success, include the following[57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69] :

• Surgical excision of the lesion (may be curative in cases of localized disease)
• Radiation therapy
• Cytotoxic T lymphocytes
• Antiviral therapy
• Intravenous gamma globulin (IVIG)
• Interferon alfa

Adoptive immunotherapy with cytotoxic T lymphocytes (CTLs) is based on the understanding that EBV PTLD arises in SOT or HSCT patients from disruption of the normal balance between latently infected B cells and the anti–EBV-specific T-cell response. In 1994, Papadopoulous et al reported on the infusion of donor leukocytes to treat PTLD that had developed in 5 patients following T-cell–depleted allogeneic HSCT. Infusions of unirradiated leukocytes from EBV-seropositive donors resulted in complete clinical responses of PTLD in all patients. However, the infusion of CTLs was complicated in some patients by the development of graft versus host disease.

Since these early experiments, adoptive immunotherapy techniques have been refined and continue to show promise as a therapy for PTLD. Donor EBV-specific CTLs administered to HSCT recipients with PTLD have been reported to achieve an overall response rate of 68% and without inducing graft versus host disease.[69] A prospective study of rituximab treatment followed by donor or autologous EBV-CTL infusion reported 5-year overall survival of 70.7% and progression-free survival of 68.9%.[70]

The use of donor CTLs is more problematic for SOT patients. This prompted the development of “third party” EBV cytotoxic T lymphocytes, which could potentially be available from a bank of HLA-typed EBV-specific T-cell lines.[71, 72] Tabelecleucel (Ebvallo) an allogeneic EBV-specific T-cell immunotherapy, has been approved by the European Medicines Agency.[73]

Antiviral therapy (eg, acyclovir, ganciclovir, foscarnet) in the absence of reduction of immunosuppression is not considered effective treatment for PTLD. While these drugs have not shown efficacy as single-agent therapy for PTLD, they have been used together with reduction of immunosuppression as a first step in management.[74] However, current European guidelines recommend against the use of antiviral drugs for treatment of PTLD.[75]

IVIG has been used as adjunctive therapy in the management of PTLD. Deficiency or absence of antibody against one of the EBNAs in patients post transplantation has been associated with the subsequent development of PTLD. Decreasing EBV viral load has been reported to be associated with increased levels of antibody against Epstein-Barr nuclear antigens (EBNAs). These 2 factors provide the rationale for the use of IVIG in the management of PTLD.

IVIG or anti-cytomegalovirus (CMV) immunoglobulin (CytoGam), which contains anti-EBV antibodies, is most commonly used in conjunction with antiviral therapy as prophylaxis against CMV in SOT or HSCT patients. This anti-CMV prophylactic regimen may also provide some protection against developing EBV-PTLD.[76] In clinical practice, this strategy is frequently initiated in SOT patients with rising EBV viral loads who are considered to be at risk for developing PTLD.

Interfen alfa has historically been considered a potential therapeutic option in the treatment of B-cell PTLD.[34, 59, 60] Interferon alfa is recognized to inhibit the outgrowth of EBV-transformed B cells, and it decreases the oropharyngeal shedding of EBV. It also is known to inhibit T helper cells, which release cytokines (ie, interleukin [IL]–4, IL-6, IL-10) that promote B-cell proliferation. Interferon functions as both a proinflammatory and antiviral agent. Because no prospective clinical trials have been conducted to date, many of the reports of its success in the management of PTLD are anecdotal.

Clinical trials are underway to evaluate the efficacy of EBV-specific inhibitors, including inhibitors of EBNA1 and EBNA2 as well as heat-shock protein 90 (HSP90) inhibitors, but none have been approved for clinical use.[77]

For patients with CNS involvement of PTLD, the prognosis remains poor even with aggressive therapy. High-dose methotrexate has been reported to be a tolerable and effective therapy for CNS PTLD.[78, 79, 67] Intrathecal therapy is also considered advisable because many systemic chemotherapy agents and monoclonal antibodies do not cross the blood-brain barrier adequately. Small series have described treatment of isolated CNS PTLD with intrathecal rituximab.[80] In one study, 7 of 8 children with isolated CNS PTLD responded after a median of 2 courses of rituximab, and the therapy was generally well tolerated.[81] Radiotherapy also remains an effective modality for the treatment of CNS PTLD.[82]

Prophylaxis

In 2012, an international multidisciplinary panel of experts published a consensus statement on the classification and risk factors for PTLD and outlined approaches to minimize the risk of developing PTLD.[2] The first of these recommendations from the Seville Workshop group is that EBV status of both the donor and the recipient should be ascertained prior to donor selection, and whenever possible, EBV-negative recipients should receive grafts from EBV-negative donors.

The next suggestion is to minimize upfront immunosuppression as much as possible and potentially to use reactivation of other viruses, such as CMV or BK virus, as cues to reduce immunosuppression. Although antiviral therapy has not proven to be an effective treatment for PTLD, in selected high-risk patients prophylactic or preemptive antiviral therapy may be considered. However, current European guidelines recommend against the use of antiviral drugs for preemptive therapy.[75]

An alternative approach to antiviral prophylaxis is to administer IVIG or CytoGam to maintain high titers of anti-EBV antibodies that may help prevent the development of EBV PTLD.

The last recommendation from the Seville Workshop is to consider preemptive treatment for those patients who appear to be developing PTLD. A rising EBV viral load in a high-risk patient may warrant preemptive reduction in immunosuppression, while continuing to monitor closely for allograft dysfunction.

In patients with significant EBV DNA blood levels but without clinical symptoms of EBV disease, European guidelines recommend preemptive treatment with rituximab, combined with reduction in immunosuppression if possible. Consideration of EBV-CTL therapy, if available, is recommended.[75] A retrospective study by Stocker et al in 219 high-risk recipients undergoing allogeneic HSCT concluded that preemptive rituximab is associated with a low incidence of EBV-PTLD and may be considered worthwhile, despite inducing a delayed B-cell reconstitution and a high risk of neutropenia.[83]

Surgical management alone is rarely the sole mode of therapy for posttransplant lymphoproliferative disease (PTLD). Obtaining tissue for histologic examination is necessary for diagnosis in patients with clinical concern for PTLD. Occasionally, when PTLD is localized to a single nodal or extranodal site (eg, in localized small bowel lesions that present as intussusception), the diagnostic surgical procedure may remove the only site of disease. In such a situation, the decision as to whether the patient will benefit from adjuvant therapy (reduction of immunosuppression, rituximab, or chemotherapy) depends on the pathologic diagnosis and the patient’s individual risk factors.

Following the Epstein-Barr virus (EBV) viral load provides useful information regarding disease status and response to treatment. Green at the University of Pittsburgh recommends weekly monitoring of EBV viral titers in the peripheral blood of patients with posttransplant lymphoproliferative disease (PTLD),[84]  although for lower-risk patients the interval of monitoring may appropriately be monthly to every 3 months.

Declining viral load may suggest a response to treatment. Persistently high or rising viral load may suggest development of PTLD or disease progression. However, serial physical examinations, radiological evaluation, and assessment for any evidence of allograft rejection are essential in conjunction with EBV viral load monitoring, as some patients have wide fluctuations in blood EBV levels that do not clinically correlate with PTLD.

Management of PTLD remains a balancing act between eradication and cure of the disease and preservation of graft function. The degree of graft loss that can be tolerated depends on the graft in question. Reduction or withdrawal of immunosuppression has to be tailored accordingly, after multidisciplinary discussion, and with due consideration of other treatment options. At this time, treatment has not been standardized beyond the generally accepted belief that reduction or withdrawal of immunosuppression is initial step in PTLD management, following which the treatment is usually tailored to the specific needs of the patient.

Guidelines for the management of PTLD have been published by the following groups:

• National Comprehensive Cancer Network (NCCN)
• American Society of Transplantation (AST)
• Sixth European Conference on Infections in Leukemia (ECIL-6), a joint venture of the Infectious Diseases Working Party of the European Society of Blood and Marrow Transplantation (EBMT-IDWP), the Infectious Diseases Group of the European Organization for Research and Treatment of Cancer (EORTC-IDG), the International Immunocompromised Host Society (ICHS) and the European Leukemia Net (ELN)

Diagnosis

The NCCN guideline for PTLD, which is integrated into the larger B-cell lymphoma guideline, provides the following algorithm to diagnose the disorder with steps categorized as either "essential" or "useful under certain circumstances."[54]

Essential steps include the following:

• Adequate immunophenotyping by immunohistochemistry (IHC) panel, with or without cell surface marker analysis by flow cytometry
• Epstein-Barr virus (EBV) evaluation by EBV latent membrane protein 1 (LMP1) or Epstein–Barr encoding region in situ hybridization (EBER-ISH); EBER-ISH recommended if EBV-LMP1 is negative

Tests useful under certain circumstances include the following:

• Additional immunophenotyping
• Molecular analysis to detect IgH gene rearrangements

AST guidelines recommend the following for the diagnosis of PTLD[85]

• Cytomegalovirus (CMV) status testing
• Viral-load testing for EBV
• Total body CT scan (head to pelvis) 
• Immunophenotyping to determine lineage and therapy dependent markers (ie, CD20) 
• Molecular genetic markers of antigen-receptor genes to assess clonality
• Donor versus recipient origin

ECIL-6 guidelines recommend quantitative PCR of whole blood, plasma, or serum to screen for EBV DNA in allogeneic hematopoietic stem cell transplant (HSCT) recipients and to monitor EBV DNA-emia.[75] Screening should begin no later than 4 weeks after HSCT, with consideration of earlier screening in patients with several risk factors. Testing should be once weekly in high-risk EBV PCR-negative patients and perhaps more frequently in patients with rising EBV DNA levels. Screening should end at least 4 months after HSCT; longer monitoring is recommended in patients with poor T-cell reconstitution, as follows:

• On treatment for severe acute or chronic graft versus host disease
• After haploidentical HSCT
• In T-cell–depleted HSCT
• After conditioning with anti-thymocyte globulin (ATG)/alemtuzumab
• With early EBV reactivation

ECIL-6 recommendations for PTLD diagnosis are as follows[75] :

• Diagnosis of EBV-PTLD must be based on clinical presentation plus detection of EBV by an appropriate method in a specimen from involved tissue.
• Noninvasive diagnostic methods include quantitative EBV DNA measurement in blood, plasma, or serum and PET-CT or CT; PET-CT is preferred over CT in extranodal disease
• Invasive methods include biopsy of lymph nodes and/or other suspected sites
• Definitive diagnosis requires biopsy and histological examination with EBV detection.
• In situ hybridization for EBER transcripts or detection of viral antigens is necessary.

Treatment

According to the NCCN guidelines, treatment depends on the PTLD subtype and has varying rates of response. Reduction in immunosuppression (RI) is the initial treatment in nearly all cases. However, response is variable and patients should be closely monitored. Additional treatment options vary according to the World Health Organization (WHO) classification.[54]

Non-destructive lesions

• For patients with a complete response (CR), re-escalation of immunosuppression should be individualized, with monitoring of EBV viral load by PCR assays and graft organ function
• For patients with persistent or progression, second-line treatment with rituximab and monitor EBV by PCR

AST guidelines concur with the recommendations above, but also suggest complete or partial surgical resection, as well as local radiotherapy as adjunctive therapy along with reduced immunosuppression.[85]

Monomorphic PTLD (B-cell type)

 RI if possible, with or without one of the following:

• Rituximab
• Rituximab as part of a concurrent or sequential chemoimmunotherapy regimen

Polymorphic PTLD

For patients with localized disease, RI if possible plus one of the following:

• Rituximab
• Radiation therapy (RT) with or without rituximab
• Surgery with or without rituximab

For patients with systemic disease, RI if possible plus one of the following:

• Rituximab
• Rituximab as part of a concurrent or sequential chemoimmunotherapy regimen

ECIL-6 guidelines

ECIL-6 recommendations for first-line therapy of EBV PTLD are as follows[75] :

• Rituximab, 375 mg/m ² once weekly
• Always consider RI combined with rituximab
• Adoptive immunotherapy with in vitro–generated donor or third-party EBV-specific T lymphocytes (CTLs), if available

ECIL-6 recommendations for second-line therapy of EBV PTLD are as follows[75] :

• Cellular therapy with EBV-specific CTLs or donor lymphocyte infusion
• Consider chemotherapy with or without rituximab, if other methods fail
• Surgery, intravenous immunoglobulin, and antiviral agents are not recommended

For preemptive therapy, the ECIL-6 guidelines recommend significant EBV DNA-emia without clinical related clinical manifestations as an indication, although no specific EBV DNA level can be recommended as a treatment threshold. Rituximab once weekly, 1-4 doses until EBV DNA levels are negative, is recommended; if possible, rituximab should be combined with RI. Donor or third-party EBV specific cytotoxic CTLs, if available, should be considered. Antiviral drugs are not recommended.[75]

Immunosuppressive agents are often an important part of graft maintenance and posttransplant care. Nevertheless, when faced with posttransplant lymphoproliferative disease (PTLD), it is important to reduce or modify the immunosuppressive regimen if at all possible. Because of the complexity and variety of immunosuppressive regimens, there are no standard approaches to achieve this reduction in immunosuppression. It is left to the combined judgement of the transplanting and PTLD-treating physicians. Any reduction in immunosuppression warrants close monitoring for the possibility of allograft dysfunction or rejection.

In addition to immunosuppression, a variety of additional therapeutic measures have been used, each with varying degrees of success. These include antiviral, immunomodulator, and chemotherapy agents.

Class Summary

These agents inhibit key factors that mediate immune reactions, which, in turn. decrease inflammatory responses.

Cyclosporine (Sandimmune, Neoral)

Cyclosporine is a cyclic polypeptide that suppresses some humoral immunity and, to a greater extent, cell-mediated immune reactions, such as delayed hypersensitivity, allograft rejection, experimental allergic encephalomyelitis, and graft versus host disease for a variety of organs.

For children and adults, base dosing on ideal body weight.

Tacrolimus (Prograf)

Tacrolimus suppresses humoral immunity (T-lymphocyte) activity.

Mycophenolate (CellCept)

Mycophenolate inhibits inosine monophosphate dehydrogenase (IMPDH) and suppresses de novo purine synthesis by lymphocytes, thereby inhibiting their proliferation. It inhibits antibody production.

Prednisone (Deltasone, Orasone, Meticorten, Sterapred)

Prednisone is used as an immunosuppressive, anti-inflammatory agent and as a component of both CHOP and ProMACE-CytaBOM chemotherapeutic regimens, which have been used to treat PTLD.

It may decrease inflammation by reversing increased capillary permeability and suppressing PMN activity. It stabilizes lysosomal membranes and suppresses lymphocytes and antibody production.

Class Summary

These are nucleoside analogs phosphorylated by viral thymidine kinase to form a nucleoside triphosphate. These nucleoside triphosphates inhibit herpes simplex virus (HSV) polymerase with 30-50 times more than they inhibit human alpha-DNA polymerase.

Acyclovir (Zovirax)

Acyclovir inhibits activity of both HSV-1 and HSV-2. It has affinity for viral thymidine kinase and, once phosphorylated, causes DNA chain termination when acted on by DNA polymerase.

It is routinely used to treat infections with HSV, mainly HSV-1 and HSV-2. EBV also is a herpes virus, but its use as prophylaxis against and treatment for EBV-related illness posttransplantation is controversial. If used for these purposes, doses and duration of treatment are variable and are determined by the clinician.

Ganciclovir (Cytovene)

Ganciclovir is a synthetic guanine derivative active against CMV. It is an acyclic nucleoside analog of 2'-deoxyguanosine that inhibits replication of herpes viruses both in vitro and in vivo.

Levels of ganciclovir-triphosphate are as much as 100-fold greater in CMV-infected cells than in uninfected cells, possibly due to preferential phosphorylation of ganciclovir in virus-infected cells.

For patients who experience progression of CMV retinitis while receiving a maintenance treatment with either dosage form of ganciclovir, the re-induction regimen should be administered.

Class Summary

Rituximab (anti-CD20 monoclonal antibody) has successfully treated PTLD. Other monoclonal antibodies, such as anti-CD21, CD24, and immunomodulatory agents such as interferon alfa have also been used in the treatment of PTLD.

Rituximab (Rituxan)

Rituximab is an antibody genetically engineered. It is a chimeric murine/human monoclonal antibody directed against the CD20 antigen found on surface of normal and malignant B lymphocytes. The antibody is an IgG1 kappa immunoglobulin containing murine light- and heavy-chain variable region sequences and human constant region sequences.

Immune globulin intravenous (Gamimune, Gammagard S/D, Sandoglobulin)

Immune globulin intravenous neutralizes circulating myelin antibodies through anti-idiotypic antibodies. It down-regulates proinflammatory cytokines, including INF-gamma. It blocks Fc receptors on macrophages. It suppresses inducer T and B cells and augments suppressor T cells. It blocks the complement cascade, promotes remyelination, and may increase CSF IgG (10%).

Interferon alfa-2b (Intron A)

Interferon alfa-2b is a protein product manufactured by recombinant DNA technology. The mechanism of antitumor activity is not understood clearly; however, direct antiproliferative effects against malignant cells and modulation of host immune response may play important roles.

Doses and duration of treatment are as determined by the involved clinicians.

Class Summary

These agents disrupt DNA replication or cell division, inhibiting cell growth and proliferation. Prednisone (listed above) also can be included in this category.

Cyclophosphamide (Cytoxan, Neosar)

Cyclophosphamide is chemically related to nitrogen mustards. As an alkylating agent, the mechanism of action of the active metabolites may involve cross-linking of DNA, which may interfere with growth of normal and neoplastic cells. It is a component of CHOP and ProMACE-CytaBOM chemotherapeutic regimens.

Doxorubicin (Adriamycin, Rubex)

Doxorubicin inhibits topoisomerase II and produces free radicals, which may cause the destruction of DNA. The combination of these 2 events can, in turn, inhibit the growth of neoplastic cells. It is a component of the CHOP and ProMACE-CytaBOM chemotherapeutic regimens.

Vincristine (Oncovin, Vincasar PFS)

Vincristine's mechanism of action is uncertain. It may involve a decrease in reticuloendothelial cell function or an increase in platelet production. However, neither of these mechanisms would fully explain the effect in thrombotic thrombocytopenic purpura and hemolytic-uremic syndrome. It is a component of the CHOP and ProMACE-CytaBOM chemotherapeutic regimens.

Etoposide (Toposar, VePesid)

Etoposide inhibits topoisomerase II and causes DNA strand breakage, causing cell proliferation to arrest in late S or early G2 portion of the cell cycle. It is a component of the ProMACE-CytaBOM regimen.

Bleomycin (Blenoxane)

Bleomycin is a glycopeptide antibiotic that inhibits DNA synthesis. It is used as a palliative measure in the management of several neoplasms. It is a component of the ProMACE-CytaBOM regimen.

Methotrexate (Folex PFS, Rheumatrex)

Methotrexate is an antimetabolite that inhibits dihydrofolate reductase, thereby hindering DNA synthesis and cell reproduction in malignant cells. Satisfactory response is observed 3-6 weeks following administration. Adjust the dose gradually to attain a satisfactory response. It is a component of the ProMACE-CytaBOM regimen.

Fludarabine (Fludara, Oforta)

Fludarabine is a purine analog can be given PO or IV. It interferes with ribonucleotide reductase and DNA polymerase. It is active against both resting and dividing cells.