Pathogenesis Of Post-transplant Lymphoproliferative Disorders Research Articles

Natural Killer Cells in Post-Transplant Lymphoproliferative Disorders.

Simple SummaryPost-transplant lymphoproliferative disorders (PTLDs) are life-threatening complications. The clinical and pathological spectrum of PTLD is broad; however, most cases of PTLD are associated with Epstein–Barr virus (EBV) infection and the use of immunosuppression treatment required to prevent graft rejection. While T-cell impairment is known to play a critical role in the immunopathogenesis of EBV complications post-transplantation, the role of natural killer (NK) cells remains more elusive. NK cells are key elements of the innate immune system that use a sophisticated array of activating, costimulatory, and inhibitory receptors to kill virally infected and/or cancerous cells. In this review we highlight the role of NK cells in the pathogenesis of PTLD, and also identify future avenues for NK cell therapy research. Post-transplant lymphoproliferative disorders (PTLDs) are life-threatening complications arising after solid organ or hematopoietic stem cell transplantations. Although the majority of these lymphoproliferations are of B cell origin, and are frequently associated with primary Epstein–Barr virus (EBV) infection or reactivation in the post-transplant period, rare cases of T cell and natural killer (NK) cell-originated PTLDs have also been described. A general assumption is that PTLDs result from the impairment of anti-viral and anti-tumoral immunosurveillance due to the long-term use of immunosuppressants in transplant recipients. T cell impairment is known to play a critical role in the immune-pathogenesis of post-transplant EBV-linked complications, while the role of NK cells has been less investigated, and is probably different between EBV-positive and EBV-negative PTLDs. As a part of the innate immune response, NK cells are critical for protecting hosts during the early response to virus-induced tumors. The complexity of their function is modulated by a myriad of activating and inhibitory receptors expressed on cell surfaces. This review outlines our current understanding of NK cells in the pathogenesis of PTLD, and discusses their potential implications for current PTLD therapies and novel NK cell-based therapies for the containment of these disorders.

Post-transplant Lymphoproliferative Disorders: Single Center Case Series and Literature Review

Post-transplant lymphoproliferative disorders (PTLD) are referred to lymphoid and/or plasmacytic proliferation which occur as result of immunosuppression therapy in patient underwent solid organ or allogeneic hematopoietic stem cell transplantation. Among solid organ transplant recipients, it accounts approximately 20% of all cancers. Epstein-Barr virus (EBV) has been linked to the pathogenesis of PTLD when EBV was identified in the tumor biopsies. In most affected patients, PTLD occurs as a result of proliferation of EBV positive B cell following immunosuppression and impaired Tcell immune activity. EBV negative PTLD has been documented but no clear etiology has been confirmed, however theories of previous exposure to EBV which is completely cleared at the time of PTLD is diagnosed, different viruses or chronic antigenic stimulation all been considered as provoking factors for tumor development. Clinical symptomatology at presentation of patients with PTLD is usually indistinct from de novo lymphoproliferative disorders and chemotherapy protocols in this group of patients are generally similar to the standard of care of lymphoma treatment according to the subtype in addition to cessation or dose reduction of immunosuppressive therapy. We report our experience and outcome analysis in PTLD management among 23 patients from our institution treated between 2011 and 2019.

Comparative analysis of post-transplant lymphoproliferative disorders after solid organ and hematopoietic stem cell transplantation reveals differences in the tumor microenvironment

Post-transplant lymphoproliferative disorders (PTLD) occur after solid organ transplantation (SOT) or hematopoietic stem cell transplantation (HCT) and are frequently associated with Epstein-Barr virus (EBV). Because of the complex immune setup in PTLD patients, the tumor microenvironment (TME) is of particular interest to understand PTLD pathogenesis and elucidate predictive factors and possible treatment options. We present a comparative study of clinicopathological features of 48 PTLD after HCT (n = 26) or SOT (n = 22), including non-destructive (n = 6), polymorphic (n = 23), and monomorphic (n = 18) PTLD and classic Hodgkin lymphoma (n = 1). EBV was positive in 35 cases (73%). A detailed examination of the TME with image analysis-based quantification in 22 cases revealed an inflammatory TME despite underlying immunosuppression and significant differences in its density and composition depending on type of transplant, PTLD subtypes, and EBV status. Tumor-associated macrophages (TAMs) expressing CD163 (p = 0.0022) and Mannose (p = 0.0016) were enriched in PTLD after HCT. Double stains also showed differences in macrophage polarization, with more frequent M1 polarization after HCT (p = 0.0321). Higher counts for TAMs (CD163 (p = 0.0008) and cMaf (p = 0.0035)) as well as in the T cell compartment (Granzyme B (p = 0.0028), CD8 (p = 0.01), and for PD-L1 (p = 0.0305)) were observed depending on EBV status. In conclusion, despite the presence of immunosuppression, PTLD predominantly contains an inflammatory TME characterized by mostly M1-polarized macrophages and cytotoxic T cells. Status post HCT, EBV positivity, and polymorphic subtype are associated with an actively inflamed TME, indicating a specific response of the immune system. Further studies need to elucidate prognostic significance and potential therapeutic implications of the TME in PTLD.

Genomic variations in EBNA3C of EBV associate with posttransplant lymphoproliferative disorder.

Epstein-Barr Virus (EBV) is a ubiquitous virus linked to a variety of lymphoid and epithelial malignancies. In solid organ and hematopoietic stem cell transplant recipients, EBV is causally associated with posttransplant lymphoproliferative disorder (PTLD), a group of heterogeneous lymphoid diseases. EBV+ B cell lymphomas that develop in the context of PTLD are generally attributed to the immunosuppression required to promote graft survival, but little is known regarding the role of EBV genome diversity in the development of malignancy. We deep-sequenced the EBV genome from the peripheral blood of 18 solid organ transplant recipients, including 6 PTLD patients. Sequences from 6 EBV+ spontaneous lymphoblastoid B cell lines (SLCL) were similarly analyzed. The EBV genome from PTLD patients had a significantly greater number of variations than EBV from transplant recipients without PTLD. Importantly, there were 15 nonsynonymous variations, including 8 in the latent cycle gene EBNA3C that were associated with the development of PTLD. One of the nonsynonymous variations in EBNA3C is located within a previously defined T cell epitope. These findings suggest that variations in the EBV genome can contribute to the pathogenesis of PTLD.

Epstein-Barr Virus-Associated Post-Transplant Lymphoproliferative Disorders after Hematopoietic Stem Cell Transplantation: Pathogenesis, Risk Factors and Clinical Outcomes

Epstein-Barr virus (EBV) is a ubiquitous virus belonging to the human γ-herpes virus subfamily. After primary infection, EBV maintains a life-long latent infection. A major concern is that EBV can cause a diverse range of neoplasms and autoimmune diseases. In addition, patients undergoing hematopoietic stem cell transplantation or solid organ transplantation can experience post-transplant lymphoproliferative disorders (PTLDs) due to dysfunction or suppression of host’s immune system, or uncontrolled proliferation of EBV-infected cells. In recent years, the number of EBV-associated PTLD cases has increased. This review focuses on the current understandings of EBV-associated PTLD pathogenesis, as well as the risk factors and clinical outcomes for patients after allogeneic stem cell transplantation.

Recurrent JAK3 Mutations in Post-Transplant Lymphoproliferative Disorder

Post-transplant lymphoproliferative disorders (PTLD) are diverse in morphology, immunophenotype and clinical outcomes, occurring in immune compromised patients after hematopoietic stem cell transplantation (HCT) or single organ transplantation (SOT). Predicting the outcome of PTLDs is complex. In most of cases PTLD is driven by Epstein-Barr virus (EBV) infection, though the etiology of EBV negative cases is unknown. There have been only few studies investigating the mutational landscape and gene expression of PTLD. In our study we performed targeted DNA deep sequencing and RNA expression profiling (RNA-Seq) on the hematopoietic lymphoid tissues of 15 post-transplant patients with PTLD and 16 post-transplant patients with follicular hyperplasia (FH) after HCT or SOT. Recurrent mutations in JAK3 were identified in five of 16 PTLD cases and one of 16 follicular hyperplasia cases. We additionally identified mutations in the PIK3CD gene in four PTLD cases and NOTCH1 mutations in 2 PTLD and four FH cases. 17 other genes were commonly mutated in both groups. Gene expression profiles differed between both groups. Genes involved in regulating the immune system were unregulated in PTLD. We also saw a stronger expression of T-cells and a weaker expression of B-cells within the PTLD group when compared with FH. Our findings suggest an important role for immune regulatory pathways in the pathogenesis of PTLD, with the JAK/STAT pathway being a central recurrent key metabolic pathway. Figure Disclosures Ohgami: Agilent technologies: Other: received support/funding.

Non-Destructive Post-Transplant Lymphoproliferative Disorder Shows Distinct Clinical Features with HHV-6 Reactivation in a Subset

Background: Post-transplant lymphoproliferative disorders (PTLD) are a diverse group of lymphoid neoplasms arising after solid organ (SOT) or hematopoietic stem cell transplant (HSCT). They may resemble lymphomas in immunocompetent patients (monomorphic); present as destructive, heterogeneous lesions (polymorphic); or be non-destructive but mass-forming (non-destructive, previously known as 'early lesions'). PTLDs are often associated with Epstein-Barr virus (EBV) infection or reactivation, which likely functions as a driver in these cases. Non-destructive lesions may be challenging to diagnose in the absence of EBV. However, a significant fraction of PTLDs are EBV(-), including a subset of non-destructive lesions. The pathogenesis of these remains to be fully elucidated. Human herpesvirus 6 (HHV-6) reactivation is common during transplantation, occurring in up to 70% of HSCT and 80% of SOT patients. While it has been hypothesized that HHV-6 may play a role in PTLD, particularly in EBV(-) cases, the role of HHV-6 in the pathogenesis of PTLD remains underexplored. Methods: Following IRB approval, pathology archives at Weill Cornell/New York Presbyterian Hospital were searched to identify patients with a tissue diagnosis of PTLD and correlated with clinical information. A tissue microarray was constructed from blocks with sufficient tissue. Standard immunohistochemistry (IHC) and in situ hybridization for EBV-encoded RNA (EBER) were performed, and IHC conditions were optimized for antibodies against HHV-6 gp60/110 (Millipore Sigma, MAB853) and p41 (Santa Cruz Biotechnologies, 9A5D12). Statistical comparisons utilized Student's t-test and Fisher's exact test. Results: We identified 79 patients from 6/23/94-6/6/19 with a tissue diagnosis of PTLD. Morphologic subtype could be determined in 74, including 8 patients with non-destructive lesions. Compared to monomorphic or polymorphic, patients with non-destructive PTLDs were significantly younger (median age 9, p<0.01) and more likely status post heart transplant (5/8 cases, p<0.001, Table 1). By contrast, polymorphic PTLDs occurred relatively more frequently in HSCT patients (25/33 cases, p<0.01). All morphologic subtypes were frequently EBV+ (63-88%) but EBV positivity was greater in polymorphic than monomorphic lesions (p<0.05). While tissue testing failed on blocks > 20 years old, 13 PTLD cases from 12 patients had sufficient evaluable material for HHV-6 testing (Table 2). 9/12 patients had a SOT (8 renal, 1 heart) and 3 had HSCT. HHV-6 was identified in 5/13 lesions (38%), all of which were in SOT patients (Figure 1). EBER was detected in 8/13 lesions (62%). When stratified by PTLD subtype, 3/3 non-destructive PTLDs were HHV-6+ (100%). Two were EBER(-) and the remaining case only showed scattered EBER+ cells. Unfortunately, material was not available for testing the EBER+ non-destructive lesions. 2/9 of the monomorphic lesions tested (22%) were positive for both HHV-6 and EBER. While the series is limited, the association between HHV-6 and non-destructive lesions, compared to other types of PTLD, appears significant (p<0.05). EBV did not appear to be associated with HHV-6 status (p = 0.29). Conclusions: Non-destructive, monomorphic and polymorphic PTLDs show significant differences in timing, patient demographics and transplant type, suggesting that they may differ in associated risk factors affecting their pathogenesis. Confirming prior studies, we find non-destructive lesions tend to occur in younger patients, often following SOT as compared to HSCT and with a median time from transplantation of ~50 months. In addition, the presence of HHV-6 in the absence of (or with limited) EBV in the 3 non-destructive lesions tested raises the possibility that HHV-6 may play a role in the pathogenesis of this PTLD subtype that has yet to be defined. These findings are intriguing given the frequent reactivation of HHV-6 seen in transplant populations. HHV-6 has been variably associated with post-transplant complications including pneumonitis, delayed hematopoietic engraftment and graft versus host disease and thus received relatively more attention in the HSCT population. A causative role for HHV-6 in PTLD remains speculative, but these findings raise the possibility that clinicians treating the SOT population should also consider aggressively identifying and treating HHV-6 reactivation. Disclosures Rutherford: AstraZeneca: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Heron: Consultancy, Honoraria; Janssen Scientific Affairs: Consultancy, Honoraria; Juno Therapeutics Inc: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria; Verastem: Consultancy, Honoraria; Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Immune activation, immune senescence and levels of Epstein Barr Virus in kidney transplant patients: Impact of mTOR inhibitors.

Post-transplant lymphoproliferative disorders (PTLD) represent a severe complication in transplanted patients and Epstein-Barr Virus (EBV) is the main driver. Besides immunodepression, immune activation/chronic inflammation play an important role in both virus reactivation and expansion of EBV-positive B cells. The aim of this study was to assess the impact of immunosuppressive strategies on factors involved in the PTLD's pathogenesis. 124 kidney transplanted patients were enrolled in this study: 71 were treated with mycophenolic acid (MPA) and 53 treated with mTOR inhibitor (mTORi), both in combination with different doses of calcineurin inhibitor. At the time of the transplant (T0), profile of inflammation/immune activation and immune senescence didn't differ between the two groups, but after one year of treatment (T1) markers were significantly higher in MPA-treated patients; their immunosenescence process was supported by the greater erosion of telomeres despite their younger age. Percentages of activated B cells and levels of EBV-DNA significantly increased in MPA-treated patients, and at T1 were significantly higher in MPA- than in mTORi-treated patients. Overall, these findings indicate that mTOR inhibitors constrain the inflammation/immune activation and senescence status, thus reducing the expansion of EBV-infected B cells and the risk of virus-associated PTLD in kidney transplant recipients.

Management of post-transplant lymphoproliferative disorders.

Management of post-transplant lymphoproliferative disorders.

The pathology of epstein-barr virus lymphoproliferations.

The pathology of epstein-barr virus lymphoproliferations.

Clinicopathologic spectrum and EBV status of post-transplant lymphoproliferative disorders after allogeneic hematopoietic stem cell transplantation

Post-transplant lymphoproliferative disorders (PTLDs) are serious, life-threatening complications of solid-organ transplantation (SOT) and bone marrow transplantation, and are associated with high mortality. PTLDs represent a heterogeneous group of lymphoproliferative diseases, which show a spectrum of clinical, morphologic, and molecular genetic features ranging from reactive polyclonal lesions to frank lymphomas. We describe clinicopathologic features of 17 cases of PTLD after allogeneic hematopoietic stem cell transplantation (allo-HSCT), which were analyzed by in situ hybridization for EBV and a panel of antibodies directed against numerous antigens, including CD20, PAX5, CD3, bcl-6, CD10, MUM-1/IRF4, CD138, Kappa, Lambda, CD30, CD15, and Ki67. The cases included 13 males and 4 females with a median age of 31years (range 9-49years) and the PTLDs developed 1.5-19months post-transplant (mean 4.7months). The histological types indicated five cases of early lesions, two of plasmacytic hyperplasia and three of infectious mononucleosis-like PTLD. Eight cases were polymorphic PTLD, and four were monomorphic PTLD, including three of diffuse large B cell lymphoma, and one of plasmablastic lymphoma. Foci and sheets of necrosis were observed in five cases. The infected ratio of EBV was 88.2%. Some cases were treated by reduction of immunosuppression, antiviral therapy, donor lymphocyte infusion, or anti-CD20 monoclonal rituximab. Eight cases died. The first half year after allo-HSCT is very important for the development of PTLD. The diagnosis of PTLD relies on morphology and immunohistochemistry, and EBV plays an important role in the pathogenesis of PTLD. The prognosis of PTLD is poor, and, notably, PTLD after allo-HSCT exhibits some features different from those of PTLD after SOT.

Post-transplant lymphoproliferative disorder: a clinicopathologic study of 15 cases

To study the clinical and histopathologic features, diagnosis, pathogenesis and therapy of post-transplant lymphoproliferative disorders (PTLD). The clinical and pathologic features of 15 cases of PTLD were retrospectively analyzed by light microscopy, immunohistochemistry and in-situ hybridization, according to the updated 2008 WHO classification of tumors of hematopoietic and lymphoid tissues. Amongst the 15 cases studied, 14 cases had received allogenic hematopoietic stem cell transplantation (AHSCT) and 1 case had received renal transplantation. There were altogether 12 males and 3 females. The male-to-female ratio was 4:1. The mean age was 30.4 years and the median age was 31 years (range from 9 to 60 years). PTLD developed 1.5 to 132 months after transplantation (median 13.0 months). The mean age of the 14 patients with AHSCT was 28.3 years (range from 9 to 45 years) and PTLD developed 1.5 to 19 months after transplantation (mean 4.5 months). Major clinical presentation included fever and lymphadenopathy. Twelve cases involved mainly lymph nodes and the remaining 3 cases involved tonsils, stomach and small intestine, respectively. The histologic types in 4 cases represented early lesions, including plasmacytic hyperplasia (n = 1) and infectious mononucleosis-like PTLD (n = 3). Seven cases were polymorphic PTLD, with 4 cases containing a predominance of large cells. Graft-versus-host disease was also seen in the case of small intestinal involvement. Four cases were monomorphic PTLD, 3 of which were diffuse large B-cell lymphoma, 1 was plasmablastic lymphoma and 1 was a mixture of monomorphic and polymorphic PTLD. Foci of necrosis were seen in 5 cases. The proliferating index of Ki-67 was high. The positive rate of EBV-encoded RNA in AHSCT was 92.9%. The duration of PTLD onset was shorter in EBV-positive cases (range from 1.5 to 7 months) than EBV-negative cases (range from 19 and 132 months). Some cases were treated by reduction of immunosuppression, antiviral agents or anti-CD20 monoclonal antibody Rituximab. The duration of follow-up in 14 patients ranged from 0 to 8 months. Five of the patients died of the disease. The diagnosis of PTLD relies on morphologic examination and immunohistochemistry. Most of them are of B-cell origin. EBV plays an important role in the pathogenesis of PTLD. The duration of disease onset is shorter in EBV-positive cases. PTLD in AHSCT cases occurs in younger age group, with shorter duration of onset, as compared to solid organ transplantation. The prognosis of PTLD is poor. The modalities of treatment include reduction of immunosuppression, antiviral agents or anti-CD20 monoclonal antibody Rituximab.

Posttransplant Lymphoproliferative Disorders

Posttransplant lymphoproliferative disorders (PTLDs) are a group of diseases that range from benign polyclonal to malignant monoclonal lymphoid proliferations. They arise secondary to treatment with immunosuppressive drugs given to prevent transplant rejection. Three main pathologic subsets/stages of evolution are recognised: early, polymorphic, and monomorphic lesions. The pathogenesis of PTLDs seems to be multifactorial. Among possible infective aetiologies, the role of EBV has been studied in depth, and the virus is thought to play a central role in driving the proliferation of EBV-infected B cells that leads to subsequent development of the lymphoproliferative disorder. It is apparent, however, that EBV is not solely responsible for the “neoplastic” state. Accumulated genetic alterations of oncogenes and tumour suppressor genes (deletions, mutations, rearrangements, and amplifications) and epigenetic changes (aberrant hypermethylation) that involve tumour suppressor genes are integral to the pathogenesis. Antigenic stimulation also plays an evident role in the pathogenesis of PTLDs. Plasmacytoid dendritic cells (PDCs) that are critical to fight viral infections have been thought to play a pathogenetically relevant role in PTLDs. Furthermore, regulatory T cells (Treg cells), which are modulators of immune reactions once incited, seem to have an important role in PTLDs where antigenic stimulation is key for the pathogenesis.

Post-transplant lymphoproliferative disorders and Epstein-Barr virus DNAemia in a cohort of lung transplant recipients

BackgroundPost-transplant lymphoproliferative disorders (PTLD) are serious complications in lung transplant recipients. No consensus on EBV DNAemia levels predictive of PTLD has been reached. In addition, in many instances EBV DNAemia is determined in patients with suggestive symptoms only.MethodsThe characteristics of five patients with PTLD as well as the prevalence of EBV DNAmia in a cohort of 137 consecutive patients receiving lung transplantation are described.ResultsTwenty-six out of 137 patients (18.9%) were excluded from the analysis because lost at follow-up or dead from PTLD-independent reasons within three months of transplantation. EBV DNA in peripheral blood mononuclear cells (PBMC) was determined in 83/111 patients (74.8%) because of potential PTLD-related symptoms, while 28 patients (25.2%) showed no symptoms and were not examined. EBV DNAemia was positive in 53/83 patients (63.8%), and negative in 30/83 patients (36.2%). PTLD was diagnosed in five (4.5%) patients at a median time of 270 (range 120-870) days following transplantation. All five PTLD (three large B-cell lymphomas, one Hodgkin lymphoma and one possible pre-neoplastic lesion) were potentially associated with EBV infection. However, only 3/5 patients with PTLD had detectable EBV DNAemia: < 1,000 copies EBV DNA/1 × 105 PBMC in one patient and > 1,000 copies EBV DNA/1 × 105 PBMC in two patients.ConclusionA systematic multidisciplinary (clinical, radiologic, virologic and histologic) approach is mandatory for the diagnosis and management of PTLD in lung transplant recipients, while monitoring of symptomatic patients only may provide an incomplete or late picture of the clinical problem. In addition, staining for EBV antigens and quantification of EBV DNA in biopsy specimens should always be performed to understand the role of EBV infection in the pathogenesis of PTLD.

IL-6 and IL-10 in post-transplant lymphoproliferative disorders development and maintenance: a longitudinal study of cytokine plasma levels and T-cell subsets in 38 patients undergoing treatment

IL-6 and IL-10 have previously been implicated in the pathogenesis of post-transplant lymphoproliferative disorders (PTLD) and, like peripheral lymphocyte populations, are markers of immune status that are amenable to study in vivo. Thus, we analyzed cytokine plasma levels as well as lymphocyte subsets in a longitudinal analysis of 38 adult transplant recipients undergoing treatment for PTLD. Pretherapeutically, we found significantly elevated IL-6 (13.8 pg/ml) and IL-10 plasma levels (54.7 pg/ml) - in the case of IL-10, even higher in treatment nonresponders than in responders (116 vs. 14 pg/ml). Over time, however, IL-10 levels did not correlate with the course of disease, whereas those of IL-6 did, falling in responders and rising in nonresponders. These findings were independent of histological EBV-status, treatment type, and total peripheral T-cell counts, which were significantly reduced in patients with PTLD. Our observations support the idea that although IL-10 is important for creating a permissive environment for post-transplant lymphoma development, IL-6 is associated with PTLD proliferation. The analysis of lymphocyte subsets further identified HLA-DR+ CD8+ lymphocyte numbers as significantly different in non-PTLD controls (33%), treatment responders (44%) and nonresponders (70%). Although the specificity of these cells is unclear, their increase might correlate with the impaired tumor-specific cytotoxic-T-lymphocyte (CTL)-response in PTLD.

Posttransplant Lymphoproliferative Disorder Following Kidney Transplant

Posttransplant Lymphoproliferative Disorder Following Kidney Transplant

Molecular characterization of post‐transplant lymphoproliferative disorders of donor origin occurring in liver transplant recipients

Post-transplant lymphoproliferative disorders (PTLDs) represent a frequent complication of solid organ transplantation. Although most PTLDs arise from recipient lymphoid cells, a considerable fraction of cases may arise from donor B-cells. In an attempt to clarify the histogenesis and pathogenesis of PTLDs derived from donor B-cells, monoclonal PTLDs occurring in liver transplant recipients were chosen as a model to compare donor (D-PTLDs) versus recipient PTLDs (R-PTLDs). The tumour panel included nine D-PTLDs and six R-PTLDs. D-PTLDs were early-onset, EBV-infected lymphoproliferations classified as polymorphic PTLD (P-PTLD; n = 7) or diffuse large B-cell lymphoma (DLBCL; n = 2) with tumour localization confined to the hepatic hilum. All R-PTLDs were late-onset DLBCLs and showed extrahepatic localization. A BCL-6(-)/MUM1(+)/CD138(+/-) phenotype, consistent with a post-germinal centre (GC) stage of pre-terminal B-cell differentiation, was observed in all D-PTLDs and in 2/6 R-PTLDs, whereas a BCL6(+)/MUM1(-)/CD138(-) profile, reminiscent of GC B-cells, was detected in 4/6 R-PTLDs. The presence of somatic IGHV hypermutation was observed in 6/9 D-PTLDs and in 4/6 R-PTLDs, suggesting derivation from antigen-experienced B-cells. IGHV4-39 was the IGHV gene most frequently encountered, being rearranged in 3/9 D-PTLDs. Among IGHV-mutated PTLDs, a mutational profile suggesting antigen stimulation and/or selection was observed in 4/6 D-s and in 2/4 R-PTLDs. The presence of ongoing IGHV mutations was detected in 2/4 D-PTLDs. Aberrant SHM was detected in 10/15 (66.7%) PTLDs, including 6/9 D-PTLDs and 4/6 R-PTLDs. Our findings suggest that (i) D-PTLDs show a clinical presentation distinct from R-PTLDs; (ii) immunophenotypic and genetic features of D-PTLDs are consistent with mature, GC-experienced B-cells; (iii) transformed donor-derived B-cells may experience antigen-driven stimulation and selection, and may acquire genetic lesions during neoplastic expansion in the recipient environment; and (iv) EBV infection and expression of viral oncoproteins may be relevant in the pathogenesis of D-PTLDs.

Post-Transplant Lymphoproliferative Disorders: Role Of Viral Infection, Genetic Lesions And Antigen Stimulation In The Pathogenesis Of The Disease

Post-transplant lymphoproliferative disorders (PTLD) are a life-threatening complication of solid organ transplantation or, more rarely, hematopoietic stem cell transplantation. The majority of PTLD is of B-cell origin and associated with Epstein–Barr virus (EBV) infection. PTLD generally display involvement of extranodal sites, aggressive histology and aggressive clinical behavior. The molecular pathogenesis of PTLD involves infection by oncogenic viruses, namely EBV, as well as genetic or epigenetic alterations of several cellular genes. At variance with lymphoma arising in immunocompetent hosts, whose genome is relatively stable, a fraction of PTLD are characterized by microsatellite instability as a consequence of defects in the DNA mismatch repair mechanism. Apart from microsatellite instability, molecular alterations of cellular genes recognized in PTLD include alterations of cMYC, BCL6, TP53, DNA hypermethylation, and aberrant somatic hypermutation of protooncogenes. The occurrence of IGV mutations in the overwhelming majority of PTLD documents that malignant transformation targets germinal centre (GC) B-cells and their descendants both in EBV–positive and EBV–negative cases. Analysis of phenotypic markers of B-cell histogenesis, namely BCL6, MUM1 and CD138, allows further distinction of PTLD histogenetic categories. PTLD expressing the BCL6+/MUM1+/-/CD138− profile reflect B-cells actively experiencing the GC reaction, and comprise diffuse large B-cell lymphoma (DLBCL) centroblastic and Burkitt lymphoma. PTLD expressing the BCL6−/MUM1+/CD138− phenotype putatively derive from B-cells that have concluded the GC reaction, and comprise the majority of polymorphic PTLD and a fraction of DLBCL immunoblastic. A third group of PTLD is reminiscent of post-GC and preterminally differentiated B-cells that show the BCL6−/MUM1+/CD138+ phenotype, and are morphologically represented by either polymorphic PTLD or DLBCL immunoblastic.

Genetic and phenotypic analysis of B‐cell post‐transplant lymphoproliferative disorders provides insights into disease biology

B-cell post-transplant lymphoproliferative disorders (PTLD) are classified as early lesions, polymorphic lymphomas (P-PTLD) and monomorphic lymphomas (M-PTLD). These morphologic categories are thought to reflect a biologic continuum, although supporting genetic data are lacking. To gain better insights into PTLD pathogenesis, we characterized the phenotypes, immunoglobulin (Ig) gene alterations and non-Ig gene (BCL6, RhoH/TTF, c-MYC, PAX5, CIITA, BCL7A, PIM1) mutations of 21 PTLD, including an IM-like lesion, 8 P-PTLD and 12 M-PTLD. Gene expression profile analysis was also performed in 12 cases. All PTLD with clonal Ig rearrangements showed evidence of germinal centre (GC) transit based on the analysis of Ig and BCL6 gene mutations, and 74% had a non-GC phenotype (BCL6 +/- MUM1+). Although surface Ig abnormalities were seen in 6/19 (32%) PTLD, only three showed 'crippling' Ig mutations indicating other etiologies for loss of the B-cell receptor. Aberrant somatic hypermutation (ASHM) was almost exclusively observed in M-PTLD (8/12 vs. 1/8 P-PTLD) and all three recurrent cases analysed showed additional mutations in genes targeted by ASHM. Gene expression analysis showed distinct clustering of PTLD compared to B-cell non-Hodgkin lymphomas (B-NHL) without segregation of P-PTLD from non-GC M-PTLD or EBV+ from EBV- PTLD. The gene expression pattern of PTLD appeared more related to that of memory and activated B-cells. Together, our results suggest that PTLD represent a distinct type of B-NHL deriving from an antigen experienced B-cell, whose evolution is associated with accrual of genetic lesions.

Evidence for Genetic Susceptibility Towards Development of Posttransplant Lymphoproliferative Disorder in Solid Organ Recipients

Posttransplant lymphoproliferative disorder (PTLD) is a life-threatening complication after organ transplantation. The identification of risk factors for PTLD development is important for disease management. It has been shown that cytokine gene polymorphisms are associated with lymphoma and Epstein-Barr virus (EBV)-associated diseases in nonimmunosuppressed patients. In the present case-control study, we analyzed the impact of -1082 interleukin (IL)-10, -308 tumor necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta1 (codon 10, 25), and +874 interferon (IFN)-gamma gene single-nucleotide polymorphisms on the late onset EBV-associated PTLD. Out of 1,765 solid organ recipients, 38 patients with late-onset EBV-associated PTLD and 408 matched solid organ recipients were selected and enrolled in the study. Single nucleotide polymorphisms (SNPs) for -1082IL-10, -308TNF-alpha, TGF-beta1 (codon 10, 25), and +874IFN-gamma genes were analyzed by a sequence specific primer polymerase chain reaction and were related to the PTLD development, and the disease course and outcome. The TGF-beta1 (codon 25) GG genotype was detected more frequently in controls than in PTLD patients (odds ratio=0.34, 95% confidence interval: 0.17-0.69, P=0.0022). The frequency of -1082 IL-10 GG genotype was also significantly higher in controls than in PTLD patients (odds ratio=0.5, 95% confidence interval: 0.25-1.0, P=0.044). There were no associations between -308TNF-alpha, TGF-beta1 codon 10, and +874IFN-gamma SNPs and PTLD. Disease course and outcome were not associated with any cytokine SNPs. Polymorphisms in two key anti-inflammatory cytokines, IL-10 and TGF-beta, are associated with susceptibility to EBV-associated PTLD, suggesting that a shift in pro-/anti-inflammatory response is involved in the pathogenesis of PTLD.