EBV-positive Post-transplant Lymphoproliferative Disorders Research Articles

Epidemiology of Epstein-Barr Virus Chronic High Viral Load in Kidney Transplant Recipients

Background. Epstein-Barr virus (EBV) chronic high viral load (CHVL) may be defined by >16 000 copies/mL whole blood or >200 copies/105 peripheral blood mononuclear cells in >50% samples exceeding 6 mo. EBV CHVL has only been characterized in a few small pediatric studies, with heterogeneous results and unclear clinical significance. Methods. This single-center observational study evaluated adult and pediatric kidney transplant recipients transplanted between 2010 and 2021 on tacrolimus/mycophenolate-based/prednisone immunosuppression. The primary outcome was EBV CHVL prevalence. Secondary outcomes included recipient characteristics, DNAemia kinetics, and posttransplant lymphoproliferative disorder (PTLD) in recipients with EBV CHVL versus low-grade DNAemia or no DNAemia. Results. Five hundred forty-one recipients had a mean follow-up of 4.6 y. Fourteen recipients (2.6%) developed EBV CHVL, 70 (12.9%) had low-grade EBV DNAemia, and 457 (84.5%) had no EBV DNAemia. EBV CHVL was more common in recipients who were Caucasian (P = 0.04), younger (P = 0.04), received induction immunosuppression (P = 0.02), and had high-risk donor–recipient EBV serologic mismatch (P < 0.0001). CHVL patients had a higher first viral load (P = 0.03), longer time to maximum viral load (P = 0.02), and did not achieve sustained DNAemia clearance versus low-grade DNAemia. Three EBV-positive PTLD cases occurred in recipients with a history of EBV DNAemia. PTLD was present in 7.1% (1/14) CHVL versus 2.9% (2/70) low-grade DNAemia patients (P = 0.002). EBV DNAemia developed in 32 EBV seronegative recipients (32/59; 54%); clearance was achieved in 70% (14/20) with low-grade DNAemia but no CHVL (0/12; P = 0.0001). Conclusions. CHVL was uncommon and appeared to occur after primary EBV infection. Future studies should explore other potentially modifiable risk factors for PTLD, including optimal management of EBV DNAemia.

Growth Transformation of B Cells by Epstein-Barr Virus Requires IMPDH2 Induction and Nucleolar Hypertrophy.

The in vitro growth transformation of primary B cells by Epstein-Barr virus (EBV) is the initial step in the development of posttransplant lymphoproliferative disorder (PTLD). We performed electron microscopic analysis and immunostaining of primary B cells infected with wild-type EBV. Interestingly, the nucleolar size was increased by two days after infection. A recent study found that nucleolar hypertrophy, which is caused by the induction of the IMPDH2 gene, is required for the efficient promotion of growth in cancers. In the present study, RNA-seq revealed that the IMPDH2 gene was significantly induced by EBV and that its level peaked at day 2. Even without EBV infection, the activation of primary B cells by the CD40 ligand and interleukin-4 increased IMPDH2 expression and nucleolar hypertrophy. Using EBNA2 or LMP1 knockout viruses, we found that EBNA2 and MYC, but not LMP1, induced the IMPDH2 gene during primary infections. IMPDH2 inhibition by mycophenolic acid (MPA) blocked the growth transformation of primary B cells by EBV, leading to smaller nucleoli, nuclei, and cells. Mycophenolate mofetil (MMF), which is a prodrug of MPA that is approved for use as an immunosuppressant, was tested in a mouse xenograft model. Oral MMF significantly improved the survival of mice and reduced splenomegaly. Taken together, these results indicate that EBV induces IMPDH2 expression through EBNA2-dependent and MYC-dependent mechanisms, leading to the hypertrophy of the nucleoli, nuclei, and cells as well as efficient cell proliferation. Our results provide basic evidence that IMPDH2 induction and nucleolar enlargement are crucial for B cell transformation by EBV. In addition, the use of MMF suppresses PTLD. IMPORTANCE EBV infections cause nucleolar enlargement via the induction of IMPDH2, which are essential for B cell growth transformation by EBV. Although the significance of IMPDH2 induction and nuclear hypertrophy in the tumorigenesis of glioblastoma has been reported, EBV infection brings about the change quickly by using its transcriptional cofactor, EBNA2, and MYC. Moreover, we present here, for the novel, basic evidence that an IMPDH2 inhibitor, namely, MPA or MMF, can be used for EBV-positive posttransplant lymphoproliferative disorder (PTLD).

Primary CNS EBV-positive post-transplant lymphoproliferative disorder with polymorphic and classic Hodgkin lymphoma features: A case report and literature review.

Post-transplant lymphoproliferative disorders (PTLD) are typically Epstein-Barr virus (EBV)-associated lymphoid or plasmacytic proliferations that occur when immunosuppressed after transplantation. Only 2 cases of primary central nervous system (PCNS) classic Hodgkin lymphoma PTLD and 1 case of PCNS Hodgkin lymphoma-like PTLD have been previously reported. A 59-year-old male presented with malaise, headaches, and dizziness; neuroimaging revealed a 1.7-cm right cerebellar mass and a 0.6-cm right frontal mass. Microscopic examination demonstrated a perivascular and parenchymal polymorphous infiltrate composed of lymphocytes (CD3-positive T cells and CD20-positive B cells), plasma cells, and macrophages. Focally, macrophages had a spindled morphology with a fascicular arrangement amounting to poorly formed granulomata. Mitoses were seen. Scattered large atypical cells were visualized with irregular hyperchromatic nuclei, reminiscent of lacunar cells, mononuclear Hodgkin and binucleate Reed-Sternberg (RS) cells. EBV in situ highlighted a significant number of small lymphoid cells as well as many large atypical forms. Large atypical cells were seen to co-express CD15 and CD30. To our knowledge, this is the first such case with hybrid polymorphic PTLD and classic Hodgkin lymphoma features and the first such case to arise following liver transplantation. This case highlights the histological and immunophenotypic spectrum of these lymphoid proliferations and the resulting challenges in diagnosis and definitive subtyping.

Genomic Landscaping of Post-Transplant Lymphoproliferative Disorders Using Circulating Tumor DNA

Introduction: Identification of post-transplant lymphoproliferative disorder (PTLD) is a diagnostic challenge. The current diagnostic framework usually require analysis on tissue and imaging through PET/CT scans. Yet, these methods have diagnostic limitations. For instance due to missing information on tumor heterogeneity and relatively low sensitivity of PET/CT scans. Measuring cell free DNA (cfDNA) represents a novel, minimally-invasive method for tumor detection and genotyping. In this study we explored the feasibility of plasma cfDNA tumor genotyping with targeted deep sequencing to detect copy number variation (CNV), single nucleotide variants (SNVs) and Epstein Bar Virus (EBV) sequences, using a new in-house variant calling pipeline. Methods: In this multi-center observational cohort study of 18 patients diagnosed with monomorphic PTLD, we performed deep targeted NGS for SNV detection and low-coverage whole genome sequencing (lcWGS) for CNV detection. NGS was targeted on a panel of genes (n = 39) known to be recurrently mutated in B-cell lymphomas (244 kbp). All patients gave written informed consent. The R package CNAclinic was used to identify genome wide CNAs and fraction of the genome altered (FGA). The ichorCNA package was used to determine the ctDNA fraction in cfDNA samples. SNVs were called using a pipeline, combining 4 (cfDNA) SNV calling tools: Mutect2, LoFreq, VarDict & SiNVICT. Variants were filtered on presumed functional effect (Synonymous-, UTR-, Intron-, 2 kb upstream-, 2 kb downstream variants were removed). A "panel of normals” (variant blacklist) was generated from variants found in cfDNA from healthy controls, which was used to produce a variant blacklist to filter out technical artifacts and SNPs. Finally, variants with a variant allele frequency (VAF) < 1% and variants with an allele frequency (AF) of > 1% in 1000Genomes and/or gnomAD were filtered out. Results: The cohort included 18 PTLD patients. The majority of patients had stage IV disease (n=13, 76%). All of the PTLD cases were histologically proven, of diffuse large B cell lymphoma type and 10/18 patients (55%) were considered to have an EBV-negative tumor. EBV copy number in plasma was performed by qPCR in 12 patients and was considered to be elevated in four patients (>5000 copies/mL). The fraction of reads mapping to the EBV genome was compared between controls (n=8), EBV-negative PTLD (n=10) and EBV-positive PTLD (n=8). The median fraction of EBV reads for controls and EBV-negative PTLD were similar with a fraction of 1.5x10-4 (Q1-Q3: 1.3x10-4-1.6x10-4) and 1.8x10-4 (Q1-Q3: 1.4x10-4-3.9x10-4), respectively (p>0.05, Kruskal-Wallis). In EBV-positive tumors, the median fraction of EBV reads was 3-log higher, 5.3x10-1 (Q1-Q3: 7.3 x10-2-2) (p≤0.05, Kruskal-Wallis) CNVs were detected in cfDNA of 11/18 (61%) patients. Two of the patients in whom no CNVs were detected had initiated therapy at the time of plasma sampling. Median fraction of genome altered was higher in EBV-negative tumors (0.03, Q1-Q3: 2.5x10-4-0.12) than in EBV-positive tumors (5x10-4, Q1-Q3: 0-0.03), but this difference was not significant (p>0.05, Mann-Whitney U) (Figure 1). In the top 15 recurrently mutated genes (described below), across all 18 samples, 180 SNVs were found in total. The median amount of SNVs found in each sample was 11 (range 6-25). The most frequently mutated gene was CREPPB (12/18, 67%), in which the most prevalent mutation was p.K1497T (8/18, 44%). ARID1A was mutated in 10/18 cases (56%), followed by KMT2D (9/18, 50%). TET2, TP53, MEF2B & TNFAIP3 were all mutated in 7/18 cases (39%). SPEN & IRF2BP2 were mutated in 6/18 cases (33%). PIM1, NOTCH1, ATM, CARD11, DDX3X, IGLL5 were all mutated in 5/18 cases (28%) (Figure 2). Few of the mutations that are called in the top 15 recurrently mutated genes (8/180, 0.04%) lay close to 50% VAF, which might imply that these are SNPs not filtered out by the pipeline. Our SNV findings are comparable with previous literature, in which KMT2D, TET2 & TP53 are also amongst top-mutated genes (Menter et al., 2017), highlighting disruption of epigenetic regulation, NFkB pathway, and DNA damage response as drivers of PTLD. Conclusion: lcWGS in combination deep targeted sequencing allows for detection of CNV an SNV in diagnostic samples of patients with monomorphic PTLD. These data can potentially be used at screening, diagnosis and response monitoring, which is part of the ongoing prospective study NTR 7402. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

ABCL-542 Prevalence and Characterization of EBV-Negative Post-Transplant Lymphoproliferative Disorder

A common and potentially fatal post-solid organ transplant (SOT) complication is post-transplant lymphoproliferative disorder (PTLD). While the association of Epstein-Barr virus (EBV) with the development of PTLD is well recognized, little is known about the etiology of EBV-negative PTLD. To characterize the differences between EBV-positive and EBV-negative PTLD to better understand the pathogenesis of EBV-negative PTLD. This was a retrospective study that reviewed patients who developed PTLD after SOT. Data was collected on PTLD patients (>18 years) from 2011 to 2021. Data included demographics, SOT type, time from SOT to PTLD development, tumor EBV status, PTLD site, histopathological morphology, tumor markers, and overall survival. EBV tumor status was compared using χ2, Fisher's Exact, and Wilcoxon Rank Sum tests. A single, large academic center. A total of 99 patients met the inclusion criteria. Of those, 45.5% were EBV-positive and 54.5% were EBV-negative. Tumor EBV status did not differ by race, but males were more likely to be EBV-negative (63% v. 44.4% of females, p=0.065). EBV tumor status did not differ by SOT type. The interval between SOT and PTLD diagnosis was longer in EBV-negative patients (8.68 years) than EBV-positive patients (4.93 years) (p=0.001). There was a significant difference in EBV-negative PTLD site (p=0.046) and PTLD subtypes (p=0.002). Site of EBV-negative PTLD consisted of gastrointestinal (38.9%), head/neck (11.1%), liver/spleen (11.1%), CNS (3.7%), other lymph (24%), and other non-lymph (11.1%). Monomorphic (65.5%) and T-cell (90%) subtypes were more common in EBV-negative patients, compared to the polymorphic (15.4%) and plasmablastic/plasmacytic (27.3%) subtypes. Expression of CD20 did not differ by tumor EBV status. Those who were CD30-negative were more likely to be EBV-negative (90% v. 25%, p=<0.001). Those who were CD138-negative were more likely to be EBV-negative (75% v. 26.7%, p=0.012). There was no difference in overall survival by tumor EBV status. This single institution retrospective review shows that there are distinct differences between EBV-negative and EBV-positive PTLD. EBV-negative PTLD has a longer interval from SOT to PTLD development, is more likely to occur in the gastrointestinal tract, and is more likely to have a monomorphic morphology.

RAPIDLY PROGRESSIVE POST-TRANSPLANT LYMPHOPROLIFERATIVE DISORDER AFTER LUNG TRANSPLANTATION

TYPE: Case Report TOPIC: Transplantation INTRODUCTION: Posttransplant lymphoproliferative disorder (PTLD) after lung transplantation occurs due to immunosuppressant therapy, which limits antiviral host immunity and permits Epstein-Barr viral (EBV) replication and B cell transformation. CASE PRESENTATION: A 67-year-old female, bilateral lung transplant recipient (07/2019) with a history of idiopathic pulmonary fibrosis developed rapidly progressive PTLD in the early postoperative course. DISCUSSION: At 6-months after transplant, chest radiographs revealed left hilar fullness; subsequent chest CT showed evidence of fluid in the fissure, with no obvious lymphadenopathy. Chest CT performed just two weeks later revealed a left hilar mass measuring 6.1 × 2.7 cm with encasement of the left main pulmonary artery and partial collapse of the left main bronchus. Endobronchial ultrasound-guided biopsies led to the diagnosis of CD20-positive, EBV-positive PTLD. Initial PET scan demonstrated multiple hilar lymph nodes, the largest measuring 2.2 × 2.8 cm. Immunosuppression was reduced, and because the PTLD involved only one area in an EBV-mismatched bilateral lung transplant recipient, she qualified for single-agent rituximab once/week. Unfortunately, repeat imaging after five doses of rituximab demonstrated significant disease progression. She required repeat bronchoscopy with tumor debulking for a necrotic, endobronchial tumor occluding up to one-half of the left lower lobe bronchus. She started R-mini-CHOP, and after three cycles of chemotherapy, complete response was observed on the most recent PET/CT scan. CONCLUSIONS: PTLD may progress quickly, as the initial chest CT scan showing fluid collection advanced rapidly into a hilar mass. Reduction in immunosuppression, anti-CD20 biologic therapy, and chemotherapy for refractory PTLD remains an appropriate treatment paradigm. DISCLOSURE: Nothing to declare. KEYWORD: Post-lung transplant lymphoproliferative disorder

The Diagnostic Dilemma of a Nodule Appearing as a Mass

<h3>Introduction</h3> Posttransplant lymphoproliferative disorder (PTLD) after lung transplantation (LT) occurs due to immunosuppressant therapy that limits antiviral host immunity and permits Epstein-Barr viral (EBV) replication and transformation of B cells. <h3>Case Report</h3> We present the case of a 61-year-old man with history of bone marrow transplant (08/2009) for non-Hodgkin's lymphoma and bilateral LT (04/2021) for idiopathic pulmonary fibrosis - EBV status: D+/R+. The transplant was complicated by prolonged duration of ECMO and a lengthy post-LT hospital stay for renal failure, significant deconditioning, and acute on chronic respiratory failure requiring tracheostomy and fiberoptic bronchoscopic airway stent placement to limit atelectasis. He also required a muscle flap for right bronchial anastomotic dehiscence in this time course. After 4 months, he was finally discharged to home with tracheostomy in place, requiring oxygen supplementation based only on physical demands. On his return to clinic, an inspection bronchoscopy was performed to assess the airway stent, which was found to have purulent secretions concerning for infection. Bronchoalveolar lavage was positive for pan-sensitive <i>Pseudomonas</i> requiring intravenous antibiotic therapy. At the same time, a chest CT revealed multiple pulmonary nodules in the right lung base, the largest measuring 2.8 cm in diameter. A CT-guided biopsy confirmed EBV-positive polymorphic PTLD. Immunostaining yielded positive results for CD20, CD138, CD79a, MUM1, and EBV-encoded RNAs (EBER) (<b>Figure 1</b>). Notably, this patient never had a positive EBV-viral load. Given his undetectable EBV levels, a decision was made to reduce immunosuppression and follow imaging and blood counts as appropriate. <h3>Summary</h3> EBV-positive PTLD with negative EBV viremia is an unusual combination in an EBV-seropositive recipient with donor non-mismatch. The patient's tenuous respiratory status requiring intensive corticotherapy likely contributed significant risk for PTLD in this case.

Post-transplant Lymphoproliferative Disorder Following Cardiac Transplantation.

Post-transplant lymphoproliferative disorder (PTLD) is a spectrum of lymphoid conditions frequently associated with the Epstein Barr Virus (EBV) and the use of potent immunosuppressive drugs after solid organ transplantation. PTLD remains a major cause of long-term morbidity and mortality following heart transplantation (HT). Epstein-Barr virus (EBV) is a key pathogenic driver in many PTLD cases. In the majority of PTLD cases, the proliferating immune cell is the B-cell, and the impaired T-cell immune surveillance against infected B cells in immunosuppressed transplant patients plays a key role in the pathogenesis of EBV-positive PTLD. Preventive screening strategies have been attempted for PTLD including limiting patient exposure to aggressive immunosuppressive regimens by tailoring or minimizing immunosuppression while preserving graft function, anti-viral prophylaxis, routine EBV monitoring, and avoidance of EBV seromismatch. Our group has also demonstrated that conversion from calcineurin inhibitor to the mammalian target of rapamycin (mTOR) inhibitor, sirolimus, as a primary immunosuppression was associated with a decreased risk of PTLD following HT. The main therapeutic measures consist of immunosuppression reduction, treatment with rituximab and use of immunochemotherapy regimens. The purpose of this article is to review the potential mechanisms underlying PTLD pathogenesis, discuss recent advances, and review potential therapeutic targets to decrease the burden of PTLD after HT.

Distinct immunopathological mechanisms of EBV-positive and EBV-negative posttransplant lymphoproliferative disorders

EBV-positive and EBV-negative posttransplant lymphoproliferative disorders (PTLDs) arise in different immunovirological contexts and might have distinct pathophysiologies. To examine this hypothesis, we conducted a multicentric prospective study with 56EBV-positive and 39EBV-negative PTLD patients of the K-VIROGREF cohort, recruited at PTLD diagnosis and before treatment (2013-2019), and compared them to PTLD-free Transplant Controls (TC, n=21). We measured absolute lymphocyte counts (n=108), analyzed NK- and T cell phenotypes (n=49 and 94), and performed EBV-specific functional assays (n=16 and 42) by multiparameter flow cytometry and ELISpot-IFNγ assays (n=50). EBV-negative PTLD patients, NK cells overexpressed Tim-3; the 2-year progression-free survival (PFS) was poorer in patients with a CD4 lymphopenia (CD4+ <300 cells/mm3 , p<.001). EBV-positive PTLD patients presented a profound NK-cell lymphopenia (median=60 cells/mm3 ) and a high proportion of NK cells expressing PD-1 (vs. TC, p=.029) and apoptosis markers (vs. TC, p<.001). EBV-specific T cells of EBV-positive PTLD patients circulated in low proportions, showed immune exhaustion (p=.013 vs. TC) and poorly recognized the N-terminal portion of EBNA-3A viral protein. Altogether, this broad comparison of EBV-positive and EBV-negative PTLDs highlight distinct patterns of immunopathological mechanisms between these two diseases and provide new clues for immunotherapeutic strategies and PTLD prognosis.

Novel Risk Factors for Epstein-Barr Virus (EBV) Reactivation and Post-Transplant Lymphoproliferative Disorder (PTLD) after Allogeneic Hematopoietic Stem Cell Transplant

Background: Epstein-Barr virus (EBV) reactivation and EBV-positive post-transplant lymphoproliferative disorder (EBV+PTLD) are often fatal complications following allogeneic hematopoietic stem cell transplant (HSCT). Risk factors for the development of EBV+PTLD after HSCT include human leukocyte antigen (HLA) mismatches, T cell depletion (TCD) and older age, among others. It has been proposed that specific HLA haplotypes are associated with the development of EBV+PTLD after solid organ transplant (SOT) and that pre-emptive use of Rituximab can reduce the risk of EBV+PTLD. We present a retrospective analysis of patients who underwent HSCT in order to determine the impact of donor source, TCD, age, HLA-A1 haplotypes and use of Rituximab prior to HSCT on the development of EBV reactivation and EBV+PTLD after HSCT.Methods: We retrospectively reviewed patients who had undergone allogeneic HSCT at our Institution between January 2012 and December 2016. We analyzed the cumulative incidence of EBV reactivation and EBV+PTLD and the impact of known and novel risk factors.Results: Of 474 patients, 10% developed EBV reactivation and 6% were diagnosed with EBV+PTLD. The cumulative incidence of EBV reactivation and EBV+PTLD was 15% and 11%, respectively, after haplo-cord HSCT, compared to 3% and 2% after MRD HSCT and 8% and 5% after MUD HSCT (p=0.0004 and p=0.02, respectively). EBV reactivation was associated with receipt of ATG compared to Alemtuzumab (p=0.03) and age older than 60 years (p=0.004). The presence of HLA-A1 haplotype in the recipient did not affect the incidence of EBV reactivation and EBV+PTLD (p=0.2 and p=0.9, respectively). The cumulative incidence of EBV reactivation and EBV+PTLD was lower in patients with Rituximab exposure prior to HSCT, although this did not reach statistical significance (p=0.2 and p=0.15, respectively). In a multivariate analysis, donor source (haplo-cord vs HLA-matched) and age older than 60 were found to be independent risk factors for the development of EBV reactivation (HR 1.9; p=0.038 and HR 2.6; p=0.002, respectively) and EBV+PTLD (HR 1.9; p=0.020 and HR 3.5; p=0.004, respectively). PTLD was fatal in 8 of 27 patients (29%) with PTLD and 17% of all patients with EBV reactivation. One patient died of disseminated EBV disease without PTLD.Conclusions: The cumulative incidence of EBV viremia and PTLD after allogeneic HSCT was 10% and 6%, respectively, and led to death in 2% of the patients. Haplo-cord grafts and age older than 60 years were independent risk factors for EBV reactivation and EBV+PTLD. Rituximab use prior to HSCT may confer protection against EBV reactivation and its use in the conditioning regimen of patients undergoing HSCT, at high risk for EBV reactivation, should be studied prospectively. Further studies are needed to determine the impact of HLA haplotypes on the incidence of EBV reactivation in patients after HSCT. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Treatment of posttransplant lymphoproliferative disorder with poor prognostic features in children and young adults: Short-course EPOCH regimens are safe and effective.

No guidelines exist for which intensive chemotherapy regimen is best in pediatric or young adult patients with high-risk posttransplant lymphoproliferative disorder (PTLD). We retrospectively reviewed patients with PTLD who received interval-compressed short-course etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin (SC-EPOCH) regimens at our institution. Eight patients were included with median age of 12years. All patients achieved a complete response with a manageable toxicity profile. Two patients developed second, clonally unrelated, EBV-positive PTLD and one patient had recurrence at 6months off therapy. No graft rejection occurred during therapy. All eight patients are alive with median follow-up of 29months.

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.

EBV-Positive Primary CNS Lymphomas in Older Patients: Incidence, Characteristics, Tumor Pathology, and Outcomes across a Large Multicenter Cohort

EBV-Positive Primary CNS Lymphomas in Older Patients: Incidence, Characteristics, Tumor Pathology, and Outcomes across a Large Multicenter Cohort

Post‐transplant lymphoproliferative disorders, Epstein‐Barr virus infection, and disease in solid organ transplantation: Guidelines from the American Society of Transplantation Infectious Diseases Community of Practice

These updated guidelines from the American Society of Transplantation Infectious Diseases Community of Practice review the diagnosis, management, and prevention of post-transplant lymphoproliferative disorders (PTLD) and other Epstein-Barr virus (EBV) syndromes after solid organ transplantation. PTLD are a heterogeneous spectrum of predominantly B-cell disorders, often extra-nodal, with complex distinct pathogeneses and variable clinical presentations determined by pathologic subtype. Recent epidemiologic studies report a decrease in early EBV-positive (+) PTLD and an increase in late EBV-negative (-) PTLD. Pre-transplant EBV-seronegativity and primary EBV infection, often from donor-transmitted infection, are an important risk factors for EBV syndromes and early EBV+PTLD. Low-quality evidence supports preemptive prevention strategies for early EBV+PTLD in EBV-seronegative recipients that involve EBV DNA measurement in peripheral blood using assays requiring further result harmonization, combined with interventions to lower viral load. Reduction in immunosuppression (RIS) is the best validated intervention. WHO pathology classification of a tissue biopsy remains the gold standard for PTLD diagnosis; optimal staging procedures are uncertain. Treatment of CD20+ PTLD with the response-dependent sequential use of RIS, rituximab, and cytotoxic chemotherapy is recommended. Evidence gaps requiring future research and alternate treatment strategies including immunotherapy are highlighted.

Isolated gastrointestinal posttransplant lymphoproliferative disorder in a child

We report a 12-year-old male renal allograft recipient, 10 months following the transplant surgery, who presented with diffuse abdominal pain, constipation, vomiting, and weight loss of 5 kg in 2 months. He was on standard maintenance immunosuppression (tacrolimus/mycophenolate mofetil/prednisolone) and had not received induction agents. Upper gastrointestinal (GI) endoscopy showed nodularity in the body of the stomach, and colonoscopy showed deep ulcers in the sigmoid colon and descending colon. Biopsy from the lesions showed dense infiltration of atypical lymphoid cells (B-cell markers, CD20, CD30, and BCL2 being positive), with immunohistochemistry showing strong positivity for Epstein–Barr virus (EBV). EBV DNA polymerase chain reaction in the blood was strongly positive (2200 copies/mL), confirming EBV-positive posttransplant lymphoproliferative disorder (PTLD). Positron emission tomography–computed tomography and bone marrow biopsy confirmed that the disease was confined to the GI tract. It is a rare complication seen after solid organ transplantation occurring usually in the 1st year after transplant. The most important risk factor is a higher degree of immunosuppression; tacrolimus levels remained high in spite of repeated revisions of the drug dose in our case. The patient was treated with injection rituximab (dose: 375 mg/m2), weekly injections of four doses, and his immunosuppressive medications were reduced by 50%. He had a good symptomatic relief. PTLD confined to GI tract is rare; constipation as a presenting manifestation is very rare. A high index of suspicion is essential to make an early diagnosis.

Lymphoma Virome Dynamics Revealed By Cell-Free DNA Sequencing

Lymphoma Virome Dynamics Revealed By Cell-Free DNA Sequencing

The Changing Epidemiology of Posttransplant Lymphoproliferative Disorder in Adult Solid Organ Transplant Recipients Over 30 Years: A Single-center Experience.

Posttransplant lymphoproliferative disorders (PTLD) are a complication of solid organ transplantation (SOT) associated with Epstein-Barr virus (EBV). We analyzed the incidence of and risk factors for PTLD among adult SOT recipients at our center over 30 years (1984-2013). We also compared PTLD incidence before and after a prevention strategy of EBV viral load monitoring in EBV serology mismatched patients was adapted in 2001 (ie, transplant era 1 [1983-2001] vs era 2 [2002-2013]). Among 4171 SOT patients, 109 developed PTLD. Cumulative incidence at 1, 10, and 20 years posttransplant was 0.95, 2.3, and 3.5 per 100 person-years, respectively. Beyond the first year peak of almost exclusively EBV-positive PTLD, a lower incidence of PTLD, predominantly EBV negative, persisted for 20 years. Thoracic transplant (hazard ratio [HR], 2.1; P = 0.007) and negative EBV serology (HR, 7.7; P < 0.001) were independent risk factors for PTLD on multivariate Cox regression analysis. EBV seronegativity significantly increased risk of early (HR, 18.5) and EBV-positive PTLD (HR, 14.2), as well as late (HR, 4.9) and EBV-negative PTLD (HR, 3.6) on univariate analyses. Risk of early PTLD was significantly reduced in the recent transplant era (0.8% era 2 vs 1.9% era 1 at 5 years, P = 0.002); this reduction was seen in recent era EBV seropositive (P = 0.035 at 5 years) but not seronegative recipients (P = 0.90 year 5), suggesting lack of impact of viral load monitoring. Adult SOT recipients face a prolonged risk of late PTLD, whereas risk of early PTLD may have declined in recent years.

Delineation of the Viral and Host Cell Genomic Alterations in EBV-positive PTLD

Post-transplant lymphoproliferative disorder (PTLD) is a potentially fatal complication of organ transplantation characterized by abnormal proliferation of lymphoid cells in the setting of immunosuppression after transplantation. Epstein-Barr virus (EBV) is responsible for abnormal lymphocyte proliferation in 50–80% of PTLDs, particularly in early-onset disease. Latent membrane protein 1 (LMP1) is the chief oncogenic protein of EBV and has been shown to be critical in the transformation of human B cells by the virus. LMP1 may activate several cellular signal transduction pathways including MAPK/ERK, p38/JNK, PI3K/AKT and NF-kB/AP-1. The ability to usurp and dysregulate signaling pathways may explain how LMP1 promotes cell transformation, survival, and proliferation. Our laboratory has shown that LMP1 isolated from EBV-associated B cell lymphoma lines of PTLD patients contains gain-of-function mutations at AA212 (G-S) and AA366 (S-T) that result in sustained ERK signaling, c-Fos activation, and AP-1 activity. In this study, we asked whether these mutations, or other genetic alterations, are present in primary EBV+ PTLD tumors themselves. DNA was isolated from formalin-fixed paraffin-embedded tissue sections of EBV+ PTLD tumors (n=8). Nested PCR was used to amplify LMP1, and the PCR products generated were cloned and sequenced. The presence or absence of gain-of-function mutations at AA212 (G-S) and AA366 (S-T) were assessed. 7 out of the 8 EBV+ PTLD tumors demonstrated both gain-of-function mutations in LMP1. Furthermore, 6 out of the 8 tumors contained an extra repeat of 8 amino acids within the LMP1 signaling tail corresponding to a putative JAK3 binding motif. We have previously identified this repeat in 3 of 6 EBV+ B cell lymphoma lines from PTLD patients. LMP1 was also cloned from the whole blood of a pediatric small bowel PTLD patient. In addition to these 2 mutations being present, the 8 amino acid repeat was also present but in triplicate, suggesting this motif may be crucial to the oncogenic activity of LMP1. In order to assess host cell mutations in EBV+ PTLD tumors, a qBiomarker Mutation PCR array was performed for the PI3K/AKT/mTOR pathway, known to be important in human malignancies. While mutations in PI3K/AKT/mTOR were identified in EBV+ B cell lymphoma lines from PTLD patients, there were significantly more mutations in the primary EBV+ PTLD tumors, with 25 distinct mutations found within the PTEN, PI3K, and STK11 molecules. 3 distinct mutations were identified in more than 1 tumor, and 4 distinct mutations were shared between a cell line and tumor. Our findings clearly demonstrate that key gain-of-functions mutations in LMP1 detected in blood and cell lines are also detected in the primary tumor, suggesting a role in tumorigenesis and great potential as biomarkers of EBV+ PTLD. Moreover, host cell mutations may also contribute to dysregulation of key signal transduction pathways in EBV+ PTLD.

Elucidation of the miRNome in EBV-positive and EBV-negative PTLD

Post-transplant lymphoproliferative disorder (PTLD) is a potentially fatal complication of organ transplantation characterized by abnormal proliferation of lymphoid cells in the setting of immunosuppression after transplantation. Epstein-Barr virus (EBV) is responsible for abnormal lymphocyte proliferation in 50–80% of PTLDs, particularly in early-onset disease. The remaining PTLDs (20–50%) are EBV-negative, with etiology currently unknown. EBV was the first human virus found to encode microRNAs (miRNA). In addition to encoding miRNAs, EBV has also been found to modulate expression and regulation of host-cell-encoded miRNAs. The purpose of this study was to define the miRNome of EBV+ and EBV- PTLD. RNA was isolated from formalin-fixed paraffin-embedded tissue sections of EBV+ and EBV- PTLD tumors (n=5 for each). miRNA-microarray profiling was used to evaluate expression levels of miRNA. One-way ANOVA analysis was used to identify the miRNAs differentially modulated within the groups. 118 miRNAs in total were found to be differentially expressed, in particular 76 up-regulated and 42 down-regulated. In order to validate these initial results, RNA was isolated from additional FFPE tissue sections of EBV+ (n=10) and EBV- (n=5) PTLD tumors, and real-time qPCR was used. Thus far, three miRNAs (hsa-miR-155, hsa-miR-17, and hsa-miR-106a) have been shown to be statistically significantly regulated. Specifically, expression levels of hsa-miR-155 were 2.8-fold greater in EBV+ PTLD tumors (p=0.05). In contrast, levels of hsa-miR-17 and has-miR-106a were 2.2 fold (p=0.04) and 2.1 fold (p=0.05) higher, respectively in EBV- tumors. EBV-encoded miRNAs were also analyzed, and 35 viral miRNAs were detected in EBV+ PTLD. The EBV miRNAs include EBV-miR-BART7-3P, EBV-miR-BART1-5p, EBV-miR-BART14-3p, EBV-miR-BART8-3P, and EBV-miR-BART11-5P. In conclusion, our findings demonstrate that the miRNA profile of EBV+ PTLD tumors is distinguishable from EBV- PTLD tumors and suggest that miRNA may be useful as biomarkers in PTLD.

Posttransplant Lymphoproliferative Disorder Presenting as Cardiogenic Shock With Tamponade

SESSION TITLE: Transplantation SESSION TYPE: Affiliate Case Report Poster PRESENTED ON: Tuesday, October 31, 2017 at 01:30 PM - 02:30 PM INTRODUCTION: Post-transplant lymphoproliferative disorders (PTLD) are malignancies associated with solid organ transplantation or allogeneic hematopoietic cell transplantation requiring immunosuppression. Most cases are associated with EBV positivity, but less commonly EBV negative PTLD occurs. CASE PRESENTATION: A 55-year-old Caucasian male presented with cardiogenic shock due to pericardial effusion with tamponade. His history was significant for type I diabetes mellitus leading to ESRD, two failed renal transplants on immunosuppression, and current peritoneal dialysis dependence. Echocardiogram upon admission demonstrated pericardial effusion with right ventricular collapse. Pericardiocentesis and pericardial drain placement were performed. Approximately 2.4L of fluid was drained. Cytology of the cell block was negative for malignancy. He improved after drainage and was discharged on day 16. Two days after discharge he returned complaining of weakness. A left sided pleural effusion was detected and drained via catheter. Cell block cytology was remarkable for monomorphic PTLD, plasma cell neoplasm. Five days after pleural catheter placement he developed multifactorial shock that led to a whole-body CT with contrast. This revealed dilation of the jejunum measuring 5.1cm, as well as a mass that measured 8.4 x 5.6. x 5.4 cm. The mass demonstrated atypical plasma cells consistent with the cell block. Discontinuing immunosuppression was the initial step in treatment in this case. Further work up for plasmacytoma or multiple myeloma demonstrated negative SPEP and negative lambda ratio confirming a solitary plasmacytoma. A skeletal survey was completed with no signs of lytic lesions or evidence of multiple myeloma. A bone marrow biopsy was unable to be completed as the patient expired. DISCUSSION: The lesion consisted of atypical plasma cells similar to cases of EBV-negative PTLD previously mentioned in the literature. This is currently a designation not clearly defined by the World Health Organization. These findings call for further investigation to discover the mechanism of disease and promote effective therapy. Therapy in EBV-negative PTLD does not seem to respond to the established algorithm for EBV-positive PTLD. Reducing immunosuppression often makes no difference. Some cases require high dose immunosupression to achieve therapeutic success, which is the opposite of the standard treatment for PTLD. Defining the mechanism behind these tumors is imperative for better classification as well as directed therapy. CONCLUSIONS: This case is remarkable for PTLD with CD20 negativity; additionally, PCR for EBV was negative. EBV is typically the primary driver of PTLD. The mechanism of EBV negative PTLD remains unknown. Reference #1: Swerdlow SH, Harris NL, Pileri S, Stein H, Jaffe ES. The 2008 WHO classification of lymphoid neoplasms and beyond: evolving concepts and practical applications. Blood. 2011;117(19):5019-5032. doi:10.1182/blood-2011-01-293050. DISCLOSURE: The following authors have nothing to disclose: Matthew Boulos, Anil Singh, David Aljadir, Tina Dudney No Product/Research Disclosure Information