ALBERT

Description: Background: Despite the increase in patients' survival over the last years, multiple myeloma (MM) remains incurable, being persistence of cancer stem cells (CSCs) a probable cause of drug resistance and disease relapse. It is possible to isolate these cells using surface antigen expression pattern (CD19+/CD34+/CD138-) and the activity of an enzyme from aldehyde dehydrogenase (ALDH) family (Boucher et al., 2012). We believe that using CD19 as potential marker of MM-CSCs makes CAR-T cell therapy against CD19 an option to eradicate residual MM disease. Aims: To isolate and characterize immunophenotypically, functionally and by gene expression the MM-CSCs derived from bone marrow (BM) samples of newly-diagnosed MM patients, focusing on identification of possible therapeutic targets. Methods: BM aspirates were collected and CD138+ cells were separated by magnetic sorting. The remaining cells were submitted to sorting by flow cytometry on FACSAria II (Becton, Dickinson and Company, Franklin Lakes, NJ, USA), labeled with anti-CD19 Pacific Blue (Invitrogen, Carlsbad, CA, USA), anti-CD34 PE Cy7 and anti-CD138 APC (both from Becton, Dickinson and Company, Franklin Lakes, NJ, USA) antibodies, in addition to Aldefluor™ reagent (StemCell Technology, Vancouver, British Columbia, Canada). RNA was extracted and pre-amplified for PCR array analysis using the RT² Profiler™ PCR Array Human Cancer Stem Cells(Qiagen, Hilden, Germany) to assess the expression profile of 84 genes related to cancer stem cells, and the results were evaluated with the online software provided by the platform manufacturer. Results: MM-CSCs (CD34+/CD19+/CD138-/ALDH1+) were isolated by flow cytometry from MM samples and presented median of 1,748.5 events (ranging from 56 to 16,633, n = 16). For comparison purposes, CD138+ MM tumor cells were isolated and used as "control group" (median of events 72,904, ranging from 1,536 to 312,504, n = 15). RNA from 16 MM-CSC samples and 6 controls were analyzed by qPCR. Considering 2-ΔCt calculation (GAPDH as normalizer) and fold change of 2, 11 genes were considered differentially expressed in MM-CSCs when compared to tumor plasma cells (p

Description: Introduction Little is known about the incidence and clinical features of Multiple Myeloma (MM) in Latin America. A clinical registry of Latin American (LA) patients with MM represents an opportunity to gain insight into the prevalence of the disease in this region, the patterns of care and the current treatment status in different LA countries. Objective To characterize the demographic and clinical features of patients with multiple myeloma from five LA countries (Brazil, Argentina, Chile, Mexico and Peru) and to create a LA database on MM; in addition to investigating the patterns of care for MM patients in Latin America. Patients and Methods This is an observational, non-intervention study, with a prospective evaluation of data. Eligible patients were diagnosed with multiple myeloma, between January 1, 2005, and December 31, 2007, at any one of the participating centers, regardless of disease stage or treatment modality. The follow-up period extended to at least 5 years for each patient (December 31, 2012). Results Eight hundred and seventy six patients were included. The median age was 60 years old (25-97), 53.4% male and 46.6% female. The median follow-up was 31.4 months, and the median overall survival was 57 months. The median overall survival to patients who received high-dose chemotherapy was 77 months and for patients who received conventional chemotherapy was 48 months (p

Description: Introduction: Follicular lymphoma (FL) is the second most prevalent non-Hodgkin lymphoma worldwide, and is characterized by an indolent course and frequent relapses. Better understanding of FL has shown that angiogenesis (AG) has an important role in its progression to a more aggressive form. The most important mediator of AG is the vascular endothelial growth factor (VEGF), which is encoded by a polymorphic gene. It is already known that allele C of the VEGF 2578C/A polymorphism (rs699947) is related to higher serum concentration of VEGF compared to the A allele. The roles of this genetic polymorphism in clinical manifestations and outcome of FL are still unknown, and therefore these were the aims of the present study. Methods: Our analysis included 86 consecutive FL patients seen at diagnosis at the university hospital. The patients were treated with 6 to 8 cicles of R-CHOP. The clinical data of these patients were obtained from medical records. Genomic DNA was extracted from peripheral blood samples, and genotyping of the VEGF -2578C/A (rs699947) polymorphism was performed with real-time qPCR. Overall-survival (OS) was defined as time from diagnosis until death from any cause or last follow-up. The differences between groups were analyzed by the logistic regression model. Kaplan-Meier and log-rank analyses were used to assess survival information from the patients’ data. Furthermore, we examined survival data using univariate Cox proportional hazards regression, with the respective hazard ratios (HR) and 95% Confidence Intervals (CIs). Adjustement of Cox regression for clinical features was also performed. A p-value smaller than 0.05 was used to denote statistical significance. Results: The FL patients had a mean age of 56.2 years at diagnosis. The majority of the patients were caucasians (87.6%), and there was an equivalent distribution between genders: 43 male and 43 female patients (50% each). Thirty-nine (45.3%) patients had B-symptoms and 48 (54.7%) had no B-symptoms at diagnosis. Sixty-three (73.2%) patients presented at diagnosis with tumors of III or IV Ann Arbor stage, and 30 (34.8%) had bone marrow infiltration. Follicular Lymphoma International Prognostic Index (FLIPI) was assessed and patients were classified as follows: low-risk FLIPI (≤ 1 point) was seen in 34 cases (39.5%), medium-risk FLIPI (2 points) in 28 cases (32.5%), and high-risk FLIPI ( ≥ 3 points) was seen in 24 patients (27.9%). The VEGF 2578CC genotype was more common in patients with B-symptoms at diagnosis than in those without B-symptoms (51.2% versus 29.7%, p=0.04). The frequency of VEGF 2578CC genotype was also higher in patients with high-risk FLIPI than in those with medium and low-risk FLIPI (58.3% versus 32.2%, p=0.03). We found no association between genotypes and bone marrow infiltration at diagnosis. Considering clinical course of the patients, Kaplan Meier curves of OS showed that at 60 months of follow up, B-symptoms at diagnosis (65.2% of OS versus 87.1%, p=0.02), high-risk FLIPI (52.5% of OS versus 90%, p=0.002), and 2578CC genotype (62.1% of OS in 2578CC patients versus 89.4% in 2578CA plus 2578AA patients, p=0.01) had negative impacts on outcome. In univariate Cox analysis, presence of B-symptoms (p=0.03, HR=3.2, 95% CI: 1.09-9.38), high-risk FLIPI (p=0.01, HR= 3.91, 95% CI: 1.38-11.03) and 2578CC genotype (p=0.02, HR=3.70, 95% CI: 1.16-11.82) were also associated with a worse outcome. The prognostic role of the abovementioned polymorphism was still present after adjustment for age (p=0.04, HR=3.23, 95% CI:1.01-10.36) and Ann Arbor stage (p=0.02, HR=3.58, 95% CI:1.1-11.6) in univariate regression. After adjustment for FLIPI status as a whole, only a trend of association of 2578CC genotype with a worse outcome was found in the study (p=0.05, HR=3.24, 95% CI: 0.95-13.05). Conclusion: Our results present, for the first time, preliminary evidence that FL patients with the VEGF 2578CC genotype, related to higher production of VEGF, are more likely to develop aggressive form of the disease at diagnosis, and to succumb earlier to death. We recognize, however, that our study is based on a small sample size and that more numerous cohorts should be analyzed in order to assess additional associations. Also, further studies, such as correlations between relevant AG genes and tumor vascularization, should be done in order to clarify this issue in the context of FL. Disclosures No relevant conflicts of interest to declare.

Description: Introduction: Post-transplant lymphoproliferative disorders are a rare heterogenous group of diseases occurring in the setting of post-transplant immunossupression (IS). Clinically, extranodal involvement is common, and it occurs in the central nervous system (CNS) in approximately 7-15% of cases. Most data on PTLD-CNS are based on case series/reports and due to paucity of data no treatment algorithms have been established. In our series, we retrospectively analyzed 23 consecutively cases of PTLD-CNS in Hospital do Rim, Sao Paulo, Brazil. Methods: We retrospectively reviewed all cases of PTLD-CNS diagnosed between 2001 and 2014 at Hospital do Rim (HRim), a school hospital from the Federal University of São Paulo, Brazil. HRim is considered the leading renal transplant hospital in the world for the past 15 years. For this study, only PTLD patients with tumors whose histology could be confirmed by hemopathologist review, EBV-association established and whose clinical, epidemiological and laboratorial parameters could be retrieved were included in this study. Response was defined as complete (CR) or less than CR (partial response or refractory disease). Event was defined as treatment related mortality, progression (defined as time for initiation of second-line therapy) or relapse. Results: From 2001 to 2014, from a total of 98 PTLD patients, 23 patients (23%) were diagnosed with PTLD-CNS. Median age at time of diagnosis was 36, with a male:female ratio of 0.9:1. Fifteen (65%) patients received anti-thymocyte globulin (ATG) at the time of transplant and 17 (74%) had at least one episode of acute rejection. The most common immunossupressive regimen (IR) consisted of cyclosporine or tacrolimus associated with prednisone and azathioprine (15, 65%). Median time from transplant to PTLD diagnosis was 31 months (ranging from 8.4 to 153). EBV was positive in tumor lymphocytes by in situ hybridization in 21 cases (91%). Monomorphic cases were diagnosed in 21 (91%) patients. All patients had their IR reduced, usually with suspension of azathioprine and calcineurin inhibitors and change from prednisone to dexamethasone, 19 (82%) patients underwent WBRt (from 25 to 40 Gy) together with intra-thecal (IT) chemotherapy with methotrexate (MTX) 12mg and dexamethasone 2mg. Only 1 patient received high dose MTX and died due to treatment-related toxicities 1 month after diagnosis and 2 patients died before starting WBRt due to disease progression and poor performance status (PS). For those patients who received WBRt together with IT chemotherapy, fourteen patients (74%) had CR, 2 patients (10%) had refractory disease and 3 patients (16%) had relapsed disease within 2 years. Overall Survival (OS) for the group treated with WBRt and IT chemotherapy was 62% in 5 years (CI95% 69-82%). In our series, induction therapy with ATG and acute rejection were associated with increased risk of PTLD-CNS. Age and PS at diagnosis were the only 2 factors predictive of survival. No serious cognitive impairment was identified among the survivors. Conclusions: The current study demonstrated that PTLD-CNS is a serious late EBV-induced B-cell lymphoma, mostly monomorphic with an incidence of 23%, higher than previously described in the post renal transplant setting. Treatment with immunossupression reduction, intrathecal chemotherapy and whole-brain radiotherapy showed a high CR rate with favorable survival in many cases. Disclosures No relevant conflicts of interest to declare.

Description: Background: Sickle cell anemia (SCA) is a complex disease, associated with hemolysis, vaso-occlusion, vascular inflammation and endothelial activation. Significant morbidity and premature mortality are hallmarks of the disease and elevation of tricuspide regurgitant velocity (TRV), determined by doppler-echocardiography, has been associated with higher risk. The identification of biomarkers associated with severity in SCA is desirable. Circulating serum microRNAs (miRNA) are molecular targets studied in different diseases as diagnostic or prognostic markers, however there are few studies in SCA. Purpose: to identify specific signatures of miRNAs in plasma samples of SCA patients according to severity indexes. Methods: Screening of the miRNAs expression was performed in 8 patients. These patients were classified by TRV measure (referred as TRV Score): 4 samples with TVR ≥ 2.5 m/s and 4 with TRV 〈 2.5 m/s. After extraction of total RNA, the samples were analyzed by real-time PCR using Megaplex RT Human Pool A and Megaplex RTHuman Pool Blife (Thermofisher) comprising 667 distinct miRNAs. Expression Suite Software (Thermofisher) and SPSS were used for analysis. Seventeen miRNAs were differentially expressed between the two groups (p 〈 0.05) and miR16 was considered as the endogenous candidate. Five differentially expressed miRNAs (miR15b, miR502, miR510, miR544, miR629) were selected for validation in 56 patient samples using TRV Score. Another two severity scores were also used: (a) Organ Injury Score (SLO) - based on the presence or absence of 5 lesions: stroke, TRV ≥ 2.5 m/s, leg ulcers, osteonecrosis, and microalbuminuria (adapted from Afenyi-Annan et al. Transfusion 2008; 48:197) and (b) NIH Bayesian score - available online (http://bios.ugr.es/dss-calculator/). The ROC curve was used to analyze the data of relative expression (2-ΔCT) of each miRNA. Results: Two out of five miRNA, miR510 and miR629, were significantly decreased in more severe patients. The miR510 expression allowed the discrimination of the patients according to TRV Score at the cutoff 0.000331043, sensitivity 71.4%, specificity 73.3% and AUC of 0.825 (95% CI: 0.689-0.962, p = 0.001). The same miRNA was also a good discriminant in the SLO at a cutoff of 0.000331043, sensitivity 77.8%, specificity 72.2% and area under the curve (AUC) of 0.769 (95% CI: 0.666-0.931), p = 0.008. The miR629 was related to severity according to the Bayesian Score at the cutoff of 0.0009854449, sensitivity 66.7%, specificity 75% and AUC of 0.729 (95% CI: 0.552-0.907, p = 0.027). The other miRNAs, miR15b, miR502 and miR544, showed no significant results. Discussion and Conclusions: This is the first study which looks into plasma miRNA as a biomarker of SCA severity. The miR510 regulate Peroxirredoxin-1 (PRDX1), a protein involved in the stress-oxidation. Increased oxidative stress presented in SCA might imply that miR510 has a role in this mechanism. The miR629 appears to be involved in the AKT1 signaling pathway related to Endothelin-1. As it is known, endothelin-1 is increased in SCA and is important in pulmonary hypertension. The miR510 and miR629 appear to be hypoexpressed in patients with more severe SCA, probably with greater importance in the regulation of clinical manifestations associated with vascular disease, although more studies seem to be necessary. Disclosures No relevant conflicts of interest to declare.

Description: Cancer-testis (CT) antigens are expressed in a variety of malignant tumors but in normal adult tissues solely in testicular germ cells. Based on this tumor-associated expression pattern, CT antigens are considered valuable targets for immunotherapy. While CT antigens have been studied in many solid tumors, much less is known about their presence in neoplasms of the hematopoetic system such as Non Hodgkin’s Lymphoma (NHL). Aims: To analyze the expression of 10 CT antigens in NHL by immunohistochemistry (IHC) using tissue microarray (TMA) technology, and to correlate its expression with histologic subtypes. Patients and Methods: Formalin-fixed paraffin-embedded tissues of 116 NHL cases dating from June 2003 to June 2007 were retrieved from the archives of the Department of Pathology, Federal University of São Paulo, Brazil. Two TMA blocks were generated comprising three cores/NHL case, as well as positive (testis) and negative (reactive lymph nodes) control tissue cores. The following monoclonal antibodies (mAbs) (to the following antigens) were used for IHC analysis: MA454 (MAGE-A1), M3H67 (MAGE-A3), 57B (MAGE-A4), CT7-33 (CT7/MAGE-C1), CT10#5 (CT10/MAGE-C2), E978 (NY-ESO-1), 219-510-23 (LAGE-1 and NY-ESO-1), #26 (GAGE) and SSX-2#7 (SSX-2). Two observers scored all slides independently and cases with loss of all three punches and/or missing tumor tissue were excluded from analysis. Results: The number of cases which could not be evaluated due to loss of tissue core and/or missing tumor differed, ranging between 8 for mAb #26 to 12 for mAbs MA454, CT10#5 and 219-510-23. 7/116 NHL cases were missing in all slides. 14/109 (12.8%) NHL cases expressed at least one CT antigen: diffuse large B-cell lymphoma (DLBCL) 9/58 (15.5%); anaplastic large cell lymphoma (ALCL) 1/3; follicular lymphoma (FL) 1/10 (10%); lymphoplasmacytic lymphoma (LPL) 1/3; peripheral T-cell lymphoma (PTCL) 1/9 and histiocytic lymphoma (HL) 1/1. No CT antigen expression was present in lymphoblastic lymphoma (5), mantle cell lymphoma (2), small lymphocytic lymphoma (6), marginal zone lymphoma (4), MALT lymphoma (5) and mycosis fungoides (3). The most frequently expressed CT antigens were: GAGE 6/108 (5.5%), NY-ESO-1 5/105 (4.7%), CT7 5/106 (4.7%), MAGE-A1 4/104 (3.8%) and MAGE-A3 4/107 (3.7%). According to cell origin, positivity of CT antigens was more prevalent in B-cell NHL (11/91; 12.1%) than in T-cell NHL (1/15; 6.7%). NY-ESO1 5/91(5.5%) was the most frequently expressed CT in B-cell NHL, followed by GAGE 4/91 (4.4%), MAGE-A1 3/91 (3.3%), MAGE-A3 3/91 (3.3%), CT7 3/91 (3.3%) and LAGE 3/91 (3.3%). The only positive T-cell lymphoma case expressed MAGE-A1 and GAGE. Conclusions: Expression of CT antigens in NHL is limited. In the present TMA-based IHC analysis, only 12% of NHLs express at least one CT antigen, with DLBCL showing the highest incidence and NY-ESO-1 being the most prevalent antigen. In spite of the low incidence, presence of CT antigens could offer the opportunity of vaccine-based immunotherapy in selected cases of NHL.

Description: Introduction: Cancer testis (CT) antigens have become the most extensively studied antigen group in the field of tumor immunology. CT45 antigen expression was described in colon adenocarcinomas, germ cell tumors, Hodgkin’s lymphomas and, more recently, in multiple myeloma (MM). Aims: This study aims to analyze the expression of CT45 in normal tissues and in plasma cell disorders and to identify possible associations with clinical data and prognosis in MM. Patients and Methods: The expression of CT45 was studied in twenty normal tissues (testis, placenta, skeletal muscle, bladder, lung, spleen, heart, brain, fetal brain, thymus, uterus, stomach, mammary gland, pancreas, prostate, small intestine, kidney, adrenal gland, spinal cord, colon and one pool of ten normal bone marrow samples) and in bone marrow aspirates from three monoclonal gammopathies of undetermined significance (MGUS), five solitary plasmacytomas, 61 newly diagnosed MM patients and MM cell line U266 by RT-PCR. Results: CT45 was positive in three out of 20 (15%) normal tissues tested: lung, brain (both fetal and adult) and spinal cord. Among monoclonal gammopathies, CT45 was positive in two out of five (40%) solitary plasmacytomas’ bone marrow aspirates, 10 out of 61 (16%) MM bone marrow aspirates and in the U266 MM cell line. Six out of 10 (60%) CT45 positive MM cases were classified as International Staging System (ISS) 3 (p = 0.009). Six CT45-positive cases were classified as plasmacytic (PC) and four as polymorphic (PM). Median OS of the MM group was 21 months. Nine patients were submitted to autologous stem cell transplantation. All of the transplanted cases were CT45-negative. Univariate analysis showed that Durie-Salmon Staging System (Durie-Salmon IIIA: N = 35, median OS = 40 months; Durie-Salmon IIIB: N = 19, median OS = 12 months; log-rank p= 0.0139), b2microglobulin (b2microglobulin £ 5.5 mg/L: N = 27, median OS = 40 months; b2microglobulin 〉 5.5 mg/L: N = 24, median OS = 12 months, log-rank p= 0.0520, Breslow p = 0.0352, Tarone-Ware p = 0.0399), plasma cell morphology (PC: N = 38, median OS = not reached; PM: N = 11, median OS = 12 months; PB: N = 5, median OS = 1 month; log-rank p= 0.0037), transplantation proceedings (transplanted patients: N = 9, median OS = not reached; non-transplanted patients: N = 47, median OS = 14 months; p = 0.0064) and CT45 expression (CT45 expression negative: N = 46, median OS = 25 months; CT45 expression positive: N = 10, median OS = 3 months, log-rank p = 0.038 for all patients and CT45 expression negative: N = 37, median OS = 19 months; CT45 expression positive: N = 10, median OS = 3 months, p = 0.0245, only non-transplanted patients) had impact on OS. Cox Regression Model showed that only plasma cell morphology (p = 0.029, RR 5.288, CI 1.77704–15.7988), transplant proceedings (p = 0.0742, RR 0.1582, CI 0.0209–1.1976) and CT45 expression (p = 0.0016, RR 7.0403, CI2.0978–23.6278) were independent prognostic factors in MM patients survival. CT45-positive cases were associated with poor outcome and presented 7 times more chance of worse evolution then the negative ones. Conclusions: CT45 was expressed in only 16% of MM patients. However, we demonstrated for the first time that positive expression of CT45 was associated with high ISS scores and poor outcome in MM

Description: Serial analysis of gene expression (SAGE) allows a comprehensive profiling of gene expression within a given tissue and also an assessment of transcript abundance. Objectives: We generated SAGE libraries from normal and neoplastic plasma cells to identify genes differentially expressed in multiple myeloma (MM). Material and Methods: Normal plasma cells were obtained from palatine tonsils and MM SAGE library was generated from bone marrow plasma cells of MM patients. Results: We obtained 29,918 SAGE tags from normal and 10,340 tags from tumor libraries. Computer-generated genomic analysis identified 46 upregulated genes in the MM library. Ten upregulated genes were selected for further investigation. Differential expression was validated by quantitative real-time PCR in purified plasma cells of 31 patients and three controls. P53CSV, DDX5, MAPKAPK2, RANBP2 were found to be upregulated in at least 50% of the MM cases tested. All of them were also found upregulated in MM when compared to normal plasma cells in a meta-analysis using ONCOMINE microarray database. Antibodies specific to DDX5, RANBP2 and MAPKAPK2 were used in a TMA containing 57 MM cases and confirmed the expression of these proteins in 74, 96, and 21% of the MM samples, respectively. Conclusions: Analysis of differential expression using SAGE could identify new genes important for myeloma tumorigenesis (P53CSV, DDX5, MAPKPK2 and RANBP2) and that could potentially be useful as therapeutic targets.

Description: Introduction: HSP70 has an integrative role in protein degradation due to the interaction with many pathways, such as ubiquitin proteasome (UPS), unfolded protein response (UPR) and autophagy. In multiple myeloma (MM) HSP70 is overexpressed and helps to prevent proteotoxic stress and cell death caused by overload of unfolded/misfolded proteins produced by tumor cells. Aims: To explore the role of HSP70 inhibition, isolated or in association with proteasome inhibitor, as therapeutic strategy for MM through in vitro and in vivo analyses. Methods: RPMI8226-LUC-PURO and U266-LUC-PURO bioluminescent cell lines were treated with HSP70 inhibitor (VER155008- 50 μM or 80μM) and proteasome inhibitor (bortezomib 100nM) for evaluation of apoptosis induction by flow cytometry using annexin V and propidium iodide. NOD.Cg-rkdcscid Il2rgtm1Wjl/SzJ immunodeficient mice were used for plasmacytoma xenograft model and treated with intravenous VER155008 (40mg/kg) and bortezomib (1mg/kg), immediately after transplant of RPMI8226-LUC-PURO and U266-LUC-PURO bioluminescent cell lines (N=3 for each group, including controls, bortezomib, VER155008, and combination of bortezomib and VER155008). Bioluminescence was measured in IVIS Kinetic (Capiler Life Science) once a day for seven days. Results: Bortezomib used as single treatment was able to induce apoptosis in RPMI8226-LUC-PURO cell line: the best result for in vitro studies RPMI8226-LUC-PURO was 65% of late apoptosis after treatment with bortezomib. On the other hand, U266-LUC-PURO cell line presented higher percentage of apoptosis when treated with bortezomib and VER155008 combination: U266-LUC-PURO cell line presented more than 60% of late apoptosis after VER155008 (80μM) combined with bortezomib, showing that inhibition of HSP70 could overcome U266-LUC-PURO resistance to bortezomib alone. Mice treated with VER155008, alone or in combination with bortezomib, showed complete inhibition of tumor growth (absence of bioluminescence) for both cell lines when compared with control group after one week of treatment (p

Description: Introduction: Post-Transplant Lymphoproliferative Disease (PTLD) is a rarewell-recognized group of lymphoid and or plasmacytic proliferations that occur following both solid organ (SOT) and allogenic hematopoietic stem cell transplantation (HSCT) as a result of immunossupression. A continuum of disease has been described, including early lesions, polymorphic PTLD and monomorphic PTLD. Epstein-bar virus (EBV) is considered the most important causative factor, with EBV positivity observed within up 90% of tumor lymphocytes. Optimal risk stratifications specific to kidney transplantation are still lacking. Patient and Methods: In our series, we retrospectively analyzed 98 consecutively cases of PTLD diagnosed between 2001 and 2014 at Hospital do Rim (HRim), a school hospital from the Federal University of São Paulo, São Paulo, Brazil. HRim is considered the leading renal transplant hospital in the world for the past 15 years. For this study, only PTLD patients with tumors whose histology could be confirmed by hemopathologist review, EBV-association established and whose clinical, epidemiological and laboratorial parameters could be retrieved were included in this study. Response was defined as complete (CR) or less than CR (partial response or refractory disease). Event was defined as treatment related mortality, progression (defined as time for initiation of second-line therapy) or relapse. Patients with conflicted data or loss of follow up were excluded. Results: A total of 98 patients were diagnosed with PTLD from 7665 renal transplants performed in this period, with a incidence of 1,3%. Two patients were excluded from the analysis due to conflicting clinical data. Median age at diagnosis was 41.4 years (range from 4-74), with a 0,6:1 Female:Male ratio. Median time of transplant to PTLD diagnosis was 56 months (range 1-967). Thirty-six patients (37.5%) received anti-thymocyte globulin (ATG), and the most common immunossupressive regimen (IR) consisted of cyclosporine or tacrolimus associated with prednisone and azathioprine (66, 69%). Monomorphic PTLD was observed in 75 (78%) patients, and two patients presented with HL. EBV positivity was seen in 67 patients (70.5%), being more frequent within PTLD cases diagnosed during the first 3 years of transplantation (90%). All patients had their IR reduced after PTLD diagnosis. The incidence of loss of engraftment following IR reduction was 19% (20 patients). Overall responses were: CR in 67% (n=64) and Partial response/Refractory disease in 33% (n=32); 30 patients died due to treatment-related toxicity and/or disease progression. Overall Survival (OS) for the entire group was 63% in 5 years (CI95% 73-83%), with a progression free survival (PFS) of 58% (CI95% 91-97%). EBV positivity was seen in 67 patients (70.5%). Median number of extra nodal sites involved was 1,2. The median overall survival time was 97 months. Patient survival following diagnosis was 77% at 1 year, 63% at 5 years and 38% at 10 years. The most common extra-nodal sites involved were: Gastro-intestinal Tract (43, 45%) followed by Central Nervous System (23, 24%). Extra nodal involvement was correlated with poorer outcome (p 0.014) as was the loss of engraftment (p