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Background Hemostatic resuscitation strategy using blood components with a balanced ratio is adopted in the civilian trauma setting. However, there is usually limited availability of blood components in the austere setting. Warm fresh whole blood (WFWB) has been used for trauma patients with life-threatening hemorrhage necessitating massive transfusions in the Okinawa Islands, Japan. The purpose of this study was to evaluate the safety and feasibility of WFWB use in the austere civilian trauma setting. Methods We conducted a retrospective cohort study between January 1999 and June 2019, including trauma patients who received WFWB within 24 hours of admission. Immediately after WFWB was collected from blood donors, the sample was typed and screened for transmissible infectious diseases. Approximately half of the study population received irradiated WFWB to prevent graft versus host disease. We evaluated the incidence of transfusion-associated adverse events. Transfusion requirements and patient outcomes were compared between early and late WFWB use. Results A total of 28 patients from three civilian institutions were eligible. Of those, 93% sustained blunt trauma. The median Injury Severity Score was 37 (interquartile range, 32-49). All patients required operative hemostatic intervention, and half of the patients required both operative and endovascular hemostatic interventions. Patients received a median of 1,800 mL WFWB transfusions from seven volunteer blood donors. None of our subjects developed hemolytic reactions, transmissible infectious diseases, or graft versus host disease. Early WFWB use (within 4 hours of admission) was associated with a significant reduction in platelet transfusion requirement compared with the late WFWB group in univariate analysis (16 units vs. 47 units, p = 0.002). Conclusion Warm fresh whole blood use is safe and feasible in an austere civilian trauma setting. Prospective studies with larger cohorts are necessary to determine whether early WFWB use will affect patient outcomes, transfusion requirement, and treatment cost. Level of evidence Therapeutic, Level IV.

To investigate the use of high-dose therapy and autologous stem cell transplantation (ASCT) for relapsed or high-risk diffuse large B-cell lymphoma (DLBCL) between 1990 and 2007, we conducted a nationwide survey using the registry database of the Japan Society for Hematopoietic Cell Transplantation. Of the 1222 patients in the database, 576 (47%) received ASCT in first complete remission (CR1), 140 (12%) in first partial remission, 281 (23%) in sensitive relapse, 150 (12%) in resistant or sensitivity-unknown relapse, and 75 (6%) in primary refractory status. With a median follow-up of 22 months, the 2-year overall survival (OS) and progression-free survival rates were 71% and 68%, respectively. The cumulative incidences of 2-year non-relapse mortality and relapse/progression were 6% and 26%, respectively. Relapse/progression after ASCT in the rituximab era (2002-2007) was significantly lower than that in the pre-rituximab era (1990-2001; P < 0.001). Older age, male gender, poor performance status at ASCT, non-CR1 at ASCT, ASCT performed in 1990-2001, and LEED or MCEC regimen were adverse predictors of OS. Because ASCT for newly diagnosed high-risk DLBCL has not been performed recently, a registry database study to assess the impact of ASCT for relapsed or refractory DLBCL in the rituximab era is warranted.

Aggressive adult T‐cell leukemia/lymphoma (ATL) has an extremely poor prognosis and is hyperendemic in Okinawa, Japan. This study evaluated two prognostic indices (PIs) for aggressive ATL, the ATL‐PI and Japan Clinical Oncology Group (JCOG)‐PI, in a cohort from Okinawa. The PIs were originally developed using two different Japanese cohorts that included few patients from Okinawa. The endpoint was overall survival (OS). Multivariable Cox regression analyses in the cohort of 433 patients revealed that all seven factors for calculating each PI were statistically significant prognostic predictors. Three‐year OS rates for ATL‐PI were 35.9% (low‐risk, n = 66), 10.4% (intermediate‐risk, n = 256), and 1.6% (high‐risk, n = 111), and those for JCOG‐PI were 22.4% (moderate‐risk, n = 176) and 5.3% (high‐risk, n = 257). The JCOG‐PI moderate‐risk group included both the ATL‐PI low‐ and intermediate‐risk groups. ATL‐PI more clearly identified the low‐risk patient subgroup than JCOG‐PI. To evaluate the external validity of the two PIs, we also assessed prognostic discriminability among 159 patients who loosely met the eligibility criteria of a previous clinical trial. Three‐year OS rates for ATL‐PI were 34.5% (low‐risk, n = 42), 9.2% (intermediate‐risk, n = 109), and 12.5% (high‐risk, n = 8). Those for JCOG‐PI were 22.4% (moderate‐risk, n = 95) and 7.6% (high‐risk, n = 64). The low‐risk ATL‐PI group had a better prognosis than the JCOG‐PI moderate‐risk group, suggesting that ATL‐PI would be more useful than JCOG‐PI for establishing and examining novel treatment strategies for ATL patients with a better prognosis. In addition, strongyloidiasis, previously suggested to be associated with ATL‐related deaths in Okinawa, was not a prognostic factor in this study., 論文

To investigate the use of high-dose therapy and autologous stem cell transplantation (ASCT) for relapsed or high-risk diffuse large B-cell lymphoma (DLBCL) between 1990 and 2007, we conducted a nationwide survey using the registry database of the Japan Society for Hematopoietic Cell Transplantation. Of the 1222 patients in the database, 576 (47%) received ASCT in first complete remission (CR1), 140 (12%) in first partial remission, 281 (23%) in sensitive relapse, 150 (12%) in resistant or sensitivity-unknown relapse, and 75 (6%) in primary refractory status. With a median follow-up of 22 months, the 2-year overall survival (OS) and progression-free survival rates were 71% and 68%, respectively. The cumulative incidences of 2-year non-relapse mortality and relapse/progression were 6% and 26%, respectively. Relapse/progression after ASCT in the rituximab era (2002-2007) was significantly lower than that in the pre-rituximab era (1990-2001; P 0.001). Older age, male gender, poor performance status at ASCT, non-CR1 at ASCT, ASCT performed in 1990-2001, and LEED or MCEC regimen were adverse predictors of OS. Because ASCT for newly diagnosed high-risk DLBCL has not been performed recently, a registry database study to assess the impact of ASCT for relapsed or refractory DLBCL in the rituximab era is warranted.

To investigate the use of high-dose therapy and autologous stem cell transplantation (ASCT) for relapsed or high-risk diffuse large B-cell lymphoma (DLBCL) between 1990 and 2007, we conducted a nationwide survey using the registry database of the Japan Society for Hematopoietic Cell Transplantation. Of the 1222 patients in the database, 576 (47%) received ASCT in first complete remission (CR1), 140 (12%) in first partial remission, 281 (23%) in sensitive relapse, 150 (12%) in resistant or sensitivity-unknown relapse, and 75 (6%) in primary refractory status. With a median follow-up of 22 months, the 2-year overall survival (OS) and progression-free survival rates were 71% and 68%, respectively. The cumulative incidences of 2-year non-relapse mortality and relapse/progression were 6% and 26%, respectively. Relapse/progression after ASCT in the rituximab era (2002–2007) was significantly lower than that in the pre-rituximab era (1990–2001; P

Although anti-T lymphocyte globulin-Fresenius (ATG-F) is commonly used as prophylaxis for graft-versus-host disease (GVHD), the appropriate dosage of ATG-F in the setting of a reduced-intensity conditioning (RIC) regimen has not been determined. In the present study, we retrospectively analyzed the clinical outcomes of 103 patients after unrelated bone marrow transplant (uBMT) with RIC regimens. RIC regimens consisted of purine analogue plus busulfan with low-dose TBI or ATG-F (5–10 mg/kg in total). Median age was 57 years (range 20–68). The incidence of grade II–IV acute GVHD and chronic GVHD with ATG-F was significantly lower than that with TBI 2 Gy (15 vs. 61 %, P < 0.05; 33 vs. 57 %, P < 0.05). The incidence of 2-year NRM with ATG-F was significantly lower than that with TBI 2 Gy (6 vs. 28 %, P < 0.05). There was no statistically significant difference in the cumulative incidence of 2-year relapse between the ATG-F and TBI 2 Gy groups (37 vs. 20 %, P = 0.13). In conclusion, the addition of low-dose ATG-F to GVHD prophylaxis in patients who received uBMT resulted in decreased incidence of acute and chronic GVHD, which led to a significantly reduced risk of NRM without compromising overall survival. The beneficial effect of low-dose ATG-F should be assessed in a prospective clinical trial.

Okinawa Prefecture, located in the subtropics, is an area of endemic adult T-cell leukemia–lymphoma (ATL) in Japan. We retrospectively analyzed 659 patients with aggressive ATL in seven institutions in Okinawa between 2002 and 2011. The median patient age was 68 years. More patients were aged ≥90 years (2.6 %), in this study, than in a nationwide survey (

The outcome after allogeneic hematopoietic stem cell transplantation (allo-HCT) for diffuse large B-cell lymphoma (DLBCL) associated with follicular lymphoma (FL), which includes DLBCL with pre- or co-existing FL, remains controversial, and few previous reports have compared the outcomes after allo-HCT for FL, DLBCL associated with FL, and de novo DLBCL. We retrospectively analyzed 97 consecutive patients with FL (n = 46), DLBCL associated with FL (n = 22), or de novo DLBCL (n = 29) who received allo-HCT at our institute between 2000 and 2010. With a median follow-up of 53 months, the 5-year overall survival (OS) and progression-free survival (PFS) were, respectively, 77% and 70% for FL, 62% and 57% for DLBCL associated with FL, and 26% and 23% for de novo DLBCL. The 5-year cumulative incidences of non-relapse mortality and disease progression/relapse were, respectively, 16% and 15% for FL, 19% and 24% for DLBCL associated with FL, and 36% and 41% for de novo DLBCL. By a multivariate analysis, the OS and PFS for DLBCL associated with FL were significantly better than those for de novo DLBCL, whereas they were not significantly different from those for FL. These results suggest that allo-HCT may be a promising option for patients with not only advanced FL but also DLBCL associated with FL.

A20 is a negative regulator of the NF-kappaB pathway and was initially identified as being rapidly induced after tumour-necrosis factor-alpha stimulation. It has a pivotal role in regulation of the immune response and prevents excessive activation of NF-kappaB in response to a variety of external stimuli; recent genetic studies have disclosed putative associations of polymorphic A20 (also called TNFAIP3) alleles with autoimmune disease risk. However, the involvement of A20 in the development of human cancers is unknown. Here we show, using a genome-wide analysis of genetic lesions in 238 B-cell lymphomas, that A20 is a common genetic target in B-lineage lymphomas. A20 is frequently inactivated by somatic mutations and/or deletions in mucosa-associated tissue lymphoma (18 out of 87; 21.8%) and Hodgkin's lymphoma of nodular sclerosis histology (5 out of 15; 33.3%), and, to a lesser extent, in other B-lineage lymphomas. When re-expressed in a lymphoma-derived cell line with no functional A20 alleles, wild-type A20, but not mutant A20, resulted in suppression of cell growth and induction of apoptosis, accompanied by downregulation of NF-kappaB activation. The A20-deficient cells stably generated tumours in immunodeficient mice, whereas the tumorigenicity was effectively suppressed by re-expression of A20. In A20-deficient cells, suppression of both cell growth and NF-kappaB activity due to re-expression of A20 depended, at least partly, on cell-surface-receptor signalling, including the tumour-necrosis factor receptor. Considering the physiological function of A20 in the negative modulation of NF-kappaB activation induced by multiple upstream stimuli, our findings indicate that uncontrolled signalling of NF-kappaB caused by loss of A20 function is involved in the pathogenesis of subsets of B-lineage lymphomas.

e19036 Background: Aggressive adult T-cell leukemia/lymphoma (aATL) has an extremely poor prognosis (median OS, 8-10 months). Okinawa, Japan’s only subtropical region, is hyperendemic for aATL. Recently, we demonstrated poorer outcomes among aATL patients in Okinawa compared with patients elsewhere in Japan, and a possible association of strongyloidiasis with ATL-related death. Two prognostic indices (PIs)—ATL-PI and JCOG-PI—have been developed using a database of national surveys and clinical trials. However, aATL patients in Okinawa were not included. This study aimed to validate these PIs using an Okinawa database. We also investigated the impact of strongyloidiasis on aATL patient survival. Methods: We constructed a clinical database of aATL patients from 7 institutions in Okinawa diagnosed between January 2002 and December 2011. The study endpoint was OS. Standard survival analysis methods (Kaplan-Meier method, log-rank test, and Cox proportional-hazards model) were used. Results: The study involved 433 evaluable patients (median OS, 6 months). Risks according to the two PIs in each patient were not always consistent (Table), but both PIs stratified aATL patients by risk. Three-year OS rates for ATL-PI were 35.9% (low-risk, n=66), 10.4% (intermediate-risk, n=256), and 1.6% (high-risk, n=111); rates for JCOG-PI were 22.4% (moderate-risk, n=176) and 5.3% (high-risk, n=257). Strongyloidiasis had little impact on OS (HR and 95% CI from univariate Cox analysis, 1.22 and 0.90-1.66, respectively). Multivariable Cox analysis returned almost the same factors as had been screened out in the previous studies to construct the two PIs. Conclusions: ATL-PI and JCOG-PI were well reproducible in Okinawa database. Strongyloidiasis did not affect prognosis in aATL patients. ATL-PI identifies low-risk aATL patients more clearly than JCOG-PI, and both identify high-risk patients with extremely poor prognosis. These will be useful to devise novel treatment strategies based on risk stratification of all Japan/world aATL patients. [Table: see text]

An increasing incidence of Pneumocystis jiroveci pneumonia (PCP) in patients with B-cell non-Hodgkin lymphoma (B-NHL) receiving rituximab treatment has been reported. We reviewed patients with B-NHL who underwent chemotherapy from 2004 to 2008 at our institution to identify risk factors for PCP development during and after chemotherapy. Among 297 patients with B-NHL, six developed PCP. Of 121 patients (41%) who received PCP prophylaxis with sulfamethoxazole–trimethoprim during chemotherapy, none developed PCP (0%), while among 176 patients (59%) who had no prophylaxis, six (3.4%) developed PCP at a median of 2 months (range: 1–3 months) after starting chemotherapy. Patients with CD4+ lymphocyte counts ≤200/mm3 before chemotherapy had a higher risk of developing PCP (p=0.045), while a history of rituximab treatment was not related to PCP. CD4+ lymphocyte counts ≤200/mm3 during and after chemotherapy were observed in 18.9% of patients.

Abstract 2318 Introduction: ES has been well recognized to increase non-relapse mortality (NRM) in autologous hematopoietic cell transplantation (HCT). However, little information is available on ES in allogeneic HCT, particularly after reduced-intensity conditioning. To determine the incidence, risk factors and outcome of ES after RIST, we retrospectively reviewed the medical records of 285 patients (pts) who underwent RIST at our institution between 1999 and 2007. Patients and Methods: After excluding 37 pts who received cord blood transplantation or a second or subsequent allogeneic HCT, and those who experienced primary graft failure, 248 pts (male 153, female 95) with a median age of 55 y (range, 21–68) fulfilled the criteria of this study. The underlying diagnosis included lymphoma (106 pts; 43%), acute myeloid leukemia (59; 24%), myelodysplastic syndrome (51; 21%), and others (32; 13%). Ninety-two pts (37%) had a standard-risk disease such as acute leukemia or lymphoma in CR1, and 156 (63%) had a high-risk disease. The donor and stem cell source were related PBSC in 170 pts (69%), related BM in 8 (3%), and unrelated BM in 70 (28%). All of the pts received reduced-intensity conditioning with a purine analog and a busulfan-based regimen. With the use of G-CSF after HCT, neutrophil engraftment (ANC ≥500/μ l) was achieved at a median of 12 days (range, 5–28). ES was diagnosed when the patient showed at least 2 of the following symptoms within 96 h of neutrophil engraftment: (1) fever (>38.0°C) without an identifiable source of infection, (2) skin rash (>25% of body surface area) that was not due to a drug reaction, (3) weight gain (>2.5% of baseline body weight), and (4) hypoxia (sPO2 Results: Among the 248 pts, 45 (18%) developed ES at a median of 14 days (6-23) after allogeneic HCT. Symptoms consisted of fever (100%), skin rash (53%), weight gain (62%) and hypoxia (33%). The incidence of ES in recipients of unrelated donor grafts was significantly higher than that in recipients of related donor grafts (28% vs 14%, p=0.02), and male pts had a significantly higher incidence of ES than female pts (22% vs 12%, p=0.04). No significant difference in the incidence of ES was observed between groups stratified according to age, stem cell source, TNC or CD34+ cell dose, disease risk, GVHD prophylaxis, and the timing of engraftment. By a multivariate analysis, use of an unrelated donor (HR 2.3, 95%CI 1.3–4.1, p=0.007), pts with lymphoid malignancy (HR 2.0, 95%CI 1.1–3.7, p=0.03) and male pts (HR 2.0, 95%CI 1.0–4.0, p=0.04) were associated with a significantly increased risk of ES. Among the 40 pts who required systemic corticosteroid therapy, 31 (78%) achieved a complete response, and the remaining 9 (23%) responded partially. There was a trend toward a higher incidence of grade III-IV acute GVHD (27% vs 18%, p=0.10) in pts with ES, and the incidence of extensive chronic GVHD (69% vs 62%, p=0.04) was significantly higher in pts with ES. Overall survival (OS) and non-relapse mortality (NRM) were not significantly different between the two groups, after a median follow-up of 53 months (range, 2–108) in surviving patents. A multivariate analysis showed that pts with high-risk disease (NRM, HR 1.9, 95%CI 1.1–3.3, p=0.01; OS, HR 2.5, 95%CI 1.6–3.8, p Conclusions: ES is a rather common complication after RIST, especially in recipients of unrelated donor grafts. Although ES shows an initial good response to corticosteroid therapy, ES may have a negative impact on survival, especially in pts with standard-risk disease, and this effect may be related to the occurrence of GVHD. The development of an optimal treatment for ES may improve the outcome of pts who have received RIST by reducing GVHD-related mortality. Disclosures: No relevant conflicts of interest to declare.

Abstract 3508 Background: Allo-HCT is a therapeutic option for patients (pts) with relapsed or refractory B-cell non-Hodgkin lymphoma (B-NHL). However, the outcome of allo-HCT with a reduced-intensity conditioning (RIC) regimen for TL remains controversial, and no previous reports have compared the outcomes after allo-HCT for FL, TL, and DLBCL in the rituximab era. Patients and Methods: We retrospectively analyzed 73 consecutive pts with FL (n=33), TL (n=18), or DLBCL (n=22) who received allo-HCT at our institute between January 2000 and December 2008. We defined TL as DLBCL that was histologically proven in pts with pre- (n=8) or co- (n=10) existing FL. The median age of the 73 pts was 47 years (range, 26–67). The median duration from diagnosis to HCT was 38 months (range, 6–175). The median number of prior chemotherapy regimens was 4 (range, 1–10): 57 pts (78%) had received prior rituximab, and 23 (32%) had received high-dose chemotherapy with autologous HCT prior to allo-HCT. The disease status at allo-HCT was CR or PR/refractory; 23 (32%)/50 (68%) for all pts, 10/23 for FL, 7/11 for TL, and 6/16 for DLBCL. The age-adjusted international prognostic index (aaIPI) at HCT was high or high-intermediate risk in 21 pts (28%), and FLIPI at HCT was high risk in 11 pts with FL (15%). The median level of serum albumin at HCT was 4.2 g/dL (range, 2.7–5.1). A myeloablative conditioning regimen was used for 14 pts (19%), and a RIC regimen was used for 59 pts (81%). The donor and stem cell source were related peripheral blood stem cells in 44 pts (60%), related bone marrow in 2 (3%), unrelated bone marrow in 21 (29%), and cord blood in 6 (8 %). Results: With a median follow-up of 68 months in surviving pts, the 5-year estimated overall survival (OS; Figure) and progression-free survival (PFS) were 58% and 54% for all pts, 80% and 71% for FL, 67% and 67% for TL, and 20% and 17% for DLBCL, respectively. The 5-year cumulative incidences of relapse/disease progression (PD) and non-relapse mortality were 25% and 39% for all pts, 10% and 22% for FL, 24% and 13% for TL, and 50% and 67% for DLBCL, respectively. Grade III-IV acute GVHD occurred in 25% of all pts, and OS was significantly worse in such pts [hazard ratio (HR) 2.5 (95%CI 1.2–5.3), p=0.02]. Extensive chronic GVHD (cGVHD) occurred in 53% of pts who survived 100 days or longer, and OS for patients with extensive cGVHD was significantly worse than that in pts without extensive cGVHD [HR 4.5 (1.3-15.6), p=0.02]. The cause of death included PD in 10 pts, GVHD in 5, infection in 4, non-infectious lung complication in 5, cerebral infarction or hemorrhage in 2, and unknown in 3. No pts with FL died of PD, and no pts with TL who had survived for 7 months after HCT relapsed thereafter. By a multivariate analysis, the OS for DLBCL was significantly worse than that of TL [HR 4.8 (1.7-13.2), p=0.002]. OS for FL was not significantly different from that of TL [HR 0.74 (0.2-2.4), p=0.6]. Other factors that influenced OS were aaIPI at HCT [high or high-intermediate risk, HR 3.7 (1.6-8.5), p=0.002] and the serum albumin level at HCT [ Conclusion: In the rituximab era, pts with TL showed more favorable outcomes after allo-HCT compared to those with DLBCL, and there was no significant difference in OS between FL and TL. AaIPI and the serum albumin level at HCT may be useful surrogate markers that influence OS for pts with B-NHL after allo-HCT. Allo-HCT, mainly with the use of a RIC regimen, might be a promising option for pts with relapsed or refractory FL and TL. Disclosures: No relevant conflicts of interest to declare.

Abstract 971 NFκB is a tightly regulated transcription factor of lymphocyte activation, proliferation and development. Controlled activity of NF κ B signaling pathway plays critical roles in coordination of immune and inflammatory response. Constitutive NFκB activation is recognized as a key pathological feature in several subsets of B-cell malignant lymphoma, and it is well known that lymphoma frequently occurred in association with chronic inflammation. Recently, our group showed frequent inactivation of A20, a negative regulator of NF κ B, in B-lineage malignant lymphomas. However, the molecular mechanism underlying the aberrant NF κ B activation in lymphomagenesis has not fully understood. In this study, to clarify the genetic basis of the aberrant NFκB activation, we performed genome-wide analysis of copy number alterations as well as allelic imbalances of primary B-lineage lymphoma specimens using Affymetrix GeneChip 250K genomic microarray with the CNAG/AsCNAR algorithm. We also searched for possible mutations in CARD11, CYLD, IKK and TRAF family genes and IκB genes, to obtain comprehensive registry of lesions in genes regulating NFκB pathway. This study included 238 primary lymphoma samples, including 64 samples of diffuse large B-cell lymphomas (DLBCL), 52 of follicular lymphomas (FL), 35 of mantle cell lymphomas (MCL), and 87 of mucosa-associated tissue (MALT) lymphomas. Five Hodgkin lymphoma-derived cell line (KM-H2, L1236, HDLM2, RPMI1666, L540) was also analyzed. Through a genome-wide analysis, we identified that each histology type had a unique genomic signature, suggesting a distinctive underlying molecular pathogenesis for different histology types. In contrast to the fact that A20 mutation was highly associated with loss of heterozygosity at 6q23.3, mutations of CARD11 (5 cases of DLBCL, 2 cases of MALT lymphoma) and IκB family genes (2 cases of DLBCL and 1 cases of MALT lymphoma) had no association with copy number abnormality at the locus of the genes. In total, mutations and copy number alterations in genes regulating NFκB pathway were found in more than 40% of B-cell lymphomas, which underpinned the importance of aberrant NFκB activation in lymphomagenesis. To also assess the role of uncontrolled signaling of NFκB pathway in lymphomagenesis, we re-expressed wild-type A20 in two lymphoma-derived cell lines without normal functional A20 alleles (KM-H2 and L1236). In both cells, re-expression of wild-type A20 resulted in suppression of cell growth and induction of apoptosis, accompanied by down-regulation of NFκB activation. The A20-deficient KM-H2 stably generated tumors in immunodeficient mice, whereas the tumorigenicity was effectively suppressed by re-expression of A20. We further investigated the role of A20 inactivation during clonal expansion of lymphoma by competitive proliferation assays using A20-deficient lymphoma-derived cell lines with or without re-expression of A20. The proportion of A20-expressing cells gradually decreased during competitive cell culture, and A20-expressing cells outgrew control cells in NOG mice, indicating the importance of A20 inactivation in clonal evolution of lymphoma. We demonstrated that uncontrolled NFκB signaling caused by alterations of multiple genes is a common feature of B-lineage lymphomas. Considering the physiological function of NFκB activation induced upon a variety of upstream stimuli, our observations provide an intriguing insight into and the pathogenesis of lymphoma. Our study also indicated that NFκB inhibition may have a role in lymphoma therapeutics. Disclosures: No relevant conflicts of interest to declare.

Abstract 3353 Poster Board III-241 [Background] Renal dysfunction is a life-threatening complication after hematopoietic stem cell transplantation, and the incidence of acute and chronic renal dysfunction after non-myeloablative transplantation is reportedly, 40 to 50% and 16 to 35%, respectively. In this study, we evaluated this complication after reduced-intensity stem cell transplantation (RIST) at a single institute. [Patients and Methods] We retrospectively reviewed the medical records of 286 patients (median age, 54 years; range, 21-68) with various hematological disorders who underwent RIST between 1999 and 2007, using a conditioning regimen that consisted of busulfan (oral 8 mg/kg or iv 6.4 mg/kg) and fludarabine (180 mg/m2, n=214) or cladribine (0.66 mg/kg, n=72). Sixty-seven patients also received 2-4 Gy of total body irradiation (TBI). The diagnosis included AML (n=77), ALL (n=9), MDS (n=56), malignant lymphoma (n=116), CML (n=14), and other (n=14). Whereas 188 patients were transplanted from a related donor (BM n=8, PB n=180), 98 were transplanted from an unrelated donor (BM n=80, PB n=1, CB n=17). GVHD prophylaxis consisted of cyclosporine (CSP, starting dose 3 mg/kg/day civ, target whole blood conc. 250-350 ng/ml, n=235) or tacrolimus (starting dose 0.03mg/kg/day civ, target whole blood conc. 10-20 ng/ml, n=51), with (n=131) or without (n=155) methotrexate, and 71 patients also received anti-human T-lymphocyte immunoglobulin (ATG, Fresenius, 5-10 mg/kg). Renal function was assessed in terms of the serum creatinine concentration and the estimated glomerular filtration rate (eGFR) as calculated by the Modification of Diet in Renal Disease equation (MDRD) for our population (eGFR=0.741*175*Age-0.203*Cr-1.154, *0.742, if female). Acute renal failure (ARF) within 100 days was categorized as grade 0 (decrease in eGFR of 25% but [Results] The median follow-up in surviving patients was 746 days (87-3291). The numbers of patients who developed grade 1, 2 and 3 ARF were 139, 72 and 9, respectively. The cumulative incidence of grade 2-3 ARF was 29 % at 100 days after RIST. The overall survival was significantly better in patients with grade 0-1 ARF than in those with grade 2-3 ARF (62% vs 42% at 2 years, p [Conslusion] We found that ARF was significantly associated with a higher mortality after busulfan-based RIST, and that preceding grade 2-3 ARF led to the occurrence of CKD in surviving patients, which highlights the importance of preventing ARF. This study also indicated that the incidence of acute and chronic renal complications was lower in our population compared to previously reported data from Western countries. Therefore, ethnicity should be taken into account when designing future international clinical trials. Disclosures: No relevant conflicts of interest to declare.

Abstract 270 BACKGROUNDS: A20 (also called TNFAIP3) gene encodes an ubiquitin-modifying enzyme involved in termination of NF-kB responses, and that is negative regulator of NF-kB. We previously reported that A20 is a common genetic target in B-cell lymphomas by using a genome–wide analysis of genetic lesions in 238 B-cell lymphomas. [Kato et al., Nature 459; 712, 2009] In the study, we showed that three Hodgkin's lymphoma (HL)-derived cell lines have deletion of both alleles or loss of one allele and a mutated allele of A20 gene by high-density typing using CNAG/AsCNAR algorism, which enabled accurate determination of allele-specific copy numbers, and thus allowed for sensitive detection of loss of heterozygosity (LOH) without apparent copy-number reduction, even in the presence of up to 70–80% normal cell contamination. Twenty-four primary samples from HL were also analyzed for the mutations and microdissected CD30-positive tumor cells (Hodgkin-Reed-Sternberg (HRS) cells) revealed one intronic and four missense mutations. However, we might have missed the homozygous deletions, which are shown in other B-cell lymphoma subtypes, and underestimates the frequency of involvement of A20 gene in primary HL cases. MATERIALS AND METHODS: The objects were 12 of the 24 samples of our previous cohort. The samples included one case that showed mutation in the previous study. We tried to detect A20 gene deletion in HRS cells with the combination of anti-CD30 immunofluorescence with FISH (fluorescence immunophenotyping and interphase cytogenetics as a tool for the investigation of neoplasm: FICTION) method. BAC clones suitable for FISH analysis of A20 gene were screened. A BAC clone RP11-783B20 (TNFAIP3 locus, 6q23) labeled with spectrum orange by nick translation, was selected because of the best signal/noise ratio. CEP 6 Spectrum Green Probe was used to detect centromere of chromosome 6 (6p11.1-q11). Approximately 4 micrometer thick slides were immunostained with anti-CD30 antibody and FISH was performed. As the diameter of HRS cells is known to be several times more than the 4 micrometer thickness of the section slides suitable for FISH, ratio of A20/CEP 6 signals were calculated and the A20 gene status was evaluated in CD30 positive cells. A20 and CEP6 signals were also counted in CD30 negative cells and used as a control. We counted signals of 30(15–55) CD30 positive cells and 50 CD30 negative cells. RESULTS: In 9 cases among 12 cases, signals were successfully counted. The average number of CEP6 signals on the CD30 negative cells was 1.53, which showed that a whole cell is located on the section slides. The ratio of A20/CEP6 signals on these CD30 negative cells were approximately 1.0 confirming the quality of FISH signals. As predicted, the average number of the CEP6 signals on the HRS cell was approximately 1.21, which suggested that one HRS cell is cut in the middle and divided into pieces during preparation of the section slides. In these CD30 positive cells, the A20/CEP6 ratio was less than 1.0; the cells contained less A20 gene signals than CEP6 signals. The relative numbers of signals determined by FICTION in 9 cases are shown in the table. We knew that the case #3 had A 20 gene mutation determined by direct sequence analysis of micro-dissected CD30 positive cells. As the sequence analysis showed no residual wild A20 genes, the relative ratio of 0.71 was considered as a consequence of loss of normal allele. Thus, the cut-off level of 0.71 was set to define deletion of at least one allele. Two cases showed that the A20/CEP6 signal ratio was more than this figure (0.90 in case #1, and 0.80 in case #4). Except for these 2 cases, 7 out of 9 cases were judged to have deletions of A20 gene. Notably case #6 had the ratio of 0.14, which suggested that virtually complete loss of A20 gene had occurred. The sequence analysis had missed the deletions probably due to the contaminated surrounding cells. CONCLUSIONS: We found a frequent deletion of A20 gene in HL cases including a case with a complete loss. The involvement A20 gene deletion associated with the pathogenesis of HL might be more frequent than previously suggested by the sequence analysis. Disclosures: No relevant conflicts of interest to declare.

Abstract 3370 Poster Board III-258 Although the outcome of patients (pts) with adult T-cell leukemia-lymphoma (ATL) remains poor when they are treated with conventional chemotherapy, we previously showed in a multi-center prospective study that one-third of pts who underwent RIST from a related donor in CR or PR could survive without disease for more than 2 years (Tanosaki R et al., BBMT 2008). In this retrospective study, we reviewed our single-center experience with RIST for ATL pts, focusing on the outcome of those who underwent RIST in non-remission status or who relapsed after RIST. A total of 24 pts underwent RIST from a related donor between 2001 and 2008. The median age was 54 years (range, 44-65). Of the 14 males and 10 females, 19 were acute type and 5 were lymphoma type. Disease status at transplantation was 5 CR, 10 PR, 8 NC and 1 PD. Donors were siblings in 18 and children in 6, including 5 HTLV-1 healthy carriers. HLA in serology was 6/6 in 19 and 5/6 in 5. Stem cell sources were PBSC in 22 and BM in 2. Conditioning regimens were fludarabine (30 mg/m2 iv days -8 to -3) and busulfan (3.2 mg/kg iv days -6 and -5) with (n=8) or without (n=16) rabbit anti-T-lymphocyte globulin (ATG, Fresenius; 2.5 mg/kg iv days -2 and -1). All patients received cyclosporine alone for GVHD prophylaxis. Engraftment was rapid in all 24 pts (neutrophil>500/uL; median 12 days, range 10 -19), with no graft failure. There were 3 non-relapse mortalities; respiratory failure from bronchiolitis obliterans at 21 months (mos), interstitial pneumonitis at 47 mos, and pneumococcal sepsis at 38 mos. Notably, 10 of the 19 pts who were non-CR at RIST survived without disease progression to a median of 53 mos (range, 20 to 85). All of these pts were acute type, and had circulating ATL cells in the peripheral blood (PB) immediately before RIST (average 33% of WBC, range 5-73). Circulating ATL cells decreased to below 5% within 1 mo in 8 pts. A total of 12 pts relapsed within 16 mos; 7 (58%) within 3 mos, and 11 (92%) within 12 mos. Two patients who had relapsed after RIST showed a significant but transient response to the withdrawal of immunosuppression (CR 1, PR 1). Donor lymphocyte infusion was performed in 6 pts without significant benefits. Seven pts who relapsed at a single site, which was confirmed by CT scan or FDG-PET, were treated with local irradiation alone, and 3 whose HTLV-1 proviral load in PB had become negative at relapse survived to 48, 64 and 77 mos; 1 pt required 2 courses of irradiation because of immediate relapse at the margin of the preceding radiation field, and another pt underwent surgical resection of a residual mass since a biopsy revealed a viable lesion at the irradiated site. The 5-year overall and progression-free survival of all pts were 52% (95% CI, 38-66%) and 37% (95% CI, 22-52%), respectively, at a median follow-up of 59 mos (range, 12 to 85) in surviving pts. HTLV-1 proviral load in PB was examined using real-time PCR for tax in 208 samples from 21 evaluable pts, and it became negative at least once in 15 pts (71%), including 1 pt whose donor was an HTVL-1 carrier; proviral load remained negative in 7 pts at a median follow-up of 32 mos (range, 3 to 84). Since HTLV-1 tax is a promising target molecule for identifying the immunological mechanism, HLA-restricted tax-specific CTLs were examined in HLA-A2- and/or A24-positive pts using tax tetramers by taking blood samples periodically after informed consent was obtained from each pt. A total of 80 samples in 13 pts were analyzed. The number of tax tetramer-positive (tax+) cells did not change significantly up to at least 1 year after RIST, while the clinical responses and decrease/disappearance of HTLV-1 proviral load were observed within 3 mos in most cases. An increase in tax+ cells was observed after 1 year in 2 pts who had achieved CR. In conclusion, about half of the acute-type ATL pts with a significant involvement of ATL cells in PB at RIST could survive for a long time in our cohort. ATL pts who relapsed at a single site after RIST still have a chance to be cured with local treatment using irradiation alone or surgical resection with the aid of HTLV-1 proviral load as a marker for minimum residual disease. Since most ATL pts had already become resistant to chemotherapy and the intensity of conditioning was reduced, potent GV-ATL and GV-HTLV-1 effects might have played a key role in disease control. However, tax-specific CTL kinetics did not correlate with either clinical responses or the HTLV-1 proviral load, which suggested that other molecules may be immunologically targeted. Our results might contribute to the establishment of the cure-oriented treatment strategy for ATL pts. Disclosures: No relevant conflicts of interest to declare.

Background: The management for recurrent lymphoma after allogeneic hematopoietic stem cell transplantation (HSCT) is challenging and it is not clear concerning its outcomes. Since potent graft-versus-lymphoma (GVL) effect is observed in some patients (pts) with follicular lymphoma (FL), peripheral T-cell lymphoma (PTCL) or adult T-cell leukemia/lymphoma (ATL) after allogeneic HSCT using reduced-intensity conditioning (Tanosaki et al, ASH 2006, BBMT2008;14:702), we hypothesized that it could also be exerted in relapsed patients. Hence, we retrospectively analyzed the outcomes among patients with lymphoma who relapsed after allogeneic HSCT. Patients and Methods: Between January 1999 and December 2007, 164 patients with refractory lymphoma, who were deemed incurable with standard or high-dose chemotherapy, underwent allogeneic HSCT in our single institution. Seventy-five pts remain in CR, 33 died associated with non-relapse mortality, 4 were lost follow-up, and 52 pts relapsed after the median of 71 days (range 1–1457), including 1 graft failure. Therefore, 51 pts were analyzed in this study. Results: Histological subtypes were diffuse large B-cell lymphoma (DLBCL) 13, ATL 12, lymphoblastic lymphoma 8, PTCL 7, FL 4, Hodgkin lymphoma (HL) 3, mantle cell lymphoma (MCL) 2, extranodal NK/T-cell lymphoma 2. Ten pts had an early relapse within 30 days, while 41 pts had a systemic (18 pts) or solitary relapse (23 pts) after the median of 98 days (range 33–1457). Relapse-oriented interventions included withdrawal of immunosuppressants (WIS), irradiation or surgical resection of localized lesion, donor lymphocyte infusion (DLI) and chemotherapy, which were used singly or in combination according to the clinical conditions; the breakdown and its outcomes were shown in Table. At the time of relapse, 46 pts (90%) had immunosuppressants, and WIS was first tried in 31 pts who didn’t have GVHD. Overall survival rate (OS) at 2 years after relapse in pts with relapse >day 30 and ≤day 30 was 37±8% and 0% (p Conclusions: These results suggested that some patients with chemo-refractory lymphoma with an immunogenic property could achieve a long durable disease-free survival after relapse without aggressive treatment modalities. Interventions and Outcomes Total Interventions Response WIS Ope/RT DLI Chemo CR (CCR)* PR NC/PD *CCR; continuous complete remission after relapse. Relapse >Day 30 Solitary 23 10 12 2 1 10 (5) 0 11 Systemic 18 15 0 1 1 6 (4) 1 11 Relapse ≤ Day 30 10 6 0 2 2 0 (0) 1 9 Total 51 31 12 5 4 16 (9) 2 31

Background: Ranimustine (MCNU), a derivative of nitrosourea that was developed in Japan, shows good penetration into cerebrospinal fluid and might be expected to decrease CNS relapse of lymphoid malignancies when used before transplantation (Takaue Y, et al. Cancer67:1830, 1991). However, its feasibility and efficacy have not been extensively analyzed in adult patients with lymphoma. Patients and Methods: We retrospectively evaluated an MCEC regimen which consisted of MCNU (200 mg/m2 on days -8 and -3), carboplatin (300 mg/m2 on days -7 through -4), etoposide (500 mg/m2 on days -6 through -4) and cyclophosphamide (50 mg/kg on days -3 and -2) in 68 patients with lymphoma (median age, 48 yr: range, 20–65 yr) who underwent autologous peripheral blood stem cell transplantation (PBSCT) at our institution between January 1999 and February 2008. The diagnosis included diffuse large B-cell lymphoma (n=32), T-cell lymphoma (12), Hodgkin lymphoma (9), follicular lymphoma (14), including 6 transformants, and intravascular lymphoma (1). The median time from diagnosis to PBSCT was 20 months (4–198 mo), and the median number of prior chemotherapy regimens was 3 (2–7). Sixty-six patients (97%) had ECOG PS< 2, 28 (41%) had prior XRT, 3 (4%) had bulky disease, 42 (62%) had stage III–IV disease, 19 (28%) had IPI at relapse of high (H) or high-intermediate (H-I), and one had IPS >4. The disease status at transplantation was 1st CR/PR in 10 (15%), ≥2nd CR/PR in 49 (72%), and NC/PD in 9 (13%). Results: The median number of days to neutrophils> 500 /μL and platelets> 20,000 /μL was 10 (8–19) and 10 (4–30), respectively. The platelet count did not decrease below 20,000 /μL in 9 patients. Grade (G) 4 non-hematological toxicities (CTCAE ver. 3.0) included elevated transaminase (1), hyponatremia of 116 mEq/L (1) and hypokalemia of 2.3 mEq/L (1). GI and CNS toxicities consisted of mucositis (G3, n=10), diarrhea (G3, n=27) and seizure (G2, n=1). G1 interstitial pneumonitis was observed in 5 patients. There were no grade 4 cardiac, pulmonary or renal toxicities except for one patient who died of treatment-related multi-organ failure. The cumulative incidence of relapse at 2 yrs was 49 %. Of 33 relapses, 2 occurred newly in the CNS without any previous history. With a median follow-up of 24 months (3–80) after transplantation for surviving patients, the 2-year overall survival (OS) and progression-free survival (PFS) were 68% (95% CI 55–81%) and 50% (95% CI 38–63%), respectively. In univariate analyses, T-cell phenotype (P Conclusion: The results suggested that autologous PBSCT with the MCEC regimen is a feasible and effective treatment option for relapsed/refractory lymphoma.

Majority of malignant lymphoma arising from the ocular adnexae are ocular adnexal MALT lymphomas (OAL). Several genetic abnormalities, including t(14;18)(q32;q21), trisomy 18 and trisomy 3 have been reported in OAL. However, none of them are found in more than half of cases with OAL by conventional methods. High density Single Nucleotide Polymorphism (SNP) array analysis with CNAG/AsCNAR algorithm allows high-resolution and genome-wide detection of both loss of heterozygosity (LOH) and copy number abnormality, especially uniparental disomy (UPD), without depending on the availability of paired normal DNA (Yamamoto et al, Am J Hum Genet. 2007; 81:114–26). UPD is acquired by somatic recombination and therefore not detected by conventional cytogenetic analysis or array CGH. In this study we analyzed DNA from OAL for the presence of LOH with or without copy number changes. Tissue samples from patients with OAL at our institute between 1995 and 2003 (N=32) were subject to SNP-array (250K NspI) analysis with CNAG/AsCNAR algorithm. The patients included 21 males and 11 females, and the median age of 56.5 years at the time of diagnosis (range, 15–90 years). Clinical stage was I (29 cases), II (1 case), and IV (2 cases). Trisomy of 3, 18, and 21 were found in 9, 7 and 1 cases, respectively. Overall, LOH due to UPD more than one chromosomal band were found in 16 cases (50%). Recurrent one allele deletion was detected at 6q23-24, 12p13 and 17q21 (N=15, 15 and 21, respectively). In total, 28 (82%) among 32 cases showed one or more of the above changes. Characterization of these recurrent genetic abnormalities might reveal the pathogenesis of OAL.

Background The TNFAIP3 gene, which encodes a ubiquitin-modifying enzyme (A20) involved in the negative regulation of NF-κB signaling, is frequently inactivated by gene deletions/mutations in a variety of B-cell malignancies. However, the detection of this in primary Hodgkin lymphoma (HL) specimens is hampered by the scarcity of Hodgkin Reed-Sternberg (HR-S) cells even after enrichment by micro-dissection. Methods We used anti-CD30 immunofluorescence with fluorescence in-situ hybridization (FISH) to evaluate the relative number of TNFAIP3/CEP6 double-positive signals in CD30-positive cells. Results From a total of 47 primary classical Hodgkin lymphoma (cHL) specimens, 44 were evaluable. We found that the relative numbers of TNFAIP3/CD30 cells were distributed among three groups, corresponding to those having homozygous (11%), heterozygous (32%), and no (57%) deletions in TNFAIP3. This shows that TNFAIP3 deletions could be sensitively detected using our chosen methods. Conclusions Comparing the results with mutation analysis, TNFAIP3 inactivation was shown to have escaped detection in many samples with homozygous deletions. This suggests that TNFAIP3 inactivation in primary cHL specimens might be more frequent than previously reported.