Clinicopathological Analysis of 320 Cases of Diffuse Large B-cell Lymphoma Using the Hans Classifier.

The estimation of clinical prognosis for diffuse large B-cell lymphoma (DLBCL) with a quick, cost-efficient method is necessary because of the clinical heterogeneity of this disease, which leads to death, relapsed or refractory disease in approximately 40% of patients. We analyzed 320 cases diagnosed from 2007 to 2013 treated with R-CHOP therapy at Tokai University Hospital and associated institutions. DLBCL was classified according to the cell-of-origin using the Hans algorithm [germinal center B-cell-like (GCB) vs non-GCB subtypes], and into 6 subgroups derived from combinations of CD10, BCL6 and MUM1 markers. The percentage of GCB and non-GCB (NGCB) subtypes was 35% and 65%, respectively. GCB-DLBCL was characterized by lower BCL2 immunohistochemical expression, extranodal sites ＜1, better therapeutic response, and favorable overall survival (OS) and progression free survival (PFS) (P＜0.01). The most frequent subgroup was NGCB-1 (CD10-BCL6+MUM1+, 51%) followed by GCB-1 (CD10+BCL6+or-MUM1+, 21%), NGCB-2 (CD10-BCL6-MUM1+, 13%), GCB-2 (CD10+BCL6+or-MUM1-, 10%), GCB-3 (CD10-BCL6+MUM1-, 4%) and NGCB-3 (CD10-BCL6-MUM1-, 2%). In comparison with GCB-2 and GCB-3 (both MUM1-), the GCB-1 (MUM1+) was characterized by favorable PFS (5-year PFS 84% vs 65%, OR 0.368, P＜0.05), independent of high LDH (associated with unfavorable PFS, OR 7.04, P＜0.01) in the multivariate analysis. This predictive value of MUM1 was independent of CD10. Interestingly, triple-negative NGCB-3 tended to have a more favorable prognosis than the other NGCB subgroups. In conclusion, the Hans classifier is a valid method to evaluate the prognosis of DLBCL NOS. In the GCB subtypes, GCB subtypes, MUM1-positivity is associated with a more favorable outcome (PFS).

Autologous G-CSF-mobilized peripheral blood CD34+ cell therapy for diabetic patients with chronic nonhealing ulcer.

Recently, animal studies have demonstrated the efficacy of endothelial progenitor cell (EPC) therapy for diabetic wound healing. Based on these preclinical studies, we performed a prospective clinical trial phase I/IIa study of autologous G-CSF-mobilized peripheral blood (PB) CD34(+) cell transplantation for nonhealing diabetic foot patients. Diabetic patients with nonhealing foot ulcers were treated with 2 × 10(7) cells of G-CSF-mobilized PB CD34(+) cells as EPC-enriched population. Safety and efficacy (wound closure and vascular perfusion) were evaluated 12 weeks posttherapy and further followed for complete wound closure and recurrence. A total of five patients were enrolled. Although minor amputation and recurrence were seen in three out of five patients, no death, other serious adverse events, or major amputation was seen following transplantation. Complete wound closure was observed at an average of 18 weeks with increased vascular perfusion in all patients. The outcomes of this prospective clinical study indicate the safety and feasibility of CD34(+) cell therapy in patients with diabetic nonhealing wounds.

Effect of liposome-encapsulated hemoglobin on antigen-presenting cells in mice.

Liposome-encapsulated hemoglobin (LEH) is removed from the circulation and degraded in the reticuloendothelial system, including dendritic cells (DCs) and macrophages. Therefore, LEH at a large dose may overload the system, cause a competitive inhibition in antigen-presenting activity, and impair the immune response of the host. Changes in cellularity of immunocompetent cells were monitored serially up to 4 weeks by flow cytometry in wild-type mice receiving 20 mL/kg of LEH, syngeneic red blood cells (RBCs), or saline. DCs were collected from the host spleen 1, 7, and 28 days after receiving the solution and were cocultured with naïve cluster of differentiation 4 T cells from T-cell receptor transgenic mice in the absence or presence of third-party antigens. After LEH administration, the cellularity of DCs and macrophages in the recipient spleen remained unchanged from control mice receiving RBCs or saline. While subset populations and costimulatory molecule expressions were different, DCs from LEH-administered mice expressed high levels of interleukin-2 production and helper T-cell activation in response to a third-party antigen and superantigens, as did the DCs from control mice receiving RBCs or saline. The results suggest that 20 mL/kg of LEH does not greatly alter antigen-presenting activity to third-party antigens.

Outcome of medium-dose VP-16/CY/TBI superior to CY/TBI as a conditioning regimen for allogeneic stem cell transplantation in adult patients with acute lymphoblastic leukemia.

The choice of conditioning regimen before allogeneic stem cell transplantation (SCT) in patients with acute lymphoblastic leukemia (ALL) is important. We retrospectively compared outcomes of medium-dose VP-16/cyclophosphamide/total body irradiation (VP/CY/TBI) regimen and CY/TBI. Five hundred and twenty-nine patients (VP/CY/TBI: n = 35, CY/TBI: n = 494) who met all of the following criteria were compared: first time for SCT, aged 15-59 years; first or second complete remission at SCT; bone marrow or peripheral blood as stem cell source; and HLA phenotypically matched donor. Median age of the patients was 34 years, and patients who received VP/CY/TBI were younger (28 vs. 34 years, P = 0.02). Cumulative incidences of relapse and non-relapse mortality (NRM) were higher for patients who received CY/TBI (P = 0.01 for relapse, P < 0.01 for NRM). After a median follow-up period of 36.9 months, 5-year overall survival (OS) rates were 82.2% in the VP/CY/TBI group and 55.2% in the CY/TBI group. OS, and disease-free survival (DFS) in the VP/CY/TBI group were shown to be significantly better by multivariate analysis [hazard ratio: 0.21 (95% confidence interval: 0.06-0.49) for DFS, hazard ratio: 0.25 (95% confidence interval: 0.08-0.59) for OS]. VP/CY/TBI was associated with a lower relapse rate and no increase in NRM, resulting in better survival than that in CY/TBI for adult ALL patients.

Incidence, risk factors and outcomes of bronchiolitis obliterans after allogeneic stem cell transplantation.

Bronchiolitis obliterans (BO) after allogeneic stem cell transplantation (allo-SCT) is a late-onset, life-threatening respiratory complication that significantly reduces a patient's quality of life. We retrospectively analysed the incidence of and risk factors for BO in allo-SCT recipients. In 2087 patients who underwent allo-SCT between January 1994 and June 2005 and survived >90 days after transplantation, 57 patients developed BO with a 5-year cumulative incidence of 2.8%. The median time interval from transplantation to BO diagnosis was 335 days (range 83-907 days). The 5-year cumulative incidence of BO was 1.62% in bone marrow transplantation (BMT) from related donors, 3.83% in peripheral blood stem cell transplantation (PBSCT) from related donors (R-PBSCT), 2.91% in BMT from unrelated donors and 2.65% in unrelated cord blood transplantation. The incidence of BO after R-PBSCT was significantly higher than that after any other type of allo-SCT (p = 0.02). R-PBSCT (p = 0.019) and preceding chronic graft-versus-host disease (GVHD) (p < 0.001) were BO-associated risk factors. Overall 5-year survival of patients with BO from the time of diagnosis was 45.4%, significantly less than those without (77.5% from day 335, p < 0.001). R-PBSCT recipients with existent chronic GVHD have a high risk of developing BO, and need extensive care and repeated pulmonary function tests.

[Autologous transfusion for patients with increased risk of massive and/or critical bleeding--indication, efficacy and limitations].

The "Guideline for Management of Critical Bleeding in Obstetrics" was implemented by five obstetrics-related societies in April 2010 to improve management strategies and outcomes of massive bleeding in obstetrics. Besides emergency transfusion replacing acute blood loss, the Guideline contains a flow chart of instructions for autologous blood transfusion in obstetric patients with rare blood types, irregular antibodies, or with an increased risk and/or history of massive bleeding. In this chapter, based on the characteristics of bleeding in the field of obstetrics, indication, contraindication, patient selection, preparation, and pitfalls regarding actual practices of autologous blood transfusion are detailed in terms of efficacy and limitations. The pros and cons of autologous blood transfusion are discussed together with the feasibility of intra/post-operative blood salvage and hemodilutional autologous blood transfusion during bleeding to reduce the total amount of transfusion and thereby improve the outcome of critical bleeding in obstetrics.

Peripheral blood stem cell versus bone marrow transplantation from HLA-identical sibling donors in patients with leukemia: a propensity score-based comparison from the Japan Society for Hematopoietic Stem Cell Transplantation registry.

We retrospectively analyzed the results of 707 adult patients who underwent myeloablative peripheral blood stem cell transplantation (PBSCT) (n = 365) and myeloablative bone marrow transplantation (BMT) (n = 342) for leukemia from HLA-identical sibling donors between 2000 and 2005 using the propensity score method. The results were obtained from the Japan Society for Hematopoietic Cell Transplantation registry. Multivariate Cox analysis showed that PBSCT was associated with lower overall survival (OS) in standard-risk patients [adjusted hazard ratio (aHR) = 1.83; 95% confidence interval (CI) 1.04-3.23; P = 0.036], but not in high-risk patients (aHR = 1.11; 95% CI 0.76-1.61; P = 0.599). Hematopoietic recovery was significantly faster after PBSCT. The risk of acquiring grade III-IV acute graft-versus-host disease (GVHD) (aHR = 2.23; P = 0.040) and extensive chronic GVHD (aHR = 1.93; P = 0.001) were significantly higher after PBSCT. PBSCT was associated with higher non-relapse mortality in standard-risk patients (aHR = 2.30; 95% CI 1.08-4.88; P = 0.030), but not in high-risk patients (aHR = 1.29; 95% CI 0.65-2.54; P = 0.468). Relapse after transplantation did not differ between PBSCT and BMT either in standard-risk group or in high-risk group (aHR = 1.17; 95% CI 0.55-2.52; P = 0.684 and aHR = 0.81; 95% CI 0.52-1.28; P = 0.370, respectively). In this retrospective analysis, OS was significantly lower after PBSCT in standard-risk patients, but not in high-risk patients. PBSCT was associated with significant risks of grade III-IV acute GVHD and extensive chronic GVHD.

Phase I/II study of humanized anti-CD33 antibody conjugated with calicheamicin, gemtuzumab ozogamicin, in relapsed or refractory acute myeloid leukemia: final results of Japanese multicenter cooperative study.

The primary objective of this study was to investigate the tolerability, efficacy and pharmacokinetic profile of gemtuzumab ozogamicin (GO) in patients with relapsed and/or refractory CD33-positive acute myeloid leukemia (AML). Patients received 2-h infusions of GO twice with an interval of approximately 14 days. Tolerability was assessed using the National Cancer Institute Common Toxicity Criteria Version 2.0. Samples for pharmacokinetics were taken on day 1 and day 8 of the first treatment cycle. The dose was increased stepwise and, in each cohort, patients were treated at the same dose. Forty patients, median age 58 years (range 28-68) were treated; 20 and 20 patients were enrolled to the phase I and II parts, respectively. In the phase I part, dose-limiting toxicities (DLTs) were hepatotoxicities, and the recommended dose was established as 9 mg/m2 given as two intravenous infusions separated by approximately 14 days. The pharmacokinetic study revealed that Cmax and AUC were equivalent to those of non-Japanese patients. In the phase II part, complete remission was observed in 5 patients, and one patient had complete remission without platelet recovery. Four of these 6 in remission and one in the phase I are long-term survivors (alive for at least 44 months). GO is safe and effective as a single agent among Japanese CD33-positive AML patients. Remission lasted longer in a subset of patients than in non-Japanese patients in earlier studies. Further studies of this agent are warranted to establish standard therapy.

Possible role of artificial oxygen carriers in transfusion medicine: a retrospective analysis on the current transfusion practice.

Artificial oxygen carriers (AOC) are under development as a substitute for red blood cells (RBC) in homologous transfusion (Tx). The lack of surface antigen in AOC makes ABO-typing and antibody-screening (T/S) unnecessary. Pathogen elimination renders it much safer, and long-term stability allows ubiquitous storage for emergency use. To delineate the utility of AOC, we retrospectively examined current Tx practices in Tokai University and the Japanese Red Cross Society. The emergency department of Tokai University Hospital has been using O(+)Rh(+) RBC in patients with hemorrhagic shock before Tx becomes available. Those who received the RBCs within 60 min of injury had a significantly higher survival rate than those who received it later (> or =60 min). The Red Cross Blood Center provided 411 units of RBC for 138 urgent requests for rare blood types. Our analysis suggests that if an AOC were available for the initial six units, 96% of such requests could have been covered to avoid urgent donor allocation, preparation, and Tx. Among 2079 surgical cases who ordered T/S, only 29% actually required Tx, rendering >70% of the T/S unnecessary. Because only 7.4% required nine units or more, more than 92% of T/S and Tx could have been avoided in retrospect if an AOC were available for the initial eight units. The results suggest that an AOC might be useful in various situations to alleviate problems, concerns, and technical burden in the current Tx practices. Because the expected utility is based mainly on physical characteristics, AOC may remain advantageous even when biogenetically derived RBC becomes available.

[Critical massive blood transfusion--O type and variant conformity blood transfusion].

Variant conformity blood is often used in transfusion at massive unpredictable hemorrhage under an operation or at hemorrhagic shock in a lifesaving emergency arena. This is because of difficulty in performing blood grouping, because of difficulty in deciding blood types in the laboratory, or because of lack in store of the same blood type transfusion as a pharmaceutical preparation needed for the patients. In performing variant conformity transfusion in the hospital, it is vitally important that in-house system be well-organized and staff be thoroughly informed about variant conformity transfusion. This chapter explains system arrangements of organization for variant conformity transfusion in hospital settings based on our experience in the urgent use of O type RCC-LR (Red Cells Concentrates-Leukocytes Reduce, "Nisseki") pharmaceutical preparation.

Allogeneic hematopoietic stem cell transplantation for Epstein-Barr virus-associated T/natural killer-cell lymphoproliferative disease in Japan.

Epstein-Barr virus (EBV)-associated T/NK-cell lymphoproliferative disease (LPD) has been linked to several different disorders. Its prognosis is generally poor and a treatment strategy has yet to be established. There are reports, however, that hematopoietic stem cell transplantation (HSCT) can cure this disease. To clarify the current situation regarding allogeneic hematopoietic stem cell transplantation (allo-HSCT) for EBV-associated T/NK-LPD, a nationwide survey was performed in Japan. Data for 74 patients were collected. There were 42 cases of chronic active EBV infection (CAEBV), 10 cases of EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH), and 22 cases of EBV-associated lymphoma/leukemia (EBV-lymphoma/leukemia). Of those with CAEBV, 54% had the EBV-infected T-cell type and 59% with EBV-lymphoma/leukemia had the EBV-infected NK-cell type. Most patients with EBV-HLH and EBV-lymphoma/leukemia received allo-HSCT within 1 year after onset compared to only 14% of patients with CAEBV. The event-free survival (EFS) rate following allo-HSCT was 0.561 +/- 0.086 for CAEBV, 0.614 +/- 0.186 for EBV-HLH, and 0.309 +/- 0.107 for EBV-lymphoma/leukemia. The EFS of allo-HSCT with conventional conditioning was 0.488 +/- 0.074 and with reduced-intensity conditioning was 0.563 +/- 0.124. Thus, in a substantial number of cases, EBV-associated T/NK-LPD can be cured by either allogeneic conventional stem cell transplantation or reduced-intensity stem cell transplantation.

Remission of lymphoma after withdrawal of methotrexate in rheumatoid arthritis: relationship with type of latent Epstein-Barr virus infection.

Rheumatoid arthritis (RA) is associated with an increased risk of developing lymphoma. Although the pathogenesis is still unclear, the increased risk appears to be related to the high inflammatory activity of RA, immunosuppressive agents, or Epstein-Barr virus (EBV) infection. We investigated the relationship between EBV latent infection and methotrexate (MTX)-associated lymphoma in RA patients. Nine patients were diagnosed with non-Hodgkin's lymphoma (NHL) during MTX treatment for RA in a multicenter study. The pathologic findings were consistent with diffuse large B-cell lymphoma in 8 patients and peripheral T-cell lymphoma, unspecified in 1. EBV infection was detected in 3 patients by in situ hybridization. Among all 9 patients who were initially treated by MTX withdrawal alone, 2 obtained spontaneous complete response (CR), 1 had partial response, 2 had stable disease (SD), and 4 had progressive disease. Both patients who had a CR and 1 who had SD were positive for EBV. Further examination of the latent EBV infection patterns revealed that 2 patients who obtained a CR had latency Type III, and the other with SD had latency Type II. These results demonstrate that immunodeficiency caused by MTX treatment is associated with the development of EBV-related NHL in RA patients. In patients who were treated by MTX for RA and developed NHL, remission can be observed following MTX withdrawal especially in NHL with latency Type III EBV infection. The analysis of EBV infection, including the latency types, is useful to decide the optimum therapeutic strategy.

[Allogeneic bone marrow transplantation for chemotherapy-resistant T-prolymphocytic leukemia].

A 34-year-old female was referred to our hospital for the evaluation of atypical lymphocytosis. Leukocyte count at diagnosis was 17,900/microl with 58% atypical lymphocytes having a convoluted nucleus and prominent nucleoli. Because the leukocyte count increased to 43,600/microl, the patient was treated with 2'deoxycoformycin followed by CHOP combination chemotherapy. However, both treatments failed to achieve remission. We planned an allogeneic bone marrow transplantation from an HLA-matched unrelated donor. The patient was treated with Ara-C and etoposide before conditioning to decrease the high leukemia burden. After administration of total body irradiation (12 Gy in six fractions) and cyclophosphamide (total dose of 120 mg/kg) unmanipulated marrow cells were infused. Under prevention of GVHD by CsA and short-term MTX, leukocyte engraft was prompt at day 16, and acute GVHD grade II was observed. Because 9.4% of residual recipient type T-cells was seen with STR analysis on day 22, we decreased the dose of Cs'A. After the occurrence of mild acute GVHD, the residual T-cell number decreased. The patient is still in complete remission for up to 22 months after BMT. We conclude that allogeneic SCT is effective for the treatment of T-PLL.

Clinical effects of infusing anti-Epstein-Barr virus (EBV)-specific cytotoxic T-lymphocytes into patients with severe chronic active EBV infection.

Immune cell therapy with autologous Epstein-Barr virus (EBV)-specific cytotoxic T-lymphocytes (CTLs) or lymphokine-activated killer (LAK) cells was performed in 2 adults with severe chronic active EBV infection (SCAEBV). The patient in case 1, who had complications of pancytopenia, high fever, and massive splenomegaly, was treated with 13 doses of LAK cell infusion followed by 4 doses of autologous CTL infusion. The patient in case 2, who had liver dysfunction due to natural killer cell-type infection, was treated with 4 doses of autologous CTL infusion. In case 1, the LAK cell infusions were effective in lowering the viral load and improving several biochemical parameters (lactate dehydrogenase, soluble interleukin 2 receptor) and resulted in complete amelioration of the high fever. Subsequent infusions of autologous CTLs reduced the viral load only temporarily and were accompanied by an increase in frequency of EBV-specific T-cells in the blood. However, the patient's main problem of pancytopenia was not resolved. In case 2, infusion of autologous CTLs did not improve the patient's hepatic dysfunction or viral load but caused a significant increase in autoantibody levels. Thus the effect of auto-CTL treatment was limited or deteriorative in SCAEBV patients.