Masato Obara

Hypogammaglobulinemia with a selective delayed recovery in memory B cells and an impaired isotype expression after rituximab administration as an adjuvant to autologous stem cell transplantation for non-Hodgkin lymphoma.

Abstract:  Objectives: Some studies have indicated patients who received rituximab as adjuvant to stem cell transplantation had an increased risk of developing severe hypogammaglobulinemia. The mechanism of this hypogammaglobulinemia is unknown, although investigators have hypothesized a further delay in the B‐cell recovery as one potential etiology. The aim of this study is to clarify the mechanism(s) of this hypogammaglobulinemia. Methods: A total of 14 patients with high‐risk CD20+ lymphoma underwent an autologous peripheral blood stem cell transplantation (APBSCT). After a hematological recovery, rituximab was given weekly for up to four doses as an adjuvant therapy. Results: After a median follow up of 33.5 months, we found six patients (group A) who had hypogammaglobulinemia, while the eight other patients (group B) had normal serum immunoglobulin levels. A phenotypical analysis revealed that group A patients had already achieved B‐cell recovery. However, we found a severe delay in the recovery of CD27+ memory B cells, especially in the IgD−/CD27+ switched populations in group A, but CD27 negative naive B‐cells reverted to a normal range in both groups. Consistent with this, reverse transcriptase‐polymerase chain reaction studies with peripheral blood mononuclear cells revealed that most patients in group A lacked more than two classes of isotype transcripts. Conclusions: Abnormal repertoires and impaired isotype expression are seen in patients with common variable immunodeficiency, these data suggested that rituximab after APBSCT can affect not only the B‐cell quantities, but also the recovery of the B‐cell repertoires.

Excellent Outcome of Allogeneic Hematopoietic Stem Cell Transplantation Using a Conditioning Regimen with Medium-Dose VP-16, Cyclophosphamide and Total-Body Irradiation for Adult Patients with Acute Lymphoblastic Leukemia

We retrospectively evaluated the outcomes of 37 adult patients with acute lymphoblastic leukemia (ALL) undergoing allogeneic hematopoietic stem cell transplantation (allo-SCT) conditioned with medium-dose VP-16 (VP, 30 mg/kg), cyclophosphamide (CY, 120 mg/kg), and fractionated total-body irradiation (TBI, 12 Gy) (medium-dose VP/CY/TBI). The median age of the patients was 26 years. Thirteen patients underwent transplantation from HLA-matched related donors (MRD), 18 patients underwent transplantation from HLA-matched unrelated donors (MUD), and 6 patients underwent transplantation from HLA-mismatched donors (MMD). Thirty-two patients received bone marrow and 4 patients received peripheral blood stem cells. Ten patients were Philadelphia chromosome-positive (Ph(+)) and 35 patients were in complete remission (CR) at transplantation. All of the patients achieved engraftment, and grade 3 organ toxicity before engraftment occurred in 27 patients. Grade II-III acute graft-versus-host disease (GVHD) and chronic GVHD (cGVHD) occurred in 15 and 18 patients, respectively. No patient developed grade IV acute GVHD (aGVHD) or died of GVHD. At median follow-up of 35.1 months, 32 patients were alive and all Ph(+) patients were alive. Three patients died of relapse and 2 died of transplant-related mortality (TRM). The actuarial 3-year overall survival (OS) rate, relapse rate, and TRM rate were 89.2%, 8.1%, and 5.4%, respectively. Non-CR at transplantation, MRD, and no aGVHD were significant adverse prognostic factors for survival. Medium-dose VP/CY/TBI for adult ALL patients was associated with lower relapse rate and no increase in toxicity, resulting in better survival.

Delayed redistribution of CD27, CD40 and CD80 positive B cells and the impaired in vitro immunoglobulin production in patients with non-Hodgkin lymphoma after rituximab treatment as an adjuvant to autologous stem cell transplantation

Recent studies have indicated that patients who received rituximab as an adjuvant to stem cell transplantation (SCT) demonstrated an increased risk of developing severe hypogammaglobulinaemia, which was found to be a result of delayed recovery of CD27 positive memory B cells and impaired isotype expression. It appears that rituximab influences both the quantity and quality of B‐cell redistribution. Precisely how the B‐cell repertoire regenerates after anti‐CD20‐mediated transient B‐cell depletion in patients with non‐Hodgkin lymphoma (NHL) remains to be elucidated. This study performed a phenotypical analysis of B cells in 17 NHL patients who received rituximab as an adjuvant to autologous SCT. The median period after final administration of rituximab was 36 months (range, 12–43 months). Surface antigen expression of CD27, CD40 and CD80 in NHL patients was statistically significantly different from healthy controls (n = 14). Moreover, B cells from NHL patients showed significantly impaired IgG and IgA production upon engagement of surface immunoglobulin receptors in the presence of interleukin (IL)‐2, IL‐10 and CD40 ligand in comparison with samples from healthy controls. The delayed recovery of memory B cells with an abnormal cell marker expression and function demonstrates that naive B cells may fail to differentiate into plasma cells, resulting in hypogammaglobulinaemia after autologous SCT and rituximab therapy.

Persistent panhypogammaglobulinemia with selected loss of memory B cells and impaired isotype expression after rituximab therapy for post-transplant EBV-associated autoimmune hemolytic anemia

To the Editor: Miles and McGratten (1) reported persistent panhypogammaglobulinemia after CHOP-rituximab for HIV-related lymphoma. Prolonged hypogammaglobulinemiawith rituximab has also been seen in patients with post-transplant Epstein–Barr virus (EBV)-associated lymphoproliferative disorder (2, 3), as well B-cell lymphoma when rituximab is used as maintenance after stem cell transplantation (4, 5). We report a case of severe persistent panhypogammaglobulinemia after treatment of a posttransplant EBV-associated autoimmune hemolytic anemia (AIHA) with rituximab. The patient is a 32-yr-old Japanese woman with severe aplastic anemia who underwent non-myeloablative allogeneic peripheral blood stem cell transplantation from her HLA-matched sister in September 2000 (6). On days 90 and 150, chronic graft versus host disease occurred, but was alleviated with tacrolimus and predonisolone. In April 2002, 19 months after transplantation, the serum immunoglobulin M (IgM) level increased and immuno-electrophresis analysis revealed monoclonality of IgM. A remarkable proliferation of EBV-DNA also occurred. Tacrolimus and predonisolone were rapidly tapered off and the IgM level and EBV-DNA decreased. At that time, the IgG level was 757 mg/ dL (normal range 870–1700); low but stable after tapering of these immunosuppressive agents. In July 2002, severe AIHA occurred (7). Due to the existence of predictive factors of post-transplant lymphoproliferative disease (PTLD), conventional immunosuppressive therapy for AIHA was not considered. We gave rituximab (Chugai Pharmaceutical, Tokyo, Japan), 375 mg/m once a week for a total of four doses. The clinical and laboratory signs of hemolysis rapidly improved and the hemoglobin level began to improve 2 wk after the start of rituximab treatment. B cells in her peripheral blood rapidly disappeared, as expected. But all serum immunoglobulin levels kept on decreasing and remained extremely low (IgG < 200 mg/dL, IgA < 10 mg/dL and IgM < 10 mg/dL) unless intravenous immunoglobulin (IVIG) was administered for more than 2 yr. She developed repeated bacterial infections, such as pneumonia and sepsis, and was treated with IVIG and antibiotics. In February 2005, 30 months after last administration of rituximab, we found her blood B-cell number reverted to a normal range (8% of total lymphocytes, 180/lL), despite the still severe hypogammaglobulinemia. Flow cytometry analysis revealed that her B cells were composed with only CD27 negative naı̈ve B cells. CD27 positive memory B cells, both IgD-positive non-switched and IgDnegative switched populations, were hardly seen (Fig. 1A). In addition, we evaluated immunoglobulin isotype production by analyzing immunoglobulin transcripts with reverse transcriptase-polymerase chain reaction as described (8). Her peripheral blood mononuclear cells (PBMC) only expressed the transcriptions of IgM, IgG1, IgA1 and IgA2, but no IgG2, IgG3 and IgG4 (Fig. 1B). Rituximab is known to induce B-cell depletion for up to 6 months post-treatment (9). Moreover, in patients treated with high dose chemotherapy and maintenance rituximab, it takes longer (18– 24 months) for B-cell recovery (4). This delay of B-cell recovery is likely to contribute to hypogammaglobulinemia, especially when rituximab is used for treatment of EBV-PTLD (2, 3, 10). But this does not seem to be the case in our patient who Eur J Haematol 2005: 75: 527–529 doi:10.1111/j.0902-4441.2005.t01-1-EJH2327.x All rights reserved Copyright Blackwell Munksgaard 2005

Peripheral blood stem cell mobilization following CHOP plus rituximab therapy combined with G-CSF in patients with B-cell non-Hodgkin's lymphoma

Summary:We mobilized peripheral blood stem cells (PBSC) following CHOP plus rituximab (CHOP-R) therapy, and compared with the findings following CHOP therapy without rituximab. All patients were given G-CSF starting from day 11 after CHOP therapy. Patients in the CHOP-R group (n=8) were given rituximab on day 12. Target CD34+ cells number was collected in a single leukapheresis on day 14, from all the eight patients in the CHOP-R group. PBSC mobilization kinetics, CD34+ cells yield and colony-forming ability in the graft collection, toxicity during mobilization, and engraftment after transplantation of CHOP-R group were not significantly different from those in the CHOP group (n=8). In all patients given CHOP-R therapy, CD20+ cells and immunoglobulin heavy chain (IgH) rearrangement in the graft collection were undetectable by flow-cytometric analysis and Southern blot analysis, respectively, but with PCR analysis two of eight grafts were positive for IgH rearrangement. While further studies are needed to evaluate the efficacy of purging and the outcome of patients undergoing autologous transplantation, CHOP-R therapy can be safely and effectively used in the mobilization phase of PBSC collection, without excess clinical toxicity or deleterious effect on PBSC mobilization kinetics or engraftment time.

FCGR3A-158V/F polymorphism may correlate with the levels of immunoglobulin in patients with non-Hodgkin’s lymphoma after rituximab treatment as an adjuvant to autologous stem cell transplantation

Objectives:  Recent studies have indicated that patients who receive stem cell transplantation (SCT) and rituximab demonstrate an increased risk of developing hypogammaglobulinemia. Such hypogammaglobulinemia has been found to be due to delayed recovery of memory B cells with an abnormal cell marker expression and impaired immunoglobulin production in vitro. However, no predictive factors for the levels of immunoglobulin after autologous SCT and rituximab therapy have been reported. The aim of this study is to clarify the relationships between the FCGR3A‐158V/F genotype and the levels of serum immunoglobulin after SCT.

Efficacy of folinic acid in preventing oral mucositis in allogeneic hematopoietic stem cell transplant patients receiving MTX as prophylaxis for GVHD

As the safety of folinic acid administration and its efficacy for reducing the toxicity of MTX remain controversial, we assessed the effect of folinic acid administration after MTX treatment for GVHD prophylaxis on the incidence of oral mucositis and acute GVHD. We retrospectively analyzed data for 118 patients who had undergone allogeneic hematopoietic SCT and had received MTX for GVHD prophylaxis. Multivariate analysis showed that systemic folinic acid administration significantly reduced the incidence of severe oral mucositis (odds ratio (OR)=0.13, 95% confidence interval (CI) 0.04–0.73, P=0.014). There was also a tendency for a lower incidence of severe oral mucositis in patients who received folinic acid mouthwash (OR=0.39, 95%CI 0.15–1.00, P=0.051). No significant difference was observed in the incidence of acute GVHD between patients who received systemic folinic acid administration and those who did not (P=0.88). Systemic folinic acid administration and mouthwash appear to be useful for reducing the incidence of severe oral mucositis in patients who have received allogeneic hematopoietic SCT using MTX as GVHD prophylaxis.

DOCK2 regulates cell proliferation through Rac and ERK activation in B cell lymphoma

DOCK2; a member of the CDM protein family, regulates cell motility and cytokine production through the activation of Rac in mammalian hematopoietic cells and plays a pivotal role in the modulation of the immune system. Here we demonstrated the alternative function of DOCK2 in hematopoietic tumor cells, especially in terms of its association with the tumor progression. Immunostaining for DOCK2 in 20 cases of human B cell lymphoma tissue specimens including diffuse large B cell lymphoma and follicular lymphoma revealed the prominent expression of DOCK2 in all of the lymphoma cells. DOCK2-knockdown (KD) of the B cell lymphoma cell lines, Ramos and Raji, using the lentiviral shRNA system presented decreased cell proliferation compared to the control cells. Furthermore, the tumor formation of DOCK2-KD Ramos cell in nude mice was significantly abrogated. Western blotting analysis and pull-down assay using GST-PAK-RBD kimeric protein suggested the presence of DOCK2-Rac-ERK pathway regulating the cell proliferation of these lymphoma cells. This is the first report to clarify the prominent role of DOCK2 in hematopoietic malignancy.

Case report: clearance of hepatitis C virus after changing the HAART regimen in a patient infected with hepatitis C virus and the human immunodeficiency virus.

The effect of highly active antiretroviral therapy (HAART) on hepatitis C virus (HCV) infection remains uncertain. This report describes the case of a man with hemophilia with HIV–HCV coinfection with persistent disappearance of HCV RNA after changing the HAART regimen. He had been treated with zidovudine, lamivudine, and indinavir for initial HAART and the HIV RNA level had been undetectable for more than 8 years. He had suffered from chronic active hepatitis. The HAART regimen was changed to emtricitabine/tenofovir, atazanavir, and ritonavir because the patient preferred a once daily regimen. The HCV RNA level fell immediately and thereafter became undetectable by quantitative and qualitative assay at 5 and 7 months after the change of the HAART regimen, respectively. In contrast to other reported cases, he experienced neither increase of CD4+ T cells count nor ALT flare‐ups before HCV RNA clearance. The HCV RNA disappearance in this case may be due to the direct effect of HAART against HCV rather than restoration of cellular immunity to HCV. J. Med. Virol. 81:979–982, 2009.

Complete remission of splenic marginal zone lymphoma after an acute flare-up of hepatitis B in a hepatitis B virus carrier.

A 44-year-old female presented with asymptomatic leukocytosis and moderate splenomegaly. The diagnosis of splenic marginal zone lymphoma (SMZL) was made by a splenectomy. A virological examination revealed the patient to be a hepatitis B virus (HBV) carrier. The lymphocyte count in her peripheral blood decreased after splenectomy, but remained high for 2 years and bone marrow infiltration was obvious. Two years after the splenectomy, she was admitted for an acute flare-up of hepatitis B. The liver dysfunction improved without any medication and thereafter returned to the normal range within a few weeks. At the same time, the lymphocyte count in her peripheral blood rapidly decreased to normal levels. Atypical lymphocytes disappeared from the peripheral blood and bone marrow aspirates and biopsy specimen revealed complete remission of SMZL, including the disappearance of the clonal rearrangement of IgH-JH. There has been no recurrence of acute hepatitis and she has been in complete remission for SMZL for more than 6 years. The clinical course of this patient suggests that an immune response against HBV also affects the clearance of lymphoma cells. This is the first report that a complete remission was achieved in a patient with SMZL after a hepatitis B flare-up.