Successful allogeneic peripheral blood stem cell transplantation for an aggressive variant of T-cell large granular-lymphocyte leukemia: A case report.

The aggressive variant of large granular lymphocyte (LGL) leukemia is very rare and the prognosis of this disease is poor. A 47-year-old woman with progressive pancytopenia and severe liver damage visited our institute. Upon hospitalization, about 30% LGL was detected in her peripheral blood and bone marrow samples. Flow cytometry was conducted to analyze lymphocytes in the bone marrow, which revealed the presence of CD3 and T-cell receptor (TCR) α/ß and absence of CD4, CD8, CD16, CD56, CD22, CD79a, and terminal deoxynucleotidyl transferase (TdT). Southern blotting was performed, which revealed the presence of rearrangement of TCR-Cß1 and Jγ. We made a diagnosis of the aggressive variant of T-LGL leukemia, and performed myeloablative allogeneic peripheral stem cell transplantation (allo-HSCT) from an HLA-matched sibling for primary refractory disease of CHOP and hyper CVAD therapy. She is alive in remission with donor-derived T-LGL lymphocytosis in peripheral blood for 7 years after allo-HSCT. Overall, Allo-HSCT could be active against the aggressive variant of LGL leukemia and induce graft-versus-leukemia effect.

Clinical significance of cancer-related fatigue in multiple myeloma patients.

Cancer-related fatigue (CRF) is one of the adverse events in multiple myeloma (MM) patients treated with cytotoxic agents, proteasome inhibitors (PIs), and immunomodulatory drugs (IMiDs) such as bortezomib, lenalidomide, and thalidomide. The aims of our study were to prospectively analyze the clinical significance of CRF, and to evaluate the cumulative incidence of CRF and the survival rates of 16 MM patients who were treated with PIs and IMiDs. Reactivation of salivary human herpes virus (HHV)-6 and HHV-7 was analyzed using real-time quantitative polymerase chain reaction (qPCR). CRF was evaluated using a visual analog scale (VAS). Eleven newly diagnosed multiple myeloma (NDMM) and five relapsed or refractory MM patients were enrolled in this study. The cumulative incidence of CRF was 54.9%. The treatment types were not associated with the CRF incidence. The cumulative incidence of reactivation of HHV-6 and HHV-7 was 73.1% and 45.6%, respectively. However, the reactivation of HHV-6 and HHV-7 was not related to CRF. The overall survival (OS) and progression-free survival (PFS) in NDMM patients with CRF was significantly shorter than in those without CRF. In conclusion, CRF was one of the major symptoms in MM patients, and predicted shorter OS and PFS in NDMM patients.

Epstein-Barr Virus-associated Lymphoproliferative Disorder with Encephalitis Following Anti-thymocyte Globulin for Aplastic Anemia Resolved with Rituximab Therapy: A Case Report and Literature Review.

Epstein-Barr virus (EBV)-associated lymphoproliferative disorders (LPDs) sometimes occur following Anti-thymocyte globulin (ATG) administration for allogenic stem cell transplantation but are rare in aplastic anemia (AA) patients. A 55-year-old woman with AA following ATG developed refractory fever and was diagnosed with EBV-LPD. She was successfully treated with weekly rituximab monotherapy; however, she developed EBV encephalitis. She was admitted to the intensive care unit and finally recovered from unconsciousness. EBV-LPD should be considered after ATG for AA when symptoms appear. Because EBV-LPD following ATG for AA can rapidly progress, weekly monitoring of EBV-DNA and early intervention may be necessary.

Clinical significance of granule-containing myeloma cells in patients with newly diagnosed multiple myeloma.

The clinical features and prognostic significance of myeloma cells containing granules remain unclear. The purpose of this retrospective study was to investigate the clinical significance of granule-containing myeloma cells in patients with newly diagnosed multiple myeloma (NDMM). We retrospectively analyzed the records of 122 patients diagnosed with NDMM between January 2007 and December 2013. Granule-containing myeloma cells were defined as myeloma cells that exhibited three or more granules in their cytoplasm by May-Giemsa staining. The patients were classified into two groups, the granule-containing myeloma (GM) and nongranule-containing myeloma (non-GM) groups, depending on the proportion of myeloma cells that contained granules (cut-off value: 10%). There were 25 (20.5%) patients in the GM group. Patients in the GM group displayed significantly higher CD56 and CD49e expression than those in the non-GM group (t-test, P = 0.027 and 0.042). None of the patient characteristics differed significantly between the two groups. There was no significant difference in the chemotherapy profiles of the two groups, and the overall response rates of the two groups were similar. During the median follow-up period of 33.9 months, the overall survival (OS) in the GM group was similar to that in the non-GM group; 4-year OS of the GM and non-GM groups were 78.5% and 51.9%, respectively (P = 0.126). We concluded that cases of NDMM involving granule-containing myeloma cells are not infrequent. Moreover, CD56 and CD49e expression was significantly higher in the presence of myeloma cell populations, and the presence of granules did not affect survival.

Combination of a Selective HSP90α/ß Inhibitor and a RAS-RAF-MEK-ERK Signaling Pathway Inhibitor Triggers Synergistic Cytotoxicity in Multiple Myeloma Cells.

Heat shock protein (HSP)90 inhibitors have shown significant anti-tumor activities in preclinical settings in both solid and hematological tumors. We previously reported that the novel, orally available HSP90α/ß inhibitor TAS-116 shows significant anti-MM activities. In this study, we further examined the combination effect of TAS-116 with a RAS-RAF-MEK-ERK signaling pathway inhibitor in RAS- or BRAF-mutated MM cell lines. TAS-116 monotherapy significantly inhibited growth of RAS-mutated MM cell lines and was associated with decreased expression of downstream target proteins of the RAS-RAF-MEK-ERK signaling pathway. Moreover, TAS-116 showed synergistic growth inhibitory effects with the farnesyltransferase inhibitor tipifarnib, the BRAF inhibitor dabrafenib, and the MEK inhibitor selumetinib. Importantly, treatment with these inhibitors paradoxically enhanced p-C-Raf, p-MEK, and p-ERK activity, which was abrogated by TAS-116. TAS-116 also enhanced dabrafenib-induced MM cytotoxicity associated with mitochondrial damage-induced apoptosis, even in the BRAF-mutated U266 MM cell line. This enhanced apoptosis in RAS-mutated MM triggered by combination treatment was observed even in the presence of bone marrow stromal cells. Taken together, our results provide the rationale for novel combination treatment with HSP90α/ß inhibitor and RAS-RAF-MEK-ERK signaling pathway inhibitors to improve outcomes in patients with in RAS- or BRAF-mutated MM.

Selective and potent Akt inhibition triggers anti-myeloma activities and enhances fatal endoplasmic reticulum stress induced by proteasome inhibition.

The PI3K/Akt pathway plays a crucial role in the pathogenesis of multiple myeloma (MM) in the bone marrow (BM) milieu. However, efficacy of selective and potent Akt inhibition has not yet been fully elucidated. In this study, we, therefore, examined the biologic impact of selective and potent Akt inhibition by a novel allosteric inhibitor TAS-117. TAS-117 induced significant growth inhibition, associated with downregulation of phosphorylated Akt (p-Akt), selectively in MM cell lines with high baseline p-Akt. Cytotoxicity of TAS-117 was also observed in patient MM cells, but not in normal peripheral blood mononuclear cells. Importantly, TAS-117 induced significant cytotoxicity in MM cells even in the presence of BM stromal cells, associated with inhibition of IL6 secretion. Oral administration of TAS-117 significantly inhibited human MM cell growth in murine xenograft models. TAS-117 triggered apoptosis and autophagy, as well as induction of endoplasmic reticulum (ER) stress response with minimal expression of C/EBP homologous protein (CHOP), a fatal ER stress marker. Importantly, TAS-117 enhanced bortezomib-induced cytotoxicity, associated with increased CHOP and PARP cleavage and blockade of bortezomib-induced p-Akt, suggesting that TAS-117 augments bortezomib-induced ER stress and apoptotic signaling. Carfilzomib-induced cytotoxicity was similarly enhanced by TAS-117. Importantly, TAS-117 enhanced bortezomib-induced cytotoxicity in vivo, associated with prolonged host survival. Our results show that selective and potent Akt inhibition by TAS-117 triggers anti-MM activities in vitro and in vivo, as well as enhances cytotoxicity of proteasome inhibition, providing the preclinical framework for clinical evaluation of selective Akt inhibitors, alone and in combination with proteasome inhibitors in MM.

Dual inhibition of canonical and noncanonical NF-κB pathways demonstrates significant antitumor activities in multiple myeloma.

PURPOSE: NF-κB transcription factor plays a key role in the pathogenesis of multiple myeloma in the context of the bone marrow microenvironment. Both canonical and noncanonical pathways contribute to total NF-κB activity. Recent studies have shown a critical role for the noncanonical pathway: selective inhibitors of the canonical pathway present a limited activity, mutations of the noncanonical pathway are frequent, and bortezomib-induced cytotoxicity cannot be fully attributed to inhibition of canonical NF-κB activity. EXPERIMENTAL DESIGN: Multiple myeloma cell lines, primary patient cells, and the human multiple myeloma xenograft murine model were used to examine the biologic impact of dual inhibition of both canonical and noncanonical NF-κB pathways. RESULTS: We show that PBS-1086 induces potent cytotoxicity in multiple myeloma cells but not in peripheral blood mononuclear cells. PBS-1086 overcomes the proliferative and antiapoptotic effects of the bone marrow milieu, associated with inhibition of NF-κB activity. Moreover, PBS-1086 strongly enhances the cytotoxicity of bortezomib in bortezomib-resistant multiple myeloma cell lines and patient multiple myeloma cells. PBS-1086 also inhibits osteoclastogenesis through an inhibition of RANK ligand (RANKL)-induced NF-κB activation. Finally, in a xenograft model of human multiple myeloma in the bone marrow milieu, PBS-1086 shows significant in vivo anti-multiple myeloma activity and prolongs host survival, associated with apoptosis and inhibition of both NF-κB pathways in tumor cells. CONCLUSIONS: Our data show that PBS-1086 is a promising dual inhibitor of the canonical and noncanonical NF-κB pathways. Our preclinical study therefore provides the framework for clinical evaluation of PBS-1086 in combination with bortezomib for the treatment of multiple myeloma and related bone lesions.

Blockade of XBP1 splicing by inhibition of IRE1α is a promising therapeutic option in multiple myeloma.

Multiple myeloma (MM) cells are characterized by high protein synthesis resulting in chronic endoplasmic reticulum (ER) stress, which is adaptively managed by the unfolded protein response. Inositol-requiring enzyme 1α (IRE1α) is activated to splice X-box binding protein 1 (XBP1) mRNA, thereby increasing XBP1s protein, which in turn regulates genes responsible for protein folding and degradation during the unfolded protein response. In this study, we examined whether IRE1α-XBP1 pathway is a potential therapeutic target in MM using a small-molecule IRE1α endoribonuclease domain inhibitor MKC-3946. MKC-3946 triggered modest growth inhibition in MM cell lines, without toxicity in normal mononuclear cells. Importantly, it significantly enhanced cytotoxicity induced by bortezomib or 17-AAG, even in the presence of bone marrow stromal cells or exogenous IL-6. Both bortezomib and 17-AAG induced ER stress, evidenced by induction of XBP1s, which was blocked by MKC-3946. Apoptosis induced by these agents was enhanced by MKC-3946, associated with increased CHOP. Finally, MKC-3946 inhibited XBP1 splicing in a model of ER stress in vivo, associated with significant growth inhibition of MM cells. Taken together, our results demonstrate that blockade of XBP1 splicing by inhibition of IRE1α endoribonuclease domain is a potential therapeutic option in MM.

Rapid progression and unusual premortal diagnosis of mucormycosis in patients with hematologic malignancies: analysis of eight patients.

Mucormycosis is a rare but emerging group of life-threatening opportunistic mycoses. We described experience of eight patients who developed mucormycosis. These patients had developed hematologic malignancies, and none achieved complete remission. Six of the eight patients presented with neutropenia, five received corticosteroid, and four had concomitant hyperglycemia. The most frequent physical finding was fever, and five patients complained of facial pain, headache, or chest pain. Four patients presented with concomitant bacterial infection, pulmonary aspergillosis, or intestinal candidiasis. Premortal diagnosis of mucormycosis was made in only one patient. Postmortem biopsy or autopsy was the diagnostic tool for the other patients. Although patients who were treated with amphotericin B survived longer than those treated with micafungin or voriconazole, all patients died due to the progression of mucormycosis. Estimated median survival was 23 days. Premortal diagnosis was rarely achieved as biopsy of infected tissues was the only diagnostic tool, and four patients who revealed dual infection were diagnosed with aspergillosis or bacterial infections. In patients with a high risk of mucormycosis presenting with pain and uncontrollable fever, mucormycosis should be included in the differential diagnosis. High dosages of liposomal amphotericin B should be given and surgical debridement should be performed promptly in cases highly suggestive of mucormycosis.

Two cases of mediastinal gray zone lymphoma.

Mediastinal gray zone lymphoma (MGZL) represents a range of tumors possessing characteristics of both nodular sclerosis classical Hodgkin lymphoma (NSHL) and mediastinal large B-cell lymphoma (MLBCL). Here we report two patients with MGZL. Patient 1 was a 30-year-old woman and patient 2 was a 22-year-old man. Both patients had a mediastinal mass, were initially diagnosed with NSHL and exhibited resistance to first-line chemotherapy. Re-biopsy of the relapsed tumors or the residual lesion was performed and based on the findings the tumors were diagnosed as MGZL. In patient 1, the morphological features of the tumor resembled those of NSHL, but the immunophenotypic features indicated MLBCL. In patient 2, the tumor was a composite lymphoma with both NSHL and MLBCL components. Both the patients received high-dose chemotherapy followed by autologous peripheral-blood stem-cell transplantation. Although there is an overlap in the biological and morphological features between NSHL and MLBCL, the therapeutic approaches to NSHL and MLBCL are quite different. The development of effective therapies for MGZL is therefore extremely critical.

[High-dose methotrexate followed by whole-brain irradiation for primary central nervous system lymphoma patients--a retrospective study in a single institute].

This study analyzed retrospectively the clinical efficacy of combined therapy consisting of high-dose methotrexate (MTX), administered at a dose of 4 g/m2 every 2 weeks (maximum of 4 courses), followed by whole-brain irradiation for newly diagnosed primary central nervous system lymphoma (PCNSL) patients. Fifteen patients (median age: 59 years old; range: 26-79) were diagnosed by histological examinations or imaging techniques in our hospital. Of 15 patients, 12 (6: complete response; 6: partial response) achieved objective response, and the response rate was 80% (95% CI, 51.9-95.7%). The median follow-up time was 20 (range: 3-81) months, and the 3-year survival rate was 76%. The overall survival time was 71 months (95% CI, 23. 7-118.3 months), and the progression free survival was 15 months (95% CI, 0-43.8 months). The major toxicity (grade>or=3) of high-dose MTX included cytopenia (20%), acute respiratory distress syndrome (6.7%), and liver damage (6.7%). No patient evidenced complicated leukoencephalopathy in the follow-up time. The combined therapy of high-dose MTX followed by whole-brain irradiation showed a substantial antitumor efficacy in PCNSL patients. Prospective studies are required to determine the suitable treatment schedule for MTX and irradiation.

Fractionated administration of gemtuzumab ozogamicin for refractory acute myeloid leukemia.

It is difficult to decide an appropriate treatment strategy for elderly leukemia patients with other complications. We encountered 2 cases of refractory acute myeloid leukemia and safely treated the patients with fractionated administration of gemtuzumab ozogamicin (GO). Standard induction therapies were not effective for these patients. Moreover, they suffered from complications due to which their treatment options were restricted. Fractionated administration of GO (GO 3 mg/m(2) on days 1, 3 and 5) was accomplished safely and alleviated the patients' conditions. After treatment, these patients were followed by outpatient basis. We consider that this is an impressive treatment because fractionated administration of GO is potentially less toxic. Further, it will be helpful to maintain or improve the QOL of patients who are unable to receive intensive chemotherapy. These cases were significant because fractionated GO treatment is potentially less toxic and it will be helpful to maintain or improve the QOL of patients who can not receive intensive chemotherapy.

Retrospective comparison of mobilization methods for autologous stem cell transplantation in multiple myeloma.

The combination of cyclophosphamide and granulocyte-colony stimulating factor (G-CSF) has widely been used to mobilize hematopoietic stem cells (HSCs) for autologous stem cell transplantation (ASCT) for multiple myeloma (MM). Recently, however, alternative approaches such as G-CSF alone or etoposide followed by G-CSF have been investigated. We, therefore, retrospectively analyzed the effects of these mobilization methods on collection yield and disease outcome in ASCT for MM. We reviewed 146 MM patients from whom we intended to collect stem cells. For mobilization, 67, 58, and 21 patients received cyclophosphamide and G-CSF, etoposide and G-CSF, and G-CSF alone (including nonmyelosuppressive chemotherapy followed by G-CSF), respectively. Among them, 136 achieved the target number of HSCs (at least 2 x 10(6)/kg). Lower creatinine and higher albumin levels at diagnosis were significantly associated with successful yield. A lower number of infused HSCs, use of the etoposide for mobilization and high ISS were associated with delayed hematopoietic recovery. The mobilization methods did not significantly affect either the successful collection of more than 2 x 10(6) CD34-positive cells/kg or PFS after ASCT. G-CSF alone was sufficient for stem cell mobilization for a single ASCT. The optimal approach to collect HSCs in MM remains to be elucidated.

Gemtuzumab ozogamicin (GO) in relapsed/refractory patients with acute myeloid leukemia.

OBJECTIVE: Gemtuzumab ozogamicin (GO) is a humanized anti-CD33 antibody, linked to calicheamicin, which has been approved in Japan recently. We conducted to evaluate the efficacy and toxicity of GO in our patients with relapsed or refractory AML retrospectively. PATIENTS AND METHODS: Data were collected between March 1, 2000, and March 1, 2006, on 10 patients with relapsed or refractory AML(excluding FAB: M3). Scheduled treatment was two doses of GO monotherapy, 14-28 days apart. RESULTS: Of the 10 assessable patients, two patients achieved CR. CR duration of one patient lasted for 52 months with post-remission treatment. Grade 4 neutropenia occurred in 9 patients, and the incidence of grade 3 or 4 thrombocytopenia was 100%, with no severe bleeding events. Two patients developed infusion-related adverse events that included grade 3 allergic reaction with shock status. Liver damage (grade 3 or 4) were observed in 40% of patients after GO treatment. No patient developed hepatic veno-occlusive disease including 2 patients who underwent HSCT. CONCLUSION: GO is a valuable new treatment option for relapsed or refractory AML patients, however, the benefit from single agent appears insufficient. On going clinical trials including combination with other antileukemic agents might better define the role of GO.

Elevated serum levels of human matrix metalloproteinase-9 (MMP-9) during the induction of peripheral blood stem cell mobilization by granulocyte colony-stimulating factor (G-CSF).

To investigate the role of matrix metalloproteinases (MMPs) in the mobilization of peripheral blood stem cells stimulated by granulocyte colony-stimulating factor (G-CSF), we analyzed MMP serum levels in 11 healthy donors and 9 patients who had hematological malignancies or germ cell tumors. A dose of 5-10 microg/kg per day of G-CSF (lenograstim) was administered for 4-8 days to each subject. The serum levels of MMP-2, and MMP-9; interleukin-3, -6, -8, and -10; stem cell factor; interferon-gamma; and tumor necrosis factor-alpha were measured both before and during G-CSF administration. MMP-9 was found to be increased in both the cancer patients and the healthy donor group. In contrast, the levels of each of the other factors tested were unchanged. No significant positive correlation was observed between the MMP-9 levels and the number of CD34+ cells. Hence, we found no significant role for MMPs during the mobilization of peripheral blood stem cells stimulated by G-CSF.

Long-term follow-up of autologous stem cell transplantation for patients with aggressive non-Hodgkin lymphoma who had bone marrow involvement at initial diagnosis in the pre-rituximab era.

BACKGROUND: High-dose therapy (HDT) followed by autologous stem cell transplantation (ASCT) is an important treatment option for selected patients with aggressive non-Hodgkin lymphoma; however, the effectiveness of HDT for patients with bone marrow (BM) involvement of lymphoma cells is not well defined. PATIENTS AND METHODS: Between February 1991 and December 2001, 57 patients with aggressive non-Hodgkin lymphoma were treated with HDT and ASCT. Thirteen of 57 patients who had BM infiltration at initial diagnosis were analyzed. RESULTS: Median follow-up was 11.5 years. Eleven of 13 patients (85%) exhibited complete remission after HDT. The overall survival (OS) at 10 years was 49%, and the median survival time was 74.3 months. Meanwhile, the probability of OS at 10 years for 44 patients who did not have BM disease was 60%. There was no significant difference in OS (P=0.895) between patients with or without BM disease at initial diagnosis. CONCLUSION: High-dose therapy treatment followed by ASCT might save some groups of patients with lymphoma regardless of BM involvement at initial diagnosis.

Pseudo-Gaucher cell proliferation associated with myelodysplastic syndrome.

We report an extremely rare case of pseudo-Gaucher cell proliferation with myelodysplastic syndrome (MDS). A 77-year old Japanese man was referred to our hospital with splenomegaly and thrombocytopenia, and subsequent bone marrow aspiration revealed infiltrates of foamy vacuolated macrophages without any evidence of other morphologic abnormalities. A karyotype analysis showed the presence of 46,XY,del(20)(q11) in 20 of 20 examined bone marrow cells. We performed a splenectomy, and the resulting pathologic findings revealed massive infiltration of foamy vacuolated macrophages, which were morphologically compatible with Gaucher cells. The activities of beta-glucosidase and acid sphingomyelinase were within normal ranges; therefore, the foamy vacuolated macrophages were considered pseudo-Gaucher cells. A diagnosis of MDS, subclassified as refractory anemia, was then made according to World Health Organization classification guidelines. Pseudo-Gaucher cell proliferation and infiltration might therefore be observed in other patients presenting with MDS.

Purification and characterization of C-terminal truncated forms of histone H2A in monocytic THP-1 cells.

Histones are key components of chromatin. We investigated histone H2A-immunoreactive proteins in acute monocytic leukemia THP-1 cells using three polyclonal antibodies raised against peptides corresponding to distinct regions of H2A. Two unknown immunoreactive proteins (9- and 12-kDa proteins), H2A (14kDa) and ubiquitinated H2A (23kDa) were found in the cell lysates prepared by immediate direct addition of SDS-PAGE sample buffer to the cells as well as in the nuclear and chromatin fractions. However, they were not found in the cytoplasmic fraction. The unknown proteins were successfully purified by immunoaffinity chromatography from the cell nucleus extract and identified as 9-kDa H2A(1-87) and 12-kDa H2A(1-114), suggesting that both were produced by limited proteolysis of intact H2A(1-129). The truncated forms of H2A probably persisted as chromatin constituents, since the stability of H2A(1-87) in the chromatin fraction was sensitive to treatment with micrococcal nuclease, and H2A(1-114) was solubilized with lower ionic strength from the chromatin fraction obtained by micrococcal nuclease treatment. Truncated H2A proteins in THP-1 cells were transiently increased in amount by short-term treatment with phorbol 12-myristate 13-acetate or all-trans-retinoic acid, both of which induce macrophage-like differentiation. Furthermore, these increases were suppressed by preceding treatment with carbobenzoxy-l-leucyl-l-leucyl-l-leucinal (MG132) but not with carbobenzoxy-l-isoleucyl-gamma-t-butyl-l-glutamyl-l-alanyl-l-leucinal (PSI), both of which are generally known as proteasome inhibitors. Our results suggest that histone H2A is cleaved at least at two sites by protease(s) that remain obscure, and might affect chromatins in the early stage of THP-1 cell differentiation.