We retrospectively evaluated long-term outcomes of high dose chemotherapy followed by autologous stem cell transplant (HDC/ASCT) in patients with diffuse large B-cell lymphoma (DLBCL). Between 2004 and 2020, 46 DLBCL patients received HDC/ASCT in our institution, including 12 patients (26.1%), who received as an upfront setting (UFS). At a median follow-up time of 69 months (range, 2-169 months), the 5-year progression-free survival (PFS) rates were 82.5% (95%CI, 46.1-95.3%) in the UFS, and 57.8% (95%CI, 38.1-73.2%) in the relapsed or refractory (R/R) patients (n=34), respectively. The 5-year PFS rates were 62.3% (95%CI, 34.0-81.3%) in primary resistant (n=13) or early relapsing (within 1 year from the initial diagnosis) patients (n=4), and 53.3% (95%CI, 25.9-74.6%) in those relapsing >1 year after the initial diagnosis (n=17), with no statistically significant difference (p=0.498). In R/R patients, multivariate analysis showed that the remission status before HDC/ASCT was an independent poor prognostic factor for progression-free survival (hazard ratio [HR], 17.0; 95%CI, 3.35-86.6; p=0.000630) and high-risk category in the international prognostic index for OS (HR, 9.39; 95%CI, 1.71-51.6; p=0.0100). The incidence of non-relapse mortality by 5 years, and 10 years were 12.2%, and 15.2%, respectively. Eleven patients (23.9%) developed second malignancies, which was the most frequent late complication after HDC/ASCT, with 5-year, and 10-year cumulative incidence of 16.9%, 22.5%, respectively. In conclusion, HDC/ASCT is effective for chemo-sensitive R/R DLBCL regardless of the timing and lines of therapy. However, careful observation is required, considering the long-term complications such as secondary malignancies.

We previously reported that the antipsychotic drug chlorpromazine (CPZ), which inhibits the formation of clathrin-coated vesicles (CCVs) essential for endocytosis and intracellular transport of receptor tyrosine kinase (RTK), inhibits the growth/survival of acute myeloid leukemia cells with mutated RTK (KIT D816V or FLT3-ITD) by perturbing the intracellular localization of these molecules. Here, we examined whether these findings are applicable to epidermal growth factor receptor (EGFR). CPZ dose-dependently inhibited the growth/survival of the non-small cell lung cancer (NSCLC) cell line, PC9 harboring an EGFR-activating (EGFR exon 19 deletion). In addition, CPZ not only suppressed the growth/survival of gefitinib (GEF)-resistant PC9ZD cells harboring T790 M, but also restored their sensitivities to GEF. Furthermore, CPZ overcame GEF resistance caused by Met amplification in HCC827GR cells. As for the mechanism of CPZ-induced growth suppression, we found that although CPZ hardly influenced the phosphorylation of EGFR, it effectively reduced the phosphorylation of ERK and AKT. When utilized in combination with trametinib (a MEK inhibitor), dabrafenib (an RAF inhibitor), and everolimus (an mTOR inhibitor), CPZ suppressed the growth of PC9ZD cells cooperatively with everolimus but not with trametinib or dabrafenib. Immunofluorescent staining revealed that EGFR shows a perinuclear pattern and was intensely colocalized with the late endosome marker, Rab11. However, after CPZ treatment, EGFR was unevenly distributed in the cells, and colocalization with the early endosome marker Rab5 and EEA1 became more apparent, indicating that CPZ disrupted the intracellular transport of EGFR. These results suggest that CPZ has therapeutic potential for NSCLC with mutated EGFR by a novel mechanism different from conventional TKIs alone or in combination with other agents.

C-type lectin-like receptor 2 (CLEC-2) expressed on megakaryocytes plays important roles in megakaryopoiesis. We found that CLEC-2 was expressed in about 20% of phenotypical long-term hematopoietic stem cells (LT-HSCs), which expressed lower levels of HSC-specific genes and produced larger amounts of megakaryocyte-related molecules than CLEC-2low LT-HSCs. Although CLEC-2high LT-HSCs had immature clonogenic activity, cultured CLEC-2high LT-HSCs preferentially differentiated into megakaryocytes. CLEC-2high HSCs yielded 6.8 times more megakaryocyte progenitors (MkPs) and 6.0 times more platelets 2 weeks and 1 week after transplantation compared with CLEC-2low LT-HSCs. However, platelet yield from CLEC-2high HSCs gradually declined with the loss of MkPs, while CLEC-2low HSCs self-renewed long-term, indicating that CLEC-2high LT-HSCs mainly contribute to early megakaryopoiesis. Treatment with pI:C and LPS increased the proportion of CLEC-2high LT-HSCs within LT-HSCs. Almost all CLEC-2low LT-HSCs were in the G0 phase and barely responded to pI:C. In contrast, 54% of CLEC-2high LT-HSCs were in G0, and pI:C treatment obliged CLEC-2high LT-HSCs to enter the cell cycle and differentiate into megakaryocytes, indicating that CLEC-2high LT-HSCs are primed for cell cycle entry and rapidly yield platelets in response to inflammatory stress. In conclusion, CLEC-2high LT-HSCs appear to act as a reserve for emergent platelet production under stress conditions.

We previously examined the utility of rituximab-bendamustine (RB) in patients with follicular lymphoma (FL) exhibiting less than optimal responses to 2 cycles of the R-CHOP chemotherapy regimen. The aim of this study was to identify molecular biomarkers that can predict prognosis in RB-treated patients in the context of the prospective cohort. We first analyzed the mutational status of 410 genes in diagnostic tumor specimens by target capture and Sanger sequencing. CREBBP, KMT2D, MEF2B, BCL2, EZH2, and CARD11 were recurrently mutated as reported before, however none was predictive for progression-free survival (PFS) in the RB-treated patients (n = 34). A gene expression analysis by nCounter including 800 genes associated with carcinogenesis and/or the immune response showed that expression levels of CD8+ T-cell markers and half of the genes regulating Th1 and Th2 responses were significantly lower in progression of disease within the 24-mo (POD24) group (n = 8) than in the no POD24 group (n = 31). Collectively, we selected 10 genes (TBX21, CXCR3, CCR4, CD8A, CD8B, GZMM, FLT3LG, CD3E, EOMES, GZMK), and generated an immune infiltration score (IIS) for predicting PFS using principal component analysis, which dichotomized the RB-treated patients into immune IIShigh (n = 19) and IISlow (n = 20) groups. The 3-y PFS rate was significantly lower in the IISlow group than in the IIShigh group (50.0% [95% CI: 27.1-69.2%] vs. 84.2% [95% CI: 58.7-94.6%], P = .0237). Furthermore, the IIS was correlates with absolute lymphocyte counts at diagnosis (r = 0.460, P = .00355). These results suggest that the T-cell-associated immune markers could be useful to predict prognosis in RB-treated FL patients. (UMIN:000 013 795, jRCT:051 180 181).

We recently treated a chronic myeloid leukemia (CML) patient with liver and renal dysfunction, who was undergoing hemodialysis (HD). He was treated with 50 mg dasatinib (DAS) once daily just before HD. The maximum plasma concentration of DAS was 227 ng/mL on a non-HD day and 46.9 ng/mL on a HD day. He was subsequently treated with 200 mg bosutinib (BOS) once daily. The plasma concentration of BOS changed from 74.5 ng/mL before HD to 58.8 ng/mL after HD. Our results indicate that close monitoring of the plasma tyrosine kinase inhibitor concentrations should be considered in CML patients with organ impairment.

Despite duodenal-type follicular lymphoma (DTFL) being morphologically, immunophenotypically and genetically indistinguishable from nodal FL (nFL), this entity typically shows a significantly better prognosis. Here, we analysed the tumour immune microenvironments of diagnostic specimens from patients with DTFL (n = 30), limited-stage FL (LSFL; n = 19) and advanced-stage FL (ASFL; n = 31). The mean number of CD8+ tumour-infiltrating lymphocytes (TILs) in the neoplastic follicles was higher in DTFL (1,827/mm2 ) than in LSFL (1,150/mm2 ) and ASFL (1,188/mm2 ) (P = 0·002, P = 0·002, respectively). In addition, CD8+ PD1-  T cells with non-exhausting phenotype were more abundant in the peripheral blood (PB) of DTFL than in LSFL and ASFL, indicating that DTFL may exhibit a better and longer-lasting T cell-mediated immune response. Moreover, whereas FOXP3+ CTLA-4+ effector regulatory T cells (eTregs) were rarely observed in the neoplastic follicles of DTFL (mean: 12/mm2 ), they were more abundant in LSFL (78/mm2 ) and ASFL (109/mm2 ) (P = 2·80 × 10-5 , P = 4·74 × 10-8 , respectively), and the numbers of eTregs correlated inversely with those of CD8+ TILs (r = -0267; P = 0·018). Furthermore, DTFL showed significantly fewer circulating FOXP3hi CD45RA- CD25hi eTregs (0·146%) than ASFL (0·497%) and healthy controls (0·639%) (P = 0·0003, P = 6·79 × 10-7 , respectively). These results suggest that the augmented anti-tumour immune reactions may contribute to a better prognosis on DTFL.

Patients with mycosis fungoides (MF), the most common subtype of primary cutaneous T-cell lymphoma, have an increased risk of developing secondary malignancies. We herein report two rare cases of MF concurring with diffuse large B cell lymphoma (B lymphoid lineage) and acute myeloid leukemia (myeloid lineage) in two otherwise healthy elderly patients. Potential etiologic factors, including the impact of the therapy-associated inflammatory response on the development of secondary tumors in patients with MF, are discussed. Further clinical, experimental and genetic studies are needed to elucidate possible physiopathogenic associations among the three concurrent malignancies occurring in the cases presented here.

Myelodysplastic syndrome (MDS) is a heterogeneous group of clonal disorders of hematopoietic stem cells, characterized by dysplastic hematopoiesis and dysregulated immune system resulting in various clinical conditions. Paraneoplastic inflammatory syndromes, which are well known to be associated with MDS, show response to immune-modulated therapy and often disappear during the course of hematologic management. Azacitidine (5-Aza) was shown to prolong survival of high-risk MDS patients, however, the effects of 5-Aza on paraneoplastic inflammation in MDS have yet to be elucidated. 5-Aza was administered to a 60-year-old man with MDS accompanying Sweet's syndrome at a dose of 75 mg/m2/daily subcutaneously for 7 days every 28 days. 5-Aza was not only effective in controlling systemic symptoms caused by paraneoplastic inflammation, but hematologic improvements were also observed after four cycles of the 5-Aza treatment. Immune profiling in peripheral blood before and after 5-Aza treatment revealed that the effector and naive regulatory T cells in lymphocytes drastically increased after the 5-Aza treatment, i.e., 5-Aza might induce a shift in lymphocytic populations toward immunosuppression in this patient. Our results raised the immune-mediated effect of 5-Aza on both dysplastic hematopoiesis and paraneoplastic inflammation in myelodyplastic syndromes.

急性骨髄性白血病に対する同種骨髄移植後に一旦造血機能の回復を認めたものの汎血球減少を来し、二次性生着不全と判断してドナーリンパ球輸注(donor lymphocyte infusion,DLI)を施行したところ著効を示した症例を経験したので報告する。症例は64歳女性。急性骨髄性白血病の部分寛解に対して非血縁者間同種骨髄移植を施行した。速やかに生着し、complete remission with incomplete blood count recovery(CRi)に到達したが、day 110から汎血球減少症が出現した。血液検査および骨髄穿刺では、再発や血球貪食症候群は認めないものの、キメリズム解析でドナー比率が経時的に低下したため、二次性生着不全と診断した。Granulocyte-colony stimulating factor(G-CSF)の投与と輸血にて経過観察したが、造血の改善は認めなかった。患者が再移植を希望しなかったため、DLIを施行した(CD3陽性細胞:1.0×107/kg、単回投与)ところ、有害事象を認めることなく、血球は速やかに改善した。(著者抄録)

Myeloproliferative neoplasms (MPNs), including polycythemia vera and essential thrombocythemia, are frequently associated with thrombotic complications. Prevention of thrombotic events is thus a primary aim of the current treatment for these disorders. Although it is known that microparticles (MPs), which are small vesicles released from cell membranes and circulate in the blood, directly contribute to thrombosis via their procoagulant activity, potential associations between plasma levels of MPs and the risk of thrombotic events in MPNs have not been reported. In the present study, we characterized plasma levels of MPs and assessed their potential association with the occurrence of thrombotic events in 59 patients with MPNs. Plasma levels of procoagulant MPs expressing tissue factor (TF+ MPs) were significantly higher in patients suffering thrombotic events than in patients without such events (median/mu l plasma: 33.8 vs 47.2, p = 0.02). Among patients who developed thrombotic events, irrespective of patients' blood counts, TF+ MP were significantly higher in patients without cytoreductive therapy than in those receiving cytoreductive therapy (101.2 vs. 42.5, p < 0.001). These results suggest that elevated levels of TF+ MP may be considered as a novel surrogate marker for thrombotic events in MPN patients. Further studies are needed to clarify the mechanism involved.

Here we report a case of secondary graft failure that was effectively treated with donor lymphocyte infusion (DLI). A 64-year-old female patient with acute myeloid leukemia obtained partial remission with azacitidine therapy and subsequently underwent unrelated allogeneic bone marrow transplantation (BMT). After confirming successful engraftment and achieving complete remission with incomplete blood count recovery, she was subsequently followed up at an outpatient clinic. A routine test performed by day 110 after BMT revealed the presence of pancytopenia. A bone marrow aspirate did not reveal any evidence of disease relapse or hemophagocytic syndrome but demonstrated hematopoietic insufficiency. Donor chimerism also declined over time; thus, the patient was diagnosed with secondary graft failure. Supportive treatment, including granulocyte-colony stimulating factor and blood transfusion, failed to improve the blood parameters. Because the patient refused a second BMT, we performed DLI on day 147 after BMT (CD3+ cells: 1.0×107/kg, single dose). Consequently, the blood cell count improved promptly and dramatically without adverse events. Following this, we discussed the case and analyzed the related literature.

A 68-year-old man was referred to our hospital due to a high fever and pancytopenia. Neither tumors nor infectious lesions were detected. Hemophagocytosis was observed on the bone marrow (BM) smear, although without abnormal cells. Prednisolone therapy was ineffective for the patient's high fever. Later on, we obtained the results of a BM biopsy indicating the presence of infiltration of atypical Reed-Sternberg cells, leading to a diagnosis of HIV-negative primary bone marrow Hodgkin lymphoma (PBMHL). However, the patient died of multiple organ failure before receiving chemotherapy. As the clinical course of PBMHL is rapid, physicians must keep in mind its possibility in similar cases.

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired stem cell disorder characterized by intravascular hemolysis and thrombosis. The most serious complication is thrombosis, the risk of which is augmented by the hyper-coagulable state that occurs during pregnancy despite this risk, however, young female PNH patients often desire to have a baby. We recently experienced two successful deliveries in PNH patients, who were treated with anticoagulant therapy during their pregnancies. Meanwhile, given the potential benefit of eculizumab (Soliris), a humanized monoclonal antibody against C5, in reducing thrombosis and hemolysis, it represents a promising therapeutic option for the treatment of pregnant PNH patients in combination with, or in replacement of, anticoagulant therapy. © The Japanese Society of Hematology 2013.