[Safety and efficacy of 7-day administration of azacitidine and venetoclax combination therapy (7+7 therapy)].

Acute myelogenous leukemia (AML) has a poor prognosis in patients who are ineligible for intensive chemotherapy. The combination of azacitidine and venetoclax has been shown to have high overall efficiency and remission rates, even in patients ineligible for aggressive chemotherapy. However, myelosuppression is often prolonged after treatment, and infection can also occur. Severe myelosuppression is often addressed by dose titration, but specific dose titration methods have not been clarified. We used the standard induction therapy with azacitidine plus venetoclax, and if blasts decreased to 20% or less, switched to 7+7 therapy to shorten venetoclax to 7 days starting from the next cycle. In the 19 patients we treated (median age 80 years), response rate above MLFS was 100%, CR 57.9%, CRc (CR+CRi) 78.8%, median OS 693 days, median PFS 458 days, and median OS was not reached in previously untreated patients. This indicates that 7+7 is a highly effective and well-tolerated treatment.

Clinicopathologic Characteristics and A20 Mutation in Primary Thyroid Lymphoma.

BACKGROUND: Primary thyroid lymphoma (PTL) is a rare disease frequently arising against a background of autoimmune thyroiditis. It has recently been reported that the inactivation of the NF-κB negative regulator A20 by deletion and/or mutation could be involved in the pathogenesis of subsets of B-cell lymphomas. This study investigated the clinicopathologic characteristics and A20 mutation in patients with PTL. METHODS: We analyzed the characteristics of 45 PTL patients (14 men and 31 women), with a median age of 71 (range, 35-90) years. A20 mutations were analyzed in DNA extracted from 20 samples consisting of 19 tumor tissue samples and 1 sample from Hashimoto's thyroiditis. RESULTS: Thirty-five patients (82%) had a history of Hashimoto's thyroiditis, and 29 (64%) had diffuse large B-cell lymphoma (DLBCL) and presented with larger tumors including bulky mass, elevated soluble interleukin-2 receptor levels, and a longer history of Hashimoto's thyroiditis than that of patients with mucosa-associated lymphoid tissue (MALT) lymphoma (n=16). A20 mutations were identified in 3 of 19 PTL patients (16%), in 2 of the 10 (20%) with DLBCL and in 1 of the 9 (11%) with MALT lymphoma. Interestingly, all patients with A20 mutations had Hashimoto's thyroiditis. Furthermore, they had a common missense variant in exon 3 (rs2230926 380T>G; F127C), which reduces the ability of A20 to inhibit NF-κB signaling. CONCLUSION: Our study suggests that the histological features of PTL affect clinical outcomes and that A20 mutations are related to PTL pathogenesis in some patients with Hashimoto's thyroiditis.

Copper deficiency-induced pancytopenia after taking an excessive amount of zinc formulation during maintenance hemodialysis.

Erythropoiesis-stimulating agent (ESA) has been recognized as an effective way in the treatment of anemia due to chronic kidney disease, but we sometimes see intractable hemodialysis (HD) patients. The causes of ESA-resistant anemia in HD patients include deficiency of trace elements. We report the case of an 89-year-old male who developed pancytopenia after taking an excessive amount of zinc formulation for ESA-resistant anemia during maintenance dialysis. He was prescribed zinc acetate hydrate formulation about 6 months before his presentation. He was found to have pancytopenia 1 month before his presentation, at which point he was introduced to our hospital. We suspected a copper deficiency at the first visit and stopped zinc and added copper, and his condition subsequently improved without being handicapped. Zinc antagonizes copper, so we must take care to diagnose patients ingesting zinc supplements.

Effects of the chelating agent DTPA on naturally accumulating metals in the body.

Diethylenetriaminepentaacetate (DTPA) is the most widely used chelating agent for Pu and Am. Volunteers were assigned to receive intravenous injections or aerosol inhalations of 1 g of DTPA on days 1-4; volunteers received once daily injections of CaDTPA or ZnDTPA, CaDTPA inhalation as an aerosol, or CaDTPA injection on day 1 and ZnDTPA on days 2-4. CaDTPA injection or inhalation increased the excretion rates of Zn in urine with concomitantly reduced levels of serum Zn. Injection of CaDTPA reduced activities of serum alkaline phosphatase (AP) in parallel with the kinetics of Zn, whereas CaDTPA and ZnDTPA injection reduced activities of lactate dehydrogenase (LDH), and reduced activities of creatinine kinase (CK) were observed upon CaDTPA injection and its inhalation. Intravenous administration of CaDTPA and ZnDTPA enhanced excretion rates of Mn in urine, whereas transient reduction of Mn levels in serum was detected only via CaDTPA injection. Both CaDTPA and ZnDTPA transiently reduced levels of Mg in serum without affecting the excretion rates. On the other hand, both DTPAs increased excretion rates of toxic metals such as Pb and Cd, and CaDTPA also increased the rates of Hg. These results suggest that DTPA, and especially CaDTPA, removes essential metals and that the activities of these metalloenzymes are good indicators for the imbalance of essential metals during the DTPA administration. Our results also show that CaDTPA injection is more potent for removing these metals than ZnDTPA and inhalation of CaDTPA, and DTPA may be useful for the treatment of acute heavy metal poisoning with Pb, Cd, or Hg.

SLAMF3-Mediated Signaling via ERK Pathway Activation Promotes Aggressive Phenotypic Behaviors in Multiple Myeloma.

The signaling lymphocytic activation molecule family 3 (SLAMF3) is a member of the immunoglobulin superfamily expressed on T, B, and natural killer cells and modulates the activation and cytotoxicity of these cells. SLAMF3 is also expressed on plasma cells from patients with multiple myeloma (MM), although its role in MM pathogenesis remains unclear. This study found that SLAMF3 is highly and constitutively expressed on MM cells regardless of disease stage and that SLAMF3 knockdown/knockout suppresses proliferative potential and increases drug-induced apoptosis with decreased levels of phosphorylated ERK protein in MM cells. SLAMF3-overexpressing MM cells promote aggressive myeloma behavior in comparison with cytoplasmic domain-truncated SLAMF3 (ΔSLAMF3) cells. SLAMF3 interacts directly with adaptor proteins SH2 domain-containing phosphatase 2 (SHP2) and growth factor receptor bound 2 (GRB2), which also interact with each other. SLAMF3 knockdown, knockout, ΔSLAMF3, and SHP2 inhibitor-treated MM cells decreased phosphorylated ERK protein levels. Finally, serum soluble SLAMF3 (sSLAMF3) levels were markedly increased in advanced MM. Patients with high levels of sSLAMF3 progressed to the advanced stage significantly more often and had shorter progression-free survival times than those with low levels. This study revealed that SLAMF3 molecules consistently expressed on MM cells transmit MAPK/ERK signals mediated via the complex of SHP2 and GRB2 by self-ligand interaction between MM cells and induce a high malignant potential in MM. Furthermore, high levels of serum sSLAMF3 may reflect MM disease progression and be a useful prognostic factor. IMPLICATIONS: SLAMF3 may be a new therapeutic target for immunotherapy and novel agents such as small-molecule inhibitors.

Atypical chronic myeloid leukemia achieving good response with azacitidine.

The prognosis of atypical chronic myeloid leukemia (aCML) patients is poor, but some patients with a suitable donor can be treated with allogeneic hematopoietic stem cell transplantation (HSCT). However, many of these patients cannot be treated with HSCT due to their age. The effectiveness of decitabine was recently indicated in case reports; however, the effectiveness of azacitidine (AZA) has not yet been reported. We report the case of a aCML patient successfully treated with AZA. A 66-year-old man with no remarkable medical history was admitted to our hospital because of leukocytosis. We diagnosed his disease as aCML and administered hydroxyurea (HU) and AZA. After four courses of AZA, his blood cell values improved, and he no longer needed transfusions and was able to stop HU. He continued receiving AZA without any severe complications. This is the first report that AZA is effective for the treatment of aCML.

Clinical impact of serum soluble SLAMF7 in multiple myeloma.

The signaling lymphocytic activation molecule family (SLAMF7; also known as CS1 or CD319) is highly expressed on plasma cells from multiple myeloma (MM) as well as natural killer (NK) cells and is a well-known therapeutic target of elotuzumab. The objective of this study was to evaluate the clinical significance of serum soluble SLAMF7 (sSLAMF7) levels in patients with MM (n=103) and furthermore the impact of sSLMF7 on the antitumor activity of anti-SLAMF7 antibody. Thirty-one percent of MM patients, but not patients with monoclonal gammopathy of undetermined significance and healthy controls, had detectable levels of serum sSLAMF7, which were significantly increased in advanced MM patients. Further, MM in sSLAMF7-postive patients exhibited aggressive clinical characteristics with shorter progression-free survival times in comparison with sSLAMF7-negative patients. In responders to MM therapy, the levels of sSLAMF7 were undetectable or decreased compared with those before treatment. In addition, the anti-SLAMF7 antibody-mediated antibody-dependent cellular cytotoxicity of NK cells against MM cell lines was inhibited by recombinant SLAMF7 protein. Thus, our findings suggest that high concentrations of sSLAMF7, which could transiently suppress the therapeutic effects of elotuzumab, may be a useful indicator of disease progression in MM patients.

Treatment-free remission after two-year consolidation therapy with nilotinib in patients with chronic myeloid leukemia: STAT2 trial in Japan.

The purpose of this trial was to evaluate the efficacy of 2-year consolidation therapy with nilotinib, at a dose of 300 mg twice daily, for achieving treatment-free remission in chronic myeloid leukemia patients with a deep molecular response (BCR-ABL1IS ≤0.0032%). Successful treatment-free remission was defined as no confirmed loss of deep molecular response. We recruited 96 Japanese patients, of whom 78 sustained a deep molecular response during the consolidation phase and were therefore eligible to discontinue nilotinib in the treatment-free remission phase; of these, 53 patients (67.9%; 95% confidence interval: 56.4-78.1%) remained free from molecular recurrence in the first 12 months. The estimated 3-year treatment-free survival was 62.8%. Nilotinib was readministered to all patients (n=29) who experienced a molecular recurrence during the treatment-free remission phase. After restarting treatment, rapid deep molecular response returned in 25 patients (86.2%), with 50% of patients achieving a deep molecular response within 3.5 months. Tyrosine kinase inhibitor withdrawal syndrome was reported in 11/78 patients during the early treatment-free remission phase. The treatment-free survival curve was significantly better in patients with undetectable molecular residual disease than in patients without (3-year treatment-free survival, 75.6 versus 48.6%, respectively; P=0.0126 by the log-rank test). There were no significant differences in treatment-free survival between subgroups based on tyrosine kinase inhibitor treatment before the nilotinib consolidation phase, tyrosine kinase inhibitor-withdrawal syndrome, or absolute number of natural killer cells. The results of this study indicate that it is safe and feasible to stop tyrosine kinase inhibitor therapy in patients with chronic myeloid leukemia who have achieved a sustained deep molecular response with 2 years of treatment with nilotinib. This study was registered with UMIN-CTR (UMIN000005904).

Retrospective analysis of prognostic factors for WaldenstrÓ§m macroglobulinemia: a multicenter cooperative study in Japan.

Although population-based cancer registries have reported lower incidence of WaldenstrÓ§m macroglobulinemia (WM) in East Asia than in Western countries, previous retrospective analyses have found the clinical features of WM to be similar in these two populations. To clarify the characteristics of Japanese WM patients, we retrospectively analyzed clinical and laboratory characteristics, treatments, outcomes, and prognostic factors in 93 patients with WM. Based on the Second International Workshop on WM (IWWM-2) criteria, symptomatic WM was found in 73 (78.5%) and asymptomatic WM in 20 (21.5%) of cases examined. The median overall survival (OS) was similar to that in reports from Western countries. Patients receiving treatment regimens including rituximab exhibited significantly better survival than those not given rituximab. Although prognostic factors for WM in Western countries may not apply to Japanese patients, our finding that newly diagnosed WM patients with pleural effusion have a poorer prognosis suggests that this may be a novel predictor of adverse prognosis in symptomatic WM.

Incidence and clinical background of hepatitis B virus reactivation in multiple myeloma in novel agents' era.

There are some reports regarding hepatitis B virus (HBV) reactivation in patients with myeloma who are HBV carriers or who have had a resolved HBV infection, and there is no standard prophylaxis strategy for these patients. We performed a retrospective multicenter study to determine the incidence and characteristics of HBV reactivation in patients with multiple myeloma. We identified 641 patients with multiple myeloma who had been treated using novel agents and/or autologous stem cell transplantation with high-dose chemotherapy between January 2006 and June 2014 at nine Japanese hospitals. The patients' characteristics, laboratory data, and clinical courses were retrieved and statistically analyzed. During a median follow-up of 101 weeks, one of eight (12.5 %) HBV carriers developed hepatitis and 9 of 99 (9.1 %) patients with resolved HBV infection experienced HBV reactivation; the cumulative incidences of HBV reactivation at 2 years (104 weeks) and 5 years (260 weeks) were 8 and 14 %, respectively. The nine cases of reactivation after resolved HBV infection had received entecavir as preemptive therapy or were carefully observed by monitoring their HBV DNA levels, and none of these cases developed hepatitis. Among patients with multiple myeloma, HBV reactivation was not rare. Therefore, long-term monitoring of HBV DNA levels is needed to prevent hepatitis that is related to HBV reactivation in these patients.

[Evaluation of the enhanced International Prognostic Index (NCCN-IPI) for cases with diffuse large B-cell lymphoma].

The NCCN-International Prognostic Index (IPI) is reported to be more powerful than the former IPI for predicting survival in the rituximab era. To evaluate the NCCN-IPI in our institutions, we analyzed 188 patients with diffuse large B-cell lymphoma treated with rituximab plus CHOP or THP-COP chemotherapy. The 5-year overall survival rates of patients with low, low-intermediate, high-intermediate, and high risk were 90%, 76%, 64%, and 34%, respectively. Although there was no difference in overall survival between patients 61-75 and those >75 years of age, the NCCN-IPI is useful for classifying prognostically relevant subgroups of Japanese patients.

Oral administration of imatinib to P230 BCR/ABL-expressing transgenic mice changes clones with high BCR/ABL complementary DNA expression into those with low expression.

The effect of imatinib on myeloproliferative disease in transgenic (Tg) mice expressing the P230 BCR/ABL transcript is unknown. To investigate this issue, we administered imatinib (30 mg/kg per day) orally to P230 BCR/ABL-expressing Tg mice for 30 days. Following imatinib administration, the enlarged spleen was significantly reduced to within the normal size range. Infiltrating megakaryocytes in the long-axis section of the spleen were also significantly reduced. However, the cellularity of the bone marrow was not affected. Fluorescence-activated cell-sorting analysis revealed that infiltrating mature granulocytes in the spleen were reduced in number. The numbers of infiltrating CD34, CD117, CD61, and CD11b populations were also reduced in immature populations of the spleen. Real-time quantitative polymerase chain reaction analysis of messenger RNA revealed a dramatic reduction in the p230 BCR/ABL transcript for CD34, CD117, CD61, and CD11b populations in both bone marrow cells and spleen cells. Western blotting and immunoprecipitation analysis also revealed a marked reduction in P230 BCR/ABL protein expression in both bone marrow cells and spleen cells. Thus, imatinib administration had the intriguing effect of replacing clones with high expression of p230 BCR/ABL complementary DNA with clones with very low expression. These data show that imatinib may still be capable of eliminating and eradicating clones with high p230 BCR/ABL expression and healing the disease phenotype in Tg mice. Pluripotent clones with very low p230 BCR/ABL expression still survive as immature CD34, CD117, CD61, and CD11b populations.

Down-regulation and growth inhibitory role of C/EBPalpha in breast cancer.

PURPOSE: CCAAT/enhancer binding proteins (C/EBP) are a family of transcription factors that regulate proliferation and differentiation in a variety of tissues. The purpose of this study was to explore the possibility that C/EBPalpha is involved in breast cancer. EXPERIMENTAL DESIGN: We quantified C/EBPalpha mRNA expression levels in 24 primary breast tumors, 16 normal breast samples, and 8 breast cancer cell lines using quantitative real-time reverse transcription-PCR assay. C/EBPalpha protein levels were further determined by immunohistochemical analysis. To examine the consequence of C/EPBalpha expression in breast cancer, we stably transfected an inducible C/EPBalpha expression vector into three breast cancer cell lines. RESULTS: Low expression of C/EBPalpha mRNA was found in 83% of primary breast cancer samples. Immunohistochemical study further showed either a markedly reduced or undetectable expression of C/EBPalpha protein in 30% of breast cancer specimens. The other 70% of breast cancers had C/EBPalpha expression in both the cytoplasm and nucleus; in control, C/EBPalpha was localized to the nucleus in the normal ductal cells. C/EBPalpha expression was associated with estrogen- and progesterone receptor-negative status. Induction of C/EBPalpha expression in these cell lines resulted in growth inhibition accompanied by G0-G1 cell cycle arrest and reduced anchorage-independent cell growth. C/EBPalpha expression was associated with down-regulation of c-myc and up-regulation of p21, PPARgamma, and the breast epithelial differentiation marker, maspin. CONCLUSIONS: These results suggest that reduced expression of C/EBPalpha may play a role in the development and/or progression of breast cancer.

C/EBPdelta expression in a BCR-ABL-positive cell line induces growth arrest and myeloid differentiation.

CCAAT/enhancer-binding proteins (C/EBPs) are a family of highly conserved transcription factors that have important roles in normal myelopoiesis as well as associated with myeloid disorders. The chronic myelogenous leukemia (CML) cell lines, KCL22 and K562, express exceptionally low levels of endogenous C/EBPs and provide a good model to test the effects of C/EBPs on myeloid differentiation. To explore the possibility that C/EBPdelta can promote differentiation in BCR-ABL-positive cells, we generated stable KCL22 and K562 clones that expressed an inducible C/EBPdelta gene. C/EBPdelta expression resulted in G0/G1 proliferative arrest and a moderate increase in apoptosis of the KCL22 and the K562 cells. Within 4 days of inducing expression of C/EBPdelta, myeloid differentiation of the CML blast cells occurred as shown by morphologic changes and induction of secondary granule-specific genes. We also showed that during granulocytic differentiation of KCL22 cells, the C/EBPdelta protein was detected in immunocomplexes with both Rb and E2F1. Furthermore, expression of C/EBPdelta was associated with downregulation of c-Myc and cyclin E and upregulation of the cyclin-dependent kinase inhibitor p27(Kip1) in both the KCL22 and K562 cell lines. These results show that expression of C/EBPdelta in BCR-ABL-positive leukemic cells in blast crisis is sufficient for neutrophil differentiation and point to the therapeutic potential of ectopic induction of C/EBPdelta in the acute phase of CML.

Identifying progression-associated genes in adult T-cell leukemia/lymphoma by using oligonucleotide microarrays.

Adult T-cell leukemia/lymphoma (ATL) is associated with human T-lymphotropic virus type-1 (HTLV-1). To understand the changes in expression that occur in the progression of chronic phase of ATL to acute crisis, the gene expression profiles of fresh ATL cells were compared in 4 pairs of samples (progression of chronic to acute phase in 3 patients, as well as 1 typical chronic phase sample vs. 1 typical acute phase sample) using high-density oligonucleotide DNA arrays. We identified 203 genes that were commonly upregulated in acute vs. chronic phase samples including ribosomal proteins, proteosome subunits, eukaryotic translation factors, immunophilins, heat shock proteins and genes important for DNA replication. Additionally, we identified 91 commonly downregulated genes including immune molecules related to MHC and a phosphatase. Several of the genes were previously identified to be associated with the Tax protein of HTLV-1. Some of the upregulated genes were located in amplified regions identified by comparative genomic hybridization in the corresponding chronic/acute ATL sample. Using real-time quantitative PCR, we confirmed the array-results in those specimens analyzed by microarray. These results demonstrated that distinct sets of genes that are known to be critical in cellular transformation and/or activation are up- or down-regulated during the transition to the acute phase of ATL.

Insulin-like growth factor binding protein-3 antagonizes the effects of retinoids in myeloid leukemia cells.

Insulin-like growth factor binding protein-3 (IGFBP-3) can cause growth suppressive and proapoptotic effects on retinoids in many types of cancer cells. However, the expression and effects of IGFBP-3 in myeloid leukemia cells have not been elucidated. In this study, we found no IGFBP-3 expression in the human myeloid leukemia cell lines either at baseline or after stimulation with all-trans retinoic acid (ATRA). Human recombinant IGFBP-3 induced growth arrest and apoptosis of HL-60 and NB4 cells. We have previously identified RXR alpha as a nuclear receptor for IGFBP-3 and have proceeded to examine further the role of this interaction in leukemia cell lines. In signaling assays, IGFBP-3 potently suppressed RAR- and VDR-mediated signaling while enhancing RXR signaling. Interestingly, when IGFBP-3 was administered to these cells in combination with an RAR-selective ligand, the ability of these retinoids to induce differentiation was blunted. On the other hand, IGFBP-3 enhanced the effect of an RXR-selective ligand to induce differentiation of HL-60 and NB4 cells. Further studies showed that IGFBP-3 down-regulated (at the transcriptional level) the retinoid-induced expression of C/EBP epsilon in NB4 cells. Taken together, these results indicate that IGFBP-3 has antiproliferative activity against myeloid leukemia cells; while it enhances signaling through RXR/RXR, it blunts signaling by activated RAR/RXR.