Makiko Takizawa

Long non-coding RNA MALAT1 is an inducible stress response gene associated with extramedullary spread and poor prognosis of multiple myeloma

Extramedullary myeloma (EMM) occurs when myeloma develops outside the bone marrow; it often develops after chemotherapy and is associated with the acquisition of chemo‐resistance and a fatal course. The mechanisms underlying extramedullary spread have not yet been fully elucidated. MALAT1 is a highly abundantly and ubiquitously expressed long non‐coding RNA that plays important roles in cancer metastasis. The aims of this study were to clarify the association of MALAT1 with EMM and to elucidate the underlying mechanism of EMM formation under chemotherapeutic pressure. MALAT1 expression was significantly higher in multiple myeloma (MM) than in monoclonal gammopathy of undetermined significance. Furthermore, MALAT1 expression was markedly higher in EMM compared with that in corresponding intramedullary myeloma cells. A higher MALAT1 level was associated with shorter overall and progression‐free survival. MALAT1 expression level was positively correlated with expression of HSP90AA1, HSP90AB1 and HSP90B1 but not with TP53 expression. MALAT1 was significantly upregulated by bortezomib and doxorubicin. Considering the known functions of MALAT1, our results suggest that it acts as a stress response gene that is upregulated by chemotherapy, thereby linking chemotherapy to EMM formation. Elucidating the biological implication of long non‐coding RNA contributes to deeper understanding concerning the pathogenesis and investigation of novel therapeutic targets for MM.

Characterization of CD56+ dendritic-like cells: a normal counterpart of blastic plasmacytoid dendritic cell neoplasm?

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare hematological malignancy. Plasmacytoid DCs (pDCs), which are defined as lineage marker (Lin)−HLA−DR+CD56−CD123+CD11c− cells, are considered to be the normal counterpart of BPDCNs. However, BPDCN can be distinguished from pDCs by uniform expression of CD56. In this study, to identify a normal counterpart of BPDCN, we searched for a Lin−HLA−DR+CD56+ population and focused on a minor subpopulation of Lin−DR+CD56+CD123+CD11c− cells that we designated as pDC-like cells (pDLCs). pDLC constituted 0.03% of peripheral blood mononuclear cells (PBMCs), and the pDLC/pDC ratio was higher in bone marrow cells than in PBMCs. pDLC clearly expressed BDCA2, BDCA4, and myeloid antigens, which are frequently expressed by BPDCN. pDLCs exhibited modest expression of Toll-like receptors and produced less interferon-α after CpG stimulation, but presented very low endocytic ability unlike mDCs. These functional differences were attributed to the expression profile of transcriptional factors. After in vitro culture with Flt3-ligand and GM-CSF, pDLCs expressed CD11c and BDCA1. These data suggested that pDLCs are a distinct subpopulation, with an immunophenotype similar to BPDCNs. Moreover, our results indicate that pDLCs might be immature DCs and might contribute to the immunophenotypical diversity of BPDCNs.

Sacroiliitis as an initial manifestation of acute myelogenous leukemia

Sacroiliitis is the most pathognomonic and earliest manifestation of ankylosing spondylitis.We herein report a 28-year-old female patient who presented with sacroiliitis as an initial manifestation of acute myelogenous leukemia (AML). She had a 3-month history of anemia and walking difficulty. Bone marrow findings revealed an increase of blasts with trilineage dysplasia. Although she was initially diagnosed with myelodysplastic syndrome (MDS), blasts rapidly increased and AML developed 1 month after the diagnosis of MDS with sacroiliitis. Induction chemotherapy failed to induce a complete remission of AML, but it did effectively treat the sacroiliitis. However, the sacroiliitis relapsed when the leukemia cells progressed thereafter. Oral corticosteroids helped ameliorate the sacroiliitis. She underwent bone marrow transplantation (BMT) from an HLA-identical sister during a nonremission period; however, the leukemic cells began to rapidly increase from day 30 after BMT. The close relationship between the occurrence of sacroiliitis and AML suggested that autoimmune sacroiliitis was a paraneoplastic phenomenon of AML in this patient. Although autoimmune disorders develop in a substantial number of MDS patients, they are rarely observed in de novo AML. No previous report has described sacroiliitis as the initial manifestation of de novo AML.

Pericardial graft vs. host disease in a patient with myelodysplastic syndrome following peripheral blood stem cell transplantation

Abstract:  A patient with myelodysplastic syndrome developed pericardial effusion 20 month after allogenic peripheral blood stem cell transplantation. Sclerotic and erythematous skin lesions were observed over the face and extremities, and a diagnosis of chronic graft vs. host disease (GVHD) was made based on skin biopsy findings. Pericardial fluid contained numerous CD8+/HLA‐DR+ lymphocytes, but no leukaemic cells. Tumour necrosis factor alpha (TNFα) and soluble Fas (sFas) levels were highly elevated in both the effusion and serum. The patient was treated with methylprednisolone and tacrolimus. Skin GVHD improved rapidly associated with resolution of pericardial effusion and reductions in cytokine levels. We concluded that pericardial effusion was due to pericarditis and was a manifestation of chronic GVHD in this patient, and that cytotoxic lymphocytes and specific cytokines played significant roles.

Effect of cytokines on growth and differentiation of leukaemic cells with translocation t(6;9)(p23;q34)

The translocation t(6;9)(p23;q34) is detected infrequently in subtypes of haematological malignancies including acute myelogenous leukaemia (AML) and myelodysplastic syndrome (MDS). Although the t(6;9) leukaemia is commonly associated with bone marrow basophilia, the cytological characteristics of leukaemic cells are unclear. In the current study, we examined the in vitro effects of several cytokines on growth and differentiation of t(6;9) leukaemic cells. Isolated bone marrow mononuclear cells from four patients with t(6;9) (two MDS and two AML) were cultured for 14 d in the presence or absence of each cytokine. At the end of culture, viable cells were counted, and their histology was examined. Bone marrow cells obtained from 22 patients (10 AML, six AML from MDS, six MDS) lacking t(6;9) were used as controls. Compared with control cultures, significantly higher numbers of blasts appeared in the culture of bone marrow cells from t(6;9)‐positive patients in response to stimulation with granulocyte colony‐stimulating factor (G‐CSF), granulocyte–macrophage CSF (GM‐CSF) or interleukin 3 (IL‐3). Stem cell factor (SCF) had little effect. Neutrophil counts were also significantly increased in the presence of G‐CSF or IL‐3. SCF and IL‐3 were potent in increasing basophil counts from t(6;9)‐positive cultures. These findings suggest that bone marrow cells obtained from t(6;9) patients are highly sensitive to growth‐ and/or differentiation‐promoting cytokines. Special attention should be paid to the use of ‘therapeutic’ cytokines in these patients.

Overexpression of B-cell lymphoma 6 alters gene expression profile in a myeloma cell line and is associated with decreased DNA damage response

B‐cell lymphoma 6 (BCL6) attenuates DNA damage response (DDR) through gene repression and facilitates tolerance to genomic instability during immunoglobulin affinity maturation in germinal center (GC) B cells. Although BCL6 expression is repressed through normal differentiation of GC B cells into plasma cells, a recent study showed the ectopic expression of BCL6 in primary multiple myeloma (MM) cells. However, the functional roles of BCL6 in MM cells are largely unknown. Here, we report that overexpression of BCL6 in a MM cell line, KMS12PE, induced transcriptional repression of ataxia telangiectasia mutated (ATM), a DDR signaling kinase, which was associated with a reduction in γH2AX formation after DNA damage. In contrast, transcription of known targets of BCL6 in GC B cells was not affected, suggesting a cell type‐specific function of BCL6. To further investigate the effects of BCL6 overexpression on the MM cell line, we undertook mRNA sequence analysis and found an upregulation in the genomic mutator activation‐induced cytidine deaminase (AID) with alteration in the gene expression profile, which is suggestive of de‐differentiation from plasma cells. Moreover, interleukin‐6 exposure to KMS12PE led to upregulation of BCL6 and AID, downregulation of ATM, and attenuation of DDR, which were consistent with the effects of BCL6 overexpression in this MM cell line. Taken together, these results indicated that overexpression of BCL6 alters gene expression profile and confers decreased DDR in MM cells. This phenotypic change could be reproduced by interleukin‐6 stimulation, suggesting an important role of external stimuli in inducing genomic instability, which is a hallmark of MM cells.

PDCD1 and CTLA4 polymorphisms affect the susceptibility to, and clinical features of, chronic immune thrombocytopenia

Programmed death‐1 (PD‐1, PDCD1) and cytotoxic T lymphocyte‐associated antigen‐4 (CTLA‐4, CTLA4) play central roles in immune checkpoint pathways. Single nucleotide polymorphisms (SNPs) of PDCD1 and CTLA4 have been reported to be associated with susceptibility to some autoimmune diseases. However, the potential association between SNPs in these immune checkpoint genes and risk of chronic immune thrombocytopenia (cITP) remain controversial and obscure. The aims of this study were to clarify the influence of PDCD1 and CTLA4 SNPs on the risk of developing cITP and its clinical features. We obtained genomic DNA from 119 patients with cITP and 223 healthy controls; their genotypes were determined by the polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) method. Patients with cITP had a significantly higher frequency of the PDCD1 +7209 TT genotype compared with healthy controls. The CTLA4 −1577 GG genotype and CT60 GG genotype showed higher frequencies of platelet count <5 × 109/l at diagnosis, minimum platelet count <5 × 109/l, and bleeding symptoms. Moreover, the PDCD1 −606 AA genotype and +63379 TT genotype were significantly associated with a lower number of patients who achieved a complete response to prednisolone treatment. Our results suggest that the immune checkpoint polymorphisms may affect the susceptibility to the clinical features of cITP, and treatment response of the affected patients.

Association between OGG1 S326C CC genotype and elevated relapse risk in acute myeloid leukemia

Recent studies have shown that tumors of relapsed acute myeloid leukemia (AML) present additional genetic mutations compared to the primary tumors. The base excision repair (BER) pathway corrects oxidatively damaged mutagenic bases and plays an important role in maintaining genetic stability. The purpose of the present study was to investigate the relationship between BER functional polymorphisms and AML relapse. We focused on five major polymorphisms: OGG1 S326C, MUTYH Q324H, APE1 D148E, XRCC1 R194W, and XRCC1 R399Q. Ninety-four adults with AML who achieved first complete remission were recruited. Genotyping was performed with the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The OGG1 S326C CC genotype (associated with lower OGG1 activity) was observed more frequently in patients with AML relapse [28.9 vs. 8.9%, odds ratio (OR) = 4.10, 95% confidence interval (CI) = 1.35–12.70, P = 0.01]. Patients with the CC genotype exhibited shorter relapse-free survival (RFS). Moreover, the TCGA database suggested that low OGG1 expression in AML cells is associated with a higher frequency of mutations. The present findings suggest that the OGG1 S326C polymorphism increased the probability of AML relapse and may be useful as a prognostic factor for AML relapse risk.

IL17A and IL23R gene polymorphisms affect the clinical features and prognosis of patients with multiple myeloma

Single nucleotide polymorphisms (SNPs) in interleukin 17 (IL17A) and IL‐23 receptor (IL23R) are involved in the pathogenesis of many cancers and autoimmune diseases. We investigated the influence of IL17A and IL23R SNPs on the risk of developing multiple myeloma (MM) and its clinical features. We obtained genomic DNA from 120 patients with MM and 201 healthy controls and detected IL17A −197 G/A (rs2275913) and IL23R H3Q (rs1884444) genotypes using the polymerase chain reaction‐restriction fragment length polymorphism method. There were no significant differences in the genotype and allele frequencies of IL17A −197 G/A and IL23R H3Q between the controls and patients with MM. Compared with the GG and GA genotypes, the IL17A AA genotype was significantly associated with lower hemoglobin levels. The IL23R HH genotype was significantly associated with higher frequency of bone lesions and plasmacytoma than the HQ and QQ genotypes. We observed significant differences in overall survival (OS) between patients treated with thalidomide and/or bortezomib and those treated conventionally. Therefore, we also examined the effect of IL17A and IL23R polymorphisms on the clinical variables and OS in patients treated with thalidomide and/or bortezomib. We observed that the IL23R HH genotype was significantly associated with poor survival compared with the QH and HH genotypes in these patients. Our findings indicate that IL17A −197 G/A and IL23R H3Q are not associated with susceptibility to MM. However, IL‐17 and IL‐23R polymorphisms may affect severity, bone lesions, and extra‐medullary disease in patients with MM. Moreover, IL23R polymorphisms may contribute to poor prognosis in patients with MM treated with thalidomide and/or bortezomib.