Hayato Tamai

Activated K-Ras protein accelerates human MLL/AF4-induced leukemo-lymphomogenicity in a transgenic mouse model

Activated K-Ras protein accelerates human MLL/AF4-induced leukemo-lymphomogenicity in a transgenic mouse model

AAV8 vector expressing IL24 efficiently suppresses tumor growth mediated by specific mechanisms in MLL/AF4-positive ALL model mice

Mixed-lineage leukemia (MLL)/AF4-positive acute lymphoblastic leukemia (ALL) is a common type of leukemia in infants, which is associated with a high relapse rate and poor prognosis. IL24 selectively induces apoptosis in cancer cells and exerts immunomodulatory and antiangiogenic effects. We examined the effects of adeno-associated virus type 8 (AAV8) vector-mediated muscle-directed systemic gene therapy in MLL/AF4-positive ALL using IL24. In a series of in vitro studies, we examined the effects of AAV8-IL24-transduced C2C12 cell-conditioned medium. We also examined the effects of AAV8-IL24 in MLL/AF4 transgenic mice. The results revealed the effects of AAV8-IL24 in MLL/AF4-positive ALL both in vitro and in vivo. With regard to the mechanism of therapy using AAV8-IL24 in MLL/AF4-positive ALL, we demonstrated the antiangiogenicity and effects on the ER stress pathway and unreported pathways through inhibition of S100A6 and HOXA9, which is specific to MLL/AF4-positive ALL. Inhibition of S100A6 by IL24 was dependent on TNF-α and induced acetylation of p53 followed by activation of the caspase 8-caspase 3 apoptotic pathway. Inhibition of HOXA9 by IL24, which was independent of TNF-α, induced MEIS1 activation followed by activation of the caspase 8-caspase 3 apoptotic pathway. Thus, gene therapy using AAV8-IL24 is a promising treatment for MLL/AF4-positive ALL.

RCSD1-ABL1-positive B lymphoblastic leukemia is sensitive to dexamethasone and tyrosine kinase inhibitors and rapidly evolves clonally by chromosomal translocations

Recently, RCSD1 was identified as a novel gene fusion partner of the ABL1 gene. The RCSD1 gene, located at 1q23, is involved in t(1;9)(q23;q34) translocation in acute B lymphoblastic leukemia. Here we describe RCSD1-ABL1-positive B-cell acute lymphoblastic leukemia (ALL) followed by rapid clonal evolution exhibiting three rare reciprocal translocations. We performed breakpoint analysis of the transcript expressed by the RCSD1-ABL1 fusion gene. RT-PCR and sequence analyses detected transcription of a single RCSD1-ABL1 fusion gene variant, which had breakpoints in exon 3 of RCSD1 and exon 4 of ABL1. The RCSD1 portion of the RCSD1-ABL1 fusion transcript consists of exons 1, 2, and 3. Tyrosine kinase inhibitors, imatinib and dasatinib, coadministered with dexamethasone achieved transient clinical effects in the present RCSD1-ABL1-positive ALL. However, leukemic cells rapidly became refractory to this treatment following the subsequent development of three additional reciprocal chromosomal translocations, t(5;16)(q33;q24), dic(18;20)(p11.2;q11.2) and t(10;19)(q24;p13.3). The present RCSD1-ABL1-positive ALL may represent a state of high chromosomal instability.

Complex molecular genetic abnormalities involving three or more genetic mutations are important prognostic factors for acute myeloid leukemia

We conducted a comprehensive analysis of 28 recurrently mutated genes in acute myeloid leukemia (AML) in 271 patients with de novo AML. Co-mutations were frequently detected in the intermediate cytogenetic risk group, at an average of 2.76 co-mutations per patient. When assessing the prognostic impact of these co-mutations in the intermediate cytogenetic risk group, overall survival (OS) was found to be significantly shorter (P=0.0006) and cumulative incidence of relapse (CIR) significantly higher (P=0.0052) in patients with complex molecular genetic abnormalities (CMGAs) involving three or more mutations. This trend was marked even among patients aged ⩽65 years who were also FLT3-ITD (FMS-like tyrosine kinase 3 internal tandem duplications)-negative (OS: P=0.0010; CIR: P=0.1800). Moreover, the multivariate analysis revealed that CMGA positivity was an independent prognostic factor associated with OS (P=0.0007). In stratification based on FLT3-ITD and CEBPA status and ‘simplified analysis of co-mutations’ using seven genes that featured frequently in CMGAs, CMGA positivity retained its prognostic value in transplantation-aged patients of the intermediate cytogenetic risk group (OS: P=0.0002. CIR: P<0.0001). In conclusion, CMGAs in AML were found to be strong independent adverse prognostic factors and simplified co-mutation analysis to have clinical usefulness and applicability.

Clinical features of adult acute leukemia with 11q23 abnormalities in Japan: a co-operative multicenter study

To clarify the clinical features of adult patients with acute leukemia (AL) with 11q23 abnormalities, we performed a retrospective analysis of data from 58 adult Japanese patients: 51 with acute myeloid leukemia (AML), and 7 with acute lymphoblastic leukemia (ALL). The incidences according to fusion partners in AML were: t(9;11), 31.3%; t(11;19), 27.4%; t(6;11), 21.5%. The incidence of patients with t(11;19) was higher than those in the US and Europe, and the incidence of t(4;11) was lower than that in childhood. The results indicated the poor prognosis of AML with 11q23 abnormalities regardless of the fusion partners. AML patients with 11q23 aged <60 years in the first CR who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) showed a more favorable outcome than those treated without allo-HSCT, although the differences were not statistically significant (P = 0.322 for DFS, P = 0.138 for OS). This result suggests that treatment strategies including allo-HSCT may be considered in the first CR in cases of AML with 11q23 abnormalities. However, further studies involving a large number of cases are required to assess the effect of allo-HSCT on adult AL with 11q23 abnormalities.

Resistance of MLL–AFF1-positive acute lymphoblastic leukemia to tumor necrosis factor-alpha is mediated by S100A6 upregulation

Mixed-lineage leukemia (MLL)–AFF1 (MLL–AF4)-positive acute lymphoblastic leukemia (ALL) is associated with poor prognosis, even after allogeneic hematopoietic stem cell transplantation (allo-HSCT). The resistance to graft-versus-leukemia (GVL) effects may be responsible for the poor effect of allo-HSCT on MLL–AFF1-positive ALL. Cytotoxic effector mechanisms mediated by tumor necrosis factor-alpha (TNF-α) was reported to contribute to the GVL effect. We showed that MLL–AFF1-positive ALL cell lines are resistant to TNF-α. To examine the mechanism of resistance to TNF-α of MLL–AFF1-positive leukemia, we focused on S100A6 as a possible factor. Upregulation of S100A6 expression and inhibition of the p53–caspase 8–caspase 3 pathway were observed only in MLL–AFF1-positive ALL cell lines in the presence of TNF-α. The effect of S100A6 on resistance to TNF-α by inhibition of the p53–caspase 8–caspase 3 pathway of MLL–AFF1-positive ALL cell lines were also confirmed by analysis using small interfering RNA against S100A6. This pathway was also confirmed in previously established MLL–AFF1 transgenic mice. These results suggest that MLL–AFF1-positive ALL escapes from TNF-α-mediated apoptosis by upregulation of S100A6 expression, followed by interfering with p53–caspase 8–caspase 3 pathway. These results suggest that S100A6 may be a promising therapeutic target for MLL–AFF1-positive ALL in combination with allo-HSCT.

Clinical characteristics and prognosis of acute myeloid leukemia associated with DNA-methylation regulatory gene mutations.

In recent years, it has been reported that the frequency of DNA-methylation regulatory gene mutations – mutations of the genes that regulate gene expression through DNA methylation – is high in acute myeloid leukemia. The objective of the present study was to elucidate the clinical characteristics and prognosis of acute myeloid leukemia with associated DNA-methylation regulatory gene mutation. We studied 308 patients with acute myeloid leukemia. DNA-methylation regulatory gene mutations were observed in 135 of the 308 cases (43.8%). Acute myeloid leukemia associated with a DNA-methylation regulatory gene mutation was more frequent in older patients (P<0.0001) and in patients with intermediate cytogenetic risk (P<0.0001) accompanied by a high white blood cell count (P=0.0032). DNA-methylation regulatory gene mutation was an unfavorable prognostic factor for overall survival in the whole cohort (P=0.0018), in patients aged ≤70 years, in patients with intermediate cytogenetic risk, and in FLT3-ITD-negative patients (P=0.0409). Among the patients with DNA-methylation regulatory gene mutations, 26.7% were found to have two or more such mutations and prognosis worsened with increasing number of mutations. In multivariate analysis DNA-methylation regulatory gene mutation was an independent unfavorable prognostic factor for overall survival (P=0.0424). However, patients with a DNA-methylation regulatory gene mutation who underwent allogeneic stem cell transplantation in first remission had a significantly better prognosis than those who did not undergo such transplantation (P=0.0254). Our study establishes that DNA-methylation regulatory gene mutation is an important unfavorable prognostic factor in acute myeloid leukemia.

Identification of TINF2 gene mutations in adult Japanese patients with acquired bone marrow failure syndromes.

Haematologica, 118, 27–29. Finnegan, D.P., Kettle, P., Drake, M., Matthews, C., Alexander, H.D., Popat, R., Cavanagh, J.D., Wachsman, W. & Morris, T. (2006) Bortezomib is effective in primary plasma cell leukemia. Leukaemia & Lymphoma, 47, 1670–1673. Garcia-Sanz, R., Orfao, A., Gonzalez, M., Tabernero, M.D., Blade, J., Moro, M.J., Fernandez-Calvo, J., Sanz, M.A., PerezSimon, J.A., Rasillo, A. & Miguel, J.F. (1999) Primary plasma cell leukemia: clinical, immunophenotypic, DNA ploidy, and cytogenetic characteristics. Blood, 93, 1032–1037. Gemmel, C., Cremer, F.W., Weis, M., Witzens, M., Moldenhauer, G., Koniczek, K.H., Imbach, U., Ho, A.D., Moos, M. & Goldschmidt, H. (2002) Anti-CD20 antibody as consolidation therapy in a patient with primary plasma cell leukemia after high-dose therapy and autologous stem cell transplantation. Annals of Hematology, 81, 119–123. Oka, S., Yokote, T., Akioka, T., Hara, S., Yamano, T., Tsuji, M. & Hanafusa, T. (2006) Successful treatment of multi-agent chemotherapy with rituximab for IgM plasma cell leukemia. Leukemia Research, 30, 1581–1583. RuizArguelles, G.J. & San Miguel, J.F. (1994) Cell surface markers in multiple myeloma. Mayo Clinic Proceedings, 69, 684–690. Saccaro, S., Fonseca, R., Veillon, D.M., Cotelingam, J., Nordberg, M.L., Bredeson, C., Glass, J. & Munker, R. (2005) Primary plasma cell leukemia: report of 17 new cases treated with autologous or allogeneic stem-cell transplantation and review of the literature. American Journal of Hematology, 78, 288–294. Tanioka, F., Tamashima, S., Shimizu, S., Kobayashi, H., Kobayashi, Y. & Sugimura, H. (2003) A case of primary plasma cell leukemia with hairycell morphology and k -type Bence-Jones protein. Immunohistochemical and molecular analysis. Japanese Journal of Clinical Oncology, 33, 232–237. Walters, M., Olteanu, H., Van Tuinen, P. & Kroft, S.H. (2010) CD23 expression in plasma cell myeloma is specific for abnormalities of chromosome 11, and is associated with primary plasma cell leukaemia in this cytogenetic sub-group. British Journal of Haematology, 149, 292–293.

Treatment of relapsed acute myeloid leukemia with MLL/AF6 fusion after allogeneic hematopoietic stem cell transplantation with gemtuzumab ozogamicin with a long interval followed by donor lymphocyte infusion

Treatment of relapsed acute myeloid leukemia with MLL/AF6 fusion after allogeneic hematopoietic stem cell transplantation with gemtuzumab ozogamicin with a long interval followed by donor lymphocyte infusion

correspondence: Identification of TINF2 gene mutations in adult Japanese patients with acquired bone marrow failure syndromes

Haematologica, 118, 27–29. Finnegan, D.P., Kettle, P., Drake, M., Matthews, C., Alexander, H.D., Popat, R., Cavanagh, J.D., Wachsman, W. & Morris, T. (2006) Bortezomib is effective in primary plasma cell leukemia. Leukaemia & Lymphoma, 47, 1670–1673. Garcia-Sanz, R., Orfao, A., Gonzalez, M., Tabernero, M.D., Blade, J., Moro, M.J., Fernandez-Calvo, J., Sanz, M.A., PerezSimon, J.A., Rasillo, A. & Miguel, J.F. (1999) Primary plasma cell leukemia: clinical, immunophenotypic, DNA ploidy, and cytogenetic characteristics. Blood, 93, 1032–1037. Gemmel, C., Cremer, F.W., Weis, M., Witzens, M., Moldenhauer, G., Koniczek, K.H., Imbach, U., Ho, A.D., Moos, M. & Goldschmidt, H. (2002) Anti-CD20 antibody as consolidation therapy in a patient with primary plasma cell leukemia after high-dose therapy and autologous stem cell transplantation. Annals of Hematology, 81, 119–123. Oka, S., Yokote, T., Akioka, T., Hara, S., Yamano, T., Tsuji, M. & Hanafusa, T. (2006) Successful treatment of multi-agent chemotherapy with rituximab for IgM plasma cell leukemia. Leukemia Research, 30, 1581–1583. RuizArguelles, G.J. & San Miguel, J.F. (1994) Cell surface markers in multiple myeloma. Mayo Clinic Proceedings, 69, 684–690. Saccaro, S., Fonseca, R., Veillon, D.M., Cotelingam, J., Nordberg, M.L., Bredeson, C., Glass, J. & Munker, R. (2005) Primary plasma cell leukemia: report of 17 new cases treated with autologous or allogeneic stem-cell transplantation and review of the literature. American Journal of Hematology, 78, 288–294. Tanioka, F., Tamashima, S., Shimizu, S., Kobayashi, H., Kobayashi, Y. & Sugimura, H. (2003) A case of primary plasma cell leukemia with hairycell morphology and k -type Bence-Jones protein. Immunohistochemical and molecular analysis. Japanese Journal of Clinical Oncology, 33, 232–237. Walters, M., Olteanu, H., Van Tuinen, P. & Kroft, S.H. (2010) CD23 expression in plasma cell myeloma is specific for abnormalities of chromosome 11, and is associated with primary plasma cell leukaemia in this cytogenetic sub-group. British Journal of Haematology, 149, 292–293.