ABSTRACT
Recent genetic analysis has identified frequent mutations in ten-eleven translocation 2 (TET2), DNA methyltransferase 3A (DNMT3A), isocitrate dehydrogenase 2 (IDH2) and ras homolog family member A (RHOA) in nodal T-cell lymphomas, including angioimmunoblastic T-cell lymphoma and peripheral T-cell lymphoma, not otherwise specified. We examined the distribution of mutations in these subtypes of mature T-/natural killer cell neoplasms to determine their clonal architecture. Targeted sequencing was performed for 71 genes in tumor-derived DNA of 87 cases. The mutations were then analyzed in a programmed death-1 (PD1)-positive population enriched with tumor cells and CD20-positive B cells purified by laser microdissection from 19 cases. TET2 and DNMT3A mutations were identified in both the PD1+ cells and the CD20+ cells in 15/16 and 4/7 cases, respectively. All the RHOA and IDH2 mutations were confined to the PD1+ cells, indicating that some, including RHOA and IDH2 mutations, being specific events in tumor cells. Notably, we found that all NOTCH1 mutations were detected only in the CD20+ cells. In conclusion, we identified both B- as well as T-cell-specific mutations, and mutations common to both T and B cells. These findings indicate the expansion of a clone after multistep and multilineal acquisition of gene mutations.
Subject(s)
Biomarkers, Tumor , Lymphoma, Extranodal NK-T-Cell/genetics , Mutation , Alleles , Amino Acid Substitution , B-Lymphocytes/metabolism , B-Lymphocytes/pathology , DNA Methyltransferase 3A , Gene Rearrangement, T-Lymphocyte , Genetic Predisposition to Disease , Humans , Immunoglobulin Heavy Chains/genetics , Immunohistochemistry , Immunophenotyping , Lymphoma, Extranodal NK-T-Cell/metabolism , Lymphoma, Extranodal NK-T-Cell/pathology , Organ Specificity/genetics , Phenotype , Sequence Analysis, DNA , V(D)J Recombination , rhoA GTP-Binding Protein/genetics , rhoA GTP-Binding Protein/metabolismABSTRACT
In leukemogenesis, Notch signaling can be up and downregulated in a context-dependent manner. The transcription factor hairy and enhancer of split-1 (Hes1) is well-characterized as a downstream target of Notch signaling. Hes1 encodes a basic helix-loop-helix-type protein, and represses target gene expression. Here, we report that deletion of the Hes1 gene in mice promotes acute myeloid leukemia (AML) development induced by the MLL-AF9 fusion protein. We then found that Hes1 directly bound to the promoter region of the FMS-like tyrosine kinase 3 (FLT3) gene and downregulated the promoter activity. FLT3 was consequently upregulated in MLL-AF9-expressing immortalized and leukemia cells with a Hes1- or RBPJ-null background. MLL-AF9-expressing Hes1-null AML cells showed enhanced proliferation and ERK phosphorylation following FLT3 ligand stimulation. FLT3 inhibition efficiently abrogated proliferation of MLL-AF9-induced Hes1-null AML cells. Furthermore, an agonistic anti-Notch2 antibody induced apoptosis of MLL-AF9-induced AML cells in a Hes1-wild type but not a Hes1-null background. We also accessed two independent databases containing messenger RNA (mRNA) expression profiles and found that the expression level of FLT3 mRNA was negatively correlated with those of HES1 in patient AML samples. These observations demonstrate that Hes1 mediates tumor suppressive roles of Notch signaling in AML development, probably by downregulating FLT3 expression.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/genetics , Gene Expression Regulation, Leukemic , Homeodomain Proteins/genetics , Leukemia, Myeloid, Acute/genetics , fms-Like Tyrosine Kinase 3/genetics , Animals , Basic Helix-Loop-Helix Transcription Factors/deficiency , Cell Proliferation , Disease Models, Animal , Extracellular Signal-Regulated MAP Kinases/genetics , Extracellular Signal-Regulated MAP Kinases/metabolism , Humans , Immunoglobulin J Recombination Signal Sequence-Binding Protein/deficiency , Immunoglobulin J Recombination Signal Sequence-Binding Protein/genetics , Leukemia, Myeloid, Acute/metabolism , Leukemia, Myeloid, Acute/mortality , Leukemia, Myeloid, Acute/pathology , Mice , Mice, Transgenic , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Phosphorylation , Promoter Regions, Genetic , Protein Binding , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Notch/genetics , Receptors, Notch/metabolism , Signal Transduction , Survival Analysis , Transcription Factor HES-1 , fms-Like Tyrosine Kinase 3/metabolismABSTRACT
A simple method for the detection and spectrodensitometric determination of a number of trichothecene mycotoxins on silica gel layers based on a colour reaction between 4-(p-nitrobenzyl)pyridine and the 12,13-epoxy group in the trichothecene nucleus is described. The detection limits for the twelve trichothecenes examined were 0.025--0.2 micrograms per spot. Further, six of the twelve trichothecenes could be determined spectrodensitometrically in the range from ca. 0.05--0.2 to 10 micrograms per spot with a coefficient of variation of ca. 5%.