Contributions
Abstract: PB2024
Type: Publication Only
Background
Idiopathic leukocytosis and erythrocytosis are hematological disorders without specific causes. Frequent V617F mutations on the JAK2 gene have been reported in patients with polycythemia vera (PV), essential thrombocythemia, and primary myelofibrosis. We also found JAK2 V617F mutations in one of 11 patients with idiopathic erythrocytosis. Mutations of the CSF3R, SETBP1, and ETNK1 genes have been found in chronic neutrophilic leukemia and atypical chronic myeloid leukemia (CML). Furthermore an autosomal mutation was found in the CSF3R gene in a family with chronic neutrophilia. However, little is known about mutations associated with idiopathic leukocytosis.
Aims
We previously analyzed the JAK2, CSF3R, CALR, SETBP1, and ETNK1 genes in 10 patients with idiopathic leukocytosis (EHA20). To elucidate the relevance of genetic alterations, we extended the analysis with 17 genes known to be involved in hematological neoplasms in 16 patients with idiopathic leukocytosis.
Methods
Leukocytosis is defined as a total white blood cell count more than two standard deviations above the mean, or a value greater than 11,000/μL. Those patients who satisfied the following criteria were included in the study: leukocytosis (predominantly neutrophils); the absence of apparent causes of leukocytosis; and documentation of the leukocytosis over a prolonged period of time. The period of observation was 1 year or longer in most patients. Sixteen patients with idiopathic leukocytosis were analyzed in the study. Neutrophils or mononuclear cells were collected after obtaining written informed consent from the 16 patients. Neutrophils from peripheral blood were purified by dextran sedimentation followed by hypotonic lysis and centrifugation with Ficoll-Conray. Mononuclear cells were isolated from bone marrow by Ficoll-Conray gradient centrifugation. Genomic DNA was extracted using the QIAamp DNA blood mini kit (Qiagen, Valencia, CA, USA). Mutations within hot spots of the CSF3R, JAK2, CALR, SETBP1, ETNK1, CBL, TET2, ASXL1, EZH2, IDH1/IDH2, DNMT3A, U2AF1, and CEBPA genes were analyzed by direct sequencing in both directions using a 3730xL DNA Analyzer (Life technologies, Carlsbad, CA, USA) and/or allele specific polymerase chain reaction analysis. Total RNA extraction and reverse transcriptase polymerase chain reaction (RT-PCR) were performed between the ETV6 and ABL1 genes in 10 patients. BCR/ABL1 gene was analyzed by RT-PCR or fluorescence in situ hybridization in 8 patients. The current study was conducted within the guidelines and with the approval of ethical committee.
Results
JAK2 V617F mutations were found in one of the 16 patients with idiopathic leukocytosis. No mutations were found in the other genes in the 16 idiopathic leukocytosis patients. ETV6-ABL1 fusion gene was detected in one of the 10 patients. No BCR/ABL1 fusion gene was detected in the 8 patients. One idiopathic leukocytosis patient with JAK2 V617F mutation has developed PV. Another patient with sustained leukocytosis for 20 years showed cytogenetic abnormalities during observation and has developed Philadelphia chromosome negative CML (Ph-CML). ETV6-ABL1 fusion gene was detected in this patient. Another patient with normal karyotype progressed to blast crisis of Ph-CML accompanied by cytogenetic abnormalities. Of the remaining 13 patients with idiopathic leukocytosis, one resolved the disease and twelve had a stable disease.
Conclusion
Idiopathic leukocytosis comprises heterogeneous conditions. JAK2 mutations and cytogenetic abnormalities are predictive of progression to hematological neoplasms.
Session topic: 15. Myeloproliferative neoplasms - Biology
Keyword(s): Myeloproliferative disorder, mutation analysis, Hematological malignancy, Cytogenetic abnormalities
Abstract: PB2024
Type: Publication Only
Background
Idiopathic leukocytosis and erythrocytosis are hematological disorders without specific causes. Frequent V617F mutations on the JAK2 gene have been reported in patients with polycythemia vera (PV), essential thrombocythemia, and primary myelofibrosis. We also found JAK2 V617F mutations in one of 11 patients with idiopathic erythrocytosis. Mutations of the CSF3R, SETBP1, and ETNK1 genes have been found in chronic neutrophilic leukemia and atypical chronic myeloid leukemia (CML). Furthermore an autosomal mutation was found in the CSF3R gene in a family with chronic neutrophilia. However, little is known about mutations associated with idiopathic leukocytosis.
Aims
We previously analyzed the JAK2, CSF3R, CALR, SETBP1, and ETNK1 genes in 10 patients with idiopathic leukocytosis (EHA20). To elucidate the relevance of genetic alterations, we extended the analysis with 17 genes known to be involved in hematological neoplasms in 16 patients with idiopathic leukocytosis.
Methods
Leukocytosis is defined as a total white blood cell count more than two standard deviations above the mean, or a value greater than 11,000/μL. Those patients who satisfied the following criteria were included in the study: leukocytosis (predominantly neutrophils); the absence of apparent causes of leukocytosis; and documentation of the leukocytosis over a prolonged period of time. The period of observation was 1 year or longer in most patients. Sixteen patients with idiopathic leukocytosis were analyzed in the study. Neutrophils or mononuclear cells were collected after obtaining written informed consent from the 16 patients. Neutrophils from peripheral blood were purified by dextran sedimentation followed by hypotonic lysis and centrifugation with Ficoll-Conray. Mononuclear cells were isolated from bone marrow by Ficoll-Conray gradient centrifugation. Genomic DNA was extracted using the QIAamp DNA blood mini kit (Qiagen, Valencia, CA, USA). Mutations within hot spots of the CSF3R, JAK2, CALR, SETBP1, ETNK1, CBL, TET2, ASXL1, EZH2, IDH1/IDH2, DNMT3A, U2AF1, and CEBPA genes were analyzed by direct sequencing in both directions using a 3730xL DNA Analyzer (Life technologies, Carlsbad, CA, USA) and/or allele specific polymerase chain reaction analysis. Total RNA extraction and reverse transcriptase polymerase chain reaction (RT-PCR) were performed between the ETV6 and ABL1 genes in 10 patients. BCR/ABL1 gene was analyzed by RT-PCR or fluorescence in situ hybridization in 8 patients. The current study was conducted within the guidelines and with the approval of ethical committee.
Results
JAK2 V617F mutations were found in one of the 16 patients with idiopathic leukocytosis. No mutations were found in the other genes in the 16 idiopathic leukocytosis patients. ETV6-ABL1 fusion gene was detected in one of the 10 patients. No BCR/ABL1 fusion gene was detected in the 8 patients. One idiopathic leukocytosis patient with JAK2 V617F mutation has developed PV. Another patient with sustained leukocytosis for 20 years showed cytogenetic abnormalities during observation and has developed Philadelphia chromosome negative CML (Ph-CML). ETV6-ABL1 fusion gene was detected in this patient. Another patient with normal karyotype progressed to blast crisis of Ph-CML accompanied by cytogenetic abnormalities. Of the remaining 13 patients with idiopathic leukocytosis, one resolved the disease and twelve had a stable disease.
Conclusion
Idiopathic leukocytosis comprises heterogeneous conditions. JAK2 mutations and cytogenetic abnormalities are predictive of progression to hematological neoplasms.
Session topic: 15. Myeloproliferative neoplasms - Biology
Keyword(s): Myeloproliferative disorder, mutation analysis, Hematological malignancy, Cytogenetic abnormalities