Contributions
Abstract: PB1666
Type: Publication Only
Background
Acute myeloid leukemia (AML) is a heterogeneous clonal disorder with the presence of diverse genetic abnormalities in hematopoietic stem cells. The most frequent alterations in normal karyotype AML (NK AML) are mutations in exon 12 of nucleophosmin gene (NPM1). Until now 56 different mutations of NPM1 exon 12 have been described, mostly insertions. The NPM protein plays an important role in cell cycle and apoptosis control. It cooperates with several proteins, among them with p53 and ARF. The median levels of functional nuclear p53 protein are reduced in NPM1 and FLT3 ITD mutant samples. TP53 encodes a tumor suppressor protein which consists of transactivation, DNA-binding and oligomerization domains. Due to alternative splicing it may exist in 13 different isoforms. Alternative splicing of intron 9 leads to production of 2 different proteins, p53β and p53γ, without oligomerization domain (stop codon is localized in exon 9b). These isoforms can be present in acute myeloid leukemia (AML) cells. p53β binds to BAX promoter and can induce apoptosis independent from p53 wt. p53 has influence on activation of CEBPA which is associated with cell cycle regulation, especially cell cycle arrest and plays also role in cell differentiation. Generally, it is a transcription factor expressed during myeloid lineage development, from progenitor cells to mature granulocytes. Various mutations of CEBPA gene are described. Among them N-terminal and C-terminal mutations, mostly insertions and deletions, are often present.
Aims
The goal of the study was to assess mutational status of NPM1, CEBPA and FLT3 in association with TP53beta and TP53gamma expression levels.
Methods
75 NK AML patients were included in the study. NPM1, CEBPA and FLT3 gene mutations were analyzed by direct sequencing. TP53β and TP53γ expression levels were assessed with real time PCR. Expression levels were analyzed with ΔΔCt method, with ABL as a control gene and K562 cell line as a calibrator.
Results
In all 75 cases, TP53β and TP53γ transcripts were detected. 36 patients had NPM1 mutations, 25 had CEBPA mutations or known polymorphisms, and 25 had FLT3 ITD mutation. Assessed median expression level of TP53β was much higher (ΔΔCt 43,11) than TP53γ (ΔΔCt 10,85; p<0,05). Furthermore, expression level of TP53γ in CEBPA mutated group (ΔΔCt 11,4) was significantly lower than in CEBPA wt group (ΔΔCt 17,7) (p=0,03). We have not found any other important correlation between mutations of studied genes and TP53γ or TP53β expression. We also classified patients, according to median expression value of TP53, to two groups: with overexpression or with low expression. Haematological and clinical features, such as white blood cells count (WBC), blasts count in bone marrow or patient age did not depend on TP53 isoform expressions. However, statistical analysis showed important difference between WBC count in NPM1mutated and NPM1wt groups.
Conclusion
Obtained results may suggest a clinical importance of simultaneous analysis of TP53 isoform expression and mutations in CEBPA gene. It may be hypothesized that a changed sequence of the latter gene might influence TP53 isoform expression and in consequence regulate the cell cycle.
Session topic: 3. Acute myeloid leukemia - Biology
Keyword(s): P53, Acute Myeloid Leukemia
Abstract: PB1666
Type: Publication Only
Background
Acute myeloid leukemia (AML) is a heterogeneous clonal disorder with the presence of diverse genetic abnormalities in hematopoietic stem cells. The most frequent alterations in normal karyotype AML (NK AML) are mutations in exon 12 of nucleophosmin gene (NPM1). Until now 56 different mutations of NPM1 exon 12 have been described, mostly insertions. The NPM protein plays an important role in cell cycle and apoptosis control. It cooperates with several proteins, among them with p53 and ARF. The median levels of functional nuclear p53 protein are reduced in NPM1 and FLT3 ITD mutant samples. TP53 encodes a tumor suppressor protein which consists of transactivation, DNA-binding and oligomerization domains. Due to alternative splicing it may exist in 13 different isoforms. Alternative splicing of intron 9 leads to production of 2 different proteins, p53β and p53γ, without oligomerization domain (stop codon is localized in exon 9b). These isoforms can be present in acute myeloid leukemia (AML) cells. p53β binds to BAX promoter and can induce apoptosis independent from p53 wt. p53 has influence on activation of CEBPA which is associated with cell cycle regulation, especially cell cycle arrest and plays also role in cell differentiation. Generally, it is a transcription factor expressed during myeloid lineage development, from progenitor cells to mature granulocytes. Various mutations of CEBPA gene are described. Among them N-terminal and C-terminal mutations, mostly insertions and deletions, are often present.
Aims
The goal of the study was to assess mutational status of NPM1, CEBPA and FLT3 in association with TP53beta and TP53gamma expression levels.
Methods
75 NK AML patients were included in the study. NPM1, CEBPA and FLT3 gene mutations were analyzed by direct sequencing. TP53β and TP53γ expression levels were assessed with real time PCR. Expression levels were analyzed with ΔΔCt method, with ABL as a control gene and K562 cell line as a calibrator.
Results
In all 75 cases, TP53β and TP53γ transcripts were detected. 36 patients had NPM1 mutations, 25 had CEBPA mutations or known polymorphisms, and 25 had FLT3 ITD mutation. Assessed median expression level of TP53β was much higher (ΔΔCt 43,11) than TP53γ (ΔΔCt 10,85; p<0,05). Furthermore, expression level of TP53γ in CEBPA mutated group (ΔΔCt 11,4) was significantly lower than in CEBPA wt group (ΔΔCt 17,7) (p=0,03). We have not found any other important correlation between mutations of studied genes and TP53γ or TP53β expression. We also classified patients, according to median expression value of TP53, to two groups: with overexpression or with low expression. Haematological and clinical features, such as white blood cells count (WBC), blasts count in bone marrow or patient age did not depend on TP53 isoform expressions. However, statistical analysis showed important difference between WBC count in NPM1mutated and NPM1wt groups.
Conclusion
Obtained results may suggest a clinical importance of simultaneous analysis of TP53 isoform expression and mutations in CEBPA gene. It may be hypothesized that a changed sequence of the latter gene might influence TP53 isoform expression and in consequence regulate the cell cycle.
Session topic: 3. Acute myeloid leukemia - Biology
Keyword(s): P53, Acute Myeloid Leukemia