The Clinical Significance of Death-Associated Protein Kinase(DAP-Kinase)Expression in Leukemia

• Main Authors: Chen, Hui-Chen, 陳蕙珍
• Other Authors: 林亮音
• Format: Others
• Language: zh-TW
• Published: 2003
• Online Access: http://ndltd.ncl.edu.tw/handle/76960019634844462636

碩士 === 國立臺灣大學 === 醫事技術學研究所 === 91 === Abstract Death—associated protein kinase (DAP-Kinase) is a Ca2+ / Calmodulin -dependent, cytoskeletal-associated serine / threonine protein kinase, which has death-inducing functions. DAP-Kinase involves in the apoptosis which was induced by IFN-γ, Fas Ligand, TNF-α and TGF-. In the recent studies, loss of DAP-Kinase expression was noted in various cancer cell lines as a result of hypermethylation in the promoter region. On the other hand, it also conveys selective advantage during the transition of lung carcinoma cells from a low-metastastic to a more aggressive high-metastastic phenotype. These findings imply the ability of DAP-Kinase in tumor suppression and metastasis. Although efforts had been made, the information about the relationship between tumor formation and DAP-Kinase was not enough. Here, we perform a large-scale screening by using several methods to analyze the expression status of DAP-Kinase in leukemic cell lines, normal blood cells and different leukemias. We hope these studies can help us to clarify the relationship between leukemia and DAP-Kinase. First of all, we wanted to understand whether leukemic cell lines had aberrant methylation in the promoter region on DNA. The results of the Methylation-Specific PCR (MSP) indicated that only Raji cell had methylated copies. The quantitative data from Reverse-Transcriptase PCR (RT-PCR) and Real-Time PCR showed all the leukemic cell lines had the expression of mRNA except Raji cell. The expression level of mRNA in leukemic cell lines was: U937 > K562 > Jurkat > HL-60. On the other hand, we also try to detect the protein expression in the Jurkat cell by immunocytochemical staining. The result showed that Jurkat cell did express the DAP-Kinase protein. According to the analysis of MSP, RT-PCR and Real-Time PCR, normal bone marrow cells with unmethylated copies had the expression of mRNA. The result of immunocytochemical staining showed the protein expression level was extremely low without any apoptotic stimuli. We found that there was no protein expression in three acute myelocytic leukemia (AML) patients who had M1, M3 and M4 subtypes respectively. The mRNA quantitative study showed 9 fresh AML patients had overexpression of mRNA. Only 2 patients who had M1 and M3 subtypes respectively had methylated copies among 53 cases. In the similar situation, there was no protein expression in acute lymphocytic leukemia (ALL) patients studied, one fresh and the other refractory. All fresh and remission ALL patients had mRNA overexpression as fresh AML patients did. Those patients only had unmethylated copies. Analysis of 52 myelodysplastic syndrome (MDS) patients indicated that 4 of the 7 chronic myelomonocytic leukemia (CMMoL) patients showed aberrant methylation. There were also 2 other cases had methylated copies, one had refractory anemia (RA), and the other had RA with progression to RA with excess blasts (RAEB). Our findings suggested there were no significant hypermethylation of DAP-Kinase in AML and ALL. Although leukemia patients would have the overexpression of mRNA under stimuli, they had no obvious protein expression. Finally, it seemed that the hypermethylation of DAP-Kinase was highly correlated to the CMMoL.