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Method for quantifying expression of functionally active topoisomerase II in patients with leukaemia
A R Cattan1, D Levett, E A Douglas
1Department of Haematology, University of Newcastle upon Tyne, Royal Victoria, Infirmary.
Journal of Clinical Pathology
|October 1, 1996
Summary
Researchers developed a method to measure active topoisomerase II in human cells. Topoisomerase II activity was higher in leukaemic cells than normal cells, with no change observed between initial diagnosis and relapse.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Topoisomerase II is a crucial enzyme involved in DNA replication and repair.
- Dysregulation of Topoisomerase II activity is implicated in various cancers, including leukemia.
- Accurate quantification of enzymatically active Topoisomerase II is essential for understanding its role in disease.
Purpose of the Study:
- To establish a reliable method for measuring and quantifying enzymatically active Topoisomerase II.
- To compare Topoisomerase II activity in normal human cells versus neoplastic cells.
- To investigate changes in Topoisomerase II activity in acute lymphoblastic leukemia (ALL) during disease progression.
Main Methods:
- Preparation of crude cell lysate from mononuclear cells (peripheral blood or bone marrow).
- Decatenation of kinetoplast DNA minicircles using the cell lysate.
- Separation and visualization of DNA by agarose gel electrophoresis and ethidium bromide/UV fluorescence.
Main Results:
- Established optimal conditions (cell number, sample volume, drug concentration) for reproducible assay results.
- Quantified active Topoisomerase II in matched samples from ALL patients (presentation and relapse).
- Developed a unit scale to compare Topoisomerase II activity in leukemic and normal cells.
Conclusions:
- Topoisomerase II activity did not significantly change between initial diagnosis and relapse in ALL patients.
- Leukaemic blast populations exhibited significantly higher Topoisomerase II activity compared to normal cells.
- The developed method allows for the quantification and comparison of active Topoisomerase II in different cellular contexts.