Purification of DNA topoisomerase I from the spleen of a patient with non-Hodgkin's lymphoma.

DNA topoisomerase I (topo I) was purified 124-fold from the spleen of a patient which was diffusely infiltrated by malignant lymphoma. Extracts prepared from the lymphoma tissue demonstrated elevated topo I activity. The purification uses a new two step chromatographic method involving hydroxylapatite and Fast Protein Liquid Chromatography mono S. The purified enzyme has a molecular weight of 67 kilodaltons as determined by sodium dodecyl sulfate denaturing gel electrophoresis. Western blotting confirms the identity of the protein as topo I. The purified lymphoma topo I has a similar specific activity as topo I isolated by the same procedure from normal placenta. By using a new quantitative radiolabeled DNA relaxation assay, it was estimated that each molecule of topo I is able to relax 9.6 molecules of supercoiled DNA per min at 30C. In addition it was found that both the placental topo I and the lymphoma topo I were each 50% inhibited by 8 microns camptothecin and that the drug stimulates the lymphoma topo I to cleave supercoiled plasmid DNA. These findings indicate that the elevation of topo I measured in crude extracts of this human malignant lymphoma can be entirely accounted for by the increase in topo I protein in the tumor. Furthermore, the sensitivity of the lymphoma topo I towards camptothecin suggests that therapy with topo I directed agents might be beneficial in this tumor.