Broadly defined, the research area of this project is a cytogenetic study of the inhibition of topoisomerase l (topo l) in human lymphocytes. This topic addresses not only the basic cell process of maintenance of correct DNA topography, but also has implications for cancer induction and treatment. Preliminary studies seem to indicate a similar pattern of induction of chromosomal alterations with inhibition of topo l and that observed in patients with Bloom Syndrome (BS), a genetic disease with DNA processing abnormalities and strong predisposition to cancer. This study is designed to use human lymphocyte cultures because they provide a synchronous cell population (at least initially) and are easily monitored by end point measurements of chromosomal aberrations (CA) and sister chromatid exchanges (SCE). In an effort to gain information concerning the role(s) of topo in maintaining the physical integrity of DNA as it is opened up in transcription and replication, a comparative study of the effects upon these end point measurements (CA and SCE) resulting from inhibition with camptothecin (CPT) at different phases of the cell cycle will be undertaken. The effects upon the progression of cells through the cell cycle will also be measured by using the end-point measurement of Proliferation Index. Preliminary studies indicate that such an approach is feasible and has the potential to give informative data that may be useful in understanding the activity of topo l in the cell. The preliminary studies by the principal investigator indicating a marked similarity in the cytogenetic changes induced by CPT inhibition of topo l in human lymphocytes treated in S phase of the cell cycle to the cytogenetic pattern observed in lymphocytes from patients with BS will be further studied. The cytogenetic features shared in common include a high incidence of chromosome breakage, presence of allocyclic chromosomes, high SCEs, and the induction of putative homologous interchanges. While data obtained from G banding of these induced interchanges seems to mimic the unique pattern of specific chromosomes involved in homologous interchanges in BS patients, a much quicker and more certain testing would use the technique of chromosome painting by fluorescent in situ hybridization (FISH) with known probes for the identification of the particular chromosomes involved in the interchanges. One of the major thrusts of this proposed project is to study the conditions which optimize the formation of these apparent homologous chromosome interchanges and then to identify the particular chromosomes involved by the FISH technique.
NIH ExPorter
NIH ExPorter