Replication synchrony-PCR: a sampling-time-independent assay for replication synchrony in human tissues and tumors in situ.
Replication synchrony within a cell population can be demonstrated by pulse-labeling followed by PCR amplification of immunoprecipitated 5-iodo-2'-deoxyuridine (IdUrd)-labeled DNA from cells of otherwise indeterminant kinetic stages. This replication synchrony-PCR approach may be valuable in understanding the dynamics of human normal tissue or solid tumor replication in situ where access for repeated sampling is severely limited. IdUrd labeling provides a sampling-time-independent method for assessing the replicative status of a cell population at the time when the label was presented. Using genes whose time of replication in S phase is already known, the presence of a cell in early or late S phase can be determined and a qualitative measure made of replication synchrony in the population. This approach was evaluated in synchronous and random cultures of Ej cells using the early replicating PGK-1 gene to identify cells in early S phase at the time of labeling and the late replicating factor IX gene to identify cells that were in late S phase. To test the feasibility of clinical application of this technique, human tumor cells from patients with advanced cancers, given IdUrd therapeutically at specified times of the day, were evaluated. In some patients, replication synchrony-PCR provided evidence of parasynchronous DNA replication in tumor cells. This technique could be appended to existing clinical studies in which BrdUrd or IdUrd is being given to patients either diagnostically or therapeutically.
De Francesco
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Department of Biology, Beckman Research Institute of the City of Hope, Duarte, CA 91010, USA.
Klevecz
R R
RR
D002455:Cell Division; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis*; D004261:DNA Replication; D006801:Humans; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods*
D004247:DNA