Imaging: Glowing success
Researchers have found a bioluminescent method of monitoring the in vivo efficacy of drugs designed to inhibit the cyclin-dependent kinase CDK2.
Although many targeted anticancer drugs prove effective when tested in biochemical assays, it is another matter to determine how they affect their molecular targets in vivo. In the June issue of Nature Medicine, William Kaelin's lab reports a bioluminescent method of monitoring the in vivo efficacy of drugs designed to inhibit the cyclin-dependent kinase CDK2.
CDK2 regulates cell-cycle progression and is therefore a potential anticancer-drug target. Among its many substrates, it phosphorylates the cyclin-dependent kinase inhibitor p27 (also known as KIP1). This phosphorylation leads to ubiquitylation and eventual proteolytic degradation of p27. Kaelin's group created a plasmid vector that expressed a fusion of p27 and the enzyme luciferase, which can be tracked in vivo with bioluminescent imaging. This protein, termed p27Luc, behaves in a manner similar to p27 and can therefore be used as a marker of CDK2 activity.
The authors transfected several tumour cell lines with the p27Luc-expressing vector. They observed that the luciferase activity in these cells increased when cells were treated with CDK2-inhibitory proteins, peptides or small inhibitory RNA, meaning that p27Luc was no longer degraded. Treatment of p27 Luc-expressing cells with CDK2-inhibitory drugs such as flavopiridol and R-roscovitine caused a dose-dependent increase in luciferase activity in these cell lines, whereas they had no effect in cells that simply expressed the luciferase gene.
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But can this system be used to monitor CDK2 activity in vivo? Kaelin's group injected p27Luc-expressing lung carcinoma cells subcutaneously into nude mice, and imaged the resulting tumours 6 weeks later (see figure). They observed that flavopiridol induced p27Luc-mediated luminescence at the xenograft site. A particularly useful feature of this approach is the ability to take repeated measurements over extended time periods, allowing analysis of tumour growth and spread without sacrificing animals. So, luciferase reporters will be useful in monitoring the pharmacokinetics of CDK2 inhibitors, as well as other targeted therapeutics.
Kristine Novak
References
- Zhang, G. -J. et al. Bioluminescent imaging of Cdk2 inhibition in vivo. Nature Med. 10, 643–648 (2004)
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