Induction of cyclin E and inhibition of DNA synthesis by the novel acronycine derivative S23906-1 precede the irreversible arrest of tumor cells in Sphase leading to apoptosis

S23906-1 is a diester derivative of 1,2-dihydrobenzo[b]acronycine with an u nknown mechanism of action. This cytotoxic compound was 20-fold more potent than acronycine in inhibiting the proliferation of six tumor cell lines. U sing a clonogenic assay of cell survival, the HT29 human colon carcinoma ce ll line was 100-fold more sensitive to S23906-1 than acronycine. Cell cycle analysis, by flow cytometry, showed that S23906-1 induced a partially reve rsible arrest of HT29 cells in G(2)+M at 1 muM and below and an irreversibl e arrest in S phase at 2.5 muM and above. These cell cycle effects were fol lowed by cell death through apoptosis, quantified by annexin-V labeling. In hibition of DNA synthesis was observed by complete prevention of bromodeoxy uridine (BrdU) incorporation after only 4 h of incubation with 5 muM S23906 -1. Interestingly, under the same experimental conditions, a significant in crease of cyclin E protein level was observed without any modification of c yclins D1, D2, D3, or A. This overexpressed cyclin E protein was not comple xed with Cdk2, as shown by western blotting for Cdk2 in immunoprecipitates of cyclin E. Similar inhibition of BrdU incorporation and elevation of cycl in E protein were observed after treatment with cytosine arabinoside, which reversibly inhibited progression into S phase, but not after DNA damage in duced by cisplatin. S23906-1 thus has a novel mechanism of action. A cell l ine resistant to S23906-1 showed that overexpression of cyclin E was implic ated in the novel cytotoxic activity of this compound.