The Cyclin Dependent Kinase 2, 7 and 9 Inhibitor SNS-032 Has Single Agent Activity in Acute Myeloid Leukaemia Cells and Is Highly Synergistic with Cytarabine.

Abstract 1059

Poster Board I-81

Cyclin dependent kinases (CDKs) are a family of serine/threonine kinases that require activation by binding to cyclin partners. These complexes of CDK-cyclin play an essential role in the regulation of cell cycle progression (e.g. CDK2) and some have also been implicated in the regulation of mRNA transcription (e.g. CDK9). CDK7 is involved in both these processes. Cyclins, CDK complexes and other cell cycle regulators have been shown to be mechanistically involved in the development of human tumours making them tractable drug targets. SNS-032 (BMS-387032) is a 2-aminothiazole derivative that has been shown to function as a selective inhibitor of CDK2, 7 and 9. Blocking the action of CDK2 and 7 is thought to block cell cycle progression in tumour cells. Additionally blocking CDK7 and 9 may also reduce transcription of survival factors in the tumour cells by abrogating their ability to promote transcription. Primary mononuclear cells isolated from acute myeloid leukaemia patients at diagnosis (n=87) were treated with SNS-032 for 48h resulting in a mean LD₅₀ of 0.14μM ± 0.2 as measured by MTS assay. By comparison, cytarabine (AraC) was more than 35 times less potent in the same cohort (mean LD₅₀ = 4.88μM ± 5.02). In combination with AraC (ratio SNS-032 1:1.9 cytarabine) showed a mean combination index (CI) of 0.13 in the 25 primary samples tested indicating strong synergism between SNS-032 and AraC. SNS-032 induces apoptosis as evidenced by dose-dependent increases in AnnexinV/propidium iodide (PI) staining and caspase-3 activation. SNS-032 affects activity of CDK 2 and 9 at the expression level as quantitative PCR showed that levels of CDK 2 and 9 mRNA were decreased at six hours of treatment to SNS-032 (CDK2 to 20% untreated levels (0.26μM), CDK9 to 36% untreated levels) but CDK7 mRNA levels were not significantly altered. However, CDK7 phosphorylation was reduced at 6 hours suggesting the SNS-032 modulates CDK7 at the post-translational level by suppressing activation. Treatment of the cell lines with the combination of SNS-032 and AraC resulted in reduced mRNA levels of CDK2 and 9. In contrast, AraC alone did not induce these reductions in mRNA expression. Additionally, the pro-survival proteins XIAP and Bcl-2 and Mcl-1 mRNA levels were decreased in SNS-032 treated cells, increased in AraC treated cells and reduced in cells treated with the combination. In conclusion, SNS-032 is effective in AML cells in vitro as a single agent and has also demonstrated a remarkable degree of synergy with AraC. One putative mode of action of this synergy is the SNS-032-induced decrease in pro-survival proteins. This may sensitise AML cells to the actions of cytotoxic agents LIKE AraC leading to the levels of synergy seen in this study.