Abstract 1626: CDK9 inhibition enhances apoptosis of TP53 mutated AML when combined with standard chemotherapy

Abstract Background: The prognosis of patients with acute myeloid leukemia (AML) remains poor for those with mutations or deletions of the TP53 gene. Inhibiting CDK9 is a strategy to trigger apoptosis downstream of p53 with the goal of overcoming the poor prognostic nature of mutant p53. Here we test dinaciclib, a multi-CDK and potent CDK9 inhibitor, alone and with azacitidine and venetoclax to demonstrate that CDK9 inhibition as monotherapy and in combination leads to apoptosis in TP53 mutated AML. Methods: To determine the effect of inhibiting CDK9 on the cytotoxicity of AML and normal cells, we treated p53 mutated AML cell lines (THP1, NOMO1, and U937), wild type (WT) p53 AML cell lines (MV4-11 and HL-60) and peripheral blood mononuclear cells (PBMCs) from healthy donors with azacitidine, venetoclax, and dinaciclib. We performed viability assays to examine cell death and determine the IC50 and optimal exposure time of these drugs. Apoptosis was measured through flow cytometry with Annexin-V and 7-AAD staining, and through the quantification of apoptotic proteins with immunoblotting. Results: Treatment of mutant and WT p53 AML cell lines with azacitidine, venetoclax, and dinaciclib as monotherapy demonstrated a dose-dependent decrease in cell viability. After testing various concentrations and exposure times of these drugs, we determined concentrations of 200nM azacitidine, 6000nM venetoclax, and 10nM dinaciclib at an exposure time of 8-hours were highly effective with dinaciclib monotherapy and in the triple-drug combination. When used in the mutant p53 cell line, THP1, and WT p53 cell line, MV4-11, the addition of dinaciclib to azacitidine and venetoclax showed increased synergy (scores of 29.6 and 12.9, respectively, using the HSA reference model) compared to azacitidine and venetoclax alone (synergy scores of -0.98 and 1.4, respectively). The addition of dinaciclib to THP1 cells resulted in increased apoptosis (55.2%) when compared to the two-drug combination of azacitidine and venetoclax (17.0%) as measured by the quantification of c-PARP expression normalized to beta-actin through immunoblotting. When this drug combination was tested in PBMCs of healthy donors, they were notably less sensitive and had lower synergy scores (1.2) than the AML cell lines. Conclusions: We have shown that the addition of a CDK9 inhibitor to azacitidine and venetoclax results in enhanced cytotoxicity of mutant p53 and WT p53 AML cell lines at low nanomolar concentrations. These results suggest that inhibition of CDK9 sensitizes AML cells independently of TP53, with less pronounced cytotoxicity in normal PBMCs of healthy donors. We plan to further examine the effect of CDK9 inhibition using a highly selective CDK9 inhibitor and validate these results with primary AML and normal bone marrow samples. Ultimately, we hope these experiments will guide the development of human clinical trials to benefit those patients with p53 mutated AML. Citation Format: Elizabeth C. Trull, David B. Sykes, Philip C. Amrein. CDK9 inhibition enhances apoptosis of TP53 mutated AML when combined with standard chemotherapy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1626.