Abstract
MJ and JF provided equal contribution as senior authors
Acute myeloid leukemia (AML) is associated with poor survival and characterized by an accumulation of immature myeloid blasts in the bone marrow. To efficiently target AML, new therapies directed to leukemia stem cells (LSCs), a self-renewing population that constitutes a chemo-resistant reservoir responsible for disease relapse, are warranted.
Cyclin-dependent kinase 8 inhibitors' (CDK8i) anti-cancer activity has been demonstrated in human acute myeloid leukemia (AML) cell lines. Efficacy has been associated with activation of super enhancer regions and reduction of STAT5 S726 phosphorylation in sensitive cells (Pelish, Nature, 2015). SEL120-34A (Selvita, Poland), a selective low nanomolar CDK8/CDK19 inhibitor, has been shown to have antileukemic effect in a panel of AML cell lines (Rzymski, Oncotarget 2017).
To evaluate whether primitive AML cells, enriched for LSCs, are sensitive to CDK8 inhibition, we tested the CDK8i SEL120-34A and Senexin B on TEX cells, an AML cell line that exhibits a hierarchical organization and can be used as a LSC model (Warner, Leukemia, 2005). Treatment of TEX cells with SEL120-34A or Senexin B resulted in an inhibition of cell growth (IC50 of 8 and 31 nM respectively at 10 days of culture), associated with reduced activation of STAT5 and STAT1. Both STAT proteins were previously identified as biomarkers for SEL120-34A activity (Rzymski, Oncotarget 2017). In addition, RNA sequencing followed by gene-set enrichment analysis (GSEA) revealed a loss of a LSC signature.
To functionally address the effects of CDK8i on LSCs, we used a murine dsRed+ AML model driven by retroviral MLL-AF9 expression. This model has a well-defined LSC population and initiates AML with a short latency, enabling rapid follow-up experiments in syngeneic hosts. Treatment of c-Kit+ AML cells with SEL120-34A or Senexin B in vitro resulted in strong inhibition of leukemia cell growth (IC50 of 119 and 143 nM, respectively, at 7 days of culture), associated with increased apoptosis and reduced cycling of the cells. To address the in vivo therapeutic efficacy of SEL120-34A, mice injected with AML cells 10 days earlier were treated orally for 12 consecutive days using doses of 20 and 40 mg/kg before sacrificed. No tolerability issues were observed with mice maintaining a stable weight through the treatment. Whereas the control group had 87% leukemia cells in the peripheral blood at the end-point analysis, SEL120-34A treated animals showed a dose-dependent selective anti-leukemic activity with a corresponding 78% (20 mg/kg) and 67% (40 mg/kg, p=0.011) of leukemia cells. Similarly, a significant selective dose-dependent anti-leukemic activity of SEL120-34A was observed also in the bone marrow. In addition, a dose-dependent reduced white blood cell count and smaller spleen size upon SEL120-34A treatment was observed, demonstrating that CDK8 inhibition has selective anti-leukemic activity in vivo. Notably, SEL120-34A treatment resulted in granulocytic (Gr1+CD11b-) differentiation of the AML cells (5.8% of the AML cells in the control group; 21.9% at 20 mg/kg, p=0.03; and 22.3%, p=0.0037 at 40 mg/kg). Moreover, SEL120-34A treatment resulted in strong inhibition of Stat5 S726 and Stat1 S727 phosphorylation in AML bone marrow cells harvested from the mice.
To test the efficacy of CDK8 inhibition on AML patient cells, four AML patient derived xenografts were treated with SEL120-34A ex vivo. In all four samples, two of which contained activating mutations in FLT3, SEL120-34A treatment resulted in a significant antileukemic activity with decreased number of AML cells and an increase in apoptotic cells.
Taken together, our data from murine and human AML models indicate that CDK8 inhibition has therapeutic efficacy in primitive AML cells. SEL120-34A treatment in vivo resulted in reduced leukemia cell burden in both blood and bone marrow accompanied by granulocytic differentiation. Treatment of AML cells in cultures consistently resulted in a reduction in AML cell number and increased apoptosis. Ongoing and future experiments will address whether SEL120-34A treatment also extends survival of mice with AML in syngeneic and patient-derived xenograft models. These data highlight CDK8 as a promising therapeutic target in AML and provides preclinical proof of concept for anti-leukemic efficacy of the clinical candidate SEL120-34A in relevant AML models.
Disclosures
Mazan: Selvita S.A.: Employment. Majewska:Selvita S.A.: Employment. Wiklik:Selvita S.A.: Employment. Combik:Selvita S.A.: Employment. Masiejczyk:Selvita S.A.: Employment. Fiedor:Selvita S.A.: Employment. Obacz:Selvita S.A.: Employment. Bialas:Selvita S.A.: Employment. Chesy:Selvita S.A.: Employment. Gabor-Worwa:Selvita S.A.: Employment. Brzózka:Selvita S.A.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Rzymski:Selvita S.A.: Employment, Equity Ownership. Fioretos:Cantargia: Equity Ownership, Membership on an entity's Board of Directors or advisory committees.
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