Potential Role of Ubiquitin E3 Ligase, FBXO9, As a Tumor Suppressor in AML

Hematopoietic stem cells (HSCs) give rise to all cells of the blood. The balance between HSC self-renewal and differentiation is critical to maintaining hematopoiesis throughout the life of the organism, and disruptions to this balance often result in malignancies, including acute myeloid leukemia (AML). Multiple transcription factor networks are known to be important regulators of HSCs and, when mutated, can lead to leukemia development. More recently, protein degradation by the ubiquitin proteasome system (UPS) has been shown to regulate HSC self-renewal, differentiation, and survival. The role of this pathway in leukemogenesis is less clear.

We recently performed a screen in embryonic stem cells to identify members of the UPS involved in cell fate decisions. From this, we identified the Ubiquitin E3 ligase FBXO9, which is a substrate recognition component of the Skp1-Cullin-F-box (SCF)-type E3 ligase complex. Interestingly, FBXO9 is highly expressed in hematopoietic stem and progenitor cells, but is decreased in patients with AML including patients with Inv(16) chromosomal abnormality. In order to study FBXO9 in normal and malignant hematopoiesis, we utilized a novel method using CRISPR/Cas9 to generate a conditional knockout model in collaboration with the Mouse Transgenic Core at UNMC. We bred our mice to transgenic mice expressing Cre Recombinase from the Mx1 promoter, which allows for PolyIC inducible expression of Cre in all hematopoietic cells including HSC. Our preliminary data shows alterations in hematopoiesis, including less competitive repopulation capability of the HSC in vivo and decreased differentiation potential of the hematopoietic progenitor population (HPC). To test if Fbxo9 plays a role in leukemia development, we crossed mice with the conditional allele of Fbxo9 with a knockin mouse model of AML that expresses the leukemia fusion gene Cbfb-MYH11 . In a small cohort of mice, we found that deletion of Fbxo9 in the context of Cbfb-MYH11 caused markedly accelerated leukemia development with >50% myeloid blasts in the peripheral blood by week 6. These findings suggest that the ubiquitin E3 ligase, Fbxo9, is a novel tumor suppressor and can regulate self-renewal, differentiation, and malignant transformation. In addition The dynamic reversibility of the ubiquitin modification (by kinases, phosphatases, E3 ligases, and de-ubiquitinases) and the recent development of both general proteasome inhibitors as well as specific antagonists of ligase function suggest that manipulation of proteins by ubiquitin E3 ligases could provide potential targets for drug discovery and stem cell therapy.