Session: 602. Myeloid Oncogenesis: Basic: Poster III
Hematology Disease Topics & Pathways:
AML, Acute Myeloid Malignancies, Research, Fundamental Science, Translational Research, genomics, Diseases, Myeloid Malignancies, Biological Processes, molecular biology, pathogenesis
To assess the leukemic potential of UBTF-TD in vivo, we expressed UBTF-TD in cbCD34+ cells and transplanted them into humanized NSG-SGM3 mice (hSCF hGM-CSF, IL-3) to assess the leukemic potential of UBTF-TD. We observed a rapid in vivo expansion of UBTF-TD expressing cells with an average latency of 11.7 weeks. Peripheral blood immunophenotyping of these mice identified an expansion of myeloid cells (CD33+, CD11b+) and splenomegaly compared to wild-type controls (p-value = 0.0002). These data suggest that UBTF-TD is sufficient to promote myeloid expansion in vivo.
To test whether human cbCD34+ myeloid cell expansion is dependent on UBTF-TD expression, we developed an in vitro inducible degradation model by integrating a targetable protein (FKBP12F36V) into our standard lentiviral overexpression vector. This system (dTAG) allows for rapid and direct control of UBTF-TD protein expression. Degradation is achieved by treating these cells with small molecules (i.e., dTAG-13) that recruit the FKBP12F36V-HA-UBTF-TD fusion protein to E3-ubiquitin ligase complex for ubiquitination and subsequent proteasome degradation. Our data shows that the FKBP12F36V tag has no effect on proliferation and replating capacity of UBTF-TD cbCD34+ cells. Treatment with 1 uM dTAG-13 quickly depleted FKBP12F36V-HA-UBTF-TD protein within 1 hour of treatment (fold change when compared to DMSO > 75%); this protein was completely undetectable by western blot after 4hrs. These cells displayed a reduction of CD117 surface marker expression (p-value < 0.0001) as measured by flow cytometry—highlighting a loss in stemness. Furthermore, long-term depletion of UBTF-TD resulted in decreased proliferation (p-value < 0.0001) and cell viability (p-value < 0.0001). This coincided with an increase in apoptosis as measured by Annexin V staining (p-value < 0.0001).
Collectively, our data suggest that UBTF-TD expression is sufficient to promote myeloid cell expansion and a leukemic expression profile. This study will help guide therapeutic development in AML as our data shows that directly targeting UBTF-TD for degradation is sufficient to reverse UBTF-TD myeloid cell expansion.
Umeda, M., Ma, J., Huang, B. J., Hagiwara, K., Westover, T., Abdelhamed, S., Klco, J. M. (2022). Integrated genomic analysis identifies UBTF tandem duplications as a recurrent lesion in pediatric acute myeloid leukemia. Blood Cancer Discov. doi:10.1158/2643-3230.BCD-21-0160
Disclosures: No relevant conflicts of interest to declare.
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