PS917 SAFE TARGETING OF T-CELL ACUTE LYMPHOBLASTIC LEUKEMIA BY SUBUNIT SELECTIVE GAMMA-SECRETASE INHIBITION

Abstract

Background: SPI1 fusion genes are recurrently found in Early Thymic Precursor cell Acute Lymphoblastic Leukemia (ETP-ALL) cases and represent a distinct subgroup with high frequency of RAS mutations and dismal prognosis. SPI1 (PU.1) is an important transcription factor in hematopoietic cells and its dysregulation contributes to several types of leukemia. Forced overexpression of SPI1 in T-cell progenitors is known to inhibit T-cell differentiation. Conversely, knock-out of TCF7, which is highly expressed in the earliest thymic progenitors, also leads to a T-cell differentiation block. However, the role of SPI1 fusions in driving and maintaining T-ALL remains unknown. Aims: In this study we aimed to elucidate the role of the TCF7-SPI1 fusion and its potential collaboration with RAS mutation in driving and maintaining T-ALL. Methods: We performed whole genome, transcriptome and nanopore sequencing on a patient sample with the TCF7-SPI1 fusion. Primary mouse pro-T cells were cultured on Dll4-coated plates in the presence of stem cell factor (Scf) and interleukin-7 (Il7). In vivo bone marrow transplant (BMT) assays were performed using hematopoietic stem and progenitor cells (HSPCs) isolated from CD2-Cre C57BL/6J mice and transduced with retroviral vectors for inducible expression of the TCF7-SPI1 fusion and/or NRAS (G12D) mutant within developing T-cells of recipient mice. Results: Combined short read and long read RNA-sequencing characterized the exon structure of the TCF7-SPI1 fusion transcript and confirmed the presence of a NRAS (G12D) mutation in the patient sample. Expression of TCF7-SPI1 and NRAS (G12D) transformed pro-T cells to Scf and interleukin-7 (Il7) independent growth. Using an inducible in vivo BMT model, conditional expression of TCF7-SPI1 and NRAS (G12D) within developing T-cells resulted in an aggressive immature acute T-ALL characterized by hyperleukocytosis (CD4-/CD8-, CD3+, CD117+), splenomegaly but no thymus enlargement. This was in contrast to either conditional TCF7-SPI1 alone that did not develop leukemia, or with NRAS (G12D) alone that developed a mature CD4+/CD8+ lymphoma with normal white blood cell count and no peripheral leukemic invasion of the bone marrow and spleen. Notably, a deletion construct of TCF7-SPI1, lacking the N-terminal beta-catenin binding domain of TCF7 lost its oncogenic properties. Summary/Conclusion: Here, we show that TCF7-SPI1 cooperates with NRAS (G12D) to drive an aggressive immature T-cell leukemia in vivo that is dependent on the N-terminal beta-catenin binding domain of TCF7.