Understanding the tumor evolution of PAX5-driven B-cell leukemias

• Applicant: Dr. Franziska Auer
• Subject Area: Hematology, Oncology ; Pediatric and Adolescent Medicine
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 547295412

PAX5 is a well-known master regulator for B-cell development. Consequently, somatic PAX5 deregulation is the most frequent feature present in B-cell precursor acute lymphoblastic leukemia (BCP-ALL) – the most common hematological malignancy in childhood. Here, PAX5alt and PAX5 P80R have recently been identified as two novel PAX5-driven subtypes. Moreover, others and I could previously show that predisposing PAX5 germline variants can act as an initiating factor for leukemia development. However, the mechanisms, which direct the stepwise progression from a cell with deregulated PAX5 expression - via pre-leukemia - to leukemia, remain elusive. My preliminary data in Pax5 heterozygous mice show that PAX5 reduction affects B-cell differentiation and proper B-cell receptor light chain rearrangement and creates a susceptible precursor population in the bone marrow, which displays high Myc expression at the pre-leukemic stage. Thus, my central hypothesis is that PAX5 alterations cause a pre-leukemic state with abnormal B-cell differentiation, which evolves to BCP-ALL throughout distinct transformation stages. Therefore, in Aim 1 of this proposal we will characterize the B-cell differentiation in mouse models of germline (p.G183S) and somatic (p.P80R) PAX5-driven leukemias to identify pre-leukemic precursor B-cell subsets prone for malignant transformation. This will be carried out by flow cytometry as well as single-cell RNA-Sequencing (scRNA-Seq) analyses, coupled with in-vitro cell culture assays. Aim 2 will focus on the generation and elucidation of two newly generated mouse models, which mimic the stepwise tumor evolution of PAX5-driven BCP-ALLs, by combining the Pax5 variants from Aim 1 with an inducible deletion of the remaining Pax5 wild type allele. Based on these models, transcriptional, epigenetic, signaling as well as metabolomic changes will be assessed to elucidate mechanistic insights of gradual PAX5 reduction. To complete the spectrum of malignant transformation, additional p19ARF tumor suppressor deletion and activating oncogenic mutations in Jak3 or Nras will be introduced in-vitro. Lastly, Aim 3 will depict the similarity between germline and somatic PAX5-driven BCP-ALLs. Although families harboring predisposing PAX5 germline variants are extremely rare, I have access to the material of multiple patients. Thus, we will perform scRNA-Seq analyses and compare the obtained data to somatic PAX5-driven BCP-ALLs to identify common denominators. Consequently, these patient derived models will aid in understanding somatic PAX5 alterations and their role in >10% of BCP-ALL across all ages. Together, all three aims will complement each other for the overall goal to identify new avenues for strategic intervention in PAX5-driven B-cell leukemias.

• DFG Programme: Research Grants