Thursday, April 3, 2025
4/3/2025
Dissecting mechanistic differences between EZH2 gain-of-function events in B-cell lymphoma - Project Summary Non-Hodgkin Lymphoma (NHL) is one of the most common cancers in the United States, with approximately 80,000 new cases and 20,000 deaths occurring each year. The two most common types of NHL are Diffuse Large B Cell Lymphoma (DLBCL) and Follicular Lymphoma (FL). Up to 30% of germinal center- derived (GC)-DLBCL and FL cases exhibit gain-of-function events in the histone methyltransferase EZH2, including a heterozygous hotspot mutation (EZH2Y641F/+) or overexpression of wild-type protein (EZH2OE). This proposal addresses the major gaps in knowledge of (1) whether EZH2Y641F/+ and EZH2OE are functionally equivalent or distinct gain-of-function events and (2) whether they drive lymphoma as early or late events in the B lineage. We hypothesize that EZH2Y641F and EZH2OE are non-equivalent gain-of-function events that drive lymphoma by distinct mechanisms, and that the timing of each event influences its oncogenic activity. The first aim of this proposal investigates the impacts of EZH2Y641F/+ and EZH2OE in B cells. To achieve this goal, we will use genetically engineered mouse models of EZH2Y641F/+ and EZH2OE driven by the B cell- specific CD19-Cre and employ a combination of flow cytometric analysis of key GC phenotypes (i.e. proliferation, apoptosis, and differentiation), gene expression analysis by RNA sequencing, and chromatin profiling of histone methylation and EZH2 binding with CUT&Tag sequencing. These mechanistic studies will identify downstream targets that may function as biomarkers or lead to new therapeutic approaches in patients with these alterations. The second aim of the proposal is to determine whether EZH2 alterations drive lymphoma as early or late events in the B lineage. To achieve this goal, we will model EZH2Y641F/+ and EZH2OE in the early B lineage using CD19-Cre and in the later GC stage using Cγ1-Cre, overexpress the lymphoma oncoprotein BCL2 in bone marrow from each cohort, and transplant the cells into lethally irradiated recipient mice. The rates of lymphoma onset will be determined by monitoring each recipient group for disease and analyzing the resulting tumors by flow cytometry and histologic analysis. This aim will address a gap in knowledge about whether EZH2Y641F/+ and EZH2OE represent early or late events in DLBCL or FL pathogenesis. The proposed research will take place at Washington University in St. Louis, an outstanding environment to support the trainee’s development for a research career in cancer epigenetics. By completing the research tasks, the trainee will acquire or master technical skills including multi-channel flow cytometry and panel design, analysis and interpretation of genomic sequencing data, and mouse bone marrow transplantation, among others. The trainee will publish the results in a first-author manuscript and will present this work at national and international conferences, which will develop the trainee’s skills of scientific writing and communication. Finally, the trainee will mentor undergraduate students during the proposed research to develop the skill of mentorship.
HHS’ Tracking Accountability in Government Grants System (TAGGS) website provides access to detailed descriptions of grants, loans, aggregated direct payments and other types of financial assistance awarded by the HHS Operating Divisions (OPDIVs) and the Office of the Secretary Staff Divisions (STAFFDIVs).