Molecular Characterization of elF4B

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Diffuse large B-cell lymphoma (DLBCL) represents the most common subtype of non-Hodgkin lymphoma (NHL), accounting for about 40% of all newly diagnosed cases in the United States. Despite the relative success of upfront R-CHOP therapy, the frequent occurrence of relapsed/refractory cases and the limitations to treating patients with co-morbidities has provided the impetus to discover novel actionable molecular targets to improve outcomes. Two of these rewired metabolic gene signatures of fatty acid synthase (FASN) & LKB1 (Liver Kinase B1) and the protein translational machinery components are emerging as putative candidates for therapeutic intervention in DLBCL. Supported by robust data from several independent laboratories, identification of unique molecular, metabolic signatures such as perturbation in fatty acid metabolism and depletion of LKB1 signaling are associated with DLBCL survival and underlying lymphomagenic mechanism(s). However, limited success in the clinical and pre-clinical arena targeting FASN and LKB1 due to pharmacological limitations has motivated investigators to identify downstream effectors, providing alternative actionable candidates with applicability in suppressing heterogenous DLCBL tumor growth. To point, exciting preliminary data from our lab identified eIF4B (Eukaryotic initiation factor 4B), a critical translational machinery component, is activated by enhanced FASN activity. It is broadly accepted that unregulated FASN activity is strongly correlated with multi-drug resistance, a prominent feature observed in patients with R-CHOP resistant disease. Further, we reported that FASN (an essential enzyme in the altered metabolome of DLBCL) directly regulates PI3K/S6Kinase mediated USP11 (Ubiquitin Specific Protease 11) driven eIF4B activity, which enhances oncogene expression in DLBCL. Unfortunately, there are no available murine models to delineate their physiological roles in B-cell development and homeostasis. Another critical question raised from our earlier published work was interrogating the molecular partners associated with the eIF4B ubiquitination machinery. Preliminary findings revealed PARK2 as a potential E3 Ligase, polyubiquitinating eIF4B. Further expanding our observation of the rewired metabolic impact on eIF4B driven translation, we found that LKB1 phosphorylates eIF4B, hindering eIF4B- sensitive gene expression. To address these findings in-depth, we will pursue the following three specific aims: Specific Aim 1: Define the molecular role of PARK2 in polyubiquitination of eIF4B and DLBCL proliferation, Specific Aim 2: Determine the impact of LKB1 activity on eIF4B-dependent translation and Specific Aim 3: Determine the molecular dependence of eIF4B in B-cells. While we have acquired compelling cell-based data demonstrating the functional importance of eIF4B in DLBCL, we will expand our mechanistic understanding by characterizing the physiological inputs of eIF4B in B-cells using engineered mouse models as well as crossing them with other clinically relevant oncogenic drivers (Myc) to assess their contribution to the development and progression of lymphomas. We have synthesized complementary mouse models (knock- out and transgenic/overexpression) to establish prima facia in vivo evidence, which will establish the pathophysiological impact of eIF4B expression in B-cell lymphomagenesis. Further, using molecular, cellular, and in vivo tools, we aim to validate how our proposed putative E3 ligase, PARK2, and the energetics kinase, LKB1, impact eIF4B functionality. Interestingly, we noted a significant positive correlation between PARK2 and LKB1 expression in the DLBCL dataset. We anticipate that the proposed studies will shed light on the functional and molecular events of eIF4B-dependent and independent roles in the ontogeny and pathophysiology of B- cells. In addition, molecular studies related to PARK2 and LKB1 will significantly enhance our understanding of this crucial post-transcriptional/translational mechanism(s) regulating eIF4B-driven lymphomagenesis. Ultimately, the successful completion of these experiments will lead to the identification and validation of novel actionable molecular target(s) in DLBCL and related lymphoid malignancies.