Introduction and Background: The “CBM” complex is composed of the scaffolding protein CARMA1, the adaptor protein BCL10, and the effector protein MALT1. This complex performs multiple pivotal functions as a mediator of antigen receptor-dependent induction of the NF-κB transcription factor and subsequent lymphocyte activation. The key downstream effector of the CBM complex, MALT1, regulates downstream signaling via both scaffolding and proteolytic functions. Inappropriate activation of MALT1, which can result from somatic gain-of-function mutation in upstream regulators of MALT1 or chromosomal translocation involving the MALT1 gene, underlies the pathogenesis of a variety of lymphoid malignancies including activated B cell type-diffuse large B cell lymphoma (ABC-DLBCL) and mucosa associated lymphoid tissue (MALT) lymphoma. We recently discovered that G-protein-coupled receptor kinase 2 (GRK2) binds to MALT1 and inhibits MALT1 scaffold and proteolytic activities. Further, we found that knockdown of GRK2 in ABC-DLBCL enhances tumor growth in vivo, suggesting that GRK2 may act as a tumor suppressor in MALT1-dependent lymphomas. Interestingly, we found that GRK2 mRNA levels are markedly lower in a subset of DLBCL cases in comparison to normal B-cell controls and that lower GRK2 level is associated with reduced survival in ABC-DLBCL patients. We thus sought to investigate how GRK2 expression is regulated in ABC-DLBCL. Methods and Results: We first investigated whether mutations/deletions in the ADRBK1 gene (GRK2) account for the lower levels of GRK2 expression in a subset of DLBCL cases. Only one mutation among 6 published DLBCL sequencing datasets (334 cases total) was identified. We next considered that other mechanisms, such as regulation by microRNA (miRNA), could play a role in downregulating GRK2. Using DICER-deficient HEK 293T cells, we demonstrated that GRK2 protein level significantly increases when miRNA processing is impaired. We then used mirDIP, a microRNA Data Integration Portal, to screen for candidate miRNAs predicted to target the 3’UTR of GRK2. We identified 4 top candidate hits, 2 of which were also identified in bioinformatic analysis as miRNA whose level inversely correlates with GRK2 mRNA level in ABC-DLBCL patient tumor specimens. Using a GRK2 3’ UTR reporter system, we confirmed direct targeting of GRK2 by these miRNAs. Next, we generated stable ABC-DLBCL cell lines (TMD8, OCI-Ly3) overexpressing candidate miRNAs. Our results demonstrate that candidate miRNA overexpression results in reduced GRK2 expression and increased MALT1 scaffolding and proteolytic activities, NF-κB transcriptional activity and ABC-DLBCL cell proliferation. Conclusions: Together, our data suggest that miRNAs down-regulate expression of GRK2 in ABC-DLBCL which in turn enhances MALT1 scaffolding and proteolytic activities, leading to increased tumor cell proliferation. Future studies are aimed at identifying potential miRNA inhibitors which enhance GRK2 expression and thereby suppress MALT1 pro-tumorigenic activity in ABC-DLBCL.

Citation Format: Jing Cheng, Mei Smyers, Matt Trotta, Neil M Carleton, Lisa M Maurer, Peter C Lucas, Linda M McAllister-Lucas. GRK2, an inhibitor of MALT1-dependent oncogenic signaling, is downregulated by microRNA in ABC-DLBCL [abstract]. In: Proceedings of the Third AACR International Meeting: Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2022 Jun 23-26; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2022;3(5_Suppl):Abstract nr A18.