PB2050 USING A FUNCTIONAL SCREEN TO IDENTIFY NOVEL ANTI‐PROLIFERATIVE MIRNAS IN DIFFUSE LARGE B‐CELL LYMPHOMA (DLBCL)

Background: Diffuse large B‐cell lymphoma (DLBCL) is the most common B‐cell lymphoma, accounting for around 40% of all newly diagnosed non‐Hodgkin lymphoma (NHL) malignancies worldwide. Thanks to R‐CHOP therapy, the majority of DLBCL patients survive their disease. However, around 15% of patients treated have refractory disease, and of the patients that do respond to therapy, an additional 20‐25% will relapse. Proliferation is an important hallmark of cancer and in DLBCL has been associated with poor prognostic outcome. Consequently, several studies have focused on targeting molecules responsible for DLBCL proliferation as a novel therapeutic approach. MicroRNAs (miRNAs) are small non‐coding RNA molecules that act as key regulators of many biological and pathological processes including lymphomagenesis. miRNAs have been shown to be involved in many aspects of DLBLCL pathogenesis including initialization and progression of the disease, as well as having prognostic value. Aims: Identify miRNAs responsible the sustainment of proliferative signaling in DLBCL with the goal to develop novel anti‐lymphoma therapies. Methods: DLBCL cell lines (RIVA, SU‐DHL‐10, SU‐DHL‐4 and Oci‐ly‐10) were infected with a lentiviral library encoding for 698 miRNAs. We used a pooled‐approach using a MOI of 50. An empty vector was used as the control. Stable cell lines were produced by puromycin selection and cells maintained in culture for 4 months in triplicate. Both DNA and RNA were extracted at time 0, 2 months and 4 months post‐infection and the resultant DNA sequenced by NGS (Ion Torrent PGM). Differentially expressed miRNAs were detected by differential expression analysis (P < 0.05). For validation, selected miRNAs were infected individually in cell lines and proliferation rate (CFSE assay) and apoptosis (AnnexinV/TO‐PRO‐3) were determined by flow cytometry. Results: More than eighty miRNAs were identified as being differentially expressed between time 0 and time 4 months in each cell and five of these were differentially expressed in all cell lines. We validated five miRNAs by infecting cell lines with individual lentiviral vectors. Summary/Conclusion: We have demonstrated that the use of functional miRNA screens is a reliable tool for elucidating the phenotype‐genotype interface of cancers such as DLBCL. From our experiments, we propose that miRNA over‐expression could be used to decrease cell proliferation in DLBCL.