Implication of the Long Non-Coding RNA Crnde in Multiple Myeloma

Multiple myeloma (MM) is a malignancy of antibody-secreting plasma cells which remains incurable, despite significant improvements in treatment and patient care. MM is characterized by a wide clinical and prognostic spectrum, even within groups bearing the same primary initiating cytogenetic event, for which the molecular mechanisms responsible remain poorly understood. Long non-coding RNAs (lncRNAs; broadly defined as non-coding RNAs of > 200 nt) have recently emerged as an important class of regulatory molecules, exercising diverse functions in normal cells and tissues, and are increasingly implicated in tumorigenesis and cancer progression. However at present the contribution of lncRNAs to the progression and clinical variability of MM is largely unknown.

In order to indentify lncRNAs potentially implicated in the etiology of MM, we have taken a data mining approach to compare the expression profile of lncRNAs between CD138+ plasmocytes from healthy individuals with those from MM patients. Patients were stratified according to the presence of the t(4;14) or t(11;14) translocations and a third group bearing neither of these cytogenetics events. This analysis revealed 110 lncRNAs that are differentially expressed in CD138+ plasma cells from newly diagnosed MM patients compared to control subjects. An interesting candidate to emerge from this analysis is Colorectal Neoplasia Differentially Expressed (CRNDE), a lncRNA that has previously been implicated in other hematological malignancies and diverse solid tumors, by promoting growth and inhibiting apoptosis potentially via a physical association with epigenetic modifying complexes (PRC2 and CoREST). Consistent with these observations in other malignancies, our findings suggest that CRNDE is also likely to have a role in the progression of MM. Expression of CRNDE increases with disease progression from the pre-malignant MGUS stage, through the stage of smoldering myeloma and reaches highest levels at progression to overt symptomatic myeloma. Further, survival analysis indicates that high expression of CRNDE is associated with poor overall survival, in particular in high risk t(4;14)+ myeloma patients.

To directly test the role of CRNDE in MM and to investigate downstream oncogenic pathways, we are using CRISPR/Cas9 to delete the major nuclear and cytoplasmic isoforms of CRNDE in MM cell lines. Our first results indicate that the deletion of CRNDE exerts a dose-dependent repressive effect on myeloma cell growth, supporting the idea that CRNDE is an important determinant of disease progression in MM. CRISPER/Cas9 deletion is being carried out in both a t(4;14) and non t(4;14) context. We have further investigated the effect of CRNDE deletions on the sensitivity of MM cells to therapeutic agents and are studying the interaction of CRNDE with Polycomb group proteins.

In summary, our results using CRISPR/Cas9-mediated deletion of the lncRNA CRNDE in MM cell lines together with the clinical data on patient survival strongly suggest that CRNDE plays an important role in MM disease progression and outcome. This work has the potential to translate into clinical benefit in a number of ways, notably by providing a useful biomarker of MM. Further, the development of molecular inhibitors of lncRNAs is a rapidly expanding field opening the way to the development of novel therapeutic strategies based on lncRNAs.