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Role of DNA methylation in the regulation of lincRNAs in Hematological Malignancies

Final Report Summary - LINCMHEM (Role of DNA methylation in the regulation of lincRNAs in Hematological Malignancies)

Traditionally, cancer researchers have focused their best efforts on studying the genetic alterations affecting the protein coding genes that lead to human tumors. In the last years, the different studies have shown that in addition to genetic alterations, epigenetic alterations affecting coding genes play an important role in tumor development. Epigenetic mechanisms refer to heritable changes in gene expression that occur in a cell without producing changes in the genomic sequence. In the last decade, largely due to the ability to sequence the human genome and the observation that much of the genome is transcribed but not translated, genes known as long non-coding RNAs (lncRNAs) have attracted great interest in relation to their biological functions, and their role in human diseases, especially cancer. But our knowledge of the mechanisms leading to the deregulation of the expression of these lncRNAs and real function of these lncRNAs in hematological malignancies is very limited.

The overall aim of LincMHeM project was to analyze the role of epigenetic regulation of lncRNAs in the pathogenesis and prognosis of B cell hematological malignancies (multiple myeloma, lymphoma and acute leukemia). To know more clearly the expression alteration of all types of lncRNAs in B cell hematological malignancies, first, we analyzed and re-annotated the all expressed lncRNAs in different normal B cell subsets that cover the human humoral immune response. We observed more than 15.000 lncRNAs expressed in normal B cells. Interestingly, some of these novel lncRNAs were transcribed from specific enhancer regions defined in B cell subpopulations and demethylated during B cell differentiation. Their expression was correlated with the expression of protein coding genes directly related with the B cell subsets identity and differentiation, indicating that lncRNAs play an important functional role. In the case of MM, a plasma cell neoplasm, we observed a highly heterogeneous DNA methylation pattern globally characterized by regional DNA hypermethylation embedded in extensive hypomethylation. The hypomethylation of the DNA occur in intergenic regions where some of the lncRNAs are located. We analyzed the whole transcriptome of plasma cells of 50 patients with MM and detected more than 30.000 lncRNAs expressed in MM patients. We observed that the inhibition of the expression of the candidate lncRNA, decrease the proliferation and increase the apoptosis of MM cells. These results show that the candidate lncRNA play an important role in the pathogenesis of MM and suggest that could be interesting therapeutic targets to MM. We are analyzing different strategies to inhibit its expression specifically in vivo in human plasma cells to use as a therapy for patients with this disease. In the case of lymphoma (DLBCL), we detected 47 lncRNAs up regulated and 27 lncRNAs down regulated between patient samples with Germinal Center type of DLBCL (GCB-DLBCL) and patient samples with Activated type of DLBCL (ABC-DLBCL). We performed the new analysis in 90 samples from DLBCL patients and 82 Follicular lymphoma (FL) patients and we are analyzing the data obtained to continue with the functional studies of lncRNAs. Finally, in acute leukemias, we observed that G9a (histone methyltransferase) and DNMTs are altered changing the expression of coding genes and lncRNAs. We designed and synthesized potent novel, selective and reversible chemical probes that simultaneously inhibit the G9a and DNMTs methyltransferase activity. The lead compound CM-272, inhibited cell proliferation and promoted apoptosis in vitro and significantly prolonged survival of in vivo AML, ALL and DLBCL xenogeneic models. Our results represent the discovery of first-in- class dual inhibitors of G9a/DNMTs and establish this chemical series, as a promising therapeutic tool for unmet needs in hematological tumors.

Potential Impact
The results and achievement of these objectives in the LincMHeM project, will tell us about the alteration and functional involvement of lncRNAs in hematological malignancies. The results obtained, will allow us to design and develop new strategies to modulate the expression and aberrant function of lncRNAs, enabling us to develop more appropriate treatments to improve the quality of life of patients with B cell hematological malignancies.