DNA methylation of microRNA and long non-coding RNA in multiple myeloma

Abstract

Multiple myeloma is one form of hematological malignancy characterized by the accumulation and patchy infiltration of the bone marrow by neoplastic plasma cells. It is often preceded by an entirely asymptomatic state, monoclonal gammopathy of undetermined significance (MGUS), that progresses into symptomatic myeloma at the rate of 1% per year. Currently, multiple myeloma remains an incurable disease. Therefore, further insight into the pathogenesis of disease is imperative. Non-coding RNAs (ncRNAs) are RNA molecules with no protein-coding capacity. Functional ncRNAs include microRNAs (miRNAs) and long non-coding RNAs (lncRNAs). miRNAs contain 19~25 nucleotides and will target and suppress the expression of their target genes by complementary binding to the 3’-UTR of the target mRNAs. lncRNAs are novel class of RNA molecules containing more than 200 nucleotides, which regulate gene expression at both transcriptional and post transcriptional levels by binding to transcription factors, genomic DNAs and mRNAs. Oncogenic or tumor suppressive roles of various miRNAs and lncRNAs have been proved in multiple cancer types, including myeloma. DNA methylation refers to the addition of a methyl group to carbon five position of the cytosine ring in a CpG dinucleotide. For regulation of ncRNAs, promoter DNA methylation at the CpG island is one of the most well-established mechanisms. Indeed, methylation has been implicated in regulation of numerous miRNAs and lncRNAs in myeloma. The thesis aimed to investigate the role of promoter DNA methylation in the regulation of candidate ncRNAs in myeloma. Moreover, the prognostic impact of DNA methylation and tumor suppressive function of ncRNAs were studied. Promoter DNA methylation of miRNAs (miR-340-5p, miR-342-3p and miR-28-5p) and lncRNAs (BM742401 and KIAA0495) were studied in myeloma. Epigenetic silencing of these ncRNAs by tumor-specific methylation were confirmed in human myeloma cell lines (HMCLs). For miR-28-5p, we proved for the first time to our knowledge that it is epigenetically regulated by promoter of its host gene. For miR-342-3p and miR-340-5p, comparable methylation frequencies were detected in primary bone marrows at MGUS and active myeloma at diagnosis and relapse, hence possibly an early event in myeloma pathogenesis. Moreover, methylation of miR-340-5p in diagnostic myeloma was correlated with significantly shorter overall survival (OS), which may serve as an adverse prognosis biomarker. Furthermore, the tumor suppressive roles of miR-340-5p were demonstrated by restoration of miR-340-5p in HMCL, leading to decreased cell proliferation via suppressing MAPK signaling and increased apoptosis via directly targeting X-linked inhibitor of apoptosis protein (XIAP). Besides, lncRNAs, including KIAA0495 and BM742401, are novel targets of DNA methylation in myeloma. In particular, methylation of BM742401 was detected at consecutive stages of myeloma pathogenesis, hence likely an early event, and it was correlated with significantly shorter OS. However, methylation of KIAA0495, as well as miR-28-5p, were rarely detected in primary samples, suggesting methylation of these ncRNAs in HMCLs were acquired during in vitro cell culture. In conclusion, methylation of miR-340-5p, miR-342-3p and BM742401 are implicated in pathogenesis of myeloma, and methylation of miR-340-5p and BM742401 in prognosis of myeloma.