Background: B-cell chronic lymphocytic leukemia (CLL) is the most common adult leukemia in the western countries. Long non-coding RNAs (lncRNAs) emerge as a new class of ncRNAs measuring longer than 200 nucleotides in length, and implicated in a wide range of biological processes including normal development, differentiation or carcinogenesis [1]. Recently, deregulated expression of lncRNAs has been found in many human cancers, indicating its potential oncogenic or tumor suppressor role in carcinogenesis [2]. However, data on the role of lncRNA in CLL is scanty. BM742401 is a tumor suppressor lncRNA downregulated in gastric cancer [3]. As the promoter region and the entire transcript are embedded in a CpG island, we postulated that BM742401 is a tumor suppressor lncRNA inactivated by DNA methylation in CLL.

Methods: Methylation of BM742401 was studied by methylation-specific polymerase chain reaction (MSP) in nine healthy donor normal controls including three bone marrow, three peripheral blood buffy coats, and three CD19-sorted peripheral B-cells, in addition to seven CLL cell lines, and ninety-eight diagnostic CLL marrow samples.

Results: By MSP, promoter of BM742401 was unmethylated in normal controls, but methylated in four out of seven (57%) CLL cell lines. MSP statuses, including complete methylation (MM), partial methylation (MU), and complete unmethylation (UU), were verified by quantitative bisulfite pyrosequencing. Methylation of BM742401 was inversely correlated with expression. In the completely methylated WAC3CD5+ CLL cells, treatment with 5-Aza-2’-deoxycytidine led to promoter demethylation and re-expression of BM742401 transcript. Functionally, stable overexpression of BM742401 by lentiviral transduction resulted in inhibition of cellular proliferation and enhanced apoptosis through caspase-9-dependent intrinsic apoptosis pathway, suggesting a tumor suppressor role of BM742401 in CLL. In primary CLL samples, methylation of BM742401 was detected in 43/98 (44%) of patients. Moreover, among CLL patients with standard-risk cytogenetic aberrations, methylation of BM742401 correlated with advanced Rai stage (≥ stage 2) (P = 0.002). Furthermore, methylation of BM742401 was significantly associated with methylation of miR-129-2 (P = 0.034).

Conclusions: BM742401 is a tumor suppressor lncRNA frequently methylated in CLL. The functional mechanism of BM742401 in triggering tumor suppressive phenotype and regulation of other protein-coding genes warrant further studies.

References:

1. Esteller, M., Non-coding RNAs in human disease. Nature Reviews Genetics, 2011. 12(12): p. 861-874.

2. Prensner, J.R. and A.M. Chinnaiyan, The emergence of lncRNAs in cancer biology. Cancer discovery, 2011. 1(5): p. 391-407.

3. Park, S.-M., et al., A known expressed sequence tag, BM742401, is a potent lincRNA inhibiting cancer metastasis. Experimental & Molecular Medicine, 2013. 45(7): p. e31.

Citation Format: Lu Qian Wang, Kwan Yeung Wong, Zhen Hai Li, Chor Sang Chim. Epigenetic silencing of tumor suppressor long non-coding RNA BM742401 in chronic lymphocytic leukemia. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 946.