A microRNA Cluster as a Target of Genomic Amplification in B-Cell Lymphomas.

Background: Genomic gain/amplification of 13q31-q32 is frequently observed in malignant lymphomas. C13orf25, recently established as a candidate gene in malignant lymphoma via 13q31-q32 genomic amplification, encodes two variant transcripts by alternative splicing (Ota et al, Cancer Res 2004). Seven microRNA genes (miR-17-5p, miR-17-3p, miR-18, miR-19a, miR-19b, miR-20 and miR-92) are clustered in C13orf25 transcript variant 2 (C13orf25 v2). Because microRNAs display dynamic temporal and spatial expression patterns, disruption of these microRNAs may be associated with tumorigenesis.

Purpose: The purposes of this study are i) to reveal frequencies of the 13q gain/amplification in various lymphoma types, and ii) to examine the expression of C13orf25 v2 and seven microRNAs using various lymphoma cell lines and tumors with and without 13q gain/amplification.

Experimental Design: We analyzed genomic alterations of chromosome 13 for 12 malignant lymphoma cell lines (eight B-cell and four T-cell lymphomas), and 214 cases of B-cell lymphomas (136 cases of diffuse large B-cell lymphoma (DLBCL), 27 cases of sporadic Burkitt’s lymphoma (sBL), 29 of mantle cell lymphoma (MCL), 22 of follicular lymphoma (FCL)) and 20 cases of T-cell lymphoma by using array-based comparative genomic hybridization. The expression levels of seven microRNAs using 12 lymphoma cell lines with (four) and without (eight) 13q gain/amplification were examined by Northern-blot and quantitative real-time PCR (RQ-PCR) analyses. RQ-PCR for C13orf25 v2 (microRNA cluster) was also conducted for 21 cases of DLBCL (eight cases with 13q gain/13 cases without), 10 cases of sBL (four cases with 13q gain/amp/six cases without) and 10 cases of mantle cell lymphoma.

Results: Frequent (> 20%) gain/amplification of 13q were detected in DLBCL (31 cases, 23%) and Burkitt’s lymphoma (8 cases, 30%) but no gain/amplification at 13q was found in MCL, FCL and T-cell lymphomas. Genomic amplification of 13q31-q32 was observed in four cases of DLBCL and two cases of sBL, four of which were c-MYC rearranged (two cases of DLBCL and two cases of sBL). RQ-PCR and Northern blot analyses revealed that five of the seven mature microRNAs displayed overexpression in lymphoma cell lines with 13q31 genomic gain/amplification but not in those without. RQ-PCR analysis for 21 cases of DLBCL demonstrated that the cases with 13q gain/amplification (8 cases) showed significantly higher expression of C13orf25 v2 than those without (13 cases) (Mann Whitney U test, P < 0.05). Significant higher levels of the five microRNAs in sBL with 13q gain/amplification were also confirmed by Northern blot analysis. Lower expression levels of the microRNAs were found in T cell lymphoma cell lines and tumors.

Conclusion: These results suggest that the microRNA cluster (C13orf25 v2) is a target of 13q/13q31 genomic gain/amplification in DLBCL and sBL.