Differential micro-RNA expression in primary CNS and nodal diffuse large B-cell lymphomas

Abstract

Most primary CNS lymphomas (PCNSL) are diffuse large B-cell lymphomas (DLBCL). However, clinical behavior and prognosis differ considerably from those for nodal DLBCL (nDLBCL), and their pathogenesis is still not fully understood. Micro-RNAs (miRNAs) have been associated with cancer development and progression. We investigated a large miRNA panel for differential expression in PCNSL and nDLBCL, to determine new mechanisms potentially involved in PCNSL pathogenesis. Using paraffin-embedded biopsy specimens from 21 HIV-negative patients with newly diagnosed PCNSL (n = 11) and nDLBCL (n= 10), we measured the expression of 365 miRNA species by quantitative real-time PCR using low-density PCR arrays. We found that 18 miRNAs were differentially expressed: median expression levels of 13 miRNAs were 2.1-13.1 times higher in PCNSL, and median expression levels of 5 miRNAs were 2.6-3.3 times higher in nDLBCL. MiRNAs upregulated in PCNSL were associated with the Myc pathway (miR-17-5p, miR-20a, miR-9), with blocking of terminal B-cell differentiation (miR-9, miR-30b/c), or with upregulation by inflammatory cytokines (miR-155). Putative tumor-suppressor miRNAs (miR-199a, miR-214, miR-193b, miR-145) were downregulated in PCNSL. There was no overlap of miRNAs dysregulated in PCNSL with those differentially expressed between immunohistologically defined germinal center B cell-like (GCB) and non-GCB types or, apart from miR-9, with miRNAs known to be overexpressed in human brain. We conclude that PCNSL exhibits a distinct pattern of miRNA expression compared with nDLBCL. This argues for the involvement of different molecular mechanisms in the pathogenesis of these two lymphoma types.

Fig. 1.

Expression levels (relative to RNU44) of the 15 most abundant miRNAs in PCNSL and their respective expressions in nDLBCL. Bars depict median expression and interquartile range.

Fig. 2.

(A) Differential expression of specific miRNAs in PCNSL and nDLBCL: examples of significant overexpression (miR-17-5p and miR-20a) and downregulation (miR-214 and miR-145) in PCNSL (expression levels relative to RNU48; the line indicates the median, all P < .05). (B) MiR-155 is overexpressed in PCNSL compared with nDLBCL, whereas no significantly different expression for miR-155 is seen between GCB- and non–GCB-type lymphomas (expression levels relative to RNU44; the line indicates the median).

Fig. 3.

Supervised cluster analysis of miRNA levels in PCNSL and nDLBCL. Heatmap of differentially expressed miRNAs. Data normalized to RNU44 (dC_t) were hierarchically clustered (Pearson correlation, complete linkage). Red indicates a decrease relative to all data in this set; blue indicates an increase relative to all data in this set.

Fig. 4.

Expression of brain-enriched miRNAs in PCNSL and nDLBCL. MiR-9 was significantly higher expressed in PCNSL (P = .006), and miR-124a was nonsignificantly higher in PCNSL (P = .05); expressions of miR-125b and miR-128b were similar in both entities (P = .48 and 0.73).

Fig. 5.

Immunohistochemistry for Myc protein. Representative sections of a nodal DLBCL (A) and a PCNSL (B) (40×) demonstrate Myc positivity in both entities.